Breast cancer is one of the most diagnosed cancers and the leading cause of cancer death in women worldwide. Breast cancer cells need activated receptor tyrosine kinases (RTKs) to invade, proliferate, and metastasize. Increased activity and overexpression of RTKs is associated with poor prognosis for breast cancer patients. Therefore, multiple RTKs have emerged as attractive therapeutic targets. However, resistance to therapies is a persistent problem for the development of therapeutics targeting RTKs. New and innovative approaches to effectively target these receptors are required. Recently S-palmitoylation has been identified as an important post-translational modification that regulates signal transduction, protein trafficking and degradation of specific RTKs in breast cancer. The substrate scope, role, extent of dysregulation and the enzymes responsible for the S-palmitoylation of specific substrates in breast cancer is largely unknown. This is partially caused by a lack of chemical biological methods to study these processes. Therefore we have worked towards the development of two novel methodologies to study protein S-palmitoylation; 1) Development of a methodology to study the extent of palmitoylation on substrate proteins. 2) Development of a methodology for the identification of direct substrates for the enzymes that attach these lipids to proteins (palmitoyl transferases).
A methodology to determine the site specific palmitoylation stoichiometry will be a important tool for the analysis of drugs that prevent palmitoylation or depalmitoylation. To this aim, we have synthesized a cysteine reactive tool that will facilitate the study of site specific palmitoylation stoichiometry by mass-spectrometry. The research fellow is currently still working on the further development of this methodology at Imperial College London.
To elucidate the substrate scope of individual palmitoyl transferases (zDHHCs), we have succesfully developed a chemical genetic methodology that enables zDHHC specific lipid transfer. Combination of this methodology with mass spectrometry-based proteomics provides a platform to aid identification of palmitoyl transferase substrates and may assist zDHHC characterisation breast cancer.
