US 20110105421Al (19) United States (12) Patent Application Publication (10) Pub. No.: US 2011/0105421 A1 Mathee et al. (43) Pub. Date: May 5, 2011 (54) ELLAGITANNINS AS INHIBITORS OF (86) PCT No.: PCT/US09/37163 BACTERIAL QUORUM SENSING § 371 (00)’ (2), (4) Date: Dec. 14, 2010 (75) Inventors: Kalai Mathee, Miami, FL (US); _ _ Allison L‘ Adonizio’ Cambridge, Related US. Application Data MA (U S); Frederick AllSllbel, (60) Provisional application No. 61/036,812, ?led on Mar. Boston, MA (US); Jon Clardy, 14, 2008. Boston, MA (US); Bradley . Bennett’ Miami’ FL (Us); Kelsey Publication Classi?cation DoWnum, Arlington, TX (US) (51) Illt- Cl A61K 31/70 (2006.01) A61K 31/7048 (2006.01) (73) Assignee: The Florida International A611) 31/04 (200601) UPiVeFsitY 30m‘ "f Trustees’ (52) us. Cl. .............................. .. 514/29; 514/23; 514/39 M1am1, FL (US) (57) ABSTRACT (21) APPL NO; 12/922 555 Described herein are materials and methods for the inhibition ’ of bacterial QS. Methods of treating bacterial infections by administration of one or more ellagitannins in amount effec (22) PCT Filed: Mar. 13, 2009 tive to inhibit bacterial QS is also provided. Patent Application Publication May 5, 2011 Sheet 1 0f 4 US 2011/0105421 A1 Figure 1 Patent Application Publication May 5, 2011 Sheet 2 0f 4 US 2011/0105421 Al Bicmssny I B MBic'as-ssy Bimnsszy; kietiimml spH I) l fi C IMEIOBSMEY1 Figure 2 Patent Application Publication May 5, 2011 Sheet 3 0f 4 US 2011/0105421 A1 Panel I Panei II Figure 3 Patent Application Publication May 5, 2011 Sheet 4 0f 4 US 2011/0105421 A1 Auewba'xceatm'n “C, F'ifeutiai Tirna Ernie‘! Figure 4 US 2011/0105421A1 May 5,2011 ELLAGITANNINS AS INHIBITORS OF bacterial density, the concentration of the signaling molecule BACTERIAL QUORUM SENSING reaches a critical threshold to activate virulence genes. [0006] In Gram negative bacteria, for example, the signal CROSS-REFERENCE TO RELATED molecule is an acylated homoserine lactone (AHSL), often APPLICATION referred to as the autoinducer, Which interacts With a protein of a quorum regulon. A quorum regulon includes tWo pro [0001] The present application claims the bene?t of priority teins, the autoinducer synthase (the I protein) and the regula of US. Provisional Application No. 61/036,812, ?led Mar. tor (the R protein), Which, upon binding of the autoinducer, 14, 2008, the disclosure of Which is incorporated herein by activates the transcription of numerous genes. In Pseudomo reference in its entirety. nas aeruginosa, tWo quorum regulons have been identi?ed. One quorum regulon is knoWn as the LasIR system and is FIELD OF THE INVENTION mediated by a 3-oxo-dodecanoyl homoserine lactone (3-oxo [0002] The present application is directed to the use of C12-HSL) signal molecule. The other quorum regulon is knoWn as the Rhl1R system and is mediated by a butyryl ellagitannins for the inhibition of bacterial quorum sensing. homoserine lactone (C4-HSL) signal molecule. BACKGROUND OF THE INVENTION [0007] In recent years it has become apparent that many Gram-negative bacteria employ one or more quorum sensing [0003] Many microbial pathogens cause tremendous dam systems. The quorum sensing system is an attractive anti age WorldWide, in humans as Well as in animals and crop bacterial target because it is not found in humans and is plants. The continuing emergence of multiple-drug-resistant critical for high level bacterial virulence. Bacterial quorum pathogen strains has necessitated ?nding neW compounds sensing systems comprise AHL derivatives With different that can be used in antimicrobial treatment. In general, tWo acyl side chains to regulate, in a cell-density dependent man strategies exist for controlling pathogens, either kill the ner, a Wide variety of physiological processes unique to the pathogen or attenuate its virulence such that it does not dam life-cycle of each microbe. These processes include: sWarm age the host. ing, motility, bio?lm formation, conjugation, biolumines [0004] Many bacteria use autoinducer ligands to monitor cence and/or production of pigments, antibiotics and their population densities in a phenomenon called quorum enZymes. For example, in P aeruginosa quorum sensing sensing. See Fuqua & Greenberg, Nature Reviews Molecular pathWays affect the expression of various exoenZymes, bio Cell Biology, 3:685-695, 2002; or de Kievit et al., Infection & ?lm formation and cell-cell spacing. Other bacteria react to Immunity, 68: 4839-4849, 2000, for a revieW of the Quorum quorum sensing stimulation by expressing proteases and pec sensing system in pathogenic bacteria. Bacteria use quorum tinases, expressing pili, entering stationary phase, emerging sensing to regulate a variety of phenotypes, such as bio?lm from lag phase and initiating cell division. formation, toxin production, exopolysaccharide production, [0008] Bio?lms are dense extracellular polymeric matrices virulence factor production, and motility, Which are essential in Which the bacteria embed themselves. Bio?lms alloW bac for the successful establishment of a symbiotic or pathogenic teria to create a microenvironment that attaches the bacteria to relationship With their respective eukaryotic hosts (Marketon the host surface and Which contains excreted enZymes and et al., J. Bacteriol., 185:325-331, 2003; Ohtani et al., Mol. other factors alloWing the bacteria to evade host immune Microbiol., 44:171-179, 2002; Quinones et al., Mol. Plant responses including antibodies and cellular immune Microbe Interact. 18:682-693, 2005; Rice et al., J. Bacteriol., responses. Such bio?lms can also exclude antibiotics. Fur 187:3477-3485, 2005; Suntharalingam et al., Trends Micro ther, bio?lms can be extremely resistant to removal and dis biol., 1313-6, 2005). At high cell densities, bacteria use this infection. For individuals suffering from cystic ?brosis, the chemical signaling process to sWitch from a nomadic exist formation of bio?lms by R aeruginosa is eventually fatal. ence to that of a multicellular community. This lifestyle Other bacteria also respond to quorum sensing signals by sWitch is signi?cant, as numerous pathogenic bacteria use producing bio?lms. Bio?lms are inherent in dental plaques, quorum sensing to turn on virulence pathWays and form drug and are found on surgical instruments, food processing and impervious communities called bio?lms that are the basis of agriculture equipment and Water treatment and poWer gener a myriad chronic infections. Over 80% of bacterial infections ating machinery and equipment. in humans involve the formation of bio?lms, as exempli?ed in [0009] Because of the virulence factors it triggers, the bac lung infections by Pseudomonas aeruginosa, Which is the terial quorum-sensing system offers a target for use in modu primary cause of morbidity in cystic ?brosis patients. The lating the virulence of pathogenic bacteria. All acyl-ho treatment of infections by pathogens that form bio?lms costs moserine lactone quorum-sensing systems described to date, over $1 billion/year in the US alone. Studies With animal except that of V harveyi, utiliZe AI synthases encoded by a models have shoWn that strains With inactivated quorum sens gene homologous to luxI of V?scheri. The response to the ing genes shoW reduced virulence. Disrupting quorum sens autoinducer is mediated by a transcriptional activator protein ing may interfere With the ability of bacteria to form robust encoded by a gene homologous to luxR of V?scheri (Bassler bio?lms and thus render the bacteria more sensitive to anti and Silverman, in TWo Component Signal Transduction, bacterial agents and the host’s immune response. Hoch et al., eds., Am. Soc. Microbiol. Washington DC, pp. [0005] Quorum sensing is mediated by a signal molecule 431-435,1995). that binds to a cognate transcriptional activator to cause either [0010] Gram-negative bacteria represent numerous rel upregulation or repression of genes that increase virulence evant pathogens using quorum-sensing pathWays. Besides R factors, Which include exotoxins, proteases, alginates, aeruginosa, other gram-negative quorum sensing bacteria lipopolysaccharides, pyocyanin and rhamnolipids. At loW include: Aeromonas hydrophila, A. salmonicida, Agrobacle bacterial cell density, the concentration of the signaling mol rium Zumefaciens, Burkholderia cepacia, Chromobaclerium ecule does not activate the virulence genes, While at higher violaceum, Enlerobacler agglomeran, Erwinia carolovora, US 2011/0105421A1 May 5,2011 E. chrysanlhemi, Escherichia coli, Nilrosomas europaea, malian subject, the method comprising administering to the Obesumbaclerium proleus, Panloea slewarlii, Pseudomonas subject an ellagitannin in an amount effective to treat the aureofaciens, P syringae, Ralslonia solanacearum, Rhiso infection. In one embodiment, the infection an infection bium elli, R. leguminosarum, Rhodobacler sphaeroides, Ser caused by a bacterium is selected from the group consisting ralia liguefaciens, S. marcescens, [?brio anguillarum, V?s of. Aeromonas hydrophila, Aeromonas salmonicida, Agro cheri, V cholerae, Xenorhabdus nemalophilus, Yersinia baclerium Zumefaciens, Burkholderia cepacia, Chromobac enlerocolilica, I’. peslis, I’. pseudoluberculosis, I’. medievalis, Zerium violaceum, Enlerobacler agglomeran, Erwinia caro and I’. ruckeri. Zovora, Erwinia chrysanlhemi, Escherichia coli, Nilrosomas [0011] In addition to their pathogenic costs, quorum sens europaea, Obesumbaclerium proleus, Panloea slewarlii, ing bacteria also have signi?cant economic impact in indus Pseudomonas aureofaciens,