Abstract
The MAPK p38 became a focal point of inflammatory research when it was recognized that it played a key role in the production of the pro-inflammatory molecules TNF-alpha, IL-beta, and cyclooxygenase-2 (COX-2). The pharmaceutical industry devoted enormous efforts to creating p38 inhibitors, because blocking p38 had the potential of downregulating a group of pro-inflammatory mediators, and thus, one drug could have a cocktail effect. The market potential seemed to be clearly established (Bonafede et al., Clinicoecon Outcomes Res 6:381–388, 2014) with a multiplicity of TNF-alpha antibodies and a soluble receptor (Mewar and Wilson, Br J Pharmacol 162:785–791, 2011) already on the market, although the relationship between TNF-alpha production and p38 activation is a complicated two-way (Sabio and Davis, Semin Immunol 26:237–245, 2014) signal transduction process. With the discovery that activated p38 stabilizes (Mancini and Di Battista, Inflamm Res 60:1083–1092, 2011) COX-2 mRNA and upregulates expression of IL-beta (Bachstetter and Van Eldik, Aging Dis 1:199–211, 2010) probably in a similar manner, inhibiting p38 appeared to be a way of blocking TNF-alpha, COX-2, and IL-beta simultaneously. At Locus Pharmaceuticals we jumped on this opportunity, because we believed that our fragment-based drug discovery approach was ideally suited for making a potent small molecule p38 inhibitor that did not bind in the ATP site, but also had the solubility, lack of planarity, and low molecular weight required of a clinical candidate. Just to be clear, in our experience highly planar compounds often result in poor pharmacokinetics, because they tend to bind strongly to plasma proteins. At Locus we typically repeated assays by adding increasing amounts of plasma to check for potency degradation in the presence of blood. We found this to be a useful early indicator of pharmacokinetics and in vivo behavior. It became clear from our work and the work of others that binding to the ATP site resulted in nonspecific isoform toxicities, but binding in the adjacent allosteric DFG-site resulted in molecules that were too planar and too hydrophobic. Applying the computational method of Simulated Annealing of Chemical Potential (SACP) to this problem, we at Locus were able to come up with surprising fragment substitution patterns that led to potent non-ATP p38 inhibitors with the solubility and lack of planarity that resulted in potent in vivo efficacy in rodents with 33 % oral bioavailability. By using the simulations, we made only a small number of molecules and created a high quality clinical candidate. We also did extensive co-crystallography work, which demonstrated that the compounds bound in the mode predicted by the simulations. Unfortunately, all p38 programs ultimately shut down, because compelling evidence emerged that inhibiting p38 had no long-term clinical (Genovese, Arthritis Rheum 60:317–320, 2009) benefit. Devoting a large amount of limited resources to a target that ultimately turns out to be a mistake because it was not properly validated is a fatal error for a small company, and this is one of the reasons that Locus ultimately failed.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Bonafede M, Joseph GJ, Shah N, Princic N, Harrison DJ (2014) Cost of tumor necrosis factor blockers per patient with rheumatoid arthritis in a multistate Medicaid population. Clinicoecon Outcomes Res 6:381–388
Mewar D, Wilson AG (2011) Treatment of rheumatoid arthritis with tumour necrosis factor inhibitors. Br J Pharmacol 162:785–791
Sabio G, Davis RJ (2014) TNF and MAP kinase signalling pathways. Semin Immunol 26:237–245
Mancini AD, Di Battista JA (2011) The cardinal role of the phospholipase A(2)/cyclooxygenase-2/prostaglandin E synthase/prostaglandin E(2) (PCPP) axis in inflammostasis. Inflamm Res 60:1083–1092
Bachstetter AD, Van Eldik LJ (2010) The p38 MAP kinase family as regulators of proinflammatory cytokine production in degenerative diseases of the CNS. Aging Dis 1:199–211
Genovese MC (2009) Inhibition of p38: has the fat lady sung? Arthritis Rheum 60:317–320
Han J, Lee JD, Bibbs L, Ulevitch RJ (1994) A MAP kinase targeted by endotoxin and hyperosmolarity in mammalian cells. Science 265:808–811
Lee JC, Laydon JT, McDonnell PC, Gallagher TF, Kumar S, Green D, McNulty D, Blumenthal MJ, Heys JR, Landvatter SW et al (1994) A protein kinase involved in the regulation of inflammatory cytokine biosynthesis. Nature 372:739–746
Dumas J, Hatoum-Mokdad H, Sibley R, Riedl B, Scott WJ, Monahan MK, Lowinger TB, Brennan C, Natero R, Turner T, Johnson JS, Schoenleber R, Bhargava A, Wilhelm SM, Housley TJ, Ranges GE, Shrikhande A (2000) 1-Phenyl-5-pyrazolyl ureas: potent and selective p38 kinase inhibitors. Bioorg Med Chem Lett 10:2051–2054
Regan J, Pargellis CA, Cirillo PF, Gilmore T, Hickey ER, Peet GW, Proto A, Swinamer A, Moss N (2003) The kinetics of binding to p38MAP kinase by analogues of BIRB 796. Bioorg Med Chem Lett 13:3101–3104
Pargellis C, Tong L, Churchill L, Cirillo PF, Gilmore T, Graham AG, Grob PM, Hickey ER, Moss N, Pav S, Regan J (2002) Inhibition of p38 MAP kinase by utilizing a novel allosteric binding site. Nat Struct Biol 9:268–272
Long DL, Loeser RF (2010) p38gamma mitogen-activated protein kinase suppresses chondrocyte production of MMP-13 in response to catabolic stimulation. Osteoarthritis Cartilage 18:1203–1210
Pogozelski AR, Geng T, Li P, Yin X, Lira VA, Zhang M, Chi JT, Yan Z (2009) p38gamma mitogen-activated protein kinase is a key regulator in skeletal muscle metabolic adaptation in mice. PLoS One 4:e7934
Bradbury MW, Stump D, Guarnieri F, Berk PD (2011) Molecular modeling and functional confirmation of a predicted fatty acid binding site of mitochondrial aspartate aminotransferase. J Mol Biol 412:412–422
Guarnieri F (2004) Computational protein probing to identify binding sites. US Patent No. 6,735,530. Issued 11 May 2004
Clark M, Guarnieri F, Shkurko I, Wiseman J (2006) Grand canonical Monte Carlo simulation of ligand-protein binding. J Chem Inf Model 46:231–242
Moore WR Jr (2005) Maximizing discovery efficiency with a computationally driven fragment approach. Curr Opin Drug Discov Devel 8:355–364
Kulp JL III, Blumenthal SN, Wang Q, Bryan RL, Guarnieri F (2012) A fragment-based approach to the SAMPL3 challenge. J Comput Aided Mol Des 26:583–594
Kulp JL III, Kulp JL Jr, Pompliano DL, Guarnieri F (2011) Diverse fragment clustering and water exclusion identify protein hot spots. J Am Chem Soc 133:10740–10743
Guarnieri F, Mezei M (1996) Simulated annealing of chemical potential: a general procedure for locating bound waters. Application to the study of the differential hydration propensities of the major and minor grooves of DNA. J Am Chem Soc 118:8493–8494
Michelotti EL, Moffett KK, Nguyen D, Kelly MJ, Shetty R, Chai X, Northrop K, Namboodiri V, Campbell B, Flynn GA, Fujimoto T, Hollinger FP, Bukhtiyarova M, Springman EB, Karpusas M (2005) Two classes of p38alpha MAP kinase inhibitors having a common diphenylether core but exhibiting divergent binding modes. Bioorg Med Chem Lett 15:5274–5279
Qin Z (2012) The use of THP-1 cells as a model for mimicking the function and regulation of monocytes and macrophages in the vasculature. Atherosclerosis 221:2–11
Oprea TI (2002) Current trends in lead discovery: are we looking for the appropriate properties? Mol Divers 5:199–208
Moffett K, Konteatis Z, Nguyen D, Shetty R, Ludington J, Fujimoto T, Lee KJ, Chai X, Namboodiri H, Karpusas M, Dorsey B, Guarnieri F, Bukhtiyarova M, Springman E, Michelotti E (2011) Discovery of a novel class of non-ATP site DFG-out state p38 inhibitors utilizing computationally assisted virtual fragment-based drug design (vFBDD). Bioorg Med Chem Lett 21:7155–7165
Zhou JS, Xing W, Friend DS, Austen KF, Katz HR (2007) Mast cell deficiency in Kit(W-sh) mice does not impair antibody-mediated arthritis. J Exp Med 204:2797–2802
Cohen SB, Cheng TT, Chindalore V, Damjanov N, Burgos-Vargas R, Delora P, Zimany K, Travers H, Caulfield JP (2009) Evaluation of the efficacy and safety of pamapimod, a p38 MAP kinase inhibitor, in a double-blind, methotrexate-controlled study of patients with active rheumatoid arthritis. Arthritis Rheum 60:335–344
Stroes E, Colquhoun D, Sullivan D, Civeira F, Rosenson RS, Watts GF, Bruckert E, Cho L, Dent R, Knusel B, Xue A, Scott R, Wasserman SM, Rocco M (2014) Anti-PCSK9 antibody effectively lowers cholesterol in patients with statin intolerance: the GAUSS-2 randomized, placebo-controlled phase 3 clinical trial of evolocumab. J Am Coll Cardiol 63:2541–2548
Robinson JG, Nedergaard BS, Rogers WJ, Fialkow J, Neutel JM, Ramstad D, Somaratne R, Legg JC, Nelson P, Scott R, Wasserman SM, Weiss R (2014) Effect of evolocumab or ezetimibe added to moderate- or high-intensity statin therapy on LDL-C lowering in patients with hypercholesterolemia: the LAPLACE-2 randomized clinical trial. JAMA 311:1870–1882
Koren MJ, Lundqvist P, Bolognese M, Neutel JM, Monsalvo ML, Yang J, Kim JB, Scott R, Wasserman SM, Bays H (2014) Anti-PCSK9 Monotherapy for Hypercholesterolemia: The MENDEL-2 Randomized, Controlled Phase III Clinical Trial of Evolocumab. J Am Coll Cardiol 63:2531–2540
Blom DJ, Hala T, Bolognese M, Lillestol MJ, Toth PD, Burgess L, Ceska R, Roth E, Koren MJ, Ballantyne CM, Monsalvo ML, Tsirtsonis K, Kim JB, Scott R, Wasserman SM, Stein EA (2014) A 52-week placebo-controlled trial of evolocumab in hyperlipidemia. N Engl J Med 370:1809–1819
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Guarnieri, F. (2015). Designing an Orally Available Nontoxic p38 Inhibitor with a Fragment-Based Strategy. In: Klon, A. (eds) Fragment-Based Methods in Drug Discovery. Methods in Molecular Biology, vol 1289. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2486-8_15
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2486-8_15
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2485-1
Online ISBN: 978-1-4939-2486-8
eBook Packages: Springer Protocols