Publications by authors named "Jay A Bertrand"

7 Publications

  • Page 1 of 1

Structure of human Bloom's syndrome helicase in complex with ADP and duplex DNA.

Acta Crystallogr D Biol Crystallogr 2014 May 30;70(Pt 5):1465-75. Epub 2014 Apr 30.

Vertex Pharmaceuticals (Europe), Abingdon, Oxfordshire, England.

Bloom's syndrome is an autosomal recessive genome-instability disorder associated with a predisposition to cancer, premature aging and developmental abnormalities. It is caused by mutations that inactivate the DNA helicase activity of the BLM protein or nullify protein expression. The BLM helicase has been implicated in the alternative lengthening of telomeres (ALT) pathway, which is essential for the limitless replication of some cancer cells. This pathway is used by 10-15% of cancers, where inhibitors of BLM are expected to facilitate telomere shortening, leading to apoptosis or senescence. Here, the crystal structure of the human BLM helicase in complex with ADP and a 3'-overhang DNA duplex is reported. In addition to the helicase core, the BLM construct used for crystallization (residues 640-1298) includes the RecQ C-terminal (RQC) and the helicase and ribonuclease D C-terminal (HRDC) domains. Analysis of the structure provides detailed information on the interactions of the protein with DNA and helps to explain the mechanism coupling ATP hydrolysis and DNA unwinding. In addition, mapping of the missense mutations onto the structure provides insights into the molecular basis of Bloom's syndrome.
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http://dx.doi.org/10.1107/S139900471400501XDOI Listing
May 2014

Structural insight into maternal embryonic leucine zipper kinase (MELK) conformation and inhibition toward structure-based drug design.

Biochemistry 2013 Sep 4;52(37):6380-7. Epub 2013 Sep 4.

Nerviano Medical Sciences , Viale Pasteur 10, 20014 Nerviano, Milan, Italy.

Maternal embryonic leucine zipper kinase (MELK) is upregulated in several types of tumor, including breast, prostate, and brain tumors. Its expression is generally associated with cell survival, cell proliferation, and resistance to apoptosis. Therefore, the potential of MELK inhibitors as therapeutic agents is recently attracting considerable interest. Here we report the first structures of MELK in complex with AMP-PNP and with nanomolar inhibitors. Our studies shed light on the role of the MELK UBA domain, provide a characterization of the kinase active site, and identify key residues for achieving high potency, laying the groundwork for structure-based drug design efforts.
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http://dx.doi.org/10.1021/bi4005864DOI Listing
September 2013

Tetramerization dynamics of C-terminal domain underlies isoform-specific cAMP gating in hyperpolarization-activated cyclic nucleotide-gated channels.

J Biol Chem 2011 Dec 17;286(52):44811-20. Epub 2011 Oct 17.

Department of Biology and Consiglio Nazionale delle Ricerche-Istituto di Biofisica, University of Milan, Via Celoria 26, 20133 Milan, Italy.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are dually activated by hyperpolarization and binding of cAMP to their cyclic nucleotide binding domain (CNBD). HCN isoforms respond differently to cAMP; binding of cAMP shifts activation of HCN2 and HCN4 by 17 mV but shifts that of HCN1 by only 2-4 mV. To explain the peculiarity of HCN1, we solved the crystal structures and performed a biochemical-biophysical characterization of the C-terminal domain (C-linker plus CNBD) of the three isoforms. Our main finding is that tetramerization of the C-terminal domain of HCN1 occurs at basal cAMP concentrations, whereas those of HCN2 and HCN4 require cAMP saturating levels. Therefore, HCN1 responds less markedly than HCN2 and HCN4 to cAMP increase because its CNBD is already partly tetrameric. This is confirmed by voltage clamp experiments showing that the right-shifted position of V(½) in HCN1 is correlated with its propensity to tetramerize in vitro. These data underscore that ligand-induced CNBD tetramerization removes tonic inhibition from the pore of HCN channels.
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http://dx.doi.org/10.1074/jbc.M111.297606DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3247997PMC
December 2011

Structural basis for CARM1 inhibition by indole and pyrazole inhibitors.

Biochem J 2011 Jun;436(2):331-9

Bristol-Myers Squibb Research and Development, P.O. Box 4000, Princeton, NJ 08543-4000, USA.

CARM1 (co-activator-associated arginine methyltransferase 1) is a PRMT (protein arginine N-methyltransferase) family member that catalyses the transfer of methyl groups from SAM (S-adenosylmethionine) to the side chain of specific arginine residues of substrate proteins. This post-translational modification of proteins regulates a variety of transcriptional events and other cellular processes. Moreover, CARM1 is a potential oncological target due to its multiple roles in transcription activation by nuclear hormone receptors and other transcription factors such as p53. Here, we present crystal structures of the CARM1 catalytic domain in complex with cofactors [SAH (S-adenosyl-L-homocysteine) or SNF (sinefungin)] and indole or pyazole inhibitors. Analysis of the structures reveals that the inhibitors bind in the arginine-binding cavity and the surrounding pocket that exists at the interface between the N- and C-terminal domains. In addition, we show using ITC (isothermal titration calorimetry) that the inhibitors bind to the CARM1 catalytic domain only in the presence of the cofactor SAH. Furthermore, sequence differences for select residues that interact with the inhibitors may be responsible for the CARM1 selectivity against PRMT1 and PRMT3. Together, the structural and biophysical information should aid in the design of both potent and specific inhibitors of CARM1.
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http://dx.doi.org/10.1042/BJ20102161DOI Listing
June 2011

Thieno[3,2-c]pyrazoles: a novel class of Aurora inhibitors with favorable antitumor activity.

Bioorg Med Chem 2010 Oct 25;18(19):7113-20. Epub 2010 Jul 25.

Nerviano Medical Sciences-Oncology, via Pasteur 10, 20014 Nerviano, Milan, Italy.

A novel series of 3-amino-1H-thieno[3,2-c]pyrazole derivatives demonstrating high potency in inhibiting Aurora kinases was developed. Here we describe the synthesis and a preliminary structure-activity relationship, which led to the discovery of a representative compound (38), which showed low nanomolar inhibitory activity in the anti-proliferation assay and was able to block the cell cycle in HCT-116 cell line. This compound demonstrated favorable pharmacokinetic properties and good efficacy in the HL-60 xenograft tumor model.
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http://dx.doi.org/10.1016/j.bmc.2010.07.048DOI Listing
October 2010

Crystal structures of anaplastic lymphoma kinase in complex with ATP competitive inhibitors.

Biochemistry 2010 Aug;49(32):6813-25

Nerviano Medical Sciences S.r.l., Viale Pasteur 10, 20014 Nerviano (MI), Italy.

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase involved in the development of several human cancers and, as a result, is a recognized target for the development of small-molecule inhibitors for the treatment of ALK-positive malignancies. Here, we present the crystal structures of the unphosphorylated human ALK kinase domain in complex with the ATP competitive ligands PHA-E429 and NVP-TAE684. Analysis of these structures provides valuable information concerning the specific characteristics of the ALK active site as well as giving indications about how to obtain selective ALK inhibitors. In addition, the ALK-KD-PHA-E429 structure led to the identification of a potential regulatory mechanism involving a link made between a short helical segment immediately following the DFG motif and an N-terminal two-stranded beta-sheet. Finally, mapping of the activating mutations associated with neuroblastoma onto our structures may explain the roles these residues have in the activation process.
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http://dx.doi.org/10.1021/bi1005514DOI Listing
August 2010

First Cdc7 kinase inhibitors: pyrrolopyridinones as potent and orally active antitumor agents. 2. Lead discovery.

J Med Chem 2009 Jan;52(2):293-307

Nerviano Medical Sciences Srl, Viale Pasteur 10, 20014 Nerviano, Milano, Italy.

Cdc7 kinase is a key regulator of the S-phase of the cell cycle, known to promote the activation of DNA replication origins in eukaryotic organisms. Cdc7 inhibition can cause tumor-cell death in a p53-independent manner, supporting the rationale for developing Cdc7 inhibitors for the treatment of cancer. In this paper, we conclude the structure-activity relationships study of the 2-heteroaryl-pyrrolopyridinone class of compounds that display potent inhibitory activity against Cdc7 kinase. Furthermore, we also describe the discovery of 89S, [(S)-2-(2-aminopyrimidin-4-yl)-7-(2-fluoro-ethyl)-1,5,6,7-tetrahydropyrrolo[3,2-c]pyridin-4-one], as a potent ATP mimetic inhibitor of Cdc7. Compound 89S has a Ki value of 0.5 nM, inhibits cell proliferation of different tumor cell lines with an IC50 in the submicromolar range, and exhibits in vivo tumor growth inhibition of 68% in the A2780 xenograft model.
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http://dx.doi.org/10.1021/jm800977qDOI Listing
January 2009