Mitochondrial Functions, Energy Metabolism and Protein Glycosylation are Interconnected Processes Mediating Resistance to Bortezomib in Multiple Myeloma Cells.

Barbara Tavazzi,Angela Maria Amorini,Cesarina Giallongo,Alessandro Barbato,Miriam Wissam Saab,Giuseppe Lazzarino,Nunzio Vicario,Giovanni Li Volti,Francesco Di Raimondo,Alessandra Romano,Rosalba Parenti,Maria Violetta Brundo,Fabrizio Puglisi,Renata Mangione,Daniele Tibullo,Giuseppe Alberto Palumbo,Giacomo Lazzarino

doi:10.3390/biom10050696

Abstract

The proteasome inhibitor bortezomib (BTZ) has emerged as an effective drug for the treatment of multiple myeloma even though many patients relapse from BTZ therapy. The present study investigated the metabolic pathways underlying the acquisition of bortezomib resistance in multiple myeloma. We used two different clones of multiple myeloma cell lines exhibiting different sensitivities to BTZ (U266 and U266-R) and compared them in terms of metabolic profile, mitochondrial fitness and redox balance homeostasis capacity. Our results showed that the BTZ-resistant clone (U266-R) presented increased glycosylated UDP-derivatives when compared to BTZ-sensitive cells (U266), thus also suggesting higher activities of the hexosamine biosynthetic pathway (HBP), regulating not only protein O- and N-glycosylation but also mitochondrial functions. Notably, U266-R displayed increased mitochondrial biogenesis and mitochondrial dynamics associated with stronger antioxidant defenses. Furthermore, U266-R maintained a significantly higher concentration of substrates for protein glycosylation when compared to U266, particularly for UDP-GlcNac, thus further suggesting the importance of glycosylation in the BTZ pharmacological response. Moreover, BTZ-treated U266-R showed significantly higher ATP/ADP ratios and levels of ECP and also exhibited increased mitochondrial fitness and antioxidant response. In conclusions, our findings suggest that the HBP may play a major role in mitochondrial fitness, driving BTZ resistance in multiple myeloma and thus representing a possible target for new drug development for BTZ-resistant patients.

Highlights

The ubiquitin-proteasome pathway plays a key role in protein processing and degradation, transcriptional regulation, cellular stress responses and antigen presentation [1,2]
Our results showed that the BTZ-resistant clone (U266-R) presented increased glycosylated UDP-derivatives when compared to BTZ-sensitive cells (U266), suggesting higher activities of the hexosamine biosynthetic pathway (HBP), regulating protein O- and N-glycosylation and mitochondrial functions
We first evaluate the activity of the ubiquitin–proteasome system in U266-R versus U266-S cells, finding that under basal conditions, it was significantly increased in U266-R compared to in U266-S (p < 0.001, Figure S1)

Summary

Introduction

The ubiquitin-proteasome pathway plays a key role in protein processing and degradation, transcriptional regulation, cellular stress responses and antigen presentation [1,2]. Proteasome inhibition causes an imbalance between the production and degradation of proteins, leading to the accumulation of unfolded proteins and endoplasmic reticulum (ER) stress, with the activation of the unfolded protein response (UPR) These mechanisms lead to the dysregulation of Ca2+ homeostasis and the activation of cell apoptosis [3,4,5]. Among PI, bortezomib (BTZ) is successfully used to treat newly diagnosed and relapsed MM patients, but its efficacy is reduced by the occurrence of drug resistance [7] The evidence for such is associated with impaired binding of the drug to proteasome subunits, the up-regulation of the proteasomal machinery and an increase in the ratio of BTZ-bound to BTZ-free proteasomal subunits, as a form of acquired proteasomal subunit immunity [8,9,10,11]

Methods

Results

Discussion

Conclusion