Profiling Global Kinome Signatures of the Radioresistant MCF-7/C6 Breast Cancer Cells Using MRM-based Targeted Proteomics

Ionizing
radiation is widely used in cancer therapy; however, cancer
cells often develop radioresistance, which compromises the efficacy
of cancer radiation therapy. Quantitative assessment of the alteration
of the entire kinome in radioresistant cancer cells relative to their
radiosensitive counterparts may provide important knowledge to define
the mechanism(s) underlying tumor adaptive radioresistance and uncover
novel target(s) for effective prevention and treatment of tumor radioresistance.
By employing a scheduled multiple-reaction monitoring analysis in
conjunction with isotope-coded ATP affinity probes, we assessed the
global kinome of radioresistant MCF-7/C6 cells and their parental
MCF-7 human breast cancer cells. We rigorously quantified 120 kinases,
of which 1/3 exhibited significant differences
in expression levels or ATP binding affinities. Several kinases involved
in cell cycle progression and DNA damage response were found to be
overexpressed or hyperactivated, including checkpoint kinase 1 (CHK1),
cyclin-dependent kinases 1 and 2 (CDK1 and CDK2), and the catalytic
subunit of DNA-dependent protein kinase. The elevated expression of
CHK1, CDK1, and CDK2 in MCF-7/C6 cells was further validated by Western
blot analysis. Thus, the altered kinome profile of radioresistant
MCF-7/C6 cells suggests the involvement of kinases on cell cycle progression
and DNA repair in tumor adaptive radioresistance. The unique kinome
profiling results also afforded potential effective targets for resensitizing
radioresistant cancer cells and counteracting deleterious effects
of ionizing radiation exposure.