Mcl-1 dependence predicts response to vorinostat and gemtuzumab ozogamicin in acute myeloid leukemia.

Older adults with acute myeloid leukemia (AML) are commonly considered for investigational therapies, which often only benefit subsets of patients. In this study, we assessed whether BH3 profiling of apoptotic functionality could predict outcomes following treatment with vorinostat (histone deacetylase inhibitor) and gemtuzumab ozogamicin (GO; CD33-targeted immunoconjugate). Flow cytometry of BH3 peptide priming with Noxa (anti-apoptotic protein Mcl-1 modulator) correlated with remission induction (p=.026; AUC=0.83 [CI: 0.65-1.00; p=.00042]: AUC=0.88 [CI:0.75-1.00] with age adjustment) and overall survival (p=.027 logistic regression; AUC=0.87 [0.64-1.00; p=.0017]). This Mcl-1-dependence suggests a pivotal role of Bcl-2 family protein-mediated apoptosis to vorinostat/GO in AML patients.

Mcl-1 mediates TWEAK/Fn14-induced non-small cell lung cancer survival and therapeutic response.

UNLABELLED: Insensitivity to standard clinical interventions, including chemotherapy, radiotherapy, and tyrosine kinase inhibitor (TKI) treatment, remains a substantial hindrance towards improving the prognosis of patients with non-small cell lung cancer (NSCLC). The molecular mechanism of therapeutic resistance remains poorly understood. The TNF-like weak inducer of apoptosis (TWEAK)-FGF-inducible 14 (TNFRSF12A/Fn14) signaling axis is known to promote cancer cell survival via NF-κB activation and the upregulation of prosurvival Bcl-2 family members. Here, a role was determined for TWEAK-Fn14 prosurvival signaling in NSCLC through the upregulation of myeloid cell leukemia sequence 1 (MCL1/Mcl-1). Mcl-1 expression significantly correlated with Fn14 expression, advanced NSCLC tumor stage, and poor patient prognosis in human primary NSCLC tumors. TWEAK stimulation of NSCLC cells induced NF-κB-dependent Mcl-1 protein expression and conferred Mcl-1-dependent chemo- and radioresistance. Depletion of Mcl-1 via siRNA or pharmacologic inhibition of Mcl-1, using EU-5148, sensitized TWEAK-treated NSCLC cells to cisplatin- or radiation-mediated inhibition of cell survival. Moreover, EU-5148 inhibited cell survival across a panel of NSCLC cell lines. In contrast, inhibition of Bcl-2/Bcl-xL function had minimal effect on suppressing TWEAK-induced cell survival. Collectively, these results position TWEAK-Fn14 signaling through Mcl-1 as a significant mechanism for NSCLC tumor cell survival and open new therapeutic avenues to abrogate the high mortality rate seen in NSCLC. IMPLICATIONS: The TWEAK-Fn14 signaling axis enhances lung cancer cell survival and therapeutic resistance through Mcl-1, positioning both TWEAK-Fn14 and Mcl-1 as therapeutic opportunities in lung cancer.

BH3 profiling discriminates response to cytarabine-based treatment of acute myelogenous leukemia.

As acute myelogenous leukemia (AML) patient response to cytarabine-based standard-of-care treatment is variable, stratification into subgroups by biomarker-predicted response may lead to improved clinical outcomes. Here, we assess cell mitochondrial depolarization to proapoptotic signaling BH3-only peptides as a surrogate for the function of Bcl-2 family proteins to address clinical response to cytarabine-based therapy in patients with AML (N = 62). Peripheral blood mononuclear cell (PBMC) or bone marrow aspirate specimens were obtained from newly diagnosed patients with AML, viably preserved, and assayed by flow cytometry following BH3 profile assay with individual BH3 peptides. Mann-Whitney analysis indicates biomarker correlation with response to induction therapy: Notably, BIM priming was highly significant (P = 2 × 10(-6)) with a compelling sensitivity/specificity profile [area under curve (AUC) = 0.83; 95% confidence interval (CI), 0.73-0.94; P = 2 × 10(-10)]. Multivariate analysis indicates improved profiles for BIM readout + patient age (AUC = 0.89; 95% CI, 0.81-0.97) and BIM + patient age + cytogenetic status (AUC = 0.91; 95% CI, 0.83-0.98). When patients were stratified by cytogenetic status, BIM readout was significant for both intermediate (P = 0.0017; AUC = 0.88; 95% CI, 0.71-1.04) and unfavorable (P = 0.023; AUC = 0.79; 95% CI, 0.58-1.00) risk groups, demonstrating predictive power independent of cytogenetics. Additional analyses of secondary clinical endpoints displayed correlation between overall survival (P = 0.037) and event-free survival (P = 0.044) when patients were stratified into tertiles by BIM peptide response. Taken together, these results highlight the potential utility of BH3 profiling in personalized diagnostics of AML by offering actionable information for patient management decisions.

Hydroxyquinoline-derived compounds and analoguing of selective Mcl-1 inhibitors using a functional biomarker.

Anti-apoptotic Bcl-2 family proteins are important oncology therapeutic targets. To date, BH3 mimetics that abrogate anti-apoptotic activity have largely been directed at Bcl-2 and/or Bcl-xL. One observed mechanism of resistance to these inhibitors is increased Mcl-1 levels in cells exposed to such therapeutics. For this reason, and because Mcl-1 is important in the onset of lymphoid, myeloid, and other cancers, it has become a target of great interest. However, small molecule inhibitors displaying potency and selectivity for Mcl-1 are lacking. Identifying such compounds has been challenging due to difficulties in translating the target selectivity observed at the biochemical level to the cellular level. Herein we report the results of an HTS strategy coupled with directed hit optimization. Compounds identified have selective Mcl-1 inhibitory activity with greater than 100-fold reduced affinity for Bcl-xL. The selectivity of these compounds at the cellular level was validated using BH3 profiling, a novel personalized diagnostic approach. This assay provides an important functional biomarker that allows for the characterization of cells based upon their dependencies on various anti-apoptotic Bcl-2 proteins. We demonstrate that cells dependent on Mcl-1 or Bcl-2/Bcl-xL for survival are commensurately responsive to compounds that genuinely target those proteins. The identification of compound 9 with uniquely validated and selective Mcl-1 inhibitory activity provides a valuable tool to those studying the intrinsic apoptosis pathway and highlights an important approach in the development of a first-in-class cancer therapeutic.