tory effect and finally, a progression to disease. The Sequential isolates from four patients under zidotheoretical likelihood of the emergence of resistance vudine (ZDV) therapy were obtained using PBMC is increased with a large burden of replicating units, coculture in the absence or in the presence of chronicity of infection, and underlying immunodefi-0.25 M ZDV. PBMC-based HIV susceptibility ciency. Each of these three conditions increases the assay demonstrated the emergence of ZDV-renumber of replicative events in which virus mutations sistance in the sequential isolates from the four leading to resistance can occur. The existence of all patients. Except in one case, the isolates obtained these conditions in patients with advanced HIV infecin the presence of ZDV did not exhibit a greater tion prompted a search for mutants resistant to both resistance pattern than their counterparts obnucleoside and non-nucleoside reverse transcriptase tained in the absence of the drug. In parallel, (RT) inhibitors: zidovudine (ZDV) [Larder and Kemp, partial reverse transcriptase gene sequence was 1989; Kellam et al., 1992] ddI [St. Clair et al., 1991], determined directly on amplified products from ddC [Fitzgibbon et al., 1992], nevirapine [Richmann et proviral DNA. In addition to the mutations preal., 1992; Nunberg et al., 1991], and others. viously described at the critical codons 41, 67, Treatment of individuals infected by HIV with ZDV 70, 215, and 219, numerous additional mutations was started in 1986 [Fischl et al., 1987; Fischl et al., were found in either ZDV-sensitive or ZDV-resis-1990]. The first report on the emergence of ZDV-resistant isolates. The mutation Thr215Tyr was not tant strains was reported by Larder and Kemp [1989; observed in a case of highly resistant virus (ZDV Larder et al., 1989]. These reports were founded on IC 50 Ͼ 6.25 M), while the mutation Lys70Arg the phenotypic and molecular analysis of HIV strains was found in either resistant or sensitive ones.
isolated from patients with AIDS or ARC who had re-The analysis of additional mutations did not received zidovudine therapy for at least 6 months. The veal any clear pattern for ZDV resistance but decrease in ZDV sensitivity, expressed by means of ZDV pointed out the existence of highly variable re-IC 50 and IC 95 values, was associated with regularly ocgions neighbouring the five critical codons. Neicurring mutations at five codons of the reverse tranther nucleotide sequence nor PBMC-based susscriptase (RT) gene which induced the following amino ceptibility assay provided unambiguous data acids substitutions: Met41Leu, Asp67Asn, Lys70Arg, about pretherapy isolates or early on-therapy iso-Thr215Tyr, or Thr215Phe and Lys219Gln. Despite the lates which could predict the emergence of ZDV presence of these substitutions, RT was still functional resistance in further samples.