Promoter usage regulating the surface density of CAR molecules may modulate the kinetics of CAR-T cells in vivo

Abstract

Although chimeric antigen receptor T (CAR-T) cell therapy achieves high remission rates, challenges (e.g., toxicity management and relapse prevention) remain. The major risks are cytokine release syndrome and related neurological toxicity. The influence of the CAR surface density on the efficacy/safety of CAR-T cell therapy and the factors determining CAR density were not elucidated comprehensively. Here, we discovered that the use of the MND promoter increased the transduction rate and reduced the CAR surface density. Additionally, MND-driven CAR-T cells had prolonged antileukemia activity in a mouse model. In an initial dual-armed anti-CD19 CAR-T cell pilot study (ClinicalTrials.gov: NCT03840317 ), eight and six subjects were infused with MND and EF1α promoter-driven autologous CAR-T cells (3 × 10 5 CAR-T cells/kg), respectively. MND subjects developed mild fever and lower interferon gamma (IFN-γ) concentrations than in the EF1A19 group. All but one subject in each cohort reached minimal residual disease (MRD)-negative complete remission after the first month of evaluation. These results represent the first comprehensive study on the promoter-driven modulation of CAR-T cell functionality. These findings encourage further evaluation of the potential of the MND promoter to drive CAR-T cells as a broadly applicable cellular product for anticancer immunotherapy.

CS1 CAR-T targeting the distal domain of CS1 (SLAMF7) shows efficacy in high tumor burden myeloma model despite fratricide of CD8+CS1 expressing CAR-T cells

“…In all of the studies presented so far, CAR expression was driven by the EF1α promoter since it drives efficient CAR expression in human T-cells [ 21 ]. However, VIVEBiotech found viral titers were seven times higher with MND-containing promoters compared to EF1α (not shown).This phenomenon has recently been reported by a separate group [ 22 ]. Since MND is being used clinically in an effective FDA approved product (BCMA-CAR-T, idecabtagene vicleucel [ 23 ]) and higher titer virus would enable treatment of more patients, we sought to assess efficacy of MND-Luc90-BBz CAR-T (MND-Luc90-CAR-T); (Fig.…”

CAR Density Influences Antitumoral Efficacy of BCMA CAR T cells and Correlates with Clinical Outcome

“…These promoters induce high levels of CAR expression that can lead to tonic signaling and premature exhaustion. The possibility of reducing gene expression via weaker promoters as well as through more physiological promoters has been associated with decreased tonic signaling (22,23,52). In fact, the use of the MND promoter has been shown to reduce CAR surface density, while EF1a promoter increased its density, leading to a higher cytotoxic activity, cytokine production and expression of exhaustion markers (23).…”

“…The possibility of reducing gene expression via weaker promoters as well as through more physiological promoters has been associated with decreased tonic signaling (22,23,52). In fact, the use of the MND promoter has been shown to reduce CAR surface density, while EF1a promoter increased its density, leading to a higher cytotoxic activity, cytokine production and expression of exhaustion markers (23). Thus, the use of physiological promoters would be a strategy to prevent an increase in CAR High T cells, thus limiting exhaustion of CAR-T and favoring long term persistence and antitumor efficacy (22).…”

“…For instance, recent studies have demonstrated that a more physiological expression of the CAR, by the integration of the transgene in the TRAC locus or the use of different promoters, can be associated with improved efficacy and reduced toxicity (21)(22)(23). These results also suggest that the density of CAR molecule in the membrane of CAR T cells might influence CAR signaling, affecting their antitumoral efficacy (23,24). However, this hypothesis and the impact on clinical efficacy has not been formally explored.…”

Programmable Mixed-Signal Biocomputers in Mammalian Cells

“…An additional benefit of the CREATE platform is its use of recombinase logic to drive such analog outputs, creating a memory device in cells. This type of memory response would be useful for differentiating cells toward a particular phenotype defined by differential expression of the same protein, such as varying FOXP3 expression in naïve and regulatory T cell subsets [41] [42] [43] [44] or adjusting chimeric antigen receptor expression to match different antigen density profiles [45] [46].…”