Immunogenicity of personalized dendritic-cell therapy in HIV-1 infected individuals under suppressive antiretroviral treatment: interim analysis from a phase II clinical trial

Abstract

Background: We developed a personalized Monocyte-Derived Dendritic-cell Therapy (MDDCT) for HIV-infected individuals on suppressive antiretroviral treatment and evaluated HIV-specific T-cell responses.

Methods: PBMCs were obtained from 10 HIV+ individuals enrolled in trial NCT02961829. Monocytes were differentiated into DCs using IFN-α and GM-CSF. After sequencing each patient's HIV-1 Gag and determining HLA profiles, autologous Gag peptides were selected based on the predicted individual immunogenicity and used to pulse MDDCs. Three doses of the MDDCT were administered every 15 days. To assess immunogenicity, patients' cells were stimulated in vitro with autologous peptides, and intracellular IL-2, TNF, and interferon-gamma (IFN-γ) production were measured in CD4+ and CD8+ T-cells.

Results: The protocol of ex-vivo treatment with IFN-α and GM-CSF was able to induce maturation of MDDCs, as well as to preserve their viability for reinfusion. MDDCT administration was associated with increased expression of IL-2 in CD4+ and CD8+ T-cells at 15 and/or 30 days after the first MDDCT administration. Moreover, intracellular TNF and IFN-γ expression was significantly increased in CD4+ T-cells. The number of candidates that increased in vitro the cytokine levels in CD4+ and CD8+ T cells upon stimulation with Gag peptides from baseline to day 15 and from baseline to day 30 and day 120 after MDDCT was significant as compared to Gag unstimulated response. This was accompanied by an increasing trend in the frequency of polyfunctional T-cells over time, which was visible when considering both cells expressing two and three out of the three cytokines examined.

Conclusions: MDDC had a mature profile, and this MDDCT promoted in-vitro T-cell immune responses in HIV-infected patients undergoing long-term suppressive antiretroviral treatment. Trial registration NCT02961829: (Multi Interventional Study Exploring HIV-1 Residual Replication: a Step Towards HIV-1 Eradication and Sterilizing Cure, https://www.clinicaltrials.gov/ct2/show/NCT02961829 , posted November 11th, 2016).

Keywords: Dendritic-cell therapy; HIV GAG; HIV cure research; HLA Haplotypes; Precision medicine.

Conflict of interest statement

ILS, AS, and RSD have a patent application for Dendritic-cell therapy.

© 2021. The Author(s).

Figures

Fig. 1

Phenotypic characterization of immature and mature MDDCs used for MDDCT. Panels A,B) PBMCs were isolated from the total blood of the enrolled individuals and induced to MDDC differentiation/maturation as depicted in Additional file 1: Fig. S1. Both immature MDDCs (iMDDC) and mature MDDCs (mMDDCs) were analyzed by flow cytometry for the expression of the activation/maturation markers HLA-DR, CD80, CD86, CD83, CD40, and CCR7 according to the gating strategy depicted in Additional file 1: Fig. S7A. Box plots and whiskers represent the median fluorescence intensity of each marker (A) or the percentage of cells expressing each marker (B). Data are expressed as median ± min/max and were analyzed by the non-parametric Wilcoxon test (N of patients = 10).** p < 0.01

Fig. 2

Viability of MDDCs used for each MDDCT dose preparation. Viability was assessed by flow cytometry following staining with a LIVE/DEAD fixable stain according to the gating strategy shown in (A). The horizontal line in the graph of panel B indicates the median (N of patients for each dose = 10). Data were analyzed by Two-Way ANOVA. * p < 0.05

Fig. 3

Immunogenicity of MDDCT. Ten recipients (Table 1) received three doses of the personalized MDDCT. PBMCs were collected upon administration of the first MDDCT dose (day 0), upon dose 2 (day 15), and dose 3 administration (day 30), as well as during the post-therapy follow-up (day 120). Isolated PBMCs were stimulated in vitro with the autologous Gag peptides that were used for MDDCT. Production of IFN-γ, IL-2, and TNF by CD4+ and CD8+ T-lymphocytes was evaluated by flow cytometry. After applying the logit transformation to restore normality, data of patients for whom all time points were available for a given cytokine were analyzed by one-way ANOVA followed by Dunnet’s post-test. *p < 0.05; **p < 0.01; ***p < 0.001

Fig. 4

Frequency of polyfunctional CD4+ and CD8+ T-lymphocytes following MDDCT. PBMCs were collected upon administration of the first MDDCT dose (day 0), upon dose 2 (day 15), and dose 3 administration (day 30), as well as during the post-therapy follow-up (day 120). Isolated PBMCs were stimulated in vitro with the autologous Gag peptides that were used for MDDCT. Production of IFN-γ, IL-2, and TNF by CD4+ and CD8+ T-lymphocytes was evaluated by flow cytometry. Panels A,B) Frequency of cells expressing one or more immune-mediator cytokine (IL-2, TNF and IFN-γ). Pie charts show the relative percentage, among cells expressing at least one cytokine, of CD4+ (A) and CD8+ (B) T-lymphocytes expressing one, two, or three cytokines. Panels C, D) Bar graphs showing the absolute frequency of CD4+ (C) and CD8+ (D) T-lymphocytes expressing each cytokine combination analyzed. Data are expressed as mean ± SD and were analyzed by two-way ANOVA. (N of patients for each dose = 10)

Fig. 5

The proportion of patients displaying an undetectable HIV-1 DNA level in rectal biopsies at the end of all treatments. Panels A,B) MDDCT recipients were subjected to rectal biopsy before all investigational interventions (baseline) and at the end of all treatments. Patients were stratified based on the concordance (or lack thereof, i.e. concordance for 50% or less of the epitopes administered) of predicted epitopes between the online tools NetMHCpan (http://www.cbs.dtu.dk/services/NetMHCpan/) and Custommune (www.custommune.com). The relative risk (RR) is referred to the risk of having detectable viral DNA as calculated by intention-to-treat (A) and per-treatment (B) analysis