This study describes the development of hydrocarbon-stapled BH3 peptides (SAHBs) that enhance the helical structure and biological activity of BCL-2 family proteins, which are critical regulators of apoptosis. The BH3 domain of BID, a pro-apoptotic protein, was modified using hydrocarbon stapling to create SAHBs that are more stable, cell-permeable, and capable of binding to BCL-2 family proteins with higher affinity. These SAHBs effectively activated the apoptotic pathway in leukemia cells and inhibited the growth of human leukemia xenografts in vivo. The hydrocarbon staple, created through ruthenium-catalyzed olefin metathesis, stabilizes the α-helix of the BH3 domain, increasing its helical content and protecting it from proteolytic degradation. SAHBs demonstrated enhanced serum stability and cellular uptake, as shown by confocal microscopy and fluorescence-activated cell sorting (FACS) analyses. They localized to mitochondria and triggered cytochrome c release, a key step in apoptosis. SAHBs were tested in Jurkat T cell leukemia cells and showed significant apoptosis induction, with 50% of cells treated with 5 μM SAHB displaying annexin V staining after 20 hours. However, BCL-2 overexpression reduced this effect, indicating that SAHBs act at the BCL-2 control point. In vivo, SAHBs suppressed leukemia growth in immunodeficient mice, extending survival and reducing tumor burden. They were effective against multiple leukemia cell lines, including T cell (Jurkat), B cell (REH), and mixed lineage (MLL) cells. The study highlights the potential of hydrocarbon-stapled peptides as a strategy for modulating protein-protein interactions in signaling pathways, offering a new approach to target intracellular interactions that are challenging for small-molecule drugs. SAHBs represent a promising therapeutic tool for cancer treatment and other diseases involving apoptosis regulation.This study describes the development of hydrocarbon-stapled BH3 peptides (SAHBs) that enhance the helical structure and biological activity of BCL-2 family proteins, which are critical regulators of apoptosis. The BH3 domain of BID, a pro-apoptotic protein, was modified using hydrocarbon stapling to create SAHBs that are more stable, cell-permeable, and capable of binding to BCL-2 family proteins with higher affinity. These SAHBs effectively activated the apoptotic pathway in leukemia cells and inhibited the growth of human leukemia xenografts in vivo. The hydrocarbon staple, created through ruthenium-catalyzed olefin metathesis, stabilizes the α-helix of the BH3 domain, increasing its helical content and protecting it from proteolytic degradation. SAHBs demonstrated enhanced serum stability and cellular uptake, as shown by confocal microscopy and fluorescence-activated cell sorting (FACS) analyses. They localized to mitochondria and triggered cytochrome c release, a key step in apoptosis. SAHBs were tested in Jurkat T cell leukemia cells and showed significant apoptosis induction, with 50% of cells treated with 5 μM SAHB displaying annexin V staining after 20 hours. However, BCL-2 overexpression reduced this effect, indicating that SAHBs act at the BCL-2 control point. In vivo, SAHBs suppressed leukemia growth in immunodeficient mice, extending survival and reducing tumor burden. They were effective against multiple leukemia cell lines, including T cell (Jurkat), B cell (REH), and mixed lineage (MLL) cells. The study highlights the potential of hydrocarbon-stapled peptides as a strategy for modulating protein-protein interactions in signaling pathways, offering a new approach to target intracellular interactions that are challenging for small-molecule drugs. SAHBs represent a promising therapeutic tool for cancer treatment and other diseases involving apoptosis regulation.