ABSTRACT
The design of potent Pin1 inhibitors has been challenging because its active site specifically recognizes a phospho-protein epitope. The de novo design of phosphate-based Pin1 inhibitors focusing on the phosphate recognition pocket and the successful replacement of the phosphate group with a carboxylate have been previously reported. The potency of the carboxylate series is now further improved through structure-based optimization of ligand-protein interactions in the proline binding site which exploits the H-bond interactions necessary for Pin1 catalytic function. Further optimization using a focused library approach led to the discovery of low nanomolar non-phosphate small molecular Pin1 inhibitors. Structural modifications designed to improve cell permeability resulted in Pin1 inhibitors with low micromolar anti-proliferative activities against cancer cells.
Subject(s)
Benzimidazoles/pharmacology , Carboxylic Acids/pharmacology , Enzyme Inhibitors/pharmacology , Peptidylprolyl Isomerase/antagonists & inhibitors , Phosphates/chemistry , Benzimidazoles/chemical synthesis , Benzimidazoles/chemistry , Carboxylic Acids/chemical synthesis , Carboxylic Acids/chemistry , Catalytic Domain/drug effects , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Humans , Molecular Structure , NIMA-Interacting Peptidylprolyl Isomerase , Peptidylprolyl Isomerase/metabolism , Structure-Activity RelationshipABSTRACT
The P21-activated kinases (PAK) are emerging antitumor therapeutic targets. In this paper, we describe the discovery of potent PAK inhibitors guided by structure-based drug design. In addition, the efflux of the pyrrolopyrazole series was effectively reduced by applying multiple medicinal chemistry strategies, leading to a series of PAK inhibitors that are orally active in inhibiting tumor growth in vivo.
Subject(s)
Antineoplastic Agents/chemical synthesis , Pyrazoles/chemical synthesis , Pyrroles/chemical synthesis , p21-Activated Kinases/antagonists & inhibitors , Administration, Oral , Amides/chemical synthesis , Amides/pharmacokinetics , Amides/pharmacology , Animals , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/pharmacology , Carbamates/chemistry , Carbamates/pharmacokinetics , Carbamates/pharmacology , Crystallography, X-Ray , Dogs , Humans , Hydrogen Bonding , Mice , Models, Molecular , Molecular Conformation , Permeability , Pyrazoles/pharmacokinetics , Pyrazoles/pharmacology , Pyrroles/pharmacokinetics , Pyrroles/pharmacology , Rats , Stereoisomerism , Structure-Activity Relationship , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
Despite abundant evidence that aberrant Rho-family GTPase activation contributes to most steps of cancer initiation and progression, there is a dearth of inhibitors of their effectors (e.g., p21-activated kinases). Through high-throughput screening and structure-based design, we identify PF-3758309, a potent (K(d) = 2.7 nM), ATP-competitive, pyrrolopyrazole inhibitor of PAK4. In cells, PF-3758309 inhibits phosphorylation of the PAK4 substrate GEF-H1 (IC(50) = 1.3 nM) and anchorage-independent growth of a panel of tumor cell lines (IC(50) = 4.7 +/- 3 nM). The molecular underpinnings of PF-3758309 biological effects were characterized using an integration of traditional and emerging technologies. Crystallographic characterization of the PF-3758309/PAK4 complex defined determinants of potency and kinase selectivity. Global high-content cellular analysis confirms that PF-3758309 modulates known PAK4-dependent signaling nodes and identifies unexpected links to additional pathways (e.g., p53). In tumor models, PF-3758309 inhibits PAK4-dependent pathways in proteomic studies and regulates functional activities related to cell proliferation and survival. PF-3758309 blocks the growth of multiple human tumor xenografts, with a plasma EC(50) value of 0.4 nM in the most sensitive model. This study defines PAK4-related pathways, provides additional support for PAK4 as a therapeutic target with a unique combination of functions (apoptotic, cytoskeletal, cell-cycle), and identifies a potent, orally available small-molecule PAK inhibitor with significant promise for the treatment of human cancers.
Subject(s)
Cell Proliferation/drug effects , Models, Molecular , Neoplasms/metabolism , Pyrazoles/pharmacology , Pyrroles/pharmacology , Signal Transduction/drug effects , p21-Activated Kinases/antagonists & inhibitors , Cell Line, Tumor , Cell Survival/drug effects , Crystallography , Guanine Nucleotide Exchange Factors/metabolism , Humans , Neoplasms/drug therapy , Phosphorylation/drug effects , Pyrazoles/chemistry , Pyrazoles/metabolism , Pyrroles/chemistry , Pyrroles/metabolism , Rho Guanine Nucleotide Exchange FactorsABSTRACT
Following the discovery of a novel series of phosphate-containing small molecular Pin1 inhibitors, the drug design strategy shifted to replacement of the phosphate group with an isostere with potential better pharmaceutical properties. The initial loss in potency of carboxylate analogs was likely due to weaker charge-charge interactions in the putative phosphate binding pocket and was subsequently recovered by structure-based optimization of ligand-protein interactions in the proline binding site, leading to the discovery of a sub-micromolar non-phosphate small molecular Pin1 inhibitor.
Subject(s)
Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Peptidylprolyl Isomerase/antagonists & inhibitors , Peptidylprolyl Isomerase/metabolism , Binding Sites , Crystallography, X-Ray , Drug Design , Humans , Models, Molecular , NIMA-Interacting Peptidylprolyl Isomerase , Peptidylprolyl Isomerase/chemistry , Protein Binding , Structure-Activity RelationshipABSTRACT
Pin1 is a member of the cis-trans peptidyl-prolyl isomerase family with potential anti-cancer therapeutic value. Here we report structure-based de novo design and optimization of novel Pin1 inhibitors. Without a viable lead from internal screenings, we designed a series of novel Pin1 inhibitors by interrogating and exploring a protein crystal structure of Pin1. The ligand efficiency of the initial concept molecule was optimized with integrated SBDD and parallel chemistry approaches, resulting in a more attractive lead series.
Subject(s)
Enzyme Inhibitors/chemistry , Peptidylprolyl Isomerase/antagonists & inhibitors , Amino Acid Sequence , Binding Sites , Combinatorial Chemistry Techniques , Computer Simulation , Crystallography, X-Ray , Drug Design , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Humans , NIMA-Interacting Peptidylprolyl Isomerase , Peptidylprolyl Isomerase/metabolism , Structure-Activity RelationshipABSTRACT
Novel tricyclic benzimidazole carboxamide poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors have been synthesized. Several compounds were found to be powerful chemopotentiators of temozolomide and topotecan in both A549 and LoVo cell lines. In vitro inhibition of PARP-1 was confirmed by direct measurement of NAD+ depletion and ADP-ribose polymer formation caused by chemically induced DNA damage.
