Volume 5, Issue 6, Pages 788-797 (June 2002) A Phase II Randomized Study of HIV-Specific T-Cell Gene Therapy in Subjects with Undetectable Plasma Viremia on Combination Antiretroviral Therapy  Steven G. Deeks, Bridget Wagner, Peter A. Anton, Ronald T. Mitsuyasu, David T. Scadden, Christine Huang, Catherine Macken, Douglas D. Richman, Cindy Christopherson, Carl H. June, Richard Lazar, David F. Broad, Sayeh Jalali, Kristen M. Hege  Molecular Therapy  Volume 5, Issue 6, Pages 788-797 (June 2002) DOI: 10.1006/mthe.2002.0611 Copyright © 2002 American Society for Gene Therapy Terms and Conditions

FIG. 1 Cumulative incidence of viral rebound in plasma. Viral rebound was defined as either a single isolated increase in plasma HIV RNA to > 40 copies/ml (“blip”) or consecutive increases in plasma HIV RNA (“viral relapse”). The bars show the cumulative number of patients in the gene-modified and unmodified groups who experienced any rebound (blip + relapse) before infusion and at monthly intervals following the first T-cell infusion. Both groups experienced viral blips before infusion (8–12 weeks). There was a trend toward fewer patients with viral rebound in the gene-modified group through week 20 after infusion (P = 0.07). Molecular Therapy 2002 5, 788-797DOI: (10.1006/mthe.2002.0611) Copyright © 2002 American Society for Gene Therapy Terms and Conditions

FIG. 2 Change from baseline in HIV reservoir assays following T-cell infusion. For all assays, baseline values represent an average of all preinfusion measurements, and data were log10-transformed before analysis. Charts show mean log change from baseline over time in the CD4ζ gene–modified (filled triangle) and unmodified (filled square) groups. Arrows designate T-cell infusions. (A) Limiting-dilution quantitative HIV coculture. Analysis was conducted at weeks −4, 0, 8, 12, and 24. The assay range was 0.01 to 20 infectious units per million (IUPM) with a mean SD equal to 0.27 log. (B) HIV DNA in blood mononuclear cells by DNA PCR. Analysis was conducted at weeks −8, −6, −4, −2, 0, 2, 4, 8, 12, 16, 20, and 24. Assay sensitivity was 10 copies/106 cells. (C) HIV DNA in rectal mucosal biopsy samples by DNA PCR. (D) HIV mRNA in rectal mucosal biopsy samples by RNA RT-PCR. Rectal biopsies were done at weeks −4, −2, 8, 12, and 24. Assay sensitivity was 10 copies/µg mRNA or 2 × 106 copies β-globin DNA. Significant decreases from baseline were seen in the gene-modified group at week 24 in the HIV coculture (–0.36 log, P = 0.02) and rectal HIV DNA (–0.50 log; P = .007) assays. Molecular Therapy 2002 5, 788-797DOI: (10.1006/mthe.2002.0611) Copyright © 2002 American Society for Gene Therapy Terms and Conditions

FIG. 3 Change in CD4+ cell count. Both the gene-modified (filled triangle) and unmodified (filled square) groups showed an increase in CD4+ cells over time, with an absolute increase at 24 weeks of 41 and 37 cells/mm3, respectively. There were no significant differences between groups. Arrows designate T-cell infusions. Molecular Therapy 2002 5, 788-797DOI: (10.1006/mthe.2002.0611) Copyright © 2002 American Society for Gene Therapy Terms and Conditions

FIG. 4 Impact of viral-load rebound on latent reservoirs and CD4+ cell counts. We conducted a subset analysis on patients with stable viral-load suppression throughout the study (“never blip,” filled triangle) versus those with evidence of transient or sustained viral-load rebound (“blip + relapse,” filled square). (A) Log change from baseline in quantitative HIV coculture (IUPM). A consistent decay was only noted in the “never blip” group (–0.24 log, P = 0.05, week 8; −0.20 log, P = 0.08, week 12; −0.27 log, P = 0.04, week 24). (B) Log change from baseline in rectal biopsy HIV DNA (copies/106 cells). A significant decay was only noted in the “never blip” group (–0.16 log, P = 0.06, week 12; −0.38 log, P = 0.02, week 24). (C) Change from baseline in CD4+ cell counts. A significant increase in CD4+ cells was only seen in the “blip + relapse” subgroup, with significant between-group differences noted at week 8 (45 versus −11, P = 0.04), week 16 (62 versus 10, P = 0.05), week 20 (74 versus 8, P = 0.07), and week 24 (81 versus −3, P = 0.02). Molecular Therapy 2002 5, 788-797DOI: (10.1006/mthe.2002.0611) Copyright © 2002 American Society for Gene Therapy Terms and Conditions

FIG. 5 Persistence of CD4ζ-modified T cells and transgene expression in vivo. Analyses were done on subjects receiving gene-modified cells only. Arrows designate T-cell infusions. (A) All patients were monitored for levels of CD4ζ T cells in PBMCs before infusion, at week 0 (30 minutes after infusion), and weeks 2, 4, 8, 12, 16, and 24 by quantitative Tagman DNA PCR. All subjects had undetectable levels before infusion and detectable levels postinfusion at all time points at or after week 2. The lower limit of detection of this assay is 117 copies/106 cells (dashed line). Mean log10-transformed values are shown. (B) In a subset of six subjects, CD4+ and CD8+ T cells were purified before DNA PCR analysis [21]. Mean log10-transformed values (copies/106 cells) of CD4ζ in selected CD4+ (filled triangle) and CD8+ (filled square) T-cell populations are shown, demonstrating stable persistence of both gene-modified CD4+ and CD8+ T cells. (C) CD4ζ RNA expression over time was quantified in a subset of six subjects by quantitative Taqman RT-PCR on PBMCs [21]. All subjects showed relatively stable CD4ζ RNA levels (log10 copies/106 cells) over time, although two subjects had undetectable levels at isolated time points (signified by small arrows). Molecular Therapy 2002 5, 788-797DOI: (10.1006/mthe.2002.0611) Copyright © 2002 American Society for Gene Therapy Terms and Conditions