Novel Peptide-drug Conjugate Melflufen Efficiently Eradicates Bortezomib-resistant Multiple Myeloma Cells Including Tumor-initiating Myeloma Progenitor Cells

Abstract

Introduction of the proteasome inhibitor bortezomib has dramatically improved clinical outcomes in multiple myeloma. However, most patients become refractory to bortezomib-based therapies. On the molecular level, development of resistance to bortezomib in myeloma cells is accompanied by complex metabolic changes resulting in increased protein folding capacity, and less dependency on the proteasome. In this study, we show that aminopeptidase B, encoded by the RNPEP gene, is upregulated in bortezomib-resistant myeloma cell lines, and in a murine in vivo model. Moreover, increased RNPEP expression is associated with shorter survival in multiple myeloma patients previously treated with bortezomib-containing regimens. Additionally, expression is increased in plasma cell precursors, a B-lymphoid compartment previously associated with myeloma stem cells. We hypothesized that increased aminopeptidase B expression in aggressive myeloma clones may be used therapeutically toward elimination of the cells via the use of a novel peptide-drug conjugate, melphalan flufenamide (melflufen). Melflufen, a substrate of aminopeptidase B, efficiently eliminates bortezomib-resistant myeloma cells in vitro and in vivo, and completely suppresses clonogenic myeloma growth in vitro at subphysiological concentrations. Thus, melflufen represents a novel treatment option that is able to eradicate drug-resistant myeloma clones characterized by elevated aminopeptidase B expression.

Pharmacokinetics and Metabolism of Melflufen, an Alkylating Peptide–Drug Conjugate, in Patients with Relapsed Refractory Multiple Myeloma

“…The disposition of melflufen in tumor cells in vitro is characterized by a rapid uptake followed by almost direct peptide hydrolysis to melphalan or via an intermediate step where desethyl‐melflufen is formed by esterases, then metabolized to melphalan by aminopeptidases 2 . The hydrolysis may be mediated by various aminopeptidases, such as aminopeptidase N 3 ; aminopeptidase B, an enzyme demonstrated to confer resistance to bortezomib 4 ; and/or a set of aminopeptidases shown to be overexpressed in multiple myeloma cells (leucine aminopeptidase 3, leukotriene‐A4 hydrolase, arginyl aminopeptidase) 5 . Various in vitro and in vivo studies using different cancer models have consistently shown significantly higher activity of melflufen versus melphalan at equimolar concentrations/doses 6 .…”