A Humanized Anti-IGF-1R Monoclonal Antibody (AVE1642) Synergizes with Bortezomib To Induce Apoptosis in CD45neg Myeloma Cells.

IGF-1 is a well-known growth factor for human myeloma cells. Thus, therapeutic strategies targeting IGF-1R have been proposed for the treatment of multiple myeloma. In this study, we investigated the effect of a humanized antagonistic monoclonal antibody to the IGF-1R (AVE1642). We showed that AVE1642 selectively inhibits IGF-1R but not insulin signaling in human myeloma cell lines (HMCL). Since we had previously shown the functional relevance of CD45 expression in the growth of myeloma cells, and the association of CD45 negative (CD45neg) status with a less favorable clinical outcome, both CD45 positive (CD45pos) and CD45neg HMCL were studied. We found that AVE1642 strongly inhibited the growth of CD45neg HMCL, leading to a G1 growth arrest, whereas it had almost no effect on the growth of CD45pos HMCL. Indeed, by cell counting over four days in standard culture conditions, AVE1642 induced a significant growth inhibition of the CD45neg LP-1 HMCL of 90% whereas it had no effect on the growth of the CD45pos XG-1 HMCL. Furthermore, AVE1642 binding induced a significant reduction of IGF-1R expression. In western blot, a decrease of 60% the IGF-1R expression of the LP-1 CD45neg HMCL was observed. This decrease was also confirmed by flow cytometry analysis. Finally, a stable CD45 silencing XG-1 cell line was generated and became sensitive to AVE1642. By cell counting over four days in standard culture conditions, AVE1642 inhibited the growth of the shCD45neg XG-1 HMCL of 56% whereas it had almost no effect on the growth of the shLuci-XG-1 HMCL. Thus, for the first time, we provided direct evidence that the expression of CD45 renders myeloma cells resistant to IGF1R inhibition. Furthermore, AVE1642 in combination with Bortezomib strongly increased the apoptosis induced by Bortezomib alone on CD45negHMCL. The CD45neg LP-1 HMCL was considered as a modestly sensitive cell line since 10nM of Bortezomib, which is in the range of therapeutic concentrations, induced 40±16% of apoptosis. Bortezomib in combination with AVE1642 was found to induce 80±1% of apoptosis in the CD45neg LP-1 HMCL. Apoptosis induction by AVE1642/Bortezomib combination was associated with an important induction of Noxa, a BH3-only pro-apoptotic protein that was increased by the addition of AVE1642. We also found a strong increase of caspase-3, -8 activation under this combination compared to Bortezomib alone. Taken together, these results support that therapy directed against IGF-1R associated with Bortezomib, could be beneficial in treating CD45neg patients, a population known for poor clinical outcome.