Background: Lung cancer is a highly heterogeneous disease in which molecular characte-rization has become essential for guiding personalized therapies. The implementation of next-generation sequencing (NGS) allows the simultaneous detection of multiple genomic alterations, improving tumor profiling and therapeutic decision-making. This study aimed to characterize the molecular landscape of lung cancer using NGS and to evaluate its as-sociation with histological subtypes and programmed death-ligand 1 (PD-L1) expression. Methods: A retrospective observational study was conducted on 96 patients diagnosed with lung cancer between 2023 and 2025. Molecular profiling was performed using the Action OncoKitDx panel. Associations between genetic alterations, histological subtypes, and PD-L1 expression were analyzed using Fisher’s exact test, with p < 0.05 considered statistically significant. Results: Adenocarcinoma was the most common histological subtype (67.7%), followed by squamous cell carcinoma (26%). The most common mutations were KRAS (34.4%), TP53 (29.2%), and EGFR (11.5%). KRAS mutations were significantly associated with adenocar-cinoma (p = 0.001), while the absence of detectable mutations was associated with squa-mous cell carcinoma (p = 0.002). Co-mutations were identified in 22.9% of cases, with KRAS–TP53 being the most common combination. Tumors harboring EGFR mutations showed a significantly lower frequency of co-mutations (p = 0.012). No significant asso-ciations were found between PD-L1 expression and either histological subtypes or the analyzed genetic alterations. Conclusions: Lung cancer exhibits marked molecular heterogeneity, with a predominance of KRAS mutations in adenocarcinoma. The low frequency of co-mutations in EGFR-mutated tumors supports their role as dominant driver alterations. The lack of asso-ciation between PD-L1 expression and genomic alterations highlights the complexity of its regulation and suggests the involvement of multiple biological factors. These findings reinforce the clinical value of NGS in comprehensive tumor profiling and in the develop-ment of precision medicine strategies.