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Crystallography, biophysical and cheminformatics studies for next-generation kinase
inhibitor design
R. A. Engh, Dept of Chemistry, UiT
Background: The human genome contains over 500 homologous protein kinases, which control
cellular signalling processes. They have become one of the most important target classes for the
design of therapeutic inhibitors; many diseases are caused by dysregulation of cellular signalling
processes. Most therapeutic inhibitors approved to now are for cancer therapies, but this focus is
now broadening. Emerging targets of interest are relevant e.g. for neurodegenerative diseases,
inflammation, and other areas. Further protein kinase targets are of non-human origin, with impacts
for antibacterial, antifungal, and antiparasitic therapies.
Research into protein kinase inhibitor drugs has been progressing for the past 20-30 years with
many high-throughput tools, including synthesis, screening, and structural studies. As a result, an
enormous body of relevant data exists, including three dimensional structures, binding strengths and
selectivity determinants. Further, many and growing numbers of highly active kinase inhibitors are
now coming off patent for the first time, including the first approved protein kinase drugs. This
creates a drug discovery environment with unparalleled richness. Protein kinase selectivity
determinants are known that can retarget such substances to similar but distinct kinases with
relatively minor changes. Due to the complexities of drug-target interactions, and especially in
connection with the natural flexibility of protein kinases, inhibitor binding properties can be
predicted with only limited success. Taken together, these facts highlight the future of protein kinase
inhibitor drug discovery: 1) increased interest in new targets, 2) cheminformatics driven and rapid
refunctionalization of known binders to these targets, and 3) ongoing discovery of key selectivity
determinants.
Project goals and activities
• (Application writing phase) Evaluation of up-to-date inhibitor and target data to select a
protein kinase target area of interest.
• Selection of specific constructs for protein production, based on strategies for successful
crystallography, variations to identify key binding interactions, and possible related target
interactions (to optimize selectivity profiles, avoid toxicities, and forestall drug resistance)
• Protein production, crystallography and biophysical binding studies, studying mechanisms
of ligand binding strengths and kinetics.
• Cheminformatics studies to identify the most promising inhibitor design approaches;
modelling and possible synthesis of new compounds
The research environment
• The project is based at the Norwegian Center for Structural Biology (NORSTRUCT),
Department of Chemistry, Tromsø, Norway
• NOSTRUCT facilities include state-of-the-art recombinant protein production facilities
allowing both prokaryotic and eukaryotic expression with scalable chromatographic
purification, and also recently upgraded (2017) systems for automated liquid handling,
crystallization, X-ray data collection, and biophysical ligand binding studies (SPR, ITC,
MST)
• The Department of Chemistry hosts and encourages collaboration with the Center for
Computational and Theoretical Chemistry (Norwegian Center of Excellence) and the
Norwegian node for the pan-European research infrastructure Elixir
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The strategic research focus of the Department of Chemistry prioritizes therapeutic
target/ligand interaction studies with a Drug Discovery and Design technology and
networking platform, with ongoing projects in antibiotic, anticancer, and other drug
discovery areas.
Tromsø is a center for marine bioprospecting studies, with potential impact for this projects
The project supervisor (https://www.linkedin.com/in/richard-a-engh-77949a88) has
extensive experience and a wide ranging publication record beginning with very early
protein kinase crystallography efforts (see publications below)
Tromsø is a vibrant and unique city north of the Polar Circle with a large international
community, often cited in "top ten" lists of places to experience (northern lights, midnight
sun, whale watching, unspoiled nature).
Selected relevant publications (see also http://www.researchgate.net/profile/Richard_Engh/)
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Probing the ATP-binding pocket of protein kinase DYRK1A with benzothiazole fragment molecules; U. Rothweiler, W. Stensen, B.O.
Brandsdal, J. Isaksson, F.A. Leeson, R.A. Engh, J.S.M. Svendsen (2016) J. Med.Chem, in press.
Addresssing the glycine-rich loop of protein kinases by a multi-dimensional interaction network: inhibition of PKA and a PKB mimic,
B.S. Lauber, L.A. Hardegger, A.K. Asraful, B. A. Lund, O.C. Dumele, M. Harder, B. Kuhn, R.A. Engh, F. Diederich (2015) Chemistry
doi: 10.1002/chem.201503552.
Assessing protein kinase target similarity: examples comparing sequence, structure, and cheminformatics approaches, O.A. Gani, B.
Thakkar, D. Narayanan, K.A. Alam, P. Kyomuhendo, U. Rothweiler, V. Tello-Franco, R.A. Engh, (2015) BBA - Proteins and
Proteomics, 1854, 1605-1616.
The crystal structure of a DYRK1A PKC412 complex reveals disulphide bridge formation with the anomalous catalytic loop HRD(HCD)
cysteine, M. Alexeeva, E. Åberg, R.A. Engh, U. Rothweiler, (2015) Acta Cryst D. 71, 1207-1215.
Luciferin and derivatives as a DYRK selective scaffold for the design of protein kinase inhibitors, U. Rothweiler, J. Eriksson, W.
Stensen, F. Leeson, R.A. Engh, J.S. Svendsen, (2015), Eur. J. Med. Chem. 2015 Feb 25; 94:140-148. doi:
10.1016/j.ejmech.2015.02.035.
Evaluating the predictivity of virtual screening for Abl kinase inhibitors to hinder drug resistance, O. Gani, D. Narayanan, R.A. Engh,
Chemical Biology and Drug Design (2013), 82, 506-519.
Structural origins of AGC protein kinase inhibitor selectivities: PKA as a drug discovery tool. E. Åberg, B. Lund, A. Pflug, O.A.B.S.M.
Gani, U. Rothweiler, T.M. de Oliveira, R.A. Engh, Biol. Chem. Hoppe Seyler, (2012), 393, 1121-1129.
Anomalous dispersion analysis of inhibitor flexibility-- A case study of the kinase inhibitor H-89. A. Pflug, K.A. Johnson, R.A. Engh,
Acta Cryst. F. (2012), 68, 873-7.
Differential sensitivity of ErbBB2 kinase domain mutations towards lapatinib, R. K. Kancha, N. von Bubnoff, N. Bartosch, C. Peschel,
R. A. Engh, J. Duyster, PLoS One (2011), 6, e26762, doi: 10.1371/journal.pone.0026760.
Dynamics of the emergence of dasatinib and nilotinib resistance in imatinib resistant CML patients, F.X. Gruber, T. Ernst, K. Porkka,
R.A. Engh, I. Mikkola, J. Maier, T. Lange, A. Hochhaus, Leukemia (2011), doi: 10.1038/leu.2011.187. [Epub ahead of print].
Mutants of protein kinase A that mimic the ATP-binding site of Aurora kinase, A. Pflug, T. M. de Oliveira, D. Bossemeyer, R.A. Engh,
Biochem. J. (2011), 440, 85-93.
p38 MAP kinase dimers with swapped activation segments and a novel catalytic loop conformation, U. Rothweiler, E. Åberg, K.A.
Johnson, T.E. Hansen, J.B. Jørgensen, R.A. Engh, J. Mol. Biol. (2011), 411, 474-85. Epub DOI: 10.1016/j.jmb.2011.06.013.
VX680 Binding in Aurora A: π-π Interactions Involving the Conserved Aromatic Amino Acid of the Flexible Glycine-Rich Loop, T.M.
Oliveira, R. Ahmad, R. A. Engh, J. Phys. Chem. (2011), 54, 312-319. Epub 2010 Dec 3.
Phthalazinone pyrazoles as potent, selective and orally bioavailable inhibitors of Aurora-A kinase, M.E. Prime, S.M. Courtney, F.A.
Brookfield, R.W. Marston, V. Walker, J. Warne, A. E. Boyd, N.A. Kairies, W. von der Saal, A. Limberg, G. Georges, R.A. Engh, B.
Goller, P. Rueger & M. Rueth, J. Med. Chem. (2011), Epub DOI: 10.1021/jm101346r.
Diversity of bisubstrate binding modes of adenosine analogue-oligoarginine conjugates in protein kinase A and implications for protein
substrate interactions, A. Pflug, J. Rogozina, D. Lavogina, E. Enkvist, A. Uri, R.A. Engh, D. Bossemeyer, J. Mol. Biol. (2010), 403, 6677.
Protein kinase inhibition of clinically important staurosporine analogues, O.A. Gani & R.A.Engh, Nat. Prod. Rep. (2010), 27, 489-498.
Crystallography for protein kinase drug design: PKA and SRC case studies; C.B.Breitenlechner, D.Bossemeyer & R.A.Engh; Biochim.
Biophys. Acta, Prot. & Proteom. (2005), 1754, 38-49.
Protein kinase A in complex with Rho-kinase inhibitors Y-27632, Fasudil (HA-1077) and H-1152P: Structural basis of selectivity;
C.B.Breitenlechner, M.Gaßel, H.Hidaka, V.Kinzel, R.Huber, R.A.Engh, & D.Bossemeyer; Structure (2003) 11; 1595-1607.
Structural aspects of protein kinase control—Role of conformational flexibility; R.A.Engh & D. Bossemeyer; Pharmacology and
Therapeutics (2002) 93; 99-111.
Phosphorylation and flexibility of cyclic-AMP dependent protein kinase (PKA) using P-31 NMR Spectroscopy; M.H.J.Seifert,
C.B.Breitenlechner, D.Bossemeyer, R.Huber, T.A.Holak & R.A.Engh; Biochemistry (2002) 41; 5968-5977.
Crystal Structures of Catalytic Subunit of cAMP-dependent Protein Kinase in Complex with Isoquinolinesulfonyl Protein Kinase
Inhibitors H-7, H-8, H-89; R.A.Engh, A.Girod, V.Kinzel, R.Huber & D. Bossemeyer; J. Biol. Chem. (1996) 271; 26157-26164.
Phosphotransferase and substrate binding mechanism of the cAMP-dependent protein kinase catalytic subunit from porcine heart as
deduced from the 2.0 Å structure of the complex with manganese(2+) adenylyl imidodiphosphate and inhibitor peptide PKI(5-24);
D.Bossemeyer, R.A.Engh, V.Kinzel, H.Ponstingl & R.Huber; EMBO Journal (1993) 12; 849-59.