Potent Cdk4 Inhibitor Research Articles

Molecular modeling studies of [4-(3H-benzoimidazol-5-yl)-pyrimidin-2-yl]-amine-based CDK4 inhibitors.

Aim: CDK4 is a promising target for breast cancer therapy. This study aimed to explore the structure-activity relationship of CDK4 inhibitor abemaciclib analogs and design potent CDK4 inhibitors for breast cancer treatment. Methods & results: A faithful 3D quantitative structure-activity relationship model was established by molecular docking, comparative molecular field analysis and comparative molecular similarity index analysis based on 56 abemaciclib analogs. Molecular dynamics simulation studies revealed the key residues of the interaction between CDK4 and inhibitors. Four novel inhibitors with satisfactory predicted binding affinity to CDK4 were designed. Conclusion: The 3D quantitative structure-activity relationship and molecular dynamics simulation studies provide valuable insight into the development of novel CDK4 inhibitors.

Design, synthesis and biological assessment of novel CDK4 inhibitor with potent anticancer activity

Efforts toward finding potent CDK4 inhibitor for cancer therapy, a series of fluorine substituted pyrrolo[2,3-d]pyrimidine derivatives were designed, synthesized, and evaluated. Among them, the optimal lead compound 18i was discovered with potent activity against CDK4 at the nanomolar level (IC50 = 2.5 nM) and exquisite selectivity which demonstrated only modest activity against 3 out of the 394 protein kinases. 18i exhibited a much greater in vitro antiproliferative activity against several human cancer cell lines than that of the approved drug ribociclib. Further mechanism studies revealed that 18i effectively stimulated cancer cell cycle arrest in G1 phase and induced tumor cell apoptosis. In the comparison of in vivo therapeutic effects in xenograft mouse models of breast cancer, oral administration of 18i showed a significantly better degree of inhibitory effect to ribociclib without obvious toxicity. All of the results indicated that 18i could be a promising CDK4 inhibitor for treating malignancies.

Insight into the Interactions between Novel Isoquinolin-1,3-Dione Derivatives and Cyclin-Dependent Kinase 4 Combining QSAR and Molecular Docking

Several small-molecule CDK inhibitors have been identified, but none have been approved for clinical use in the past few years. A new series of 4-[(3-hydroxybenzylamino)-methylene]-4H-isoquinoline-1,3-diones were reported as highly potent and selective CDK4 inhibitors. In order to find more potent CDK4 inhibitors, the interactions between these novel isoquinoline-1,3-diones and cyclin-dependent kinase 4 was explored via in silico methodologies such as 3D-QSAR and docking on eighty-one compounds displaying potent selective activities against cyclin-dependent kinase 4. Internal and external cross-validation techniques were investigated as well as region focusing, bootstraping and leave-group-out. A training set of 66 compounds gave the satisfactory CoMFA model (q 2 = 0.695, r 2 = 0.947) and CoMSIA model (q 2 = 0.641, r 2 = 0.933). The remaining 15 compounds as a test set also gave good external predictive abilities with r 2 pred values of 0.875 and 0.769 for CoMFA and CoMSIA, respectively. The 3D-QSAR models generated here predicted that all five parameters are important for activity toward CDK4. Surflex-dock results, coincident with CoMFA/CoMSIA contour maps, gave the path for binding mode exploration between the inhibitors and CDK4 protein. Based on the QSAR and docking models, twenty new potent molecules have been designed and predicted better than the most active compound 12 in the literatures. The QSAR, docking and interactions analysis expand the structure-activity relationships of constrained isoquinoline-1,3-diones and contribute towards the development of more active CDK4 subtype-selective inhibitors.

Abstract A66: Potent CDK4 inhibitor in human breast cancer therapy

Abstract Deregulated cell cycle is a common feature in many human cancers. Overexpression of cyclinD1 oncogene and highly activation of the cyclinD1/Cdk4Cdk6 kinase modulate the tumor initiation, tumor progression and the chemo-sensitivity of cancer cells. Targeting G1 phase of cell cycle leading cancer cell into senescence or quiescence stage or causing cell death has become a promising approach in cancer treatment. Here, we report the in vitro anti-tumor functions of the synthetic Cdk4 inhibitor, Naphtho [2, 1-α] pyrrolo [3, 4-c] carbazole-5, 7 (6H, 12H)-dione (NPCD) in a panel of human breast cancer cell lines. NPCD is able to inhibit cancer cell growth, arrest cancer cells at G1 phase and cause cell death as results of diminished Cdk4Cdk6 phosphorylation of the Rb protein without disrupting the cyclinD1/Cdk4Cdk6 complex formation or affecting G1 phase proteins intracellular localization. The sensitivity to NPCD is associated with the intracellular cyclinD1 and Cdk4 or Cdk6 proteins level. Upon NPCD treatment, the changes in several G1-S phase proteins levels differ between cell lines which could because different cell lines develop different signaling networks responding to same stimuli. The dramatically decreased p27kip protein is a common feature between cell lines we studied upon NPCD treatment; it could be either a result of NPCD induced G1 arrest and inactivation of cyclinD1/Cdk4Cdk6 or could be an upstream mediator to cause Cdk4Cdk6 inactivation and G1 arrest. In recent years, the functions of p27kip protein have been extended involving in cell cycle regulation, embryonic development, tumor genesis and tumor progression, consequently, pursuing the mechanisms resulting rapidly decreased p27kip protein in responding to NPCD may help us to better understand the function of this protein in deregulated cell cycle system. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A66.

Recent Research in Selective Cyclin-Dependent Kinase 4 Inhibitors for Anti-Cancer Treatment

It is well known that cyclins and cyclin-dependent kinases (CDKs) play essential roles in regulation of the cell cycle. In past two decades, the scientific researches suggest that the cyclin D1/ CDK4 complex is a key regulator of the transition through the G1 phase of the cell cycle. Moreover, deregulation of the cyclin D /CDK4 pathway has been identified in multiple tumor types. Thus, CDK4 is a genetically validated therapeutic target; hence, there has been a surge of interests in finding selective CDK4 inhibitors as anti-cancer agents. This review will give the recent progress in the studies of structure, functions of CDK4 and highly selective and potent CDK4 inhibitors.

Structure-based generation of a new class of potent Cdk4 inhibitors: new de novo design strategy and library design.

As a first step in structure-based design of highly selective and potent Cdk4 inhibitors, we performed structure-based generation of a novel series of Cdk4 inhibitors. A Cdk4 homology model was constructed according to X-ray analysis of an activated form of Cdk2. Using this model, we applied a new de novo design strategy which combined the de novo design program LEGEND with our in-house structure selection supporting system SEEDS to generate new scaffold candidates. In this way, four classes of scaffold candidates including diarylurea were identified. By constructing diarylurea informer libraries based on the structural requirements of Cdk inhibitors in the ATP binding pocket of the Cdk4 model, we were able to identify a potent Cdk4 inhibitor N-(9-oxo-9H-fluoren-4-yl)-N'-pyridin-2-ylurea 15 (IC(50) = 0.10 microM), together with preliminary SAR. We performed a docking study between 15 and the Cdk4 model and selected a reasonable binding mode which is consistent with the SAR. Further modification based on the proposed binding mode provided a more potent compound, N-[(9bR)-5-oxo-2,3,5,9b-tetrahydro-1H-pyrrolo[2,1-a]isoindol-9-yl]-N'-pyridin-2-ylurea 26a (IC(50) = 0.042 microM), X-ray analysis of which was accomplished by the soaking method. The predicted binding mode of 15 in Cdk4 was validated by X-ray analysis of the Cdk2-26a complex.

Solution structure of p21Waf1/Cip1/Sdi1 C-terminal domain bound to Cdk4

Cyclin-dependent kinase (Cdk) inhibitor p21Waf1/Cip1/Sdi1 a multifunctional protein, has a major role as tumor suppressor, mediating G1/S arrest through inhibition of Cdks. Recent biological studies of Cyclin D1/Cdk4 have proposed that p21 C-terminal domain (p21CT) plays a key role as a potent Cdk4 inhibitor. We report here solution structures of p21CT for both the free and Cdk4-bound forms using 2D transferred NOE spectroscopy and dynamical simulated annealing calculations. Even though p21CT peptide is very flexible in the free state, when it bound to Cdk4, the structure becomes well structured in the binding domain. Therefore we propose that p21CT experiences an extensive conformational change upon Cdk4 binding. This structural change of p21CT may suggest the molecular mechanism of p21 for specificity and inhibition mode to assemble different cyclin-Cdk complexes. Especially, our data suggests that the D149FYHSKRR156 region of p21 is critical for Cdk4 binding, indicating that the major driving force for complex originates from hydrophobic interaction between p21 and Cdk4.

A Novel Approach for the Development of Selective Cdk4 Inhibitors: Library Design Based on Locations of Cdk4 Specific Amino Acid Residues

Identification of a selective inhibitor for a particular protein kinase without inhibition of other kinases is critical for use as a biological tool or drug. However, this is very difficult because there are hundreds of homologous kinases and their kinase domains including the ATP binding pocket have a common folding pattern. To address this issue, we applied the following structure-based approach for designing selective Cdk4 inhibitors: (1) identification of specifically altered amino acid residues around the ATP binding pocket in Cdk4 by comparison of 390 representative kinases, (2) prediction of appropriate positions to introduce substituents in lead compounds based on the locations of the altered amino acid residues and the binding modes of lead compounds, and (3) library design to interact with the altered amino acid residues supported by de novo design programs. Accordingly, Asp99, Thr102, and Gln98 of Cdk4, which are located in the p16 binding region, were selected as first target residues for specific interactions with Cdk4. Subsequently, the 5-position of the pyrazole ring in the pyrazol-3-ylurea class of lead compound (2a) was predicted to be a suitable position to introduce substituents. We then designed a chemical library of pyrazol-3-ylurea substituted with alkylaminomethyl groups based on the output structures of de novo design programs. Thus we identified a highly selective and potent Cdk4 inhibitor, 15b, substituted with a 5-chloroindan-2-ylaminomethyl group. Compound 15b showed higher selectivity on Cdk4 over those on not only Cdk1/2 (780-fold/190-fold) but also many other kinases (>430-fold) that have been tested thus far. The structural basis for Cdk4 selective inhibition by 15b was analyzed by combining molecular modeling and the X-ray analysis of the Cdk4 mimic Cdk2-inhibitor complex. The results suggest that the hydrogen bond with the carboxyl group of Asp99 and hydrophobic van der Waals contact with the side chains of Thr102 and Gln98 are important. Compound 15b was found to cause cell cycle arrest of the Rb(+) cancer cell line in the G(1) phase, indicating that it is a good biological tool.

Pyrido[2,3-d]pyrimidin-7-one inhibitors of cyclin-dependent kinases.

The identification of 8-ethyl-2-phenylamino-8H-pyrido[2, 3-d]pyrimidin-7-one (1) as an inhibitor of Cdk4 led to the initiation of a program to evaluate related pyrido[2, 3-d]pyrimidin-7-ones for inhibition of cyclin-dependent kinases (Cdks). Analysis of more than 60 analogues has identified some clear SAR trends that may be exploited in the design of more potent Cdk inhibitors. The most potent Cdk4 inhibitors reported in this study inhibit Cdk4 with IC(50) = 0.004 microM ([ATP] = 25 microM). X-ray crystallographic analysis of representative compounds bound to the related kinase, Cdk2, reveals that they occupy the ATP binding site. Modest selectivity between Cdks is exhibited by some compounds, and Cdk4-selective inhibitors block pRb(+) cells in the G(1)-phase of the cell division cycle.