RNA sequencing reveals that withaferin A reduces overexpression of Bruton's tyrosine kinase in glucocorticoid resistant multiple myeloma cells

Multiple myeloma (MM) is an incurable hematological malignancy characterized by the uncontrolled growth of plasma cells in the bone marrow. The major barrier in treating MM is the occurrence of primary and acquired therapy resistance to multiple existing anti-cancer drugs. Often, this therapy resistance is associated with constitutive activation of Bruton tyrosine kinase (BTK) dependent B-cell receptor (BCR) signaling. Novel kinase inhibitors covalently targeting BCR signaling, including the clinically approved BTK inhibitor ibrutinib (IBR) and the preclinical phytochemical Withaferin A (WA), have therefore gained pharmaceutical interest. Remarkably, glucocorticoid (GC)-resistant MM cells overexpressing BTK are more sensitive to WA then to IBR. To further characterize the kinase inhibitor profiles of WA and IBR in GC-resistant MM cells, we applied phosphopeptidome- and transcriptome-specific tyrosine kinome profiling. Our results demonstrate that WA treatment triggers dual inhibition of BCR signaling by transcriptional downregulation and covalent cysteine-dependent kinase inhibition of BTK. Covalent interaction between WA and BTK could further be confirmed by biotin-based affinity purification and confocal imaging microscopy. Altogether, we show that covalent BCR-BTK kinase inhibition by WA represents an attractive strategy to treat GC-resistant MM. mRNA profiles of glucocorticoid sensitive MM1.S and glucocorticoid resistant MM1.R cell lines after treatment with withaferin A