Recombination of a dual-CAR-modified T lymphocyte to accurately eliminate pancreatic malignancy

Abstract

BackgroundThe therapeutic application of T cells endowing with chimeric antigen receptors (CARs) is faced with “on-target, off-tumor” toxicity against solid tumors, particularly in the treatment of the pancreatic cancer. To our best knowledge, the pancreatic cancer cell line AsPC-1 often highly expressed some distinct tumor-associated antigens, such as carcino-embryonic antigen (CEA) and mesothelin (MSLN). Therefore, in this research, we have characterized dual-receptor CAR-modified T cells (dCAR-T) that exert effective and safe cytotoxicity against AsPC-1 cells.MethodsBased on the dual signaling pathway of wild T cells, we designed a novel dCAR diagram specific for CEA and MSLN, which achieved comparable activity relative to that of conventional CAR-T cells (CEA-CAR T or MSLN-CAR T). In this dCAR, a tandem construct containing two physically separate structures, CEA-CD3ζ and MSLN-4/1BB signaling domains were effectively controlled with tumor antigens CEA and MSLN, respectively. Finally, the activity of dCAR-T cells has been verified via in vitro and in vivo experiments.ResultsIn the presence of cognate tumor cells (AsPC-1) expressing both CEA and MSLN, dCAR-T cells exerted high anti-tumor activity relative to that of other single-receptor CAR-T cells bearing only one signaling pathway (e.g., Cζ-CAR and MBB-CAR). In a xenograft model, dCAR-T cells significantly inhibited the growth of AsPC-1 cells yet no effect on the growth of non-cognate tumor cells. Furthermore, the released cytokines and T cell persistence in mice were comparable with that of conventional CAR-T cells, obtaining specific and controllable cytotoxicity.ConclusionsA novel type of CAR-T cells, termed dCAR-T, was designed with specific activities, that is, significant cytotoxicity for two antigen-positive tumor cells yet no cytotoxicity for single antigen-positive tumor cells. Dual-targeted CAR-T cells can be precisely localized at the tumor site and can exert high cytotoxicity against tumor cells, alleviating “on-target, off-tumor” toxicity and enabling accurate application of CAR-T cell therapy.Electronic supplementary materialThe online version of this article (10.1186/s13045-018-0646-9) contains supplementary material, which is available to authorized users.

PD-L1 chimeric costimulatory receptor improves the efficacy of CAR-T cells for PD-L1-positive solid tumors and reduces toxicity in vivo

“…However, there have been several clinical trials involving TAA-targeting CAR-T cells within solid tumors, in which on-target off-tumor recognition of bystander tissue cells has led to life-threatening adverse effects caused by engineered T cells that attack essential normal tissues [6,7]. Therefore, many researchers have attempted to improve the speci city of CAR-T cells; thus, the logic-gated CAR [8,[12][13][14], masked CAR [9], synNotch receptor [10,28,29] and SUPA CAR [11] were developed to reduce on-target off-tumor toxicity. Among these tools, logic-gated CAR-engineered T cells are fully activated only when both a suboptimal CAR and a CCR simultaneously recognize two TAAs on tumor cells, which markedly increases speci city and thus prevents nontumor cells from attacking.…”

“…Among these tools, logic-gated CAR-engineered T cells are fully activated only when both a suboptimal CAR and a CCR simultaneously recognize two TAAs on tumor cells, which markedly increases speci city and thus prevents nontumor cells from attacking. Nevertheless, recent reports involving logic-gated CARs have shown that two different target antigens, CAR-targeted antigens and CCR-targeted antigens, must be carefully chosen to treat different solid tumors [8,[12][13][14], which is time consuming and laborious.…”

“…this system consists of a zipCAR and a leucine zipper-fused zipFv [11]. Among these strategies, the feasibility of a logic-gated CAR was con rmed in many preclinical studies, including studies of HER2 + MUC1 + breast cancer [12], PSCA + PSMA + prostate cancer [8], GPC3 + ASGR1 + hepatocellular carcinoma [13] and CEA + MSLN + pancreatic cancer [14]; however, this strategy will impede the broad application of CAR-T cells because of tumor antigen heterogeneity. Many tumor antigens [e.g., EGFR, HER2, MSLN and MUC1] identi ed in solid tumors are not broadly expressed on different types of tumors or the same tumor cells, e.g., MSLN was found in 25 ~ 30% of breast cancers, 40 ~ 45% of colon cancers and 80 ~ 85% of pancreatic cancers [15], so simultaneously targeting two TAAs expressed on the same tumor cells with current logic-gated CAR is more di cult than using a CAR targeting a single tumor antigen.…”

Characterization of novel dual tandem CD19/BCMA chimeric antigen receptor T cells to potentially treat multiple myeloma

“…CD69 is a standard marker for T-cell activation [20,21]. To demonstrate antigen-specific activation of tan-CAR-transduced T cells, cells from the K562 human leukemia line were transfected with lentiviruses encoding CD19 and/or BCMA in order to generate K562-CD19, K562-BCMA, or K562-CD19+BCMA cells.…”

Adoptive immunotherapy with CAR modified T cells in cancer current landscape and future perspectives

“…In order to overcome antigen escape risk and inter-patient variability, CARs co-targeting two or even three antigens are being developed in several solid malignancies, including breast, pancreatic and brain tumors (110)(111)(112)(113). Conversely, other investigators are testing dual constructs requiring both targets to be expressed on tumor cells in order to exert their cytotoxic activity (114) and inhibitory CARs able to redirect T cells activity from healthy tissues (115), with the aim of increasing CAR specificity and of preventing off-tumor side effects.…”