EA037069B1 - Pharmaceutical composition comprising bacteria of the genus blautia for treating diseases of the intestine - Google Patents

Abstract

The invention relates to a pharmaceutical composition comprising a bacterial strain of Blautia hydrogenotrophica DSMZ 14294 having the 16s rRNA sequence of SEQ ID NO: 5 and/or a bacterial strain of the genus Blautia having a 16s rRNA sequence that is at least 95% identical to SEQ ID NO: 5 for treating or preventing diarrhea and/or constipation in a subject.

Description

(54) PHARMACEUTICAL COMPOSITION CONTAINING BACTERIA OF THE GENUS BLAUTIA FOR THE TREATMENT OF INTESTINAL DISEASES

(31) 1612190.7; 1616016.0; 1616018.6; 1703552.8; 1703548.6 (32) 2016.07.13; 2016.09.20; 2016.09.20; 2017.03.06; 2017.03.06 (33) GB (43) 2019.04.30 (86) PCT / GB2017 / 052077 (87) WO 2018/011594 2018.01.18 (71) (73) Applicant and patentee: (56) US-A1-2016184370 US-A1-2010247489 WO-A1-2012142605 WO-A1-0207741 WO-A1-0185187 WO-A1-2016057671 C. LIU ET AL: Reclassification of Clostridium coccoides, Ruminococcus hansenii, Ruminococcus hydrogenotrophicus, Ruminococcus luti, Ruminococcus productus and Ruminococcus schinkii as Blautia coccoides gen. nov., 4D PHARMA PIELS (GB) comb, nov., Blautia hansenii comb, nov., (72) Inventor: Blautia hydrogenotrophica comb, nov., Blautia luti comb, nov, INTERNATIONAL JOURNAL Bernalier-Donadiy Annick, Cruse Loreen, Abuzy Cloe (FR) OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY, vol. 58, no. 8, 1 August (74) Representative: 2008 (2008-08-01), pages 1896-1902, XP055005236, ISSN: 1466-5026, D01: 10.1099 / ijs.0.65208-0, the Medvedev V.N. (RU) whole document

037069 B1 (57) The invention relates to a pharmaceutical composition containing the bacterial strain Blautia hydrogenotrophica DSMZ 14294 having the sequence of the 16s rRNA gene SEQ ID NO: 5 and / or a bacterial strain of the genus Blautia having the sequence of the 16s rRNA gene at least 95% identical SEQ ID NO: 5, for treating or preventing diarrhea and / or constipation in a subject.

037069 Bl

Technology area

This invention relates to the field of compositions containing bacterial strains isolated from the digestive tract of mammals, and the use of such compositions for the treatment of disease.

Prior art

It is believed that the human intestine is sterile in utero, but immediately after birth it is exposed to a large number of microorganisms from the mother and from the environment. This is followed by a dynamic period of microbial colonization and succession, depending on factors such as mode of birth, environment, nutrition, and host genotype, which collectively affect the composition of the gut microbiota, especially during early life. Subsequently, the microbiota is stabilized and becomes similar to the microbiota of adults [1]. The human intestinal microbiota contains more than 1500 different phylotypes, among which two main bacterial types (phyla) are quantitatively dominant - Bacteroidetes and Firmicutes [2-3]. Successful symbiotic relationships resulting from bacterial colonization of the human intestine have led to the development of a wide range of metabolic, structural, protective and other beneficial functions. The increased metabolic activity of the colonized intestine allows for the degradation of otherwise indigestible food components to form by-products that are an important source of nutrients for the host and provide additional health benefits. Likewise, the immunological importance of the gut microbiota is generally recognized and has been confirmed in microbial-free animals that have a dysfunctional immune system that is functionally restored after administration of commensal bacteria [4-6].

Significant changes in microbiota composition have been documented in gastrointestinal disorders such as inflammatory bowel disease (IBD). For example, the levels of bacteria of the Clostridium XIVa cluster and the Clostridium XI cluster (F. prausnitzii) in patients with IBD are reduced, while the number of E. coli increases, which suggests a shift in the balance between symbionts and pathobionts in the intestine [7-11].

In recognition of the potential beneficial effect that certain bacterial strains can have on the intestines of an animal, different strains have been proposed for use in the treatment of various diseases (see, for example, [12-15]). A number of strains, including mainly Lactobacillus and Bifidobacterium strains, have been proposed for use in the treatment of various intestinal disorders (see review [16]). Strains of the genus Blautia have also been proposed for use in modulating the bacterial balance of the digestive ecosystem in patients with IBS (Irritable Bowel Syndrome) (WO 01/85187). However, the relationships between different bacterial strains and different diseases and the exact effects of specific bacterial strains on the gut and at the systemic level and on any specific types of disease are poorly understood.

There is a need to study the potential effects of gut bacteria in order to develop new treatments using gut bacteria.

US 2010/0247489 describes the use of mineral nutrients for the treatment of digestive disorders. US '789 also suggests the possible use of numerous different genera of bacteria to prevent a wide range of gastrointestinal symptoms, but does not associate any bacteria with any disease or symptom.

WO 2016/086206 suggests that bacteria of the order Clostridiales can be used to treat or prevent various conditions, but does not associate any bacteria with any disease or symptom.

WO 2012/142605 suggests the possibility of treating a number of different diseases with a combination of microorganisms. WO'605 suggests the possible use of a large number of bacterial species, but does not contain a description of how any of the bacterial species proposed in WO'605 can be used to treat any of the proposed diseases.

WO 02/07741 suggests the use of various bacteria of the Clostridia class for the treatment of gastrointestinal disorders, but does not associate any bacteria with any disease or symptom. WO 2016/086209 suggests the possible use of a large number of bacteria of the order Clostridiales to reduce the secretion of pro-inflammatory cytokines and / or increase the secretion of anti-inflammatory cytokines, but does not describe how any of the proposed bacterial species can be used to treat any of the proposed diseases.

The essence of the invention

The inventors have developed new therapies for the treatment and prevention of diarrhea and / or constipation. In particular, the inventors have determined that bacterial strains from the genus Blautia can effectively reduce diarrhea and / or constipation. As described in the examples, oral administration of compositions containing Blautia hydrogenotrophica can reduce diarrhea and / or constipation in patients with irritable bowel syndrome (IBS). Thus, in a first embodiment, the invention provides a composition comprising a bacterial strain of the genus Blautia for use in a method for treating or preventing diarrhea and / or constipation.

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In preferred embodiments, the invention provides a composition comprising a bacterial strain of the genus Blautia for use in a method for treating or preventing diarrhea and / or constipation in a subject diagnosed with Crohn's disease, ulcerative colitis, or more preferably IBS. In some embodiments, a subject diagnosed with IBS has IBS with manifestations of both diarrhea and constipation. In some embodiments, a subject diagnosed with IBS has IBS with diarrhea but no constipation or only mild constipation. In some embodiments, a subject diagnosed with IBS has IBS with constipation but no diarrhea or only mild diarrhea.

In preferred embodiments of the invention, the bacterial strain in the composition belongs to Blautia hydrogenotrophica. Closely related strains can also be used, such as bacterial strains having a 16s pRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% identical to the 16s pRNA sequence of bacterial strain Blautia hydrogenotrophica. Preferably, the bacterial strain has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 5. Most preferably, the bacterial strain in of the composition is a Blautia hydrogenotrophica strain deposited with accession number DSM 10507/14294.

In additional embodiments of the invention, the bacterial strain in the composition belongs to Blautia stercoris. Closely related strains can also be used, such as bacterial strains having a 16s pRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% identical to the 16s pRNA sequence of bacterial strain Blautia stercoris. Preferably, the bacterial strain has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 1 or 3. Preferably, sequence identity is defined with respect to SEQ ID NO: 3. Preferably, the bacterial strain for use in the invention has the 16s pRNA sequence represented by SEQ ID NO: 3.

In additional embodiments of the invention, the bacterial strain in the composition belongs to Blautia wexlerae. Closely related strains can also be used, such as bacterial strains having a 16s pRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% identical to the 16s pRNA sequence of bacterial strain Blautia wexlerae. Preferably, the bacterial strain has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 2 or 4. Preferably, sequence identity is defined with respect to SEQ ID NO: 4. Preferably, the bacterial strain for use in the invention has the 16s pRNA sequence represented by SEQ ID NO: 4.

In certain embodiments, a composition of the invention is for oral administration. Oral administration of the strains of the invention can be effective in treating diarrhea and / or constipation. Also, oral administration is convenient for patients and practitioners and allows delivery to and / or partial or complete colonization of the intestine.

In certain embodiments, a composition of the invention comprises one or more pharmaceutically acceptable excipients or carriers.

In certain embodiments, a composition of the invention comprises a lyophilized bacterial strain. Lyophilization is an efficient and convenient technique for the preparation of stable compositions that allow delivery of bacteria, and has demonstrated the possibility of obtaining effective compositions in the examples.

In certain embodiments, the invention provides a food product comprising a composition as described above.

In certain embodiments, the invention provides a vaccine composition comprising a composition as described above.

Additionally, the invention provides a method for treating or preventing diarrhea and / or constipation, comprising administering a composition comprising a bacterial strain of the genus Blautia.

In preferred embodiments, the invention provides a composition comprising a bacterial strain of the genus Blautia for use in a method for treating or preventing a disease associated with an Enterobacteriaceae infection, such as E. coli infection. In certain embodiments, the compositions of the invention are for use in the treatment or prevention of diarrhea, gastroenteritis, urinary tract infection, or neonatal meningitis.

In further preferred embodiments, the compositions of the invention are intended for use to reduce the level of Enterobacteriaceae in the gastrointestinal tract, preferably E. coli, in the treatment or prevention of IBS, Crohn's disease, ulcerative colitis, functional dyspepsia, diarrhea, gastroenteritis, urinary infection pathways or meningitis of newborns.

In particularly preferred embodiments, the compositions of the invention are for use in a method of treating or preventing diarrhea associated with an Enterobacteriaceae infection, such as E. coli infection, or are intended to be used to reduce

- 2,037,069 levels of Enterobacteriaceae in the gastrointestinal tract, preferably E. coli, in the treatment or prevention of diarrhea.

Brief description of graphic materials

FIG. 1: Changes in patient symptoms over the dosing period (days 1-16) of phase I clinical trials.

FIG. 2: Changes in patient symptoms over the washout period of phase I clinical trials.

FIG. 3: Results of C _max determination by hydrogen breath test on Day 1, Day 2, Day 15 and Day 16 for IBS patients treated with Blautix (FIG. 3a) and IBS patients treated with placebo (FIG. 3b). FIG. 3c is a graph comparing the percentage of Blautix-treated patients with a decrease in hydrogen between the mean for days 15 and 16 and the mean for days 1 and 2, with the percentage of placebo-treated patients with a decrease in hydrogen between these time points.

FIG. 4a: Paired data from unadjusted and corrected hydrogen breath tests on Day 1 and Day 15 for IBS patients treated with Blautix; fig. 4b: Graph comparing mean uncorrected hydrogen breath test results on Day 1 and Day 15 for the Blautix treatment group; fig. 4c: Graph comparing the mean corrected hydrogen breath test results on Day 1 and Day 15 for the Blautix treatment group.

FIG. 5a: Paired data from unadjusted and corrected hydrogen breath tests on Day 1 and Day 15 for IBS patients receiving placebo; fig. 5b: Graph comparing mean uncorrected hydrogen breath test results on Day 1 and Day 15 for the placebo group; fig. 5c: Graph comparing the adjusted mean adjusted hydrogen breath test results on Day 1 and Day 15 for the placebo group.

FIG. 6: Graphs comparing the average hydrogen breath test results on Day 1 and Day 15 for the Blautix treatment group (Verum (formulation with active ingredients)) and the placebo group (Fig.6a: uncorrected hydrogen level; Fig.6b: corrected hydrogen level ).

Disclosure of invention

Bacterial strains

The compositions according to the invention contain a bacterial strain of the genus Blautia. The examples demonstrate that bacteria of this genus are useful in the treatment or prevention of diarrhea and / or constipation. Preferred bacterial strains belong to the species Blautia hydrogenotrophica, Blautia stercoris and Blautia wexlerae. Other preferred bacterial strains for use in the invention are Blautia producta, Blautia coccoides and Blautia hansenii.

Examples of Blautia strains for use in the invention include Blautia hydrogenotrophica, B. stercoris, B. faecis, B. coccoides, B. glucerasea, B. hansenii, B. luti, B. producta, B. schinkii and B. wexlerae. Blautia species are gram-positive immotile bacteria that can be coccoid or oval, and all are obligate anaerobes that produce acetic acid as the main end product of glucose fermentation [17]. Blautia can be isolated from the human intestine, although B. producta was isolated from a septic sample.

Blautia hydrogenotrophica (formerly Ruminococcus hydrogenotrophicus) was isolated from the intestines of mammals, is strictly anaerobic, and metabolizes H ₂ / CO ₂ to acetate, which may be important for human nutrition and health. A typical Blautia hydrogenotrophica strain is S5a33 = DSM 10507 = JCM 14656. The 16S rRNA gene sequence of Blautia hydrogenotrophica S5a36 has GenBank accession number X95624.1 (cited herein as SEQ ID NO: 5). This typical Blautia hydrogenotrophica strain is described in [17] and [18]. Strain S5a33 and strain S5a36 correspond to two subclones of a strain isolated from a fecal sample from a healthy subject. They show the same morphology, physiology, and metabolism, and have identical 16s rRNA sequences. Thus, in some embodiments, Blautia hydrogenotrophica for use in the invention has a 16s rRNA sequence in accordance with SEQ ID NO: 5.

The Blautia hydrogenotrophica bacterium deposited with accession number DSM 10507 as well as accession number DSM 14294 has been tested in the examples and is also referred to herein as strain BH. The BH strain was deposited in the German Collection of Microorganisms and Cell Cultures (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Mascheroder Weg 1b, 38124 Braunschweig, Germany) on January 26, 1996 as Ruminococcus hydrogenotrophicus with accession number DSMZ 10507, and 1429 with accession number DSMZ as S5a33 on 14 May 2001. The depositary was INRA Laboratoire de Microbiologie CR de Clermont-Ferrand / Theix 63122 Saint Genes Champanelle, France. Ownership of the deposited materials passed to 4D Pharma Plc by assignment.

The 16S rRNA gene sequence of Blautia stercoris strain GAM6-1T has GenBank accession number HM62 6177 (disclosed herein as SEQ ID NO: 1). A typical example of the Blautia stercoris strain is described in [19]. A typical Blautia wexlerae strain is WAL 14507 = ATCC BAA-1564 = DSM 19850 [17]. The gene sequence of the 16S pRNA of the WAL 14507 T Blautia wexlerae strain has GenBank accession number EF036467 (disclosed herein as SEQ ID NO: 2). This typical strain

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Blautia wexlerae is described in [17].

A preferred Blautia stercoris strain is the strain deposited with accession number NCIMB 42381, which is also referred to herein as strain 830. The 16s sequence of pRNA of strain 830 is shown as SEQ ID NO: 3. Strain 830 was deposited with the international depository NCIMB, Ltd. (Ferguson Building, Aberdeen, AB21 9YA, Scotland) by GT Biologics Ltd. (Life Sciences Innovation Building, Aberdeen, AB25 2ZS, Scotland) on 12 March 2015 as Blautia stercoris 830 and assigned accession number NCIMB 42381. GT Biologics Ltd. subsequently changed its name to 4D Pharma Research Limited.

A preferred Blautia wexlerae strain is the strain deposited with accession number NCIMB 42486, which is also referred to herein as strain MRX008. The sequence of 16s pRNA of strain MRX008 is shown as SEQ ID NO: 4. Strain MRX008 was deposited with the international depository NCIMB, Ltd. (Ferguson Building, Aberdeen, AB21 9YA, Scotland) by 4D Pharma Research Ltd. (Life Sciences Innovation Building, Aberdeen, AB25 2ZS, Scotland) November 16, 2015 as Blaqutia / Ruminococcus MRx0008 and assigned accession number NCIMB 42486.

It is expected that bacterial strains closely related to the strain tested in the examples will also be effective in treating or preventing diarrhea and / or constipation. In certain embodiments, the bacterial strain for use in the invention has a 16s rRNA sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% identical to the 16s rRNA sequence of the Blautia bacterial strain hydrogenotrophica. Preferably, the bacterial strain for use in the invention has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 5.

In certain embodiments, the bacterial strain for use in the invention has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% identical to the 16s rRNA sequence of the Blautia bacterial strain stercoris. Preferably, the bacterial strain for use in the invention has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 1 or SEQ ID NO: 3. Preferably, sequence identity is determined with respect to SEQ ID NO: 3. Preferably, the bacterial strain for use in the invention has the 16s rRNA sequence represented by SEQ ID NO: 3. In certain embodiments, the bacterial strain for use in the invention has the sequence 16s rRNA is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to the 16s rRNA sequence of the bacterial strain Blautia wexlerae. Preferably, the bacterial strain for use in the invention has a 16s rRNA sequence at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to SEQ ID NO: 2 or SEQ ID NO: 4. Preferably, sequence identity is determined with respect to SEQ ID NO: 4. Preferably, the bacterial strain for use in the invention has the 16s rRNA sequence represented by SEQ ID NO: 4.

Bacterial strains that are biotypes of bacteria deposited with accession number DSM 10507/14294 or biotypes of bacteria deposited with accession numbers NCIMB 42381 and NCIMB 42486 are also expected to be effective in the treatment or prevention of diarrhea and / or constipation. A biotype is a closely related strain that has the same or very similar physiological and biochemical characteristics.

Strains that are biotypes of the bacterium deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486 and are suitable for use in the invention can be identified by sequencing other nucleotide sequences of the bacterium deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486. For example, a substantially complete genome can be sequenced and a biotype strain for use in the invention can have at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9 % sequence identity in at least 80% of its entire genome (for example, at least 85%, 90%, 95% or 99% or in its entire genome). For example, in some embodiments, a biotype strain has at least 98% sequence identity in at least 98% of its genome, or at least 99% sequence identity in 99% of its genome. Other suitable sequences for use in identifying biotype strains may include hsp60 or repeated sequences such as BOX, ERIC, (GTG) ₅ or REP or [20]. Strains of the biotype may have sequences with at least 95%, 96%, 97%, 98%, 99%, 99.5%, or 99.9% sequence identity with the corresponding sequence of the bacterium deposited with DSM accession number 10507/14294, NCIMB 42381 or NCIMB 42486. In some embodiments, the biotype strain has a sequence with at least 97%, 98%, 99%, 99.5%, or 99.9% sequence identity with the corresponding sequence of the Blautia hydrogenotrophica strain deposited as DSM 10507/14294 and contains a 16s rRNA sequence that is at least 99% identical (e.g., at least 99.5% or at least 99.9% identical) to SEQ ID NO: 5. In some embodiments, the biotype strain has sequence with at least 97%, 98%, 99%, 99.5% or 99.9% sequence identity with the corresponding sequence

- 4 037069 with the identity of the Blautia hydrogenotrophica strain deposited as DSM 10507/14294 and has the 16s pRNA sequence according to SEQ ID NO: 5.

Alternatively, strains that are biotypes of bacteria deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486 and are suitable for use in the invention can be identified by using deposit accession number DSM 10507/14294, deposit accession number NCIMB 42381 or deposit with accession number NCIMB 42486 and performing restriction fragment analysis and / or PCR analysis, for example, by fluorescence analysis of amplified fragment length polymorphism (FAFLP) and fingerprinting using PCR amplification of repeated sequences of intergenic palindromes (repetitiveR DNA element (rep) -PCR) fingerprinting), or protein profiling, or partial 16S or 23s rDNA sequencing. In preferred embodiments, such techniques can be used to identify other strains of Blautia hydrogenotrophica, Blautia stercoris, or Blautia wexlerae.

In certain embodiments, strains that are biotypes of a bacterium deposited with accession number DSM 10507/14294, NCIMB 42381, or NCIMB 42486 and suitable for use in the invention are strains that provide the same pattern as the bacterium deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486, when analyzing by restriction analysis of amplified ribosomal DNA (ARDRA), for example, using the restriction enzyme Sau3AI (examples of methods and guidance are given, for example, in [21]). Alternatively, strains of the biotype are identified as having the same carbohydrate fermentation pattern as the bacterium deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486.

Other Blautia strains suitable for use in the compositions and methods of the invention, such as biotypes of bacteria deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486, can be identified using any suitable method or strategy, including the assays described in examples. For example, strains for use in the invention can be identified by culturing bacteria and administering to rats for testing by a distension assay. In particular, bacterial strains having growth characteristics, metabolic type and / or surface antigens similar to bacteria deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486 may be suitable for use in the invention. A useful strain will have comparable microbiota modulating activity with DSM 10507/14294, NCIMB 42381, or NCIMB 42486. In particular, the biotype strain will have an effect on diarrhea and / or constipation comparable to the effects shown in the examples, which can be identified using the protocols cultivation and introduction described in the examples.

A particularly preferred strain according to the invention is the Blautia hydrogenotrophica strain deposited with accession number DSM 10507/14294. It is a typical BH strain that has been used for testing in the examples and has been shown to be effective in treating disease. Thus, the invention provides a cell, such as an isolated cell, of the Blautia hydrogenotrophica strain deposited with accession number DSM 10507/14294, or a derivative thereof, for use in therapy, in particular in the diseases described herein.

Derivatives of the strain deposited with accession number DSM 10507/14294, NCIMB 42381 or NCIMB 42486 may be a daughter strain (progeny) or a strain grown (subcloned) from the parent strain. The derived strain according to the invention can be modified, for example, at the genetic level, without loss of biological activity. In particular, the derived strain of the invention is therapeutically active. The derived strain will have a microbiota modulating activity comparable to the original strain DSM 10507/14294, NCIMB 42381 or NCIMB 42486. In particular, the derived strain will have an effect on diarrhea and / or constipation comparable to the effects shown in the examples, which can be identified with using the cultivation and administration protocols described in the examples. Derivative DSM 10507/14294 will usually be the biotype of strain DSM 10507/14294. Derivative NCIMB 42381 will usually be the biotype of strain NCIMB 42381. Derivative NCIMB 42486 will usually be the biotype of strain NCIMB 42486.

References to cells of the Blautia hydrogenotrophica strain deposited with accession number DSM 10507/14294 encompass any cells having the same safety and therapeutic efficacy characteristics as strains deposited with accession number DSM 10507/14294, and such cells are within the scope of the invention. References to cells of the Blautia stercoris strain deposited with accession number NCIMB 42381 encompass any cells having the same safety and therapeutic efficacy characteristics as strains deposited with accession number NCIMB 42381, and such cells are within the scope of the invention. References to cells of the Blautia wexlerae strain deposited with accession number NCIMB 42486 encompass any cells having the same safety and therapeutic efficacy characteristics as strains deposited with accession number NCIMB 42486, and such cells are within the scope of the invention.

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In preferred embodiments, the bacterial strains in the compositions of the invention are viable and capable of partially or completely colonizing the intestine.

Therapeutic Uses

In preferred embodiments, the compositions of the invention are for use in the treatment of diarrhea and / or constipation. In some embodiments, the composition is for use in treating a patient having both diarrhea and constipation. In some embodiments, the composition is for use in treating a patient having diarrhea without constipation. In some embodiments, the composition is for use in treating a patient having constipation without diarrhea. Although patients often have diarrhea or constipation, the same patient may have both conditions at different times. Diarrhea and / or constipation is common in patients with IBS and / or inflammatory bowel disease. Accordingly, the invention provides a composition of the invention for use in treating a patient diagnosed or suspected of having IBS or inflammatory bowel disease. For example, IBS patients may experience alternating cycles of diarrhea and constipation, or an IBS patient may be characterized as an IBS patient with diarrhea or an IBS patient with constipation. In some embodiments, a composition of the invention can be used to treat a patient with IBS, a patient with IBS with diarrhea, or a patient with IBS with constipation. For example, in some embodiments, a subject diagnosed with IBS has IBS with manifestations of both diarrhea and constipation. In some embodiments, a subject diagnosed with IBS has IBS with diarrhea but no constipation or only mild constipation. In some embodiments, a subject diagnosed with IBS has IBS with constipation but no diarrhea or only mild diarrhea. In some embodiments, the inflammatory disease refers to the small intestine, colon, or rectum. Diarrhea is a symptom of Crohn's disease and ulcerative colitis. Accordingly, the composition of the invention can be used to treat a patient with Crohn's disease or ulcerative colitis.

As demonstrated in the examples, the bacterial compositions of the invention can effectively reduce diarrhea and / or constipation.

In preferred embodiments, the compositions of the invention are for use in the treatment or prevention of diarrhea and / or constipation associated with Crohn's disease, ulcerative colitis, or more preferably IBS. In preferred embodiments, the compositions of the invention are for use in treating or preventing diarrhea and / or constipation in a subject diagnosed with Crohn's disease, ulcerative colitis, or more preferably IBS. In preferred embodiments, the compositions of the invention are for use in the treatment or prevention of diarrhea and / or constipation in the treatment of Crohn's disease, ulcerative colitis, or more preferably IBS. In some embodiments, the patient has abdominal pain and / or bloating in addition to diarrhea and / or constipation. In such embodiments, the patient can have any combination of two, three, or all four of these conditions. For example, a patient may have diarrhea, constipation, abdominal pain, and flatulence; diarrhea, constipation, and abdominal pain; diarrhea, constipation and flatulence; diarrhea, abdominal pain and / or flatulence without constipation; constipation, abdominal pain and / or flatulence without diarrhea.

In certain embodiments, the composition may be in the form of a bacterial culture. In some embodiments, the composition may preferably be a lyophilisate.

In some embodiments, the composition of the invention is for use in the treatment of diarrhea and / or constipation in a subject having an increased level of hydrogen in their breath as compared to a healthy subject. In some embodiments, a composition of the invention is for use in reducing respiratory hydrogen levels in a subject having diarrhea and / or constipation. The subject is preferably a subject diagnosed with IBS and / or inflammatory bowel disease. In some embodiments, the inflammatory disease refers to the small intestine, colon, or rectum. In some embodiments, the subject has been diagnosed with Crohn's disease, ulcerative colitis, or more preferably IBS, such as one of the types of IBS described herein. Without being bound by theory, it is assumed that the elevated hydrogen levels are due to mechanisms underlying inflammation in the gut. The examples show that treatment with a composition of the invention reduces the level of hydrogen detected in breath hydrogen tests. Accordingly, hydrogen levels are preferably assessed using a hydrogen breath test. The hydrogen breath test is well known in the art and thus the skilled person will know how to perform such a test. In some embodiments, lactulose is administered to the patient as a test substrate.

The breath hydrogen test is also a useful tool for monitoring the efficacy or likely efficacy of a treatment or prophylaxis using a composition of the invention. For example, a decrease in the level of hydrogen detectable in a subject's breath following treatment or prophylaxis using a composition of the invention may indicate that the treatment is having a therapeutic or prophylactic effect. Accordingly, in some variants, real

The methods and use of the invention further comprise monitoring the respiration hydrogen level of a subject during and / or after treatment or prophylaxis using a composition of the invention and thereby assessing the efficacy or likely efficacy of the treatment or prevention. For example, hydrogen levels can be monitored at one or more (eg, 1, 2, 3, 4, or more than 4) points in time, for example including time points before treatment, at the beginning of treatment, during treatment, at the end of treatment and / or after treatment as desired. In some embodiments, the level of hydrogen in the subject's breath at the end and / or after the dosing period (during which the subject is administered the composition) is compared to the level at the beginning and / or before the dosing period, and a decrease in the level indicates the effectiveness or likely effectiveness of the treatment or prevention ... For example, in embodiments in which the dosing period is 16 days, it may be desirable to take measurements on Day 1 and Day 16, or for example, on Day 1, Day 2, Day 15, and Day 16. In some embodiments, multiple measurements are taken and determine the average of such measurement results (for example, the average for day 1 and day 2 and the average for day 15 and day 16). In some embodiments, the hydrogen reduction Cmax level of at least 40 ppm (mn ^-1) indicates that the treatment or prevention are effective or probably effective. In some embodiments, the respiration hydrogen level of the subject is measured only once, such as at the end of or after treatment, and a level value equal to or close to a predetermined level indicates that the treatment or prevention was likely to be effective. The breath hydrogen test is a standard assay and thus predetermined levels are known to those skilled in the art.

In certain embodiments, compositions of the invention are intended for use to prevent diarrhea and / or constipation in a subject receiving, or receiving, or intending to receive (e.g., later that day, the next day, or for 2, 3, 4, 5, 6, 7, 10, 14 or 21 days) antibiotic treatment, or suffering from or had recovered from bacterial gastroenteritis. Antibiotic treatment and bacterial gastroenteritis are associated with changes in the gut microbiota that may precede diarrhea and / or constipation and which can be prevented by the compositions of the invention. The compositions of the invention can be administered concurrently with, separately from, or sequentially with antibiotic treatment. For example, the compositions of the invention can be administered concurrently with antibiotic treatment.

In preferred embodiments, treatment with compositions of the invention results in a decrease in diarrhea and / or constipation.

Treatment or prevention of diarrhea and / or constipation can refer, for example, to a decrease in the severity of symptoms or to a decrease in the frequency of exacerbations, or to a decrease in the set of factors (triggers) that are problematic for the patient.

In certain embodiments, the compositions of the invention are for use in the treatment or prevention of a disease associated with an Enterobacteriaceae infection, such as E. coli infection. In certain embodiments, the compositions of the invention are for use in the treatment or prevention of diarrhea, gastroenteritis, urinary tract infection, or neonatal meningitis. In such embodiments, the Enterobacteriaceae may be a pathogenic strain.

In certain embodiments, compositions of the invention are for use in the treatment or prevention of a disease associated with elevated levels of Enterobacteriaceae, such as E. coli. In certain embodiments, the compositions of the invention are for use in the treatment or prevention of diarrhea, gastroenteritis, urinary tract infection, or neonatal meningitis. In such embodiments, the Enterobacteriaceae may be a commensal or non-pathogenic strain.

In preferred embodiments, the compositions of the invention are for use to reduce the level of Enterobacteriaceae in the gastrointestinal tract, preferably E. coli, in the treatment or prevention of a disease associated with elevated levels of Enterobacteriaceae, such as IBS, Crohn's disease, ulcerative colitis. , functional dyspepsia, diarrhea, gastroenteritis, urinary tract infection, or neonatal meningitis. It is known that Enterobacteriaceae, and in particular E. coli, are potential initiators or, as is known, lead to exacerbation of Crohn's disease and ulcerative colitis [22-24], therefore, the effect demonstrated in the examples for the compositions of the invention may be beneficial for treating these conditions.

In certain embodiments, the compositions of the invention are for use in the treatment or prevention of IBS, Crohn's disease, ulcerative colitis, functional dyspepsia, diarrhea, gastroenteritis, urinary tract infection, or neonatal meningitis by reducing the level of Enterobacteriaceae in the gastrointestinal tract.

Methods of administration

Preferably, the compositions of the invention are intended to be administered to the gastrointestinal tract to provide delivery and / or partial or complete colonization of the intestine with the bacterial strain of the invention. Typically, the compositions of the invention are administered orally, but

- 7 037069 they can be administered rectally, intranasally or by buccal or sublingual routes.

In certain embodiments, the compositions of the invention may be administered as a foam, spray, or gel.

In certain embodiments, the compositions of the invention can be administered as a suppository, such as a rectal suppository, for example, as a composition with cocoa butter, synthetic hard fat (e.g. suppocire, witepsol), glycerol gelatin, polyethylene glycol or glycerol soap (soap glycerin).

In certain embodiments, the composition of the invention is administered into the gastrointestinal tract through a tube such as a nasogastric tube, orogastric tube, gastric tube, jejunostomy tube (J tube), percutaneous endoscopic gastrostomy (PEG), or an opening such as an opening in the chest wall cells that provide access to the stomach, jejunum, and other suitable access openings.

The compositions of the invention may be administered in a single dose, or they may be administered sequentially in a treatment regimen. In certain embodiments, the compositions of the invention are for daily administration. The examples demonstrate that administration provides successful colonization and clinical benefit in the treatment of diarrhea and / or constipation.

In certain embodiments, the compositions of the invention are administered regularly, for example daily, every other day, or weekly, over an extended period of time, such as for at least one week, two weeks, one month, two months, six months, or one year. The examples demonstrate that VL administration may not lead to permanent colonization of the intestine, therefore, regular administration over long periods of time can provide great therapeutic benefits.

In some embodiments, compositions of the invention are administered over 7 days, 14 days, 16 days, 21 days, or 28 days, or no more than 7 days, 14 days, 16 days, 21 days, or 28 days. For example, in some embodiments, compositions of the invention are administered over 16 days.

In certain embodiments of the invention, treatment in accordance with the invention is accompanied by an assessment of the patient's gut microbiota. Treatment can be repeated if delivery and / or partial or complete colonization with the strain of the invention is not achieved and thus no effect is observed, or treatment can be discontinued if delivery and / or partial or complete colonization is successful and the effect is observed.

In certain embodiments, a composition of the invention can be administered to a pregnant animal, eg, a mammal, such as a human, to prevent the development of diarrhea and / or constipation in a child in utero and / or after birth.

The compositions of the invention can be administered to a patient diagnosed with diarrhea and / or constipation, or a disease or condition associated with diarrhea and / or constipation, or identified as at risk of diarrhea and / or constipation. The compositions can also be administered as a prophylactic measure to prevent the development of diarrhea and / or constipation in a healthy patient.

The compositions of the invention can be administered to a patient who has been identified as having abnormal gut microbiota. For example, a patient may have decreased colonization of Blautia, and in particular Blautia hydrogenotrophica, Blautia stercoris or Blautia wexlerae, or no colonization thereof.

The compositions of the invention can be administered in the form of a food product, such as a nutritional supplement.

In general, the compositions of the invention are intended for the treatment of humans, although they may be used for the treatment of animals, including monogastric mammals such as poultry, pigs, cats, dogs, horses, or rabbits. The compositions of the invention may be useful for improving the growth and performance of animals. When administered to animals, gavage may be employed.

In some embodiments, the subject to which the composition is to be administered is an adult. In some embodiments, the subject to which the composition is to be administered is a human infant.

Compositions

In general, the composition of the invention contains bacteria. In preferred embodiments of the invention, the composition is prepared in lyophilized form. For example, the composition according to the invention may contain granules or gelatin capsules, for example hard gelatin capsules, containing the bacterial strain according to the invention.

Preferably, the composition according to the invention contains lyophilized bacteria. Lyophilization of bacteria is a well-established procedure and recommendations are given, for example, in [25-27]. Lyophilisate compositions can be particularly effective. In preferred embodiments, the compositions of the invention comprise lyophilized bacteria and are intended for the treatment of diarrhea and / or constipation associated with IBS.

- 8 037069

Alternatively, the composition of the invention may comprise a live, active bacterial culture. The examples demonstrate that the bacterial cultures of the invention are therapeutically effective.

In some embodiments, the bacterial strain in the composition of the invention has not been inactivated, for example, has not been inactivated by heat. In some embodiments, the bacterial strain in the composition of the invention has not been killed, for example, has not been killed by heat. In some embodiments, the bacterial strain in the composition of the invention has not been attenuated, such as not attenuated by heat. For example, in some embodiments, the bacterial strain in the composition of the invention has not been killed, inactivated, and / or weakened. For example, in some embodiments, the bacterial strain in the composition of the invention is live. For example, in some embodiments, the bacterial strain in the composition of the invention is viable. For example, in some embodiments, a bacterial strain in a composition of the invention is capable of partially or completely colonizing the intestine. For example, in some embodiments, the bacterial strain in the composition of the invention is viable and capable of partially or completely colonizing the intestine.

In some embodiments, the composition comprises a mixture of live bacterial strains and killed bacterial strains.

In preferred embodiments, the composition of the invention is encapsulated to allow delivery of the bacterial strain to the intestine. Encapsulation protects the composition from degradation prior to delivery to the target area by, for example, rupture due to chemical or physical attack, such as pressure, enzymatic activity, or physical degradation, which can be initiated by a change in pH. Any suitable encapsulation method can be used. Typical examples of encapsulation techniques include entrapment in a porous matrix, attachment or adsorption on solid support surfaces, self-aggregation by flocculation or with crosslinking agents, and mechanical retention behind a microporous membrane or microcapsule. Encapsulation guidelines that may be useful for preparing compositions of the invention are given, for example, in references [28-29].

The composition can be administered orally and can be in the form of a tablet, capsule or powder. Encapsulated products are preferred because Blautia are anaerobic. Other ingredients (such as, for example, vitamin C) can be included as oxygen scavengers and prebiotic substrates to improve delivery and / or partial or complete colonization and in vivo survival. Alternatively, the probiotic composition of the invention can be administered orally as a food or nutritional product, such as a fermented milk product based on milk or whey, or as a pharmaceutical product.

The composition can be formulated as a probiotic.

The composition of the invention comprises a therapeutically effective amount of a bacterial strain of the invention. A therapeutically effective amount of the bacterial strain is sufficient to have a beneficial effect on the patient. A therapeutically effective amount of the bacterial strain may be sufficient to provide delivery and / or partial or complete colonization of the patient's intestine.

A suitable daily dose of bacteria, for example, for an adult, may be from ^{about 1 x 10 3} to about 1 x 10 ¹¹ colony forming units (CFU); for example, from about ^{1 x 10 7} to about 1 x 10 ¹⁰ CFU; in another example, from about ^{1 x 10 6} to about 1 x 10 ¹⁰ CFU; in another example, from ^{about 1 x 10 7} to about 1 x 10 ¹¹ CFU; in another example, from ^{about 1 x 10 8} to about 1 x 10 ¹⁰ cfu; in another example, from about 1x 108 to about 1x 10 ¹¹ CFU.

In certain embodiments, the dose of bacteria is at least 10 ⁹ cells per day, such as at least 10 ¹⁰ , at least 10 ¹¹ , or at least 10 ¹² cells per day.

In certain embodiments, the composition comprises a bacterial strain in an amount of from about 1x10 ⁶ to about 1x10 ¹¹ CFU / g, based on the weight of the composition; for example, from about 1 x 108 to about 1 x 10 ¹⁰ cfu / g. The dose can be, for example, 1 g, 3 g, 5 g and 10 g.

Typically, a probiotic, such as a composition of the invention, is optionally combined with at least one suitable prebiotic compound. The prebiotic compound is usually a non-digestible carbohydrate such as an oligo- or polysaccharide or sugar alcohol that is not degraded or absorbed in the upper digestive tract. Known prebiotics include commercial products such as inulin and transgalacto-oligosaccharides.

In certain embodiments, the probiotic composition of this invention comprises a prebiotic compound in an amount of from about 1 to about 30% by weight based on the total weight of the composition (eg, 5 to 20% by weight). Carbohydrates can be selected from the group consisting of: fructo-oligosaccharides (or FOS), short-chain fructooligosaccharides, inulin, isomaltoligosaccharides, pectins, xylo-oligosaccharides (or XOS), chitosanoligosaccharides (or COS), modified aravable and aravable glucans, starches, poly

- 9 037069 dextrose, D-tagatose, acacia fibers, carob, oat and citrus fibers. In one aspect, the prebiotics are short-chain flucto-oligosaccharides (referred to below for simplicity cc-FOS (FOSs-cs)); these cc-FOS are non-digestible carbohydrates, usually obtained by the conversion of beet sugar, and contain a sucrose molecule to which three glucose molecules are linked.

Compositions of the invention may contain pharmaceutically acceptable excipients or carriers. Examples of such suitable excipients are given in reference [30]. Suitable carriers or diluents for therapeutic use are well known in the pharmaceutical industry and are described, for example, in the source [31]. Examples of suitable carriers include lactose, starch, glucose, methylcellulose, magnesium stearate, mannitol, sorbitol, and the like. Examples of suitable diluents include ethanol, glycerin, and water. The choice of pharmaceutical carrier, excipient or diluent can be made in accordance with the intended route of administration and standard pharmaceutical practice. The pharmaceutical compositions may contain, as or in addition to the carrier, excipient or diluent, any suitable binder (s), lubricant (s), suspending agent (s), coating agent (s), solubilizer (s). Examples of suitable binders include starch, gelatin, natural sugars such as glucose, anhydrous lactose, bulk lactose, betalactose, corn sugars, natural and synthetic gums such as gum arabic, tragacanth or sodium alginate, carboxymethyl cellulose and polyethylene glycol. Examples of suitable lubricants include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like. Preservatives, stabilizers, colorants, and even flavoring agents can be included in the pharmaceutical composition. Examples of preservatives include sodium benzoate, sorbic acid, cysteine, and p-hydroxybenzoic acid esters, for example, in some embodiments, the preservative is selected from sodium benzoate, sorbic acid, and poxybenzoic acid esters. Antioxidants and suspending agents can also be used. An additional example of a suitable carrier is sucrose. An additional example of a preservative is cysteine.

The compositions of the invention can be formulated as a food product. For example, a food product can provide, in addition to the therapeutic effect of the invention, a nutritional benefit such as a nutritional supplement. Likewise, the food composition can be formulated to enhance the taste of the composition of the invention or make the composition more palatable by being more similar to a conventional food ingredient than to a pharmaceutical composition. In certain embodiments, the composition of the invention is formulated as a milk-based product. The term milk-based product means any liquid or semi-solid milk or whey product having a different fat content. A milk-based product can be, for example, cow's milk, goat's milk, sheep's milk, skim milk, whole milk, milk reconstituted from milk powder and whey without any processing, or a processed product such as yogurt, curdled milk, curdled milk , sour milk, sour whole milk, buttermilk, and other sour dairy products. Another important group includes dairy beverages such as whey drinks, fermented milk products, condensed milk, breast milk substitutes (infant) or baby milk (baby milks); flavored milks, ice cream; milk-containing foods such as sweets.

In some embodiments, the compositions of the invention contain one or more bacterial strains of the genus Blautia and do not contain bacteria of any other genus, or contain only a minimal (de minimis) or biologically insignificant amount of bacteria of another genus.

In certain embodiments, the compositions of the invention comprise a single bacterial strain or species and do not contain any other bacterial strains or species. Such compositions may contain only minimal or biologically insignificant amounts of other bacterial strains or species. Such compositions can be a culture that is substantially free of other species of organism. In some embodiments, such compositions may be a lyophilisate substantially free of other types of organisms.

In certain embodiments, the compositions of the invention comprise one or more bacterial strains of the genus Blautia, such as Blautia hydrogenotrophica, and do not contain any other bacterial genus or contain only minimal or biologically insignificant amounts of bacteria of another genus. In certain embodiments, the compositions of the invention contain one Blautia species, eg, Blautia hydrogenotrophica, and do not contain any other bacterial species or contain only minimal or biologically insignificant amounts of another species of bacteria. In certain embodiments, the compositions of the invention comprise a single Blautia strain, such as Blautia hydrogenotrophica, and do not contain any other bacterial strains or species, or contain only minimal or biologically insignificant amounts of bacteria of a different strain or species.

In some embodiments, the compositions of the invention comprise more than one bacterium

- 10 037069 aal strain or species. For example, in some embodiments, compositions of the invention comprise more than one strain from the same species (e.g., more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30 , 35, 40 or 45 strains) and optionally does not contain any other species of bacteria. In some embodiments, compositions of the invention comprise less than 50 strains from the same species (e.g., less than 45, 40, 35, 30, 25, 20, 15, 12, 10, 9, 8, 7, 6, 5, 4 or 3 strains) and optionally does not contain any other species of bacteria. In some embodiments, the compositions of the invention comprise 1-40, 1-30, 1-20, 1-19, 1-18, 1-15, 1-10, 1-9, 1-8, 1-7, 1- 6, 1-5, 1-4, 1-3, 1-2, 2-50, 2-40, 2-30, 2-20, 2-15, 2-10, 2-5, 6-30, 6-15, 16-25, or 31-50 strains are from the same species and, optionally, do not contain bacteria of any other species. In some embodiments, compositions of the invention comprise more than one species from the same genus (e.g., more than 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 17, 20, 23 , 25, 30, 35 or 40 species) and optionally does not contain bacteria of any other kind. In some embodiments, compositions of the invention comprise less than 50 species from the same genus (e.g., less than 50, 45, 40, 35, 30, 25, 20, 15, 12, 10, 8, 7, 6, 5, 4 or 3) and, optionally, does not contain bacteria of any other kind. In some embodiments, the compositions of the invention comprise 1-50, 1-40, 1-30, 1-20, 1-15, 1-10, 1-9, 1-8, 1-7, 1-6, 1- 5, 1-4, 1-3, 1-2, 2-50, 2-40, 230, 2-20, 2-15, 2-10, 2-5, 6-30, 6-15, 16- 25 or 31-50 species are from the same genus and optionally do not contain bacteria of any other genus. The invention includes any combination of the above.

In some embodiments, the composition comprises a microbial consortium. For example, in some embodiments, the composition comprises a Blautia bacterial strain in a microbial consortium. For example, in some embodiments, the Blautia bacterial strain is present in combination with one or more (e.g., at least 2, 3, 4, 5, 10, 15, or 20) other bacterial strains from other genera with which it can exist symbiotically in vivo in the intestine. For example, in some embodiments, the composition comprises a Blautia hydrogenotrophica bacterial strain in combination with a different genus bacterial strain. In some embodiments, the microbial consortium comprises two or more bacterial strains obtained from a faecal sample from one organism, such as a human. In some embodiments, the microbial consortium does not occur together in nature. For example, in some embodiments, the microbial consortium comprises bacterial strains obtained from fecal samples from at least two different organisms. In some embodiments, the two different organisms are of the same species, such as two different humans. In some embodiments, the two different organisms are a human infant and an adult. In some embodiments, the two different organisms are a human and a non-human mammal.

In some embodiments, the composition of the invention further comprises a bacterial strain having the same safety and therapeutic efficacy characteristics as the Blautia hydrogenotrophica strain deposited with accession number DSM 10507/14294, but not the Blautia hydrogenotrophica strain deposited with accession number DSM 10507 / 14294, or non-Blautia hydrogenotrophica, or non-Blautia.

In some embodiments, in which the composition of the invention contains more than one bacterial strain, species or genus, individual bacterial strains, species or genera can be designed for separate, simultaneous or sequential administration. For example, the composition can contain all of more than one bacterial strain, species or genus, or bacterial strains, species or genera can be stored separately and administered separately, simultaneously or sequentially. In some embodiments, more than one of the bacterial strains, species, or genera are kept separate but mixed prior to use.

In some embodiments, the bacterial strain for use in the invention is obtained from the faeces of an adult. In some embodiments in which the composition of the invention contains more than one bacterial strain, all bacterial strains are obtained from adult feces or, if other bacterial strains are present, they are present in only minimal amounts. The bacteria can be grown in culture after being obtained from adult feces and used in the composition of the invention.

In some embodiments, one or more Blautia bacterial strains is / are the only therapeutically active agent (s) in the composition of the invention. In some embodiments, the bacterial strain (s) in the composition is / are the only therapeutically active agent (s) in the composition of the invention.

Compositions for use in accordance with the invention may or may not require marketing authorization.

In certain embodiments, the invention provides a pharmaceutical composition as defined above, wherein said bacterial strain is lyophilized. In certain embodiments, the invention provides a pharmaceutical composition as defined above wherein said bacterial strain has been spray dried. In certain embodiments, the invention provides the above pharmaceutical composition

- 11 037069 in which the bacterial strain has been lyophilized or spray dried and in which it is alive. In certain embodiments, the invention provides a pharmaceutical composition as defined above in which the bacterial strain has been lyophilized or spray dried and in which it is viable. In certain embodiments, the invention provides the aforementioned pharmaceutical composition in which the bacterial strain is lyophilized or spray dried and in which it is capable of partially or completely colonizing the intestine. In certain embodiments, the invention provides the aforementioned pharmaceutical composition in which the bacterial strain is lyophilized or spray dried and in which it is viable and capable of partially or completely colonizing the intestine.

In some cases, the lyophilized or spray dried bacterial strain is reconstituted prior to administration. In some cases, reconstitution is carried out using a diluent as described herein.

Compositions of the invention may contain pharmaceutically acceptable excipients, diluents or carriers.

In certain embodiments, the invention provides a pharmaceutical composition comprising: a bacterial strain of the invention; and a pharmaceutically acceptable excipient, carrier or diluent; moreover, the amount of bacterial strain is sufficient to treat the disorder when administered to a subject in need thereof; and the disorder is diarrhea and / or constipation such as diarrhea and / or constipation associated with Crohn's disease, ulcerative colitis, or more preferably IBS.

In certain embodiments, the invention provides the aforementioned pharmaceutical composition, wherein the amount of the bacterial strain is from about 1x10 ³ to about 1x1011 CFU per gram of weight of the composition.

In certain embodiments, the invention provides a pharmaceutical composition as defined above, the composition being administered in a dose of 1 g, 3 g, 5 g, or 10 g.

In certain embodiments, the invention provides a pharmaceutical composition as defined above, the composition being administered in a manner selected from the group consisting of oral, rectal, subcutaneous, nasal, buccal, and sublingual.

In certain embodiments, the invention provides a pharmaceutical composition as defined above comprising a carrier selected from the group consisting of lactose, starch, glucose, methylcellulose, magnesium stearate, mannitol, and sorbitol.

In certain embodiments, the invention provides the aforementioned pharmaceutical composition comprising a diluent selected from the group consisting of ethanol, glycerol and water.

In certain embodiments, the invention provides the above pharmaceutical composition comprising an excipient selected from the group consisting of starch, gelatin, glucose, anhydrous lactose, bulk lactose, beta-lactose, corn sugar, gum arabic, tragacanth, sodium alginate, carboxymethylcellulose, polyethylene sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate and sodium chloride.

In certain embodiments, the invention provides the aforementioned pharmaceutical composition further comprising at least one of a preservative, an antioxidant, and a stabilizer.

In certain embodiments, the invention provides the above pharmaceutical composition comprising a preservative selected from the group consisting of sodium benzoate, sorbic acid, and p-hydroxybenzoic acid esters.

In certain embodiments, a pharmaceutical composition of the invention is provided that does not contain any minerals or, more specifically, does not contain any metals with an atomic number greater than 33, for example, does not contain any minerals from the group consisting of selenium, molybdenum, tungsten , selenium compounds, molybdenum compounds and tungsten compounds.

In certain embodiments, the invention provides a pharmaceutical composition as defined above, wherein said bacterial strain is lyophilized.

In certain embodiments, the invention provides the above pharmaceutical composition, wherein the composition, when stored in a sealed container at about 4 ° C or about 25 ° C and placed in an atmosphere of 50% relative humidity, retains at least 80% of the bacterial strain when measured in colony forming units after a period of time of at least about: 1 month, 3 months, 6 months, 1 year, 1.5 years, 2 years, 2.5 years or 3 years.

In some embodiments, the composition of the invention is provided in a hermetically sealed container containing the composition described herein. In some varieties

- 12 037069 anth of realization The hermetically sealed container is a sachet or bottle. In some embodiments, a composition of the invention is provided in a syringe containing a composition as described herein.

The composition of this invention may, in some embodiments, be provided as a pharmaceutical composition. For example, the composition can be in the form of a tablet or capsule. In some embodiments, the capsule is a gel-cap.

In some embodiments, the compositions of the invention are administered orally. Oral administration may include swallowing, whereby the formulation enters the gastrointestinal tract, and / or buccal, lingual, or sublingual administration, in which the formulation enters the bloodstream directly from the mouth.

Pharmaceutical compositions suitable for oral administration include solid plugs, solid microdispersed materials, semi-solid and liquid materials (including multiphase or dispersed systems) such as tablets; soft or hard capsules containing multiparticulate materials or nanoparticulate materials, liquids (eg, aqueous solutions), emulsions or powders; sucking tablets (including those filled with liquid); gum; gels; fast-dispersing dosage forms; films; caplets (ovules); sprays; and buccal / mucoadhesive patches.

In some embodiments, the pharmaceutical composition is an enteric formulation, i.e. a gastro-resistant composition (eg, resistant to gastric pH), suitable for delivering a composition of the invention to the intestine by oral administration. Enteric compositions can be especially useful in cases where the bacteria or other component of the composition is acid sensitive, for example, prone to degradation under conditions of the stomach.

In some embodiments, the enteric composition comprises an enteric coating. In some embodiments, the composition is an enteric coated dosage form. For example, the composition can be an enteric coated tablet or an enteric coated capsule and the like. The enteric coating may be a conventional enteric coating, for example a conventional tablet, capsule, and the like coating for oral delivery. The composition may include a film coating, such as a thin film layer of an enteric polymer, such as an acid insoluble polymer.

In some embodiments, the enteric composition is enteric in nature, eg, gastro-resistant, without the need for an enteric coating. Thus, in some embodiments, the composition is an enteric composition that does not contain an enteric coating. In some embodiments, the composition is a capsule made from a thermogelling material. In some embodiments, the thermosetting material is a cellulosic material such as methylcellulose, hydroxymethylcellulose, or hydroxypropylmethylcellulose (HPMC). In some embodiments, the capsule includes a shell that does not contain any film-forming polymer. In some embodiments, the capsule includes a shell, and the shell contains hydroxypropyl methylcellulose and does not contain any film-forming polymer (see, for example, [32]). In some embodiments, the composition is an inherently enteric capsule (eg, Vcaps® from Capsugel).

In some embodiments, the composition is a soft capsule. Soft capsules are capsules which can, due to the addition of softeners such as, for example, glycerin, sorbitol, maltitol and polyethylene glycols, present in the capsule shell, have a certain elasticity and deformability. Soft capsules can be prepared, for example, based on gelatin or starch. Gelatin soft capsules are commercially available from various suppliers. Depending on the route of administration, such as, for example, oral or rectal, soft capsules can have various shapes, they can be, for example, round, oval, oblong or torpedo-shaped. Soft capsules can be prepared by conventional methods such as, for example, using the Scherer process (pressing method), the Accogel process (pressing method), or a drip or blowing process.

Cultivation methods

Bacterial strains for use in this invention can be cultured using standard microbiological techniques, as described, for example, in references [33-35].

The solid or liquid medium used for the culture can be, for example, YCFA (yeast extract-casein fatty acid hydrolyzate) agar or YCFA medium. YCFA medium may contain (per 100 ml, approximate values): casiton (1.0 g), yeast extract (0.25 g), NaHCO ₃ (0.4 g), cysteine (0.1 g), K ₂ HPO ₄ (0.045 g), KH2PO4 (0.045 g), NaCl (0.09 g), (NH ₄ ) ₂ SO ₄ (0.09 g), MgSO ₄ -7H ₂ O (0.009 g), CaCl ₂ (0.009 g ), resazurin (0.1 mg), hemin (1 mg), biotin (1 μg), cobalamin (1 μg), p

- 13 037069 aminobenzoic acid (3 μg), folic acid (5 μg) and pyridoxamine (15 μg).

General Provisions

The practice of this invention will employ, unless otherwise indicated, conventional methods of chemistry, biochemistry, molecular biology, immunology, and pharmacology available to those skilled in the art. Such techniques are described in detail in the literature. See, for example, sources [36-43], etc.

The term “containing” encompasses an inclusive as well as consisting of, for example, a composition containing X, may consist exclusively of X, or may further include something, such as X + Y.

The term about in relation to the numerical value of x is optional and means, for example, x ± 10%.

An expression as such does not completely exclude, for example, a composition that is essentially free of Y may be completely free of Y. If necessary, an expression as such can be omitted from the claims (definition of the invention).

References to percent sequence identity between two nucleotide sequences means that when aligned, the percentage of nucleotides when comparing the two sequences will be the same. Such alignment and percent homology or sequence identity can be determined using computer programs known to those skilled in the art, such as those described in section 7.7.18 of the source [44]. The preferred alignment is determined by the Smith-Waterman homology search algorithm using an affine gap search with a gap opening penalty of 12 and a gap extension penalty of 2, the BLOSUM 62 matrix. The Smith-Waterman homology search algorithm is disclosed in [ 45].

Unless specifically indicated, a multi-step process or method may include additional steps at the beginning or end of the process, or may include additional intermediate steps. Also, the steps can be combined, skipped or performed in an alternative order, if necessary.

Various embodiments of the invention are described herein. It should be understood that the features specified in each embodiment may be combined with other specified features to provide additional options for implementation. In particular, embodiments identified herein as suitable, typical, or preferred may be combined with each other (unless they are mutually exclusive).

Modes for Carrying Out the Invention

Example 1 Changes in Patient Symptoms During Phase I Clinical Trials

Phase I clinical trials were conducted in which Blautia hydrogenotrophica (Blautix, strain deposited with accession number DSM 10507 as well as accession number DSM 14294) was administered to patients with irritable bowel syndrome (IBS). Patients were administered Blautix over a dosing period (days 1-16) with a washout period on days 19-23. Blautix has been found to be safe and well tolerated. Four symptoms were monitored, two of which were diarrhea and constipation. The studies recorded whether patients experienced an improvement, no change, or worsening of each of these symptoms. Results for patients receiving Blautix were compared with those obtained for patients receiving placebo. Symptom control was performed at three time points: day 1, day 15/16 and at the end of the trial. The results are shown in FIG. 1 and 2.

When comparing patient reported symptoms on day 16 with baseline on day 1, 82% of 17 IBS patients treated with Blautix reported an improvement in symptoms (FIG. 1). Improvement in symptoms, two of which were diarrhea and constipation, is evidence in favor of using Blautix to treat or prevent diarrhea and / or constipation.

50% of patients receiving placebo reported an improvement in symptoms (Fig. 1). High response rates to placebo are well established in clinical trials of IBS. Xifaxan was recently approved for the treatment of IBS based on significantly less improvement over placebo (see: http://www.accessdata.fda.gov/spl/data/5ab6fceb4d22-4480-81fc-8bc28c16770d/5ab6fceb -4d22-4480-81fc-8bc28c 16770d.xml).

Based on the description provided here, a worsening of symptoms was expected at study completion (days 19-23) compared to symptoms at completion of dosing (day 16). This worsening of symptoms was observed in phase I clinical trials: 41% of IBS patients reported worsening of symptoms after discontinuation of Blautix (FIG. 2). Worsening of symptoms, two of which are diarrhea and constipation, after discontinuation of Blautix thus also supports the use of Blautix in the treatment or prevention of diarrhea and / or constipation.

Example 2. Results of a breath hydrogen test

Respiratory hydrogen levels are a biomarker of Blautix activity - its mechanism of action (MoA) is linked to the metabolism of endogenous H2 to produce acetate. People were injected with lactulose and hydrogen (H ₂ ) levels (C _max ) were determined in samples taken at four time points: day 1, day 2,

- 14 037069 day 15 and day 16. The uncorrected results for hydrogen were converted to corrected results for hydrogen.

Some patients were excluded from the analysis. There were three reasons why subjects were not included in the hydrogen breath test analysis:

1) they had a CMAX hydrogen breath test <20 on one of four days of sampling and were therefore considered not responding to the test;

2) they were methane producers (defined as producing more methane than hydrogen according to the results of a breath test), this affects the hydrogen response; and / or

3) a highly deviating value (232 ppm) was obtained in the hydrogen breath test. Subjects 3.12 (Blautix), 3.24 (Blautix), 4.07 (Blautix) were excluded as not responding. Subjects 3.03 (Blautix) and 3.08 (placebo) were excluded as methane producers (4.07, indicated as excluded above, was also a methane producer). Subject 4.09 (placebo) was excluded due to highly outliers.

The corrected hydrogen analysis results for the end of the dosing period (days 15/16) were compared to baseline (days 1/2). In 10 of 12 patients (83%) treated with Blautix, there was a decrease in hydrogen levels during this period (Fig. 3a and 3c). In contrast, 3 out of 6 (50%) placebo patients had decreased hydrogen levels (Figs. 3b and 3c). These percentages are similar to the percentages of patients showing improvement in symptoms after treatment with Blautix or placebo.

FIG. 4 shows uncorrected and corrected hydrogen results for the Blautix (Verum (active ingredient formulation)) group, together with a statistical analysis of the results. It was found that the mean values for the uncorrected and corrected H2 differ between day 1 and day 15. After 13.5 days of treatment was detected a statistically significant (p <0,05) decrease in _H2 breath test definition Cmax after stimulation lactulose. In contrast, in the placebo group, the mean values for day 1 and day 15 were equivalent (p> 0.05) (Fig. 5). Thus, the mean in the treatment group (also called the VERUM group) declines between day 1 and day 15, while the mean in the placebo group is equivalent on day 1 and day 15 for both uncorrected hydrogen test and corrected test results. hydrogen (Fig. 6).

Example 3. Tests for stability

The composition described herein, containing at least one bacterial strain described herein, is stored in a sealed container at a temperature of 25 ° C or 4 ° C and the container is placed in an atmosphere with a relative humidity of 30%, 40%, 50%, 60%, 70%, 75%, 80%, 90% or 95%. After 1 month, 2 months, 3 months, 6 months, 1 year, 1.5 years, 2 years, 2.5 years or 3 years, at least 50%, 60%, 70%, 80% or 90 should be retained % bacterial strain as measured by colony forming units according to standard protocols.

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Sequences

SEQ ID NO: 1 (Gene 16s ribosomal RNA strain GAM6-1 Blautia stercoris, partial sequence - HM626177)

1 tgcaagtcga gcgaagcgct tacgacagaa ccttcggggg aagatgtaag ggactgagcg 61 gcggacgggt gagtaacgcg tgggtaacct gcctcataca gggggataac agttggaaac 121 ggctgctaat accgcataag cgcacggtat cgcatgatac agtgtgaaaa actccggtgg 181 tatgagatgg acccgcgtct gattagctag ttggaggggt aacggcccac caaggcgacg 241 atcagtagcc ggcctgagag ggtgaacggc cacattggga ctgagacacg gcccagactc 301 ctacgggagg cagcagtggg gaatattgca caatggggga aaccctgatg cagcgacgcc 361 gcgtgaagga agaagtatct cggtatgtaa acttctatca gcagggaaga aaatgacggt 421 acctgactaa gaagccccgg ctaactacgt gccagcagcc gcggtaatac gtagggggca 481 agcgttatcc ggatttactg ggtgtaaagg gagcgtagac ggaagagcaa gtctgatgtg 541 aaaggctggg gcttaacccc aggactgcat tggaaactgt ttttcttgag tgccggagag 601 gtaagcggaa ttcctagtgt agcggtgaaa tgcgtagata ttaggaggaa caccagtggc 661 gaaggcggct tactggacgg taactgacgt tgaggctcga aagcgtgggg agcaaacagg 721 attagatacc ctggtagtcc acgccgtaaa cgatgaatac taggtgttgg ggagcaaagc 781 tcttcggtgc cgcagcaaac gcaataagta ttccacctgg ggagtacgtt cgcaagaatg 841 aaactcaaag gaattgacgg ggacccgcac aagcggtgga gcatgtggtt taattcgaag 901 caacgcgaag aaccttacca agtcttgaca tcgatctgac cggttcgtaa tggaaccttt 961 ccttcgggac agagaagaca ggtggtgcat ggttgtcgtc agctcgtgtc gtgagatgtt 1021 gggttaagtc ccgcaacgag cgcaacccct atcctcagta gccagcaggt gaagctgggc 1081 actctgtgga gactgccagg gataacctgg aggaaggcgg ggacgacgtc aaatcatcat 1141 gccccttatg atttgggcta cacacgtgct acaatggcgt aaacaaaggg aagcgagccc 1201 gcgaggggga gcaaatccca aaaataacgt cccagttcgg actgcagtct gcaactcgac 1261 tgcacgaagc tggaatcgct agtaatcgcg aatcagaatg tcgcggtgaa

tacgttcccg

1321 ggtcttgtac acaccgcccg tcacaccatg ggagtcagta acgcccgaag tc

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SEQ ID NO: 2 (16s ribosomal RNA gene of WAL 14507 Blautia wexlerae strain, partial sequence - EF036467)

1 caagtcgaac gggaattant ttattgaaac ttcggtcgat ttaatttaat tctagtggcg 61 gacgggtgag taacgcgtgg gtaacctgcc ttatacaggg ggataacagt cagaaatggc 121 tgctaatacc gcataagcgc acagagctgc atggctcagt gtgaaaaact ccggtggtat 181 aagatggacc cgcgttggat tagcttgttg gtggggtaac ggcccaccaa ggcgacgatc 241 catagccggc ctgagagggt gaacggccac attgggactg agacacggcc cagactccta 301 cgggaggcag cagtggggaa tattgcacaa tgggggaaac cctgatgcag cgacgccgcg 361 tgaaggaaga agtatctcgg tatgtaaact tctatcagca gggaagatag tgacggtacc 421 tgactaagaa gccccggcta actacgtgcc agcagccgcg gtaatacgta gggggcaagc 481 gttatccgga tttactgggt gtaaagggag cgtagacggt gtggcaagtc tgatgtgaaa 541 ggcatgggct caacctgtgg actgcattgg aaactgtcat acttgagtgc cggaggggta 601 agcggaattc ctagtgtagc ggtgaaatgc gtagatatta ggaggaacac cagtggcgaa 661 ggcggcttac tggacggtaa ctgacgttga ggctcgaaag cgtggggagc aaacaggatt 721 agataccctg gtagtccacg ccgtaaacga tgaataacta ggtgtcgggt ggcaaagcca 781 ttcggtgccg tcgcaaacgc agtaagtatt ccacctgggg agtacgttcg caagaatgaa 841 actcaaagga attgacgggg acccgcacaa gcggtggagc atgtggttta attcgaagca 901 acgcgaagaa ccttaccaag tcttgacatc cgcctgaccg atccttaacc ggatctttcc 961 ttcgggacag gcgagacagg tggtgcatgg ttgtcgtcag ctcgtgtcgt gagatgttgg 1021 gttaagtccc gcaacgagcg caacccctat cctcagtagc cagcatttaa ggtgggcact 1081 ctggggagac tgccagggat aacctggagg aaggcgggga tgacgtcaaa tcatcatgcc 1141 ccttatgatt tgggctacac acgtgctaca atggcgtaaa caaagggaag cgagattgtg 1201 agatggagca aatcccaaaa ataacgtccc agttcggact gtagtctgca acccgactac 1261 acgaagctgg aatcgctagt aatcgcggat cagaatgccg cggtgaatac gttcccgggt 1321 cttgtacaca ccgcccgtca caccatggga gtcagtaacg cccgaagtca gtgacctaac 1381 tgcaaagaag gagctgccga aggcgggacc gatgactggg gtgaagtcgt

aacaaggt

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SEQ ID NO: 3 (consensus sequence 16s pPHK strain 830 Blautia stercoris) TTTKGTCTGGCTCAGGATGAACGCTGGCGGCGTGCTTAACACATGCAAGTCGAGCGAAGCGCTT ACGACAGAACCTTCGGGGGAAGATGTAAGGGACTGAGCGGCGGACGGGTGAGTAACGCGTGGGT AACCTGCCTCATACAGGGGGATAACAGTTGGAAACGGCTGCTAATACCGCATAAGCGCACAGTA TCGCATGATACAGTGTGAAAAACTCCGGTGGTATGAGATGGACCCGCGTCTGATTAGCTAGTTG GAGGGGTAACGGCCCACCAAGGCGACGATCAGTAGCCGGCCTGAGAGGGTGAACGGCCACATTG GGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGGGAA ACCCTGATGCAGCGACGCCGCGTGAAGGAAGAAGTATCTCGGTATGTAAACTTCTATCAGCAGG GAAGAAAATGACGGTACCTGACTAAGAAGCCCCGGCTAACTACGTGCCAGCAGCCGCGGTAATA CGTAGGGGGCAAGCGTTATCCGGATTTACTGGGTGTAAAGGGAGCGTAGACGGAAGAGCAAGTC TGATGTGAAAGGCTGGGGCTTAACCCCAGGACTGCATTGGAAACTGTTTTTCTTGAGTGCCGGA GAGGTAAGCGGAATTCCTAGTGTAGCGGTGAAATGCGTAGATATTAGGAGGAACACCAGTGGCG AAGGCGGCTTACTGGACGGTAACTGACGTTGAGGCTCGAAAGCGTGGGGAGCAAACAGGATTAG ATACCCTGGTAGTCCACGCCGTAAACGATGAATACTAGGTGTTGGGGAGCAAAGCTCTTCGGTG CCGCAGCAAACGCAATAAGTATTCCACCTGGGGAGTACGTTCGCAAGAATGAAACTCAAAGGAA TTGAC GGGGACCCGCACAAGCGGTGGAGCATGTGGTTTATTCGAAGCAACGCGAAGAACCTTAC CAAGTCTTGACATCGATCTGACCGGTTCGTAATGGAACCTTTCCTTCGGGACAGAGAAGACAGG TGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAAC CCCTATCGTCAGTAGCCAGCAGGTAAAGCTGGGCACTCTGAGGAGACTGCCAGGGATAACCTGG AGGAAGGCGGGGACGACGTCAAATCATCATGCCCCTTATGATTTGGGCTACACACGTGCTACAA TGGCGTAAACAAAGGGAAGCGAGCCCGCGAGGGGGAGCAAATCCCAAAAATAACGTCCCAGTTC GGACTGCAGTCTGCAACTCGACTGCACGAAGCTGGAATCGCTAGTAATCGCGAATCAGAATGTC GCGGTGAATACGTTCCCGGGTCTTGTACACACCGCCCGTCACACCATGGGAGTCAGTAACGCCC GAAGTCAGTGACCCAACCTTAGGGAGGGAGCTGCCGAAGGCGGGATTGATAACTGGGGTGAAGT CTAGGGGGT

SEQ ID NO: 4 (consensus sequence 16s pPHK strain MRX008 Blautia wexlerae) TTCATTGAGACTTCGGTGGATTTAGATTCTATTTCTAGTGGCGGACGGGTGAGTAACGCGTGGG TAACCTGCCTTATACAGGGGGATAACAGTCAGAAATGGCTGCTAATACCGCATAAGCGCACAGA GCTGCATGGCTCAGTGTGAAAAACTCCGGTGGTATAAGATGGACCCGCGTTGGATTAGCTTGTT GGTGGGGTAACGGCCCACCAAGGCGACGATCCATAGCCGGCCTGAGAGGGTGAACGGCCACATT GGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGGGA AACCCTGATGCAGCGACGCCGCGTGAAGGAAGAAGTATCTCGGTATGTAAACTTCTATCAGCAG GGAAGATAGTGACGGTACCTGACTAAGAAGCCCCGGCTAACTACGTGCCAGCAGCCGCGGTAAT ACGTAGGGGGCAAGCGTTATCCGGATTTACTGGGTGTAAAGGGAGCGTAGACGGTGTGGCAAGT CTGATGTGAAAGGCATGGGCTCAACCTGTGGACTGCATTGGAAACTGTCATACTTGAGTGCCGG AGGGGTAAGCGGAATTCCTAGTGTAGCGGTGAAATGCGTAGATATTAGGAGGAACACCAGTGGC GAAGGCGGCTTACTGGACGGTAACTGACGTTGAGGCTCGAAAGCGTGGGGAGCAAACAGGATTA GATACCCTGGTAGTCCACGCCGTAAACGATGAATACTAGGTGTCNGGGGAGCATGGCTCTTCGG TGCCGTCGCAAACGCAGTAAGTATTCCACCTGGGGAGTACGTTCGCAAGAATGAAACTCAAAGG AATTGACGGGGACCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCT TAC CAAGTCTTGACATCCGCCTGACCGATCCTTAACCGGATCTTTCCTTCGGGACAGGCGAGAC AGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGC AACCCCTATCCTCAGTAGCCAGCATTTAAGGTGGGCACTCTGGGGAGACTGCCAGGGATAACCT GGAGGAAGGCGGGGATGACGTCAAATCATCATGCCCCTTATGATTTGGGCTACACACGTGCTAC AATGGCGTAAACAAAGGGAAGCGAGATCGTGAGATGGAGCAAATCCCAAAAATAACGTCCCAGT TCGGACTGTAGTCTGCAACCCGACTACACGAAGCTGGAATCGCTAGTAATCGCGGATCAGAATG CCGCGGTGAATACGTTCCCGGGTCTTGTACACACCGCCCGTCACACCATGGGAGTCAGTAACGC

СCGAAGTCAGTGAGСTAACTGCAAAGAAGGAGСTGСCGAA

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SEQ ID NO: 5 (Gene 16s ribosomal RNA strain S5a36 Blautia hydrogenotrophica, partial sequence - X95624.1)

1 gatgaacgct c jgcggcgtgc ttaacacatg caagtcgaac gaagcgatag agaacggaga 61 tttcggttga agttttctat tgactgagtg gcggacgggt gagtaacgcg tgggtaacct 121 gccctataca gggggataac agttagaaat gactgctaat accgcataag cgcacagctt 181 cgcatgaagc ggtgtgaaaa actgaggtgg tataggatgg acccgcgttg gattagctag 241 ttggtgaggt aacggcccac caaggcgacg atccatagcc ggcctgagag ggtgaacggc 301 cacattggga ctgagacacg gcccaaactc ctacgggagg cagcagtggg gaatattgca 361 caatggggga aaccctgatg cagcgacgcc gcgtgaagga agaagtatct cggtatgtaa 421 acttctatca gcagggaaga aagtgacggt acctgactaa gaagccccgg ctaattacgt 481 gccagcagcc gcggtaatac gtaaggggca agcgttatcc ggatttactg ggtgtaaagg 541 gagcgtagac ggtttggcaa gtctgatgtg aaaggcatgg gctcaacctg tggactgcat 601 tggaaactgt cagacttgag tgccggagag gcaagcggaa ttcctagtgt agcggtgaaa 661 tgcgtagata ttaggaggaa caccagtggc gaaggcggcc tgctggacgg taactgacgt 721 tgaggctcga aagcgtgggg agcaaacagg attagatacc ctggtagtcc acgctgtaaa 781 cgatgaatac taggtgtcgg gtggcaaagc cattcggtgc cgcagcaaac gcaataagta 841 ttcccacctg gggagtacgt tcgcaagaat gaaactcaaa ggaattgacg gggacccgca 901 caagcggtgg agcatgtggt ttaattcgaa gcaacgcgaa gaaccttacc aaatcttgac 961 atccctctga ccgggaagta atgttccctt ttcttcggaa cagaggagac aggtggtgca 1021 tggttgtcgt cagctcgtgt cgtgagatgt tgggttaagt cccgcaacga gcgcaaccct 1081 tattcttagt agccagcagg tagagctggg cactctaggg agactgccag ggataacctg 1141 gaggaaggtg gggatgacgt caaatcatca tgccccttat gatttgggct acacacgtgc 1201 tacaatggcg taaacaaagg gaagcgaagg ggtgacctgg agcaaatctc aaaaataacg 1261 tctcagttcg gattgtagtc tgcaactcga ctacatgaag ctggaatcgc tagtaatcgc 1321 gaatcagaat gtcgcggtga atacgttccc gggtcttgta cacaccgccc gtcacaccat 1381 gggagtcagt aacgcccgaa gtcagtgacc caaccnaaag gagggagctg ccgaaggtgg 1441 gactgataac t iggggtga

- 19 037069

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Claims (11)

1. A pharmaceutical composition containing the bacterial strain Blautia hydrogenotrophica DSM 14294 having the sequence of the 16s pRNA gene SEQ ID NO: 5, and / or a bacterial strain of the genus Blautia having the sequence of the 16s pRNA gene at least 95% identical to SEQ ID NO: 5 to treat or prevent diarrhea and / or constipation in a subject. 2. A pharmaceutical composition according to claim 1, characterized in that diarrhea and / or constipation are associated - 21 037069 with irritable bowel syndrome IBS and / or inflammatory bowel disease. 3. A pharmaceutical composition according to claim 2, wherein the inflammatory bowel disease is a disease of the small intestine or the large intestine, in particular of the rectum. 4. A pharmaceutical composition according to any one of claims 1 to 3, characterized in that the inflammatory bowel disease is Crohn's disease or ulcerative colitis. 5. A pharmaceutical composition according to any one of claims 1 to 4, characterized in that the subject suffers from abdominal pain and / or flatulence. 6. Pharmaceutical composition according to any one of claims 1 to 5, characterized in that the bacterial strain of the genus Blautia has the sequence of the 16s pRNA gene at least 97%, 98%, 99%, 99.5% or 99.9% identical SEQ ID NO: 5. 7. A pharmaceutical composition according to any one of claims 1 to 6 for oral administration. 8. A pharmaceutical composition according to any one of claims 1 to 7, comprising one or more pharmaceutically acceptable excipients, in particular carriers. 9. A pharmaceutical composition according to any one of claims 1 to 8, characterized in that the bacterial strain is lyophilized or native. 10. A pharmaceutical composition according to any one of claims 1 to 9, characterized in that the composition contains one strain of bacteria of the genus Blautia. 11. A pharmaceutical composition according to any one of claims 1 to 8, characterized in that the composition comprises a bacterial strain Blautia hydrogenotrophica DSM 14294 having a 16s pRNA gene sequence SEQ ID NO: 5 and a bacterial strain of the genus Blautia having a 16s pRNA gene sequence according to at least 95% identical to SEQ ID NO: 5.