Cancer immunotherapy, including immune checkpoint blocking antibodies are important components for treatment of patients with various types of cancer as they enhance the ability of the immune system to fight tumours. However, tumor cells have the ability to develop resistance to a variety of transitional therapies such as chemotherapy. In this study, in vitro Tumour-Immune co-culture system (TICS) has been developed to evaluate the impact on the antitumor activity of the primary human lymphocytes and response to PD-1 (nivolumab) and PD-L1 (durvalumab) checkpoint blocking antibodies against acquired chemotherapy resistance cancer cell lines. Using paired ovarian and neuroblastoma cancer cell lines obtained prior to chemotherapy (naïve) and after chemotherapy resistance, the results show that resistant ovarian cancer cells have differential effect on activation of lymphocytes and respond poorly to nivolumab and durvalumab, compared to chemotherapy naïve cells. On the other hand, chemotherapy neuroblastoma resistance cells show to respond to PD-1/L1 blockade therapy in TICS. Furthermore, blocking important molecular interactions between cancer cells and human lymphocytes such as HLA-ABC, HLA-DR and IFN-γ receptor compromises response to immune checkpoint blockade. In accordance, deletion of programmed death ligand 1 (PD-L1) on cancer cells by the CRISPR/Cas9 system significantly increases antitumor activity of immune cells in TICS. Moreover, deletion of beta-2-microglobulin (B2M) on human cancer cells resulted in substantial downregulation of HLA-ABC, which influenced immune activation induced by PD-1 blockade. Together, these findings demonstrate that chemotherapy resistance in human cancer cells could limit efficient response of PD-1/L1 blockade and thus immune checkpoint therapy could be more effective in early stage cancers.