Rat anti-Mouse IL-24 monoclonal antibody, Biotin conjugated

Melanoma differentiation-associated-7 (mda-7)/interleukin-24 (IL-24), a unique cytokine-tumor suppressor, exerts tumor-selective killing activity in numerous types of cancer cell. Although eukaryotically and prokaryotically expressed recombinant human (rh)IL-24 proteins have been previously shown to produce potent antitumor effects, to the best of our knowledge, no side-by-side study has been conducted that compares the two proteins directly. In the present study, rhIL-24 protein was expressed in BL21 Escherichia coli transformed with the pET-21a(+)-hIL-24 plasmid by isopropyl-beta-D-1-thiogalactopyra noside induction. Following a denaturing and renaturing process, the soluble rhIL-24 was purified using a Q-Sepharose column. rhIL-24 protein was also expressed in Chinese hamster ovary mammalian cells stably transfected with the pcDNA3-hIL-24 plasmid. The in vitro antitumor efficacies of the two treatments were compared using the MDA-MB-231 human breast cancer cell line. Furthermore, the therapeutic efficacies of the bacteria-derived rhIL-24 protein and the liposome-coated pcDNA3-hIL-24 naked plasmid were evaluated in athymic nude mice with subcutaneously xenografted MDA-MB-231 cell tumors. The prokaryotically expressed/purified rhIL-24 protein and the eukaryotically expressed rhIL-24 in the cell supernate were revealed to be capable of efficiently suppressing MDA-MB-231 tumor growth in vitro. Similarly, the administration of bacteria-derived rhIL-24 protein and pcDNA3-hIL-24 naked plasmid also provided therapeutic benefits in the treatment of in vivo MDA-MB-231 xenografted tumors. The retarded in vitro and in vivo breast cancer growth elicited by rhIL-24 was closely associated with the upregulation of the ratio of anti-apoptotic B cell lymphoma 2 (Bcl-2) to pro-apoptotic Bcl-2-associated X protein (Bax), as well as the activation of caspase-3 followed by marked induction of apoptosis, and the notable inhibition of tumor angiogenesis. Thus, the results of the present study indicate that prokaryotically expressed rhIL-24 protein may be an alternate and promising antitumor agent in human breast cancer or other types of cancer.

Recombinant human interleukin 24 (rhIL24) is a member of the interleukin 10 (IL10) family of cytokines with novel therapeutic properties. Human IL24 possesses three N glycosylation sites and a disulfide bridge. The cost and composition of culture media is critical for commercial-scale production of recombinant proteins in E. coli. Addition of yeast extract and glucose to medium enhances rhIL24 production, and the use of lactose instead of IPTG for induction drops the cost and decreases toxicity. In addition, a two-step denaturing and one-step refolding (2DR) strategy improves rhIL24 production. The 2DR strategy replaces a more conventional approach for protein solubilization and refolding. LC-MS/MS provides definitive identification and quantitative information on rhIL24. Single-step purified rhIL24 displayed biological activity on HepG2 hepatocellular carcinoma cells, but no effect on L02 cells. Proliferation analysis suggests that rhIL24 may have potential use as a medication. In the present study, we developed a simple process for producing quality product with high purity. The expression and purification of rhIL24 described here may be a step towards inexpensive large-scale production.