Inhibition of Bcr-Abl in Human Leukemic Cells with a Coiled-Coil Protein Delivered by a Leukemia-Specific Cell-Penetrating Peptide

摘要

The oncoprotein Bcr-Abl is the cause of chronic myeloid leukemia (CML).(1) Current therapies target the tyrosine kinase domain of Bcr-Abl, but resistance to these drugs is common.(2) Bcr-Abl homooligomerization via its N-terminal coiled-coil (CC) domain is required for tyrosine kinase activity.(3) Our previous work has shown that it is possible to inhibit Bcr-Abl activity by targeting the CC domain with a peptidomemetic known as CCmut3, delivered as a plasmid.(4) In this study, CCmut3 is delivered to cells as a protein by utilizing a leukemia-specific cell-penetrating peptide (CPP).(5) Here, recombinant CPP-CCmut3 was expressed, purified, and tested for its antioncogenic activity. CPP-CCmut3 was able to enter two leukemic cell lines (K562 and Ba/F3-P210) and inhibit Bcr-Abl activity as shown by induction of necrosis/apoptosis via 7-AAD/Annexin V staining, reduction of oncogenic potential in colony forming assays, reduction of cell proliferation, and inhibition of Bcr-Abl phosphorylation (kinase activity). Further, CPP-CCmut3 did not enter nonleukemic cell lines (HEK293 and MCF-7). While CPP-CCmut3 was able to enter the parental, nonleukemic Bcr-Abl(-) Ba/F3 pro-B cell line, it revealed no signs of activity in the assays performed, as expected. These results indicate the feasibility of using CPP-CCmut3 as a therapeutic against CML.