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The development of effective cancer treatments depends on the availability of cell lines that faithfully recapitulate the cancer in question. This study definitively re-assigns the histologic identities of two ovarian cancer cell lines, COV434 (originally described as a granulosa cell tumour) and TOV-112D (originally described as grade 3 endometrioid carcinoma), both of which were recently suggested to represent small cell carcinoma of the ovary, hypercalcemic type (SCCOHT), based on their shared gene expression profiles and sensitivity to EZH2 inhibitors. For COV434 and TOV-112D, we re-reviewed the original pathology slides and obtained clinical follow-up on the patients, when available, and performed immunohistochemistry for SMARCA4, SMARCA2 and additional diagnostic markers on the original formalin-fixed, paraffin-embedded (FFPE) clinical material, when available. For COV434, we further performed whole exome sequencing and validated SMARCA4 mutations by Sanger sequencing. We studied the growth of the cell lines at baseline and upon re-expression of SMARCA4 in vitro for both cell lines and evaluated the serum calcium levels in vivo upon injection into immunodeficient mice for COV434 cells. The available morphological, immunohistochemical, genetic, and clinical features indicate COV434 is derived from SCCOHT, and TOV-112D is a dedifferentiated carcinoma. Transplantation of COV434 into mice leads to increased serum calcium level. Re-expression of SMARCA4 in either COV434 and TOV-112D cells suppressed their growth dramatically. COV434 represents a bona fide SCCOHT cell line. TOV-112D is a dedifferentiated ovarian carcinoma cell line. • The original tumour of COV434 cells displayed morphological, immunohistochemical, genetic, and clinical features of SCCOHT. • The original tumour of TOV-112D cells was re-diagnosed as a dedifferentiated ovarian carcinoma. • Re-expression of SMARCA4 suppressed the growth of COV434 and TOV-112D cells dramatically. [ABSTRACT FROM AUTHOR]

Within the last decade, molecular advances have provided insights into the genetics of several ovarian sex cord–stromal tumours that have otherwise been enigmatic. Chief among these advances are the identification of FOXL2, DICER1 and CTNNB1 mutations in adult granulosa cell tumours, Sertoli–Leydig cell tumours (SLCTs), and microcystic stromal tumours (MCSTs), respectively. As access to molecular diagnostic laboratories continues to become more widely available, the potential roles for tumour mutation testing in the pathological diagnosis of these tumours merit discussion. Furthermore, links to inherited cancer susceptibility syndromes may exist for some women with SLCT (DICER1 syndrome) and MCST [familial adenomatous polyposis (FAP)]. This review will address practical issues in deciding when and how to apply mutation testing in the diagnosis of these three sex cord–stromal tumours. The pathologist's role in recommending referral for formal risk assessment for DICER1 syndrome and FAP will also be discussed. [ABSTRACT FROM AUTHOR]

Background: The newly developed Proactive Molecular Risk Classifier for Endometrial Cancer (ProMisE) has consistently been shown to be prognostically significant in endometrial carcinomas (EC). Recently, we and others have demonstrated L1 cell-adhesion molecule (L1CAM) to be a significant indicator of high-risk disease in EC. In the current study, it was our aim to determine the prognostic significance of aberrant L1CAM expression in ProMisE subgroups in a large, single centre, population-based EC cohort.Methods: ProMisE (POLE; MMR-D; p53 wt/NSMP; p53 abn) classification results from a cohort of 452 EC were available for analysis. L1CAM expression was studied by immunohistochemistry on whole slides. Correlations between clinicopathological data and survival were calculated.Results: Expression of L1CAM was most frequent in p53 abnormal tumours (80%). L1CAM status was predictive of worse outcome among tumours with no specific molecular profile (p53 wt/NSMP) (p < 0.0001). Among p53 wt/NSMP EC, L1CAM remained a significant prognosticator for disease-specific survival after multivariate analysis (p = 0.035).Conclusion: L1CAM status was able to significantly stratify risk among tumours of the large p53 wt/NSMP ProMisE subgroup of EC. Furthermore, our study confirms a highly significant correlation between mutation-type p53 immunostaining and abnormal L1CAM expression in EC. [ABSTRACT FROM AUTHOR]

Aims: Gynandroblastoma is a rare ovarian sex cord‐stromal tumour characterised by the presence of both male (Sertoli and/or Leydig cells) and female (granulosa cells) components. We investigated the mutational status of DICER1, FOXL2 and AKT1 genes at hot‐spot regions that are known to be the key driving events in the development of Sertoli–Leydig cell tumour (SLCT), adult granulosa cell tumour (aGCT) and juvenile granulosa cell tumour (jGCT), respectively, to gain insights into the molecular pathogenesis of gynandroblastoma. Methods and results: Sixteen cases of gynandroblastoma were studied. All contained SLCT or Sertoli cell tumour components. aGCT and jGCT components were identified in seven and 10 cases, respectively, with one presenting both components. Heterozygous hot‐spot mutations in the RNase IIIb domain of DICER1 were discovered in three cases, including one case with heterologous mucinous elements, all of which were composed of moderately or poorly differentiated SLCT and jGCT components, and harboured the mutations in both histological components. None of the 16 cases displayed mutations at the p.C134W (c.402C→G) of FOXL2 or within the pleckstrin‐homology domain of AKT1. All cases showed FOXL2 immunostaining in both male and female components. Conclusion: DICER1 hot‐spot mutation is the key‐driving event in a subset of gynandroblastomas containing components of SLCT and jGCT. Gynandroblastomas composed of SLCT and jGCT may represent morphological variants of SLCT. The molecular basis of gynandroblastoma containing a component of aGCT is different from pure aGCT. [ABSTRACT FROM AUTHOR]

Preclinical models are relatively underutilized and underfunded resources for modeling the pathogenesis and prevention of ovarian cancers. Several reviews have detailed the numerous published models of ovarian cancer. In this review, we will provide an overview of experimental model systems, their strengths and limitations, and use selected models to illustrate how they can be used to address specific issues about ovarian cancer pathogenesis. We will then highlight some of the preclinical prevention studies performed to date and discuss experiments needed to address important unanswered questions about ovarian cancer prevention strategies. [ABSTRACT FROM AUTHOR]

Non-small cell lung carcinoma (NSCLC) is the leading cause of cancer-related death worldwide, with an overall 5-year survival rate of only 10-15%. Deregulation of the Ras pathway is a frequent hallmark of NSCLC, often through mutations that directly activate Kras. p53 is also frequently inactivated in NSCLC and, because oncogenic Ras can be a potent trigger of p53 (ref. 3), it seems likely that oncogenic Ras signalling has a major and persistent role in driving the selection against p53. Hence, pharmacological restoration of p53 is an appealing therapeutic strategy for treating this disease. Here we model the probable therapeutic impact of p53 restoration in a spontaneously evolving mouse model of NSCLC initiated by sporadic oncogenic activation of endogenous Kras. Surprisingly, p53 restoration failed to induce significant regression of established tumours, although it did result in a significant decrease in the relative proportion of high-grade tumours. This is due to selective activation of p53 only in the more aggressive tumour cells within each tumour. Such selective activation of p53 correlates with marked upregulation in Ras signal intensity and induction of the oncogenic signalling sensor p19ARF (ref. 6). Our data indicate that p53-mediated tumour suppression is triggered only when oncogenic Ras signal flux exceeds a critical threshold. Importantly, the failure of low-level oncogenic Kras to engage p53 reveals inherent limits in the capacity of p53 to restrain early tumour evolution and in the efficacy of therapeutic p53 restoration to eradicate cancers. [ABSTRACT FROM AUTHOR]

A key difference among the three structurally similar pRB family members is that only pRB is a tumor suppressor. Identification of distinctive functional differences between pRB and p107/p130 therefore holds promise for a better understanding of the tumor suppression mechanisms of pRB. Enigmatically, pRB and p107 have been shown to have indistinguishable growth suppression activities when studied in the pRB-deficient Saos-2 cell system. In this study, we discovered that, when expressed at physiologically relevant levels, pRB and p107 had distinctive effects in causing growth suppression. pRB induced cellular p130 levels while p107 repressed them. p107, but not pRB, blocked cells inside S phase in addition to G1 arrest. In contrast, no qualitative differences were identified in their abilities to repress the expression of a set of suspected pRB/E2F repression target genes. These results indicate that pRB and p107 possess different growth suppression effects, despite the fact that they have similar E2F repression effects. Oncogene (2000) 19, 3878–3887 [ABSTRACT FROM AUTHOR]

Small cell carcinoma of the ovary of hypercalcemic type (SCCOHT) is an extremely rare, aggressive cancer affecting children and young women. We identified germline and somatic inactivating mutations in the SWI/SNF chromatin-remodeling gene SMARCA4 in 75% (9/12) of SCCOHT cases in addition to SMARCA4 protein loss in 82% (14/17) of SCCOHT tumors but in only 0.4% (2/485) of other primary ovarian tumors. These data implicate SMARCA4 in SCCOHT oncogenesis. [ABSTRACT FROM AUTHOR]

A correction to the article "Small cell carcinoma of the ovary, hypercalcemic type, displays frequent inactivating germline and somatic mutations in SMARCA4," published online in the March 23, 2014 issue, is presented.

Aims Dedifferentiated endometrial carcinoma ( DDEC) is defined by the presence of an undifferentiated carcinoma together with an endometrioid carcinoma. Inactivation of SMARCA4 ( BRG1) and inactivation of SMARCB1 ( INI1) were recently described as potential mechanisms underlying the histological dedifferentiation. The aim of this study was to characterize the immunophenotypic features of DDECs, particularly in cases with prototypical histological and molecular features ( BRG1/ INI1 deficiency). Methods and results We evaluated PAX8, oestrogen receptor ( ER) and p53 immunostaining in the endometrioid and the undifferentiated components of 20 BRG1/ INI1-deficient DDECs and 15 BRG1/ INI1-intact DDECs, and compared the results with those of 23 grade 3 endometrioid carcinomas. The differentiated endometrioid component was positive for PAX8 and/or ER in 19 of 20 BRG1/ INI1-deficient DDECs, whereas the corresponding undifferentiated component of all 20 tumours showed a complete absence of PAX8 and ER staining. All except one of the BRG1/ INI1-deficient tumours showed a wild-type p53 staining pattern. PAX8 and ER expression in the undifferentiated component was absent in 67% and 80% of BRG1/ INI1-intact DDECs, respectively, whereas 47% of the BRG1/ INI1-intact DDECs showed a mutated p53 staining pattern. In comparison, absent PAX8 expression and absent ER expression were each observed in the more solid area of 48% and 48% of grade 3 endometrioid carcinomas. Conclusions The consistent absence of PAX8 and ER expression in molecularly defined ( BRG1/ INI1-deficient) DDECs suggests that the loss of PAX8 and ER expression is a fundamental feature of dedifferentiation. The frequent findings of a mutated p53 staining pattern in BRG1/ INI1-intact DDECs indicate that BRG1/ INI1-intact DDECs may be biologically different from BRG1/ INI1-deficient tumours. [ABSTRACT FROM AUTHOR]

Today, one of the most popular and controversial topics in medicine is undoubtedly the rapidly developing field of stem cell research. Some of the controversy in this field arises from lack of uniform terminology and different interpretation of concepts such as brain tumor stem cells. In addition, lack of reliable and universal markers that can identify stem cells and define precursor cells in a particular differentiation pathway further confounds the interpretation of results in many studies. Stem cells are undoubtedly critical in normal cellular development as well as tumor biology and better characterization of these cells is likely to have profound influence on the classification schemes of tumors. In this manuscript, we present the generally accepted definitions of key concepts in stem cell biology and review some of the related molecular pathways. In addition, we put forth our position on how progress in this field should be affecting the future classification schemes of central nervous system neoplasia. We strongly believe that the ever increasing knowledge in the field of neural and brain tumor stem cells should be influential in the subsequent attempts to classify brain tumors. [ABSTRACT FROM AUTHOR]

Kras is the most frequently mutated ras family member in lung carcinomas, whereas Hras mutations are common in tumors from stratified epithelia such as the skin. Using a Hras knock-in mouse model, we demonstrate that specificity for Kras mutations in lung and Hras mutations in skin tumors is determined by local regulatory elements in the target ras genes. Although the Kras 4A isoform is dispensable for mouse development, it is the most important isoform for lung carcinogenesis in vivo and for the inhibitory effect of wild-type (WT) Kras on the mutant allele. Kras 4A expression is detected in a subpopulation of normal lung epithelial cells, but at very low levels in lung tumors, suggesting that it may not be required for tumor progression. The two Kras isoforms undergo different post-translational modifications; therefore, these findings can have implications for the design of therapeutic strategies for inhibiting oncogenic Kras activity in human cancers. [ABSTRACT FROM AUTHOR]

Background: TP53 mutations occur in more than 50% of cancers. We sought to determine the effect of the intragenic P72R single nucleotide polymorphism (SNP; rs1042522) on the oncogenic properties of mutant p53.Methods: P72R allelic selection in tumors was determined from genotype calls and a Gaussian distributed mixture model. The SNP effect on mutant p53 was determined in p53-negative cancer cell lines. RNA-sequencing, chromatin immunoprecipitation, and survival analysis were performed to describe the SNP effect. All statistical tests were 2-sided.Results: Among 409 patients with germline heterozygous P72R SNP who harbored somatic mutations in TP53, we observed a selection bias against missense TP53 mutants encoding the P72 SNP (P = 1.64 x 10-13). Exogenously expressed hotspot p53 mutants with the P72 SNP were negatively selected in cancer cells. Gene expression analyses showed the enrichment of p53 pathway genes and inflammatory genes in cancer cells transduced with mutants encoding P72 SNP. Immune gene signature is enriched in patients harboring missense TP53 mutations with homozygous P72 SNP. These patients have improved overall survival as compared with those with the R72 SNP (P = .04).Conclusion: This is the largest study demonstrating a selection against the P72 SNP. Missense p53 mutants with the P72 SNP retain partial wild-type tumor-suppressive functions, which may explain the selection bias against P72 SNP across cancer types. Ovarian cancer patients with the P72 SNP have a better prognosis than with the R72 SNP. Our study describes a previously unknown role through which the rs1042522 SNP modifies tumor suppressor activities of mutant p53 in patients. [ABSTRACT FROM AUTHOR]

Background: PTEN loss is a putative driver in histotypes of ovarian cancer (high-grade serous (HGSOC), endometrioid (ENOC), clear cell (CCOC), mucinous (MOC), low-grade serous (LGSOC)). We aimed to characterise PTEN expression as a biomarker in epithelial ovarian cancer in a large population-based study.Methods: Tumours from 5400 patients from a multicentre observational, prospective cohort study of the Ovarian Tumour Tissue Analysis Consortium were used to evaluate associations between immunohistochemical PTEN patterns and overall survival time, age, stage, grade, residual tumour, CD8+ tumour-infiltrating lymphocytes (TIL) counts, expression of oestrogen receptor (ER), progesterone receptor (PR) and androgen receptor (AR) by means of Cox proportional hazard models and generalised Cochran-Mantel-Haenszel tests.Results: Downregulation of cytoplasmic PTEN expression was most frequent in ENOC (most frequently in younger patients; p value = 0.0001) and CCOC and was associated with longer overall survival in HGSOC (hazard ratio: 0.78, 95% CI: 0.65-0.94, p value = 0.022). PTEN expression was associated with ER, PR and AR expression (p values: 0.0008, 0.062 and 0.0002, respectively) in HGSOC and with lower CD8 counts in CCOC (p value < 0.0001). Heterogeneous expression of PTEN was more prevalent in advanced HGSOC (p value = 0.019) and associated with higher CD8 counts (p value = 0.0016).Conclusions: PTEN loss is a frequent driver in ovarian carcinoma associating distinctly with expression of hormonal receptors and CD8+ TIL counts in HGSOC and CCOC histotypes. [ABSTRACT FROM AUTHOR]

Cancer patient-derived organoids (PDOs) grow as three dimensional (3D) structures in the presence of extracellular matrix and have been found to represent the original tumor's genetic complexity. In addition, PDOs can be grown and subjected to drug sensitivity testing in a shorter time course and with lesser expense than patient-derived xenograft models. Many patients with recurrent ovarian cancer develop malignant effusions that become refractory to chemotherapy. Since these same patients often present for palliative aspiration of ascites or pleural effusions, there is a potential opportunity to obtain tumor specimens in the form of multicellular spheroids (MCS) present in malignant effusion fluids. Our objective was to develop a short duration culture of MCS from ovarian cancer malignant effusions in conditions selected to support organoid growth and use them as a platform for empirical drug sensitivity testing. In this study, malignant effusion specimens were collected from patients with high-grade serous ovarian carcinoma (HGSOC). MCS were recovered and subjected to culture conditions designed to support organoid growth. In a subset of specimens, RNA-sequencing was performed at two time points during the short-term culture to determine changes in transcriptome in response to culture conditions. Organoid induction was also characterized in these specimens using Ki67 staining and histologic analysis. Drug sensitivity testing was performed on all specimens. Our model describes organoids formed within days of primary culture, which can recapitulate the histological features of malignant ascites fluid and can be expanded for at least 6 days. RNA-seq analysis of four patient specimens showed that within 6 days of culture, there was significant up-regulation of genes related to cellular proliferation, epithelial-mesenchymal transition, and KRAS signaling pathways. Drug sensitivity testing identified several agents with therapeutic potential. Short duration organoid culture of MCS from HGSOC malignant effusions can be used as a platform for empiric drug sensitivity testing. These ex vivo models may be helpful in screening new or existing therapeutic agents prior to individualized treatment options. • Patient-derived organoids (PDOs) can be derived from malignant ascites and pleural effusions. • PDOs can be used for empirical drug testing of novel therapeutics. • RNA-sequencing analysis indicates gene signature associated with proliferation upon organoid induction. • PDOs represent an efficient model that recapitulates histological features of malignant ascites and pleural effusions. [ABSTRACT FROM AUTHOR]

Aims Vulvar squamous cell carcinoma ( VSCC) can be subdivided by human papillomavirus ( HPV) status into two clinicopathological entities. Studies on the prognostic significance of HPV in VSCC are discordant. Methods and results We performed a retrospective analysis of overall survival ( OS), disease-specific survival ( DSS) and progression-free survival ( PFS) in 217 patients with VSCC. Cases were extracted from an era of more aggressive en-bloc radical dissections (1985-95) and more localized radical surgery through separate vulvar and groin excisions (1996-2005). p16 immunohistochemistry was used as a surrogate for HPV status. HPV status could be determined in 197 tumours, 118 HPV-independent and 79 HPV-associated tumours. Patients with HPV-associated tumours were younger (mean 58.8 versus 71.6 years for HPV-independent tumours, P < 0.0001) and more likely to have prior abnormal cervical cytology (41.1 versus 5.6% for HPV-independent tumours, P < 0.0001). In univariable analysis, patients with HPV-associated tumours had superior PFS [hazard ratio ( HR): 0.37, 95% confidence interval ( CI): 0.18-0.70], DSS ( HR: 0.19, 95% CI: 0.08-0.41) and OS ( HR: 0.35, 95% CI: 0.21-0.59). This was driven by worse outcomes ( PFS, DSS and OS) for patients with HPV-independent tumours compared with HPV-associated tumours who underwent surgery after 1995. After adjusting for age and stage in multivariable analysis, patients with HPV-associated tumours showed superior PFS ( HR: 0.25, 95% CI: 0.07-0.77) and DSS ( HR: 0.21, 95% CI: 0.04-0.78). Conclusions VSCC can be stratified into two prognostically different diseases based on p16 immunostaining. HPV status was associated only with prognosis in the cohort that underwent surgery after 1995, suggesting that more conservative surgery may have led to worse outcomes for patients with HPV-independent tumours. [ABSTRACT FROM AUTHOR]

Background: Classification of endometrial carcinomas (ECs) by morphologic features is irreproducible and imperfectly reflects tumor biology. The authors developed the Proactive Molecular Risk Classifier for Endometrial Cancer (ProMisE), a molecular classification system based on The Cancer Genome Atlas genomic subgroups, and sought to confirm both feasibility and prognostic ability in a new, large cohort of ECs.Methods: Immunohistochemistry (IHC) for the presence or absence of mismatch repair (MMR) proteins (to identify MMR deficiency [MMR-D]), sequencing for polymerase-ɛ (POLE) exonuclease domain mutations (POLE EDMs), and IHC for tumor protein 53 (p53) (wild type vs null/missense mutations; p53 wt and p53 abn, respectively) were performed on 319 new EC samples. Subgroups were characterized and assessed relative to outcomes. The prognostic ability of ProMisE was compared with that of current risk-stratification systems (European Society of Medical Oncology [ESMO]).Results: ProMisE decision-tree classification achieved categorization of all cases and identified 4 prognostic subgroups with distinct overall, disease-specific, and progression-free survival (P < .001). Tumors with POLE EDMs had the most favorable prognosis, and those with p53 abn the worst prognosis, and separation of the 2 middle survival curves (p53 wt and MMR-D) was observed. There were no significant differences in survival between the ESMO low-risk and intermediate-risk groups. ProMisE improved the ability to discriminate outcomes compared with ESMO risk stratification. There was substantial overlap (89%) between the p53 abn and high-risk ESMO subgroups; but, otherwise, there were no predictable associations between molecular and ESMO risk groups.Conclusions: Molecular classification of ECs can be achieved using clinically applicable methods and provides independent prognostic information beyond established clinicopathologic risk factors available at diagnosis. Consistent, biologically relevant categorization enables stratification for clinical trials and/or targeted therapy, identification of women who are at increased risk of having Lynch syndrome, and may guide clinical management. Cancer 2017;123:802-13. © 2016 American Cancer Society. [ABSTRACT FROM AUTHOR]

Aims Molecular investigation of small-cell carcinoma of the ovary, hypercalcaemic type ( SCCOHT) has revealed that it is a monogenetic tumour characterized by alteration of SMARCA4 ( BRG1), encoding a member of the switch/sucrose non-fermentable ( SWI/ SNF) chromatin remodelling complex. A large majority of cases show loss of expression of the corresponding SMARCA4/ BRG1 protein. Furthermore, three cases of SCCOHT with retained SMARCA4 protein expression showed loss of SMARCB1/ INI1 expression. The aim of this study was to assess the sensitivity and specificity of loss of SMARCA4 expression as a diagnostic test for SCCOHT. Methods and results We performed SMARCA4 and SMARCB1 staining in 245 tumours, many of which were potentially in the differential diagnosis of SCCOHT. We also stained 56 cases of SCCOHT for SMARCA4 and 37 of these for SMARCB1. Fifty-four of the SCCOHT cases showed complete absence of SMARCA4 expression. The two cases with retained expression showed molecular alteration of SMARCA4. Of the 217 other neoplasms with interpretable staining, all retained SMARCA4 expression. Although the majority showed diffuse, strong nuclear expression, a heterogeneous, typically weak staining pattern was present in 13% of cases. All 37 cases of SCCOHT tested and all other neoplasms, apart from three malignant rhabdoid tumours, showed retained nuclear SMARCB1 expression. Loss of SMARCA4 expression had a sensitivity of 96.55% and specificity of 100%. Conclusions Loss of SMARCA4 expression is sensitive and specific for SCCOHT. Although some mimics show heterogeneous expression, there is retention of nuclear staining in at least a part of the tumour; therefore, only complete loss of staining should be regarded as being supportive of SCCOHT. [ABSTRACT FROM AUTHOR]

Objective Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is an aggressive tumor, with long term survival at ~ 30% in early stage disease. SCCOHT is caused by germline and somatic SMARCA4 mutations, but the effect of the mutation type on patients remains unknown. Furthermore, the rarity of SCCOHT has resulted in varied treatment, with no standardized protocols. We analyzed 293 cases to determine the effect of treatment modalities and SMARCA4 mutations on patient diagnosis and outcome. Methods In 293 SCCOHT patients we collected information on age and stage at diagnosis, treatment modality (surgery, chemotherapy, radiotherapy, and/or high-dose chemotherapy with autologous stem cell rescue (HDC-aSCR)), SMARCA4 mutation origin (germline/somatic), and overall survival. Cox analysis and log-rank tests were performed on 257 cases with available survival data. Results The strongest prognostic factors were stage at diagnosis ( p = 2.72e‐ 15) and treatment modality ( p = 3.87e‐13). For FIGO stages II–IV, 5-year survival was 71% for patients who received HDC-aSCR, compared to 25% in patients who received conventional chemotherapy alone following surgery ( p = 0.002). Patients aged ≥ 40 had a worse outcome than younger patients ( p = 0.04). Twenty-six of 60 tested patients carried a germline SMARCA4 mutation, including all patients diagnosed < 15 years; carriers presented at a younger age than non-carriers ( p = 0.02). Conclusions Stage at diagnosis is the most significant prognostic factor in SCCOHT and consolidation with HDC-aSCR may provide the best opportunity for long-term survival. The large fraction of SMARCA4 germline mutations carriers warrants genetic counseling for all patients. [ABSTRACT FROM AUTHOR]

Many women with ovarian endometrioid carcinoma present with concurrent endometrial carcinoma. Organ-confined and low-grade synchronous endometrial and ovarian tumors (SEOs) clinically behave as independent primary tumors rather than a single advanced-stage carcinoma. We used 18 SEOs to investigate the ancestral relationship between the endometrial and ovarian components. Based on both targeted and exome sequencing, 17 of 18 patient cases of simultaneous cancer of the endometrium and ovary from our series showed evidence of a clonal relationship, ie, primary tumor and metastasis. Eleven patient cases fulfilled clinicopathological criteria that would lead to classification as independent endometrial and ovarian primary carcinomas, including being of FIGO stage T1a/1A, with organ-restricted growth and without surface involvement; 10 of 11 of these cases showed evidence of clonality. Our observations suggest that the disseminating cells amongst SEOs are restricted to physically accessible and microenvironment-compatible sites yet remain indolent, without the capacity for further dissemination. [ABSTRACT FROM AUTHOR]

We have sequenced the genomes of 110 small cell lung cancers (SCLC), one of the deadliest human cancers. In nearly all the tumours analysed we found bi-allelic inactivation of TP53 and RB1, sometimes by complex genomic rearrangements. Two tumours with wild-type RB1 had evidence of chromothripsis leading to overexpression of cyclin D1 (encoded by the CCND1 gene), revealing an alternative mechanism of Rb1 deregulation. Thus, loss of the tumour suppressors TP53 and RB1 is obligatory in SCLC. We discovered somatic genomic rearrangements of TP73 that create an oncogenic version of this gene, TP73Δex2/3. In rare cases, SCLC tumours exhibited kinase gene mutations, providing a possible therapeutic opportunity for individual patients. Finally, we observed inactivating mutations in NOTCH family genes in 25% of human SCLC. Accordingly, activation of Notch signalling in a pre-clinical SCLC mouse model strikingly reduced the number of tumours and extended the survival of the mutant mice. Furthermore, neuroendocrine gene expression was abrogated by Notch activity in SCLC cells. This first comprehensive study of somatic genome alterations in SCLC uncovers several key biological processes and identifies candidate therapeutic targets in this highly lethal form of cancer. [ABSTRACT FROM AUTHOR]

Background: Mucinous ovarian tumors represent a distinct histotype of epithelial ovarian cancer. The rarest (2-4 % of ovarian carcinomas) of the five major histotypes, their genomic landscape remains poorly described. We undertook hotspot sequencing of 50 genes commonly mutated in human cancer across 69 mucinous ovarian tumors. Our goals were to establish the overall frequency of cancer-hotspot mutations across a large cohort, especially those tumors previously thought to be "RAS-pathway alteration negative", using highly-sensitive next-generation sequencing as well as further explore a small number of cases with apparent heterogeneity in RAS-pathway activating alterations. Methods: Using the Ion Torrent PGM platform, we performed next generation sequencing analysis using the v2 Cancer Hotspot Panel. Regions of disparate ERBB2-amplification status were sequenced independently for two mucinous carcinoma (MC) cases, previously established as showing ERBB2 amplification/overexpression heterogeneity, to assess the hypothesis of subclonal populations containing either KRAS mutation or ERBB2 amplification independently or simultaneously. Results: We detected mutations in KRAS, TP53, CDKN2A, PIK3CA, PTEN, BRAF, FGFR2, STK11, CTNNB1, SRC, SMAD4, GNA11 and ERBB2. KRAS mutations remain the most frequently observed alteration among MC (64.9 %) and mucinous borderline tumors (MBOT) (92.3 %). TP53 mutation occurred more frequently in carcinomas than borderline tumors (56.8 % and 11.5 %, respectively), and combined IHC and mutation data suggest alterations occur in approximately 68 % of MC and as many as 20 % of MBOT. Proven and potential RAS-pathway activating changes were observed in all but one MC. Concurrent ERBB2 amplification and KRAS mutation were observed in a substantial number of cases (7/63 total), as was co-occurrence of KRAS and BRAF mutations (one case). Microdissection of ERBB2-amplified regions of tumors harboring KRAS mutation suggests these alterations are occurring in the same cell populations, while consistency of KRAS allelic frequency in both ERBB2 amplified and non-amplified regions suggests this mutation occurred in advance of the amplification event. Conclusions: Overall, the prevalence of RAS-alteration and striking co-occurrence of pathway "double-hits" supports a critical role for tumor progression in this ovarian malignancy. Given the spectrum of RAS-activating mutations, it is clear that targeting this pathway may be a viable therapeutic option for patients with recurrent or advanced stage mucinous ovarian carcinoma, however caution should be exercised in selecting one or more personalized therapeutics given the frequency of non-redundant RAS-activating alterations. [ABSTRACT FROM AUTHOR]

Objectives: Relatively little is known about the genomic landscape of vulvovaginal squamous cell carcinoma (VSC). We sought to explore this landscape and the possibility of molecularly driven outcomes in a large database of VSC. Methods: VSC tumor samples were analyzed using next-generation sequencing of DNA (TruSeq, 45 genes; NextSeq, 592 genes and NovaSeq, whole-exome sequencing [WES]) and RNA (NovaSeq). PD-L1 expression was tested by immunohistochemistry (IHC) using SP142. Microsatellite instability (MSI) was tested by fragment analysis, IHC, and NGS. Tumor mutational burden (TMB) was measured by counting all somatic mutations per tumor (TMB-high: ≥ 10 mutations/MB). HPV16/18 status was determined using WES. Real-world overall survival (OS) was obtained from insurance claims data and Kaplan-Meier estimates were calculated for molecularly defined patient cohorts as the time of first treatment of Pembrolizumab and Nivolumab (IO) to last day of contact. Significance was tested using Chi-square and adjusted for multiple comparisons (q-value <0.05). Results: A total of 651 VSC tumors were included in this analysis, of which 67% were primary tumors. The top five most commonly mutated genes were: TP53 (45.8%), CDKN2A (23.7%), PIK3CA (22.6%), TERT (19.8%) and KMT2D (12.9%). The top five most commonly active pathways were: TP53 (45.2%), Chromatin Remodeling (27%), PI3K (27%), Cell Cycle (26.3%), and RTK-Ras (25.7%). Among the tumors identified, 78.9% of tumors were PD-L1+ and 1.8% of tumors were MSI-H. Since HPV is linked to the oncogenesis of VSC and 16 and 18 are the most common high-risk variants of HPV, we compared HPV16/18+ VSC tumors (n =62) with VSC tumors that were both HPV16/18-negative and TP53 -mutant (n =59). CDKN2A mutations were exclusive to HPV16/18-/ TP53mut VSC tumors (49.2%, q<0.001). HPV16/18+ tumors had significantly increased alterations in the PI3K pathway (45.2% vs 11.9%) but lower alterations in the Telomerase Maintenance (9.7% vs 61%) and Cell Cycle (8.1% vs 54.2%) pathways (q<0.001). CDKN2A (HR: 3.26, 95% CI: 0.94-11.2, p=0.049) and TP53 (HR: 3.62, 95% CI: 1.07-12.2, p=0.028) mutations each were significantly associated with worse OS after IO. Combined survival analysis of VSC patients with CDKN2A or TP53 mutations showed a trend toward worse survival after IO compared to CDKN2A and TP53 wildtype (WT) VSC patients (HR: 3.12, 95% CI: 0.92-10.6, p=0.054) (Figure 1). Interestingly, CDKN2A and TP53 WT VSC patients had improved OS with IO treatment compared to non-IO treatments (HR: 0.37, 95% CI: 0.14-1.03, p=0.047), but there was no such improvement in OS in patients with either a CDKN2A or TP53 mutation in their VSC (p=0.985). Conclusions: HPV16/18+ VSC has significantly higher PI3K pathway activity, while HPV16/18-/ TP53 mut VSC has a significantly higher activity of Telomerase Maintenance and Cell Cycle pathways, suggesting the potential of targeted therapy. TP53 and CDKN2A mutations in VSC appear limited to HPV16/18-negative tumors and might signal the lack of benefit from IO treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

: The base excision repair (BER) that repairs oxidative damage is upregulated as an adaptive response in maintaining tumorigenesis of RAS-transformed cancer cells. [ABSTRACT FROM AUTHOR]

The principal reason for failure of targeted cancer therapies is the emergence of resistant clones that regenerate the tumor. Therapeutic efficacy therefore depends on not only how effectively a drug inhibits its target, but also the innate or adaptive functional redundancy of that target and its attendant pathway. In this regard, the Myc transcription factors are intriguing therapeutic targets because they serve the unique and irreplaceable role of coordinating expression of the many diverse genes that, together, are required for somatic cell proliferation. Furthermore, Myc expression is deregulated in most "perhaps all" cancers, underscoring its irreplaceable role in proliferation. We previously showed in a preclinical mouse model of non-small-cell lung cancer that systemic Myc inhibition using the dominant-negative Myc mutant Omomyc exerts a dramatic therapeutic impact, triggering rapid regression of tumors with only mild and fully reversible side effects. Using protracted episodic expression of Omomyc, we now demonstrate that metronomic Myc inhibition not only contains Ras-driven lung tumors indefinitely, but also leads to their progressive eradication. Hence, Myc does indeed serve a unique and nondegenerate role in lung tumor maintenance that cannot be complemented by any adaptive mechanism, even in the most aggressive p53-deficient tumors. These data endorse Myc as a compelling cancer drug target. [ABSTRACT FROM AUTHOR]

A wealth of evidence has now demonstrated that the microenvironment in which a tumorigenic cell evolves is as critical to its evolution as the genetic mutations it accrues. However, there is still relatively little known about how signals from the microenvironment contribute to the early events in the progression to malignancy. To address this question, we used a premalignant mammary model to examine how fibroblasts, and the extracellular matrix (ECM) proteins they secrete, influence progression to malignancy. Their effect on metastatic malignant cells was also assessed for comparison. We found that carcinoma-associated fibroblasts, and the distinct aligned ECM they deposit, can cause both premalignant and malignant mammary epithelial cells to assume a mesenchymal morphology that is associated with increased dissemination and metastasis, while benign reduction mammoplasty fibroblasts favor the maintenance of an epithelial morphology and constrain early dissemination, tumor growth, and metastasis. Our results suggest that normalizing the organization of the ECM could be effective in limiting systemic dissemination and tumor growth. [ABSTRACT FROM AUTHOR]

The ubiquitous deregulation of Myc in human cancers makes it an intriguing therapeutic target, a notion supported by recent studies in Ras-driven lung tumors showing that inhibiting endogenous Myc triggers ubiquitous tumor regression. However, neither the therapeutic mechanism nor the applicability of Myc inhibition to other tumor types driven by other oncogenic mechanisms is established. Here, we show that inhibition of endogenous Myc also triggers ubiquitous regression of tumors in a simian virus 40 (SV40)-driven pancreatic islet tumor model. Such regression is presaged by collapse of the tumor microenvironment and involution of tumor vasculature. Hence, in addition to its diverse intracellular roles, endogenous Myc serves an essential and nonredundant role in coupling diverse intracellular oncogenic pathways to the tumor microenvironment, further bolstering its credentials as a pharmacological target. [ABSTRACT FROM AUTHOR]

Small-cell lung cancer (SCLC) is an aggressive neuroendocrine subtype of lung cancer for which there is no effective treatment. Using a mouse model in which deletion of Rb1 and Trp53 in the lung epithelium of adult mice induces SCLC, we found that the Hedgehog signaling pathway is activated in SCLC cells independently of the lung microenvironment. Constitutive activation of the Hedgehog signaling molecule Smoothened (Smo) promoted the clonogenicity of human SCLC in vitro and the initiation and progression of mouse SCLC in vivo. Reciprocally, deletion of Smo in Rb1 and Trp53-mutant lung epithelial cells strongly suppressed SCLC initiation and progression in mice. Furthermore, pharmacological blockade of Hedgehog signaling inhibited the growth of mouse and human SCLC, most notably following chemotherapy. These findings show a crucial cell-intrinsic role for Hedgehog signaling in the development and maintenance of SCLC and identify Hedgehog pathway inhibition as a therapeutic strategy to slow the progression of disease and delay cancer recurrence in individuals with SCLC. [ABSTRACT FROM AUTHOR]

Summary: The molecular basis for p53-mediated tumor suppression remains unclear. Here, to elucidate mechanisms of p53 tumor suppression, we use knockin mice expressing an allelic series of p53 transcriptional activation mutants. Microarray analysis reveals that one mutant, p5325,26, is severely compromised for transactivation of most p53 target genes, and, moreover, p5325,26 cannot induce G1-arrest or apoptosis in response to acute DNA damage. Surprisingly, p5325,26 retains robust activity in senescence and tumor suppression, indicating that efficient transactivation of the majority of known p53 targets is dispensable for these pathways. In contrast, the transactivation-dead p5325,26,53,54 mutant cannot induce senescence or inhibit tumorigenesis, like p53 nullizygosity. Thus, p53 transactivation is essential for tumor suppression but, intriguingly, in association with a small set of novel p53 target genes. Together, our studies distinguish the p53 transcriptional programs involved in acute DNA-damage responses and tumor suppression—a critical goal for designing therapeutics that block p53-dependent side effects of chemotherapy without compromising p53 tumor suppression. [ABSTRACT FROM AUTHOR]

Mutational activation of BRAF is the earliest and most common genetic alteration in human melanoma. To build a model of human melanoma, we generated mice with conditional melanocyte-specific expression of BRafV600E. Upon induction of BRafV600E expression, mice developed benign melanocytic hyperplasias that failed to progress to melanoma over 15–20 months. By contrast, expression of BRafV600E combined with Pten tumor suppressor gene silencing elicited development of melanoma with 100% penetrance, short latency and with metastases observed in lymph nodes and lungs. Melanoma was prevented by inhibitors of mTorc1 (rapamycin) or MEK1/2 (PD325901) but, upon cessation of drug administration, mice developed melanoma, indicating the presence of long-lived melanoma-initiating cells in this system. Notably, combined treatment with rapamycin and PD325901 led to shrinkage of established melanomas. These mice, engineered with a common genetic profile to human melanoma, provide a system to study melanoma's cardinal feature of metastasis and for preclinical evaluation of agents designed to prevent or treat metastatic disease. [ABSTRACT FROM AUTHOR]

Summary: Deregulated Myc triggers a variety of intrinsic tumor suppressor programs that serve to restrain Myc''s oncogenic potential. Since Myc activity is also required for normal cell proliferation, activation of intrinsic tumor suppression must be triggered only when Myc signaling is oncogenic. However, how cells discriminate between normal and oncogenic Myc is unknown. Here we show that distinct threshold levels of Myc govern its output in vivo: low levels of deregulated Myc are competent to drive ectopic proliferation of somatic cells and oncogenesis, but activation of the apoptotic and ARF/p53 intrinsic tumor surveillance pathways requires Myc overexpression. The requirement to keep activated oncogenes at a low level to avoid engaging tumor suppression is likely an important selective pressure governing the early stages of tumor microevolution. [Copyright &y& Elsevier]

Myc is a pleiotropic basic helix–loop–helix leucine zipper transcription factor that coordinates expression of the diverse intracellular and extracellular programs that together are necessary for growth and expansion of somatic cells. In principle, this makes inhibition of Myc an attractive pharmacological approach for treating diverse types of cancer. However, enthusiasm has been muted by lack of direct evidence that Myc inhibition would be therapeutically efficacious, concerns that it would induce serious side effects by inhibiting proliferation of normal tissues, and practical difficulties in designing Myc inhibitory drugs. We have modelled genetically both the therapeutic impact and the side effects of systemic Myc inhibition in a preclinical mouse model of Ras-induced lung adenocarcinoma by reversible, systemic expression of a dominant-interfering Myc mutant. We show that Myc inhibition triggers rapid regression of incipient and established lung tumours, defining an unexpected role for endogenous Myc function in the maintenance of Ras-dependent tumours in vivo. Systemic Myc inhibition also exerts profound effects on normal regenerating tissues. However, these effects are well tolerated over extended periods and rapidly and completely reversible. Our data demonstrate the feasibility of targeting Myc, a common downstream conduit for many oncogenic signals, as an effective, efficient and tumour-specific cancer therapy. [ABSTRACT FROM AUTHOR]

High-grade serous carcinoma (HGSC), the most lethal subtype of epithelial ovarian cancer (EOC), is characterized by widespread TP53 mutations (>90%), most of which are missense mutations (>70%). The objective of this study was to investigate differential transcriptional targets affected by a common germline P72R SNP (rs1042522) in two p53 hotspot mutants, R248Q and R248W, and identify the mechanism through which the P72R SNP affects the neomorphic properties of these mutants. Using isogenic cell line models, transcriptomic analysis, xenografts, and patient data, we found that the P72R SNP modifies the effect of p53 hotspot mutants on cellular morphology and invasion properties. Most importantly, RNA sequencing studies identified CXCL1 a critical factor that is differentially affected by P72R SNP in R248Q and R248W mutants and is responsible for differences in cellular morphology and functional properties observed in these p53 mutants. We show that the mutants with the P72 SNP promote a reversion of the EMT phenotype to epithelial characteristics, whereas its R72 counterpart promotes a mesenchymal transition via the chemokine CXCL1. These studies reveal a new role of the P72R SNP in modulating the neomorphic properties of p53 mutants via CXCL1, which has significant implications for tumor invasion and metastasis. [ABSTRACT FROM AUTHOR]