Date of Award

12-2007

Document Type

Thesis

Degree Name

Master of Science (MS)

Program

Pharmaceutical Sciences

Research Advisor

Isaac O. Donkor, Ph.D.

Committee

John K. Buolamwini, Ph.D Wei Li, Ph.D Duane D. Miller, Ph.D. Evgueni Pinkhassik, Ph.D.

Abstract

Calpain is a class of intracellular cytoplasmic cysteine proteases.1 The enzyme participates in different intracellular signaling pathways that are mediated by Ca2+.2 The two major isoforms of calpain universally distributed in most mammalian tissues are calpain 1 (µ-calpain) and calpain 2 (m-calpain). The exact in vivo function of the enzyme is not clear, but calpain has been implicated in a variety of physiological and pathological conditions,3 such as cancer, stroke, cardiac ischaemia, muscular dystrophy, cataract and Alzheimer’s disease. Calpain inhibitors are therefore of interest as therapeutic agents and as biomedical tools.

Several potent calpain inhibitors isolated from natural sources as well as synthesized in the laboratory have been reported (Chapter 1.4). Unfortunately, most of the inhibitors show poor calpain selectivity, metabolic stability and cell permeability. In an attempt to develop potential calpain inhibitors based on the X-ray crystal structure of the µ-calpain, NCI compound library was screened by virtual screening method and diazosulfonamide 1 (Ki = 1.0 ± 0.02 µM) was identified as a new nonpeptide competitive inhibitor of µ-calpain. Analogues of 1 were synthesized to explore structure requirements of 1 (Chapter 2). In order to test the hypothesis that derivatives of diazosulfonamide 1 with an electrophilic group can have a covalent interaction with cysteine at the active site of calpain, novel sulfonamide-based peptidomimetic analogues of 1 were synthesized and assayed for their ability to inhibit µ-calpain utilizing a kinetic fluorescence assay and for their anti-tumor ability by SRB colorimetric assay (Chapter 3). Introduction of the electrophilic functionality significantly enhanced calpain inhibition. From 13 target compounds, 7 compounds had better calpain 1 inhibition (Ki ranging from 9 to 500nM) than 1 and 5 showed good anticancer activity (GI50 ranging from 4 to 22µM). Sulfonamide-based peptidomimetic analogue 19 with Ki of 9 nM is over 100-fold more potent than the lead diazosulfonamide 1. Compound 16 was the most effective anticancer agent (GI50 4µM) of the series.

DOI

10.21007/etd.cghs.2007.0361

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