Elucidating the role of liver enzymes as markers and regulators in ovarian cancer: a synergistic approach using Mendelian randomization, single-cell analysis, and clinical evidence.

OBJECTIVE: To investigate the association between liver enzymes and ovarian cancer (OC), and to validate their potential as biomarkers and their mechanisms in OC. Methods Genome-wide association studies for OC and levels of enzymes such as Alkaline phosphatase (ALP), Aspartate aminotransferase (AST), Alanine aminotransferase, and gamma-glutamyltransferase were analyzed. Univariate and multivariate Mendelian randomization (MR), complemented by the Steiger test, identified enzymes with a potential causal relationship to OC. Single-cell transcriptomics from the GSE130000 dataset pinpointed pivotal cellular clusters, enabling further examination of enzyme-encoding gene expression. Transcription factors (TFs) governing these genes were predicted to construct TF-mRNA networks. Additionally, liver enzyme levels were retrospectively analyzed in healthy individuals and OC patients, alongside the evaluation of correlations with cancer antigen 125 (CA125) and Human Epididymis Protein 4 (HE4). RESULTS: A total of 283 single nucleotide polymorphisms (SNPs) and 209 SNPs related to ALP and AST, respectively. Using the inverse-variance weighted method, univariate MR (UVMR) analysis revealed that ALP (P = 0.050, OR = 0.938) and AST (P = 0.017, OR = 0.906) were inversely associated with OC risk, suggesting their roles as protective factors. Multivariate MR (MVMR) confirmed the causal effect of ALP (P = 0.005, OR = 0.938) on OC without reverse causality. Key cellular clusters including T cells, ovarian cells, endothelial cells, macrophages, cancer-associated fibroblasts (CAFs), and epithelial cells were identified, with epithelial cells showing high expression of genes encoding AST and ALP. Notably, TFs such as TCE4 were implicated in the regulation of GOT2 and ALPL genes. OC patient samples exhibited decreased ALP levels in both blood and tumor tissues, with a negative correlation between ALP and CA125 levels observed. CONCLUSION: This study has established a causal link between AST and ALP with OC, identifying them as protective factors. The increased expression of the genes encoding these enzymes in epithelial cells provides a theoretical basis for developing novel disease markers and targeted therapies for OC.

Analysis of potential biomarkers for diabetic kidney disease based on single-cell RNA-sequencing integrated with a single-cell sequencing assay for transposase-accessible chromatin.

Diabetic kidney disease (DKD) is a renal microvascular disease caused by hyperglycemia that involves metabolic remodeling, oxidative stress, inflammation, and other factors. The mechanism is complex and not fully unraveled. We performed an integrated single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) and single-cell RNA-sequencing (scRNA-seq) analyses of kidneys from db/db and db/m mice to identify differential open chromatin regions and gene expression, particularly in genes related to proximal tubular reabsorption and secretion. We identified 9,776 differentially expressed genes (DEGs) and 884 cell type-specific transcription factors (TFs) across 15 cell types. Glucose and lipid transporters, and TFs related to the circadian rhythm in the proximal tubules had significantly higher expression in db/db mice than in db/m mice (P<0.01). Crosstalk between podocytes and tubular cells in the proximal tubules was enhanced, and renal inflammation, oxidative stress, and fibrosis pathways were activated in db/db mice. Western blotting and immunohistochemical staining results showed that Wfdc2 expression in the urine and kidneys of DKD patients was higher than that in non-diabetic kidney disease (NDKD) controls. The revealed landscape of chromatin accessibility and transcriptional profiles in db/db mice provide insights into the pathological mechanism of DKD.

Secreted proteins MDK, WFDC2, and CXCL14 as candidate biomarkers for early diagnosis of lung adenocarcinoma.

BACKGROUND: Early diagnosis of lung adenocarcinoma (LUAD), one of the most common types of lung cancer, is very important to improve the prognosis of patients. The current methods can't meet the requirements of early diagnosis. There is a pressing need to identify novel diagnostic biomarkers. Secretory proteins are the richest source for biomarker research. This study aimed to identify candidate secretory protein biomarkers for early diagnosis of LUAD by integrated bioinformatics analysis and clinical validation. METHODS: Differentially expressed genes (DEGs) of GSE31210, gene expression data of early stage of LUAD, were analyzed by GEO2R. Upregulated DEGs predicted to encode secreted proteins were obtained by taking the intersection of the DEGs list with the list of genes encoding secreted proteins predicted by the majority decision-based method (MDSEC). The expressions of the identified secreted proteins in the lung tissues of early-stage LUAD patients were further compared with the healthy control group in mRNA and protein levels by using the UALCAN database (TCGA and CPTAC). The selected proteins expressed in plasma were further validated by using Luminex technology. The diagnostic value of the screened proteins was evaluated by receiver operating characteristic (ROC) analysis. Cell counting kit-8 assay was carried out to investigate the proliferative effects of these screened proteins. RESULTS: A total of 2183 DEGs, including 1240 downregulated genes and 943 upregulated genes, were identified in the GSE31210. Of the upregulated genes, 199 genes were predicted to encode secreted proteins. After analysis using the UALCAN database, 16 molecules were selected for further clinical validation. Plasma concentrations of three proteins, Midkine (MDK), WAP four-disulfide core domain 2 (WFDC2), and C-X-C motif chemokine ligand 14 (CXCL14), were significantly higher in LUAD patients than in healthy donors. The area under the curve values was 0.944, 0.881, and 0.809 for MDK, WFDC2, and CXCL14, 0.962 when combined them. Overexpression of the three proteins enhanced the proliferation activity of A549 cells. CONCLUSIONS: MDK, WFDC2, and CXCL14 were identified as candidate diagnostic biomarkers for early-stage LUAD and might also play vital roles in tumorigenesis.

A bioinformatic analysis of WFDC2 (HE4) expression in high grade serous ovarian cancer reveals tumor-specific changes in metabolic and extracellular matrix gene expression.

Human epididymis protein-4 (HE4/WFDC2) has been well-studied as an ovarian cancer clinical biomarker. To improve our understanding of its functional role in high grade serous ovarian cancer, we determined transcriptomic differences between ovarian tumors with high- versus low-WFDC2 mRNA levels in The Cancer Genome Atlas dataset. High-WFDC2 transcript levels were significantly associated with reduced survival in stage III/IV serous ovarian cancer patients. Differential expression and correlation analyses revealed secretory leukocyte peptidase inhibitor (SLPI/WFDC4) as the gene most positively correlated with WFDC2, while A kinase anchor protein-12 was most negatively correlated. WFDC2 and SLPI were strongly correlated across many cancers. Gene ontology analysis revealed enrichment of oxidative phosphorylation in differentially expressed genes associated with high-WFDC2 levels, while extracellular matrix organization was enriched among genes associated with low-WFDC2 levels. Immune cell subsets found to be positively correlated with WFDC2 levels were B cells and plasmacytoid dendritic cells, while neutrophils and endothelial cells were negatively correlated with WFDC2. Results were compared with DepMap cell culture gene expression data. Gene ontology analysis of k-means clustering revealed that genes associated with low-WFDC2 were also enriched in extracellular matrix and adhesion categories, while high-WFDC2 genes were enriched in epithelial cell proliferation and peptidase activity. These results support previous findings regarding the effect of HE4/WFDC2 on ovarian cancer pathogenesis in cell lines and mouse models, while adding another layer of complexity to its potential functions in ovarian tumor tissue. Further experimental explorations of these findings in the context of the tumor microenvironment are merited.

MicroRNA-325-3p Targets Human Epididymis Protein 4 to Relieve Right Ventricular Fibrosis in Rats with Pulmonary Arterial Hypertension.

BACKGROUND: Pulmonary arterial hypertension (PAH) usually causes right ventricular dysfunction, which is closely related to cardiac fibrosis. But cardiac fibrosis mechanism remains unclear. Our purpose was to explore microRNA-325-3p (miR-325-3p)/human epididymis protein 4's (HE4) role in the occurrence and development of right ventricular fibrosis in PAH. METHODS: The right ventricular fibrosis model of rats with PAH was constructed, and miR-325-3p was overexpressed to explore miR-325-3p's effect on myocardial fibrosis in rats with PAH. Pearson correlation coefficient examined the correlation between HE4 and miR-325-3p. We separated and identified the primary rat myocardial fibroblasts from the heart tissue. Then, the Ang II-treated myocardial fibroblast fibrosis model was constructed. miR-325-3p mimics and si-HE4 and oe-HE4 cell lines were constructed to investigate miR-325-3p and HE4 effects on myocardial cell fibrosis. Then, we added PI3K inhibitor LY294002 to study the effects of HE4 in cell fibrosis by the PI3K/AKT pathway. Starbase was used to predict miR-325-3p and HE4 binding sites. Dual-luciferase reporter assay verified whether miR-325-3p and HE4 were targeted. RESULTS: Overexpression of miR-325-3p alleviated myocardial fibrosis in rats with PAH. Pearson correlation coefficient showed that HE4 was negatively correlated with miR-325-3p expression. Starbase predicted that miR-325-3p had binding sites with HE4. Dual-luciferase reporter assay demonstrated that miR-325-3p targeted HE4. Overexpression of miR-325-3p downregulated HE4 and inhibited myocardial fibroblast fibrosis. HE4 knockdown inhibited myocardial fibroblast fibrosis. HE4 promoted myocardial fibroblast fibrosis and activated the PI3K/AKT pathway. In addition, HE4 affected myocardial fibroblast fibrosis through the PI3K/AKT pathway. CONCLUSIONS: miR-325-3p could target HE4 to relieve right ventricular fibrosis in rats with PAH. This study could provide new targets and strategies for right ventricular fibrosis in PAH.

Detection of plasma exosomal miRNA-205 as a biomarker for early diagnosis and an adjuvant indicator of ovarian cancer staging.

BACKGROUND: Ovarian cancer (OC) is one of the serious threats to the health of women worldwide, and accurate biomarkers are urgently demanded for early diagnosis of OC. We have previously confirmed that miR-205 promotes the invasion and metastasis of OC cells by inhibiting the expression of the tumor suppressor gene TCF21. In this study, we used liquid biopsy technology to detect the expression levels of the four genes, miR-205, CA125, HE4 and TCF21, in the exosomes of plasma of OC patients. Combined with analysis of clinicopathological parameters of OC patients, we aimed to provide efficient and non-invasive laboratory biomarkers for early diagnosis of OC. METHODS: 36 OC patients who were diagnosed in local hospitals from September 2020 to July 2021 were selected as OC group, 31 cases of surgically diagnosed with ovarian benign lesions were selected as benign group, and 32 healthy people who underwent physical examination during the same period were selected as a control group. We employed transmission electron microscope (TEM), Western blotting (WB), and nanoparticle tracking analysis (NTA) to identify biomarkers in the exosomes extracted from plasma of the three groups. The RNA levels of miR-205, CA125, HE4 and TCF21 genes in plasma exosomes were detected by real-time quantitative PCR (qRT-PCR) method. We used clinical pathological parameters and the Receiver Operating Characteristic (ROC) curves to evaluate the diagnostic efficacy for the genes detected in plasma exosomes. RESULTS: We found that the expression level of miR-205 in plasma exosomes of the OC group was significantly higher than that of the benign and control groups (P <  0.05), and the level of miR-205 was elevated during the III-IV periods of OC and lymph node metastasis. CONCLUSION: The level of miR-205 in plasma exosomes is a valuable tumor biomarker to improve OC diagnosis.

WFDC2 Promotes Spasmolytic Polypeptide-Expressing Metaplasia Through the Up-Regulation of IL33 in Response to Injury.

BACKGROUND & AIMS: WAP 4-disulfide core domain protein 2 (WFDC2), also known as human epididymis protein 4, is a small secretory protein that is highly expressed in fibrosis and human cancers, particularly in the ovaries, lungs, and stomach. However, the role of WFDC2 in carcinogenesis is not fully understood. The present study aimed to investigate the role of WFDC2 in gastric carcinogenesis with the use of preneoplastic metaplasia models. METHODS: Three spasmolytic polypeptide-expressing metaplasia (SPEM) models were established in both wild-type and Wfdc2-knockout mice with DMP-777, L635, and high-dose tamoxifen, respectively. To reveal the functional role of WFDC2, we performed transcriptomic analysis with DMP-777-treated gastric corpus specimens. RESULTS: Wfdc2-knockout mice exhibited remarkable resistance against oxyntic atrophy, SPEM emergence, and accumulation of M2-type macrophages in all 3 SPEM models. Transcriptomic analysis revealed that Wfdc2-knockout prevented the up-regulation of interleukin-33 (IL33) expression in the injured mucosal region of SPEM models. Notably, supplementation of recombinant WFDC2 induced IL33 production and M2 macrophage polarization, and ultimately promoted SPEM development. Moreover, long-term treatment with recombinant WFDC2 was able to induce SPEM development. CONCLUSIONS: WFDC2 expressed in response to gastric injury promotes SPEM through the up-regulation of IL33 expression. These findings provide novel insights into the role of WFDC2 in gastric carcinogenesis.

Human epididymis protein 4 promotes P­glycoprotein­mediated chemoresistance in ovarian cancer cells through interactions with Annexin II.

The aim of the present study was to investigate the effects of human epididymis protein 4 (HE4) on drug resistance and its underlying mechanisms. The associations among proteins were detected by immunoprecipitation and immunofluorescence assays. Then, stably transfected cell lines CAOV3­HE4­L and CAOV3­A2­L expressing HE4 short hairpin (sh)RNAs and ANXA2 shRNAs, respectively, were constructed. MTT assay, immunocytochemistry, western blotting, reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and flow cytometry were employed to examine drug sensitivity, as well as the expression and activity of P­glycoprotein (P­gp). HE4 and P­gp in epithelial ovarian cancer tissues were assessed via immunohistochemistry. MicroRNAs that targeted the P­gp gene, ABCB1, were predicted using bioinformatics methods, and their expression was evaluated by RT­qPCR. The common signaling pathways shared by HE4, ANXA2 and P­gp were selected by Gene Set Enrichment Analysis (GSEA). The interaction of HE4, ANXA2 and P­gp were confirmed. P­gp expression was positively associated with HE4 and ANXA2 expression, respectively. Moreover, it was observed that there was no significant rescue of P­gp expression in CAOV3­A2­L cells following the administration of active HE4 protein. In addition, the expression of HE4 and P­gp in ovarian cancer tissues of drug­resistant patients were higher compared with that of the drug­sensitive group (P<0.05). Furthermore, the results revealed that hsa­miR­129­5p was significantly increased accompanied by decreased HE4 or ANXA2 expression and P­gp expression in CAOV3­HE4­L and CAOV3­A2­L cells. GSEA analyses disclosed that HE4, ANXA2 and P­gp genes were commonly enriched in the signaling pathway involved in regulating the actin cytoskeleton. These results indicated that HE4 promotes P­gp­mediated drug resistance in ovarian cancer cells through the interactions with ANXA2, and the underlying mechanism may be associated with decreased expression of hsa­miR­129­5p and dysregulation of the actin cytoskeleton signaling pathway.

MRPL15 is a novel prognostic biomarker and therapeutic target for epithelial ovarian cancer.

PURPOSE: To analyze the role of six human epididymis protein 4 (HE4)-related mitochondrial ribosomal proteins (MRPs) in ovarian cancer and selected MRPL15, which is most closely related to the tumorigenesis and prognosis of ovarian cancer, for further analyses. METHODS: Using STRING database and MCODE plugin in Cytoscape, six MRPs were identified among genes that are upregulated in response to HE4 overexpression in epithelial ovarian cancer cells. The Cancer Genome Atlas (TCGA) ovarian cancer, GTEX, Oncomine, and TISIDB were used to analyze the expression of the six MRPs. The prognostic impact and genetic variation of these six MRPs in ovarian cancer were evaluated using Kaplan-Meier Plotter and cBioPortal, respectively. MRPL15 was selected for immunohistochemistry and GEO verification. TCGA ovarian cancer data, gene set enrichment analysis, and Enrichr were used to explore the mechanism of MRPL15 in ovarian cancer. Finally, the relationship between MRPL15 expression and immune subtype, tumor-infiltrating lymphocytes, and immune regulatory factors was analyzed using TCGA ovarian cancer data and TISIDB. RESULTS: Six MRPs (MRPL10, MRPL15, MRPL36, MRPL39, MRPS16, and MRPS31) related to HE4 in ovarian cancer were selected. MRPL15 was highly expressed and amplified in ovarian cancer and was related to the poor prognosis of patients. Mechanism analysis indicated that MRPL15 plays a role in ovarian cancer through pathways such as the cell cycle, DNA repair, and mTOR 1 signaling. High expression of MRPL15 in ovarian cancer may be associated with its amplification and hypomethylation. Additionally, MRPL15 showed the lowest expression in C3 ovarian cancer and was correlated with proliferation of CD8+ T cells and dendritic cells as well as TGFßR1 and IDO1 expression. CONCLUSION: MRPL15 may be a prognostic indicator and therapeutic target for ovarian cancer. Because of its close correlation with HE4, this study provides insights into the mechanism of HE4 in ovarian cancer.

HE4 Overexpression by Ovarian Cancer Promotes a Suppressive Tumor Immune Microenvironment and Enhanced Tumor and Macrophage PD-L1 Expression.

Ovarian cancer is a highly fatal malignancy characterized by early chemotherapy responsiveness but the eventual development of resistance. Immune targeting therapies are changing treatment paradigms for numerous cancer types but have had minimal success in ovarian cancer. Through retrospective patient sample analysis, we have determined that high human epididymis protein 4 (HE4) production correlates with multiple markers of immune suppression in ovarian cancer, including lower CD8+ T cell infiltration, higher PD-L1 expression, and an increase in the peripheral monocyte to lymphocyte ratio. To further understand the impact that HE4 has on the immune microenvironment in ovarian cancer, we injected rats with syngeneic HE4 high- and low-expressing cancer cells and analyzed the differences in their tumor and ascites immune milieu. We found that high tumoral HE4 expression promotes an ascites cytokine profile that is rich in myeloid-recruiting and differentiation factors, with an influx of M2 macrophages and increased arginase 1 production. Additionally, CTL activation is significantly reduced in the ascites fluid, and there is a trend toward lower CTL infiltration of the tumor, whereas NK cell recruitment to the ascites and tumor is also reduced. PD-L1 expression by tumor cells and macrophages is increased by HE4 through a novel posttranscriptional mechanism. Our data have identified HE4 as a mediator of tumor-immune suppression in ovarian cancer, highlighting this molecule as a potential therapeutic target for the treatment of this devastating disease.

High levels of human epididymis protein 4 mRNA and protein expression are associated with chemoresistance and a poor prognosis in pancreatic cancer.

Pancreatic cancer is associated with an exceedingly poor prognosis, warranting the development of novel therapeutic strategies and discovery of prognostic predictors. Given that chemoresistance­related molecules are reportedly associated with the poor prognosis of pancreatic cancer, the present study aimed to identify molecules that could be efficacious therapeutic targets for pancreatic cancer. First, 10 patient­derived xenografts (PDXs) were established from patients with pancreatic cancer. Subsequently, after treating tumor tissue generated from the PDXs with standard drugs, next­generation sequencing (NGS) was performed using these tissues. The results of NGS analysis and immunohistochemical analysis on 80 pancreatic cancer tissues revealed that human epididymis protein 4 (HE4) expression in the anticancer drug­treated PDX group was higher than that in the untreated PDXs. In addition, chemoresistance ability was observed in tumor cell lines overexpressing HE4. Furthermore, Kaplan­Meier analysis of tumor tissues from 80 patients with pancreatic cancer was performed and it was found that patients with a high HE4 expression level had a poor survival rate compared with those who had a low HE4 expression level. Multivariate analysis also indicated the high expression level of HE4 was an independent poor prognostic biomarker. Thus, it was concluded that high gene and protein expression levels of HE4 mediate chemoresistance and are independent prognostic factors for pancreatic cancer.

Meta-analysis based gene expression profiling reveals functional genes in ovarian cancer.

BACKGROUND: Ovarian cancer causes high mortality rate worldwide, and despite numerous attempts, the outcome for patients with ovarian cancer are still not well improved. Microarray-based gene expressional analysis provides with valuable information for discriminating functional genes in ovarian cancer development and progression. However, due to the differences in experimental design, the results varied significantly across individual datasets. METHODS: In the present study, the data of gene expression in ovarian cancer were downloaded from Gene Expression Omnibus (GEO) and 16 studies were included. A meta-analysis based gene expression analysis was performed to identify differentially expressed genes (DEGs). The most differentially expressed genes in our meta-analysis were selected for gene expression and gene function validation. RESULTS: A total of 972 DEGs with P-value < 0.001 were identified in ovarian cancer, including 541 up-regulated genes and 431 down-regulated genes, among which 92 additional DEGs were found as gained DEGs. Top five up- and down-regulated genes were selected for the validation of gene expression profiling. Among these genes, up-regulated CD24 molecule (CD24), SRY (sex determining region Y)-box transcription factor 17 (SOX17), WFDC2, epithelial cell adhesion molecule (EPCAM), innate immunity activator (INAVA), and down-regulated aldehyde oxidase 1 (AOX1) were revealed to be with consistent expressional patterns in clinical patient samples of ovarian cancer. Gene functional analysis demonstrated that up-regulated WFDC2 and INAVA promoted ovarian cancer cell migration, WFDC2 enhanced cell proliferation, while down-regulated AOX1 was functional in inducing cell apoptosis of ovarian cancer. CONCLUSION: Our study shed light on the molecular mechanisms underlying the development of ovarian cancer, and facilitated the understanding of novel diagnostic and therapeutic targets in ovarian cancer.

ZNF703 promotes tumor progression in ovarian cancer by interacting with HE4 and epigenetically regulating PEA15.

BACKGROUND: It is known that the transcription factor zinc finger protein 703 (ZNF703) plays an important role in physiological functions and the occurrence and development of various tumors. However, the role and mechanism of ZNF703 in ovarian cancer are unclear. MATERIALS AND METHODS: Immunohistochemistry was used to analyze the expression of ZNF703 in ovarian cancer patients and to assess the effect of ZNF703 expression on the survival and prognosis of ovarian cancer patients. ZNF703 overexpression and suppression expression experiments were used to evaluate the effect of ZNF703 on malignant biological behavior of ovarian cancer cells in vitro. Detecting the interaction between HE4 and ZNF703 by immunofluorescence colocalization and coprecipitation, and nuclear translocation. Chromatin immunoprecipitation-sequencing (ChIP-Seq), dual luciferase reporter assay, ChIP-PCR, in vivo model were applied to study the molecular mechanism of ZNF703 affecting the development of ovarian cancer. RESULTS: ZNF703 was highly expressed in ovarian cancer tissues, and its expression level is related to the prognosis of ovarian cancer patients. In vivo and in vitro experiments confirmed that ZNF703 overexpression/inhibition expression will promoted/inhibited the malignant biological behavior of ovarian cancer. Mechanically, ZNF703 interacted with HE4, and HE4 promoted nuclear translocation of ZNF703. ChIP-Seq identified multiple regulatory targets of ZNF703, of which ZNF703 directly binds to the enhancer region of PEA15 to promote the transcription of PEA15 and thereby promoted the proliferation of cancer cells. CONCLUSION: The results showed that ZNF703 as an oncogene played an important role in the epigenetic modification of ovarian cancer proliferation, and suggested that ZNF703 as a transcription factor may become a prognostic factor and a potential therapeutic target for ovarian cancer.

Use of a human embryonic stem cell model to discover GABRP, WFDC2, VTCN1 and ACTC1 as markers of early first trimester human trophoblast.

Human placental development during early pregnancy is poorly understood. Many conceptuses are lost at this stage. It is thought that preeclampsia, intrauterine growth restriction and other placental syndromes that manifest later in pregnancy may originate early in placentation. Thus, there is a need for models of early human placental development. Treating human embryonic stem cells (hESCs) with BMP4 (bone morphogenic protein 4) plus A83-01 (ACTIVIN/NODAL signaling inhibitor) and PD173074 (fibroblast growth factor 2 or FGF2 signaling inhibitor) (BAP conditions) induces differentiation to the trophoblast lineage (hESCBAP), but it is not clear which stage of trophoblast differentiation these cells resemble. Here, comparison of the hESCBAP transcriptome to those of trophoblasts from human blastocysts, trophoblast stem cells and placentas collected in the first-third trimester of pregnancy by principal component analysis suggests that hESC after 8 days BAP treatment most resemble first trimester syncytiotrophoblasts. To further test this hypothesis, transcripts were identified that are expressed in hESCBAP but not in cultures of trophoblasts isolated from term placentas. Proteins encoded by four genes, GABRP (gamma-aminobutyric acid type A receptor subunit Pi), WFDC2 (WAP four-disulfide core domain 2), VTCN1 (V-set domain containing T-cell activation inhibitor 1) and ACTC1 (actin alpha cardiac muscle 1), immunolocalized to placentas at 4-9 weeks gestation, and their expression declined with gestational age (R2 = 0.61-0.83). None are present at term. Expression was largely localized to syncytiotrophoblast of both hESCBAP cells and placental material from early pregnancy. WFDC2, VTCN1 and ACTC1 have not previously been described in placenta. These results support the hypothesis that hESCBAP represent human trophoblast analogous to that of early first trimester and are a tool for discovery of factors important to this stage of placentation.

The biomarker HE4 (WFDC2) promotes a pro-angiogenic and immunosuppressive tumor microenvironment via regulation of STAT3 target genes.

Epithelial ovarian cancer (EOC) is a highly lethal gynecologic malignancy arising from the fallopian tubes that has a high rate of chemoresistant recurrence and low five-year survival rate. The ovarian cancer biomarker HE4 is known to promote proliferation, metastasis, chemoresistance, and suppression of cytotoxic lymphocytes. In this study, we sought to examine the effects of HE4 on signaling within diverse cell types that compose the tumor microenvironment. HE4 was found to activate STAT3 signaling and promote upregulation of the pro-angiogenic STAT3 target genes IL8 and HIF1A in immune cells, ovarian cancer cells, and endothelial cells. Moreover, HE4 promoted increases in tube formation in an in vitro model of angiogenesis, which was also dependent upon STAT3 signaling. Clinically, HE4 and IL8 levels positively correlated in ovarian cancer patient tissue. Furthermore, HE4 serum levels correlated with microvascular density in EOC tissue and inversely correlated with cytotoxic T cell infiltration, suggesting that HE4 may cause deregulated blood vessel formation and suppress proper T cell trafficking in tumors. Collectively, this study shows for the first time that HE4 has the ability to affect signaling events and gene expression in multiple cell types of the tumor microenvironment, which could contribute to angiogenesis and altered immunogenic responses in ovarian cancer.

Prognostic value of serum HE4 in patients with advanced ovarian, fallopian tube, and peritoneal carcinoma.

PURPOSE: In this study, the prognostic value of serum HE4 was investigated in patients with advanced ovarian, fallopian tube, and peritoneal carcinoma. METHODS: Serum HE4 and CA125 levels were measured in both patients and controls, and the response of treatment and the detection of recurrence were evaluated by serum HE4 and CA125 levels in the patients. RESULTS: The results showed that the levels of serum HE4 and CA125 were significantly higher in advanced patients than those seen in benign disease controls (p < 0.001). Compared with CA125, HE4 had higher specificity, but lower sensitivity. Furthermore, serum HE4 was closely associated with the response of treatment and recurrence, the effective response rate for therapy treatment showed by HE4 was higher than CA125, and a serum HE4 level was correlated with a sensitivity of 82.8% and a specificity of 99%, a positive predictive value (PPV) of 97.7%, and a negative predictive value (NPV) of 77.9% to show the presence of recurrence; the accuracy of HE4 for recurrence prediction after treatment was 88.6%. CONCLUSIONS: Our study indicated that serum HE4 levels are effective for diagnosis, evaluating the response of treatment and predicting recurrence in patients with advanced ovarian, fallopian tube, and peritoneal carcinoma.

WFDC2 gene deletion in mouse led to severe dyspnea and type-I alveolar cell apoptosis.

HE4 (Human Epididymis Protein 4) encoded by the wfdc2 gene was first identified as a highly expressed factor in human epididymis. HE4 expression levels in malignant lesions are correlated with the clinical manifestations of gynecologic cancers. HE4 serum test has been widely used for the triage of patients suspected of gynecologic cancers, prognosis of cancer patients, and monitoring cancer recurrence. While it is reported that HE4 may actively participate in the regulation of cancer cell proliferation, migration and drug sensitivity, the physiological role(s) of HE4 in embryo development remains unknown. We applied the TALEN-based strategy to generate wfdc2 gene deletion mice for observation of HE4 function in organogenesis. While heterozygous mice were normal in terms of birth weight, reproductivity, and general behaviors, all the neonates with homozygous wfdc2 deletion suffered severe dyspnea and died in 10 h after birth. Biopsy detected pale-colored lungs, and mechanistic studies indicated increased apoptosis in type-I alveolar cells in lung tissues, which caused hypovascular lung tissue, then led to severe dyspnea in wfdc2-/- neonates. The HE4 knockout mouse has provided an in vivo model for studying the patho-physiological function and relevant molecular pathways of HE4 for the development of respiratory system.

Comparisons of efficacy and safety between docetaxel + cisplatin and paclitaxel + cisplatin and their effects on serum HE4, CA125 and ROMA indicators in patients with ovarian carcinoma.

PURPOSE: To explore the efficacy and safety of docetaxel (DTX) + cisplatin (DDP) and paclitaxel (TAX) + DDP and their effects on serum human epididymis protein 4 (HE4), carbohydrate antigen 125 (CA125) and risk of ovarian malignancy algorithm (ROMA) index in the treatment of ovarian carcinoma. METHODS: A total of 90 patients admitted and treated in our hospital from February 2017 to June 2018, with definitely diagnosed ovarian carcinoma via pathological biopsy were selected. The included patients were randomly divided into two groups: DTX+DDP group (n=45) and TAX+DDP group (n=45). With 3 weeks as a course, the treatments lasted for 6 consecutive courses. The changes in serum vascular endothelial growth factor (VEGF), matrix metalloproteinase 2 (MMP-2), HE4, CA125 and ROMA index were detected before and after treatments. Moreover, the incidence of adverse reactions was observed, and the clinical therapeutic efficacy was assessed. RESULTS: The clinical efficacy in both groups revealed that there were 39 and 34 cases obtained complete remission or partial remission in the DTX+DDP group and in the TAX+DDP group, respectively. Overall efficiencies were 86.67 and 75.56%, respectively, showing statistically significant differences between the two groups (p<0.05). The incidence rate of adverse reactions in DTX+DDP group was significantly lower than that in TAX+DDP group (p<0.05). The VEGF and MMP-2 levels in both DTX+DDP and TAX+DDP group were decreased compared with those before treatment (183.35±25.26 vs. 279.18±27.75 pg/mL and 228.22±40.21 vs. 316.11±33.6 pg/mL (p<0.05). The serum HE4 and CA125 levels and ROMA index in both groups were lower than those before treatment (121.19±14.14 vs. 159.43±18.15 pmol/L) (p<0.05), 239.45±25.37 vs. 288.37±30.36 pmol/L (p<0.05) and 58.02±6.61 vs. 76.23±11.58 (p<0.05), respectively). The above indicators were decreased in the DTX+DDP group to a significant extent (p<0.05). CONCLUSIONS: Both DTX+DDP and TAX+DDP treatments are effective for the patients with ovarian carcinoma. However, DTX+DDP is more efficacious in lowering indicators such as serum CA125, HE4 and MMP-2 and ROMA index and adverse reactions, thus providing a more efficient practice scheme with lower toxic side effects for the clinical treatment of ovarian carcinoma.

Downregulation of Rab23 inhibits proliferation, invasion, and metastasis of human ovarian cancer.

Previously, we reported that the expression of human epididymis protein (HE4) was correlated with the expression of RAB23 in ovarian cancer cells. Rab23 is a member of the Ras-related small GTPase superfamily, which plays a key role in the Sonic Hedgehog (Shh) signaling pathway. However, the function of Rab23 in ovarian cancer remains unclear. In this study, we explored the location and expression of Rab23 in ovarian cancer tissues and cells (CaoV3 and A2780), and further investigated the function and potential mechanism of Rab23 in malignant biological behaviors including the epithelial-mesenchymal transition (EMT) process in ovarian cancer for the first time. Rab23 is highly expressed in ovarian cancer tissues and associated with advanced stage, and shortened overall survival time of ovarian cancer patients. We are the first to report that human epididymis protein (HE4) can regulate the expression of the Rab23 protein, and that knockdown of RAB23 decreases the proliferation, invasion, and migration abilities as well as inhibits the epithelial-mesenchymal transition (EMT) process in ovarian cancer cells. Furthermore, downregulation of Rab23 significantly inhibited Shh-Gli1 and PI3K-AKT signaling pathways. Collectively, our results indicate that Rab23 plays a critical role in the malignant biological behavior of ovarian cancer and may serve as a potential biomarker and therapeutic target for ovarian cancer.

Cervical, Ovarian and Endometrial Tumor Markers: Potential Clinical Value.

Tumors markers can be described as molecular products expressed by neoplasia tissues (immunohistochemistry), or metabolized and secreted by tumor and characterized biochemically in body fluids such as blood and urine. They may have utility as indicators of tumor stage and grade as well useful for monitoring responses to treatment and predicting recurrence, progression, development of metastases, or even patient survival. Unfortunately, in some cases they may have no identified clinical potential. Several investigations have been carried out, especially in the last decade, using biotechnological methods, in order to identify new potential tumor markers. By translating these findings into clinical use one may facilitate accurate diagnosis and prognostic prediction, and contribute to individualized treatment. The objective of this review is to describe some biomarkers with potential use in clinical settings of uterine cervix, ovary, and endometrium carcinomas.