Human Epididymis Protein 4 Promotes Events Associated with Metastatic Ovarian Cancer via Regulation of the Extracelluar Matrix

Abstract

Human epididymis protein 4 (HE4) has received much attention recently due to its diagnostic and prognostic abilities for epithelial ovarian cancer. Since its inclusion in the Risk of Ovarian Malignancy Algorithm (ROMA), studies have focused on its functional effects in ovarian cancer. Here, we aimed to investigate the role of HE4 in invasion, haptotaxis, and adhesion of ovarian cancer cells. Furthermore, we sought to gain an understanding of relevant transcriptional profiles and protein kinase signaling pathways mediated by this multifunctional protein. Exposure of OVCAR8 ovarian cancer cells to recombinant HE4 (rHE4) promoted invasion, haptotaxis toward a fibronectin substrate, and adhesion onto fibronectin. Overexpression of HE4 or treatment with rHE4 led to upregulation of several transcripts coding for extracellular matrix proteins, including SERPINB2, GREM1, LAMC2, and LAMB3. Gene ontology indicated an enrichment of terms related to extracellular matrix, cell migration, adhesion, growth, and kinase phosphorylation. LAMC2 and LAMB3 protein levels were constitutively elevated in cells overexpressing HE4 and were upregulated in a time-dependent manner in cells exposed to rHE4 in the media. Deposition of laminin-332, the heterotrimer comprising LAMC2 and LAMB3 proteins, was increased in OVCAR8 cells treated with rHE4 or conditioned media from HE4-overexpressing cells. Enzymatic activity of matriptase, a serine protease that cleaves laminin-332 and contributes to its pro-migratory functional activity, was enhanced by rHE4 treatment in vitro. Proteomic analysis revealed activation of focal adhesion kinase signaling in OVCAR8 cells treated with conditioned media from HE4-overexpressing cells. Focal adhesions were increased in cells treated with rHE4 in the presence of fibronectin. These results indicate a direct role for HE4 in mediating malignant properties of ovarian cancer cells and validate the need for HE4-targeted therapies that will suppress activation of oncogenic transcriptional activation and signaling cascades.

Inhibition of DUSP6 sensitizes ovarian cancer cells to chemotherapeutic agents via regulation of ERK signaling response genes

“…Quantitative RT-PCR was performed as previously described [10]. Validated primers for DUSP6, EGR1, c-JUN, and ERBB3 were purchased from https:// www.realtimeprimers.com.…”

“…Inclusion of preoperative serum HE4 levels into the diagnostic Risk of Ovarian Malignancy Algorithm (ROMA) results in demonstrably improved specificity and sensitivity in detection and monitoring of the disease over Cancer Antigen 125 (CA 125), pelvic sonography, and menopausal status alone [5]. Research has also shown its ability to support EOC pathogenesis, including the promotion of proliferation, chemoresistance, antiestrogen resistance, adhesion, invasion, and migration [6][7][8][9][10][11][12][13][14][15][16][17]. One oncogenic pathway that has consistently been shown to interact with HE4 in several studies is the extracellular signal regulated kinase (ERK) pathway.…”

Beyond the Biomarker: Understanding the Diverse Roles of Human Epididymis Protein 4 in the Pathogenesis of Epithelial Ovarian Cancer

“…Interestingly, there was no change in adhesion to collagen I, IV, laminin I, and fibrinogen matrices, suggesting that HE4 has a specific effect on fibronectin adhesion. Haptotaxis toward a fibronectin substrate also was increased in the ovarian cancer cells treated with recombinant HE4 by 1.72-fold ( 60 ).…”

Gene Expression Profile of Active HE4 Stimulation in Epithelial Ovarian Cancer Cells: Microarray Study and Comprehensive Bioinformatics Analysis

“…In recent years, a growing number of investigations have gradually found that HE4 promotes cell proliferation, adhesion, invasion, migration, and chemoresistance in ovarian cancer [20][21][22][23][24][25][26][42][43][44][45][46][47]. It was found that HE4 overexpression or rHE4 treatment in EOC cells resulted in upregulation of many transcripts coding for extracellular matrix proteins, including LAMC2, LAMB3, SERPINB2 and GREM1; moreover, in cells overexpressing HE4 or exposed to rHE4 in culture medium, the protein levels of LAMC2 and LAMB3 were continously increased, and in the presence of bronectin, the focal adhesions were elevated in cells treated with rHE4 [22].…”

“…Furtherly, it was found in vitro that HE4 may regulate the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase/AKT signal transduction (PI3K/AKT) pathways to produce tumor-suppressing effect [18,19]. Recently, emerging studies have been carried out to investigate the association between HE4 and tumorigenesis as well as chemotherapeutic resistance in EOC, whereas the studies were inconclusive due to inconsistencies in results [20][21][22][23][24].…”