The prime objective for every life form is to deliver its genetic material, intact and unchanged, to the next generation. This must be achieved despite constant assaults by endogenous and environmental agents on the DNA. To counter this threat, life has evolved several systems to detect DNA damage, signal its presence and mediate its repair. Such responses, which have an impact on a wide range of cellular events, are biologically significant because they prevent diverse human diseases. Our improving understanding of DNA-damage responses is providing new avenues for disease management.

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The DNA-damage response in human biology and disease

Head and neck squamous cell carcinoma (HNSCC) is a common, morbid, and frequently lethal malignancy. To uncover its mutational spectrum, we analyzed whole-exome sequencing data from 74 tumor-normal pairs. The majority exhibited a mutational profile consistent with tobacco exposure; human papillomavirus was detectable by sequencing DNA from infected tumors. In addition to identifying previously known HNSCC genes (TP53, CDKN2A, PTEN, PIK3CA, and HRAS), our analysis revealed many genes not previously implicated in this malignancy. At least 30% of cases harbored mutations in genes that regulate squamous differentiation (for example, NOTCH1, IRF6, and TP63), implicating its dysregulation as a major driver of HNSCC carcinogenesis. More generally, the results indicate the ability of large-scale sequencing to reveal fundamental tumorigenic mechanisms.

/pdf/the-mutational-landscape-of-head-and-neck-squamous-cell-oc5uxdfntk.pdf

The Mutational Landscape of Head and Neck Squamous Cell Carcinoma

The long non-coding RNA MALAT1, also known as MALAT-1 or NEAT2, is a highly conserved nuclear ncRNA and a predictive marker for metastasis development in lung cancer. To uncover its functional importance, we developed a MALAT1 knockout model in human lung tumor cells by genomically integrating RNA destabilizing elements using Zinc Finger Nucleases. The achieved 1000-fold MALAT1 silencing provides a unique loss-of-function model. Proposed mechanisms of action include regulation of splicing or gene expression. In lung cancer, MALAT1 does not alter alternative splicing but actively regulates gene expression including a set of metastasis-associated genes. Consequently, MALAT1-deficient cells are impaired in migration and form fewer tumor nodules in a mouse xenograft. Antisense oligonucleotides blocking MALAT1 prevent metastasis formation after tumor implantation. Thus, targeting MALAT1 with antisense oligonucleotides provides a potential therapeutic approach to prevent lung cancer metastasis with MALAT1 serving as both, predictive marker and therapeutic target. Lastly, regulating gene expression, but not alternative splicing is the critical function of MALAT1 in lung cancer metastasis. In summary, ten years after the discovery of the lncRNA MALAT1 as a biomarker for lung cancer metastasis, our loss-of-function model unravels the active function of MALAT1 as a regulator of gene expression governing hallmarks of lung cancer metastasis.

/pdf/the-noncoding-rna-malat1-is-a-critical-regulator-of-the-1hkni2t23y.pdf

The Noncoding RNA MALAT1 Is a Critical Regulator of the Metastasis Phenotype of Lung Cancer Cells

Oesophageal cancers are prominent worldwide; however, there are few targeted therapies and survival rates for these cancers remain dismal. Here we performed a comprehensive molecular analysis of 164 carcinomas of the oesophagus derived from Western and Eastern populations. Beyond known histopathological and epidemiologic distinctions, molecular features differentiated oesophageal squamous cell carcinomas from oesophageal adenocarcinomas. Oesophageal squamous cell carcinomas resembled squamous carcinomas of other organs more than they did oesophageal adenocarcinomas. Our analyses identified three molecular subclasses of oesophageal squamous cell carcinomas, but none showed evidence for an aetiological role of human papillomavirus. Squamous cell carcinomas showed frequent genomic amplifications of CCND1 and SOX2 and/or TP63, whereas ERBB2, VEGFA and GATA4 and GATA6 were more commonly amplified in adenocarcinomas. Oesophageal adenocarcinomas strongly resembled the chromosomally unstable variant of gastric adenocarcinoma, suggesting that these cancers could be considered a single disease entity. However, some molecular features, including DNA hypermethylation, occurred disproportionally in oesophageal adenocarcinomas. These data provide a framework to facilitate more rational categorization of these tumours and a foundation for new therapies.

/pdf/integrated-genomic-characterization-of-oesophageal-carcinoma-13ov4kwcqk.pdf

Integrated genomic characterization of oesophageal carcinoma.

The biology and life-cycle of human papillomaviruses.

Human papillomaviruses (HPV) are believed to be the primary causal agents for development of pre-neoplastic and malignant lesions of the uterine cervix, and high-risk types such as type 16 and 18 are associated with more than 90% of all cervical carcinomas. The E6 and E7 genes of HPV are thought to play causative roles, since E6 promotes the degradation of p53 through its interaction with E6AP, an E3 ubiquitin ligase, whereas E7 binds to the retinoblastoma protein (pRb) and disrupts its complex formation with E2F transcription factors. Although prophylactic vaccines have become available, it is still necessary to clarify the mechanisms of HPV-induced carcinogenesis because of the widespread nature of HPV infection. Approximately 493,000 new cases of cervical cancer are diagnosed each year with approximately 274,000 mortalities due to invasive cervical cancer. In the present article, the mechanisms of HPV16 E6- and E7-induced multistep carcinogenesis and recently identified functions of these onco-proteins are reviewed.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1349-7006.2007.00546.x

Basic mechanisms of high-risk human papillomavirus-induced carcinogenesis: roles of E6 and E7 proteins.

The E6 protein of cervical cancer-associated human papillomaviruses (HPVs) is known to suppress keratinocyte differentiation through unidentified mechanisms. Notch1 is a determinant of keratinocyte differentiation and functions as a tumor suppressor in mammalian epidermis. Here, we report that the Notch1 gene is a novel target of p53 and can be down-regulated by E6 through p53 degradation in normal human epithelial cells. Thus, inactivation of p53 by E6 or short-hairpin RNA (shRNA) resulted in reduced Notch1 expression at the transcription level, and a p53-responsive element could be identified in the Notch1 promoter. The expression of E6, p53 shRNA, or Notch1 shRNA suppressed both spontaneous keratinocyte differentiation in culture and its induction upon DNA damage. Furthermore, the induction of Notch1 and differentiation makers as well as thickening of the epidermal layer upon UV irradiation was observed in wild-type but not in p53-deficient mouse skin. Together, our findings not only demonstrate a novel link between p53 and Notch1 in keratinocyte differentiation upon genotoxic stress but also suggest a novel tumor suppressor mechanism of p53 in the development of squamous cell carcinomas, including HPV-induced tumors.

Regulation of Notch1 gene expression by p53 in epithelial cells.

NRF2 Mutation Confers Malignant Potential and Resistance to Chemoradiation Therapy in Advanced Esophageal Squamous Cancer

The activity of human Cdt1 is negatively regulated by multiple mechanisms. This suggests that Cdt1 deregulation may have a deleterious effect. Indeed, it has been suggested that overexpression of Cdt1 can induce rereplication in cancer cells and that rereplication activates Ataxia-telangiectasia-mutated (ATM) kinase and/or ATM- and Rad3-related (ATR) kinase-dependent checkpoint pathways. In this report, we highlight a new and interesting aspect of Cdt1 deregulation: data from several different systems all strongly indicate that unregulated Cdt1 overexpression at pathophysiological levels can induce chromosomal damage other than rereplication in non-transformed cells. The most important finding in these studies is that deregulated Cdt1 induces chromosomal damage and activation of the ATM-Chk2 DNA damage checkpoint pathway even in quiescent cells. These Cdt1 activities are negatively regulated by cyclin A/Cdks, probably through modification by phosphorylation. Furthermore, we found that deregulated Cdt1 induces chromosomal instability in normal human cells. Since Cdt1 is overexpressed in cancer cells, this would be a new molecular mechanism leading to carcinogenesis.

/pdf/deregulation-of-cdt1-induces-chromosomal-damage-without-21h266udfl.pdf

Deregulation of Cdt1 induces chromosomal damage without rereplication and leads to chromosomal instability.

Ovarian surface epithelium (OSE) is considered to give rise to epithelial ovarian carcinomas (EOCs). To elucidate early processes contributing to the development of EOCs from the OSE, two batches of primary human OSE cells were transduced with non-viral human genes (mutant Cdk4, cyclinD1 and hTERT) so as to efficiently establish normal diploid OSE cells without chromosomal instability. Then defined genetic alterations frequently observed in EOCs were transduced into the OSE cells. A combination of p53 inactivation and oncogenic Kras transduction did not confer tumor-forming ability in immunodeficient mice, though additional transduction of Akt or combined transduction of c-myc with bcl-2 did result in tumor formation. In the latter case, tumors demonstrated phenotypes reminiscent of human EOCs, including cytokeratin expression, a highly aggressive phenotype, metastatic behavior and formation of ascites. These results indicate that inactivation of p53 and activation of the Ras pathway play critical roles in ovarian carcinogenesis in co-operation with the Akt or c-myc pathways. This first in vitro model system faithfully recapitulating the development of EOCs using normal human OSE cells should greatly facilitate further studies of EOCs.

/pdf/oncogenic-transformation-of-human-ovarian-surface-epithelial-7qh7puwym6.pdf

Mako Narisawa-Saito

Papers

Basic mechanisms of high-risk human papillomavirus-induced carcinogenesis: roles of E6 and E7 proteins.

Regulation of Notch1 gene expression by p53 in epithelial cells.

NRF2 Mutation Confers Malignant Potential and Resistance to Chemoradiation Therapy in Advanced Esophageal Squamous Cancer

Deregulation of Cdt1 induces chromosomal damage without rereplication and leads to chromosomal instability.

Oncogenic transformation of human ovarian surface epithelial cells with defined cellular oncogenes.