An Investigational Proteasome Inhibitor MLN9708 (MLN2238) Induces Apoptosis In Human Multiple Myeloma Cells In Vitro

Abstract 3066

Targeting the proteasome has proven to be one of the most effective therapeutic strategies in the treatment of multiple myeloma (MM), and the proteasome inhibitor bortezomib is approved for treatment of MM. However its clinical efficacy is compromised by the acquired resistance in patients, necessitating the development of new therapeutics. Several new proteasome inhibitors are under investigation for their therapeutic efficacy in MM. MLN9708 (Millennium Pharmaceuticals, Inc., Cambridge, MA) is a proteasome inhibitor which shows refined pharmacokinetic and pharmacodynamic properties in preclinical studies and is currently in Phase I clinical development. Upon exposure to aqueous solutions or plasma, MLN9708 rapidly hydrolyzes to MLN2238, the biologically active form. MLN2238 was used for all of the studies reported here, in which we report the efficacy of MLN2238 on three established MM cell lines-KMS11, OPM2 and U266. MLN2238 was found to inhibit the chymotrypsin-like proteasomal activity of all MM cell lines in a dose dependent manner. Investigation of the IC₅₀ of MLN2238 on these cell lines demonstrated that KMS11 is the most sensitive (IC₅₀ of 15.9 nM) while U266 was found to be the least sensitive cell line (IC₅₀ of 511 nM). OPM2 cells also showed intermediate sensitivity with an IC₅₀ of 58.6 nM. MLN2238 induced apoptosis in KMS11 cells as evidenced by annexin V staining and PARP-1 cleavage. Cleavage of caspases 9 and 3 suggested activation of the intrinsic apoptotic pathway by MLN2238. Furthermore, MLN2238 treatment was shown to increase the mitochondrial outer membrane permeability (MOMP) and decrease BCL-2 levels. Evaluation of the expression of PSMB5, the preferred proteasomal subunit target for both bortezomib and MLN2238, revealed that it is expressed at approximately 3 fold more in KMS11 cells as compared to U266, suggesting a possible mechanism for higher sensitivity of KMS11 to the proteasomal inhibitor, MLN2238. This preclinical evaluation confirms the anti-myeloma effects of MLN2238, warranting further in-depth evaluation in both in vitro and in vivo models of MM.