Publication Date

2018-07-17

Availability

Open access

Embargo Period

2018-07-17

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Chemistry (Arts and Sciences)

Date of Defense

2018-06-08

First Committee Member

James Wilson

Second Committee Member

Francisco Raymo

Third Committee Member

Rajeev Prabhakar

Fourth Committee Member

James Baker

Abstract

The EGFR/ERBB family of tyrosine kinase receptors (ERBB1-4) are involved in regulation of intracellular communications such as cell proliferation, metabolism, and numerous biological responses. The dysregulation of their signaling pathways contributes to development of cancer malignancies. This receptor family is a conventional chemotherapeutic target and serves as diagnostic modality. ERBB2, also known as HER2, is a prognostic biomarker implicated in about 20 % of invasive breast cancer incidence. Hence, the status of ERBB2 is a crucial factor in deciding treatment type for breast cancer patients. Common immunohistochemistry assays analyzing the ERBB2 receptor tyrosine kinase rely on use of antibodies. These assays often require multi-step specimen preparation such as cell fixation or cell lysis, resulting in observation of a biologically stopped system. In this context, we aimed to develop small molecular fluorescent probes for ERBB2 to explore the dynamic information related to the signaling activation at the live single cell level. Hence, we generated turn-on fluorescent kinase inhibitors targeting the ATP binding pocket of the ERBB2 by structurally modifying clinically relevant EGFR/ERBB inhibitors. The fluorescent kinase inhibitors are capable of acquiring dynamic information in live cell with minimized specimen preparation comparing to immunohistochemistry assays. The turn-on fluorescent kinase inhibitors that we have developed are capable of identifying the ERBB2 (+) breast cancer cells from ERBB2 (-) cells as well as tracking rapid responses to perturbation factors at the level of individual breast cancer cells including ERBB2 activation and internalization. Our live cell compatible kinase probes serve as a novel analytic tool providing complimentary information to immunohistochemical analysis in ERBB2 (+) breast cancer biology.

Keywords

Fluorescent inhibitor; Receptor tyrosine kinase activation; Dynamic imaging; ERBB2

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