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Thesis

English

ID: <

10402/era.43894

>

Where these data come from
Delineating the biology of Anaplastic Lymphoma Kinase (ALK) and its resistance to crizotinib

Abstract

Degree: Doctor of Philosophy Abstract: Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase receptor that was initially identified as a potent oncogenic driver in anaplastic large-cell lymphoma (ALCL) in the form of NPM-ALK fusion protein. Various forms of oncogenic ALK proteins have been identified, subsequently, in various types of human cancers. The focus of this thesis was to get more insight into the ALK biology and how this could impact cancer phenotypes. 1) Crizotinib is an ALK inhibitor that has been found to be therapeutically useful against a subset of ALK-positive tumors. However, clinical resistance to this drug has been well known and the mechanism of this resistance is incompletely understood. In this part of my thesis, I hypothesized that the crizotinib—ALK binding is a determining factor for the sensitivity to crizotinib in ALK-positive cell lines derived from various types of cancers. Using the cellular thermal shift assay (CETSA), I measured the crizotinib—ALK binding in a panel of ALK-positive cell lines, and correlated the findings with the ALK structure and its interactions with specific binding proteins. I found that the biological response to crizotinib treatment significantly correlated with crizotinib—ALK binding. Additionally, I found that the crizotinib-resistant cell lines expressed higher protein levels of β-catenin. Furthermore, the siRNA knockdown of β-catenin in these cells restored crizotinib—ALK binding and was associated with a significant lowering of IC50. Taken together, this part of the thesis showed that the crizotinib—ALK binding measurable by CETSA is useful in predicting crizotinib sensitivity in ALK-positive cancer cells and crizotinib—ALK binding is dictated by the structure of ALK and some of its binding partners. 2) Since I have highlighted the importance of the physical binding between crizotinib and ALK in determining the crizotinib sensitivity, here I asked if these observations hold true for the stem-like cells in neuroblastoma (NB) cells, which were purified based on their responsiveness to a Sox2 reporter. NB is the most common pediatric extra-cranial solid tumor and the survival for metastatic NB remains 50% of the tumors. Importantly, patients with tumors that did not express ALK-I19 and lacked MYCN amplification had an excellent clinical outcome, with 19/19 patients survived at 5-years as compared to only 10/18 patients with tumors carrying ALK-I19 and/or MYCN amplification. This data suggests that the absence of ALK-I19 and MYCN amplification is a useful prognostic marker for NB patients. To conclude, this thesis has provided insights into the molecular mechanism underlying the biology of ALK and a novel mechanism of resistance, which may provide valuable therapeutic targets in neuroblastoma and other ALK-positive cancers.

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