Immunogenicity of AGS-004 Dendritic Cell Therapy in Patients Treated During Acute HIV Infection

Abstract

AGS-004 consists of matured autologous dendritic cells co-electroporated with in vitro transcribed RNA encoding autologous HIV antigens. In an open-label, single arm sub-study of AGS-004-003, AGS-004 was administered monthly to suppressed participants who started antiretroviral therapy (ART) during acute HIV infection. HIV-1 specific T cell responses were measured by multicolor flow cytometry after 3-4 doses. The frequency of resting CD4+ T-cell infection (RCI) was measured by quantitative viral outgrowth assay. Participants demonstrating increased immune response postvaccination were eligible for analytic treatment interruption (ATI). AGS-004 induced a positive immune response defined as ≥2-fold increase from baseline in the number of multifunctional HIV-1 specific CD28+/CD45RA- CD8+ effector/memory cytoxic T-lymphocytes (CTLs) in all six participants. All participants underwent ATI with rebound viremia at a median of 29 days. Immune correlates between time to viral rebound and the induction of effector CTLs were determined. Baseline RCI was low in most participants (0.043-0.767 IUPM). One participant had a >2-fold decrease (0.179-0.067 infectious units per million [IUPM]) in RCI at week 10. One participant with the lowest RCI had the longest ATI. AGS-004 dendritic cell administration increased multifunctional HIV-specific CD28+/CD45RA- CD8+ memory T cell responses in all participants, but did not permit sustained ART interruption. However, greater expansion of CD28-/CCR7-/CD45RA- CD8+ effector T cell responses correlated with a longer time to viral rebound. AGS-004 may be a useful tool to augment immune responses in the setting of latency reversal and eradication strategies.

Keywords: HIV; HIV eradication; acute HIV infection; analytic treatment interruption; dendritic cell vaccine.

Conflict of interest statement

C.G. has received research support from Bristol Myers Squibb, Gilead Sciences, Abbott, and Janssen (formerly Tibotec Therapeutics). C.H. has received grant support and/or consulting/honoraria from BMS, GSK, Merck, Tibotec Therapeutics, Gilead, Myriad Pharmaceuticals, and Pfizer. D.M. has received research support from Bristol Myers Squibb, Gilead Sciences, and Janssen, has consulted for Merck, and holds common stock in Gilead Sciences. J.E. receives research support from ViiV Healthcare and is a consultant to Bristol Myers Squibb, Merck, Gilead, Janssen, and ViiV Healthcare. M.D., I.T., E.V., A.G., W.L., and C.N. are employees of Argos Therapeutics, Inc., and M.D., E.V., I.T., and C.N have been granted stock options in the company. A.C., J.K., K.M., and M.M. have no competing financial interests.

Funding Sources: This project has been funded in whole or in part by Federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under contract no. N01-AI-60019, AI50410 to the UNC CFAR, and in part by NIH U19-AI096113 to D.M.M. Single-copy assays were funded by intramural NIH funding.

Prior presentation of interim data by: Gay C, Archin N, Tcherepanova I, Villiard E, Hicks C, Kearney M, Coffin J, DeBenedette, M, Eron J, Nicolette C, Margolis DM. Immunogenicity of AGS-004 Dendritic Cell Therapy in Patients Treated during Acute HIV Infection. 21st Conference on Retroviruses and Opportunistic Infections 2014, Boston, MA. March 2–6, Abstract 344.

Figures

FIG. 1.

Study schema. ART, antiretroviral therapy; ATI, analytic treatment interruption; BL, baseline leukapheresis; PS, prescreen; S, screen.

FIG. 2.

Determination of multifunctional T cell responses. PBMC collected pre and post-treatment were cultured in vitro with autologous AGS-004 DCs. The following gating strategy was used to identify central/memory, effector/memory, and effector CTLs. (A) After in vitro stimulation multicolor flow cytometry identified AGS-004-induced HIV Ag-reactive CTL subsets by first gating on live CD3-positive cells present in the lymphocyte gate forward scatter (FSC) vs. side scatter (SSC). CD8-positive cells were then gated through the CD28 histogram and the CD28-positive and CD28-negative subsets were further gated to identify the CCR7, CD27 CD45RA-positive and negative subsets. Boolean gating was used to identify CTL subsets based on the combinatorial expression of the surface markers CD28, CCR7, CD27, and CD45RA. (B) To identify functional cells/ml within each CTL subset the expression of functional markers was determined by gating on cells within the gates for cytokines (IFN-γ, TNF-α, and IL-2), cytolytic markers (Grb and CD107) and proliferation (BrdU). Trucount beads were identified in the FSC versus SSC gate. DC, dendritic cell; IL, interleukin; PBMC, peripheral blood mononuclear cells; FSC, forward scatter; SSC, side scatter.

FIG. 3.

CD8+ T cell responses in participants receiving AGS-004. Box and whisker plots show the distribution of the number of CD8+ T cells (cells/ml) proliferating (BrdU), expressing IFN-γ, CD107a, Grb, IL-2, or TNF-α determined before or post AGS-004 administration for participants. Statistically significant differences between the GFP background response (open bar and whisker plots) and the total antigen payload (GNVR)-specific response (red bar and whisker plots) are shown. Statistically significant differences between the total antigen payload (GNVR) responses determined pre- and post-AGS-004 administration are shown. *p < .05; **p < .01. GNVR, Gag, Nef, Vpr, and Rev; NS, not statistically significant.

FIG. 4.

Multifunctional immune responses to the total antigen RNA payload in participants treated with AGS-004. CD8 T cell responses at baseline (W0) or after AGS-004 administration (W12 or W16) were measured after in vitro stimulation with AGS-004 autologous DCs and functional marker expression was detected by multicolor flow cytometry. (A)Dot plot overlays from one representative participant before AGS-004 administration (W0) and after AGS-004 administration (W12) show the distribution of cellular events representing each functional marker (red dots) BrdU, CD107a, Grb, IFN-γ, IL-2, and TNF-α overlaid on the total CD8+ T cell population (blue dots) gated on the CD28 (y-axis) and CD45RA (x-axis) plots. The numbers are representative of cells/ml expressing each functional marker (red dots). (B–G) The total number of CD28+/CD45RA− CTL (cells/ml, Y-axis) expressing any functional marker after stimulation with DCs encoding GFP or the final product HIV payload (GNVR, GNR, or GVR) was calculated at baseline (W0) and post-AGS-004 (W12 or W16). Total numbers of cells/ml were determined by adding the numbers of cells/ml measured for each individual functional marker. (B) 51–100 to GNVR, (C) 51–102 to GNVR, (D) 54–100 to GNVR, (E) 54–101 to GNR, (F) 054–102 GNR, and (G) 054–104 to GVR. Statistically significant increases in the total number of cells/ml expressing functional markers over baseline are shown. NS, not significant. The contribution of each functional marker (cells/ml, y-axis) within the total response at baseline (W0) or post-AGS-004 (W12 or W16) for each participant is shown, (H) 51–100 to GNVR, (I) 51–102 to GNVR, (J) 54–100 to GNVR, (K) 54–101 to GNR, (L) 054–102 GNR, and (M) 054–104 to GVR. BrdU , CD107a , Grb , IFN-γ , IL-2 , and TNF-α . Starred (*) markers indicate statistically significant increased CTL responses from pre- to post-AGS-004 administration. GNR, Gag, Nef, and Rev; GVR, Gag, Vpr and Rev.

FIG. 5.

Increased numbers of effector CTL within the CD28−/CCR7−/CD45RA− CTL subset correlates with longer time to viral rebound. CD8+ T cell responses at baseline (W0) or after AGS-004 administration (W12 or W16) were measured after in vitro stimulation with AGS-004 autologous DCs and functional marker expression was detected by multicolor flow cytometry. (A) Absolute numbers of CTL (cells/mL) for each effector marker (BrdU •, CD107a ▲, Grb □, IFN-γ X, IL-2 ◯, and TNF-α △) were calculated by subtracting the number of CTL determined at baseline (W0) from the number of CTL determined after AGS-004 administration (W12 or W16). Absolute numbers of CTL with effector marker expression for the CD28+/CD45RA− CTL subset (A), the CD28+CCR7+CD27−CD45RA− CTL subset (B), and the CD28−CCR7−CD45RA− CTL subset (C) were plotted versus time to viral rebound (days) for each participant. For each subset only those effector markers that had either a positive correlate with time to viral rebound or an inverse correlate with time to viral rebound are shown. Correlates were determined by nonparametric bivariate Spearman's Rho statistical analysis.

FIG. 6.

Multifunctional immune responses to the individual HIV antigens present in the administered AGS-004 payload. The total number of CD28+/CD45RA− CTL (cells/ml, X-axis) expressing the any functional marker after in vitro stimulation with DCs encoding the single HIV antigens are expressed as the values (cells/ml) greater than twofold over baseline. (A) Participant responses to GAG, (C) participant responses to Nef, (E) participant responses to Vpr, and (G) participant responses to Rev. The percent contribution of each individual functional marker to the total response is represented by pie charts for participant responses to (B) GAG, (D) Nef, (F) Vpr, and (H) Rev. Pie charts of representative responses to BrdU , CD107a , Grb , IFN-γ , IL-2 , TNF-α are shown clockwise starting at 12 o'clock.

FIG. 7.

Measurement of viral load and CD4 T cell counts in participants receiving AGS-004. Plasma HIV RNA levels (red lines) and CD4 cell counts (blue lines) are shown for participants treated with AGS-004. The blue shaded areas indicate the period of analytic treatment interruption (ATI) across study visits and overlying arrows indicate the duration of ATI (days). AGS-004 administrations are denoted by black arrows. Participant 51–102 is not shown given drug monitoring revealed ongoing ART administration during the ATI period. ART, antiretroviral therapy; PS, prescreen; S, screen.