IMAT-ITCR Collaboration: Development of a high-resolution mapping platform for HPV DNA integration in premalignant lesions

/Summary Viruses play a role in 10 to 20% of human cancers. Tying specific viruses to human cancers remains a fundamental biomedical and technological problem. One virus family, the human papillomaviruses (HPVs), comprises over 225 known types and causes lesions ranging from benign warts to highly lethal, invasive carcinomas. HPV-induced tumors occur at several anatomical sites including nearly 100% of cervical cancers, 90% of anal cancers, 30 to 60% of head & neck tumors, and roughly one-fourth to one-half of vaginal, vulvar & penile cancers. In most HPV-induced tumors, at least part of the viral DNA genome has become integrated into the human genome, presumably as a consequence of aberrant host cell DNA repair processes. Our laboratory work has been focused on the development of accurate, sensitive, broad specificity technology based on DNA hybridization capture plus massively parallel sequencing to detect and structurally analyze integrated HPV DNA in precancerous lesions and invasive tumors. Ongoing studies are also developing fluorescent microscopy technologies for detection of both integrated HPV DNA and HPV RNA transcripts, as these provide highly specific, potential diagnostic tools to detect the presence of HPV-induced tumor cells, including post-therapy residual disease in clinical samples. Our work encompasses long-range, nanopore DNA sequencing plus RNA sequencing to confirm any complex structural rearrangements of the HPV and human genomes and to elucidate potential viral effects on viral and human gene transcription. Key to these studies has been a very successful, collaborative effort by the Haas group (Broad Institute at MIT) with the Lenz (Albert Einstein College of Medicine) and Montagna (Rutgers Cancer Institute of New Jersey) labs to expand the computational tools of the Trinity Cancer Transcriptome Analysis Toolkit (CTAT). This has led to the development of expanded CTAT components for detection of integrated HPV DNA, structural assembly of integrated viral DNA segments, and overall transcriptional analysis of HPV-induced tumors and precancerous lesions. The CTAT-virus insertion finder (CTAT-VIF) is available on GitHub. The IMAT-ITCR collaboration proposed here will substantially expand the ongoing collaboration by pursuing three additional aims. Aim 1 will expand CTAT-VIF to analyze the presence of ~13,000 different viruses and investigate if any viral DNAs are integrated and/or expressed by screening all The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression Project (GTEx) datasets. Aim 2 is to identify and analyze novel virus-human fusion transcripts generated from virus DNA insertions. Aim 3 will broaden our in-situ analysis of HPVinduced cervical cancers and precancerous lesions to include spatial transcriptomic analysis, including expanding CTAT for this technology. These studies should improve computational and laboratory technologies for understanding the roles of viruses in human cancer.