Alterations In Endometrial Cancer Research Articles

Inhibition of ATM with KU-55933 Sensitizes Endometrial Cancer Cell Lines to Olaparib.

Endometrial cancer (EC) is one of the most prevalent gynecologic cancers, which poses a serious threat to women's health worldwide. Olaparib, the first FDA-approved PARP inhibitor for the treatment of BRCA-mutated breast, ovarian and pancreatic cancers, triggers apoptosis of cancer cells through synthetic lethality by inhibiting PARP1/2 enzymatic activity and BRCA1/2-dependent homologous recombination (HR) repair deficiency. However, the synergistic lethal effects between Olaparib and inhibitors of other DNA damage response proteins, such as ATM, PTEN and RAD51, are still unknown. Exploring the synergistic lethal effect between Olaparib and KU-55933 on EC. The GEPIA database was used to test EC patient survival rate. CCK8 was used for cell viability assays. Western blot was used for examining gene levels. The wound healing assay was used to detect cell migration ability. Flow cytometry was used for detecting the apoptosis rate. All experimental conditions were repeated independently in triplicate and analyzed in three separate experiments. In this study, we discovered that the frequency of ATM alterations in endometrial cancer reaches nearly 20% and that there is a positive correlation between ATM alterations and prognosis. Furthermore, we discovered that endometrial cells with low expression levels of ATM are sensitive to Olaparib. Treatment with KU-55933, a specific inhibitor of ATM, significantly enhanced the sensitivity of endometrial cancer cells to Olaparib, as evidenced by colony formation, cell migration and apoptosis assay. Further analysis revealed that KU-55933 potentiates Olaparib-induced cell apoptosis by inhibiting ATM phosphorylation. Our study demonstrates that inhibiting ATM could enhance the sensitivity of endometrial cancer to Olaparib, thereby providing a potential alternative treatment for the clinical treatment of endometrial cancer.

Role of genetic ancestry in endometrial cancers: Understanding disparities in black women.

e17631 Background: Endometrial cancer is the most common gynecological malignancy in the United States. Endometrial cancer disproportionately affects black women in terms of incidence and survival despite advancement in treatment, and preventive strategies. Identifying distinct tumor markers in patients with African ancestry will help distinguish biological determinants underlying the disparities in endometrial cancer. In this study, we investigated the genetic alterations in endometrial cancer in individuals of African (AFR) ancestry using The Cancer Genome Atlas (TCGA). Methods: The genomic and clinical data of TCGA PanCancer Atlas uterine corpus endometrial carcinoma (UCEC) were explored using CBioportal (http://www.cbioportal.org/). We utilized The Cancer Genome Ancestry Atlas (TCGAA) and LAMP for estimates of genetic ancestry and quantitative ancestral composition. This dataset contains 529 patients, including 357 self-reported whites and 107 blacks. For each case the proportion of European, West African, East Asian, Native American ancestry was estimated. The dominant ancestry was defined as ≥50% of admixture from one reference population. Differences in gene mutation frequency were analyzed based on AFR ancestry proportion. The Kaplan-Meier curves were generated, and Cox regression analyses were performed. Results: Global genetic ancestry analysis identified 115 AFR ancestry cases with mean ancestry of 80.4%. The dominant AFR ancestry subject matched the self-reported race with 94% accuracy. We identified 23 subjects with ≥90% AFR ancestry, 43 subjects with 80-90% AFR ancestry, 34 subjects with 70-80% AFR ancestry, and 15 subjects with 50-70% AFR ancestry. TP53 was the most frequently mutated gene in patients with AFR ancestry(49.5%). ≥90% AFR ancestry had the highest rate of TP53 mutations (52.2%) compared with 80-90% AFR ancestry (51.2%), 70-80% AFR ancestry (43.8%), and 50-70% AFR ancestry (41.7%). PTEN was less commonly mutated in patients with higher proportion of AFR ancestry (47.8% in ≥90% AFR ancestry, 39.0% in 80-90% AFR ancestry, 59.4% in 70-80% AFR ancestry, and 66.7% in 50-70% AFR ancestry). PIK3CA mutation frequency was almost identical in all AFR ancestry groups studied. No significant differences in overall survival was observed between AFR ancestry groups. Conclusions: We aimed to investigate the etiology of endometrial cancer disparities by analyzing the effect of ancestry on genetic alterations in endometrial cancer. This study demonstrated differences in the mutation frequency among patients with AFR ancestry. We have shown that TP53 mutations were seen more frequently in patients with higher AFR ancestry, which may confer a poorer prognosis. Nonetheless, to validate the findings of this study, future studies should be conducted with larger sample sizes and more diverse ancestral groups to explore how genetic ancestry impacts tumorigenesis and cancer progression.

An MRI-Based Radiomic Prognostic Index Predicts Poor Outcome and Specific Genetic Alterations in Endometrial Cancer.

Integrative tumor characterization linking radiomic profiles to corresponding gene expression profiles has the potential to identify specific genetic alterations based on non-invasive radiomic profiling in cancer. The aim of this study was to develop and validate a radiomic prognostic index (RPI) based on preoperative magnetic resonance imaging (MRI) and assess possible associations between the RPI and gene expression profiles in endometrial cancer patients. Tumor texture features were extracted from preoperative 2D MRI in 177 endometrial cancer patients. The RPI was developed using least absolute shrinkage and selection operator (LASSO) Cox regression in a study cohort (n = 95) and validated in an MRI validation cohort (n = 82). Transcriptional alterations associated with the RPI were investigated in the study cohort. Potential prognostic markers were further explored for validation in an mRNA validation cohort (n = 161). The RPI included four tumor texture features, and a high RPI was significantly associated with poor disease-specific survival in both the study cohort (p < 0.001) and the MRI validation cohort (p = 0.030). The association between RPI and gene expression profiles revealed 46 significantly differentially expressed genes in patients with a high RPI versus a low RPI (p < 0.001). The most differentially expressed genes, COMP and DMBT1, were significantly associated with disease-specific survival in both the study cohort and the mRNA validation cohort. In conclusion, a high RPI score predicts poor outcome and is associated with specific gene expression profiles in endometrial cancer patients. The promising link between radiomic tumor profiles and molecular alterations may aid in developing refined prognostication and targeted treatment strategies in endometrial cancer.

Molecular expression characteristics confirm the malignancy concealed by morphological alterations in endometrial cancer after fertility-preserving treatment

Fertility-preserving treatment (FPT) has been widely used for young patients with early stage endometrial cancer (EC). However, the literature on the effectiveness and safety of FPT remains controversial. The aim of this study was to investigate malignant transformation in EC after FPT by immunohistochemistry (IHC). A retrospective analysis of pre- and post-treatment biopsy specimens from 24 patients with grade 1 endometrioid adenocarcinoma (EAC) or complex atypical hyperplasia (CAH) was performed. The expression levels of ARID1A, PTEN, and β-catenin were assessed by IHC. The protein expression levels of ARID1A, PTEN, and β-catenin were not significantly different between pre- and post-treatment specimens. However, there was a significant difference between pre-treatment and normal specimens as well as between post-treatment and normal specimens. The protein expression of β-catenin was significantly increased in patients with progression compared with those without progression after FPT. The morphologic normalization of patients with EC after FPT may not be accompanied by the absence of tumor malignancy, and β-catenin may serve as a biomarker for the response to FPT. These results may contribute to a better understanding of the malignant transformation of EC after FPT and the optimization of treatment strategies for young patients with birth plans.

Molecular Modifiers of Hormone Receptor Action: Decreased Androgen Receptor Expression in Mismatch Repair Deficient Endometrial Endometrioid Adenocarcinoma.

Endometrial endometrioid carcinoma is related to estrogen excess and expression of estrogen and progesterone receptors. Epidemiological evidence suggests that exposure to elevated androgens, as in polycystic ovarian syndrome, increases the risk of endometrial cancer. Factors impacting androgen receptor (AR) expression are not well studied. Mismatch repair (MMR) deficiency due to MLH1 gene methylation is one of the most common molecular alterations in endometrial cancer, occurring in 15% to 20% of cases. MLH1 methylation can be associated with decreased expression of other genes, so we examined the effect of MMR status on AR expression. As NF-κB is known to induce AR, this transcription factor was also examined. Three hundred forty-four unselected endometrial carcinomas were evaluated for DNA MMR. Loss of expression of MLH1 with MLH1 methylation was defined as MMR deficient, and positive expression of MMR proteins was defined as MMR intact. A case-control cohort of 96 grade 2 endometrioid carcinomas was studied from this set (47 MMR deficient, 49 MMR intact). Cases were matched for histotype, grade, and age. AR and NF-κB immunohistochemical expression were evaluated by 2 different scoring systems (CAP/ASCO and Allred) used for estrogen receptor. Despite higher levels of NF-κB, MMR deficiency was associated with a significantly lower mean percentage of AR expression. The MMR deficient group had more variable AR expression, with more cases scoring on the lower end of the spectrum. These findings have implications for clinical trials of AR antagonists in gynecologic cancers.

CTCF Expression is Essential for Somatic Cell Viability and Protection Against Cancer.

CCCTC-binding factor (CTCF) is a conserved transcription factor that performs diverse roles in transcriptional regulation and chromatin architecture. Cancer genome sequencing reveals diverse acquired mutations in CTCF, which we have shown functions as a tumour suppressor gene. While CTCF is essential for embryonic development, little is known of its absolute requirement in somatic cells and the consequences of CTCF haploinsufficiency. We examined the consequences of CTCF depletion in immortalised human and mouse cells using shRNA knockdown and CRISPR/Cas9 genome editing as well as examined the growth and development of heterozygous Ctcf (Ctcf+/−) mice. We also analysed the impact of CTCF haploinsufficiency by examining gene expression changes in CTCF-altered endometrial carcinoma. Knockdown and CRISPR/Cas9-mediated editing of CTCF reduced the cellular growth and colony-forming ability of K562 cells. CTCF knockdown also induced cell cycle arrest and a pro-survival response to apoptotic insult. However, in p53 shRNA-immortalised Ctcf+/− MEFs we observed the opposite: increased cellular proliferation, colony formation, cell cycle progression, and decreased survival after apoptotic insult compared to wild-type MEFs. CRISPR/Cas9-mediated targeting in Ctcf+/− MEFs revealed a predominance of in-frame microdeletions in Ctcf in surviving clones, however protein expression could not be ablated. Examination of CTCF mutations in endometrial cancers showed locus-specific alterations in gene expression due to CTCF haploinsufficiency, in concert with downregulation of tumour suppressor genes and upregulation of estrogen-responsive genes. Depletion of CTCF expression imparts a dramatic negative effect on normal cell function. However, CTCF haploinsufficiency can have growth-promoting effects consistent with known cancer hallmarks in the presence of additional genetic hits. Our results confirm the absolute requirement for CTCF expression in somatic cells and provide definitive evidence of CTCF’s role as a haploinsufficient tumour suppressor gene. CTCF genetic alterations in endometrial cancer indicate that gene dysregulation is a likely consequence of CTCF loss, contributing to, but not solely driving cancer growth.

Abstract 5728: Metabolomic pathway alterations in endometrial cancer

Abstract Introduction: Endometrial cancer is the most frequent gynecological cancer diagnosed in United States, accounting for 76,000 deaths every year. Up to now, it has been broadly described the proteomic and the transcriptomic profiles of endometrial cancer (EC). However, the metabolomic content of EC tissues is largely unknown. In this work, we aim to characterize the pathways dysregulated in EC tumorigenesis and the changes that the metabolites experience along the progression of the disease. Methods: We performed an untargeted discovery using liquid chromatography coupled with electro-spray quadrupole time of flight mass spectrometry (UPLC-ESI-QTOF-MS). We analyzed the metabolomic and lipidomic profile of a set of 56 tumor tissues, corresponding to 39 different stages of endometrioid EC tumors (10 FIGO stages IA, 9 stages IB, 10 stages II and 10 stages III), and 17 benign endometrium tissues. Verification of initiation and progression markers was performed by MS/MS. Furthermore the dysregulated pathways were validated by Real Time quantitative PCR (RTqPCR) and Immunohistochemistry (IHQ) in an independent cohort of 60 and 109 patients, respectively. Results: A total of 8,146 features were detected in the positive and 7,558 in the negative electrospray ionization mode. Among those, a list of 40 significant differentially expressed metabolites (FC&amp;gt;2) was verified. The lipid metabolism appeared highly upregulated; with an important number of PCs, PSs, PEs, PIs and PGs. Linoleic acid, 3-Deoxyvitamin D3, and UDP-N-acetyl-D-galactosamine were also upregulated in EC. Differently, the peptide Glu Phe Arg Trp, some amides (palmitamide, stearamide and oleamide), PA (18:0/18:1), PE (20:1/22:6), PE (22:6/P-18:1), PG (19:0/22:4), inosine and picolinic acid were downregulated in the tumor tissue. Regarding tumoral progression, we showed a significant and non-gradual dysregulation of PC (16:0/20:5), the PC (16:0/22:6), PE (16:0/22:6), PE (22:6/P-18:1) and PE (18:1/22:6) at different stages of cancer progression. A significant decrease of the levels of UDP-N-acetyl-D-galactosamine and Arachidonic acid at advanced stages of the tumor was detected. We confirmed by RTqPCR and by IHQ the downregulation of the 3 adenosine-to-inosine editing enzymes (ADAR1, ADAR2 and ADARB3) in EC patients compared to the control group. We also observed a higher expression of the enzymes in the most aggressive (serous EC tumors) compared to the less aggressive tumors (endometrioid EC tumors). We are currently performing studies to understand the role of the RNA editing pathway in EC. Conclusions: In this study we described the metabolome and lipidome of EC initiation and progression. We verified and validated several metabolites that are significantly differentially found in control subjects compared to EC patients and along tumoral progression. These results also depictured the metabolism of EC and deciphered, for the first time, an alteration in the adenosine-to-inosine editing pathway in EC. Note: This abstract was not presented at the meeting. Citation Format: Tatiana Altadill. Metabolomic pathway alterations in endometrial cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5728. doi:10.1158/1538-7445.AM2017-5728

Genetic alterations in endometrial cancer by targeted next-generation sequencing

Many genetic factors play important roles in the development of endometrial cancer. The aim of this study was to investigate genetic alterations in the Taiwanese population with endometrial cancer. DNA was extracted from 10 cases of fresh-frozen endometrial cancer tissue. The exomes of cancer-related genes were captured using the NimbleGen Comprehensive Cancer Panel (578 cancer-related genes) and sequenced using the Illumina Genomic Sequencing Platform. Our results revealed 120 variants in 99 genes, 21 of which were included in the Oncomine Cancer Research Panel used in the National Cancer Institute Match Trial. The 21 genes comprised 8 tumor suppressor candidates (ATM, MSH2, PIK3R1, PTCH1, PTEN, TET2, TP53, and TSC1) and 13 oncogene candidates (ALK, BCL9, CTNNB1, ERBB2, FGFR2, FLT3, HNF1A, KIT, MTOR, PDGFRA, PPP2R1A, PTPN11, and SF3B1). We identified a high frequency of mutations in PTEN (50%) and genes involved in the endometrial cancer-related molecular pathway, which involves the IL-7 signaling pathway (PIK3R1, n=1; AKT2, n=1; FOXO1, n=1). We report the mutational landscape of endometrial cancer in the Taiwanese population. We believe that this study will shed new light on fundamental aspects for understanding the molecular pathogenesis of endometrial cancer and may aid in the development of new targeted therapies.

Increased angiogenesis is associated with a 32-gene expression signature and 6p21 amplification in aggressive endometrial cancer.

Angiogenesis is a hallmark of cancer. The aim of this study was to explore whether microvessel proliferation is associated with gene expression profiles or copy number alterations in endometrial cancer. A prospective series of endometrial carcinomas was studied for angiogenesis markers, gene expression profiles, and gene copy number data. For validation, an independent series of endometrial carcinomas as well as an external cohort of endometrial cancer patients were examined by gene expression microarrays. Increased microvessel proliferation (MVP) was associated with aggressive tumor features and reduced survival, and a 32-gene expression signature was found to separate tumors with high versus low MVP. An increased 32-gene signature score was confirmed to associate with high-grade tumor features and reduced survival by independent cohorts. Copy number studies revealed that amplification of the 6p21 region was significantly associated with MVP, a high 32-gene score, as well as reduced survival. Increased MVP was significantly associated with aggressive endometrial cancer and reduced survival. Integrated analyses demonstrated significant associations between increased vascular proliferation, amplification of the 6p21 region, VEGF-A mRNA expression, and the 32-gene angiogenesis signature. Our findings indicate amplification of 6p21 as a possible driver of tumor vascular proliferation in endometrial cancer.

Mig-6 suppresses endometrial cancer associated with Pten deficiency and ERK activation.

PTEN mutations are the most common genetic alterations in endometrial cancer. Loss of PTEN and subsequent AKT activation stimulate estrogen receptor α-dependent pathways that play an important role in endometrial tumorigenesis. The major pathologic phenomenon of endometrial cancer is the loss of ovarian steroid hormone control over uterine epithelial cell proliferation and apoptosis. However, the precise mechanism of PTEN/AKT signaling in endometrial cancer remains poorly understood. The progesterone signaling mediator MIG-6 suppresses estrogen signaling and it has been implicated previously as a tumor suppressor in endometrial cancer. In this study, we show that MIG-6 also acts as a tumor suppressor in endometrial cancers associated with PTEN deficiency. Transgenic mice, where Mig-6 was overexpressed in progesterone receptor-expressing cells, exhibited a relative reduction in uterine tumorigenesis caused by Pten deficiency. ERK1/2 was phosphorylated in uterine tumors and administration of an ERK1/2 inhibitor suppressed cancer progression in PR(cre/+)Pten(f/f) mice. In clinical specimens of endometrial cancer, MIG-6 expression correlated inversely with ERK1/2 phosphorylation during progression. Taken together, our findings suggest that Mig-6 regulates ERK1/2 phosphorylation and that it is crucial for progression of PTEN-mutant endometrial cancers, providing a mechanistic rationale for the evaluation of ERK1/2 inhibitors as a therapeutic treatment in human endometrial cancer.

High concordance of molecular tumor alterations between pre-operative curettage and hysterectomy specimens in patients with endometrial carcinoma

ObjectiveMolecular alterations in endometrial cancer have been shown to be prognostically significant but have not yet been implemented in the current clinical risk assessment. Few studies have investigated the reliability of molecular alterations in pre-operative specimens. Therefore, the objective was to determine whether molecular analysis of pre-operative endometrial cancer samples accurately reflects those alterations in the subsequent hysterectomy specimens. MethodsPaired pre-operative and hysterectomy specimens of 48 patients diagnosed with endometrial carcinoma, 42 endometrioid (EEC) and 6 non-endometrioid (NEEC) carcinomas, were analyzed for immunohistochemical expression of p53, PTEN and β-catenin. Tumor DNA was isolated and analyzed for microsatellite instability (MSI), TP53 mutations and somatic hot spot mutations in 13 genes. ResultsIn EEC patients, loss of PTEN, nuclear β-catenin and p53-mutant expression was found in 43%, 7% and 12%, respectively. No nuclear β-catenin was found in 5 of 6 NEEC patients, all serous cancers, whereas a p53-mutant expression was present in all serous cases. MSI was found in 19.5%, all EEC. Concordance for PTEN, β-catenin, p53 expression and MSI status was found in 79%, 92%, 79% and 93.5%, respectively. We detected 65 hot spot mutations in 39/48 (81%) tumors. Overall concordance of the GynCarta multigene analysis was 99.8%. ConclusionsThe results confirm the reliability of immunohistochemical and DNA-based techniques in the evaluation of molecular alterations in pre-operative endometrial specimens and high concordance rates with the definitive hysterectomy specimens. The resulting molecular signature provides initial pre-operative diagnostic information on the status of oncogenic pathways, which may contribute to individualized treatment strategies.

GALR1 Methylation in Vaginal Swabs Is Highly Accurate in Identifying Women With Endometrial Cancer

Endometrial cancer has become the most common gynecological cancer in developed countries. Postmenopausal bleeding is indicative of the disease in only 1 of 10 women with this symptom. A noninvasive tool to identify women with cancer would be highly desirable. We analyzed more than 27,000 CpGs in normal endometrial tissue (n = 23) and endometrial cancers (n = 64) and found that DNA methylation of GALR1 is among the most frequent epigenetic alterations in this cancer. We then developed a real-time polymerase chain reaction-based GALR1 methylation test and applied this test to vaginal swabs from 79 women who presented with postmenopausal bleeding. The receiver operating characteristics area under the curve, describing sensitivity and specificity to correctly identify the 41 women with both premalignant and malignant endometrial changes, was 0.93 (95% confidence interval, 0.87-0.97; P < 0.0001).GALR1 DNA methylation is one of the most common molecular alterations in endometrial cancer, and the presence of GALR1 methylation in vaginal swabs from women with postmenopausal bleeding indicates the presence of endometrial malignancy with a sensitivity of 92.7% and a specificity of 78.9%.

Molecular targets and targeted therapeutics in endometrial cancer

Endometrial cancer is the most common gynaecological malignancy in the western world. Two clinicopathological subtypes are recognized: type I (endometrioid) and type II (nonendometrioid) carcinomas. This review describes the molecular alterations in endometrial cancer and how this knowledge is leading to the development of novel treatments in this area. Molecularly targeted agents have entered clinical trials in endometrial cancer. So far, mechanistic target of rapamycin (mTOR) inhibitors and antiangiogenic agents appear promising and are being pursued further in addition to other targeted approaches. The clinicopathological and molecular heterogeneity of endometrial cancer needs to be taken into account in the design of future clinical trials as well as the incorporation of robust biomarkers for the success of therapeutic strategies in endometrial cancer.

Detection of mosaic pattern of mitochondrial DNA alterations in different populations of cells from the same endometrial tumor

Somatic mitochondrial DNA (mtDNA) alterations including point mutations and microsatellite instability (MSI) have been frequently detected in human cancers. To further explore the extensiveness of mtDNA alterations, we have analyzed the occurrence of somatic mtDNA mutations in different populations of endometrial cancer cells from the same tumor tissues as compared with adjacent non-tumor cells. Laser-captured micro-dissection was used to harvest endometrial cancer cells from separated areas of the same tumor and adjacent normal cells. Total DNA isolated from micro-dissected cells was PCR amplified and analyzed for mtDNA alterations by polyacrylamide gel electrophoresis and DNA sequencing. Multiple mtDNA alterations were detected in different portions of the same tumor. Different populations of endometrial cancer cells carried different patterns of mtDNA mutations. Interestingly, unlike previous reports, most mutations were found to be heteroplasmic. We have demonstrated the occurrence of hyper-variability of mtDNA alterations in a single piece of tumor tissue. Our observations support the hypothesis that the accumulation of mtDNA alterations is random and expands independently. The data presented here showed the heterogeneity of cancer cells in terms of mtDNA alterations in endometrial cancer.

Chromosomal abnormalities and microsatellite instability in sporadic endometrial cancer

Defective DNA mismatch repair and nonfunctional mechanisms controlling the proper progression of the cell cycle have been proposed as being responsible for the genomic instability and accumulation of karyotypic alterations in endometrial cancer (EC). To assess whether numerical chromosomal anomalies (aneuploidy) and microsatellite instability (MSI) might be representative of distinctive tumour behaviour, paraffin-embedded tissue samples from 86 patients with sporadic EC were evaluated by both fluorescence in situ hybridisation (FISH) and microsatellite analysis, using free nuclei and genomic DNAs (respectively). Approximately one-third of the tumours analysed (24/74; 32%) exhibited MSI, whereas 38/86 (44%) of the EC samples displayed aneuploidy. The majority of the unstable cases (15/24; 63%) were from advanced-stage patients. Conversely, 23 (61%) out of the 38 tumours with aneuploidy were from early-stage patients. No apparent correlation was found between MSI and aneuploidy, whereas the immunohistochemical (IHC) analysis revealed that inactivation of the MLH1 mismatch repair gene may be involved in the majority of the MSI+ sporadic ECs. No genetic or cytogenetic alteration analysed here seems to add any significant predictive value to the stage of disease.