Genetic characterization of rebounding human immunodeficiency virus type 1 in plasma during multiple interruptions of highly active antiretroviral therapy

Abstract

Various strategies of interrupting highly active antiretroviral therapy (HAART) are being investigated for the treatment of human immunodeficiency virus (HIV) infection. Interruptions of greater than 2 weeks frequently result in rebound of plasma HIV RNA. In order to discern changes in the viral population that might occur during cycles of treatment interruption, we evaluated the homology of HIV-1 envelope gene sequences over time in 12 patients who received four to seven cycles of 4 weeks off HAART followed by 8 weeks on HAART by using the heteroduplex tracking assay and novel statistical tools. HIV populations in 9 of 12 patients diverged from those found in the first cycle in at least one subsequent cycle. The substantial genetic changes noted in HIV env did not correlate with increased or decreased log changes in levels of plasma HIV RNA (P > 0.5). Thus, genetic changes in HIV env itself did not contribute in a systematic way to changes in levels of plasma viremia from cycle to cycle of treatment interruption. In addition, the data suggest that there may be multiple compartments contributing to the rebound of plasma viremia and to viral diversity from cycle to cycle of intermittent therapy.

FIG. 1.

Week 4 HTA and plasma HIV RNA levels from multiple HAART interruptions. Plasma HIV RNA was determined by branched-chain DNA assay and is reported as the number of copies per milliliter (•). Nine of 12 patients (patients 101, 103, 104, 106, 107, 108, 114, 115, and 121) had heterogeneous HIV env during four to seven cycles of treatment interruptions (A), while 3 patients (patients 102, 105, and 111) maintained similar levels of HIV env as determined by HTA (B). The sample from which the probe was made is indicated by an asterisk.

FIG. 1.

Week 4 HTA and plasma HIV RNA levels from multiple HAART interruptions. Plasma HIV RNA was determined by branched-chain DNA assay and is reported as the number of copies per milliliter (•). Nine of 12 patients (patients 101, 103, 104, 106, 107, 108, 114, 115, and 121) had heterogeneous HIV env during four to seven cycles of treatment interruptions (A), while 3 patients (patients 102, 105, and 111) maintained similar levels of HIV env as determined by HTA (B). The sample from which the probe was made is indicated by an asterisk.

FIG. 2.

Reproducibility of HTA results. Independent PCRs for paired samples from week 4 of multiple cycles of HAART interruption were evaluated in three patients to analyze the reproducibility of variant population sampling (A). The OC for each pair is provided. In addition, the effects of different numbers of input HIV RNA copies on HTA were evaluated by serial dilutions of a single sample from patient 101 (B). Two independent PCRs were performed for each sample. The OC for each sample pair is provided. The overall OC was 0.19.

FIG. 3.

HTA and plasma HIV RNA levels for patients with multiple HIV env isolates within cycles off HART. Plasma HIV RNA was determined by branched-chain DNA assay and is reported as the number of copies per milliliter (•). Four patients (patients 101, 104, 105, and 115) had weekly evaluations during each of 4 weeks off HAART during a cycle of 4 weeks off HAART followed by 8 weeks on HAART. The first sample shown for each cycle is the first sample with >50 copies of HIV RNA per ml. The sample from which the probe was made is indicated by an asterisk.

FIG. 4.

OC plots for patients with divergent HIV env variants within cycles off HAART. The plot reduces the complexity of the gel data to a single comparative measure to assist in interpretation of the gels. The OC, a measure of divergence in mobility and intensity of bands between pairs of lanes, is calculated and displayed for each pair of lanes or time points. Warm (more red) colors indicate greater divergence of the banding patterns between two lanes, while cool (more blue) colors indicate greater homogeneity of banding patterns. cyc, cycle; Pt, patient.