Chimeric antigen receptor T (CAR T) cell therapy is a new pillar in cancer therapeutics, and has been successfully used for the treatment of cancers, including acute lymphoblastic leukemia and solid cancers. Following immune attack, many tumors upregulate inhibitory ligands which bind to inhibitory receptors on T cells. For example, the interaction between programmed cell death protein 1 (PD-1) on activated T cells and its ligands (widely known as PD-L1) on a target tumor limits the efficacy of CAR T cells therapy against poorly responding tumors. Here, we use mesothelin (MSLN)-expressing human ovarian cancer cells (SKOV3) and human colon cancer cells (HCT116) to investigate whether PD-1-mediated T cell exhaustion affects the anti-tumor activity of MSLN-targeted CAR T cells. We utilized cell-intrinsic PD-1-targeting shRNA overexpression strategy, resulting in a significant PD-1 silencing in CAR T cells. The reduction of PD-1 expression on T cell surface strongly augmented CAR T cell cytokine production and cytotoxicity towards PD-L1-expressing cancer cells in vitro. This study indicates the enhanced anti-tumor efficacy of PD-1-silencing MSLN-targeted CAR T cells against several cancers and suggests the potential of other specific gene silencing on the immune checkpoints to enhance the CAR T cell therapies against human tumors.