USOO9345245B2

(12) United States Patent (10) Patent No.: US 9,345,245 B2 Cutler et al. (45) Date of Patent: May 24, 2016

(54) SYNTHETIC COMPOUNDS FOR Cutler et al., “Abscisic Acid. Emergence of a Core Signaling Net VEGETATIVE ABA RESPONSES work”. Annual Review of Plant Biology, vol. 61, pp. 651-679 (2010). Iyer et al., "Adaptations of the helix-grip fold for ligand binding and (71) Applicant: THE REGENTS OF THE catalysis in the START domain superfamily”. Protens: Structure, UNIVERSITY OF CALIFORNLA, Function, and Bioinformaticis, vol.43, No. 2, pp. 134-144 (2001). Oakland, CA (US) Melcher et al., “Thirsty plants and beyond: structural mechanisms of abscisic acid perception and signaling'. Current Opinion in Struc (72) Inventors: Sean R. Cutler, Riverside, CA (US); tural Biology, vol. 20, No. 6, pp. 722-729 (2010). Masanori Okamoto, Riverside, CA Notman, “Organic compound comes to the aid of thirsty plants'. (US) Royal Society of Chemistry (May 1, 2009) http://www.rsc.org/ chemistry world/News/2009/May/01050901.asp (downloaded on (73) Assignee: THE REGENTS OF THE Jun. 29, 2015). UNIVERSITY OF CALIFORNLA, Parket al., Abscisic Acid Inhibits Type 2C Protein Phosphatases via Oakland, CA (US) the PYR/PYL Family of START Proteins, Science, vol. 324, No. 5930, pp. 1068-1071 (2009). (*) Notice: Subject to any disclaimer, the term of this Ponting etal. “START: a lipid-binding domain in StAR, HD-ZIP and patent is extended or adjusted under 35 signalling proteins”. Trends Biochem, vol. 24. No. 4, pp. 130-132 U.S.C. 154(b) by 0 days. (1999). Radauer, “The Bet v 1 fold: an ancient, versatile scaffold for binding (21) Appl. No.: 14/385,695 of large, hydrophobic ligands'. BMC Evol. Biol., vol. 8, pp. 286 (2008). (22) PCT Filed: Mar. 15, 2013 Rademacher W. Maisch R, Liessegang J, Jung J (1987) Water con Sumption and yield formation in crop plants under the influence of (86). PCT No.: PCT/US2O13/032281 Synthetic analogues of abscisic acid. In: Hawkins AF, Stead AD, Pinfield NJ (eds)Plantgrowth regulators for agricultural and amenity S371 (c)(1), use. BCPC Monograph 36:53-66. (2) Date: Sep. 16, 2014 Database accession No. 1340941-81-2, Database Registry, Chemical Abstract Service, Columbus, Ohio, US (Nov. 4, 2011). (87) PCT Pub. No.: WO2013/148339 Database accession No. 1331274-04-4. Database Registry, Chemical PCT Pub. Date: Oct. 3, 2013 Abstract Service, Columbus, Ohio, US (Sep. 11, 2011). Database accession No. 1324695-29-5. Database Registry, Chemical (65) Prior Publication Data Abstract Service, Columbus, Ohio, US (Aug. 28, 2011). Database accession No. 121 1832-29-9. Database Registry, Chemical US 2015/OO47O73 A1 Feb. 12, 2015 Abstract Service, Columbus, Ohio, US (Mar. 19, 2010). Yang et al., “Narrowing Down the Targets: Towards Successful Related U.S. Application Data Genetic Engineering of Drought-Tolerant Crops”. Mol. Plant, vol. 3, (60) Provisional application No. 61/618,386, filed on Mar. No. 3, pp. 469-490 (2010). 30, 2012. Wang et al., “Molecular tailoring of farnesylation for plant drought tolerance and yield protection'. The Plant Journal, vol.43, No.3, pp. (51) Int. Cl. 413-424 (2005). AOIN 43/42 (2006.01) Tamura et al., Mega5: Molecular Evolutionary Genetics Analysis C07D 215/227 (2006.01) Using maximum Likelihood, Evolutionary Distance, and Maximum (52) U.S. Cl. Parsimony Methods, Molecular Biology and Evolution, vol. 28, No. CPC ...... A0IN 43/42 (2013.01); C07D 215/227 10, pp. 2731-2739 (2011). (2013.01) (Continued) (58) Field of Classification Search CPC ...... A01N 43/42; C07D 215/227 See application file for complete search history. Primary Examiner — Johann R Richter Assistant Examiner — Andriae M Holt (56) References Cited (74) Attorney, Agent, or Firm — Kilpatrick Townsend & Stockton LLP U.S. PATENT DOCUMENTS 5,498,755 A 3, 1996 Chandraratna et al. (57) ABSTRACT 6,127,382 A 10, 2000 Beard et al. 2011/023O350 A1 9/2011 Frackenpohl et al. 2011/0271408 A1 11/2011 Cutler et al. The present invention provides agonist compounds that active 2013,0045952 A1 2/2013 Xu et al. ABA receptors, and agricultural formulations comprising the agonist compounds. The agricultural formulations are useful FOREIGN PATENT DOCUMENTS for inducing ABA responses in plant vegetative tissues, reducing abiotic stress in plants, and inhibiting germination WO 2010/093954 A2 8, 2010 of plant seeds. The compounds are also useful for inducing WO 2011, 1397.98 A2 11/2011 expression of ABA-responsive genes in cells that express OTHER PUBLICATIONS endogenous or heterologous ABA receptors. Bartel et al., “Analyzing protein-protein interactions using two-hy brid system'. Methods Enzymol. vol. 254, pp. 241-263 (1995). 19 Claims, 27 Drawing Sheets US 9,345,245 B2 Page 2

(56) References Cited Umezawa et al., “Type 2C protein phosphataes directly regulate abscisic acid-activated protein kinases in Arabidopsis'. Proceedings OTHER PUBLICATIONS of the National Academy of Sciences of the USA, vol. 106, No. 41, pp. 17588-17593 (2009). Peterson et al., “Structural basis for selective activation of ABA Nambara et al., “A Screen for genes that function in abscisic acid receptors', Nature Structural & Molecular Biology, vol. 17. No. 9. signaling in Arabidopsis thaliana. Genetics, vol. 161, No. 3, pp. pp. 1109-1113 (2010). Vidal et al., “Reverse two-hybrid and one hybrid systems to detect 1247-1255 (2002). dissociation of protein-protein and DNA-protein interactions'. Pro Matsui et al., “Arabidopsis Transcriptome Analysis under Drought, ceeding of the national Academy of Sciences of the USA, vol. 93, No. Cold, High-Salinity and ABA Treatment Conditions using a Tiling 19, pp. 103 15-10320 (1996). Array”, Plant Cell Physiol, vol. 49, No. 8, pp. 1135-1149 (2008). Gassner et al., “Accelerating the discovery of biologically active EP13715055.3, “Office Action”. Dec. 18, 2015, 4 pages. Small molecules using a high-throughput yeast halo assay, Journal CN201380018656.X, “Office Action”, Sep. 15, 2015, 8 pages. of Natural Products, vol. 70, No. 3, pp. 383-390 (2007). Liu Bulin (ed.) “Processing Technology for Pesticide Dosage Mosquna et al., Potent and selective activation of abscisic acid reep tors in vivo by mutational stabilization of their agonist-bound con Forms.” Beijing: Chemical Industry Press, Oct. 1998, pp. 17.hou, et formation, PNAS, vol. 108, No. 51, pp. 20838-20843 (2011). al., “Synthesis and Plant Growth Regulation Activity of Pyrabactin”. Fujii et al., “In vitro Reconstitution of an ABA Signaling Pathway”. Agrochemicals, vol. 49, No. 7, Jul. 2010, pp. 484-485. Nature, vol. 462, No. 7273, pp. 660-664 (2009). Zhou, et al., “Synthesis and Plant Growth Regulation Activity of Leon-KloosterZiel et al., “Isolation and characterization of abscisic Pyrabactin”, Agrochemicals, vol. 49, No. 7, Jul. 2010, pp. 484-485. acid-deficient Arabidopsis mutants at two new loci', Plant J. vol. 10, Patent Examination Report No. 1, Australian Pat. Applin. No. No. 4, pp. 655-661 (1996). 201324.0193, dated Feb. 9, 2016, 4 pages. U.S. Patent May 24, 2016 Sheet 1 of 27 US 9,345,245 B2

H O H O O Nss Ng7 CC w O - COOH - O- ) 11 C 10 (+)-ABA) Pyrabactin (PyrA) NO.1 3B r NO.24H

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U.S. Patent May 24, 2016 Sheet 2 of 27 US 9,345,245 B2

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FIG. 2A U.S. Patent May 24, 2016 Sheet 3 of 27 US 9,345,245 B2

DMSO Pyrabactin ABA Quinabactin

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