AEGiS-12IAC: HIV maturation and stability in the presence of protease inhibitors in vitro.

12th International AIDS Conference

Geneva, Switzerland - June 28-July 3, 1998

HIV maturation and stability in the presence of protease inhibitors in vitro.

Int Conf AIDS 1998 Jun 28-Jul 3; 12:9 (abstract no. 11133)

Donovan RM, Friedman DA, Caceres HJ, Baxa DM, Bush CE, Markowitz NP;;; Henry Ford Hospital, Detroit, MI, USA.

BACKGROUND: Protease inhibitors (PIs) act as competitive inhibitors of the HIV viral protease. The objective of this study was to compare virucidal versus virustatic properties of the currently used PIs by examining the ability of immature HIV virions to mature, and the stability of RNA in immature virions.

METHODS: HeLa/LAV cells constitutively producing HIV were grown in the presence or absence of saquinavir, indinavir, nelfinavir or ritonavir to generate immature HIV virions. Immature HIV virions were incubated for time periods ranging from minutes to several days, under a variety of conditions of temperature and pH. The resultant HIV Gag protein forms were then analyzed by western blot, and a novel antigen capture ELISA for Pr55Gag. HIV RNA was measured by RT-PCR.

RESULTS: Western blots showed that all cultures containing 1 microM or more of any of the 4 PIs had a strong Pr55Gag band, and a fainter high molecular weight Gag-Pol band. Cultures without drugs had strong p24 and p17 bands and faint or absent bands of immature forms. Quantitative results from antigen capture ELISA on immature virions incubated at 37 degrees C for 48 hours showed that immature Gag forms remained at 85%, 84%, 98%, and 96% of initial values for saquinavir, indinavir, nelfinavir, and ritonavir, respectively. Removal of residual PI by repeated pelleting of the virion at 23,000 x g, or by ultrafiltration did not cause the immature HIV virions to mature. Incubation of the virions under conditions optimized for the activity of the HIV protease similarly did not lead to detectable maturation of the viral Gag proteins. The average HIV RNA copy number in immature virions after incubation for 48 hours at 37 degrees C was 80% of their initial value, while the average of mature virions was 90%. HIV virions disrupted by detergent immediately had RNA copy numbers fall to less than 1% of their initial value.

CONCLUSIONS: We found no evidence that immature HIV virions produced in the presence of any of the drugs used as protease inhibitors were able to undergo maturation in vitro. However, the HIV RNA genome in immature virions does appear to be stable for relatively long time periods.

Keywords: AEGIS, HIV, HIV Protease, Protease Inhibitors, Saquinavir, Gene Products, gag, Virion, Nelfinavir, Indinavir, HIV Protease Inhibitors, Blotting, Western, HIV Envelope Protein gp41, In Vitro, immunology, ICA12
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