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Modular synthetic approaches for the inhibition and assay of proteases involved in transcription factor processing and cell differentiation

Reference	BB/D005957/1
Principal Investigator / Supervisor	Professor Brian Walker
Co-Investigators / Co-Supervisors	Professor Brendan Gilmore, Dr ALEXANDRA IRVINE, Professor Stella Martin, Professor Chris Shaw
Institution	Queen's University of Belfast
Department	Sch of Pharmacy
Funding type	Research
Value (£)	211,743
Status	Completed
Type	Research Grant
Start date	01/12/2005
End date	30/11/2008
Duration	36 months

Abstract

The controlled action of intracellular proteases is pivotal for a diverse range of processes including peptide hormone and neurotransmitter processing, transcriptional control, cellular differentiation and apoptosis. The purpose of this proposal is to develop modular synthetic approaches (including 'click chemistry' and chemo selective ligation) for the generation and intracellular delivery of internally quenched fluorescent substrate sequences and small molecule inhibitory modules. The latter will be based upon [5,5] trans-lactone templates and peptidyl diphenylphosphonates. As a paradigm, we propose to develop synthetic approaches for the generation of inhibitor and substrates modules for proteases that process some members of the Stat (signal transducers and activators of transcription) family of proteins. The Stat proteins play crucial roles in a broad spectrum of cellular processes including differentiation, growth and survival of myeloid progenitor cells, and the clonally expansion of immune cells. It is anticipated that our approaches will afford an opportunity to relate changes in cellular function/fate of living cells to the deliberate manipulation of intracellular protease activity, rather than having to depend on current 'post event' activity measurements, which are almost exclusively performed on sub-cellular fractions or whole lysates, obtained from disrupted cells.

Summary

Proteases are Nature's molecular scissors. They catalyse the cutting (hydrolysis) of peptides and proteins, with exquisite precision. A broad spectrum of processes within the cells that make up our tissues and organs, are controlled by the regulated action of proteases that are also located within various compartments of these cells. Examples of such processes are cell division, cell growth, programmed cell death (apoptosis), and the conversion (differentiation) of stem cells into cells with specialised function. At present, there are very few available methods that permit researchers interested in these processes to view or observe the 'action' of these proteases in living cells. Also, in some instances, it would be highly desirable to inhibit or block the action of these proteases in living cells. The ability to do so would enable researchers to pinpoint precisely the contribution an individual protease makes to cell viability and the fate of the cell. The purpose of this work is to develop chemical approaches that will lead to the generation of smart methods for the delivery of reagents into living cells. These reagents would give a visible signal of the action of a single protease within a group of cells. Other reagents that block the action of key proteases will also be developed. These protease inhibitors will be investigated for their ability to block the differentiation of stem cells. Ultimately, such reagents might find application in preserving bone marrow stem cells in their immature state, so as to protect them from harmful drugs used in the treatment of cancer patients.

Committee	Closed Committee - Biochemistry & Cell Biology (BCB)
Research Topics	Synthetic Biology, Technology and Methods Development
Research Priority	X – Research Priority information not available
Research Initiative	Selective Chemical Intervention In Biological Systems (SCIBS) [2005]
Funding Scheme	X – not Funded via a specific Funding Scheme

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