US 2008 OO15352A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2008/0015352 A1 Piccariello (43) Pub. Date: Jan. 17, 2008

(54) METAL COORDINATED COMPOSITIONS CD7C 233/00 (2006.01) C07C 247/00 (2006.01) (76) Inventor: Thomas Piccariello, Blacksburg, VA C07D 205/00 (2006.01) (US) C07F I/08 (2006.01) C07F 15/02 (2006.01) Correspondence Address: C07F 15/06 (2006.01) REED SMITH LLP C07F 9/02 (2006.01) 2SOO ONE LIBERTY PLACE C7F 3/06 (2006.01) 16SO MARKET STREET C07F 15/04 (2006.01) PHILADELPHIA, PA 19103 (US) C07F II/00 (2006.01) C07D 495/00 (2006.01) (21) Appl. No.: 11/824,411 C07D 209/04 (2006.01) 1-1. C07D 217/00 (2006.01) (52) U.S. Cl...... 544/265; 546/155: 548/432: Related U.S. Application Data 548/953; 549/23: 549/59; 552/8; (63) Continuation-in-part of application No. 1 1/257,504, 556/110; 556/118; 556/138; filed on Oct. 24, 2005. 556/42; 556/57; 562/21562/433; 562/449; 562/456; 562/553; (60) Provisional application No. 60/621,747, filed on Oct. 564/155; 564/162; 564/165 25, 2004. (57) ABSTRACT Publication Classification A metal coordination complex of a biologically active moiety and a metal is disclosed. The complex confers to the (51) Int. Cl. biologically active moiety an improved performance which CO7D 473/00 (2006.01) can include potency, stability, absorbability, targeted deliv C07C 229/00 (2006.01) ery, and combinations thereof.

Base

// OH RO O 8 W P s W

2 P- O O

R = Rest of RNA molecule Base = Purine or pyrimidine

Inner sphere RNA:Magnesium Coordination Complex Patent Application Publication Jan. 17, 2008 Sheet 1 of 20 US 2008/OO15352 A1

FIG. 1 (Prior Art)

-Pr 4 Be i-Pr MQM N N

-Pr -Bu i-Pr FIG. 2 (Prior Art) Patent Application Publication Jan. 17, 2008 Sheet 2 of 20 US 2008/OO15352 A1

FIG. 3 (Prior Art)

FIG. 4 (Prior Art) Patent Application Publication Jan. 17, 2008 Sheet 3 of 20 US 2008/OO15352 A1

HDO-----Mg 21 H H 4 N1 RO \ O - Y- O tP\-b- N H N P / Base O2 \ 1. / N O : RO O HO, S p––o OR R = Rest Of RNA molecule Bage 9 Base = Purine or pyrimidine

Outer sphere RNA:Magnesium Coordination Complex

FIG. 5 (Prior Art) Patent Application Publication Jan. 17, 2008 Sheet 4 of 20 US 2008/OO15352 A1

Base O

OR // O 'OH RO e N W Old- s \u-OR O- O >e -Mg-o - N 2'-O O

R = Rest of RNA molecule Base = Purine or pyrimidine

Inner sphere RNA:Magnesium Coordination Complex

FIG. 6 Patent Application Publication Jan. 17, 2008 Sheet 5 of 20 US 2008/OO15352 A1

Base

R = Rest Of RNA molecule P = Rest of Peptide Base = Purine or pyrimidine

RNA:Magnesium: Arginine Coordination Complex

FIG. 7 Patent Application Publication Jan. 17, 2008 Sheet 6 of 20 US 2008/0015352 A1 H

O NHN2V

RN O 2.. HN NH /

Protective Polymer Shell FIG. 9 Patent Application Publication Jan. 17, 2008 Sheet 7 of 20 US 2008/OO15352 A1

-- H3C-N-- IM 9

O-CH FIG. 10

10 8 6 4 2 PPM Proton NMR of Bis(triiodothyroninato)- bis(dimethylsulfoxide)magnesium FIG 11 Patent Application Publication Jan. 17, 2008 Sheet 8 of 20 US 2008/OO15352 A1

9 8 7 6 5 4. 3 2 1 PPM Proton NMR of Triiodothyronine (T3) FIG. 12 (Prior Art)

10 8 6 4 2 PPM Proton NMR of Bis(triiodothyroninato)zinc FIG. 13 Patent Application Publication Jan. 17, 2008 Sheet 9 of 20 US 2008/OO15352 A1

7 6 5 4 3 2 PPM Proton NMR of Dimethylbiguanide:Zinc Complex FIG. 14

7 6 5 4. 3 2 1 PPM Proton NMR of Dimethylbiguanide FIG. 15 (Prior Art) Patent Application Publication Jan. 17, 2008 Sheet 10 of 20 US 2008/OO15352 A1

7 6 5 4 3 2 1 PPM Proton NMR of Tetracycline FIG. 16 (Prior Art)

8 7 6 5 4. 3 2 - PPM Proton NMR of Bis(tetracyclinato)magnesium FIG. 17 Patent Application Publication Jan. 17, 2008 Sheet 11 of 20 US 2008/OO15352 A1

7 6 5 4 3 2 PPM Proton NMR of Tetracycline-magnesium complex with 1 NHCl added. FIG. 18 Patent Application Publication Jan. 17, 2008 Sheet 12 of 20 US 2008/OO15352 A1

...) 7 6 5 4 3 2 ''PE Proton NMR of Tetracyline with 1 NHCl added.

FIG. 19 Patent Application Publication Jan. 17, 2008 Sheet 13 of 20 US 2008/OO15352 A1

8 7 6 5 4 3 PPM Proton NMR of Hydrochlorothiazide FIG. 20 (Prior Art)

8 7 6 5 4 3 2 PPM Proton NMR of Hydrochlorothiazide-Zinc Complex FIG 21 Patent Application Publication Jan. 17, 2008 Sheet 14 of 20 US 2008/OO15352 A1

--- 8 7 6 5 4 3 2 1 PPM Proton NMR of Hydrochlorothiazide-Zinc Complex with 1 NHCl added FIG. 22

| - 1 O 8 6 4 2 PPM Proton NMR Bis(acycloguanosinato)magnesium FIG. 23 Patent Application Publication Jan. 17, 2008 Sheet 15 of 20 US 2008/OO15352 A1

Bis(triiodothyroninato)-bis(dimethylsulfoxide)magnesium FIG. 24 Patent Application Publication Jan. 17, 2008 Sheet 16 of 20 US 2008/OO15352 A1

OH

HO

Bis(triiodothyroninato)zinc FIG. 25 Patent Application Publication Jan. 17, 2008 Sheet 17 of 20 US 2008/OO15352 A1

Bis(minocyclinato)magnesium FIG. 26

Bis(tetracyclinato)magnesium FIG. 27 Patent Application Publication Jan. 17, 2008 Sheet 18 of 20 US 2008/OO15352 A1

CH Dimethylbiguanide-zinc complex FIG. 28

NH

r s \ -O Q-M9, 2-()N >s \ N-o HN \- OH Bis(acycloguanosinato)magnesium FIG. 29 Patent Application Publication Jan. 17, 2008 Sheet 19 of 20 US 2008/OO15352 A1

T3Mg vs. T3/ZnO vs. T37n vs. T3, no outliers -0-T3, free acid --T3Mg -A-T3/ZnO

Hours Pharmacokinetic Profile of T3, T3Mg, T3Zn in a Rat Animal Model FIG. 30 Patent Application Publication Jan. 17, 2008 Sheet 20 of 20 US 2008/0015352 A1 0.5uM 1.0 uM 1.5uM 1.0 uM

Reo Ro A W A W A W A W C

lonic MRNA COValent M:RNA

|EF evaluation of magnesium and zinc iRNA complexes. F.G. 31 US 2008/OO15352 A1 Jan. 17, 2008

METAL COORDINATED COMPOSITIONS covalency, and this is particularly true with the transition 0001. This application claims priority from U.S. applica metals combined with nitrogenous ligands, in most cases it tion Ser. No. 1 1/257,504 filed on Oct. 24, 2005, which is a preferred embodiment of this invention that the active claims priority from provisional application No. 60/621,747, agent chelate with the metal, particularly if the metal is filed Oct. 25, 2004. magnesium.

FIELD OF THE INVENTION 0008. It is an embodiment of this invention that the active agents that make the best candidates for complexing with 0002 This invention relates to novel metal coordinated magnesium and calcium are those that have a proton on a complexes of biologically active molecules. heteroatom (i.e., oxygen, nitrogen or sulfur) with a pKa BACKGROUND OF THE INVENTION slightly greater than water or lower than water and have an additional heteroatom in close proximity to the first proto 0003. It is desirable to improve the properties of known, nated heteroatom Such that it can participate in the bonding, biologically active molecules by modifying their structures. or otherwise chelate, with the metal. Drugs that have this The goal of Such modifications is a molecule that is arrangement of functional groups are most likely going to improved in some way, such as potency, stability, reduced bond with a metal, where the resultant metal coordinated side effects, or targeted delivery. This improvement is active agent will be stable enough in a biological system and achieved without sacrificing the molecule's desirable prop survive hydrolysis therein, such that the performance of the erties. While this goal is easily stated, it is difficult to achieve active agent will be sufficiently modulated. This hydrolytic in actual practice, as the effects of any particular modifica stability imparted by multidentate ligands is Supported by tion is often highly unpredictable. the fact that they can lower the pKa’s of the ligand such that even amides can be deprotonated with weak bases, such as SUMMARY OF THE INVENTION triethylamine, in the presence of coordinating metals. There 0004. According to the current invention, the structure of fore, active agents with protons on heteroatoms, which known biologically active molecules is modified to result in normally would not be ionized in typical biological pH, can new molecules known as metal coordinated complexes. have the proton replaced with a covalently coordinated These new molecules have unexpectedly superior proper metal, where covalency is enhanced by the additional che ties. The metal coordinated complexes of the current inven lation from participating heteroatoms. It is a preferred tion include complexes of thyronine, tetracycline antibiotics, embodiment of this invention that at least one of the het oxycodone and hydrocodone, and complexes of their deriva eroatoms on the active agent that will bind to magnesium or tives. calcium be oxygen or Sulfur. Magnesium forms unusually 0005 Chelation is a critical component in the stabiliza strong bonds with phosphates and phosphonates and, there fore, it is an additional embodiment of this invention that the tion of a metal coordinated complex. For the s-block metals, active agent coordinated with magnesium is an organophos this is particularly true for calcium and magnesium. For phate or organophosphonate compound. example, it can be seen that the log K of the acetic acid-magnesium complex is 0.47. With the incorporation of 0009. It is an embodiment of this invention that the active a single amino group on the molecule (i.e., glycine) the log agents that make the best candidates for complexing with K, increased to 1.34. Magnesium typically prefers chela Zinc and the p-block metals are the same as those with the tion with oxygen over nitrogen and this effect can be seen by s-block metals with the additional flexibility that if the active comparing the log K of adenine (log K =2.08) with that agent has two nitrogens, a nitrogen and a mercaptain or two of 6-hydroxypurine (log K =6.65). Magnesium forms par mercaptains in a proper chelation arrangement, then the ticularly strong bonds with oxidized phosphorus, such as presence of a proton on a heteroatom is not necessary to phosphates, as is revealed by comparing the log K of form a stable metal coordinated complex. It is a further adenosine (log K =0.50) with that of adenosine-5'-mono embodiment of this invention that transition metals have phosphoric acid (log K =1.80). further ligation flexibility in that chelation is even less 0006. In general, zinc complexes are more stable then the required for their covalent coordination complexes if the comparable magnesium complexes. This is particularly true ligands have at least one nitrogen or mercapto group. if the ligand bears nitrogen or sulfur. (This may not be the case for ligands with oxygen only and even less So if the 0010. The active agents which are embodied in this ligand is a phosphate.) Using the glycine example above, the invention can be divided into chemical classes as shown in log K for the glycine-zinc complex is 4.85. The strength of Table 1 (actually they may be divided into combinations of the Zinc sulfur bond versus the oxygen bond is manifest in chemical classes to reflect the heterogenous chelation poten the relative log K values for the zinc complexes of hydrox tial). The drugs listed in Table 1 are not intended to be an ypropanoic acid (log K =0.86) and mercaptopropanoic acid exhaustive list of all drugs that satisfy the embodiment of (log K =6.43). Comparisons of log K values with other this invention but a representation of the chemical classes metals and ligands reveal that this chelation stabilization that exist in pharmaceuticals and that other pharmaceuticals prevails in metal coordination chemistry. that are of the same class listed in Table 1 or have arrange 0007 Whereas it may not be required that chelation occur ments of atoms that is satisfied by the embodiments of this to form a stable metal coordinated complex with inherent invention are also claimed by this invention. US 2008/OO15352 A1 Jan. 17, 2008

TABLE 1. Biologically active molecules that form coordination complexes in accordance with the Invention. Chemical Class or Functional Group Combination Therapeutic Classes Drug Examples Guanide or diamine Antidiabetic, AntiGERD, Metformin, Famotidine, Antineoplastic, Antiviral, Mitoxantrone, Adefovir, Antihypertensive Hydralazine, Zanamivir Amine or amide with GERD, Diuretic, Famotidine, Sulfonamide Antimigraine, Antidiabetic Hydrochlorothiazide, Sumatriptan, Glipizde, Glyburide, Torsemide Amine or amide with azole GERD, Antiviral, Lansoprazole, antimigraine, Antiurolithic, Zolmitriptan, Rabeprazole, Antihypertensive, Omeprazole, Analgesic, Anitemetic Esomeprazole, Ribavarin, Allopurinol, Clonidine, Granisetron Amine or amide with alcohol Antineoplastic, Antiviral, Mitoxantrone, Saquinavir, Bone resorption inhibitor, Alendronate, Albuterol, Antibiotic, Bronchodilator, Ephedrine, Epinephrine, Antithrombotic, Analgesic, Dipyramidole, Oxycodone, Antihypertensive, Oxymorphone, Anxiolytic, Anticonvulsant Tetracycline, Minocycline, Doxycycline, Labetalol, Lorazepam, Oxazepam 3-diketone, C-diketone, Antibiotic, Antineoplastic, Tetracycline, Minocycline, ketophenol, C.-ketoalcohol Antiinflammatory, Doxycycline, B-ketoalcohol Multiple sclerosis Mitoxantrone, Atovaquone, treatinent Betamethasone, Paclitaxel, Docetaxel, Methylprednisolone, Prednisone, Idarubicin 3-ketoacid Antibiotic Levofloxacin, Ofloxacin, Norfloxacin Ureide iviral, Tenofovir, Acyclovir, iparkinsonian, Cabergoline, Theophylline, Bronchodilator Valgancyclovir Amine or amide with acid An ihypertensive, Quinapril, Ramipril, Hormone replacement, Trandolopril, Enalipril iparkinsonian, Diuretic, Lisinopril, Thyroxine, ipsoriatic, Liothyronine, DOPA, ineoplastic, Furosemide, Methotrexate, irheumatic, Antibiotic, Penicillin, Amoxicillin, iepileptic, Cefotetan, Captopril, idepressant, Analgesic Gabapentin, Ketorolac Alcohol with azole giotensin II receptor LOSartan, agonist, Phosphonate or phosphate Bone resorption inhibitor, Alendronate, Etidronate, iviral Fosamprenavir Phosphonateor phosphate with A. iepileptic Fosphenytoin amide Diol or polyol Bronchodilator, Nutritional Albuterol, Epinephrine, Supplements, Contrast Myoinositol, Chiroinositol, imager odixanol Mercaptain with acid Antiasthmatic, Antibiotic Montelukast, Cefazolin, Cefotetan Mercaptain with amine or ipsychotic, Olanzapine, Captopril amide A. ihypertensive Amine with amide Hormone deficiency, Tabimorelin, Amoxicillin, ibiotic Loracarbef, Iodochlorohydroxyquin Alcohol with acid algesia, Cholesterol Salicylic acid, owering, Antihypertensive Atorvastatin, Mesalamine, inflammatory Pravastatin, Sitofloxacin, Trovafloxacin Dicarboxylic acid ineoplastic Pemetrexed Amine with N-oxide ialopecia agent Minoxidi Alcohol with Nitrites ibiotic Metronidazole Diene with alcohol, amine, iacne, Antineoplastic Retinoic acid, Fenretinde amide or acid Oligonucleotide (polyureide Gene therapy, Anti-AMD iRNA, Pegaptainib or polyphosphate) US 2008/OO15352 A1 Jan. 17, 2008

TABLE 1-continued Biologically active molecules that form coordination complexes in accordance with the Invention. Chemical Class or Functional Group Combination Therapeutic Classes Drug Examples Oligopeptide (polyamide) Immunosuppressant, Cyclosporin, Epoetin, Antianemic, Antiviral, Inteferon, Atrial Natriuretic Antineoplastic, Diuretic Peptide, Abarelix Oligosaccharide (polyol) Anticoagulant, Heparin, Acarbose, Antidiabetic, Antibiotic Gentamycin, Tobramycin GERD = Gastroesophageal Reflux Disease AMD = Age-related Macular Degeneration

0011. As illustrated in Table 1, a suitable biologically ered to be a triamine but since only two of the amino groups active moiety may have two functional heteroatom groups, are necessary for chelating with a metal, the guanide and each of which is capable of participating in the formation of diamine are grouped together as a single chemical class. a metal coordination bond. Further, as can bee seen from This is the same argument for the reason beta-diketone is Table 1 the two functional groups may be in a spatial grouped with beta-ketoalcohol, and diol is grouped with relationship to each other to permit chelation to the same polyol. metal atom by those coordination bonds. According to a preferred embodiment, the coordination bond forms a 4 to 8 0016 Compounds that may be used in the embodiments atom ring encompassing the metal and the heteroatom of the of the invention, such as having two functional heteroatom functional groups and the ring usually does not include a groups and are capable of forming stable metal coordination trans double bond. The biologically active moiety and metal bonds with a metal include, but are not limited to: Cladrib atom form a stable coordination complex as illustrated by ine, Acetalzolamide, Eliprodil, (R.S)-3-(2-carboxypiper (1) Table 1: (2) the examples set forth herein, such as azin-4-yl)-propyl-1-phosphonic acid (CPP), Ifenprodil, (R)- Triiodothyronine (T3); (3) the figures, such as FIGS. 7-10 4-oxo-5-phosphononorvaline (MDL 100453), and 24-29; (4) the discussion of stability herein; and (5) Dihydroxyphenylglycine, (S)-(+)-a-amino-4-carboxy-2-me general ring structure and Stability principles known to one thylbenzeneacetic acid (LY367385), Eglumegad of ordinary skill in this art, see for example, Martell, et al., (LY354740), (2S,2R,3R)-2-(2',3'-dicarboxycyclopropyl)g- Coordination Chemistry Reviews, (1994) 133:39-65. lycine (DCG), Remacemide, Fingolimod, Teriflunomide, Laquinomod, Azathioprine, CloraZepate, Lorazepam, 0012. The following discussion illustrates specific Temazepam, Rufinamide, Tiagabine, Progabide, Phenace embodiments within the general principles discussed above. mide, Lamotrigine, EthoXZolamide, Zonisamide, Etoposide, 0013 Whereas it may not be required that chelation occur Doxorubicin, Vorinostat (SAHA), Bicalutamide, 7-phenyl to form a stable metal coordinated complex with inherent 2.4.6-hepta-trienal hydroxamic acid, Goserelin, Naltrexone, covalency, and this is particularly true with the transition Fentanyl, Piritramide, Acadesine, Acarbose, Acebutolol, metals combined with nitrogenous ligands, in most cases it Acecarbromal, Acetylpheneturide, Acitretin, Adrafinil, is a preferred embodiment of this invention that the biologi Albendazole, Alexidine, Aliskiren, Alprenolol. Althiazide, cally active moiety chelate with the metal, particularly if the Alvimopan, Ambuphylline, Amcamprosate, Amfenac, metal is magnesium. Amidephrine, Amidinomycin, Amiloride, 4-Amino-3-phe nylbutyric acid, Aminophylline, Amlexanox, Amosulalol, 0014. The biologically active moieties that have two Amprenavir, Arotinolol, Atorvastatin, AZidamfenicol, functional heteroatom groups that are capable of participat Baclofen, Balsalazide, Bambuterol, Bamethan, Befunolol, ing in the formation of a stable metal coordination bond are Benzthiazide, Betaxolol, Bevantolol, Bisantrene, Bitolterol, embodied in this invention and include, but are not limited Brinzolamide, Bromfenac, Bromhexine, 5-Bromosalicylhy to the biologically active moieties set forth in Table 1. The droxamic Acid, Bucillamine, Bucindolol, Bucumolol, Bufe biologically active moieties listed in Table 1 are not intended niode, Bufetolol, Bufexamac, Buformin, Bufuralol, to be an exhaustive list of all biologically active moieties Bumadizon, Bunitrolol, Bupranolol, Buramate. Butanilic that satisfy the embodiment of this invention. The examples aine. Butazolamide, Butoctamide, Calcium N-Carbamoylas provided in Table 1 represent various groups of biologically partate, Capreomycin, Capuride, Carazolol, Carbazochrome active moieties that exist in pharmaceuticals or have Sodium Sulfonate, Carbimazole, Carisoprodol, Carmustine, arrangements of atoms that satisfy the embodiments of this Carteolol, Carticaine, Carubicin, Carvedilol, Catechin, invention. Chloraminophenamide, Chlorguanide, Chlorphenesin Car 0.015 The biologically active moieties listed in Table 1 bamate, Chlorproguanil, Chlorpropamide, Choline Alfoscer have the attributes that make them a member of the group in ate, Cidofovir, Clodronic Acid, Clonixin, Cloranolol, Clo which they are listed. They also have the functional groups razepic Acid, Clorprenaline, Closantel, Cynarine, that define that group and are in close enough proximity to Dacarbazine, Delapril, Delavirdine, Denopamine, Diazi each other to be able to chelate to a metal. Each group is quone, 3,5-Dibromo-L-tyrosine, Diclofenac, Didanosine, defined by the specific combination of functional groups Dideoxyadenosine, Digitalin, Digitoxin, Dioxethedrine, listed. For example, a guanide is a diamine but is also a Dobutamine, Docarpamine, Docetaxel, Dorzolamide, specific kind of diamine. A guanide can actually be consid Drotebanol, Droxidopa, Dyphylline, Ebrotidine, Ecabapide, US 2008/OO15352 A1 Jan. 17, 2008

Ecgonidine, Edatrexate, Eflornithine, Ellagic Acid, Endrala Mefenamic acid, 6-Mercaptopurine, Melagatran, Myco Zine, Enfenamic Acid, Entacapone, Epalrestat, Ephedrine, pheolate mofetil, Pregabalin, Quinocide, Rilmazafone, Taf Epinephrine, Erdosteine, Ergotamine, Eritadenine, Esapra enoquine, Tilarginine, Tolfenamic acid, Cidofovir, Zole, Etanidazole, Ethylmethylthiambutene, Etidronic Acid, Didanosine, Dideoxyadenosine, Etidronic acid, Moroxy Etodolac, Exifone, Fenbendazole, Fendosal, Fenethylline, dine, Nelfinavir, Pamidronic acid, Risedronic acid and Fenoldopam, Fenoterol, Fenpentadiol, Fentiazac, Fepradi Zoledronic acid. nol, Flavopiridol, Fludrocortisone, Flufenamic Acid, Flunixin, Fluocortolone, Fluvastatin, Formoterol, Fosfosal, BRIEF DESCRIPTION OF THE DRAWINGS Gancliclovir, Gentisic Acid, Glafenine, Glibornuride, Gli clazide, Glimepiride, Glipizide, Gliquidone, Glisoxepid, 0018 For the present invention to be clearly understood Glyburide, Glybuthiazole, Guanabenz, Guanfacine, Hydro and readily practiced, the present invention will be described cortisone, Isoetharine, Isoflupredone, Isoladol, LaZabemide, in conjunction with the following figures, wherein: Levobunolol, Lidamidine, Lopinavir, Lorazepam, Lorimetazepani, Lotrafiban, Mefenamic Acid, Meglutol, 0019 FIG. 1 illustrates the structure of Magnesocene in Melagatran, Melphalan, Mepindolol, 6-Mercaptopurine, accordance with the prior art. Metaproterenal, Methazolamide, Methisazone, Methocar 0020 FIG. 2 illustrates the structure of (Cyclopentadi bamol, Methoxamine, Methylergonovine, Metipranolol. enyl)-butylmethylbis(N,N'-(2,6-diisopropylphenylamidi Metoprolol, Midodrine, MitoguaZone, Mitoxantrone, nate)magnesium in accordance with the prior art. MivaZerol, Mizoribine, Modafinil, Mopidamol, Moprolol, Moroxydine, Mycophenolate mofetil, Nadolol, Nadoxolol, 0021 FIG. 3 illustrates the structure of magnesium: sali Nalbuphine, Nalmefene, Naloxone, Nateglinide, Nebivolol. cylaldehyde complex in accordance with the prior art. Nelfinavir, Nialamide, Nifenalol, Ninopterin, Nipradilol. 0022 FIG. 4 illustrates the structure of Magnesium Nitazoxanide, Nithiazide, Nolatrexed, Nordefrin, Norepi phthalocyanine in accordance with the prior art. nephrine, Norfenefrine, Norpseudoephedrine, Nylidrin, Octopamine, Omapatrilat, Onapristone, OraZamide, 0023 FIG. 5 illustrates an outer sphere RNA: magnesium Osalmid, Orotic acid, Orthocaine, Oseltamivir, Oxazepam, coordination complex in accordance with the prior art. Oxycinchophen, Oxyfedrine, Oxymorphone, Paclitaxel, Pamabrom, Pamidronic acid, Paramethasone, Penciclovir, 0024 FIG. 6 illustrates an inner sphere RNA: magnesium Penicillamine, Perfosfamide, Phenazopyridine Hydrochlo coordination complex in accordance with the present inven ride, Pheneturide, Phenformin, Phenylephrine Hydrochlo tion. ride, Phenyramidol, Phosphocreatine, Pindolol, Pipradrol, 0025 FIG. 7 illustrates an RNA:magnesium:arginine Pirarubicin, Pirbuterol, Piroxicam, Practolol, Prednylidene, coordination complex in accordance with the present inven Pregabalin, Prenalterol, Procaterol, Procodazole, Proglu tion. mide, Pronethalol, Propafenone, Propranolol, Protokylol, Pseudoephedrine, Quercetin, Quinocide, Raltitrexed, 0026 FIG. 8 illustrates a substituted arginine:magnesium Rebamipide, Rebeccamycin, Reproterol, Ribavirin, Ril complex in accordance with the present invention. mazafone, Risedronic Acid, Ritodrine, Romurtide, Rufina 0027 FIG. 9 illustrates salicylic acid and polymer bound mide, Salacetamide, Salicylamide, Sapropterin, Saquinavir, arginine complexed with magnesium in the inner sphere and Sivelestat, Sotalol, Soterenol, Stepronin, Tafenoquine, Tali peptides encapsulating the ligand.metal complex in the outer nolol, Taltirelin, Tegaserod, Temazepam, Temozolomide, Tenoxicam, Terbutaline, Tertatolol. Theophylline. Thiami sphere in accordance with the present invention. prine. Thioguanine, Tiaprofenic Acid, Tilisolol, Tilarginine, 0028 FIG. 10 illustrates a magnesium:oxycodone com Timolol, Timonacic, Tioclomarol, Tixocortol, Tocainide, plex in accordance with the present invention. Tolazamide, Tolbutamide, Tolcyclamide, Tolfenamic Acid, Toliprolol, Tolrestat, Torsemide, Tretoquinol, Triamcino 0029 FIG. 11 illustrates the proton NMR of Bis(triiodot lone, Tulobuterol, Ubenimex, Velnacrine, Vidarabine, hyroninato)-bis(dimethylsulfoxide)magnesium in accor Vigabatrin, Voglibose, Xamoterol, and Zoledronic Acid. dance with the present invention. Each compound listed above has other chemical properties 0030 FIG. 12 illustrates the proton NMR of Triiodothy that may require special conditions during the complexation ronine (T3) in accordance with the prior art. reaction. The list of biologically active moieties and examples set forth herein provide a general protocol for the 0031 FIG. 13 illustrates the proton NMR of Bis(triiodot biologically active moieties listed and those related therein, hyroninato)Zinc in accordance with the present invention. but minor modifications to the applicable general protocols may be necessary for specific drugs, but are within the 0032 FIG. 14 illustrates the proton NMR of Dimethyl ordinary skill in the art. biguanide:Zinc complex in accordance with the present invention. 0017 Preferrably, compounds that may be used in the embodiments of the invention, such as having two func 0033 FIG. 15 illustrates the proton NMR of Dimethyl tional heteroatom groups and are capable of forming stable biguanide in accordance with the prior art. metal coordination bonds with a metal include, but are not 0034 FIG. 16 illustrates the proton NMR of Tetracycline limited to: Acetalzolamide, Vorinostat, Aliskiren, Alvimo in accordance with the prior art. pan, Bicalutamide, Baclofen, Balsalzide, Brinzolamide, Chlorproguanil, Diclofenac, Dorzolamide, Droxidopa, 0035 FIG. 17 illustrates the proton NMR of Bis(tetracy Clonixin, Ebrotidine, Enfenamic acid, Ethylmethyltham clinato)magnesium in accordance with the present inven butene, Etodolac, Flufenamic acid, Fosfosal, Lazabemide, tion. US 2008/OO15352 A1 Jan. 17, 2008

0036 FIG. 18 illustrates the proton NMR of Tetracycline AG=-RTInK Equation 1 magnesium complex with 1 NHCl added in accordance with 0.052 Where AG is the Gibbs free energy and indicates the present invention. the thermodynamic stability of the compound. The more 0037 FIG. 19 illustrates the proton NMR of Tetracycline negative AG is the more stable the compound. R is the gas with 1N HCl added in accordance with the present inven constant, T is the absolute temperature and K is the equi tion. librium constant. The equilibrium constant is expressed as a ratio of products over reactants. In the case of coordination 0038 FIG. 20 illustrates the proton NMR of Hydrochlo compounds for the reaction: rothiazide in accordance with the prior art. M+xLe XML 0039 FIG. 21 illustrates the proton NMR of Hydrochlo 0053 K is expressed in equation 2: rothiazide-Zinc complex in accordance with the present invention. K-MLMMILL Equation 2 0054 Thus the increasing thermodynamic stability of a 0040 FIG. 22 illustrates the proton NMR of Hydrochlo compound is directly related to the increasing value of the rothiazide-Zinc complex with 1 NHCl added in accordance equilibrium constant. with the present invention. 0055. In certain cases it is advantageous to express the 0041 FIG. 23 illustrates the proton NMR Bis(acyclogua equilibrium constant in terms the dissociation potential of a nosinato)magnesium in accordance with the present inven tion. metal-ligand bond. The reaction is thus: 0.042 FIG. 24 illustrates the structure of Bis(triiodothy rominato)-bis(dimethylsulfoxide)magnesium in accordance 0056. The dissociation constant, K is shown in equa with the present invention. tion 3: 0043 FIG. 25 illustrates the structure of Bis(triiodothy K=MILL/ML Equation 3 rominato)Zinc in accordance with the present invention. 0057 Whereas it is commonly accepted that ionic bonds nearly completely dissociate in Solution, most covalent 0044 FIG. 26 illustrates the structure of Bis(minocycli bonds, do not dissociate at all. Thus, determining bond nato)magnesium in accordance with the present invention. strength in solution, through measurement of the dissocia 004.5 FIG. 27 illustrates the structure of Bis(tetracycli tion constant or the more commonly expressed parameter nato)magnesium in accordance with the present invention. the equilibrium constant, is a method of discerning the bond type. For coordination compounds, which involve bonding 0046 FIG. 28 illustrates the structure of Dimethylbigu between metals and ligands from Groups 15-17, the ther anide-Zinc complex in accordance with the present inven modynamic stabilities have not been firmly established. tion. 0058 Examination of the literature reveals that cova 0047 FIG. 29 illustrates the structure of Bis(acyclogua lency of organometallic bonds can be determined from nosinato)magnesium in accordance with the present inven spectroscopic data (i.e. NMR and MS), ab initio molecular tion. mechanics calculations or a combination of the two. In 0.048 FIG. 30 illustrates the relative pharmacokinetic general, covalency is most likely to occur with transition metals, with nitrogen and Sulfur ligand atoms (in preference profile of T3, T3Mg, T37n in a rat animal model in accor to oxygen) and with increasing bond order or haptivity dance with the present invention. (designated “m) from the ligand. Organometallic com 0049 FIG. 31 illustrates the IEF profiles of magnesium pounds with ligands that having multiple haptivities are and zinc iRNA complexes in accordance with the present described as chelates. Amongst the metals in groups 1 and invention. The rows labeled “A” refer to complexes pre 2, the so called S-block main group elements, only beryllium pared in anhydrous conditions. The rows labeled 'W' refer and magnesium are considered to be important chelate to complexes prepared in water. forming elements.

DETAILED DESCRIPTION OF THE 0059 Recent research, using ab initio theoretical calcu INVENTION lations, has further qualified the nature of the coordination bond in terms of the ionic vs. covalent nature of the 0050 Chemical bonds exist in three basic forms: ionic, ligand-metal bond. Pierloot applied the CASSCF (complete covalent and coordination or the so-called Werner com active-space self-consistent field) model to a series of plexes, which are typically larger than inorganic metal salts. Werner complexes to measure the degree of covalency of (It should be pointed out that Werner complexes are con these organometallic complexes. Her general conclusions sidered to have neutral ligands.) The differences between the were that a trend exists wherein the static correlation energy, three bond types can be attributed, in part, to the thermo obtained from the CASSCF calculations, correlates well dynamic stability of the bond, particularly in solution. with covalency of the metal-ligand bond. She further con Conversely, the stability of a compound can be expressed as cluded, that for the same metal, the metal-ligand covalency the propensity for the atoms of the molecule to separate or and related correlation effects increase in the following order dissociate in Solution. of ligands: 0051. The thermodynamic stability of a compound is expressed in terms of it free energy of formation according 0060. This was in agreement with the nephelauxetic to equation 1: effect described by Jorgensen. The magnitude of this effect US 2008/OO15352 A1 Jan. 17, 2008

was directly correlated to the reduction of the interelectronic argument that main group metallocenes have significant repulsion of a transition metal upon coordination in a ligand covalent bond elements has been made as well. The lack of field. This reduction depended on the ligands and was d-orbital participation in the metal-ligand bonding may expressed as the ratio of the Racah parameter, B, in the reduce the stability of the compound but does not preclude complex and in the free metal ion (B=Benie/Bs). The the notion that main group metals, particularly magnesium, reduction in B resulted from a decrease in electron-electron can form bonds with ligands that are more covalent than repulsion of the free metal ion after ligands were added to ionic. It is generally known that the formation of 6-coordi form the metal complex; a large reduction in B indicates a nate magnesium complexes upon their crystallization is due strong nephelauxetic effect. Thus ionic ligands, such as F. to spid hybridization. So it is conceivable that in certain give a small reduction in B and have larger f3 values. Based situations even the d-orbitals of magnesium can participate on spectroscopic measurements, ligands were ordered in bonding of the coordination complex. according to decreasing B values generating the nephelaux etic series: 0066 One can see a large difference in stability when comparing the equilibrium constants of a Werner complex (CO)? NCSS-Cls-CN >Bris-N-I-S? with a metal halide. For example, log KMidi-2.08 and s(CHO)PS’ >diarsine log KMc12s-1.0. Keeping in mind that pyridine is a neutral ligand; this difference in log K can only be due to the 0061 The correlation of the two series is supported by covalency of the magnesium-pyridine bond vis-a-vis the the similarities between the effects that both techniques ionic nature of the magnesium-chloride bond. Another described. In the former case, the CASSCF calculation example of a stable magnesium complex is seen with gauges the contribution of the metal d-orbital to the metal magnesium-Salicylaldehyde (SA) complex, with a log KM ligand bond. Racah parameter reduction by complex forma sA2=6.80 (FIG. 3). The stability of this bond is remarkable tion (B1) is caused by delocalization of the transition in that the ligand bonding atom is oxygen, which typically metal d-orbital electron cloud on the ligands, which is tend to form ionic bonds with metals. However, the exist indicative of covalent bond formation. ence of chelating oxygen stabilized the complex beyond 0062 Bonding between Li" and Be" with Cp ligands is what a pure ionic bond would do. Although nitrogen will mostly ionic due to the low energy state of the contributing form stronger bonds to magnesium than calcium, typically metal bond relative to the Cp bond. In addition, the ionic oxygen is a stronger chelator of magnesium than nitrogen. radius of these elements is too small to allow more than one 0067. The equilibrium constants of chelates are typically Cp ligand to bond. very large (e.g., log K for magnesium ethylenediamine-N, 0063. Theoretical calculations of magnesocene (FIG. 1), N'-disuccinate complex is 6.09) and may not reveal the Cp2Mg, reveal that the structure of the compound resembles extent of covalency between the neutral part of the ligand that of Cp Ca, CpSr and Cp2Ba but the d-orbital popula and the metal. However, it is the equilibrium constant that tions of Mg were found to be negligible in CpMg. How dictates the stability of any coordination compound and that ever, the Mulliken charge for Mg in CpMg using the is an important criterion for determining the nature of the density functional theory (DFT) model predicted 0.66; a chemical entity and how it will perform in particular appli value close to 2 is expected for a compound with a large cations. The existence of a covalent bond within the com dissociation constant such as MgCl2. This is in agreement plex and its contribution to the stability of chelates can with a paper by Faegri, Almlöf and Lithi, who, according to explain their very large log K and may also contribute to ab initio MO-LCAO calculations conclude that the charge the rigidity of the molecular structure. It should be pointed separation of magnesocene is only slightly higher than that out that, in many cases, covalency is the most important of ferrocene (Cp2Fe), a known covalent coordination com contributor to the stability of a coordination complex. pound. These data would suggest that the Mg-Cp bond is 0068 The magnesium porphyrin complexes or chelates somewhat covalent. The Cp moiety contributes its covalent are likely the most well known organomagnesium com bonding partly from its negative charge and partly from pounds; chlorophyll is a magnesium porphine. Phthalocya II-bonding from the double bonds. This combination of nine is a porphyrin representing the basic elements of that anionic and II-bonding with metals would also occur in class of compound and is used extensively as a model retinoic acid and its analogs. Thus forming metal coordi system to study metal-porphyrin bonds. It has been deter nated compounds of retinoic acid is an embodiment of this mined that transition metals form complexes with phthalo invention. cyanine (FIG. 4) very easily but because alkali and alkali 0064. The reactivity of CpMg and related Mg-Cp com earth salts dissociate so completely in water and other protic pounds was studied by Winter, et al., and they found that solvents, no solvent has been found, so far, which is suitable magnesium forms stable bonds with amidinate ligands. for direct introduction of Li", Na', K", Sr" and Ba" from Perhaps most telling was the stability of CpMg(m- solutions of their salts. As predicted from the ease of "BuC(N(2,6-PrCH)))), which was sublimed unchanged complexation of Mg" and Be", only these two s-block with 80% recovery at 180° C./0.05 torr (FIG. 2). Thus an elements along with Ca", can be directly introduced into example of a stable magnesium-amidinate compound has phthalocyanine, which is typically done from their iodide or been reported, which provides further support to the cova perchlorate salts in pyridine. lency of Such compounds. This is important for compounds that contain amidinate functionalities such as the purine and 0069. The synthesis, structure, stability and physical arginine containing compounds. properties of metal-porphyrin complexes have been well studied. The structure and the physical properties of mag 0065. Whereas it is commonly accepted that transition nesium phthalocyanine have been further elucidated using a metallocenes have strong covalent character, a legitimate variety of techniques, most recently, near-IR absorption and US 2008/OO15352 A1 Jan. 17, 2008

X-ray crystallography. The recurring conclusion is that the ligands attached to the metal, impact the structure and the magnesium-porphyrin chelate represents an extremely stability of the coordination complex. stable example of a metal coordinated compound. 0075 Additional ligands, other than the drug, can stabi 0070 Certain magnesium-ligand complexes are indeed lize the metal-drug complex. For example the K of the covalent in nature and not ionic and thus are new compo glycine (G) magnesium bond is 1.34. If, however, salicyla sitions of matter and not merely new salt forms. Metal ldehyde is added to the complex, the equilibrium for the organic ligand compounds, covalency in the nature of their reaction bonds. 0071. It is an embodiment of this invention that the formation of a coordination complex is favored when the is 4.77. Clearly salicylaldehyde adds a stabilizing effect to ligand has direct bonding opportunity to the inner sphere of magnesium glycine bond. It is an embodiment of this the metal, preferably magnesium. This is accomplished by invention, that adjuvants, like Salicylaldehyde are incorpo using anhydrous magnesium and non-protic solvents (or if rated into the drug: metal complexes to impart beneficial the solvent is protic it should be bulky). This concept is physicochemical properties. It is a further embodiment of supported by the fact that the catalytic reactivity of a metal this invention that the benefit of adjuvants is to stabilize the ion is reduced in its hydrated form. Complex formation in drug: metal complex in certain environments, such as in aqueous systems is a delicate balance between hydrogen aqueous solutions. bonds between ligand and water and the competition for 0076. There are very few examples of coordination com binding sites on the metal by hydration and complexation plexes with transition metals found in the Physician’s Desk capability of the ligand. It follows that complexation of a Reference (“PDR) and include 1) insulin modified by zinc: ligand with the inner sphere of metal is also reduced in 2) carboplatin contains platinum; 3) niferex is a polysac aqueous systems. It further follows that the converse is charide-iron complex; 4) pyrithione Zinc, used as the active true—that is, the rate of chelation or complexation of metals ingredient in anti-dandruff shampoo. In addition, some nutri with ligands in non-aqueous systems is accelerated vis-a-vis tional Supplements are described as complexes. Chromium aqueous Systems. picolinate is one example, where three picolinic acid groups 0072 A composition comprising an organic active agent are bound to a single Cr' in an octahedron (the nitrogens bound to a metal as a stable metal-ligand coordination provide the three other binding sites). Whereas it is an compound with inherent covalency is as a new molecular embodiment of this invention that the metal is selected from entity. In another preferred embodiment of the invention, the the group representing transition metals in a more preferred metal is selected from the main group elements. In yet a embodiment of the invention the metal is selected from the further embodiment of the invention, the metal is selected S-block main group elements, groups 1 and 2. In a most from the s-block elements. In a preferred embodiment of the preferred embodiment of the invention the metal is magne invention, the metal is magnesium. S1. 0073. Furthermore, it is an embodiment of this invention 0077. The patent literature cites some examples of novel that virtually any drug-magnesium complex with a K>1.0 magnesium coordinated drugs, which include: 1) Trilisate(R) has enough inherent stability to modulate the pharmacoki a stable, Solid choline magnesium salicylate composition netics of dissolution, absorption, distribution, metabolism mentioned above for treating arthritic pain; 2) magnesium and excretion. Given that the dissociation constant of salts of 2-descarboxy-2-(tetrazol-5-yl)-11-desoxy-15-substi Mg(OH), is -11.5, it is not surprising to discover that most tuted-omega-pentanorprostaglandins imparting greater tis magnesium complexes are much more stable in alkaline Sue specificity and ease of purification and compounding conditions than in acid. Thus the stability of the drug into medicaments; 3) magnesium Vanadate with insulinomi magnesium complex in the Small intestines is likely to metic properties with utility in treating insulin resistance modulate the pharmacokinetics of drug absorption. For syndromes; 4) a crystalline magnesium-taurine compound those metal-drug complexes that are acid labile, it is an for treatment of thrombotic or embolic stroke and prophy embodiment of this invention that protection of the complex lactic treatment of pre-eclampsiadeclampsia and acute car from the acidic milieu of the stomach be accomplished by a diac conditions; 5) the magnesium omeprazole'salt” deriva coating or encapsulation material that releases the complex tives mentioned above to treat GERD. upon entry into the small intestines. It is a further embodi 0078. The science of pharmaceuticals salts is a well ment of the invention that the encapsulation agent is a ligand studied area and selection of the salt form can impact a given or group of ligands forming an outer coordination sphere. pharmaceutical's performance. Examples of effects that the 0074 Another important concept of this invention is that salt form can have on a drug include dissolution rate, simple combinations of metal with ligands in solution do not solubility, organoleptic properties, stability, formulation always produce the same product. It is recognized that effects, absorption modulation and pharmacokinetics. The several, if not many, patents claim various salts as dependent periodical, Drug Delivery, published an article in October, claims without any Support in the Subject matter. This is 2003, citing three excipient applications using metals, pre accepted because the salt of an organic acid is easily Sumably forming salts, to stabilize pharmaceutical agents. prepared by treating it with a base and a metal salt where the Human Growth Hormone is complexed with zinc to reduce expected product is the metal salt of the organic acid; a its hydrophilicity and thereby slow the drug release; stabi method known by anyone skilled in the art. However, when lization of proteins against the acidic environment produced coordination chemistry contributes to the bonding between by degradation of encapsulating polymers was accom the organic acid and the metal, a variety of conditions, such plished by adding magnesium hydroxide to the formulation; as solvent, temperature and, perhaps most importantly, Zinc carbonate was used to stabilize Vinca alkaloids from US 2008/OO15352 A1 Jan. 17, 2008 acid hydrolysis. Whereas these products clearly use metals rin. As stated earlier chlorophyll is a porphine structure to stabilize the pharmaceutically active agent, the latter two Surrounding magnesium. Some enzymes require metals in do not claim to have modified the structure of the active order for them to be active. That is the reason why trace agent. metals, such as copper, Zinc, chromium, etc. are important 0079. Further it is well established that by simply chang for proper nutrition. Even some antibodies have transition ing salt forms, the pharmacokinetics of absorption in the metals associated with them. The metal is required for Small intestines is significantly modulated. For example, the enzyme activity due to the metal locking the peptide struc chemical structures of both the phosphate salt of tetracycline ture of the enzyme in a conformation through the formation and tetracycline hydrochloride differ in that portion of the of a coordination complex. salt form which is not the pharmacophore and one would 0091. The concept of incorporating computer aided expect that the relative physical properties of each would not design of drugs has gained popularity in recent years. This have a great influence on their relative bioavailabilities. But technique, which has been referred to as in silico, has in fact, the phosphate salt is absorbed twice as much as the developed to the point that through the understanding of hydrochloride salt. Conversely, the bioavailability of the free allosteric, coulombic and non-covalent interactions between acid of warfarin is nearly equivalent to its sodium salt, which the Substrate and the receptor, lead drug candidates have is unexpected because the dissolution rate of the warfarin been identified by computer modeling, before any material sodium tablet is 350 times faster than the tablet containing has been produced. It is an embodiment of this invention that the free acid. If different salt forms can confer such changes by including metal coordination in the computer simulated in the kinetics of absorption, than a complex with an S-block molecule, new and improved lead compounds can be iden main group element may have even a more pronounced tified. It is a further embodiment of this invention that in effect on absorption. silico derived lead compounds will have altered docking thermodynamics when the incorporation of a metal as a 0080. It is known that the bioavailability of tetracycline complex of the lead compound is included in the calcula antibiotics is mainly influenced by the physicochemical tions. It is a yet further embodiment of this invention that properties of their metal complexes that will most likely compounds previously removed from consideration based form in the GI tract. This is clearly an indication of drug on unsatisfactory in silico analyses will become important :metal bond formation in vivo. The formation of a covalent lead compounds when reanalyzed with the incorporation of bond between a drug and a metal in Vivo is even a more a metal complex into the calculations. It is a preferred reasonable expectation when the drug contains nitrogen and embodiment of this invention that the metal used for the the metal is in Groups 10-12 (e.g., nickel, copper, Zinc). It is revised in silico calculations as described above are selected an embodiment of this invention that modulation of a drugs from the main group elements. In a more preferred embodi performance is imparted by a formulation of the drug and the ment of this invention the metal is selected from the s-block metal, which will facilitate formation of a stable complex main group elements. It is a preferred embodiment of this between the drug and the metal. invention that the metal is magnesium. 0081. It is an embodiment of this invention that the 0092. In terms of direct application of the complex to a following benefits can be conferred upon a drug when biological system, it is an embodiment of this invention that complexed or coordinated with a metal: active agents that require ligand-receptor binding are 0082) 1. Improved water solubility, which can equate to imparted enhanced biological activity by virtue of the active better bioavailability (see discussion below); agent's conformational structure being locked in place through complexation with a metal. The receptor can be 0.083 2. Enhanced lipophilicity for improved absorption membrane-associated, within the cytoplasm or circulating in through the cell membrane; the body. It is an embodiment of this invention that metals 0084 3. Locking the pharmacophore into a conformation be incorporated into injectable drugs to lock the drug into a for improved receptor binding: conformation that provides optimum interaction with its target receptor. It is a preferred embodiment of this invention 0085 4. Ameliorating formulation problems due to poly that the metal be considered safe for injection. It is yet an morphism (see discussion below); even further embodiment of this invention that the metal be 0.086 5. Acid absorption properties for protection from selected from the list of aluminum, bismuth, magnesium, degradation in acidic environments, such as the stomach; calcium, iron or zinc. It is yet a further preferred embodi ment of this invention that the active agent is selected from 0087 6. The stability of the coordination complex may the list of injectable drugs, including, but not limited to, infera delay in absorption of the active pharmacophore. This vaccines, antineoplastics, antidiabetic drugs, and antisense is important for drugs like liothyronine, where rapid absorp RNA or other metabolic modulators. tion of the drug increases toxicity potential. 0093. The metal coordination technology of this inven 0088 7. Bioadhesion properties for sustained absorption tion could also advance current research in vaccine design. of the active pharmacophore; For example, a new cancer vaccine being developed com bines a lipoprotein adjuvant, a peptide antigen with a car 0089 8. Prevent abuse of narcotic analgesics by binding bohydrate antigen specific for cancer cells. The three com the pharmacophore of the narcotic through an organometal ponents of the vaccine construct are joined together lic complex to render the narcotic inert unless ingested. covalently via linkers. This method of constructing the 0090 Nature provides many examples of how transition vaccine is common in bioconjugate chemistry. Metal coor metals are transported, stored and utilized. Perhaps the most dination can be used as a scaffold to bind the different well known example is hemoglobin, which is iron porphy components of a bioconjugate such as Pegaptainib, whose US 2008/OO15352 A1 Jan. 17, 2008 combined components are an aptamer, polyethylene glycol viral vectors or plasmid vectors. The latter technique is and a lipid. It is an embodiment of this invention that the particularly interesting in that the plasmid vectors cause the components of a bioconjugate can be combined in a single siRNA to adopt a “hairpin' structure and these iRNA molecular entity by complexing each component to a central variants have been given the name of short hairpin RNA or metal. It is a further embodiment of this invention that metal shRNA. These shRNA molecules have enhanced silencing coordination serves as a general technique in bioconjugate capacity. Moreover, there is a body of evidence that suggests chemistry. that transfection agents are not necessarily required for the 0094) Of particular note is the remarkable affinity that siRNA molecules to enter the cell. The recent discovery of magnesium has for nucleic acids. With the advent of anti pulmonary applications of siRNA and viroids are two sense RNA, interference RNA and aptamers as therapeutic reported phenomena wherein naked RNA can enter the cell agents it will be increasingly important to incorporate deliv and silence gene products. As a matter of fact, it is well ery technologies for these nucleic acid drugs. Some of the known by scientists in this field that the secondary structure drug delivery techniques that are currently being investi of the RNA does not seem to impact its gene silencing gated include pegylation, liposomes or anionic clays. Inter effects. estingly, a recent study released by Howard Hughes Medical 0099 RNA is an oligonucleotide with multiple phosphate Research Lab indicated that montmorillonite clay facilitated groups. Magnesium forms very strong bonds with phos entry of RNA into lipid vesicles. There are a variety of clays phates and so RNA-Mg complexes are likely to have the that vary in the amount of alumina, silica, magnesia, iron magnesium atoms bound to the phosphate groups. By com and potassium. Thus, forming a magnesium complex of bining magnesium with RNA under anhydrous conditions, a RNA may facilitate RNAs entry into vesicles, which are covalent bond is formed, which, theoretically, would considered to be a laboratory model of cellular membranes. increase the lipophilicity of that portion of the RNA mol 0.095 The efficacy of the magnesium-nucleic acid com ecule. Furthermore, the magnesium center can bind multiple plex can be evaluated vis-a-vis the nucleic acid alone using phosphate groups, theoretically, causing the formation of the in silico techniques described above. Thus it becomes an hairpin structure mentioned above. This hairpin structure important embodiment of this invention that nucleic acid would not only manifest a lipophilic residue but would also drugs' efficacy is enhanced by their coordination with met provide greater resistance to attack from nucleases, which als. It is a further embodiment of this invention that the would lead to greater stability. nucleic acid be combined with a metal to form a coordina 0100 Since RNA would have other phosphate groups in tion complex prior to administration. A significant portion of excess of what is bound to magnesium, that portion of the the complexes in simple combination of a metal salt with a RNA molecule would retain its water solubility. This novel nucleic acid in aqueous systems will be outer sphere coor form of RNA would have the desired amphiphilic properties dinated ligands (FIG. 5) and may not provide the optimum that are important for mass transfer (hydrophilicity) and conformation for receptor binding, particularly for mem absorption (lipophilicity). For further discussion on this brane transport applications. A major premise of this inven point see the “Improved solubility” section below. tion is that metal-ligand complex structure is impacted significantly if the ligand has the opportunity to be an inner 0101. A typical process would entail combining RNA sphere ligand (FIG. 6) in preference to being an outer sphere with a magnesium salt in an anhydrous solvent. A Suitable ligand. solvent may be DMSO or perhaps an ionic liquid. An advantage of ionic liquids is that recovery of the magne 0096. It is a further premise of this invention that inner sium-RNA complex would merely entail adding the solution sphere ligand formation is promoted by using anhydrous to an ionic liquid miscible non-solvent such as alcohol (or in conditions to prepare the metal-ligand complex. It is an some cases Supercritical CO may work), where the desired embodiment of this invention; therefore, that the metal product would precipitate out. The ionic liquid could then be ligand complex is prepared under anhydrous conditions and recycled for the next reaction by distilling off the alcohol. that reconstituting the complex in water will produce a coordination complex with greater covalency, greater sta 0102) The above process would likely be applicable to bility, greater cell permeability and modulated biological any water soluble biologically active agent. Thus it is an performance relative to the complex prepared in water. This embodiment of the invention that the biologically active system is very amenable to incorporating adjuvants, such as agent is any saccharide, peptide or nucleotide. In a preferred polyarginine to enhance transfection efficiency, within the embodiment of the invention the biologically active agent is inner coordination sphere (FIG. 7). a nucleotide. In a more preferred embodiment of the inven tion the biologically active agent is an antisense RNA, 0097 Perhaps the most important development in recent interference RNA or an aptamer. It is a preferred embodi times in gene therapy was the discovery and advanced ment of the invention that the metal is selected from the research on interference RNA (“iRNA). Unlike antisense main group elements. It is a further preferred embodiment of RNA, iRNA is recycled by the cell's biochemical machinery the invention that the metal is selected from the s-block main to further silence gene products encoded by mRNA. This group elements. It is recognized that magnesium binds to results in increased efficiency of the gene silencing. The nucleic acids more tightly than calcium, thus it is a most major problems associated with iERNA include their perme preferred embodiment of the invention that the metal is ability into cells and their stability, particularly in the magnesium. presence of nucleases. Improved Solubility/Permeability 0098. These problems have been addressed by the incor poration of pulmonary surface active material ("SAM), 0103) In quantifying drug absorption it is useful to apply lipid or amine based transfection agents, electroporation, the term bioavailability. This is defined as the fraction (F) of US 2008/OO15352 A1 Jan. 17, 2008

the dose that reaches the systemic circulation. Thus, in the this invention to bind the active agent to a transition metal extreme cases, F=0 in drugs which are not absorbed at all in or alkaline earth metal to form a new composition of matter the GI tract while for drugs that are completely absorbed that has improved solubility while retaining its permeability. (and not metabolized by a first pass effect) F=1. The bio Since the new metal ligand bond is covalent, it is preferable availability can be calculated from the area under the curve that the metal have additional ligands (e.g., amino acid) (AUC) of the serum level vs. time plot. It depends on many attached to it to counterbalance the lipophilic nature of the factors and some of these factors differ between normal newly formed covalent metal center. individuals. In terms of bioavailability, drugs have been 0.108 Due to the covalent nature the stability of the classified into four categories according to the table below. metal-active agent complex is retained up to transport to the water film coating of the brush border membrane. When the complex reaches the membrane the metal and the drug are Class Solubility Permeability Bioavailability Expectation separated by the lipids in the membrane accepting the lipophilic active agent and rejecting the hydrophilic metal. I High High Very high bioavailability but is rare due to the requirement for active This is imparted through physicochemical action and, in transport contrast to the earlier methods of increasing solubilities of II Low High Reasonable bioavailability if drugs, does not require enzymes. solubility problem is not too severe III High Low Low permeability is difficult to 0.109 Drugs are applied to the skin to elicit an effect to overcome and drugs may be shelved for the 1) skin Surface, 2) an effect within the stratum corneum, this reason. 3) an effect requiring deeper penetration into the epidermis IV Low Low Very low or no bioavailability. Drugs in this class are usually not developed and dermis or 4) a systemic effect through penetration to the any further. vasculature. The aim of this research is to design a new transdermal drug delivery (TDD) system that will allow penetration of the drug through the epidermis or into the 0104. As can be seen a delicate balance between cell vasculature. The desired level of penetration will depend on membrane permeability and solubility needs to be struck for the drug. a drug to become a viable candidate for further development. 0110. The stratum corneum provides an effective barrier The reason for this is because physical properties that and prevents water and chemicals from penetrating to the enhance solubility (i.e. hydrophilicity) are usually orthogo epidermis and beyond. It has been proposed that the struc nal to those properties enhancing permeability (i.e. hydro tural organization of the lipids in the stratum corneum is an phobicity or lipophilicity). important factor in preventing fast transport of water and 0105 The interaction between metals and tetracycline chemicals. This organization of lipids results in a liquid antibiotics has been shown to reduce the bioavailability of crystal morphology and penetration though this matrix is both the drug and the metal. As stated earlier, the bioavail caused by destabilization of the liquid crystal through a ability of tetracycline antibiotics are mainly influenced by disordering of the lipid hydrocarbon chains. This is the the physicochemical properties of the metal complexes that mechanism that has been proposed for the hydrotropes prevail in the GI tract. Electric charge has the greatest impact ability to enhance penetration of topically applied drugs. on bioavailability since neutral species are more likely to 0.111 Some of the classes of chemicals that are used to readily absorb into the phospholipid membrane of the intes enhance skin permeability include alcohols, alkyl methyl tinal cells. A lipophilic metal coordinated complex should Sulfoxides, pyrrolidones, Surfactants (anionic, cationic and serve to allow greater bioavailability vis-a-vis metal salts of nonionic), and fatty acids and alcohols. In addition, lauro the drugs, which carry electric charges. Thus it is an embodi capram, urea, calcium thioglyclate, acetone and dimethyl ment of this invention that by administering lipophilic m-toluamide have been used to enhance skin penetration of metal-antibiotic covalent complexes, physicochemical prop specific bioactive reagents. Most of these drug vehicles erties of the antibiotic can be controlled and, further, may effect is by virtue of their hydrotropic properties. In chemi prevent the metals in the GI tract to impact the dynamics of cal terms, many of them have a large dipole moment; that is metal interaction with the drug and ultimately absorption. It they have a lipophilic portion and a hydrophilic portion. It is a further embodiment of this invention that the above is this large dipole moment which is a major contributing stated principle is generally applicable to all drugs. factor that causes these chemicals to disorder the lipids in the 0106 This technology can also be used to increase the Stratum COrneum. lipophilicity of highly water soluble drugs, or the so called 0112 Many drugs do not intrinsically possess enough Class III drugs. In this case, the conversion of an ionic skin penetrative ability to be used topically. Thus, virtually center, Such as a phosphate or Sulfate group, is converted to every topically applied pharmaceutical requires a formula a covalent bond. This change in bonding between metal and tion that includes a vehicle or TDD enhancer in order to ligand is known to decrease water Solubility and increase achieve the desired efficacy. Aside from the standard organic solvent solubility or lipophilicity of the ligand. requirements of safety and efficacy to which all pharmaceu 0107) If a drug is poorly soluble but is readily permeable ticals must comply, topically applied drugs need to be one way its solubility can be enhanced is by covalently soluble and stable in the vehicle, the formulation must have attaching water soluble entities such as amino acids or content uniformity, the formulation must have proper vis carbohydrates, to the drug. Alternatively, by forming a cosity and dispersion characteristics and must maximize metal-ligand complex between the drug and an ionized patient compliance, which means it must not be uncomfort metal center a new chemical entity is formed that now has able to apply, have an unpleasant odor or cause skin irrita inherent hydrophilicity imparted to it. It is an embodiment of tion. Most notably, the lag time for the drugs penetration US 2008/OO15352 A1 Jan. 17, 2008 into the epidermis, which relies on its ability to partition Most of these technologies make use of an encapsulation from the vehicle into the stratum corneum, has presented technique or bead technology wherein the active ingredient significant obstacles during the development of TDD for is encapsulated or “trapped inside a polymeric sphere. This mulations. Previous reports show that this lag time can be polymeric sphere can exist as a micelle, as a self assembled anytime between minutes to several days. Thus, a major molecular rod or ball or a coating around the active ingre impediment of the development of TDD systems has been dient. The drug is released by solvation or swelling from the these additional considerations unique to this application encapsulating agent as it circulates through the blood or and, historically, the development time for transdermal traverses the gut. The main advantage of modulating the pharmaceuticals has often been viewed as exorbitant. delivery of the drug is to extend its release, modulate the 0113 Enhanced transdermal permeability of a drug com blood levels for improved safety or enhance its absorption plex according to this invention relies on the stability of the for improved efficacy. Thus it is an embodiment of this complex coupled with its amphiphilic properties. Thus it is invention that drug-metal complex release is modulated in an embodiment that the formation of a covalent metal-drug Vivo by physicochemical action on the complex itself. bond converts the drug into an effective hydrotrope capable 0118. In certain cases it may be beneficial to enhance the of enhancing TDD of the drug itself. It is a further embodi stability of the active agent-metal complex by encapsulating ment of the invention that, if a TDD enhancer is still the drug-metal complex within a porphine, peptide or poly required, the metal will act as an anchor for the vehicle and meric matrix. This is particularly true if the active agent does the entire complex will behave as a single molecular entity. not contain the necessary elements for forming a stable The advantage with this is that drug release from the complex with a metal. Such as with the primary amines or complex no longer requires differential partition coefficients alcohols mentioned above. It is a preferred embodiment of between the vehicle and the lipid matrix of the epidermis. the invention that the matrix be a porphine derivative, 0114. Due to its covalent nature, the stability of the modified, if necessary, to allow bonding of the active agent metal-active agent complex is retained should be retained to the metal. It is an embodiment of this invention that during transport through the stratum corneum. When the drug-metal complex release is modulated in vivo by physi complex is in the epidermis the metal and the drug are cochemical action on the porphine, peptide or polymeric separated by the lipids in the membrane accepting the matrix. It is a further preferred embodiment of this invention lipophilic active agent and rejecting the hydrophilic metal. that the matrix be a compound found naturally in the Small This is imparted through physicochemical action and, in intestines. In yet a further preferred embodiment of this contrast to the earlier methods of increasing solubilities of invention the porphine matrix is bilirubin or a derivative drugs, does not require enzymes. thereof. 0115 Converting a drug to a metal coordination complex 0119) It is a further preferred embodiment of the inven also facilitates entry into the eye. It has been shown that tion that the encapsulating matrix is an amino acid or converting Sulfonamides for treating intraocular pressure dipeptide, wherein amino acids or multiple dipeptides can be (IOP) to their metal coordination complexes increased their added to coordinate with or self assemble about the metal IOP reduction effect. It is believed that this is due, in part, ligand complex. Histidine is an ideal amino acid due to the to the increased presence of the Sulfonamide in the eye and strong metal binding capacity of the imidazole moiety in that this, in turn, is due to the right balance between lipo- and histidine. Arginine is another amino acid well Suited for hydrosolubility of the metal coordinated complex. Drugs to complexation with magnesium through amidinate ligation of treat eye diseases can be improved by converting them into the guanidine portion of peptide bound arginine (FIG. 8). In a metal coordination complex according to this invention. a related embodiment of the invention, magnesium, due to This is very important to treat age-related macular degen its complexing and acid neutralizing, would stabilize argi eration (AMD), where the current therapies rely on injection nine in the stomach and increase it potency. This is good for of the drug behind the eye. An eye drop application of a drug when arginine is used as a NO source to help with COPD to treat AMD greatly improves patient compliance; coordi and related disease states. nating the AMD drug with a metal accomplish this. 0.120. The use of amino acids as secondary ligands on the Controlling Polymorphism metal is to stabilize the inner coordination sphere, create a hydrophobic shell about the inner sphere and thus prevent 0116 Polymorphism contributes a significant portion to ing hydrolysis of the metal-drug bond. Thus, it is an embodi the variability in dosages in part due to variation in Solu ment of the invention that amino acids, dipeptides or oli bility. Historically speaking, an inherent physical property of gopeptides act as secondary ligands or adjuvants on the organometallic compounds is that stable crystalline forms metal-drug complex to stabilize the complex, particularly in are relatively easy to prepare. Thus it is a further embodi aqueous systems. It is a preferred embodiment of the inven ment of this invention that polymorphism is overcome by tion that the secondary ligand is a dipeptide. It is another converting the active agent into a metal complex and Sub preferred embodiment of the invention that the secondary jecting the complex to recrystallization processes by meth ligand is an amino acid. In yet another preferred embodi ods commonly known by those skilled in the art. In so doing ment of the invention the amino acid is selected from the the active moiety is “locked' into a desired polymorph. group histidine and arginine. Modulating Drug Absorption 0121 Organometallic complexes that have a free amino 0117. In recent times there has been a flurry of activity to group (e.g. having an amino acid as part of the complex Such improve drugs by modulating how the drugs are delivered. as histidine) can initiate polymerization of an amino acid Drug delivery technology spans over all forms of adminis NCA to form a polypeptide, conformationally protecting the tration from oral to injectable to implants to skin patches. organometallic complex. It is a further advantage of this US 2008/OO15352 A1 Jan. 17, 2008

technique to allow the amino acid NCA’s to self assemble important for adhesive functions of integrins, thus it is about the organometallic complex and then coacervating the reasonable to expect that a magnesium or calcium salt or polypeptide into its self assembled structure upon initiation complex of a drug in the intestinal tract would enhance of polymerization. bioadhesion of the drug to integrins expressed on the brush 0122) It is an embodiment of this invention to combine border membrane of the intestinal lining. And since bioad the encapsulation technology with the covalent technology hesion translates into slower transit time in the gut, these to form an inner sphere covalent bond between the active complexes will confer sustained period of absorption in the agent and the transition metal, thus making a new compo gut. Therefore, it is an embodiment of this invention that sition of matter, and then encapsulating the complex with Sustained absorption of a drug will be enhanced by com outer sphere coordination within a polymer matrix to pro plexing the drug with magnesium or calcium. It is a further vide a stable complex. FIG. 9 illustrates an active agent (for embodiment of the invention that said sustained release is structural simplicity salicylic acid is the example used), and conferred upon the magnesium or calcium drug complex by polymer bound arginine bond to magnesium in the inner virtue of stronger bioadhesive properties. sphere and peptides encapsulating the complex in the outer Prevent Abuse of Narcotics sphere. 0.126 Narcotics are very effective analgesics but also can 0123. It is a further embodiment of the invention that the be very addictive. There have been many reports in the last active agent only be released when the encapsulating matrix few years describing the abuse of OxyContin by opiate swells or is dissolved by water, oil, emulsions or biologic addicts and recreational drug users. Typically the drug fluids such as gastric juices. It is an embodiment of the abuser will break the tablet matrix down mechanically or invention that the active agent cannot be released from the chemically, by adding water for example, thus making the encapsulating matrix by virtue of the strong bond between full 12 hour dose available all at once. In addition, this type the encapsulating agent and the active agent, Such as what of abuse, which usually starts with oral administration, can would occur with an antibody-antigen complex. In some often lead the abuser to snort or inject the concentrated cases it would be beneficial to have the release of the drug narcotic. from the encapsulating agent be modulated by digestive enzymes. It is a preferred embodiment of the invention that 0127. Analysis of the structure of oxymorphone and the active agent is released from the encapsulating agent by oxycodone reveals that the molecules are ideal candidates its chemical breakdown by enzymes Secreted in the intes for chelation with a metal. The B-hydroxyl at the 9-position tines, within the cell membrane or circulating in the blood and the nitrogen are positioned in Such a way that complex stream. It is preferred embodiment of the invention that the ing a metal between the two would form a highly thermo active agent is bound to aluminum, magnesium, calcium, dynamically favored 5-member ring. It is preferred that the iron, bismuth, silicon or zinc. In another most preferred 9-hydroxyl is deprotonated to form an anionic alkoxide embodiment of the invention the encapsulating agent is an (FIG. 10). The nitrogen's lone pair of electrons may con antibody raised against the metal-ligand complex. In yet tribute enough electron density to stabilize the metal chelate. another embodiment of the invention the complex comprises Further stabilization can be imparted by adding secondary an active agent-metal complex and the encapsulating agent ligands or adjuvants to the complex in the manner of the case is self-assembled from the combinations of amino acids, where Salicylaldehyde Stabilizes the glycine-magnesium porphines, carbohydrates or mixtures thereof. In a most complex. It is a further embodiment of the invention that the preferred embodiment of the invention the active agent metal-narcotic complex is encapsulated within the matrix as metal-encapsulating agent complex is a pharmaceutical. described above. 0.124. In another embodiment of the invention, the coor 0128. Thus it is an embodiment of the invention that by dination complex is a metal selected from all metals that can virtue of the narcotic being complexed with a metal that the form such complexes, and the drug is selected from the narcotic is released from the complex slowly through physi group of all biologically or pharmacologically active agents. cochemical action. This means that the narcotic is not In a preferred embodiment of the invention the pharmaco available immediately or all at once. Furthermore, it is an logically active agent requires a specific conformation for embodiment of this invention that the metal-narcotic com biological activity. The activity could be dependant on the plex is unable to pass the blood brain barrier, rendering the active agents ability to cross cell membranes and the narcotic ineffective until release from the complex has coordinating metal provides the correct structure for mem occurred. Since the kinetics of release is slow the amount of brane translocation of the active agent. In a preferred narcotic available for transport across the blood brain barrier embodiment the pharmaceutically active agent is selected at any one time is much less than the dose administered and from the group consisting of Small molecules, peptides, so no euphoric effect is achieved. It is a further embodiment carbohydrates, DNA or RNA, the latter two being used in of the invention that the kinetics of narcotic release can be gene therapy, as aptamers or in antisense nucleotide thera slowed even more by incorporating secondary ligands, peutic applications. In a preferred embodiment the metal is encapsulating agents or a combination of both. In a preferred selected from the group consisting of aluminum, bismuth, embodiment the metal is selected from the group consisting calcium, magnesium, iron, silicon and zinc. of aluminum, bismuth, calcium, magnesium, iron, silicon and zinc. In a more preferred embodiment of the invention Bioadhesion Properties the metal is selected from the main group elements. In an 0125 There are a variety of ways that incorporating a even more preferred embodiment of the invention the metal magnesium or calcium ion into the molecular formula of a is selected from the S-block main group elements. In a most drug would infer bioadhesive properties to the drug. For preferred embodiment of the invention the metal is magne example, it is known that magnesium and calcium are S1. US 2008/OO15352 A1 Jan. 17, 2008

Selection of Metals headaches, ADHD), and so from a prophylactic point of view magnesium may have significant benefit. For example, 0129 Reference has been made to the preferred metals to triptan magnesium may be an ideal candidate for this be used in the coordination complexes. In pharmaceutical technology. applications, the safety of the entire metal coordinated pharmaceutical needs to be considered when selecting the 0.135 Calcium is absorbed in the intestines by an active metal used in complexes of this invention. Although to transport mechanism, whereas magnesium is transported practice this invention many metals can be used, it is a passively. Magnesium (as a salt) and Furosemide's intestinal preferred embodiment of this invention that the metal be transport were facilitated when both were co-administered selected from a shortlist that would be generally regarded as orally. Thus Furosemide, a poorly absorbed drug, represents safe (GRAS). One criterion for selecting the metal is to another compelling candidate for this technology. review the list of mineral supplements currently on the market and select the ones whose dosages would far exceed 0.136. It is a most preferred embodiment of this invention the dose likely to be included as a coordination complex that the metal is magnesium. with the drugs listed in Table 1. From the PDR for Non Selection of Solvents prescription Drugs and Dietary Supplements a list of 7 0.137 As stated earlier, the selection of solvent for the metals (excluding alkaline metals, i.e. sodium, potassium, complexation reaction has an impact on the structure and etc) with amounts greater than 2 mg/dose is shown in Table stability of the metal coordinated compound. Magnesium 2. forms strong bonds with water and the coordination sphere hydrated magnesium will have an impact on the kinetics of TABLE 2 product formation as well as the structure and stability of the Coordination complex candidates from product. Because of the strong nitrogen-transition metal the PDR for Non-prescription Drugs bond, in those cases where nitrogen containing ligands are reacted with transition metals, such as Zinc, the presence of Metal Compound Brand Name Amt. metal dose water in the reaction mixture will usually not have as strong Aluminum Aluminum Hydroxide Maalox 400 mg an impact on the structure and stability of the metal coor Bismuth Bismuth Subsalicylate Pepto-Bismol 525 mg dinated product. Calcium Calcium Carbonate Caltrate 600 mg Iron Ferrous Fumarate Ferretts 106 mg 0.138. In some cases, depending on the ligand, the metal Magnesium Magnesium Hydroxide Maalox 400 mg Silicon Sodium Metasilicate One-A-Day 6 mg and the desired product water may be the solvent of choice. Zinc Zinc Oxide One-A-Day 15 mg The majority of the products will dictate that an anhydrous organic solvent will be the best selection. Some suitable solvents include alcohol, acetone or THF. The most prefer 0130. In a preferred embodiment of the invention the able solvent is DMSO because it is an excellent universal metal is selected from the group consisting of aluminum, solvent that dissolves virtually every pharmaceutical or bismuth, calcium, iron, magnesium, silicon and zinc. nutraceutical and also will dissolve most metal halides Whereas it is the embodiment of the invention that a new including magnesium chloride. This allows for single phase composition of matter is formed through the formation of a reactions. In addition, a stable metal coordinated pharma covalent bond between a pharmaceutical and any metal, ceutical can be isolated by a process similar to coacervation, including the lanthanides, actinides, the transition metals, which typically will include simply adding a non-solvent to and the main group metals (S- and p-block), it is a preferred the reaction mixture. embodiment of the invention that the metal be selected from 0.139. DMSO can form complexes with metals, including the S-block main group elements. The reason for this is that magnesium, in situ, setting up the DMSO-metal complex to the s-block elements are more likely to be GRAS and are react with the drug ligand, thereby displacing the DMSO more often used in OTC drug products and vitamin Supple ligand at the metal center. DMSO can then serve as a ments than the transition metals or p-block main group transient protecting group in those reactions where adjuvants elements (lanthanides or actinides are never used in OTC are to be included in the complex. This in-process reaction products). There are several reasons for selecting magne scheme is facilitated by the fact that the DMSO-metal sium over the other S-block elements, such as calcium, complex cannot form outer coordination spheres like water which are: does due to the lack of hydrogen bonding between the 0131 Calcium shows larger variability with respect to DMSO ligands. This makes the metal center easily acces coordination number with 8>7>6>9 in order of preference. sible by incoming ligands. Depending on the metal coordi Magnesium, being Smaller than calcium, is almost exclu nated complex formed, the final product may or may not sively octahedral, which simplifies the synthetic strategies retain DMSO as a ligand. If DMSO is attached to the ligand, and will more likely give a higher yield of a single product it is unlikely that a situation will exist Such that the dosing instead of a mixture of products. of DMSO will ever reach anywhere close to toxic levels. 0132) Magnesium can form covalent bonds with chelat 0140. It is the premise of this invention that by merely ing ligands more readily than the other S-block elements; adding a metal salt to an aqueous Solution of a biologically active ligand the coordination complex formed is not the 0.133 Magnesium forms a more stable bond with proteins same as if the combination of the reagents were done under and nucleic acids than calcium and thus provides enhanced anhydrous conditions. Furthermore, by reconstituting the stabilization of biologic pharmaceuticals. dried coordination complexes in the same aqueous environ 0134 Magnesium deficiency has been implicated in sev ment the structure of the two complexes would be different. eral disease states (e.g., cardiovascular related, migraine To that end, several coordination complexes can be prepared US 2008/OO15352 A1 Jan. 17, 2008 with FDA approved pharmaceuticals, demonstrating that the T3-Zn examples), the reaction is repeated except water is complexes are stable. The products will be characterized as included in the reaction medium as described in the completely as possible and bioavailability studies will be examples below. The reaction is worked up and dried as conducted. The metal coordinated complexes can be pre before. pared in water and organic solvents. It is expected that, in 0147 Adding water to the T3-Mg complexation reaction many cases, the respective products will have differences in clearly had an impact on the isolated product. The T3-Mg stability, structure or biological activity. compounds prepared in DMSO alone, showed line broad Selection of Drugs ening in the aliphatic region only, with sharp aromatic peaks, in its "H NMR spectrum (FIG. 11). The "H NMR of T3, 0141 Complexes of almost any drug that can form a which shows the sharp peaks in the aliphatic region, is stable complex with a metal is enabled by this invention. The shown in FIG. 12. By comparison, the "H NMR of the drugs selected for examples below represent a cross section T3-Mg product prepared in the presence of water revealed of chemical and therapeutic classes as shown in Table 2. extensive line broadening throughout the "H NMR spectra. Furthermore, the magnesium content (1.62%) of the anhy TABLE 3 drously prepared T3-Mg product very closely matched that of bis(triiodothyroninato)-bis(dimethylsulfoxide)-magne Drugs selected as examples in the invention sium. In contrast, the T3-Mg complex prepared in the Drug Therapeutic Class Chemical Class presence of water had a magnesium content of only 0.96%. Trilidothyronine Hypothyroid drug Amino Acid It also had 0.23% potassium in it, whereas no potassium was Minocycline Antibiotic B-diketone detected in the bis(triiodothyroninato)-bis(dimethylsulfox Tetracycline Antibiotic B-diketone ide)-magnesium product. Hydrochlorothiazide Diuretic Sulfonamide Metformin Diabetes drug Biguanide 0.148 Tetracycline has 3-diketone and B-ketophenol Acycloguanosine Antiviral Ureide functionalities and will form stable complexes with magne iRNA Gene therapy Oligonucleotide sium. Adding water to the reaction in DMSO has very little effect on the solubility, the "H NMR spectra or the metal content of the respective products. In fact the "H NMR Small Molecule Discussions spectrum of the product isolated from a reaction done in Synthesis water alone does not differ significantly from the product isolated from DMSO alone or in a 5:1 DMSO: water mixture. 0142. Other drug-magnesium complexes that may be There is a trend of lower magnesium content with higher important include Triptan-Mg due to the importance of water content in the reaction solvent, but that may be due, magnesium for headache relief and Oxycodone-Mg because primarily, to extent of hydration in the product. of the importance of abuse resistant narcotics. 0.149 Since zinc forms a very stable bond with nitrogen 0143 Typically an anhydrous metal halide (iodide, bro containing compounds, water does not interfere with com mide or chloride) is added to a mixture of the drug and plexation between Zinc and the ligand but may impact the KO'Bu in "BuOH/DMSO. Alternatively, the metal halide can complex structure. The product resulting from reacting a be added to the drug plus a solution of a tertiary amine (e.g. metal and a drug in anhydrous DMSO typically yielded well triethylamine) in DMSO. Yet another option is where the characterized coordination complexes. For those com metal halide can be added to the drug plus KH in THF. The pounds that were either not well defined structurally (i.e. metals of choice are magnesium and Zinc and the halide of HCTZ-Zinc) or were somewhat unstable (i.e., dimethylbigu choice is chloride. Zinc chloride is soluble in DMSO, anide-Zinc complex), a Zinc coordination complex was isol acetone or ethanol and are the solvents of choice for Zinc able and was, at least partially, characterized. The same complexation, particularly with nitrogen containing ligands. complexes prepared in the presence of water, had higher solubility in polar solvents. The difference in solubilities of 0144. The product is isolated by precipitation, is sepa the products from the respective methods of preparation rated from the liquid by Suction filtration or centrifugation, clearly established a difference in the products themselves. washed and then dried under high vacuum to remove the last It is believed that the Zinc products prepared in aqueous traces of moisture. The drug:metal complex may form a Solvent systems produced ionic salts, outer coordination hydrate and all of the water may not be removable under complexes, hydrated complexes or combinations thereof. high vacuum. Alternatively, the added water may not dis This appeared to be the case in the dimethylbiguanide-Zinc place the remaining DMSO ligands on the metal formed in complex prepared in 5:1 DMSO: water mixture, where the situ. Consequently, the product may be a drug: metal:DMSO H NMR spectrum revealed resonances at 2.85 and 2.80 complex. ppm, which correspond to the N-methyl groups and indicate 0145 Certain drugs, where their dissociation constants free (or ionic) and complexed dimethylbiguanide, respec are high when bound to magnesium, favor complexation tively. with DMSO. Formation of ternary complexes in situ would Characterization further stabilize the complexes and would retain their 0150. Each product synthesized were characterized by molecular integrity during the process of absorption after NMR, MS (either TOF or FAB) and ICP. The NMR spec oral administration. It is for this reason that for most drugs trum confirms the integrity of the sample and shows that a when reacted with magnesium halide, DMSO is the solvent metal is complexed by the presence of line broadening, peak of choice. shifts or multiple resonances. 0146). As a comparator for the complexation reactions in 0151 Metal content for most of the complexes prepared the examples (excluding the acycloguanosine-Mg and in anhydrous DMSO were consistent with a complex of two US 2008/OO15352 A1 Jan. 17, 2008

drugs bound to a metal. In addition, the metal content the imidazole nitrogen; the resonance at 10.6 ppm, which is remained constant from batch to batch. The complexes of missing in the NMR spectrum of the complex, is assigned to Dimethylbiguanide had variable metal content depending on the amide proton. the method of isolation and never had consistent drug:metal 0157 The mass spectrum revealing a significant presence ratios. The metal content was determined by the ICP analysis of the coordination complex is an important indicator of the and based on that data, in conjunction with the NMR and stability of the complex. Thus, molecular ions were found MS, the ratio of drug to metal can be calculated. for bis(T3)Mg, bis(T3)Zn, bis(minocyclinato)Mg, bis(tetra 0152 The "H NMR spectra of the T3 complexes showed cyclinato)Mg, and bis(acycloguanosinato)Mg. Two molecu Some line broadening and upfield shifting in the aliphatic lar ions with zinc isotope patterns were observed in the region indicative of complex formation with the amino acid MALDI spectrum of hydrochlorothiazide-zinc complex. It is portion of the molecule. This can be seen by comparing the not known, at this time, a structure corresponding to those region between 2.5 ppm and 3.5 ppm in the "H NMR spectra masses. Dimethylbiguanide-Zinc complex did not have a of bis(T3)bis(DMSO)Mg, T3, free acid and bis(T3)Zn, Zinc containing molecular ion in its MALDI spectrum. This which are shown in FIGS. 11, 12 and 13, respectively. is believed to be due to the compounds instability. 0153. The H NMR spectrum of the dimethylbiguanide 0158 FTIR studies may be used to determine whether, complexes showed large upfield shifts of the NH reso for a particular complex that has been found, the ligand nances indicative of complex formation with the nitrogen bonding atom and if the complex is coordinated with atoms. In addition, a 0.05 ppm upfield shift of N-dimethyl DMSO, water, or not solvated at all. groups was observed in the dimethylbiguanide-Zinc com plex spectrum (FIG. 14) relative to the spectrum of dimeth Stability ylbiguanide (FIG. 15) 0159. Equilibrium constants for the coordination com 0154) The "H NMR spectrum of the minocycline and plexes made have been estimated from literature precedents tetracycline complexes resembled polymeric structures with of similar compounds. For example, the equilibrium con very large line broadening and manifestation of many new stant, log K for T3-Zn is estimated to be between 4 and 5 resonances throughout the entire spectrum. To demonstrate based on another amino acid Zinc complex, phenylalanine that these spectra were due to dynamic isomeric mixtures zinc. Likewise, the log K for dimethylbiguanide-zinc is and not decomposition or polymerization, 12 NHCl was estimated to be between 5 and 7. The log K for hydro added to the NMR samples of tetracycline and its magne chlorothiazide-zinc is difficult to estimate from literature sium complex and the spectra retaken. FIGS. 16-18 show the values. Stability constants for tetracycline-magnesium in "H NMR spectra of Tetracycline, bis(tetracyclinato) mag water at various pH values have been reported and the nesium and the complex with HCl added, respectively. As expected log K for tetracycline-magnesium is between 4 can be seen in the spectra series the magnesium complex and 5. The log K for acycloguanosine-magnesium is esti reverted back to the reference tetracycline compound. Inter mated to be 1.6. The log K for T3-Mg is difficult to estingly, when HCl was added to tetracycline, a considerable estimate due to the lack of a good comparator but the log K. amount of decomposition could be seen in the aromatic for glycine, which like T3 is also an amino acid, is 1.34. Due region of the NMR spectrum (FIG. 19), the magnitude of to T3's much greater hydrophobicity relative to glycine, the which was not seen in the commensurate spectra of the log K for T3-Mg is expected to be much larger than 1.34. tetracycline-magnesium complex. This indicates an acid 0.160 Another indicator of drug-metal or drug-metal protective effect imparted by complexation with magnesium adjuvant stability is their binding constants, which is related and could become an important attribute of this technology to K, but further shows the stepwise stability for multi for those drugs that are unstable in acid environments. Such dentate ligands. The cumulative binding constant, B, for the as in the stomach. maximum binding between the metal and the ligand is given O155 The H NMR spectrum of the hydrochlorothiazide by Equation 4. complexes also resembled polymeric structures with the same kind of line broadening and new nondescript reso nances seen in the spectra of the antibiotic-metal complexes. Equation 4 FIGS. 20-22 show the "H NMR spectrum of hydrochlorothi azide, hydrochlorothiazide-Zinc complex and the complex with HCl added, respectively. As can be seen the line broadening was reverted back to the sharp resonances (0161) This difference can be seen in the measured log K. observed in the reference drug. This proves that the line for phenylalanine-zinc and its approximated B value of 8.5. broadening and additional resonances observed in the "H This f value may also more closely reflect the stability of NMR spectrum of the respective complexes were due to the T3-Zn complex. multiple stereochemical and geometric isomers of the com 0162 There are several ways in which binding constants plex in solution. Moderately slow interchange between the of metal-drug complexes in different environments can be isomers in solution could also contribute to the line broad estimated. Optical absorption spectroscopy, NMR spectros ening observed. copy, mass spectrometry, reaction kinetics, potentiometry 0156 Spectral data on the acycloguanosine-magnesium and chromatography are several Such methods. complex data showed that a complex was formed. Compari son of the "H NMR spectra of acycloguanosine with that of Partition Coefficient and Distribution Coefficient its magnesium complex (FIG. 23) suggested that the com 0.163 The partition coefficient is a constant and is defined plexation site on acycloguanosine was the amide oxygen and as the ratio of concentration of a neutral compound in US 2008/OO15352 A1 Jan. 17, 2008 aqueous phase to the concentration in an immiscible organic 0182 LogD is related to LogP and the pKa by the phase, as shown in Equation 5. following equations: Partition Coefficient, P=Organic Aqueous Equation 5 Log DH=log P-log 1+10(PHPisa) for acids Equation 9 0164. In practice the Log P. defined in Equation 6, will Log DH=log P-log 1+10(PKPH for bases Equation 10 vary according to the conditions under which it is measured 0183 So, when the pH is adjusted such that ionization is especially pH since at a low pH bases will be ionized and at minimized, the log D will be nearly equivalent to the log P. a high pH acids will be ionized. Under those conditions, then, log D is a reliable indicator of Log P=log 10 (Partition Coefficient) Equation 6 the bioavailability of a drug in a particular application. In terms of a metal coordinated drug, increases in log D of the 0165 Thus, a Log P=1 means 10:1 Organic: Aqueous, a drug-metal complex relative to the reference drug will not Log P=0 means 1:1 Organic: Aqueous and a Log P=-1 only indicate an increase in lipophilicity but will also means 1:10 Organic:Aqueous. demonstrate its stability in water, as well. The log D's at pH 0166 Naturally, ionized compounds will partition pref 7.4 for tetracycline, bis(tetracyclinato)magnesium, triiodot erentially into the aqueous phase, thereby lowering their log hyronine and bis(triiodothyroninato)Zinc were determined P. For neutral molecules that are bases they will remain and are shown in Table 4 along with their pKa's. neutral when the pH is greater than 2 units above its pKa and for neutral acids when the pH is 2 units below its pKa. TABLE 4 0167 The choice of partitioning solvent will also have an pKa and Log D for selected reference drugs impact on log P. Most log P measurements will use the and their metal coordination complexes. octanol: water system. Ion pairing effects impact the log P Compound Method pKa Log D74 measurements and should be accounted for, especially with Tetracycline DPAS 8.95, 7.04, -1.2 metal coordinated compounds such as those embodied in 3.34 this invention. Bis(tetracyclinato)magnesium DPAS 8.33, 7.17, O.O1 3.36 0168 Interms of pharmaceutical applications the follow Triiodothyronine Potentiometric 8.41, 8.07 2.80 ing guidelines have been used in determining the method of Bis(triiodothyroninato)Zinc Potentiometric 6.66, 6.42 3.43 administration, formulation and dosage forms: DPAS = Dip Probe Absorption Spectroscopy 0169 Low Log P (below 0) Injectable 0.184 These results run counter to what the prior art 0170 Medium (0-3) Oral teaches; that is combining the anion of a neutral compound 0171 High (3-4) Transdermal with a metal salt should produce a compound with less lipophilicity and a reduced log D. Perhaps more interest 0172 Very High (4-7) Toxic build up in fatty tissues ingly, by application of the technology in this invention, tetracycline was theoretically transformed from an inject 0173 And within the realm of orally administered drugs able drug (log D-0) to an oral drug (log DO) and T3 was these guidelines have been used: transformed from an oral drug (log D-3) to a transdermal 0.174 1. For optimum CNS penetration, Log P=2+/- drug (log D3). This latter manifestation has important 0.7 (Hansch rules) implications in increasing the safety of T3 products by administering the drug in a slow release transdermal depot. 0.175 2. For optimum oral absorption, Log P=1.8 Bioavailability Studies 0176 3. For optimum intestinal absorption, Log P-135 0185. Overview: A preliminary study in a rat model to observe the effects that coordinating a metal with a drug will 0.177 4. For optimum colonic absorption, Log P=1.32 have on the absorbance of the reference drug was conducted. The reference drug selected for this particularly study was 0.178 5. For optimum sub lingual absorption, Log triiodothyronine (T3), which is the active ingredient in P-55 Cytomel and Thyrolar. Both Cytomel and Thyrolar are 0.179 6. For optimum Percutaneous penetration, Log currently used to treat hypothyroidism. Cytomel has also been indicated in the treatment of certain psychological P=2.6 (& low mw) disorders. 0180. The distribution coefficient (D) is the ratio of unionized compound in the organic phase to the total amount 0186 Study Design: T3-Zinc and T3-Magnesium were of compound in the aqueous phase given by Equation 7. tested for bioavailability, relative to the reference drug over a 5 hour time period. The three compounds were formulated, D=Unionised (o), Unionised (aq)+Ionised (aq) Equation 7 separately, into gelatin capsules, with a total dose of 108-12 ug/kg administered. In order to avoid acid degradation of the 0181 Log D is the log distribution coefficient at a par test compounds in the rat Stomach, metal oxides were added ticular pH (Equation 8). This is not constant and will vary to the formulation. Another T3 control was included, where according to the protogenic nature of the molecule. Log D Zinc oxide was added to T3, free acid. Each of the formu at pH 7.4 is often quoted to give an indication of the lated gelatin capsules were orally administered directly to lipophilicity of a drug at the pH of blood plasma. the esophagus of respective rats and blood samples were Log D=log 10 (Distribution Coefficient) Equation 8 collected at pre-dosing and at 0.5, 1, 2, 2.5, 5 hours after US 2008/OO15352 A1 Jan. 17, 2008 dosing. Serum triiodothyronine levels were analyzed by an DMSO-magnesium complex. The FTIR spectrum showed independent laboratory, using an industry standard assay an extra stretch at 954 cm, which is indicative of an method. S=O Mg stretch. FTIR of T3 complexes were examined 0187. Results: The data shown in the FIG. 30 reveal that for the presence of S=O Mg, C=O and N—H stretches. all four drug formulations were readily absorbed by the rat, 0.195 The following examples illustrate compounds hav with a rapid rise in serum T3 levels up to the 2.5 hour time point and a general leveling off in the metal coordinated T3 ing lower water solubility and carboxylic acid functionalities fed rats after that. The T3, free acid fed rats indicated that the when complexed with a metal. DMSO is a preferred solvent, serum T3 levels may still be rising at the 5 hour time point. to which a strong or weak base is added. Such as potassium The rats that were administered metal complexed T3 and T3 t-butoxide or potassium carbonate, followed by the salt of with zinc oxide added had an increase in serum T3 blood the complexing metal, preferably the chloride salt. The levels that were approximately 65-85% greater that of T3, preparation also includes water precipitation with or without free acid at the 5 hour time point. concentrating in vacuo. This method can be applied to the 0188 Conclusions: The data clearly indicate that com amido acid and thio acid. plexing a metal with T3 increased the absorbance of orally Preparation of Bis(triiodothyroninato)-bis(dimethylsulfoxi administered T3 nearly two fold over T3 alone. It is believed de)magnesium that the T3, free acid complexed with the Zinc oxide prior to absorption of T3 in the animals fed the formulation of T3 0196) Triiodothyronine or T3 (218 mg) was dissolved in and Zinc oxide. 4 mL of anhydrous DMSO, after which 0.34 mL of 1 M 0189 This study represents a significant advance in drug potassium t-butoxide int-butanol was added and the Solution delivery technology and that coordinating metals with phar stirred for 10 minutes. Magnesium chloride (16 mg) was maceuticals can be applied to and improve the performance added and the solution stirred overnight. The solution was of drugs with bioavailability limitations. poured into 10 mL of deionized water to precipitate the product, which was suction filtered and air dried. After an Large Molecule Protocols overnight drying under high vacuum the yield was 164 mg Synthesis of a light beige powder. The product structure was charac 0190. Double stranded iRNA of a defined size were terized by H NMR, FAB-MS and ICP. H NMR (DMSO): prepared according to the standard protocol described in the & 7.83 (s), 7.05 (d), 6.82 (d), 6.62 (dd), 3.26 (bm), 3.15 (bd), Examples section. The iRNA was then reacted with mag 2.71 (bm), 2.54 (s). The presence of the broad multiplets in nesium or Zinc under anhydrous conditions or in the pres the side chain region indicates that as the site for magnesium ence of water as described in the Examples section. binding (see FIG. 11). FAB-MS: Molecular ion at 1325 indicative of bis(triiodothyroninato)magnesium (after loss 0191 The biological activity of iRNA can be modulated of DMSO ligands). FTIR (neat): cm 1596 (C=O stretch), in various ways by complexing it with other metals such as 1014 (S=O stretch), 949 (S=O Mg stretch); absence of calcium, Zinc, cobalt and manganese. In addition, combina N—H stretch at 1633. Magnesium analysis (ICP): Expected tions of multiple metals, such as including Cu or Ag to 1.68%; Found 1.62%. This confirms the structure as shown facilitate binding of the purine/pyrimidine groups along with in FIG. 24 and is about 96% pure, where most of the the phosphate groups can improve the transfection efficiency contamination is probably due to water as seen in the "H and stability of iRNA. NMR spectrum. Characterization Preparation of Bis(triiodothyroninato)Zinc 0192 The metal:RNA products were tested for changes in ionic/covalent behavior using Isoelectric focusing (IEF) 0197) Triiodothyronine or T3 (192 mg) was dissolved in gel following the standard protocol described in the 4 mL of anhydrous DMSO, after which 0.30 mL of 1 M Examples section. The advantage of the IEF gel is that it potassium t-butoxide int-butanol was added and the Solution provides an inexpensive tool by which to monitor changes in stirred for 10 minutes. Zinc chloride in diethyl ether (0.16 the charge distribution on the target RNA molecule. Data mL of 1M solution) was added and the solution stirred from the IEF studies were represented as ApKa for each overnight. The solution was poured into 10 mL of deionized iRNA complex prepared. water to precipitate the product, which was suction filtered and air dried. After an overnight drying under high vacuum Conclusion of IEF Experiments the yield was 140 mg of a light beige powder. The product 0193 The presence of an RNA molecule from the anhy structure was characterized by "H NMR, FAB-MS and ICP. drous DMSO reactions with a different isoelectric point H NMR (DMSO): 8 7.82 (s), 7.02 (d), 6.80 (d), 6.61 (dd), (pKa) indicates the presence a new RNA molecule (FIG. 3.49 (bm), 3.22 (bm), 2.67 (bm). The presence of the broad 31). Since this is not due to RNA degradation means that the multiplets in the side chain region indicates that as the site new RNA molecule is a stable complex between the metal for zinc binding (FIG. 13). FAB-MS: Molecular ion at 1364 (magnesium or Zinc) and RNA. In addition, because its pKa and 1366 indicative of bis(triiodothyroninato)Zinc with iso is lower than the native RNA supports the formation of a topic abundance pattern consistent with zinc. FTIR (neat): covalent bond between the metal and RNA. cm 1582 (C=O stretch); absence of N-H stretch at 1633. EXAMPLES Zinc analysis (ICP): Expected 4.8%; Found 4.3%. This supports the structure as shown in FIG. 25 and is about 90% FTIR Analysis of DMSO-Magnesium Complex pure, where most of the contamination is probably due to 0194 In order to determine which atom of DMSO binds water and may be a hydrate as seen in the "H NMR to magnesium and FTIR spectrum was collected of a spectrum. US 2008/OO15352 A1 Jan. 17, 2008

Preparation of Magnesium Triiodothyronine Complex in the vacuo at 30° C., after which 0.5 mL of deionized water was Presence of Water. added and the mixture triturated and transferred to a 2 mL microcentrifuge tube. The product was separated from the 0198 Triiodothyronine or T3 (188 mg) was dissolved in liquid by centrifuging at 8,000 rpm for 6 minutes and the 3.5 mL of anhydrous DMSO, after which 0.29 mL of 1 M Supernatant was decanted. The pellet was washed by adding potassium t-butoxide int-butanol was added and the Solution 0.5 mL of water, vortex mixed, centrifuged and the super stirred for 10 minutes. Magnesium chloride (16 mg) in 0.5 natant decanted. The washing procedure was repeated. After mL of water was added and the solution stirred overnight. an overnight drying under high vacuum the yield was 72 mg The solution was poured into 10 mL of deionized water to of a deep yellow powder. The "H NMR spectrum (FIG. 20) precipitate the product, which was suction filtered and air resembled a polymeric structure, which contained broad dried. After an overnight drying under high vacuum the yield multiplets between 8.8 and 10.1 ppm, 6.4 and 7.8 ppm, 4.2 was 188 mg of a light beige powder. The product structure and 5.0 ppm and 1.1 and 3.1 ppm. The product structure was characterized by "H NMR, FAB-MS and ICP. H NMR could not be accurately characterized by "H NMR, possibly (DMSO): 8 7.81 (bs), 7.02 (bs), 6.81 (bd), 6.59 (dd). The due to the different permutations of bidentate complex forms other resonances were hiding behind the solvent and water possible with anionic Tetracycline and magnesium and the peaks. The sample formed a cloudy suspension in DMSO. moderately slow equilibrium between those isomeric com FAB-MS: Molecular ion at 652 indicative of protonated T3 plex forms. Confirmation was observed by adding approxi with no bound Mg. FTIR (neat): cm 1633 (N-H stretch), mately 1 equivalent of 12 NHCl to the NMR sample and 1535 (C=O stretch), 1012 (S=O stretch), 949 (S=O Mg reanalyzing by "H NMR, which revealed reversion of the stretch). The strength of the DMSO related stretches relative magnesium complex back to tetracycline and, presumably, to the T3 related stretches indicated a mixture of magnesium chloride (FIG. 18). The product was further (DMSO). Mg, cation and T3 anion. Magnesium analysis characterized by MALDI-ES and ICP. MALDI-ES: Molecu (ICP): Expected 1.8%; Found 0.96%. Potassium analysis lar ion at 911.3 indicative of bis(tetracyclinato)magnesium. (ICP): Found 0.23%. From the ICP data it appears that this Magnesium analysis (ICP): Expected 2.67%: Found 2.53%. product is a mixture of the magnesium salt, the potassium FTIR does not indicate presence of a DMSO ligand. This salt and the Zwitterion. indicates that there are two Tetracycline molecules per 0199 The following examples illustrate the use as start magnesium atom. According to NMR evidence from previ ing materials compounds that may be unstable. Examples of ously published studies of tetracycline-magnesium com such compounds are the B-diketone, C.-diketone, ketophe plexation in aqueous systems, a likely structure for bis(tet nol, C.-ketoalcohol, or B-ketoalcohol chemical classes. racyclinato)magnesium is shown in FIG. 27. The product DMSO is a preferred solvent, to which a strong base is about 95% pure, which most of the contamination is prob added, such as potassium t-butoxide, followed by the salt of ably due to solvents as seen in the "H NMR spectrum. the complexing metal, preferably the chloride salt. The Preparation of Magnesium Tetracycline Complex in Water preparation also includes water precipitation with or without concentrating in vacuo. 0202 Tetracycline (89 mg) was dissolved in 1.5 mL of water, after which 0.2 mL of 1 M potassium t-butoxide in Preparation of Bis(minocyclinato)magnesium t-butanol was added and the solution stirred for 10 minutes. 0200 Minocycline (104 mg) was dissolved in 3 mL of Magnesium chloride (0.11 mL of 1 M solution) was added anhydrous DMSO, after which 0.44 mL of 1 M potassium and the solution stirred for 3 hours. The resultant precipitant t-butoxide int-butanol was added and the solution stirred for was separated from the water by centrifuging at 8,000 rpm 10 minutes. Magnesium chloride (11 mg) was added and the for 6 minutes and the supernatant was decanted. The pellet solution stirred overnight. The solution was poured into 10 was washed by adding 1 mL of water, Vortex mixed, mL of deionized water to precipitate the product, which was centrifuged and the Supernatant decanted. After an overnight Suction filtered and air dried. After an overnight drying drying under high vacuum the yield was 65 mg of a deep under high vacuum the yield was 52 mg of a deep yellow yellow powder. The "H NMR spectrum very closely powder. The product structure could not be characterized by resembled the complex prepared in anhydrous DMSO. Mag "H NMR, possibly due to the different permutations of nesium analysis (ICP): Expected 2.67%: Found 2.42%. It bidentate complex forms possible with anionic Minocycline appears that performing the complexation in water versus and magnesium. The product was characterized, then, by under anhydrous conditions has a minor impact on the FAB-MS and ICP, FAB-MS: Molecular ion at 937.4 indica stability and structure of the tetracycline-magnesium com tive of bis(minocyclinato)magnesium. Magnesium analysis plex. (ICP): Expected 2.68%; Found 2.61%. This confirms that 0203 The following examples illustrate the use as start there are two Minocycline molecules per magnesium atom, ing materials compounds that belong to any of the amine, as represented by a likely structure shown in FIG. 26 (see diamine, guanide, diamide, certain purine, Sulfonamide, and discussion for Tetracycline below). The product about 97% oligopeptide chemical class. DMSO is a preferred solvent, to pure, which most of the contamination is probably due to which a strong base is added. Such as potassium t-butoxide, water as seen in the "H NMR spectrum. followed by the salt of the complexing metal, preferably the Preparation of Bis(tetracyclinato)magnesium chloride salt. The preparation also includes alcohol precipi tation with or without concentrating in vacuo. 0201 Tetracycline (89 mg) was dissolved in 0.5 mL of Preparation of Hydrochlorothiazide Zinc Complex anhydrous DMSO, after which 0.2 mL of 1 M potassium t-butoxide int-butanol was added and the solution stirred for 0204 Hydrochlorothiazide (120 mg) was dissolved in 0.5 10 minutes. Magnesium chloride (11 mg) was added and the mL of anhydrous DMSO, after which 0.4 mL of 1 M solution stirred for 3 hours. The solution was concentrated in potassium t-butoxide int-butanol was added and the Solution US 2008/OO15352 A1 Jan. 17, 2008

stirred for 10 minutes. Zinc chloride in diethyl ether (0.2 mL was repeated. After an overnight drying under high vacuum of 1M solution) was added and the solution stirred for 4 the yield was 57 mg of a free flowing white powder. "H hours. The solution was concentrated in vacuo at 30° C., NMR (DMSO): & 4.90 (s), 4.66 (s), 4.50 (s), 2.80 (s). The after which 0.5 mL of methanol was added and the mixture large upfield shifts of —NH protons relative to dimethyl triturated and transferred to a 2 mL microcentrifuge tube. biguanide indicate complexation with zinc. A small upfield The product was separated from the liquid by centrifuging at shift of 0.05 ppm in the dimethyl groups was also observed 8,000 rpm for 6 minutes and the supernatant was decanted. in the dimethylbiguanide-zinc complex. FIGS. 14 and 15 The pellet was washed by adding 0.5 mL of methanol, show the NMR spectra of dimethylbiguanide-zinc and dim Vortex mixed, centrifuged and the Supernatant decanted. The ethylbiguanide, respectively. The product was further char washing procedure was repeated. After an overnight drying acterized by ICP. Zinc analysis (ICP): Found 8.14%. FTIR under high vacuum the yield was 104 mg of a free flowing data did not indicate presence of a DMSO ligand. NMR and white powder. The product was apparently hygroscopic due ICP data clearly indicate the formation of a dimethylbigu to the powder turning gummy after a few minutes exposure anide-zinc complex. FIG. 28 represents the biguanide-metal to ambient air. The "H NMR spectrum resembled a poly complex prepared. MALDI-ES analysis did not reveal a zinc meric structure (FIG. 21), which contained broad multiplets containing compound. between 6.3 and 8.0 ppm, and 4.4 and 5.9 ppm. The product structure could not be accurately characterized by H NMR, Preparation of Zinc Dimethylbiguanide Complex in the possibly due to the different permutations of complex forms Presence of Water possible with anionic hydrochlorothiazide and zinc, and the 0207. This procedure followed the analogous anhydrous moderately slow equilibrium between those isomeric com preparation exactly except the Zinc chloride was added to plex forms. Confirmation was observed by adding approxi 100 uL of water and the ether was allowed to evaporate prior mately 1 equivalent of 12 NHCl to the NMR sample and to adding to the reaction mixture. Work up and drying was reanalyzing by H NMR, which revealed reversion of the followed in the same manner to yield 47 mg of a free flowing Zinc complex back to hydrochlorothiazide and, presumably, white powder. The compound was sparingly soluble in Zinc chloride (FIG. 22). The product was further character DMSO resulting in low signal to noise ratio in the NMR. "H ized by MALDI-ES and ICP. MALDI-ES: Molecular ions at NMR (DMSO): & 4.90 (bs), 4.66 (bs), 4.50 (bs), 2.85 (s), 705.9 and 932.9 with typical zinc isotopic abundances but 2.80 (s). The singlet at 2.85 ppm indicates the presence of not indicative of any particular hydrochlorothiazide zinc dimethylbiguanide that is not complexed with zinc. Judging complex structure. Zinc analysis (ICP): Found 10.4%. FTIR from the integration of the two peaks at 2.85 and 2.80 ppm, (neat): cm 1027 (S=O stretch), 953 (S=O-Zn stretch): the ratio of free dimethylbiguanide to the zinc complex is absence of N H stretch at 1646. NMR, MALDI and ICP about 1:1. data clearly indicate the formation of a hydrochlorothiazide Zinc complex. It seems reasonable that the site of complex Preparation of Zinc Dimethylbiguanide Complex in Water ation may be on one or both of the Sulfonamide nitrogens of 0208 Dimethylbiguanide (33 mg) was dissolved in 1 mL hydrochlorothiazide. FTIR data suggest the presence of of water, after which 0.44 mL of 1 M potassium t-butoxide DMSO ligands, which by "H NMR integration the ratio of in t-butanol was added and the solution stirred for 10 HCTZ:DMSO is 1:1. Chemical shift of the DMSO methyl minutes. Zinc chloride in diethyl ether (0.22 mL of 1M groups of 0.08 ppm Suggests O-bonding between Zinc and solution) was added and the solution stirred for 5 hours. The DMSO. resultant precipitant was separated from the liquid by cen Reaction between Zinc and Hydrochlorothiazide in the trifuging at 8,000 rpm for 6 minutes and the supernatant was Presence of Water decanted. The pellet was washed by adding 1 mL of water, Vortex mixed, centrifuged and the Supernatant decanted. 0205 This procedure followed the analogous anhydrous After an overnight drying under high vacuum the yield was preparation exactly except the Zinc chloride was added to 20 mg of a free flowing white powder which contained no 100 uL of water and the ether was allowed to evaporate off organic material by "H NMR. The isolated product was zinc prior to adding to the reaction mixture. No product was salts in various hydrated forms. isolated because the entire mixture slowly dissolved in methanol during the precipitation step. This was putatively Preparation of Bis(acycloguanosinato)magnesium due to the formation of methanol soluble hydrated com 0209 Acycloguanosine (45 mg) was dissolved in 0.5 mL plexes. of anhydrous DMSO, after which 0.2 mL of 1 M potassium Preparation of Dimethylbiguanide Zinc Complex t-butoxide int-butanol was added and the solution stirred for 10 minutes. Magnesium chloride (11 mg) was added and the 0206 Dimethylbiguanide (66 mg) was dissolved in 1 mL solution stirred for 4 hours. The solution was concentrated in of anhydrous DMSO, after which 0.88 mL of 1 M potassium vacuo at 30° C., after which 0.5 mL of methanol was added t-butoxide int-butanol was added and the solution stirred for and the mixture triturated and transferred to a 2 mL micro 10 minutes. Zinc chloride in diethyl ether (0.22 mL of 1M centrifuge tube. The product was separated from the liquid solution) was added and the solution stirred for 3 hours. The by centrifuging at 8,000 rpm for 6 minutes and the super solution was concentrated in vacuo at 35°C., after which 0.5 natant was decanted. The pellet was washed by adding 0.5 mL of ethanol was added and the mixture triturated and mL of methanol, Vortex mixed, centrifuged and the Super transferred to a 2 mL microcentrifuge tube. The product was natant decanted. The washing procedure was repeated. After separated from the liquid by centrifuging at 8,000 rpm for 6 an overnight drying under high vacuum the yield was 15 mg minutes and the Supernatant was decanted. The pellet was of a beige coarse powder. The product structure was char washed by adding 0.5 mL of ethanol, vortex mixed, centri acterized by H NMR, MALDI-ES and ICP. H NMR fuged and the Supernatant decanted. The washing procedure (DMSO): 8 7.70 (s), 6.75 (bs), 5.30 (s), 4.64 (bs), 3.42 (s). US 2008/OO15352 A1 Jan. 17, 2008 20

MALDI-ES: Molecular ion at 473.1 is suggestive of bis(a- was heated at reflux for 18 hours. The reaction was cooled cycloguanosine)magnesium. Other molecular ions at 666, to room temperature and the precipitate was filtered thru a 702, 893 and 1065 have not been assigned to a particular fritted funnel (medium frit). The solid was washed with structure. Magnesium analysis (ICP): Expected 5.14%; EtOH causing the Suspension to pass thru the frit leaving an Found 4.16%. FTIR data did not indicate presence of a insoluble solid behind. The EtOH fraction was collected and DMSO ligand. From a combination of the absence of the concentrated to afford product. (Salicylato)zinc. "H NMR amide proton at 10.6 ppm, the ICP analysis and the MALDI (DMSO-d6) d7.46 (brs, 1H), 6.84 (brs, 1H), 6.36 (brs, 1H), ES analysis, the structure for bis(acycloguanosinato)magne 6.14 (brs, 1H). sium is shown in FIG. 29. 0215. A sample of the zinc salt of salicylic acid was 0210. The following examples illustrate the use as start prepared by reacting sodium salicylate (2 equiv) with ZnCl2 ing materials compounds that belong to any of the thio azole, (1 equiv) in HO at room temperature overnight. The azole amine, azole amide, ureide or certain purine chemical precipitate was filtered, washed with HO and EtOH to classes. DMAC is a preferred solvent, to which the base and afford zinc salicylate. "H NMR (DMSO-d6) d 7.76 (dd. 1H), the complexing metal are added as a single reagent (e.g., 7.30 (m. 1H), 6.76 (m. 2H), in agreement with literature Magnesium tert-butoxide). The mixture is heated, preferably values under reflux conditions and followed by removal of the Solvent in vacuo. 0216) The structure of (Salicylato)Zinc is as follows: Preparation of Magnesium Azathioprine Complex 0211 Azathioprine (azaH, 50 mg, 0.18 mmol) was dis solved in dimethylacetamide (5 mL) and Mg(OBut) (20.2 mg, 0.09 mmol, 90% purity) was added. The mixture was heated to reflux for 2.5 hours. The reaction turned orange. The reaction was cooled to room temperature and concen trated under reduced pressure to afford a solid material. Analysis by H NMR indicated formation of the chelate. "H NMR (DMSO-d6)d 8.35 (s, 1H), 8.20 (s, 1H), 8.18 (s, 1H). The H NMR (DMSO-d6) of azah is: d 8.60 (s, 1H, H2), 0217. The following examples illustrate the use as start 8.57 (s, 1H, H8), 8.26 (s, 1H, H11). ing materials compounds that belong to any of the thio acid, amino acid and certain B-ketoacid chemical classes. DMF is 0212. The structure of azathioprine is as follows: a preferred solvent, to which the base and the complexing metal are added as a single reagent (e.g., Magnesium tert butoxide). The mixture is stirred at room temperature and N NO followed by removal of the solvent in vacuo. CC Preparation of Bis(furosemide)magnesium N S 0218. Furosemide was obtained from Kemprotec, Ltd. HC 6 7 Solvents and reagents were obtained from Sigma-Aldrich N1 N N and were used without further purification. Oasis Hydro philic-Lipophilic Balanced (HLB) reversed phase Solid 2 ly2 Phase Extraction (SPE) cartridges were obtained from N. Waters Corporation. NMR spectra were obtained on a Varian 400 MHz instrument. Elemental analyses were performed at Galbraith Laboratories. Inductively Coupled Plasma (ICP) 0213 The following examples illustrate the use as start spectrometry was performed at REI Consultants. Reagents ing materials compounds that are very stable and have two for the calorimetric titration of Mg were obtained from Hach functional groups, both of which can be deprotonated. Such Corporation. as acid alcohols, diols, polyols (e.g. polysaccharides), dicar 0219. To a 25 mL round bottom flask equipped with boxylic acids and certain B-ketoacids. DMAC is a preferred magnetic stirrer was added 1.00 g (3.02 mmol) of furo Solvent, to which a strong base is added, such as potassium semide and 10 mL of anhydrous DMF. To this solution was t-butoxide, followed by the salt of the complexing metal, added 277 mg (1.51 mmol) of Mg(t-BuO). The suspension preferably the chloride salt, or the metal oxide. The mixture was stirred for 18 hat room temperature. During the course is then heated, preferably under reflux conditions. The of the reaction, the Suspension turned opaque. Solvent was preparation also includes concentrating the reaction mixture removed under reduced pressure at 50° C. and the yellow oil in vacuo, extracting the metal coordinated product with was further dried in a vacuum oven (50° C., 3 torr) over alcohol, and removing the alcohol in vacuo. night. The semi-solid material was dissolved in 3 mL (Salicylato)Zinc MeOH. Water (57 mL) was added and this solution was applied to an Oasis HLB solid phase extraction cartridge (35 0214 Sodium salicylate (1.60 g, 10 mmol) was dissolved cc. 6 g). The column was eluted sequentially with MeOH/ in DMAC (100 mL) and a solution of potassium tert HO fractions of increasing solvent strength. The fractions butoxide in THF (10 mL, 10 mmol. 1 N) was added were concentrated under reduced pressure as above to dropwise with stirring at room temperature. To this suspen afford: Fraction 1 (20% MeOH, 60 mL, 17 mg), fraction 2 sion was added ZnO (408 mg, 5.0 mmol) and the mixture (40% MeOH, 120 mL, 1.01 g), fraction 3 (60% MeOH, 60 US 2008/OO15352 A1 Jan. 17, 2008

mL, 18 mg), fraction 4 (80% MeOH 60 mL, 5.6 mg), 0225. The spectrophotometric method used a fiber optics fraction 5 (100% MeOH, 60 mL, 5 mg). Fraction 2 con dip probe, a UV light source (pulsed deuterium lamp) and a tained product as an off-white solid, 97% yield. Unreacted photodiode array detector to automatically capture the furosemide eluted in fraction 4. absorption spectra of the sample solution in the course of 0220 Melting point (Mp) decomposed above 200° C. "H adding an acid or base solution. NMR (DMSO-d6)D 8.42 (s, 1H), 7.54 (s, 1H), 7.07 (s. 2H), 0226 Up to a 10 mM stock solution was prepared by 6.75 (s, 1H), 6.35 (m, 1H), 6.26 (m. 1H), 4.39 (d. J=6.4 Hz, dissolving several 0.5 mg samples in 0.5-1.0 mL of water or 2H). C (DMSO-d6)d 170.8 (C12), 152.0 (C9), 151.8 (C6), cosolvent. An adequate amount of stock is pipetted into the 142.2 (C1), 133.8 (C10 or 7), 133.6 (C4), 125.6 (C8), 115.1 vial for titration. The dip probe is in the assay vial and (C10 or 7), 112.2 (C2 or 3), 110.3 (C11), 107.2 (C2 or 3), aqueous 0.15 M KCl solution is added to cover the dip 39.5 (C5). Metals analysis (total metals by ICP) 3.3000% probe. Acid or base is added to bring the pH to the desired Mg: Theory (Mgfur.H2O)3.41% Mg. Total hardness analy starting value. Over the chosen pH range, the spectra sis (EDTA titration, Calmagite indicator) 3.3%. Elemental changes due to ionization were captured by the photodiode analysis (C, H, N, Cl) 41.10, 3.20, 8.00, 9.72%: Theory array detector for Subsequent analysis. Target Factor Analy (Mgfur,.HO) 41.08, 3.16, 7.98, 10.10%. sis (TFA) was applied to deduce the pK values of the 0221) The structure of Bis(furosemide)magnesium is as sample and resolve the major absorptivity spectra of the follows: reducing species. The aqueous pKa was determined by extrapolation using the Yasuda-Shedlovsky technique. The pK values obtained from spectrophotometric experiments are in excellent agreement with those derived from poten tiometric titrations. C Bioavailability Study 0227 Title: Assessment of absorbance and effect of a HNOS ope hormone complex Supplementation in Sprague Dawley rats SONH2 0228 Test subjects: Fifteen young female Sprague Daw ley rats (180–225 gms) were used. These rats were obtained from a commercial source (Harlan Laboratory Animals, Dublin, Va.), housed in the Vivarium at Litton Reeves Hall (Division of Laboratory Animal Resources), in groups of three in polypropylene shoebox cages. Water was available ad libitum. Rats were fed certified rodent chow ad libitum. Partition Coefficients, Distribution Coefficients and pK for After arrival, the health of rats was assessed and animals T3 and Tetracycline Complexes and Reference Drugs were placed in quarantine for a minimum of five days, 0222 Determination of pK and log P was done by during which time, general health was assessed. At the end potentiometry and spectrophotometry. The potentiometric of quarantine, rats were moved to permanent animal quarters method includes the use of expert software to calculate pKa for access and study. and log P from simple acid and base titrations of the 0229 Study design: This is a study to compare the analytes. The pK was first determined by weighing approxi absorption and effect of three hormones administered to rats mately 2 mg of pure Substance into an assay vial. Ionic orally directly into the esophagus. T3-Zinc and T3-Magne strength was adjusted with 0.15M KCl and water was added sium were synthesized, which are the two test compounds. to dissolve the compound followed by an acid or base titrant Triiodothyronine, free acid (T3) was the positive control. to drop or raise the pH to the desired starting value. The The three compounds were formulated, separately, into solution was thentitrated with acid (0.5N HCl) or base (0.5N gelatin capsules, with a total dose of 108-12 Lug/kg admin NaOH) to the final pH. Approximate pK values were istered. In order to avoid acid degradation of the test displayed and later refined to exact data. compounds in the rat Stomach, metal oxides were added to 0223) For (T3),Zn, which is sparingly soluble in water, the formulation—to the T3-magnesium compounds, the pK was determined in mixtures of water and DMSO 1.08+0.13 mg of magnesium oxide was added and to the cosolvent. A minimum of three ratios of water/DMSO was T3-zinc compounds, 106-13 lug of zinc oxide was added. titrated to obtain pKa (apparent pKa in the presence of Another T3 control was included, where 145-50 ug of zinc cosolvent). The aqueous PK was determined by extrapola oxide was added to T3, free acid. tion using the Yasuda-Shedlovsky technique. 0230 For dosing and blood collection, three rats per 0224. The log P was determined by a titration in the replicate were anesthetized. A baseline blood sample was presence of octanol (water saturated). The pKa in water and collected, by retroorbital sampling. Then, compound was the apparent pKa in the presence of octanol (pKa) were administered orally by gavage tube. Following administra compared and the log P determined. Ion-pairing (partition tion, blood samples (500 uL) were collected at 0.5, 1, 2, 2.5, ing of a charged species into octanol, termed log P or log 5 hours after dosing. Blood samples were centrifuged, the P) were determined with an additional titration in the sera removed and the Sera kept on ice and then analyzed for presence of another Volume of octanol. Using experimen serum triiodothyronine levels. tally determined pKa and log P, a drug lipophilicity profile (log DVS. pH) was calculated. The log D7 was determined 0231 Analysis of samples: Serum triiodothyronine levels from this profile at pH 7.4. were determined by RIA. US 2008/OO15352 A1 Jan. 17, 2008 22

0232 Results: The individual serum T3 levels from each Preparation of Zinc:RNA Outer Coordination (Ionic) Com group of rats were averaged and a plot of T3 concentration plex (ng/mL) vs. hour was produced. The plot is shown in FIG. 0238. The preparation for the ionic zinc:RNA complex 3O. followed the procedure for the covalent analog exactly except the stock Zinc chloride solution was prepared in Large Molecule Example RNAse free water instead of anhydrous DMSO. The result ant colorless pellet was air-dried for several minutes before Preparation of Interference RNA testing in the isoelectric focusing gel. 0233 Interference RNA was prepared using a modified Isoelectric Focusing Gel Experiment New England Biolabs Litmus 281 RNAi bidirectional tran scription vector. A 922 bp bovine serum albumin cDNA 0239). This is a novel approach of using IEF gels to fragment was introduced into the Bgl II and Stul sites of the monitor for the anticipated modification of the iRNA target. Litmus RNAi vector. The target RNAi transcript was pro FIG. 31 shows the results from the initial experiment. duced by in vitro transcription with T7 RNA Polymerase to Interpretation of IEF Experiments yield 1 mg/ml. The RNA was then divided into 50 ug samples and freeze dried. 0240 The IEF experiment showed that magnesium RNA complexes prepared in anhydrous (A) conditions with three 0234. The following examples illustrate the use as start concentrations of magnesium chloride produced covalent ing materials compounds that belong to any of the oligo complexes in approximately 50% yield. Magnesium RNA nucleotide, phosphate or phosphonate chemical classes. The complexes prepared in aqueous (W) conditions with three drug will usually be available as the Sodium, potassium or concentrations of magnesium chloride produced ionic com quaternary ammonium salt. DMSO is a preferred solvent plexes. The zinc RNA complex prepared in anhydrous (A) and depending on the drug, gentle heating may be required, conditions with Zinc chloride produced a covalent complex followed by the addition of a buffer solution and precipita in approximately 50% yield. The Zinc RNA complex pre tion from alcohol. pared in aqueous (W) conditions with Zinc chloride pro Preparation of RNA: Magnesium Inner Coordination (Cova duced an ionic complex. lent) Complex 0235 Approximately 50 ug of iRNA was dissolved in We claim: 100 uL of anhydrous DMSO at 50° C. A stock solution of 4 1-36. (canceled) mM magnesium chloride was prepared by dissolving 19 mg 37. A metal coordination complex, comprising: of anhydrous magnesium chloride in anhydrous DMSO. a biologically active moiety and Three separate reactions were run where 15 uL, 30 uL and 60 uL of 4 mM magnesium chloride was added to three a metal atom, separate solutions of iRNA in DMSO. The solutions were allowed to set at room temperature with occasional Vortex wherein the biologically active moiety comprises a first mixing for 90 minutes at which time 20 uL of 7.5 M aqueous functional group having a heteroatom and second ammonium chloride followed by 400 uL of RNAse free functional group having a heteroatom, ethanol was added and vortex mixed. The product was wherein the first functional group and the second allowed to precipitate out of solution over 1 hour, centri functional group are capable of forming a metal fuged and the liquid decanted from the pellet. The pellet was coordination bond, washed with 100 uL of RNAse free ethanol, vortex mixed, centrifuged and the ethanol Supernatant decanted off the wherein the first functional group and the second pellet. The resultant colorless pellet was air-dried for several functional group are in a spatial relationship Such minutes before testing in the isoelectric focusing gel. that the first functional group and the second functional group are capable of participating in the Preparation of Magnesium:RNA Outer Coordination (Ionic) formation of a stable chelate with the metal atom, Complex and 0236. The preparation for the ionic magnesium:RNA wherein the biologically active moiety is not DOPA, complex followed the procedure for the covalent analog acycloguanosine, or tetracycline antibiotics. exactly except the Stock magnesium chloride solution was 38. The metal coordination complex of claim 37, wherein prepared in RNAse free water instead of anhydrous DMSO. the biologically active moiety is selected from the group The resultant colorless pellet was air-dried for several min consisting of Acetalzolamide, Vorinostat, Aliskiren, Alvimo utes before testing in the isoelectric focusing gel. pan, Bicalutamide, Baclofen, Balsalzide, Brinzolamide, Preparation of RNA:Zinc Inner Coordination (Covalent) Chlorproguanil, Diclofenac, Dorzolamide, Droxidopa, Complex Clonixin, Ebrotidine, Enfenamic acid, Ethylmethyltham butene, Etodolac, Flufenamic acid, Fosfosal, Lazabemide, 0237) The preparation for the covalent zinc:RNA com Mefenamic acid, 6-Mercaptopurine, Melagatran, Myco plex followed the procedure for the magnesium analog pheolate mofetil, Pregabalin, Quinocide, Rilmazafone, Taf exactly except a stock Zinc chloride solution was prepared enoquine, Tilarginine, Tolfenamic acid, Cidofovir, instead of a stock magnesium chloride solution. The result Didanosine, Dideoxyadenosine, Etidronic acid, Moroxy ant colorless pellet was air-dried for several minutes before dine, Nelfinavir, Pamidronic acid, Risedronic acid, and testing in the isoelectric focusing gel. Zoledronic acid. US 2008/OO15352 A1 Jan. 17, 2008

39. The metal coordination complex of claim 37, wherein 50. A method of modulating the properties of a biologi the metal coordination bond forms a 4 to 8 atom ring cally active moiety, comprising: encompassing the metal atom, the first functional group, and the second functional group. forming a metal coordination complex with the biologi 40. The metal coordination complex of claim 39, wherein cally active moiety and a metal atom, the 4 to 8 atom ring is free of trans double bonds. 41. The metal coordination complex of claim 37, wherein wherein the biologically active moiety comprises a first at least one of the heteroatoms is selected from the group functional group having a heteroatom and second consisting of oxygen and Sulfur when the metal atom is functional group having a heteroatom, selected from the group consisting of magnesium and cal cium. wherein the first functional group and the second 42. The metal coordination complex of claim 37, wherein functional group are capable of forming a metal the first functional group comprises a heteroatom selected coordination bond, from the group consisting of nitrogen and Sulfur. wherein the first functional group and the second 43. The metal coordination complex of claim 37, wherein functional group are in a spatial relationship Such the first functional group and the second functional group that the first functional group and the second comprise a heteroatom selected from the group consisting of functional group are capable of participating in the nitrogen and Sulfur. formation of a stable chelate with the metal atom, 44. The metal coordination complex of claim 37, wherein and the heteroatom is selected from the group consisting of oxygen, Sulfur, and nitrogen. wherein the biologically active moiety is not DOPA, 45. The metal coordination complex of claim 37, wherein acycloguanosine, or tetracycline antibiotics. the metal atom is selected from the group consisting of 51. The method of claim 50, wherein the metal atom is aluminum, bismuth, calcium, iron, magnesium, silicon, Zinc, selected from the group consisting of aluminum, bismuth, Vanadium, chromium, cobalt, nickel, and copper. calcium, iron, magnesium, silicon, Zinc, Vanadium, chro 46. A metal coordination complex, comprising: mium, cobalt, nickel, and copper. a biologically active moiety and 52. The method of claim 50, wherein the biologically active moiety is selected from the group consisting of a metal atom, Acetalzolamide, Vorinostat, Aliskiren, Alvimopan, Bicaluta wherein the biologically active moiety comprises a first mide, Baclofen, Balsalzide, Brinzolamide, Chlorproguanil, protonated heteroatom and a second protonated het Diclofenac, Dorzolamide, Droxidopa, Clonixin, Ebrotidine, eroatom, Enfenamic acid, Ethylmethylthambutene, Etodolac, Flufe namic acid, Fosfosal, Lazabemide, Mefenamic acid, 6-Mer wherein the biologically active moiety has a pKa of captopurine, Melagatran, Mycopheolate mofetil, Pregabalin, approximately equal to water or lower than water, Quinocide, Rilmazafone, Tafenoquine, Tilarginine, Tolfe wherein the first protonated heteroatom and the namic acid, Cidofovir, Didanosine, Dideoxyadenosine, second protonated heteroatom are in a spatial Etidronic acid, Moroxydine, Nelfinavir, Pamidronic acid, relationship such that the first protonated heteroa Risedronic acid, and Zoledronic acid. tom and the second protonated heteroatom are 53. A method of claim 50, further comprising capable of participating in the formation of a adding an adjuvant selected from the group consisting of stable chelate with the metal atom, and amino acids, peptides, carbohydrates and lipids that wherein the biologically active moiety is not DOPA, confer desired performance parameters to the biologi acycloguanosine, or tetracycline antibiotics. cally active moiety. 47. The metal coordination complex of claim 46, wherein 54. The method of claim 50, wherein the properties to be the metal atom is selected from the group consisting of modulated are selected from the group consisting of potency, transition metals, p-block metals, and S-block metals. stability, absorbability, targeted delivery, and combinations 48. The metal coordination complex of claim 46, wherein thereof. the metal atom is selected from the group consisting of 55. A method of modulating the properties of a biologi aluminum, bismuth, calcium, iron, magnesium, silicon, Zinc, cally active moiety, comprising: Vanadium, chromium, cobalt, nickel, and copper. 49. The metal coordination complex of claim 46, wherein forming a metal coordination complex with the biologi the biologically active moiety is selected from the group cally active moiety and a metal atom, consisting of Acetalzolamide, Vorinostat, Aliskiren, Alvimo pan, Bicalutamide, Baclofen, Balsalzide, Brinzolamide, wherein the biologically active moiety comprises a first Chlorproguanil, Diclofenac, Dorzolamide, Droxidopa, functional group having a heteroatom and second Clonixin, Ebrotidine, Enfenamic acid, Ethylmethyltham functional group having a heteroatom, butene, Etodolac, Flufenamic acid, Fosfosal, Lazabemide, wherein the first functional group and the second Mefenamic acid, 6-Mercaptopurine, Melagatran, Myco functional group are capable of participating in the pheolate mofetil, Pregabalin, Quinocide, Rilmazafone, Taf formation of a metal coordination bond, enoquine, Tilarginine, Tolfenamic acid, Cidofovir, Didanosine, Dideoxyadenosine, Etidronic acid, Moroxy wherein the first functional group and the second dine, Nelfinavir, Pamidronic acid, Risedronic acid, and functional group are in a spatial relationship Such Zoledronic acid. that the first functional group and the second US 2008/OO15352 A1 Jan. 17, 2008 24

functional group are capable of participating in the wherein the metal is selected from the group consisting formation of a stable chelate with the metal atom, of aluminum, bismuth, calcium, iron, magnesium, wherein the biologically active moiety is selected from silicon, Zinc, Vanadium, chromium, cobalt, nickel, the group consisting of Acetalzolamide, Vorinostat, and copper. Aliskiren, Alvimopan, Bicalutamide, Baclofen, Bal 56. A method of claim 55, further comprising Salzide, Brinzolamide, Chlorproguanil, Diclofenac, adding an adjuvant selected from the group consisting of Dorzolamide, Droxidopa, Clonixin, Ebrotidine, amino acids, peptides, carbohydrates, and lipids that Enfenamic acid, Ethylmethylthambutene, Etodolac, confer desired performance parameters to the biologi Flufenamic acid, Fosfosal, Lazabemide, Mefenamic cally active moiety. acid, 6-Mercaptopurine, Melagatran, Mycopheolate 57. The method of claim 55, wherein the properties to be mofetil, Pregabalin, Quinocide, Rilmazafone, Taf modulated are selected from the group consisting of potency, enoquine, Tilarginine, Tolfenamic acid, Cidofovir, stability, absorbability, targeted delivery, and combinations Didanosine, Dideoxyadenosine, Etidronic acid, thereof. Moroxydine, Nelfinavir, Pamidronic acid, Risedronic acid, and Zoledronic acid, and