HOOKING TRAIL-R2 TO ¿EDUCATE¿ IMMUNE SYSTEM TO RECOGNIZE CANCER CELLS

Use of recombinant human tumor necrosis factor (TNF) related apoptosis inducing ligand (rhTRAIL) or TRAIL-receptors agonistic monoclonal antibodies (mAbs) promotes apoptosis in most cancer cells preserving normal cells. Furthermore, TRAIL-R2, one of the receptor that can trigger cell apoptotic machinery, is expressed at higher levels in tumor tissue as compared to normal tissues. Agonistic mAbs could mimic the tumor killing properties of rhTRAIL, with the advantage to have a prolonged half-life in vivo and to avoid binding of decoy TRAIL-receptors unable to transmit apoptotic signals. Although mAbs have proven to be highly effective, their use still unsolved the problem of TRAIL-resistance that some tumor cells are able to develop. To increase the anti-tumor activity of some mAbs, in last decades, bispecific antibodies (BsAbs) able to redirect cytotoxic activity of tumor resident/ circulating T cells against tumor cells in an MHC-independent fashion have been developed. In this study, we isolated a BsAb able to bind the CD3, invariant component of the T-cell receptor, and TRAIL-R2. Among the different isolated BsAbs, one in particular presented good thermodynamic characteristics and showed a very good stability even after 2 years its purification. The BsAb was biochemically and functionally well characterized and the production method was optimized. We demonstrated that the BsAb could act with two mechanisms: retargeting T-cells to lyse tumor cells when acting as BsAb and activating apoptotic pathway by triggering TRAIL-R2 when acting as agonistic mAb. T-cell activation was confirmed by up-regulation of CD69 and CD25 and by production of inflammatory cytokines without off target toxicity. In vitro we demonstrated that, after BsAb-mediated retargeting, T cells were able to lyse tumors of different histotypes, like melanoma, breast and ovarian cancer while sparing normal cells.