Features of HPV+ OPSCC predictive of recurrence identified by transcriptional profiling of a case-control cohort.

6055 Background: De-escalating therapy for HPV+ oropharyngeal cancers (OPSCCs) is hampered by poor ability to predict recurrence, and limited insight into the biologic traits predisposing to recurrence impedes personalizing therapy. We aimed to (1) define biologic features underlying therapy resistance by transcriptional profiling and (2) evaluate the features for prognostic utility. Methods: A single institution cohort of 851 HPV+ OPSCC patients undergoing transoral robotic surgery during 2007-2020 was used to identify 50 cases that recurred locoregionally in the adjuvant RT field (n=14) and/or at distant sites (n=43). As controls, we used 49 recurrence-free patients with long-term follow up and similar stage, smoking history, and adjuvant therapy. RNAseq of pretreatment tumors was used to compare individual gene expression and Hallmark/Kegg pathway activity between cases and controls using unpaired t-test (p<.05) and Gene Set Enrichment Analysis (p-adj<.05), respectively. Activity of the significant pathways was quantified in individual tumors using Gene Set Variation Analysis (GSVA), and a regression-based composite of GSVA scores was developed to distinguish cases from controls by ROC analysis. The composite score was used to test for stratification of recurrence free survival (RFS) in external cohorts using Youden Index and logrank test. Results: The21 Hallmark/Kegg pathways downregulated in cases indicated suppression of anti-tumor immunity, and the 20 upregulated ones revealed increased biosynthetic function and tumor cell proliferation. The 1472 upregulated mRNAs contained several components of the ATR-chk1 DNA repair pathway and trans-lesion synthesis polymerases, suggesting that mitigation of DNA replication stress enhanced tumor growth. The 958 downregulated mRNAs suggested reductions in the cytoplasmic dsDNA sensing and downstream NF-kB signals that lead to replication stress-driven anti-tumor immunity. Two GSVA scores were created to quantify the key tumor-intrinsic and immune suppressive features separately in each tumor. The scores correlated with each other (R=0.6, p<.001), and a regression-based combination of them distinguished cases from controls (AUC=.76, p<.001). This composite score also stratified RFS in three external cohorts: TCGA (n=52, p<0.001) and two single institution cohorts containing both surgical and nonsurgical cases (n=46, p=.002; n=81, p<0.001). Conclusions: HPV+ OPSCCs failing primary surgical therapy show evidence of reduced replication stress mediating tumor progression and low anti-tumor immunity. These features appear generalizable to heterogeneously treated external cohorts, where they predicted recurrence. Our results provide a new basis for creating transcriptomic predictors of treatment response and suggest targetable molecular mechanisms to overcome therapy resistance.