Sustained HIV remission after allogeneic hematopoietic stem cell transplantation with wild-type CCR5 donor cells

Abstract

HIV cure has been reported for five individuals who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) with cells from CCR5Δ32 homozygous donors. By contrast, viral rebound has occurred in other people living with HIV who interrupted antiretroviral treatment after undergoing allo-HSCT, with cells mostly from wild-type CCR5 donors. Here we report the case of a male individual who has achieved durable HIV remission following allo-HSCT with cells from an unrelated HLA-matched (9 of 10 matching for HLA-A, HLA-B, HLA-C, HLA-DRB1 and HLA-DQB1 alleles) wild-type CCR5 donor to treat an extramedullary myeloid tumor. To date, plasma viral load has remained undetectable for 32 months after the interruption of antiretroviral treatment. Treatment with ruxolitinib has been maintained during this period to treat chronic graft-versus-host disease. Low levels of proviral DNA were detected sporadically after allo-HSCT, including defective but not intact HIV DNA. No virus could be amplified in cultures of CD4⁺ T cells obtained after antiretroviral treatment interruption, while CD4⁺ T cells remained susceptible to HIV-1 infection in vitro. Declines in HIV antibodies and undetectable HIV-specific T cell responses further corroborate the absence of viral rebound after antiretroviral treatment interruption. These results suggest that HIV remission could be achieved in the context of allo-HSCT with wild-type CCR5.

Fig. 1. Treatment course and immuno-virological evolution.

Timeline of clinical events, CD4⁺ T cell counts, CD4/CD8 ratio and plasma viral load since HIV-1 diagnosis and until last follow-up. The successive immunosuppressants and antiretroviral regimens are also shown (detailed information can be found in Supplementary Fig. 1). Undetectable plasma viral load concentrations are represented by empty symbols at the threshold of detection, which varied throughout the follow-up. Hexagons represent different episodes of GvHD.

Fig. 2. Viral markers before and after allo-HSCT.

a,b, Evolution of residual low-level viremia measured with an ultrasensitive viral load (VL) assay in plasma (a) and HIV DNA associated with PBMCs, purified blood CD4⁺ T cells or bone marrow (BM) cells (b) before and after allogeneic HSCT. The empty symbols represent undetectable levels and are shown at the threshold of the techniques, which varied depending on the amount of material analyzed. The time of allo-HSCT and of analytical treatment interruption (ATI) is indicated as a dashed vertical line. c, Summary of the outcome of IPDA analyses in blood and tissues at different timepoints before and after allo-HSCT. The number of cells per amount of material tested is indicated for each analysis. ^aDefective in 3′ or 5′. ^bDefective in 3′.

Fig. 3. Phenotype and HIV susceptibility of CD4⁺ T cells.

a,b, Percentage of CD4⁺ T cells from IciS-34 expressing activation markers (CD38⁺HLA-DR⁺, Ki67⁺) (a) and the HIV-1 coreceptors CCR5 and CXCR4 (b) at different times after allo-HSCT (samples designated as the month after allo-HSCT when they were obtained; purple). The proportions of CD4⁺ T cells from an independent HIV-negative (HIVneg) blood donor (gray) and one person with HIV on ART (black) are depicted for reference. c, Dynamics of viral replication in purified CD4⁺ T cells from IciS-34 (M59, purple) and one unrelated blood donor upon infection in vitro with HIV-1_BaL. The data are shown as the mean ± s.d. (n = 3 replicates) of p24 levels in culture supernatants. d, Proportion of infected (GFP⁺) CD4⁺ T cells from IciS-34 3 days after challenge with HIV_NL4.3GFPΔEnv-VSV-G particles (purple). The negative control is depicted as a dashed line. The rate of infected CD4⁺ T cells from one unrelated blood donor is provided as a reference (gray).

Fig. 4. Antibody response after ART interruption.

a, Results of immunoblots for HIV-1 antibodies in plasma samples from IciS-34 at different times before and after allo-HSCT. The arrows indicate the antigens against which reactivity was progressively lost. On the right are indicated the reactivity ratings that were automatically attributed for each antigen on the strips. b, ELISA graphs (left) comparing the reactivity of purified plasma IgG antibodies from IciS-34 (values for samples obtained at three different timepoints (M53, M55, M65) are depicted) and early- and late-treated PLWH (eART (n = 6) and lART (n = 6), respectively) against HIV-1 p24 and Env proteins. HIV-1-seronegative (Ctr−) sera (n = 5) were used as negative controls. Dot plots comparing the area under the curve (AUC) values calculated from the titration curves are shown on the right (horizontal lines indicate the median values). c, Heatmap showing the ELISA binding analysis of purified serum IgG antibodies from IciS-34 at two timepoints against consensus subtype B overlapping Env peptides. Darker colors indicate higher reactivity (absorbance values); white, no binding. Sera from eART (CO108), lART (CO107), post-treatment controller (PTC005002) and elite controller (pt3) individuals were used as positive controls. d, Heatmap comparing the in vitro neutralizing activity (as percentages) of purified serum IgG antibodies from IciS-34 (3 timepoints), CO108, CO107, PTC005002 and pt3 against selected clade B tier 1 and 2 viruses as measured in the TZM-bl assay. e, Heatmap comparing the percentage of CEM.NKR-CCR5 cells infected by laboratory-adapted (YU2 and AD8) and transmitted/founder viruses (CH058, REJO and THRO) bound by purified serum IgG antibodies from IciS-34 (2 timepoints), eART (CO42 and CO108), lART (CO105 and CO107), PTC005002 and pt3. HIV-1-seronegative (Ctr−) serum was used as negative control (c,e).

Fig. 5. T cell reactivity and HIV-specific T cell response.

a, Percentage of CD8⁺ T cells from IciS-34 (M45) and an HIV-negative donor producing IFNγ and TNF after 6 h of stimulation with PMA/ionomycin, or with pools of HCMV pp65 peptides and HIV-1 Gag, Nef and Pol peptides. Unstimulated (Unstim) cells were used as control. Results are depicted as standard pseudocolor dot plots. b, Percentage of CD8⁺ T cells from IciS-34 (M45) and an HIV-negative donor that proliferated (CFSE^low) after 6 days of stimulation with anti-CD3/CD28, pools of HCMV pp65 peptides, and HIV-1 Gag, Nef and Pol peptides and were able to produce IFNγ upon short polyclonal or antigen-specific restimulation. c, Heatmap comparing the percentage of CD8⁺ T cells from IciS-34 (two samples) and two HIV-negative donors that produced IFNγ and/or TNF and IFNγ and/or expressed CD107 after 6 h or 6 days (d) of polyclonal (positive control, Pos) or antigen-specific stimulation. Darker colors indicate higher percentages. White, undetectable; gray, not done. The percentages are indicated after subtraction of the background from the unstimulated condition. d, Comparison of the level of infection of CD4⁺ T cells from IciS-34 (M45) infected in vitro with HIV-1_BaL cultured alone or in the presence of autologous CD8⁺ T cells (1:1 ratio). The results are shown as the levels of p24 in culture supernatants at day 7 after infection in vitro (mean ± s.d. of 3 experiments). Similar results were obtained at M54, M56, M57 and M59. e, Percentage of CD8⁺ T cells from an HIV-negative donor and IciS-34 at time of ruxolitinib discontinuation (M51), 15 days after ruxolitinib discontinuation (M52-1), 4 weeks after ruxolitinib discontinuation, time of reinitiation (M52-2) and 15 days after ruxolitinib reinitiation (M53), producing IFNγ 6 h after polyclonal stimulation with PMA/ionomycin (top). Heatmap comparing the percentages of CD4⁺ and CD8⁺ T cells from the HIV-negative donor and IciS-34 at different timepoints that produced IFNγ and/or TNF and IFNγ and/or expressed CD107 after 6 h of polyclonal or antigen-specific stimulation.

Fig. 6. Phenotype and antiviral activity of NK cells.

a, Expression of CD16 and CD56 on NK cells from one person with HIV on ART and IciS-34 (M52-2). b, Proportion of CD16⁻CD56⁺⁺, CD16⁺CD56⁺ and CD16⁺CD56⁻ NK cells in the sample from the person on ART and from IciS-34 at six different timepoints (left). Expression of KIR2DL2/3 and/or KIR3DL1/S1 and CD69 and/or CD57 in the three NK cell subsets from IciS-34 (M52-2) (right). c, Expression of CD57 on NK cell subsets from IciS-34 (M52-2) defined on the absence or expression of different combinations of KIRs. MFI, median fluorescence intensity. d, Comparison of the level of infection of CD4⁺ T cells from IciS-34 (M59) infected in vitro with HIV-1_BaL cultured alone or in the presence of autologous NK cells (1:1 and 1:3 ratios). The results are shown as the levels of p24 in culture supernatants at day 3 after infection in vitro (mean ± s.d. of three experiments). Similar results were obtained at M54.