Abstract 3815: Synergistic mechanisms of oncolytic reovirus with bortezomib in overcoming therapy resistance of multiple myeloma
Authors: Thirukkumaran Chandini M. , Shi Zhong-Qiao , Luider Joanne , Kopciuk Karen , Neri Paola , Bahlis Nizar , Morris Don 
Published: August 2015

Abstract
Introduction: Multiple myeloma (MM) is an incurable hematologic malignancy that accounts for 10% of hematopoietic neoplasms. The potential of reovirus (RV) as a novel therapeutic agent for MM under in vitro, in vivo and ex vivo conditions has been demonstrated by us and a phase I clinical trial of MM with RV therapy has shown indications of efficacy. Recently we have shown that RV could synergize with several MM drugs such as bortezomib (Bort), cafilzomib, dexamethasone and perifosine and importantly, enhance its therapeutic potential in therapy resistant MM. Here we show that RV/Bort synergy in MM is mediated via enhanced apoptosis than augmented viral progeny production.
Methods: MM cell lines RPMI 8226, U266, NCIH929, U266, INA-6, KMS11 and OPM2 were incubated with live or dead RV at a MOI of 40 for 24, 48, and 72 hours respectively and cell death was assessed via the WST assay. RV resistant OPM2 and moderately resistant KMS-11 were used in combination therapy and treated with RV and drugs at various concentrations to evaluate Effective Dose 50% (ED50) values and synergism was determined by the Chou-Talalay method. To assess viral progeny production, MM cells were treated with RV+/- drugs and harvested at various time points and frozen. Following 3 freeze thaw cycles viral progeny was assessed via plaque titration on L929 cells. To assess apoptosis, cells treated with RV or Bort singly or in combination were harvested 48 hours and stained with AnnexinV/FITC and 7AAD and analyzed by flow cytometry.
Results: RV sensitive RPMI8226 cells demonstrated the maximal viral progeny production. Log-fold increases in viral plaques forming units (PFU)/ml detected at 72h was 3.04 for RV+Bort treated cells and 3.58 for RV alone treated cells indicating productive virus replication in both treatments. RV resistant OPM2 cells demonstrated the least amounts of virus progeny (0.99-1.28 PFU/ml fold increase) in RV+Bort treated or RV alone treated cells respectively. KMS11 cells treated with RV alone showed a log-fold increase in virus progeny of 2.32PFU/ml at 72h. Interestingly, when KMS11 cells were treated with RV+Bort no significant increase in RV progeny was seem at 72h indicating minimal RV replication in these cells in the presence of Bort.
The synergistic cytotoxicity observed with RV+Bort in all 3 cell lines was due to enhanced apoptosis where significantly augmented apoptosis was seen in combination treatment than in single agent treatment.
Conclusion: Not all MM cell lines are amenable to RV mediated cell death. This has important implications for the future use of RV as a therapeutic agent. Mechanisms such as autophagy and necroptosis that may contribute to RV/drug mediated cytotoxicity is currently been investigated and experimentation in animal models is also underway. Understanding the signalling pathways of resistant tumour will help develop a more personalized approach of RV therapy for MM patients in the future.
Citation Format: Chandini M. Thirukkumaran, Zhong-Qiao Shi, Joanne Luider, Karen Kopciuk, Paola Neri, Nizar Bahlis, Don Morris. Synergistic mechanisms of oncolytic reovirus with bortezomib in overcoming therapy resistance of multiple myeloma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3815. doi:10.1158/1538-7445.AM2015-3815