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Background: Inhibitors of B-cell CLL/lymphoma 2 (Bcl-2) family proteins have shown hope as antitumor drugs. While the notion that it is efficient to coordinate, balance, and neutralize both arms of the anti-apoptotic Bcl-2 family has been validated in many cancer cells, the weights of the two arms contributing to apoptosis inhibition have not been explored. This study analyzed the best combination ratio for different Bcl-2 selective inhibitors. Methods: We used a previously established mathematical model to study the weights of Bcl-2 (representing both Bcl-2 and Bcl-xL in this study) and myeloid cell leukemia-1 (Mcl-1). Correlation and single-parameter sensitivity analysis were used to find the major molecular determinants for Bcl-2 and Mcl-1 dependency, as well as their weights. Biological experiments were used to verify the mathematical model. Results: Bcl-2 protein level and Mcl-1 protein level, production, and degradation rates were the major molecular determinants for Bcl-2 and Mcl-1 dependency. The model gained agreement with the experimental assays for ABT-737/A-1210477 and ABT-737/compound 5 combination effect in MCF-7 and MDA-MB-231. Two sets of equations composed of Bcl-2 and Mcl-1 levels were obtained to predict the best combination ratio for Bcl-2 inhibitors with Mcl-1 inhibitors that stabilize and downregulate Mcl-1, respectively. Conclusions: The two sets of equations can be used as tools to bypass time-consuming and laborious experimental screening to predict the best drug combination ratio for treatment. [ABSTRACT FROM AUTHOR]

The chaperone heat shock protein 70 (Hsp70) is crucial for avoiding protein misfolding under stress, but is also up-regulated in many kinds of cancers, where its ability to buffer cellular stress prevents apoptosis. Previous research has suggested Hsp70 interacts with pro-apoptotic Bcl-2 family proteins, including Bim and Bax. However, a definitive demonstration of this interaction awaits, and insights into the structural basis and molecular mechanism remain unclear. Earlier studies have identified a Bcl-2 homology 3 (BH3) domain present in Bcl-2 family members that engages receptors to stimulate apoptosis. We now show that Hsp70 physically interacts with pro-apoptotic multidomain and BH3-only proteins via a BH3 domain, thereby serving as a novel BH3 receptor, using in vitro fluorescent polarization (FP), isothermal titration calorimetry (ITC), and cell-based co-immunoprecipitation (co-IP) experiments, µH-15N-transverse relaxation optimized spectroscopy (TROSY-HSQC), trypsin proteolysis, ATPase activity, and denatured rhodanese aggregation measurements further demonstrated that BimBH3 binds to a novel allosteric site in the nucleotide-binding domain (NBD) of Hsp70, by which Bim acts as a positive co-chaperone to promote the ATPase activity and chaperone functions. A dual role of Hsp70's anti-apoptotic function was revealed that when it keeps Bim in check to inhibit apoptosis, it simultaneously stabilizes oncogenic clients including AKT and Raf-1 with the aid of Bim. Two faces of Bim in cell fate regulation were revealed that in opposite to its well-established pro-apoptotic activator role, Bim could help the folding of oncogenic proteins. [ABSTRACT FROM AUTHOR]

No α-helical mimetic that exhibits Bcl-2/MDM2 dual inhibition has been rationally designed due to the different helicities of the α-helixes at their binding interfaces. Herein, we extracted a one-turn α-helix-mimicking ortho-triarene unit from o-phenylene foldamers. Linking benzamide substrates with a rotatable C-N bond, we constructed a novel semirigid pyramid-like scaffold that could support its two-turn α-helix mimicry without aromatic stacking interactions and could adopt the different dihedral angles of the key residues of p53 and BH3-only peptides. On the basis of this universal scaffold, a series of substituent groups were installed to capture the key residues of both p53TAD and BimBH3 and balance the differences of the bulks between them. Identified by FP, ITC, and NMR spectroscopy, a compound 6e (zq-1) that directly binds to Mcl-1, Bcl-2, and MDM2 with balanced submicromolar affinities was obtained. Cell-based experiments demonstrated its antitumor ability through Bcl-2/MDM2 dual inhibition simultaneously. [ABSTRACT FROM AUTHOR]