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(12) Patent Application Publication (10) Pub. No.: US 2016/0000837 A1 Rey Et Al US 2016.0000837A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2016/0000837 A1 Rey et al. (43) Pub. Date: Jan. 7, 2016 (54) COMPOSITIONS AND METHODS TO ALTER Related U.S. Application Data GUT MICROBAL FERMENTATION USING (60) Provisional application No. 61/852.221, filed on Mar. SULFATE-REDUCING BACTERA 15, 2013, provisional application No. 61/765,991, filed on Feb. 18, 2013. (71) Applicant: WASHINGTON UNIVERSITY, St. Publication Classification Louis, MO (US) (51) Int. Cl. A6135/74 (2006.01) (72) Inventors: Federico E. Rey, St. Louis, MO (US); A2.3L I/29 (2006.01) Mark Gonzalez, St. Louis, MO (US); A2.3L I/30 (2006.01) Jeffrey I. Gordon, St. Louis, MO (US) A 6LX3L/737 (2006.01) (52) U.S. Cl. CPC ............. A61K 35/741 (2013.01); A61 K3I/737 (21) Appl. No.: 14/768,394 (2013.01); A23L I/293 (2013.01); A23L I/3014 (2013.01); A61K 2035/1 15 (2013.01) (22) PCT Filed: Feb. 18, 2014 (57) ABSTRACT (86). PCT No.: PCT/US14/16883 The present invention provides combinations and methods for changing the representation of at least one Sulfate-reduc S371 (c)(1), ing bacterial species in a Subject's gut, thereby changing (2) Date: Aug. 17, 2015 microbial fermentative activity in the gut in the subject. Patent Application Publication Jan. 7, 2016 Sheet 1 of 20 US 2016/0000837 A1 Patent Application Publication Jan. 7, 2016 Sheet 2 of 20 US 2016/0000837 A1 Patent Application Publication Jan. 7, 2016 Sheet 3 of 20 US 2016/0000837 A1 Patent Application Publication Jan. 7, 2016 Sheet 4 of 20 US 2016/0000837 A1 on 90 ESCheriChia Coli 70 Y. CoilinSeiia aerofaciefs SO 30 - 20 10 Fi-S F.G. 2A Patent Application Publication Jan. 7, 2016 Sheet 5 of 20 US 2016/0000837 A1 is iii-S 2,8-3-O-tetan tricianohydrolase, C3.2.185 Fructokinase, EC2.7.1.4 5-dehydro-2-deoxygluconokinase, EC2.7.1.92 sorbitoi-6-phosphate dehydrogenase, C1,1,140 raitose-6-phosphate glicosidase, EC32, 22 -taitrate deydratase, EC42.32 1,3-propanedici dehydrogenase, EC 1202 N-acetyinetiatinate lyase, EC4.3.3 N-acetylglucosamine-8-statase, EC3.1.3.4 Hyaianuridase, EC3.2.1.35 glicioisoglycosaminoglycan lyase, EC4.2.2.1 3-N-acetylhexosaminidase, EC23, 1.48 4---giugan 4-giidaroydroiase, EC3.2.1.4 Ayopatiaase, EC3.2, 4 poiy gaiactufotate) hydroiase, EC3.3.87 Acetyxyla esterase, EC3, 2 Aaina endo--O--aabiatase, EC32.99 -attorate diehydratase, EC42, 7 -bulose 5-phosphate 4-epimerase, EC5, 3.4 Ributokinase, EC2.7.1.16 6-phosphoglucoseiaotonase, EC3.3 -3 -2 Patent Application Publication Jan. 7, 2016 Sheet 8 of 20 US 2016/0000837 A1 N WS × N S WSS: S. S. s SS SS S S. S. R & ? s x s s s N xx 3:1; 3 KSs Patent Application Publication Jan. 7, 2016 Sheet 9 of 20 US 2016/0000837 A1 FG 3B Patent Application Publication Jan. 7, 2016 Sheet 10 of 20 US 2016/0000837 A1 L?????????????????????????????????????????????????????????????????????????????????????????? |-100 p-1000 Patent Application Publication Jan. 7, 2016 Sheet 11 of 20 US 2016/0000837 A1 Feca FIP Ceca LFIHPP Feca FIS 400- T & Ceca FFS 2 5 O F.G. 3D Patent Application Publication Jan. 7, 2016 Sheet 12 of 20 US 2016/0000837 A1 3&tixias Cytopias: es Ot. -er acetate i/ y A. As FG 4 Patent Application Publication Jan. 7, 2016 Sheet 13 of 20 US 2016/0000837 A1 mutant library wild-type mutant library wild-type (LF/HPP) (LF/HPP) (HF/HS) (HFIHS) F.G. 5 atent Application Publication Jan. 7, 2016 Sheet 14 of 20 US 2016/0000837 A1 ç. H„slevels(mm) F.G. 6A Patent Application Publication Jan. 7, 2016 Sheet 16 of 20 US 2016/0000837 A1 D. gigsrauliance % of cost sity y 3.3%, Sos sspierrettes sit: cys & &is 838. SRs 3.1% S&s &c. &Q 3% hors. S. ^a : & & S. 3. 3.2%, Sö 3.3% $gos 8.2% SRs 88, 38:18, 8.3: FIG 7 Patent Application Publication Jan. 7, 2016 Sheet 17 of 20 US 2016/0000837 A1 & w ;: 88 ixiosis: &x::gs. &cy grasps - &yieattie caretire A Extery& *8. *::::::::::::::::::::::::::::::: ::::::::::::::::::::::::::::::::::::::::::::: ...::* s: Sy ::- sŠ s: :N-4 S: ses-&:8 so six 8:is & &S's is& :8 &8 ::::::::3: & N '''Pwywww. : *- ;:is ANGme Notass see-: f N 1 a. a a r. - Vis "'s Nes & 88: Patent Application Publication Jan. 7, 2016 Sheet 18 of 20 US 2016/0000837 A1 C. aerofaciens -8 FG 9A Norialized relative a undance - S - O - 5 O O.S O 15 3-Hydroxybutyrate Š& $xŠxŠ Lactate ŠxŠ Šx& Cysteine $x& SNSNS Aspartate &x. SNSNS Glutamate &x$ SwŠxŠ Histidire Šx& Šwsw Phenylacetate Š& &x&xŠ 4-Hydroxyphenylacetate Šx& ŠxŠxs Lyxosefxylose & S&S Fructose Š $x& Galactose &S ŠxŠxŠ Giucuronate S& Sw & 8 enter GalactOSannie $x& &x&x& $ 8 member N-Acetyl-Galactosamine Š Š& + D. piger Galactosamine 8-Stifate Š & FG 9B Patent Application Publication Jan. 7, 2016 Sheet 19 of 20 US 2016/0000837 A1 chondroitin ABC endolyase chondroitin sulfate ABC exolyase choline sulfatase C-glucuronidase W-glutamyltranspeptidase endochitinase 2, 1-3-D-fructan lyase malate dehydrogenase Mr WWOOOOOOOD YYYYYYYXXXX XXX 3-hexosaminidase uroporphyrinogen decarboxylase -12-08-04 00 0.4 0.8 12 FG 9C Patent Application Publication Jan. 7, 2016 Sheet 20 Of 20 US 2016/0000837 A1 8 member 8 member D. piger Acetate Butyrate Propionate FG 9D H US 2016/0000837 A1 Jan. 7, 2016 COMPOSITIONS AND METHODS TO ALTER SUMMARY OF THE INVENTION GUT MICROBAL FERMENTATION USING 0007. The present invention encompasses a combination SULFATE-REDUCING BACTERIA comprising a sulfated polysaccharide and an effective amount of at least one isolated Desulfo vibrio species. The at least one CROSS REFERENCE TO RELATED isolated Desulfo vibrio species comprises at least one nucleic APPLICATIONS acid with at least 80% identity to a nucleic acid selected from the group consisting of DpigGOR1. 1496 (SEQ ID NO: 1), 0001. This application claims the priority of PCT applica DpigGOR1. 1497 (SEQID NO: 2), DpigGOR1 0739 (SEQ tion No. PCT/US2014/016883, filed Feb. 18, 2014, which ID NO:3), DpigGOR1 0740 (SEQID NO:4), DpigGOR1 claims priority to U.S. provisional application No. 61/765, 1393 (SEQ ID NO. 5), DpigGOR1. 1398 (SEQID NO: 6), 991, filed Feb. 18, 2013, and U.S. provisional application No. DpigGOR1 0741 (SEQID NO:7), DpigGOR1 0744 (SEQ 61/852.221, filed Mar. 15, 2013, each of which is hereby ID NO:8), DpigGOR1 0790 (SEQID NO:9), DpigGOR1 incorporated by reference in its entirety. 0792 (SEQID NO: 10), DpigGOR1 0170 (SEQID NO: 11), and DpigGOR1 0174 (SEQID NO: 12). The isolated Des GOVERNMENTAL RIGHTS ulfo vibrio species may comprise any combination of 1,2,3,4, 5, 6, 7, 8, 9, 10, 11 or 12 nucleic acids. The sulfated polysac 0002 This invention was made with government support charide may be naturally occurring or synthetic, including but under DK78669, DK70977, DKO78669, and P30-AG028716 not limited to pentosan polysulfate, a fucoidan, a carrag awarded by the National Institutes of Health. The government eenan, a Sulfated glycosaminoglycan, or derivatives thereof. has certain rights in the invention. Optionally, the combination may further comprises an effec tive amount of at least one additional probiotic. FIELD OF THE INVENTION 0008. The present invention also encompasses a combina tion comprising a Sulfated polysaccharide and an effective 0003. The present invention encompasses compositions amount of at least one isolated SRB species selected from the and methods for changing the representation of sulfate-reduc group consisting of a D. piger and a bacterial species with at ing bacteria in a Subject's gut, thereby changing the microbial least one comparable in vivo fitness determinant to D. piger, fermentative activity in the gut and changing adiposity in the wherein the at least one comparable in vivo fitness determi Subject. nant is selected from the group consisting of DpigGOR1 1496 (SEQ ID NO: 1), DpigGOR1. 1497 (SEQID NO: 2), REFERENCE TO SEQUENCE LISTING DpigGOR1 0739 (SEQID NO:3), DpigGOR1 0740 (SEQ ID NO:4), DpigGOR1. 1393 (SEQID NO:5), DpigGOR1 0004. A paper copy of the sequence listing and a computer 1398 (SEQ ID NO: 6), DpigGOR1 0741 (SEQID NO: 7), readable form of the same sequence listing are appended DpigGOR1 0744 (SEQID NO:8), DpigGOR1 0790 (SEQ below and herein incorporated by reference. The information ID NO: 9), DpigGOR1 0792 (SEQ ID NO: 10), Dpig recorded in computer readable form is identical to the written GOR1 0170 (SEQ ID NO: 11), and DpigGOR1 0174 sequence listing, according to 37 C.F.R. 1.821 (f). (SEQ ID NO: 12). The isolated SRB species may comprise any combination of 1,2,3,4,5,6,7,8,9, 10, 11 or 12 nucleic BACKGROUND OF THE INVENTION acids. The Sulfated polysaccharide may be naturally occur ring or synthetic, including but not limited to pentosan 0005. In the gut, fermentation is one digestive process that polysulfate, a fucoidan, a carrageenan, a sulfated glycosami extracts energy from the available nutrient sources. Prior to noglycan, or derivatives thereof. Optionally, the combination the present invention, it was known in the art that clearing may further comprises an effective amount of at least one hydrogen gas generated by fermenting microbial communi additional probiotic. ties through mechanisms that produce methane (methanogen 0009. The present invention also encompasses a method esis), acetate (acetogenesis), or hydrogen Sulfide (via Sulfate for increasing microbial fermentative activity in the gut of a reduction), affects energy extraction from available nutrient Subject in need thereof. The method comprises administering Sources in the gut.
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