In cancer treatment, programmed death-1 (PD-1) and programmed death ligand-1 (PD-L1) inhibitors are thriving. Activated T lymphocytes expressed PD-1, it works with its ligand PD-L1 to limit T lymphocyte activation and prevent autoimmune disease. The expression of molecular biomarkers and PD-L1 in lung cancer determines the appropriate treatment strategy for patients with lung cancer. The purpose of this study was to look at the prevalence of molecular biomarkers and PD-L1 expression in a large group of Tunisian patients with advanced non-small cell lung cancer. We conducted an observational retrospective study in which medical/treatment history data were extracted retrospectively from medical records and archived tissue samples between January 1st 2019 and December 31st 2021. We gathered 157 patients who had recently been diagnosed with non-small cell lung carcinoma. In 36.9%of the cases, there was no molecular genotyping. EGFR (28.6%), KRAS (5.73%), and ALK gene rearrangement were the most common genotyping mutations (3.8%). ROS1 rearrangement was not present. There was a link between EGFR and gender, HER and age, and KRAS and biopsy tissue origin. Six of the tested cases with PD-L1 met the cut-off (³50%). PD-L1 positivity was more common in solid type adenocarcinoma (1.9%) than in acinar or papillary adenocarcinoma. There were no significant differences in PD-L1 expression across clinical and demographic parameters. High PD-L1 expression and molecular abnormalities were found in 1 case of EGFR, 1 case of BRAF, and 1 case of KRAS (3 cases). All of the other specimens with abnormalities had a PD-L1 <50%. ALK, ROS1, BRAF, KRAS, and MET were found to be significantly associated with PD-L1 expression. Our study is one of the country's largest, describing a large panel of biomarkers and their clinicopathologic/histopathologic associations in Tunisian lung cancer patients. We have the same molecular profile as European patients with an EGFR mutation, which is not the most common genotype abnormality in Tunisian patients. There is only one mutation at any given time. The expression of PD-L1 is determined by the histologic type and the origin of the biopsy tissue.