Abstract
The phosphatidylinositol 3-kinase (PI3K) pathway is involved in many cellular functions including cell growth, metabolism, and transformation. Hyperactivation of this pathway contributes to tumorigenesis, therefore, PI3K is a major target for anticancer drug discovery. Since the PI3Kα isoform is implicated mostly in cancer, we conducted a high-throughput screening (HTS) campaign using a 3-step PI3K homogenous time-resolved fluorescence assay against this isoform bearing the H1047R mutation. A total of 288,000 synthetic and natural product-derived compounds were screened and of which, we identified 124 initial hits that were further selected by considering the predicted binding mode, relationship to known pan-assay interference compounds and previous descriptions as a lipid kinase inhibitor. A total of 24 compounds were then tested for concentration-dependent responses. These hit compounds provide novel scaffolds that can potentially be optimized to create novel PI3K inhibitors.
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References
Williams R, Berndt A, Miller S, Hon WC, Zhang X. Form and flexibility in phosphoinositide 3-kinases. Biochem Soc Trans. 2009;37:615–26.
Asati V, Mahapatra DK, Bharti SK. PI3K/Akt/mTOR and Ras/Raf/MEK/ERK signaling pathways inhibitors as anticancer agents: Structural and pharmacological perspectives. Eur J Med Chem. 2016;109:314–41.
Katso R, Okkenhaug K, Ahmadi K, White S, Timms J, Waterfield MD. Cellular function of phosphoinositide 3-kinases: implications for development, homeostasis, and cancer. Annu Rev Cell Dev Biol. 2001;17:615–75.
Fruman DA, Meyers RE, Cantley LC. Phosphoinositide kinases. Annu Rev Biochem. 1998;67:481–507.
Engelman JA, Luo J, Cantley LC. The evolution of phosphatidylinositol 3-kinases as regulators of growth and metabolism. Nat Rev Genet. 2006;7:606–19.
Vanhaesebroeck B, Leevers SJ, Panayotou G, Waterfield MD. Phosphoinositide 3-kinases: a conserved family of signal transducers. Trends Biochem Sci. 1997;22:267–72.
Curran E, Smith SM. Phosphoinositide 3-kinase inhibitors in lymphoma. Curr Opin Oncol. 2014;26:469–75.
Flanagan JU, Shepherd PR. Structure, function and inhibition of the phosphoinositide 3-kinase p110alpha enzyme. Biochem Soc Trans. 2014;42:120–4.
Yuan TL, Cantley LC. PI3K pathway alterations in cancer: variations on a theme. Oncogene. 2008;27:5497–510.
Huang CH, Mandelker D, Schmidt-Kittler O, Samuels Y, Velculescu VE, Kinzler KW, et al. The structure of a human p110alpha/p85alpha complex elucidates the effects of oncogenic PI3Kalpha mutations. Science. 2007;318:1744–8.
Vanhaesebroeck B, Guillermet-Guibert J, Graupera M, Bilanges B. The emerging mechanisms of isoform-specific PI3K signalling. Nat Rev Mol Cell Biol. 2010;11:329–41.
Samuels Y, Wang Z, Bardelli A, Silliman N, Ptak J, Szabo S, et al. High frequency of mutations of the PIK3CA gene in human cancers. Science. 2004;304:554.
Burke JE, Perisic O, Masson GR, Vadas O, Williams RL. Oncogenic mutations mimic and enhance dynamic events in the natural activation of phosphoinositide 3-kinase p110alpha (PIK3CA). Proc Natl Acad Sci USA. 2012;109:15259–64.
Zhao L, Vogt PK. Helical domain and kinase domain mutations in p110alpha of phosphatidylinositol 3-kinase induce gain of function by different mechanisms. Proc Natl Acad Sci USA. 2008;105:2652–7.
Hon WC, Berndt A, Williams RL. Regulation of lipid binding underlies the activation mechanism of class IA PI3-kinases. Oncogene. 2012;31:3655–66.
Mandelker D, Gabelli SB, Schmidt-Kittler O, Zhu J, Cheong I, Huang CH, et al. A frequent kinase domain mutation that changes the interaction between PI3Kalpha and the membrane. Proc Natl Acad Sci USA. 2009;106:16996–7001.
Chaussade C, Cho K, Mawson C, Rewcastle G, Shepherd P. Functional differences between two classes of oncogenic mutation in the PIK3CA gene. Biochem Biophys Res Commun. 2009;381:577–81.
Chaussade C, Rewcastle GW, Kendall JD, Denny WA, Cho K, Gronning LM, et al. Evidence for functional redundancy of class IA PI3K isoforms in insulin signalling. Biochem J. 2007;404:449–58.
Costa C, Ebi H, Martini M, Beausoleil SA, Faber AC, Jakubik CT, et al. Measurement of PIP3 levels reveals an unexpected role for p110beta in early adaptive responses to p110alpha-specific inhibitors in luminal breast cancer. Cancer Cell. 2015;27:97–108.
Elkabets M, Vora S, Juric D, Morse N, Mino-Kenudson M, Muranen T, et al. mTORC1 inhibition is required for sensitivity to PI3K p110alpha inhibitors in PIK3CA-mutant breast cancer. Sci Transl Med. 2013;5:196ra99.
Jamieson S, Flanagan JU, Kolekar S, Buchanan C, Kendall JD, Lee WJ, et al. A drug targeting only p110alpha can block phosphoinositide 3-kinase signalling and tumour growth in certain cell types. Biochem J. 2011;438:53–62.
Knight SD, Adams ND, Burgess JL, Chaudhari AM, Darcy MG, Donatelli CA, et al. Discovery of GSK2126458, a highly potent inhibitor of PI3K and the mammalian target of rapamycin. ACS Med Chem Lett. 2010;1:39–43.
Furet P, Guagnano V, Fairhurst RA, Imbach-Weese P, Bruce I, Knapp M, et al. Discovery of NVP-BYL719 a potent and selective phosphatidylinositol-3 kinase alpha inhibitor selected for clinical evaluation. Bioorg Med Chem Lett. 2013;23:3741–8.
Gong GQ, Kendall JD, Dickson JMJ, Rewcastle GW, Buchanan CM, Denny WA, et al. Combining properties of different classes of PI3Kalpha inhibitors to understand the molecular features that confer selectivity. Biochem J. 2017;474:2261–76.
Zheng Z, Amran SI, Zhu J, Schmidt-Kittler O, Kinzler KW, Vogelstein B, et al. Definition of the binding mode of a new class of phosphoinositide 3-kinase alpha-selective inhibitors using in vitro mutagenesis of non-conserved amino acids and kinetic analysis. Biochem J. 2012;444:529–35.
Fairhurst RA, Gerspacher M, Imbach-Weese P, Mah R, Caravatti G, Furet P, et al. Identification and optimisation of 4,5-dihydrobenzo[1,2-d:3,4-d]bisthiazole and 4,5-dihydrothiazolo[4,5-h]quinazoline series of selective phosphatidylinositol-3 kinase alpha inhibitors. Bioorg Med Chem Lett. 2015;25:3575–81.
Fairhurst RA, Imbach-Weese P, Gerspacher M, Caravatti G, Furet P, Zoller T, et al. Identification and optimisation of a 4′,5-bisthiazole series of selective phosphatidylinositol-3 kinase alpha inhibitors. Bioorg Med Chem Lett. 2015;25:3569–74.
Gerspacher M, Fairhurst RA, Mah R, Roehn-Carnemolla E, Furet P, Fritsch C, et al. Discovery of a novel tricyclic 4H-thiazolo[5',4':4,5]pyrano[2,3-c]pyridine-2-amino scaffold and its application in a PI3Kα inhibitor with high PI3K isoform selectivity and potent cellular activity. Bioorg Med Chem Lett. 2015;25:3582–4.
Hayakawa M, Kawaguchi K, Kaizawa H, Koizumi T, Ohishi T, Yamano M, et al. Synthesis and biological evaluation of sulfonylhydrazone-substituted imidazo[1,2-a]pyridines as novel PI3 kinase p110alpha inhibitors. Bioorg Med Chem. 2007;15:5837–44.
Kendall JD, Giddens AC, Tsang KY, Frederick R, Marshall ES, Singh R, et al. Novel pyrazolo[1,5-a]pyridines as p110alpha-selective PI3 kinase inhibitors: exploring the benzenesulfonohydrazide SAR. Bioorg Med Chem. 2012;20:58–68.
Bruce I, Akhlaq M, Bloomfield GC, Budd E, Cox B, Cuenoud B, et al. Development of isoform selective PI3-kinase inhibitors as pharmacological tools for elucidating the PI3K pathway. Bioorg Med Chem Lett. 2012;22:5445–50.
Heffron TP, Heald RA, Ndubaku C, Wei B, Augistin M, Do S, et al. The Rational design of selective benzoxazepin inhibitors of the alpha-Isoform of phosphoinositide 3-kinase culminating in the identification of (S)-2-((2-(1-isopropyl-1H-1,2,4-triazol-5-yl)-5,6-dihydrobenzo[f]imidazo[1,2-d][1,4]oxazepin-9-yl)oxy)propanamide (GDC-0326). J Med Chem. 2016;59:985–1002.
Nacht M, Qiao L, Sheets MP, St Martin T, Labenski M, Mazdiyasni H, et al. Discovery of a potent and isoform-selective targeted covalent inhibitor of the lipid kinase PI3Kalpha. J Med Chem. 2013;56:712–21.
Janku F, Tsimberidou AM, Garrido-Laguna I, Wang X, Luthra R, Hong DS, et al. PIK3CA mutations in patients with advanced cancers treated with PI3K/AKT/mTOR axis inhibitors. Mol Cancer Ther. 2011;10:558–65.
Luo J, Sobkiw CL, Hirshman MF, Logsdon MN, Li TQ, Goodyear LJ, et al. Loss of class IA PI3K signaling in muscle leads to impaired muscle growth, insulin response, and hyperlipidemia. Cell Metab. 2006;3:355–66.
Taniguchi CM, Kondo T, Sajan M, Luo J, Bronson R, Asano T, et al. Divergent regulation of hepatic glucose and lipid metabolism by phosphoinositide 3-kinase via Akt and PKCλ/ζ. Cell Metab. 2006;3:343–53.
Irwin JJ, Shoichet BK. Docking screens for novel ligands conferring new biology. J Med Chem. 2016;59:4103–20.
Kitchen DB, Decornez H, Furr JR, Bajorath J. Docking and scoring in virtual screening for drug discovery: methods and applications. Nat Rev Drug Discov. 2004;3:935–49.
Verdonk ML, Giangreco I, Hall RJ, Korb O, Mortenson PN, Murray CW. Docking performance of fragments and druglike compounds. J Med Chem. 2011;54:5422–31.
Dickson JM, Lee WJ, Shepherd PR, Buchanan CM. Enzyme activity effects of N-terminal His-tag attached to catalytic sub-unit of phosphoinositide-3-kinase. Biosci Rep. 2013;33:e00079.
Jones G, Willett P, Glen R, Leach A, Taylor R. Development and validation of a genetic algorithm for flexible docking. J Mol Biol. 1997;267:727–48.
Kendall JD, O'Connor PD, Marshall AJ, Frederick R, Marshall ES, Lill CL, et al. Discovery of pyrazolo[1,5-a]pyridines as p110alpha-selective PI3 kinase inhibitors. Bioorg Med Chem. 2012;20:69–85.
Gkeka P, Papafotika A, Christoforidis S, Cournia Z. Exploring a non-ATP pocket for potential allosteric modulation of PI3Kalpha. J Phys Chem B. 2015;119:1002–16.
Baell JB, Holloway GA. New substructure filters for removal of pan assay interference compounds (PAINS) from screening libraries and for their exclusion in bioassays. J Med Chem. 2010;53:2719–40.
Giordanetto F, Kull B, Dellsén A. Discovery of novel class 1 phosphatidylinositide 3-kinases (PI3K) fragment inhibitors through structure-based virtual screening. Bioorg Med Chem Lett. 2011;21:829–35.
Yang L, Li G, Ma S, Zou C, Zhou S, Sun Q, et al. Structure–activity relationship studies of pyrazolo[3,4-d]pyrimidine derivatives leading to the discovery of a novel multikinase inhibitor that potently inhibits FLT3 and VEGFR2 and evaluation of its activity against acute myeloid leukemia in vitro and in vivo. J Med Chem. 2013;56:1641–55.
Acknowledgements
We are indebted to Qiang Shen and Jie-jie Deng for technical assistance. This work was partially supported by grants from the Ministry of Science and Technology of China (to MWW: 2014DFG32200), the Shanghai Science and Technology Development Fund (to MWW: 15DZ2291600), the Thousand Talents Program in China (to MWW), and the New Zealand Health Research Council (to PRS HRC 13/1019). The funders had no role in study design, data collection, and analysis; decision to publish; or manuscript preparation.
Author contribution
PRS and MWW designed research. JW, GQG, YZ, WJL, CMB, JMJD, and JUF performed research. JW, GQG, YZ, WAD, GWR, JDK, JUF, DHY, PRS, and MWW analyzed data. GQG, CMB, JUF, and MWW wrote the paper.
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Wang, J., Gong, G.Q., Zhou, Y. et al. High-throughput screening campaigns against a PI3Kα isoform bearing the H1047R mutation identified potential inhibitors with novel scaffolds. Acta Pharmacol Sin 39, 1816–1822 (2018). https://doi.org/10.1038/s41401-018-0057-z
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DOI: https://doi.org/10.1038/s41401-018-0057-z
