Construction of recombinant human IFNα-2b BCG and its antitumor effects on bladder cancer cells in vitro

Abstract

ABSTRACT. We constructed recombinant Bacille Calmette-Guérin (rBCG) that secreted human interferon alpha 2b (hIFNa-2b), and investigated its antitumor effects on bladder cancer cells in vitro. The recombinant plasmid phIFN-a-2b was constructed using pMAO-4 and transformed into BCG. The supernatant was collected at various times and IFN-g, interleukin (IL)-12, and tumor necrosis factor (TNF)-a were detected using an enzyme-linked immunosorbent assay. EJ cells were cultivated for 24, 48, and 72 h, together with rBCG, wild-type BCG (wBCG), or wBCG+IFN-a-2b. rBCG capable of secreting cytokine IFNa-2b was constructed. On the 4th day of culture, the IFNa-2b secreted by rBCG reached a maximum. wBCG and rBCG showed no significant difference on cell growth rate over 7 days of incubation in 7H9 medium. wBCG and rBCG were both positive for acid-fast staining, and showed mycobacterial characteristics of intercellular connection in clusters with no clear abnormalities. Higher levels Antitumor effects of recombinant IFNa-2b BCG on EJ cells of IFN-g, TNF-a, and IL-12 were induced by rBCG compared with wBCG or MAO4-rBCG (P < 0.05). rBCG may induce lymphocyte proliferation; the proliferation ratio was higher than those induced by wBCG and wBCG+IFN. rBCG had direct anti-proliferative effects on EJ cells. An MTT assay showed that rBCG inhibited the proliferation of bladder cancer cells and had more activity compared with wBCG (P < 0.05). The highest anti-tumor activity of lymphocytes was stimulated by rBCG (20.31-51.22%). rBCG-IFNa-2b induces enhanced cytotoxicity against bladder cancer cells in vitro and may be used as an alternative to BCG for bladder cancer patients.

Role of Interferons in the Development of Diagnostics, Vaccines, and Therapy for Tuberculosis

“…BCG secreting IFN- α -2b induced higher levels of IFN- γ , TNF- α , IL-12, and lymphoproliferation as well as increased antiproliferative and cytotoxic effect on bladder cancer cells in vitro [72]. …”

Mycobacteria-Derived Agents for the Treatment of Urological and Renal Cancers

“…Recombinant BCG expressing human interferon-alpha 2b (hIFNα-2b) showed higher antiproliferative capacity on EJ BC cells after infection compared to wild-type BCG. Furthermore BCG-hIFNα-2b-stimulated lymphocytes trigger the production of higher levels of IFN-γ, tumor necrosis factor (TNF)-α, and IL-12, together with higher cytotoxic activity against BC cells, compared to those treated with BCG [36].…”

Intramuscular Boosting with hIFN-Alpha 2b Enhances BCGphipps-Induced Protection in a Murine Model of Leprosy

“…We decided in this study to further investigate the protection against murine leprosy induced by vaccination with M. bovis BCGphipps (BCGph) and a pharmaceutical presentation of human IFN-α 2b [51]. In recent studies, a recombinant BCG that expresses human IFN-α 2b was used to successfully treat bladder cancer [51][52][53][54][55]. Consistent with these data, our goal in this study was to analyze the protection induced by BCGph priming and hIFN-α 2b boosting in a murine model of leprosy.…”