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(12) United States Patent (10) Patent No.: US 9,168,269 B2 Koyuncu Et Al

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(12) United States Patent (10) Patent No.: US 9,168,269 B2 Koyuncu Et Al USOO9168269B2 (12) United States Patent (10) Patent No.: US 9,168,269 B2 Koyuncu et al. (45) Date of Patent: Oct. 27, 2015 (54) INHIBITORS OF LONG AND VERY LONG (58) Field of Classification Search CHAIN EATTY ACID METABOLISMAS None BROAD SPECTRUMANTI-VIRALS See application file for complete search history. (75) Inventors: Emre Koyuncu, Princeton, NJ (US); (56) References Cited Joshua D. Rabinowitz, Princeton, NJ (US); Thomas Shenk, Princeton, NJ U.S. PATENT DOCUMENTS (US) 2001/0033864 A1 10, 2001 Colonno et al. 2008/0045482 A1 2/2008 Robbins et al. (73) Assignee: THE TRUSTEES OF PRINCETON 2009/0304634 A1 12/2009 Erickson-Miller et al. UNIVERSITY, Princeton, NJ (US) FOREIGN PATENT DOCUMENTS *) Notice: Subject to anyy disclaimer, the term of this patent is extended or adjusted under 35 EP 1914237 A3 4, 2008 U.S.C. 154(b) by 0 days. WO 95.29688 A1 11, 1995 OTHER PUBLICATIONS (21) Appl. No.: 13/579,967 Rider et al. PLoS ONE 2011 (6) 1-15.* (22) PCT Filed: Feb. 18, 2011 Munger et al. Nature Biotechnology 2008 (26) 1179-1186.* PCT International Search Report and Written Opinion for PCT/ US 1 1/25558 mailed Aug. 10, 2011. (86). PCT No.: PCT/US2011/025558 Morikawa et al., "Complete inhibition of Human Immunodeficiency S371 (c)(1) Virus Gag Myristoylation is Necessary for Inhibition of Particle . Budding”, JBC 1996, 271(5): 2868-2873. (2), (4) Date: Apr. 1, 2013 Yao et al., "Long chain acyl-CoA synthetase 3-mediated phosphalidylcholine synthesis is required for assembly of very low (87) PCT Pub. No.: WO2011/103516 density lipoproteins in human hepatoma Huh? cells”. J. Biol Chem 2006, 283(2):849-54. PCT Pub. Date: Aug. 25, 2011 Gortmaker et al., “Effect of Combination Therapy Including Protease 65 Prior Publication D Inhibitors on Mortality Among Children and Adolescents Infected (65) rior Publication Data with HIV-1, N Engl J Med 2001, 345:1522-8. US 2013/O190381 A1 Jul. 25, 2013 Tornodo et al., “Evidence for an Essential Role of Long9. Chian Acyly CoA Synthetase in Animal Cell Proliferation'. The Journal of Bio O O logical Chemistry 1991, 266(7):4214-9. Related U.S. Application Data Pub Chem CID:9576787. Oct. 24, 2006. Triascinc-compound sum (60) Provisional application No. 61/305,862, filed on Feb. mary. 18, 2010. * cited by examiner (51) Int. Cl. A6 IK3 L/45 (2006.01) Primary Examiner — Jeffrey S Lundgren A6 IK3I/40 (2006.01) Assistant Examiner — Michael Schmitt A6 IK3I/713 (2006.01) (74) Attorney, Agent, or Firm — Kenyon & Kenyon LLP A6 IK3I/536 (2006.01) A6 IK 45/06 (2006.01) (57) ABSTRACT A6 IK3I/4I55 (2006.01) The present invention provides methods and compounds for A6 IK3I/439 (2006.01) treating viral infections using modulators of host cell A6 IK3I/445 (2006.01) enzymes relating to long chain fatty acid and lipid droplet A6 IK3 L/46 (2006.01) metabolism. It includes a method of treating viral infections (52) U.S. Cl. using triacsin C and its relatives, analogs and derivatives as CPC ........... A61 K3I/713 (2013.01); A61K 31/4155 well as other inhibitors of long chain fatty acid metabolism (2013.01); A61 K3I/439 (2013.01); A61 K and lipid droplet metabolism. 3 1/445 (2013.01); A61 K3I/46 (2013.01); A6 IK3I/536 (2013.01); A61K 45/06 (2013.01) 4 Claims, 18 Drawing Sheets U.S. Patent Oct. 27, 2015 Sheet 1 of 18 US 9,168,269 B2 32% U.S. Patent Oct. 27, 2015 Sheet 2 of 18 US 9,168,269 B2 Triacsin C Cell viability Wirus production e-eCMV-F Serum+ 3OOE--O5 r&YHCMV+ Serum+ 251E05 OnM 25 nM 50 nM 100 nM 250 nM Triacsin C HCMV HSV-1 Adenovirus influenza A 1. 2 O 1. O O 80 6.67X 20 27.1X N 12.6X 17.0X 28.5 x 0nM 100nM 25 OM 100M 250nM 100nM 250nM 100nM 250M Triacsin C concentration MRC-5 cells Fig. 3 U.S. Patent Oct. 27, 2015 Sheet 3 of 18 US 9,168,269 B2 3 r r i U.S. Patent Oct. 27, 2015 Sheet 4 of 18 US 9,168,269 B2 Aminoxyacetic acid - Influenza A MDCK cells - influenza A - WSN - 0.001 pfu/cell 0.5 mM AOAA 1 mM AOAA SS S &S SSS 48 hpi 72 hpi 8 8s s :* Š8. 1.0003 NY. Š saSa...S. & : *::::::::::::::: 1.00E00 or s-------------- Indulum sh 23 Fig. 5 U.S. Patent Oct. 27, 2015 Sheet 5 of 18 US 9,168,269 B2 SS S.S s e L. s Iufni AWOH U.S. Patent Oct. 27, 2015 Sheet 6 of 18 US 9,168,269 B2 is s S.S s u/n WWOH U.S. Patent US 9,168,269 B2 !|901100'z í801309z ?pornost |301100't |501300's ~40011000 Iu/n WWOH U.S. Patent Oct. 27, 2015 Sheet 9 of 18 US 9,168,269 B2 PF-1052 10 M S& Š S S Phase $8 S.Š Š Š S: S. & SS pUL99-GFP Virus production s s 2 D d O s Y 5.90E0 2.5, SM GM PF-1052 Fig. 10 U.S. Patent Oct. 27, 2015 Sheet 10 of 18 US 9,168,269 B2 1.00E (9 proc S. Sas: Yes . i.00E08 or N Y. I (JOE-07 - Es-- w w w y w S E.00E+05 ir .00E404 ic NWehicle 100E03 to wPF 1052 (5 M) S s E.00E-02 ir E1. OE- icswsa - - - - ------------------------------------------------- ---------------------------------- g s s s is s: s 'll Fig.11 1, 3 ····················································································· 1OLM Fig. 12 is is EGM Fig. 13 U.S. Patent US 9,168,269 B2 waL-oxapunoduopXII-opuapunoduuoo XII-opul? U.S. Patent Oct. 27, 2015 Sheet 16 of 18 US 9,168,269 B2 r s & s R s s s R s s s & s R s s s R s s s S. D Š s s s s s r s s s s s s i d s s Ln s n s s s u/n AWOH U.S. Patent Oct. 27, 2015 Sheet 17 of 18 US 9,168,269 B2 |d?OZT96ZZ8tvtvzZTW ------- (pou/WWOH) (esueup pop)ao US 9, 168,269 B2 1. 2 INHIBITORS OF LONG AND VERY LONG cycloalkyl. When the carbonatoms of R' are substituted, it is CHAIN FATTY ACID METABOLISMAS preferred that from 0-8 hydrogenatoms along the chain may BROAD SPECTRUMANTI-VIRALS be replaced by a substituent selected from halo, OR, SR, lower alkyl, and cycloalkyl, wherein R is H, C, alkyl, and STATEMENT OF GOVERNMENT INTEREST C. cycloalkyl. In certain preferred embodiments, R' is unsubstituted (i.e., R' is unbranched, and none of the hydro This invention was made with government Support under gens have been replaced by a substituent). In preferred Grants #1 R01 AI 078063 and #5 R01 CA085786 awarded by embodiments for compounds of the formula I, R' has a chain the National Institutes of Health. The government has certain length of 8 to 12 atoms. More preferably, R has a total chain rights in the invention. 10 length of R' has a chain length of 9 to 11 atoms. Most pref CROSS-REFERENCE TO RELATED erably R' has a chain length of 10 atoms. In other preferred APPLICATIONS embodiments, R' has 2 to 4 double bonds. In anotherembodi ment of the invention, the compound of Formula I is a triacsin. This application is a 371 of International Patent Applica 15 In yet another embodiment of the invention, the compound of tion No.: PCT/US2011/025558 filed Feb. 18, 2011, which Formula I is triacsin C. In still another embodiment of the claims the benefit of priority to U.S. patent application Ser. invention, the compound is an inhibitor of acyl-CoA syn No. 61/305,862 filed Feb. 18, 2010, all of which are incorpo thetase long-chain family member 1 (ACSL1). In still another rated herein by reference in their entireties. embodiment of the invention, the compound is an inhibitor of another enzyme inhibited by triacsin C, including without FIELD OF THE INVENTION limitation arachidonoyl-CoA synthase or enzymes of triglyc eride (TG) or cholesterol ester (CE) synthesis. The present invention provides methods and compounds Compounds of Formula I are broad spectrum antivirals for treating viral infections using modulators of host cell useful for treating or preventing infection by a wide range of enzymes relating to long chain fatty acid and lipid droplet 25 viruses. In one embodiment, the compounds of Formula I are metabolism. It includes a method of treating viral infections used to treat or prevent viral infection by an enveloped virus. using triacsin C and its relatives, analogues and derivatives as In certain embodiments of the invention, the compounds are well as other inhibitors of long chain fatty acid metabolism used to treat or prevent infection by human cytomegalovirus and lipid droplet metabolism. (HCMV), herpes simplex virus-1 (HSV-1), influenza A, or 30 hepatitis C virus. BACKGROUND OF THE INVENTION The invention also provides a pharmaceutical composition There is a great unmet medical need for agents that more for treatment or prevention of a viral infection comprising a safely, effectively, and reliably treat viral infections, from therapeutically effective amount of a composition comprising HIV to the common cold. This includes a major need for 35 a compound or prodrug thereof, or pharmaceutically accept better agents to treat human cytomegalovirus (where current able salt of said compound or prodrug; and a pharmaceuti agents Suffer from significant toxicity and lack of efficacy), cally acceptable carrier, wherein the compound is a com herpes simplex virus (where current agents are beneficial but pound of Formula I.
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