Our Science – Lountos Website
George T. Lountos, Ph.D.
Field of Degree: Chemistry
Degree Institution: Georgia Institute of Technology
Date Degree Granted: 2005
One area of research that I am involved in is in the structure-assisted design of novel Chk2 inhibitors. Human checkpoint kinase 2 (Chk2) is a ser/thr kinase that is involved in the ATM/Chk2 checkpoint pathway that is activated by genomic instability and DNA damage and results in the arrest of the cell cycle to allow for either DNA repair to occur, or to trigger apoptosis if the DNA damage is severe. As a tumor suppressor, Chk2 plays an important role in maintaining genomic integrity by acting as a signal transducer of DNA damage. Chk2 is endogeneously activated in precancerous lesions with genomic instability and in cancer cells grown in culture. In addition, studies have shown that activated Chk2 in tumor cells plays an important role in the expression of survivin, which is crucial for tumor survival. Chk2 has been identified as a promising target for anti-cancer drug design, but very few selective Chk2 inhibitors have been identified to date. Thus, the development of novel Chk2 inhibitors is of great interest, not only from a drug development viewpoint, but also to provide molecular tools with which the biological functions of Chk2 can be further elucidated. There are two rationales for the development of Chk2 inhibitors. First, selective inhibition in p53-defective tumor cells may provide chemo/radiosensitization because Chk2 is activated in tumor cells by a wide range of chemotherapeutic drugs and ionizing radiation. Thus, selective inhibition of Chk2 could increase the therapeutic indices of DNA-targeted agents in these cells. Second, inhibition of Chk2 in normal cells may also protect normal tissues from p53-induced apoptosis during chemotherapy or radiation therapy. We have initiated a drug design effort targeting Chk2 by screening compounds from the NCI Open Repository Library and identified a bis-guanylhydrazone, NSC109555, as a potent and selective inhibitor with an IC50=240 nM. A 2.05 Angstrom co-crystal structure of Chk2 complexed with NSC109555 has been solved, thereby establishing an opportunity for structure-assisted drug design and optimization. After several rounds of structure-based optimization of the lead compound and analogs, we have identified several compounds with increased potency against Chk2, the most potent exhibiting an IC50 of 41 pM. Co-crystal structures of Chk2 with 9 of these analogs have been solved at resolutions up to 1.7 Angstroms, yielding structural information on the protein-inhibitor interactions that will guide future optimization of these compounds.
The second area of research involves structure determinations of proteins involved in the type III secretion system of Yersinia pestis. Recently, the atomic resolution structure of the cytoplasmic domain of Yersinia pestis YscU, a regulatory switch involved in type III secretion was determined. Crystal structures of cleaved and uncleaved forms of the YscU cytoplasmic domain, an essential component of the type III secretion system (T3SS) in Yersinia pestis, have been solved by single-wavelength anomalous dispersion and refined with X-ray diffraction data extending up to atomic resolution (1.13 Angstroms). These crystallographic studies provide structural insights into the conformational changes induced upon auto-cleavage of the cytoplasmic domain of YscU. The structures indicate that the cleaved fragments remain bound to each other. The conserved NPTH sequence that contains the site of the N263-P264 peptide bond cleavage is found on a beta-turn which, upon cleavage, undergoes a major reorientation of the loop away from the catalytic N263, resulting in altered electrostatic surface features at the site of cleavage. The YscU structures determined here also correlate well with the auto-cleavage mechanism described for the flagellar homolog FlhB and E. coli EscU.
We have also recently solved the X-ray crystal structure of the human dual-specificity phosphatase 14 at 1.88 Angstroms resolution and also of the human dual-specificity phosphatase 27. Dual-specificity phosphatases are a class of protein tyrosine phosphatases that are able to dephosphorylate either phosphotyrosine, phosphoserine, or phosphothreonine residues and are involved a number of physiological processes such as cell development, differentiation, and transformation. Although many dual-specificity phosphatases are still poorly characterized, many are being identified as potential therapeutic targets against cancer, infectious disease, inflammatory disorders, and diabetes. The availability of high resolution crystal structures should enable structure-guided experiments to further characterize these enzymes at the molecular level and to discover inhibitors.
I am also working on a structure-based drug design project targeting the Yersinia pestis YOPH, a protein tyrosine phosphatase virulence factor as well as the dual-specificity phosphatase variola H1 phosphatase from smallpox.
This page was last updated on 8/4/2011.