title: Lentivirus-mediated RNA Interference Targeting LAPTM4B Inhibits Human Ovarian Cancer Cell Invasion In Vitro.
abstract: LAPTM4B (lysosome-associated protein transmembrane 4 beta) play an important role in several human carcinomas. We examines the effects of RNA interference mediated downregulation of human lysosomal-associated protein transmembrane 4 beta expression on the biological behavior of the human serous adenocarcinoma cell line NIH:OVCAR3. This study investigated the expression level of lysosomal-associated protein transmembrane 4 beta in several ovarian cancer cell lines. RNA interference mediated by recombinant lentiviral vectors expressing an artificial lysosomal-associated protein transmembrane 4 beta miRNA was used to induce long-lasting downregulation of lysosomal-associated protein transmembrane 4 beta gene expression in NIH:OVCAR3 cells. Lysosomal-associated protein transmembrane 4 beta expression as well as the motility, migration potential, and proliferation of the tumor cells was measured by flow cytometry, real-time polymerase chain reaction, Western blot analysis, transwell migration assays, wound healing assays, and cell counting kit-8 assays. In addition, the cell cycle analysis utilized fluorescence-activated cell sorting. Four recombinant plasmid expression vectors encoding premiRNAs against lysosomal-associated protein transmembrane 4 beta (pcDNA-LAPTM4B-miR-1, -2, -3, and-4) were constructed and transfected into 293T cells, which overexpress lysosomal-associated protein transmembrane 4 beta. The recombinant lentiviral vector for lysosomal-associated protein transmembrane 4 beta RNA interference was packaged with pcDNA-LAPTM4B-miR-3, which had the highest interfering efficiency, thereby successfully generating stable transfectants. Compared with the control cells, the LAPTM4B-miRNA-transfected NIH:OVCAR3 cells exhibited significant decreases in cell motility and invasion. Furthermore, LAPTM4B depletion resulted in a significant decrease in proliferating cell nuclear antigen, vascular endothelial growth factor, MMP2, MMP9, and CDK12 expression. We propose that lysosomal-associated protein transmembrane 4 beta expression may be an oncogene-inducing feature of invasive ovarian cancer cells and may be a potential therapeutic target for ovarian cancer treatment.
authors.affiliation: Department of Gynecology, The Affiliated Tumor Hospital, Harbin Medical University, Harbin, 150081, China.
authors.forename: Fanling
authors.initials: F
authors.lastname: Meng
authors.affiliation: Department of Gynecology, The Affiliated Tumor Hospital, Harbin Medical University, Harbin, 150081, China.
authors.forename: Xiuwei
authors.initials: X
authors.lastname: Chen
authors.affiliation: Department of Pathology, The Affiliated Tumor Hospital, Harbin Medical University, 150 Ha Ping Road, Nangang District, Harbin, Heilongjiang Province, 150081, China.
authors.forename: Hongtao
authors.initials: H
authors.lastname: Song
authors.affiliation: Department of Gynecology, The Affiliated Tumor Hospital, Harbin Medical University, Harbin, 150081, China.
authors.forename: Ge
authors.initials: G
authors.lastname: Lou
authors.affiliation: Laboratory of Medical Genetics, Harbin Medical University, 194 Xue Fu Road, Nangang District, Harbin, Heilongjiang Province, 150086, China.
authors.forename: Songbin
authors.initials: S
authors.lastname: Fu
mesh_terms: D002453:Cell Cycle; D049109:Cell Proliferation; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D016086:Lentivirus / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics*; D035683:MicroRNAs / Q000235:genetics; D009363:Neoplasm Proteins / Q000378:metabolism; D015513:Oncogene Proteins / Q000235:genetics*; D010051:Ovarian Neoplasms / Q000473:pathology*; D034622:RNA Interference
chemical_list: C477083:LAPTM4B protein, human; D008565:Membrane Proteins; D035683:MicroRNAs; D009363:Neoplasm Proteins; D015513:Oncogene Proteins
keywords: LAPTM4B; RNA interference; angiogenesis; invasion; ovarian cancer; proliferation