Abstract

The PIM-1 kinase is a serine/threonine kinase that has been implicated in the development of many human cancers, including leukemias, lymphomas, and prostate cancer. We have endeavored to identify and characterize a selective inhibitor of the PIM- 1 kinase. Such an inhibitor would have utility as a laboratory tool for the study of PIM-1 kinase function and as a template for the design of molecular therapeutics for diseases in which PIM-1 kinase activity is dysregulated.

Using phage display techniques we identified the peptide, HGVKKRPHNPYG, as a probable pseudosubstrate of the PIM-1 kinase with inhibitory activity in the low micromolar range in in vitro kinase assays. However, the peptide lacked sufficient potency for our objectives; hence we employed a second approach - the computational design of a novel PIM-1 kinase antagonist.

We created a homology model of the PIM-1 kinase that was subsequently determined to be structurally similar to a recently published crystal structure of the PIM-1 kinase. Docking studies using the PIM-1 kinase model demonstrated that the AFFINITY module of Insightll, under the parameters employed, was unable to accurately predict the potency of known PIM-1 kinase inhibitors. Consequently, we utilized comparative molecular field analysis, which resulted in a strong correlation of predicted IC50 vs. experimental IC50 values for known flavonoid inhibitors of the PIM-1 kinase. Thus we have created the first predictive model that may be used for the rational design of small molecule inhibitors of the PIM-1 kinase.

We have also performed an extensive screen of small molecules with structures similar to known kinase inhibitors and identified quercetagetin as a potent inhibitor of the PIM-1 kinase, with an in vitro IC50 of 0.34 pM. Lineweaver-burk analyses of the reaction kinetics demonstrated that quercetagetin is an ATP competitive inhibitor the PIM-1 kinase. We have also determined that quercetagetin is selective for the PIM-1 kinase over many other kinases and that quercetagetin successfully inhibits PIM-1 kinase activity in mammalian cells. Therefore quercetagetin may be an effective tool for the laboratory study of the PIM-1 kinase, and may serve as a template for the design of molecular therapeutics directed against the PIM-1 kinase.

LLU Discipline

Microbiology

Department

Microbiology

School

Graduate School

First Advisor

Michael B. Lilly

Second Advisor

Carlos Casiano

Third Advisor

Penelope Duerken-Hughes

Fourth Advisor

Mark Johnson

Fifth Advisor

Jonathan Neidigh

Sixth Advisor

Lawrence Sowers

Degree Name

Doctor of Philosophy (PhD)

Degree Level

Ph.D.

Year Degree Awarded

2005

Date (Title Page)

9-2005

Language

English

Library of Congress/MESH Subject Headings

Protein Kinases; Proto-Oncogene Proteins c-pim-1 -- antagonists and inhibitors; Neoplasms -- genetics.

Type

Dissertation

Page Count

xv; 132

Digital Format

PDF

Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Collection

Loma Linda University Electronic Theses and Dissertations

Collection Website

http://scholarsrepository.llu.edu/etd/

Repository

Loma Linda University. Del E. Webb Memorial Library. University Archives

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Microbiology Commons

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