Benzamide capped peptidomimetics as non-ATP competitive inhibitors of CDK2 using the REPLACE strategy

Abstract

Inhibition of Cyclin Dependent Kinase 2 (CDK2) in complex with cyclin A in G1/S phase of the cell cycle has been shown to promote selective apoptosis of cancer cells through the E2F1 pathway. An alternative approach to catalytic inhibition is to target the substrate recruitment site also known as the cyclin binding groove (CBG) to generate selective non-ATP competitive inhibitors. The REPLACE strategy has been applied to identify fragment alternatives and substituted benzoic acid derivatives were evaluated as a promising scaffold to present appropriate functionality to mimic key peptide determinants. Fragment Ligated Inhibitory Peptides (FLIPs) are described which potently inhibit both CDK2/cyclin A and CDK4/cyclin D1 and have preliminary anti-tumor activity. A structural rationale for binding was obtained through molecular modeling further demonstrating their potential for further development as next generation non ATP competitive CDK inhibitors.

Cyclin-Dependent Kinase 2 Inhibitors in Cancer Therapy: An Update

“…Pentapeptides (e.g., TAALS) having a similar mechanism of action have also been described . In addition, drug design strategies have been used to identify peptidomimetic inhibitors (e.g., HAKRRLIF based on a sequence found in the endogenous CDK inhibitor, p27) that act at the cyclin groove substrate recruitment site identified on the cyclin subunit − or the protein–protein interaction interface of CDK2 and its regulatory cyclin partners (C4, derived from amino acids 285–306 in the α5-helix of cyclin A) to develop non-ATP-competitive CDK2 inhibitors (Figure A). However, the use of these peptides is limited by their poor pharmacokinetic properties.…”

Targeting Protein-Protein Interactions to Inhibit Cyclin-Dependent Kinases

“…CDK2 requires binding to the Cyclin A or Cyclin E family of cyclin proteins [11]. To date, efforts to target PPI for CDK2 has focused on the interactions between CDK2 and Cyclin A [16][17][18]. Much of the effort in this area has focused on a strategy to target either the p21Waf naturally occurring inhibitor of CDK2 or the cyclin binding groove located on CDK2 [11,19].…”

“…Much of the effort in this area has focused on a strategy to target either the p21Waf naturally occurring inhibitor of CDK2 or the cyclin binding groove located on CDK2 [11,19]. The strategy called REPLACE (Replacement with Partial Ligand Alternatives using Computation Enrichment) has resulted in lead molecules that disrupt CDK2-Cyclin A interactions and a decrease in CDK2 enzymatic activity [16][17][18][19]. The strategy involves (1) identifying and/or determining the three-dimensional (3D) structure of a lead peptide in contact with the cyclin binding groove, (2) identify structural and chemical features of the peptide-protein interactions, (3) truncate the peptide starting at the N-terminus, (4) use computational techniques to identify small-molecule alternatives to the truncated peptide (partial ligand alternatives, PLAs) that retain key binding characteristics to the target CDK, (5) identify commercial sources for or synthesize small molecule alternatives (PLAs), (6) synthesize fragment-ligated peptides (FLIP) by ligating the small molecule alternative (PLA) to the truncated peptide, (7) test the efficacy of the candidate FLIPs in an in vitro binding assay and/or functional assay, and (8) evaluate the efficacy of the candidate FLIPs in a cell viability assay [19].…”

Identification of β-strand mediated protein–protein interaction inhibitors using ligand-directed fragment ligation

“…kinases. [31][32][33][34][35] Subsequently, extension of this concept to a-helix mediated PPIs has been achieved by screening peptide-small molecule hybrids in silico, then preparing promising candidates for experimental validation using click chemistry. [36][37][38] In this work, we instead use reversible hydrazone formation 39 between peptide hydrazones and an aldehyde library together with dynamic ligation screening 40,41 to identify peptide-fragment hybrids.…”