Patients with multiple myeloma (MM) are treated with chemotherapy and recently with the proteasome inhibitor Bortezomib. However, there are patients who either do no respond to such treatments or develop resistance suggesting the need of novel therapeutic approaches. We have developed an anti-human transferrin receptor IgG3-avidin fusion protein (anti-hTfR .IgG3-Av) to target the TfRwith the objective of targeting the transferrin receptor on tumor cells and induceinducing tumor cell killing with a biotinyalated toxin. We have found, however, that anti-hTfR .IgG3-Av treatment alone could inhibit tumor cell proliferation and was also cytotoxic to some but not all MM cell lines studied.. We hypothesized that the cytotoxic potential of anti-hTfR .IgG3-Av may alter the anti-apoptotic pathway of tumor cells reducingand reduce the anti-apoptotic threshold and sensitizingthus can sensitize tumor cells to apoptosis by chemotherapeutic drugs. This hypothesis was examined with IM-9 and 8226S 2 MM cell lines both of which are resistant to CDDP-induced apoptosis. Treatment of MM cells with various suboptimal concentrations of anti-hTfR .IgG3-Av followed by treatment with CDDP (non-biotinylated) resulted in significant potentiation of apoptosis and synergy was achieved. We examined the underlying mechanism of synergy. Cell treatment with anti-hTfR .IgG3-Av resulted in significant mitochondrial depolarization, activation of caspase 9 and inhibition of the anti-apoptotic gene products XIAP and Bcl-xL. The combination treatment with CDDP enhanced these effects and resulted in caspase 3 activation and apoptosis. The inhibitioninhibiton of Bcl-xL and XIAP by anti-hTfR .IgG3-Av suggested that it may inhibit survival pathways that regulate the transcription of these gene products. It has been reported that Bcl-xL and XIAP are transcriptionally regulated by the NF-κB pathway and Bcl-xL function is regulated by the Akt pathway through phosphorylation of Bad and the inhibition of Bad-Bcl-xL complexes. In addition, the Akt pathway cross-talks and also regulates the NF-κB pathway. We demonstrate that treatment of MM cells with anti-hTfR.IgG3-Av inhibited the constitutively activated Akt pathway suggesting its role in the downstream regulation of XIAP and Bcl-xL and anti-hTfR .IgG3-Av-mediated chemosensitization. These findings demonstrate that the combination of anti-hTfR .IgG3-Av and CDDP results in activation of the mitochondrial pathway for apoptosis, inhibition of XIAP and Bcl-xL which leads to synergistic apoptosis. InhibitorsInhibitors of the Akt pathway have been used clinically (example: rapamycin) and many rapamycin analogues are currently in clinical trials. The inhibition of the Akt pathway by anti-hTfR .IgG3-Av provides a novel Akt inhibitor and its potential use in cancer therapy.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA