阅读说明：本技术 凝结芽孢杆菌和枯草芽孢杆菌用于预防和治疗功能性胃肠道病症 (Bacillus coagulans and bacillus subtilis for preventing and treating functional gastrointestinal tract diseases ) 是由 R·戴巴杰罗 W·提斯 T·万奴特塞尔 L·沃特斯 于 2020-03-27 设计创作，主要内容包括：本发明总体上涉及益生菌用于控制功能性胃肠道病症的用途。更具体地,本发明涉及凝结芽孢杆菌或枯草芽孢杆菌或者两种菌株的组合用于预防或治疗功能性胃肠道病症,例如功能性消化不良的用途。本发明还公开了凝结芽孢杆菌或枯草芽孢杆菌或者两种菌株的组合作为胃酸抑制治疗的替代疗法或辅助疗法的用途。(The present invention relates generally to the use of probiotics for the control of functional gastrointestinal disorders. More specifically, the present invention relates to the use of bacillus coagulans or bacillus subtilis or a combination of both strains for the prevention or treatment of functional gastrointestinal disorders, such as functional dyspepsia. The invention also discloses the use of bacillus coagulans or bacillus subtilis or a combination of the two strains as a replacement therapy or an adjuvant therapy for gastric acid inhibition therapy.)

1. A combination comprising bacillus subtilis and bacillus coagulans spores; for use in the prevention and/or treatment of a functional gastrointestinal disorder in a subject.

2. The combination for use according to claim 1, wherein the functional gastrointestinal disorder is selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or bloating.

3. The combination for use according to claim 1 or 2, wherein the functional gastrointestinal disorder is functional dyspepsia.

4. A combination for use as defined in any one of claims 1 to 3; wherein the ratio of the bacillus subtilis spores to the bacillus coagulans spores is 1:1, 1:2 or 2: 1.

5. A combination for use as defined in any one of claims 1 to 4; wherein the Bacillus subtilis spores are derived from strain MY02 deposited with BCCM under accession number LMG P-31319.

6. A combination for use as defined in any one of claims 1 to 5; wherein the Bacillus coagulans spores are derived from strain MY01 deposited with BCCM under accession number LMG P-31318.

7.A pharmaceutical composition for preventing and/or treating a functional gastrointestinal disorder in a subject; wherein the composition comprises a combination as defined in any one of claims 1 to 6, together with one or more pharmaceutically acceptable carriers, diluents or excipients.

8. The composition for use according to claim 7, wherein the functional gastrointestinal disorder is selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or bloating.

9. The composition for use according to claim 7 or 8, wherein the functional gastrointestinal disorder is functional dyspepsia.

10. A composition for use as defined in any one of claims 7 to 9; wherein the composition further comprises one or more ingredients selected from the group consisting of plant extracts, proteins, vitamins, minerals, and digestive enzymes.

11. A composition for use as defined in any one of claims 7 to 10; wherein the composition is in the form of a tablet, capsule, sachet, bar, or any other edible form including, but not limited to, functional food, juice, beverage, or candy.

12. The composition for use as defined in any one of claims 7 to 11, wherein said composition is a product of the dairy industry, the beverage industry or the pharmaceutical industry, or is a natural product.

13. The combination for use as defined in any one of claims 1 to 6, or the pharmaceutical composition for use as defined in any one of claims 7 to 12, wherein the subject is a human or an animal; preferably, wherein the subject is a human.

14. The combination for use as defined in any one of claims 1 to 6 or claim 13, or the pharmaceutical composition for use as defined in any one of claims 7 to 13, further in combination with standard therapy for the treatment and/or prevention of functional gastrointestinal disorders.

15. The combination or pharmaceutical composition for use according to claim 14, wherein the standard therapy for the treatment and/or prevention of functional gastrointestinal disorders is selected from the group consisting of anti-acid therapy (proton pump inhibitors and histamine 2 receptor antagonists) antibiotics; antiemetic, prokinetic, antispasmodic, antidepressant, antipsychotic, digestive enzymes and plant extracts.

16. A bacillus subtilis strain deposited with BCCM under accession number LMG P-31319.

17.A bacillus coagulans strain deposited with BCCM under accession number LMG P-31318.

18. Use of a combination comprising bacillus subtilis and bacillus coagulans spores for enhancing gastrointestinal tract function in a subject.

19. The use according to claim 18, wherein the ratio of bacillus subtilis to bacillus coagulans spores is 1:1, 1:2 or 2: 1.

20. Use according to claim 18 or 19, wherein the bacillus subtilis spores are derived from strain MY02 deposited with BCCM under accession number LMG P-31319.

21. Use according to any one of claims 18 to 20; wherein the Bacillus coagulans spores are derived from strain MY01 deposited with BCCM under accession number LMG P-31318.

22. Use of a composition comprising a combination as defined in any one of claims 18 to 21 and one or more pharmaceutically acceptable carriers, diluents or excipients for enhancing the gastrointestinal tract function in a subject.

23. Use according to claim 22, wherein the composition further comprises one or more ingredients selected from plant extracts, proteins, vitamins, minerals and digestive enzymes.

24. Use according to claim 22 or 23, wherein the composition is in the form of a tablet, capsule, sachet, stick or any other edible form including but not limited to functional food, juice, beverage or candy.

25. Use according to any one of claims 22 to 24, wherein the composition is a product of the dairy industry, the beverage industry or the pharmaceutical industry, or is a natural product.

26. Use according to any one of claims 18 to 25, wherein the subject is a human subject suffering from a functional gastrointestinal disorder.

27. Use according to any one of claims 18 to 25, wherein the subject is a healthy subject.

Technical Field

The present invention relates generally to the use of probiotics for the control of functional gastrointestinal disorders. More specifically, the present invention relates to the use of Bacillus coagulans (Bacillus coagulousns) or Bacillus subtilis (Bacillus subtilis) or a combination of both strains for the prevention or treatment of functional gastrointestinal disorders, such as functional dyspepsia. The invention also discloses the use of bacillus coagulans or bacillus subtilis or a combination of the two strains as a replacement therapy or an adjuvant therapy for gastric acid inhibition therapy.

Background

Functional Gastrointestinal (GI) disorders, also known as disorders of gut-brain interactions, include a number of independent idiopathic disorders affecting different parts of the GI tract and involving visceral hypersensitivity and dyskinesias. Examples of functional GI disorders include functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distention, reflux hypersensitivity, irritable bowel syndrome, functional constipation, functional diarrhea.

Functional dyspepsia is a Functional GI disorder, a chronic or recurrent upper Gastrointestinal (GI) condition that originates in the gastroduodenal region, and has a significant impact on the life of patients (Talley NJ et al 2015.Functional dyspsia. n Engl J Med 373,1853-63). Organic causes include peptic ulcer, Helicobacter pylori (Helicobacter pylori) infection, biliary tract disease, and esophageal gastric cancer. According to roman standards, Functional Dyspepsia (FD) includes the diagnostic categories of Epigastric Pain Syndrome (EPS) with epigastric pain or burning sensation and Postprandial Distress Syndrome (PDS) with meal-related satiety or early satiety, which are not interpretable by routine examination, including upper GI endoscopy (Stanghellini V et al 2016. gastroendodal disorders 150,1380-92). Although FD often occurs in up to 15% of the general population, the underlying pathophysiology is unclear and the first line treatment option is antacid (Moayyedi PM et al 2017, ACG and CAG Clinical guidelines: Management of Dyspsia. am J gastroenterol112, 988-1013). Although gastroesophageal reflux disease (GERD) may coexist with FD, one population-based study in the meio Clinic (Mayo Clinic) concluded that FD is often misdiagnosed and misidentified as GERD. Recent Cochrane analysis concluded that Proton Pump Inhibitors (PPIs) were more effective in FD than placebo and histamine 2 receptor antagonists (Pinto-Sanchez MI et al 2017.Proton pump inhibitors for functional dyspysia. Cochrane Database Syst Rev 3, CD 011194). PPIs, however, are also associated with potential long-term adverse events, such as increased Risk of GI infection and Small intestine Bacterial Overgrowth (Lo W et al, 2013.Proton Pump Inhibitor Use and the Risk of Small Intestinal Bacterial Overgrowth: amino-analysis. clin Gastroenterol Heapatol 11,483-90). Recent studies have also reported that stomach (Tsuda A et al 2015. infiluence of Proton-Pump Inhibitors on the Luminal Microbiota in the Gastric specific tract of protein microorganisms 6, e 89; Paroni Sterbini F et al 2016.Effects of Proton Pump Inhibitors on the Gastric Mucosa-Associated Microbiota in Dysporadic Patients. applied Environ Microbiol 82,6633-44) and feces (Jackson MA et al 2016.Proton Pump Inhibitors on the composition of the gut microbial barriers Gut 65,749-56; Imhann F et al 2016. Propton microorganisms after the Gastric acid barrier 539) are most likely to be inhibited by Gastric acid in the oral microbial flora, Gastric acid inhibition group 65,740. PPI may affect the Gastric microbiome by increasing the relative abundance of Streptococcus (Streptococcus), whether or not helicobacter pylori is present, which may contribute to the persistence of FD symptoms with PPI or cause long-term adverse Effects (Paroni Sterbini F et al 2016.Effects of Proton Pump Inhibitors on the Gastric Mucosa-Associated Microbiol in chromatographic Patients. apple Environ Microbiol 82,6633-44). In addition, inappropriate use of PPIs can bring significant costs to the healthcare budget (Boghessian TA et al 2017. suppression conversion reception of respiratory proton inhibitor use in addends. Cochrane Database Syst Rev 3, CD 011969).

Several publications have demonstrated increased permeability of duodenal mucosa and mild inflammation in FD patients, which is associated with diet-related symptoms (kit S et al 2009.Immune dysfunction in tissues with functional organization disorders. neurogastrotriol Motil 21,389-98; Vanhel H et al 2014. Imperial dual mucosal integration and low-gradient simulation in functional dyspepsia. Gut63,262-71; Cirillo C et al 2015. identification for neural and structural changes in submucous tissue of functional tissue with functional dynamic steric diffusion. am JGastrunol 110,1205-15). The reasons for barrier defects and immune activation are not clear, but candidate factors include Psychological stress, luminal food composition, (bile) acids and microbiota (Vanheel H et al 2014. affected dual dietary intake and low-grade intake in functional gravity diet 63,262-71; Vanuytsel T et al 2014. psychopharmacological stress and physiological stretch-refining one intake of dietary intake in human by a cell-dependent dietary intake. Gut63, 1293-9). A preliminary report involving 9 patients with FD showed that the Streptococcus (Streptococcus) and total bacterial load in the duodenum of FD was increased compared to healthy controls, which was related to the severity of diet-related symptoms and quality of life (Zhong L et al 2017. Dyspessia and the microbiome: time to focus on the small intestine. gut 66,1168-9).

Few clinical trials have tested probiotics for the treatment of functional dyspepsia. Recently, probiotic treatment was found to be beneficial for postprandial dyspepsia symptoms (Igarashi M et al 2017 Alteration in the systemic microbiota and its restriction by probiotics in tissues with functional dyspeptia. BMJ open Gastroenterol 4, e000144), possibly by changes in the flora, although this study did not examine this hypothesis.

Bacillus (Bacillus) is a gram-positive strain capable of forming endospores highly resistant to gastric acid (Elshaghabe FMF et al 2017.Bacillus As Potential Probiotics: Status, conccerns, and Future perspectives. front Microbiol 8,1490). In addition, spore-forming probiotics or "sporozoites" are stable at room temperature and deliver more viable bacteria to the small intestine than traditional probiotic supplements (Cutting sm.2011.bacillus probiotics. food Microbiol 28,214-20). Oral administration of a combination of spore-based probiotics (bacillus indicus, b. subtilis), bacillus coagulans, bacillus licheniformis (b. licheniformis) and bacillus clausii (b. clausii)) was shown to reduce postprandial endotoxemia, possibly due to increased intestinal permeability (McFarlin BK et al 2017.Oral spore-based biological supplementation was associated with increased with reduced involvement of post-metabolic endixin, trigycerins, and disease biological manufacturers J structural nutrition 8, 117-.

The present invention describes the use of bacillus coagulans or bacillus subtilis or a combination of both strains for the prevention and/or treatment of functional GI disorders, such as functional dyspepsia. In addition, the invention also discloses the application of the bacillus coagulans or the bacillus subtilis or the combination of the two strains as a replacement therapy or an adjuvant therapy of gastric acid inhibition therapy. In another aspect, the present invention discloses the use of bacillus coagulans or bacillus subtilis spores, or a combination of both strains, for the prevention or treatment of a further disease selected from the group consisting of: metabolic syndrome, such as high cholesterol levels and high blood glucose levels; immune system disorders such as chronic inflammatory, allergic and autoimmune diseases; mental health disorders, such as depression and anxiety; and genitourinary disorders, such as urogenital infections.

Summary of The Invention

The present invention describes the use of bacillus coagulans or bacillus subtilis or a combination of both strains for the prevention/prevention and/or treatment of a functional gastrointestinal disorder in a subject. In particular, the present invention relates to a combination comprising bacillus subtilis and bacillus coagulans spores for use in the prevention and/or treatment of a functional Gastrointestinal (GI) disorder in a subject. The functional GI disorder is selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distension, reflux hypersensitivity, irritable bowel syndrome, functional constipation, functional diarrhea. In a further embodiment, the functional GI disorder is selected from functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or bloating. Thus, in an even more preferred embodiment, the present invention provides a combination comprising bacillus subtilis and bacillus coagulans spores for use in the prevention and/or treatment of functional dyspepsia in a subject.

Additionally, the present invention relates to a combination comprising bacillus subtilis and bacillus coagulans spores for use as a replacement therapy or adjunct therapy for gastric acid inhibitory treatment of a subject, in particular a subject having a functional Gastrointestinal (GI) disorder. The functional GI disorder is selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distension, reflux hypersensitivity, irritable bowel syndrome, functional constipation, functional diarrhea. In a further embodiment, the functional GI disorder is selected from functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or bloating. Thus, in an even more preferred embodiment, the present invention provides a combination comprising bacillus subtilis and bacillus coagulans spores for use as a replacement therapy or adjunct therapy for gastric acid inhibitory treatment of a subject, in particular a patient suffering from a functional Gastrointestinal (GI) disorder.

Further, in one aspect of the invention, the combination of bacillus subtilis and bacillus coagulans spores for use according to all embodiments of the invention is provided in combination with standard therapy for functional gastrointestinal disorders. In a further aspect, the standard of care therapy is selected from the group consisting of acid-inhibitory therapy (proton pump inhibitors and histamine 2 receptor antagonists), antibiotics, antiemetics, prokinetic drugs, antispasmodics, antidepressants, antipsychotics, digestive enzymes, and plant extracts.

In another aspect of the invention, the combination for use according to the different embodiments of the invention comprises bacillus subtilis and bacillus coagulans spores, wherein the bacillus subtilis and bacillus coagulans spores are in a ratio of 1:1, 1:2 or 2: 1.

In a further aspect, the combination for use according to the invention comprises bacillus subtilis and bacillus coagulans, wherein the bacillus subtilis spores are derived from strain MY02 deposited with BCCM under accession number LMG P-31319. Said strain has been deposited with the coordinated collections of microorganisms (BCCM) Belgium at 2019, 3, 14, under accession number LMG P-31319 (Universal Gent, KL Ledeganckstraat 35,9000Gent, Belgium) and is further denoted MY02 strain herein. In another aspect, the combination for use according to the invention comprises bacillus subtilis and bacillus coagulans, wherein the bacillus coagulans spores are derived from strain MY01 deposited with BCCM under accession number LMG P-31318. Said strain has been deposited with the coordinated collections of microorganisms (BCCM) Belgium at 14/3.2019 under the accession number LMG P-31318 (Universal Gent, KL Ledeganckstraat 35,9000Gent, Belgium) and is further denoted MY01 strain herein.

In a still further aspect, the combination for use according to the invention comprises bacillus subtilis and bacillus coagulans, wherein the bacillus subtilis spores are derived from strain MY02 deposited with BCCM under accession number LMG P-31319 and wherein the bacillus coagulans spores are derived from strain MY01 deposited with BCCM under accession number LMG P-31318.

In another aspect, the present invention provides a pharmaceutical composition for preventing and/or treating a functional gastrointestinal disorder in a subject. The pharmaceutical composition comprises a combination comprising bacillus subtilis and bacillus coagulans spores according to various embodiments as described above. The pharmaceutical composition further comprises one or more pharmaceutically acceptable carriers, diluents or excipients. In another embodiment, the pharmaceutical composition according to the invention is for use in the prevention and/or treatment of a functional gastrointestinal disorder selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distension, reflux hypersensitivity, irritable bowel syndrome, functional constipation, functional diarrhea; preferably selected from functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distension. In an even more preferred embodiment, the pharmaceutical composition according to the invention is for use in the prevention and/or treatment of functional dyspepsia in a subject.

In another embodiment, the present invention discloses a pharmaceutical composition comprising bacillus subtilis and bacillus coagulans spores and one or more pharmaceutically acceptable carriers for use as a replacement therapy or adjunct therapy for gastric acid inhibition treatment of a subject, in particular a subject having a functional Gastrointestinal (GI) disorder. The functional GI disorder is selected from the group consisting of functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or distension, reflux hypersensitivity, irritable bowel syndrome, functional constipation, functional diarrhea. In a further embodiment, the functional GI disorder is selected from functional dyspepsia, functional chest pain, functional heartburn, hiccup disorder, nausea and vomiting disorder, functional abdominal bloating or bloating. Thus, in an even more preferred embodiment, the present invention provides a pharmaceutical composition comprising a combination of bacillus subtilis and bacillus coagulans spores for use as a replacement therapy or adjunct therapy for gastric acid suppression therapy in a subject, in particular a patient suffering from a functional Gastrointestinal (GI) disorder.

Further, in one aspect of the present invention, there is provided a pharmaceutical composition comprising a combination of bacillus subtilis and bacillus coagulans spores for use in accordance with all embodiments of the present invention, in combination with standard therapy for functional gastrointestinal disorders. In another aspect, the standard of care therapy is selected from the group consisting of proton pump inhibitors, acid suppressive therapy (proton pump inhibitors and histamine 2 receptor antagonists), antibiotics, antiemetics, prokinetic drugs, antispasmodics, antidepressants, antipsychotics, digestive enzymes, and plant extracts.

The pharmaceutical composition for use according to the different embodiments of the present invention may further comprise one or more ingredients selected from the list comprising plant extracts, such as ginger, artichoke, proteins, vitamins, minerals or digestive enzymes, such as amylase, cellulase, lipase, beta-galactosidase, papain, bromelain and protease.

In another embodiment, the pharmaceutical composition for use in various embodiments according to the present invention is in the form of a tablet, capsule, stick or any other edible form including, but not limited to, functional food, juice, beverage or candy.

In another embodiment, the pharmaceutical composition of the invention is a product of the dairy industry, the beverage industry, the food industry or the pharmaceutical industry, or it is a natural product.

In another aspect of the present invention, the combination or pharmaceutical composition according to the different embodiments of the present invention is for use in the prevention and/or treatment of a functional gastrointestinal disorder, such as functional diarrhea, in a subject. The subject may be a human subject, such as an adult or an infant. In another embodiment, the subject may be an animal, preferably a pet or a production animal. Preferably, the subject is a human.

The invention also provides a bacillus subtilis strain. The Bacillus subtilis strain was deposited at Belgium Coordinated Collections of Microorganisms (BCCM) (Universal Gent, KL Ledeganckstraat 35,9000Gent, Belgium) at 14/3.2019 under accession number LMG P-31319.

The invention additionally provides a bacillus coagulans strain. The Bacillus coagulans strain was deposited at 14.3.2019 with the accession number LMG P-31318 at the Belgian Coordinated Collections of Microorganisms (BCCM) (Universal Gent, KL Ledeganckstraat 35,9000Gent, Belgium).

In another aspect, the invention features the use of bacillus subtilis or bacillus coagulans or a combination of both strains for enhancing gastrointestinal function in a subject. More specifically, the use of a combination comprising bacillus subtilis and bacillus coagulans spores for enhancing gastrointestinal tract function in a subject is disclosed. The subject is a human or a mammal; preferably a human subject. In particular embodiments, the subject is a human suffering from a functional gastrointestinal disorder; particularly those with functional dyspepsia. In another embodiment, the subject is a healthy subject.

In another aspect, and in the uses, the bacillus subtilis and bacillus coagulans spores are present in the combination in a ratio of 1:1, 1:2, or 2: 1.

In yet another aspect and in the uses, the Bacillus subtilis spores are derived from strain MY02 deposited with BCCM under accession number LMG P-31319. In another aspect, and in the use, the bacillus coagulans spores are derived from strain MY01 deposited with BCCM under accession number LMG P-31318.

Another aspect discloses the use of a composition comprising a combination comprising bacillus subtilis and bacillus coagulans spores according to any of the embodiments and one or more pharmaceutically acceptable carriers, diluents or excipients for enhancing the gastrointestinal function in a subject. The composition may further comprise one or more ingredients selected from the group consisting of plant extracts, proteins, vitamins, minerals, and digestive enzymes. In another aspect, the composition is in the form of a tablet, capsule, pouch, stick, or any other edible form, including but not limited to functional food, juice, beverage, or candy. In yet another aspect, the composition is a product of the dairy industry, the beverage industry, or the pharmaceutical industry, or is a natural product.

Brief description of the drawings

With specific reference now to the drawings in detail, it is stressed that the particulars shown are by way of example and for purposes of illustrative discussion of the various embodiments of the present invention only. They are presented to provide what is believed to be the most useful and readily described of the principles and conceptual aspects of the invention. In this regard, no attempt is made to show structural details of the invention in more detail than is necessary for a fundamental understanding of the invention. The description taken with the drawings making apparent to those skilled in the art how the several forms of the invention may be embodied in practice.

FIG. 1: study protocol to study the effect of sporulating probiotics of the present invention on dyspeptic symptoms and clinical parameters.

Fig. 2 group 1FD (with PPI) mean LPDS scores reported between baseline (1 week before the start of the blind phase) and cycle 13. Week 13 corresponds to week 5 of the open label phase, in which all patients received sporozoites of the present invention (2 capsules per day). ES: early satiety; PPF: postprandial satiety; UAB: abdominal distension in the upper abdomen; UAP: epigastric pain; EGB: upper abdominal burning; and NS: nausea; BC: burping; HB: heartburn; FT: fatigue; PPNS: postprandial nausea; PPUAP: epigastric pain after meals.

Fig. 3 group 2FD (no PPI) mean LPDS scores reported between baseline (1 week before the start of the blind phase) and cycle 13. Week 13 corresponds to week 5 of the open label phase, in which all patients received sporozoites of the present invention (2 capsules per day). ES: early satiety; PPF: postprandial satiety; UAB: abdominal distension in the upper abdomen; UAP: epigastric pain; EGB: upper abdominal burning; and NS: nausea; BC: burping; HB: heartburn; FT: fatigue; PPNS: postprandial nausea; PPUAP: epigastric pain after meals.

Fig. 4 sum of mean LPDS scores for group 1FD and 2FD during baseline and week 13. Week 13 corresponds to week 5 of the open label phase, in which all patients received sporozoites of the present invention (2 capsules per day). ES: early satiety; PPF: postprandial satiety; UAB: abdominal distension in the upper abdomen; UAP: epigastric pain; EGB: upper abdominal burning; and NS: nausea; BC: burping; HB: heartburn; FT: fatigue; PPNS: postprandial nausea; PPUAP: epigastric pain after meals.

Detailed Description

The present invention is based on the following findings: the combination comprising bacillus subtilis and bacillus coagulans is particularly useful for the treatment and/or prevention of functional gastrointestinal disorders, such as functional dyspepsia, and for enhancing gastrointestinal function in humans. In particular, the inventors have found that a combination comprising bacillus subtilis and bacillus coagulans spores is useful for the treatment and/or prevention of functional gastrointestinal diseases, such as functional dyspepsia, in humans. Furthermore, combinations comprising bacillus subtilis and bacillus coagulans spores may also be used to enhance general gastrointestinal tract function, in particular to stimulate gastrointestinal digestion and to reduce general discomfort of postprandial fullness, epigastric bloating or early satiety after eating a meal.

Furthermore, the inventors have found that a combination comprising bacillus coagulans and bacillus subtilis spores reduces symptoms of upper abdominal pain and burning, chest pain and heartburn. These disorders are usually treated by gastric acid inhibitory therapy (e.g. proton pump inhibitors). Thus, in a further embodiment, the present invention provides a combination or pharmaceutical composition comprising bacillus coagulans spores and bacillus subtilis spores according to various embodiments of the present invention, which is also used as a medicament in a subject; particularly gastric acid inhibitory treatments such as alternative or adjunctive therapies to proton pump inhibitors in subjects suffering from functional gastrointestinal disorders.

Accordingly, the present invention provides a combination or pharmaceutical composition comprising bacillus coagulans and bacillus subtilis for the treatment of a functional gastrointestinal disorder. Furthermore, the use of the combination or the composition for enhancing the general gastrointestinal function in a healthy subject or a subject suffering from a functional gastrointestinal disorder, such as functional dyspepsia, is disclosed.

In the context of the present invention, an improvement in gastrointestinal function is observed in healthy subjects as well as in subjects suffering from functional gastrointestinal disorders. Also in the context of the present invention, an improvement in gastrointestinal tract function is any improvement observed in the activity and functionality of the gastrointestinal tract. In particular, after eating, gastrointestinal function is improved, for example, as indicated by a reduction in general discomfort such as postprandial fullness, epigastric bloating, or early satiety after eating. In a particular aspect, the combination or composition according to the different embodiments of the present invention is used to stimulate gastrointestinal digestion to reduce incidental digestive symptoms such as hiccups, epigastric bloating, epigastric pain or burning, heartburn and gastroesophageal reflux in otherwise healthy subjects.

The bacteria are usually in the form of bacterial spores or as vegetative bacterial cells, or a mixture of both. In a particularly preferred aspect of the invention, the bacteria are in the form of bacterial spores which can germinate in the gastrointestinal tract. In another embodiment, they may have been treated such that they cannot germinate. For example, the spores may be treated by heating and may, for example, be autoclaved to prevent germination. In another embodiment, the bacteria are in the form of vegetative cells. In one embodiment, the spores or vegetative cells can be provided in an isolated form. In yet another embodiment, the microbial material may be an extract of bacterial cells. In yet another embodiment, the microbial material may be a metabolite of bacterial cells released during cell growth in the culture medium. In another other embodiment, the microbial material may be any combination of bacterial spores, vegetative bacterial cells, extracts of bacterial cells, or metabolites of bacterial cells. In a preferred embodiment, the bacteria in the different embodiments of the present invention are provided as viable spores capable of germinating in the gastrointestinal tract.

The preparations or pharmaceutical preparations according to the different embodiments of the present invention require isolation of the bacteria bacillus subtilis and bacillus coagulans or their corresponding spores from the final growth medium. The skilled person is well aware of techniques which can be used to separate viable bacteria or spores from the growth medium, for example using centrifugation, filtration, micromanipulation and the like. For medium and long term storage, the isolated bacteria are preferably kept in a dry state, for example using freeze drying or spray drying. In one embodiment of the invention, the bacillus subtilis and bacillus coagulans used for the manufacture of the composition or the pharmaceutical composition are spray-dried bacillus subtilis and bacillus coagulans, obtained by spray-drying bacteria using a sugar protectant, in particular trehalose, as protectant. Exemplary methods suitable for spray drying the bacteria of the present invention are available, for example, from Sunny-Roberts and Knorr (int. Diary J.,19(2009)209- "214).

Once isolated, the Bacillus subtilis and Bacillus coagulans strains of the present invention may be prepared by any known or otherwise effective method for pharmaceutical formulation or manufacture of the selected product form. Methods of preparing pharmaceutical compositions according to the invention can be found in "Remington's pharmaceutical Sciences", mid.

For example, the compositions or pharmaceutical compositions according to the various embodiments of the present invention may be formulated with common excipients, diluents or carriers and formed into oral tablets, capsules, sprays or oral liquids (e.g., suspensions, solutions, emulsions), powders or any other suitable dosage forms.

Non-limiting examples of suitable and pharmaceutically acceptable Excipients, diluents and carriers can be found in the Handbook of Pharmaceutical Excipients, Second Edition, American Pharmaceutical Association,1994, and may include: fillers and extenders such as starch, sugars, mannitol, and silicon derivatives; binders such as carboxymethyl cellulose and other cellulose derivatives, alginates, gelatin, and polyvinyl pyrrolidone; humectants such as glycerol; disintegrants such as calcium carbonate and sodium bicarbonate; dissolution retarding agents such as paraffin; resorption accelerators such as quaternary ammonium compounds; surfactants such as acetyl alcohol, glyceryl monostearate, adsorption carriers such as kaolin and bentonite; carriers such as propylene glycol and ethanol, and lubricants such as talc, calcium stearate and magnesium stearate, and solid polyethylene glycols.

The combinations, compositions and pharmaceutical compositions according to the invention are primarily suitable for adults and infants. However, they are also suitable for animals, in particular pets and production animals. Examples of these include dogs, cats, rabbits, horses, cattle, pigs, goats, sheep and poultry. Thus, the term "subject" as used herein includes humans and animals.

The composition used according to the invention may be in the form of a tablet, capsule, sachet, stick or any other edible form including, but not limited to, functional food, juice, beverage or candy. The term "edible form" as used herein is intended to encompass all consumer products, especially food products, and it may be solid, jelly or liquid. The term encompasses ready-to-use products and products produced using the probiotic composition as a sole starter or in combination with conventional starters or other probiotics. The food product may for example be a product of the dairy industry or the beverage industry. Alternatively, it may be a natural product.

In the present invention, "dairy" refers to any liquid or semi-solid milk or whey-based product with different fat content. The dairy content may be, for example, cow's milk, goat's milk, sheep's milk, skim milk, whole milk, dairy combined from milk powder and whey without any processing, or processed products such as yogurt, curd milk, curd, yogurt, whole yogurt, buttermilk, other yogurt products such as villi, fillings for snack bars, and the like. Another important class includes milk beverages such as whey beverages, fermented milk, condensed milk, baby milk, ice cream; milk-containing foods, such as candies.

Products according to various embodiments of the invention may also be concentrated and used as ingredients. In addition, the product may also be dried and used in the form of a powder or a lyophilizate. The product is also applied in capsules, pills or tablets. The product can also be used for preparing functional food, edible product for promoting health and promoting health, or other corresponding products. It can also be in animal feed. Possible forms are capsules, pills or tablets, e.g. manufactured in a conventional process for preparing such products, e.g. in the pharmaceutical industry. For this, the form of each of the food, food material and/or pharmaceutical product and animal feed is not particularly limited.

Examples

The following examples illustrate the invention. The examples should not be construed as limiting the claims in any way.

Example 1

I. Research settings

Purpose of the experiment

Our aim was to study the effect of sporulating probiotics compared to placebo on dyspeptic symptoms and blood, saliva and stool parameters in FD patients with and without acid inhibition by Proton Pump Inhibitors (PPIs).

Test registration

The study was approved by the ethical committee for KU Leuven (belgium) and was registered in the clinical trial. gov database under accession number NCT 04030780.

Primary endpoint

The LPDS questionnaire was used to assess the effect of sporogens (a combination of bacillus coagulans MY01 and bacillus subtilis MY02 spores) on clinical symptoms compared to placebo.

Secondary endpoint

Effect of sporozoite (combination of bacillus coagulans MY01 and bacillus subtilis MY02 spores) on blood, saliva and stool parameters compared to placebo. The exploratory endpoint was the effect of the sporozoite compared to placebo on the bile acid breath test to assess bacterial overgrowth (group 1 FD).

Design of experiments and flow charts

A prospective randomized placebo-controlled study was performed in 2FD groups with an open label expansion phase (fig. 1):

group 1FD (with PPI): study procedure when admitted (week 0 or week-1) (1) and 8 weeks after PPI + sporozoite or placebo (2) and then 8 weeks with PPI + sporozoite (open label);

group 2FD (no PPI): study procedure on inclusion (week 0 or-1) (1) and 8 weeks after sporozoite or placebo (2) and then 8 weeks after sporozoite (open label).

The experimental flow chart shown in table 1 was used.

TABLE 1 Experimental flow sheet for the study

Test drugs

a) Study drug product and dosing regimen

FD patients (with or without PPI) were treated randomly with sporulating probiotics or placebo. The spore forming probiotic in this study was a combination of bacillus coagulans MY01 and bacillus subtilis MY02, and the spore forming probiotic was encapsulated twice a day in 1 capsule (2.5 x10 per capsule) over an 8 week period⁹A CFU; in a ratio of 50: 50). Placebo treatment was administered in a similar capsule, also twice daily. In the open labelThe sigextension phase, sporozoite was administered for a further 8 weeks.

b) Drug management responsibility

The remaining capsules were counted every 8 weeks.

c) Subject compliance (compliance)

Each month the patient is contacted by email to verify compliance.

d) Concomitant medication (non-IMP)

Patients are required to avoid taking illicit drugs at the same time.

Subject selection and withdrawal

a) Inclusion criteria

Patients were eligible for study if all of the following criteria were met:

patients >18 years of age diagnosed as FD (roman IV standard):

-abdominal pain or burning (EPS) 1 or more days per week

-postprandial satiety or early satiety severe enough to affect normal activity (PDS) for 3 or more days per week

FD group 1: daily PPI treatment (of any type and dose) over the past 4 weeks

FD group 2: no PPI treatment over the past 8 weeks

Male or female (not pregnant or lactating and using contraceptive measures or post-menopause)

Normal defecation habit (defecation once every 3 days, up to 3 times a day)

Witness written informed consent

-use of a household refrigerator (-18 to-20 ℃)

To be able to understand and comply with research requirements

b) Exclusion criteria

Patients were excluded from the study if any of the following criteria were met:

any active somatic or psychiatric condition that may explain the symptoms of dyspepsia (allowing a single antidepressant in a stable dose for psychiatric indications, without limitation for other indications)

-the main symptoms of gastroesophageal reflux disease (GERD) or Irritable Bowel Syndrome (IBS)

-use of immunosuppressive or antibiotic agents for <3 months

History of major abdominal operations (appendectomy, cholecystectomy or splenectomy)

-personal history of type 1 diabetes, celiac disease or inflammatory bowel disease

Type 2 diabetes (including therapy)

Active malignant tumors (including therapy)

Known HIV, HBV or HCV infections (including therapy)

Magnetizable objects (e.g. cochlear implants, neurostimulators, pacemakers, metal fragments or implants) or claustrophobia (MRI safety standard, with consent only attached to group 2FD, other procedures still possible)

-bulk alcohol consumption (>10 units/week)

c) Participant selection

FD patients were recruited in outpatients or endoscopy rooms for general and functional GI disorders. Advertisements are distributed through the magazine "Metro" and hung as posters in hospitals and universities, and also uploaded under the "openep students" of the UZ Leuven intranet website. Patients responding to this advertisement are invited to study visits. Patients may also be recruited from a referral hospital (referring hospital) if all inclusion criteria and non-exclusion criteria are met.

d) Randomization procedure

The randomization list between sporozoites and placebo was done by the sponsor using an online randomization tool (http:// www.randomization.com /), with a chunk size of 5. The randomized list of groups 1FD and 2FD is kept by the sponsor. Study treatments were numbered according to a randomized list and were provided by the sponsor to a study blinded to the randomized code.

e) Subject withdrawal

Patients were entitled to withdraw from the study at any time, without affecting future management and treatment. Patients may be withdrawn from the study if the following occurs:

the patient experiences any adverse event the investigator deems harmful to the patient's health

Patient violation of eligibility criteria for study protocol

Patient start of concomitant medication with disablement

Patient non-compliance with required study procedures, e.g. filling in an entry or follow-up (monthly) questionnaire

Data from patients who exited the study may still be available, depending on the procedure performed and the questionnaire filled out.

f) Predicted test time

Study duration was 16 weeks. The duration of the entire trial was expected to be 24 months from the first visit of the first patient to the last visit of the last patient.

LPDS diary

Throughout the study, subjects filled out the Luxen Postprandial Distress Scale (LPDS) daily. LPDS diaries have been validated for assessment of dyspepsia and heartburn symptoms (carbon et al, 2016; animal Pharmacol Ther 44: 989-. The LPDS scale is a diary tool with 10 epigastric symptoms (early satiety, postprandial satiety, epigastric bloating, epigastric pain, epigastric burning, nausea, hiccups, heartburn, postprandial nausea, postprandial epigastric pain) and fatigue as additional quality of life parameters. The rating of the items is expressed as a verbal description (five levels per item, never present to very severe) accompanied by a "smiley face"See table 2 for an example of an LPDS diary. For ease of analysis, the linguistic descriptors are converted to numerical values as follows: absent-0; mild ═ 1; medium ═ 2; severe ═ 3; very severe ═ 4.

Table 2. example LPDS diary problem. The daily questionnaire consisted of 10 questions for upper abdominal symptoms and 1 question for general fatigue as a quality of life parameter.

Results II

At this stage, the study results are only available for the open label stage and for a limited number of patients. The study is still ongoing at present and more patients will be recruited and the data further analyzed. Nevertheless, the currently available data have shown a positive effect of a combination comprising bacillus subtilis and bacillus coagulans on the prevention and/or treatment of functional gastrointestinal disorders.

Patients began recording their gastrointestinal symptoms daily in the LPDS diary 7 days (week-1) prior to the start of the placebo-controlled, randomized blind phase (week 0) of the trial. LPDS diaries were completed throughout the clinical trial (blind phase followed by open label phase). The average results recorded daily during the-1 st cycle were taken as the baseline value for each symptom. After completion of the blinding phase with 8-week placebo or 8-week sporozoite (combination of bacillus coagulans MY01 and bacillus subtilis MY02 spores) as the test drug administration, patients were included in an open label phase in which all patients were supplemented daily with sporozoite (combination of bacillus coagulans MY01 and bacillus subtilis MY02 spores).

Figure 2 shows baseline symptom values at week-1 compared to the most recently available mean LPDS score reported by 18 patients in group 1FD (with PPI) during week 13 of the trial (corresponding to week 5 of the open label phase). During week 13 of the trial, the severity of all dyspepsia symptoms was significantly reduced in group 1FD (with PPI) except for postprandial nausea (PPNS).

Unexpectedly, there was a nearly 50% reduction in gastric acid related symptoms of epigastric burning (EGB) and Heartburn (HB). Since in group 1FD gastric acid inhibition therapy (proton pump inhibitor PPI) was combined with administration of sporozoites of the invention, epigastric and heartburn were expected to be rare symptoms in this population due to PPI therapy. However, even with PPI therapy, dyspepsia patients appear to suffer from these annoying symptoms. Notably, administration of the sporogens of the present invention further reduces the symptoms of epigastric and heartburn. Thus, these data indicate that the combination of bacillus coagulans and bacillus subtilis sporogens improves functional gastrointestinal symptoms when the sporogens are administered in combination with PPIs. Thus, the present data provide sufficient evidence that the combination of bacillus coagulans and bacillus subtilis sporogens may be used to treat functional gastrointestinal disorders, as well as in combination with PPI as an adjunct therapy for dyspeptic symptoms.

Figure 3 shows baseline symptom values at week-1 compared to the recently available mean LPDS score reported by 16 patients in group 2FD (non-PPI) during week 13 of the trial (corresponding to week 5 of the open label phase). At week 13 of the trial, the severity of all dyspepsia symptoms in group 2FD (no PPI) was significantly reduced compared to the starting point of the study (week-1). Thus, these data indicate that the combination of bacillus coagulans and bacillus subtilis spores is an effective treatment (no other type of therapy) for functional gastrointestinal disorders.

Figure 4 shows the sum of the mean baseline symptom scores for cohorts 1FD and 2FD (week-1 at the start) compared to the sum of the mean LPDS scores reported during the 13 th cycle of the trial. The data presented show that the combination of bacillus coagulans and bacillus subtilis spores significantly reduced the symptoms of fatigue in all dyspepsia symptoms (problems in LPDS diary) and in patients receiving additional acid suppression therapy (with PPI) as well as in patients not receiving additional treatment (without PPI).