The loss of function of key components of growth regulatory pathways results in unprogrammed proliferation, the primary characteristic of human cancer. The overall goal of this proposal is to identify and characterize promising lead compounds that may be utilized as models to develop therapeutic agents for cancer. Potential targets for such inhibitors include those regulatory molecules that provide functions necessary for growth; one such class of proteins is the cyclin family and their associated catalytic subunits, the cyclin-dependent kinases (cdk). Cyclin/cdk functions are responsible for regulating key checkpoints that allow traverse through and transitions between different stages of the cell cycle, and loss of cdk4 and cdk2 functions have been demonstrated to result in a loss of growth and reversion of a transformed phenotype in a number of cell models. This proposal seeks to identify peptide sequences and other structural motifs that specify binding to and inhibition of cyclin-cdk activity to develop clinically useful therapeutics. This will be accomplished through the completion of four Specific Aims. In the first aim, peptides that interact with cyclin/cdk subunits will be identified from a phage of display peptide library, and their sequences will be utilized to develop targeted combinatorial from a phage display peptide library, and their sequences will be utilized to develop targeted combinatorial libraries. In the second aim, peptides that interact specifically with cyclin/cdk subunits, second generation combinatorial libraries based on their conserved structural motifs and additional combinatorial libraries developed in Project 1 will be screened by a high throughput biochemical assay to identify those molecules capable of inhibiting enzymatic activity.
The third aim will utilize human tumor cell culture models to ascertain the specificity and selectivity of potential inhibitors in whole cells. In the fourth aim, those compounds which have proven to be effective in whole cells will be subjected to analysis in tumorigenic models, both in cell cultures and in nude mice, to assess their ability to inhibit growth and revert transformation.
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