Development of a novel interferon-[alpha]2b gene construct with a repetitive hypoxia-inducible factor binding site and its suppressive effects on human renal cell carcinoma cell lines in vitro

• Published in: International journal of clinical oncology Vol. 19; no. 3; p. 497
• Main Authors: Fukui, Naotaka, Kageyama, Yukio, Higashi, Yotsuo, Kihara, Kazunori, Kizaka-kondoh, Shinae, Hiraoka, Masahiro, Shinojima, Toshiaki, Suzuki, Kenjiro, Oya, Mototsugu
• Format: Journal Article
• Language: English
• Published: Tokyo Springer Nature B.V 01-06-2014
• Subjects: Binding sites; Gene therapy; Hypoxia; Interferon; Kidney cancer; Oncology
• ISSN: 1341-9625; 1437-7772
• DOI: 10.1007/s10147-013-0568-z

Despite the advent of targeted therapies, interferon-alpha (IFN-[alpha]) remains a therapeutic option for advanced renal cell carcinoma (RCC), especially in Japan, with a treatment response rate of 15-20 %. To improve the efficacy of IFN-[alpha]-based therapies, we evaluated a novel treatment strategy for RCC using an IFN-[alpha]2b gene construct with a repetitive hypoxia-inducible factor binding site. We constructed an expression plasmid designated 5HREp-IFN-[alpha]2b containing the coding region of the IFN-[alpha]2b gene. Five copies of the hypoxia-response element (HRE) sequences were inserted upstream of the IFN-[alpha]2b gene, and the construct was transfected into human RCC cell lines ACHN, 786-O and KU19-20. The concentrations of IFN-[alpha]2b in the conditioned media were measured by enzyme-linked immunosorbent assay. Cell viabilities were determined by MTS assays. Construct-induced IFN-[alpha] secretion was confirmed in all three cell lines. IFN-[alpha] production was significantly enhanced by the hypoxia-mimicking agent deferoxamine mesylate in cell lines expressing the wild-type von Hippel-Lindau (VHL) gene (KU19-20 and ACHN) compared with cells expressing the mutant VHL gene (786-O). The construct exerted significant suppressive effects on the viabilities of all RCC cell lines. This is the first study to report on the construction of a cytokine gene with a repetitive hypoxia-inducible factor binding site and its application in the suppression of human cancer cells. Gene therapy using this IFN-[alpha]2b gene construct with HREs may represent a novel treatment modality for advanced RCC.[PUBLICATION ABSTRACT]