Protease inhibitors: current status and future prospects.

One hope to maintain the benefits of antiviral therapy against the human immunodeficiency virus type 1 (HIV-1), despite the development of resistance, is the possibility that resistant variants will show decreased viral fitness. To study this possibility, HIV-1 variants showing high-level resistance (up to 1,500-fold) to the substrate analog protease inhibitors BILA 1906 BS and BILA 2185 BS have been characterized. Active-site mutations V32I and I84V/A were consistently observed in the protease of highly resistant viruses, along with up to six other mutations. In vitro studies with recombinant mutant proteases demonstrated that these mutations resulted in up to 10(4)-fold increases in the Ki values toward BILA 1906 BS and BILA 2185 BS and a concomitant 2,200-fold decrease in catalytic efficiency of the enzymes toward a synthetic substrate. When introduced into viral molecular clones, the protease mutations impaired polyprotein processing, consistent with a decrease in enzyme activity in virions. Despite these observations, however, most mutations had little effect on viral replication except when the active-site mutations V32I and I84V/A were coexpressed in the protease. The latter combinations not only conferred a significant growth reduction of viral clones on peripheral blood mononuclear cells but also caused the complete disappearance of mutated clones when cocultured with wild-type virus on T-cell lines. Furthermore, the double nucleotide mutation I84A rapidly reverted to I84V upon drug removal, confirming its impact on viral fitness. Therefore, high-level resistance to protease inhibitors can be associated with impaired viral fitness, suggesting that antiviral therapies with such inhibitors may maintain some clinical benefits.

Impaired fitness of human immunodeficiency virus type 1 variants with high-level resistance to protease inhibitors.

It is currently well established that HIV-1 Vpr augments viral replication in primary human macrophages. In its virion-associated form, Vpr has been suggested to aid efficient translocation of the proviral DNA into the cell nucleus. Although Vpr growth-arrests dividing T cells, the relevance of this biological activity in nondividing macrophages is unclear. Here we use Vpr-mutants to demonstrate that the molecular determinants involved in G2-arresting T cells are also involved in increasing viral transcription in macrophages, even though these cells are refractive to the diploid DNA status typical of G2 phase. Our results suggest that the two phenotypes, namely the nuclear localization and the G2-arrest activity of the protein, segregate functionally among the late and early functions of Vpr. The nuclear localization property of Vpr correlates with its ability to effectively target the proviral DNA to the cell nucleus early in the infection, whereas the G2-arrest phenotype correlates with its ability to activate viral transcription after establishment of an infection. These two functions may render Vpr's role essential and not accessory under infection conditions that closely mimic the in vivo situation, that is, primary cells being infected at low viral inputs.

https://rupress.org/jem/article-pdf/187/7/1103/1116092/97-2244.pdf

Human immunodeficiency virus type 1 Vpr is a positive regulator of viral transcription and infectivity in primary human macrophages

Organocatalytic Enantioselective γ-Aminoalkylation of Unsaturated Ester: Access to Pipecolic Acid Derivatives

Objectives: To assess the in-vitro drug susceptibility of a panel of five well-characterized drug-resistant HIV variants to recently developed anti-HIV compounds including seven reverse transcriptase (RT) inhibitors and seven protease inhibitors. Methods: Drug-resistant viral strains were selected on the basis of the prevalence of these mutants in patient samples from local area HIV clinics. The isolates included one multinucleoside-resistant virus containing the Q151M mutation, and four clinical isolates containing multiple RT and protease resistance mutations. The activity of the experimental compounds against these isolates was determined using drug susceptibility assays and measuring the viral antigen p24 end-point. Results: These clinically relevant highly drug-resistant viruses were resistant to many of the new compounds in clinical development. In most cases the resistance mutations of the clinical isolate were different from those selected in vitro for the particular experimental compound. Conclusions: It is critical to expand the preclinical development of new drugs to include the assessment of their activity against currently circulating highly drug-resistant clinical strains, in order to develop appropriate salvage therapies for patients harboring resistant strains.

Highly drug-resistant HIV-1 clinical isolates are cross-resistant to many antiretroviral compounds in current clinical development

Palinavir is a potent inhibitor of the human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) proteases. Replication of laboratory strains (HIV-1, HIV-2, and simian immunodeficiency virus) and HIV-1 clinical isolates is inhibited by palinavir with 50% effective concentrations ranging from 0.5 to 30 nM. The average cytotoxic concentration of palinavir (35 microM) in the various target cells indicates a favorable therapeutic index. Potent antiviral activity is retained with increased doses of virus and with clinical isolates resistant to zidovudine (AZT), didanosine (ddI), or nevirapine. Combinations of palinavir with either AZT, ddI, or nevirapine demonstrate synergy or additivity in the inhibition of HIV-1 replication. Palinavir retains anti-HIV-1 activity when administered postinfection until times subsequent to the reverse transcription step. In chronically infected CR-10 cells, palinavir blocks Gag precursor polyprotein processing completely, reducing greater than 99% of infectious particle production. The results indicate that the antiviral activity of palinavir is specific to inhibition of the viral protease and occurs at a late stage in the replicative cycle of HIV-1. On the basis of the potent in vitro activity, low-level cytotoxicity, and other data, palinavir was selected for in-depth preclinical evaluation.

E Cohen

Papers

Antiviral properties of palinavir, a potent inhibitor of the human immunodeficiency virus type 1 protease.