Background: Solid malignancies are heterogeneous diseases with variable clinical outcomes. Cancer mortalities are caused by the progression to metastasis and the development of treatment resistance. Recent advancements in sequencing technologies and computational capabilities have enhanced the molecular characterization of clinically actionable genetic alterations and the identification of genomic biomarkers [tumor mutational burden (TMB) and microsatellite instability (MSI)] that have therapeutic implications in solid tumors. Tissue biopsy-based Next Generation Sequencing (NGS), including testing for MSI and/or TMB, was performed to evaluate the molecular landscape of 205 prostate cancer (PC), 132 non-small cell lung cancer (NSCLC), 19 colorectal cancer (CRC) and 10 cutaneous melanoma (CM) patients, and to inform the selection of targeted therapies and immunotherapy. Additionally, liquid biopsy-based NGS bioinformatics data from 19 breast cancer (BC) patients were analyzed to inform treatment selection.
Aims: In aim 1, NGS was employed using the ForeSENTIATM tumor profile panels, which assess specific genetic alterations, to evaluate the genomic landscape of 205 PC, 132 NSCLC, 19 CRC, and 10 CM patients, giude diagnosis and prognosis, and inform targeted therapy selection. In aim 2, we expanded tissue biopsy testing by adding additional genes and implementing the expanded assay to assess MSI and TMB biomarkers, aiding in the identification of patients who could benefit from immunotherapy. In aim 3, liquid biopsy-based NGS bioinformatics data and NeothetisTM breast/gynecological panel were used to detect clinically significant SNVs/Indels in BC patients, guiding diagnosis and prognosis, and informing targeted therapy selection.
Materials and Methods: DNA was extracted from FFPE tissue samples of PC, NSCLC, CRC and CM patients, and then subjected to DNA library preparation, target capture enrichment, and NGS. Bioinformatics tools were applied for demultiplexing, alignment to the human reference genome and variant calling. Variant curation, classification and interpretation were performed using the
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Cancer Genome Interpreter (CGI) and Varsome Clinical, or CGI and Franklin by genoox interpretation platforms, in accordance with AMP guidelines. Cross-referencing was performed using the COSMIC database.
Results: The comprehensive molecular profiling of 385 cancer patients has shown clinically significant SNVs/Indels, gene amplifications and/or gene rearrangements in driver genes associated with oncogenic pathways (MAPK, PI3K/AKT, and Wnt/β-catenin), and DNA repair pathways (MMR and HRR), contributing to tumorigenesis. Our findings exhibited 6 PC and 10 CRC patients with MSI-H tumors who could benefit from the FDA-approved immunotherapy, pembrolizumab. We also detected 11 NSCLC patients with TMB-L tumors who could derive some clinical benefit from pembrolizumab, based on previous studies, despite its lack of FDA approval for this group. Pembrolizumab remains investigational for TMB-L patients and requires further research. Furthermore, we identified several patients eligible for FDA-approved targeted therapies and novel potential therapeutic targets, which need further evaluation. Importantly, we highlighted two main challenges: tumor heterogeneity and treatment resistance.
Conclusion: The application of NGS tumor profile panels, including optional MSI and/or TMB testing, enabled comprehensive molecular profiling of five cancer cohorts and the identification of patients who could benefit from targeted therapy and/or immunotherapy. Multiple clinically actionable genetic alterations were detected, highlighting the importance of NGS-based targeted and broader gene panels in the molecular characterization of solid tumors.