Drug design with Cdc7 kinase: a potential novel cancer therapy target

Abstract

Identification of novel molecular targets is critical in development of new and efficient cancer therapies. Kinases are one of the most common drug targets with a potential for cancer therapy. Cell cycle progression is regulated by a number of kinases, some of which are being developed to treat cancer. Cdc7 is a serine-threonine kinase originally discovered in budding yeast, which has been shown to be necessary to initiate the S phase. Inhibition of Cdc7 in cancer cells retards the progression of the S phase, accumulates DNA damage, and induces p53-independent cell death, but the same treatment in normal cells does not significantly affect of less than viability. Low-molecular-weight compounds that inhibit Cdc7 kinase with an IC(50) 10 nM have been identified, and shown to be effective in the inhibition of tumor growth in animal models. Thus Cdc7 kinase can be recognized as a novel molecular target for cancer therapy.

Keywords: ATP-binding pocket; Cdc7 kinase; DNA damages; cell cycle; genome stability; kinase inhibitor; replication fork.

Figure 1

Homology analyses of human Cdc7 kinase. A)The human kinome tree. Kinase family members were classified into the following eight subgroups on the basis of their primary structures: CAMK (calcium/calmodulin-dependent kinase group), TK (tyrosine kinase group), RGC (receptor guanylyl cyclase group), TKL (tyrosine kinase-like group), STE (sterile phenotype kinase group), CK1 (cell kinase 1/casein kinase 1 group), AGC (protein kinases A, G and C group), CMGC (cyclin-dependent-kinase [CDK], mitogen-activated-kinase [MAPK], glycogen-synthase-kinase [GSK] and CDK-like kinase group). B) A phylogenetic tree of Cdc7 and some human protein kinases which are most similar to Cdc7.

Figure 2

Scheme of initiation of eukaryotic DNA replication and action of Cdc7 kinase. Eukaryotic DNA replication is initiated by binding of ORC (origin recognition complex) at a replication origin. With the aid of Cdc6 and Cdt1 proteins, Mcm (minichromosome maintenance) is delivered at the origin, generating pre-RC (pre-replicative complex). Cdc45 associates with the pre-RC, followed by GINS complex. Phosphorylation by Cdk and Cdc7 is required for this step. It was reported that phosphorylation of the N-terminal tails of Mcm2, Mcm4, and Mcm6 proteins facilitates association of Cdc45 and other proteins with Mcm (Masai et al 2006; Sheu et al 2006). Active replication forks are generated by association of three DNA polymerases at the origin.

Figure 3

Schematic drawing of Cdc7 kinase. Human Cdc7 (574 amino acids) are composed of kinase conserved domains (white segments) and three kinase insert sequences (KI-1, -2, and -3; gray segments). Roman numerals indicate the kinase subdomains defined previously (Hanks et al 1988). KI-1 is not obvious in human Cdc7, but is longer in Cdc7 from other species. Although the length and sequences of kinase inserts are less conserved between species, they are located at the conserved positions among the kinase subdomains. The gatekeeper residue for the ATP-binding pocket (methionine 134) is also indicated.

Figure 4

Schematic representation of general binding mode of ATP; interactions with specific segments of protein kinases: A) solvent accessible region, B) hydrophobic region II, C) adenine-binding region, D) sugar pocket, E) hydrophobic region I, F) phosphate-binding region.

Figure 5

A homology model of Cdc7 kinase. A) Mapping of important residues within the ATP-binding pocket of Cdc7 kinase based on a homology model. The dotted circle area marks the hydrophobic area (adenine-binding region) formed by a hydrophobic sandwich with two residues Met118 and Ile64. Possible hydrogen bonds between an inhibitor and the ATP-binding site of Cdc7 are indicated by the arrows. B) A model of staurosporine bound to the ATP-binding site of Cdc7 shown by Van der Waals surfaces. The negatively and positively charged regions are shown in pink and blue, respectively, on the Van der Waals surfaces. Oxygen and nitrogen are shown in red and blue, respectively. The carbon atoms in the ligand (staurosporine) and in the protein are shown in white and green, respectively.

Figure 6

Cdc7 kinase inhibitors reported by Nerviano Medical Sciences. 1, PHA-767491 (IC₅₀ =10 nM), 2, 3-aminopyrimidine analog (IC₅₀ = 7 nM), 3, (S)-2-(2-aminopyrimidiny-4-yl)-7-(2-fluoroethyl)-1,5,6,7-tetrahydropyrrolo[3,2-c]pyridinone (IC₅₀ =2 nM), and 4, the pyrrolopyridinone analog (IC₅₀ = 3 nM).

Figure 7

Structures of other Cdc7 inhibitors. 5, Tricyclic Cdc7 inhibitor (Roche); 6, indazolylpyrimidin-2(1H)-one inhibitor (Novartis); 7, thienopyrazole-base inhibitor (Pfizer); 8, 2-pyrimidyl-5-amidothiophene analog (Novartis Vaccines and Diagnostics); 9, imidazolone-based inhibitor (Sanofi-Aventis).