Pre-clinical validation of a novel pan-cancer CAR-T cell immunotherapy against nfP2X7

Abstract

Chimeric antigen receptor (CAR)-T immunotherapy is a novel treatment that genetically modifies the patient’s own T cells to target and kill malignant cells. CAR-T cells demonstrated robust clinical activity against certain B-cell malignancies. However, identification of tumour-specific antigens expressed on multiple cancer types, especially on solid cancers, remains a major challenge. P2X purinoceptor 7 (P2X7) is an ATP gated cation channel that forms homotrimers and heterotrimers at the cell surface. When functioning normally, it controls ion transport in response to ATP. A dysfunctional version of P2X7, named nfP2X7, has been identified on cancer cells from a range of tissues, while being undetectable on healthy cells. We generated prototype nfP2X7-targeting human CAR-T cells, which demonstrated effective antigen-specific cytotoxicity against twelve solid cancer types including breast, prostate, lung, colorectal, brain and skin in vitro. In preclinical xenograft mouse models of aggressive breast and prostate cancer, CAR-T cells targeting nfP2X7 exhibited robust anti-tumour efficacy. These data indicate CAR-T cells targeting nfP2X7 have potential as a novel broad-spectrum cancer immunotherapy for solid tumours in humans.

Engineered CAR‐T cells targeting the non‐functional P2X purinoceptor 7 (P2X7) receptor as a novel treatment for ovarian cancer

“…The nfP2X7 peptide-binding domain 16,19,22 was cloned into a well characterised second-generation CAR lentiviral backbone encoding a IgG4 hinge/linker and intracellular domains from 41BB and CD3 zeta, connected by a T2A self-cleaving peptide to a truncated EGFR (EGFRt) reporter as described previously 20 (Supplementary figure 1). Previous work confirmed that the nfP2X7-CAR-T cells act specifically via the P2X7 receptor gene locus and possess on-target specificity and limited off target cytotoxicity.…”

“…Previous work confirmed that the nfP2X7-CAR-T cells act specifically via the P2X7 receptor gene locus and possess on-target specificity and limited off target cytotoxicity. 20 Deletion of the P2X7R gene in prostate cancer cell line (PC3), significantly reduced the CAR-T cell-mediated cytotoxicity, when compared with the wild-type PC3 cells. 20 Significant cytotoxicity was observed against cell lines that express nfP2X7 receptor but little cytotoxicity against cell lines that express P2X7 receptor only, or no expression of both nfP2X7 and P2X7 receptor.…”

“…20 Deletion of the P2X7R gene in prostate cancer cell line (PC3), significantly reduced the CAR-T cell-mediated cytotoxicity, when compared with the wild-type PC3 cells. 20 Significant cytotoxicity was observed against cell lines that express nfP2X7 receptor but little cytotoxicity against cell lines that express P2X7 receptor only, or no expression of both nfP2X7 and P2X7 receptor. 20 There was no in vitro cytotoxicity against normal healthy peripheral blood mononuclear cells 20 that express only wild-type P2X7 receptor.…”

“…Our previous work showed that CAR-T cells targeting nfP2X7 receptor (nfP2X7-M CAR-T cells) were highly cytotoxic in vitro against 12 different solid cancer types, including ovarian cancer. 20 In addition, significant tumour inhibition was observed against both breast and prostate cancer xenograft mouse models in vivo. 20 In this study, we show efficacy of the nfP2X7-targeting CAR-T cells against ovarian cancer cells in monolayer culture, 3D spheroid culture and in patient-derived explant assays.…”

“…20 In addition, significant tumour inhibition was observed against both breast and prostate cancer xenograft mouse models in vivo. 20 In this study, we show efficacy of the nfP2X7-targeting CAR-T cells against ovarian cancer cells in monolayer culture, 3D spheroid culture and in patient-derived explant assays. 17,21 Furthermore, nfP2X7-targeting CAR-T cells significantly reduced tumour burden in an ovarian cancer (OVCAR3) xenograft mouse model in vivo.…”