PROTEASES AND CHEMOTHERAPY INDUCED APOPTOSIS

Project: Research project

Project Details

Description

Apoptosis is a morphologically and biochemically distinct form of cell
death that has been observed during development, after withdrawal of
trophic hormones or treatment with chemotherapeutic agents, and during the
course of cell-mediated immunotoxicity. The present proposal seek funds
for collaborative studies by two laboratories with a long-standing
interest in the mechanism of chemotherapy-induced apoptosis. Previous
studies from these laboratories have demonstrated that l) topoisomerase-
directed agents and a variety of other clinically useful antineoplastic
agents induce apoptosis in a variety of cell types, 2) the morphologic
changes of apoptosis are accompanied by the early, quantitative cleavage
of a number of nuclear proteins including poly(ADP-ribose polymerase) and
the nuclear lamins, and 3) the morphological and biochemical events of
apoptosis can be recapitulated with synchrony and high fidelity in a
unique cell-free assay developed using the cytosol from cells committed to
apoptosis. Using this cell-free system, we have more recently identified
two distinct protease activities that appear to play pivotal roles during
the course of apoptosis. One, termed priCE (protease resembling
interleukin-1-beta converting enzyme), cleaves the nuclear enzyme
poly(ADP-ribose) polymerase prior to the earliest detectable cleavage of
DNA into nucleosomal fragments. Inhibitors of priCE prevent all of the
morphological and biochemical events associated with apoptosis (including
cleavage of protein and DNA), suggesting that activation of priCE occurs
very early in the process of apoptosis. A second protease with a distinct
inhibitor profile is responsible for lamin degradation that occurs later
in apoptosis and is required for the nuclear reorganization that results
in apoptotic bodies. We now propose a series of experiments designed to
characterize these proteases, investigate their structure and regulation,
and further evaluate their role in controlling apoptosis. in particular,
we propose to l) clone the cDNA for priCE using one of three different
strategies, 2) utilize a novel fluorogenic assay and immunological probes
against priCE to study the regulation of priCE in tissue culture cells and
clinical tumor specimens that differ in their intrinsic ability to undergo
apoptosis, 3) identify additional priCE substrates, 4) characterize in
greater detail the lamin protease, and 5) isolate and characterize a
recently demonstrated naturally-occurring inhibitor of apoptosis. These
studies should provide insight into the structure and regulation of
biochemical activities that appear to play a pivotal role in the control
of chemotherapy-induced cell death.
StatusFinished
Effective start/end date4/1/955/31/11

Funding

  • National Cancer Institute: $336,291.00
  • National Cancer Institute: $329,695.00
  • National Cancer Institute: $320,095.00
  • National Cancer Institute: $316,927.00
  • National Cancer Institute: $328,884.00
  • National Cancer Institute: $340,987.00
  • National Cancer Institute: $331,053.00

ASJC

  • Medicine(all)

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