Abstract
Velcade is one of the inescapable drug to treat patient suffering from multiple myeloma (MM) and resistance to this drug represents a major drawback for patients. However, the mechanisms underlying velcade resistance remain incompletely understood. We derived several U266 MM cell clones that resist to velcade. U266-resistant cells were resistant to velcade-induced cell death but exhibited a similar sensitivity to various proapoptotic stimuli. Careful analysis of proteosomal subunits and proteasome enzymatic activities showed that neither the composition nor the activity of the proteasome was affected in velcade-resistant cells. Elimination of velcade-induced poly-ubiquitinated proteins and protein aggregates was drastically stimulated in the resistant cells and correlated with increased cell survival. Inhibition of the lysosomal activity in velcade-resistant cells resulted in an increase of cell aggregates and decrease survival, indicating that aggregates are eliminated through lysosomal degradation. In addition, pangenomic profiling of velcade-sensitive and resistant cells showed that the small heat shock protein HSPB8 was overexpressed in resistant cells. Finally, gain and loss of function experiment demonstrated that HSPB8 is a key factor for velcade resistance. In conclusion, HSPB8 plays an important role for the elimination of aggregates in velcade-resistant cells that contributes to their enhanced survival.
Highlights
The ubiquitin-proteasome pathway plays an essential role in the degradation of 80% of ubiquitin-tagged intracellular proteins, many of which play a regulatory role in cell proliferation, cell survival, and signaling processes [1,2,3]
Generation and characterization of multiple myeloma cell lines resistant to velcade To gain insights into the molecular mechanisms of resistance to conventional therapies in MM, we generated U266 cell clones resistant to velcade, one of the leading drug to treat patients suffering from MM
To disprove that increased expression of HSPB8 could be a clonal particularity of the R6 cell line, we looked for its expression in the bulk of U266 resistant cells and in the different U266 cell clones
Summary
The ubiquitin-proteasome pathway plays an essential role in the degradation of 80% of ubiquitin-tagged intracellular proteins, many of which play a regulatory role in cell proliferation, cell survival, and signaling processes [1,2,3]. Proteasome inhibition leads to the accumulation of damaged proteins in the intracellular environment which causes endoplasmic reticulum overload integrated as an ER stress [9]. If the accumulation of damaged proteins is excessive, cell can at last triggers apoptosis through CHOP, caspase-4 and caspase-12 activation. It was reported that proteasome inhibition triggers apoptosis by affecting the levels of many regulatory proteins, resulting in inhibition of nuclear factor κB (NF-κB), increased activity P53 and Bax leading to the activation of c-Jun NH2 –terminal kinase (JNK), which in turn activates caspase-8 and caspase-3 [10]
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