Integrated Clinico-Molecular Characterization Identifies RAS Activation and CDKN2A Deletion As Independent Adverse Prognostic Factors in Cancer of Unknown Primary

Background: Cancer of unknown primary (CUP) denotes a malignancy with histologically confirmed metastatic spread while the primary tumor remains elusive. Here, we address prognostic and therapeutic implications of mutations and copy number variations (CNVs) detected in tumor tissue in the context of a comprehensive clinical risk assessment. Methods: Targeted panel sequencing was performed in 252 CUP patients. 71.8% of patients had unfavorable CUP according to ESMO guidelines. 74.7% were adeno- and 13.7% squamous cell carcinomas. DNA was extracted from micro-dissected formalin-fixed, paraffin-embedded tissues. For library preparation, mostly multiplex PCR-based Ion Torrent AmpliSeqTM technology with Oncomine comprehensive assays was used. Findings: Most frequent genetic alterations were mutations/deletions of TP53 (49.6%), CDKN2A (19.0%) and NOTCH1 (14.1%) as well as oncogenic activation of KRAS (23.4%), FGFR4 (14.9%) and PIK3CA (10.7%). KRAS activation was predominantly found in adenocarcinomas (P=0.01), PIK3CA activation in squamous cell carcinomas (P=0.03). Male sex, high ECOG score, unfavorable CUP, higher number of involved organs and RAS activation predicted decreased event-free and overall survival in multivariate analysis. Deletions of CDKN2A were prognostically adverse regarding overall survival. TP53 mutations did not significantly influence prognosis in the overall cohort, but worsened prognosis in otherwise favorable CUP subtypes. Although not standard in CUP, for 17/198 (8.6%) patients molecularly targeted treatment was recommended and 10 patients (5.1%) were treated accordingly. Interpretation: Besides the identification of drug targets, panel sequencing in CUP is prognostically relevant, with RAS activation and CDKN2A deletion emerging as novel independent risk factors in a comprehensive assessment with clinico-pathological data. Funding: None. Declaration of Interest: TB and AK work as study oncologists for the CUPISCO trial, which is sponsored by Roche, and have received reimbursement for study‐related travels as well as remuneration for their work as study oncologists for the benefit of their employer. VE: advisory board and lecture fees from AstraZeneca and ThermoFisher. ALV: lecture fees from AstraZeneca. PS: advisory board honoraria from Pfizer, Roche, Novartis and AstraZeneca as well as speaker's honoraria and research funding from Roche, AstraZeneca and Novartis. WW: advisory board honoraria from MSD, BMS, Roche, AZ, Novartis, Celgene, Merck, Pfizer; speaker's honoraria from MSD, BMS, Roche, AZ, Boehringer, Lilly, Amgen, Takeda, Novartis; research funding (to institution): Roche, MSD, BMS. AS: advisory board honoraria from Bayer, Novartis, BMS, AstraZeneca, ThermoFisher, Illumina; speaker's honoraria from Takeda, Roche, BMS, Illumina, AstraZeneca, Novartis, ThermoFisher, Bayer, MSD and research funding from Chugai and BMS. All remaining authors have declared no conflicts of interest. Ethical Approval: Ethics board approval from the University of Heidelberg was obtained and patients signed informed consent.