Robust CAR-T memory formation and function via hematopoietic stem cell delivery

Abstract

Due to the durability and persistence of reservoirs of HIV-1-infected cells, combination antiretroviral therapy (ART) is insufficient in eradicating infection. Achieving HIV-1 cure or sustained remission without ART treatment will require the enhanced and persistent effective antiviral immune responses. Chimeric Antigen Receptor (CAR) T-cells have emerged as a powerful immunotherapy and show promise in treating HIV-1 infection. Persistence, trafficking, and maintenance of function remain to be a challenge in many of these approaches, which are based on peripheral T cell modification. To overcome many of these issues, we have previously demonstrated successful long-term engraftment and production of anti-HIV CAR T cells in modified hematopoietic stem cells (HSCs) in vivo. Here we report the development and in vivo testing of second generation CD4-based CARs (CD4CAR) against HIV-1 infection using a HSCs-based approach. We found that a modified, truncated CD4-based CAR (D1D2CAR) allows better CAR-T cell differentiation from gene modified HSCs, and maintains similar CTL activity as compared to the full length CD4-based CAR. In addition, D1D2CAR does not mediate HIV infection or stimulation mediated by IL-16, suggesting lower risk of off-target effects. Interestingly, stimulatory domains of 4-1BB but not CD28 allowed successful hematopoietic differentiation and improved anti-viral function of CAR T cells from CAR modified HSCs. Addition of 4-1BB to CD4 based CARs led to faster suppression of viremia during early untreated HIV-1 infection. D1D2CAR 4-1BB mice had faster viral suppression in combination with ART and better persistence of CAR T cells during ART. In summary, our data indicate that the D1D2CAR-41BB is a superior CAR, showing better HSC differentiation, viral suppression and persistence, and less deleterious functions compared to the original CD4CAR, and should continue to be pursued as a candidate for clinical study.

Allogeneic gene-edited HIV-specific CAR-T cells secreting PD-1 blocking scFv enhance anti-HIV activity in vitro

“…In comparison with UTD cells, there was no significant difference among the CD62L expression of 3B, 3BE and 3BE/TCR-CAR-T cells (figure 5A). The data showed that the CD62L+ cells in all the groups of anti-HIV CAR-T cells we manufactured were over 60%, which indicated that our CAR-T cells were mainly composed of naive and central memory cells, a phenotype associated with greater in vivo anti-HIV activity 35,50 (figure 5A). The expression of CD45RA was also tested and the percentages of CD45RA+ cells were lower than 7% in all the anti-HIV CAR-T cells (figure 5A).…”

Preclinical Evaluation of CAR T Cell Function: In Vitro and In Vivo Models

“…CD19-CAR T cell infusion in this model resulted in tumor regression, together with induction of TAM-secreted cytokines and Treg activity, which resembles clinical observations [ 158 ]. A similar BLT-HM model without introduction of oncogene was used to evaluate the antiviral function of HIV-targeted CAR T cells [ 159 ]. Additionally, CARs targeting one, two, or three HIV epitopes were examined in a PBL-HM model, revealing that multi-specific CARs are required for anti-HIV therapy [ 160 ].…”

Cancer-Homing CAR-T Cells and Endogenous Immune Population Dynamics

“…This may allow creating a more favorable environment for CAR-T cells and longer CAR-T cell persistence and efficacy [ 97 ]. This may favor CAR-T cells acquisition of memory cell’s function [ 98 , 99 ], which may provide long-term immune recognition of cancer [ 100 ] ( Figure 5 ).…”

“…The interaction of CAR-T cells with immunoregulatory environments has been shown by CAR-T memory formation and function via hematopoietic stem cells (HSC) [ 98 ], which were shown to be common in multiple cancer types and appeared supported by CAF-originated cytokines [ 136 ]. Tumor-associated myelopoiesis was recently demonstrated, and this was shown to generate immune-suppressive myeloid populations [ 164 ].…”