Publication Date

2014-08-06

Availability

Embargoed

Embargo Period

2016-08-05

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Chemistry (Arts and Sciences)

Date of Defense

2014-07-07

First Committee Member

James N. Wilson

Second Committee Member

Angel E. Kaifer

Third Committee Member

Rajeev Prabhakar

Fourth Committee Member

Ralf Landgraf

Abstract

Fluorescent probes are indispensable tools for biologists owing to the safe and non-invasive method of detection they offer with minimum perturbation of the cell under investigation. Despite all the advantages they offer, there are certain limitations for the use of fluorescent probes for live cell imaging such as phototoxicity to the cell, photobleaching or photoinstability, etc. Additionally, they require labeling with specific recognition units such as biomolecules like DNA, RNA or an antibody to render them specific to the target cell. In this context, “turn-on” fluorescent probes are advantageous, as they remain silent or inactive and become emissive only under special circumstances or in the event of a biochemical process/reaction. This can overcome the challenges arising from the use of fluorescent probes and more importantly, these probes do not require the additional rinsing steps which prevent the access to the target cell dynamics. Dysregulation of ERBB is encountered in several malignancies including head, neck, breast and ovarian cancers, thereby making it an extensive area of research. In this context, development of novel strategies to study the receptor activation will be very beneficial for designing effective small molecule inhibitors. In this project, we have designed and developed novel fluorescent “turn-on” probes targeting the ATP binding pocket of ERBB receptor which can provide deeper insights into the receptor activation, receptor conformation at the time of ligand binding, subsequent signaling pathways activated upon ligand binding, etc. DMAQ (6-(4-dimethyl-aminostyryl)-N-benzylquinazolin-4-amine) was the first “turn-on” fluorescent probe designed in this direction which was followed by a series of 15 kinase inhibitors with binding affinities comparable to gefitinib, a well-known anti-cancer drug. Further, we also evaluated the effects of extension of conjugation and the presence of electron donating/withdrawing groups at the 6-position of quinazoline core on the optical properties of the probes. We confirmed from our studies that substitution at the 6-position of quinazoline core does not perturb the binding affinity of the probe to the receptor. Overall, our probes can serve as a good platform for designing effective small molecule inhibitors with variable substitution at the 6-position to improve the hydrophilicity and prevent issues arising from aggregation inside the cells besides illuminating the receptor activation and cellular dynamics.

Keywords

Fluorescent; Kinase; Turn-on; Inhibitor

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