THE HUMAN PAPILLOMAVIRUS TYPE 16 E7 (HPV-16 E7) ONCOPROTEIN AND THE HOST CELL DNA DAMAGE RESPONSE

High-risk human papillomaviruses (HPVs), such as HPV-16, are the etiological agents of squamous cell carcinomas (SCCs) of the anogenital tract and a subset of oropharyngeal cancers. High-risk HPVs encode two oncoproteins, E6 and E7, which promote unscheduled host cell proliferation by targeting the p53 and pRB tumor suppressor proteins, respectively. HPV-16 E7 has been shown to stimulate structural chromosomal instability and DNA breakage. These findings raise several important questions. First, how does HPV-16 E7 induce DNA damage? Second, what are the precise consequences of HPV-16 E7-induced DNA damage for host cell genomic integrity, and lastly, how do HPV-16 E7-expressing cells maintain proliferation despite activated DNA damage checkpoints? Here, we show that HPV-16 E7 activates the Fanconi Anemia (FA) pathway, a branch of the host cell DNA damage response that primarily responds to stalled DNA replication forks. Importantly, we show that HPV-16 E7 expression in FA-deficient cells accelerates the formation of structural chromosomal alterations, which may help to explain the heightened susceptibility of FA patients to HPV-associated tumors. However, we also provide evidence that HPV-16 E7-induced FA pathway activation in FA-proficient cells may contribute to evasion of anti-proliferative host cell barriers by promoting alternative lengthening of telomeres (ALT). Finally, we demonstrate that HPV-16 E7 circumvents DNA damage checkpoint control and promotes aberrant mitotic entry by increasing the proteolytic turnover of claspin, which plays a role in the ATR/CHK1-mediated replication stress response. Collectively, our results underscore that HPV-16 E7 interferes with host cell genome integrity by inducing DNA replication stress. The detrimental effects of HPV-16 E7 on the genomic integrity of host cells with a deficient FA pathway support the notion that this DNA damage response pathway is crucial to prevent HPV-16 E7-induced genomic instability and malignant progression. However, we also provide evidence that HPV-16 E7 can exploit the FA pathway to promote cellular immortalization. Future experiments to explore these events for cancer therapy and/or prevention are warranted

PIK3CA mutant tumors depend on oxoglutarate dehydrogenase

Oncogenic PIK3CA mutations are found in a significant fraction of human cancers, but therapeutic inhibition of PI3K has only shown limited success in clinical trials. To understand how mutant PIK3CA contributes to cancer cell proliferation, we used genome scale loss-of-function screening in a large number of genomically annotated cancer cell lines. As expected, we found that PIK3CA mutant cancer cells require PIK3CA but also require the expression of the TCA cycle enzyme 2-oxoglutarate dehydrogenase (OGDH). To understand the relationship between oncogenic PIK3CA and OGDH function, we interrogated metabolic requirements and found an increased reliance on glucose metabolism to sustain PIK3CA mutant cell proliferation. Functional metabolic studies revealed that OGDH suppression increased levels of the metabolite 2-oxoglutarate (2OG). We found that this increase in 2OG levels, either by OGDH suppression or exogenous 2OG treatment, resulted in aspartate depletion that was specifically manifested as auxotrophy within PIK3CA mutant cells. Reduced levels of aspartate deregulated the malate-aspartate shuttle, which is important for cytoplasmic NAD + regeneration that sustains rapid glucose breakdown through glycolysis. Consequently, because PIK3CA mutant cells exhibit a profound reliance on glucose metabolism, malate-aspartate shuttle deregulation leads to a specific proliferative block due to the inability to maintain NAD + /NADH homeostasis. Together these observations define a precise metabolic vulnerability imposed by a recurrently mutated oncogene. Keyword: PIK3CA; 2OG; OGDH; TCA cycle; glycolysisDamon Runyon Cancer Research Foundation (HHMI Fellowship

Genomic sequencing of colorectal adenocarcinomas identifies a recurrent VTI1A-TCF7L2 fusion

Prior studies have identified recurrent oncogenic mutations in colorectal adenocarcinoma1 and have surveyed exons of protein-coding genes for mutations in 11 affected individuals2,3. Here we report whole-genome sequencing from nine individuals with colorectal cancer, including primary colorectal tumors and matched adjacent non-tumor tissues, at an average of 30.7× and 31.9× coverage, respectively. We identify an average of 75 somatic rearrangements per tumor, including complex networks of translocations between pairs of chromosomes. Eleven rearrangements encode predicted in-frame fusion proteins, including a fusion of VTI1A and TCF7L2 found in 3 out of 97 colorectal cancers. Although TCF7L2 encodes TCF4, which cooperates with β-catenin4 in colorectal carcinogenesis5,6, the fusion lacks the TCF4 β-catenin–binding domain. We found a colorectal carcinoma cell line harboring the fusion gene to be dependent on VTI1A-TCF7L2 for anchorage-independent growth using RNA interference-mediated knockdown. This study shows previously unidentified levels of genomic rearrangements in colorectal carcinoma that can lead to essential gene fusions and other oncogenic events

Genomic sequencing of colorectal adenocarcinomas identifies a recurrent VTI1A-TCF7L2 fusion

Prior studies have identified recurrent oncogenic mutations in colorectal adenocarcinoma and have surveyed exons of protein-coding genes for mutations in 11 affected individuals. Here we report whole-genome sequencing from nine individuals with colorectal cancer, including primary colorectal tumors and matched adjacent non-tumor tissues, at an average of 30.7× and 31.9× coverage, respectively. We identify an average of 75 somatic rearrangements per tumor, including complex networks of translocations between pairs of chromosomes. Eleven rearrangements encode predicted in-frame fusion proteins, including a fusion of VTI1A and TCF7L2 found in 3 out of 97 colorectal cancers. Although TCF7L2 encodes TCF4, which cooperates with β-catenin in colorectal carcinogenesis, the fusion lacks the TCF4 β-catenin–binding domain. We found a colorectal carcinoma cell line harboring the fusion gene to be dependent on VTI1A-TCF7L2 for anchorage-independent growth using RNA interference-mediated knockdown. This study shows previously unidentified levels of genomic rearrangements in colorectal carcinoma that can lead to essential gene fusions and other oncogenic events