SUMMARY MYC contributes to the pathogenesis of a majority of human cancers, yet strategies to modulate the function of the c-Myc oncoprotein do not exist. Toward this objective, we have targeted MYC transcription by interfering with chromatin-dependent signal transduction to RNA polymerase, specifically by inhibiting the acetyl-lysine recognition domains (bromodomains) of putative co-activator proteins implicated in transcriptional initiation and elongation. Using a selective small-molecule bromodomain inhibitor, JQ1, we identify BET bromodomain proteins as regulatory factors for c-Myc. BET inhibition by JQ1 downregulates MYC transcription, followed by genome-wide downregulation of Myc-dependent target genes. In experimental models of multiple myeloma, a Myc-dependent hematologic malignancy, JQ1 produces a potent antiproliferative effect associated with cell cycle arrest and cellular senescence. Efficacy of JQ1 in three murine models of multiple myeloma establishes the therapeutic rationale for BET bromodomain inhibition in this disease and other malignancies characterized by pathologic activation of c-Myc.

The NEDD8 activating enzyme (NAE) is upstream of the 20S proteasome in the ubiquitin/proteasome pathway and catalyzes the first step in the neddylation pathway. NEDD8 modification of cullins is required for ubiquitination of cullin-ring ligases (CRLs), which regulate degradation of a distinct subset of proteins. The more targeted impact of NAE on protein degradation prompted us to study MLN4924, an investigational NAE inhibitor, in preclinical multiple myeloma (MM) models. In vitro treatment with MLN4924 led to dose-dependent decrease of viability (EC50=25–150nM) in a panel of human MM cell lines. MLN4924 was similarly active against a bortezomib-resistant ANBL-6 subline and its bortezomib-sensitive parental cells. MLN4924 had sub-μM activity (EC50 values <500nM) against primary CD138+ MM patient cells and exhibited at least additive effect when combined with dexamethasone, doxorubicin and bortezomib against MM.1S cells. The bortezomib-induced compensatory up-regulation of transcripts for ubiquitin/proteasome was not observed with MLN4924 treatment, suggesting distinct functional roles of NAE vs 20S proteasome. MLN4924 was well tolerated at doses up to 60mg/kg 2x daily and significantly reduced tumor burden in both a subcutaneous and an orthotopic mouse model of MM. These studies provide the framework for the clinical investigation of MLN4924 in MM.

Cell cycle regulators, such as cyclin-dependent kinases (CDKs), are appealing targets for multiple myeloma (MM) therapy given the increased proliferative rates of tumor cells in advanced vs. early stages of MM. We hypothesized that a multi-targeted CDK inhibitor with a different spectrum of activity compared to existing CDK inhibitors could trigger distinct molecular sequelae with therapeutic implications for MM. We therefore studied the small molecule heterocyclic compound NVP-LCQ195/AT9311 (LCQ195), which inhibits CDK1, CDK2 and CDK5, as well as CDK3 and CDK9. LCQ195 induced cell cycle arrest and eventual apoptotic cell death of MM cells, even at sub-μM concentrations; spared non-malignant cells; and overcome the protection conferred to MM cells by stroma or cytokines of the bone marrow milieu. In MM cells, LCQ195 triggered decreased amplitude of transcriptional signatures associated with oncogenesis, drug resistance, and stem cell renewal, including signatures of activation of key transcription factors for MM cells e.g. myc, HIF-1α, IRF4. Bortezomib-treated MM patients whose tumors had high baseline expression of genes suppressed by LCQ195 had significantly shorter progression-free and overall survival than those with low levels of these transcripts in their MM cells. These observations provide insight into the biological relevance of multi-targeted CDK inhibition in MM.