Expression Of Lentivirus - Mediated DR5 Chimeric Antibody And Its Antitumor Activity

TNF-related apoptosis-inducing ligand (TRAIL) belongs to the TNF superfamily which inducing tumor cells apoptosis via death receptors of the target cell without adverse effects. Five receptors have been identified of which two death receptors, TRAIL receptor 1 (death receptor 4, DR4) and TRAIL receptor 2 (death receptor 5, DR5), containing death domain (DD). They interact with FADD to recruit several cytolic proteins to formate the death-inducing signaling complex (DISC) which can trigger apoptosis pathway. Other three receptors, Decoy receptor 1 (DcR1), Decoy receptor 2 (DcR2) and osteoprotegerin (OPG) display high sequence homology in their extracellular domains with DR4 and DR5, but have not a completed DD, are therefore unable to induce apoptosis.In the previous study, we developed a novel mouse anti-human DR5 monoclonal antibody AD5-10 inducing various tumor cells apoptosis in vitro and inhibiting tumor growth in vivo. More research found that AD5-10 did not induce apoptosis of human normal hepatocytes and primary peripHeral-blood lympHocytes.The abilities of Lentivirus (LV) to transduce nondividing cells and integrate to the genome for long-terrn expression of the target cell have prompted many different gene therapy applications. Moreover, the capacity of LV to accommodate relatively large transgenes is an advantage over adeno-associated viral vectors (AAVs).To investigate the prospect of our m Ab in the gene therapy of tumor, we constructed a lentivirus vector which expresses the full-lenth chimeric mAb based on AD5-10, Zaptuzumab, by linking the heavy chain and the light chain with 2A/Furin self-processing peptide derived from the foot-and-mouth disease virus.Transient transfection in HEK293 cells with a lentiviral expression vector encoding the chimeric antibody resulted in 12±0.7μg/ml of Zaptuzumab expression in medium for over 6 months. Zaptuzumab in the medium could bind to its antigen DR5 specifically as demonstrated by ELISA and Western Blotting assay, and possessed a significant apoptosis-inducing activity in various tumor cell lines over 50%. Antibody-dependent cell-mediated cytotoxicity (ADCC) and the complement dependent cytotoxicity (CDC) of Zaptuzumab were also detected.The results showed that peripHeral blood mononuclear cells activated by IL-2 exhibited Zaptuzumab-mediated ADCC activity against lung cancer cell lines. And packaged virus, Lenti-H2AL, showed great apoptosis-inducing activity in vitro. Meanwhile, it inhibited human tumor conformation in nude mice significantly. Moreover, cooperated with Mitomycin can prolong the life span of the lung A549 tumor-bearing nude mice. This recombinant vector provides a feasible chimeric antibody delivery approach and a novel strategy for cancer antibody therapy.The plastic adherent cells from the bone marrow (BM) referred to as mesenchymal stem cells (MSC) are capable of self-renewing and have the potential to differentiate into mesenchymal and nonmesenchymal tissues. The ability of MSC to migrate to the areas of injury and to tumours has encouraged investigation of MSC as therapeutic tools. In the second part of the research, we transfected the mesenchymal stem cells(MSC) with Lenti-H2AL, and demonstrated the expression and the tumor- killing-ability of MSC-H2AL. The result of the use of Lenti-H2AL transfected MSC provide new data in Stem Cell therapy.Studies on Lenti-H2AL may help us to well understand on the functions of the antibody as potential cancer therapeutic agent and also provide a novel strategy for cancer biotherapy.