The study investigates the impact of chimeric antigen receptor (CAR) affinity on the sensitivity of CAR-T cells to PD-1/PD-L1-mediated inhibition, a key factor in T-cell suppression within solid tumors. Using preclinical models, the researchers found that low-affinity (LA) CAR-T cells were more sensitive to PD-L1-mediated inhibition compared to high-affinity (HA) CAR-T cells. CRISPR/Cas9-mediated PD-1 knockout (KO) enhanced the cytokine secretion and polyfunctionality of LA CAR-T cells in vitro and improved antitumor effects in vivo, while having no significant impact on HA CAR-T cells. This effect was observed across different co-stimulatory domains (CD28, ICOS, and 4-1BB), but 4-1BB co-stimulated CAR-T cells were less sensitive to PD-L1 inhibition regardless of affinity. The study also explored the role of target antigen density and CAR expression levels, finding that HA CAR-T cells became more susceptible to PD-L1-mediated inhibition at low antigen densities. Additionally, the findings suggest that PD-1 KO may not increase the antitumor efficacy of HA CAR-T cells co-stimulated with 4-1BB. The study highlights the importance of CAR affinity in modulating the sensitivity of CAR-T cells to PD-1/PD-L1-mediated inhibition, providing insights for the design and optimization of CAR-T therapies for solid tumors.The study investigates the impact of chimeric antigen receptor (CAR) affinity on the sensitivity of CAR-T cells to PD-1/PD-L1-mediated inhibition, a key factor in T-cell suppression within solid tumors. Using preclinical models, the researchers found that low-affinity (LA) CAR-T cells were more sensitive to PD-L1-mediated inhibition compared to high-affinity (HA) CAR-T cells. CRISPR/Cas9-mediated PD-1 knockout (KO) enhanced the cytokine secretion and polyfunctionality of LA CAR-T cells in vitro and improved antitumor effects in vivo, while having no significant impact on HA CAR-T cells. This effect was observed across different co-stimulatory domains (CD28, ICOS, and 4-1BB), but 4-1BB co-stimulated CAR-T cells were less sensitive to PD-L1 inhibition regardless of affinity. The study also explored the role of target antigen density and CAR expression levels, finding that HA CAR-T cells became more susceptible to PD-L1-mediated inhibition at low antigen densities. Additionally, the findings suggest that PD-1 KO may not increase the antitumor efficacy of HA CAR-T cells co-stimulated with 4-1BB. The study highlights the importance of CAR affinity in modulating the sensitivity of CAR-T cells to PD-1/PD-L1-mediated inhibition, providing insights for the design and optimization of CAR-T therapies for solid tumors.