Anti-PD1 efficacy in metastatic urothelial cancer patients associates with intra-tumoral juxtaposition of T helper-type 1 and CD8+ T-cells.

Purpose: PD1 inhibition results in durable anti-tumor responses in a proportion of metastatic urothelial cancer (mUC) patients. The majority of patients, however, does not experience clinical benefit. In this study, we aimed to identify early changes in T-cell subsets that underlie anti-PD1 efficacy in mUC patients. Experimental design: Paired samples were collected from peripheral blood, plasma, and metastatic lesions of 56 mUC patients at baseline and weeks 6 and 12 after initiating pembrolizumab treatment (200mg intravenously, q3w). Samples were analyzed using multiplex flow cytometry, ELISA and in situ stainings, including cellular network analysis. Treatment response was evaluated as best overall response according to RECIST v1.1, and patients were classified as responder (complete or partial response) or non-responder (progressive disease). Results: In responders, baseline fractions of CD4+ T-cells expressing co-signaling receptors were higher compared to non-responders. The fraction of circulating PD1+ CD4+ T-cells decreased at week 6 and 12, whereas the fraction of 4-1BB+ CD28+ CD4+ T-cells increased at week 12. In metastatic lesions of responders, the baseline density of T helper-type 1 (Th1) cells, defined as T-bet+ CD4+ T-cells, was higher as compared to non-responders. Upon treatment, Th1 cells became localized in close proximity to CD8+ T-cells, CD11b+ myeloid cells, and tumor cells. Conclusions: A decrease in the fraction of circulating PD1+ CD4+ T-cells, and juxtaposition of Th1, CD8+, and myeloid cells was associated with response to anti-PD1 treatment in mUC patients.