Insertional oncogenesis by HPV70 revealed by multiple genomic analyses in a clinically HPV‐negative cervical cancer

Abstract

Cervical carcinogenesis, the second leading cause of cancer death in women worldwide, is caused by multiple types of human papillomaviruses (HPVs). To investigate a possible role for HPV in a cervical carcinoma that was HPV‐negative by PCR testing, we performed HPV DNA hybridization capture plus massively parallel sequencing. This detected a subgenomic, URR‐E6‐E7‐E1 segment of HPV70 DNA, a type not generally associated with cervical cancer, inserted in an intron of the B‐cell lymphoma/leukemia 11B (BCL11B) gene in the human genome. Long range DNA sequencing confirmed the virus and flanking BCL11B DNA structures including both insertion junctions. Global transcriptomic analysis detected multiple, alternatively spliced, HPV70‐BCL11B, fusion transcripts with fused open reading frames. The insertion and fusion transcripts were present in an intraepithelial precursor phase of tumorigenesis. These results suggest oncogenicity of HPV70, identify novel BCL11B variants with potential oncogenic implications, and underscore the advantages of thorough genomic analyses to elucidate insights into HPV‐associated tumorigenesis.

Structure and transcription of integrated HPV DNA in vulvar carcinomas

“…2 ). HPV DNA insertions usually occur in conjunction with deletion of human sequences between the human-virus DNA junction pair, and with microhomology or small DNA insertions at the junctions with no flanking repeats 21 , similar to the observations made for Tumor 5 in this study (Fig. 2 ).…”

“…Since the breadth of effectiveness of GP5+/GP6+ primers with less common HPV types is uncertain 22 , and some HPV-induced cancers contain integrated, subgenomic HPV DNA segments lacking the GP5+/GP6+ target segment 12 , we performed HC + SEQ on all 13 samples to define the identity and structure of HPV DNA in each 21 . HPV DNA enrichment was accomplished through the utilization of a custom-designed capture probe set, consisting of oligonucleotide probes complementary to 143 different HPV types (HPV1 through HPV143 10 ).…”

“…In contrast, the late genes were situated upstream of the URR and HPV transcription start sites, and thus improperly positioned to be transcribed from the viral URR or late (in E7) promoters. Such an overall structure is characteristic of HPV DNA integrations in other HPV induced tumors 21 , 31 , and presumably is important for expression of the viral E6 and E7 oncogenes.…”

“…The extent of HPV DNA integration in HPV-dependent vulvar carcinomas, and whether it resembles the patterns observed in cervical cancers and other HPV-associated tumors, particularly head and neck squamous cell carcinomas (HNSCCs), remained unknown. Hence, our objective was to assess HPV DNA integration, genome structure, and transcription in a cohort of vulvar carcinomas using a custom, hybridization capture (HC) method capable of detecting multiple HPV types, followed by Illumina massively parallel high-throughput sequencing (HC + SEQ) 21 . To further enhance our analysis, we employed long-range nanopore DNA sequencing and Illumina RNA-seq to examine the tumor transcriptomes.…”

Accurate Detection of HPV Integration Sites in Cervical Cancer Samples Using the Nanopore MinION Sequencer Without Error Correction

“…There was only few newly published paper using nanopore platform to identify HPV integration sites ( Quan et al, 2019 ; Van Arsdale et al, 2019 ). The first study revealed that using nanopore sequencing could simultaneously detect HPV infection and microbiota composition promptly and accurately.…”