Purpose: In locally advanced p16+ oropharyngeal squamous cell carcinoma (OPSCC), (i) to investigate kinetics of human papillomavirus (HPV) circulating tumor DNA (ctDNA) and association with tumor progression after chemoradiation, and (ii) to compare the predictive value of ctDNA to imaging biomarkers of MRI and FDG-PET.Experimental Design: Serial blood samples were collected from patients with AJCC8 stage III OPSCC (n = 34) enrolled on a randomized trial: pretreatment; during chemoradiation at weeks 2, 4, and 7; and posttreatment. All patients also had dynamic-contrast-enhanced and diffusion-weighted MRI, as well as FDG-PET scans pre-chemoradiation and week 2 during chemoradiation. ctDNA values were analyzed for prediction of freedom from progression (FFP), and correlations with aggressive tumor subvolumes with low blood volume (TVLBV) and low apparent diffusion coefficient (TVLADC), and metabolic tumor volume (MTV) using Cox proportional hazards model and Spearman rank correlation.Results: Low pretreatment ctDNA and an early increase in ctDNA at week 2 compared with baseline were significantly associated with superior FFP (P < 0.02 and P < 0.05, respectively). At week 4 or 7, neither ctDNA counts nor clearance were significantly predictive of progression (P = 0.8). Pretreatment ctDNA values were significantly correlated with nodal TVLBV, TVLADC, and MTV pre-chemoradiation (P < 0.03), while the ctDNA values at week 2 were correlated with these imaging metrics in primary tumor. Multivariate analysis showed that ctDNA and the imaging metrics performed comparably to predict FFP.Conclusions: Early ctDNA kinetics during definitive chemoradiation may predict therapy response in stage III OPSCC.

Mucoepidermoid Carcinomas (MEC) represent the most common malignancies of salivary glands. Approximately 50% of all MEC cases are known to harbor CRTC1/3-MAML2 gene fusions, but the additional molecular drivers remain largely uncharacterized. Here, we sought to resolve controversy around the role of human papillomavirus (HPV) as a potential driver of mucoepidermoid carcinoma. Bioinformatics analysis was performed on 48 MEC transcriptomes. Subsequent targeted capture DNA sequencing was used to annotate HPV content and integration status in the host genome. HPV of any type was only identified in 1/48 (2%) of the MEC transcriptomes analyzed. Importantly, the one HPV16+ tumor expressed high levels of p16, had high expression of HPV16 oncogenes E6 and E7, and displayed a complex integration pattern that included breakpoints into 13 host genes including PIK3AP1, HIPI, OLFM4,SIRT1, ARAP2, TMEM161B-AS1, and EPS15L1 as well as 9 non-genic regions. In this cohort, HPV is a rare driver of MEC but may have a substantial etiologic role in cases that harbor the virus. Genetic mechanisms of host genome integration are similar to those observed in other head and neck cancers.

Merkel cell carcinoma (MCC) is an aggressive disease with poor survival outcomes and increasing incidence. There is a clear and present need for enhanced understanding of cellular mechanisms of tumorigenesis, validation of robust genetic signatures predictive of aggressive disease, and novel informatics tools to simplify analysis of Merkel cell polyomavirus (MCPyV)–host genome interactions. Genomic DNA was harvested from 54 MCC tumors for exome sequencing and in-depth genetic profiling of a 226-gene panel. We further developed a robust informatics package (MCPyViewer) optimized for MCPyV integration site analysis with graphical output to simplify usability for end users. Finally, we assessed the prognostic impact of specific genetic signatures on MCC-specific survival in our cohort. Our study included 54 patients (n = 44 MCPyV positive), 11 (20.4%) of whom had died of MCC at last follow-up. Human genes altered at high frequency included LRP1B (n = 10, 18.5%), FAT1 (n = 9, 16.7%), KMT2D (n = 9, 16.7%), and RB1 (n = 7, 13.0%). In 36 of 44 (81.8%) MCPyV-positive tumors, we identified viral integration into the human genome with a median of two events per tumor. In six tumors, MCPyV integrated into Catalogue of Somatic Mutations in Cancer tier 1 or tier 2 cancer-related human genes. Implications: A combined genomics score incorporating tumor mutational burden and copy-number variation was strongly prognostic of MCC-specific survival controlling for lymph node metastases and tumor MCPyV status; thus, our study adds critical understanding to prognostic markers and tumorigenic mechanisms in MCC.

Next generation sequencing quality control data, HPViewer read counts and somatic alteration burden

Lollipop plots illustrating the non-synonymous alterations identified in the MCPyV Large T and Small T coding regions for each VP-MCC tumor.

Survival analyses for prognostic genetic signatures in MCC cohort. A-B. Kaplan-Meier curves for MCC-specific survival by t-TMB category in N+ patients (A) and VP-MCC patients (B). Kaplan-Meier curves for MCC-specific survival by CNV score category in N+ patients (C) and VP-MCC patients (D).