CD40 costimulation enhances CAR-T cell activation

The clinical use of chimeric antigenic receptor (CAR)-T cells yields impressive clinical responses in the treatment of B cell leukemias and lymphomas. Yet, clinical benefit in other types of cancer, especially solid tumors, is limited, underscoring the need in improving the functional potency of CAR-T cells. In attempt to overcome these limitations, great effort is put in recent years into optimizing the signaling moieties incorporated into the intracellular portion of 2nd and 3rd generation CARs, which largely govern the clinical outcome of CAR-T cell therapy.

Among the different costimulatory signaling elements capable of augmenting T cell function and persistence explored so far, the most widely used are undoubtedly CD28 and 4-1BB. In a number of publications we have recently reported that different derivatives of the signaling domain of CD40, a member of the TNF receptor family mainly expressed by professional antigen-presenting cells, exert remarkable phenotypic and functional enhancing effects in human T cells. We hypothesized that the incorporation of the CD40 signaling domain as a costimulatory element in 2nd and 3rd CARs can similarly improve the functional properties of antitumor CAR-T cells.

To test our hypothesis we have constructed a series of new anti-HLA-A2 CARs harboring either the CD40 or the 4-1BB element, with or without CD28, and tested these in mRNA-electroporated human CD8 T cells. Indeed, CD40 consistently and reproducibly exhibited marked superiority over 4-1BB in upregulating costimulatory markers and activating the NFkB pathway and in antigen-specific induction of proinflammatory cytokines secretion, and was equally effective in mediating direct target cell killing.

We conclude that new CARs harboring the CD40 signaling element can endow gene-modified T cells with superior functionality compared to current CARs and potentially improve the clinical efficacy of CAR T cell therapy.