Inhibition of nucleophosmin/B23 sensitizes ovarian cancer cells to immune check-point blockade via PD-L1 in ovarian cancer.

BACKGROUND: Immune checkpoint inhibitors (ICIs) that against programmed cell death protein-1 (PD-1) and its ligand PD-L1 have been approved as a promising treatment of many human cancers. However, the responses to these ICIs were limited in patients with ovarian cancer. Studies have indicated that the response to PD-1/PD-L1 blockade might be correlated with the PD-L1 expression level in cancer cells. Nucleophosmin (NPM/B23) was found to be a potential target for immunotherapy. Whether NPM/B23 plays a role in cancer-associated immunity, such as PD-1/PD-L1 axis, and its underlying mechanisms remain largely unknown in ovarian cancer. METHODS: We applied ovarian cancer cell lines as research models. The effect of modulating PD-L1 by NPM/B23 was subsequently confirmed via Western blot, flow cytometry, qRT-PCR, luciferase reporter assays, and immunoprecipitation. Protein stability and ubiquitin assay assays were used to analyze the interplay between NPM/B23 and NF-ĸB/p65 in PD-L1 regulation. The MOSEC/Luc xenograft mouse model was used to validate the role of NPM/B23-PD-L1 through tumor growth in vivo. RESULTS: Our results revealed that NPM/B23 negatively regulates PD-L1 expression via a protein complex with NF-κB/p65 and through an IFN-γ pathway. Moreover, NPM/B23 inhibitor/modulator sensitized ovarian cancer cells to the anti-PD-1 antibody by regulating PD-L1 expression in the immunocompetent mouse model. Compared to anti-PD-1 antibody alone, a combination of anti-PD-1 antibody and NPM/B23 inhibitor/modulator showed reduced tumorigenesis and increased CD8+ T-cell expansion, thus contributing to prolonged survival on MOSEC/Luc-bearing mouse model. CONCLUSION: Targeting NPM/B23 is a novel and potential therapeutic approach to sensitize ovarian cancer cells to immunotherapy.

Metabolomic profiling of maternal plasma identifies inverse associations of acetate and urea with anti-SARS-CoV-2 antibody titers following COVID-19 vaccination during pregnancy.

We conducted a comprehensive metabolomic analysis of plasma samples obtained from pregnant women who displayed varying post-vaccination antibody titers after receiving mRNA-1273-SARS-CoV-2 vaccines. The study involved 62 pregnant women, all of whom had been vaccinated after reaching 24 weeks of gestation. To quantify post-vaccination plasma antibody titers, we employed binding antibody units (BAU) in accordance with the World Health Organization International Standard. Subsequently, we classified the study participants into three distinct BAU/mL categories: those with high titers (above 2000), medium titers (ranging from 1000 to 2000), and low titers (below 1000). Plasma metabolomic profiling was conducted using 1H nuclear magnetic resonance spectroscopy, and the obtained data were correlated with the categorized antibody titers. Notably, in pregnant women exhibiting elevated anti-SARS-CoV-2 antibody titers, reduced plasma concentrations of acetate and urea were observed. A significant negative correlation between these compounds and antibody titers was also evident. An analysis of metabolomics pathways revealed significant inverse associations between antibody titers and four distinct amino acid metabolic pathways: (1) biosynthesis of phenylalanine, tyrosine, and tryptophan; (2) biosynthesis of valine, leucine, and isoleucine; (3) phenylalanine metabolism; and (4) degradation of valine, leucine, and isoleucine. Additionally, an association between the synthesis and degradation pathways of ketone bodies was evident. In conclusion, we identified different metabolic pathways that underlie the diverse humoral responses triggered by COVID-19 mRNA vaccines during pregnancy. Our data hold significant implications for refining COVID-19 vaccination approaches in expectant mothers. KEY MESSAGES : Anti-SARS-CoV-2 antibody titers decline as the number of days since COVID-19 vaccination increases. Anti-SARS-CoV-2 antibody titers are inversely associated with acetate, a microbial-derived metabolite, and urea. Amino acid metabolism is significantly associated with SARS-CoV-2 antibody titers.

Comparison of immediate germline sequencing and multi-step screening for Lynch syndrome detection in high-risk endometrial and colorectal cancer patients.

OBJECTIVE: Lynch syndrome (LS) is a hereditary cancer predisposition syndrome with a significantly increased risk of colorectal and endometrial cancers. Current standard practice involves universal screening for LS in patients with newly diagnosed colorectal or endometrial cancer using a multi-step screening protocol (MSP). However, MSP may not always accurately identify LS cases. To address this limitation, we compared the diagnostic performance of immediate germline sequencing (IGS) with MSP in a high-risk group. METHODS: A total of 31 Taiwanese women with synchronous or metachronous endometrial and colorectal malignancies underwent MSP which included immunohistochemical staining of DNA mismatch repair (MMR) proteins, MLH1 promoter hypermethylation analysis, and germline sequencing to identify pathogenic variants. All patients who were excluded during MSP received germline sequencing for MMR genes to simulate IGS for the detection of LS. RESULTS: Our findings indicate that IGS surpassed MSP in terms of diagnostic yield (29.0% vs. 19.4%, respectively) and sensitivity (90% vs. 60%, respectively). Specifically, IGS successfully identified nine LS cases, which is 50% more than the number detected through MSP. Additionally, germline methylation analysis revealed one more LS case with constitutional MLH1 promoter hypermethylation, bringing the total LS cases to ten (32.3%). Intriguingly, we observed no significant differences in clinical characteristics or overall survival between patients with and without LS in our cohort. CONCLUSION: Our study suggests that IGS may potentially offer a more effective approach compared to MSP in identifying LS among high-risk patients. This advantage is evident when patients have been pre-selected utilizing specific clinical criteria.

Radiomics analysis for the prediction of locoregional recurrence of locally advanced oropharyngeal cancer and hypopharyngeal cancer.

PURPOSE: By radiomic analysis of the postcontrast CT images, this study aimed to predict locoregional recurrence (LR) of locally advanced oropharyngeal cancer (OPC) and hypopharyngeal cancer (HPC). METHODS: A total of 192 patients with stage III-IV OPC or HPC from two independent cohort were randomly split into a training cohort with 153 cases and a testing cohort with 39 cases. Only primary tumor mass was manually segmented. Radiomic features were extracted using PyRadiomics, and then the support vector machine was used to build the radiomic model with fivefold cross-validation process in the training data set. For each case, a radiomics score was generated to indicate the probability of LR. RESULTS: There were 94 patients with LR assigned in the progression group and 98 patients without LR assigned in the stable group. There was no significant difference of TNM staging, treatment strategies and common risk factors between these two groups. For the training data set, the radiomics model to predict LR showed 83.7% accuracy and 0.832 (95% CI 0.72, 0.87) area under the ROC curve (AUC). For the test data set, the accuracy and AUC slightly declined to 79.5% and 0.770 (95% CI 0.64, 0.80), respectively. The sensitivity/specificity of training and test data set for LR prediction were 77.6%/89.6%, and 66.7%/90.5%, respectively. CONCLUSIONS: The image-based radiomic approach could provide a reliable LR prediction model in locally advanced OPC and HPC. Early identification of those prone to post-treatment recurrence would be helpful for appropriate adjustments to treatment strategies and post-treatment surveillance.

Hypoxia-associated genetic signature in ovarian steroid cell tumor NOS.

Steroid cell tumors, not otherwise specified (SCT-NOS), are uncommon ovarian neoplasms accompanied by virilization symptoms due to hyperandrogenism, which are malignant in approximately one-third of the cases. Given the rarity of SCT-NOS, their molecular underpinnings have not yet been studied in depth. In this case series, we performed the first comprehensive analysis of the genetic landscape of this rare ovarian tumor. A detailed clinicopathological description of an index case is also provided. Over a 20-year period, a total of eight patients were seen at our institution. Total nucleic acids (RNA and DNA) were extracted from evaluable formalin-fixed, paraffin-embedded tumor specimens (n = 7) and subjected to TruSight Oncology 500 testing and/or exome sequencing. The results identified pathogenic variants in several hypoxia-related genes - including HIF1A, VHL, SDHB, SRC, IDH2, and FOXO4. As the first comprehensive genetic analysis of SCT-NOS, this study shows that dysregulation in the hypoxia signaling pathway is a key molecular feature of this rare tumor. Clinically, long-term follow-up with periodic measurements of androgen levels should be pursued in all cases since recurrences may occur several years after the initial diagnosis.

Small cell neuroendocrine carcinoma of the cervix: From molecular basis to therapeutic advances.

Small cell neuroendocrine carcinoma of the cervix (SCNECC) is an uncommon but aggressive uterine malignancy, the cause of which is generally associated with human papillomavirus (HPV) infection. A lack of clinical trials and evidence-based treatment guidelines poses therapeutic challenges to this rare tumor. At present, published data remain limited to case series and case reports. While clinical management has traditionally followed those of small cell neuroendocrine (SCNE) lung cancer relying on surgery, chemoradiation, and systemic chemotherapy, the prognosis remains dismal. Immune checkpoint inhibitors (ICIs), such as monoclonal antibodies that target programmed death-1 (PD-1) or programmed death-ligand 1 (PD-L1), atezolizumab and durvalumab have proven effective in extensive-stage SCNE lung cancer. Moreover, pembrolizumab has also proven beneficial effects when added onto chemotherapy in metastatic and recurrent HPV-associated non-SCNE cervical cancer. It holds promise to use ICIs in combination with chemoradiation to improve the clinical outcomes of patients with SCNECC. Future advances in our understanding of SCNECC biology - associated with the study of its genomic and molecular aberrations as well as knowledge from SCNE of lung and other extrapulmonary sites- would be helpful in discovering new molecular targets for drug development. Collaborative efforts and establishment of a SCNECC-specific biobank will be essential to achieve this goal.

Starvation-inactivated MTOR triggers cell migration via a ULK1-SH3PXD2A/TKS5-MMP14 pathway in ovarian carcinoma.

ABBREVIATIONS: AMPK: AMP-activated protein kinase; CHX: cycloheximide; RAD001: everolimus; HBSS: Hanks' balanced salt solution; LC-MS/MS: liquid chromatography-mass spectrometry/mass spectrometry; MMP14: matrix metallopeptidase 14; MTOR: mechanistic target of rapamycin kinase; MAPK: mitogen-activated protein kinase; RB1CC1/FIP200: RB1 inducible coiled-coil 1; PtdIns3P: phosphatidylinositol-3-phosphate; PX: phox homology; SH3: Src homology 3; SH3PXD2A/TKS5: SH3 and PX domains 2A; SH3PXD2A-[6A]: S112A S142A S146A S147A S175A S348A mutant; ULK1: unc-51 like autophagy activating kinase 1.

Targeted next-generation sequencing for the detection of cancer-associated somatic mutations in adenomyosis.

Adenomyosis is a condition characterised by the invasion of endometrial tissues into the uterine myometrium, the molecular pathogenesis of which remains incompletely elucidated. Lesion profiling with next-generation sequencing (NGS) can lead to the identification of previously unanticipated causative genes and the detection of therapeutically actionable genetic changes. Using an NGS panel that included 275 cancer susceptibility genes, this study examined the occurrence and frequency of somatic mutations in adenomyotic tissue specimens collected from 17 women. Extracted DNA was enriched using targeted formalin-fixed paraffin-embedded tissue cores prior to the identification of lesion-specific variants. The results revealed that KRAS and AT-rich interactive domain 1A (ARID1A) were the two most frequently mutated genes (mutation frequencies: 24% and 12%, respectively). Notably, endometrial atypical hyperplasia did not involve adenomyotic areas. We also identified, for the first time, two potentially pathogenic mutations in the F-box/WD repeat-containing protein 7 (FBXW7) and cohesin subunit SA-2 (STAG2) genes. These findings indicate that mutations in the KRAS, ARID1A, FBXW7 and STAG2 genes may play a critical role in the pathogenesis of adenomyosis. Additional studies are needed to assess whether the utilisation of oncogenic driver mutations can inform the surveillance of patients with adenomyosis who had not undergone hysterectomy.Impact statementWhat is already known on this subject? Although somatic point mutations in the KRAS oncogene have been recently detected in adenomyosis, the molecular underpinnings of this condition remains incompletely elucidated. Lesion profiling with next-generation sequencing (NGS) can lead to the identification of previously unanticipated causative genes and the detection of therapeutically actionable genetic changes.What do the results of this study add? The results of NGS revealed that KRAS and AT-rich interactive domain 1A (ARID1A) were the two most frequently mutated genes (mutation frequencies: 24% and 12%, respectively). We also identified, for the first time, two potentially pathogenic mutations in the F-box/WD repeat-containing protein 7 (FBXW7) and cohesin subunit SA-2 (STAG2) genes.What are the implications of these findings for clinical practice and/or further research? The utilisation of oncogenic driver mutations has the potential to inform the surveillance of patients with adenomyosis who had not undergone hysterectomy.

Analysis of endometrial lavage microbiota reveals an increased relative abundance of the plastic-degrading bacteria Bacillus pseudofirmus and Stenotrophomonas rhizophila in women with endometrial cancer/endometrial hyperplasia.

The pathogenic influences of uterine bacteria on endometrial carcinogenesis remain unclear. The aim of this pilot study was to compare the microbiota composition of endometrial lavage samples obtained from women with either endometrial hyperplasia (EH) or endometrial cancer (EC) versus those with benign uterine conditions. We hypothesized that specific microbiota signatures would distinguish between the two groups, possibly leading to the identification of bacterial species associated with endometrial tumorigenesis. A total of 35 endometrial lavage specimens (EH, n = 18; EC, n = 7; metastatic EC, n = 2; benign endometrial lesions, n = 8) were collected from 32 women who had undergone office hysteroscopy. Microbiota composition was determined by sequencing the V3-V4 region of 16S rRNA genes and results were validated by real-time qPCR in 46 patients with EC/EH and 13 control women. Surprisingly, we found that Bacillus pseudofirmus and Stenotrophomonas rhizophila - two plastic-degrading bacterial species - were over-represented in endometrial lavage specimens collected from patients with EC/EH. Using computational analysis, we found that the functional profile of endometrial microbiota in EC/EH was associated with fatty acid and amino acid metabolism. In summary, our hypothesis-generating data indicate that the plastic-degrading bacteria Bacillus pseudofirmus and Stenotrophomonas rhizophila are over-represented within the endometrial lavage microbiota of women with EC/EH living in Taiwan. Whether this may be related to plastic pollution deserves further investigation.

Management and Prognosis of Patients with Recurrent or Persistent/Progressive Uterine Carcinosarcoma.

Uterine carcinosarcoma (UCS) is a highly aggressive gynecologic malignancy. Recurrent or persistent/progressive disease is usually fatal. We aimed to investigate the management and prognosis of these patients. Clinical records of UCS patients from June 1987 to April 2020 were retrospectively reviewed. The stage was re-assigned with the FIGO 2009 staging system. Univariate and multivariate analyses were used to identify the independent predictors of survival after recurrence (SAR) and cancer-specific survival (CSS). Of the 168 patients, 98 experienced treatment failure. The median time to treatment failure (TTF) was 8.1 months (range: 0.0-89.1). The median follow-up time of censored patients was 32.0 months (range: 16.8-170.7). The 5-year SAR rates of those with recurrent or persistent/progressive disease were 7.6%. On multivariate analysis, salvage therapy mainly using radiotherapy (HR 0.27, 95% CI: 0.10-0.71) or chemotherapy (HR 0.41, 95% CI: 0.24-0.72) or chemoradiotherapy (CRT) (HR 0.33, 95% CI: 0.15-0.75) were associated with improved SAR, whereas disseminated recurrence was associated with significantly worse SAR (HR 3.94, 95% CI: 1.67-9.31, p = 0.002). Salvage therapy using radiotherapy or chemotherapy or CRT significantly improved SAR. Surgery significantly improved CSS but not SAR, adjusting for confounding factors.

A case of adult granulosa cell tumor of the ovary with long-term survival after multiple recurrences.

OBJECTIVE: To illustrate the clinical course of a rare case of recurrent adult granulosa cell tumor (AGCT) and discuss the features and management for recurrences. CASE REPORT: A 56-year-old female was first diagnosed with AGCT in 2008 and had uneventful, regular follow-ups until 2013. Recurrence was suspected and proven by computed tomography-guided biopsy. After undergoing complete cytoreductive surgery (CRS) followed by adjuvant megestrol acetate then leuprolide acetate, another recurrence sprouted at the presacral area in 2017. On both occasions, CRS with no visible residual tumor were attained. The patient has remained in complete remission to date with progestin therapy. CONCLUSION: There are currently no standardized tumor markers, imaging exams, or therapies for managing AGCT recurrences. Whole exome sequencing analysis of our patient suggested possible association with triosephosphate isomerase 1 mutation. Regular follow-ups with at least two types of imaging exams and indefinite hormone therapy are crucial for this patient's remission.

Towards less invasive molecular diagnostics for endometrial cancer: massively parallel sequencing of endometrial lavage specimens in women attending for an office hysteroscopy.

We aimed to detect endometrial cancer (EC)-associated mutations in endometrial lavage specimens collected in an office setting and to compare the detected mutations with those identified in tissue samples. Participants included 16 women attending for an office hysteroscopy because of suspected EC between July 2020 and October 2021. Massively parallel sequencing was conducted using the targeted 72 cancer-associated genes. Endometrial lavage specimens, endometrial tissue samples, and blood samples were simultaneously sequenced to establish the concordance of genetic alterations. In this study, the vast majority of EC-associated mutations identified in lavage samples (R2 = 0.948) were identical to those detected in endometrial tissues. Of the 13 patients with EC, 12 (92.3%) had at least one mutation identified in endometrial lavage samples. Notably, no mutations in lavage samples were identified in the two patients with a previous history of EC but no actual endometrial lesions, supporting a high negative predictive value of the test. A patient previously diagnosed with EC and with current evidence of atypical hyperplasia showed persisting PTEN, PIK3R1, and KRAS mutations in her endometrial lavage specimen. PTEN was the most commonly mutated gene, followed by PIK3R1, ARID1A, PIK3CA, CTNNB1, and KRAS. In conclusions, our study provides pilot evidence on the actionability of uterine lavage samples sequencing to detect EC-associated mutations in women with suspected endometrial lesions. In a precision medicine framework, the high mutational concordance between uterine lavage samples and tissue specimens may help inform less invasive diagnostic protocols and the need for ongoing surveillance in patients with EC who wished for fertility-preserving treatment. KEY MESSAGES: • Sequencing of uterine lavage samples collected by office hysteroscopy is feasible. • Most EC mutations identified in lavage were identical to endometrial tissues. • Sequencing of uterine lavage samples may help inform diagnostic protocols for EC. • This approach can be used for recurrence surveillance in patients with EC.

Predicting Hazardous Alcohol Drinking Behaviors in Family Members of Hazardous Alcohol-Drinker Patients.

Family members of hazardous or harmful alcohol drinkers suffer many consequences of their relative's alcohol-drinking behaviors and risk developing their own hazardous alcohol drinking behaviors. Studies of alcohol-related healthcare problems have mainly focused on patients, with few studies on their family members. This cross-sectional study explored factors predicting hazardous alcohol drinking behaviors in family members of hazardous alcohol-drinker patients. Participants were recruited from four randomly chosen hospitals in Taiwan. Data were collected using self-report questionnaires on family members' alcohol use, perceived stress, coping mechanisms, social support, health, quality of life, protective factors against hazardous alcohol drinking, facilitative factors for hazardous alcohol drinking, and demographics. The 318 family members who participated in this study were divided by their Chinese-version Alcohol Use Disorders Identification Test scores into two groups: hazardous alcohol drinkers (score ≥ 8) and non-hazardous alcohol drinkers (score < 8). Significant factors predicting hazardous alcohol drinking behaviors were found by logistic regression to be the frequency of using general coping mechanisms (OR = 1.29, p < 0.01), the frequency of using strategies to cope with patients' drinking-related behaviors (OR = 0.89, p < 0.01), factors protecting against hazardous alcohol drinking (OR = 0.76, p < 0.01) and factors facilitating hazardous alcohol drinking (OR = 1.52, p < 0.01). Interventions should be designed for family members of hazardous alcohol drinkers to address these four significant predictors.

Rare Subtype of Endometrial Cancer: Undifferentiated/Dedifferentiated Endometrial Carcinoma, from Genetic Aspects to Clinical Practice.

Endometrial cancer (EC) is one of the most common gynecologic cancers worldwide. There were 417,367 newly diagnosed cases and 97,370 deaths due to this disease worldwide in 2020. The incidence rates have increased over time, especially in countries with rapid socioeconomic transitions, and EC has been the most prevalent gynecologic malignancy in Taiwan since 2012. The new EC molecular classifications of The Cancer Genome Atlas (TCGA) Research Network include clear-cell carcinoma, serous carcinoma, and carcinosarcoma, while undifferentiated/dedifferentiated EC (UDEC) is not mentioned, and most previous clinical trials for EC have not included UDEC. UDEC is rare, has an aggressive growth pattern, tends to be diagnosed at an advanced stage, and is resistant to conventional chemotherapy. In this review, case series or case reports on the clinical features and genomic/epigenetic and expression profiles on UDEC data are summarized in order to identify potential molecular targets for current and future research.

JAK2-Mediated Phosphorylation of Stress-Induced Phosphoprotein-1 (STIP1) in Human Cells.

Stress-induced phosphoprotein-1 (STIP1)-a heat shock protein (HSP)70/HSP90 adaptor protein-is commonly overexpressed in malignant cells, where it controls proliferation via multiple signaling pathways, including JAK2/STAT3. We have previously shown that STIP1 stabilizes the protein tyrosine kinase JAK2 in cancer cells via HSP90 binding. In this study, we demonstrate that STIP1 may act as a substrate for JAK2 and that phosphorylation of tyrosine residues 134 and 152 promoted STIP1 protein stability, induced its nuclear-cytoplasmic shuttling, and promoted its secretion into the extracellular space. We also found that JAK2-mediated STIP1 phosphorylation enhanced cell viability and increased resistance to cisplatin-induced cell death. Conversely, interference STIP1 with JAK2 interaction-attained either through site-directed mutagenesis or the use of cell-penetrating peptides-decreased JAK2 protein levels, ultimately leading to cell death. On analyzing human ovarian cancer specimens, JAK2 and STIP1 expression levels were found to be positively correlated with each other. Collectively, these results indicate that JAK2-mediated phosphorylation of STIP-1 is critical for sustaining the JAK2/STAT3 signaling pathway in cancer cells.

MicroRNAs as Predictors of Future Uterine Malignancy in Endometrial Hyperplasia without Atypia.

The histological criteria for classifying endometrial hyperplasia (EH) are based on architectural crowding and nuclear atypia; however, diagnostic agreement among pathologists is poor. We investigated molecular biomarkers of endometrial cancer (EC) risk in women with simple hyperplasia or complex hyperplasia without atypia (SH/CH-nonA). Forty-nine patients with EC preceded by SH/CH-nonA were identified, of which 23 were excluded (15 with complex atypical hyperplasia (CAH), six not consenting, one with a diagnosis <6 months prior, and one lost to follow-up). The EH tissues of these patients were compared with those of patients with SH/CH-nonA that did not progress to EC (control) through microRNA (miRNA) array analysis, and the results were verified in an expanded cohort through reverse transcription-quantitative polymerase chain reaction (RT-qPCR). MiRNA arrays analyses revealed 20 miRNAs that differed significantly (p < 0.05, fold change >4) between the control (n = 12) and case (n = 6) patients. Multiplex RT-qPCR for the 20 miRNAs in the expanded cohort (94 control and 25 case patients) led to the validation of miR-30a-3p (p = 0.0009), miR-141 (p < 0.0001), miR-200a (p < 0.0001), and miR-200b (p < 0.0001) as relevant biomarkers, among which miR-141, miR-200a, and miR-200b regulate the expression of phosphatase and tensin homolog (PTEN). For the prediction of EC, the area under the curve for miR-30a-3p, miR-141, miR-200a, and miR-200b was 0.623, 0.754, 0.783, and 0.704, respectively. The percentage of complete PTEN loss was significantly higher in the case group than in the control group (24% vs. 0%, p < 0.001, Fisher's exact test). A combination of complete PTEN loss and miR-200a provided optimal prediction performance (sensitivity = 0.760; specificity = 1.000; positive predictive value = 1.000; negative predictive value = 0.937; accuracy = 0.947). MiR-30a-3p, miR-141, miR-200a, miR-200b, and complete PTEN loss may be useful tissue biomarkers for predicting EC risk among patients with SH/CH-nonA.

Massively parallel sequencing of endometrial lavage specimens for the detection of cancer-associated mutations in atypical and non-atypical endometrial hyperplasia.

Background: Endometrial hyperplasia (EH), particularly with atypia, is considered an antecedent of endometrial adenocarcinoma. In this study, we aimed to apply massively parallel sequencing of endometrial lavage specimens for the detection of cancer-associated mutations in atypical (AEH) and non-atypical endometrial hyperplasia (NEH). The identified alterations were compared with those detected in tissue samples. Materials and methods: Endometrial lavage specimens and parallel biopsy samples (n = 11 for AEH and n = 9 for NEH) were obtained from 18 women (9 with AEH and 9 with NEH) who received an office hysteroscopy for suspected endometrial lesions. All samples were tested for somatic mutations in hotspot regions of 72 cancer-associated genes by massively parallel sequencing. Results: On analyzing sequencing data, the presence of at least one cancer-associated gene mutation was identified in 72.7 and 44.4% of endometrial lavage specimens obtained from women with AEH and NEH, respectively (p = 0.362, 95% confidence interval = 0.72-3.70). The concordance rates between mutations identified in endometrial lavage specimens and endometrial biopsies were 54.5 and 0% from women with AEH and NEH, respectively (p = 0.014). A patient with NEH harbored mutations in endometrial lavage with the same mutations found in the tissue specimen at low allele frequency below detection cutoff, raising the suspicion of missed focal atypia. Conclusion: Endometrial hyperplasia is characterized by a high burden of cancer-associated mutations, particularly in the presence of atypia. Our study, albeit performed with a relatively small number of samples, indicates that their detection by massively parallel sequencing of endometrial lavage is feasible. Our findings may allow tailoring of endometrial biopsies to the individual risk of AEH; additionally, they can pave the way toward less invasive surveillance protocols in patients with known EH.

Glucose Activates Lysine-Specific Demethylase 1 through the KEAP1/p62 Pathway.

Endometrial cancer incidence increases annually. Several risk factors, including high glucose intake, are associated with endometrial cancer. We investigated whether glucose affects lysine-specific demethylase 1 (LSD1) expression and the responsible molecular mechanisms. A high concentration of glucose stimulated p62 phosphorylation and increased LSD1 protein expression. Knockdown of p62 or treatment with mammalian target of rapamycin (mTOR), transforming growth factor-ß activated kinase 1 (TAK1), casein kinase 1 (CK1), and protein kinase C (PKC) inhibitors abrogated glucose-regulated LSD1 expression. Unphosphorylated p62 and LSD1 formed a complex with Kelch-like ECH-associated protein 1 (KEAP1) and were degraded by the KEAP1-dependent proteasome. Phosphorylated p62 increased LSD1 protein expression by escaping the KEAP1 proteasome complex. LSD1 and KEAP1 interaction was enhanced in the presence of the nuclear factor erythroid 2-related factor 2 (NRF2) protein. LSD1 also participated in antioxidant gene regulation with NRF2. In diabetic mice, increasing LSD1and phospho-p62 expression was observed in uterine epithelial cells. Our results indicate that glucose induces p62 phosphorylation through mTOR, TAK1, CK1, and PKC kinases. Subsequently, phospho-p62 competitively interacts with KEAP1 and releases NRF2-LSD1 from the KEAP1 proteasome complex. Our findings may have public health implications for the prevention of endometrial cancer.

A Patient-Derived Xenograft Model of Dedifferentiated Endometrial Carcinoma: A Proof-of-Concept Study for the Identification of New Molecularly Informed Treatment Approaches.

Conventional treatment of dedifferentiated endometrial carcinoma (DEC)-an uncommon and highly aggressive uterine malignancy-is beset by high failure rates. A line of research that holds promise to overcome these limitations is tailored treatments targeted on specific molecular alterations. However, suitable preclinical platforms to allow a reliable implementation of this approach are still lacking. Here, we developed a patient-derived xenograft (PDX) model for preclinical testing of investigational drugs informed by molecular data. The model-termed PDX-mLung was established in mice implanted with lung metastatic lesions obtained from a patient with DEC. Histologic and whole-exome genetic analyses revealed a high degree of identity between PDX-mLung and the patient's parental lesions (both primary DEC and lung metastases). Interestingly, molecular analyses revealed that PDX-mLung harbored druggable alterations including a FGFR2 mutation and CCNE2 amplification. Targeted combined treatment with the FGFR inhibitor lenvatinib and the cell cycle inhibitor palbociclib was found to exert synergistic therapeutic effects against in vivo tumor growth. Based on the results of RNA sequencing, lenvatinib and palbociclib were found to exert anti-tumor effects by interfering interferon signaling and activating hormonal pathways, respectively. Collectively, these data provide proof-of-concept evidence on the value of PDX models for preclinical testing of molecularly informed drug therapy in difficult-to-treat human malignancies. Further clinical research is needed to examine more rigorously the potential usefulness of the lenvatinib and palbociclib combination in patients with DEC.

Serum levels of alpha1-antitrypsin isoforms in patients with ovarian clear cell carcinoma: An exploratory study.

BACKGROUND: Ovarian clear cell carcinoma (OCCC) is frequently associated with endometriosis. Since serum levels of cancer antigen 125 (CA125) have limited diagnostic and prognostic value in this malignancy, there is an unmet need for reliable and specific biomarkers. Previous findings indicated that alpha 1-antitrypsin isoforms (isoAAT) are significantly increased in the peritoneal fluid of patients with endometriosis. This study was undertaken to examine whether serum isoAAT levels in patients with OCCC differ from those measured in women with endometriosis or benign ovarian tumors. We also investigated whether this biomarker may be useful for predicting survival in OCCC. METHODS: Paired serum samples before and after debulking surgery were collected from 27 patients with OCCC. All sera from patients with endometriosis (n = 44) and benign ovarian tumors (n = 32) were obtained in the pretreatment phase. Serum isoAAT levels were assayed using a proprietary ELISA kit. RESULTS: The highest levels of serum isoAAT (median, range) were identified in patients with OCCC (preoperative values: 160.9 ng/mL, range, 101.4-1098.8 ng/mL), followed by patients with endometriosis (125.0 and 83.4-473.2 ng/mL), and those with benign tumors (125.2 and 60.5-191.3 ng/mL). The differences in serum isoAAT levels between patients with OCCC and benign tumors were significant (p = 0.041). Debulking surgery of OCCC resulted in a significant decrease in serum isoAAT levels compared with the preoperative period (median, 160.9 versus 113.0 ng/mL, respectively, p = 0.012). As for prognostic prediction, we found that none of the nine patients with OCCC and serum isoAAT levels ≤130 ng/mL died of disease. CONCLUSION: Serum isoAAT levels may be diagnostically useful to distinguish OCCC from benign ovarian tumors and could also serve as a potential prognostic marker.