Activating the Intrinsic Pathway of Apoptosis Using BIM BH3 Peptides Delivered by Peptide Amphiphiles with Endosomal Release.

Mathew R Schnorenberg,James L Labelle,Matthew V Tirrell,Joseph A Bellairs,Handan Acar,Ravand Samaeekia

doi:10.3390/ma12162567

Abstract

Therapeutic manipulation of the BCL-2 family using BH3 mimetics is an emerging paradigm in cancer treatment and immune modulation. For example, peptides mimicking the BIM BH3 helix can directly target the full complement of anti- and pro-apoptotic BCL-2 proteins to trigger apoptosis. This study has incorporated the potent BH3 α-helical death domain of BIM into peptide amphiphile (PA) nanostructures designed to facilitate cellular uptake and induce cell death. This study shows that these PA nanostructures are quickly incorporated into cells, are able to specifically bind BCL-2 proteins, are stable at physiologic temperatures and pH, and induce dose-dependent apoptosis in cells. The incorporation of a cathepsin B cleavable linker between the BIM BH3 peptide and the hydrophobic tail resulted in increased intracellular accumulation and mitochondrial co-localization of the BIM BH3 peptide while also improving BCL-2 family member binding and apoptotic reactivation. This PA platform represents a promising new strategy for intracellular therapeutic peptide delivery for the disruption of intracellular protein:protein interactions.

Highlights

The BCL-2 family of proteins forms a complex protein-protein interaction (PPI) network that regulates cellular life and death decisions, which contributes to organismal development, cancer ontogeny and chemoresistance, hematopoiesis, and immune regulation [1,2,3,4]
BH3 death domain of BIM (BIMA BH3; Figure 1). This peptide sequence is sparingly soluble in water, so five C-terminal lysines were added to increase the charge and solubility of the peptide (BIMA,K )
A cathepsin-cleavable linker was incorporated between the therapeutic peptide and the C-terminal lysines to form a cathepsin-cleavable peptide amphiphile (PA), BIMA,cath,K PA2, which we hypothesized would allow for release of the peptide cargo following PA uptake into the cell

Summary

Introduction

The BCL-2 family of proteins forms a complex protein-protein interaction (PPI) network that regulates cellular life and death decisions, which contributes to organismal development, cancer ontogeny and chemoresistance, hematopoiesis, and immune regulation [1,2,3,4]. Members of the BCL-2 family known as BH3-only proteins (e.g., BIM, BID, PUMA, NOXA) serve as cellular stress sentinels and can trigger irreversible activation of apoptosis through their α-helical BH3 death domains. These pro-apoptotic signals are normally held in check by the multidomain anti-apoptotic family members (e.g., BCL-XL, BCL-2, BCL-W, MCL-1) through sequestering PPIs. when pro-apoptotic signals outweigh anti-apoptotic signals, the multidomain pro-apoptotic effectors, BAX and BAK, are activated and trigger the cell death cascade by oligomerizing in the mitochondrial membrane, leading to mitochondrial outer membrane permeabilization (MOMP), cytochrome c release, apoptosome formation, and effector caspase activation [1]. Cancer cells often push the BCL-2 family’s PPI balance toward an anti-apoptotic state to avoid cell death despite cellular stress and damage, for example by upregulating anti-apoptotic members or downregulating

Methods

Results

Conclusion