Background: Next generation sequencing (NGS) can be used to detect tumor-specific genomic alterations, with the aim to identify actionable genomic alteration and provide additional treatment options for patients with advanced cancer. While smaller sequencing panels are being applied broadly, controversy exists around the routine implementation of comprehensive genomic profiling with large multigene NGS panels. Indeed, there is an ongoing discussion about the indications as well as timing of comprehensive genomic profiling in patients with advanced solid tumors. Therefore, there is a need for more data to increase understanding of the potential indications and added clinical benefit of routine implementation of broad comprehensive genomic profiling. However, available studies have often been reporting on a small patient population, which complicates the interpretation and translation to the clinical setting. This study aims to describe the feasibility of performing broad NGS panels in patients with advanced cancer in an academic hospital setting, as well as the targetable alterations detected. Methods: In this retrospective study, we analysed genomic alterations in solid tumor biopsies of 464 patients with locally advanced or metastatic cancer. Next generation sequencing was performed using the Foundation Medicine assay (FoundationOneCDx). The therapeutic implications of the genomic alterations were determined using the OncoKB classification. Results: The FoundationOne CDx was successfully applied in 464/521 patients (89%) with a range of cancer types. The most common altered genes were TP53 (61%), followed by KRAS (20%), CDKN2A (20%), TERT (16%), APC (16%), PTEN (14%), PIK3CA (14%) and CDKN2B (14%). Among 419 patients with successfully analyzed tumor mutational burden (TMB), the median TMB was 2.52 (range 0 – 67) mutations/Mb (mut/Mb). Forty-three patients presented with a high-TMB ( > = 10 mut/Mb) of which eight had microsatellite instability. Out of 126 patients with an actionable target (e.g. ERBB2, KRAS, PIK3CA, FGFR2), 40 patients received matched treatment (8.6%), of which 17 were included in clinical trials. Conclusions: This single-center retrospective study shows that broad application of NGS is feasible in an academic center, increases the detection of actionable genomic alterations, and opens opportunities to provide treatment with immune checkpoint inhibitors in case of high tumor mutational burden and microsatellite instability status regardless of tumor histology. The study brings to attention that strategies such as inclusion in clinical (basket) trials and development of new targeted drugs are necessary to improve matched treatment rates. Further research is needed to assess the possible clinical benefit of broad application of NGS and genotype matched treatment.