Disruption of Bcr-Abl Coiled Coil Oligomerization by Design

Abstract

Oligomerization is an important regulatory mechanism for many proteins, including oncoproteins and other pathogenic proteins. The oncoprotein Bcr-Abl relies on oligomerization via its coiled coil domain for its kinase activity, suggesting that a designed coiled coil domain with enhanced binding to Bcr-Abl and reduced self-oligomerization would be therapeutically useful. Key mutations in the coiled coil domain of Bcr-Abl were identified that reduce homo-oligomerization through intermolecular charge-charge repulsion yet increase interaction with the Bcr-Abl coiled coil through additional salt bridges, resulting in an enhanced ability to disrupt the oligomeric state of Bcr-Abl. The mutations were modeled computationally to optimize the design. Assays performed in vitro confirmed the validity and functionality of the optimal mutations, which were found to exhibit reduced homo-oligomerization and increased binding to the Bcr-Abl coiled coil domain. Introduction of the mutant coiled coil into K562 cells resulted in decreased phosphorylation of Bcr-Abl, reduced cell proliferation, and increased caspase-3/7 activity and DNA segmentation. Importantly, the mutant coiled coil domain was more efficacious than the wild type in all experiments performed. The improved inhibition of Bcr-Abl through oligomeric disruption resulting from this modified coiled coil domain represents a viable alternative to small molecule inhibitors for therapeutic intervention.

Improved Coiled-Coil Design Enhances Interaction with Bcr-Abl and Induces Apoptosis

“…3, 4 We have previously reported a modified version of this coiled-coil domain, CCmut2, which exhibits disruption of Bcr-Abl oligomeric complexes and results in decreased proliferation of CML cells and induction of apoptosis. 5 A major contributing factor to these enhanced capabilities is the destabilization of the CCmut2 homo-dimers, increasing the availability to interact with and inhibit Bcr-Abl. Here, we included an additional mutation (K39E) that could in turn further destabilize the mutant homo-dimer.…”

Microtubule association of EML proteins and the EML4-ALK variant 3 oncoprotein require an N-terminal trimerization domain

“…In oncogenic ALK-fusions oligomerization conferred by the fusion partner mimics the native oligomerization-dependent activation mechanism of receptor tyrosine kinases and engenders constitutive activation of ALK [25]. The use of rationally-designed peptide and peptidomimetic inhibitors is under investigation for the disruption of several coiled-coil interactions that are implicated in disease and clinically useful therapeutics have been developed using this approach [27,28,29]. The TD structures may enable the development of protein-protein interaction inhibitors targeting the trimerization interface of the EML4 TD, another possible avenue towards therapeutic intervention in EML4-ALK NSCLC and one that would be applicable to all variants.…”

HMGB2 regulates satellite-cell-mediated skeletal muscle regeneration through IGF2BP2

“…Previous studies have demonstrated that all of them are validated for use (Aguilera et al, 2011;Averous et al, 2012;Della Pietra et al, 2015;Dixon et al, 2011;Gong et al, 2015;Kim et al, 2011;Li et al, 2012;Murphy et al, 2011;Nissar et al, 2012;Rao et al, 2014;Sarkar and Zohn, 2012;Takeda et al, 2014;Tan et al, 2011;Taniguchi et al, 2011;Wu et al, 2010;Zhang et al, 2011).…”