Overcoming bortezomib resistance in human B cells by anti-CD20/rituximab-mediated complement-dependent cytotoxicity and epoxyketone-based irreversible proteasome inhibitors
1 Department of Rheumatology, VU University Medical Center, Amsterdam, The Netherlands
2 The Fred Wyszkowski Cancer Research Laboratory, Faculty of Biology, The Technion-Israel Institute of Technology, Haifa, Israel
3 Department of Hematology, VU University Medical Center, Amsterdam, The Netherlands
4 Department of Medical Oncology, VU University Medical Center, Amsterdam, The Netherlands
5 Department of Pathology, VU University Medical Center, Amsterdam, The Netherlands
6 Onyx Pharmaceuticals, South San Francisco, CA, USA
Experimental Hematology & Oncology 2013, 2:2 doi:10.1186/2162-3619-2-2Published: 10 January 2013
In clinical and experimental settings, antibody-based anti-CD20/rituximab and small molecule proteasome inhibitor (PI) bortezomib (BTZ) treatment proved effective modalities for B cell depletion in lymphoproliferative disorders as well as autoimmune diseases. However, the chronic nature of these diseases requires either prolonged or re-treatment, often with acquired resistance as a consequence.
Here we studied the molecular basis of acquired resistance to BTZ in JY human B lymphoblastic cells following prolonged exposure to this drug and examined possibilities to overcome resistance by next generation PIs and anti-CD20/rituximab-mediated complement-dependent cytotoxicity (CDC).
Characterization of BTZ-resistant JY/BTZ cells compared to parental JY/WT cells revealed the following features: (a) 10–12 fold resistance to BTZ associated with the acquisition of a mutation in the PSMB5 gene (encoding the constitutive β5 proteasome subunit) introducing an amino acid substitution (Met45Ile) in the BTZ-binding pocket, (b) a significant 2–4 fold increase in the mRNA and protein levels of the constitutive β5 proteasome subunit along with unaltered immunoproteasome expression, (c) full sensitivity to the irreversible epoxyketone-based PIs carfilzomib and (to a lesser extent) the immunoproteasome inhibitor ONX 0914. Finally, in association with impaired ubiquitination and attenuated breakdown of CD20, JY/BTZ cells harbored a net 3-fold increase in CD20 cell surface expression, which was functionally implicated in conferring a significantly increased anti-CD20/rituximab-mediated CDC.
These results demonstrate that acquired resistance to BTZ in B cells can be overcome by next generation PIs and by anti-CD20/rituximab-induced CDC, thereby paving the way for salvage therapy in BTZ-resistant disease.