US 20130245070A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2013/0245070 A1 Ratan et al. (43) Pub. Date: Sep. 19, 2013
(54) COMPOUNDS FOR ENHANCING ARGINASE A63L/36 (2006.01) ACTIVITY AND METHODS OF USING SAME A613 L/352 (2006.01) A613 L/484 (2006.01) (75) Inventors: Rajiv R. Ratan, Scarsdale, NY (US); A63L/05 (2006.01) Marie T. Filbin, New York, NY (US) A63L/22 (2006.01) (52) U.S. Cl. (73) Assignee: THE RESEARCH FOUNDATION OF CPC ...... A6 IK3I/4439 (2013.01); A61K3I/05 THE CITY UNIVERSITY OF NEW (2013.01); A61 K3I/085 (2013.01); A61 K YORK, New York, NY (US) 31/122 (2013.01); A61 K3I/352 (2013.01); A6 IK3 L/484 (2013.O1): 46 IK3I/36 (21) Appl. No.: 13/416,885 ( ); (2013.01) USPC ...... 514/338; 514/733; 514/720; 514/680; (22) Filed: Mar. 9, 2012 514/453: 514/395; 514/456; 514/455 Related U.S. Application Data (57) ABSTRACT (63) Continuation of PE i 1i. filed O The present invention relates to a method for enhancing argi Riis.O3, s 8 opp ication No. nase activity in a damagedor injured cell. In other aspects, the On Jul. TRS, invention provides a method for treating a disorder that can be (60) Provisional application No. 60/807,661, filed on Jul. treated by enhancing arginase activity in a human in need 18, 2006. thereof, the method comprising administering to the human an effective amount of a compound that enhances arginase Publication Classification activity. Such disorders include ischemia, hypoxia, neurode generative disease or condition, stroke or trauma of the ner (51) Int. Cl. Vous system. In yet another aspect, the invention provides A6 IK3I/4439 (2006.01) methods for promoting regeneration of a neural cell in a A6 IK3I/085 (2006.01) human in need thereof. Patent Application Publication Sep. 19, 2013 Sheet 1 of 38 US 2013/0245070 A1
Figure 1
Chemical Reference Formula
Pinosylvin C14H2O2 J. Ann. Chen, Soc., 1940, 62:3512
J. Chen, Soc., 1959, 2679 Phytochem istry 1968, 7:701 Phytochem istry 1972, 1:089 Methoxyvone Patent Application Publication Sep. 19, 2013 Sheet 2 of 38 US 2013/0245070 A1
Dehydrovariabilin Phytochern istry 1978, 17:1417
Chrysophanol Phytochen istry 1972, 11:2122 Patent Application Publication Sep. 19, 2013 Sheet 3 of 38 US 2013/0245070 A1
MOLENAME rom ocular Wit OO 102007 Formononetn C15H12O4 268.3
2- Apigenin triacetate C21H16O8 396.36 fold DO2005O2 2- Xanthone C3H8O2. 196.2 fold OO200523 90-47-1 2- Daidzeir C15H10O4 254.2
Acacetin diacetate C20H16O7 368.3 OO2OO833 270.2 OO200846 Apigenin
C14H12O2 212.3 OO2O1066 Pinosylvin oozoos Pinosytvin methyl ether C15H14O2 226.3
C21H18O6 366.4 002O1650 Derrubone
OO2O296 Genistein C5H10O5 270.2
OO210454 Robustone
OO210554 4,7-dimethoxyflavone C7H14O4. 282,3
2- Dehydrovariabilin C17H4O4. 282.3 fold 00210658 2- 4'-methoxychalcone C15H14O2 238.3
fold OO211475 2- 5,7-dimethoxyisoflavone C17H1404 26964-35 fold OO240576 282.3 2 17 2- Retusin 7-methyl ether C17H14O5 298.3 fold OO24O645
OO2.40651 2-hydroxyxanthone
5,7,4'-trimethoxyflavone C18H16O5 312.3 Patent Application Publication Sep. 19, 2013 Sheet 4 of 38 US 2013/0245070 A1
Chemo MOENAME Forua
OO3OO545 Chrysophanol
2 old 3-methylcholanthrene C2H16 268.4
praflavone C18H16O3 fold 3. Methoxyvone C17H4O3
2 f o d oratorm site
o15.00351 IndoProfen C17H15NO3
O1500473 Phenazopyridine CH12CN5 hydrochloride
Penindice C15H10O2 222.2
Ferbedazole C15H13N3O2S
o1501.016
O1502198. Anisidine C6H12O3 242.3 O154044 Resver atrol 4'-methyl ether C15H14C3
o1504132 -dimethoxyflavone C 1714O4. 282,3
Dibenzoylmethane 224.3 OSO5311 120-146-7 Tranilast (aka MK-341) 53902-12. O1505.333 8
Tilbore C25H34N2O3
101 OOOO3 Biochanin a Patent Application Publication Sep. 19, 2013 Sheet 5 of 38 US 2013/0245070 A1
I creou roma ow a fold | | | | | Patent Application Publication Sep. 19, 2013 Sheet 6 of 38 US 2013/0245070 A1
Figure 3
better than Chern Name Wel control x2 activity in Tilorone primary screen AO2 O2300009 x2 activity in Phenindione primary Screen . AO3 O1500477 not quite 2x data O1501139 Pamoxine in primary hydrochloride screers 3. x2 activity in Indoprofen primary screen O1500351 Phenazopyridine x2 activity in hydrochloride primary screen 5 AO6 O1500473 not quite 2x data in primary Piperine Sctees 6 AO7 O1500B73 x2 activity in 6,3'-dimethoxyflavone primary screen 7 AO8 O1504132 x2 activity in Anisindione primary screen 8 AO9 O1502198. not quite 2x data in primary 5,4'-dimethoxyflavone Screens A10 O0211227 x2 activity in primary screen X x2 activity in Deriustone primary Screen 11 BO2 X XX x2 activity in 4,7-dimethoxyflavone primary screen 2 BO3 00210554 x2 activity in Daidzeir primary screen 13 B04 00200789 X
x2 activity in 4'-methoxychalcone primary screen O02.11475 X x2 activity in Traniast primary screen 15 B06 O1505.333 X x2 activity in Biochanin a diacetate primary screen
Resveratrol 4'-methyl x4 activi Patent Application Publication Sep. 19, 2013 Sheet 7 of 38 US 2013/0245070 A1
Western Blot RT-PCR better than Numwa control X4 activity in Derrubone primary screen BC 9 002O1650 x2 activity in Chlorpropham primary screer B10 003.30051
x2 activity in . Genistein primary screen 20 B11 O0210296 x2 activity in
Dehydrovariabilin primary screen CO2 00210658 X X X x2 activity in Retusin 7-methyl ether primary screen 22 CO3 0024O645 x2 activity in Xanthone primary screen 23 C4 OO2O0523
x2 activity in Pinosylwin methyl ether primary screen 24 CO5 002O1067 x2 activity in Chrysophanol primary screen 25 CO8 00300545 X X X x2 activity in Apigenin primary screen CO7 00:200846 not quite 2x data in primary 2-methoxyxanthone SCfeeS 27 CO8 0024.0736 x2 activity in Apigenin triacetate primary screen 28 CO9002005O2 x2 activity in Fenbendazole primary screen 29 C10 O150.1016 x2 activity in Dibenzoylmethane primary screen 30 0.150531 x2 activity in Methoxyvone primary Screen 31 DO2 O1400666 X X
x4 activity in Ginkgetin, k salt primary screen 32 DO3 00200436 x4 activity in Methyl robustone primary screen 014O401 not quite 2x data Liquiritigenin dinnethyl in primary ether Screens O60.0561 x2 activity in
primary screen DOS 014O1406 Patent Application Publication Sep. 19, 2013 Sheet 8 of 38 US 2013/0245070 A1
x2 activity in primary screen
lot quite 2x data in primary 4'-methoxyflavone SC eS x2 activity in Acacetin diacetate primary screen Patent Application Publication Sep. 19, 2013 Sheet 9 of 38 US 2013/0245070 A1
12 3 415 1617 18 9 20
3.33. Patent Application Publication Sep. 19, 2013 Sheet 10 of 38 US 2013/0245070 A1
Figure 5
2222222228.2939CSix s 3 ------32 33 34 35 36 37 38 39 40 C ES t fg i : "...rge
Patent Application Publication Sep. 19, 2013 Sheet 11 of 38 US 2013/0245070 A1
Figure 6
Arg-Screen/Custom Plate/HT22 COmOund #1-10
l 4 e l e Z Y E
s S
an s
e (v.
w Patent Application Publication Sep. 19, 2013 Sheet 12 of 38 US 2013/0245070 A1
Figure 7
Arg-Screen/Custom Plate Compound #11-20
4
3
2
Patent Application Publication Sep. 19, 2013 Sheet 13 of 38 US 2013/0245070 A1
Figure 8
+ Argil-Screen/Custom Plate Compound #21-30 Patent Application Publication Sep. 19, 2013 Sheet 14 of 38 US 2013/0245070 A1
Figure 9
Argl-Screen/Custom Plate Compound #31-40
4
3
2
Patent Application Publication Sep. 19, 2013 Sheet 15 of 38 US 2013/0245070 A1
Figure 10
Overcoming MAG inhibition in vitro
Post-treatment
Acetaminphen Pinosylvin Resver atol 4-methyl ether
4. Chrysophanol (low dose) 5 daidzein
dehydrovariabilin
CAPE
11 deriustone Epicatechin pentaacetate
Lansoprazole N/A Patent Application Publication Sep. 19, 2013 Sheet 16 of 38 US 2013/0245070 A1
Figure 11
burke drugs 10 mm
control dimso rho 2 4. 6 10 Patent Application Publication Sep. 19, 2013 Sheet 17 of 38 US 2013/0245070 A1
Figure 12
12
OO
oo:: Patent Application Publication Sep. 19, 2013 Sheet 18 of 38 US 2013/0245070 A1
Figure 13
Overcoming MAG inhibition in vitro
Prstreate Post treatnet
daidzein Yes dehydrovariabilin None Patent Application Publication Sep. 19, 2013 Sheet 19 of 38 US 2013/0245070 A1
Figure 14A
Patent Application Publication Sep. 19, 2013 Sheet 20 of 38 US 2013/0245070 A1
Figure 14B
Patent Application Publication Sep. 19, 2013 Sheet 21 of 38 US 2013/0245070 A1
Figure 14C
Niethopyvone spil on AAG. Patent Application Publication Sep. 19, 2013 Sheet 22 of 38 US 2013/0245070 A1
Figure 14D
is a Daidzein 2pati See als. . . . e. . . Patent Application Publication Sep. 19, 2013 Sheet 23 of 38 US 2013/0245070 A1
Figure 14E
anzoprazole 20pm on iMAG as wers arrass 8x8 wrx . . a. s. were Patent Application Publication Sep. 19, 2013 Sheet 24 of 38 US 2013/0245070 A1
Figure 14F
Patent Application Publication Sep. 19, 2013 Sheet 25 of 38 US 2013/0245070 A1
Figure 15
Arginase Actin Patent Application Publication Sep. 19, 2013 Sheet 26 of 38 US 2013/0245070 A1
Figure 16
MOLENAME At nates Structure
2-hydroxyxanthone 2-hydroxy- 9H xanther- 9- one
2-methoxyxanthone 2-methoxy- 9H xanthen- 9- one
3 3- methyl-1,2- methylcholanthrene dihydrocyclopentaijitetr aphene
4,7- 7-methoxy-2-(4- dimethoxyflavone methoxyphenyl)-4H chronen-4-one (IUPAC)
4'- (2E)-1-(4- methoxychalcone methoxyphenyl)-3- phenylprop-2-en-1-one (IUPAC), 1-(4- Tethoxyphenyl)-3- phenylprop-2-en-1-one (IUPAC) Patent Application Publication Sep. 19, 2013 Sheet 27 of 38 US 2013/0245070 A1
MOLENAME Structure 4'-methoxyflavone 2-(4-methoxyphenyl)- 4-chromen-4-one (IUPAC),
5,4'- 5-methoxy-2-(4- dimethoxyflavone methoxyphenyl)-4-h- chromen-4-one (IUPAC)
5,7,4'- 5,7-dimethoxy-2-(4- trimethoxyflavone methoxyphenyl)-4H chromen-4-one (IUPAC)
5,7- 5,7-dimethoxy-3- dimethoxyisoflavon phenyl-4H-chromen-4- e one (IUPAC)
6,3'- 6-methoxy-2-(3- dimethoxyflavone methoxyphenyl)-4H chromen-4-one (IUPAC)
Acacetin diacetate 2-(4-methoxyphenyl)-4- oxo-4H-chronene-5,7- diyi diacetate Patent Application Publication Sep. 19, 2013 Sheet 28 of 38 US 2013/0245070 A1
MOENAME Alt names
Anisindone 2-(4-methoxyphenyl)- 1H-indene-1,3(2H) - diore
Apigenin 4,5,7-trihydroxyflavone (chemical), 4H-1- benzopyran-4- one, 5,7-dihydroxy-2- (4-hydroxyphenyl)- (chemical), 5,7,4'- trihydroxy-flavone (chemical), 5,7- dihydroxy-2-(4- hydroxyphenyl)- 4H chromen- 4- one (IUPAC), 83244 (NSC), apigenin (common, vendor, MeSH, primary common), apigenine (common), apigenol (cornthon), C. l. natural yellow 1 (common), flavone,4,5,7- trihydroxy- (chemical), NP-000448 (extract), pelargidenon 1449 (common), spigenin (common), versulin (common)
Apigenin triacetate 2-(4-acetoxyphenyl)- 4-oxo- 4H-- chromene 5,7-diyl diacetate Patent Application Publication Sep. 19, 2013 Sheet 29 of 38 US 2013/0245070 A1
MOLENAME Structure
5,7-diydroxy-3-(4- methoxyphenyl)- 4H chronet- 4- one
Biochanin a 3-(4-methoxyphenyl)-4- diacetate oxo-4H-chronene-5,7- diyl diacetate
Chlorpropham isopropy (3- chlorophenyl) caroarrate
Chrysophanol 1,8-dihydroxy-3-methyl 9,10-anthraquinone Patent Application Publication Sep. 19, 2013 Sheet 30 of 38 US 2013/0245070 A1
Daidzein 4,7-dihydroxy-iso flavone (chemical), 4',7- dihydroxyisoflavone (chemical), 4H-1- benzopyran-4-one,7- hydroxy-3-(4- hydroxyphenyl)- (chemical), 7,4'- dihydroxyisoflavone (chemical), 7-hydroxy 3-(4-hydroxyphenyl)-4- benzopyrone (chemical), 7-hydroxy 3-(4-hydroxyphenyl)- 4H-1-benzopyran-4- one (chemical), 7 hydroxy-3-(4- hydroxyphenyl)-4H chromen-4-ore (IUPAC), 7-hydroxy-3- (4-hydroxyphenyl) chromen-4-one (chemical), daidzein (common, vendor, MeSH, primary common), daidzeol (common)
Dehydrovariabilin 3,9-dimethoxy-6H 1)benzofuro3,2- cchromene Patent Application Publication Sep. 19, 2013 Sheet 31 of 38 US 2013/0245070 A1
MOLENAWE Alt names
Derrubone 3-(1,3-benzodioxol-5- yl)-5,7-dihydroxy-6-(3- methylbut-2-en-1-yl)- 4H-chronen-4-one
Derrusnin 3-(1,3-benzodioxol-5- yl)-4,5,7-trimethoxy-2H chofne-2-one
Deriustone 3-(1,3-benzodioxol-5- yl)-5,7-dimethoxy-4H chromen-4-one
Dibenzoylmethane 1,3-diphenylpropane 1,3-dione
Fenbendazole methyl 6-(phenylthio) - 1H-benzimidazol-2- yl)carbamate Patent Application Publication Sep. 19, 2013 Sheet 32 of 38 US 2013/0245070 A1
MOLENAME At names Structure Formononetr 7-hydroxy-3-(4- methoxyphenyl)- 4H chromen- 4- one
Genistein 36586 (NSC), 4'5,7- trihydroxyisoflavone (chemical), 4H-1-. benzopyran-4-one, 5,7- dihydroxy-3-(4- hydroxyphenyl)- (chemical), 5,7,4'- trihydroxyisoflavone (chemical), 5,7- dihydroxy-3-(4- hydroxyphenyl)-4- benzopyrone (chemical), 5,7- dihydroxy-3-(4- hydroxyphenyl)-4H chromen-4-one (IUPAC), differenol A (common), genistein (common, MeSH, primary-common), genisteol (common), genisterin (common), isoflavone,4,5,7- trihydroxy-(chemical), NP-001561 (extract), prunetol (common), sophoricol (common)
Patent Application Publication Sep. 19, 2013 Sheet 34 of 38 US 2013/0245070 A1
MOLENAME Alt names Structure
Lansoprazole 2- (3-methyl-4- (prevacid) (2,2,2- trifluoroethoxy) pyridin-2- yl)methyl)sulfinyl)-1H benzinidazole (IUPAC), 2-(S)- {(3- methyl-4-(2,2,2- trifluoroethoxy) pyridin 2-yl)methyl)sulfinyl 1H-benzimidazole (IUPAC)
Liquiritigenin (2R) - 7-methoxy-2- dimethyl ether (4-methoxyphenyl)- 2,3-dihydro-4H chromen-4-one) (note: liquiritigenin is also called 7,4'- dihydroxyflavarone and 5-deoxyflavone Patent Application Publication Sep. 19, 2013 Sheet 35 of 38 US 2013/0245070 A1
Structure OLENAME At ranes
Methoxyvone 7-methoxy-5-methyl-2- phenyl-4H-chromen-4- one
Methyl robustone 7-(1,3-benzodioxol-5- yl)-5-methoxy-2,2- dimethyl-2H,6H pyrano3,2-gchromen 6-one
Phenazopyridine 3-(phenyldiazeny) hydrochloride pyridine-2,6-diamine (IUPAC)
Phenindone 2-phenyl-1H-indene 1,3(2H)-dione (IUPAC)
Pinosylwin 5-(E)-2- phenyvinylbenzene 1,3-diol (IUPAC)
Pinosylwin methyl 3-methoxy-5-(E)-2- ether phenylvinylphenol (IUPAC) Patent Application Publication Sep. 19, 2013 Sheet 36 of 38 US 2013/0245070 A1
NOENAME Structure
Piperine 1-(2E,4E)-5-(1,3- benzodioxol-5-yl)penta 2,4-dienoylpiperidine (IUPAC), 1-(5-(1,3- benzodioxol-5-yl)penta 2,4-dienoylpiperidine (IUPAC),
Pramoxine 25573 (NSC), 4-3-(4- hydrochloride butoxyphenoxy) propylimorpholine (IUPAC),
Resvei atrol 4'- 5-(E)-2-(4- methyl ether methoxyphenyl) vinyl)benzene-1,3-diol
Retusin 7-methyl 8-hydroxy-7-methoxy ether 3-(4-methoxyphenyl)- 4-chrone-4-core (UPAC) Patent Application Publication Sep. 19, 2013 Sheet 37 of 38 US 2013/0245070 A1
MOLENAME At farmes Structure
Robustone 7-(1,3-benzodioxol-5- yl)-5-hydroxy-2,2- dimethyl-2H,6H pyrano3,2-gchromen 6-one (IUPAC)
Spironolactone S (7R,8R,9S,10R,13S, 14 S,17R)-10,13-dimethyl 3,5'-dioxo 1,2,3,4,5,6,7,8,9,10,11 ,12,13,14,15, 16 hexadecahydro-3'H- spirocyclopertaalphen anthrene-17,2'-furan-7- yl)ethanethioate (IUPAC)
Tilloroe 2,7-bis(2-(diethylamino) ethoxy-9H-fluoren-9- one (UPAC) Patent Application Publication Sep. 19, 2013 Sheet 38 of 38 US 2013/0245070 A1
Tranilast (aka MK- 2-((2E)-3-(3,4- 341) dimethoxyphenyl)prop 2-enoyl)amino)benzoic acid (IUPAC)
Xanthone 9H-xanthen-9-one (IUPAC) US 2013/0245070 A1 Sep. 19, 2013
COMPOUNDS FOR ENHANCING ARGINASE method comprises administering to a human in need thereof ACTIVITY AND METHODS OF USING SAME with an effective amount of one of the following compounds: 2-hydroxyXanthone; 2-methoxyxanthone; 3-methylcholan CROSS REFERENCE TO RELATED threne; 4,7-dimethoxyflavone; 4'-methoxychalcone; 4'-meth APPLICATIONS oxyflavone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyfla vone; 5,7-dimethoxyisoflavone; 6.3'-dimethoxyflavone: 0001. This application is a continuation of U.S. Ser. No. Acacetin diacetate; Anisindione: Apigenin; Apigenin triac 12/309,500, filed on Jun. 23, 2009, abandoned, which is a 371 etate; Biochanina, Biochanin a diacetate; Chlorpropham; application of PCT/US2007/016335, filed Jul. 18, 2007, Chrysophanol; Daidzein, Dehydrovariabilin: Derrubone; which claims the benefit of U.S. Provisional Application Ser. Derrusnin; Derrustone; Dibenzoylmethane: Fenbendazole; No. 60/807,661, filed on Jul. 18, 2006. The contents of the Formononetn; Genistein; Ginkgetin; Indoprofen; Iprafla aforementioned prior applications are incorporated herein by vone: Liquiritigenin dimethyl ether; Methoxyvone; Methyl reference in their entireties. robustone; Phenazopyridine hydrochloride; Phenindione: 0002. The invention was made with funds from New York Pinosylvin; Pinosylvin methyl ether; Piperine; Pramoxine State Department of Health, contract number CO19772. New hydrochloride: Resveratrol 4'-methyl ether; Retusin 7-methyl York State has certain rights in this invention. ether; Robustone; Spironolactone; Tilorone; Tranilast; or BACKGROUND OF THE INVENTION Xanthone. 0003. It is reported that the adult mammalian central ner 0011. In another aspect, the invention provides a method Vous system (CNS) shows little spontaneous regeneration for enhancing arginase activity in a damaged or injured cell. after injury despite that fact that there are many molecules The method comprises administering to a human in need present which promote nerve and axonal growth. In contrast thereof an effective amount of Lansoprazole. to the CNS, the adult peripheral nervous system (PNS) is 0012. In another aspect, the invention provides a method capable of regenerating to Some extent. for treating a disorder that can be treated by enhancing argi 0004. It is believed that the lack of regeneration in the CNS nase activity in a human in need thereof. The method com is caused by the presence of molecules which actively prevent prises administering to the human an effective amount of a or inhibit regeneration. Such molecules include Nogo (an compound that enhances arginase activity, wherein the com antigen of the IN-1 antibody), myelin-associated glycopro pound is any one of the following: 2-hydroxyXanthone; tein, and myelin-oligodendrocyte glycoprotein. 2-methoxyxanthone: 3-methylcholanthrene; 4,7-dimethoxy 0005 Arginase is an enzyme that catalyzes the conversion flavone; 4'-methoxychalcone; 4'-methoxyflavone; 5,4'- of the amino acid arginine to urea and ornithine. Arginase has dimethoxyflavone: 5,7,4'-trimethoxyflavone; 5,7- been reported to reverse the inhibition of neural regeneration dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin in the central and peripheral nervous system. Thus, enhancing diacetate; Anisindione: Apigenin; Apigenin triacetate; Bio arginase activity would be beneficial for reversing the inhibi chanina, Biochanin a diacetate; Chlorpropham; Chrysopha tion of neural regeneration. nol; Daidzein: Dehydrovariabilin: Derrubone; Derrusnin; 0006 Arginase I is a 35- to 38-kDa cytoplasmic protein Derrustone; Dibenzoylmethane; Fenbendazole; Formonon that cleaves arginine into urea and ornithine. Arginine is the etn; Genistein, Ginkgetin; Indoprofen; Ipraflavone; Liquiriti only Substrate capable of donating the guanidine group nec genin dimethyl ether; Methoxy vone; Methyl robustone: essary for nitric oxide production. Nitric oxide is produced Phenazopyridine hydrochloride; Phenindione; Pinosylvin; from arginine by three nitric oxide synthase (NOS) isoforms. Pinosylvin methyl ether, Piperine: Pramoxine hydrochloride: Nitric oxide production can be regulated by modulating the Resveratrol 4'-methyl ether; Retusin 7-methyl ether; Robus levels of arginine. Arginase I can limit the pool of arginine tone; Spironolactone; Tilorone; Tranilast; or Xanthone. available for nitric oxide synthase (NOS), thereby influencing 0013. In yet another aspect, the invention provides a the production of nitric oxide. method for treating a disorder that can be treated by enhanc 0007 Neuronal damage can be caused by excess levels of ing arginase activity in a human in need thereof. The method nitric oxide (NO). NO is a diffusible neuronal second mes comprises administering to the human an effective amount of senger synthesized in the nervous system by three enzymes: Lansoprazole. neuronal NO synthase, endothelial NO synthase, and induc ible NO synthase. Excess NO generated by NO synthase is 0014. In a further aspect, the invention provides a method promoting regeneration of a neural cell in a human in need associated with various neurodegenerative diseases and con thereof. The method comprises administering to the human ditions, such as multiple Sclerosis, dementia, Huntington's an effective amount of a compound that enhances arginase disease, Alzheimer's disease, etc. activity, wherein the compound is any one of the following: 0008. The amino acid arginine is the only endogenous 2-hydroxyXanthone; 2-methoxyxanthone; 3-methylcholan substrate of NO synthase. It is reported that arginase can threne; 4,7-dimethoxyflavone; 4'-methoxychalcone; 4'-meth reduce cell death in the nervous system by competing with oxyflavone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyfla NO synthase for their common Substrate, arginine. vone; 5,7-dimethoxyisoflavone; 6.3'-dimethoxyflavone: 0009. Therefore, enhancing arginase activity would be Acacetin diacetate; Anisindione: Apigenin; Apigenin triac beneficial for promoting neural regeneration or reducing neu etate; Biochanina, Biochanin a diacetate; Chlorpropham; ral damage in diseases and conditions associated with neural Chrysophanol; Daidzein, Dehydrovariabilin: Derrubone; damage. Derrusnin; Derrustone; Dibenzoylmethane: Fenbendazole; Formononetn; Genistein; Ginkgetin; Indoprofen; Iprafla SUMMARY OF THE INVENTION vone: Liquiritigenin dimethyl ether; Methoxyvone; Methyl 0010. The present invention provides a method for robustone; Phenazopyridine hydrochloride; Phenindione: enhancing arginase activity in a damaged or injured cell. The Pinosylvin; Pinosylvin methyl ether; Piperine; Pramoxine US 2013/0245070 A1 Sep. 19, 2013
hydrochloride: Resveratrol 4'-methyl ether; Retusin 7-methyl 0031 FIG. 14C. P5 cerebellar neurons were plated on ether; Robustone; Spironolactone; Tilorone; Tranilast; or substrate inhibitor MAG expressing CHO cells and treated Xanthone. with methoxyvone (5 uM). Picture is representative of BIII 0015. In yet a further aspect, the invention provides a tubulin positive cells. method for promoting regeneration of a neural cellina human 0032 FIG. 14D. P5 cerebellar neurons were plated on in need thereof. The method comprises administering to the substrate inhibitor MAG expressing CHO cells and treated human an effective amount of Lansoprazole. with daidzein (20 uM). Picture is representative of BIII tubu lin positive cells. BRIEF DESCRIPTION OF THE FIGURES 0033 FIG. 14E. P5 cerebellar neurons were plated on 0016 FIG.1. Chemical structures of compounds. substrate inhibitor MAG expressing CHO cells and treated 0017 FIG. 2. Table listing compounds that upregulated with Lanzoprazole (20 uM). Picture is representative of BIII arginase I at least above or near 2-fold. tubulin positive cells. 0018 FIG. 3. Table listing compounds that were tested in 0034 FIG. 14F. P5 cerebellar neurons were plated on quantitative RT-PCR and immunoblot analysis. CONT and substrate inhibitor (MAG) expressing CHO cells. 0019 FIG. 4. Western blot results that measured level of Neurons were plated either with DMSO (0.1%) on control enhanced arginase 1 protein associated with the compounds CHO monolayers, or on MAG and treated with DMSO listed in FIG. 3. (0.1%), methoxy vone (5uM), daidzein (20 uM) and Lanzo 0020 FIG. 5. Western blot results that measured level of prazole (20 uM). Graph depicts the average length of the enhanced arginase 1 protein associated with the compounds longest neurite (percentage of the longest neurite of the con listed in FIG. 3. trol). At least 400 neurons were measured in each assay and 0021 FIG. 6. Quantitative RT-PCR results presented in a the experiment was carried out at least twice. graph showing levels of arginase I messenger RNA (mRNA) 0035 FIG. 15. Western blot and immunostaining results upregulation associated with compounds I-10 listed in FIG.3. for arginase I protein. Neurons were treated with, diazein (20 0022 FIG. 7. Quantitative RT-PCR results presented in a uM), methoxyvone (5 uM) or lanzoprazole (20 uM) for 18 graph showing levels of arginase I messenger RNA (mRNA) hours and were then lysed and Subjected to gel electrophore upregulation associated with compounds II-20 listed in FIG. sis, followed by western blotting and immunostaining for 3. Arginase I protein. As a positive control, neurons were treated 0023 FIG. 8. Quantitative RT-PCR results presented in a with 1 mM db cAMP. graph showing levels of arginase I messenger RNA (mRNA) 0036 FIG. 16. The chemical formula, structures, and ref upregulation associated with compounds 21-30 listed in FIG. erences for a genus of compounds useful in the methods of the 3. invention. 0024 FIG. 9. Quantitative RT-PCR results presented in a graph showing levels of arginase I messenger RNA (mRNA) DETAILED DESCRIPTION OF THE INVENTION upregulation associated with compounds 31-40 listed in FIG. 3. Method for Enhancing Arginase Activity 0025 FIG. 10. Table summarizing results of compounds tested for their ability to overcome myelin-associated glyco 0037. In one aspect, the invention provides a method for protein (MAG) inhibition in P7 rat cerebellar neurons, as enhancing arginase activity in a damaged or injured cell in a compared with rho kinase inhibitor. The compounds were human in need thereof. Any isoform of arginase can be administered after (post-treatment) the neurons were plated. enhanced by the methods of the present invention. Examples 0026 FIG. 11. Graph showing results from testing the of arginase isoforms include arginase I and arginase II. compounds (10 nM) listed in FIG. 10 for their ability to 0038. The method comprises administering to the human overcome myelin-associated glycoprotein (MAG) inhibition an effective amount of a member of the genus of compounds in P7 rat cerebellar neurons, as compared with rho kinase selected from: 2-hydroxyXanthone; 2-methoxyXanthone; inhibitor. 3-methylcholanthrene; 4,7-dimethoxyflavone; 4'-methoxy 0027 FIG. 12. Graph showing results from testing the chalcone; 4'-methoxyflavone; 5,4'-dimethoxyflavone; 5.7,4'- compounds (40 nM) listed in FIG. 10 for their ability to trimethoxyflavone: 5,7-dimethoxyisoflavone; 6,3'- overcome myelin-associated glycoprotein (MAG) inhibition dimethoxyflavone; Acacetin diacetate; Anisindione; in P7 rat cerebellar neurons, as compared with rho kinase Apigenin; Apigenin triacetate; Biochanin a; Biochanin a inhibitor. diacetate; Chlorpropham; Chrysophanol; Daidzein: Dehy 0028 FIG. 13. Table summarizing results of compounds drovariabilin; Derrubone; Derrusnin, Derrustone; Diben tested for their ability to overcome myelin-associated glyco Zoylmethane: Fenbendazole; Formononetn; Genistein; protein (MAG) inhibition in P7 rat cerebellar neurons, as Ginkgetin, Indoprofen; Ipraflavone; Liquiritigenin dimethyl compared with rho kinase inhibitor. The compounds were ether; Methoxyvone; Methyl robustone; Phenazopyridine administered before (pretreatment) and after (post treatment) hydrochloride; Phenindione; Pinosylvin; Pinosylvin methyl the neurons were plated. ether; Piperine; Pramoxine hydrochloride; Resveratrol 0029 FIG. 14A. P5 cerebellar neurons were plated on 4'-methyl ether; Retusin 7-methyl ether; Robustone; Spirono CONT expressing CHO cells with DMSO (0.1%) on control lactone; Tilorone; Tranilast; or Xanthone. CHO monolayers. Picture is representative of BIII tubulin 0039. The terms "enhancing arginase activity” or positive cells. "enhanced arginase activity” refer to an increased level of 0030 FIG. 14B. P5 cerebellar neurons were plated on measurable arginase activity in a given assay in the presence substrate inhibitor MAG expressing CHO cells and treated of a candidate compound relative to the measurable level of with DMSO (0.1%). Picture is representative of BIII tubulin arginase activity in the absence of the candidate compound, positive cells. when tested under the same conditions. US 2013/0245070 A1 Sep. 19, 2013
0040 Activity is considered enhanced according to the 0049. In another embodiment, the method comprises invention if it is enhanced at least about 10% greater, prefer administering to a human in need thereofan effective amount ably at least about 25% greater, more preferably at least about of a compound comprising a 9H-Xanthen-9-one selected from 50% greater, even more preferably at least about 75% greater, a group consisting of 2-hydroxyXanthone, 2-methoxyxan most preferably at least about 90% greater, or more than in the thone, and Xanthone. absence of the candidate compound. 0050. In yet another embodiment, the method comprises 0041 Arginase activity as used herein can be enhanced by administering to a human in need thereofan effective amount any mechanism. For example, arginase activity could be of a compound comprising a 4H-chromen-4-one selected enhanced by transcriptional induction of its cognate messen from a group consisting of 4,7-dimethoxyflavone; 4'-meth ger RNA (mRNA), increased stability of its mRNA, increased oxyflavone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyfla translation of mRNA into protein, increased stability of argi vone; 5,7-dimethoxyisoflavone; 6.3'-dimethoxyflavone: nase protein, increased arginase activity (in the presence or Acacetin diacetate; Apigenin; Apigenin triacetate; Biochanin absence of increased protein), or any other mechanism. a; Biochanin a diacetate; Daidzein, Derrubone; Derrustone; Increases in arginase activity could be realized by the ability Formononetn; Genistein, Ginkgetin; Ipraflavone; Liquiriti of a compound to increase the affinity of the arginase enzyme genin dimethyl ether; Methoxyvone; and Retusin 7-methyl for its prototypical Substrate, arginine. ether. 0042. The arginase activity is enhanced in any damaged or 0051. In a further embodiment, the method comprises injured cell that benefits from enhanced arginase or from a administering to a human in need thereofan effective amount reduction of nitric oxide or arginine. The damage or injury of a compound comprising a (4-methoxyphenyl)-4H may be to any part of a cell. Such as to membranes, DNA, chromene-4-one selected from a group consisting of 4.7- RNA, and ribosomes. dimethoxyflavone; 4'-methoxyflavone; 5,4'-dimethoxyfla 0043. Examples of cells that may be damaged or injured vone; 5.7,4'-trimethoxyflavone; Acacetin diacetate; include cells of the central nervous system (CNS) or periph Biochanin a: Biochanin a diacetate; Formononetn; and eral nervous system (PNS), including neurons, ganglia, Retusin 7-methyl ether. Schwann cells, astrocytes, oligodendrocytes, microglia cells, 0052. In still another embodiment, the method comprises endothelial cells, immune cells (e.g., macrophages, T cells, B administering to a human in need thereof a compound com cells, and neutrophils), etc. In one embodiment, the damaged prising a 1,3-benzodioxol Selected from a group consisting or injured cell is in a human. of Derrubone; Derrusnin: Derrustone; Methyl robustone: 0044) In one embodiment, the method comprises admin Piperine; and Robustone. istering to a human in need thereofan effective amount of any 0053. In yet a further embodiment, the method comprises one or any combination of the following compounds: Pino administering to a human in need thereofan effective amount Sylvin; Derrustone; Methoxy vone; DehydroVariabilin; or of a compound selected from any one or a combination of Chrysophanol. compounds listed in FIG. 2 and/or FIG. 3. 0045. In another embodiment, the method comprises 0054. In another aspect of the invention, the method com administering to a human in need thereofan effective amount prises administering to the human in need thereofan effective of any one or any combination of the following compounds: amount of Lansoprazole. Resveratrol 4'-methyl ether; Derrubone; Ginkgetin; or Method for Treating a Disorder that can be Treated by Methyl robustone. Enhancing Arginase Activity 0046. In yet another embodiment, the method comprises 0055. In another aspect, the invention provides a method administering to a human in need thereofan effective amount for treating a disorder that can be treated by enhancing argi of any one or any combination of the following compounds: nase activity in a human in need thereof. The method includes Tilorone; Phenindione: Pramoxine hydrochloride; Indopro administering to the human an effective amount of a com fen; Phenazopyridine hydrochloride; Piperine; 6.3'- pound that enhances arginase activity, wherein the compound dimethoxyflavone; Anisindione; 5,4'-dimethoxyflavone: is any one of the following: 2-hydroxyXanthone, 2-methox Pinosylvin; Derrustone; 4,7-dimethoxyflavone; Daidzein; yxanthone: 3-methylcholanthrene; 4,7-dimethoxyflavone: 4'-methoxychalcone; Tranilast; Biochanin a diacetate; Res 4'-methoxychalcone; 4'-methoxyflavone; 5,4'-dimethoxyfla Veratrol 4'-methyl ether; Derrubone; Chlorpropham; vone; 5,7,4'-trimethoxyflavone; 5,7-dimethoxyisoflavone: Genistein: Dehydrovariabilin; Retusin 7-methyl ether; Xan 6.3'-dimethoxyflavone; Acacetin diacetate; Anisindione; thone; Pinosylvin methyl ether: Chrysophanol: Apigenin: Apigenin; Apigenin triacetate; Biochanin a; Biochanin a 2-methoxyxanthone: Apigenin triacetate; Fenbendazole; diacetate; Chlorpropham; Chrysophanol; Daidzein: Dehy Dibenzoylmethane; Methoxyvone; Ginkgetin, k salt; Methyl drovariabilin; Derrubone; Derrusnin, Derrustone; Diben robustone; Liquiritigenin dimethyl ether; Derrusnin; Biocha Zoylmethane: Fenbendazole; Formononetn; Genistein; nina; 5,7-dimethoxyisoflavone; Formononetin; 4'-methoxy Ginkgetin, Indoprofen; Ipraflavone; Liquiritigenin dimethyl flavone; or Acacetin diacetate. ether; Methoxyvone; Methyl robustone; Phenazopyridine 0047. In a further embodiment, the method comprises hydrochloride; Phenindione; Pinosylvin; Pinosylvin methyl administering to a human in need thereofan effective amount ether; Piperine; Pramoxine hydrochloride; Resveratrol of any one or any combination of the following compounds: 4'-methyl ether; Retusin 7-methyl ether; Robustone; Spirono Pinosylvin; Derrustone; Daidzein; 4'-methoxychalcone; Tra lactone; Tilorone; Tranilast; or Xanthone. nilast; Biochanin a diacetate; Resveratrol 4'-methyl ether; 0056. In one embodiment, the method comprises admin Dehydrovariabilin: Chrysophanol; or Methoxyvone. istering to the human an effective amount of any one or any 0048. In yet a further embodiment, the method comprises combination of the following compounds: Pinosylvin; Der administering to a human in need thereofan effective amount rustone; Methoxyvone; Dehydrovariabilin; or Chrysophanol. of any one or any combination of the following compounds: 0057. In another embodiment, the method comprises Daidzein or Methoxy vone. administering to the human an effective amount of any one or US 2013/0245070 A1 Sep. 19, 2013 any combination of the following compounds: Resveratrol 0067. Disorders and diseases in which enhancing arginase 4'-methyl ether; Derrubone; Ginkgetin; or Methyl robustone. activity is desired for treatment include ischemia, hypoxia, 0058. In yet another embodiment, the method comprises neurodegenerative disease or condition, or stroke. Additional administering to the human an effective amount of any one or disorders and diseases in which enhancing arginase activity is any combination of the following compounds: Tilorone; desired for treatment traumatic disorders (including but not Phenindione; Pramoxine hydrochloride; Indoprofen; limited to spinal cord injuries, spinal cord lesions, or other Phenazopyridine hydrochloride; Piperine; 6.3'-dimethoxy CNS pathway lesions), Surgical nerve lesions, damage sec flavone; Anisindione; 5,4'-dimethoxyflavone; Pinosylvin; ondary to infarction, infection, exposure to toxic agents, Derrustone; 4,7-dimethoxyflavone; Daidzein; 4'-methoxy malignancy, paraneoplastic syndromes, or patients with vari chalcone; Tranilast; Biochanin a diacetate; Resveratrol ous types of neurodegenerative disorders of the central ner 4'-methyl ether; Derrubone; Chlorpropham; Genistein; Vous system. Dehydrovariabilin: Retusin 7-methyl ether, Xanthone; Pino Sylvin methyl ether; Chrysophanol: Apigenin, 2-methoxyx Method for Treating Ischemia anthone: Apigenin triacetate; Fenbendazole; Dibenzoyl 0068. In one embodiment, the invention provides a methane; Methoxyvone; Ginkgetin, k salt; Methyl robustone; method for treating ischemia in a human in need thereof. The Liquiritigenin dimethyl ether; Derrusnin; Biochanina; 5,7- method comprises administering to the human a compound dimethoxyisoflavone; Formononetin; 4'-methoxyflavone; or that enhances arginase activity as described above, including Acacetin diacetate. those compounds grouped in the various genera and Sub 0059. In a further embodiment, the method comprises genera. administering to the human an effective amount of any one or 0069. Any mammal suffering from ischemia can be any combination of the following compounds: Pinosylvin; treated in accordance with the method of the present inven Derrustone; Daidzein; 4'-methoxychalcone; Tranilast; Bio tion. Ischemia generally refers to a condition of decreased chanin a diacetate; Resveratrol 4'-methyl ether, Dehydrova blood flow to an organ, tissue and/or cell. The decrease in riabilin; Chrysophanol; or Methoxyvone. blood flow can be caused by, for example, constriction (e.g., 0060. In yet a further embodiment, the method comprises hypoxemic vasoconstriction) or obstruction (e.g., clot, ath administering to the human an effective amount of any one or erosclerotic plaque) of a blood vessel. any combination of the following compounds: Daidzein or 0070 Ischemia can occur in any cell, organ, and/or tissue. Methoxyvone. Examples of cells, organs, and/or tissues which can be Sub 0061. In another embodiment, the method comprises jected to ischemia include neuronal cells (e.g., neurons, gan administering to the human an effective amount of a com glia, Schwann cells, astrocytes, oligodendrocytes and micro pound comprising a 9H-Xanthen-9-one selected from a group glia), brain, spinal cord, intestinal cells, kidney cells, heart consisting of 2-hydroxyXanthone, 2-methoxyxanthone, and and cardiac muscle cells such as myocytes, etc. Xanthone. 0062. In yet another embodiment, the method comprises Method for Treating Hypoxia administering to the human an effective amount of a com 0071. In yet another embodiment, the invention provides a pound comprising a 4H-chromen-4-one selected from a method for treating hypoxia in a human in need thereof. The group consisting of 4.7-dimethoxyflavone; 4'-methoxyfla method includes administering to the human a compound that vone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyflavone; 5,7- enhances arginase activity as described above, including dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin diac those compounds grouped in the genera and various Sub etate; Apigenin; Apigenin triacetate; Biochanina, Biochanin genera. a diacetate; Daidzein, Derrubone; Derrustone; Formononetn; 0072 Any mammal suffering from hypoxia can be treated Genistein; Ginkgetin; Ipraflavone; Liquiritigenin dimethyl in accordance with the method of the present invention. ether; Methoxy vone; and Retusin 7-methyl ether. Hypoxia generally refers to a lack of oxygen to cells, organs, 0063. In a further embodiment, the method comprises and/or tissues. Hypoxia can be caused by, for example, administering to the human an effective amount of a com ischemia, anemia and chemical modification of blood, Such pound comprising a (4-methoxyphenyl)-4H-chromene-4- as carboxyhemoglobin, etc. one selected from a group consisting of 4.7-dimethoxyfla 0073 Hypoxia can occur in any cell, organ, and/or tissue. vone; 4'-methoxyflavone; 5,4'-dimethoxyflavone; 5.7,4'- Examples of cells, organs, and/or tissues which can be Sub trimethoxyflavone; Acacetin diacetate; Biochanin a; jected to hypoxia include neuronal cells (e.g., neurons, gan Biochanin a diacetate; Formononetn; and Retusin 7-methyl glia, Schwann cells, astrocytes, oligodendrocytes and micro ether. glia), brain, spinal cord, kidney cells, intestinal cells, heart 0064. In still another embodiment, the method comprises and cardiac muscle cells such as myocytes, skin cells, etc. administering to the human an effective amount of a com pound comprising a 1,3-benzodioxol Selected from a group Method for Treating Neurodegenerative Disease or consisting of Derrubone; Derrusnin; Derrustone; Methyl Condition robustone; Piperine; and Robustone. 0074. In still another embodiment, the invention provides 0065. In yet a further embodiment, the method comprises a method for treating a neurodegenerative disease or condi administering to the human an effective amount of a com tion in a human in need thereof. The method includes admin pound selected from any one or a combination of compounds istering to the human a compound that enhances arginase listed in FIG. 2 and/or FIG. 3. activity as described above, including those compounds 0066. In another aspect of the invention, the method com grouped in the genera and various Sub-genera. prises administering to the human an effective amount of 0075. A neurodegenerative disease or condition typically Lansoprazole. refers to a disorder generally characterized by gradual and US 2013/0245070 A1 Sep. 19, 2013 progressive loss of cells, tissue and/or organ of the central or I0083. Any type of trauma to the nervous system may be peripheral nervous system. Examples of Such cells, tissues treated by the methods of the claimed invention. As described and organs include, the brain, spinal cord, neurons, ganglia, above, trauma of the CNS or PNS include, but are not limited Schwann cells, astrocytes, oligodendrocytes and microglia. to, spinal cord injuries, spinal cord lesions, other CNS path 0076 Any mammal suffering from any neurodegenerative way lesions, as well as injuries to the PNS, such as injuries to disease or condition can be treated in accordance with the a nerve or neuron of the PNS and axon damage resulting in method of the present invention. For example, the neurode demyelination of the PNS. Such trauma can arise from either generative disease or condition can be an acute condition. physical injury or disease. Any mammal Suffering from a Acute conditions generally occur as a result of trauma to a trauma of the CNS or PNS can be treated in accordance with cell, tissue and/or organ of the nervous system. The trauma the methods of the present invention. can, for example, partially or completely block blood flow to I0084. For example, spinal cord injury refers to any dam the cell, tissue and/or organ. Examples of acute neurodegen age to the spinal cord. The damage typically results in loss of erative conditions include head injury and brain injury. function, Such as mobility or feeling. Damage to the spinal 0077 Alternatively, the neurodegenerative disease or con cord can occur, for example, as a result or trauma (car acci dition can be a chronic neurodegenerative condition. dent, gunshot, falls, etc.) or disease (polio, spina bifida, Frie Examples of chronic neurodegenerative diseases and condi dreich's Ataxia, etc). tions include Parkinson's disease, Alzheimer's disease, Hun I0085. Any injury to the spinal cord can be treated in accor tington's disease and Amyotrophic Lateral Sclerosis (also dance with the method of the present invention. For example, known as Lou Gehrig's disease). the injury can be a complete injury to the spinal cord. Com 0078. Additional examples of neurodegenerative disor plete injury typically refers to the lack of function (e.g., no ders and diseases that can be treated by the invention include sensation and no voluntary movement) below the site of but are not limited to Alexander disease, Alper's disease, injury. Both sides of the body are usually affected. Alzheimer's disease, Amyotrophic lateral Sclerosis, Ataxia I0086 Alternatively, the injury may be an incomplete telangiectasia, Batten disease (also known as Spielmeyer injury to the spinal cord. An incomplete injury generally Vogt-Sjogren-Batten disease), Bovine spongiform encephal refers to some function below the site of injury. For instance, opathy (BSE), Canavan disease, Cockayne syndrome, Corti a person with an incomplete injury may be able to move one cobasal degeneration, Creutzfeldt-Jakob disease, Huntington limb more than another, may be able to feel parts of the body disease, HIV-associated dementia, Kennedy's disease, that cannot be moved, or may have more functioning on one Krabbe disease, Lewy body dementia, Machado-Joseph dis side of the body than the other, etc. ease (Spinocerebellar ataxia type 3), Multiple sclerosis, Mul tiple System Atrophy, Neuroborreliosis, Parkinson disease, Method for Promoting Regeneration of a Neural Cell in a Pelizaeus-Merzbacher Disease, Pick's disease, Primary lat Human in Need Thereof eral sclerosis, Prion diseases, Refsum’s disease, Sandhoff I0087. In another aspect, the invention provides a method disease, Schilder's disease, Schizophrenia, Spielmeyer-Vogt for promoting regeneration of a neural cell in a human in need Sjogren-Batten disease (also known as Batten disease), thereof. As described in Lange, et al., J. Nutr 2004 October; Spinocerebellar ataxia (multiple types with varying charac 134(10 Suppl):28125-28175; discussion 2818S-2819S, argi teristics), Spinal muscular atrophy, Steele-Richardson nase has a role in the axonal regeneration pathway. Arginase Olszewski disease, Tabes dorsalis, and other dementias. also has neuroprotective properties. Arginase is Sufficient in protecting neurons against several apoptosis-inducing Method for Treating Stroke stimuli. Moreover, arginase acts as a nitric oxide-independent 0079. In a further aspect, the invention provides a method inhibitor of neuronal apoptosis. for treating stroke in a human in need thereof. The method I0088. Several investigators have found that molecules or includes administering to the human a compound that drugs that prevent injury in the PNS or CNS have no effect on enhances arginase activity as described above, including or worse, negatively impact the ability of the nervous system those compounds grouped in the genera and various Sub to regenerate or repair. There is thus an urgent need to identify genera. targets whose activation would provide an environment that is 0080. Any mammal suffering from stroke can be treated in simultaneously instructive for neuronal protection and repair. accordance with the method of the present invention. Stroke It is proposed that Such an intervention would provide greater is a type of cardiovascular disease that generally involves the latitude in the timing of initiation of treatment. interruption of blood flow to and/or within the brain. The I0089 Arginine can be metabolized by nitric oxide syn interruption of blood flow can be due to, for example, a thase to produce nitric oxide. It can also be degraded by blockage or rupture of an artery or vessel. The blockage arginase to produce urea and ornithine, which in turn is a typically occurs from a blood clot. As a result of the interrup precursor for the synthesis of polyamines. These two path tion of blood flow, the brain does not receive sufficient ways compete for arginine. Arginase thus produces amounts of blood. polyamines at the expense of nitric oxide. I0081 Method for Treating Trauma of the CNS or PNS 0090 Polyamines have been implicated in neuronal 0082 Instill a further embodiment, the invention provides growth and development, axonal regeneration after injury, a method for treating trauma of the central nervous system and in would healing outside of the CNS. Arginase I is (CNS) or peripheral nervous system (PNS) in a human in upregulated and polyamine synthesis increases in neurons in need thereof. The method includes administering to the response to either dbCAMP or BDNF. human a compound that enhances arginase activity as 0091 By producing polyamines, arginase can overcome described above, including those compounds grouped in the the effects of myelin-associated glycoprotein (MAG) and genera and various Sub-genera. myelin on neurite outgrowth. Arginase is an enzyme that US 2013/0245070 A1 Sep. 19, 2013 mediates repair by reducing its Substrates (L-arginine) and Liquiritigenin dimethyl ether; Derrusnin; Biochanina; 5,7- increasing its ultimate products (polyamines), respectively. dimethoxyisoflavone; Formononetin; 4'-methoxyflavone; or 0092. As toxic levels of nitric oxide (NO) are implicated in Acacetin diacetate. acute ischemic cortical injury and motor neuron loss due to 0099. In a further embodiment, the method comprises absence of trophic factors, arginase can mediate neuroprotec administering to the human an effective amount of any one or tion. By producing polyamines, arginase can overcome the any combination of the following compounds: Pinosylvin; effects of myelin-associated glycoprotein (MAG) and myelin Derrustone; Daidzein; 4'-methoxychalcone; Tranilast; Bio on neurite outgrowth. Arginase is thus a bi-functional enzyme chanin a diacetate; Resveratrol 4'-methyl ether, Dehydrova that mediates neuroprotection or repair by reducing its Sub riabilin; Chrysophanol; or Methoxyvone. strates (L-arginine) and increasing its ultimate products 0100. In yet a further embodiment, the method comprises (polyamines), respectively. administering to the human a an effective amount of n effec 0093. As used herein, the phrase “neural cell includes tive amount of any one or any combination of the following nerve cells (i.e., neurons, e.g., uni-, bi-, or mulipolar neurons) compounds: Daidzein or Methoxyvone. and their precursors and glial cells (e.g., macroglia Such as 0101. In another embodiment, the method comprises astrocytes, oligodendrocytes, ependymal cells, radial glia, administering to the human an effective amount of a com Schwann cells, Satellite cells, and microglia) and their pre pound comprising a 9H-Xanthen-9-one selected from a group cursors. Microglia are specialized macrophages capable of consisting of 2-hydroxyXanthone, 2-methoxyxanthone, and phagocytosis that protect neurons of the central nervous sys Xanthone. tem. The term “precursor refers to cells which are capable of 0102. In yet another embodiment, the method comprises developing into a specific cell type. For example, a neural cell administering to the human an effective amount of a com precursor is a cell which is capable of developing into a pound comprising a 4H-chromen-4-one selected from a mature neural cell (i.e., a cell having the characteristic mor group consisting of 4.7-dimethoxyflavone; 4'-methoxyfla phology and function of a neural cell). vone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyflavone; 5,7- 0094. Accordingly, the claimed invention provides meth dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin diac ods for promoting regeneration of a neural cell in a human in etate; Apigenin; Apigenin triacetate; Biochanina, Biochanin need thereof. a diacetate; Daidzein, Derrubone; Derrustone; Formononetn; 0095. The method includes administering to the human an Genistein; Ginkgetin; Ipraflavone; Liquiritigenin dimethyl effective amount of a compound that enhances arginase activ ether; Methoxy vone; and Retusin 7-methyl ether. ity, wherein the compound is any one of the following: 2-hy 0103) In a further embodiment, the method comprises droxyXanthone; 2-methoxyxanthone: 3-methylcholanthrene; administering to the human an effective amount of a com 4,7-dimethoxyflavone; 4'-methoxychalcone; 4'-methoxyfla pound comprising a (4-methoxyphenyl)-4H-chromene-4- vone; 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyflavone; 5,7- one selected from a group consisting of 4.7-dimethoxyfla dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin diac vone; 4'-methoxyflavone; 5,4'-dimethoxyflavone; 5.7,4'- etate; Anisindione: Apigenin; Apigenin triacetate; Biochanin trimethoxyflavone; Acacetin diacetate; Biochanin a; a; Biochanin a diacetate; Chlorpropham; Chrysophanol: Biochanin a diacetate; Formononetn; and Retusin 7-methyl Daidzein: Dehydrovariabilin: Derrubone; Derrusnin; Derrus ether. tone; Dibenzoylmethane: Fenbendazole; Formononetn; 0104. In still another embodiment, the method comprises Genistein; Ginkgetin; Indoprofen; Ipraflavone; Liquiritige administering to the human an effective amount of a com nin dimethyl ether; Methoxyvone; Methyl robustone; pound comprising a 1,3-benzodioxol selected from a group Phenazopyridine hydrochloride; Phenindione; Pinosylvin; consisting of Derrubone; Derrusnin; Derrustone; Methyl Pinosylvin methyl ether; Piperine: Pramoxine hydrochloride: robustone; Piperine; and Robustone. Resveratrol 4'-methyl ether; Retusin 7-methyl ether; Robus 0105. In yet a further embodiment, the method comprises tone; Spironolactone; Tilorone; Tranilast; or Xanthone. administering to the human an effective amount of a com 0096. In one embodiment, the method comprises admin pound selected from any one or a combination of compounds istering to the human an effective amount of any one or any listed in FIG. 2 and/or FIG. 3. combination of the following compounds: Pinosylvin; Der 0106. In another aspect of the invention, the method com rustone; Methoxyvone; Dehydrovariabilin; or Chrysophanol. prises administering to the human an effective amount of 0097. In another embodiment, the method comprises Lansoprazole. administering to the human an effective amount of any one or any combination of the following compounds: Resveratrol Methods for Protecting a Neural Cell in a Human in Need 4'-methyl ether; Derrubone; Ginkgetin; or Methyl robustone. Thereof 0098. In yet another embodiment, the method comprises 0107. In yet another aspect, the invention provides a administering to the human an effective amount of any one or method for protecting a neural cellina human in need thereof. any combination of the following compounds: Tilorone; The method includes administering to the human a compound Phenindione; Pramoxine hydrochloride; Indoprofen; that enhances arginase activity, wherein the compound com Phenazopyridine hydrochloride; Piperine; 6.3'-dimethoxy prises any one of the following compounds: 2-hydroxyXan flavone; Anisindione; 5,4'-dimethoxyflavone; Pinosylvin; thone: 2-methoxyxanthone; 3-methylcholanthrene; 4.7- Derrustone; 4,7-dimethoxyflavone; Daidzein; 4'-methoxy dimethoxyflavone; 4'-methoxychalcone; 4'-methoxyflavone: chalcone; Tranilast; Biochanin a diacetate; Resveratrol 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyflavone; 5,7- 4'-methyl ether; Derrubone; Chlorpropham; Genistein; dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin diac Dehydrovariabilin: Retusin 7-methyl ether, Xanthone; Pino etate; Anisindione: Apigenin; Apigenin triacetate; Biochanin Sylvin methyl ether; Chrysophanol: Apigenin, 2-methoxyx a; Biochanin a diacetate; Chlorpropham; Chrysophanol: anthone: Apigenin triacetate; Fenbendazole; Dibenzoyl Daidzein: Dehydrovariabilin: Derrubone; Derrusnin; Derrus methane; Methoxyvone; Ginkgetin, k salt; Methyl robustone; tone; Dibenzoylmethane: Fenbendazole; Formononetn; US 2013/0245070 A1 Sep. 19, 2013
Genistein; Ginkgetin; Indoprofen; Ipraflavone; Liquiritige 0115 Throughout this specification, parameters are nin dimethyl ether; Methoxyvone; Methyl robustone; defined by maximum and minimum amounts. Each minimum Phenazopyridine hydrochloride; Phenindione; Pinosylvin; amount can be combined with each maximum amount to Pinosylvin methyl ether; Piperine: Pramoxine hydrochloride: define a range. Resveratrol 4'-methyl ether; Retusin 7-methyl ether; Robus tone; Spironolactone; Tilorone; Tranilast; or Xanthone. Administration 0108. In another aspect of the invention, the method com 0116. The compounds are administered to a human. The prises administering to the human an effective amount of compound is administered to the human in an amount effec Lansoprazole. The method includes administering to the tive in achieving its purpose. The effective amount of the human any one or a combination of compounds that enhances compound to be administered can be readily determined by arginase activity included in the genera and Sub-genera of those skilled in the art during pre-clinical trials and clinical compounds described above. trials by methods familiar to physicians and clinicians. Typi cal daily doses include approximately 1 mg to 1000 mg. Compounds 0117. Any method known to those in the art for contacting a cell, organ or tissue with a compound may be employed. 0109 Compounds useful in the methods of the present Suitable methods include invitro, ex vivo, or in vivo methods. invention include 2-hydroxyXanthone, 2-methoxyxanthone; In vitro methods typically include cultured samples. For 3-methylcholanthrene; 4,7-dimethoxyflavone; 4'-methoxy example, a cell can be placed in a reservoir (e.g., tissue culture chalcone; 4'-methoxyflavone; 5,4'-dimethoxyflavone; 5.7,4'- plate), and incubated with a compound under appropriate trimethoxyflavone: 5,7-dimethoxyisoflavone; 6,3'- conditions suitable for enhancing arginase activity. Suitable dimethoxyflavone; Acacetin diacetate; Anisindione; incubation conditions can be readily determined by those Apigenin; Apigenin triacetate; Biochanin a; Biochanin a skilled in the art. diacetate; Chlorpropham; Chrysophanol; Daidzein: Dehy 0118. Ex vivo methods typically include cells, organs or drovariabilin; Derrubone; Derrusnin, Derrustone; Diben tissues removed from a mammal, such as a human. The cells, Zoylmethane: Fenbendazole; Formononetn; Genistein; organs or tissues can, for example, be incubated with the Ginkgetin, Indoprofen; Ipraflavone; Liquiritigenin dimethyl compound under appropriate conditions. The contacted cells, ether; Methoxyvone; Methyl robustone; Phenazopyridine organs or tissues are normally returned to the donor, placed in hydrochloride; Phenindione; Pinosylvin; Pinosylvin methyl a recipient, or stored for future use. Thus, the compound is ether; Piperine: Pramoxine hydrochloride: Resveratrol generally in a pharmaceutically acceptable carrier. 4'-methyl ether; Retusin 7-methyl ether; Robustone; Spirono 0119. In vivo methods are typically limited to the admin lactone; Tilorone; Tranilast; or Xanthone. istration of a compound, Such as those described above, to a mammal, preferably a human. The compounds useful in the 0110. Another compound useful in the methods of the methods of the present invention are administered to a mam present invention includes Lansoprazole. mal in an amount effective in enhancing arginase activity or 0111. Further examples compounds useful in the methods treating the mammal. The effective amount is determined of the present invention include any one or a combination of during pre-clinical trials and clinical trials by methods famil compounds listed in FIG. 2 and/or FIG. 3. iar to physicians and clinicians. 0112 These compounds are known in the art. The chemi 0.120. An effective amount of a compound useful in the cal formula, structures, and references for a genus of com methods of the present invention, preferably in a pharmaceu pounds described above are shown in FIG. 16. tical composition, may be administered to a mammal in need thereof by any of a number of well-known methods for 0113. The chemical formula, structures, and references for administering pharmaceutical compounds. The compound Pinosylvin, Derrustone, Methoxyvone, Dehydrovariabilin, may be administered systemically or locally. and Chrysophanol are shown in FIG. 1. I0121 For example, the compound may be administered 0114. The compounds can be in the form of a pharmaceu orally, intravenously, intranasally, intramuscularly, Subcuta tically acceptable salt. The term “pharmaceutically accept neously, or transdermally. Other routes of administration able salt” refers to a well-tolerated, nontoxic salt prepared include intracerebroventricularly or intrathecally. Intracere from any basic or acidic compound mentioned above, and an broventiculatly refers to administration into the ventricular acid or base, respectively. The acids may be inorganic or system of the brain. Intrathecally refers to administration into organic acids of any one of the compounds mentioned above. the space under the arachnoid membrane of the spinal cord. Examples of inorganic acids include hydrochloric, hydrobro Thus intracerebroventricular or intrathecal administration mic, nitric hydroiodic, Sulfuric, and phosphoric acids. may be preferred for those diseases and conditions which Examples of organic acids include carboxylic and Sulfonic affect the organs or tissues of the central nervous system. acids. The radical of the organic acids may be aliphatic or 0.122 The compounds useful in the methods of the inven aromatic. Some examples of organic acids include formic, tion may also be administered to mammals by Sustained acetic, phenylacetic, propionic, succinic, glycolic, glucu release, as is known in the art. Sustained release administra ronic, maleic, furoic, glutamic, benzoic, anthranilic, Salicylic, tion is a method of drug delivery to achieve a certain level of phenylacetic, mandelic, embonic (pamoic), methanesulfonic, the drug over a particular period of time. The level typically is ethanesulfonic, panthenoic, benzenesulfonic, Stearic, Sulfa measured by serum or plasma concentration. nilic, alginic, tartaric, citric, gluconic, gulonic, arylsulfonic, I0123. A description of methods for delivering a compound and galacturonic acids. Appropriate organic bases may be by controlled release can be found in international PCT selected, for example, from N,N-dibenzylethylenediamine, Application No. WO 02/083106. The PCT application is chloroprocaine, choline, diethanolamine, ethylenediamine, incorporated herein by reference in its entirety. Other con meglumine (N-methylglucamine), and procaine. trolled release agents are described, for example, in U.S. Pat. US 2013/0245070 A1 Sep. 19, 2013
Nos. 5,567,439; 6,838,094; 6,863,902; and 6,905,708. The CollectionTM from MicroSource Discovery System, Inc. controlled release agents and methods for making them in (Groton, Conn.). The 2000 compounds in the library are these patents are incorporated herein by reference. primarily Food and Drug Administration (FDA)-approved 0.124. Any formulation known in the art of pharmacy is compounds or natural products. An alphabetical list of the suitable for administration of the compounds useful in the compounds is available at the MicroSource Discovery web methods of the present invention. For oral administration, site at www.msdiscovery.com/spect.html. The compounds liquid or Solid formulations may be used. Some examples of are supplied as 10 mM solutions in dimethyl sulfoxide formulations include tablets, gelatin capsules, pills, troches, elixirs, Suspensions, syrups, wafers, chewing gum and the (DMSO). like. The compounds can be mixed with a suitable pharma 0.130. The library was screened using murine hippocampal ceutical carrier (vehicle) or excipient as understood by prac HT22 cells transfected with a luciferase-arginase 1 construct titioners in the art. Examples of carriers and excipients on 96 well plates. Total protein was also measured to use in include starch, milk, Sugar, certain types of clay, gelatin, normalization. The luciferase assay result for each compound lactic acid, Stearic acid or salts thereof, including magnesium is normalized to protein content and is expressed as or calcium Stearate, talc, vegetable fats or oils, gums and luciferase/mg protein. The fold increase was measured rela glycols. tive to untreated control, represented as sample #1 in each 0.125 For systemic, intracerebroventricular, intrathecal, HT22 Arg plate. topical, intranasal, Subcutaneous, or transdermal administra I0131 Ratios of the results from the “luciferase assay” and tion, formulations of the compounds useful in the methods of “protein assay” were tabulated. A comparison of these nor the present inventions may utilize conventional diluents, car malized responses indicates amount of enhanced arginase riers, or excipients etc.. Such as those known in the art to activity. Compounds and their respective Chem ID numbers deliver the compounds. For example, the formulations may that upregulated arginase 1 above or near 2 fold were identi comprise one or more of the following: a stabilizer, a Surfac fied. See FIG. 2. The protocol that was used to screen for the tant, preferably a nonionic Surfactant, and optionally a salt Arginase I upregulators is described in detail below. and/or a buffering agent. The compound may be delivered in the form of an aqueous solution, or in a lyophilized form. Materials 0126 The stabilizer may, for example, be an amino acid, (0132) T75 flask, Corning 430641 vented Such as for instance, glycine; or an oligosaccharide, such as 96 well plate (for tissue culture), Mictotest Primaria, flat for example. Sucrose, tetralose, lactose or a dextran. Alterna bottom, #35-3872 tively, the stabilizer may be a Sugar alcohol. Such as for 96 well plate (for chemical compound), MictotestTM 96, flat instance, mannitol; or a combination thereof. Preferably the bottom, #35-3072 stabilizer or combination of stabilizers constitutes from about Dulbecco's Phosphate Buffered Saline (1xPBS), from Gibco 0.1% to about 10% weight for weight of the compound. The H 14190-144 Surfactant is preferably a nonionic Surfactant, such as a 0.25% Trypsin-EDTA solution, from Sigma #T4049 polysorbate. Some examples of Suitable surfactants include DMEM (Dulbecco's Modified Eagle's Medium), from Gibco Tween20, Tween80; a polyethylene glycol or a polyoxyeth H 11995-040 & #11965-092 ylene polyoxypropylene glycol, such as Pluronic F-68 at from Fetal Bovine Serum (FCS), from Gibco #10082-147 about 0.001% (w/v) to about 10% (w/v). Penicillin 10,0001 U/ml & Streptomycin 10,000 ug/ml (P/S) 0127. The salt or buffering agent may be any salt or buff from Cellgro #30-002-C1 ering agent, such as for example, sodium chloride, or Sodium/ Puromycin 1 mg/ml stock potassium phosphate, respectively. Preferably, the buffering 10 mM stock of chemical compounds in DMSO, The Spec agent maintains the pH of the pharmaceutical composition in trum CollectionTM from MicroSource Discovery System. Inc: the range of about 5.5 to about 7.5. The salt and/or buffering Stored at -80° C. agent is also useful to maintain the osmolality at a level suitable for administration to a human or an animal. Prefer Lysis Reagent (5x), from Promega i E153A ably the salt or buffering agent is presentata roughly isotonic Luciferase Assay Substrate, from Promega i E151A concentration of about 150 mM to about 300 mM. Luciferase Assay Buffer, from Promega i E152A 0128. The formulations of the compounds useful in the LimaxII from Molecular Device methods of the present invention may additionally contain one or more conventional additive. Some examples of Such 70% Ethanol additives include a solubilizer Such as, for example, glycerol; (0.133 96-well NUNC 236107 white plate (Luciferase an antioxidant Such as for example, benzalkonium chloride (a Assay) mixture of quaternary ammonium compounds, known as UV Plate for plate reader (Protein Assay) “quats'), benzyl alcohol, chloretone or chlorobutanol; anaes thetic agent Such as for example a morphine derivative; or an Reagents isotonic agent etc.. Such as described above. As a further precaution against oxidation or other spoilage, the pharma DMEM for HT22 4.8Kb Arg-Puro ceutical compositions may be stored under nitrogen gas in vials sealed with impermeable stoppers. 0134) EXAMPLES
Example 1 Ingredient Stock Final conc. Volume Screening for Arginase I Upregulators DMEM (#11965-092) 445 ml 0129. A 2000 compound library was tested to identify FCS 10% 50 ml arginase-1 upregulators. The library tested was The Spectrum US 2013/0245070 A1 Sep. 19, 2013
-continued Day 2. Chemical Treatment Ingredient Stock Final conc. Volume Materials: PS 3 ml O150 Puromycin 1 mg/ml 4 g/ml 2 ml
Total 500 ml Lysis Reagent (10 ml per plate) 3x 96 well plates (#35-3072) 3x Chemical plates Lysis Reagent Sx 1X 8 ml Repeat pipette & tips cAMP stock (2.5 mM) (positive control) ddH2O 32 ml 10 ultips (12x regular and 3x long “reach') Medium (for HT22 Luc) Total 40 ml Steps 0151. 1. Check if T75 flasks reach 50-70% confluence Day 1. Harvest Cell and Seeding under microscope. 0152 2. Thaw chemical plates at room temperature for Materials: 10 minutes 0153. 3. On a sterile 96 well plate, add 100 ul/well 0135) DMEM medium (for HT22 Luc) 0154 4. Prepare 500 uM secondary chemical stocks by adding 5 ul original 10 mM chemical stocks to 100 ul 12x 96 well plates (#35-3872) Two types of DMEM medium DMEM medium (1:20) 8x T75 Flasks Trypsin Solution 0.155 5. Add 2 ul/well chemical to duplicate plates (col 2x Glass sterile pipette Timer umn #2-11) and two cell lines (HT22 Luc and Sterile pipette (25 ml, 10 ml, 5 ml) HT22Arg); the final concentration is 10 uM (1:50) 6x 50 ml centrifuge tubes 0156 6. Treat and prepare the other two chemical plates Repeat pipette & tips the same way (O157 7. Use repeat pipette to add 2 ul/well 2.5 mM cAMP stock (positive control fic. 50 uMDFO) to col Steps umni 12 for all plates; columni 1 is negative control (Blank). 0.136 1. Remove old medium from T4s flask by suction 0158 8. Label and incubate at 37° C. incubator for 24 hours 0.137 2. Rinse once with 5 ml 1xPBS: discard 1XPBS 0159. 9. Wrap and store two secondary stock plates at 4 0.138. 3. Add 3 ml Trypsin to treat cells and incubate at C. refrigerator 37° C. incubator for 3 min 0.160 10. Leave original chemical plates at basement 0.139 4. Observe under microscope to see if all cells -80° C. freezer detached 0140) 5. Stop trypsin reaction by adding 10 ml DMEM Day 3. Lysis and Read and then transfer the solution into a 50ml centrifuge tube Materials: 0141 6. Centrifuge at 980 rpm/4 min (0161 0142 7. Prepare three T75 by adding 10 ml fresh medium to each flask 0.143 8. Prepare two 50 ml centrifuge tubes by adding 2x Glass sterile pipette 1x Lysis Reagent (10 ml for each plate) 30 ml fresh medium to each tube 12x 96-well NUNC white Luciferase Assay Substrate & Buffer plate 0144. 9. Discard supernatant by suction and add 8 ml 12x 96-well UV plate (1 set per plate) fresh DMEM into the tube to resuspend the cell pellet 10 ultips (24x regular) Repeat pipette & tips 0145 10. Mix and Titrate cell suspension and add 2 ml Timer cell suspension into three T75 flasks (total 12 ml) and 1 ml to each 50 ml centrifuge tube. 0146 11. Use repeat pipette and sterile tips to dispense Steps 100 ul/well to six 96-well plates (for duplicate three 0162. 1. After 24hrs incubation, remove all medium by chemical plate) from two 50 ml centrifuge tubes suction and add 100 ul/well 1x Lysis Reagent
0166 5. Prepare Substrate solution by mixing enhanced arginase activity. Phosphate buffered saline (PBS) Luciferase Assay Substrate (keep in freezer) with 10 ml was used as a negative control, and cAMP was used as a Luciferase Assay Buffer (thaw at room temperature) positive control. The sample numbers (e.g., c1, c2) listed on 0.167 6. Use multiple channel pipette to transfer 10 the bottom of each graph of FIGS. 6 through 9 correspond to ul/well cell lysate/supernatant to a 96-well NUNC white the sample number and compound listed in FIG. 3. plate for reading with Luciferase Assay (0189 The protocols used for the quantitative RT-PCR and 0168 7. Use multiple channel pipette to transfer 2.5 Immunoblot analyses are described in detail below. ul/well cell lysate/supernatant to a 96-well UV plate for (0190. Quantitative RT-PCR reading with Protein Assay 0191 Total RNA was prepared from primary mixed cor (0169 8. Stored the plate in 4° C. refrigerator or base tical neurons using TriZOL (Invitrogen) and cDNA generat ment freezer ing using a SuperScript III First-Strand Synthesis System for (0170 9. Set up and run LimaxII ATP Assay: RT-PCR kit (Invitrogen), according to the manufacturers (0171 Dentistry, twelve compounds were tested, using the materials 0207. In experiments performed in the Marie Filbin labo and methods described by Venkatesh, et al., J Neurosci. 2005 ratory at Hunter College on p5 rat cerebellar neurons, twelve Jan. 26; 25(4):808-22. The compounds and their respective compounds were tested, using the materials and methods post-treatment results are listed in FIG. 10. described by Mukhopadhyay, eta. Neuron. 1994 September; 0197) The twelve compounds tested were Acetami 13(3):757-67. The compounds and their respective post-treat nophen; Pinosylvin; Resveratol 4-methyl ether: Chrysopha ment results are listed in FIG. 13. nol (low dose); Daidzein; Anisomycin; Methoxyvone; Dehy 0208. The twelve compounds were Acetaminophen; Pino drovariabilin; Phenethyl caffeate (cape); Fenbendazole; Sylvin; Resveratol 4-methyl ether: Chrysophanol (low dose): DerruStone; Epicatechin pentaacetate; Lansoprazole. Daidzein; Anisomycin; Methoxyvone; DehydroVariabilin; 0198 The twelve compounds were tested in two different Phenethyl caffeate (cape); Fenbendazole; Derrustone: Epi dilutions (10 nM and 40 nM final concentration). FIG. 11 catechin pentaacetate; Lansoprazole. Two experiments were shows the assay results for the compounds at 10 nM and FIG. done. 12 shows the assay results for the compounds at 40 nM. The 0209. In a first experiment, the twelve compounds were compound sample numbers listed on the bottom of FIGS. 11 tested for their ability to overcome inhibition by MAG in and 12 correspond to the compound sample numbers listed in culture. The compounds were tested for their ability to inhibit FIG 10. neurite outgrowth of MAG-expressing CHO cells. Cerebellar (0199 MAG-overexpressing CHO cells and control CHO neurons were cultured on MAG-expressing CHO cells and feeder layers were used in the assay. All experiments were neurite length is compared to neurons growing on control done with Percoll purified P7 rat cerebellar neurons. The CHO cells, not expressing MAG. twelve compounds were tested in a “post-treatment manner. 0210. The cells were assessed for their ability to overcome Post-treatment refers to administering the compounds after MAG inhibition when added directly to the cultures at a range the neurons were plated. of concentrations from 2-20 uM. Neurons were plated at a 0200. After 24 h in culture, cells were fixed and stained density of about 10,000 neurons per well of an 8-well culture with Tulj 1 (cells with neurites longer than 1 cell body diam dish containing a monolayer of either MAG-expressing or eter were quantified). Untreated cells (first column of FIGS. control CHO cells. The co-cultures were, incubated for 16-18 11 and 12) and DMSO only (second column of FIGS. 11 and hand then immunostained for BIII tubulin. See FIG. 14 A-E. 12) were used as a negative controls. RHO-kinase inhibitor Neurite outgrowth was quantified as previously described Y27632 (15 microM) (third column of FIGS. 11 and 12) was (Mukhopadhyay et al., 1994). When added directly to the used as a positive control. All experiments have been repeated co-cultures, none of the compounds had any effect on inhibi 3-5 times independently. tion by MAG. MAG inhibited neurite outgrowth as potently 0201 In FIGS. 11 and 12, there are two bars for each as when the compound was absent. compound tested. One bar represents the results on the CHO 0211 Next, the compounds were tested for their ability to MAG cells (labeled “MAG”), and the other bar (labeled “r2) overcome inhibition by MAG when the neurons were is the result on control CHO cells. exposed to the compounds prior to being exposed to MAG-a 0202 The following compounds were found to have no procedure termed, “priming Neurons were primed with the effect on releasing MAG inhibition: #1 (Acetaminophen), #3 individual compounds at various concentrations ranging (Resveratol 4-methyl ether), #5 (Daidzein), #7 (Methox from 2-20 uM overnight and were then plated onto either the yvone), #9 CAPE, #11 (Derrustone), and #12 (epicatechin MAG-expressing or control CHO cell monolayers. Of the 12 pentaacetate). compounds tested, three were able to overcome inhibition by 0203 The experiment was repeated three times with the MAG completely: Daidzein (20 uM), Lanzoprazole (20 uM) most promising compounds, which were compound #6 (Ani and Methoxyvone (5uM). See FIGS. 13 and 14. somycin), #8 (DehydroVariabilin), #10 (Fenbendazole), #2 (Pinosylvin) and #4 (Chrysophanol at low doses). 0212. In FIG. 14, P5 cerebellar neurons were plated on CONT and MAG expressing CHO cells. See FIG. 14A-E, Example 4 which are pictures representative offIII tubulin positive cells. Neurons are plated either with DMSO (0.1%) on control MAG Inhibition Analysis CHO monolayers (FIG. 14A), or on substrate inhibitor (MAG) (FIG. 14B-E) and treated with DMSO (0.1%) (FIG. 0204 MAG has been identified as an inhibitor of axonal 14B), methoxy vone (5uM) (FIG.14C), daidzein (20M) and regeneration and neurite outgrowth. A consequence of Lanzoprazole (20 uM). Graph in FIG. 14 depicts the average elevated cAMP is the synthesis of polyamines, resulting from length of the longest neurite (percentage of the longest neurite an up-regulation of Arginase I, a key enzyme in their synthe of the control). At least 400 neurons were measured in each sis Inhibiting polyamine synthesis blocks the cAMP effect on assay and the experiment was carried out at least twice. regeneration. Either over-expression of Arginase I or exog 0213. The other compounds did not show as strong of an enous polyamines can overcome inhibition by MAG and by effect in overcoming MAG inhibition as the compounds myelin in general, as described by Cai, et al., Neuron. 2002 Daidzein (20 uM), Lanzoprazole (20 uM) and Methoxyvone Aug. 15:35(4):711-9. (5 uM). See FIG. 13. The overcoming of MAG inhibition 0205 Compounds were tested for their ability to over occurred when the cells were “preconditioned.” Precondi come myelin-associated glycoprotein (MAG) inhibition in tioning treatment means that neurons were first treated over primary rat neurons. night with the compound and then trypsinized, plated, and 0206. Inhibiting polyamine synthesis blocks the cAMP allowed to grow for neurite extension for 16-18h on the top of effect on regeneration. Either over-expression of Arginase I or the CHO monolayers. Neurons that were directly treated with exogenous polyamines can overcome inhibition by MAG and the different compounds on the top of the monolayers did not by myelin in general. show any overcoming of MAG inhibition. US 2013/0245070 A1 Sep. 19, 2013 0214. In a second experiment, the twelve compounds were ing of Derrubone; Derrusnin; Derrustone; Methyl robustone: tested for their ability to upregulate arginase I protein. Neu Piperine; and Robustone; or a pharmaceutically acceptable rons were treated with, diazein (20uM), methoxyvone (5uM) salt of any Such compound. or lanzoprazole (20 uM) for 18 hours and were then lysed and 7. A method for enhancing arginase activity in a damaged subjected to gel electrophoresis, followed by western blotting or injured cell in a human in need thereof, the method com and immunostaining for Arginase I protein. See FIG. 15. As a prising administering to the human an effective amount of positive control, neurons were treated with 1 mM db cAMP Lansoprazole; or a pharmaceutically acceptable salt of Such (FIG. 15). Db-cAMP was previously shown to increased compound. Arginase I protein 3-fold as described by Cai, et a..., Neuron. 8. A method for treating a disorder that can be treated by 2002 Aug. 15:35(4):711-9. enhancing arginase activity in a human in need thereof, the 0215. As was shown previously (Cai et al., 2002), Argin method comprising administering to the human an effective ase I is up-regulated in response to treatment with dbcAMP. amount of a compound that enhances arginase activity, As shown presently, Arginase I protein level is also up-regu wherein the compound is any one of the following: 2-hydrox lated after the treatment with diadzein (20 uM), methoxyvone yXanthone; 2-methoxyxanthone: 3-methylcholanthrene; 4.7- (5uM) or Lanzoprazole (20 uM). dimethoxyflavone; 4'-methoxychalcone; 4'-methoxyflavone: 5,4'-dimethoxyflavone; 5,7,4'-trimethoxyflavone; 5,7- What is claimed is: dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin diac 1. A method for enhancing arginase activity in a damaged etate; Anisindione: Apigenin; Apigenin triacetate; Biochanin or injured cell in a human in need thereof, the method com a; Biochanin a diacetate; Chlorpropham; Chrysophanol: prising administering to the human an effective amount of Daidzein: Dehydrovariabilin: Derrubone; Derrusnin; Derrus one of the following compounds: 2-hydroxyXanthone; tone; Dibenzoylmethane: Fenbendazole; Formononetn; 2-methoxyxanthone: 3-methylcholanthrene; 4,7-dimethoxy Genistein; Ginkgetin; Indoprofen; Ipraflavone; Liquiritige flavone; 4'-methoxychalcone; 4'-methoxyflavone; 5,4'- nin dimethyl ether; Methoxyvone; Methyl robustone; dimethoxyflavone: 5,7,4'-trimethoxyflavone; 5,7- Phenazopyridine hydrochloride; Phenindione; Pinosylvin; dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin Pinosylvin methyl ether, Piperine: Pramoxine hydrochloride: diacetate; Anisindione: Apigenin; Apigenin triacetate; Bio Resveratrol 4'-methyl ether; Retusin 7-methyl ether; Robus chanina, Biochanin a diacetate; Chlorpropham; Chrysopha tone; Spironolactone; Tilorone; Tranilast; or Xanthone; or a nol; Daidzein: Dehydrovariabilin: Derrubone; Derrusnin; pharmaceutically acceptable salt of any Such compound. Derrustone; Dibenzoylmethane; Fenbendazole; Formonon etn; Genistein; Ginkgetin: Indoprofen; Ipraflavone; Liquiriti 9. A method according to claim 8, wherein the disorder is genin dimethyl ether; Methoxy vone; Methyl robustone: ischemia. Phenazopyridine hydrochloride; Phenindione; Pinosylvin; 10. A method according to claim 8, wherein the disorder is Pinosylvin methyl ether; Piperine: Pramoxine hydrochloride: hypoxia. Resveratrol 4'-methyl ether; Retusin 7-methyl ether; Robus 11. A method according to claim 8, wherein the disorder is tone; Spironolactone; Tilorone: Tranilast; or Xanthone; or a a neurodegenerative disease or condition. pharmaceutically acceptable salt of any such compound. 12. A method according to claim 11, wherein the neurode generative disease or condition is any one of the following: 2. A method according to claim 1, wherein the compound Alexander disease, Alper's disease, Alzheimer's disease, is Pinosylvin; Derrustone; Methoxyvone; DehydroVariabilin; Amyotrophic lateral Sclerosis, Ataxia telangiectasia, Batten or Chrysophanol; or a pharmaceutically acceptable salt of any disease (also known as Spielmeyer-Vogt-Sjogren-Batten dis Such compound. ease), Bovine spongiform encephalopathy (BSE), Canavan 3. A method according to claim 1, wherein the compound disease, Cockayne syndrome, Corticobasal degeneration, comprises a 9H-Xanthen-9-one selected from a group consist Creutzfeldt-Jakob disease, Huntington disease, HIV-associ ing of 2-hydroxyXanthone, 2-methoxyxanthone, and Xan ated dementia, Kennedy's disease, Krabbe disease, Lewy thone; or a pharmaceutically acceptable salt of any such com body dementia, Machado-Joseph disease (Spinocerebellar pound. ataxia type 3), Multiple sclerosis, Multiple System Atrophy, 4. A method according to claim 1, wherein the compound Neuroborreliosis, Parkinson disease, Pelizaeus-Merzbacher comprises a 4H-chromen-4-one selected from a group con Disease, Pick's disease, Primary lateral sclerosis, Prion dis sisting of 4,7-dimethoxyflavone; 4'-methoxyflavone; 5,4'- eases, Refsum’s disease, Sandhoff disease, Schilder's dis dimethoxyflavone: 5,7,4'-trimethoxyflavone; 5,7- ease, Schizophrenia, Spielmeyer-Vogt-Sjogren-Batten dis dimethoxyisoflavone; 6.3'-dimethoxyflavone; Acacetin ease (also known as Batten disease), Spinocerebellar ataxia diacetate; Apigenin; Apigenin triacetate; Biochanina, Bio (multiple types with varying characteristics), Spinal muscu chanin a diacetate; Daidzein; Derrubone; Derrustone; For laratrophy, Steele-Richardson-Olszewski disease, Tabes dor mononetn; Genistein, Ginkgetin; Ipraflavone; Liquiritigenin salis, or other dementias. dimethyl ether; Methoxy vone; and Retusin 7-methyl ether; or 13. A method according to claim 8, wherein the disorder is a pharmaceutically acceptable salt of any such compound. stroke. 5. A method according to claim 4, wherein the compound 14. A method according to claim 8, wherein the disorder is comprises a (4-methoxyphenyl)-4H-chromene-4-one trauma to the central nervous system or peripheral nervous selected from a group consisting of 4.7-dimethoxyflavone; system. 4'-methoxyflavone: 5,4'-dimethoxyflavone; 5,7,4'-tri 15. A method according to claim 8, wherein the compound methoxyflavone; Acacetin diacetate; Biochanina; Biochanin is Pinosylvin; Derrustone; Methoxyvone; DehydroVariabilin; a diacetate; Formononetn; and Retusin 7-methyl ether; or a or Chrysophanol; or a pharmaceutically acceptable Salt of any pharmaceutically acceptable salt of any such compound. Such compound. 6. A method according to claim 1, wherein the compound 16. A method for treating a disorder that can be treated by comprises a 1,3-benzodioxol Selected from a group consist enhancing arginase activity in a human in need thereof, the US 2013/0245070 A1 Sep. 19, 2013 method comprising administering to the human an effective hydrochloride; Phenindione; Pinosylvin; Pinosylvin methyl amount of Lansoprazole; or a pharmaceutically acceptable ether; Piperine; Pramoxine hydrochloride; Resveratrol salt of Such compound. 4'-methyl ether; Retusin 7-methyl ether; Robustone; Spirono 17. A method for promoting regeneration of a neural cell in lactone; Tilorone; Tranilast; or Xanthone; or a pharmaceuti a human in need thereof, the method comprising administer cally acceptable salt of any such compound. ing to the human an effective amount of a compound that 18. A method according to claim 17, wherein the com enhances arginase activity, wherein the compound is any one pound is Pinosylvin; Derrustone; Methoxyvone; Dehydrova of the following: 2-hydroxyXanthone; 2-methoxyxanthone; riabilin; or Chrysophanol; or a pharmaceutically acceptable 3-methylcholanthrene; 4,7-dimethoxyflavone; 4'-methoxy salt of any Such compound. chalcone; 4'-methoxyflavone; 5,4'-dimethoxyflavone; 5.7,4'- 19. A method for promoting regeneration of a neural cell in trimethoxyflavone: 5,7-dimethoxyisoflavone; 6,3'- a human in need thereof, the method comprising administer dimethoxyflavone; Acacetin diacetate; Anisindione; ing to the human an effective amount of Lansoprazole; or a Apigenin; Apigenin triacetate; Biochanin a; Biochanin a pharmaceutically acceptable salt of Such compound. diacetate; Chlorpropham; Chrysophanol; Daidzein: Dehy 20. A method according to claim 19, wherein the neural cell drovariabilin; Derrubone; Derrusnin, Derrustone; Diben is a neuron. Zoylmethane: Fenbendazole; Formononetn; Genistein; 21. A method according to claim 19, wherein the neural cell Ginkgetin, Indoprofen; Ipraflavone; Liquiritigenin dimethyl is a glial cell. ether; Methoxyvone; Methyl robustone; Phenazopyridine