Abstract C205: Cytotoxicity and target modulation in pediatric solid tumors by the proteasome inhibitor carfilzomib.

Abstract

Abstract Rationale: Currently, even with escalating multimodal treatment regimens, most children with recurrent metastatic solid tumors endure unacceptably high mortality rates. Hence, there is an urgent need to identify novel targets and new therapeutic approaches. Recent studies have shown that proteasome inhibition leads to effective tumor killing in cells that have acquired treatment resistance and metastatic properties. Carfilzomib (CFZ) is a selective and potent proteasome inhibitor that binds and inhibits the 20S proteasome resulting in the accumulation of polyubiquinated proteins and consequently, cycle arrest, growth inhibition and apoptosis. Clinical trials in adult myeloma have shown activity and a high tolerability of CFZ. However, data with respect to the potential of this agent for refractory pediatric solid tumors are not yet available. Methods: Cells from a panel of lines including neuroblastoma (n=6), Ewings sarcoma (n=2), osteosarcoma (n=2), rhabdomyosarcoma (n=4) and ATRT (n=2), were treated with increasing concentrations of CFZ and cell growth inhibition was quantified by Alamar blue assay. Drug pulsing experiments were carried out by adding new drug preparations at defined time intervals. Target modulation and apoptosis analyses were done by Western blotting. Drug combination studies were interpreted using the Chou and Talalay method. Results and Discussion: CFZ showed effective cytotoxicity against all cell lines tested (mean IC50 = 7nM, range = 1-20nM) and activity in a fluorophore tagged cell based proteasome assay. More detailed target modulation studies in NB cells showed a dose dependent initial up-regulation of MCL-1 that subsequently decreased in a manner corresponding with PARP cleavage. Up-regulation of BCL-2 was also noted but required the exposure to higher drug concentrations. This indicated the initial stabilization or activation of survival pathways in response to drug treatment. Drug scheduling showed that the minimum exposure of 4 to 8 hours /day is needed for effective cumulative killing coinciding with MCL-1 regulation. Drug combination studies identified the ability of CFZ to synergistically enhance the activity of a number of chemotherapeutic agents, including etoposide, vincristine, mefloquine and the BCL-2 antagonist ABT-263. However, the extent of synergy differed greatly between cell types indicating the potentially variable relationship of proteasome functions to distinct oncogenic pathways present in different cells. Our studies provide initial in vitro data on the potential of CFZ to treat pediatric solid tumors and support further investigations in to the components of drug scheduling, biological correlates and drug combinations for future early phase clinical trials. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C205. Citation Format: Yibing Ruan, David Liu, Aarthi Jayanthan, Tony Truong, Jessica Boklan, Aru Narendran. Cytotoxicity and target modulation in pediatric solid tumors by the proteasome inhibitor carfilzomib. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C205.