(12) United States Patent (10) Patent No.: US 9,315,447 B2 Gjorstrup Et Al

USOO9315447B2

(12) United States Patent (10) Patent No.: US 9,315,447 B2 Gjorstrup et al. (45) Date of Patent: *Apr. 19, 2016

(54) COMPOSITIONS AND METHODS FOR THE (52) U.S. Cl. TREATMENT OF INFLAMMATION CPC ...... C07C 69/732 (2013.01); C07C 69/734 (2013.01) (71) Applicant: A.T. Resolve Sarl, St-Legier (CH) (58) Field of Classification Search (72) Inventors: Per Gjorstrup, Cambridge, MA (US); CPC ...... C07C 69/732; C07C 69/734 C. Eric SchwartZ, Wakefield, MA (US USPC ...... 514f63 . Lric SchwartZ, eleld, (US) See application file for complete search history. (73) Assignee: A.T. RESOLVE SARL, Lausanne (CH) (56) References Cited (*) Notice: Subject to any disclaimer, the term of this U.S. PATENT DOCUMENTS patent is extended or adjusted under 35 U.S.C. 154(b) by 0 days. 4,172,896 A 10/1979 Uno et al. 4,346,227 A 8, 1982 Terahara et al. This patent is Subject to a terminal dis- 4,444,784. A 4, 1984 Hoffman et al. claimer. 4,681,893 A 7, 1987 Roth 5,354,772 A 10, 1994 Kathawala (21) Appl. No.: 14/207,342 (Continued) (22) Filed: Mar 12, 2014 FOREIGN PATENT DOCUMENTS (65) Prior Publication Data WO 2006,105.058 A2 10, 2006 WO 2009/038671 3, 2009 US 2014/0275,247 A1 Sep. 18, 2014 (Continued) Related U.S. Application Data OTHER PUBLICATIONS (63) Continuation of application No. 13/264,155, filed as & 8 s application No. PCT/US2010/030812 on Apr. 13, Becker et al., “Hydrolyse von Carbonsaurederivaten . Organikum, 2010, now Pat No. 8673,881. pp. 414-415 (1990) (No translation available). (Continued) (60) Provisional application No. 61/174,806, filed on May on1, 2009 they isional applicauonapplication No.No 61/168,739,, /39, filedIlle Primary Examiner — Deborah D Carr (74) Attorney, Agent, or Firm — Kilpatrick Townsend & (51) Int. Cl. Stockton LLP AOIN 55/00 (2006.01) A6 IK3I/695 (2006.01) (57) ABSTRACT CO7D 303/2 (2006.01) The invention relates to novel resolvin compounds and phar C07C 59/00 (2006.01) maceutical preparations thereof. The invention further relates C07F 7/02 (2006.01) to methods of treatment using the novel resolvin compounds C07F II/00 (2006.01) of the invention. C07C 69/732 (2006.01) C07C 69/734 (2006.01) 16 Claims, 7 Drawing Sheets

A -- 1001 -- 102 E

windwavywww.www.wrwrrrmamer-rrm-www. 2 4.

. US 9,315,447 B2 Page 2

(56) References Cited Iigo et al., “Inhibitory effects of docosahexaenoic acid on colon carninoma 26 metastasis to the lung.” Br: J. Cancer, (1997), U.S. PATENT DOCUMENTS 75(5):650-655. Simopoulous et al., “Workshop on the Essentiality of and Recom 5,358,970 A 10, 1994 Ruffet al. mended Dietary Intakes for Omega-6 and Omega-3 Fatty Acids.” 5,427,798 A 6/1995 Ludwig et al. 5,541,231 A 7, 1996 Ruffet al. (1999), J. Am. Coll. Nutr., 18(5):487-489. 5,731,000 A 3, 1998 Ruffet al. International Search Reportdated Jun. 29, 2010 in related PCT Appli 5,763,493 A 6, 1998 Ruffet al. cation No. PCT/US2010/030813. 6,110,973 A 8/2000 Young European Search report dated Jul. 26, 2012 in related European RE37,314 E 8, 2001 Hirai et al. Patent Application No. 10764985.7. 6,583,124 B2 6/2003 Asgharian Mexican Office Action dated Sep. 14, 2012 in related Mexican Patent 2005, 0004074 A1 1/2005 Lyons et al. 2005/OO31697 A1 2/2005 Vehige et al. Application No. MX/a/2011/010827 (with English Translation). 2005/0059744 A1 3/2005 Donello et al. New Zealand Examination Report dated Aug. 29, 2012 in related 2005, 0080056 A1 4, 2005 Horn New Zealand Application No. 596228. 2005/0228042 A1 10, 2005 Friswad et al. U.S. Office Action dated May 22, 2013 in parent U.S. Appl. No. 2005/0228047 A1* 10, 2005 Petasis ...... 514,560 13/264,155, 5 pages. 2009,0294347 A1 12, 2009 Wochele et al. European Examination Report dated Nov. 13, 2013 in related Euro pean Patent Application No. 10764985.7. 5 pages. FOREIGN PATENT DOCUMENTS Russian Office Action dated Mar. 5, 2014 in related Russian Patent Application No. 2011 146005, 11 pages (with English Translation). WO 2009/051670 A2 4/2009 Mexican Office Action dated Apr. 24, 2014 in related Mexican Patent WO 2010/0395.61 4/2010 Application No. MX/a/2011/010827. 5 pages (with English Transla OTHER PUBLICATIONS tion). Chilean Office Action dated May 5, 2014 in related Chilean Patent Billman et al., “Prevention of Sudden Cardiac Death by Dietary Pure Application No. 2548-2011, 9 pages (with English Translation). w-3 Polyunsaturated Fatty Acids in Dogs.” Circulation, (1999), Japanese Office Action dated May 20, 2014 in related Japanese Patent 99:2452-2457. Application No. 2012-506110, 11 pages (with English Translation). Henkel-Hanke et al., “Artificial oxygen carriers: a current review.” AANA.J. (2007), 75(3):205-211. * cited by examiner U.S. Patent Apr. 19, 2016 Sheet 1 of 7 US 9,315,447 B2

p~~~~~••••••••••••••••••••••••------••••••••••~~~~~~~~--~~~~~~~~~~~~~~~.~~~~~~~);

w s w s wa (utifu) toeueolic (fruijfti) togeyua303

s o 's es se ***;" tliffju) togesteguio U.S. Patent Apr. 19, 2016 Sheet 2 of 7 US 9,315,447 B2

smas .

ac www. s: x \ {i : SN. Y. wi. S.x2 is: it c > ...... six is: C-3s. 2 is iii &f 88.

s: sa c s

$3. s U.S. Patent Apr. 19, 2016 Sheet 3 of 7 US 9,315,447 B2

s 3. s

S. S. S &s 3s & 8 Ys s^ rw --wa

U.S. Patent Apr. 19, 2016 Sheet 5 of 7 US 9,315.447 B2

Figure 5

U.S. Patent Apr. 19, 2016 Sheet 6 of 7 US 9,315,447 B2

Figure 6

U.S. Patent Apr. 19, 2016 Sheet 7 Of 7 US 9,315,447 B2

S. Figure 7

3. US 9,315,447 B2 1. 2 COMPOSITIONS AND METHODS FOR THE SUMMARY OF INVENTION TREATMENT OF INFLAMMATION The present invention provides a compound of formula I, RELATED APPLICATIONS (I) This application claims the benefit of priority to U.S. Pro visional Patent Application No. 61/168,739, filed Apr. 13, 2009, and U.S. Provisional Patent Application No. 61/174, 10 806, filed May 1, 2009, which applications are hereby incor porated by reference in their entirety. and pharmaceutically acceptable salts thereof, wherein: the stereochemistry of the carbon qq to carbon rr double BACKGROUND 15 bond is cis or trans; the stereochemistry of the carbon ss' to carbon tt'double bond is cis or trans; Supplementation of dietary omega-3 polyunsaturated fatty Re and Rfare independently selected from hydrogen, alkyl, acids (“c)-3 PUFAs) such as eicosapentaenoic acid, a com alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxyacyl, ponent of fish oils, may have beneficial effects in diseases aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl; E is a branched alkoxy Such as isopropoxy, isobutoxy, sec Such as arteriosclerosis, arthritis, asthma and cancer, which butoxy, tert-butoxy, 3-methylbutoxy, 2,2-dimethylpro may be mediated by antithrombotic, immunoregulatory and poxy, or 1.1.2-trimethylpropoxy. anti-inflammatory responses De Caterina, R. S. Endres, S. Rh and Ri are independently selected from hydrogen, alkyl, D. Kristensen, and E. B. Schmidt, editors. (1993). n-3 Fatty alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl; 25 Rs is selected from i-iv as follows: i) CHCH(R)CH, where Acids and Vascular Disease. Springer-Verlag, London; R is hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, Lands, W. E. M., editor. (1987). Proceedings of the AOCS aryl, heteroaryl, fluoro, hydroxyl or alkoxy; ii) CHC Short Course on Polyunsaturated Fatty Acids and (RR)CH, where RandR, are each independently alkyl, Eicosanoids. American Oil Chemists’ Society, Champaign, alkenyl, alkynyl, perfluoroalkyl, aryl, or fluoro, or R and 30 R, are connected together to form a carbocyclic or hetero Ill.: Iigo, M. et al. (1997) Br. J. Cancer 75:650. The potential cyclic ring; iii) CHOCH, CHC(O)CH, or CHCH; or of (D-3 PUFAs for preventative actions in cardiovascular dis iv) Rs is a carbocyclic, heterocyclic, aryl or heteroaryl ring: eases was recently supported by the finding that major dietary and Rs and R are independently selected from hydrogen, alkyl, c)-3 PUFAs, such as eicosapentaenoic acid (C20:5 co-3; EPA) alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or heteroaryl, and docosahexaenoic acid (C22:6 co-3; DHA), have a dra 35 or Rs and R are connected together to form a carbocyclic matic effect on ischemia-induced ventricular fibrillation or heterocyclic ring. Billman, G. E. et al. (1999) Circulation. 99:2452. Emer In certain embodiments, a compound of formula I is rep gence of Such possible preventative and/or therapeutic resented by formula II, actions of c)-3 PUFA supplementation in infant nutrition, 40 (II) cardiovascular diseases, and mental health has led to a call for ORf recommended dietary intakes by an international workshop Simopoulous, A. P. et al. (1999).J. Am. Coll. Nutr. 18:487. The Gruppo Italiano per lo Studio della Sopravvivense nell 45 Infarto Miocardio (GISSI) Prevenzione trial evaluated the effects of c)-3 PUFA supplementation with 11,300 patients and pharmaceutically acceptable salts thereof, wherein: surviving myocardial infarction taking ~1 g of c)-3 PUFA the stereochemistry of the carbon qq to carbon rr double daily (n=2,836) along with recommended preventive treat 50 bond is cis or trans; ments including aspirin, and reported a significant benefit the stereochemistry of the carbon ss' to carbon tt'double bond is cis or trans; with a decrease in cardiovascular death Marchioloi, R. et al. Re, Rf, Rs, and E are as defined above. (1999). Lancet. 354:447. However, the mechanisms under In certain embodiments, a compound of formula I or II is lying the protective action of dietary (D-3 PUFAs in these represented by formula III, studies and other studies including those concerned with dis 55 eases of the skin, bowel, and neural tissues are not currently understood. One of the many hypothesized elements of the (III) mechanism(s) of action of c)-3 PUFAs is that naturally occur ORf ORe O ring metabolites, formed from these PUFAs, may act as 60 mediators that provide important biological functions, but E, these metabolites may have relatively short half-lives in vivo. There remains a need for new analogues which may have greater in vivo stability than naturally occurring ()-3 PUFA 65 metabolites for settings where a longer half-life may be and pharmaceutically acceptable salts thereof, wherein: advantageous. Re, Rf, and E are as defined above. US 9,315,447 B2 3 4 In certain embodiments, the present invention provides a Re and Rfare independently selected from hydrogen, alkyl, pharmaceutical preparation Suitable for use in a human alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxyacyl, patient, comprising an effective amount of any of the com aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl, pref pounds shown above (e.g., a compound of the invention, Such erably from hydrogen, acyl (e.g., alkoxyacyl, aminoacyl), as a compound of any of formulae I-III), and one or more aminocarbonyl, and alkoxycarbonyl, most preferably pharmaceutically acceptable excipients. In certain embodi hydrogen; ments, the pharmaceutical preparations may be for use in E is a branched alkoxy Such as isopropoxy, isobutoxyt, Sec treating or preventing a condition or disease as described butoxy, tert-butoxy, 3-methylbutoxy, 2,2-dimethylpro herein. In certain embodiments, the pharmaceutical prepara poxy, or 1.1.2-trimethylpropoxy, preferably isopropoxy. tions have a low enough pyrogen activity to be suitable for 10 Rh and Ri are independently selected from hydrogen, alkyl, intravenous use in a human patient. alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl, prefer The present invention further provides methods of treating ably hydrogen or alkyl, most preferably hydrogen; or preventing bone metabolism, mucositis, cardiovascular Rs is selected from i-iv as follows: i) CHCH(R)CH, where disease, inflammatory diseases, metabolic diseases, oph 15 R is hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, thalmic conditions, immune function, pulmonary conditions, aryl, heteroaryl, fluoro, hydroxyl or alkoxy; ii) CHC gastrointestinal conditions, rheumatological conditions, der (RR)CH2, where RandR, are each independently alkyl, matological conditions, neurological conditions, cancer, alkenyl, alkynyl, perfluoroalkyl, aryl, or fluoro, or R and infectious conditions, degenerative conditions, gerontologi R, are connected together to form a carbocyclic or hetero cal conditions, and apoptotic conditions, reducing, prevent cyclic ring; iii) CHOCH, CHC(O)CH, CH, or ing or reversing organ damage, reducing and/or preventing CHCH, or iv) Rs is a carbocyclic, heterocyclic, aryl or stem cell damage and/or death, enhancing organ preservation heteroaryl ring, preferably (CH2); and and/or Survival, or enhancing stem cell preservation and/or Rs and Ro are independently selected from hydrogen, alkyl, Survival, as described herein, comprising administering a alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or heteroaryl, compound of the invention. 25 or Rs and Ro are connected together to form a carbocyclic or heterocyclic ring, preferably from hydrogen and alkyl, DETAILED DESCRIPTION OF THE DRAWINGS most preferably hydrogen. For example, the present invention provides a compound of FIG. 1 shows that comparable levels of compound 1003 formula Ia, were observed in the aqueous humor (FIG.1a), the vitreous 30 (FIG. 1b), and the cornea (FIG. 1c) upon topical ocular administration of compounds 1001 and 1002 to rabbits. (Ia) FIG. 2 shows data from measurements of ocular discom R. ORf Rh ORe Q fort on day 28 of a human dry eye study after topical admin 35 R8 ul istration of vehicle or compound 1001 at dose A, B, or C. 21a 21 S Rs E, FIG. 3 shows data from measurements of ocular discom fort on day 29 (approximately one day after last treatment) of a human dry eye study after topical administration of vehicle or compound 1001 at dose A, B, or C. FIG. 4 shows measurements of grittiness and dryness on 40 and pharmaceutically acceptable salts thereof, wherein: day 28 of a human dry eye study after topical administration Re and Rfare independently selected from hydrogen, alkyl, of vehicle or compound 1001 at dose A, B, or C. alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxyacyl, FIG.5 shows the "H NMR spectrum for compound 1001 in aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl, pref CDC1. erably from hydrogen, acyl (e.g., alkoxyacyl, aminoacyl), FIG. 6 shows the "H NMR spectrum for compound 1002 in 45 aminocarbonyl, and alkoxycarbonyl, most preferably CDC1. hydrogen; FIG. 7 shows the "H NMR spectrum for compound Z in E is a branched alkoxy Such as isopropoxy, isobutoxyt, Sec D.O. butoxy, tert-butoxy, 3-methylbutoxy, 2,2-dimethylpro poxy, or 1.1.2-trimethylpropoxy, preferably isopropoxy. DETAILED DESCRIPTION OF THE INVENTION 50 Rh and Ri are independently selected from hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl, prefer The present invention provides a compound of formula I, ably hydrogen or alkyl, most preferably hydrogen; Rs is selected from i-iv as follows: i) CHCH(R)CH, where 55 R is hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, (I) aryl, heteroaryl, fluoro, hydroxyl or alkoxy; ii) CHC (RR)CH2, where RandR, are each independently alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl, or fluoro, or R and R, are connected together to form a carbocyclic or hetero 60 cyclic ring; iii) CHOCH, CHC(O)CH, CH, or CHCH, or iv) Rs is a carbocyclic, heterocyclic, aryl or heteroaryl ring, preferably (CH2); and and pharmaceutically acceptable salts thereof, wherein: Rs and R are independently selected from hydrogen, alkyl, the stereochemistry of the carbon qq to carbon rr double alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or heteroaryl, bond is cis or trans; 65 or Rs and Ro are connected together to form a carbocyclic the stereochemistry of the carbon ss' to carbon tt'double bond or heterocyclic ring, preferably from hydrogen and alkyl, is cis or trans; most preferably hydrogen. US 9,315,447 B2 5 6 In certain preferred embodiments of formula Ia, the stere Exemplary compounds of formulae I, II, and III include ochemistry of the carbons bearing —ORf and —ORe are as compound 1001 shown in formula Ia',

1001

ORf OH O

10 \ s

and pharmaceutically acceptable salts thereof. In certain embodiments, a compound of formula I is rep In certain embodiments, the present invention provides a resented by formula II, 15 pharmaceutical preparation Suitable for use in a human patient, comprising an effective amount of any of the com pounds shown above (e.g., a compound of the invention, such

(II) as a compound of any of formulae and one or more pharma ceutically acceptable excipients. In certain embodiments, the pharmaceutical preparations may be for use in treating or preventing a condition or disease as described herein. In certain embodiments, the pharmaceutical preparations have a low enough pyrogen activity to be suitable for use in a human patient. 25 Compounds of any of the above structures may be used in and pharmaceutically acceptable salts thereof, wherein: the manufacture of medicaments for the treatment of any the stereochemistry of the carbon qq to carbon rr double diseases or conditions disclosed herein. Bone Metabolism bond is cis or trans; The present invention provides methods of treating or pre the stereochemistry of the carbon ss' to carbon tt'double bond venting bone loss in a patient comprising administering a is cis or trans; 30 compound of the invention. In certain embodiments, condi Re, Rf, Rs, and E are as defined above. tions with which the bone loss is associated include, for For example, the present invention provides a compound of example, but are not limited to any one or more of ankylosing spondylitis, renal osteodystrophy (e.g., in patients undergo formula IIa, ing dialysis), osteoporosis, glucocorticoid-induced 35 osteoporosis, Paget’s disease, abnormally increased bone (IIa) turnover, periodontitis, periodontal disease, bone fractures, ORf ORe Q rheumatoid arthritis, osteoarthritis, periprosthetic osteolysis, osteogenesis imperfecta, metastatic bone disease, hypercal 21N 21 S ls E, cemia of malignancy, multiple myeloma, bone loss associ 40 ated with microgravity, Langerhan's Cell Histiocytosis (LHC), bone loss associated with renal tubular disorders, or N bone loss associated with bed-ridden conditions. The present invention provides a method of treating or preventing osteoporosis in a patient comprising administer and pharmaceutically acceptable salts thereof, wherein: ing a compound of the invention. In certain embodiments, the Re, Rf, Rs, and E are as defined above. 45 osteoporosis is medicine-induced osteoporosis. In certain In certain embodiments, a compound of formula I or II is embodiments, the medicine-induced osteoporosis is gluco represented by formula III, corticoid-induced osteoporosis. The present invention provides a method of treating or preventing diabetic osteopenia in a patient comprising admin 50 istering a compound of the invention. (III) The present invention provides a method of treating or ORf ORe O preventing metastatic bone disease in a patient comprising administering a compound of the invention. The present invention provides a method of decreasing the incidence of 55 bone metastasis in a patient comprising administering a com pound of the invention. The present invention further pro vides a method of delaying the onset of bone metastasis in a patient comprising administering a compound of the inven and pharmaceutically acceptable salts thereof, wherein: tion. In certain embodiments, a compound of the invention is 60 administered conjointly with chemotherapy or radiation Re, Rf, and E are as defined above. therapy. In certain embodiments of Formulae I-III, E represents The present invention provides a method of treating or O—R, where R represents an alkyl group, preferably a lower preventing periodontitis in a patient comprising administer alkyl group, that is branched at the position bonded to the ing a compound of the invention. oxygen atom. Exemplary Such R moieties include —CH 65 The present invention provides a method of treating or (CH) (isopropyl), —CH(CHCH), —CH(CH) preventing gingivitis in a patient comprising administering a (CHCH) (sec-butyl), and —C(CH) (tert-butyl). compound of the invention. US 9,315,447 B2 7 8 The present invention provides a method of treating or factor-alpha, transforming growth factor-beta, bone morpho preventing ankylosing spondylitis in a patient comprising genetic protein, parathyroid hormone, osteoprotegerin, a administering a compound of the invention. fibroblast growth factor, Vitamin D, vitronectin, plasmino The present invention provides a method of treating or gen-activator inhibitor, or a protease inhibitor Such as a met preventing renal osteodystrophy (e.g., in patients undergoing alloprotease inhibitor, or elements known to be beneficial to dialysis) in a patient comprising administering a compound bone formation, such as calcium, fluoride, magnesium, of the invention. boron, or a combination thereof. In one embodiment, the method of treating or preventing In one embodiment, the method of treating or preventing bone loss may comprise administering a compound of the metastatic bone disease may comprise administering a com invention conjointly with an additional agent useful in the 10 pound of the invention conjointly with a chemotherapeutic treatment of bone loss. In certain embodiments, the com agent. Chemotherapeutic agents that may be conjointly pound of the invention may be conjointly administered with a administered with compounds of the invention include: ami bisphosphonate (e.g., ibandronate, Zolendronate, risedronate, noglutethimide, amsacrine, anastroZole, asparaginase, bcg, etidronate, or alendronate), a steroid, Such as an anabolic bicalutamide, bleomycin, buserelin, buSulfan, campothecin, steroid (e.g., testosterone, quinbolone, oxymetholone, nan 15 capecitabine, carboplatin, carmustine, chlorambucil, cispl drolone hexylphenylpropionate, Oxandrolone, testosterone atin, cladribine, clodronate, colchicine, cyclophosphamide, undecanoate, mibolerone, danozol, nandrolone decanoate, cyproterone, cytarabine, dacarbazine, dactinomycin, dauno trenbolone cyclohexylmethylcarbonate, methenolone rubicin, dienestrol, diethylstilbestrol, docetaxel, doxorubicin, acetate, methenolone enanthate, mesterolone, dihydrotest epirubicin, estradiol, estramustine, etoposide, exemestane, osterone, methandrostenolone, nandrolone undecanoate, filgrastim, fludarabine, fludrocortisone, fluorouracil, flu boldenone undecylenate, formebolone, trenbolone acetate, oxymesterone, flutamide, gemcitabine, genistein, goserelin, fluoxymesterone, nandrolone laureate, drostanolone propi hydroxyurea, idarubicin, ifosfamide, imatinib, interferon, onate, clostebol acetate, trestolone acetate, methandriol irinotecan, ironotecan, letrozole, leucovorin, leuprolide, dipropionate, methyltestosterone, furazabol, bolasterone, levamisole, lomustine, mechlorethamine, medroxyprogester norethandrolone, mepitioStane, tetrahydrogestrinone, tren 25 one, megestrol, melphalan, mercaptopurine, mesna, methotr bolone enanthate, and stanozolol), an estrogen (e.g., estra exate, mitomycin, mitotane, mitoxantrone, nilutamide, diol, estriol, estrone, equilin, or equilenin), a Substance hav nocodazole, octreotide, oxaliplatin, paclitaxel, pamidronate, ing estrogenic activity (e.g., Xenoestrogens, phytoestrogens, pentostatin, plicamycin, porfimer, procarbazine, raltitrexed, or mycoestrogens), a selective estrogen receptor modulator rituximab, Streptozocin, Suramin, tamoxifen, temozolomide, (e.g., raloxifene), or hormone treatment (e.g., calcitonin or 30 teniposide, testosterone, thioguanine, thiotepa, titanocene teriparitide). In certain embodiments, the compound of the dichloride, topotecan, trastuzumab, tretinoin, vinblastine, invention may be conjointly administered with growth factors Vincristine, vindesine, and vinorelbine. or other therapeutic agents that have a positive effect on the Many combination therapies have been developed for the growth of bone or connective tissue. Such as osteoprotegerin, treatment of cancer. In certain embodiments, compounds of interleukins, MMP inhibitors, beta glucans, integrin antago 35 the invention may be conjointly administered with a combi nists, calcitonin, proton pump inhibitors, protease inhibitors, nation therapy. Examples of combination therapies with insulin-like growth factor-1, platelet-derived growth factor, which compounds of the invention may be conjointly admin epidermal growth factor, inhibitors of transforming growth istered are included in Table 1. TABLE 1. Exemplary combinatorial therapies for the treatment of cancer. Name Therapeutic agents ABV Doxorubicin, Bleomycin, Vinblastine ABVD Doxorubicin, Bleomycin, Vinblastine, Dacarbazine AC (Breast) Doxorubicin, Cyclophosphamide AC (Sarcoma) Doxorubicin, Cisplatin AC (Neuroblastoma) Cyclophosphamide, Doxorubicin ACE Cyclophosphamide, Doxorubicin, Etoposide ACe Cyclophosphamide, Doxorubicin AD Doxorubicin, Dacarbazine AP Doxorubicin, Cisplatin ARAC-DNR Cytarabine, Daunorubicin B-CAWe Bleomycin, Lomustine, Doxorubicin, Vinblastine BCVPP Carmustine, Cyclophosphamide, Vinblastine, Procarbazine, Prednisone BEACOPP Bleomycin, Etoposide, Doxorubicin, Cyclophosphamide, Vincristine, Procarbazine, Prednisone, Filgrastim BEP Bleomycin, Etoposide, Cisplatin BIP Bleomycin, Cisplatin, Ifosfamide, Mesna BOMP Bleomycin, Vincristine, Cisplatin, Mitomycin CA Cytarabine, Asparaginase CABO Cisplatin, Methotrexate, Bleomycin, Vincristine CAF Cyclophosphamide, Doxorubicin, Fluorouracil CAL-G Cyclophosphamide, Daunorubicin, Vincristine, Prednisone, Asparaginase CAMP Cyclophosphamide, Doxorubicin, Methotrexate, Procarbazine CAP Cyclophosphamide, Doxorubicin, Cisplatin CaT Carboplatin, Paclitaxel CAV Cyclophosphamide, Doxorubicin, Vincristine US 9,315,447 B2 9 10 TABLE 1-continued Exemplary combinatorial therapies for the treatment of cancer. Name Therapeutic agents CAVE ADD CAV and Etoposide CA-VP16 Cyclophosphamide, Doxorubicin, Etoposide CC Cyclophosphamide, Carboplatin CDDPVP-16 Cisplatin, Etoposide CEF Cyclophosphamide, Epirubicin, Fluorouracil CEPP(B) Cyclophosphamide, Etoposide, Prednisone, with or without Bleomycin CEV Cyclophosphamide, Etoposide, Vincristine CF Cisplatin, Fluorouracil or Carboplatin Fluorouracil CHAP Cyclophosphamide or Cyclophosphamide, Altretamine, Doxorubicin, Cisplatin ChIVPP Chlorambucil, Vinblastine, Procarbazine, Prednisone CHOP Cyclophosphamide, Doxorubicin, Vincristine, Prednisone CHOP-BLEO Add Bleomycin to CHOP CISCA Cyclophosphamide, Doxorubicin, Cisplatin CLD-BOMP Bleomycin, Cisplatin, Vincristine, Mitomycin CMF Methotrexate, Fluorouracil, Cyclophosphamide CMFP Cyclophosphamide, Methotrexate, Fluorouracil, Prednisone CMFVP Cyclophosphamide, Methotrexate, Fluorouracil, Vincristine, Prednisone CMV Cisplatin, Methotrexate, Vinblastine CNF Cyclophosphamide, Mitoxantrone, Fluorouracil CNOP Cyclophosphamide, Mitoxantrone, Vincristine, Prednisone COB Cisplatin, Vincristine, Bleomycin CODE Cisplatin, Vincristine, Doxorubicin, Etoposide COMLA Cyclophosphamide, Vincristine, Methotrexate, Leucovorin, Cytarabine COMP Cyclophosphamide, Vincristine, Methotrexate, Prednisone Cooper Regimen Cyclophosphamide, Methotrexate, Fluorouracil, Vincristine, Prednisone COP Cyclophosphamide, Vincristine, Prednisone COPE Cyclophosphamide, Vincristine, Cisplatin, Etoposide COPP Cyclophosphamide, Vincristine, Procarbazine, Prednisone CP(Chronic Chlorambucil, Prednisone lymphocytic leukemia) CP (Ovarian Cancer) Cyclophosphamide, Cisplatin CT Cisplatin, Paclitaxel CVD Cisplatin, Vinblastine, Dacarbazine CV Carboplatin, Etoposide, Ifosfamide, Mesna CVP Cyclophosphamide, Vincristine, Prednisome CVPP Lomustine, Procarbazine, Prednisone CYVADIC Cyclophosphamide, Vincristine, Doxorubicin, Dacarbazine DA Daunorubicin, Cytarabine DAT Daunorubicin, Cytarabine, Thioguanine DAV Daunorubicin, Cytarabine, Etoposide DCT Daunorubicin, Cytarabine, Thioguanine DHAP Cisplatin, Cytarabine, Dexamethasone DI Doxorubicin, Ifosfamide DTIC, Tamoxifen Dacarbazine, Tamoxifen DVP Daunorubicin, Vincristine, Prednisone EAP oposide, Doxorubicin, Cisplatin EC oposide, Carboplatin EFP oposie, Fluorouracil, Cisplatin ELF oposide, Leucovorin, Fluorouracil EMA 86 itoxantrone, Etoposide, Cytarabine EP oposide, Cisplatin EVA oposide, Vinblastine FAC uorouracil, Doxorubicin, Cyclophosphamide FAM uorouracil, Doxorubicin, Mitomycin FAMTX Methotrexate, Leucovorin, Doxorubicin FAP uorouracil, Doxorubicin, Cisplatin F-CL, uorouracil, Leucovorin FEC uorouracil, Cyclophosphamide, Epirubicin FED uorouracil, Etoposide, Cisplatin FL utamide, Leuprolide utamide, Goserelin acetate implant HDMTX Methotrexate, Leucovorin Hexa-CAF Altretamine, Cyclophosphamide, Methotrexate, Fluorouracil CE-T fosfamide, Carboplatin, Etoposide, Paclitaxel, Mesna DMTXF6-MP Methotrexate, Mercaptopurine, Leucovorin E fosfamide, Etoposie, Mesna foVP fosfamide, Etoposide, Mesna PA fosfamide, Cisplatin, Doxorubicin US 9,315,447 B2 11 12 TABLE 1-continued Exemplary combinatorial therapies for the treatment of cancer. Name Therapeutic agents M-2 Vincristine, Carmustine, Cyclophosphamide, Prednisone, Melphalan MAC-III Methotrexate, Leucovorin, Dactinomycin, Cyclophosphamide MACC Methotrexate, Doxorubicin, Cyclophosphamide, Lomustine MACOP-B Methotrexate, Leucovorin, Doxorubicin, Cyclophosphamide, Vincristine, Bleomycin, Prednisone MAID Mesna, Doxorubicin, Ifosfamide, Dacarbazine m-BACOD Bleomycin, Doxorubicin, Cyclophosphamide, Vincristine, Dexamethasone, Methotrexate, Leucovorin MBC Methotrexate, Bleomycin, Cisplatin MC Mitoxantrone, Cytarabine MF Methotrexate, Fluorouracil, Leucovorin MICE fosfamide, Carboplatin, Etoposide, Mesna MINE Mesna, Ifosfamide, Mitoxantrone, Etoposide mini-BEAM Carmustine, Etoposide, Cytarabine, Melphalan MOBP Bleomycin, Vincristine, Cisplatin, Mitomycin MOP Mechlorethamine, Vincristine, Procarbazine MOPP Mechlorethamine, Vincristine, Procarbazine, Prednisone MOPPABV Mechlorethamine, Vincristine, Procarbazine, Prednisone, Doxorubicin, Bleomycin, Vinblastine MP (multiple Melphalan, Prednisone myeloma) MP (prostate cancer) Mitoxantrone, Prednisone MTXF6-MO Methotrexate, Mercaptopurine MTX6-MPVP Methotrexate, Mercaptopurine, Vincristine, Prednisone MTX-CDDPAdr Methotrexate, Leucovorin, Cisplatin, Doxorubicin MV (breast cancer) Mitomycin, Vinblastine MV (acute myelocytic Mitoxantrone, Etoposide leukemia) M-WAC Methotrexate Vinblastine, Doxorubicin, Cisplatin MVP Mitomycin Vinblastine, Cisplatin MVPP Mechlorethamine, Vinblastine, Procarbazine, Prednisone NFL Mitoxantrone, Fluorouracil, Leucovorin NOVP Mitoxantrone, Vinblastine, Vincristine OPA Vincristine, Prednisone, Doxorubicin OPPA Add Procarbazine to OPA. PAC Cisplatin, Doxorubicin PAC-I Cisplatin, Doxorubicin, Cyclophosphamide PA-CI Cisplatin, Doxorubicin PC Paclitaxel, Carboplatin or Paclitaxel, Cisplatin PCV Lomustine, Procarbazine, Vincristine PE Paclitaxel, Estramustine PFL Cisplatin, Fluorouracil, Leucovorin POC Prednisone, Vincristine, Lomustine ProMACE Prednisone, Methotrexate, Leucovorin, Doxorubicin, Cyclophosphamide, Etoposide ProMACE cytaBOM Prednisone, Doxorubicin, Cyclophosphamide, Etoposide, Cytarabine, Bleomycin, Vincristine, Methotrexate, Leucovorin, Cotrimoxazole PROMACEMOPP Prednisone, Doxorubicin, Cyclophosphamide, Etoposide, Mechlorethamine, Vincristine, Procarbazine, Methotrexate, Leucovorin Cisplatin, Teniposide Prednisone, Vincristine, Asparaginase Cisplatin, Vinblastine, Bleomycin Prednisone, Vincristine, Daunorubicin, Asparaginase Streptozocin, Mitomycin, Fluorouracil Mechlorethamine, Doxorubicin, Vinblastine, Vincristine, Bleomycin, Etoposide, Prednisone TCF Paclitaxel, Cisplatin, Fluorouracil TIP Paclitaxel, Ifosfamide, Mesna, Cisplatin TTT Methotrexate, Cytarabine, Hydrocortisone Topo/CTX Cyclophosphamide, Topotecan, MeSna VAB-6 Cyclophosphamide, Dactinomycin, Vinblastine, Cisplatin, Bleomycin WAC Vincristine, Dactinomycin, Cyclophosphamide WACAdr Vincristine, Cyclophosphamide, Doxorubicin, Dactinomycin, Vincristine WAD Vincristine, Doxorubicin, Dexamethasone WATH Vinblastine, Doxorubicin, Thiotepa, Flouxymesterone WBAP Vincristine, Carmustine, Doxorubicin, Prednisone VBCMP Vincristine, Carmustine, Melphalan, Cyclophosphamide, Prednisone VC Vinorelbine, Cisplatin US 9,315,447 B2 13 14 TABLE 1-continued Exemplary combinatorial therapies for the treatment of cancer. Name Therapeutic agents VCAP Vincristine, Cyclophosphamide, Doxorubicin, Prednisone VD Vinorelbine, Doxorubicin WeIP Vinblastine, Cisplatin, Ifosfamide, Mesna VIP Etoposide, Cisplatin, Ifosfamide, Mesna VM Mitomycin, Vinblastine VMCP Vincristine, Melphalan, Cyclophosphamide, Prednisone VP Etoposide, Cisplatin WTAD Etoposide, Thioguanine, Daunorubicin, Cytarabine 5 - 2 Cytarabine, Daunorubicin, Mitoxantrone 7 - 3 Cytarabine with, Daunorubicin or Idarubicin or Mitoxantrone 8 in 1 Methylprednisolone, Vincristine, Lomustine, Procarbazine, Hydroxyurea, Cisplatin, Cytarabine, Dacarbazine

In certain embodiments, a compound of the invention may In certain embodiments, the kit further comprises instruc be conjointly administered with non-chemical methods of tions for the administration of the pharmaceutical formula cancer treatment. In certain embodiments, a compound of the tion comprising a compound of the invention conjointly with invention may be conjointly administered with radiation a chemotherapeutic agent as mentioned above. In certain therapy. In certain embodiments, a compound of the inven embodiments, the kit further comprises a second pharmaceu tion may be conjointly administered with Surgery, with ther 25 tical formulation (e.g., as one or more single dosage forms) moablation, with focused ultrasound therapy, or with cryo comprising a chemotherapeutic agent as mentioned above. therapy. The present invention provides a kit comprising: In certain embodiments, different compounds of the inven a) a first pharmaceutical formulation (e.g., one or more tion may be conjointly administered with one or more other single dosage forms) comprising an agent Suitable for compounds of the invention. Moreover, Such combinations 30 the treatment or prevention of bone loss (e.g., a bispho may be conjointly administered with other therapeutic sphonate) as mentioned above; and agents, such as other agents suitable for the treatment of a b) instructions for the administration of the first pharma bone metabolism condition, Such as the agents identified ceutical formulation and a second pharmaceutical for above. mulation comprising a compound of the invention for In certain embodiments, the present invention provides a 35 treating or preventing bone loss, treating or inhibiting kit comprising: a) one or more single dosage forms of a the development of osteoporosis, treating or inhibiting compound of the invention; b) one or more single dosage the development of periodontitis, treating or inhibiting forms of a chemotherapeutic agent as mentioned above; and the development of metastatic bone disease, decreasing c) instructions for the administration of the compound of the the incidence of bone metastasis, or delaying the onset of invention and the chemotherapeutic agent. 40 bone metastasis. The present invention provides a kit comprising: The present invention provides a kit comprising: a) a pharmaceutical formulation (e.g., one or more single a) a first pharmaceutical formulation (e.g., one or more dosage forms) comprising a compound of the invention; single dosage forms) comprising a chemotherapeutic and agent as mentioned above; and b) instructions for the administration of the pharmaceutical 45 b) instructions for the administration of the first pharma formulation, e.g., for treating or preventing any of the ceutical formulation with a compound of the invention conditions discussed above, e.g., bone loss. for treating or preventing bone loss, treating or inhibit In certain embodiments, the kit further comprises instruc ing the development of osteoporosis, treating or inhib tions for the administration of the pharmaceutical formula iting the development of periodontitis, treating or inhib tion comprising a compound of the invention conjointly with 50 iting the development of metastatic bone disease, an agent Suitable for the treatment or prevention of bone loss decreasing the incidence of bone metastasis, or delaying (e.g., a bisphosphonate) as mentioned above. In certain the onset of bone metastasis. embodiments, the kit further comprises a second pharmaceu Mucositis tical formulation (e.g., as one or more single dosage forms) The present invention provides a method of treating or comprising an agent Suitable for the treatment or prevention 55 preventing mucositis comprising administering a compound of bone loss (e.g., a bisphosphonate) as mentioned above. of the invention. Mucositis, for the purposes of this applica The present invention provides a kit comprising: tion, refers to mucosal injury induced by or associated with a) a pharmaceutical formulation (e.g., one or more single the administration of radiation or drugs (chemotherapy) for dosage forms) comprising a compound of the invention; the treatment of cancer and related diseases. Mucositis typi and 60 cally manifests itself as ulcerations, tissue necrosis, and atro b) instructions for the administration of the pharmaceutical phy of the mucous membranes anywhere along the digestive formulation for treating or inhibiting the development of tract, from the mouth to the anus. For example, the present osteoporosis, treating or inhibiting the development of methods may be used to treat ulcerations and tissue necrosis periodontitis, treating or inhibiting the development of associated with radiation therapy and/or chemotherapy. metastatic bone disease, decreasing the incidence of 65 The present invention provides a method of preventing the bone metastasis, or delaying the onset of bone metasta development of chemotherapy or radiation therapy-induced S1S. mucositis comprising administering a compound of the US 9,315,447 B2 15 16 invention. In certain embodiments, a compound of the inven embodiments, the kit further comprises a second pharmaceu tion is administered conjointly with chemotherapy or radia tical formulation (e.g., as one or more single dosage forms) tion therapy. comprising a chemotherapeutic agent as mentioned above. The present invention provides a method of improving The present invention provides a kit comprising: Survival rates by reducing the incidence of therapy-induced a) a first pharmaceutical formulation (e.g., one or more mucositis comprising administering a compound of the single dosage forms) comprising an agent Suitable for invention. The rate of life-threatening severe mucositis, grade the treatment or prevention of mucositis (e.g., an anti 4 on WHO scale, would be expected to be reduced from an microbial agent, a growth factor, an agent that inhibits average incidence of 60% in untreated patients, to 20% or less the synthesis of ceramide, an agent that blocks the activ in patents receiving a Subject treatment. 10 ity of ceramide, or an agent that degrades ceramide) as In one embodiment, the method of treating or preventing mentioned above; and mucositis may comprise administering a compound of the b) instructions for the administration of the first pharma invention conjointly with an additional agent useful in the ceutical formulation with a compound of the invention, treatment of mucositis. In certain embodiments, the com e.g., for treating or preventing mucositis, preventing the pound of the invention may be conjointly administered with 15 development of chemotherapy or radiation therapy-in an antimicrobial agent. In certain embodiments, the com duced mucositis, or improving Survival rates by reduc pound of the invention may be conjointly administered with a ing the incidence of therapy-induced mucositis. growth factor. In certain embodiments, the compound of the The present invention provides a kit comprising: invention may be conjointly administered with an agent that a) a first pharmaceutical formulation (e.g., one or more inhibits the synthesis of ceramide, an agent that blocks the single dosage forms) comprising a chemotherapeutic activity of ceramide, or an agent that degrades ceramide. agent as mentioned above; and In one embodiment, the method of treating or preventing b) instructions for the administration of the first pharma mucositis may comprise administering a compound of the ceutical formulation with a compound of the invention, invention conjointly with a chemotherapeutic agent. Chemo e.g., for treating or preventing mucositis, preventing the therapeutic agents that may be conjointly administered with 25 development of chemotherapy or radiation therapy-in compounds of the invention include any Suitable chemothera duced mucositis, or improving Survival rates by reduc peutic agent or combination therapy as set forth above. ing the incidence of therapy-induced mucositis. In certain embodiments, different compounds of the inven Cardiovascular Disease tion may be conjointly administered with one or more other The present invention provides a method of treating or compounds of the invention. Moreover, Such combinations 30 preventing cardiovascular disease in a patient comprising may be conjointly administered with other therapeutic administering to said patient a compound of the invention. In agents, such as other agents suitable for the treatment or certain embodiments, the method comprises optional con prevention of mucositis, Such as the agents identified above. joint administration with a statin. In certain embodiments, the present invention provides a Cardiovascular disease refers to one or more disease states kit comprising: a) one or more single dosage forms of a 35 of the cardiovascular tree (including the heart). Diseases of compound of the invention; b) one or more single dosage the cardiovascular tree and diseases of dependent organs forms of a chemotherapeutic agent as mentioned above; and include, for example, but are not limited to any one or more c) instructions for the administration of the compound of the of: invention and the chemotherapeutic agent. disorders of the heart muscle (cardiomyopathy or myocardi The present invention provides a kit comprising: 40 tis) such as idiopathic cardiomyopathy, metabolic cardiomy a) a pharmaceutical formulation (e.g., one or more single opathy which includes diabetic cardiomyopathy, alcoholic dosage forms) comprising a compound of the invention; cardiomyopathy, drug-induced cardiomyopathy, ischemic and cardiomyopathy, and hypertensive cardiomyopathy; b) instructions for the administration of the pharmaceutical atheromatous disorders of the major blood vessels (mac formulation e.g., for treating or preventing mucositis, 45 rovascular disease) Such as the aorta, the coronary arteries, preventing the development of chemotherapy or radia the carotid arteries, the cerebrovascular arteries, the renal tion therapy-induced mucositis, or improving Survival arteries, the iliac arteries, the femoral arteries, and the rates by reducing the incidence of therapy-induced popliteal arteries; mucositis. toxic, drug-induced, and metabolic (including, but not limited In certain embodiments, the kit further comprises instruc 50 to, hypertensive and/or diabetic) disorders of small blood tions for the administration of the pharmaceutical formula vessels (microvascular disease) Such as the retinal arterioles, tion comprising a compound of the invention conjointly with the glomerular arterioles, the Vasa nervorum, cardiac arteri an agent Suitable for the treatment or prevention of mucositis oles, and associated capillary beds of the eye, the kidney, the (e.g., an antimicrobial agent, a growth factor, an agent that heart, and the central and peripheral nervous systems; and, inhibits the synthesis of ceramide, an agent that blocks the 55 plaque rupture of atheromatous lesions of major blood ves activity of ceramide, or an agent that degrades ceramide) as sels such as the aorta, the coronary arteries, the carotid arter mentioned above. In certain embodiments, the kit further ies, the cerebrovascular arteries, the renal arteries, the iliac comprises a second pharmaceutical formulation (e.g., as one arteries, the femoral arteries and the popliteal arteries. or more single dosage forms) comprising an agent Suitable for Yet other disorders that may be treated with compounds of the treatment of mucositis (e.g., an antimicrobial agent, a 60 the invention include restenosis, e.g., following coronary growth factor, an agent that inhibits the synthesis of ceramide, intervention, and disorders relating to an abnormal level of an agent that blocks the activity of ceramide, or an agent that high density and low density cholesterol. degrades ceramide) as mentioned above. In certain embodiments, the present invention provides In certain embodiments, the kit further comprises instruc methods of treating a vascular disease or disorder. In certain tions for the administration of the pharmaceutical formula 65 embodiments, the vascular disorder may include any vascular tion comprising a compound of the invention conjointly with disease or disorder which comprises an autoimmune element, a chemotherapeutic agent as mentioned above. In certain for example one which is caused by an autoimmune response. US 9,315,447 B2 17 18 Exemplary vascular disorders include one or more of 7-dimethyl-8-2-(2R,4R)-tetrahydro-4-hydroxy-6-oxo-2H Raynaud's disease and phenomenon, anterior uveitis, vascu pyran-2-ylethyl-1-naphthalenyl ester), pravastatin (BR.6R, litis, obliterative vascular disorder, atheroma formation, arte 1S,2S,6S,8S,8aR)-1,2,6,7,8,8a-hexahydro-B.B.6- riosclerosis, arteritis (e.g., Takayasu arteritis, temporal arteri trihydroxy-2-methyl-8-(2S)-2-methyl-1-oxobutoxy-1- tis/giant cell arteritis), myointimal hyperplasia (natural or 5 naphthaleneheptanoic acid), simvastatin (2,2-dimethyl following angioplasty), inflammatory and autoimmune thick butanoic acid (1S,3R,7S,8S,8aR)-1,2,3,7,8,8a-hexahydro-3, ening of the intima and/or muscular layer of blood vessels, 7-dimethyl-8-2-(2R,4R)-tetrahydro-4-hydroxy-6-oxo-2H inflammatory blood vessel lesions, atherosclerotic heart dis pyran-2-ylethyl-1-naphthalenyl ester), rosuvastatin ((3R, ease, reperfusion injury, cardiac conduction disturbances, 5S,6E)-7-4-(4-fluorophenyl)-6-(1-methylethyl)-2-methyl myocarditis, and myocardial infarction. 10 (methylsulfonyl)amino-5-pyrimidinyl-3,5-dihydroxy-6- In certain embodiments, the present invention provides a method of treating or preventing heart attack or dysrhythmia heptenoic acid), eptastatin, pitavastatin ((3R,5S,6E)-7-2- in a patient comprising administering to said patient a com cyclopropyl-4-(4-fluorophenyl)-3-quinolinyl-3,5- dihydroxy-6-heptenoic acid), cerivastatin ((3R,5S,6E)-7-4- pound of the invention. In certain embodiments, the present (4-fluorophenyl)-5-(methoxymethyl)-2,6-bis(1- invention provides a method of preventing cardiac death in a 15 patient comprising administering to said patient a compound methylethyl)-3-pyridinyl-3,5-dihydroxy-6-heptenoic acid), of the invention. In certain embodiments, the method com berivastatin ((R*.S*-(E)-7-(4-(4-fluorophenyl)spiro(2H-1- prises optional conjoint administration with a statin. benzopyran-2,1'-cyclopentan)-3-yl)-3,5-dihydroxy-ethyl ester), dalvastatin ((4R,6S)-rel-6-(1E)-2-[2-(4-fluoro-3-me Compounds of the invention are capable of resolving thylphenyl)-4,4,6,6-tetramethyl-1-cyclohexen-1-yl)ethenyl inflammation. Several aspects of cardiovascular disease, in tetrahydro-4-hydroxy-, 2H-Pyran-2-one), glenvastatin ((4R, particular the formation of atherosclerotic vessel wall 6S)-6-(1E)-2-[4-(4-fluorophenyl)-2-(1-methylethyl)-6- plaques, are believed to be intimately related to inflammation. phenyl-3-pyridinylethenyltetrahydro-4-hydroxy-2H Today it is believed that serum markers of inflammation such Pyran-2-one), RP 61969 (2S-2a(E),4?-4-(4- as CRP may be as predictive of risk of cardiovascular disease fluorophenyl)-2-(1-methylethyl)-3-2-(tetrahydro-4- as elevated levels of LDL. Thus, compounds of the invention 25 hydroxy-6-oxo-2H-pyran-2-yl)ethenyl-1(2H)- are useful in treating or preventing cardiovascular disease. In isoquinolinone), SDZ-265859, BMS-180431 ((3R,5S,6E)- certain embodiments, compounds of the invention are useful rel-9.9-bis(4-fluorophenyl)-3,5-dihydroxy-8-(1-methyl-1H for the treatment or prevention of arterial inflammation and/or tetrazol-5-yl)-6,8-Nonadienoic acid), CP-83101 ((3R,5S, artherosclerosis. 6E)-rel-3,5-dihydroxy-9,9-diphenyl-6,8-Nonadienoic acid Another mechanism by which compounds of the invention 30 may be effective in treating or preventing cardiovascular dis methyl ester), dihydromevinolin ((2S)-2-methyl-butanoic ease is by inhibiting the structural and functional modifica acid (1S,3S,4aR.7S,8S,8aS)-1,2,3,4,4a,7,8,8a-octahydro-3, tions of HDL that are an immediate effect of the acute phase 7-dimethyl-8-2-(2R,4R)-tetrahydro-4-hydroxy-6-oxo-2H response commonly seen in cardiovascular disease with pyran-2-yl)ethyl-1-naphthalenyl ester), and L-669262 (2.2- active atherosclerotic vessel wall plaques. Thus, compounds 35 dimethyl-butanoic acid (1S,7R,8R,8aR)-1,2,6,7,8,8a of the invention may increase HDL levels (or prevent the hexahydro-3,7-dimethyl-6-oxo-8-2-(2R,4R)-tetrahydro-4- decrease of HDL levels) and restore the LDL scavenging hydroxy-6-oxo-2H-pyran-2-yl)ethyl-1-naphthalenyl ester). effects of HDL. This may lead to a lower and improved serum For example, statins suitable for use in the methods of this LDL/HDL ratio. invention include statins of formula 200: In addition to increasing HDL levels, statins also demon 40 strate anti-inflammatory activity which contributes to their 200

ability to lower cardiovascular disease risk and treat cardio vascular disease. However, the full anti-inflammatory poten tial of statins cannot be utilized clinically as a monotherapy due to the high doses required, which can lead to an increased 45 rate and severity level of treatment-limiting adverse events, notably liver toxicity. Advantageously and Surprisingly, treatment or prevention of cardiovascular disease with a combination of a statin and a compound of the invention leads to a mutual enhancement of 50 both the anti-inflammatory properties and the serum HDL elevating properties of the two classes of compounds while avoiding the risks associated with high doses of statins alone. In methods of the invention, wherein a compound of the invention is administered conjointly with a statin (i.e., an 55 wherein HMG-CoA reductase inhibitor), the statin may be chosen Ro is selected from alkyl, alkenyl, alkynyl, cycloalkyl or from any statin known in the art. Statins suitable for said aralkyl; conjoint administration include, but are not limited to, meV Rao, Rao and R2 are independently selected from hydro astatin ((2S)-2-methylbutanoic acid (1S,7S,8S,8aR)-1,2,3,7, gen, halogen, alkyl, alkenyl or alkynyl: 8.8a-hexahydro-7-methyl-8-2-(2R,4R)-tetrahydro-4-hy 60 Ros and Roe are independently selected from hydrogen, droxy-6-OXO-2H-pyran-2-yl)ethyl-1-naphthalenyl ester), halogen, alkyl, alkenyl, alkynyl, cycloalkyl, aralkyl, atorvastatin (BR, 8R)-2-(4-fluorophenyl)-3,6-dihydroxy-5- alkoxy or aralkoxy; and (1-methylethyl)-3-phenyl-4-(phenylamino)carbonyl)-1H Rao is selected from hydrogen, R or M: Pyrrole-1-heptanoic acid), fluvastatin ((3R,5S,6E)-rel-7-3- Ra is a physiologically acceptable and hydrolyzable ester (4-fluorophenyl)-1-(1-methylethyl)-1H-indol-2-yl)-3,5- 65 group; and dihydroxy-6-heptenoic acid), lovastatin (2CS)-2-methyl M is a pharmaceutically acceptable cation; butanoic acid (1S,3R,7S,8S,8aR)-1,2,3,7,8,8a-hexahydro-3, or enantiomers or salts or hydrates thereof. US 9,315,447 B2 19 20 Other statins suitable for use in the methods of this inven -continued tion include statins of formula 201: F wherein A is selected from

10 / N n/ N 4.

B is selected from

R207 O

s 40 C1 and C2 are joined by a single or a double bond; Roz is selected from CORs, CONRR or CHOR, or Rao, and Roo can form a lactone; Rs is selected from Hora cationic salt moiety, or CORs 45 forms a pharmaceutically acceptable ester moiety; Ros, Roo and Rao are independently selected from H. C(O) R21 or C(O)NR2 R22. R and R2 are independently selected from H. alkyl, alk 50 enyl or alkynyl: R is selected from H or C(O)Ra; and Ra is selected from alkyl, alkenyl or alkynyl. Other statins suitable for use in the methods of this inven 55 tion include statins of formula 202:

2O2 R216 R222 60

R520 65 R218 R219 US 9,315,447 B2 21 wherein R is selected from 204 R232

HO O O HO CO A4 O OH R233 V

wherein one of R2so and R2 is

R is selected from OH, CHCO or RCO: 15 R is a branched or straight C-C alkyl, C-C alkenyl, or / s R234 C-C alkynyl: ==7. R235 R7, Ras and Rio are independently selected from H. C-Cs R236 alkyl, C-C alkenyl, C-C alkynyl or C-C acyl; and Ro is selected from H or CH. and the other is primary or secondary Calkyl, alkenyl, or Other statins suitable for use in the methods of this inven alkynyl not containing an asymmetric carbon atom, C tion include statins of formula 203: cycloalkyl or phenyl-(CH2)—; Ra is selected from hydrogen, C alkyl, C-C alkenyl, 25 C-C alkynyl. n-butyl, butyl, t-butyl, C. alkoxy, n-bu 2O3 OH toxy, i-butoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy; Rs is selected from hydrogen, C. alkyl, C-C alkenyl, R223 C-C alkynyl, Calkoxy, trifluoromethyl, fluoro, chloro, R224 phenoxy or benzyloxy; R is selected from hydrogen, C. alkyl, Calkenyl, C. ar\le \o-soH S. alkynyl, Calkoxy, fluoro or chloro; R525 m is selected from 1, 2 or 3, with the provisos that both Rs R226 and R2 must be hydrogen when R is hydrogen, R 35 must be hydrogen when Rs is hydrogen, not more than one of R and Rs is trifluoromethyl, not more than one wherein of R24 and R2ss is phenoxy, and not more than one of R2 R27 is —CH2—, —CH2—CH2—, —CH2—CH)—CH2 and Rs is benzyloxy; R is selected from hydrogen, C alkyl, C-C alkenyl, or —CH-CH(CH)–: 40 R is 1-naphthyl: 2-naphthyl, cyclohexyl, norbornenyl: 2-, C-C alkynyl. n-butyl, butyl, t-butyl, C. cycloalkyl, C 3-, or 4-pyridinyl; phenyl, phenyl substituted with fluorine, alkoxy, n-butoxy, i-butoxy, trifluoromethyl, fluoro, chloro, chlorine, bromine, hydroxyl trifluoromethyl; alkyl, alk phenoxy or benzyloxy; R is selected from hydrogen, C. alkyl, C-C alkenyl, enyl, oralkynyl of from one to four carbonatoms, alkoxy of alkynyl, C- alkoxy, trifluoromethyl, fluoro, chloro, phe from one to four carbon atoms, or alkanoyloxy of from two 45 noxy or benzyloxy, with the provisos that R must be to eight carbon atoms; hydrogen when R is hydrogen, not more than one of Either R or Rs is —CONRSR-6 where Rs and Ro RandR is trifluoromethyl, not more than one of Rs. are independently hydrogen; alkyl, alkenyl, or alkynyl of and R2 is phenoxy, and not more than one of R and from one to six carbonatoms; 2-, 3-, or 4-pyridinyl; phenyl: R2s is benzyloxy, phenyl Substituted with fluorine, chlorine, bromine, cyano, 50 Rs7 is selected from —(CH), or—CH=CH-, wherein trifluoromethyl, or carboalkoxy of from three to eight car n is 0, 1, 2 or 3: bonatoms; and the other of R or Rs is hydrogen; alkyl, Rs is selected from alkenyl, or alkynyl of from one to six carbonatoms; cyclo propyl, cyclobutyl, cyclopentyl; cyclohexyl; phenyl; or phenyl substituted with fluorine, chlorine, bromine, 55 R239 hydroxyl; trifluoromethyl; alkyl, alkenyl, or alkynyl of H H2 H2 from one to four carbon atoms, alkoxy of from one to four C-C ––c -COR240 or carbon atoms, or alkanoyloxy of from two to eight carbon OH OH atoms; and OH R is alkyl, alkenyl, or alkynyl of from one to six carbon 60 R239: atoms; cyclopropyl; cyclobutyl, cyclopentyl; cyclohexyl, or trifluoromethyl; O or the hydroxyl acids, and pharmaceutically acceptable salts thereof, derived from the opening of the lactone ring. 65 O Other statins suitable for use in the methods of this inven tion include statins of formula 204: US 9,315,447 B2 23 24 Ro is selected from hydrogen, or C. alkyl, C-C alkenyl, the dotted lines at X,Y and Zrepresent possible double bonds, or C-C alkynyl: said double bonds, when any are present, being either X Rao is selected from hydrogen, R or M: and Z in combination or X, Y or Zalone; Ra is a physiologically acceptable and hydrolyzable ester or the corresponding dihydroxy acid of formula 206a group; and 5 M is a pharmaceutically acceptable cation. Other statins suitable for use in the methods of this inven 206a tion include statins of formula 205: HO CO2H 10 205 OH HOC us HO R244 15

R242 2O wherein or a pharmaceutically acceptable salt of said acid, a Calkyl Ra is selected from ester of said acid or a phenyldimethylamino-, or acety lamino-Substituted-Calkyl ester of said acid. Other statins suitable for use in the methods of this inven O 25 tion include statins of formula 207:

HO N O R245 R349- N - O 35 wherein Ras is lower alkyl, alkenyl, alkynyl, aryl or aralkyl, each of which may have one or more Substituents; OH 40 Rae and R27 independently are selected from hydrogen, lower alkyl, alkenyl, alkynyl, oraryl, and each of said lower or ring-closed lactones, salts or esters thereof. alkyl, alkenyl, alkynyl and aryl may have one or more Substituents; Other statins suitable for use in the methods of this inven Ras is hydrogen, lower alkyl, alkenyl, alkynyl, or a cation tion include statins of formula 206: 45 capable of forming a non-toxic pharmaceutically accept able salt; 2O6 R is sulfur, oxygen, or sulfonyl, orimino which may have HO O a Substituent; and the dotted line represents the presence or absence of a double 50 bond; O or the corresponding ring-closed lactone. The synthesis of various statins is set forth in U.S. Pat. No. RE37,314 E., U.S. Pat. No. 4,444,784, U.S. Pat. No. 4,346, ul 227, U.S. Pat. No. 5,354,772, U.S. Pat. No. 4,681,893 and US 55 2005/0228.042. In another embodiment, the invention provides a method of raising serum HDL concentration (or preventing a decrease in serum HDL concentration) or decreasing the serum LDL/ HDL ratio in a patient, said method comprising administering 60 to said patient a compound of the invention, optionally in wherein combination with a statin. The patient to be treated in this R is selected from H or CH: method may have a total serum cholesterol level of greater R is selected from 1,1-dimethylpropyl: Cocycloalkyl: than 189 mg/dl., preferably higher than 200 mg/dl and most Coalkenyl; CCF-Substituted alkyl; phenyl: preferably higher than 240 mg/dl; and/or a serum LDL con halophenyl; phenyl-C-alkyl; Substituted phenyl-C 65 centration of greater than 130 mg/dl., preferably greater than alkyl in which the substituent is halo, Calkyl or 160 mg/dl., and most preferably higher than 189 mg/dl. In Calkoxy. addition to serum cholesterol and/or LDL levels, other factors US 9,315,447 B2 25 26 to be considered are the presence or absence of coronary agent, a calcium channel blocker, an antianginal drug, a disease and risk factors, such as age (45 or over for men, 55 or diuretic, a cardioplegic Solution, a cardiotonic agent, an anti over for women), family history of coronary heart disease, arrhythmic drug, a fibrinolytic agent, a Sclerosing solution, a Smoking, high blood pressure, serum HDL cholesterol level. vasoconstrictoragent, a nitric oxide donor, a potassium chan or presence of diabetes. nel blocker, a sodium channel blocker, an antihyperlipidemic In certain embodiments, the invention provides a method drug, an immunosuppressant, or a natriuretic agent. of lowering triglyceride levels in a patient, said method com Examples of a cyclooxygenase inhibitor include, but are prising administering to said patient a compound of the inven not limited to, celecoxib, rofecoxib, meloxicam, Valdecoxib, tion, optionally in combination with a statin. aspirin or indomethacin. In certain embodiments, the patient to be treated in this 10 An example of a thromboxane receptor antagonist is method of the invention may already be receiving a choles ifetroban. terol-lowering drug. In one preferred embodiment, the patient Examples of vasodilators include, e.g., bencyclane, cin is already taking a statin, such as one of the statins described narizine, citicoline, cyclandelate, cyclonicate, ebumamonine, above; and will continue to take that drug conjointly with a phenoxeZyl, flunarizine, ibudilast, ifenprodil, lomerizine, compound of the invention. Alternatively, the compound of 15 naphlole, nikamate, nosergoline, nimodipine, papaverine, the invention may be used as a replacement for the previously pentifylline, nofedoline, Vincamin, Vinpocetine, Vichizyl. administered cholesterol-lowering drug. pentoxifylline, prostacyclin derivatives (such as prostaglan In a related embodiment, the invention provides a method din E1 and prostaglandin I2), an endothelin receptor blocking of reducing the dose of a statin required to achieve a desired drug (such as bosentan), diltiazem, nicorandil, and nitroglyc increase in serum HDL, a decrease in serum LDL/HDL ratio C1. or serum total cholesterol level, and/or a decrease in triglyc Examples of the cerebral protecting drug include radical eride level. Reducing the dose of statins while maintaining Scavengers (such as edaravone, vitamin E, and vitamin C), potent serum lipid-reducing properties is highly desirable due glutamate antagonists, AMPA antagonists, kainate antago to side effects associated with certain statins. Well-known nists, NMDA antagonists, GABA agonists, growth factors, side effects include, deleterious changes in liver function, 25 opioid antagonists, phosphatidylcholine precursors, seroto muscle pain, weakness, muscle tenderness, myopathy. Other nin agonists, Na"/Ca" channel inhibitory drugs, and K" side effects of statins include reduced cognition, memory channel opening drugs. impairment, depression, irritability, non-muscle pain, periph Examples of the brain metabolic stimulants include aman eral neuropathy, sleep disorders, sexual dysfunction, fatigue, tadine, tiapride, and gamma-aminobutyric acid. dizziness, Swelling, shortness of breath, vision changes, 30 Examples of the anticoagulant include heparins (such as changes in temperature regulation, weight change, hunger, heparin Sodium, heparin potassium, dalteparin sodium, dalte breast enlargement, blood Sugar changes, dry skin, rashes, parin calcium, heparin calcium, parnaparin Sodium, reviparin blood pressure changes, nausea, upset stomach, bleeding, and Sodium, and danaparoid sodium), warfarin, enoxaparin, arga ringing in ears or other noises. troban, batroXobin, and sodium citrate. In this embodiment, the dose of a statin is reduced by at 35 Examples of the antiplatelet drug include ticlopidine least 5%, at least 10%, at least 15%, at least 20%, at least 25%, hydrochloride, dipyridamole, cilostazol, ethyl icosapentate, at least 30%, at least 40%, at least 50%, at least 60%, at least Sarpogrelate hydrochloride, dilaZephydrochloride, trapidil, a 70%, or more. The actual reduction in statin dose will depend nonsteroidal antiinflammatory agent (such as aspirin), bera upon the nature of the compound of the invention being prostsodium, iloprost, and indobufene. administered, the amount of compound of the invention being 40 Examples of the thrombolytic drug include urokinase, tis administered, and the reduction in serum lipid and/or triglyc Sue plasminogen activator (tPA), recombinant tEPA, issue-type eride level desired, as well as other factors set forth elsewhere plasminogen activators (such as alteplase, tisokinase, in this application that are typically considered in treating a nateplase, pamiteplase, monteplase, and rateplase), streptoki disease or condition. The amount of compound of the inven nase, urokinase, prourokinase, anisoylated plasminogen tion administered in this method will also depend upon the 45 streptokinase activator complex (APSAC, Eminase, Bee factors set forth above, as well as the nature and amount of cham Laboratories), animal salivary gland plasminogen acti statin being administered. In certain embodiments, the vators, and nasaruplase. amount of compound of the invention administered in this Examples of the antihypertensive drug include angiotensin method is less than 5%, less than 10%, less than 15%, less converting enzyme inhibitors (such as captopril, alacepril, than 20%, less than 25%, less than 30%, less than 40%, less 50 lisinopril, imidapril, quinapril, temocapril, delapril, than 50%, less than 60%, less than 70%, less than 80%, or less benazepril, cilaZapril, trandolapril, enalapril, ceronapril, fosi than 90% of the dose of compound of the invention required nopril, imadapril, mobertpril, perindopril, ramipril, spirapril, to produce an anti-inflammatory effect. In other embodi Zofenopril, pentopril, randolapril and salts of Such com ments, the amount of compound of the invention adminis pounds), angiotensin II antagonists (such as losartan, cande tered is over 110%, over 120%, over 130%, over 140%, over 55 Sartan, Valsartan, eprosartan, and irbesartan), calcium chan 150%, over 160%, over 170%, over 180%, over 190%, or nel blocking drugs (such as aranidipine, efonidipine, even over 200% of the dose of compound of the invention nicardipine, bamidipine, benidipine, manidipine, cilnidipine, required to produce an anti-inflammatory effect. nisoldipine, nitrendipine, nifedipine, nilvadipine, felodipine, In one embodiment, the method of treating or preventing amlodipine, diltiazem, bepridil, clentiazem, phendilin, galo cardiovascular disease according to this invention may com 60 pamil, mibefradil, prenylamine, Semotiadil, terodiline, Vera prise the additional step of conjointly administering to the pamil, cilnidipine, elgodipine, isradipine, lacidipine, lercani patient another agent suitable for treating cardiovascular dis dipine, nimodipine, cinnarizine, flunarizine, lidoflazine, ease, Such as, for example, a cyclooxygenase inhibitor, a lomerizine, bencyclane, etafenone, and perhexyline), thromboxane receptor antagonist, a prostacyclin mimetic, a B-adrenaline receptor blocking drugs (propranolol, pindolol. phosphodiesterase inhibitor, a vasodilator, a cerebral protect 65 indenolol, carteolol, bunitrolol, atenolol, acebutolol, meto ing drug, a brain metabolic stimulant, an anticoagulant, an prolol, timolol, nipradillol, penbutolol, nadolol, tilisolol, antiplatelet drug, a thrombolytic drug, an antihypertensive carvedilol, bisoprolol, betaxolol, celiprolol, bopindolol, US 9,315,447 B2 27 28 bevantolol, labetalol, alprenolol, amoSulalol, arotinolol. 6E)-7-4-(4-fluorophenyl)-6-(1-methylethyl)-2-methyl befunolol, bucumolol, bufetolol, buferalol, buprandolol. (methylsulfonyl)amino-5-pyrimidinyl-3,5-dihydroxy-6- butylidine, butofilolol, carazolol, cetamolol, cloranolol, dil heptenoic acid), eptastatin, pitavastatin ((3R,5S,6E)-7-2- evalol, epanolol, levobunolol, mepindolol, metipranolol. cyclopropyl-4-(4-fluorophenyl)-3-quinolinyl-3,5- moprolol, nadoxolol, nevibolol, Oxprenolol, practol, pron dihydroxy-6-heptenoic acid), cerivastatin ((3R,5S,6E)-7-4- etalol, Sotalol, Sufinalol, talindolol, tertalol, toliprolol, (4-fluorophenyl)-5-(methoxymethyl)-2,6-bis(1- Xybenolol, and esmolol), C.-receptor blocking drugs (such as methylethyl)-3-pyridinyl-3,5-dihydroxy-6-heptenoic acid), amoSulalol, praZosin, teraZosin, doxazosin, bunaZosin, urapi berivastatin ((R*.S*-(E)-7-(4-(4-fluorophenyl)spiro(2H-1- dil, phentolamine, arotinolol, dapiprazole, fenspiride, benzopyran-2,1'-cyclopentan)-3-yl)-3,5-dihydroxy-ethyl indoramin, labetalol, naftopidil, nicergoline, tamsulosin, 10 tolaZoline, trimaZosin, and yohimbine), sympathetic nerve ester), dalvastatin ((4R,6S)-rel-6-(1E)-2-[2-(4-fluoro-3-me inhibitors (such as clonidine, guanfacine, guanabenz, meth thylphenyl)-4,4,6,6-tetramethyl-1-cyclohexen-1-yl)ethenyl yldopa, and reserpine), hydralazine, todralazine, budralazine, tetrahydro-4-hydroxy-, 2H-Pyran-2-one), glenvastatin ((4R, and cadralazine. 6S)-6-(1E)-2-[4-(4-fluorophenyl)-2-(1-methylethyl)-6- Examples of the antianginal drug include nitrate drugs 15 phenyl-3-pyridinylethenyltetrahydro-4-hydroxy-2H (such as amyl nitrite, nitroglycerin, and isosorbide), Pyran-2-one), RP 61969 (2S-2a(E),4?-4-(4- B-adrenaline receptor blocking drugs (exemplified above), fluorophenyl)-2-(1-methylethyl)-3-2-(tetrahydro-4- calcium channel blocking drugs (exemplified above) tri hydroxy-6-oxo-2H-pyran-2-yl)ethenyl-1(2H)- metazidine, dipyridamole, etafenone, dilaZep, trapidil, nic isoquinolinone), SDZ-265859, BMS-180431 ((3R,5S,6E)- orandil, enoxaparin, and aspirin. rel-9.9-bis(4-fluorophenyl)-3,5-dihydroxy-8-(1-methyl-1H Examples of the diuretic include thiazide diuretics (such as tetrazol-5-yl)-6,8-Nonadienoic acid), CP-83101 ((3R,5S, hydrochlorothiazide, methyclothiazide, bendrofluazide, 6E)-rel-3,5-dihydroxy-9,9-diphenyl-6,8-Nonadienoic acid chlorothiazide, trichlormethiazide, benzylhydrochlorothiaz methyl ester), dihydromevinolin ((2S)-2-methyl-butanoic ide, flumethiazide, hydroflumethiazide, bendroflumethiaz acid (1S,3S4aR,7S,8S,8aS)-1,2,3,4,4a,7,8,8a-octahydro-3, ide, methylchlorthiazide, polythiazide, benzthiazide and pen 25 7-dimethyl-8-2-(2R,4R)-tetrahydro-4-hydroxy-6-oxo-2H flutizide), loop diuretics (such as furosemide, etacrynic acid, pyran-2-yl)ethyl-1-naphthalenyl ester), and L-669262 (2.2- bumetanide, piretanide, azosemide, and torasemide), K" dimethyl-butanoic acid (1S,7R,8R,8aR)-1,2,6,7,8,8a sparing diuretics (spironolactone, triamterene, amiloride, and hexahydro-3,7-dimethyl-6-oxo-8-2-(2R,4R)-tetrahydro-4- potassium canrenoate), osmotic diuretics (such as isosorbide, hydroxy-6-oxo-2H-pyran-2-yl)ethyl-1-naphthalenyl ester). D-mannitol, and glycerin), nonthiazide diuretics (such as 30 Examples of the immunosuppressant include azathioprine, meticrane, tripamide, chlorthalidone, and mefruside), and mizoribine, cyclosporine, tacrolimus, gusperimus, and meth acetazolamide. Otrexate. Examples of the cardiotonic include digitalis formulations In one embodiment, the method of treating or preventing (such as digitoxin, digoxin, methyldigoxin, deslanoside, cardiovascular disease according to this invention may com Vesnarinone, lanatoside C, and proscillaridin), Xanthine for 35 prise administering a compound of the invention conjointly mulations (such as aminophylline, choline theophylline, with non-chemical means for the treatment or prevention of diprophylline, and proxyphylline), catecholamine formula cardiovascular disease. Such as part of a regimen including tions (such as dopamine, dobutamine, and docarpamine), physical intervention (e.g., percutaneous transluminal coro PDE III inhibitors (such as aminone, olprinone, and mil nary angioplasty, coronary Surgery, or vascular Surgery). In rinone), denopamine, ubidecarenone, pimobendan, levosim 40 certain Such embodiments, the regimen including physical endan, aminoethylsulfonic acid, Vesnarinone, carperitide, intervention may further include conjointly administering and colforsin daropate. another agent Suitable for the treatment or prevention of car Examples of the antiarrhythmic drug include ajmaline, diovascular disease, such as the agents listed above. pirmenol, procainamide, cibenzoline, disopyramide, quini It should be understood that the methods of treatment or dine, aprindine, mexiletine, lidocaine, phenyloin, pilsicam 45 prevention of cardiovascular disease according to this inven ide, propafenone, flecamide, atenolol, acebutolol, Sotalol, tion may include conjointly administering one or more of the propranolol, metoprolol, pindolol, amiodarone, nifekalant, above agents either as a separate dosage form or as part of a diltiazem, bepridil, moricizine, tocamide, encamide, pro composition that also comprises a statin, a compound of the pafenone, esmolol, artilide, bretylium, clofilium, isobutilide, invention, and optionally further comprising a statin. More Sotalol, azimilide, dofetilide, dronedarone, ersentilide, ibutil 50 over, the use of a composition comprising both a statin and a ide, tedisamil, trecetilide, digitalis, adenosine, nickel chlo compound of the invention according to this invention in the ride, and magnesium ions and Verapamil. treatment of cardiovascular disease, does not preclude the Examples of the antihyperlipidemic drug include atorvas separate but conjoint administration of another statin. tatin, simvastatin, pravastatin Sodium, fluvastatin Sodium, cli The method of increasing serum HDL concentration, nofibrate, clofibrate, simfibrate, fenofibrate, bezafibrate, 55 reducing serum LDL/HDL ratio, reducing total serum cho colestimide, colestyramine, mevastatin ((2S)-2-methyl lesterol concentration, and/or lowering the triglyceride level butanoic acid (1S,7S,8S,8aR)-1,2,3,7,8,8a-hexahydro-7-me in a patient according to this invention may additionally com thyl-8-2-(2R-4R)-tetrahydro-4-hydroxy-6-oxo-2H-pyran prise administering to said patient another active ingredient 2-yl)ethyl-1-naphthalenyl ester), fluvastatin ((3R,5S,6E)- other than a statin. Such additional active ingredient may be rel-7-3-(4-fluorophenyl)-1-(1-methylethyl)-1H-indol-2- 60 selected from a non-statin cholesterol lowering reagent. Such yl-3,5-dihydroxy-6-heptenoic acid), lovastatin (2(S)-2- as bile acid sequestrants (colesevelam, cholestyramine and methyl-butanoic acid (1S,3R,7S,8S,8aR)-1,2,3,7,8,8a colestipol), niacin, fibrates (gemfibrozil, probucol and clofi hexahydro-3,7-dimethyl-8-2-(2R,4R)-tetrahydro-4- brate). hydroxy-6-oxo-2H-pyran-2-yl)ethyl-1-naphthalenyl ester), In certain embodiments, different compounds of the inven pravastatin ((BR.6R,1S,2S,6S,8S,8aR)-1,2,6,7,8,8a-hexahy 65 tion may be conjointly administered with one or more other dro-3,3,6-trihydroxy-2-methyl-8-(2S)-2-methyl-1-oxobu compounds of the invention. Moreover, Such combinations toxy-1-naphthaleneheptanoic acid), rosuvastatin ((3R,5S, may be conjointly administered with other therapeutic US 9,315,447 B2 29 30 agents, such as other agents Suitable for the treatment or statin or another agent Suitable for treating cardiovascu prevention of cardiovascular disease, such as the agents iden lar disease. Such as, for example, a cyclooxygenase tified above. inhibitor, a thromboxane receptor antagonist, a prosta In certain embodiments, the present invention provides a cyclin mimetic, a phosphodiesterase inhibitor, a vasodi kit comprising: lator, a cerebral protecting drug, a brain metabolic a) one or more single dosage forms of a compound of the stimulant, an anticoagulant, an antiplatelet drug, a invention; thrombolytic drug, an antihypertensive agent, a calcium b) one or more single dosage forms of a statin as mentioned channel blocker, an antianginal drug, a diuretic, a car above; and dioplegic Solution, a cardiotonic agent, an antiarrhyth c) instructions for the administration of the compound of 10 mic drug, a fibrinolytic agent, a Sclerosing Solution, a the invention and the statin. vasoconstrictor agent, a nitric oxide donor, a potassium The present invention provides a kit comprising: channel blocker, a sodium channel blocker, an antihy a) a pharmaceutical formulation (e.g., one or more single perlipidemic drug, an immunosuppressant, or a natri dosage forms) comprising a compound of the invention; uretic agent); and and 15 b) instructions for the administration of the first pharma b) instructions for the administration of the pharmaceutical ceutical formulation with a compound of the invention, formulation e.g., for treating or preventing cardiovascu e.g., for treating or preventing cardiovascular disease, lar disease, raising serum HDL concentration (or pre raising serum HDL concentration (or preventing a venting a decrease in serum HDL concentration), decrease in serum HDL concentration), decreasing the decreasing the serum LDL/HDL ratio, and/or lowering serum LDL/HDL ratio, and/or lowering triglyceride lev triglyceride levels. els. In certain embodiments, the kit further comprises instruc The present invention provides a kit comprising: tions for the administration of the pharmaceutical formula a) a first pharmaceutical formulation (e.g., one or more tion comprising a compound of the invention conjointly with single dosage forms) comprising a statin as mentioned an agent Suitable for treating or preventing cardiovascular 25 above; and disease, raising serum HDL concentration (or preventing a b) instructions for the administration of the first pharma decrease in serum HDL concentration), decreasing the serum ceutical formulation with a compound of the invention, LDL/HDL ratio, and/or lowering triglyceride levels (e.g., a e.g., for treating or preventing cardiovascular disease, statin or another agent Suitable for treating cardiovascular raising serum HDL concentration (or preventing a disease. Such as, for example, a cyclooxygenase inhibitor, a 30 decrease in serum HDL concentration), decreasing the thromboxane receptor antagonist, a prostacyclin mimetic, a serum LDL/HDL ratio, and/or lowering triglyceride lev phosphodiesterase inhibitor, a vasodilator, a cerebral protect els. ing drug, a brain metabolic stimulant, an anticoagulant, an General Inflammatory Diseases antiplatelet drug, a thrombolytic drug, an antihypertensive The present invention provides a method of treating or agent, a calcium channel blocker, an antianginal drug, a 35 preventing inflammatory disease in a patient comprising diuretic, a cardioplegic Solution, a cardiotonic agent, an anti administering to said patient a compound of the invention. arrhythmic drug, a fibrinolytic agent, a Sclerosing solution, a Examples of inflammatory conditions, which may be vasoconstrictoragent, a nitric oxide donor, a potassium chan treated or prevented by the administration of a compound of nel blocker, a sodium channel blocker, an antihyperlipidemic the invention include, but are not limited to, inflammation of drug, an immunosuppressant, or a natriuretic agent) as men 40 the lungs, joints, connective tissue, eyes, nose, bowel, kidney, tioned above. In certain embodiments, the kit further com liver, skin, central nervous system, vascular system and heart. prises a second pharmaceutical formulation (e.g., as one or In certain embodiments, inflammatory conditions which may more single dosage forms) comprising an agent Suitable for be treated by the current invention include inflammation due treating or preventing cardiovascular disease, raising serum to the infiltration of leukocytes or other immune effector cells HDL concentration (or preventing a decrease in serum HDL 45 into affected tissue. Other relevant examples of inflammatory concentration), decreasing the serum LDL/HDL ratio, and/or conditions which may be treated by the present invention lowering triglyceride levels (e.g., a statin or another agent include inflammation caused by infectious agents, including, Suitable for treating cardiovascular disease. Such as, for but not limited to, viruses, bacteria fungi and parasites. example, a cyclooxygenase inhibitor, a thromboxane receptor Inflammatory lung conditions include, but are not limited antagonist, a prostacyclin mimetic, a phosphodiesterase 50 to, asthma, adult respiratory distress syndrome, bronchitis, inhibitor, a vasodilator, a cerebral protecting drug, a brain pulmonary inflammation, pulmonary fibrosis, and cystic metabolic stimulant, an anticoagulant, an antiplatelet drug, a fibrosis (which may additionally or alternatively involve the thrombolytic drug, an antihypertensive agent, a calcium gastro-intestinal tract or other tissue(s)). Inflammatory joint channel blocker, an antianginal drug, a diuretic, a cardiople conditions include rheumatoid arthritis, rheumatoid gic solution, a cardiotonic agent, an antiarrhythmic drug, a 55 spondylitis, juvenile rheumatoid arthritis, osteoarthritis, fibrinolytic agent, a Sclerosing solution, a vasoconstrictor gouty arthritis and other arthritic conditions. Eye diseases agent, a nitric oxide donor, a potassium channel blocker, a with an inflammatory component include, but are not limited Sodium channel blocker, an antihyperlipidemic drug, an to, uveitis (including iritis), conjunctivitis, Scleritis, kerato immunosuppressant, or a natriuretic agent) as mentioned conjunctivitis sicca, and retinal diseases, including, but not above. 60 limited to, diabetic retinopathy, retinopathy of prematurity, The present invention provides a kit comprising: retinitis pigmentosa, and dry and wet age-related macular a) a first pharmaceutical formulation (e.g., one or more degeneration. Inflammatory bowel conditions include single dosage forms) comprising an agent Suitable for Crohn's disease, ulcerative colitis and distal proctitis. treating or preventing cardiovascular disease, raising Inflammatory skin diseases include, but are not limited to, serum HDL concentration (or preventing a decrease in 65 conditions associated with cell proliferation, such as psoria serum HDL concentration), decreasing the serum LDL/ sis, eczema and dermatitis, (e.g., eczematous dermatitides, HDL ratio, and/or lowering triglyceride levels (e.g., a topic and seborrheic dermatitis, allergic or irritant contact US 9,315,447 B2 31 32 dermatitis, eczema craquelee, photoallergic dermatitis, pho The present invention also provides methods for treating totoxic dermatitis, phytophotodermatitis, radiation dermati ischemia by administering an effective amount of a com tis, and stasis dermatitis). Other inflammatory skin diseases pound of the invention. In certain embodiments, the ischemia include, but are not limited to, Scleroderma, ulcers and ero is cardiac ischemia, cerebral ischemia, bowel ischemia (e.g., sions resulting from trauma, burns, bullous disorders, or ischemic colitis or mesenteric ischemia), or cutaneous ischemia of the skin or mucous membranes, several forms of ischemia. ichthyoses, epidermolysis bullosae, hypertrophic scars, kel The present invention provides a method of treating or oids, cutaneous changes of intrinsic aging, photoaging, fric preventing inflammatory disease in a patient comprising tional blistering caused by mechanical shearing of the skin administering to said patient a compound of the invention 10 conjointly with a glucocorticoid. and cutaneous atrophy resulting from the topical use of cor The present invention provides a method of treating or ticosteroids. Additional inflammatory skin conditions include preventing an inflammatory condition (e.g., any of the inflam inflammation of mucous membranes, such as cheilitis, matory conditions described above) comprising administer chapped lips, nasal irritation, mucositis and Vulvovaginitis. ing to said patient a compound of the invention conjointly Inflammatory disorders of the endocrine system include, 15 with a glucocorticoid. but are not limited to, autoimmune thyroiditis (Hashimoto's Compounds of the invention are capable of resolving disease). Type I diabetes, and acute and chronic inflammation inflammation. Glucocorticoids are also known for their role of the adrenal cortex. Inflammatory conditions of the cardio in treating inflammation. However, the full anti-inflammatory vascular system include, but are not limited to, coronary potential of glucocorticoids is often clinically constrained as infarct damage, peripheral vascular disease, myocarditis, vas a monotherapy due to the rate and severity of treatment culitis, revascularization of Stenosis, artherosclerosis, and limiting adverse events that accompany high or prolonged vascular disease associated with Type II diabetes. dosing regimens. For example, the administration of gluco Inflammatory condition of the kidney include, but are not corticoids can result in side effects that mimic Cushings limited to, glomerulonephritis, interstitial nephritis, lupus disease. These side effects and others associated with gluco nephritis, nephritis secondary to Wegener's disease, acute 25 corticoid use include increased appetite and weight gain, renal failure secondary to acute nephritis, Goodpasture’s Syn deposits of fat in the chest, face, upper back, and stomach, drome, post-obstructive syndrome and tubular ischemia. water and salt retention leading to Swelling and edema, high Inflammatory conditions of the liver include, but are not blood pressure, diabetes, slow healing of wounds, osteoporo limited to, hepatitis (arising from viral infection, autoimmune sis, cataracts, acne, muscle weakness, thinning of the skin, responses, drug treatments, toxins, environmental agents, or 30 increased Susceptibility to infection, Stomach ulcers, as a secondary consequence of a primary disorder), biliary increased Sweating, mood Swings, psychological problems atresia, primary biliary cirrhosis and primary sclerosing cho Such as depression, and adrenal Suppression and crisis. langitis. Advantageously, treatment of inflammatory disease with a Inflammatory conditions of the central nervous system combination of a glucocorticoid and a compound of the include, but are not limited to, multiple Sclerosis and neuro 35 invention enhances the anti-inflammatory properties of both degenerative diseases such as Alzheimer's disease, Parkin classes of compounds while reducing the effects associated son's disease, or dementia associated with HIV infection. with high doses of glucocorticoids alone. Other inflammatory conditions include periodontal dis In methods of the invention, wherein a glucocorticoid is ease, tissue necrosis in chronic inflammation, endotoxin administered conjointly with a compound of the invention, shock, Smooth muscle proliferation disorders, graft versus 40 the glucocorticoid may be chosen from any glucocorticoid host disease, tissue damage following ischemia reperfusion known in the art. Glucocorticoids suitable for said conjoint injury, and tissue rejection following transplant Surgery. administration include, but are not limited to, alclometaSone, The present invention further provides a method of treating amcinonide, beclometasone, betamethasone, budesonide, or preventing inflammation associated with post-Surgical ciclesonide, clobetasol, clobetasone, clocortolone, clopred wound healing in a patient comprising administering to said 45 nol, cortisone, cortivaZol, deflazacort, desonide, desoximeta patient a compound of the invention. Sone, desoxycortone, dexamethasone, diflorasone, diflucor It should be noted that compounds of the current invention tolone, difluprednate, fluclorolone, fludroxycortide, may be used to treat or prevent any disease which has an flumetaSone, flunisolide, fluocinolone acetonide, fluocinon inflammatory component, such as those diseases cited above. ide, fluocortin, fluocortolone, fluorometholone, fluperolone, Further, the inflammatory conditions cited above are meant to 50 fluprednidene, fluticasone, formocortal, halcinonide, halom be exemplary rather than exhaustive. etaSone, hydrocortisonefcortisol, hydrocortisone aceponate, Those skilled in the art would recognize that additional hydrocortisone buteprate, hydrocortisone butyrate, lotepred inflammatory conditions (e.g., systemic or local immune nol, medrysone, meprednisone, methylprednisolone, methyl imbalance or dysfunction due to an injury, an insult, infection, prednisolone aceponate, mometasone furoate, parametha inherited disorder, or an environmental intoxicant or pertur 55 Sone, prednicarbate, prednisone/prednisolone, prednylidene, bant to the subject's physiology) may be treated or prevented rimexolone, tiXocortol, triamcinolone, ulobetasol, mometa by compounds of the current invention. Thus, the methods of Sone, fluticasone propionate, beclomethasone dipropionate, the current invention may be used to treat or prevent any fluocinolone, flunisolide hemihydrate, mometasone furoate disease which has an inflammatory component, including, monohydrate, desoxymethasone, diflorasone diacetate, but not limited to, those diseases cited above. 60 hydrocortisone acetate, difluorocortolone, fluorocortisone, The present invention also provides methods for treating or flumethasone, flunisolide, fluorocortolone, prednisolone, preventing arthritis, inflammatory bowel disease, uveitis, prednisone, cortisol. 6a-methylprednisolone, alclometasone ocular inflammation, asthma, pulmonary inflammation, cys dipropionate, fluclorolone acetonide, fluocinolone acetonide, tic fibrosis, psoriasis, arterial inflammation, cardiovascular betamethasone benzoate, fluocoritin butyl, betamethasone diseases, multiple Sclerosis, or neurodegenerative disease by 65 dipropionate, fluocortolone preparations, betamethasone val administering an effective amount of a compound of the erate, fluprednidene acetate, flurandrenolone, clobetasol pro invention. pionate, clobetasol butyrate, hydrocortisone, hydrocortisone US 9,315,447 B2 33 34 butyrate, methylprednisolone acetate, diflucortolone valer jointly administered with other therapeutic agents, such as ate, flumethasone pivalate, or triamcinolone acetonide, or other anti-inflammatory agents. pharmaceutically acceptable salts thereof. In certain embodiments, the present invention provides a In certain embodiments, the patient to be treated by a kit comprising: a) one or more single dosage forms of a method of the invention may already be receiving an anti compound of the invention; b) one or more single dosage inflammatory drug (other than a glucocorticoid). In one pre forms of a glucocorticoid as mentioned above; and c) instruc ferred embodiment, the patient is already taking a glucocor tions for the administration of the compound of the invention ticoid, such as one of the glucocorticoids described above, and the glucocorticoid. and will continue to take that drug conjointly with a com The present invention provides a kit comprising: 10 a) a pharmaceutical formulation (e.g., one or more single pound of the invention. Alternatively, the compound of the dosage forms) comprising a compound of the invention; invention may be used as a replacement for the previously and administered anti-inflammatory drug. b) instructions for the administration of the pharmaceutical In a related embodiment, the invention provides a method formulation e.g., for treating or preventing a disorder or of reducing the dose of a glucocorticoid required to achieve a 15 condition as discussed above, e.g., inflammatory dis desired anti-inflammatory effect. Reducing the dose of glu CaSC. cocorticoid while maintaining potent anti-inflammatory In certain embodiments, the kit further comprises instruc properties is highly desirable due to side effects associated tions for the administration of the pharmaceutical formula with certain glucocorticoids. Side effects of glucocorticoids tion comprising a compound of the invention conjointly with include increased appetite and weight gain, deposits of fat in a glucocorticoid as mentioned above. In certain embodi the chest, face, upperback, and stomach, water and salt reten ments, the kit further comprises a second pharmaceutical tion leading to Swelling and edema, high blood pressure, formulation (e.g., as one or more single dosage forms) com diabetes, slow healing of wounds, osteoporosis, cataracts, prising a glucocorticoidas mentioned above. acne, muscle weakness, thinning of the skin, increased sus In certain embodiments, the kit further comprises instruc ceptibility to infection, stomach ulcers, increased Sweating, 25 tions for the administration of the pharmaceutical formula mood Swings, psychological problems such as depression, tion comprising a compound of the invention conjointly with and adrenal Suppression and crisis. an agent Suitable for the treatment or prevention of inflam In this embodiment, the dose of a glucocorticoid is reduced matory disease (e.g., a non-steroidal anti-inflammatory drug by at least 5%, at least 10%, at least 15%, at least 20%, at least (NSAID), a mast cell stabilizer, or a leukotriene modifier) as 25%, at least 30%, at least 40%, at least 50%, at least 60%, at 30 mentioned above. In certain embodiments, the kit further least 70%, at least 80%, at least 90%, or more. The actual comprises a second pharmaceutical formulation (e.g., as one reduction in glucocorticoid dose will depend upon the nature or more single dosage forms) comprising an agent suitable for and amount of the compound of the invention being admin the treatment or prevention of inflammatory disease (e.g., a istered, the reduction in inflammation desired, and other fac non-steroidal anti-inflammatory drug (NSAID), a mast cell tors set forth elsewhere in this application that are typically 35 stabilizer, or a leukotriene modifier) as mentioned above. considered in treating a disease or condition. The amount of The present invention provides a kit comprising: the compound of the invention administered in this method a) a first pharmaceutical formulation (e.g., one or more will also depend upon the factors set forth above, as well as single dosage forms) comprising an agent Suitable for the nature and amount of glucocorticoid being administered. the treatment or prevention of inflammatory disease In certain embodiments, the amount of the compound of the 40 (e.g., a non-steroidal anti-inflammatory drug (NSA/D). invention administered in this method is less than 5%, less a mast cell stabilizer, or a leukotriene modifier) as men than 10%, less than 15%, less than 20%, less than 25%, less tioned above; and than 30%, less than 40%, less than 50%, less than 60%, less b) instructions for the administration of the first pharma than 70%, less than 80%, or less than 90% of the dose of the ceutical formulation with a compound of the invention, compound of the invention required to produce an anti-in 45 e.g., for treating or preventing inflammatory disease. flammatory effect without conjoint administration with a glu The present invention provides a kit comprising: cocorticoid. a) a first pharmaceutical formulation (e.g., one or more In one embodiment, the method of treating or preventing single dosage forms) comprising a glucocorticoid as inflammatory disease according to this invention may com mentioned above; and prise the additional step of conjointly administering to the 50 b) instructions for the administration of the first pharma patient another anti-inflammatory agent, Such as, for ceutical formulation with a compound of the invention, example, a non-steroidal anti-inflammatory drug (NSAID), a e.g., for treating or preventing inflammatory disease. mast cell stabilizer, or a leukotriene modifier. Metabolic Diseases In certain embodiments, the use of a composition compris The present invention provides a method of treating or ing a compound of the invention and a glucocorticoid accord 55 preventing complex disorders having an inflammatory com ing to this invention in the treatment of inflammatory disease, ponent in a patient comprising administering to said patient a does not preclude the separate but conjoint administration of compound of the invention. In certain embodiments, the com another corticosteroid. plex disorder having an inflammatory component is type 2 In certain embodiments, the use of a composition compris diabetes or obesity. ing both a compound of the invention and a glucocorticoid 60 The present invention provides a method of treating or according to this invention in the treatment of inflammatory preventing metabolic disorders in a patient comprising disease, does not preclude the separate but conjoint adminis administering to said patient a compound of the invention. In tration of another glucocorticoid. certain embodiments, the metabolic disorder is selected from In certain embodiments, different compounds of the inven an eating disorder, dyslipidemia, hypertrigylceridemia, tion may be conjointly administered with one or more other 65 hypertension, or metabolic syndrome. compounds of the invention while conjointly administering a The present invention further provides a method of treating glucocorticoid. Moreover, Such combinations may be con or preventing type 1 diabetes in a patient, comprising admin US 9,315,447 B2 35 36 istering to said patient a compound of the invention. In certain The present invention provides a kit comprising: embodiments, the present invention provides a method of a) a pharmaceutical formulation (e.g., one or more single treating a patient at risk of developing type 1 diabetes com dosage forms) comprising a compound of the invention; prising administering to said patient a compound of the inven and tion. In certain embodiments, the present invention provides 5 b) instructions for the administration of the pharmaceutical a method of treating a patient exhibiting one or more warning formulation e.g., for treating or preventing complex dis signs of type 1 diabetes, such as extreme thirst, frequent orders having an inflammatory component (e.g., type 2 urination; drowsiness or lethargy; Sugar in urine; Sudden diabetes or obesity), treating or preventing metabolic vision changes; increased appetite; Sudden weight loss; disorders (e.g., eating disorder, dyslipidemia, hypertri 10 gylceridemia, hypertension, or metabolic syndrome), fruity, Sweet, or wine-like odor on breath; heavy, labored treating or preventing type 1 diabetes, treating a patient breathing; stupor; or unconsciousness, comprising adminis at risk of developing type 1 diabetes, treating a patient tering to said patient a compound of the invention. exhibiting warning signs of type 1 diabetes, or protect The present invention further provides a method of treating ing (e.g., promoting the growth and/or Survival of) beta or preventing type 2 diabetes in a patient, comprising admin 15 cells of Islets of Langerhans from lipid- or glucose istering to said patient a compound of the invention. In certain triggered toxicity. embodiments, the present invention provides a method of In certain embodiments, the kit further comprises instruc treating a patient at risk of developing type 2 diabetes com tions for the administration of the pharmaceutical formula prising administering to said patient a compound of the inven tion comprising a compound of the invention conjointly with tion. In certain embodiments, the present invention provides an agent suitable for the treatment or prevention of diabetes a method of treating a patient exhibiting one or more warning (e.g., Sulfonylureas (e.g., chlorpropamide, tolbutamide, gly signs of type 2 diabetes, such as extreme thirst, frequent buride, glipizide, acetohexamide, tolaZamide, gliclazide, urination; drowsiness or lethargy; Sugar in urine; Sudden gliquidone, or glimepiride), medications that decrease the vision changes; increased appetite; Sudden weight loss; amount of glucose produced by the liver (e.g., metformin), fruity, Sweet, or wine-like odor on breath; heavy, labored 25 meglitinides (e.g., repaglinide or nateglinide), medications breathing; stupor; or unconsciousness, comprising adminis that decrease the absorption of carbohydrates from the intes tering to said patient a compound of the invention. tine (e.g., alpha glucosidase inhibitors such as acarbose), The present invention further provides a method for pro medications that effect glycemic control (e.g., pramlintide or tecting, e.g., promoting the growth and/or Survival of beta exenatide), DPP-IV inhibitors (e.g., sitagliptin), insulin treat cells of Islets of Langerhans from lipid- or glucose-triggered 30 ment, or combinations of the above) as mentioned above. In toxicity in a patient comprising administering to the patient a certain embodiments, the kit further comprises a second phar compound of the invention. maceutical formulation (e.g., as one or more single dosage In certain embodiments, the methods of treating or pre forms) comprising an agent Suitable for the treatment or pre venting a complex disorder having an inflammatory compo vention of diabetes (e.g., Sulfonylureas (e.g., chlorpropamide, nent, such as type 2 diabetes, or of treating type 1 diabetes 35 tolbutamide, glyburide, glipizide, acetohexamide, tolaza according to this invention may comprise the additional step mide, gliclazide, gliquidone, or glimepiride), medications of conjointly administering to the patient another treatment that decrease the amount of glucose produced by the liver for diabetes including, but not limited to, Sulfonylureas (e.g., (e.g., metformin), meglitinides (e.g., repaglinide or nateglin chlorpropamide, tolbutamide, glyburide, glipizide, aceto ide), medications that decrease the absorption of carbohy hexamide, tolaZamide, gliclazide, gliquidone, or glime 40 drates from the intestine (e.g., alpha glucosidase inhibitors piride), medications that decrease the amount of glucose pro Such as acarbose), medications that effect glycemic control duced by the liver (e.g., metformin), meglitinides (e.g., (e.g., pramlintide or exenatide), DPP-IV inhibitors (e.g., sita repaglinide or nateglinide), medications that decrease the gliptin), insulin treatment, or combinations of the above) as absorption of carbohydrates from the intestine (e.g., alpha mentioned above. glucosidase inhibitors such as acarbose), medications that 45 In certain embodiments, the kit further comprises instruc effect glycemic control (e.g., pramlintide or exenatide), DPP tions for the administration of the pharmaceutical formula IV inhibitors (e.g., Sitagliptin), insulin treatment, or combi tion comprising a compound of the invention conjointly with nations of the above. an agent Suitable for the treatment or prevention of obesity, as In certain embodiments, the methods of treating or pre mentioned above. In certain embodiments, the kit further venting a complex disorder having an inflammatory compo 50 comprises a second pharmaceutical formulation (e.g., as one nent. Such as obesity, according to this invention may com or more single dosage forms) comprising an agent Suitable for prise the additional step of conjointly administering to the the treatment or prevention of obesity, as mentioned above. patient another treatment for obesity including, but not lim The present invention provides a kit comprising: ited to, orlistat, Sibutramine, phendimetrazine, phentermine, a) a first pharmaceutical formulation (e.g., one or more diethylpropion, benzphetamine, mazindol, dextroamphet 55 single dosage forms) comprising an agent Suitable for amine, rimonabant, cetilistat, GT 389-255, APD356, pram the treatment or prevention of obesity as mentioned lintide/AC137, PYY3-36, AC 162352/PYY3-36, oxynto above; and modulin, TM 30338, AOD 9604, oleoyl-estrone, b) instructions for the administration of the first pharma bromocriptine, ephedrine, leptin, pseudoephedrine, or phar ceutical formulation with a compound of the invention, maceutically acceptable salts thereof. 60 e.g., for treating or preventing complex disorders having In certain embodiments, different compounds of the inven an inflammatory component (e.g., type 2 diabetes or tion may be conjointly administered with one or more other obesity), treating or preventing metabolic disorders compounds of the invention. Moreover, Such combinations (e.g., eating disorder, dyslipidemia, hypertrigylceri may be conjointly administered with other therapeutic demia, hypertension, or metabolic syndrome), treating agents, such as other agents Suitable for the treatment or 65 or preventing type 1 diabetes, treating a patient at risk of prevention of metabolic disorders, such as the agents identi developing type 1 diabetes, treating a patient exhibiting fied above. warning signs of type 1 diabetes, or protecting (e.g., US 9,315,447 B2 37 38 promoting the growth and/or survival of) beta cells of —O—C(O)-heteroaryl, —O—C(O)—O—(C-C)- Islets of Langerhans from lipid- or glucose-triggered alkyl, —O—C(O)—O-aryl, —O C(O)—O-het toxicity. eroaryl, and —O C(O) N(R)(R), wherein any The present invention provides a kit comprising: alkyl, aryl or heteroaryl is optionally substituted with up a) a first pharmaceutical formulation (e.g., one or more 5 to 3 substituents independently selected from halo, (C- single dosage forms) comprising an agent Suitable for Cs)-alkyl, O—(C-C)-alkyl, hydroxyl, carboxyl, ester, the treatment or prevention of diabetes as mentioned alkoxycarbonyl, acyl, thioester, thioacyl, thioether, above; and amino, amido, acylamino, cyano, and nitro, b) instructions for the administration of the first pharma each of RandR is independently selected from hydro ceutical formulation with a compound of the invention, 10 gen, halo, (C-C)-alkyl, perfluoroalkyl, aryl, and het e.g., for treating or preventing complex disorders having eroaryl, preferably hydrogen, halo and (C-C)-alkyl, an inflammatory component (e.g., type 2 diabetes or R is selected from -phenyl, -(C-Cs)-alkyl, -(C-C,)- obesity), treating or preventing metabolic disorders cycloalkyl, C=C phenyl, -C=C (C-C)-alkyl, and (e.g., eating disorder, dyslipidemia, hypertrigylceri —O-phenyl, wherein phenyl is optionally substituted 15 with up to 3 substituents independently selected from demia, hypertension, or metabolic syndrome), treating halo, (C-C)-alkyl, O-(C-C)-alkyl, hydroxyl, car or preventing type 1 diabetes, treating a patient at risk of boxyl, ester, alkoxycarbonyl, acyl, thioester, thioacyl, developing type 1 diabetes, treating a patient exhibiting thioether, amino, amido, acylamino, cyano, and nitro, warning signs of type 1 diabetes, or protecting (e.g., and R is additionally selected from -C=CH when: promoting the growth and/or survival of) beta cells of a) X is C(R)=C(R)— or -(cyclopropyl)-; or Islets of Langerhans from lipid- or glucose-triggered b) each of R“ and R' is hydrogen or halo; or toxicity. c) each of R and R is halo; or Ophthalmic Conditions d) R is —CH2—, The present invention provides a method of treating or each R" is independently selected from hydrogen and (C- preventing an ophthalmic condition in a patient, comprising 25 Cs)-alkyl, or two occurrences of R may optionally be administering to said patient a compound of the invention taken together with the carbons to which they are (e.g., a compound of any one of formula I-III as shown attached to form a 5- or 6-membered ring: above). each of R'' and R' is independently selected from The present invention further provides a method of treating hydrogen, (C-C)-alkyl, perfluoroalkyl, O—(C-C)- or preventing an ophthalmic condition (such as dry eye) in a 30 alkyl, aryl and heteroaryl, or R'" and R'' are taken patient, comprising administering to said patient a compound together with the carbonatom to which they are bound to of formula IV, form a carbocyclic or heterocyclic ring; and each double bond is independently in an E- or a Z-con figuration. (IV) 35 In certain embodiments, R' is —OM, where M is a cation selected from ammonium, tetra-alkyl ammonium, Na, K, Mg, and Zn. In certain embodiments, R and R' together are 40 or a pharmaceutically acceptable salt thereof, wherein: X is selected from C(R)=C(R)-, -(cyclopropyl)- -(cyclobutyl)-, -(cyclopentyl)-, and -(cyclohexyl)-; C/ NSN R" is selected from OR, N(R) SO, R and 45 - N(R)(R), wherein each of R" and R is indepen dently selected from H. C-C-alkyl, aryl, aralkyl, het In certain embodiments, X is —C=C . In certain eroaryl, and heteroaralkyl, and R is selected from embodiments, X is C(R)=C(R)-, -(cyclopropyl)-, C-C-alkyl, aryl, aralkyl, heteroaryl, and heteroaralkyl; -(cyclobutyl)-, -(cyclopentyl)-, or -(cyclohexyl)-. In certain R is selected from —CH2—, —C(O) , -SO. , PO 50 embodiments, X is C(R)=C(R)-. In certain embodi (OR)—, and tetrazole; ments, X is -(cyclopropyl)-, -(cyclobutyl)-, -(cyclopentyl)-. R is selected from hydrogen and alkyl: or -(cyclohexyl)-. In certain embodiments, X is -(cyclopro R is selected from a carbocyclic ring, a heterocyclic ring, pyl)-. In certain embodiments, X is -C=C- or -C(R) —(CH), , CHC(O)CH, and —CH2—O—CH, =C(R)-. In certain embodiments wherein X is -(cyclopro wherein: 55 pyl)-, -(cyclobutyl)-, -(cyclopentyl)-, or -(cyclohexyl)-, the n is an integer from 1 to 3: olefin and the carbon bearing R* are attached to adjacent any hydrogen atom in R is optionally and indepen carbons on the -(cyclopropyl)-, -(cyclobutyl)-, -(cyclopen dently replaced by halo, (C-C)-alkyl, perfluoro tyl)-, or -(cyclohexyl)- ring system. alkyl, aryl, heteroaryl, hydroxy, or O—(C-C)-alkyl; In certain embodiments, R' is hydrogen. In certain and 60 embodiments, R' is halo, OH, O (C-Cs)-alkyl, -O- any two hydrogen atoms bound to a common carbon aryl, O-heteroaryl, —O—C(O)—(C-C)-alkyl, —O C atom in R are optionally taken together with the (O)-aryl, —O C(O)-heteroaryl, —O C(O)—O—(C- carbon atom to which they are bound to form a car Cs)-alkyl, - O C(O)—O-aryl, O C(O)—O-heteroaryl, bocyclic or heterocyclic ring; or —O C(O) N(R)(R'), wherein any alkyl, aryl or het each of R“and R' is independently selected from hydro 65 eroaryl is optionally substituted with up to 3 substituents gen, halo. —OH, -O-(C-C)-alkyl, —O-aryl, O-het independently selected from halo, (C-C)-alkyl, O—(C- eroaryl, —O—C(O)—(C-C)-alkyl, —O—C(O)-aryl, Cs)-alkyl, hydroxyl, carboxyl, ester, alkoxycarbonyl, acyl,

US 9,315,447 B2 41 42 dently replaced by halo, (C-C)-alkyl, perfluoroalkyl, aryl, In certain embodiments, each double bond is in an E-con heteroaryl, hydroxy, or O—(C-C)-alkyl. In certain embodi figuration. In certain embodiments, each double bond is in a ments wherein R is —CH2—O—CH2, any hydrogen atom Z-configuration. In certain embodiments, one double bond is in R is optionally and independently replaced by halo, (C- in an E-configuration and one double bond is in a Z-configu Cs)-alkyl, perfluoroalkyl, aryl, heteroaryl, or O—(C-C)- ration. alkyl. In certain embodiments, R is selected from In certain embodiments, the invention contemplates any —(CH), and —CH2—O—CH2, wherein n is an integer combination of the foregoing. Those skilled in the art will from 1 to 3, and up to two hydrogenatoms in Rare optionally recognize that all specific combinations of the individual and independently replaced by (C-C)-alkyl. In certain possible residues of the variable regions of the compounds as embodiments, R is selected from a carbocyclic ring, a het 10 disclosed herein, e.g. R. R. R. R. R. R. R. R7, R', erocyclic ring, and CHC(O)CH, wherein n is an integer R', R. R. R., n and X, are within the scope of the inven from 1 to 3; any hydrogen atom in R is optionally and tion. As an example, any of the various particular recited independently replaced by halo, (C-C)-alkyl, perfluoro embodiments for R* may be combined with any of the vari alkyl, aryl, heteroaryl, hydroxy, or O—(C-C)-alkyl, and 15 ous particular recited embodiments of X. any two hydrogen atoms bound to a common carbon atom in R are optionally taken together with the carbon atom to In certain embodiments, the compound is selected from which they are bound to form a carbocyclic or heterocyclic any one of ring. In certain embodiments, R" is hydrogen. In certain embodiments, R' is selected from (C-C)-alkyl, perfluo (301) roalkyl, O (C-C)-alkyl, aryl and heteroaryl, or R' is OH O taken together with R', and the carbon atom to which they are bound to form a carbocyclic or heterocyclic ring. 21Ne. In certain embodiments, R' is hydrogen. In certain 25 embodiments, R' is selected from (C-Cs)-alkyl, perfluo roalkyl, O (C-Cs)-alkyl, aryl and heteroaryl, or R' is taken together with R'' and the carbon atom to which they --a", are bound to form a carbocyclic or heterocyclic ring. In certain embodiments, R is —OR". In certain embodi 30 (302) ments, R is selected from N(R) SO Rand—N(R) OH O (R). In certain embodiments, R is N(R)-SO. R. In certain embodiments. R' is selected from —OR and —N(R)(R). In certain embodiments, R is N(R)(R). In certain embodiments, R' is selected from —OR, and 35 N(R) SO, R. In certain embodiments, R is hydrogen. In certain embodi --- O'Na', ments, R is (C-C)-alkyl or two occurrences of R may optionally be taken together with the carbons to which they (303) are attached to form a 5- or 6-membered ring. 40 OH OH O In certain embodiments, X is -C=C- and R' is hydro gen. In certain embodiments, X is -C=C- and R" is hydro gen. In certain embodiments, X is C=C , R“ is fluoro, and 45 R is fluoro. In certain embodiments, X is -C=C R' is fluoro, and R is fluoro. In certain embodiments, X is -C=C , and each of R' F --- and R' is independently selected from OH, - O (C- 50 Cs)-alkyl, O-aryl, O-heteroaryl, —O C(O)—(C-C)- (304) alkyl, O—C(O)-aryl, O C(O)-heteroaryl, and —O—C (O) N(R)(R). OH OH O In certain embodiments, X is -C=C- and R is —CH2—. 55 In certain embodiments, X is -(cyclopropyl)-, -(cyclobu tyl)-, -(cyclopentyl)-, and -(cyclohexyl)-. In certain embodi ments, X is -(cyclopropyl)-. In certain embodiments, X is C(R)=C(R)—. In certain embodiments, each of R and R is indepen 60 dently selected from Hand C-C-alkyl; R is C-C-alkyl; R (305) is selected from —(CH), and —CH2—O—CH2, wherein OH OH O n is an integer from 1 to 3, and up to two hydrogenatoms in R are optionally and independently replaced by (C-C)-alkyl; each of R“ and R' is independently selected from hydrogen, 65 halo, -OH,-O (C-C)-alkyl; and each of R'' and R' is hydrogen.

US 9,315,447 B2 45 46 -continued -continued (317) (327) HO, HC CH3 O OH OH O

5 o HO 2 O'Na',8. 21N 21 o OEt, N1 N.

O 10 (328) OH OH O

15 21N 21 OEt, F (318) O E 2O (329) 21 Nea O'Na', OH O | 21 Ne. OEt, (319) is OH F F O |

21 Ne. OEt, (330) | 30 O

65 F F US 9,315,447 B2 47 48 -continued -continued (334) (340) OH OH OH OH O an ea = OEt, 5 21Ne. OEt,

(336) OH OH O 15 F 21 Ne. OEt,

| 2O (341) OH OH O

(337) 25 21 Ne. OEt, OH OH O

21Ne. OEt, | 30

35 OCH

(338) OH OH O 40 (342) 21 Ne. OEt, OH OH O 21 Nea OEt, | 45

C (339)339 55 OH OH O

21Ne. OEt, (343) | 60 OH O

21 Nea OMe,

65 US 9,315,447 B2 49 50 -continued two hydrogenatoms in Rare independently replaced by (344) halo, (C-C)-alkyl, or O—(C-C)-alkyl: each of RandR is independently selected from hydro HO, HC CH3 O gen, (C-C)-alkyl, perfluoroalkyl, aryl, and heteroaryl, 5 preferably hydrogen and (C-C)-alkyl; HO 2 OMe, R is selected from -C=CH, -phenyl, -(C-Cs)-alkyl, N1 N. —(C-C)-cycloalkyl, C=C phenyl, -C=C(C-C) cycloalkyl, -C=C-(C-C)-alkyl, and —O-phenyl, O wherein phenyl is optionally substituted with up to 3 10 Substituents independently selected from halo, (C-C)- alkyl, O—(C-C)-alkyl, hydroxyl, carboxyl, ester, alkoxycarbonyl, acyl, thioester, thioacyl, thioether, amino, amido, acylamino, cyano, and nitro, each of Rand Rare independently selected from hydro 15 gen, —(C-C)-alkyl, -aryl, -heteroaryl, -C(O)—(C- Cs)-alkyl, —C(O)-aryl, —C(O)-heteroaryl, —C(O)— (345) O-aryl, —C(O)—O-heteroaryl, and —C(O)—N(R) OH OH O (R), wherein any alkyl, aryl or heteroaryl is optionally substituted with up to 3 substituents independently Selected from halo, (C-C)-alkyl, O—(C-C)-alkyl, hydroxyl, carboxyl, ester, alkoxycarbonyl, acyl, '-y' -- and thioester, thioacyl, thioether, amino, amido, acylamino, cyano, and nitro: (346) each of R'' and R' is independently selected from 25 hydrogen, (C-C)-alkyl, perfluoroalkyl, O—(C-C)- OH OH O alkyl, aryland heteroaryl, or R" and R'' are taken together with the carbon atom to 21\21N OMe. which they are bound to form a carbocyclic or hetero cyclic ring; and HC 30 wherein each double bond is independently in an E- or a Z-configuration. In certain embodiments, R is —OM, where M is a cation The present invention further provides a method of treating selected from ammonium, tetra-alkyl ammonium, Na, K, Mg, and Zn. or preventing an ophthalmic condition (such as dry eye) in a 35 patient, comprising administering to said patient a compound In certain embodiments, RandR together are of the formula V.

(V) 40 * asC / R1. NRN In certain embodiments, R is C(O)—. In certain 45 embodiments, R is —OR", wherein R is hydrogen or C-C- alkyl. In certain embodiments, Ris-(CH2), , whereinnis 3. In certain embodiments, R is -C=CH. In certain embodiments, R is hydrogen. In certain embodiments, R. is hydrogen. In certain embodiments, R" is hydrogen. In (VI) 50 OR OR9 certain embodiments, R' is hydrogen. In certain embodi R. E R; ments, R is —C(O) , R' is —OR", wherein R is C-C- R19. R2 alkyl, R is —(CH), , wherein n is 3, R is -C=CH, R 21\e S R31 NRI is hydrogen, R is hydrogen, R" is hydrogen, and R' is R10b s R6 hydrogen. 55 In certain embodiments, the compound is selected from any one of or a pharmaceutically acceptable salt of either of the forego ing, wherein: R" is selected from OR, N(R) SO, R and (323) 60 - N(R)(R), wherein each of R" and R is indepen dently selected from H. C-C-alkyl, aryl, aralkyl, het eroaryl, and heteroaralkyl, and R is selected from C-C-alkyl, aryl, aralkyl, heteroaryl, and heteroaralkyl; O'Na', R is selected from C(O) ,-PO(OR)—, and tetrazole; R is selected from hydrogen and alkyl: 65 R is selected from -(CH2), and —CH2—O—CH2, wherein n is an integer from 1 to 3; and optionally up to US 9,315,447 B2 51 52 -continued (324) (VIII) ORf 4N4\– Rs E, ONa,

(325) 10 and pharmaceutically acceptable salts thereof, wherein: Re, Rf, Rs, and E are as defined above. In certain embodiments, a compound of formula VII or OEt, or VIII is represented by formula IX, 15 (IX) ORf ORe O (335)

25 and pharmaceutically acceptable salts thereof, wherein: Re, Rf, and E are as defined above. Additional compounds Suitable for use in methods and The present invention further provides a method of treating compositions of the invention include those of Formula A, or preventing an ophthalmic condition (such as dry eye) in a 30 patient, comprising administering to said patient a compound of formula VII,

(VII) 35 wherein: R O R. ORf ORe each of W' and Y is a bond or a linker independently Rs o ul selected from a ring containing up to 20 atoms or a chain of up 21 2 o Rs E, to 20 atoms, provided that W and Y can independently include one or more nitrogen, oxygen, Sulfur orphosphorous 40 atoms, further provided that W and Y can independently include one or more substituents independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, and pharmaceutically acceptable salts thereof, wherein: iodo, bromo, fluoro, hydroxy, alkoxy, aryloxy, carboxy, Re and Rfare independently selected from hydrogen, alkyl, amino, alkylamino, dialkylamino, acylamino, carboxamido, alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxyacyl, 45 cyano, oxo, thio, alkylthio, arylthio, acylthio, alkylsulfonate, aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl; arylsulfonate, phosphoryl, or sulfonyl, further provided that E is a hydroxyl, alkoxy, aryloxy, amino, alkylamino, dialky W' and Y can independently contain one or more fused car lamino, or arylamino; bocyclic, heterocyclic, aryl or heteroaryl rings, and further Rh and Ri are independently selected from hydrogen, alkyl, 50 provided that when o' is 0, and V is alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl; Rs is selected from i-iv as follows: i) CHCH(R)CH, where R1001 Ra' R is hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl, heteroaryl, fluoro, hydroxyl or alkoxy; ii) CHC 21 (RR)CH, where RandR, are each independently alkyl, 55 alkenyl, alkynyl, perfluoroalkyl, aryl, or fluoro, or R and R7 are connected together to form a carbocyclic or hetero cyclic ring; iii) CHOCH, CHC(O)CH, or CHCH; or Y is connected to V via a carbon atom; iv) Rs is a carbocyclic, heterocyclic, aryl or heteroaryl ring; and 60 V is selected from Rs and R are independently selected from hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or heteroaryl, R 1001 Ra' R1001 Ra' or Rs and R are connected together to form a carbocyclic or heterocyclic ring. 65 A-X->, WX-y In certain embodiments, a compound of formula VII is represented by formula VIII, US 9,315,447 B2 53 54 -continued trans configuration, wherein each R' is independently R1 OO Ra Selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, acyl, silyl, alkoxyacyl, aminoacyl, aminocar N1N bonyl, alkoxycarbonyl, or a protecting group; R1002 Rb' or when V is R1002 Rb' R1002 , or 10

R1002 Ra' R1001

Ra' R1001 15 and V is wherein when q is 0 and V is a bond, n' is 0 or 1; otherwise R1001 Ra' n' is 1: V is selected from a bond, N

RI OO Ra' R'' and Rare both hydrogen; X is selected from CN, -C(NH)N(R") (R"), C(S)-A', A-X, Y, , or 25 C(S)R", C(O)-A", C(O) R", C(O). SR", Rb' R1002 - C(O) NH S(O). R", S(O)-A', -S(O) R", S(O)N(R")(R"), -P(O)-A'. - PO(OR")-A', -tetrazole, 2^4 n alkyltetrazole, or —CHOH, wherein , or A is selected from OR", N(R")(R") or - OM': Ra' R1001 30 each R" is independently selected from hydrogen, alkyl, R1002 aryl, arylalkyl, heteroaryl, heteroarylalkyl or a detect able label molecule, wherein any alkyl-, aryl- or het eroaryl-containing moiety is optionally Substituted with 1001 R Ra 35 up to 3 independently selected substituents; and M' is a cation; wherein: G' is selected from hydrogen, halo, hydroxy, alkyl, aryl, L' is selected from C(R')(R') , wherein each of arylalkyl, heteroaryl, heteroarylalkyl, alkoxy, aryloxy, car R'' and R'' is independently selected from hydro boxy, amino, alkylamino, dialkylamino, acylamino, carboxa gen, alkyl, alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl 40 mido or a detectable label molecule, wherein any alkyl-, aryl or heteroaryl, or R'' and R'' are connected together or heteroaryl-containing moiety is optionally Substituted with to form a carbocyclic or heterocyclic ring; when V is up to 3 independently selected substituents; o' is 0, 1, 2, 3, 4, or 5: p' is 0, 1, 2, 3, 4, or 5: R1001 Ra' 45 q' is 0, 1, or 2; and S o'+p'+q' is 1, 2, 3, 4, 5 or 6: wherein: if V is a bond, then q is 0, and V is a bond; L' is additionally selected from W'; and n' is 0 or 1; 50 V is selected from a bond or if V is

R1001 Ra R1001 Ra' 55 N

wherein: then o' is 0, V is each R'' and R' is independently for each occurrence 60 Selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, alkylaryl, alkoxy, or halo, wherein said alkyl R1001 Ra' or aryl-containing moiety is optionally Substituted with up to 3 independently selected substituents; each of R'' and R is independently for each occurrence 65 selected from —OR' or N(R'), or adjacent RandR are taken together to form an epoxide ring having a cis or US 9,315,447 B2 55 56 p' is 1 and V2 is In certain embodiments, V is selected from a bond, Rb' R100

RI OO Ra' R1001 R' 5 N A > C, s O Ra R1001

any acyclic double bond may be in a cis or a trans configu In certain embodiments, when q is 0 and V is a bond, n' is ration or is optionally replaced by a triple bond; and 10 0 or 1; otherwise n' is 1. either one In certain embodiments, p' is 0, 1, 2, 3, or 5. In certain embodiments, q' is 0 or 1. In certain embodiments, if V is N 21 Na 15 R1001 R' portion of the compound, if present, is optionally replaced by N1 N R1002 Rib' then o' is 0 or 1, p' is 1 or 2, o'+p' is 1 or 2, V, is 21 Q'-- R1001 R' N 25 A >>, Oi Oile and V is a bond. 30 In certain embodiments, if V is

R1001 R' A-y 35 A >>, portion of the compound, if present, is optionally replaced by then o' is 3, 4 or 5, p' is 0, 1 or 2, o'+p' is 4 or 5, and V is a bond. In certain embodiments, if V is a bond, then o' is 0, 3, 4 or 40 5; p' is 0, 1, 2 or 5, o'+p' is 4 or 5, q' is 0, and V is a bond. In certain embodiments, each of WandY" is independently 21 selected from a bond or lower alkyl or heteroalkyl optionally Q'-- s substituted with one or more substituents independently N selected from alkenyl, alkynyl, aryl, chloro, iodo, bromo, 45 fluoro, hydroxy, amino, or oxo. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 1, wherein Q' represents one or more substituents and each Q' is independently selected from halo, alkyl, alkenyl, alkynyl, 50 cycloalkyl, aryl, heteroaryl, alkoxy, aryloxy, alkylcarbonyl, R ORf Rh ORe arylcarbonyl, alkoxycarbonyl, aryloxycarbonyl, amino, hydroxy, cyano, carboxyl, alkoxycarbonyloxy, aryloxycarbo L 21\21N J-I nyloxy or aminocarbonyl. In certain embodiments, V is selected from "A c' (d 55 a N-2 H-G 1001 R R1001 R' R R ORg

A >>, s 21 , or 60 wherein: Carbons a' and b' are connected by a double bond or a triple R1001 R' bond; Carbons c' and d'are connected by a double bond or a triple N N bond; R1002 Rib' 65 Re, Rf, and Rg are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxya cyl, aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl; US 9,315,447 B2 57 58 Rh, Ri and Rj are independently selected from hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl; 3 I is selected from —C(O)-E, SO-E, PO(OR)-E, where E is hydroxy, alkoxy, aryloxy, amino, alkylamino, dialky ORf ORe O lamino, or arylamino; and R is hydrogen or alkyl; J. L and H are linkers independently selected from a ring containing up to 20 atoms or a chain of up to 20 atoms, provided that J. L and H can independently include one or \ 4 more nitrogen, oxygen, Sulfur or phosphorous atoms, and further provided that J. L and H can independently include 10 one or more substituents selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, iodo, bromo, wherein: fluoro, hydroxy, alkoxy, aryloxy, carboxy, amino, alky lamino, dialkylamino, acylamino, carboxamido, cyano, E. Re, Rf, and Rg are as defined above. Oxo, thio, alkylthio, arylthio, acylthio, alkylsulfonate, aryl 15 In certain embodiments, a pharmaceutically acceptable Sulfonate, phosphoryl, and Sulfonyl, and further provided salt of the compound is formed by derivatizing E, wherein E that J. L and H can also contain one or more fused carbocy is —OM, where M is a cation selected from ammonium, clic, heterocyclic, aryl or heteroaryl rings, and provided tetra-alkyl ammonium, Na, K, Mg, and Zn. that linker J is connected to the adjacent C(R)OR group via Exemplary compounds of formula 3 include compound 3a, a carbon atom; G is selected from hydrogen, alkyl, perfluoroalkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, iodo, bromo, fluoro, (3a) hydroxy, alkoxy, aryloxy, carboxy, amino, alkylamino, C(CH3)3 C(CH3)3 dialkylamino, acylamino, or carboxamido; 25 r. or pharmaceutically acceptable salts thereof. j- -- In certain embodiments, a pharmaceutically acceptable O O O salt of the compound is formed by derivatizing E, wherein E is —OM, where M is a cation selected from ammonium, 41\4\– OCH tetra-alkyl ammonium, Na, K, Mg, and Zn. 30 In certain embodiments, a compound of formula 1 is rep \ - resented by formula 2. O 35 -- ORf ORe O C(CH3)3

and compound 3b, 40

(3.b) ORg OH OH O 45 wherein: 21N 21 S OCH3. E. Re, Rf, and Rg are as defined above. In certain embodiments, a pharmaceutically acceptable \ - salt of the compound is formed by derivatizing E, wherein E is —OM, where M is a cation selected from ammonium, 50 OH tetra-alkyl ammonium, Na, K, Mg, and Zn. Exemplary compounds of formula 2 include compound 2a: Further exemplary compounds of formula 1 include Com pound X, (2a) 55 OH OH O (X) OH OH O OCH3. 60 21\21N OH, \ 4 \ 4 OH OH 65 In certain embodiments, a compound of formula 1 is rep resented by formula 3, and pharmaceutically acceptable salts and esters thereof. US 9,315,447 B2 59 60 Additional compounds suitable for use in methods and Exemplary compounds of formula 4 include compound 4a, compositions of the invention include those of Formula 4.

(4a)

10 compound 4b.

wherein: (4b) 15 A is H or —OP: P, P and Peach individually is a protecting group or hydro gen atom; R and Reach individually is a substituted or unsubstituted, branched or unbranched alkyl, alkenyl, or alkynyl group, substituted or unsubstituted aryl group, substituted or and pharmaceutically acceptable salts and esters thereof. unsubstituted, branched or unbranched alkylaryl group, Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 5, halogen atom, hydrogen atom; 25 Zis C(O)CR, C(O)NRR, C(O)H, C(NH)NR'R'',

—C(S)H, C(S)OR, C(S)NR'R, CN, preferably a carboxylic acid, ester, amide, thioester, thiocarboxamide or a nitrile; 30 each R', if present, is independently selected from hydro gen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C3-C8) cycloalkyl, cyclohexyl, (C4-C11) cycloalkyla lkyl, (C5-C10) aryl, phenyl, (C6-C16) arylalkyl, benzyl, 35 2-6 membered heteroalkyl, 3-8 membered heterocyclyl, or pharmaceutically acceptable salts thereof, wherein: morpholinyl, piperazinyl, homopiperazinyl, piperidinyl, the stereochemistry of the carboni' to carboni' bond is cis or trans; 4-11 membered heterocyclylalkyl, 5-10 membered het Ps is a protecting group or hydrogen atom; and eroaryland 6-16 membered heteroarylalkyl; 40 P. P., R and Z are as defined above in formula 4. In certain embodiments, the stereochemistry of the carbon each R', if present, is a suitable group independently selected ii' to carboni' bond is trans. from =O, OR, (C1-C3) haloalkyloxy, OCF =S, Exemplary compounds of formula 5 include compound 5a, - SR", =NR, =NOR, NRR, halogen, CF, CN, NC, OCN, -SCN, NO, NO, —N, 45

(5a)

NRR, NHC(NH)NR'R'' and NRC(NR)NR Re; 55 compound 5b, each R, if present, is independently a protecting group or R', or, alternatively, two R taken together with the nitrogen (5b) atom to they are bonded form a 5 to 8-membered hetero OH cyclyl or heteroaryl which optionally including one or 60 more additional heteroatoms and optionally substituted with one or more of the same or different R or suitable R' groups; each n independently is an integer from 0 to 3: HG each Rindependently is a protecting group or R': 65 or pharmaceutically acceptable salts thereof. and pharmaceutically acceptable salts and esters thereof. US 9,315,447 B2 61 62 Additional compounds suitable for use in methods and or pharmaceutically acceptable salts thereof, wherein: compositions of the invention include those of Formula 6, Carbons e' and fare connected by a double bond or a triple bond, and when carbon e' is connected to carbon f through a double bond the stereochemistry is cis or trans; Carbons g and h' are connected by a double bond or a triple bond and when carbong is connected to carbonh' through a double bond the stereochemistry is cis or trans; m is 0 or 1; 10 T is hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl, (C2-C6) alkynyl, (C5-C14) aryl, (C6-C16) arylalkyl, 5-14 mem bered heteroaryl, 6-16 membered heteroarylalkyl, or -CH=CHCHCH: or pharmaceutically acceptable salts thereof, wherein: T is -(CH2) - or -(CH2) O-, where q is an integer the stereochemistry of the carbongg' to carbonhh' bond is cis 15 from 0 to 6: or trans; Z is (C1-C6) alkylene optionally substituted with 1, 2, 3, 4, 5 each X represents hydrogen or taken together both X groups or 6 of the same or different halogen atoms, -(CH2) - represent one substituted or unsubstituted methylene, an O—CH2—, or —(CH), S-CH , where p is an inte oxygen atom, a Substituted or unsubstituted Natom, or a ger from 0 to 4: Sulfur atom Such that a three-membered ring is formed; and R. R. and Rs each individually is substituted or unsubsti P. P. P. R. and Z are as defined above. tuted, branched or unbranched alkyl, alkenyl, or alkynyl In certain embodiments, the stereochemistry of the carbon group, Substituted or unsubstituted aryl group, Substituted gg' to carbon hh' bond is trans. or un Substituted, branched or unbranched alkylaryl group, Exemplary compounds of formula 6 include compound 6a, 25 Calkoxy, halogen atom, —CHR, —CHRR, —CRRR, or a hydrogen atom; Ra is independently for each occurrence selected from (6a) —CN, NO, or halogen; HO OH P. P., P., and Z are as defined above. 30 In certain embodiments, carbons e' and fare connected by a cis double bond. In certain embodiments, carbons gandh' are connected by a double bond. 35 In certain embodiments, carbons e' and fare connected by a cis double bond and carbons g and h' are connected by a double bond. Exemplary compounds of formula 7 include compound 7a,

40 (7a)

(6b) COH, 45

HO 50 compound 7b, and pharmaceutically acceptable salts and esters thereof. (7b) 55

Additional compounds suitable for use in methods and compositions of the invention include those of Formula 7.

R11 OP 60

65 and pharmaceutically acceptable salts and esters thereof. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 8, US 9,315,447 B2 63 64 Additional compounds Suitable for use in methods and R compositions of the invention include those of Formula 9.

10 or pharmaceutically acceptable salts thereof, wherein: the stereochemistry of the carbon i' to carboni' bond is cis or trans; m is 0 or 1; 15 D' is CH, -CH=CHCHU or -CH=CHCHCHA; or pharmaceutically acceptable salts thereof, wherein: U is a branched or unbranched, substituted or unsubstituted Carbons k" and 1' are connected by a double bond or a triple alkyl, alkenyl, alkynyl, cycloalkyl, aryl, alkoxy, aryloxy, bond, and when carbonk' is connected to carbon 1' through alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aryloxycar a double bond the stereochemistry is cis or trans; bonyl, alkoxycarbonyloxy, and aryloxycarbonyloxy the stereochemistry of the carbon m' to carbon n' double bond group; is cis or trans; A is Hor—OP: m is 0 or 1; P. P. P. R. R. and Z are as defined above. 25 D is CH or -CH=CHCHCH: In certain embodiments, the stereochemistry of the carbon i' to carboni' bond is cis. P. P. P. R. X., and Z are as defined above. In certain embodiments, the stereochemistry of the carbon Exemplary compounds of formula 8 include compound 8a, m' to carbon n' double bond is cis. 30 In certain embodiments, carbons k" and 1' are connected by (8a) a cis double bond. In certain embodiments, the stereochemistry of the carbon m' to carbon n' double bond is cis and carbons k" and 1' are connected by a cis double bond. 35 Exemplary compounds of formula 9 include compound 9a,

(9a) OH 40 compound 8b, FN 2n 2 COH, N 21 - T (8b) 45

50 compound 9b,

(9b) compound 8c, OH 55 FN 2n 2 COH,

(8c) N 21 - T 60 OH

65 and pharmaceutically acceptable salts and esters thereof. Additional compounds Suitable for use in methods and and pharmaceutically acceptable salts and esters thereof. compositions of the invention include those of Formula 10. US 9,315,447 B2 65 66 or pharmaceutically acceptable salts thereof, wherein:

10 P. P., P, R, and Z are as defined above. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 13,

13 PO Z R 10 N

R2 15 s or pharmaceutically acceptable salts thereof, wherein: PO P. P., P., R and Z are as defined above; and U Q represents one or more Substituents and each Q individu ally, if present, is a halogen atom or a branched or or pharmaceutically acceptable salts thereof, wherein: unbranched, substituted or unsubstituted alkyl, alkenyl, P. P. R. R. U, and Z are as defined above. alkynyl, cycloalkyl, aryl alkoxy, aryloxy, alkylcarbonyl, Additional compounds Suitable for use in methods and arylcarbonyl, alkoxycarbonyl, aryloxycarbonyl, amino, compositions of the invention include those of Formula 14, hydroxy, cyano, carboxyl, alkoxycarbonyloxy, aryloxycar bonyloxy or aminocarbonyl group. 25

Additional compounds suitable for use in methods and 14 compositions of the invention include those of Formula 1 1.

11 30

35 or pharmaceutically acceptable salts thereof, wherein: P. P. R. R. Q, and Z are as defined above. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 15, 40

15 R OP or pharmaceutically acceptable salts thereof, wherein: 45 P. P. P. R. and Z areas defined above. Additional compounds suitable for use in methods and compositions of the invention include those of Formula 12, 50

12 or pharmaceutically acceptable salts thereof, wherein: P. P., and Z are as defined above. Additional compounds Suitable for use in methods and 55 compositions of the invention include those of Formula 16,

or pharmaceutically acceptable salts thereof, wherein: 65 P and Z are as defined above. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 17, US 9,315,447 B2 67 Additional compounds Suitable for use in methods and 17 compositions of the invention include those of

Formula 20

10 or pharmaceutically acceptable salts thereof, wherein: Carbons o' and p' are connected by a single or a double bond (e.g., a cis or trans double bond): Formula 21 Carbons q' and rare connected by a single or a double bond

21 (e.g., a cis or trans double bond); and 15 P. P., and Z are as defined above. Additional compounds suitable for use in methods and compositions of the invention include those of Formula 18,

Formula 22 18 22 PO OP S1S CO2R, 25 21 OP OP or pharmaceutically acceptable salts thereof, wherein: 30 Formula 23

the stereochemistry of the carbons' to carbont' double bond 23 is cis or trans; the stereochemistry of the carbon u' to carbon v' double bond is cis or trans; and 35 P. P. R. R. and Z are as defined above. Additional compounds suitable for use in methods and compositions of the invention include those of Formula 19. Formula 24

24 40

45 Formula 25 25 or pharmaceutically acceptable salts thereof, wherein: Carbons w' and X’ are connected by a single or a double bond; 50 Carbons y' and z are connected by a single or a double bond; Formula 26 and

P. P., and Z are as defined above. 26 In certain embodiments of formulae 4 to 19, each R', if 55 present, is a suitable group independently selected from =O. —OR", (C1-C3) haloalkyloxy, OCF. =S, - SR", =NR", =NOR, NRR, halogen, CF, CN, NC, OCN, —SCN, NO,-NO =N - N - S(O)R, S(O).R. S(O), OR, S(O)NRR, S(O)NRR, OS(O)R’, 60 Formula 27 –OS(O),R, OS(O),OR, OS(O)NR'R'', C(O)R, C(O)OR, C(O)NR'R'', C(NH)NR'R'', C(NR) 27 NRR, C(NOH)R’, C(NOH)NR'R'', OC(O)R’, OC(O)OR, OC(O)NR'R'', OC(NH)NR'R'', OC (NH)NR'R'', -NHC(O)IR, NRC(O)I.R., -NHC 65 (O)IOR, NHC(O)NRR, NRC(O)NRR, -NHC(NH)NRR and NRC(NR)NR'R''. US 9,315,447 B2 69 70 -continued A is selected from (C1-C6) alkylene optionally substituted Formula 28 with 1,2,3,4, 5 or 6 of the same or different halogenatoms, 28 —(CH), O—CH2— and —(CH), S-CH2—, where m is an integer from 0 to 4: 5 X is selected from —(CH), and —(CH), O—, where n is an integer from 0 to 6: Y is selected from hydrogen, (C1-C6) alkyl, (C2-C6) alk enyl, or (C2-C6) alkynyl, optionally substituted with one or 10 or pharmaceutically acceptable salts of any of the above, more of the same or different Roo groups, (C5-C14) aryl wherein: optionally substituted with one or more of the same or each P is individually selected from Hora protecting group; different Roo groups, phenyl, optionally substituted with and one or more of the same or different Roo groups, (C6-C16) R is H. Calkyl (e.g., methyl, ethyl, glycerol), Calkenyl or 15 Calkynyl. arylalkyl optionally substituted with one or more of the Exemplary compounds of formula 21 include compound same or different Roo groups, 5-14 membered heteroaryl 21a, optionally substituted with one or more of the same or different Roo groups, 6-16 membered heteroarylalkyl optionally substituted with one or more of the same or

(21a) different Roo groups and a detectable label molecule; each Roo is independently selected from an electronegative group, =O. —OR', (C1-C3) haloalkyloxy, =S. —SR', 25 =NR', =NONR', NR'R'', halogen, CF, CN, OH and pharmaceutically acceptable salts and esters thereof. Additional compounds suitable for use in methods and 30 NR'R'', C(O)R', C(O)OR, C(O)NR'R'', compositions of the invention include those of Formula 29. C(NH)NR'R'', OC(O)R', OC(O)OR', OC (O)NR'R'', OC(NH)NR'R'', NHC(O)R', NHC(O)OR', NHC(O)NR'R'' and NHC(NH) 29 NRel Rel; R102 OH R101 l 35 each R' is independently selected from hydrogen, (C1-C4) Nafna D-E.1 5 YA1 W alkyl, (C2-C4) alkenyl or (C2-C4) alkynyl; and each R is independently an R' or, alternatively, R'R'' Fl NG S 1st-XI ny taken together with the nitrogenatom to which it is bonded 40 forms a 5 or 6 membered ring. R103 OH In certain embodiments of Formula 29, when X—Y is —CHCH, then at least one of Rio, Rio or Ros is other and pharmaceutically acceptable salts, hydrates and Solvates than hydrogen. thereof, wherein: In certain embodiments, a compound of Formula 29 is D-E and F-G are independently are cis or trans-C=C- 45 represented by Formula 30, or —C=C : Ro, Ro and Ros are independently selected from hydro gen, (C1-C4) straight-chained or branched alkyl, (C2-C4) 30 alkenyl (C2-C4) alkynyl, (C1-C4) alkoxy, —CHR, R102 OH R101 Ul —CHRio Rio and —CRio RoRo; 50 each Roa is independently selected from CN, - NO and 21a 21 Y. W halogen; Fl X W is selected from —Rios —ORios —SRios and NG N Ny. —NRos Rios: 55 ROH each Ros is independently selected from hydrogen, (C1-C6) alkyl, (C2-C6) alkenyl or (C2-C6) alkynyl optionally sub stituted with one or more of the same or different R groups, and pharmaceutically acceptable salts, hydrates and Solvates (C5-C14) aryl optionally substituted with one or more of thereof, wherein: the same or different R groups, phenyl optionally Substi 60 tuted with one or more of the same or different R groups, D-E and F-G are independently are cis or trans-C=C- (C6-C16) arylalkyl optionally substituted with one or more or —C=C- and of the same or different R groups, 5-14 membered het Rg1: Ro2, Rios, Rio, WI, Rios, A, X, n, Y. Roo R',al and eroaryl optionally substituted with one or more of the same R" are as defined above. or different R groups, 6-16 membered heteroarylalkyl 65 Additional compounds Suitable for use in methods and optionally substituted with one or more of the same or compositions of the invention include those of Formulae 31 to different R groups and a detectable label molecule: 37 US 9,315,447 B2 72 Ro7 is

Additional compounds Suitable for use in methods and 10 compositions of the invention include those of Formula 38.

wherein Carbonsaa' and bb' are connected by a double bond or a triple bond; 25 Carbons cc' and dd'are connected by a double bond or a triple bond; Re, Rf, and Rg are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, acyl (e.g., alkoxya cyl, aminoacyl), aminocarbonyl, alkoxycarbonyl, or silyl; 34 30 E is hydroxyl, alkoxy, aryloxy, amino, alkylamino, dialky lamino, or arylamino; Rh, Ri and Rj are independently selected from hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl or heteroaryl; R is selected from hydrogen, alkyl, perfluoroalkyl, alkenyl, 35 alkynyl, aryl, heteroaryl, fluoro, hydroxyl, alkoxy, aryloxy; Rs is selected from i-iv as follows: i) CHCH(R)CH, where R is hydrogen, alkyl, alkenyl, alkynyl, perfluoroalkyl, 35 aryl, heteroaryl, fluoro, hydroxyl or alkoxy; ii) CHC (RR)CH2, where RandR, are each independently alkyl, 40 alkenyl, alkynyl, perfluoroalkyl, aryl, or fluoro, or R and R, are connected together to form a carbocyclic or hetero cyclic ring; iii) CHOCH, CHC(O)CH, or CHCH; or iv) Rs is a carbocyclic, heterocyclic, aryl or heteroaryl ring; and 45 Rs and Ro are independently selected from hydrogen, alkyl, 36 alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or heteroaryl, or Rs and R are connected together to form a carbocyclic or heterocyclic ring; or pharmaceutically acceptable salts thereof. 50 In certain embodiments Rs and Ro are hydrogen. In certain embodiments, a pharmaceutically acceptable salt of the compound is formed by derivatizing E, wherein E is —OM, where M is a cation selected from ammonium, 37

tetra-alkyl ammonium, Na, K, Mg, and Zn. 55 Additional compounds Suitable for use in methods and compositions of the invention include those of Formulae 39-44,

S 60 39 -SX ORf ORe O and pharmaceutically acceptable salts, hydrates and Solvates thereof, wherein: 65 Ro is —OH, —OCH —OCH(CH) or —NHCHCH: and US 9,315,447 B2 73 74 -continued 40

ORf ORe O OH OH O

21N 21 S E, 5 21N 21 ONa. \ \

SiR3 10 Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 46. ORf ORe 9 41 46 15 \ ---, ORf-----, ORe Q 42 25 or a pharmaceutically acceptable salt or prodrug thereof, \ wherein: SiR each see independently designates a double or triple bond; R", R, and R are each independently OR, OX', SR, SX, N(R), NHX, NRC(O)R, NRC(O)N(R), C(O)OR, C(O) Ri ORe Q 43 30 N(R), SOR, NRSOR, C(O)R, or SON(R): each Ris independently selected from hydrogen oran option ally substituted group selected from Caliphatic, a 3-8 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitro \ 35 gen, oxygen, or Sulfur, or, two R on the same nitrogen are taken together with the nitro Ri ORe Q 44 gento form a 5-8 membered heterocyclyl or heteroaryl ring having 1-3 heteroatoms independently selected from nitro gen, oxygen, or Sulfur, tly X-----, 40 each X is independently a suitable hydroxyl protecting group; each X is independently a suitable thiol protecting group; SiR -- each X is independently a suitable amino protecting group; 45 and R is NRC(O)R, NRC(O)N(R), C(O)CR, C(O)N(R), and pharmaceutically acceptable salts thereof, wherein: SOR, NRSOR, C(O)R, or SON(R). Re, Rf, E., Ri, Rs. Rs and Rs are as defined above. Additional compounds Suitable for use in methods and compositions of the invention include those of Formula 47: Exemplary compounds of formulae 39, 41, and 43 include: 50

45 (47) OH 55 OH,

60 and pharmaceutically acceptable salts and esters thereof. In certain embodiments, a pharmaceutically acceptable salt of the compound is formed by derivatizing E, wherein E is —OM, where M is a cation selected from ammonium, 65 tetra-alkyl ammonium, Na, K. Mg, and Zn. Examples of Such compounds include compound Z. US 9,315,447 B2 75 76 or a pharmaceutically acceptable salt or prodrug thereof, In certain embodiments, the stereochemistry of the carbon wherein: kk to carbon 11 double bond is trans, the stereochemistry of the stereochemistry of the carbonkk' to carbon ll' double bond the carbon mm' to carbon nin' double bond trans, and the is cis or trans; stereochemistry of the carbon oo' to carbon pp'double bond is the stereochemistry of the carbon mm' to carbon nin' double C1S. bond is cis or trans; In certain embodiments, a compound of formula 47 is the stereochemistry of the carbon oo' to carbon pp' double represented by compound 48a, bond is cis or trans; Y' is a bond or a linker selected from a ring containing up to 20 atoms or a chain of up to 20 atoms, provided that Y can 10 include one or more nitrogen, oxygen, Sulfur or phospho (48a) rous atoms, further provided that Y can include one or more Substituents independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, iodo, bromo, fluoro, hydroxy, alkoxy, aryloxy, carboxy, amino, 15 alkylamino, dialkylamino, acylamino, carboxamido, cyano, oxo, thio, alkylthio, arylthio, acylthio, alkylsul fonate, arylsulfonate, phosphoryl, or Sulfonyl, further pro vided that Y can contain one or more fused carbocyclic, heterocyclic, aryl or heteroaryl rings; Z is selected from –CN, -C(NH)N(R") (R"), C(S)-A', C(S)R", C(O)-A", C(O) R", C(O). SR", —C(O) NH-S(O). R", S(O)-A', S(O). R", S(O)N(R")(R"), -P(O)-A'. -PO(OR")-A', -tetrazole, 25 alkyltetrazole, or —CHOH, wherein compound 48b. A is selected from OR", N(R")(R") or - OM': each R" is independently selected from hydrogen, alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl or a detect 30 able label molecule, wherein any alkyl-, aryl- or het (48b) eroaryl-containing moiety is optionally substituted with up to 3 independently selected substituents; and M" is a cation. In certain embodiments, a compound of formula 47 is represented by formula 48, 35

(48) 40

45 compound 48c,

(48c) 50 or pharmaceutically acceptable salts and esters thereof, wherein: 55 the stereochemistry of the carbonkk' to carbon ll' double bond is cis or trans; the stereochemistry of the carbon mm' to carbon nin' double bond is cis or trans; the stereochemistry of the carbon oo' to carbon pp' double 60 bond is cis or trans. In certain embodiments, the stereochemistry of the carbon kk" to carbon 11 double bond is trans. In certain embodiments, the stereochemistry of the carbon mm' to carbon nin' double bond trans. 65 or pharmaceutically acceptable salts and esters thereof. In certain embodiments, the stereochemistry of the carbon In certain embodiments, a compound of formula 47 is oo' to carbon pp' double bond is cis. represented by formula 48d, US 9,315,447 B2 77 78 configuration, wherein each R' is independently selected (48d) from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, acyl, silyl, alkoxyacyl, aminoacyl, aminocarbonyl, alkoxy carbonyl, or a protecting group. Exemplary compounds of formula 49 include compound 49a,

(49a) 10

15 or pharmaceutically acceptable salts and esters thereof, wherein: the stereochemistry of the carbonkk' to carbon ll' double bond compound 49b, is cis or trans; the stereochemistry of the carbon mm' to carbon nin' double bond is cis or trans; (49b) the stereochemistry of the carbon oo' to carbon pp' double bond is cis or trans. 25 Additional compounds suitable for use in methods and compositions of the invention include those of Formula 49. W O

(49) 30 or pharmaceutically acceptable salts and esters thereof. The compounds above (e.g., compounds of formula A or formulae 1 to 49) are known to be useful in the treatment or prevention of inflammation or inflammatory disease. 35 Examples of Such compounds are disclosed in the following patents and applications: US 2003/0191184, WO 2004/ 0.14835, WO 2004/078143, U.S. Pat. No. 6,670,396, US 2003/023.6423, US 2005/0228047, US 2005/023.8589 and or a pharmaceutically acceptable salt or prodrug thereof, US2005/0261255. These compounds are suitable for use in wherein: 40 methods and compositions of the present invention. Y' is a bond or a linker selected from a ring containing up to Additional compounds Suitable for use in methods and 20 atoms or a chain of up to 20 atoms, provided that Y can compositions of the invention are compounds that are chemi include one or more nitrogen, oxygen, Sulfur or phospho cally similar variants to any of the compounds of formula A or rous atoms, further provided that Y can include one or formulae 1-49 or I-III set forth above. The term “chemically more Substituents independently selected from hydrogen, 45 similar variants' includes, but is not limited to, replacement alkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, iodo, of various moieties with known biosteres; replacement of the bromo, fluoro, hydroxy, alkoxy, aryloxy, carboxy, amino, end groups of one of the compounds above with a correspond alkylamino, dialkylamino, acylamino, carboxamido, ing end group of any other compound above, modification of cyano, oxo, thio, alkylthio, arylthio, acylthio, alkylsul the orientation of any double bond in a compound, the fonate, arylsulfonate, phosphoryl, or Sulfonyl, further pro 50 replacement of any double bond with a triple bond in any vided that Y can contain one or more fused carbocyclic, compound, and the replacement of one or more Substituents heterocyclic, aryl or heteroaryl rings; present in one of the compounds above with a corresponding Z is selected from –CN, -C(NH)N(R") (R"), C(S)-A', C(S)R", C(O)-A", C(O) R", C(O). SR", Substituent of any other compound. - C(O) NH S(O). R", S(O)-A', S(O). R", 55 Lipoxin compounds Suitable for use in the methods and S(O)N(R")(R"), -P(O)-A', -PO(OR")-A', -tetrazole, compositions of this invention include those of formula 50: alkyltetrazole, or —CH2OH, wherein

A is selected from OR", N(R")(R") or - OM': each R" is independently selected from hydrogen, alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl or a detect 60 able label molecule, wherein any alkyl-, aryl- or het eroaryl-containing moiety is optionally Substituted with up to 3 independently selected substituents; and M" is a cation; and each of R'' and R is independently for each occurrence 65 selected from —OR', or adjacent R'' and Rare taken together to form an epoxide ring having a cis or trans US 9,315,447 B2 79 80 wherein: wherein Yo is —OH, methyl, —SH, an alkyl of 2 to 4 X is Rao, ORao, or SR so; carbonatoms, inclusive, straight chain or branched, analkoxy Rao is of 1 to 4 carbon atoms, inclusive, or (CH),(Z), wherep+q 3. (a) a hydrogen atom; p=0 to 3, q=0 to 3 and Z is cyano, nitro or a halogen; and (b) an alkyl of 1 to 8 carbonatoms, inclusive, which may be 5 T is O or S, and pharmaceutically acceptable salts thereof. straight chain or branched; Lipoxin compounds Suitable for use in the methods and (c) a cycloalkyl of 3 to 10 carbon atoms; compositions of this invention include those of formulae 51, (d) an aralkyl of 7 to 12 carbon atoms; 52, 53 or 54: (e) phenyl: (51) (f) substituted phenyl 10

s R309 R320 R308,

15 R319 OR307 R. R314 R312 OR307 Zii (52) R318 Rs. Rsis 313 311 R307Q R300 Z Ziv R320 V R308, 2O e wherein ZZ, Z, Z, and Z are each independently R319 eOR307 R. selected from —NO. —CN, —C(=O)—Ro, —SOH, a hydrogen atom, halogen, methyl, —OR, wherein R is 1 to 8 carbon atoms, inclusive, which may be a straight chain or branched, and hydroxyl, 25 wherein when any of ZZ, Z, Z, or Z is C(=O)— Roi, said Z.Z, Z, Z, or Z is not substituted with another C(=O)—Ro. (g) a detectable label molecule; or (h) a straight or branched chain alkenyl of 2 to 8 carbon 30 atoms, inclusive; Q is (C=O), SO or (CN), provided when Q is CN, then X is absent; Q and Q are each independently O, S or NH; one of Ro and Ros is a hydrogen atom and the other is: 35 (a) H: (b) an alkyl of 1 to 8 carbonatoms, inclusive, which may be wherein: a straight chain or branched; each Ro, is independently selected from hydrogen and (c) a cycloalkyl of 3 to 6 carbon atoms, inclusive; straight, branched, cyclic, Saturated, or unsaturated alkyl hav 40 ing from 1 to 20 carbon atoms; (d) analkenyl of 2 to 8 carbonatoms, inclusive, which may Ros, Roo, Rio, Rs 19, and Roo are independently selected be straight chain or branched; or from: (e) RQR wherein Q is —O— or - S -, wherein R is (a) hydrogen; alkylene of 0 to 6 carbons atoms, inclusive, which may (b) straight, branched, cyclic, Saturated, or unsaturated be straight chain or branched and wherein R is alkyl of alkyl having from 1 to 20 carbon atoms: 0 to 8 carbon atoms, inclusive, which may be straight 45 (c) substituted alkyl having from 1 to 20 carbon atoms, chain or branched, provided when R is 0, then R is a wherein the alkyl is substituted with one or more sub hydrogen atom; stituents selected from halo, hydroxy, lower alkoxy, ary Rao.4 is loxy, amino, alkylamino, dialkylamino, acylamino, ary (a) H: lamino, hydroxyamino, alkoxyamino, alkylthio. (b) an alkyl of 1 to 6 carbonatoms, inclusive, which may be 50 arylthio, carboxy, carboxamido, carboalkoxy, aryl, and a straight chain or branched; heteroaryl; Rsos is (d) substituted aryl or heteroaryl, wherein the aryl or het eroaryl is substituted with one or more substituents Selected from alkyl, cycloalkyl, alkoxy, halo, aryl, het Zi Zii 55 eroaryl, carboxyl, and carboxamido; and (e) Z Y, wherein: Z is selected from a straight, branched, cyclic, Saturated, or Ziii, unsaturated alkyl having from 1 to 20 carbon atoms; Substi tuted lower alkyl, wherein the alkyl is substituted with one or 60 more Substituents selected from halo, hydroxy, lower alkoxy, Z Ziv aryloxy, amino, alkylamino, dialkylamino, acylamino, ary lamino, hydroxyamino, alkoxyamino, alkylthio, arylthio, car wherein ZZ, Z, Z, and Z are defined as above; boxy, carboxamido, carboalkoxy, aryl, and heteroaryl; and Rao is substituted aryl or heteroaryl, wherein the aryl or heteroaryl is (a) H: 65 substituted with one or more substituents selected from alkyl, (b) an alkyl from 1 to 4 carbon atoms, inclusive, straight cycloalkyl, alkoxy, halo, aryl, heteroaryl, carboxyl, and car chain or branched; boxamido; and US 9,315,447 B2 81 82 Y is selected from hydrogen; alkyl cycloalkyl; carboxyl: Rao is selected from: carboxamido: aryl; heteroaryl; substituted aryl or heteroaryl, wherein the aryl or heteroaryl is substituted with one or more (forms ring) Substituents selected from alkyl, cycloalkyl, alkoxy, halo, R415. aryl, heteroaryl, carboxyl, and carboxamido; and R to Rs are independently selected from: (a) hydrogen; (b) halo: (c) straight, branched, cyclic, Saturated, or unsaturated alkyl having from 1 to 20 carbon atoms: (d) substituted alkyl having from 1 to 20 carbon atoms, wherein the alkyl is substituted with one or more sub stituents selected from halo, hydroxy, lower alkoxy, ary loxy, amino, alkylamino, dialkylamino, acylamino, ary lamino, hydroxyamino, alkoxyamino, alkylthio. arylthio, carboxy, carboxamido, carboalkoxy, aryl, and heteroaryl; (e) substituted aryl or heteroaryl, wherein the aryl or het eroaryl is substituted with one or more substituents R422 R423 s Selected from alkyl, cycloalkyl, alkoxy, halo, aryl, het (forms ring) eroaryl, carboxyl, and carboxamido; or Ros to Ro are independently a bond that forms a carbon 25 carbon double bond, a carbon-carbon triple bond, or a ring with the lipoxin backbone; or any two of Ro, to Ro are taken together with the atoms to which they are bound and optionally to 1 to 6 oxygen atoms, 30 1 to 6 nitrogen atoms, or both 1 to 6 oxygen atoms and 1 to 6 Xois Rai1, OR, or SR-11; nitrogen atoms, to form a ring containing 3 to 20 atoms. R41 is (a) a hydrogen atom; Lipoxin compounds Suitable for use in the methods and (b) an alkyl of 1 to 8 carbonatoms, inclusive, which may be compositions of this invention include those of formula 55: straight chain or branched; 35 (c) a cycloalkyl of 3 to 10 carbon atoms: (d) an aralkyl of 7 to 12 carbon atoms; (55) (e) phenyl, R401 (f) substituted phenyl r 40 R402 wherein: Ziii Rao is selected from: 45 Z Ziv

HO wherein ZZ, Z, Z, and Z are each independently 50 selected from —NO. —CN, —C(=O)—R, —SOH, a hydrogen atom, halogen, methyl, —OR, wherein R is 1 to 8 carbon atoms, inclusive, which may be a straight chain or branched, and hydroxyl, wherein when any of ZZ, Z, Z, or Z is C(=O)— 55 R, said Z.Z, Z, Z, or Z is not substituted with 2. another C(=O)—R. R413a R4135 (g) a detectable label molecule; or HO OH O (h) a straight or branched chain alkenyl of 2 to 8 carbon atoms, inclusive; 60 Q is (C=O), SO, or (CN); NY (-Nulls. O Q is O, S or NH; one of R and R is a hydrogen atom and the other is selected from: (CH2),l OR.421; (a) H: 65 (b) an alkyl of 1 to 8 carbonatoms, inclusive, which can be straight chain or branched; (c) a cycloalkyl of 3 to 6 carbon atoms, inclusive; US 9,315,447 B2 83 84 (d) an alkenyl of 2 to 8 carbon atoms, inclusive, which can (c) an alkyl of 2 to 4 carbonatoms, inclusive, straight chain be straight chain or branched; or or branched; or (e) RQR wherein Q is —O— or —S—, wherein (d) an alkoxy of 1 to 4 carbon atoms, inclusive, Ra is alkylene of 0 to 6 carbons atoms, inclusive, which can be straight chain or branched and wherein R is or Yao and Yao taken together are: alkyl of 0 to 8 carbon atoms, inclusive, which can be (a) —NH; or straight chain or branched; (b)=O; Ras, and Ras, are each independently: one of Yaos or Yao is —OH, methyl, or—SH, and wherein (a) H: the other is selected from: (b) an alkyl of 1 to 8 carbonatoms, inclusive, which can be 10 straight chain or branched; (a) H (c) a cycloalkyl of 3 to 6 carbon atoms, inclusive; (b) (CH), (Z), wherein Z. p, and q are as defined above: (d) an alkenyl of 2 to 8 carbon atoms, inclusive, which can (c) an alkyl of 2 to 4 carbonatoms, inclusive, straight chain be straight chain or branched; or or branched; or (e) Ras Q-Ras, wherein Ras, Q2, and Ras are as defined 15 (d) an alkoxy of 1 to 4 carbon atoms, inclusive, above; or Yao and Yao taken together are: R44 is (a) H: (a) —NH; or (b) an alkyl of 1 to 6 carbon atoms, inclusive, can be (b)=O; straight chain or branched; R42 is Rais is (a) an alkyl of 1 to 9 carbon atoms which can be straight (a) H; or chain or branched; (b) alkyl of 1 to 8 carbon atoms; (b) —(CH)—R, Ra and Ra are each independently: wherein n=0 to 4 and R, is 25 (a) H: (i) a cycloalkyl of 3 to 10 carbon atoms, inclusive; (ii) a phenyl; or (b) a hydroxyl, or a thiol: (iii) substituted phenyl (c) a methyl or a halomethyl; (d) a halogen; or 30 (e) an alkoxy of 1 to 3 carbon atoms; Ra and Ras are each independently: (a) H:

35 (b) a hydroxyl, or a thiol: (c) a methyl or a halomethyl; (d) a halogen; wherein Z, through Z are as defined above: (e) an alkoxy of 1 to 3 carbon atoms; or (b) RQR, wherein Ra, Q, and Ras are as defined 40 (f) an alkyl or haloalkyl of 2 to 4 carbon atoms inclusive, above; which can be straight chain or branched; and (c) —C(R)(R)-R, R426 is wherein R, and Rare each independently: (a) a Substituted phenyl (i) a hydrogen atom; (ii) (CH), (Z), wherein Z. p, and q are as defined above; 45 (e) a haloalkyl of 1 to 8 carbon atoms, inclusive, and 1 to 6 halogen atoms, inclusive, straight chain or branched; R416 is (a) H: (b) an alkyl from 1 to 4 carbon atoms, inclusive, straight 50 chain or branched; (c) a halogen; one of Yao or Yao is —OH, methyl, or—SH, and wherein the other is selected from: wherein Z, through Zare as defined above: (a) H: 55 (b)(CH), (Z), where p+q3, p=0 to 3, q=0 to 3 and each Z. (b) a substituted phenoxy independently, is cyano, nitro or a halogen; (c) an alkyl of 2 to 4 carbonatoms, inclusive, straight chain or branched; or Zi Zii (d) an alkoxy of 1 to 4 carbon atoms, inclusive, 60 or Yao and Yao taken together are: (d) —NH; or (e)=O: – O -- Ziii one of Yao or Yao is —OH, methyl, or—SH, and wherein Z Ziv the other is selected from: 65 (a) H: (b) (CH),(Z), wherein Z. p, and q are as defined above; wherein Z, through Z are as defined above; or US 9,315,447 B2 85 86 (c) that J'and K" can independently include one or more substitu ents selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, chloro, iodo, bromo, fluoro, hydroxy, alkoxy, ary Zi Zii loxy, carboxy, amino, alkylamino, dialkylamino, acylamino, carboxamido, cyano, Oxo, thio, alkylthio, arylthio, acylthio. 5 alkylsulfonate, arylsulfonate, phosphoryl, and Sulfonyl, and further provided that J and K" can also contain one or more fused carbocyclic, heterocyclic, aryl or heteroaryl rings, and provided that linkers and K" are connected to the adjacent ---Z Ziv C(R)OR group via a carbon atom or a C-heteroatom bond 10 where the heteroatomis oxygen, Sulfur, phosphorous or nitro gen, wherein Z, through Z are as defined above. G is selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, Lipoxin compounds Suitable for use in the methods and heteroaryl, chloro, iodo, bromo, fluoro, hydroxy, alkoxy, ary compositions of this invention include those of formula 56: loxy, carboxy, amino, alkylamino, dialkylamino, acylamino, 15 and carboxamido. Re, Rf and Rg, are independently selected from hydrogen, (56) alkyl, aryl, heteroaryl, acyl, silyl, alkoxyacyl and aminoacyl: R502O OR50 O Roo, Roo and Roos are independently selected from hydrogen, alkyl, aryland heteroaryl, provided that R. R. and Roos can independently be connected to linkers J" or K'; Roa and Ros are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, fluoro, and provided C that Ro and Ros can be joined together to form a carbocy clic, heterocyclic or aromatic ring, and further provided that 25 Roa and Ros can be replaced by a bond to form a triple bond. Other compounds suitable for use in the methods and com positions of the invention are the oxylipins described in inter national applications WO 2006055965, WO 2007090162, and WO2008103753 the compounds in which are incorpo wherein: rated herein by reference. Examples of Such compounds are E is hydroxy, alkoxy, aryloxy, amino, alkylamino, dialky 30 those of formulae 58-132, as shown in Table 2. These com lamino or —OM, where M is a cation selected from ammo pounds include long chain omega-6 fatty acids, docosapen nium, tetra-alkyl ammonium, and the cations of sodium, taenoic acid (DPAn-6) (compounds 58-73) and docosatet potassium, magnesium and Zinc; raenoic acid (DTAn-6) (compounds 74-83), and the omega-3 W is hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, counterpart of DPAn-6, docosapentaenoic acid (DPAn-3) halo, hydroxy, alkoxy, aryloxy, carboxy, amino, alkylamino, 35 (compounds 84-97). Further compounds are the docosanoids dialkylamino, acylamino, carboxamido, or Sulfonamide; 98-115, the Y-linolenic acids (GLA) (compounds 116-122), each of Rso-Rs are independently selected from hydro and the stearidonic acids (SDA) (compounds 123-132). gen, alkyl, aryl, acyl or alkoxyacyl:

n is 0, 1 or 2; 40 TABLE 2 m is 1 or 2; and 10,17 the two Substituents on the phenyl ring are ortho, meta, or Dihydroxy para. DPA n-6 CO2H Lipoxin compounds Suitable for use in the methods and (58) compositions of this invention include those of formula 57: 45

(57) 16,17 CO2H Dihydroxy 50 DPA n-6 (59)

4,5- OH 55 Dihydroxy DPA n-6 CO2H (60) wherein:

I is selected from: C(O)-E, -SO-E, PO(OR)-E, 7,17 where E is hydroxy, alkoxy, aryloxy, amino, alkylamino, 60 Dihydroxy dialkylamino, or —OM, where M is a cation selected from DPA n-6 CO2H ammonium, tetra-alkyl ammonium, Na, K, Mg, and Zn; and (61) R is hydroxyl or alkoxy J and K" are linkers independently selected from a chain of up to 20 atoms and a ring containing up to 20 atoms, provided 65 OH that J'and K" can independently include one or more nitrogen, oxygen, Sulfur or phosphorous atoms, and further provided

US 9,315,447 B2 91 92 TABLE 2-continued TABLE 2-continued

10.11 epoxy 4.5-Epoxy DPA 17-OHDPA (112) CO2H (103) CO2H

19.20 epoxy CO2H 10 DPA

(113) 7,16,17 Trihydroxy DTAn-3 CO2H (104) 15 7-hydroxy DHA (114) CO2H

16,17 CO2H Dihidroxy DTAn-3 (105) 13,14 epoxy CO2H DPA (115)

10,16,17 OH 25 Trihydroxy DTRAn-6 CO2H (106) 6-hydroxy GLA 30 (116) HO OH

16,17 CO2H Dihydroxy DTRAn-6 10-hydroxy 35 (107) GLA (117) HO OH

7,16,17 OH Trihydroxy CO2H 40 DTRAn-6 7-hydroxy (108) GLA (118)

HO OH 45

15-epi HO OH lipoxin A4 12-hydroxy (109) CO2H GLA (119) 50

9-hydroxy 16,17-epoxy CO2H 55 GLA DHA (120) (110)

13-hydroxy 7,8-epoxy GLA DPA (121) (111) CO2H

65 US 9,315,447 B2 93 94 TABLE 2-continued TABLE 2-continued

6,13 6,16 dihydroxy dihydroxy GLA SDA (122) (132)

10 6-hydroxy SDA Other oxylipin compounds that are suitable for use in (123) methods and compositions of the invention include analogs of the compounds shown in Table 2. Such compounds include but are not limited to those analogs wherein one or more 15 double bonds are replaced by triple bonds, those wherein one or more carboxy groups are derivatized to form esters, amides 10-hydroxy or salts, those wherein the hydroxyl-bearing carbons are fur SDA ther derivatized (with, for example, a substituted or unsubsti (124) tuted, branched or unbranched alkyl, alkenyl, or alkynyl group, Substituted or unsubstituted aryl group, Substituted or unsubstituted, branched or unbranched alkylaryl group, halo gen atom) to form tertiary alcohols (or ethers, esters, or other 7-hydroxy SDA derivatives thereof), those wherein one or more hydroxyl (125) groups are derivatized to form esters or protected alcohols, or 25 those having combinations of any of the foregoing modifica tions. Further oxylipin compounds suitable for use in methods and compositions of the invention include the following: 12-hydroxy isolated docosanoids of docosapentaenoic acid (DPAn-6); SDA 30 monohydroxy, dihydroxy, and trihydroxy derivatives of (126) DPAn-6; isolated docosanoids of docosapentaenoic acid (DPAn-3); monohydroxy, dihydroxy, and trihydroxy deriva tives of DPAn-3; isolated docosanoids of docosapentaenoic acid (DTAn-6); or monohydroxy, dihydroxy, and trihydroxy 9-hydroxy 35 derivatives of DTAn-6. SDA In certain embodiments, the present invention provides (127) methods of treating or preventing an ophthalmic condition (such as dry eye), comprising administering an effective amount of any one or more of the compounds described above 40 (e.g., a compound of any of formulae I-IX, a compound of 13-hydroxy formula A, a compound of any one of formulae 1-49, a lipoxin SDA compound, or an oxylipin compound), e.g., as an aqueous (128) Solution as described herein, such as topically to the eye, e.g., as eye drops. In certain Such embodiments, the methods of 45 treating or preventing an ophthalmic condition comprise administering greater than 6 nanomoles of any of the com 15-hydroxy pounds described above (e.g., a compound of any of formulae SDA I-IX, a compound of formula A, a compound of any one of (129) formulae 1-49, a lipoxin compound, or an oxylipin com 50 pound) per treated eye per day, e.g., up to 550 nanomoles per treated eye per day, such as from 6 to 400 nanomoles, from 6 to 300 nanomoles, from 6 to 250 nanomoles, from 6 to 200 16-hydroxy nanomoles, from 6 to 150, or from 6 to 100 nanomoles of any SDA of the compounds described above (e.g., a compound of any (130) 55 of formulae I-IX, a compound of formula A, a compound of any one of formulae 1-49, a lipoxin compound, oran oxylipin compound) per treated eye per day. In certain Such embodi ments, the methods of treating or preventing an ophthalmic 6,13 condition comprise administering greater than 7 nanomoles dihydroxy 60 of any of the compounds shown above (e.g., a compound of SDA any of formulae I-IX, a compound of formula A, a compound (131) of any one of formulae 1-49, a lipoxin compound, or an oxylipin compound) per treated eye per day, Such as greater than 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, or 50 65 nanomoles of any of the compounds shown above (e.g., a compound of any of formulae I-IX, a compound of formula A, a compound of any one of formulae 1-49, a lipoxin com US 9,315,447 B2 95 96 pound, oran oxylipincompound) per treated eye per day, e.g., day, such as from 2 to 150 micrograms, from 2 to 125 micro up to 100, 150, 200, 250, 300, 400, or even up to 550 nano grams, from 2 to 100 micrograms, from 2 to 75 micrograms, moles per treated eye per day. For example, the methods of from 2 to 50 micrograms, or from 2 to 25 micrograms of treating or preventing an ophthalmic condition comprise compound 1001 per treated eye per day. In certain such administering from 7 to 550 nanomoles of any of the com 5 embodiments, the methods of treating or preventing an oph pounds described above (e.g., a compound of any of formulae thalmic condition comprise administering greater than 2.5 I-IX, a compound of formula A, a compound of any one of micrograms of compound 1001 per treated eye per day. Such formulae 1-49, a lipoxin compound, or an oxylipin com as greater than 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9,9.5, pound) per treated eye per day, such as from 7 to 400 nano 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, moles, from 7 to 300 nanomoles, from 7 to 250 nanomoles, 10 16.5, 17, 17.5, 18, 18.5, 19, 19.5, or 20 micrograms of com from 7 to 200 nanomoles, from 7 to 150, or from 7 to 100 pound 1001 per treated eye per day, e.g., up to 25, 50, 75, 100, nanomoles of any of the compounds described above (e.g., a 125, 150, or even up to 175 micrograms per treated eye per compound of any of formulae I-IX, a compound of formula A, day. In certain embodiments, the methods of treating or pre a compound of any one of formulae 1-49, a lipoxin com venting an ophthalmic condition comprise administering pound, or an oxylipin compound) per treated eye per day. In 15 from 2.5 to 175 micrograms of compound 1001 per treated certain embodiments, the methods of treating or preventing eye per day, such as from 2.5 to 150 micrograms, from 2.5 to an ophthalmic condition comprise administering from 10 to 125 micrograms, from 2.5 to 100 micrograms, from 2.5 to 75 550 nanomoles of any of the compounds described above micrograms, from 2.5 to 50 micrograms, or from 2.5 to 25 (e.g., a compound of any of formulae I-IX, a compound of micrograms of compound 1001 per treated eye per day. In formula A, a compound of any one of formulae 1-49, a lipoxin certain embodiments, the methods of treating or preventing compound, oran oxylipin compound) per treated eye per day, an ophthalmic condition comprise administering from 3 to such as from 10 to 400 nanomoles, from 10 to 300 nanomoles, 175 micrograms of compound 1001 per treated eye per day, from 10 to 250 nanomoles, from 10 to 200 nanomoles, from Such as from 3 to 150 micrograms, from 3 to 125 micrograms, 10 to 150, or from 10 to 100 nanomoles of any of the com from 3 to 100 micrograms, from 3 to 75 micrograms, from 3 pounds described above (e.g., a compound of any of formulae 25 to 50 micrograms, or from 3 to 25 micrograms of compound I-IX, a compound of formula A, a compound of any one of 1001 per treated eye per day. In certain embodiments, the formulae 1-49, a lipoxin compound, or an oxylipin com methods of treating or preventing an ophthalmic condition pound) per treated eye per day. In certain embodiments, the comprise administering from 4 to 175 micrograms of com methods of treating or preventing an ophthalmic condition pound 1001 per treated eye per day, such as from 4 to 150 comprise administering from 15 to 550 nanomoles of any of 30 micrograms, from 4 to 125 micrograms, from 4 to 100 micro the compounds described above (e.g., a compound of any of grams, from 4 to 75 micrograms, from 4 to 50 micrograms, or formulae I-IX, a compound of formula A, a compound of any from 4 to 25 micrograms of compound 1001 per treated eye one of formulae 1-49, a lipoxin compound, or an oxylipin per day. In certain embodiments, the methods of treating or compound) per treated eye per day, such as from 15 to 400 preventing an ophthalmic condition comprise administering nanomoles, from 15 to 300 nanomoles, from 15 to 250 nano 35 from 5 to 175 micrograms of compound 1001 per treated eye moles, from 15 to 200 nanomoles, from 15 to 150, or from 15 per day, such as from 5 to 150 micrograms, from 5 to 125 to 100 nanomoles of any of the compounds described above micrograms, from 5 to 100 micrograms, from 5 to 75 micro (e.g., a compound of any of formulae I-IX, a compound of grams, from 5 to 50 micrograms, or from 5 to 25 micrograms formula A, a compound of any one of formulae 1-49, a lipoxin of compound 1001 per treated eye per day. In certain embodi compound, oran oxylipin compound) per treated eye per day. 40 ments, the methods of treating or preventing an ophthalmic In certain embodiments, the methods of treating or preventing condition may comprise administering the dosages of com an ophthalmic condition may comprise administering the pound 1001 as set forth above over the course of a day through dosages of any of the compounds described above (e.g., a any Suitable dosing regimen. Suitable dosing regimens may compound of the invention, such as a compound of any of include once daily dosing, twice daily dosing, three times formulae I-IX, a compound of formula A, a compound of any 45 daily dosing, four times daily dosing, and any other Suitable one of formulae 1-49, a lipoxin compound, or an oxylipin dosing regimen, such that the net effect throughout the course compound) as set forth above over the course of a day through of the day is administering the total dosages per eye per day of any suitable dosing regimen. Suitable dosing regimens may compound 1001 as set forth above. In certain embodiments, include once daily dosing, twice daily dosing, three times Suitable dosing regimens further include once every two days daily dosing, four times daily dosing, and any other Suitable 50 dosing, such as every other day, or once every three days dosing regimen Such that the net effect throughout the course dosing, Such as every third day, such that the net effect of the day is administering the total dosages per eye per day as throughout the course of the day of dosing is administering at set forth above. In certain embodiments, Suitable dosing regi least the total dosages per eye per day of compound 1001 as mens further include once every two days dosing, such as set forth above. every other day, or once every three days dosing, such as 55 In certain embodiments, the present invention provides a every third day, such that the net effect throughout the course pharmaceutical preparation Suitable for treating or preventing of the day of dosing is administering at least the total dosages an ophthalmic condition (such as dry eye) in a human patient, per eye per day as set forth above. comprising any one or more of the compounds shown above As a particular example, the present invention provides (e.g., a compound of any of formulae I-IX, a compound of methods of treating or preventing an ophthalmic condition 60 formula A, a compound of any one of formulae 1-49, a lipoxin (such as dry eye), comprising administering an effective compound, or an oxylipin compound), and one or more phar amount of compound 1001, e.g., as an aqueous solution as maceutically acceptable excipients, e.g., as an aqueous solu described herein, such as topically to the eye, e.g., as eye tion Suitable for topical administration to the eye. Such as eye drops. In certain Such embodiments, the methods of treating drops. In certain embodiments, the pharmaceutical prepara or preventing an ophthalmic condition comprise administer 65 tion has a concentration over 90 micromolar of any of the ing greater than 2 micrograms of compound 1001 per treated compounds described above (e.g., a compound of any of eye per day, e.g., up to 175 micrograms per treated eye per formulae I-IX, a compound of formula A, a compound of any US 9,315,447 B2 97 98 one of formulae 1-49, a lipoxin compound, or an oxylipin from 30 ug/mL to 1500 g/mL, 30 ug/mL to 1000 g/mL, 30 compound), e.g., from 90 micromolar to 7000 micromolar, ug/mL to 750 ug/mL, 30 g/mL to 500 g/mL, 30 g/mL to such as from 90 to 5000 micromolar, 90 to 3000 micromolar, 400 ug/mL, or 30 ug/mL to 350 g/mL of compound 1001. In 90 to 2000 micromolar, or 90 to 1000 micromolar of any of certain embodiments, the pharmaceutical preparation has a the compounds described above (e.g., a compound of any of 5 concentration of greater than 40 ug/mL of compound 1001, formulae I-IX, a compound of formula A, a compound of any Such as greater than 50 ug/mL, greater than 60 g/mL, greater one of formulae 1-49, a lipoxin compound, or an oxylipin than 70 g/mL, greater than 75 ug/mL, greater than 100 compound). In certain embodiments, the a pharmaceutical ug/mL, greater than 125 ug/mL, greater than 150 ug/mL, preparation has a concentration of greater than 100 micromo greater than 175ug/mL, greater than 200 g/mL, greater than lar of any of the compounds described above (e.g., a com 10 225 g/mL, or greater than 250 ug/mL of compound 1001, pound of any of formulae I-IX, a compound of formula A, a e.g., up to 350 ug/mL, up to 400 ug/mL, up to 500 g/mL, up compound of any one of formulae 1-49, a lipoxin compound, to 750 ug/mL, up to 1000 ug/mL, up to 1500 ug/mL, or even or an oxylipin compound), such as greater than 150 micro up to 2000 ug/mL. For example, the pharmaceutical prepara molar, greater than 200 micromolar, greater than 250 micro tion may have a concentration from 40 ug/mL to 2000 ug/mL molar, greater than 300 micromolar, greater than 350 micro 15 of compound 1001, such as from 40 ug/mL to 1500 g/mL, 40 molar, greater than 400 micromolar, greater than 450 ug/mL to 1000 ug/mL, 40 ug/mL to 750 g/mL, 40 ug/mL to micromolar, greater than 500 micromolar, greater than 550 500 ug/mL, 40 ug/mL to 400 ug/mL, or 40 ug/mL to 350 micromolar, greater than 600 micromolar, greater than 650 ug/mL of compound 1001. In certain embodiments, the phar micromolar, greater than 700 micromolar, greater than 750 maceutical preparation has a concentration from 50 ug/mL to micromolar, or greater than 800 micromolar of any of the 2000 ug/mL of compound 1001, such as from 50 lug/mL to compounds described above (e.g., a compound of any of 1500 ug/mL, 50 g/mL to 1000 ug/mL, 50 g/mL to 750 formulae I-IX, a compound of formula A, a compound of any ug/mL, 50 g/mL to 500 g/mL, 50 ug/mL to 400 ug/mL, or one of formulae 1-49, a lipoxin compound, or an oxylipin 50 ug/mL to 350 g/mL of compound 1001. In certain compound), e.g., up to 1000 micromolar, up to 2000 micro embodiments, the pharmaceutical preparation has a concen molar, up to 3000 micromolar, up to 5000 micromolar, or 25 tration from 60 g/mL to 2000 ug/mL of compound 1001, even up to 7000 micromolar. For example, the pharmaceuti such as from 60 g/mL to 1500 lug/mL, 60 lug/mL to 1000 cal preparation may have a concentration from 100 micromo ug/mL, 60 g/mL to 750 g/mL, 60 ug/mL to 500 ug/mL, 60 lar to 7000 micromolar of any of the compounds described ug/mL to 400 g/mL, or 60 g/mL to 350 ug/mL of com above (e.g., a compound of any of formulae I-IX, a compound pound 1001. In certain embodiments, the pharmaceutical of formula A, a compound of any one of formulae 1-49, a 30 preparation has a concentration from 75 lug/mL to 2000 lipoxin compound, or an oxylipin compound). Such as from ug/mL of compound 1001, such as from 75 lug/mL to 1500 100 to 5000 micromolar, 100 to 3000 micromolar, 100 to ug/mL, 75ug/mL to 1000 ug/mL, 75ug/mL to 750Lig/mL, 75 2000 micromolar, or 100 to 1000 micromolar of any of the ug/mL to 500 g/mL, 75ug/mL to 400 ug/mL, or 75ug/mL to compounds described above (e.g., a compound of any of 350 ug/mL of compound 1001. formulae I-IX, a compound of formula A, a compound of any 35 In certain embodiments of the methods of treating or pre one of formulae 1-49, a lipoxin compound, or an oxylipin venting an ophthalmic condition (Such as dry eye), compris compound). In certain embodiments, the pharmaceutical ing administering an effective amount of any one or more of preparation has a concentration from 150 micromolar to 7000 the compounds described above (e.g., a compound of any of micromolar of any of the compounds described above (e.g., a formulae I-IX, a compound of formula A, a compound of any compound of any of formulae I-IX, a compound of formula A, 40 one of formulae 1-49, a lipoxin compound, or an oxylipin a compound of any one of formulae 1-49, a lipoxin com compound), e.g., as an aqueous Solution as described herein, pound, or an oxylipin compound), such as from 150 to 5000 Such as topically to the eye, e.g., as eye drops, compound micromolar, 150 to 3000 micromolar, 150 to 2000 micromo 1001 is the preferred compound. lar, or 150 to 1000 micromolar of any of the compounds In certain embodiments of any of the foregoing pharma described above (e.g., a compound of any of formulae I-IX, a 45 ceutical preparations (e.g., any of the pharmaceutical prepa compound of formula A, a compound of any one of formulae rations described above comprising a compound of any of 1-49, a lipoxin compound, or an oxylipin compound). In formulae I-IX, a compound of formula A, a compound of any certain embodiments, the pharmaceutical preparation has a one of formulae 1-49, a lipoxin compound, or an oxylipin concentration from 90 micromolar to 7000 micromolarofany compound, Such as compound 1001, and one or more phar of the compounds described above (e.g., a compound of any 50 maceutically acceptable excipients), the pharmaceutical of formulae I-IX, a compound of formula A, a compound of preparation is an aqueous solution Suitable for topical admin any one of formulae 1-49, a lipoxin compound, oran oxylipin istration to the eye wherein the solution has a pH in the range compound), such as 250 to 5000 micromolar, 250 to 3000 of 5.5 to 7.4, such as from 5.5 to 7.0, or from 5.5 to 6.5, or from micromolar, 250 to 2000 micromolar, or 250 to 1000 micro 5.5 to 6.0. In certain embodiments of any of the foregoing molar of any of the compounds described above (e.g., a com 55 pharmaceutical preparations (e.g., any of the pharmaceutical pound of any of formulae I-IX, a compound of formula A, a preparations described above comprising a compound of any compound of any one of formulae 1-49, a lipoxin compound, of formulae I-IX, a compound of formula A, a compound of or an oxylipin compound). any one of formulae 1-49, a lipoxin compound, oran oxylipin As a particular example, the present invention provides a compound, Such as compound 1001, and one or more phar pharmaceutical preparation Suitable for treating or preventing 60 maceutically acceptable excipients), the pharmaceutical an ophthalmic condition (such as dry eye) in a human patient, preparation is an aqueous solution Suitable for topical admin comprising compound 1001 and one or more pharmaceuti istration to the eye wherein the solution has a pH in the range cally acceptable excipients, e.g., as an aqueous solution Suit of 5.0 to 7.4, such as from 5.0 to 7.0, or from 5.0 to 6.5, such able for topical administration to the eye, Such as eye drops. In as from 5.0 to 6.0, or from 5.0 to 5.5. In certain embodiments certain embodiments, the pharmaceutical preparation has a 65 of the foregoing, the Solution further comprises one or more concentration over 30 micrograms/milliliter (ug/mL) of com Surfactants, one or more demulscents, and/or one or more pound 1001, e.g., from 30 ug/mL to 2000 ug/mL, such as emulsifiers, such as polysorbate 80, pluronic F-147, tylox US 9,315,447 B2 99 100 apol, polyvinylpyrrolidone (such as polyvinylpyrrolidone neovascularization due to penetration of the eye or contusive having an average molecular weight of 360,000, e.g., polyvi ocular injury; non-infectious uveitis; ocular herpes; ocular nylpyrrolidone K-90, Chemical Abstracts Service Registry rosacea; ophthalmic infections (e.g., corneal herpes, bacterial No. 9003-39-8), mineral oil or castor oil. In certain embodi keratitis, bacterial conjunctivitis, mycotic keratitis, acan ments, the pharmaceutical preparation is substantially free of 5 thamebic keratitis, infectious endophthalmitis, infectious preservatives. corneal ulcer, inflammation of the conjunctiva or cornea by In certain embodiments of the present invention, a phar staphylococci, Streptococci, enterococci, euterococci, bacil maceutical preparation as set forth above (e.g., having a con lus, corynebacterium, chlamydia, and neisseria); ophthalmic centration as set forth above), may be administered using any pemphigoid, optic disc drusen; optic neuritis; panuveitis; Suitable dosing regimen for the treatment or prevention of an 10 papilledema; papillitis; pars planitis; persistent macular ophthalmic condition. Suitable dosing regimens for an aque edema; phacoanaphylaxis; posterior uveitis (chorioentinitis); ous eye drop Solution include administering the pharmaceu post-operative inflammation (e.g., post-LASIK inflammation tical preparation once, twice, three times, or four times per of the cornea); proliferative diabetic retinopathy; prolifera day to an affected eye. In certain embodiments, Suitable dos tive sickle cell retinopathy; proliferative vitreoretinopathy; ing regimens for an aqueous eye drop solution further include 15 retinal artery occlusion; retinal detachment; retinal vasculitis; once every two days dosing, such as every other day, or once retinal vein occlusion; retinitis pigmentosa; retinopathy of every three days dosing, Such as every third day, to an affected prematurity; rubeosis iritis; Scleritis; Stevens-Johnson syn eye. In certain embodiments, any particular incidence of drome (erythema multiforme major); sympathetic oph administration of a pharmaceutical preparation as set forth thalmia; temporal arteritis; toxic retinopathy: uveitis (e.g., above (e.g., having a concentration as set forth above), may anterior uveitis or posterior uveitis); Vernal conjunctivitis; comprise administering one or more drops of the pharmaceu Vitamin A insufficiency-induced keratomalacia; vitreitis; and tical preparation to an affected eye. In certain Such embodi wet age-related macular degeneration. ments, any particular incidence of administration of a phar Diseases causing dry eye include Riley-Day Syndrome, maceutical preparation as set forth above (e.g., having a Shy-Drager syndrome, Sjogren syndrome, sarcoidosis, amy concentration as set forth above), may comprise administer 25 loidosis, sequela of radiotherapy, lagophthalmia, avitamino ing two, three, four, or five drops of the pharmaceutical prepa sis A, Stevens-Johnson syndrome, ocular pemphigoid, mar ration to an affected eye. The present invention further con ginal blepharitis, meibomitis, sequela of intraocular Surgery, templates any and all combinations of the foregoing. contact-lens affection, diabetic corneal epitheliopathy, dry Examples of ophthalmic conditions that may be treated by eye due to VDT operation, graft versus host disease, and the administration of a compound or formulation of the inven 30 like. Disorders caused by corneal infective disease include, tion, include, but are not limited to, AIDS-related retinal for example, viral epitheliopathy and the like. Stem cell disorders; age-related macular degeneration; alkaline erosive depletion syndromes include Stevens-Johnson syndrome, keratoconjunctivitis; allergic keratitis; anterior ischemic ocular pemphigoid, thermal or chemical burn, drug toxicity optic neuropathy; anterior uveitis (iridocyclitis); Behcet’s of idoxuridine (IDU) and therapeutic agents for glaucoma, disease; blepharitis; seborrheic blepharitis; canaliculitis; 35 and the like. cataract; central serous chorioretinopathy; chorioiditis; The present invention provides a method of inhibiting chronic uveitis; Coats disease; conjunctivitis (e.g., infectious COX-2 or TNF in the eye in a patient comprising administer conjunctivitis, neonatal conjunctivitis, non-infectious con ing to said patient a compound of the invention. The present junctivitis, and allergic conjunctivitis); contact lens-induced invention further provides a method of protecting against keratoconjunctivitis; contact eczema, corneal ulcer (e.g., 40 goblet cell loss in the eye in a patient comprising administer Mooren’s ulcer, corneal ulcer Subsequent to chronic rheuma ing to said patient a compound of the invention. toid arthritis or collagen disease, Terrien’s marginal degen Compounds as described herein may also inhibit inflam eration, catarrhal corneal ulcer, infectious corneal ulcer); matory mediators in the cornea, such as TNF, IL-1a, IL-1b. crystalline retinopathy; cyclitis; edema (e.g., cystoid macular IL-6, and IL-8. Accordingly, these compounds may be useful edema), dacryoadenitis, dacryocystitis; degenerative myo 45 in the treatment of dry eyes diseases, age-related macular pia; degenerative retinoschisis; diabetic keratophathy; dia degeneration, retinopathy of prematurity, uveitis, and glau betic macular edema; diabetic retinopathy; dry eye disease COa. (e.g., dry eye of the lacrimal system or dry eye of the cornea): Compounds as described herein may also inhibit COX-2 in dry age-related macular degeneration; endophthalmitis; epis the cornea. Accordingly, these compounds may be useful in cleritis; exudative macular edema, Fuchs’ Dystrophy: giant 50 the treatment of dry eyes diseases. cell arteritis; giant papillary conjunctivitis; glaucoma (e.g., Compounds as described herein may also guard against primary open angle glaucoma, primary angle closure glau goblet cell loss. Accordingly, these compounds may be useful coma, secondary open angle glaucoma, secondary angle clo in the treatment of dry eye diseases, age-related macular Sure glaucoma, and childhood glaucoma); glaucoma Surgery degeneration, retinopathy of prematurity, retinitis pigmen failure; graft versus host disease of the eye (often a form of 55 tosa, and glaucoma. Compounds as described herein may also dry eye); herpes Zoster (shingles); hypertensive retinopathy; induce increases in tear production and density of Superficial inflammation after cataract Surgery; iridocorneal endothelial epithelial cells, two endpoints relevant to the treatment of dry syndrome; iridocytis; iritis; keratitis (e.g., infectious keratitis, eye. non-infectious keratitis, and neuroparalytic keratitis); kerato Compounds as described herein may also reduce vascular conjunctiva sicca, keratoconjunctival inflammatory disease; 60 leakage. Accordingly, these compounds may be useful in the keratoconus; keratopathy; lattice dystrophy; map-dot-finger treatment of age-related macular degeneration. print dystrophy, necrotic keratitis; neovascular diseases Compounds as described herein may also inhibit CD11b+ involving the retina, uveal tract or cornea Such as neovascular cells. Animal models of dry eye show an increase in CD11b+ glaucoma, corneal neovascularization (inflammatory, trans cells Suggesting the increased presence of leukocytes in cor plantation, developmental hypoplasia of the iris), neovascu 65 neas. Accordingly, these compounds may be useful in the larization resulting following a combined vitrectomy and treatment of dry eye by decreasing the arrival of leukocytes lensectomy, neovascularization of the optic nerve, and induced by dry eye. US 9,315,447 B2 101 102 Compounds as described herein may also prevent pig drase inhibitors such as dorZolamide HCl; prostaglandin mented retinal epithelium destruction. Accordingly, these derivatives and analogs such as tafluprost and travoprost; compounds may be useful in the treatment of age-related NMDA antagonists such as memantine, hyaluronic acid (e.g., macular degeneration, retinopathy of prematurity, retinitis Sodium hyaluronate); corticosteroids such as loteprednoleta pigmentosa, and glaucoma. bonate, difluprednate and rimexolone; antibiotics such as In certain embodiments, different compounds of the inven doxycycline; agents that increase mucin Such as ecabet and tion may be conjointly administered with other agents Suit rebamipide; lubricants such as the combination of carboxym able for the treatment or prevention of an ophthalmic condi ethylcellulose Sodium and glycerin; A3 adenosine receptor tion. For example, the following agents or classes of agents agonists such as CF-101; immunomodulators such as thali may be conjointly administered with a compound of the 10 domide, TNFC. antagonists such as etanercept; protein kinase invention: doxocycline; decosahexanoic acid; angiogenesis C-b inhibitors such as ruboxistaurin; immunosuppressants inhibitors, e.g., VEGF inhibitors, such as pegaptainib Sodium, such as sirolimus; PARP inhibitors such as AG-014699; neu bevacizumab, ranibizumab, AV-951, Vandetanib, semaxanib, roprotective thrombolytic agents such as microplasmin, CBO-P11, axitinib, Sorafenib, Sunitinib, pazopanib, and hyaluronidase; oxidizing agents such as carbamide. Soma TIMP3; anesthetics and pain killing agents such as lidocaine 15 to statin analogs such as Octreotide acetate; angiotensin II and related compounds and benzodiazepam and related com receptor antagonists such as candesartan cilexetil, disease pounds; anti-cancer agents such as 5-fluorouracil, adriamycin modifying antirheumatic drugs such as leflunomide; and related compounds; anti-inflammatory agents such as AEB071; TNF antagonists such as adalimumab: CD11 6-mannose phosphate; anti-fungal agents such as fluconazole antagonists such as efalizumab; calcineurin inhibitors such as and related compounds; anti-viral agents such as trisodium LX211; interferons such as interferon C-2a, and human alpha phosphomonoformate, trifluorothymidine, acyclovir, ganci fetoproteins such as MM-093. clovir, DDI, DDC, and AZT cell transport/mobility impend In addition to the above agents, other agents are Suitable for ing agents such as colchicine, Vincristine, cytochalasin B, and administration to the eye and its Surrounding tissues to pro related compounds; antiglaucoma drugs such as beta-block duce a local or a systemic physiologic or pharmacologic ers: timolol, betaXol, atenalol, etc.; prostaglandins such as 25 beneficial effect. Such agents may be conjointly administered latanoprost and travoprost, etc.; immunological response with a compound of the invention. Examples of Such agents modifiers such as muramyldipeptide and related compounds; include neuroprotectants, such as nimodipine and related peptides and proteins such as cyclosporin, insulin, growth compounds; antibiotics. Such as tetracycline, chlortetracy hormones, insulin related growth factor, nerve growth factor cline, bacitracin, neomycin, polymyxin, gramicidin, oxytet (optionally in further combination with decosahexanoic 30 racycline, chloramphenicol, gentamycin, and erythromycin; acid), heat shock proteins and related compounds; estrogen antibacterials, such as Sulfonamides, Sulfacetamide, Sulfame treatments; corticosteroids such as dexamethasone, dexam thizole, and sulfisoxazole; antivirals, such as, idoxuridine: ethasone 21-phosphate, fluorometholone, medrysone, other antibacterial agents, such as nitrofuraZone and sodium betamethasone, triamcinolone, triamcinolone acetonide, propionate; antiallergenics, such as antazoline, methapy triminolone, prednisone, prednisolone, prednisolone 35 riline, chlorpheniramine, pyrilamine, and prophenpy 21-phosphate, prednisolone acetate, hydrocortisone, hydro ridamine; decongestants, such as phenylephrine, naphazo cortisone acetate, prednicarbate, deflazacort, halomethasone, line, and tetrahydrazoline; miotics and anti-cholinesterase, tiXocortol, prednylidene (21-diethylaminoacetate), predni Such as pilocarpine, eserine Salicylate, carbachol, di-isopro val, paramethasone, prednisolone, methylprednisolone, pyl fluorophosphate, phospholine iodine, and demecarium meprednisone, maZipredone, isoflupredone, halopredone 40 bromide; mydriatics, such as atropine Sulfate, cyclopentolate, acetate, halcinonide, formocortal, flurandrenolide, flupred homatropine, Scopolamine, tropicamide, eucatropine, and nisolone, flurprednidine acetate, fluperolone acetate, fluocor hydroxyamphetamine; sympathomimetics, such as epineph tolone, fluocortin butyl, fluocinonide, fluocinolone, fluocino rine; and prodrugs, such as those described in Design of lone acetonide, flunisolide, flumethasone, fludrocortisone, Prodrugs, edited by Hans Bundgaard, Elsevier Scientific Pub fluclorinide, fluoromethalone, enoxolone, difluprednate, 45 lishing Co., Amsterdam, 1985. Reference may be made to any diflucortolone, diflorasone diacetate, desoximetaSone (des standard pharmaceutical textbook Such as Remington's Phar oxymethasone), desonide, descinolone, cortivaZol, corticos maceutical Sciences (Remington's Pharmaceutical Sciences. terone, cortisone, cloprednol, clocortolone, clobetaSone, clo Mack Publishing Company, Easton, Pa., USA 1985) for the betasol, chloroprednisone, cafestol, budesonide, identity of other agents. beclomethasone, amcinonide, allopregnane acetonide, 50 In certain embodiments, different compounds of the inven alclometasone, 21-acetoxypregnenolone, tralonide, diflo tion may be conjointly administered with non-chemical rasone acetate, deacylcortivazol, RU-26988, budesonide, and methods suitable for the treatment or prevention of an oph deacylcortivazol oxetanone. All of the above-cited corticos thalmic condition. In certain embodiments, different com teroids are known compounds. Further information about the pounds of the invention may be conjointly administered with compounds may be found, for example, in The Merck Index, 55 laser treatment (e.g., photocoagulation or photodynamic Thirteenth Edition (2001), and the publications cited therein, therapy), macular translocation Surgery or with devices (e.g., the entire contents of which are hereby incorporated herein by brimonidine tartrate implant). reference. In certain embodiments, the cortico steroid is In certain embodiments, different compounds of the inven selected from fluocinolone acetonide, triamcinolone tion may be conjointly administered with one or more other acetonide, dexamethasone, and related compounds, or any 60 compounds of the invention. Moreover, Such combinations combination thereof; and carbonic anhydaze inhibitors. may be conjointly administered with other therapeutic Further examples of agents or classes of agents may be agents, such as other agents suitable for the treatment or conjointly administered with a compound of the invention prevention of an ophthalmic condition, Such as the agents include: DE-104; PF-04217329; PF-03187207; AL 37807; identified above. OPC-12759; chemotherapeutic agents such as mitomycin C: 65 In certain embodiments, the present invention provides a synthetic structural analogs of prostaglandin Such as bimato kit comprising: a) one or more single dosage forms of a prost; alpha 2 agonists such as brimonidine; carbonic anhy compound of the invention; b) one or more single dosage US 9,315,447 B2 103 104 forms of an agent suitable for the treatment or prevention of In certain embodiments, the packaged pharmaceutical an ophthalmic condition or an agent suitable for administra comprises an aqueous Solution (e.g., eye drops) packaged in tion to the eye and its Surrounding tissues to produce a local or a sealed single-use container, e.g., each container comprising a systemic physiologic or pharmacologic beneficial effect as at least 1.5 nanomoles of a compound of any of formulae I-IX, mentioned above; and c) instructions for the administration of 5 a compound of formula A, a compound of any one of formu the compound of the invention and the agent suitable for the lae 1-49, a lipoxin compound, oran oxylipin compound, such treatment or prevention of an ophthalmic condition or agent as as least 2, 3, or 6 nanomoles of a compound of any of Suitable for administration to the eye and its Surrounding formulae I-IX, a compound of formula A, a compound of any tissues to produce a local or a systemic physiologic or phar one of formulae 1-49, a lipoxin compound, or an oxylipin 10 compound, in an aqueous formulation Suitable for topical macologic beneficial effect. administration to an eye. In certain such embodiments, each The present invention provides a kit comprising: container comprises from 1.5 nanomoles to 550 nanomoles, a) a pharmaceutical formulation (e.g., one or more single such as from 1.5 to 400 nanomoles, or 1.5 to 100 nanomoles dosage forms) comprising a compound of the invention; of a compound of any of formulae I-IX, a compound of and 15 formula A, a compound of any one of formulae 1-49, a lipoxin b) instructions for the administration of the pharmaceutical compound, or an oxylipin compound, in an aqueous formu formulation e.g., for treating or preventing an oph lation Suitable for topical administration to an eye. In certain thalmic condition, inhibiting COX-2 or TNF in the eye, embodiments, the packaged pharmaceutical comprises an or protecting against goblet cell loss in the eye. aqueous solution (e.g., eye drops) packaged in a sealed In certain embodiments, the kit further comprises instruc single-use container, e.g., each container comprising at least tions for the administration of the pharmaceutical formula 0.5 micrograms of compound 1001, such as at least 0.7. 1, or tion comprising a compound of the invention conjointly with 2 micrograms of compound 1001 in an aqueous formulation an agent Suitable for the treatment or prevention of an oph Suitable for topical administration to an eye. In certain Such thalmic condition, an agent suitable for administration to the embodiments, each container comprises from 0.5 micro eye and its surrounding tissues to produce a local or a sys 25 grams to 150 micrograms of compound 1001, Such as from temic physiologic orpharmacologic beneficial effect, or non 0.5 to 100 micrograms, or 0.5 to 25 micrograms of compound chemical methods suitable for the treatment or prevention of 1001 in an aqueous formulation suitable for topical adminis an ophthalmic condition as mentioned above. In certain tration to an eye. In certain embodiments, the packaged phar embodiments, the kit further comprises a second pharmaceu maceutical comprises an aqueous solution (e.g., eye drops) tical formulation (e.g., as one or more single dosage forms) 30 packaged in a sealed single-use container, e.g., each container comprising an agent Suitable for the treatment or prevention comprising a solution with a concentration over 90 micromo of an ophthalmic condition or an agent suitable for adminis lar of a compound of any of formulae I-IX, a compound of tration to the eye and its surrounding tissues to produce a local formula A, a compound of any one of formulae 1-49, a lipoxin or a systemic physiologic or pharmacologic beneficial effect compound, oran oxylipin compound. Such as a concentration as mentioned above. 35 over 100, 150, 200, or 250 micromolar of a compound of any The present invention provides a kit comprising: of formulae I-IX, a compound of formula A, a compound of a) a first pharmaceutical formulation (e.g., one or more any one of formulae 1-49, a lipoxin compound, oran oxylipin single dosage forms) comprising an agent Suitable for compound, as an aqueous formulation Suitable for topical the treatment or prevention of an ophthalmic condition administration to an eye. In certain such embodiments, each or an agent Suitable for administration to the eye and its 40 container comprises a Solution with a concentration from 90 Surrounding tissues to produce a local or a systemic micromolar to 7000 micromolar, such as from 90 micromolar physiologic or pharmacologic beneficial effect as men to 2000 micromolar, or 90 micromolar to 1000 micromolar of tioned above; and a compound of any of formulae I-IX, a compound of formula b) instructions for the administration of the first pharma A, a compound of any one of formulae 1-49, a lipoxin com ceutical formulation with a compound of the invention, 45 pound, or an oxylipin compound, as an aqueous formulation e.g., for treating or preventing an ophthalmic condition, Suitable for topical administration to an eye. For example, in inhibiting COX-2 or TNF in the eye, or protecting certain embodiments, the packaged pharmaceutical com against goblet cell loss in the eye. prises an aqueous solution (e.g., eye drops) packaged in a The present invention further provides a packaged phar sealed single-use container, e.g., each container comprising a maceutical for delivering eyedrops comprising a compound 50 Solution with a concentration over 30 ug/mL of compound of any one of formulae I-IX, a compound of formula A, a 1001, such as a concentration over 40, 50, 60, or 75 g/mL of compound of any one of formulae 1-49, a lipoxin compound, compound 1001, as an aqueous formulation Suitable for topi oran oxylipin compound to an eye in need of treatment for an cal administration to an eye. In certain such embodiments, ophthalmic condition. In certain Such embodiments, the each container comprises a solution with a concentration packaged pharmaceutical is configured to deliver the eye 55 from 30 ug/mL to 2000 g/mL, such as from 30 ug/mL to drops in any of the daily doses (e.g., any of the molar or 1000 g/mL, or 30 lug/mL to 350 g/mL of compound 1001, weight quantities) set forth above by administering one, two, as an aqueous formulation Suitable for topical administration three, four, or five eyedrops per eye, either once, twice, three to an eye. times, or four times daily, or even administering once every In further embodiments, the packaged pharmaceutical two days, Such as every other day, or once every three days, 60 comprises an aqueous preservative-free solution comprising such as every third day. One of skill in the art will recognize a compound of any one of formulae I-IX, a compound of how to vary the drop Volume (e.g., by altering the Surface formula A, a compound of any one of formulae 1-49, a lipoxin tension and/or viscosity of the Solution and/or by modifying compound, or an oxylipin compound (e.g., eye drops) pack the physical configuration of the drop-dispensing portion of aged in a container Suitable for multidose administration to an the package), the concentration of the solution, and the dos 65 eye in need of treatment for an ophthalmic condition. In age regimen (e.g., the number drops and frequency of admin certain such embodiments, the container suitable for multi istration) to provide the desired dosage. dose administration of a preservative-free solution is US 9,315,447 B2 105 106 designed such that sterility of the solution is maintained able for modulating immune function, Suppressing immune between successive uses. Suitable dispensing devices include response, treating an autoimmune disease or autoimmune those disclosed in United States Patent Application 2009/ disorder, or treating a disease, sequela or pathological condi 294347. tion mediated by an activation of the imune system. For Immune Function example, the following immunosuppressive agents may be The present invention provides a method of inhibiting conjointly administered with a compound of the invention: immune function and/or suppressing an immune response in cyclosporin, cyclosporin A, tacrolimus, rapamycin, everoli a patient, comprising administering to said patient a com mus, FK-506, cyclophosphamide, azathioprene, methotrex pound of the invention. ate, brequinar, leflunomide, mizoribine, mycophenolic acid, The present invention provides a method of treating or 10 mycophenolate mofetil, 15-deoxyspergualine, triamcinolone preventing an autoimmune disease or an autoimmune disor acetonide, decadron, daclizumab, basiliximab, glatiramer der in a patient, comprising administering to said patient a acetate, infliximab, muromonab, octreotide, muramylic acid compound of the invention. dipeptide derivatives, levamisole, niridazole, Oxysuran, In certain embodiments, the autoimmune disease or flagyl, and sirolimus autoimmune disorder is of the type where the patients own 15 In certain embodiments, different compounds of the inven immune system damages one or more of the patient’s tissues. tion may be conjointly administered with one or more other In certain embodiments, the autoimmune disease or autoim compounds of the invention. Moreover, Such combinations mune disorder may be triggered by something within the may be conjointly administered with other therapeutic patient or something within the patient's environment. agents, such as other agents Suitable for modulating immune In certain embodiments, the autoimmune disease or function, Suppressing immune response, treating or prevent autoimmune disorder of the present invention may be one ing an autoimmune disease or autoimmune disorder, or treat which follows an initiating cause. For example, the autoim ing or preventing a disease, sequela or pathological condition mune disease or autoimmune disorder may be one which is mediated by an activation of the imune system, such as the caused by an infection and/or some other initiating cause. agents identified above. Potential initiating causes may include old age, infection 25 In certain embodiments, the present invention provides a (such as parasitic infection), treatment with steroids, repeated kit comprising: a) one or more single dosage forms of a vaccination with alum, pregnancy and/or cancers. compound of the invention; b) one or more single dosage In certain embodiments, the autoimmune disease or forms of an agent Suitable for modulating immune function, autoimmune disorder may be organ-specific or non-organ Suppressing immune response, treating or preventing an specific. Examples of Such autoimmune diseases or autoim 30 autoimmune disease or autoimmune disorder, or treating or mune disorders include multiple Sclerosis, arthritis (e.g., preventing a disease, sequela or pathological condition medi rheumatoid arthritis or juvenile arthritis), Crohn's disease, ated by an activation of the imune system as mentioned colitis ulcerosa and aplastic anemia systemic lupus erythe above; and c) instructions for the administration of the com matosus (SLE or lupus), dermatomyositis, pernicious ane pound of the invention and the agent Suitable for modulating mia, Addison's disease, ankylosing spondylitis, antiphospho 35 immune function, Suppressing immune response, treating or lipid syndrome, Churg-Strauss Syndrome, discoid lupus, preventing an autoimmune disease or autoimmune disorder, fibromyalgia, Grave's Disease, myasthenia gravis, psoriasis, or treating or preventing a disease, sequela or pathological Reiter's Syndrome, rheumatic fever, sarcoidosis, sclero condition mediated by an activation of the imune system. derma, Sjogren's Syndrome, stiff-man syndrome, thyroiditis, The present invention provides a kit comprising: uveitis, vitiligo, Wegener's granulomatosis, graft rejection, 40 a) a pharmaceutical formulation (e.g., one or more single and vascular disorders. dosage forms) comprising a compound of the invention; In certain embodiments wherein the autoimmune disease and or autoimmune disorder is graft rejection, the graft rejection b) instructions for the administration of the pharmaceutical may be chronic graft rejection. In certain embodiments of the formulation, e.g., for inhibiting immune function, Sup present invention wherein a compound of the invention is 45 pressing an immune response, treating or preventing an administered for the treatment of graft rejection, the admin autoimmune disease or an autoimmune disorder, or istration of a compound of the invention modulates immune treating or preventing a disease, sequela or pathological responses to grafts (e.g., allografts or Xenografts) where condition mediated by an activation of the immune sys untreated rejection would otherwise lead to graft loss. Thus, a tem. compound of the invention may be used as a replacement for 50 In certain embodiments, the kit further comprises instruc or in addition to the conventional immunosuppressant admin tions for the administration of the pharmaceutical formula istered prior to, during and/or after transplantation. In certain tion comprising a compound of the invention conjointly with embodiments, the graft rejection is in response to transplant an agent Suitable for modulating immune function, Suppress ing natural or artificial cells, islet cells, tissues (e.g., natural or ing immune response, treating or preventing an autoimmune artificial skin tissue), corneas, bone marrow, organs (e.g. kid 55 disease or autoimmune disorder, or treating or preventing a ney, liver, pancreas, lung, or heart), lenses, or pacemakers. disease, sequela or pathological condition mediated by an The present invention further provides a method of treating activation of the imune system as mentioned above. In certain or preventing a disease, sequela or pathological condition embodiments, the kit further comprises a second pharmaceu mediated by an activation of the immune system in a patient, tical formulation (e.g., as one or more single dosage forms) comprising administering to said patient a compound of the 60 comprising an agent Suitable for modulating immune func invention. In certain embodiments, diseases, sequelae and tion, Suppressing immune response, treating or preventing an pathological conditions mediated by an activation of the autoimmune disease or autoimmune disorder, or treating or immune system include, but are not limited to, capillary leak preventing a disease, sequela or pathological condition medi age, pulmonary failure, sepsis, endotoxic shock, or sequelae ated by an activation of the imune system as mentioned above. of tissue damage. 65 The present invention provides a kit comprising: In certain embodiments, different compounds of the inven a) a first pharmaceutical formulation (e.g., one or more tion may be conjointly administered with other agents Suit single dosage forms) comprising an agent Suitable for US 9,315,447 B2 107 108 modulating immune function, Suppressing immune more single dosage forms) comprising an agent Suitable for response, treating or preventing an autoimmune disease the treatment or prevention of a pulmonary condition. or autoimmune disorder, or treating or preventing a dis In certain embodiments, the kit further comprises instruc ease, sequela or pathological condition mediated by an tions for the administration of the one or more single dosage activation of the imune system as mentioned above; and 5 forms each comprising a compound of the invention con b) instructions for the administration of the first pharma jointly with an agent suitable for the treatment or prevention ceutical formulation with a compound of the invention, of a pulmonary condition. In certain embodiments, the kit e.g., for inhibiting immune function, Suppressing an further comprises one or more single dosage forms of an immune response, treating or preventing an autoimmune agent Suitable for the treatment or prevention of a pulmonary 10 condition. disease or an autoimmune disorder, or treating or pre The present invention provides a kit comprising: venting a disease, sequela or pathological condition a) a first pharmaceutical formulation (e.g., one or more mediated by an activation of the immune system. single dosage forms) comprising an agent Suitable for Pulmonary Conditions the treatment or prevention of a pulmonary condition; The present invention provides a method of treating or 15 and preventing a pulmonary condition in a patient comprising b) instructions for the administration of the first pharma administering a compound of the invention. In certain ceutical formulation with a compound of the invention, embodiments, the pulmonary condition may be selected from e.g., for treating or preventing a condition as discussed pulmonary inflammation, airway inflammation, asthma, above, e.g., for treating or preventing a pulmonary con chronic bronchitis, bronchiectasis, non-cystic fibrosis-related dition. bronchiectasis, cystic fibrosis, eosinophilic lung diseases Gastrointestinal Conditions (e.g., parasitic infection, idiopathic eosinophilic pneumonias, The present invention provides a method of treating or and Churg-Strauss vasculitis), allergic bronchopulmonary preventing a gastrointestinal condition in a patient compris aspergillosis, allergic inflammation of the respiratory tract ing administering a compound of the invention. In certain (e.g., rhinitis and sinusitis), bronchiolitis, bronchiolitis oblit 25 embodiments, the gastrointestinal condition may be selected erans, bronchiolitis obliterans with organizing pneumonia, from gastrointestinal inflammation, ulcerative colitis, eosinophilic granuloma, Wegener's granulomatosis, sarcoi clostridium difficile colitis, microscopic or lymphocytic coli dosis, hypersensitivity pneumonitis, idiopathic pulmonary tis, collagenous colitis, Crohn's disease, irritable bowel syn fibrosis, pulmonary manifestations of connective tissue dis drome, infectious enteritis, antibiotic associative diarrhea, eases, acute or chorionic lung injury, adult respiratory distress 30 colon polyps, familial polyps, familial polyposis syndrome, syndrome, pneumonia, emphysema, pulmonary disorders, or Gardner's syndrome, helicobacter pylori, nonspecific diar chronic obstructive pulmonary disease. rheal illnesses, intestinal cancers, distal proctitis, inflamma In certain embodiments, different compounds of the inven tory states associated with abnormal colonic muscular con tion may be conjointly administered with one or more other traction (e.g., spastic colon and mucous colitis), allergic compounds of the invention. Moreover, Such combinations 35 bowel disease (e.g., coeliac disease), esophogitis, or pancre may be conjointly administered with other therapeutic atitis. agents, such as other agents Suitable for the treatment or In certain embodiments, different compounds of the inven prevention of a pulmonary condition, Such as the conditions tion may be conjointly administered with one or more other disclosed herein. Exemplary agents suitable for the treatment compounds of the invention. Moreover, Such combinations of pulmonary conditions include corticosteroids (e.g., pred 40 may be conjointly administered with other therapeutic nisone, beclomethasone dipropionate, fluticasone propi agents, such as other agents suitable for the treatment or onate, and other Suitable corticosteroids), beta-agonists (e.g., prevention of a gastrointestinal condition, such as the condi albuterol, metaproterenol, pirbuterol, formoterol, terbutaline, tions disclosed herein. Exemplary agents Suitable for the isoetharine, levalbuterol, salmeterol Xinafoate, and other suit treatment or prevention of gastrointestinal conditions include able beta-agonists), and anti-cholinergic agents (e.g., ipratro 45 immunosuppressive agents (e.g., corticosteroids), hista pium bromide, tiotropium bromide, and other suitable anti mine-2 receptor antagonists (e.g., cimetidine, famotidine, cholinergic agents). nizatidine, and ranitidine), Sucralfate, prostaglandins (e.g., In certain embodiments, the present invention provides a misopostol), and proton pump inhibitors (e.g., omeprazole, kit comprising: a) one or more single dosage forms of a lansoprazole, esomeprazole, pantoprazole, and rabeprazole). compound of the invention; b) one or more single dosage 50 In certain embodiments, the present invention provides a forms of an agent Suitable for the treatment or prevention of a kit comprising: a) one or more single dosage forms of a pulmonary condition; and c) instructions for the administra compound of the invention; b) one or more single dosage tion of the compound of the invention and the agent Suitable forms of an agent Suitable for the treatment or prevention of a for the treatment or prevention of a pulmonary condition. gastrointestinal condition; and c) instructions for the admin The present invention provides a kit comprising: 55 istration of the compound of the invention and the agent a) a pharmaceutical formulation (e.g., one or more single Suitable for the treatment or prevention of a gastrointestinal dosage forms) comprising a compound of the invention; condition. and The present invention provides a kit comprising: b) instructions for the administration of the pharmaceutical a) a pharmaceutical formulation (e.g., one or more single formulation, e.g., for treating or preventing a condition 60 dosage forms) comprising a compound of the invention; as discussed above, e.g., a pulmonary condition. and In certain embodiments, the kit further comprises instruc b) instructions for the administration of the pharmaceutical tions for the administration of the pharmaceutical formula formulation, e.g., for treating or preventing a condition tion comprising a compound of the invention conjointly with as discussed above, e.g., a gastrointestinal condition. an agent Suitable for the treatment or prevention of a pulmo 65 In certain embodiments, the kit further comprises instruc nary condition. In certain embodiments, the kit further com tions for the administration of the pharmaceutical formula prises a second pharmaceutical formulation (e.g., as one or tion comprising a compound of the invention conjointly with US 9,315,447 B2 109 110 an agent Suitable for the treatment or prevention of a gas The present invention provides a kit comprising: trointestinal condition. In certain embodiments, the kit fur a) a pharmaceutical formulation (e.g., one or more single ther comprises a second pharmaceutical formulation (e.g., as dosage forms) comprising a compound of the invention; one or more single dosage forms) comprising an agent Suit and able for the treatment or prevention of a gastrointestinal con b) instructions for the administration of the pharmaceutical dition. formulation, e.g., for treating or preventing a condition The present invention provides a kit comprising: as discussed above, e.g., a rheumatological condition. a) a first pharmaceutical formulation (e.g., one or more In certain embodiments, the kit further comprises instruc single dosage forms) comprising an agent Suitable for tions for the administration of the pharmaceutical formula the treatment or prevention of a gastrointestinal condi 10 tion comprising a compound of the invention conjointly with tion; and b) instructions for the administration of the first pharma an agent Suitable for the treatment or prevention of a rheuma ceutical formulation with a compound of the invention, tological condition. In certain embodiments, the kit further e.g., for treating or preventing a condition as discussed comprises a second pharmaceutical formulation (e.g., as one above, e.g., a gastrointestinal condition. 15 or more single dosage forms) comprising an agent Suitable for Rheumatological Conditions the treatment or prevention of a rheumatological condition. The present invention provides a method of treating or The present invention provides a kit comprising: preventing a rheumatological condition in a patient compris a) a first pharmaceutical formulation (e.g., one or more ing administering a compound of the invention. In certain single dosage forms) comprising an agent Suitable for embodiments, the rheumatological condition may be selected the treatment or prevention of a rheumatological condi from rheumatic diseases, rheumatoid arthritis, osteoarthritis, tion; and inflammatory conditions of joints (e.g., rheumatoid arthritis, b) instructions for the administration of the first pharma rheumatoid spondylitis, osteoarthritis, gouty arthritis), anky ceutical formulation with a compound of the invention, losing spondylitis, lupus, psoriatic arthritis, myalgias, or e.g., for treating or preventing a condition as discussed chronic low back pain. 25 above, e.g., a rheumatological condition. In certain embodiments, different compounds of the inven Dermatological Conditions tion may be conjointly administered with one or more other The present invention provides a method of treating or compounds of the invention. Moreover, Such combinations preventing a dermatological condition inapatient comprising may be conjointly administered with other therapeutic administering a compound of the invention. In certain agents, such as other agents Suitable for the treatment or 30 embodiments, the dermatological condition may be selected prevention of a rheumatological condition, such as the con from Scleroderma, psoriasis, urticaria, Vasculitis, seborrheic ditions disclosed herein. Exemplary agents suitable for the dermatitis, pustular dermatosis, eczema, dermatitis, ulcers treatment or prevention of rheumatological conditions and erosions resulting from trauma, burns, bullous disorders, include non-steroidal anti-inflammatory drugs such as Sali or ischemia of the skin or mucous membranes, ichthyoses, cylates (e.g., aspirin, amoxiprin, benorilate, choline magne 35 epidermolysis bullosae, hypertrophic scars and keloids, cuta sium salicylate, diflunisal, faislamine, methyl salicylate, neous changes of intrinsic aging and photoaging, frictional magnesium salicylate, Salicyl salicylate), arylalkanoic acids blistering caused by mechanical shearing of the skin, cutane (e.g., diclofenac, aceclofenac, acemetacin, bromfenac, etod ous atrophy resulting from the topical use of corticosteroids, olac, indometacin, nabumetone, Sulindac, and tolmetin), inflammation to mucous membranes (e.g., cheilitis, chapped 2-arylpropionic acids (e.g., ibuprofen, carprofen, fenbufen, 40 lips, nasal irritation, or Vulvovaginitis), dandruff. Sunburn, fenoprofen, flurbiprofen, ketoprofen, ketorolac, loxoprofen, poison ivy, atopic dermatitis, acne, or rosacea. naproxen, Oxaprozin, tiaprofenic acid, and Suprofen), N-ary In certain embodiments, different compounds of the inven lanthranilic acids (e.g., mefenamic acid and meclofenamic tion may be conjointly administered with one or more other acid), pyrazolidine derivatives (e.g., phenylbutaZone, aza compounds of the invention. Moreover, Such combinations propaZone, metamizole, oxyphenbutaZone, and Sulfinpyra 45 may be conjointly administered with other therapeutic Zone), oxicams (e.g., piroXicam, lornoxicam, meloxicam, and agents, such as other agents suitable for the treatment or tenoxicam), COX-2 inhibitors (e.g., celecoxib, etoricoxib, prevention of a dermatological condition, Such as the condi lumiracoxib, parecoxib, rofecoxib, and Valdecoxib) and Sul tions disclosed herein. Exemplary agents Suitable for the phonanilides (e.g., nimeSulide), corticosteroids (e.g., pred treatment or prevention of dermatological conditions include nisone, cortisone, Solumedrol, and hydrocortisone), disease 50 immunosuppressive agents, such as cyclosporine or cal modifying anti-rheumatic drugs (e.g., leflunomide, oral gold, cineurin inhibitors (e.g., tacrolimus or pimecrolimus), corti Sulfasalazine, mycophenolate, cyclophosphamide, azathio costeroids (e.g., prednisone and betamethasone dipropi prine, chlorambucil, rheumatrex, minocycline, gold shots, onate), antihistamines (e.g., diphenhydramine, hydroxy Zine, cuprimine, and quineprox), pain medication (e.g., acetami and cyproheptadine), salicyclic acid, anthroline, calcipot nophen, codeine, propoxyphene, fentanyl, hydromorphone, 55 riene, and tazarotene. morphine, oxycodone, pentazocine, tramadol, and hydroc In certain embodiments, the present invention provides a odone) and biologic response modifiers (e.g., etanercept, kit comprising: a) one or more single dosage forms of a adalimumab, anakinra, abatacept, efalizumab, infliximab.l compound of the invention; b) one or more single dosage rituximab, and natalizumab) forms of an agent Suitable for the treatment or prevention of a In certain embodiments, the present invention provides a 60 dermatological condition; and c) instructions for the admin kit comprising: a) one or more single dosage forms of a istration of the compound of the invention and the agent compound of the invention; b) one or more single dosage suitable for the treatment or prevention of a dermatological forms of an agent Suitable for the treatment or prevention of a condition. rheumatological condition; and c) instructions for the admin The present invention provides a kit comprising: istration of the compound of the invention and the agent 65 a) a pharmaceutical formulation (e.g., one or more single suitable for the treatment or prevention of a rheumatological dosage forms) comprising a compound of the invention; condition. and US 9,315,447 B2 111 112 b) instructions for the administration of the pharmaceutical golide, pramipexole, ropinirole, cabergoline, apomorphine, formulation, e.g., for treating or preventing a condition and lisuride: MAO-B inhibitors, such as selegiline and rasa as discussed above, e.g., a dermatological condition. giline, or combinations thereof. In certain embodiments, the kit further comprises instruc In certain embodiments, the present invention provides a tions for the administration of the pharmaceutical formula kit comprising: a) one or more single dosage forms of a tion comprising a compound of the invention conjointly with compound of the invention; b) one or more single dosage an agent Suitable for the treatment or prevention of a derma forms of an agent Suitable for the treatment or prevention of a tological condition. In certain embodiments, the kit further neurological condition; and c) instructions for the adminis comprises a second pharmaceutical formulation (e.g., as one tration of the compound of the invention and the agent Suit or more single dosage forms) comprising an agent Suitable for 10 able for the treatment or prevention of a neurological condi the treatment or prevention of a dermatological condition. tion. The present invention provides a kit comprising: The present invention provides a kit comprising: a) a first pharmaceutical formulation (e.g., one or more a) a pharmaceutical formulation (e.g., one or more single single dosage forms) comprising an agent Suitable for dosage forms) comprising a compound of the invention; the treatment or prevention of a dermatological condi 15 and tion; and b) instructions for the administration of the pharmaceutical b) instructions for the administration of the first pharma formulation, e.g., for treating a condition as discussed ceutical formulation with a compound of the invention, above, e.g., a neurological condition. e.g., for treating or preventing a condition as discussed In certain embodiments, the kit further comprises instruc above, e.g., a dermatological condition. tions for the administration of the pharmaceutical formula Neurological Conditions tion comprising a compound of the invention conjointly with The present invention provides a method of treating or an agent Suitable for the treatment or prevention of a neuro preventing a neurological condition in a patient comprising logical condition. In certain embodiments, the kit further administering a compound of the invention. In certain comprises a second pharmaceutical formulation (e.g., as one embodiments, the neurological condition may be selected 25 or more single dosage forms) comprising an agent Suitable for from neurodegeneration or dementia associated with HIV the treatment or prevention of a neurological condition. infection, depression, Alzheimer's disease, Parkinson's dis The present invention provides a kit comprising: ease, addiction, alcohol-related disorders, decision analysis, a) a first pharmaceutical formulation (e.g., one or more degenerative neurological disorders, dementia, neurological single dosage forms) comprising an agent Suitable for disorders, neuromuscular disorders, psychiatric disorders, 30 the treatment or prevention of a neurological condition; brain injury, trauma, neuronal inflammation, or multiple scle and rosis. b) instructions for the administration of the first pharma In certain embodiments, different compounds of the inven ceutical formulation with a compound of the invention, tion may be conjointly administered with one or more other e.g., for treating or preventing a condition as discussed compounds of the invention. Moreover, Such combinations 35 above, e.g., a neurological condition. may be conjointly administered with other therapeutic Cancer agents, such as other agents Suitable for the treatment or The present invention provides a method of treating or prevention of a neurological condition, such as the conditions preventing cancer in a patient comprising administering a disclosed herein. Exemplary agents suitable for the treatment compound of the invention. In certain embodiments, the can of neurological conditions include thyroid hormone replace 40 cer may be selected from breast cancer, colon cancer, leuke ment, cholinesterase inhibitors (e.g., donepezil and tacrine), mia, lymphoma, lung cancer, or prostate cancer. antipsychotic drugs (e.g., clozapine, olanzapine, quetiapine, In certain embodiments, different compounds of the inven risperidone, Ziprasidone, aripiprazole, trifluoperazine, flu tion may be conjointly administered with one or more other penthixol, loxapine, perphenazine, chlorpromazine, halo compounds of the invention. Moreover, Such combinations peridol, fluiphenazine decanoate, and thioridazine), anxiolyt 45 may be conjointly administered with other therapeutic ics, such as benzodiazepines (e.g., lorazepam, clonazepam, agents, such as other agents suitable for the treatment or alprazolam, and diazepam) and non-benzodiazepines (e.g., prevention of cancer, such as the chemotherapeutic agents or buspirone), drugs for the treatment of Alzheimer's disease, the combination therapies as disclosed above. Such as acetylcholinesterase inhibitors (e.g., donepezil, In certain embodiments, a compound of the invention may rivastigmine, and galantamine) or NMDA receptor antago 50 be conjointly administered with non-chemical methods of nists (e.g., memantine), antidepressants, such as selective cancer treatment or prevention. In certain embodiments, a serotonin reuptake inhibitors (e.g., fluoxetine, paroxetine, compound of the invention may be conjointly administered escitalopram, citalopram, and Sertraline), serotonin and nore with radiation therapy. In certain embodiments, a compound pinephrine reuptake inhibitors (e.g., Venlafaxine and dulox of the invention may be conjointly administered with Surgery, etine), noradrenergic and specific serotonergic antidepres 55 with thermoablation, with focused ultrasound therapy, or sants (e.g., mirtazapine), norepinephrine reuptake inhibitors with cryotherapy. (e.g., reboxetine), norepinephrine and dopamine reuptake The present invention provides a kit comprising: inhibitors (e.g., bupropion), tricyclic antidepressants (e.g., a) a pharmaceutical formulation (e.g., one or more single amitriptyline and desipramine), and monoamine oxidase dosage forms) comprising a compound of the invention; inhibitors (e.g., phenelzine, moclobemide, and selegiline), 60 and drugs for the treatment of multiple Sclerosis, Such as interfer b) instructions for the administration of the pharmaceutical ons (e.g., interferon beta-1a and interferon beta-1b), glati formulation, e.g., for treating or preventing cancer. ramer acetate, mitoxantrone, natalizumab, and corticoster In certain embodiments, the kit further comprises instruc oids (e.g., methylprednisolone or any of the corticosteroids tions for the administration of the pharmaceutical formula referenced above), and drugs for the treatment of Parkinson's 65 tion comprising a compound of the invention conjointly with disease. Such as levodopa, carbidopa, benserazide, tolcapone, a chemotherapeutic agent, as described above. In certain entacapone; dopamine agonists, such as bromocriptine, per embodiments, the kit further comprises a second pharmaceu US 9,315,447 B2 113 114 tical formulation (e.g., as one or more single dosage forms) The present invention provides a kit comprising: comprising a chemotherapeutic agent, as described above. a) a pharmaceutical formulation (e.g., one or more single The present invention provides a kit comprising: dosage forms) comprising a compound of the invention; a) a first pharmaceutical formulation (e.g., one or more and single dosage forms) comprising a chemotherapeutic b) instructions for the administration of the pharmaceutical formulation, e.g., for treating or preventing a condition agent; and b) instructions for the administration of the as discussed above, e.g., an infectious condition. first pharmaceutical formulation with a compound of the In certain embodiments, the kit further comprises instruc invention, e.g., for treating or preventing cancer. tions for the administration of the pharmaceutical formula Infectious Conditions tion comprising a compound of the invention conjointly with The present invention provides a method of treating or 10 an agent Suitable for the treatment or prevention of an infec preventing an infectious condition in a patient comprising tious condition. In certain embodiments, the kit further com administering a compound of the invention. In certain prises a second pharmaceutical formulation (e.g., as one or embodiments, the infectious condition may be selected from more single dosage forms) comprising an agent Suitable for the treatment or prevention of an infectious condition. a bacterial infection, parasitic diseases, a gram negative bac 15 The present invention provides a kit comprising: terial infection, salmonella typhimurium infection, an oral a) a first pharmaceutical formulation (e.g., one or more infection, a fungal infection, or a viral infection. single dosage forms) comprising an agent Suitable for The present invention further provides a method of treating the treatment or prevention of an infectious condition; or preventing inflammation associated with infection in a and patient comprising administering a compound of the inven b) instructions for the administration of the first pharma tion. In certain Such embodiments, compounds of the inven ceutical formulation with a compound of the invention, tion may stimulate and increase production of anti-microbial e.g., for treating or preventing a condition as discussed peptides expressed by human epithelial cells (e.g., bacteri above, e.g., an infectious condition. cidal permeability increasing protein). Apoptotic Conditions In certain embodiments, different compounds of the inven 25 The present invention provides a method of inhibitingapo tion may be conjointly administered with one or more other ptosis in a patient comprising administering a compound of compounds of the invention. Moreover, Such combinations the invention. The present invention further provides a may be conjointly administered with other therapeutic method of treating or preventing a condition arising from agents, such as other agents Suitable for the treatment or apoptosis in a patient comprising administering a compound prevention of an infectious condition, such as the conditions 30 of the invention. In certain embodiments, the condition aris disclosed herein. Exemplary agents suitable for the treatment ing from apoptosis may be selected from coronary infarct or prevention of infectious conditions include aminoglyco damage, tissue necrosis in chronic inflammation, smooth sides (e.g., amikacin, gentamicin, kanamycin, neomycin, muscle proliferation disorders (e.g., restenosis following netilmicin, Streptomycin, tobramycin, and paromomycin), angioplasty), inflammatory states associated with arterial ansamycins (e.g., geldanamycin and herbimycin), carba 35 Smooth muscle constriction (e.g., coronary spasm, ischemia cephem (e.g., loracarbef), carbapenems (e.g., ertapenem, induced myocardial injury, cerebral spasm, or cerebral doripenem, imipenem, and meropenem), cephalosporins ischemia and related disorders, such as stroke), conditions (e.g., cefadroxil, cefazolin, cefalotin, cephalexin, cefaclor, associated with thrombosis (e.g., coronary thrombosis, phle cefamandole, cefoxitin, cefprozil, cefuroxime, cefixime, cef bitis, or phlebothrombosis), and neurodegenerative diseases. dinir, cefditoren, cefoperaZone, cefotaxime, cefpodoxime, 40 In certain embodiments, the present invention provides a ceftazidime, ceftibuten, ceftizoxime, ceftriaxone, cefdinir, kit comprising: a) one or more single dosage forms of a and cefepime), glycopeptides (e.g., Vancomycin), macrollides compound of the invention; b) one or more single dosage (e.g., azithromycin, clarithromycin, dirithromycin, erythro forms of an agent Suitable for the treatment or prevention of a mycin, roXithromycin, troleandomycin, tellithromycin, and condition arising from apoptosis; and c) instructions for the spectinomycin), monobactams (e.g., aztreonam), penicillins 45 administration of the compound of the invention and the (e.g., amoxicillin, amplicillin, azlocillin, carbenicillin, cloX agent Suitable for inhibiting apoptosis or for the treatment or acillin, dicloxacillin, flucloxacillin, mezlocillin, nafcillin, prevention of a condition arising from apoptosis. penicillin, piperacillin, and ticarcillin), polypeptides (e.g., The present invention provides a kit comprising: bacitracin, colistin, and polymyxin B), quinolones (e.g., a) a pharmaceutical formulation (e.g., one or more single ciprofloxacin, enoxacin, gatifloxacin, levofloxacin, lom 50 dosage forms) comprising a compound of the invention; efloxacin, moxifloxacin, norfloxacin, ofloxacin, and trova and floxacin), Sulfonamides (e.g., mafenide, prontosil, Sulfaceta b) instructions for the administration of the pharmaceutical mide, Sulfamethizole, Sulfanilimide, Sulfasalazine, formulation, e.g., for inhibiting apoptosis or treating or Sulfisoxazole, and trimethoprim), tetracyclines (e.g., deme preventing a condition arising from apoptosis. clocycline, doxycycline, minocycline, Oxytetracycline, and 55 In certain embodiments, the kit further comprises instruc tetracycline), arsphenamine, chloramphenicol, clindamycin, tions for the administration of the pharmaceutical formula lincoamycin, ethambutol, fosfomycin, fusidic acid, furazoli tion comprising a compound of the invention conjointly with done, isoniazid, lineZolid, metronidazole, mupirocin, nitro an agent Suitable for inhibiting apoptosis or for the treatment furantoin, platensimycin, pyrazinamide, quinuprisin, dalfo or prevention of a condition arising from apoptosis. In certain pristin, rifampin, and timidazole. 60 embodiments, the kit further comprises a second pharmaceu In certain embodiments, the present invention provides a tical formulation (e.g., as one or more single dosage forms) kit comprising: a) one or more single dosage forms of a comprising an agent Suitable for inhibiting apoptosis or for compound of the invention; b) one or more single dosage the treatment or prevention of a condition arising from apo forms of an agent suitable for the treatment or prevention of ptosis. an infectious condition; and c) instructions for the adminis 65 The present invention provides a kit comprising: tration of the compound of the invention and the agent Suit a) a first pharmaceutical formulation (e.g., one or more able for the treatment or prevention of an infectious condition. single dosage forms) comprising an agent Suitable for US 9,315,447 B2 115 116 inhibitingapoptosis or for the treatment or prevention of e.g., for inhibiting angiogenesis or for treating or pre a condition arising from apoptosis; and venting a condition arising from angiogenesis. b) instructions for the administration of the first pharma When administered alone or as part of a therapeutic regi ceutical formulation with a compound of the invention, men, in certain embodiments, the invention contemplates e.g., for inhibitingapoptosis or for treating or preventing administration of compounds of the present invention to treat a condition arising from apoptosis. or prevent a particular primary disease, injury, disorder, or Angiogenesis condition. In certain embodiments, the invention contem The present invention provides a method of inhibiting plates administration of compounds of the present invention angiogenesis in a patient comprising administering a com to treat or prevent symptoms secondary to the primary dis 10 ease, injury, disorder, or conditions. pound of the invention. In certain embodiments, compounds Organ Preservation of the present invention limitangiogenesis necessary for Solid The present invention provides methods for reducing, pre tumor metastasis. Since angiogenesis and neovascularization Venting or reversing organ damage or enhancing organ pres are essential steps in Solid tumor growth, inhibition of angio ervation and/or Survival comprising administering a com genesis is very useful to prevent the further growth, retard, or 15 pound of the invention to an organ donor patient prior to even regress Solid tumors. Exemplary neoplasias which may removal of the organ. The present invention provides methods be treated with compounds of the present invention include, for reducing or preventing stem cell damage and/or death or but are not limited to, gastrointestinal tumors and gliomas. enhancing stem cell Survival and/or preservation comprising In certain embodiments, additional disorders or diseases administering a compound of the invention to stem cell donor that may be treated or prevented by inhibition of angiogenesis patient prior to removal of the stem cells. In certain embodi (e.g., by administering a compound of the present invention) ments, the compound of the invention is administered to the include, but are not limited to, retinopathy associated with organ and/or stem cell donor patient less than 24 hours prior diabetes, rheumatoid arthritis, osteoarthritis, macular degen to removal of the organ, Such as less than 12, eight, six, four eration, glaucoma, Keloid formation, ulcerative colitis, or two hours prior to removal of the organ and/or stem cells. Krohn's disease, and psoriasis. 25 In certain embodiments, the compound of the invention is In certain embodiments, different compounds of the inven administered to the organ and/or stem cell donor patient tion may be conjointly administered with one or more other immediately prior to removal of the organ and/or stem cells compounds of the invention. Moreover, Such combinations (e.g., less than one hour prior to removal of the organ and/or may be conjointly administered with other therapeutic stem cells, such as less than 30, 15, or 10 minutes prior to agents, such as other agents suitable for inhibiting angiogen 30 removal of the organ and/or stem cells). In certain embodi esis or for the treatment or prevention of a condition arising ments, the organ and/or stem cell donor patient is a human. from angiogenesis, such as angiogenesis inhibitors including, In certain embodiments, the organ and/or stem cell donor but not limited to, siRNAs, aptamers, angiostatin, endosta patient is characterized by brain death. In certain embodi tin, and bevacizumab. ments, “brain death’ is defined as the total cessation of brain In certain embodiments, the present invention provides a 35 function, including brain stem function, e.g., wherein there is kit comprising: a) one or more single dosage forms of a no oxygen or blood flow to the brain, or wherein the brain no compound of the invention; b) one or more single dosage longer functions in any manner and will never function again. forms of an agent Suitable for inhibiting angiogenesis or for In certain embodiments, the organ and/or stem cell donor the treatment or prevention of a conditionarising from angio patient is not diagnosed as having a chronic, transmissible, or genesis; and c) instructions for the administration of the com 40 infectious physical ailment, e.g., for which pharmacological pound of the invention and the agent Suitable for inhibiting intervention is or would have been suitable. In certain angiogenesis or for the treatment or prevention of a condition embodiments, the organ and/or stem cell donor patient is not arising from angiogenesis. currently and/or has not been diagnosed with diabetes, can The present invention provides a kit comprising: cer, high blood pressure, kidney disease, or cardiovascular a) a pharmaceutical formulation (e.g., one or more single 45 disease, e.g., atherosclerosis or heart disease. dosage forms) comprising a compound of the invention; In certain embodiments, the method for reducing, prevent and ing or reversing organ damage or enhancing organ preserva b) instructions for the administration of the pharmaceutical tion and/or Survival comprises administering a compound of formulation, e.g., for inhibiting angiogenesis or treating the invention to an organ donor patient prior to removal of the or preventing a condition arising from angiogenesis. 50 organ further comprises the step of removing the organ from In certain embodiments, the kit further comprises instruc the organ donor patient. In certain such embodiments, the tions for the administration of the pharmaceutical formula organis selected from one or more of a kidney, a liver or a lobe tion comprising a compound of the invention conjointly with of a liver, a lung or part of a lung, a portion of pancreas, a an agent Suitable for inhibiting angiogenesis or for the treat portion of intestine, a heart, a cornea or tissue (e.g., skin, ment or prevention of a condition arising from angiogenesis. 55 blood, bone marrow, blood stem cells, or umbilical cord In certain embodiments, the kit further comprises a second blood). pharmaceutical formulation (e.g., as one or more single dos In certain embodiments, the method for reducing or pre age forms) comprising an agent Suitable for inhibiting angio venting stem cell damage and/or death or enhancing stem cell genesis or for the treatment or prevention of a condition Survival and/or preservation comprises administering a com arising from angiogenesis. 60 pound of the invention to a stem cell donor patient prior to The present invention provides a kit comprising: removal of the stem cells further comprises the step of remov a) a first pharmaceutical formulation (e.g., one or more ing the stem cells from the stem cell donor patient. single dosage forms) comprising an agent Suitable for In certain embodiments, the organ and/or stem cell donor inhibiting angiogenesis or for the treatment or preven patient is any suitable organ and/or stem cell donor patient. In tion of a condition arising from angiogenesis; and 65 certain embodiments, the organ and/or stem cell donor patient b) instructions for the administration of the first pharma is a non-human animal. For example, the organ and/or stem ceutical formulation with a compound of the invention, cell donor patient may be a pig or primate, such as a geneti US 9,315,447 B2 117 118 cally altered animal. In certain Such embodiments, the organ The present invention further provides methods for reduc and/or stem cell donor patient is an animal that has been ing or preventing stem cell damage and/or death or enhancing genetically modified Such that proteins on the Surface of the stem cell preservation and/or Survival comprising contacting animals organs and/or cells are recognised as compatible by the stem cells with a compound of the invention. a human immune system. For example, the organ and/or stem In certain embodiments, the stem cells are contacted with a cell donor patient may be an animal that has been genetically compound of the invention ex vivo (e.g., during a period of ex modified such that proteins on the surface of the animals Vivo culture and/or manipulation, for example ex vivo culture organs and/or cells are recognised as human by the human and/or manipulation for cell expansion and/or differentiation, immune system, so the organs and/or cells are not attacked during the process of cryopreservation of the stem cells, dur when transplanted. In certain embodiments, the organ donor 10 ing the process of thawing cryopreserved stem cells, or any patient is a pig. combination thereof). In certain Such embodiments, the com The present invention provides methods for reducing, pre pound of the invention is present as a component of a Suitable Venting or reversing organ damage or enhancing organ pres culture medium (e.g., any culture medium Suitable forex vivo ervation and/or Survival comprising administering a com culture and/or manipulation, cryopreservation of stem cells, pound of the invention to an organ recipient prior to organ 15 or the thawing of cryopreserved stem cells). transplantation. In certain embodiments, the stem cells are contacted with a In certain embodiments, the method for reducing, preventing compound of the invention while the stem cells are still in a or reversing organ damage or enhancing organ preservation subject’s body, during the removal of the stem cells from a and/or Survival comprising administering a compound of the subject’s body, after the stem cells are removed from a sub invention to an organ recipient prior to organ transplantation 20 ject’s body, during the process of ex vivo culture and/or further comprises the step of removing one or more organs manipulation (e.g., for expansion and/or differentiation) dur from the organ recipient. In certain Such embodiments, the ing the process of cryopreservation of the stem cells, during compound of the invention is administered to the organ the process of thawing cryopreserved stem cells, while the recipient at any point during the organ removal process. In stem cells are being transplanted into a recipient, immediately certain Such embodiments, the step of removing the one or 25 after the stem cells are transplanted into a recipient, or any more organs from the organ recipient occurs prior to admin combination thereof. istering a compound of the invention to the organ recipient. In The present invention further provides methods for reduc certain embodiments, the step of removing the one or more ing, preventing or reversing organ damage or enhancing organs from the organ recipient occurs simultaneously to organ preservation and/or Survival comprising contacting the administering a compound of the invention to the organ 30 organ with a preservation Solution wherein the preservation recipient. In certain embodiments, the step of removing the Solution comprises a compound of the invention. one or more organs from the organ recipient occurs subse In certain embodiments, the preservation solution com quent to administering a compound of the invention to the prises a compound of the invention in an amount Sufficient for organ recipient. reducing, preventing or reversing organ damage or enhancing In certain embodiments, the method for reducing, preventing 35 organ preservation and/or Survival. In certain embodiments, or reversing organ damage or enhancing organ preservation the preservation solution comprises a compound of the inven and/or Survival comprising administering a compound of the tion at a concentration of 1 nM to 1 M, e.g., from 1 uM to 1 invention to an organ recipient prior to organ transplantation mM. In certain embodiments, the organ preservation Solution further comprises the step of transplanting one or more organs further comprises potassium, Sodium, magnesium, calcium, into the organ recipient. 40 phosphate, Sulphate, glucose, citrate, mannitol, histidine, The present invention further provides methods for reduc tryptophan, alpha-ketoglutaric acid, lactobionate, raffinose, ing or preventing stem cell damage and/or death or enhancing adenosine, allopurinol, glutathione, glutamate, insulin, dex stem cell preservation and/or Survival comprising administer amethasone, hydroxyethyl starch, bactrim, trehalose, glucon ing a compound of the invention to a stem cell recipient prior ate, or combinations thereof. In certain embodiments, the to stem cell transplantation. 45 organ preservation solution comprises sodium, potassium, In certain embodiments, the method for reducing or pre magnesium, or combinations thereof. In certain embodi venting stem cell damage and/or death or enhancing stem cell ments, the organ preservation solution is free or Substantially preservation and/or Survival comprising administering a free of cells, coagulation factors, nucleic acids such as DNA, compound of the invention to a stem cell recipient prior to and/or plasma proteins. In certain embodiments, the organ stem cell transplantation further comprises the step of trans 50 preservation solution is sterile. In certain embodiments, the planting stem cells into the stem cell recipient. organ preservation Solution comprises an aqueous solution. The present invention further provides methods for reduc In certain embodiments, the organ preservation Solution com ing, preventing or reversing organ damage or enhancing prises a perfluorocarbon, such as a perfluorohydrocarbon or organ preservation and/or Survival comprising contacting the a perfluoroalkylamine. Exemplary perfluorocarbons are organ with a compound of the invention. 55 described in Transplantation, 74(12), 1804-1809, Dec. 27. In certain embodiments, the organis contacted ex vivo with 2002 and Am. Assoc. of Nurse Anesthetists Journal, 74(3): a compound of the invention. In certain embodiments, the 205-211, June 2007, the compounds in which are incorpo organ is contacted with a compound of the invention in a rated herein by reference. manner other than directly through the organ's blood Supply In certain embodiments wherein the method of the present (e.g., the organis contacted with a compound of the invention 60 invention comprises reducing, preventing or reversing organ outside of its circulatory system). In certain embodiments, the damage or enhancing organ preservation and/or Survival organis contacted with a compound of the invention while the comprising contacting the organ with a preservation Solution organ is still in a Subject’s body, during the removal of the wherein the preservation solution comprises a compound of organ from a Subject's body, after the organ is removed from the invention, the preservation solution may be any Suitable a subject’s body, while the organ is being transplanted into a 65 preservation solution known in the art. Examples of Such recipient, immediately after the organ is transplanted into a preservation solutions include, but are not limited to, Univer recipient, or any combination thereof. sity of Wisconsin solution, Krebs-Henseleit solution, Celsior US 9,315,447 B2 119 120 Solution, St. Thomas Hospital 2 solution, Ringer-lactate solu The present invention provides a method of promoting tion, Collins solution, Euro-Collins solution, Stanford solu Survival of a stem cell transplant recipient, comprising tion, Ross-Marshall citrate solution, phosphate-buffered administering a compound of the invention to a stem cell sucrose solution, Kyoto ET solution, or Bretschneider histi donor patient prior to removal of the stem cells; dine tryptophan ketoglutarate (HTK) solution. administering a compound of the invention to a stem cell The organ may be contacted with (or administered) the recipient prior to stem cell transplantation; preservation solution comprising the compound of the inven contacting the stem cells ex vivo (e.g., in a suitable culture tion at any point during the transplantation process. For medium) with a compound of the invention; example, the preservation solution comprising a compound or any combination thereof. of the invention may be administered by flushing the organ, 10 continuously perfusing the organ, or intermittently perfusing The present invention provides a method of facilitating an through the blood vessels of the organ while the organ is still organ transplant procedure and/or enhancing the Success of in a subjects body, during the removal of the organ from a an organ transplant procedure, comprising subjects body, after the organ is removed from a subjects administering a compound of the invention to an organ body, while the organ is being transplanted into a recipient, 15 donor patient prior to removal of the organ; immediately after the organistransplanted into a recipient, or administering a compound of the invention to an organ any combination thereof. In certain embodiments, the organ recipient prior to organ transplantation; preservation solution comprising the compound of the inven contacting the organ ex vivo with a compound of the inven tion is administered directly into the organ's blood Supply tion; while the organ is being blood-perfused by a cardiovascular contacting the organ ex vivo with a preservation solution system, which can be within the body of the organ donor or wherein the preservation Solution comprises a com organ recipient. pound of the invention; In certain embodiments, the organ may be any organ Suit or any combination thereof. able for transplatation, such as a kidney, liver or lobe of aliver, The present invention provides a method of facilitating a heart, lung or part of a lung, skin, intestine or portion of an 25 stem cell transplant procedure and/or enhancing the Success intestine, cornea, pancreas or portion of a pancreas, tissue of a stem cell transplant procedure, comprising (e.g., blood, bone marrow, blood stem cells, or umbilical cord administering a compound of the invention to a stem cell blood), or any combination thereof. donor patient prior to removal of the stem cells; In certain embodiments of methods of the invention, the stem administering a compound of the invention to a stem cell cell is selected from adult stem cells or embryonic stem cells. 30 recipient prior to stem cell transplantation; Exemplary stem cells include, but are not limited to, totipo contacting the stem cells ex vivo (e.g., in vitro in a Suitable tent stem cells, pluripotent stem cells, multipotent stem cells, culture medium, such as during the process of cyro unipotent stem cells, hematopoietic stem cells, adipose-de preservation and/or thawing of cyropreserved stem cells rived stem cells, endothelial stem cells, muscle stem cells, or during ex vivo culture and/or manipulation) with a bone marrow stromal cells (e.g., mesenchymal stem cells), 35 compound of the invention; neural stem cells, skin stem cells, and follicular stem cells. or any combination thereof. Embryonic stem cells include embryonic stem cells made The Success of an organ and/or a stem cell transplant pro using Somatic cell nuclear transfer, as well as embryonic stem cedure may be evaluated, for example, by the reduction of cells derived from the inner cell mass of embryos produced by side effects and/or symptoms associated with the transplan fertilization. Suitable stem cells also include induced pluri 40 tation procedure, by a reduction in hospitalization time fol potent stem cells, regardles of whether the induced pluripo lowing the organ and/or stem cell transplant procedure, by a tent stem cells are produced using integrative or non-integra reduction in the time between organ and/or stem cell trans tive vectors to express one or more reprogramming factors, plantation and resumption of normal bodily functions and and/or whether the induced pluripotent stem cells are pro processes (e.g., cessation of the need for dialysis, artificial duced using Small molecules that mimic the effects of over 45 respiration, the use of a cardiopulmonary bypass machine or expressing one or more reprogramming factors. other prosthetic devices, such as artificial hearts, etc.) or by an In certain embodiments, compounds of the present inven increased life expectancy following organ and/or stem cell tion reduce, prevent or reverse organ damage or enhance transplantation. In certain embodiments, the Success of an organ preservation by protecting the organ against reperfu organ transplant procedure may be evaluated, for example, as sion injury. 50 enhanced organ viability and/or functional longevity follow In certain embodiments, compounds of the present inven ing transplantation as compared to an untreated organ (e.g., as tion reduce, prevent or reverse organ damage, reduce or pre may be measured by a delayed need for Subsequent transplan vent stem cell damage and/or death, enhance organ preserva tation and/or other therapeutic intervention(s)). The presence tion, or enhance stem cell preservation and/or Survival by of any of the foregoing may be viewed as an enhancement in decreasing or protecting against apoptosis. 55 the Success of an organ and/or stem cell transplant procedure. The present invention provides a method of promoting The present invention provides a method of prolonging Survival of an organ transplant recipient, comprising organ viability ex vivo, comprising administering a compound of the invention to an organ administering a compound of the invention to an organ donor patient prior to removal of the organ; donor patient prior to removal of the organ; administering a compound of the invention to an organ 60 contacting the organ ex vivo with a compound of the inven recipient prior to organ transplantation; tion; contacting the organ ex vivo with a compound of the inven contacting the organ ex vivo with a preservation solution tion; wherein the preservation Solution comprises a com contacting the organ ex vivo with a preservation Solution pound of the invention; wherein the preservation Solution comprises a com 65 or any combination thereof. pound of the invention; The present invention provides a method of prolonging or any combination thereof. stem cell viability ex vivo, comprising US 9,315,447 B2 121 122 administering a compound of the invention to a stem cell allopurinol, glutathione, glutamate, insulin, dexamethasone, donor patient prior to removal of the stem cells; hydroxyethyl starch, bactrim, trehalose, gluconate, or com contacting the stem cells ex vivo (e.g., in vitro in a Suitable binations thereof. In certain embodiments, the organ preser culture medium, Such as during the process of cyro Vation solution comprises sodium, potassium, magnesium, or preservation and/or thawing of cryopreserved stem cells combinations thereof. In certain embodiments, the organ or during ex vivo culture and/or manipulation, such as preservation solution is free or substantially free of cells, for stem cell expansion and/or differentiation) with a coagulation factors, DNA, or plasma proteins. In certain compound of the invention; embodiments, the organ preservation Solution is sterile. In or any combination thereof. certain embodiments, the organ preservation solution com The present invention provides a method of enhancing the 10 prises a compound of the invention at a concentration of Success of stem cell cryopreservation and/or thawing cryo greater than 1 nM, such as greater than 10 nM, 100 nM, 1 mM, preserved stem cells, comprising one or more steps of 10 mM or 100 mM. In certain embodiments, the organ pres administering a compound of the invention to a stem cell ervation Solution comprises an aqueous solution. In certain donor patient prior to removal of the stem cells; and embodiments, the organ preservation solution comprises a contacting the stem cells ex vivo (e.g., in vitro in a Suitable 15 perfluorocarbon. culture medium, Such as during the process of cryo The present invention provides a kit comprising: preservation of the stem cells and/or thawing of cryo a) a pharmaceutical formulation (e.g., one or more single preserved stem cells) with a compound of the invention. dosage forms) comprising a compound of the invention; A patient’s body, as a whole, can typically only tolerate and much lower levels of chemo-, bio- and radiation therapy than b) instructions for the administration of the pharmaceutical many particular organs. As such, prolonged and reliable ex formulation to an organ donor patient prior to removal of vivo organ viability would provide benefits outside the con the organ for reducing, preventing or reversing organ text of organ transplantation, including providing opportuni damage or enhancing organ preservation and/or Sur ties for ex vivo therapy. Accordingly, the Subject methods vival, and/or instructions for the administration of the may be used to permit an organ to be removed from the body 25 pharmaceutical formulation to a stem cell donor patient and treated in isolation, reducing the risk of damage to other prior to removal of the stem cells for reducing or pre parts of the body. venting stem cell damage and/or death or enhancing The present invention provides an organ infused with a stem cell preservation and/or Survival. compound of the invention. In certain embodiments, the The present invention provides a kit comprising: organ is ex vivo. For example, the present invention provides 30 a) a pharmaceutical formulation (e.g., one or more single an ex vivo organ infused with a compound of the invention. In dosage forms) comprising a compound of the invention; certain embodiments, the organ comprises a concentration of and greater than 1 nM of a compound of the invention, Such as 1 b) instructions for the administration of the pharmaceutical nM to 1 M, 1 mM to 1 M, or 10 mM to 1 M. In certain formulation to an organ recipient prior to organ trans embodiments, alumen of an organ comprises a fluid having a 35 plantation for reducing, preventing or reversing organ concentration of greater than 1 nM of a compound of the damage or enhancing organ preservation and/or Sur invention, such as 1 nM to 1 M, 1 mM to 1 M, or 10 mM to 1 vival, and/or instructions for the administration of the M. pharmaceutical formulation to a stem cell recipient prior The present invention further provides an organ in contact to stem cell transplantation for reducing or preventing with, and preferably partially or wholly submersed in, an 40 stem cell damage and/or death or enhancing stem cell organ preservation solution, wherein the organ preservation preservation and/or Survival. Solution comprises a compound of the invention. In certain The present invention provides a kit comprising: embodiments, the organ preservation solution further com a) a pharmaceutical formulation (e.g., one or more single prises potassium, Sodium, magnesium, calcium, phosphate, dosage forms) comprising a compound of the invention; Sulphate, glucose, citrate, mannitol, histidine, tryptophan, 45 and alpha-ketoglutaric acid, lactobionate, raffinose, adenosine, b) instructions for contacting an organ with the pharma allopurinol, glutathione, glutamate, insulin, dexamethasone, ceutical formulation for reducing, preventing or revers hydroxyethyl starch, bactrim, trehalose, gluconate, or com ing organ damage or enhancing organ preservation and/ binations thereof. In certain embodiments, the organ preser or Survival, and/or instructions for contacting stem cells Vation solution comprises sodium, potassium, magnesium, or 50 with the pharmaceutical formulation for reducing or combinations thereof. In certain embodiments, the organ preventing stem cell damage and/or death or enhancing preservation solution is free or substantially free of cells, stem cell preservation and/or Survival. coagulation factors, DNA, and/or plasma proteins. In certain In certain embodiments, the kit further comprises instruc embodiments, the organ preservation Solution is sterile. In tions for the administration of the pharmaceutical formula certain embodiments, the organ preservation Solution com 55 tion (e.g., one or more single dosage forms) comprising a prises a compound of the invention at a concentration of compound of the invention conjointly with a second thera greater than 1 nM, such as greater than 10 nM, 100 nM, 1 mM, peutic agent, such as those mentioned above. In certain 10 mM or 100 mM. In certain embodiments, the organ pres embodiments, the kit further comprises a second pharmaceu ervation Solution comprises an aqueous Solution. In certain tical formulation (e.g., one or more single dosage forms) embodiments, the organ preservation solution comprises a 60 comprising a second therapeutic agent, such as those men perfluorocarbon. tioned above. In certain embodiments, the kit further com The present invention provides an organ preservation solu prises a second pharmaceutical formulation (e.g., one or more tion comprising a compound of the invention. In certain single dosage forms) comprising a second agent Suitable for embodiments, the organ preservation solution further com reducing, preventing or reversing organ damage or enhancing prises potassium, Sodium, magnesium, calcium, phosphate, 65 organ preservation and/or Survival. In certain embodiments, Sulphate, glucose, citrate, mannitol, histidine, tryptophan, the kit further comprises a second pharmaceutical formula alpha-ketoglutaric acid, lactobionate, raffinose, adenosine, tion (e.g., one or more single dosage forms) comprising a US 9,315,447 B2 123 124 second agent Suitable for reducing or preventing stem cell stem cell preservation and/or Survival, and licensing, to a third damage and/or death or enhancing stem cell preservation party, the rights for further development and sale of the for and/or survival. mulation. The present invention provides a kit comprising: a) a first pharmaceutical formulation (e.g., one or more DEFINITIONS single dosage forms) comprising a therapeutic agent Suitable for modulating immune function, Suppressing Angiogenesis” is defined as any enhancement of an exist immune response, treating an autoimmune disease or ing vascular bed or the formation of new vasculature which autoimmune disorder, or treating a disease, sequela or benefits tissue perfusion. This includes the formation of new 10 vessels by sprouting of endothelial cells from existing blood pathological condition mediated by an activation of the vessels or the remodeling of existing vessels to alter size, immune system, Such as those mentioned above, or an maturity, direction or flow properties to improve blood per agent Suitable for reducing, preventing or reversing fusion of tissue. organ damage or enhancing organ preservation and/or As used herein, the term “corticosteroid’ refers to any of Survival, or an agent Suitable for reducing or preventing 15 the adrenal corticosteroid hormones isolated from the adrenal stem cell damage and/or death or enhancing stem cell cortex or produced synthetically, and derivatives thereof that preservation and/or Survival; and are used for treatment of inflammatory diseases, as described b) instructions for the administration of the first pharma herein. Corticosteroids include those that are naturally occur ceutical formulation (e.g., one or more single dosage ring, synthetic, or semi-synthetic in origin, and Such com forms) and a second pharmaceutical formulation (e.g., pounds are characterized by the presence of a steroid nucleus one or more single dosage forms) comprising a com of four fused rings, e.g., as found in cholesterol, dihydroxy pound of the invention for reducing, preventing or cholesterol, Stigmasterol, and lanosterol structures. reversing organ damage or enhancing organ preserva The term “LASIK', as used herein, is an acronym for tion and/or Survival, and/or instructions for the admin LAser in SItu Keratomileusis. This is a type of refractive istration of the first pharmaceutical formulation (e.g., 25 Surgery in which the cornea is reshaped to change its optical one or more single dosage forms) and a second pharma power. Specifically, a disc of cornea is raised as a flap, then an ceutical formulation (e.g., one or more single dosage excimer laser is used to reshape the middle layer of corneal forms) comprising a compound of the invention for tissue, producing Surgical flattening. LASIK surgery may be reducing or preventing stem cell damage and/or death or used for correcting myopia, hyperopia, and astigmatism. enhancing stem cell preservation and/or Survival. 30 As used herein, “immunosuppressive agent” refers to In certain embodiments, the invention relates to a method agents that Suppress the body's ability to elicit an immuno for conducting a pharmaceutical business, by manufacturing logical response to the presence of an antigenfallergen. For a formulation of a compound of the invention, or a kit as example, the ability to fight off disease or reject a transplanted described herein, and marketing to healthcare providers the organ. Another term for these agents is anti-rejection agents. benefits of using the formulation or kit for reducing, prevent 35 Not only are they are used to treat organ rejection after trans ing or reversing organ damage or enhancing organ preserva plantation, but many other diseases of immunological etiol tion and/or Survival, or for reducing or preventing stem cell ogy Such as Crohn's disease, rheumatoid arthritis, lupus, damage and/or death or enhancing stem cell preservation multiple Sclerosis, psoriasis, and other diseases and disorders and/or survival. as described herein. In certain embodiments, the invention relates to a method 40 The term 'graft, as used herein, refers to a body part, for conducting a pharmaceutical business, by providing a organ, tissue, or cells. Grafts may comprise all or part of one distribution network for selling a formulation of a compound or more organs such as liver, kidney, heart or lung; body parts of the invention, or kit as described herein, and providing Such as bone or skeletal matrix; tissue such as skin, intestines, instruction material to patients or physicians for using the endocrine glands; or progenitor stem cells of various types. formulation for reducing, preventing or reversing organ dam 45 The term “acyl is art-recognized and refers to a group age or enhancing organ preservation and/or Survival, or for represented by the general formula hydrocarbylC(O)—, pref reducing or preventing stem cell damage and/or death or erably alkylC(O)—. enhancing stem cell preservation and/or Survival. The term “acylamino' is art-recognized and refers to an In certain embodiments, the invention comprises a method amino group Substituted with an acyl group and may be for conducting a pharmaceutical business, by determining an 50 represented, for example, by the formula hydrocarbylC(O) appropriate formulation and dosage of a compound of the NH invention for reducing, preventing or reversing organ damage The term “acyloxy” is art-recognized and refers to a group or enhancing organ preservation and/or Survival, or for reduc represented by the general formula hydrocarbylC(O)O—, ing or preventing stem cell damage and/or death or enhancing preferably alkylC(O)O—. stem cell preservation and/or Survival, conducting therapeu 55 The term “alkoxy' refers to an alkyl group, preferably a tic profiling of identified formulations for efficacy and toxic lower alkyl group, having an oxygen attached thereto. Rep ity in animals, and providing a distribution networkfor selling resentative alkoxy groups include methoxy, ethoxy, propoxy, an identified preparation as having an acceptable therapeutic tert-butoxy and the like. profile. In certain embodiments, the method further includes The term “alkoxyalkyl refers to an alkyl group substituted providing a sales group for marketing the preparation to 60 with an alkoxy group and may be represented by the general healthcare providers. formula alkyl-O-alkyl. In certain embodiments, the invention relates to a method The term “alkenyl', as used herein, refers to an aliphatic for conducting a pharmaceutical business by determining an group containing at least one double bond and is intended to appropriate formulation and dosage of a compound of the include both “unsubstituted alkenyls' and “substituted alk invention for reducing, preventing or reversing organ damage 65 enyls', the latter of which refers to alkenyl moieties having or enhancing organ preservation and/or Survival, or for reduc Substituents replacing a hydrogen on one or more carbons of ing or preventing stem cell damage and/or death or enhancing the alkenyl group. Such Substituents may occur on one or US 9,315,447 B2 125 126 more carbons that are included or not included in one or more nyls', the latter of which refers to alkynyl moieties having double bonds. Moreover, such substituents include all those Substituents replacing a hydrogen on one or more carbons of contemplated for alkyl groups, as discussed below, except the alkynyl group. Such Substituents may occur on one or where stability is prohibitive. For example, substitution of more carbons that are included or not included in one or more alkenyl groups by one or more alkyl, carbocyclyl, aryl, het 5 triple bonds. Moreover, such substituents include all those erocyclyl, or heteroaryl groups is contemplated. contemplated for alkyl groups, as discussed above, except The term “alkyl refers to the radical of saturated aliphatic where stability is prohibitive. For example, substitution of groups, including straight-chain alkyl groups, branched alkynyl groups by one or more alkyl, carbocyclyl, aryl, het chain alkyl groups, cycloalkyl (alicyclic) groups, alkyl-sub erocyclyl, or heteroaryl groups is contemplated. stituted cycloalkyl groups, and cycloalkyl-substituted alkyl 10 groups. In preferred embodiments, a straight chain or The term "amide', as used herein, refers to a group branched chain alkyl has 30 or fewer carbon atoms in its backbone (e.g., C-Clso for straight chains, C-Clso for branched chains), and more preferably 20 or fewer. Likewise, preferred cycloalkyls have from 3-10 carbon atoms in their 15 ring structure, and more preferably have 5, 6 or 7 carbons in the ring structure. Moreover, the term “alkyl (or “lower alkyl) as used throughout the specification, examples, and claims is wherein each R" independently represent a hydrogen or intended to include both “unsubstituted alkyls' and “substi tuted alkyls', the latter of which refers to alkyl moieties hydrocarbyl group, or two R' are taken together with the N having Substituents replacing a hydrogen on one or more atom to which they are attached complete a heterocycle hav carbons of the hydrocarbon backbone. Such substituents, if ing from 4 to 8 atoms in the ring structure. not otherwise specified, can include, for example, a halogen, The terms “amine' and “amino” are art-recognized and refer to both unsubstituted and substituted amines and salts a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbo 25 nyl, a formyl, or an acyl), a thiocarbonyl (such as a thioester, thereof, e.g., a moiety that can be represented by a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, a phosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine, an imine, a cyano, a nitro, an azido, a R10 or R10 Sulfhydryl, an alkylthio, a Sulfate, a Sulfonate, a Sulfamoyl, a 30 N Nt-R10 V V Sulfonamido, a Sulfonyl, a heterocyclyl, an aralkyl, or an R10 R10 aromatic or heteroaromatic moiety. It will be understood by those skilled in the art that the moieties substituted on the hydrocarbon chain can themselves be substituted, if appro wherein each R" independently represents a hydrogen or a priate. For instance, the substituents of a substituted alkyl 35 hydrocarbyl group, or two R' are taken together with the N may include Substituted and unsubstituted forms of amino, atom to which they are attached complete a heterocycle hav azido, imino, amido, phosphoryl (including phosphonate and ing from 4 to 8 atoms in the ring structure. phosphinate), Sulfonyl (including Sulfate, Sulfonamido, Sul The term “aminoalkyl, as used herein, refers to an alkyl famoyl and Sulfonate), and silyl groups, as well as ethers, group Substituted with an amino group. alkylthios, carbonyls (including ketones, aldehydes, car 40 The term “aralkyl', as used herein, refers to an alkyl group boxylates, and esters), —CF, CN and the like. Exemplary Substituted with an aryl group. substituted alkyls are described below. Cycloalkyls can be The term “aryl” as used herein include substituted or further substituted with alkyls, alkenyls, alkoxys, alkylthios, unsubstituted single-ring aromatic groups in which each atom aminoalkyls, carbonyl-substituted alkyls, —CF, —CN, and of the ring is carbon. Preferably the ring is a 5- to 7-membered the like. 45 ring, more preferably a 6-membered ring. The term “aryl The term “C” when used in conjunction with a chemical moiety, Such as, acyl, acyloxy, alkyl, alkenyl, alkynyl, or also includes polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two alkoxy is meant to include groups that contain from X to y adjoining rings wherein at least one of the rings is aromatic, carbons in the chain. For example, the term "Calkyl” refers e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, to Substituted or unsubstituted Saturated hydrocarbon groups, 50 including straight-chain alkyl and branched-chain alkyl cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. Aryl groups that contain from X toy carbons in the chain, including groups include benzene, naphthalene, phenanthrene, phenol, haloalkyl groups such as trifluoromethyl and 2.2.2-trifluoro aniline, and the like. ethyl, etc. Coalkyl indicates a hydrogen where the group is in The term “carbamate' is art-recognized and refers to a a terminal position, a bond if internal. The terms "C2-alk 55 group enyl" and “Calkynyl" refer to substituted or unsubstituted unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but that contain at O O least one double or triple bond respectively. ul R10 or ls R10 The term “alkylamino', as used herein, refers to an amino 60 group Substituted with at least one alkyl group. A. r A. o1 The term “alkylthio', as used herein, refers to a thiol group R9 R9 substituted with an alkyl group and may be represented by the general formula alkylS-. wherein R and R' independently represent hydrogen or a The term “alkynyl', as used herein, refers to an aliphatic 65 hydrocarbyl group, such as an alkyl group, or R and R' group containing at least one triple bond and is intended to taken together with the intervening atom(s) complete a het include both “unsubstituted alkynyls' and “substituted alky erocycle having from 4 to 8 atoms in the ring structure. US 9,315,447 B2 127 128 The terms “carbocycle”, “carbocyclyl', and "carbocyclic', ethoxyethyl 2-pyridyl, and trifluoromethyl are considered to as used herein, refers to a non-aromatic saturated or unsatur be hydrocarbyl for the purposes of this application, but sub ated ring in which each atom of the ring is carbon. Preferably stituents such as acetyl (which has a =O substituent on the a carbocycle ring contains from 3 to 10 atoms, more prefer linking carbon) and ethoxy (which is linked through oxygen, ably from 5 to 7 atoms. 5 not carbon) are not. Hydrocarbyl groups include, but are not The term “carbocyclylalkyl, as used herein, refers to an limited to aryl, heteroaryl, carbocycle, heterocycle, alkyl, alkyl group Substituted with a carbocycle group. alkenyl, alkynyl, and combinations thereof. The term “carbonate' is art-recognized and refers to a The term “hydroxyalkyl, as used herein, refers to an alkyl group —OCO. R', wherein R' represents a hydrocarbyl group Substituted with a hydroxy group. group. 10 The term “carboxy’, as used herein, refers to a group The term “lower when used in conjunction with a chemi represented by the formula—CO.H. cal moiety, such as, acyl, acyloxy, alkyl, alkenyl, alkynyl, or The term “ester, as used herein, refers to a group —C(O) alkoxy is meant to include groups where there are ten or fewer OR' wherein R' represents a hydrocarbyl group. non-hydrogen atoms in the Substituent, preferably six or The term “ether', as used herein, refers to a hydrocarbyl 15 fewer. A “lower alkyl, for example, refers to an alkyl group group linked through an oxygen to another hydrocarbyl that contains ten or fewer carbon atoms, preferably six or group. Accordingly, an ether Substituent of a hydrocarbyl fewer. In certain embodiments, acyl, acyloxy, alkyl, alkenyl, group may be hydrocarbyl-O . Ethers may be either sym alkynyl, oralkoxy substituents defined herein are respectively metrical or unsymmetrical. Examples of ethers include, but lower acyl, lower acyloxy, lower alkyl, lower alkenyl, lower are not limited to, heterocycle-f-heterocycle and aryl-O-het- 20 alkynyl, or lower alkoxy, whether they appear alone or in erocycle. Ethers include “alkoxyalkyl groups, which may be combination with other Substituents, such as in the recitations represented by the general formula alkyl-O-alkyl. hydroxyalkyl and aralkyl (in which case, for example, the The terms “halo' and “halogen as used herein means atoms within the aryl group are not counted when counting halogen and includes chloro, fluoro, bromo, and iodo. the carbon atoms in the alkyl substituent). The terms “hetaralkyl and "heteroaralkyl, as used herein, 25 The terms “polycyclyl”, “polycycle', and “polycyclic' refers to an alkyl group Substituted with a hetaryl group. refer to two or more rings (e.g., cycloalkyls, cycloalkenyls, The term "heteroalkyl, as used herein, refers to a saturated cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls) in or unsaturated chain of carbonatoms and at least one heteroa which two or more atoms are common to two adjoining rings, tom, wherein no two heteroatoms are adjacent. e.g., the rings are “fused rings'. Each of the rings of the The terms "heteroaryland “hetaryl' include substituted or 30 polycycle can be substituted or unsubstituted. In certain unsubstituted aromatic single ring structures, preferably 5- to embodiments, each ring of the polycycle contains from 3 to 7-membered rings, more preferably 5- to 6-membered rings, 10 atoms in the ring, preferably from 5 to 7. whose ring structures include at least one heteroatom, pref The term “silyl refers to a silicon moiety with three hydro erably one to four heteroatoms, more preferably one or two carbyl moieties attached thereto. heteroatoms. The terms "heteroaryl and “hetaryl also 35 The term “substituted” refers to moieties having substitu include polycyclic ring systems having two or more cyclic ents replacing a hydrogen on one or more carbons of the rings in which two or more carbons are common to two backbone. It will be understood that “substitution' or “sub adjoining rings wherein at least one of the rings is heteroaro stituted with includes the implicit proviso that such substi matic, e.g., the other cyclic rings can be cycloalkyls, cycloalk tution is in accordance with permitted valence of the substi enyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls. 40 tuted atom and the substituent, and that the substitution Heteroaryl groups include, for example, pyrrole, furan, results in a stable compound, e.g., which does not spontane thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, ously undergo transformation Such as by rearrangement, pyrazine, pyridazine, and pyrimidine, and the like. cyclization, elimination, etc. As used herein, the term "sub The term "heteroatom' as used herein means an atom of stituted” is contemplated to include all permissible substitu any element other than carbon or hydrogen. Preferred het- 45 ents of organic compounds. In a broad aspect, the permissible eroatoms are nitrogen, oxygen, and Sulfur. Substituents include acyclic and cyclic, branched and The terms "heterocyclyl, "heterocycle', and "heterocy unbranched, carbocyclic and heterocyclic, aromatic and non clic” refer to substituted or unsubstituted non-aromatic ring aromatic Substituents of organic compounds. The permissible structures, preferably 3- to 10-membered rings, more prefer substituents can be one or more and the same or different for ably 3- to 7-membered rings, whose ring structures include at 50 appropriate organic compounds. For purposes of this inven least one heteroatom, preferably one to four heteroatoms, tion, the heteroatoms such as nitrogen may have hydrogen more preferably one or two heteroatoms. The terms "hetero Substituents and/or any permissible Substituents of organic cyclyl and "heterocyclic” also include polycyclic ring sys compounds described herein which satisfy the valences of the tems having two or more cyclic rings in which two or more heteroatoms. Substituents can include any Substituents carbons are common to two adjoining rings wherein at least 55 described herein, for example, a halogen, a hydroxyl, a car one of the rings is heterocyclic, e.g., the other cyclic rings can bonyl (such as a carboxyl, an alkoxycarbonyl, a formyl, oran be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroar acyl), a thiocarbonyl (such as a thioester, a thioacetate, or a yls, and/or heterocyclyls. Heterocyclyl groups include, for thioformate), an alkoxyl, a phosphoryl, a phosphate, a phos example, piperidine, piperazine, pyrrolidine, morpholine, phonate, a phosphinate, an amino, an amido, an amidine, an lactones, lactams, and the like. 60 imine, a cyano, a nitro, an azido, a Sulfhydryl, an alkylthio, a The term "heterocyclylalkyl, as used herein, refers to an Sulfate, a Sulfonate, a sulfamoyl, a Sulfonamido, a Sulfonyl, a alkyl group Substituted with a heterocycle group. heterocyclyl, an aralkyl, or an aromatic or heteroaromatic The term “hydrocarbyl, as used herein, refers to a group moiety. It will be understood by those skilled in the art that the that is bonded through a carbonatom that does not have a =O moieties substituted on the hydrocarbon chain can them or =S Substituent, and typically has at least one carbon- 65 selves be substituted, if appropriate. Unless specifically hydrogen bond and a primarily carbon backbone, but may stated as “unsubstituted, references to chemical moieties optionally include heteroatoms. Thus, groups like methyl, herein are understood to include substituted variants. For US 9,315,447 B2 129 130 example, reference to an “aryl group or moiety implicitly TMS or TIPS groups), glycol ethers, such as ethylene glycol includes both substituted and unsubstituted variants. and propylene glycol derivatives and allyl ethers. The term "sulfate is art-recognized and refers to the group The term “healthcare providers' refers to individuals or —OSOH, or a pharmaceutically acceptable salt thereof. organizations that provide healthcare services to a person, The term “sulfonamide' is art-recognized and refers to the 5 community, etc. Examples of “healthcare providers' include group represented by the general formulae doctors, hospitals, continuing care retirement communities, skilled nursing facilities, Subacute care facilities, clinics, mul tispecialty clinics, freestanding ambulatory centers, home R10 health agencies, and HMO's. O R10 or 's ( 10 The term “treating refers to: preventing a disease, disorder / / So or condition from occurring in a cell, a tissue, a system, N animal or human which may be predisposed to the disease, O| \,,R VR9 disorder and/or condition but has not yet been diagnosed as having it; stabilizing a disease, disorder or condition, i.e., 15 arresting its development; and relieving one or more symp wherein R and R' independently represents hydrogen or toms of the disease, disorder or condition, i.e., causing regres hydrocarbyl, such as alkyl, or RandR' taken together with sion of the disease, disorder and/or condition. the intervening atom(s) complete a heterocycle having from 4 As used herein, a therapeutic that “prevents” a disorder or to 8 atoms in the ring structure. condition refers to a compound that, in a statistical sample, The term “sulfoxide' is art-recognized and refers to the reduces the occurrence of the disorder or condition in the group —S(O) R', wherein R' represents a hydrocarbyl. treated sample relative to an untreated control sample, or The term “sulfonate' is art-recognized and refers to the delays the onset or reduces the severity of one or more symp group SOH, or a pharmaceutically acceptable salt thereof. toms of the disorder or condition relative to the untreated The term “sulfone' is art-recognized and refers to the control sample. group—S(O), R', wherein R' represents a hydrocarbyl. 25 As used herein, a "complex disorder having an inflamma The term “thioalkyl, as used herein, refers to an alkyl tory component' is a disease where the initial pathology/ group Substituted with a thiol group. dysfunction in a particular tissue or organ that is vital for the The term “thioester, as used herein, refers to a group systems biology function of an individual will secondarily —C(O)SR'or-SC(O)R' wherein R' represents a hydro lead to systemic metabolic derangement and/or tissue stress 30 causing, or further enhancing, activation of the immune sys carbyl. tem leading to dysfunction in several organs vital for body The term “thioether, as used herein, is equivalent to an homeostasis. ether, wherein the oxygen is replaced with a Sulfur. Pharmaceutical Compositions The term “urea' is art-recognized and may be represented The compositions and methods of the present invention by the general formula 35 may be utilized to treat an individual in need thereof. In certain embodiments, the individual is a mammal Such as a human, or a non-human mammal. When administered to an O animal. Such as a human, the composition or the compound is ul preferably administered as a pharmaceutical composition A. N1 40 comprising, for example, a compound of the invention and a pharmaceutically acceptable carrier. Pharmaceutically R9 R9 acceptable carriers are well known in the art and include, for example, aqueous solutions such as water or physiologically wherein R and R' independently represent hydrogen or a buffered saline or other solvents or vehicles such as glycols, hydrocarbyl, such as alkyl, or either occurrence of R taken 45 glycerol, oils such as olive oil or injectable organic esters. In together with R" and the intervening atom(s) complete a a preferred embodiment, when such pharmaceutical compo heterocycle having from 4 to 8 atoms in the ring structure. sitions are for human administration, the aqueous solution is “Protecting group' refers to a group of atoms that, when pyrogen free, or Substantially pyrogen free. The excipients attached to a reactive functional group in a molecule, mask, can be chosen, for example, to effect delayed release of an reduce or prevent the reactivity of the functional group. Typi 50 agent or to selectively target one or more cells, tissues or cally, a protecting group may be selectively removed as organs. The pharmaceutical composition can be in dosage desired during the course of a synthesis. Examples of protect unit form Such as tablet, capsule (including sprinkle capsule ing groups can be found in Greene and Wuts, Protective and gelatin capsule), granule, powder, syrup, Suppository, Groups in Organic Chemistry, 3 Ed., 1999, John Wiley & injection or the like. The composition can also be present in a Sons, NY and Harrison et al., Compendium of Synthetic 55 transdermal delivery system, e.g., a skin patch. The compo Organic Methods, Vols. 1-8, 1971-1996, John Wiley & Sons, sition can also be present in a solution Suitable for topical NY. Representative nitrogen protecting groups include, but administration, Such as an eye drop. are not limited to, formyl, acetyl, trifluoroacetyl, benzyl, ben A pharmaceutically acceptable carrier can contain physi Zyloxycarbonyl (“CBZ), tert-butoxycarbonyl (“Boc'), trim ologically acceptable agents that act, for example, to stabilize ethylsilyl (TMS), 2-trimethylsilyl-ethanesulfonyl 60 or to increase the absorption of a compound such as a com (“TES), trity1 and substituted trityl groups, alkyloxycarbo pound of the invention. Such physiologically acceptable nyl, 9-fluorenylmethyloxycarbonyl (“FMOC), nitro-vera agents include, for example, carbohydrates, such as glucose, tryloxycarbonyl (“NVOC) and the like. Representative Sucrose or dextrans, antioxidants. Such as ascorbic acid or hydroxyl protecting groups include, but are not limited to, glutathione, chelating agents, low molecular weight proteins those where the hydroxyl group is either acylated (esterified) 65 or other stabilizers or excipients. The choice of a pharmaceu or alkylated such as benzyl and trityl ethers, as well as alkyl tically acceptable carrier, including a physiologically accept ethers, tetrahydropyranyl ethers, trialkylsilyl ethers (e.g., able agent, depends, for example, on the route of administra US 9,315,447 B2 131 132 tion of the composition. The pharmaceutical composition of the compound which produces a therapeutic effect. Gen (preparation) also can be a liposome or other polymer matrix, erally, out of one hundred percent, this amount will range which can have incorporated therein, for example, a com from about 1 percent to about ninety-nine percent of active pound of the invention. Liposomes, for example, which com ingredient, preferably from about 5 percent to about 70 per prise phospholipids or other lipids, are nontoxic, physiologi cent, most preferably from about 10 percent to about 30 cally acceptable and metabolizable carriers that are relatively percent. simple to make and administer. Methods of preparing these formulations or compositions The phrase “pharmaceutically acceptable' is employed include the step of bringing into association an active com herein to refer to those compounds, materials, compositions, pound, such as a compound of the invention, with the carrier and/or dosage forms which are, within the scope of Sound 10 and, optionally, one or more accessory ingredients. In gen medical judgment, Suitable for use in contact with the tissues eral, the formulations are prepared by uniformly and inti of human beings and animals without excessive toxicity, irri mately bringing into association a compound of the present tation, allergic response, or other problem or complication, invention with liquid carriers, or finely divided solid carriers, commensurate with a reasonable benefit/risk ratio. or both, and then, if necessary, shaping the product. The phrase “pharmaceutically acceptable carrier as used 15 Formulations of the invention suitable for oral administra herein means a pharmaceutically acceptable material, com tion may be in the form of capsules (including sprinkle cap position or vehicle. Such as a liquid or Solid filler, diluent, Sules and gelatin capsules), cachets, pills, tablets, lozenges excipient, solvent or encapsulating material. Each carrier (using a flavored basis, usually Sucrose and acacia or traga must be “acceptable' in the sense of being compatible with canth), powders, granules, or as a solution or a suspension in the other ingredients of the formulation and not injurious to an aqueous or non-aqueous liquid, or as an oil-in-water or the patient. Some examples of materials which can serve as water-in-oil liquid emulsion, or as an elixir or syrup, or as pharmaceutically acceptable carriers include: (1) Sugars, pastilles (using an inert base. Such as gelatin and glycerin, or Such as lactose, glucose and Sucrose; (2) starches, such as Sucrose and acacia) and/or as mouth washes and the like, each corn starch and potato starch; (3) cellulose, and its deriva containing a predetermined amount of a compound of the tives. Such as sodium carboxymethyl cellulose, ethyl cellu 25 present invention as an active ingredient. Compositions or lose and cellulose acetate; (4) powdered tragacanth; (5) malt, compounds may also be administered as a bolus, electuary or (6) gelatin; (7) talc.; (8) excipients, such as cocoa butter and paste. Suppository waxes; (9) oils, such as peanut oil, cottonseed oil, To prepare solid dosage forms for oral administration (cap safflower oil, sesame oil, olive oil, corn oil and soybean oil; Sules (including sprinkle capsules and gelatin capsules), tab (10) glycols, such as propylene glycol; (11) polyols, such as 30 lets, pills, dragees, powders, granules and the like), the active glycerin, Sorbitol, mannitol and polyethylene glycol, (12) ingredient is mixed with one or more pharmaceutically esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) acceptable carriers, such as sodium citrate or dicalcium phos buffering agents, such as magnesium hydroxide and alumi phate, and/or any of the following: (1) fillers or extenders, num hydroxide; (15) alginic acid, (16) pyrogen-free water; Such as starches, lactose, Sucrose, glucose, mannitol, and/or (17) isotonic saline: (18) Ringer's solution; (19) ethyl alco 35 silicic acid; (2) binders, such as, for example, carboxymeth hol; (20) phosphate buffer solutions; and (21) other non-toxic ylcellulose, alginates, gelatin, polyvinyl pyrrolidone. Sucrose compatible Substances employed in pharmaceutical formula and/or acacia; (3) humectants, such as glycerol; (4) disinte tions. grating agents. Such as agar-agar, calcium carbonate, potato A pharmaceutical composition (preparation) can be or tapioca starch, alginic acid, certain silicates, and sodium administered to a subject by any of a number of routes of 40 carbonate; (5) Solution retarding agents, such as paraffin; (6) administration including, for example, orally (for example, absorption accelerators, such as quaternary ammonium com drenches as in aqueous or non-aqueous Solutions or Suspen pounds; (7) wetting agents, such as, for example, cetyl alco sions, tablets, capsules (including sprinkle capsules and gela hol and glycerol monostearate; (8) absorbents, such as kaolin tin capsules), boluses, powders, granules, pastes for applica and bentonite clay; (9) lubricants, such a talc, calcium Stear tion to the tongue); absorption through the oral mucosa (e.g., 45 ate, magnesium Stearate, Solid polyethylene glycols, sodium Sublingually); anally, rectally or vaginally (for example, as a lauryl Sulfate, and mixtures thereof, and (10) coloring agents. pessary, cream or foam); parenterally (including intramusc In the case of capsules (including sprinkle capsules and gela lularly, intravenously, Subcutaneously or intrathecally as, for tin capsules), tablets and pills, the pharmaceutical composi example, a sterile solution or Suspension); nasally; intraperi tions may also comprise buffering agents. Solid compositions toneally; Subcutaneously; transdermally (for example as a 50 of a similar type may also be employed as fillers in Soft and patch applied to the skin); and topically (for example, as a hard-filled gelatin capsules using Such excipients as lactose or cream, ointment or spray applied to the skin, or as an eye milk Sugars, as well as high molecular weight polyethylene drop). The compound may also be formulated for inhalation. glycols and the like. In certain embodiments, a compound may be simply dis A tablet may be made by compression or molding, option solved or suspended in sterile water. Details of appropriate 55 ally with one or more accessory ingredients. Compressed routes of administration and compositions Suitable for same tablets may be prepared using binder (for example, gelatin or can be foundin, for example, U.S. Pat. Nos. 6,110,973, 5,763, hydroxypropylmethyl cellulose), lubricant, inert diluent, pre 493, 5,731,000, 5,541,231, 5,427,798, 5,358,970 and 4,172, servative, disintegrant (for example, sodium starch glycolate 896, as well as in patents cited therein. or cross-linked sodium carboxymethyl cellulose), Surface The formulations may conveniently be presented in unit 60 active or dispersing agent. Molded tablets may be made by dosage form and may be prepared by any methods well molding in a suitable machine a mixture of the powdered known in the art of pharmacy. The amount of active ingredient compound moistened with an inert liquid diluent. which can be combined with a carrier material to produce a The tablets, and other solid dosage forms of the pharma single dosage form will vary depending upon the host being ceutical compositions, such as dragees, capsules (including treated, the particular mode of administration. The amount of 65 sprinkle capsules and gelatin capsules), pills and granules, active ingredient that can be combined with a carrier material may optionally be scored or prepared with coatings and to produce a single dosage form will generally be that amount shells. Such as enteric coatings and other coatings well known US 9,315,447 B2 133 134 in the pharmaceutical-formulating art. They may also be for The ointments, pastes, creams and gels may contain, in mulated so as to provide slow or controlled release of the addition to an active compound, excipients, such as animal active ingredient therein using, for example, hydroxypropy and vegetable fats, oils, waxes, paraffins, starch, tragacanth, lmethyl cellulose in varying proportions to provide the cellulose derivatives, polyethylene glycols, silicones, bento desired release profile, other polymer matrices, liposomes 5 nites, silicic acid, talc and Zinc oxide, or mixtures thereof. and/or microspheres. They may be sterilized by, for example, Powders and sprays can contain, in addition to an active filtration through a bacteria-retaining filter, or by incorporat compound, excipients such as lactose, talc, silicic acid, alu ing sterilizing agents in the form of sterile solid compositions minum hydroxide, calcium silicates and polyamide powder, that can be dissolved in sterile water, or some other sterile or mixtures of these Substances. Sprays can additionally con 10 tain customary propellants, such as chlorofluorohydrocar injectable medium immediately before use. These composi bons and volatile unsubstituted hydrocarbons, such as butane tions may also optionally contain opacifying agents and may and propane. be of a composition that they release the active ingredient(s) Transdermal patches have the added advantage of provid only, or preferentially, in a certain portion of the gastrointes ing controlled delivery of a compound of the present inven tinal tract, optionally, in a delayed manner. Examples of 15 tion to the body. Such dosage forms can be made by dissolv embedding compositions that can be used include polymeric ing or dispersing the active compound in the proper medium. Substances and waxes. The active ingredient can also be in Absorption enhancers can also be used to increase the flux of micro-encapsulated form, if appropriate, with one or more of the compound across the skin. The rate of such flux can be the above-described excipients. controlled by either providing a rate controlling membrane or Liquid dosage forms useful for oral administration include dispersing the compound in a polymer matrix or gel. pharmaceutically acceptable emulsions, microemulsions, Ophthalmic formulations, eye ointments, powders, solu Solutions, Suspensions, syrups and elixirs. In addition to the tions and the like, are also contemplated as being within the active ingredient, the liquid dosage forms may contain inert Scope of this invention. Exemplary ophthalmic formulations diluents commonly used in the art, Such as, for example, water are described in U.S. Publication Nos. 2005/0080056, 2005/ or other solvents, Solubilizing agents and emulsifiers, such as 25 0059744, 2005/003 1697 and 2005/004074 and U.S. Pat. No. ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl 6,583,124, the contents of which are incorporated herein by acetate, benzyl alcohol, benzyl benzoate, propylene glycol, reference. If desired, liquid ophthalmic formulations have 1,3-butylene glycol, oils (in particular, cottonseed, ground properties similar to that of lacrimal fluids, aqueous humor or nut, corn, germ, olive, castor and sesame oils), glycerol, tet vitreous humor or are compatable with such fluids. A pre rahydrofuryl alcohol, polyethylene glycols and fatty acid 30 ferred route of administration is local administration (e.g., esters of sorbitan, and mixtures thereof. topical administration, such as eye drops, or administration Besides inert diluents, the oral compositions can also via an implant). include adjuvants such as wetting agents, emulsifying and The phrases “parenteral administration” and “adminis Suspending agents, Sweetening, flavoring, coloring, perfum tered parenterally as used herein means modes of adminis ing and preservative agents. 35 tration other than enteral and topical administration, usually Suspensions, in addition to the active compounds, may by injection, and includes, without limitation, intravenous, contain Suspending agents as, for example, ethoxylated isos intramuscular, intraarterial, intrathecal, intracapsular, tearyl alcohols, polyoxyethylene Sorbitol and Sorbitan esters, intraorbital, intracardiac, intradermal, intraperitoneal, tran microcrystalline cellulose, aluminum metahydroxide, bento stracheal, Subcutaneous, Subcuticular, intraarticular, Subcap nite, agar-agar and tragacanth, and mixtures thereof. 40 Sular, Subarachnoid, intraspinal and intrasternal injection and Formulations of the pharmaceutical compositions for rec infusion. tal, vaginal, or urethral administration may be presented as a Pharmaceutical compositions suitable for parenteral Suppository, which may be prepared by mixing one or more administration comprise one or more active compounds in active compounds with one or more Suitable nonirritating combination with one or more pharmaceutically acceptable excipients or carriers comprising, for example, cocoa butter, 45 sterile isotonic aqueous or nonaqueous solutions, disper polyethylene glycol, a Suppository wax or a salicylate, and sions, Suspensions or emulsions, or sterile powders which which is solid at room temperature, but liquid at body tem may be reconstituted into sterile injectable solutions or dis perature and, therefore, will melt in the rectum or vaginal persions just prior to use, which may contain antioxidants, cavity and release the active compound. buffers, bacteriostats, solutes which render the formulation Formulations of the pharmaceutical compositions for 50 isotonic with the blood of the intended recipient or suspend administration to the mouth may be presented as a mouth ing or thickening agents. wash, or an oral spray, or an oral ointment. Examples of suitable aqueous and nonaqueous carriers that Alternatively or additionally, compositions can be formu may be employed in the pharmaceutical compositions of the lated for delivery via a catheter, stent, wire, or other intralu invention include water, ethanol, polyols (such as glycerol, minal device. Delivery via such devices may be especially 55 propylene glycol, polyethylene glycol, and the like), and Suit useful for delivery to the bladder, urethra, ureter, rectum, or able mixtures thereof, vegetable oils, such as olive oil, and intestine. injectable organic esters, such as ethyl oleate. Proper fluidity Formulations which are suitable for vaginal administration can be maintained, for example, by the use of coating mate also include pessaries, tampons, creams, gels, pastes, foams rials, such as lecithin, by the maintenance of the required or spray formulations containing Such carriers as are known 60 particle size in the case of dispersions, and by the use of in the art to be appropriate. Surfactants. Dosage forms for the topical or transdermal administration These compositions may also contain adjuvants such as include powders, sprays, ointments, pastes, creams, lotions, preservatives, wetting agents, emulsifying agents and dis gels, Solutions, patches and inhalants. The active compound persing agents. Prevention of the action of microorganisms may be mixed under sterile conditions with a pharmaceuti 65 may be ensured by the inclusion of various antibacterial and cally acceptable carrier, and with any preservatives, buffers, antifungal agents, for example, paraben, chlorobutanol, phe or propellants that may be required. nol sorbic acid, and the like. It may also be desirable to US 9,315,447 B2 135 136 include isotonic agents. Such as Sugars, sodium chloride, and include, but are not limited to, the severity of the patients the like into the compositions. In addition, prolonged absorp condition, the disorder being treated, the stability of the com tion of the injectable pharmaceutical form may be brought pound, and, if desired, another type of therapeutic agent being about by the inclusion of agents that delay absorption Such as administered with the compound of the invention. A larger aluminum monostearate and gelatin. total dose can be delivered by multiple administrations of the In some cases, in order to prolong the effect of a drug, it is agent. Methods to determine efficacy and dosage are known desirable to slow the absorption of the drug from subcutane to those skilled in the art (Isselbacher et al. (1996) Harrison's ous or intramuscular injection. This may be accomplished by Principles of Internal Medicine 13 ed., 1814-1882, herein the use of a liquid Suspension of crystalline or amorphous incorporated by reference). material having poor water solubility. The rate of absorption 10 In general, a suitable daily dose of an active compound of the drug then depends upon its rate of dissolution, which, in used in the compositions and methods of the invention will be turn, may depend upon crystal size and crystalline form. that amount of the compound that is the lowest dose effective Alternatively, delayed absorption of a parenterally adminis to produce a therapeutic effect. Such an effective dose will tered drug form is accomplished by dissolving or Suspending generally depend upon the factors described above. the drug in an oil vehicle. 15 If desired, the effective daily dose of the active compound Injectable depot forms are made by forming microencap may be administered as one, two, three, four, five, six or more Suled matrices of the Subject compounds in biodegradable Sub-doses administered separately at appropriate intervals polymers such as polylactide-polyglycolide. Depending on throughout the day, optionally, in unit dosage forms. In cer the ratio of drug to polymer, and the nature of the particular tain embodiments of the present invention, the active com polymer employed, the rate of drug release can be controlled. pound may be administered two or three times daily. In pre Examples of other biodegradable polymers include poly ferred embodiments, the active compound will be (orthoesters) and poly(anhydrides). Depot injectable formu administered once daily. lations are also prepared by entrapping the drug in liposomes The patient receiving this treatment is any animal in need, or microemulsions that are compatible with body tissue. including primates, in particular humans, and other mammals For use in the methods of this invention, active compounds 25 Such as equines, cattle, Swine and sheep; and poultry and pets can be given perse or as a pharmaceutical composition con in general. taining, for example, 0.1 to 99.5% (more preferably, 0.5 to In certain embodiments, compounds of the invention may 90%) of active ingredient in combination with a pharmaceu be used alone or conjointly administered with another type of tically acceptable carrier. therapeutic agent. As used herein, the phrase "conjoint Methods of introduction may also be provided by recharge 30 administration” refers to any form of administration of two or able or biodegradable devices. Various slow release poly more different therapeutic compounds such that the second meric devices have been developed and tested in vivo in compound is administered while the previously administered recent years for the controlled delivery of drugs, including therapeutic compound is still effective in the body (e.g., the proteinacious biopharmaceuticals. A variety of biocompat two compounds are simultaneously effective in the patient, ible polymers (including hydrogels), including both biode 35 which may include synergistic effects of the two com gradable and non-degradable polymers, can be used to form pounds). For example, the different therapeutic compounds an implant for the Sustained release of a compound at a can be administered either in the same formulation or in a particular target site. separate formulation, either concomitantly or sequentially. In Actual dosage levels of the active ingredients in the phar certain embodiments, the different therapeutic compounds maceutical compositions may be varied so as to obtain an 40 can be administered within one hour, 12 hours, 24 hours, 36 amount of the active ingredient that is effective to achieve the hours, 48 hours, 72 hours, or a week of one another. Thus, an desired therapeutic response for a particular patient, compo individual who receives such treatment can benefit from a sition, and mode of administration, without being toxic to the combined effect of different therapeutic compounds. patient. This invention includes the use of pharmaceutically The selected dosage level will depend upon a variety of 45 acceptable salts of compounds of the invention in the com factors including the activity of the particular compound or positions and methods of the present invention. In certain combination of compounds employed, or the ester, salt or embodiments, contemplated salts of the invention include, amide thereof, the route of administration, the time of admin but are not limited to, alkyl, dialkyl, trialkyl or tetra-alkyl istration, the rate of excretion of the particular compound(s) ammonium salts. In certain embodiments, contemplated Salts being employed, the duration of the treatment, other drugs, 50 of the invention include, but are not limited to, L-arginine, compounds and/or materials used in combination with the benenthamine, benzathine, betaine, calcium hydroxide, cho particular compound(s) employed, the age, sex, weight, con line, deanol, diethanolamine, diethylamine, 2-(diethylamino) dition, general health and prior medical history of the patient ethanol, ethanolamine, ethylenediamine, N-methylglucam being treated, and like factors well known in the medical arts. ine, hydrabamine, 1H-imidazole, lithium hydroxide, A physician or veterinarian having ordinary skill in the art 55 L-lysine, magnesium hydroxide, 4-(2-hydroxyethyl)mor can readily determine and prescribe the therapeutically effec pholine, piperazine, potassium hydroxide, 1-(2-hydroxy tive amount of the pharmaceutical composition required. For ethyl)pyrrolidine, Sodium hydroxide, triethanolamine, example, the physician or veterinarian could start doses of the tromethamine, and Zinc hydroxide salts. In certain embodi pharmaceutical composition or compound at levels lower ments, contemplated salts of the invention include, but are not than that required in order to achieve the desired therapeutic 60 limited to, Na, Ca, K, Mg., Zn or other metal salts. effect and gradually increase the dosage until the desired The pharmaceutically acceptable acid addition salts can effect is achieved. By “therapeutically effective amount” is also exist as various Solvates, such as with water, methanol, meant the concentration of a compound that is Sufficient to ethanol, dimethylformamide, and the like. Mixtures of such elicit the desired therapeutic effect. It is generally understood Solvates can also be prepared. The source of Such solvate can that the effective amount of the compound will vary accord 65 be from the solvent of crystallization, inherent in the solvent ing to the weight, sex, age, and medical history of the Subject. of preparation or crystallization, or adventitious to Such sol Other factors which influence the effective amount may Vent. US 9,315,447 B2 137 138 Wetting agents, emulsifiers and lubricants, such as Sodium and the corresponding methyl ester compound 1002, lauryl Sulfate and magnesium Stearate, as well as coloring agents, release agents, coating agents, Sweetening, flavoring and perfuming agents, preservatives and antioxidants can 10O2 also be present in the compositions. OH OH O Examples of pharmaceutically acceptable antioxidants include: (1) water Soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, Sodium bisulfate, sodium met OMe, abisulfite, sodium sulfite and the like; (2) oil-soluble antioxi dants, such as ascorbyl palmitate, butylated hydroxyanisole 10 (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, and the like; and (3) metal chelat in the ocular tissues of Dutch-belted rabbits when adminis ing agents, such as citric acid, ethylenediamine tetraacetic tered topically to the surface of the eye was evaluated. Healthy, young adult Dutch-belted rabbits (male/female; at acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the 15 least 1.5 kg) were acclimated for at least 7 days prior to like. initiation of the study. Prior to the initiation of the study, the In certain embodiments, the invention relates to a method eyes of the rabbits were examined to ensure that the eyes were for conducting a pharmaceutical business, by manufacturing free of any defects that may affect the integrity of the study. a formulation of a compound of the invention, or a kit as Examinations were performed by a qualified observer. described herein, and marketing to healthcare providers the Animals were dosed via the topical ocular route using a benefits of using the formulation or kit for treating or prevent pipettor system which delivered a 30 uL dose to each eye. ing any of the diseases or conditions as described herein. Each animal was dosed at T=0. The right eye received a dose In certain embodiments, the invention relates to a method concentration of 300 ug/mL of compound 1001 and the left for conducting a pharmaceutical business, by providing a eye received a dose concentration of 300 g/mL of compound distribution network for selling a formulation of a compound 25 1002. At the intervals noted in Table 3, the animals were of the invention, or kit as described herein, and providing sacrificed. instruction material to patients or physicians for using the formulation for treating or preventing any of the diseases or TABLE 3 conditions as described herein. Test Test In certain embodiments, the invention comprises a method 30 # of Dose Substance Substance Sacrifice for conducting a pharmaceutical business, by determining an Group Animals Volume Right Eye Left Eye Time appropriate formulation and dosage of a compound of the 1 2 30 L 1001 10O2 0.25 hrs 2 2 30 L 1001 10O2 0.50 hrs invention for treating or preventing any of the diseases or 3 2 30 L 1001 10O2 1 hr conditions as described herein, conducting therapeutic pro 35 4 2 30 L 1001 10O2 2 hrs filing of identified formulations for efficacy and toxicity in 5 2 30 L 1001 10O2 6 hrs animals, and providing a distribution network for selling an identified preparation as having an acceptable therapeutic Animals were euthanized with an overdose of an injectable profile. In certain embodiments, the method further includes barbiturate, and the eyes were retrieved from each animal. providing a sales group for marketing the preparation to 40 Using new or newly cleaned instruments for each eye, the healthcare providers. eyes were dissected and the following tissues isolated for In certain embodiments, the invention relates to a method analysis: conjunctiva, aqueous humor, cornea, lens, vitreous, for conducting a pharmaceutical business by determining an iris/ciliary body, retina/choroid, optic nerve, and Sclera. Each tissue was Snap frozen in liquid nitrogen and stored at -80°C. appropriate formulation and dosage of a compound of the for further analysis. invention for treating or preventing any of the disease or 45 Analysis of Ocular Tissues: conditions as described herein, and licensing, to a third party, Without wishing to be bound by theory, it is expected that the rights for further development and sale of the formulation. both compounds 1001 and 1002 are metabolized to their corresponding carboxylic acid, compound 1003 EXAMPLES 50

Example 1 1003 Ocular PK (Distribution) Study in Dutch-Belted OH OH O Rabbits 55 OM, The ocular distribution of compound 1001,

1 OO1 60 wherein M is H (compound 45) or a pharmaceutically accept OH OH O able cation, Such as ammonium, tetra-alkyl ammonium, Na (compound Z), K, Mg, and Zn. Without wishing to be bound OiPr, by theory, it is believed that compound 1003 or its meta bolite(s) is primarily responsible for the biological activity 65 associated with compound 1001. Compound 1003 was extracted from ocular tissues (cornea, retina) by homogeni Zation followed by protein precipitation with 4 parts acetoni US 9,315,447 B2 139 140 trile containing IS. The resulting Supernatant was dried under The linear standard curve of area ratio to compound 1003 nitrogen and reconstituted as a concentrated Solution in concentration was generated using 1/X weighting. Mobile Phase A. Compound 1003 was extracted from aque Analysis of Ocular Fluids: ous/vitreous humor by protein precipitation with 5 parts Compound 1003 was extracted from aqueous/vitreous acetonitrile containing IS. The resulting Supernatant was 5 humor by protein precipitation with 5 parts acetonitrile con dried under nitrogen and reconstituted as a concentrated Solu taining IS. Resulting Supernatant was dried under nitrogen tion in Mobile Phase A. and reconstituted as a concentrated solution in Mobile Phase Mobile Phase A was added to rabbit cornea (-80 mg) to A. achieve a final w/v concentration of 5% solids. Corneas were Spiking solutions for standards were prepared by diluting homogenized until a homogenous suspension was obtained. 10 compound 1003 as its sodium salt (e.g., M is Na) (dissolved For blank homogenate, multiple corneas were pooled and in ethanol) in ethanol spanning the concentration range of homogenized. 0.040 to 4000 ng/mL of compound 1003. Two standard Mobile Phase A was added to rabbit retina (-75 mg) to curves were extracted and Subsequently analyzed one before achieve a final w/v concentration of 7% solids. Retinas were and one after the samples. homogenized until a homogenous suspension was obtained. 15 To 1 volume aqueous or vitreous humor (120 uL), '4' For blank homogenate, multiple retinas were pooled and volume (30 uL) of ethanol was added to samples and blanks, homogenized. and '4" volume (30 uL) of spiking solution was added to Spiking solutions for standards were prepared by diluting standards. 5 Volumes of acetonitrile containing 25 ng/mL IS compound 1003 as its sodium salt (e.g., M is Na) (dissolved (600 uL) were added, and the samples were gently mixed for in ethanol) in ethanol spanning the concentration range of 20 ~1 minute and centrifuged for 10 min at 3200 rcf. The super 0.100 to 4000 ng/mL of compound 1003. Two standard natant was removed (~450 LL), dried under nitrogen at 60° C. curves were extracted and Subsequently analyzed one before and reconstituted in 90 uL of 5 mMAmmonium Acetate. and one after the samples. Alternatively, standards were prepared by diluting com To 1 volume homogenate (100 uL), Mo' volume (10 uL) of pound 1003 as its sodium salt (e.g., M is Na) directly in blank ethanol was added to samples and blanks, and Mo' volume 25 aqueous or vitreous humor spanning the concentration range (10 uL) of spiking Solution was added to standards. 4 Volumes of 0.01.00 to 1000 ng/mL of compound 1003. This eliminated of acetonitrile containing 12.5 ng/mL IS (400 uL) were the addition of '4" volume (30 uD) of ethanol to samples and added, and the samples were gently mixed for ~ 1 minute and blanks in the Subsequent extraction. centrifuged for 10 min at 3200 rcf. The supernatant was Compound 1003 in the extracted/dried/reconstituted removed (~400 uL), dried under nitrogen at 60° C. and recon 30 samples was analyzed by LC/MS/MS using D4-PGE2 as the stituted in 100 uL of 5 mMAmmonium Acetate. internal standard. Detection was performed on an Applied Alternatively, standards were prepared by diluting com Biosystems API-4000 Mass Spectrometer using the follow pound 1003 as its sodium salt (e.g., M is Na) directly in blank ing method: homogenate spanning the concentration range of 0.0100 to LCGuard Column: Aqua C184x2.0 mm, 3 um, Phenomenex 400 ng/mL of compound 1003. This eliminated the addition 35 if AJO-7510 of /10" volume (10 uL) of ethanol to samples and blanks in Column: Luna C18(2) 30x2.0 mm, 3 um, Phenomenex the Subsequent extraction. HOOA-4251-BO Compound 1003 in the homogenized/extracted/dried/re Mobile Phase A: 5 mMAmmonium Acetate in water constituted samples was analyzed by LC/MS/MS using Mobile Phase B: 5 mMAmmonium Acetate in 95.5 aceto D4-PGE2 as the internal standard. Detection was performed 40 nitrile: water on an Applied Biosystems API-4000 Mass Spectrometer Gradient: using the following method: LCGuard Column: Aqua C184x2.0 mm, 3 um, Phenomenex if AJO-7510 t = O 5% B t = 1 min 5% B Column: Luna C18(2) 30x2.0 mm, 3 um, Phenomenex 45 t = 1.5 min 40% B HOOA-4251-BO t = 2.0 min 40% B Mobile Phase A: 5 mMAmmonium Acetate in water t = 2.75 min 100% B Mobile Phase B: 5 mMAmmonium Acetate in 95.5 aceto t = 4.15 min 100% B t = 4.25 min 5% B nitrile: water t = 4.75 min 5% B Gradient: 50 Flow Rate: 0.5 mL/min t = O 5% B Injection Volume: 50 uL t = 1 min 5% B MS Detection Scan Type: MRM t = 1.5 min 40% B 55 t = 2.0 min 40% B Polarity: Negative t = 2.75 min 100% B Ion Source Electrospray t = 4.15 min 100% B Analyte Transition: m/z. 260.95 to 114.9 t = 4.25 min 5% B IS Transition: m/z. 355.0 to 275.3 t = 4.75 min 5% B The linear standard curve of area ratio to compound 1003 60 concentration was generated using 1/x weighting. Flow Rate: 0.5 mL/min FIG. 1 shows that comparable levels of compound 1003 Injection Volume: 50 uL were observed in the aqueous humor (FIG.1a), the vitreous MS Detection Scan Type: MRM (FIG. 1b), and the cornea (FIG. 1c) upon administration of Polarity: Negative compounds 1001 and 1002 using the protocol described Ion Source Electrospray 65 above. Analyte Transition: m/z. 260.95 to 114.9 Ocular pharmacokinetic parameters (for the measured IS Transition: m/z. 355.0 to 275.3 compound 1003) following administration of either prodrug US 9,315,447 B2 141 142 compound 1001 or 1002 are shown in Table 4. Peak levels of reference solution for compound 1002 was stable for up to 3 compound 1003 following administration of compound 1001 days stored at 5°C.-3°C., protected from light. were ~12 mM in cornea with a half-life of ~1 hour and in the Ophthalmic formulations of compounds 1001 (0.994 g of a vitreous peak levels were ~15 nM with a slightly longer 52 mg/mL Solution in propylene glycol) or compound 1002 (1.034 g of a 50 mg/mL Solution in propylene glycol) were half-life of 1.3 hours. 5 prepared at 500 g/mL dose strength in potassium phosphate buffer (25 mM, pH 5.5, 0.34g) with 0.2M sodium hydroxide TABLE 4 solution (q.s.), sodium chloride (0.327 g), poly sorbate 80 Aq. Humor Cornea Vitreous (0.5% w/v), and purified water (q.s. to 100). Sample solutions 10 of compounds 1001 and 1002 were each prepared by accu Compound 1001 rately weighing approximately 20 mg of the 500 ug/mL com T/2. (h) 1.18 1.OS 1.3 pound formulation into a 1 mL amber Volumetric flask. AUC (h * ng/mL) or 2756 5.53 6.61 Approximately 0.5 mL of sample diluent was added and the (h ng/mg) Cmax (ng/mL) or (ng mg) 1055 3.56 4.36 solution gently swirled to mix. The solution was diluted to Tmax (h) O.25 O.25 O.25 15 volume with sample diluent and mixed well. Compound 1002 The following equation was used to calculate the com pound 1001 or compound 1002 purity assay (% area) in each T/2. (h) 1.3 1.17 1.35 sample: AUC (h * ng/mL) or 3.146 3.02 3.74 (h ng/mg) Analogue Content(% area)=(Area of analog Total area Cmax (ng/mL) or (ng mg) 1337 3.35 1.64 of all peakse0.10% area)x100. Tmax (h) O.25 O.25 O.25 Impurities 0.10% area were integrated and reported as % area to 2 decimal places. Tables 5 and 6 show the results of the chemical stability Example 2 study of compounds 1001 and 1002, respectively, over the 25 course of 8 weeks at 5° C. Compound 1001 displayed better Chemical Stability Study chemical stability than compound 1002, as shown by a greater percent purity over time (with less impurities). The chemical stability of compounds 1001 and 1002 was measured via HPLC analysis of the purity of these com TABLE 5 pounds over time. The purity assay of active components 30 using percent area (% area) and impurities reported as percent Chemical Stability of Compound 1001 area (% area) were measured by HPLC using the following % Area conditions: Compound % Area Column: ACE 3, Cs, 3 pin, 100x2.1 mm Time Point 1001 Impurity 35 Column Temperature: 35°C. Initial 99.55 O45 Autosampler Temperature: 5°C. 1 week Not Tested Not Tested Detection: UV at 272 nm 2 weeks 99.03 0.97 Injection Volume: 30 uL 4 weeks 99.39 O.61 Flow Rate: 0.5 mL/min 8 weeks 99.31 O.69 Run Time: 20 minutes 40 Needle Wash: 100% Methanol with extended needle wash Mobile Phase: A) Water/Trifluoroacetic acid (100/0.05 V/v), TABLE 6 pH 3.0 B) Acetonitrile?Trifluoroacetic acid (100/0.05 V/v) Chemical Stability of Compound 1002 45 % Area Compound % Area Time (min) % A % B Time Point 10O2 Impurity

O 8O 2O Initial 98.29 1.71 2 8O 2O 1 week 97.58 2.43 12 60 40 50 2 weeks 97.46 2.55 12.01 8O 2O 4 weeks 96.54 3.46 2O 8O 2O 8 weeks 94.95 5.05

Retention Times: Compound 1002, 6.3 minutes 55 Example 3 Compound 1001, 11.0 minutes Compound 1003, 2.3 minutes Study of Compound 1001 in Dry Eye A sample diluent was prepared with water/absolute ethanol (90/10 v/v). The objective of this study was to compare the efficacy of Reference solutions of compounds 1001 and 1002 were 60 compound 1001 to placebo (vehicle minus active) for the prepared by weighing approximately 10 mg (t2 mg) of a 1 treatment of the signs and symptoms of dry eye, in a multi mg/mL stock solution of compound 1001 or 1002 into a 1 mL center, double-masked, randomized, placebo-controlled amber volumetric flask. Approximately 0.5 mL of sample study in which 100 human patients were equally randomized diluent was added and the solution swirled to mix. The sample into one of three active treatment groups or placebo and all was diluted to volume with sample diluent and mixed well. 65 treatment groups were exposed to identical humidity, tem The reference solution for compound 1001 was stable for up perature, and wind conditions which exacerbated the Symp to 7 days stored at 5° C.-3°C., protected from light. The toms of dry eye. US 9,315,447 B2 143 144 During a 14 day study run-in period prior to randomization obtained on a Bruker AV-300 spectrometer at 300 MHz for (e.g., day -14 to day 0), all patients received placebo bilater proton and 75 MHz for carbon, using CDC1, DMSO-d, or ally two times per day (BID). At the time of treatment ran CDOD as the solvent. Tetramethylsilane was used as an domization (e.g., day 0), patients were exposed to 90 minutes internal reference for proton spectra and the solvent peak was of identical humidity, temperature, and wind conditions used as the reference peak for carbon spectra. Mass spectra which exacerbated the symptoms of dry eye after which were obtained on a Finnigan LCQ Duo LC-MS ion trap patients were stratified based on the following criterion: Cen electrospray ionization (ESI) mass spectrometer. Thin-layer tral corneal staining post-exposure <2.5 or >3.0. Patients chromatography (TLC) was performed on Whatman No. eligible to be randomized received one of the following treat 4500-101 silica-gel plates (250 um layer thickness). Visual ments to be administered as approximately a 30 uL drop to 10 ization of TLC plates was performed using ultraviolet light both eyes BID: 1) Compound 1001 at dose A (26.4 ug/mL) Ophthalmic Solution; 2) Compound 1001 at dose B (87.8 (254 nm) or iodine staining. Corning pH meter 430 apparatus ug/mL) Ophthalmic Solution; 3) Compound 1001 at dose C was used to determine pH. (275.6 ug/mL) Ophthalmic Solution; 4) Placebo Ophthalmic HPLC Conditions: Method A Solution (Vehicle minus active). 15 Column: Zorbax SB-CN, 4.6x250 mm, 5uM Brief Summary of Visit Schedule. 5 visits over the course of approximately 6 weeks were scheduled as follows: Visit Column Temp Ambient 1=Day -14+1 day; Visit 2-Day 0; Visit 3-Day 14+2 days; Sample Temp Ambient Visit 4-Day 28+2 days; Visit 5-Day 29-1 hour. Detection: UV at 254 nm The primary endpoint of mean ocular discomfort in the conditions for exacerbating the symptoms of dry eye using Mobile Phase A: HPLC water with 0.1% formic acid the ORA Ocular Discomfort Scale (during the 90 minute Mobile Phase B: HPLC acetonitrile with 0.1% formic acid exposure) was Summarized statistically (m, mean, standard Diluent: 50:50 waterfacetonitrile deviation, median, min and max), and analyzed with 1-sided Flow Rate: 1 mL/min t-tests comparing the best dose to placebo with a 25 p-values 0.025 considered significant. At visit 4 (e.g., day 28), patients were exposed to 90 min TABLE 7 utes of identical humidity, temperature, and wind conditions which exacerbated the symptoms of dry eye. Ocular discom Gradient fort during this exposure period was measured using the ORA 30 Ocular Discomfort Scale with the ocular discomfort score Time (min) % A % B being the average of all time points during exposure. Ocular discomfort was measured again at visit 5 (e.g., day 29). O 90 10 Scores on the ORAOcular Discomfort Scale areas follows: a 2O O 100 score of 0 indicates no discomfort; a score of 1 indicates 35 22 90 10 intermittent awareness; a score of 2 indicates constant aware 25 90 10 ness; a score of 3 indicates intermittent discomfort; and a score of 4 indicates constant discomfort. FIG. 2 shows ocular discomfort as measured on environ mental day 28 after topical administration of vehicle or com 40 pound 1001 at doses A, B, or C. The change in ocular dis Scheme 1. Preparation of Compound 2005 comfort as compared to vehicle showed approximately a 27% TMS TMS improvement when compound 1001 was administered at AlCl3, DCM dose C. -- FIG. 3 shows ocular discomfort as measured on environ 45 O O O mental day 29 (e.g., 24 hours after last treatment) after topical 2001 administration of vehicle or compound 1001 at doses A, B, or TsOH (0.1 equiv) C. The change in ocular discomfort as compared to vehicle O O IPA, 659 C. -e- showed approximately a 36% improvement when compound 86% over 2 steps 1001 was administered at dose C, indicating that the effect 50 seen upon administration of compound 1001 persists 24 2 OH hours after the last treatment. TMS Symptoms of grittiness, dryness were measured on envi 2002 ronmental day 28 on a visual analog scale of 0-5. FIG. 4 O O Noyori catalyst shows a significant change in both grittiness and dryness, as 55 compared to vehicle, when compound 1001 was topically i-PrCH, 100% administered at dose C. 2 Oi-Pir 99% ee TMS Example 4 2003 60 OH O Synthetic Protocols TBAF 84% All nonaqueous reactions were carried out under an atmo 2. Oi-Pir sphere of dry nitrogen. Reagents were purchased from com TMS mercial sources and used as received. Solvents for reactions 65 were reagent-grade unless otherwise noted in the text. Proton 2004 and carbon nuclear magnetic resonance (NMR) spectra were US 9,315, 447 B2 145 146 -continued vacuum at 35-40°C. to give compound 2002 (1,700 g, 91%) OH O as a dark brown oil which was used in the next step without further purification. O-P Synthesis of Compound 2003: 26 - 5 To a clean, dry, 12-L, four-neck, round-bottom flask H equipped with a condenser and a temperature probe were 2005 charged crude acid 2002 (1000 g), p-TsOH.H2O (89.5g, 0.1 equiv), and 2-propanol (6 L). The reaction mixture was Synthesis of Compound 2002: 10 heated at 65° C. for 24 h at which point the conversion was To a clean, dry, 100-L jacketed reactor equipped with a about 90% by HPLC. The reaction mixture was cooled and condenser, an addition funnel, and a temperature probe was 2-propanol was concentrated under vacuum at 50–55°C. The resulting residue was added to MTBE (3 L) and washed with cautiously added aluminum chloride (1285 g, 9.64 mol) fol saturated NaHCOs (3x500 mL). The organic layer was dried lowed by dichloromethane (55 L). The mixture was stirred 15 over NaSO, filtered over a packed bed of Celite on a glass and cooled to 0-5°C. A solution of glutaric anhydride (1000 fritted funnel, and washed thoroughly with MTBE. The fil g, 8.76 mol) and bis(trimethylsilyl)acetylene (1643 g, 9.64 trate was concentrated under vacuum at 40-45° C. to give mol) in dichloromethane (10 L) was cautiously added to the crude ester 2003 (1,160 g, 96%) as a dark brown oil, which was used in the next step without further purification. mixture dropwise while maintaining the temperature between Synthesis of Compound 2005: 0 and 5° C. After the addition was complete, the reaction To a clean, nitrogen-flushed, 12-L, round-bottom flask mixture was stirred at room temperature overnight. After equipped with a mechanical stirrer, temperature probe, and 12-20h, the reaction mixture was assayed to check for the nitrogen inlet were charged compound 2003 (433 g, 1.7 mol), anhydrous isopropyl alcohol (4.3 L), and compound 2010 presence of glutaric anhydride by "H NMR (CDC1). The 25 sample was prepared for HNMR analysis by adding dichlo (20.4g, 0.034 mol). After stirring at ambient temperature for 1 h, the conversion to compound 2004 was deemed complete romethane (0.5 mL) and 1 M HCl (0.25 mL) to a sample (0.25 by TLC analysis (20% EtOAc/heptane). Ammonium chloride mL) of the reaction mixture in a vial; the dichloromethane (109 g, 2.03 mol) was added followed by the streamwise was separated and removed under vacuum. The residue was 30 addition of tetrabutylammonium fluoride (2.03 L, 1 M solu diluted with CDC1 and submitted for "H NMR analysis. The tion in THF). After overnight stirring, the conversion to com reaction was deemed complete when less than 3% by wt of pound 2005 was deemed complete by TLC analysis. The glutaric anhydride remained. The reaction mixture was then reaction mixture was concentrated under vacuum to approxi slowly added to a 1M HCl solution (12 L) while maintaining mately /10 of the original Volume and the material was parti 35 tioned between MTBE (4.3 L) and saturated aqueous ammo the temperature below 10° C. The mixture was stirred for nium chloride (2 L). Phases were separated and the aqueous 30-60 min until a clear solution was observed. Two phases was extracted with MTBE (2 L). The combined organics were were separated and the organic phase was washed with brine washed with brine (2 L), dried over sodium sulfate, and con (12 L), dried over sodium sulfate, filtered over a packedbed of 40 centrated to a dark oil. Chromatographic purification gave Celite on a glass fritted funnel, and washed thoroughly with compound 2005 (216 g. 70% as pure fractions and 28 g.9% dichloromethane. The filtrate was concentrated under as mixed fractions) as an orange oil.

Scheme 2. Preparation of Noyori Catalyst i-Pir RuCl3 -- Me C EtOH wC.,x / 619/ 0. / x',W YCI ''R.Yo ir ( )- Me 2006 i-Pir 2007 KOH DCM 91% O NH2 TsCl2, NaOH Ol H2O, DCM 88% NH, NH

2008 US 9,315,447 B2 147 148 -continued f Me Nv 21 Ru N^ N H i-Pir

2010 Noyori catalyst

Synthesis of Compound 2007: 15 Scheme 3. Preparation of Compound 2018 To a nitrogen-flushed, 2-L, three-neck, round-bottom flask O were added ruthenium chloride hydrate (12g, 0.058 mol) and TBDMSCI, imidazole absolute ethanol (0.6 L) followed by p-mentha-1,5-diene (96 OH --DMAP, CHCL, D-90% mL). The mixture was stirred at reflux for 4 h. The suspension was cooled, stirred at 0-5° C. for 30 min, and filtered; the 2011 O solids were washed with absolute ethanol (3x1 bed volume) TMS, n-hexLi to yield compound 2007 (10.4g, 58%) as a dark red solid. The -e- filtrate was concentrated to 75 mL and was stored in the Auons BFEtO, THF refrigerator overnight. The solids were filtered and washed 25 2012 with absolute ethanol (3x1 bed volume) to give compound TMS OH 2007 (2g, 11%) as a dark red solid. Šs HTBDPSCI, imidazole Synthesis of Compound 2009: OTBS CHCl2, >80% over 2-steps 2013 To a 2-L, three-neck, round-bottom flask equipped with a 30 TMS OTBDPS temperature probe, magnetic stir bar, nitrogen inlet, and addi CSA Hip tion funnel were added (1S,2S)-(-)-1,2-diphenylethylenedi CHC1/CHOH amine (20g, 0.094 mol) and dichloromethane (160 mL). The Suluors mixture was cooled to 0-3°C. and a 1 M solution of sodium 2014 TMS OTBDPS hydroxide (160 mL) was added dropwise while maintaining 35 DMSO, pyo SO3, EtN the temperature below 5°C. To this mixture was added a OH -e-CHCl2, 70% over 2 steps solution of toluenesulfonyl chloride (17.9 g, 0.094 mol) in dichloromethane (320 mL) dropwise over a 2-h period. The biphasic mixture was stirred at 0-5°C. for an additional 1 h TMS OTBDPS and the reaction was deemed complete by TLC (50% EtOAc/ 40 Šs O -e- heptane, UV). Phases were separated and the organic phase was washed with water (2x320 mL) and brine (320 mL), dried H over sodium sulfate and concentrated to a crude solid. The 2016 solid was dissolved in toluene (200 mL) at 70-80° C. and 45 TMS OTBDPS heptane (300 mL) was added portionwise while maintaining CHI, this temperature. The resulting slurry was cooled and stirred -CrCl - at 20-25° C. for 1 h and then cooled and stirred at 0-5°C. for Null-N- 10 min. The solids were filtered and washed with a 50% 2017 solution of toluene in heptane (3x1 bed volume) to give TMS OTBDPS compound 2009 (30.24g, 88%) as a white powder. 50 Synthesis of Compound 2010: To a 1-L, nitrogen-flushed, three-neck, round-bottom flask 2018 equipped with a temperature probe, mechanical stirrer, addi tion funnel and nitrogen inlet were added compound 2007 55 Synthesis of Compound 2012: (10.4 g., 0.017 mol), compound 2009 (12.5 g., 0.034 mol), A three-neck, 22-L, round-bottom flask equipped with a potassium hydroxide (14.14 g., 0.252 mol), and anhydrous magnetic stirrer, a temperature probe, and an addition funnel dichloromethane (217 mL). This mixture was stirred at with a nitrogen inlet was charged with (S)-(-)-glycidol (1.0 20-25° C. for 10 min and then water (217 mL) was added kg, 13.5 mol) and DCM (3.0 L). Imidazole (1.01 kg, 14.8 dropwise while maintaining the temperature below 30° C. 60 mol) and DMAP (84g, 680 mmol) were added and the reac The resulting mixture was stirred for 15 min and the phases tion was cooled to 0°C. A solution of TBDMSC1 (2.04 kg, separated. The organic phase was washed with water (217 13.5 mol) in DCM (2.0 L) was added while maintaining the mL), dried over sodium sulfate, and filtered. The filtrate was internal temperature below 10°C. The reaction was stirred at dried over calcium hydride (4.2 g) portionwise and the solids 0° C. for 2 h and analyzed by TLC (9:1 heptane/EtOAc) were filtered over Celite and washed with anhydrous dichlo 65 which indicated that the reaction was complete. The reaction romethane. The filtrate was concentrated under vacuum to mixture was purified by Silica-gel plug in two bathes (silica give compound 2010 (20g, 98%) as a purple solid. gel: 6 kg each) using DCM as eluent to afford the desired US 9,315,447 B2 149 150 product as a colorless oil (2.2 kg, 86% yield). The "H NMR trated to dryness. The residue was purified in four batches spectrum was consistent with the assigned structure of com with a silica-gel plug (silica gel: 7 kg each) using 2% EtOAc pound 2012. in heptanes as eluent to afford the desired product as a yellow Synthesis of Compound 2013: oil (2.0 kg, 59% yield). The mixed fractions were combined A multiple-neck, 72-L, round-bottom flask equipped with and concentrated to dryness to afford a yellow oil (0.38 kg, a mechanical stirrer, a temperature probe, and an addition 11% yield). The "H NMR spectrum was consistent with the funnel with a nitrogen inlet was charged with THF (5.5 L). assigned structure of compound 2016. n-Hexyllithium (2.3M in hexane, 5.3L, 12.3 mol) was added while maintaining the internal temperature below 30°C. The Synthesis of Compound 2017: solution was cooled to -20°C. and TMS acetylene (1.98 L. 10 A three-neck, 22-L, round-bottom flask equipped with a 1.2 equiv) was added at <-50° C. The reaction was stirred at magnetic stirrer, a temperature probe, aheating mantle, and a <-55° C. for 1 h and a solution of compound 2012 (2.2 kg, nitrogen inlet was charged with compound 2016 (2.0 kg, 11.66 mol) was added over 75 min at <-55° C., followed by contained 1.42 kg of 2017 by weight assay, 3.48 mol) and the addition of BF.EtO (1.44 L, 1.0 equiv) over 110 min at ACN (7.1 L). The reaction mixture was heated to 30° C. and <-55° C. The reaction was stirred at <-60° C. for 16 hand 15 PhPCHCHO (1.38 kg, 68.6 mmol) was added in portions TLC analysis (9:1 heptane/EtOAc) indicated that the reaction while maintaining the internal temperature at <35° C. The was complete. It was quenched with 75% saturated brine reaction was heated at 30° C. for 15 h and analyzed by "H solution (11 L, 5 vol) and diluted with MTBE (11 L, 5 vol). The organic layer was concentrated to dryness to afford a NMR which indicated that the reaction was complete. It was colorless oil (3.43 kg, >100% yield). The H NMR spectrum concentrated to dryness and the residue was dissolved in was consistent with the assigned structure of compound 2013. DCM (0.71 mL). The resulting solution was diluted with Synthesis of Compound 2014: heptanes (1.42 L) and purified by silica-gel plug in two A multiple-neck, 72-L, round-bottom flask equipped with batches using 10:1 heptane/EtOAc as eluent to afford the a mechanical stirrer, a temperature probe, and an addition desired product as a pale yellow oil (1.87 kg, 88% yield). The funnel with a nitrogen inlet was charged with crude 2013 25 H NMR spectrum was consistent with the assigned structure (3.43 kg) and DCM (6.6 L). Imidazole (1.19 kg, 17.5 mol) and of compound 2017. DMAP (70 g. 580 mmol) were added and the reaction was Synthesis of Compound 2018: cooled to 0°C. TBDPSC1 (4.17 kg, 15.2 mol) was added A 5-L, three-neck, round-bottom flask was heated with a while maintaining the internal temperature below 10°C. The heat gun under vacuum (1-10 torr) three times and purged reaction was stirred at ambient temperature for 18 h and 30 under argon. Anhydrous chromium chloride (100 g, 0.814 analyzed by TLC (9:1 heptane/EtOAc) which indicated that mol) was added and the flask was again heated under vacuum the reaction was complete. The reaction mixture was concen three times. Anhydrous tetrahydrofuran (1000 mL) was trated to dryness, diluted with heptanes (11 L), and washed added and the resulting slurry was stirred for 2 hat ambient with water (2x11 L). The organic layer was concentrated to temperature. The slurry gave an exotherm to about 30-35°C. dryness to afford the desired product 2014 as a pale brown oil 35 during this period and turned a greenish color. In a separate (8.21 kg, >100 yield). The "H NMR spectrum was consistent flask, compound 2017 (57.g., 0.131 mol) was azeotroped with with the assigned structure of compound 2014. toluene three times. Iodoform (102 g, 0.273 mol) and anhy Synthesis of Compound 2015: drous tetrahydrofuran (700 mL) were added to compound A multiple-neck, 72-L, round-bottom flask equipped with 2017 under nitrogen forming a yellow solution. The solution a mechanical stirrer, a temperature probe, and a nitrogen inlet 40 was charged with crude compound 2014 (8.21 kg, approxi was then added to the slurry of chromium chloride by cannula mately 11.7 mol) and DCM (15.3 L). The reaction mixture under nitrogen over 30 min while maintaining the internal was cooled to 0°C. and a solution of CSA (2.72 kg, 11.7 mol) temperature below 5°C. After 30 min, TLC showed comple in MeOH (15.3 L) was added at <10° C. The reaction was tion of reaction. The reaction was quenched portionwise with stirred at 0°C. for 2 hand analyzed by TLC (9:1 heptane/ 45 a mixture of sodium thiosulfate (10 wt %, 570 mL) and EtOAc) which indicated that the reaction was complete. The saturated sodium bicarbonate (570 mL) while maintaining reaction was quenched with TEA (1.18 kg, 11.7 mol) and the temperature below 15° C. MTBE (2000 mL) was added: concentrated to dryness. The residue was diluted in heptanes two phases were separated and the organic phase was washed (18.5 L) and washed with water (2x18.5 L). The organic layer with brine (1140 mL), dried over magnesium sulfate, filtered, was purified by silica-gel plug in four bathes (silica gel: 5 kg 50 and concentrated to give crude 2018 as agreen residue with an each) using heptanes to eluent the impurities and 4:1 hep E/Z-ratio of 8.8:1 by "H NMR analysis. The crude material tanes/EtOAc to eluent the desired product. The fraction con was stored in diethylamine (400 mL) in the freezer overnight. taining the product was concentrated to dryness to afford a pale brown oil (3.38 kg, 70% yield over three steps). The 'H Scheme 4. Preparation of Compound 1001 NMR spectrum was consistent with the assigned structure of 55 compound 15. Synthesis of Compound 2016: A multiple-neck, 72-L, round-bottom flask equipped with TMS Nu4nen a magnetic stirrer, a temperature probe, aheating mantle, and 2018 a nitrogen inlet was charged with compound 2015 (3.38 kg, 60 8.2 mol) and DCM (16 L). It was cooled to 0°C. and DMSO OH O Pol(PPh3)4, CuI, EtNH (1.93 L) and TEA (3.79L) were added. SOspyr (3.93 kg, 24.6 -e- mol) was added while maintaining the internal temperature 2 Oi-Pir 56% over 2 steps below 10° C. The reaction was stirred at 0° C. for 3 h and 2 from compound 2017 analyzed by TLC (9:1 heptane/EtOAc) which indicated that 65 H the reaction was complete. The reaction was washed with 2005 10% citric acid (2x5 vol) and the organic layer was concen US 9,315,447 B2 151 152 -continued OTBDPS OH O Scheme 5. Preparation of Compound 1002 TBAF, NHCI eace = Oi-Pir 84% 5 OH OH O 1s p-TsOH | MeOH MS 2019 | OH OH O 10 H 21a 21 1 OO1 Oi-Pir OH OH O

15 H 1001 H --- Synthesis of Compound 2019. 10O2 A 3-L, three-neck, round-bottom flask was degassed and purged under nitrogen. To the flask was added tetrakis(triph enylphosphine)palladium(0) (7.4 g. 6 mmol) and copper Synthesis of Compound 1002: iodide (2.5 g., 13 mmol). A degassed solution of compound Ta a solution of 1001 (35 g, 115 mmol) in MeOH (350 mL) 2005 (24 g. 131 mol) in anhydrous tetrahydrofuran (55 mL) 25 was added p-TsOH.H2O (4.38 g, 23 mmol). The resulting was added under nitrogen by cannula at ambient temperature. solution was stirred at room temperature for 22 hand diluted A degassed solution of compound 2018 (0.131 mol) in with EtOAc (700 mL) The mixture was washed with 5% diethylamine (400 mL) and anhydrous tetrahydrofuran (800 aqueous sodium bicarbonate (3x100 mL). The combined mL) was then added under nitrogen by cannula. The reaction aqueous were extracted with EtOAc (3x200 mL). The organ mixture was allowed to stir at ambient temperature overnight. 30 ics were combined and washed with brine (150 mL), dried At this point the reaction was complete by HPLC analysis. over Sodium sulfate, and concentrated under vacuum. The oil The reaction was quenched with saturated aqueous ammo residue was purified by column chromatography (2:3 EtOAc/ nium chloride (20 mL) and then concentrated to a residue. n-heptane) to give 1002 (22.8 g., 71.7%, 98.1% (AUC) by MTBE (700 mL) was added and the mixture was slurried for HPLC) as a light yellow oil. The "H NMR spectrum was 10 min; the solids were filtered over a packed bed of Celite on 35 a glass fritted funnel. The filtrate was washed with saturated consistent with the assigned structure of compound 1002 as is aqueous ammonium chloride (300 mL), dried over magne shown in FIG. 6. sium sulfate, filtered, and concentrated to give 2019 (89 g, >100%) as a crude brown oil which had an E/Z-ratio of 6.3:1 Scheme 6. Preparation of Compound Z by H NMR analysis which was purified by column chroma 40 tography on silica gel followed by CombiFlash chromatog raphy. OH OH O 1) LiOH, o THF/HO Synthesis of Compound 1001: o -e- A 5-L, three-neck, round-bottom flask was charged with 21N21 Oi-Pr 2) 2 Naq HCI intermediate 2019 (284.4g, 0.46 mol), ammonium chloride 45 (74.2 g, 1.39 mol), and THF (850 mL). The mixture was stirred and cooled to 5°C. using an ice bath. To the stifling 1001 mixture was added 1 Mtetrabutylammonium fluoride in THF NaOH, (1.4 L) via an addition funnel over 15 min such that the OH OH O EtOHF reaction temperature remained below 10°C. The mixture was 50 o HO stirred for 2 hat which point TLC analysis (40% ethylacetate/ 21N21 —- heptanes) showed no starting material remained. The reaction mixture was diluted with tert-butyl methyl ether (1.4 L) and saturated ammonium chloride (1.4 L) and the layers sepa 45 rated. The organic layer was washed with Saturated ammo 55 OH OH O nium chloride (1.4L), dried over Sodium Sulfate, and concen trated to afford an oil (271.4 g). The oil was suspended in 20% 21N 21 ONa heptanes/ethyl acetate (200 mL) and dichloromethane was slowly added until a solution was formed (approximately 20 mL). The mixture was purified on silica gel (2 kg) eluting with 60 20% ethyl acetate?heptanes (1.6 L), 30% ethyl acetate?hep tanes (8L), 40% ethyl acetate/heptanes (20 L), and 50% ethyl Compound Z acetate?heptanes (16 L). The pure fractions were combined and concentrated to afford 1001 (128.3g, 91% yield): HPLC Synthesis of Compound Z: 97.7% (AUC), t-13.3 min; the "H NMR spectrum was con 65 A 5-L, three-neck, round-bottom flask was charged with sistent with the assigned structure of compound 1001 as is 1001 (335 g, 1.1 mol), THF (5 vol), and water (5 vol). LiOH shown in FIG. 5. (52.7 g, 2.2 mol) was added and the reaction was stirred at US 9,315,447 B2 153 154 ambient temperature for 4 h and analyzed by TLC which indicated that the reaction was complete. The reaction mix O (I) ture was extracted with MTBE (2x5 Vol) and the aqueous R9 Ri ORf Rh ORe layer was adjusted to pH 4.4 using 2 NHCl (900 mL). It was extracted with MTBE (6x5 vol) and the combined organic Rs 4N41)a– Rsul E, layers were washed with water (5 Vol), dried over NaSO, diluted with ethanol (10 Vol), and concentrated to five vol umes. The resulting ethanol solution was cooled to 0°C. and 6 N NaOH (1 equiv) was added. The reaction was stirred for 1 h, concentrated to dryness, and dried under vacuum for 18 10 or a pharmaceutically acceptable salt thereof, wherein: h to afford a brown oil (277 g. 88% yield): HPLC 99.2% Re and Rf are independently selected from hydrogen, (AUC), t-10.1 min; the "H NMR spectrum was consistent alkyl, alkenyl, alkynyl, aryl, heteroaryl, acyl, aminocar with the assigned structure of compound Z and indicated the bonyl, alkoxycarbonyl, or silyl, presence of residual ethanol. The brown oil (277 g) was dissolved in water (2 vol), concentrated to dryness at 40°C., 15 E is a branched alkoxy; and further dried under vacuum for 18 h to afford the desired Rh and Ri are independently selected from hydrogen, product compound Z as a brown oil (279 g): HPLC 99.0% alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl or het (AUC), to 9.9 min; the "H NMR spectrum was consistent eroaryl; with the assigned structure of compound Z as shown in FIG. 7. Rs is selected from i-iv as follows: i) CHCH(R)CH, where R is hydrogen, alkyl, alk INCORPORATION BY REFERENCE enyl, alkynyl, perfluoroalkyl, aryl, heteroaryl, fluoro, hydroxyl or alkoxy; All publications and patents mentioned herein are hereby incorporated by reference in their entirety as if each indi 25 ii) CHC(R.R.)CH, where RandR, are each indepen vidual publication or patent was specifically and individually dently alkyl, alkenyl, alkynyl, perfluoroalkyl, aryl, or indicated to be incorporated by reference. In case of conflict, fluoro, or R and R7 are connected together to form a the present application, including any definitions herein, will carbocyclic or heterocyclic ring; control. iii) CHOCH, CHC(O)CH, or CHCH; or 30 iv) Rs is a carbocyclic, heterocyclic, aryl or heteroaryl EQUIVALENTS ring; and Rs and Rare independently selected from hydrogen, alkyl, While specific embodiments of the subject invention have alkenyl, alkynyl, perfluoroalkyl, alkoxy, aryl or het been discussed, the above specification is illustrative and not eroaryl, or Rs and R are connected together to form a restrictive. Many variations of the invention will become 35 apparent to those skilled in the art upon review of this speci carbocyclic or heterocyclic ring. fication and the claims below. The full scope of the invention 4. The pharmaceutical preparation of claim3, wherein E is should be determined by reference to the claims, along with selected from isopropoxy, isobutoxy, Sec-butoxy, tert-butoxy, their full scope of equivalents, and the specification, along 3-methylbutoxy, 2,2-dimethylpropoxy, or 1,1,2-trimethyl with Such variations. 40 propoxy. 5. The pharmaceutical preparation of claim 3 or 4, wherein The invention claimed is: the compound is represented by formula II, 1. A pharmaceutical preparation Suitable for treating or preventing an ophthalmic condition in a human patient, said preparation comprising: 45 (II) one or more of: ORf ORe Q a compound of any of formulae I-IX, a compound of formula A, 21\21n ls E, a compound of any one of formulae 1-49, a lipoxin compound, or 50 an oxylipin compound, and one or more pharmaceutically acceptable excipients, wherein the pharmaceutical preparation is characterized in or a pharmaceutically acceptable salt thereof. that 6. The pharmaceutical preparation of claim 5, wherein the a) the one or more of a compound of any of formulae I-IX, 55 compound is represented by formula III, a compound of formula A, a compound of any one of formulae 1-49, a lipoxin compound, oran oxylipin com pound is present in a concentration over 90 micromolar, b) the pharmaceutical preparation is a solution with a pH in (III) the range of 5.5 to 7.4, and/or 60 ORf ORe O c) the pharmaceutical preparation further comprises one or more surfactants. 2. The pharmaceutical preparation of claim 1, wherein the pharmaceutical preparation is Substantially free of preserva tives. 65 3. The pharmaceutical preparation of claim 1 wherein said compound of formula I has the structure: or a pharmaceutically acceptable salt thereof. US 9,315,447 B2 155 156 7. The pharmaceutical preparation of claim 1 wherein said compound is represented by formula 1001, 1001 OH OH O 1001 5 OH O 41 Nay– OiPr. OH

21N 21 S OiPr, \ \ " 12. The method of claim 11, wherein greater than 2 micro grams of the compound is administered to an eye per day. 13. The method of claim 12, wherein from 2 to 175 micro grams of the compound is administered to an eye per day. or a pharmaceutically acceptable salt thereof. 14. The method of any of claims 11-13, wherein the com 8. The pharmaceutical preparation of claim 1, wherein the 15 pound is administered once, twice, three times, or four times preparation is an aqueous Solution Suitable for topical admin- daily. istration to an eye, the Solution having a concentration of 15. The method of claim 11, wherein the ophthalmic con greater than 30 micrograms/milliliter of the compound. dition is dry eye. 16. A packaged pharmaceutical for delivering eyedrops to 9. The pharmaceutical preparation of claim 8 having a 20 an eye in need of treatment for an opthalmic condition, concentration from 30 micrograms/milliliter to 2000 micro- wherein the eyedrops comprise a grams/milliliter of the compound. 10. The preparation of claim 1, wherein the ophthalmic condition is dry eye. 1001 11. A method of treating or preventing an ophthalmic con- 25 OH OH O dition comprising administering a pharmaceutical prepara ti - - - - 21N 21 1On compr1S1ng: o OiPr, one or more of: \ a compound of any of formulae I-IX, 30 a compound of formula A p s a pharmaceutical preparation comprising: a compound of any one of formulae 1-49, one or more of: a lipoxin compound, or a compound of any of formulae I-IX, lipi d. and 35 a compound of formula A, an OXyl1p1n compound, an a compound of any one of formulae 1-49, one or more pharmaceutically acceptable excipients, a lipoxin compound, or an oxylipin compound, and wherein the pharmaceutical preparation is characterized in one or more pharmaceutically acceptable excipients, that 40 wherein the pharmaceutical preparation is characterized in a) the one or more of a compound of any of formulae that I-IX, a compound of formula A, a compound of any a) the one or more of a compound of any of formulae I-IX, one of formulae 1-49, a lipoxin compound, or an a compound of formula A, a compound of any one of oxylipin compound is present in a concentration over formulae 1-49, a lipoxin compound, oran oxylipin com pound is present in a concentration over 90 micromolar, 90 micromolar, 45 b) the pharmaceutical preparation is a solution with a pH in b) the pharmaceutical preparation is a solution with a pH the range of 5.5 to 7.4, and/or in the range of 5.5 to 7.4, and/or c) the pharmaceutical preparation further comprises one or c) the pharmaceutical preparation further comprises one more surfactants. or more surfactants k . . . .