Anti-HIV Drugs (FDA Approved)
Protease Inhibitors
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Saquinavir; Ro 31-8959; Fortovase®; Invirase®
Indinavir; MK639; L-735,524; Crixivan®
Ritonavir; ABT-538; Norvir®
Nelfinavir; AG-1343; Viracept®
Amprenavir; VX-478; 141W94; Agenerase®
All of the HIV protease inhibitors that have been approved and most that are in development, are non-hydrolysable transition state peptidomimetics in which the cleavage site peptide linkage is replaced is replaced by transition state isosteres, such as statine, norstatine, hydroxyethylene, a reducedamide,hydroxyethyl, or dihydroxyethylene. Many inhibitors have been designedto be symmetrical to take advantage of the C2 symmetry of the dimeric enzyme.Although symmetrical inhibitors can result in tighter binding as well asasimpler designand synthetic pathway, symmetrical compounds may be more susceptibleto viral resistance since single mutations in the protease have multiplicativeeffects on inhibitor binding.
Rational iterative drug design based on structural studies of the
HIV protease in conjunction with molecular modeling has yielded many compounds
in a brief period of time with significant clinical potential. Four protease
inhibitors, saquivanir (Invirase®, Hoffman-LaRoche), ritonavir (Norvir®,
Abbott), indinavir (Crixivan®, Merck) and nelfinavir (Viracept®,Agouron)
have already been approved and several others are in the late stagesof clinical
development. Current efforts areunderway to develop simpler compoundswith
higher bioavailability and less susceptibility to viral resistance. Pharmacokinetic
problems are also being addressed through the development of prodrugs, the
improvement of formulations and the development of inhibitors of liver metabolizing
enzymes.
Saquinavir is a synthetic, transition state, peptidomimetic analog which inhibits the HIV protease and thereby prevents the maturation and infectivity of the viral particle. Saquinavir is active against bothHIV-1and HIV-2 protease with EC50 values in the range of 1-30 nM (Roberts,N.etal. 1990, Craig, J. et al. 1991, Ohta, Y. et al.1997). Low bioavailability(~ 4%) due to poor absorption and extensive first pass metabolism limitstheuse of saquinavir in potent triple combination therapies (Ohta, Y.et al 1997). A new soft gel formulation which increases bioavailability by 3-fold hasrecentlybeen approved (Cadman, J. 1997). The combinationof ritonavir andsaquinavirhas also been reported to dramatically increase saquinavir plasmaconcentrations(by as much as 50-fold). Ritonavir is believed to act by inhibitingcytochromeP450 (CYP 3A4), the enzyme responsible for saquinavir first pass metabolism(Kempf, D. et al 1997). Genotypic analysis of the protease genefrom invitro selected isolates indicated that the Gly48Val and Leu90Metmutantshad reduced susceptibility to saquinavir (Ohta, Y. et al 1997)
Roberts, N.; Martin, J.; Kinchington, D.; Broadhurst, A.; Craig, J.; Duncan, I.; Galpin, S.; Handa, B.; Kay, J.; Krohn, A.; Lambert, R.; Merrett, J.; Mills, J.; Parkes, K.; Redshaw, S.; Ritchie, A.; Taylor, J.; Thomas,G.;Machin, P. (1990) Rational design of peptide-based HIV proteinase inhibitors. Science. 248. 258-361.
Craig, J.; Duncan, I.; Hockley, D.; Grief, C.; Roberts, N.; Mills, J. (1991) Antiviral propeties of Ro31-8959, an inhibitor of HIV proteinase. Antiviral Res. 16. 295-305.
Ohta, Y. Shinkai, I. (1997) Saquinavir. Bioorg. Med. Chem . 5, 465-466.
Cadman, J. (1997 apr/may) Roche brings new formulationof saquinavir to FDA. GMHC Treat Issues. 11, 8.
Kempf, D. Marsh, K., Kumar., G.; Rodriguez,A.; Denissen, J.; McDonald, E.; Kukulka, M.; Hsu, A.; Granneman, G.; Baroldi, P.; Sun, E.; Pizzuti, D.; Plattner, J.; Norbeck, D; Leonard, J. (1997) Pharmacokinetic enhancement of inhibitors of the human immunodeficiency virus protease by coadministration with ritonavir. Antimicrob Agents Chemother. 41. 654-60.
Ritonavir is a potent protease inhibitor with high oral bioavailability.In rats, following an oral 10 mg/kg dose, ritonavir was 78 % bioavailableandhad a maximal plasma concentration (Cmax) of 2.62 µM, twohours after dosing. Ritonavir was soluble to 5.3 µg/mL at pH 7.4 and6.9 µg/mL at pH 4.0 and had a plasma half-life of 1.2 hours, followinga 5 mg/kg i. v.dose (Kempf, D. et al, 1995).
Kempf, D.; Marsh, K.; Denissen, J.; McDonald,E.; Vasavanonda, S.;Flentge, C.; Green, B.; Fino, L.; Park, C.; Kong, X.;Plattner, J.; Leonard, J; Norbeck, D. (1995) Abt-538 is a potent inhibitorof human immunodeficiency virus protease and has high oral bioavailabilityin humans. Proc. Natl. Acad. Sci. 92. 2484-2488.
Nelfinavir is a relatively small protease inhibitor with goodoral bioavailability
(52% in rats at 50 mg/kg; 9-42% in monkeys at 25mg/kg).Although, the
half life after 25 mg/kg i.v. and 12.5 mg/kg i.v.dosesin rats and monkeys
was only 1 and 1.5 hr respectively, plasma levelsexceeding the EC95
were maintained for over 7 hrs following oral administrationdue to slow absorptivity
(Kalish, V. et al. 1995; Shetty, B. et al, 1996).Nelfinavir also does not
appear to have much cross-resistance to either ritonaviror indinavir resistant
strains containing single mutations, but exhibitssome cross-resistance to
strains with multiple mutations, and saquinavir resistantstrains (Patick,
A. et al, 1996).
Kalish, V; Kaldor, S.; Shetty, B.; Tatlock, J.; Davies, J.; Hammond,M.; Dressman, B.; Fritz, J.; Appelt, K.; Reich,S.; Musick, L.; Wu, B.; Su,K. (1995) Iterative protein structure-based drug design and synthesisof hivprotease inhibitors. Eur. J. Med. Chem. 30 s201-s214.
Patick, A.; Markowitz, M.; Appelt, K.; Wu, B.;Musick, L.; Kalish,V.; Kaldor, S.; Reich, S.; Ho, D.; Webber, S. (1996) Antiviraland resistancestudies of ag1343, an orally bioavailable inhibitor of humanimmunodeficiencyvirus protease. Antimicrob. Agents Chemother. 40.292-297.
Shetty, B.; Kosa, M.; Khalil, D.; Webber, S. (1996) Preclinical pharmacokinetics and distribution to tissue of ag1343, an inhibitorof human immunodeficiency virus type 1 protease. Antimicrob. Agents Chemother. 40 110-114.
Protease Gene Mutations by Protease Inhibitors
Nucleoside Analogs
Zidovudine: AZT; Azidothymidine; Retrovir® from GlaxoWelcome
Didanosine; Dideoxyinosine; ddI; Videx® from Bristol-Myers Squibb
Zalcitabine; Dideoxycytidine; ddC; Hivid® from Hoffman-La Roche
Lamivudine; 3TC; Epivir® from IAF Biochem Int/Glaxo Welcome
Stavudine; 2',3'-Didehydro-3'-deoxythymidine; D4T; Zerit® from Bristol-Myers Squibb
Abacavir succinate; 1592U89 Succinate; Ziagen® from Glaxo-Welcome
Anti-HIV activity is dependent on the intracellular phosphorylation of the analog and the ability of the phosphorylated analog to interact with the HIV-RT. The rate limiting step in most cells is believed to bethe initial phosphorylation by nucleoside kinases, or in the case of AZT, the conversion of a nucleoside monophosphate to a nucleoside diphosphate. Many 2'-3' dideoxynucleoside analogs have potent inhibitory effects onviral RT when in the triphosphate form, but little effect as a nucleoside on HIV infected cells. This lackof activity is believed to be due to low affinity between the nucleoside ornucleotide analog and cellular kinases, such as thymidine kinase. Nucleosidekinase andnucleotide kinase activity varies widely between cells. Some HIV-infectablecells, such as monocytes and macrophages, when at rest, are believed to havelittle kinase activity. This could account for the inability of AZT and othernucleoside analogs to prevent transmission when given immediately after HIVexposure. Prodrugs such as methoxyglycinyl derivativesand bis[S-(2-hydroxyethylsulfidyl)-2-thioethyl] esters have been designed to form monophosphorylated nucleoside analogs intracellularly. Acyclic nucleoside phosphonates, such as 9-(2-phosphonylmethoxyethyl)adenine (PMEA), have also been used to overcome the kinase bottleneck. The acyclic phosphonateanalog, 9-(2-phosphonylmethoxypropyl)adenine (PMPA) has been reported to prevent SIV transmission in macaques, even when administered 24 hoursafterexposure (Herdewijn, P. et al, 1987).
Another approach to increase the potency of nucleoside analogshas been to use potentiating drugs to increase the amount of dideoxynucleoside triphosphates and decrease the amount of deoxynucleoside triphosphates. Ribavirin and hydroxy urea have been shown to enhance the anti-HIV activity of ddI bysuppressing the formation of dATP and facilitating the conversion of ddI toddATP without increasing the toxicity of ddI.
The major limitations of nucleoside analogs include their toxicity,lack of activity in some cell types, and susceptibility to viral resistance.Toxic side effects vary from compound to compound: anemia and/or neutropeniaare frequently seen with AZT; neutropenia and peripheral neuropathy with3TC;peripheral neuropathy with ddC, D4T and ddI; and acute pancreatitis withddI. HIV viral isolates from patients are often resistant to the nucleoside analogthat was used therapeutically in thepatient. Resistance has also been reportedin patients who have not been treated with nucleoside analogs.
The mutations responsible for viral resistance to nucleosideanalogshave all been mapped to the RT enzyme. The five following mutations in HIV-1 RT, confer a high level of resistance to AZT: 41 Met--Leu; 67 Asp--Asn;70 Lys--Arg; 215 Thr--Phe/Tyr; 219 Lys--Gln. Multiple mutations in RT havealso been reported to occur in an ordered fashion, such as: 41--41/215--41/67/215--41/67/70/215--41/67/70/215/219; with each mutation leading to accrued resistance. Although extensive structural and genetic studies have been done on AZT resistant enzymes, a biochemical explanation for AZT resistance is still lacking. Most AZT-resistant enzymes bind AZT-triphosphate with the same avidity as nonresistant enzymes. Resistance to ddI is conferred by the 74 Leu--Val mutation, the 69 Thr--Asp mutationreduces susceptibility to ddC; the 75 Val--Thr mutation confers resistanceto D4T. The 184 Met--Val mutation not only confers multiple resistance to ddC , 3TC and ddI, but also will supress the effects of AZT resistant mutations resulting in an enzyme that is once again susceptible to AZT.Enzymes containing the 184 Met toVal mutation also makes 50-fold less errorscompared to the wild-type enzyme. This increased fidelity that results from 3TC resistance, shouldalsoreduce the appearance of protease resistant strains. Currently, manytreatmentregimens consist of two nucleoside analogs (i.e. AZT and 3TC) anda proteaseinhibitor. These treatments are designed to lead to the greatestreductionin viral burden and also to prevent protease resistant strains fromappearing (Herdewijn, P. et al, 1987).
AZT was one of the first nucleoside analogs shown tohavepotent anti-HIV activity in-vitro (Mitsuya, H. et al, 1985). AZT enters cellsby passive diffusion,and appears to be phosphorylated by the same enzymes that convert thymidine (dT) to dT-5'-TP. AZT-triphosphate (AZT-TP) isthe active species, acting as a chain terminating substrate for HIV reverse transcriptase (HIV-RT) during either first or second strand DNA synthesis. Although resistanceto AZT frequently develops, the addition of 3TC (Lamivudine) to a therapeuticregimen can cause AZT-resistant strains to revert to AZT-sensitive strains (Larder, B. et al, 1995).
Mitsuya, H.; Weinhold, K.; Furman, P.; St. Clair, M.;Nusinofff-Lehrman, S.; Gallo, R.; Bolognesi, D.; Barry, D.; BroderS. (1985) 3-azido-3'- deoxythymidine (bwa509u): An antiviralagent that inhibits the infectivity and cytopathic effect of humant-lymphotropic virustype iii/ lymphadenopathy- associatedvirus in vitro. Proc. Natl. Acad. Sci. 82. 7096-7100
Larder, B.; Kemp S.; Harrigan, P. (1995) Potential mechanisimfor sustained antiretroviral efficacy of AZT-3TC combination therapy. Science , 269, 696-699.
Herdewijn, P.; Balzarini, J.; De Clercq, E.; Pauwels, R.;Baba, M.; Broder, S.; Vanderhaeghe, H. (1987) 3'-Subsituted 2',3'- dideoxynucleoside analogues as potential anti-HIV agents. J. Med. Chem. 30. 1270-1278.
Daluge, S.; Purifoy, D.; Savina, P.; St. Clair, M.; Parry,N.; Dev, I.; Novak, P.; Ayers, K.; Reardon, J.; Roberts, G.; Fyfe, J.;Blum, M.; Averett, D.; Dornsife, R.; Domin, B.; Ferone,R.; Lewis, D.; Krenitsky,T. (1994) 5-Chloro-2',3'-dideoxy-3'-fluorouridine (935U83), a selective anti-HIVagent with an improved metabolic and toxicological profile. Antimicrob.Agents Chemother. 38 1590-1603.
Non-Nucleoside Reverse Transcriptase Inhibitors; NNRTI's)
Nevirapine; BI-RG-587; Viramune® from Boehringer-Ingelheim (Roxane)
Delavirdine; BHAP; U-90152; Rescriptor® from Pharmacia Upjohn
Efavirenz; DMP-266; Sustiva® from Dupont-Merck