A dominant-negative F-box deleted mutant of E3 ubiquitin ligase, β-TrCP1/FWD1, markedly reduces myeloma cell growth and survival in mice
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Ramaswamy Sharma1,*, Paul J. Williams1,*, Anjana Gupta1, Brandon McCluskey1, Shylesh Bhaskaran1,2, Steve Muñoz1,3 and Babatunde O. Oyajobi1,4
1 Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX, USA
2 Current address: Oklahoma Medical Research Foundation, Oklahoma City, OK, USA
3 Current address: Vanderbilt Center for Bone Biology, Vanderbilt University Medical Center, Nashville, TN, USA
4 Cancer Therapy and Research Center, San Antonio, TX, USA
* These authors contributed equally to this work
Babatunde O. Oyajobi, email:
Keywords: FWD1/β-TrCP1, 5TGM1, myeloma, plasmacytoma, proteasome
Received: December 23, 2014 Accepted: April 30, 2015 Published: May 12, 2015
Treatment of multiple myeloma with bortezomib can result in severe adverse effects, necessitating the development of targeted inhibitors of the proteasome. We show that stable expression of a dominant-negative F-box deleted (∆F) mutant of the E3 ubiquitin ligase, SCFβ-TrCP/FWD1, in murine 5TGM1 myeloma cells dramatically attenuated their skeletal engraftment and survival when inoculated into immunocompetent C57BL/KaLwRij mice. Similar results were obtained in immunodeficient bg-nu-xid mice, suggesting that the observed effects were independent of host recipient immune status. Bone marrow stroma offered no protection for 5TGM1-∆F cells in cocultures treated with tumor necrosis factor (TNF), indicating a cell-autonomous anti-myeloma effect. Levels of p100, IκBα, Mcl-1, ATF4, total and cleaved caspase-3, and phospho-β-catenin were elevated in 5TGM1-∆F cells whereas cIAP was down-regulated. TNF also activated caspase-3 and downregulated Bcl-2, correlating with the enhanced susceptibility of 5TGM1-∆F cells to apoptosis. Treatment of 5TGM1 tumor-bearing mice with a β-TrCP1/FWD1 inhibitor, pyrrolidine dithiocarbamate (PDTC), significantly reduced tumor burden in bone. PDTC also increased levels of cleaved Mcl-1 and caspase-3 in U266 human myeloma cells, correlating with our murine data and validating the development of specific β-TrCP inhibitors as an alternative therapy to nonspecific proteasome inhibitors for myeloma patients.
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