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Artículo destacado 
Drugs of the Future
Bevirimat Dimeglumine (USAN) [1S-(1α,3aβ,5aα,5bβ,7aα,9β,11aβ,11bα,13aβ,13bα)]-1-Isopropenyl-9-(3,3-dimethylsuccinyloxy) 3β-(3-Carboxy-3-methylbutyryloxy)lup-20(29)-en-28-oic acid O-(3,3-Dimethylsuccinyl)betulinic acid InChI=1/C36H56O6/c1-21(2)22-12-17-36(30(40)41)19-18-34(8)23(28(22)36)
C36H56O6 CAS: 174022-42-5 EN: 246885 Abstract Despite the advances made in the treatment of HIV/AIDS, particularly with the widespread use of highly active antiretroviral therapy (HAART), side effects and the emergence of resistant strains of virus remain major problems. Therefore, a major focus of antiretroviral research is the discovery of compounds with novel structures and/or mechanism(s) of action. Bevirimat is a first-in-class maturation inhibitor derived from betulinic acid which was shown to have potent and selective activity against HIV-1, including resistant strains. Further studies revealed that its mechanism of action involves disruption of a late step in Gag processing. Clinical studies indicated good efficacy and safety after once-daily doses and phase II trials are under way. Synthesis Bevirimat can be synthesized from betulinic acid as follows: Betulinic acid (I) is extracted from the leaves of Syzigium claviflorum (1). Acylation of (I) with 2,2-dimethylsuccinic anhydride (II) in the presence of DMAP in refluxing pyridine furnishes the target 3’,3’-dimethylsuccinyl betulinic acid along with minor amounts of the undesired 2’,2’-dimethyl regioisomer (III), which is separated by means of preparative HPLC (2-5). Scheme 1.
Scheme 1: Synthesis of Bevirimat Introduction According to UNAIDS (2006 Report on the Global AIDS Epidemic [UNAIDS]), there are currently about 40 million adults and children living with HIV/AIDS worldwide, and the cumulative death toll over the first 25 years of the HIV pandemic was in excess of 25 million, as reported at the XVI International AIDS Conference in 2006. A growing number of antiretroviral agents and combinations are available for the treatment of HIV/AIDS. The current standard of care is highly active antiretroviral therapy, or HAART, a combination of 3 or more therapeutic agents, at least 2 of which are active antiretrovirals. The use of HAART has significantly reduced the morbidity and mortality associated with HIV infection (6, 7). However, although HAART is effective at reducing the levels of HIV in the blood, treatment failure leading to viral rebound remains a problem. The main reasons are poor compliance with the HAART regimen due in large part to unpleasant side effects which can lead to suboptimal therapy, and the rapid emergence of treatment-resistant strains of HIV (8-11). Moreover, it has been estimated that up to 10% of HIV-infected individuals are resistant to all classes of reverse transcriptase and protease inhibitors (12), and the main focus of current antiretroviral research is therefore the discovery of new compounds with novel structures or mechanism(s) of action. Promising agents under development include attachment inhibitors, CCR5 co-receptor antagonists, CXCR4 co-receptor antagonists and integrase inhibitors (10, 13, 14). The natural compound betulinic acid, isolated from S. claviflorum, was found to possess anti-HIV activity and chemical modifications were undertaken in an attempt to identify potent anti-HIV agents, leading to the discovery of bevirimat (PA-457, YK-FH-312) as a compound with a novel mechanism of action (1, 2). Further studies demonstrated bevirimat to be a first-in-class maturation inhibitor with potent activity against HIV-1 that acts by disrupting Gag processing (15-24). Bevirimat, which was granted fast track designation by the FDA in 2004, is presently in phase II clinical evaluation (25). Preclinical Pharmacology The activity of bevirimat in inhibiting HIV replication was studied both in vitro and in vivo. With a mean IC50 of 10.3 nM against wild-type isolates in peripheral blood mononuclear cells (PBMCs), bevirimat demonstrated antiviral activity comparable to the approved drugs zidovudine and indinavir (IC50 = 4.3 and 8.8 nM, respectively), while being significantly more active than the non-nucleoside reverse transcriptase inhibitor (NNRTI) nevirapine (IC50 = 40.0 nM). When tested against a range of resistant isolates using MT-2 cells, bevirimat retained low nanomolar inhibitory activity, in contrast to approved drugs. Its activity was also found to be specific for HIV-1, with no effect against HIV-2 or SIV. The therapeutic index for bevirimat was reported to be > 2,500 (CC50 = 25 µM). When combined with approved anti-HIV drugs, bevirimat exhibited synergy or additive activity (16, 17, 26, 27). In vivo studies were carried out in NL4-3-infected SCID-hu Thy/Liv mice over a 21-day period. Oral administration of a salt of bevirimat reduced viral load in a dose-dependent manner, causing reductions of 2.1, 0.9 and 0.3 log10 copies/ml, respectively, at doses of 100, 30 and 10 mg/kg/day (26). The antiviral mechanisms of bevirimat were extensively studied. Bevirimat was found not to act by blocking virus attachment or entry or by inhibiting reverse transcriptase, integrase or protease (2, 17, 27). Preliminary experiments using a MAGI (multinuclear-activation galactosidase indicator) assay indicated that bevirimat interferes with viral maturation by disrupting a late step in Gag processing, specifically inhibiting the conversion of the capsid (CA) precursor p25 to the mature CA protein p24, resulting in the release of noninfective viral particles (17-19). Further studies indicated that bevirimat specifically protects the CA-p2 (or CA-SP1) cleavage site from processing by the viral protease during virion maturation, a novel anti-HIV mechanism. This was confirmed in viral resistance studies with the drug (20-24, 28-30). In vitro, 5 amino acid changes (H226Y, L231M, L231F, A1V and A3V) conferring bevirimat resistance were identified; all were located at the CA-SP1 junction. However, resistance to bevirimat was not detected in vivo during a single-dose phase I/II and a 10-day multiple-dose phase II study in patients (28, 31). Also, in some cases, resistant mutants showed reduced viral fitness (20, 28, 29), and resistant viruses remained susceptible to all other classes of anti-HIV drugs (29, 30). Pharmacokinetics and Metabolism Preclinical studies carried out in rats demonstrated that orally administered bevirimat was metabolized to several glucuronide conjugates, including a di-glucuronide and two mono-glucuronides, which were mainly eliminated in bile. Cumulative biliary excretion demonstrated excretion mainly in the form of a mono-glucuronide. The metabolites displayed relatively high stability in vitro (32). To characterize the metabolic profile of bevirimat, the compound was tested in a variety of in vitro systems, and the drug was administered orally and intravenously to mice, rats, marmosets and dogs to characterize its in vivo disposition. Results of in vitro studies indicated that only rat liver microsomes significantly metabolized bevirimat. No significant inhibition of cytochrome P-450 CYP1A2, CYP2C19, CYP2D6 or CYP3A4 was seen at concentrations up to 100 µM, while CYP2C9 was inhibited with an IC50 of about 10 µM; no inhibition of human P-glycoprotein was observed (33, 34). The oral bioavailability of bevirimat was greatest in marmosets (about 60%) and lowest in dogs (about 10%), and the half-life was also longest in marmosets (about 14 h); bioavailability in rats was 14% and the terminal half-life was about 2 h (27, 34). The oral bioavailability and the main elimination routes of bevirimat were evaluated in vivo in rats administered the drug i.v. (5 mg/kg) or orally (25 mg/kg as the disodium salt); rats with chronic biliary catheters were also studied. After i.v. administration, bevirimat showed moderate systemic clearance of 0.45 ± 0.128 l/h/kg, an estimated terminal t1/2 of 1.4 h and a volume of distribution of 0.78 l/kg; after oral administration, the mean Cmax was 3.6 mg/ml and the bioavailability was 21 ± 12%. Neither bevirimat nor glucuronide was detected in urine. Although negligible amounts of unchanged drug were detected in bile, a significant amount of the dose was recovered as bevirimat after treatment of bile with β-glucuronidase. Bile collection reduced the i.v. AUC of bevirimat from 11.8 to 7.14 mg.h/ml, indicating that a significant portion of the dose may be subject to enterohepatic recycling. The results indicate that the clearance of bevirimat in rats occurs primarily via glucuronidation with subsequent biliary excretion (35). The pharmacokinetics of bevirimat were also evaluated in clinical studies. A double-blind, placebo-controlled, dose-escalation study of single doses of bevirimat (25, 50, 100 and 250 mg) was carried out in 32 healthy male volunteers. Bevirimat was rapidly absorbed, with very high plasma levels (2.3-31.1 µg/ml) and a long half-life (2.5 h at 250 mg). The results suggested that therapeutic concentrations (about 2.3 µg/ml) can be achieved with a single daily oral dose as low as 50 mg. The drug was well tolerated (36). Another double-blind, placebo-controlled, dose-escalation study assessed the pharmacokinetics of multiple oral doses (25, 50 and 100 mg once daily for 10 days) in 24 healthy male volunteers. Bevirimat was rapidly absorbed, achieving very high plasma levels (Cmax = 2.62, 6.02 and 11.09 µg/ml, respectively, on day 1, and 7.98, 15.58 and 31.58 µg/ml, respectively, on day 10) and showing a long half-life (56.3-68.9 h). Both peak plasma levels and exposure were dose-dependent, and 3-5-fold plasma accumulation was seen with multiple dosing. Target therapeutic concentrations could be safely achieved even at the lowest dose (37). Single oral doses (75, 150 and 250 mg) of bevirimat were evaluated for pharmacokinetics/pharmacodynamics (PK/PD) in a placebo-controlled study in 24 HIV-infected patients. Bevirimat exhibited linear pharmacokinetics. Volume of distribution at steady state and oral clearance were 13.3 l and 0.17 l/h, respectively, and bevirimat had a prolonged half-life of 60.3 h. The agent dose-dependently inhibited HIV replication, with a maximum inhibition of viral replication of 33.2%, 45.0% and 49.6%, respectively, for doses of 75, 150 and 250 mg. The mean concentration of bevirimat in the effect site inhibiting viral replication by 50% (Ce50) was 59.4 ng/ml and the area under the HIV RNA effect curve (AUEC) was dose-dependent (38, 39). The PK/PD of multiple doses of bevirimat were also characterized in 32 HIV-infected adults in a phase II clinical study. In this study, the patients received placebo or oral bevirimat once daily at 25, 50, 100 or 200 mg for 10 days. Bevirimat again demonstrated linear pharmacokinetics, with a long mean half-life of 62.7 h. The mean oral clearance of bevirimat was 0.21 l/h. Both AUC and trough concentrations of bevirimat were highly associated with antiviral response. Maximum antiviral effect (Emax) for bevirimat was not reached at the highest dose (200 mg) tested (40). Safety Bevirimat was well tolerated in preclinical animal studies. No teratogenic effects were observed at doses of 900 mg/kg/day in rats and 300 mg/kg/day in rabbits administered during gestation, well above the anticipated human dose of 200 mg/day (41). Clinical Studies The safety and antiviral activity of bevirimat were also reported in the above-mentioned double-blind, placebo-controlled phase II study in HIV-infected patients (40, 42). All doses of bevirimat were generally safe and well tolerated, with no grade 3 or 4 laboratory abnormalities and only mild or moderate adverse events. The study showed that doses of 100 and 200 mg significantly reduced HIV RNA load compared with placebo. Median decreases on day 11 were 0.17, 0.48 and 1.03 log10 copies/ml for the doses of 50, 100 and 200 mg, respectively. Genotypic data showed no evidence of resistance development to bevirimat. In the single-dose study in 24 HIV-infected patients, all patients treated with bevirimat (75, 100 or 250 mg) showed reductions in mean viral load compared with placebo. Bevirimat demonstrated dose-dependent antiviral activity, with maximum viral load decreases of 0.27, 0.45 and 0.51 log10 copies/ml, respectively, on doses of 75, 100 and 250 mg, compared to a reduction of 0.17 log10 copies/ml on placebo. No drug-related adverse events were seen. Pre-existing drug resistance mutations in 2 subjects remained susceptible to bevirimat (43). Drug Interactions An open-label, randomized study in 48 healthy volunteers examined the PK/PD interactions between bevirimat and atazanavir. Subjects received bevirimat 200 mg once daily for 14 days or atazanavir 400 mg once daily on days 1-21 and bevirimat 200 mg once daily on days 8-21. Bevirimat was generally well tolerated and was not associated with an increase in serum bilirubin, as seen with atazanavir. Co-administration did not significantly increase serum bilirubin levels compared to atazanavir alone and no significant pharmacokinetic interaction between the two anti-HIV agents was seen (19, 44). Also, no clinically relevant interaction between bevirimat and ritonavir has been observed (19). Source Panacos Pharmaceuticals, Inc. (US). References 1. Fujioka, T., Kashiwada, Y., Kilkuskie, R.E. et al. Anti-AIDS agents, 11. Betulinic acid and platanic acid as anti-HIV principles from Syzigium claviflorum, and the anti-HIV activity of structurally related triterpenoids. J Nat Prod 1994, 57(2): 243-7. 2. Kashiwada, Y., Hashimoto, F., Cosentino, L.M., Chen, C.-H., Garrett, P.E., Lee, K.-H. 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Anti-human immunodeficiency virus activity of YK-FH312 (a betulinic acid derivative), a novel compound blocking viral maturation. Antimicrob Agents Chemother 2001, 45(4): 1225-30. 19. Martin, D.E. Recent data on PA-457 maturation inhibitor. 14th Int Symp HIV Emerg Infect Dis (ISHEID) (June 21-23, Toulon) 2006, Abst OP 4.5. 20. Zhou, J., Chen, C.H., Aiken, C. The sequence of the CA-SPI junction accounts for the differential sensitivity of HIV-I and SIV to the small molecule maturation inhibitor 3-O-{3’,3’-dimethylsuccinyl}-betulinic acid. Retrovirology 2004, 1: 15. 21. Li, F., Zoumplis, D., Matallana, C. et al. The determinants of activity of the HIV-1 maturation inhibitor PA-457 map to residues flanking the Gag CA-SP1 cleavage site. 12th Conf Retroviruses Opportunistic Infect (Feb 22-25, Boston) 2005, Abst 256. 22. Zhou, J., Yuan, X., Dismuke, D. et al. Small-molecule inhibition of human immunodeficiency virus type 1 replication by specific targeting of the final step of virion maturation. J Virol 2004, 78(2): 922-9. 23. Zhou, J., Huang, L., Hachey, D.L., Chen, C.H., Aiken, C. Inhibition of HIV-1 maturation via drug association with the viral Gag protein in immature HIV-1 particles. J Biol Chem 2005, 280(51): 42149-55. 24. Sakalian, M., McMurtrey, C.P., Deeg, F.J., Maloy, C.W., Li, F., Wild, C.T., Salzwedel, K. 3-O-(3’,3’-Dimethylsuccinyl) betulinic acid inhibits maturation of the human immunodeficiency virus type 1 Gag precursor assembled in vitro. J Virol 2006, 80(12): 5716-22. 25. Lee, K. Recent progress in the discovery and development of plant-derived chemotherapeutic agents. Int Chem Congr Pacific Basin Soc (Dec 15-20, Honolulu) 2005, Abst 1395. 26. Allaway, G.P., Bare, J., Kilgore, N.R., Reddick, M., Martin, D.E., Wild, C.T., Stoddart, C.A. The first-in-class maturation inhibitor, PA-457, is a potent inhibitor of HIV-replication both in vitro and in vivo. 15th Annu Int AIDS Conf (July 11-16, Bangkok) 2004, Abst WePeA5643. 27. Wild, C., Kilgore, N.R., Reddick, M.S. et al. In vitro and in vivo pre-clinical analyses of PA-457, a novel betulinic acid derivative that potently inhibits HIV-1 replication. 14th Int AIDS Conf (July 7-12, Barcelona) 2002, Abst MoPeA3030. 28. Adamson, C., Salzwedel, K., Castillo, A. et al. Viral resistance to PA-457, a novel inhibitor of HIV-1 maturation. 13th Conf Retroviruses Opportunistic Infect (Feb 5-9, Denver) 2006, Abst 156. 29. Salzwedel, K., Goila-Gaur, R., Adamson, C. et al. Selection for and characterization of HIV-1 isolates resistant to the maturation inhibitor PA-457. 13th Int HIV Drug Resist Workshop: Basic Princ Clin Implic (June 8-12, Adeje), 2004, Abst 4. 30. Salzwedel, K., Goila-Gaur, R., Li, F. et al. Selection for and characterization of HIV-1 isolates resistant to the maturation inhibitor PA-457. 15th Int AIDS Conf (July 11-16, Bangkok) 2004, Abst WeOrA1276. 31. Castillo, A., Adamson, C., Doto, J., et al. Genotypic analysis of the Gag CA/SPI cleavage site in patients receiving the maturation inhibitor PA-457. 15th Int HIV Drug Resist Workshop (June 13-17, Sitges) 2006, Abst 32. 32. Wen, Z., Stern, S.T., Martin, D.E., Lee, K.-H., Smith, P.C. Structural characterization of anti-HIV drug candidate PA-457 [3-O-(3’,3’-dimethylsuccinyl)-betulinic acid] and its acyl glucuronides in rat bile and evaluation of in vitro stability in human and animal liver microsomes and plasma. Drug Metab Dispos 2006, 34(9): 1436-42. 33. Martin, D.E., Smith, P., Goila-Gaur, R. et al. Determinants of activity, in vitro metabolism and in vivo disposition of the novel maturation inhibitor PA-457. 11th Conf Retroviruses Opportunistic Infect (CROI) (Feb 8-11, San Francisco) 2004, Abst 545. 34. Martin, D.E., Smith, P., Wild, C.T., Allaway, G.P. In vitro and in vivo disposition of PA-457, a novel inhibitor of HIV-1 maturation. 15th Int AIDS Conf (July 11-16, Bangkok) 2004, Abst WePeA5644. 35. Smith, P.C., Bender, B.C., Wild, C.T., Allaway, G.P., Martin, D.E. Disposition of PA-457, a novel inhibitor of HIV-1 maturation, in rats involves extensive biliary excretion of a glucuronide. Drug Metab Rev 2003, 35(Suppl. 2): Abst 442. 36. Martin, D.E., Ballow, C.H., Blum R., Doto, J., Wild, C.T., Allaway, G.P. The safety, tolerability and pharmacokinetics of PA-457, a first-in-class HIV maturation inhibitor, in healthy volunteers. 7th Int Cong Drug Ther HIV Infect (Nov 14-18, Glasgow) 2004, Abst P312. 37. Martin, D.E., Ballow, C., Doto, J., Blum, R., Wild, C., Allaway, G. The safety, tolerability, and pharmacokinetics of multiple oral doses of PA-457, the first-in-class HIV maturation inhibitor, in healthy volunteers. 12th Conf Retroviruses Opportunistic Infect (Feb 22-25, Boston) 2005, Abst 551. 38. Ogundele, A., Smith, P., Forrest, A., Martin, D. Pharmacokinetic/pharmacodynamic effects of a novel maturation inhibitor (PA-457) in HIV-infected patients following a single oral dose. 3rd Int AIDS Soc Conf HIV Pathog Treat (July 24-27, Rio de Janeiro) 2005, Abst TuPe3.1B01. 39. Smith, P., Ogundele, A., Forrest, A., Martin, D.E. PKPD modeling & simulation of PA-457 from a single dose viral dynamic study in HIV-infected patients. 45th Intersci Conf Antimicrob Agents Chemother (ICAAC) (Dec 16-19, Washington, D.C.) 2005, Abst A-1149. 40. Smith, P., Forrest, A., Beatty, G. et al. Pharmacokinetics/pharmacodynamics of PA-457 in a 10-day multiple dose monotherapy trial in HIV-infected patients. 13th Conf Retroviruses Opportunistic Infect (CROI) (Feb 5-9, Denver) 2006, Abst 52. 41. Martin, D.E., Alexander, T., Bollinger, J. et al. PA-457, a first-in-class maturation inhibitor, is non-teratogenic in rats an rabbits. 16th Int AIDS Conf (Aug 13-18, Toronto) 2006, Abst CDA0140. 42. Beatty, G., Laleazari, J., Eron, J. et al. Safety and antiviral activity of PA-457, the first-in-class maturation inhibitor, in a 10-day monotherapy study in HIV-1 infected patients. 45th Intersci Conf Antimicrob Agents Chemother (ICAAC) (Dec 16-19, Washington, D.C.) 2005, Abst H-416d. 43. Martin, D.E., Jacobson, J., Schurmann, D., Osswald, E., Doto, J., Wild, C., Allaway, G. PA-457, the first-in-class maturation inhibitor, exhibits antiviral activity following a single oral dose in HIV-1-infected patients. 12th Conf Retroviruses Opportunistic Infect (Feb 22-25, Boston) 2005, Abst 159. 44. Martin, D.E., Gailbraith, H., Schettler, J., Ellis, C., Doto, J. Lack of a PK/PD interaction between PA-457 and atazanavir (ATV) in healthy volunteers. 46th Intersci Conf Antimicrob Agents Chemother (ICAAC) (Sept 27-30, San Francisco) 2006, Abst A-377. Y. Wang. 8106 Runnymeade Dr., Frederick, MD 21702, USA. N. Serradell. Prous Science, P.O. Box 540, 08080 Barcelona, Spain. Drugs of the Future Vol. 32, No. 1, 2007, pp. 7-11
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