Therapeutic effect of dual CAR-T targeting PDL1 and MUC16 antigens on ovarian cancer cells in mice

Abstract

Background: More favorable treatment against epithelial ovarian cancer (EOC) is urgently needed because of its insidious nature at an early stage and a low rate of five-year survival. The current primary treatment, extensive surgery combined with chemotherapy, exhibits limited benefits for improving prognosis. Chimeric antigen receptor T (CAR-T) cell technology as novel immunotherapy has made breakthrough progress in the treatment of hematologic malignancies, and there were also benefits shown in a partial solid tumor in previous research. Therefore, CART cell technology may be a promising candidate as an immunotherapeutic tool against EOC. However, there are some weaknesses in targeting one antigen from the previous preclinical assay, such as ontarget off-tumor cytotoxicity. The dual-target CART cell may be a better choice. Methods: We constructed tandem PD1-antiMUC16 dual-CAR, PD1 single-CAR, and anti-MUC16 single-CAR fragments by PCR and genetic engineering, followed by preparing CART cells via lentiviral infection. The expression of CAR molecules on single and dual CART cells was detected by flow cytometry. The killing capacity and activation of CART cells were measured by cytotoxic assays and cytokines release assays in vitro. The therapeutic capacity of CART cells was assessed by tumor-bearing mice model assay in vivo. Results: We successfully constructed CARs lentiviral expression vectors and obtained single and dual CART cells. CART cells demonstrated robust killing capacity against OVCAR-3 cells in vitro. Meanwhile, CART cells released plenty of cytokines such as interleukin-2(IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-α(TNF-α). CART cells showed a therapeutic benefit against OVCAR-3 tumor-bearing mice and significantly prolonged the survival time. Dual CART cells were shown to be two to four times more efficacious than single CART cells in terms of survival time. Conclusion: Although exhibiting a similar ability as single CART cells against OVCAR-3 cells in vitro, dual CART cells demonstrated enhanced killing capacity against OVCAR-3 cells as compared to single CART cells in vivo and significantly prolonged the survival time of tumor-bearing mice. PD1-antiMUC16 CART cells showed more potent antitumor activity than single CART cells in vivo. The present experimental data may support further research work that will have the potential to lead to clinical studies.

ImmunoPET of Ovarian and Pancreatic Cancer with AR9.6, a Novel MUC16-Targeted Therapeutic Antibody

“…Isoforms of MUC16 have also been implicated in the progression and metastasis of pancreatic ductal adenocarcinoma as well as aggressive subtypes of the disease such as squamous and basal-like carcinomas (9,10). Despite its endogenous expression in a few healthy tissues and status as a shed antigen, MUC16 remains a viable therapeutic target for antibody-based and adoptive cell therapies (11)(12)(13)(14)(15)(16)(17)(18)(19).…”

Human epidermal growth factor receptor 2 (HER2)-specific chimeric antigen receptor (CAR) for tumor immunotherapy; recent progress

“…Preliminary studies have shown that targeting overexpressed molecules like mucin 16 (MUC16), annexin 2 (ANXA2), and also HER2 can sustain high tumor cells toxicity, and so dwindle tumor burden [ 134 ]. For instance, MUC16-redirected CAR T cells exhibited a therapeutic benefit versus human breast OVCAR-3 tumor-bearing mice and also prolonged their survival time [ 135 ]. Notwithstanding, because of the inherent heterogeneity of OC concurrently high mutation multiplicity and overexpression of diverse receptors, employing individual therapeutic strategies is largely desired [ 134 ].…”

Chimeric Antigen Receptor Design and Efficacy in Ovarian Cancer Treatment

“…To overcome ‘on-target off-tumour’ cytotoxicity, the dual-target CARs may be a better choice [ 92 ]. It has been shown that dual CARs are related to the longer survival time of mice up to two times when compared to single CAR groups and control group (80 vs. 40 days) [ 83 ].…”

“…MUC16-CAR-T cells injected intravenously or intraperitoneally are able to delay OC’s progression or altogether remove tumours in mouse tumour-bearing models. Therefore, again it seems that MUC16 is an ideal antigenic target for CAR molecules [ 83 ].…”