Inhibitory Functions of SUSD2 in the Progression of High-Grade Serous Ovarian Carcinoma

• Main Author: Sheets, Jordan N.
• Language: EN
• Published: University of South Dakota 2017
• Subjects: Cellular biology|Medicine|Oncology
• Online Access: http://pqdtopen.proquest.com/#viewpdf?dispub=10633628

<p> <i>Sushi Domain Containing 2 (SUSD2)</i> encodes a type I transmembrane protein containing several functional domains inherent to adhesion molecules. A clinically annotated HGSOC tissue microarray was stained with an anti-SUSD2 antibody. Patients with tumors that had weak SUSD2 staining had a shorter median survival (31.7 months) compared to patients that had tumors with strong SUSD2 staining (49.1 months; p value = 0.0083).</p><p> To investigate the role of SUSD2 in HGSOC, stable OVCAR3, OVSAHO and KURAMOCHI cell lines were established through transfection of shRNA targeted to <i>SUSD2</i> transcripts (SUSD2 knock-down [KD] cell lines) or non-targeting shRNA (SUSD2 NT) cell lines. Boyden chamber and wound healing assays demonstrated that OVCAR3, OVSAHO and KURAMOCHI SUSD2 KD cells migrated at significantly higher rates than their SUSD2 NT counterpart cell lines. RT-qPCR and western immunoblot analysis indicated an inverse relationship between SUSD2 and well characterized mesenchymal genes, such as <i>TWIST1, ZEB1</i> and <i>CHD2.</i> In addition, OVCAR3 and KURAMOCHI SUSD2 KD spheroids displayed increased mesothelial clearance ability compared to SUSD2 NT spheroids.</p><p> To explore the potential for SUSD2 to inhibit late-stage HGSOC metastasis, female athymic nude mice were injected with either OVCAR3 NT or OVCAR3 KD cells. Fewer nodules were observed in the pancreas and omentum of the OVCAR3 NT mice when compared to the OVCAR3 KD mice. Furthermore, OVCAR3 KD mice had a significantly shorter median survival compared to OVCAR3 NT mice (175 days compared to 185.5 days, respectively; p-value = 0.0047).</p><p> KURAMOCHI lysate was immunoprecipitated for SUSD2-associated immunocomplexes and subjected to liquid chromatography, tandem mass spectrometry (LC-MS/MS) analysis, yielding a list of candidate SUSD2-interacting proteins associated with RNA processing. Immunofluorescence analysis of OVCAR3, KURAMOCHI and SKBR3 cells and western immunoblot analysis of their subcellular extracts revealed SUSD2 to be present in cell nuclei, mitochondria and cytoplasm; however, SUSD2 was relatively less abundant in SKBR3 nuclei.</p><p> Our findings suggest that increased <i>SUSD2</i> expression in HGSOC impedes metastasis, consistent with prolonged survival observed in HGSOC patients with high <i>SUSD2</i>-expressing primary tumors. The differences in subcellular distribution between HGSOC cells and breast cancer cells may explain alternate functions of SUSD2 in different cancers.</p><p>