Vladlena Kharchenko
Albert Einstein College of Medicine
Allosteric Regulation of the Pro-apoptotic Effector BAK
Apoptosis is a programmed cell death pathway whose dysregulation is a hallmark of cancer and other diseases. Mitochondrial apoptosis is controlled by the BCL-2 family of proteins, including the pro-apoptotic effectors BAK and BAX, whose activation triggers mitochondrial outer membrane permeabilization (MOMP), cytochrome c release, and caspase activation. Because BCL-2 family proteins share highly similar three-dimensional structures and canonical BH3-binding grooves, achieving selective pharmacological modulation of these proteins remains challenging. Using a combination of computational approaches and fragment-based screening, we identified a previously unrecognized cryptic pocket that enables selective allosteric regulation of pro-apoptotic BAK. Structure-guided mutational analysis of residues within this pocket revealed variants that modulate mitochondrial apoptotic priming in cancer cell lines, as measured by BH3 profiling. These mutations also alter sensitivity to BH3-mimetic drugs and influence caspase activation, cell viability, and proliferation in colorectal cancer cells. Mechanistic studies of mutation-dependent BAK activation provide new insight into the regulation of mitochondrial apoptosis and reveal an exploitable site for selective modulation of BAK function. Targeting this allosteric pocket represents a promising strategy for direct pharmacological activation of BAK and may enable new therapeutic approaches for malignancies characterized by elevated BAK expression, including colorectal and gastric cancers.
Apoptosis is a programmed cell death pathway whose dysregulation is a hallmark of cancer and other diseases. Mitochondrial apoptosis is controlled by the BCL-2 family of proteins, including the pro-apoptotic effectors BAK and BAX, whose activation triggers mitochondrial outer membrane permeabilization (MOMP), cytochrome c release, and caspase activation. Because BCL-2 family proteins share highly similar three-dimensional structures and canonical BH3-binding grooves, achieving selective pharmacological modulation of these proteins remains challenging. Using a combination of computational approaches and fragment-based screening, we identified a previously unrecognized cryptic pocket that enables selective allosteric regulation of pro-apoptotic BAK. Structure-guided mutational analysis of residues within this pocket revealed variants that modulate mitochondrial apoptotic priming in cancer cell lines, as measured by BH3 profiling. These mutations also alter sensitivity to BH3-mimetic drugs and influence caspase activation, cell viability, and proliferation in colorectal cancer cells. Mechanistic studies of mutation-dependent BAK activation provide new insight into the regulation of mitochondrial apoptosis and reveal an exploitable site for selective modulation of BAK function. Targeting this allosteric pocket represents a promising strategy for direct pharmacological activation of BAK and may enable new therapeutic approaches for malignancies characterized by elevated BAK expression, including colorectal and gastric cancers.
