Background: MPM is an aggressive cancer associated with asbestos exposure. Chemotherapy has been a mainstay of therapy and recently immunotherapy has been associated with survival improvements. Chemotherapy and immunotherapy influence the tumor microenvironment and some reports have shown increased of TIL after chemotherapy in MPM. The purpose of this study was to determine dynamic changes in the tumor immune environment during mesothelioma evolution and after systemic therapies in paired samples. Methods: We examined 20 matched formalin-fixed paraffin embedded tissue samples from 10 patients (p) with MPM taken at diagnosis and at disease progression (follow-up), including untreated and treated with chemotherapy or immunotherapy. Tissue densities (number of cells/mm2) of key immune effector cells such as total lymphocytes (CD3), cytotoxic T cells (CD8), regulatory T cells (FOXP3) and monocytes/macrophages (CD163) were quantified by image analysis using multiplexed immunohistochemistry (IHC). PD-L1 expression in both tumor cells and immune cells was analysed by singleplex IHC and quantified using the CPS score. Results: Median age was 62 years (range 53-84), 7 p were male and all p presented epithelioid histology. Three matched samples belong to patients that did not receive any systemic treatment between biopsies and the other 7 pair of samples belong to patients treated with chemotherapy +/- immunotherapy. The median interval between paired biopsies was 12 m (3-33 m). Median overall survival was 27.8 months (1 m-NR). Median densities of T cells and macrophages in diagnostic and matched follow-up samples were: CD3+:982 vs 712; CD8+:422 vs 334, FOXP3+: 22 vs 25 and CD163+: 2086 vs 1706. Median PD-L1 CPS was 2.5 and 4 in diagnostic and follow-up samples, respectively. In p receiving treatment between sample pairs, the median densities in diagnostic and follow-up matched samples were: CD3+:979 vs 407; CD8+:467 vs 225, FOXP3+: 25 vs 15, CD163+: 1583 vs 1508, and PD-L1: 2 and 1. In p without any systemic treatment between sample pairs, the median densities were CD3+:1645 vs 2397; CD8+:376 vs 1605, FOXP3+: 19 vs 121, CD163+: 2102 vs 2377, and PD-L1: 3 vs 25. Conclusions: In our small series with paired samples from MPM p, we observed immune cells infiltration changes during MPM evolution with a trend towards a decrease after systemic therapy. Further investigation in tumor microenvironment in MPM to define the most appropriate time for immunotherapy is needed.