An AsCas12f-based compact genome-editing tool derived by deep mutational scanning and structural analysis.

SpCas9 and AsCas12a are widely utilized as genome-editing tools in human cells. However, their relatively large size poses a limitation for delivery by cargo-size-limited adeno-associated virus (AAV) vectors. The type V-F Cas12f from Acidibacillus sulfuroxidans is exceptionally compact (422 amino acids) and has been harnessed as a compact genome-editing tool. Here, we developed an approach, combining deep mutational scanning and structure-informed design, to successfully generate two AsCas12f activity-enhanced (enAsCas12f) variants. Remarkably, the enAsCas12f variants exhibited genome-editing activities in human cells comparable with those of SpCas9 and AsCas12a. The cryoelectron microscopy (cryo-EM) structures revealed that the mutations stabilize the dimer formation and reinforce interactions with nucleic acids to enhance their DNA cleavage activities. Moreover, enAsCas12f packaged with partner genes in an all-in-one AAV vector exhibited efficient knock-in/knock-out activities and transcriptional activation in mice. Taken together, enAsCas12f variants could offer a minimal genome-editing platform for in vivo gene therapy.

Structure and engineering of the minimal type VI CRISPR-Cas13bt3.

Type VI CRISPR-Cas13 effector enzymes catalyze RNA-guided RNA cleavage and have been harnessed for various technologies, such as RNA detection, targeting, and editing. Recent studies identified Cas13bt3 (also known as Cas13X.1) as a miniature Cas13 enzyme, which can be used for knockdown and editing of target transcripts in mammalian cells. However, the action mechanism of the compact Cas13bt3 remains unknown. Here, we report the structures of the Cas13bt3-guide RNA complex and the Cas13bt3-guide RNA-target RNA complex. The structures revealed how Cas13bt3 recognizes the guide RNA and its target RNA and provided insights into the activation mechanism of Cas13bt3, which is distinct from those of the other Cas13a/d enzymes. Furthermore, we rationally engineered enhanced Cas13bt3 variants and ultracompact RNA base editors. Overall, this study improves our mechanistic understanding of the CRISPR-Cas13 enzymes and paves the way for the development of efficient Cas13-mediated transcriptome modulation technologies.

Structure of the miniature type V-F CRISPR-Cas effector enzyme.

RNA-guided DNA endonucleases derived from CRISPR-Cas adaptive immune systems are widely used as powerful genome-engineering tools. Among the diverse CRISPR-Cas nucleases, the type V-F Cas12f (also known as Cas14) proteins are exceptionally compact and associate with a guide RNA to cleave single- and double-stranded DNA targets. Here, we report the cryo-electron microscopy structure of Cas12f1 (also known as Cas14a) in complex with a guide RNA and its target DNA. Unexpectedly, the structure revealed that two Cas12f1 molecules assemble with the single guide RNA to recognize the double-stranded DNA target. Each Cas12f1 protomer adopts a different conformation and plays distinct roles in nucleic acid recognition and DNA cleavage, thereby explaining how the miniature Cas12f1 enzyme achieves RNA-guided DNA cleavage as an "asymmetric homodimer." Our findings augment the mechanistic understanding of diverse CRISPR-Cas nucleases and provide a framework for the development of compact genome-engineering tools critical for therapeutic genome editing.

Rel Family Transcription Factor NFAT5 Upregulates COX2 via HIF-1α Activity in Ishikawa and HEC1a Cells.

Nuclear factor of activated T cells 5 (NFAT5) and cyclooxygenase 2 (COX2; PTGS2) both participate in diverse pathologies including cancer progression. However, the biological role of the NFAT5-COX2 signaling pathway in human endometrial cancer has remained elusive. The present study explored whether NFAT5 is expressed in endometrial tumors and if NFAT5 participates in cancer progression. To gain insights into the underlying mechanisms, NFAT5 protein abundance in endometrial cancer tissue was visualized by immunohistochemistry and endometrial cancer cells (Ishikawa and HEC1a) were transfected with NFAT5 or with an empty plasmid. As a result, NFAT5 expression is more abundant in high-grade than in low-grade endometrial cancer tissue. RNA sequencing analysis of NFAT5 overexpression in Ishikawa cells upregulated 37 genes and downregulated 20 genes. Genes affected included cyclooxygenase 2 and hypoxia inducible factor 1α (HIF1A). NFAT5 transfection and/or treatment with HIF-1α stabilizer exerted a strong stimulating effect on HIF-1α promoter activity as well as COX2 expression level and prostaglandin E2 receptor (PGE2) levels. Our findings suggest that activation of NFAT5-HIF-1α-COX2 axis could promote endometrial cancer progression.

Constitutive activation of estrogen receptor α signaling in muscle prolongs exercise endurance in mice.

Estrogen is a female hormone that plays a role in various tissues, although the mechanism in skeletal muscle has not been fully clarified. We previously showed that systemic administration of estrogen for 10 weeks ameliorated decreased exercise endurance in ovariectomized mice. To assess whether a long-term and muscle-specific activation of estrogen signaling modulates muscle function, we constructed an expression plasmid for a constitutively active estrogen receptor α (caERα) under the control of muscle creatine kinase (Mck) gene promoter/enhancer. In C2C12 mouse myoblastic cells, transfection of the Mck-caERα plasmid elevated the estrogen response element-driven transcription in a ligand-independent manner. Using this construct, we generated Mck-caERα transgenic mice, in which caERα is predominantly expressed in muscle. Treadmill running test revealed that female Mck-caERα mice exhibit a prolonged running time and distance compared with the wild-type mice. Moreover, microarray expression analysis revealed that the genes related to lipid metabolism, insulin signaling, and growth factor signaling were particularly upregulated in the quadriceps femoris muscle of Mck-caERα mice. These results suggest that estrogen signaling potentiates exercise endurance in skeletal muscle through modulating the expression of metabolism-associated genes.

Depressive and anxiety symptoms among Japanese cancer survivors: Japan cancer survivorship research project.

BACKGROUND: This study aimed to clarify predictors of depressive symptoms and anxiety symptoms after cancer diagnosis among Japanese cancer survivors (CSs). METHODS: As part of a Japanese cancer survivorship research project commissioned by the Ministry of Health, Labour and Welfare (MHLW) of Japan, we conducted a web-based nationwide survey of CSs in 2018. We analyzed the risk factors for depressive and anxiety symptoms, as measured by the Hospital Anxiety and Depression Scale Japanese version (HADS). RESULTS: Of 1,234 Japanese CSs, mean score of HADS-depression and HADS-anxiety were 4.08 and 4.78, respectively. At the time of the study, the number of CSs with symptoms of depression and anxiety were 111 (9.0%) and 269 (21.8%), respectively. After multivariable analysis, CSs ≥ 60 years old (reference: ≤ 39 years old, odds ratios (OR): 0.39, 95%CI: 0.17-0.90) and those ≥ 10 years from cancer diagnosis (reference: 0-4 years, OR: 0.55, 95%CI: 0.32-0.96) had lower odds for depressive symptoms. And CSs ≥ 60 years old (reference: ≤ 39 years old, OR: 0.27, 95%CI: 0.15-0.49) and those ≥ 10 years from cancer diagnosis (reference: 0-4 years, OR: 0.62, 95%CI: 0.42-0.90) also had lower odds for anxiety symptoms. CSs who received chemotherapy (OR: 1.56, 95%CI: 1.10-2.20) had higher odds for anxiety symptoms. CONCLUSIONS: Based on manifestation of symptoms, CSs who were younger, closer to the time of cancer diagnosis, had advanced-staged cancer, or received chemotherapy may be at higher risk for depressive or anxiety symptoms. Those CSs who have higher risk for depression and anxiety symptoms, should be followed-up more carefully for better cancer survivorship, by medical professionals, companies, and society.

Fetal heart rate evolution patterns in cerebral palsy associated with umbilical cord complications: a nationwide study.

BACKGROUND: The aim of the present study was to clarify fetal heart rate (FHR) evolution patterns in infants with cerebral palsy (CP) according to different types of umbilical cord complications. METHODS: This case-control study included children born: with a birth weight ≥2000 g, at gestational age ≥33 weeks, with disability due to CP, and between 2009 and 2014. Obstetric characteristics and FHR patterns were compared among patients with CP associated with (126 cases) and without (594 controls) umbilical cord complications. RESULTS: There were 32 umbilical cord prolapse cases and 94 cases with coexistent antenatal umbilical cord complications. Compared with the control group, the persistent non-reassuring pattern was more frequent in cases with coexistent antenatal umbilical cord complications (p = 0.012). A reassuring FHR pattern was observed on admission, but resulted in prolonged deceleration, especially during the first stage of labor, and was significantly identified in 69% of cases with umbilical cord prolapse and 35% of cases with antenatal cord complications, compared to 17% of control cases (p < 0.001). CONCLUSION: Hypercoiled cord and abnormal placental umbilical cord insertion, may be associated with CP due to acute hypoxic-ischemic injury as well as sub-acute or chronic adverse events during pregnancy, while umbilical cord prolapse may be characterized by acute hypoxic-ischemic injury during delivery.

Risk factors for infertility treatment-associated harassment among working women: a Japan-Female Employment and Mental health in assisted reproductive technology (J-FEMA) study.

PURPOSE: This study aims to elucidate the risk factors of infertility treatment-associated harassment (I-harassment) among Japanese working women. METHODS: The study participants were 1103 female patients who enrolled in the Japan-Female Employment and Mental Health in artificial reproductive technology (J-FEMA) study. Of the 1727 female patients, 1103 female patients were working during the initiation of infertility treatment and were still working during the survey. Risk factors for I-harassment were analyzed using a multivariable logistic regression model. RESULTS: In this study, 82 female patients (7.4%) experienced I-harassment. The risk was significantly higher in those who had more in vitro fertilization (IVF) cycles than those who had fewer IVF cycles (OR, 1.06; 95% CI, 1.01-1.10). Similarly, those who disclosed their infertility treatment to their workplace were at significantly higher risk for I-harassment than those who did not (OR, 1.80; 95% CI, 1.03-3.15). CONCLUSION: This study found that 7.4% of female patients experienced I-harassment after infertility treatment initiation. Those female patients who "experienced more IVF cycles," and "disclosed their infertility treatment in their workplace" should be carefully followed up by healthcare professionals to prevent I-harassment.

Identification of a Novel Oncogenic Fusion Gene SPON1-TRIM29 in Clinical Ovarian Cancer That Promotes Cell and Tumor Growth and Enhances Chemoresistance in A2780 Cells.

Gene structure alterations, such as chromosomal rearrangements that develop fusion genes, often contribute to tumorigenesis. It has been shown that the fusion genes identified in public RNA-sequencing datasets are mainly derived from intrachromosomal rearrangements. In this study, we explored fusion transcripts in clinical ovarian cancer specimens based on our RNA-sequencing data. We successfully identified an in-frame fusion transcript SPON1-TRIM29 in chromosome 11 from a recurrent tumor specimen of high-grade serous carcinoma (HGSC), which was not detected in the corresponding primary carcinoma, and validated the expression of the identical fusion transcript in another tumor from a distinct HGSC patient. Ovarian cancer A2780 cells stably expressing SPON1-TRIM29 exhibited an increase in cell growth, whereas a decrease in apoptosis was observed, even in the presence of anticancer drugs. The siRNA-mediated silencing of SPON1-TRIM29 fusion transcript substantially impaired the enhanced growth of A2780 cells expressing the chimeric gene treated with anticancer drugs. Moreover, a subcutaneous xenograft model using athymic mice indicated that SPON1-TRIM29-expressing A2780 cells rapidly generated tumors in vivo compared to control cells, whose growth was significantly repressed by the fusion-specific siRNA administration. Overall, the SPON1-TRIM29 fusion gene could be involved in carcinogenesis and chemotherapy resistance in ovarian cancer, and offers potential use as a diagnostic and therapeutic target for the disease with the fusion transcript.

Transcriptomic analysis of hormone-sensitive patient-derived endometrial cancer spheroid culture defines Efp as a proliferation modulator.

Estrogen-responsive endometrial cancer (EC) is prevalent in uterine cancer. Its precise molecular mechanisms remain to be elucidated partly because of limited availability of estrogen-sensitive EC models recapitulating clinical pathophysiology. We previously established EC patient-derived cancer cell (EC-PDC) spheroid culture with high expression of estrogen receptor α (ERα). Using this EC-PDC, we study the transcriptional regulation and function of estrogen-responsive finger protein (Efp), a prototypic tripartite motif (TRIM) protein that modulates protein degradation and RNA processing. Intense estrogen-dependent EFP mRNA induction and high ERα occupancy to EFP estrogen responsive element (ERE) were observed in EC-PDC. Luciferase reporter gene assay showed that the ERE facilitates EFP transcriptional activity estrogen-dependently. siRNA-mediated Efp silencing in EC-PDC resulted in suppressed spheroid proliferation and altered gene expression profile, featuring downregulation of genes related to cell cycle (e.g., CDK6) and inflammation/immune responses (e.g., IL10RA, IL26, and IL6ST) while unaffected expression of cancer stemness-related markers. Taken together, EC-PDC spheroid culture is a powerful EC tool that enables to dissect Efp-mediated ERα signaling pathways as an estrogen-sensitive EC model. This study provides an insight into alternative EC therapeutic strategies targeting ERα-Efp axis.

Whole exome sequencing of fetal structural anomalies detected by ultrasonography.

The objective of this study was to evaluate the efficacy of whole exome sequencing (WES) for the genetic diagnosis of cases presenting with fetal structural anomalies detected by ultrasonography. WES was performed on 19 cases with prenatal structural anomalies. Genomic DNA was extracted from umbilical cords or umbilical blood obtained shortly after birth. WES data were analyzed on prenatal phenotypes alone, and the data were re-analyzed after information regarding the postnatal phenotype was obtained. Based solely on the fetal phenotype, pathogenic, or likely pathogenic, single nucleotide variants were identified in 5 of 19 (26.3%) cases. Moreover, we detected trisomy 21 in two cases by WES-based copy number variation analysis. The overall diagnostic rate was 36.8% (7/19). They were all compatible with respective fetal structural anomalies. By referring to postnatal phenotype information, another candidate variant was identified by a postnatal clinical feature that was not detected in prenatal screening. As detailed phenotyping is desirable for better diagnostic rates in WES analysis, we should be aware that fetal phenotype is a useful, but sometimes limited source of information for comprehensive genetic analysis. It is important to amass more data of genotype-phenotype correlations, especially to appropriately assess the validity of WES in prenatal settings.

Predictors of resignation and sick leave after cancer diagnosis among Japanese breast cancer survivors: a cross-sectional study.

BACKGROUND: In Japan, 55.5% of breast cancer survivors (BCSs) are of working age, so various perspectives regarding return to work (RTW) after cancer diagnosis need to be considered. Therefore, this study aimed to clarify the risk factors for resignation and taking sick leave (SL) among BCSs in continued employment at the time of diagnosis. METHODS: A web-based retrospective cross-sectional survey was conducted on BCSs using data from a 2018 Japanese national research project (Endo-Han) commissioned by the Ministry of Health, Labour and Welfare of Japan. The subjects were women aged 18-69 years who had been diagnosed with breast cancer for the first time at least 1 year previously. The risk factors for resignation and taking SL after breast cancer diagnosis, including age at diagnosis, education level, cancer stage, surgery, chemotherapy, radiotherapy, employment status, and occupational type, were then analyzed using a logistic regression model. RESULTS: In total, 40 (14.9%) of 269 BCSs quit their jobs at least 1 year after being diagnosed with breast cancer. The results of the multivariable analysis indicated that lower education level (odds ratio [OR]: 3.802; 95% confidence interval [CI]: 1.233-11.729), taking SL (OR: 2.514; 95%CI: 1.202-5.261), and younger age at diagnosis (OR: 0.470; 95%CI: 0.221-0.998) were predictors of resignation. Of 229 patients who continued working, SL was taken by 72 (31.4%). In addition, undergoing surgery was found to be a predictor of taking SL (OR: 8.311; 95%CI: 1.007-68.621). CONCLUSIONS: In total, 40 (14.9%) of 269 BCSs quit their jobs at least 1 year after being diagnosed with breast cancer. The results of this study indicated that younger age, lower education level, and taking SL were predictors of resignation after breast cancer diagnosis.

Risk factors for cerebral palsy in neonates due to placental abruption.

AIM: This study aimed to identify risk factors for the onset of cerebral palsy (CP) in neonates due to placental abruption and investigate their characteristics. METHODS: A retrospective case-control study was conducted using a nationwide registry from Japan. The study population included pregnant women (n = 122) who delivered an infant with CP between 2009 and 2015, where placental abruption was identified as the single cause of CP. The control group consisted of pregnant women with placental abruption, who delivered an infant without CP and were managed from 2013 to 2014. They were randomly identified from the prenatal database of the Japan Society of Obstetrics and Gynecology (JSOG-DB; n = 1214). Risk factors were investigated using multivariate analysis. RESULTS: Alcohol consumption (3.38, 2.01-5.68) (odds ratio, 95% confidence interval), smoking during pregnancy (3.50, 1.32-9.25), number of deliveries (1.28, 1.05-1.56), polyhydramnios (5.60, 1.37-22.6), oral administration of ritodrine hydrochloride (2.09, 1.22-3.57) and hypertensive disorders in pregnancy (2.25, 1.27-4.07) were significant risk factors. In contrast, intravenous administration of oxytocin (odds ratio, 95% confidence interval: 0.22, 0.09-0.58) and magnesium sulfate (0.122, 0.02-0.89) attenuated risk. CONCLUSION: Alcohol consumption, smoking during pregnancy, number of deliveries, polyhydramnios, oral administration of ritodrine hydrochloride and hypertensive disorders in pregnancy were identified as risk factors for CP following placental abruption. Regarding alcohol consumption and smoking during pregnancy, the results suggest the importance of educational activities targeting pregnant women to increase their awareness of placental abruption.

Analysis of severe psychological stressors in women during fertility treatment: Japan-Female Employment and Mental health in Assisted reproductive technology (J-FEMA) study.

PURPOSE: To identify risk factors for severe psychological stress in women undergoing fertility treatment. METHODS: This cross-sectional, multi-center study was conducted from August to December 2018. We recruited 1672 subjects who completed an anonymous, self-reported questionnaire regarding fertility treatment, conditions at work and home, and psychological stress using K6 score, which estimates psychological distress during the previous 30 days. We further focused our analysis on 1335 subjects who were working when starting fertility treatment. RESULTS: Of 1672 women, mean K6 score (range 0-24) was 4.8 ± 4.4, including 103 women (6.2%) with K6 score ≥ 13 (high K6), and classified as probable severe psychological distress. Multivariate logistic regression analysis showed that high K6 was strongly associated with low annual family income of ≤ USD55,700 (JPY6 million) (odds ratio [OR] 1.89, 95% confidence interval [CI] 1.04-3.42), infertility duration of ≥ 2 years (OR 1.87, 95% CI 1.08-3.25), and no experience of childbirth (OR 2.04, 95% CI 1.05-3.97). Focusing on 1335 working women, 266 (19.9%) experienced resignation from work. High K6 was strongly associated with low family income (OR 2.83, 95% CI 1.52-5.28), cessation of professional duties (OR 2.08, 95% CI 1.05-4.14), infertility-related harassment in the workplace (OR 2.07, 95% CI 1.08-3.98), and perceived difficulties to continue working during fertility treatment (OR 2.94, 95% CI 1.15-7.50). CONCLUSION: Severe psychological stressors in women during fertility treatment included low family income, long infertility duration, childlessness, infertility-related harassment, and perceived difficulty in working conditions or cessation from work. Establishment of mental health care support systems is urgently required in this population.

Analysis of vaginal and endometrial microbiota communities in infertile women with a history of repeated implantation failure.

PURPOSE: To identify specific bacterial communities in vaginal and endometrial microbiotas as biomarkers of implantation failure by comprehensively analyzing their microbiotas using next-generation sequencing. METHODS: We investigated α- and ß-diversities of vaginal and endometrial microbiotas using 16S rRNA gene sequencing and compared their profiles between 145 women with repeated implantation failure (RIF) and 21 controls who lacked the factors responsible for implantation failure with a high probability of being healthy and fertile to identify specific bacteria that induce implantation failure. RESULTS: The endometrial microbiotas had higher α-diversities than did the vaginal microbiotas (P < .001). The microbiota profiles showed that vaginal and endometrial samples in RIF patients had significantly higher levels of 5 and 14 bacterial genera, respectively, than those in controls. Vaginal Lactobacillus rates in RIF patients were significantly lower at 76.4 ± 38.9% compared with those of the controls at 91.8 ± 22.7% (P = .018), but endometrial Lactobacillus rates did not significantly differ between the RIF patients and controls (56.2 ± 36.4% and 58.8 ± 37.0%, respectively, P = .79). CONCLUSIONS: Impaired microbiota communities containing specific bacteria in both the endometrium and vagina were associated with implantation failure. The vaginal Lactobacillus rates, but not the endometrial, may be a biomarker for RIF.

Dual-specificity phosphatase 6 plays a critical role in the maintenance of a cancer stem-like cell phenotype in human endometrial cancer.

The prognosis of patients with high-grade or advanced-stage endometrial cancer remains poor. As cancer stem-like cells (CSCs) are thought to be associated with endometrial cancers, it is essential to investigate the molecular mechanisms that regulate endometrial CSCs. Dual-specificity phosphatase 6 (DUSP6) functions as a negative-feedback regulator of MAPK-ERK1/2 signaling, but its role in endometrial cancer remains unknown. We investigated whether DUSP6 is involved in cancer cell stemness using endometrial cancer cell lines and specimens from endometrial cancer patients. DUSP6 induced the expression of CSC-related genes including ALDH1, Nanog, SOX2 and Oct4A, increased the population of cells in the G0/G1 phase, and promoted sphere formation ability. DUSP6 knockdown resulted in reduced cell invasion and metastasis, whereas DUSP6 overexpression inhibited apoptosis under serum-free conditions. Moreover, DUSP6 decreased phosphorylated ERK1/2 and increased phosphorylated Akt levels, which potentially induces CSC features. In patients with endometrial cancers, DUSP6 expression was determined using immunohistochemistry, and based on the results, the patients were dichotomized into high- and low-DUSP6-expression groups. Progression-free survival and overall survival were significantly shorter in the high-DUSP6-expression group. These results suggest that DUSP6 has potential value as a biomarker of CSCs and as a target of therapies designed to eliminate CSCs in endometrial cancer.

Retraction Note to: Nonsense variants in STAG2 result in distinct sex-dependent phenotypes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Depth structure analysis by surface scanning in near-field microscopes.

High-resolution imaging of the surfaces of samples can be performed using near-field optical microscopes by scanning a small light spot; however, structures located deep beneath cannot be observed because the light spot spreads in three directions. In this study, we propose an observation technique for near-field optical microscopes that can obtain depth information within the resolution of the diffraction limit of light by analyzing interference patterns formed with divergent incident light and scattered light from a sample. We analyze depth structures by evaluating correlation coefficients between observed interference patterns and calculated reference patterns. Our technique can observe both high-resolution surface images and the diffraction-limited three-dimensional structure by scanning a near-field light source on a single plane.

Risk factors for resignation from work after starting infertility treatment among Japanese women: Japan-Female Employment and Mental health in Assisted reproductive technology (J-FEMA) study.

OBJECTIVE: To elucidate the risk factors associated with resignation from work of Japanese women undergoing infertility treatment. METHODS: A total of 1727 female patients who attended a private fertility clinic in Japan participated in the Japan-Female Employment and Mental health in Assisted reproductive technology study. Questions related to demographic, clinical and socioeconomic characteristics were employed in the questionnaire. Out of the 1727 patients, 1075 patients who were working at the time of initiating infertility treatment and felt infertility treatment incompatible with work were included in the analysis. Risk factors for resignation were assessed by using multivariable logistic regression models. RESULTS: Among 1075 working women who started infertility treatment, 179 (16.7%) subsequently resigned. Multivariable-adjusted ORs for resignation in those with lower educational background and infertility for ≥2 years were 1.58 (95% CI: 1.07 to 2.34) and 1.82 (95% CI: 1.15 to 2.89), respectively. The OR for resignation in non-permanent workers undergoing infertility treatment was 2.65 (95% CI: 1.61 to 4.37). While experiencing harassment in the workplace approached significance, lack of support from the company was significantly associated with resignation after starting infertility treatment, with ORs of 1.71 (95% CI: 0.98 to 2.99) and 1.91 (95% CI: 1.28 to 2.86), respectively. CONCLUSION: One-sixth of women resigned after starting infertility treatments. It was found that factors related to education, infertility duration and work environment were significantly associated with resignation. Reducing the physical and psychological burden endured by women, for example, by increasing employer-provided support, is vitally important in balancing infertility treatment with maintenance of work life.

Identification of novel mutations of ovarian cancer-related genes from RNA-sequencing data for Japanese epithelial ovarian cancer patients.

Ovarian cancer has the highest mortality rate among gynecological cancers. Gene mutations are involved in the carcinogenesis, metastasis, and therapeutic response in ovarian cancer. However, the variety and proportion of gene mutation is not fully analyzed in Japanese ovarian cancer patients, especially, in those with recurrent tumors. In the present study, RNA-sequencing was performed for 32 clinical ovarian specimens obtained from 24 Japanese patients (24 primary cancer specimens and 8 recurrent specimens paired with corresponding primary cancer specimens). Mutations in 24 primary specimens were analyzed by comparing the sequence data mapped on RefSeq genes with those in the public online databases BRCA Exchange, COSMIC, ClinVar, and cBioportal. Mutations were observed in TP53 in 16 specimens (67%), BRCA1 in 9 (38%), BRCA2 in 13 (54%), ARID1A in 3 (13%), PIK3CA in 2 (8%), KRAS in 1 (4%), PTEN in 1 (4%), and CTNNB1 in 1 (4%), excluding synonymous mutations. Among those identified muations, 13 of 14 mutations in TP53, 10 of 11 mutations of BRCA1, 10 of 23 mutation positions of BRCA2, none of 7 mutations of ARID1A, 1 mutation of PIK3CA, and 1 mutation of CTNNB1 were consistent with those reported in the public online databases; however, the other mutations identified were novel. Comparison between matched-paired specimens of primary and recurrent tumors revealed the changes of mutational status in expressed RNAs. RNA-sequencing-based mutation analysis will be useful to reveal ethnic differences of gene mutations in ovarian cancer and to understand the contribution of gene mutations to recurrence.