Low human immunodeficiency virus envelope diversity correlates with low in vitro replication capacity and predicts spontaneous control of plasma viremia after treatment interruptions

Abstract

Genetic diversity of viral isolates in human immunodeficiency virus (HIV)-infected individuals varies substantially. However, it remains unclear whether HIV-related disease progresses more rapidly in patients harboring virus swarms with low or high diversity and, in the same context, whether high or low diversity is required to induce potent humoral and cellular immune responses. To explore whether viral diversity predicts virologic control, we studied HIV-infected patients who received antiretroviral therapy (ART) for years before undergoing structured treatment interruptions (STI). Viral diversity before initiation of ART and the ability of the patients to contain viremia after STI and final cessation of treatment was evaluated. Seven out of 21 patients contained plasma viremia at low levels after the final treatment cessation. Clonal sequences encompassing the envelope C2V3C3 domain derived from plasma prior to treatment, exhibited significantly lower diversity in these patients compared to those derived from patients with poor control of viremia. Viral diversity pre-ART correlated with the viral replication capacity of rebounding virus isolates during STI. Neutralizing antibody activity against autologous virus was significantly higher in patients who controlled viremia and was associated with lower pretreatment diversity. No such association was found with binding antibodies directed to gp120. In summary, lower pretreatment viral diversity was associated with spontaneous control of viremia, reduced viral replication capacity and higher neutralizing antibody titers, suggesting a link between viral diversity, replication capacity, and neutralizing antibody activity.

Figures

FIG. 1.

Relationship between plasma virus concentrations and nucleotide sequence diversities observed in 16 clones isolated from each sample (A). The sequenced clones are representative of the actual viral populations present in vivo in 21 patients before antiretroviral therapy due to the high HIV copy number input into the RT-PCRs (total RNA extracted from 800 to 900 ul plasma was used) (B).

FIG. 2.

Inferred neighbor-joining phylogenetic trees of clonal HIV-1 env C2-C3 sequences derived from seven patients in the controlling group (A) and 14 patients in the noncontrolling group (B). Triangles represent the compressed subtrees containing 16 clones isolated from plasma of each patient before antiretroviral therapy. The length of the triangle corresponds to the respective intrapatient diversity. Bootstrap values are indicated on each branch. The bar denotes 5% nucleotide divergence and diversity; HXB2 was included as external reference strain.

FIG. 3.

Comparison between controlling and noncontrolling patients. Pretreatment nucleotide diversity (A) and amino acid diversity (B) within individual subjects (16 clones each). (C) HIV DNA levels in peripheral blood mononuclear cells during the first treatment interruption cycle following 1.6 to 3.9 years of continuous antiretroviral therapy. Box plots symbolize median value and interquartile range; whiskers extend over the entire range. P values were calculated by nonparametric Mann-Whitney test.

FIG. 4.

Correlation between pretreatment viral diversity and in vitro growth characteristics of virus isolated during the fifth treatment interruption. Replication capacity was derived from the profile of HIV p24 antigen production in cultures by linear regression (slope of natural logarithms of p24 levels over 6 days). Spearman correlation analysis: r2 = 0.26, P = 0.04, n = 16.

FIG. 5.

HIV-specific cellular immune responses in patients exhibiting low (lo = below median) and high (hi = above median) pretreatment viral diversity determined at week 0 (baseline of STI), week 39 (before the fifth treatment interruption) and week 50 (during the last treatment interruption). Magnitude of CD8+ cytotoxic T-cell response (CTL) expressed as total spot forming colonies (SFC) per 106 cells (A); 6-31 (median 18) different peptides were tested. The breadth of the CD8+ CTL response is indicated as the percentage of peptides tested that induced a response >50 SFC/106 cells (B). Magnitude of the HIV-specific CD4+ T-cell response expressed as SFC per 106 CD8-depleted PBMC (C).

FIG. 6.

Comparison of 70% neutralizing titers found in 7 controlling and 12 noncontrolling patients (A). Neutralization activity of patient plasma derived from the first STI cycle was tested against autologous virus isolates derived during the fifth cycle. Dashed line represents the detection limit. Lower pretreatment diversity was correlated with higher neutralizing antibody activity (B, r2 = 0.24, P = 0.03, n = 19), whereas no correlation between diversity and titers of antibodies binding to monoclonal anti-gp120 was found (C).