Identification of the Microtubule Inhibitor-Activated Bcl-xL Kinase: A Regulator of Breast Cancer Cell Chemosensitivity to Taxol

This training grant set out to define a molecular pathway involved in breast tumor resistance to the key chemotherapeutic class of drugs called microtubule inhibitors MTIs. It is also designed to train the Principal Investigator PI as a future breast cancer physician-scientist. MTIs are the most actively used agents for metastatic and adjuvant BC therapy, yet their use is limited by resistance. They activate a kinase that phosphorylates and inactivates Bcl-xL, an anti-apoptotic protein that causes resistance to chemotherapeutic agents. The goal is to identify the Bcl-xL kinase first by developing an in vitro assay for it. Data presented here shows that a 12 amino acid peptide, termed FL62, that harbors the Bcl-xL phosphorylation site Ser62, is an excellent substrate for the MTI activated kinase. Activation of this kinase correlates with Bcl-xL phosphorylation as well as mitotic arrest and cyclin B expression, implicating cyclin-dependent kinase-1 Cdk1. When tested, purified Cdk1 and cellular Cdk1cyclin B immunopurified from mitotic and vinblastine treated cells robustly phosphorylated FL62 confirming that Cdk1cyclin B is the FL62 kinase. This provides the foundation to validate Cdk1cyclin B as the MTI activate Bcl-xL kinase in vivo.