The overexpression of programmed death-1 (PD-1) and cytotoxic T cell antigen 4 (CTLA-4) receptors on T cells are among the major mechanisms of tumor immunoevasion. However, the expression pattern of these receptors on T cell subpopulations of a different activation status and at different sites is poorly characterized. Thus, we analyzed the expression of PD-1 and CTLA-4 on the naïve, activated, memory, and activated memory T cells. Bronchoalveolar lavage fluid (BALF) from the lung affected by lung cancer (clBALF), the opposite 'healthy' lung (hlBALF), and peripheral blood (PB) samples were collected from 32 patients. The cells were analyzed by multiparameter flow cytometry. The proportion of memory, activated, and activated memory CD8+ cells with the expression of PD-1 and CTLA-4 were elevated in the clBALF when compared to the hlBALF (insignificantly), but these proportions were significantly higher in the BALF when compared with the PB. The proportions of PD-1+ and CTLA-4+ T cells were elevated in the squamous cell carcinoma when compared to the adenocarcinoma patients. Also, the expression of PD-1 and CTLA-4 on T cells from the BALF was significantly higher than from PB. We report for the first time the differential expression of checkpoint molecules on CD4+ and CD8+ lymphocytes at a different stage of activation in the local environment of lung cancer. Moreover, the circulating T cells have a distinct expression of these receptors, which suggests their poor utility as biomarkers for immunotherapy.