A quadrivalent in vivo CAR-macrophage serve as cell vaccine to repress solid tumors and overcome tumor heterogeneity through antigen spreading

We designed a quadrivalent in vivo CAR-engineered macrophage for treating solid tumors by the LNP-mRNA system. Mechanically, we found that in vivo engineered CAR macrophages can promote the formation of T-cell immune memory against neoantigens through antigen spreading. To further elucidate this phenomenon, we treated orthotopic HCC model mice with PBS, LNP-GPC3 CAR, or LNP-GPC3 CAR-Super IL-2 and subsequently collected splenocytes from these three groups for transcriptomic and TCR sequencing analyses. Based on data from transcriptome and TCR sequencing, we propose a two-signal model for T-cell activation: the GPC3 CAR provides the primary signal by recognizing and presenting tumor antigens, while the secreted Super IL-2 supplies the additional signal necessary for robust T-cell clonal expansion. These findings provide novel mechanistic insights into the anti-tumor activity of in vivo engineered CAR macrophages.