Abstract
We have previously shown the potential for dramatic improvement in overall survival in immunodeficient mice bearing patient-derived xenograft (PDX) models of primary and temozolomide (TMZ)-resistant GBM when treated using a combination of intracranial therapy with ex-vivo expanded/activated MGMT-engineered γδT cells and simultaneous systemic TMZ. We have termed this approach Drug-Resistant Immunotherapy (DRI). TMZ upregulates γδT cell stress-antigen targets (NKG2DL) on primary and TMZ-resistant tumors making them more visible to effector γδT cells. In the present study, we sought to determine whether checkpoint inhibition would potentiate the effect of DRI in our PDX model system for glioma. Expanded/activated δT cells were evaluated for lysis of target cells K562 and disaggregated PDXT cells in the presence or absence of checkpoint inhibitors of PD-1, CTLA-4 and PD-L1 using a flow cytometric based killing assay. Cytotoxicity was increased by anti-CTLA-4 and anti-PD-L1 against JX22T PDXT, although a much more noticeable effect of blockade with anti-PD-1, anti-CTLA-4 and anti-PD-L1 was seen against K562. Anti-PD-1 combined with anti-CTLA-4 also showed a synergistic effect on JX22T and K562. Concurrently, we also demonstrated that although TMZ did not influence the already low expression of PD-L1 in disaggregated PDXT lines exposed to 200µM TMZ, immunohistochemical analysis of tumors from mice injected with 60mg/kg TMZ and examined 4h later showed upregulation of NKG2DL accompanied by modest upregulation of PD-L1. At the time of this writing, mice bearing JX12P and JX12T were still under examination. These early studies show that checkpoint inhibition can potentiate the cytotoxic activity of expanded/activated γδT cells. Also, TMZ may increase both the expression of NKG2DL and PD-L1 in some tumors. Taken together, our findings justify further exploration of combination DRI and checkpoint inhibition either by systemic administration, as part of the therapeutic graft, or both.