Pharmaceutical compositions comprising ribose and amino acids

Abstract

The present invention relates to a composition for use in a method of treating a bacterial infection selected from a bacterial urinary tract infection, a bacterial respiratory infection, a bacterial soft tissue infection, and a bacterial bone infection in a patient, the composition comprising ribose and the amino acids glycine, alanine, and glutamine in an aqueous solution, the method comprising orally administering the composition to a patient who is further being treated with an antibiotic.

Description

[Technical Field]

[0001] The present invention relates to compositions for use in methods of treating a bacterial infection selected from a bacterial urinary tract infection, a bacterial respiratory infection, a bacterial soft tissue infection, and a bacterial bone infection in a patient. [Background technology]

[0002] Urinary tract infections (UTIs) are the most common bacterial disease worldwide. The prevalence of community-acquired infections is 0.7%. The frequency of healthcare-associated UTIs among healthcare-associated infections is 12.9% in the United States, 19.6% in Western Europe, and 24% in developing countries. In urology, the prevalence is 5.1%. UTIs are frequently caused by both gram-negative and gram-positive bacteria, frequently Escherichia coli and Klebsiella pneumoniae. Because of high recurrence rates and increasing antibiotic resistance among urinary tract pathogens, the burden of these infections is likely to increase significantly. Summary of the Invention

[0003] The present invention proposes a new treatment for bacterial infections selected from bacterial urinary tract infections, bacterial respiratory infections, bacterial soft tissue infections, and bacterial bone infections. In particular, the present invention provides a composition for use in a method of treating a bacterial infection selected from a bacterial urinary tract infection, a bacterial respiratory infection, a bacterial soft tissue infection, and a bacterial bone infection in a patient, the composition comprising ribose and the amino acids glycine, alanine, and glutamine in an aqueous solution, the method comprising orally administering the composition to a patient who is being concomitantly treated with an antibiotic. Preferably, the infection being treated is a bacterial urinary tract infection. Ribose is preferably D-ribose, alanine is preferably racemic (D / L)-alanine, and glutamine is preferably L-glutamine. The concentration of ribose in the aqueous solution of the composition is preferably between 30 and 40 g / L. The concentration of glycine in the aqueous solution of the composition is preferably between 1.5 and 2.5 g / L. The concentration of alanine in the aqueous solution of the composition is preferably between 15 and 25 g / L. The concentration of glutamine in the aqueous solution of the composition is preferably between 20 and 30 g / L. The relative concentrations of amino acids in the aqueous solution of the composition are preferably 1:8 to 12:10 to 15 (glycine:alanine:glutamine). The relative concentrations of ribose and the amino acids glycine, alanine, and glutamine in the aqueous solution of the composition are preferably 1:0.8 to 2.0 (ribose:amino acids). The volume of the aqueous solution of the composition is preferably 5 to 50 ml, preferably 10 to 30 ml.

[0004] In a preferred embodiment, the composition is administered to the patient two or three times daily. Twice daily administration is most preferred. In an embodiment, the composition may be administered to the patient daily for 1 to 3 weeks, preferably 1 to 2 weeks. The amount of ribose administered to a patient per day is preferably between 1200 and 2400 mg. The amount of glycine administered to a patient per day is preferably between 60 and 150 mg. The amount of alanine administered to a patient per day is preferably between 600 and 1500 mg. The amount of glutamine administered to a patient per day is preferably between 800 and 1800 mg. The relative amounts of amino acids administered to a patient are preferably 1:8 to 12:10 to 15 (glycine:alanine:glutamine). The relative amounts of ribose and the amino acids glycine, alanine, and glutamine administered to a patient are preferably 1:0.8 to 2.0 (ribose:amino acids). The patient is preferably a human patient, and in particular may be an adult over the age of 18.

[0005] In one embodiment, the composition may be used in the treatment of infections caused by bacterial strains with multiple antibiotic resistance, in particular E. coli or Klebsiella pneumoniae bacterial strains with multiple antibiotic resistance. It has previously been proposed in German Patent Application Publication No. 102011105594 that compositions containing ribose, glycine, alanine, and glutamine may have beneficial effects in the treatment of infections caused by bacteria resistant to multiple antibiotics. The method generally involves treating the patient with an antibiotic in addition to simultaneously treating the patient with a composition of the present invention. The composition of the present invention is intended to enhance the effectiveness of the antibiotic. In one embodiment, the composition of the present invention is administered simultaneously with every or every other dose of the antibiotic. In another embodiment, the composition of the present invention is administered at least 30 minutes and at most 5 hours before every or every other dose of the antibiotic. This is because the composition may increase the susceptibility of bacteria to antibiotic treatment, and the composition should be allowed to act on the bacteria before administering the antibiotic.

[0006] The antibiotics administered to the patient may be penicillins, especially ampicillin, cephalosporin antibiotics such as cefazolin, fluoroquinolone antibiotics such as ciprofloxacin, and sulfonamides such as sulfamethoxazole, optionally in combination with trimethoprim. The aqueous solution forming the composition can be obtained by dissolving a mixed lyophilizate of amino acids in an aqueous solution of ribose, which can be obtained by lyophilizing a solution containing the amino acids glycine, alanine, and glutamine in the appropriate ratio. Further details and advantages of the present invention are described with reference to the following examples and drawings. [Brief explanation of the drawings]

[0007] [Figure 1a] FIG. 1 shows plots of results for Strain No. 1 and Antibiotic No. 1 from the in vitro studies described below. [Figure 1b]FIG. 1 shows plots of results from strain no. 1 and antibiotic no. 2 from the in vitro studies described below. [Figure 1c] FIG. 1 shows plots of results from strain no. 1 and antibiotic no. 3 from the in vitro studies described below. [Figure 1d] FIG. 1 shows plots of results from strain no. 1 and antibiotic no. 4 from the in vitro studies described below. [Figure 2a] FIG. 1 shows plots of results for strain no. 2 and antibiotic no. 1 from the in vitro studies described below. [Figure 2b] FIG. 1 shows plots of results for strain no. 2 and antibiotic no. 2 from the in vitro study described below. [Figure 2c] FIG. 1 shows plots of results from strain no. 2 and antibiotic no. 3 from the in vitro studies described below. [Figure 2d] FIG. 1 shows plots of results from strain no. 2 and antibiotic no. 4 from the in vitro studies described below. [Figure 3a] FIG. 1 shows plots of results for strain no. 3 and antibiotic no. 1 from the in vitro studies described below. [Figure 3b] FIG. 1 shows plots of results from strain no. 3 and antibiotic no. 2 from the in vitro studies described below. [Figure 3c] FIG. 1 shows plots of results for strain no. 3 and antibiotic no. 3 from the in vitro studies described below. [Figure 3d] FIG. 1 shows plots of results from strain no. 3 and antibiotic no. 4 from the in vitro studies described below. [Figure 4a] FIG. 1 shows plots of results for strain no. 4 and antibiotic no. 1 from the in vitro studies described below. [Figure 4b] FIG. 1 shows plots of results from strain number 4 and antibiotic number 2 from the in vitro study described below. [Figure 4c] FIG. 1 shows plots of results from strain number 4 and antibiotic number 3 from the in vitro studies described below. [Figure 4d]FIG. 1 shows plots of results from strain number 4 and antibiotic number 4 from the in vitro studies described below. [Figure 5] FIG. 10 shows result plots for in vivo study number 2. DETAILED DESCRIPTION OF THE INVENTION

[0008] composition An aqueous composition was obtained by mixing one vial of the lyophilisate as described in Table 1 below together with two ampoules of a 3.5% solution of D-ribose as described in Table 2 below. [Table 1]

[0009] [Table 2]

[0010] After mixing one vial of lyophilisate and two vials of ribose solution together, a liquid composition according to the description in Table 3 was obtained. [Table 3] The resulting composition was subjected to several in vitro and in vivo mouse model studies.

[0011] In vitro studies The effect of the composition on antibiotic susceptibility was tested in vitro on four resistant bacterial strains. This study was performed as described in Ostrosky EA, Mizumoto MK, Lima MEL, Kaneko TM, Nishikawa SO, Freitas BR Rev Bras Farmacogn (2008) 18:301-307. Each purified bacterial strain was inoculated into a nutrient broth medium and incubated overnight (18-20 hours), then a bacterial suspension was prepared and the turbidity adjusted to 0.2-0.25 at 360 nm. The concentrations of antibiotics used with the bacterial strains during the 8-day treatment are listed in Table 1. Details of the panel of different treatments in the presence of various antibiotics and compositions of the present invention against resistant strains are listed in Table 2.

[0012] [Table 4]

[0013] [Table 5]

[0014] All vials were incubated at 37° C. After the initial 24 hours, the total number of bacteria in each vial was monitored daily using a spectrophotometer at 630 nm. The results are shown in Figures 1a to 4d. In detail, Figure 1a shows the results for strain No. 1 and antibiotic No. 1. Figure 1b shows the results for strain No. 1 and antibiotic No. 2. Figure 1c shows the results for strain No. 1 and antibiotic No. 3. Figure 1d shows the results for strain No. 1 and antibiotic No. 4. Figure 2a shows the results for strain No. 2 and antibiotic No. 1. Figure 2b shows the results for strain No. 2 and antibiotic No. 2. Figure 2c shows the results for strain No. 2 and antibiotic No. 3. Figure 2d shows the results for strain No. 2 and antibiotic No. 4. Figure 3a shows the results for strain No. 3 and antibiotic No. 1. Figure 3b shows the results for strain No. 3 and antibiotic No. 2. Figure 3c shows the results for strain No. 3 and antibiotic No. 3. Figure 3d shows the results for strain No. 3 and antibiotic No. 4. Figure 4a shows the results for strain No. 4 and antibiotic No. 1. Figure 4b shows the results for strain No. 4 and antibiotic No. 2. Figure 4c shows the results for strain No. 4 and antibiotic No. 3. Figure 4d shows the results for strain No. 4 and antibiotic No. 4. As can be seen from the figure, the simultaneous treatment group shows a significant reduction in bacterial growth within 2-3 days of treatment, while treatment with antibiotics alone does not show a significant effect compared to the control. Treatment with the composition of the present invention alone also does not show a significant effect compared to the control. This suggests that the composition of the present invention helps resistant bacteria become more susceptible to antibiotic attack.

[0015] In vivo Animal Study No. 1 A group of Sprague-Dawley rats was infected with an antibiotic-resistant strain of Staphylococcus aureus (MRSA L2-15 ST 13DSMZ46320) (intraperitoneally, 1 × 10 6 CFU / rats augmented with 6% porcine mucin type III). Subgroups of 10 rats (5 males and 5 females) were either untreated, or treated with 10 mg / kg / day (oral) of the antibiotic rifampicin for 10 consecutive days, or co-treated with 10 mg / kg / day (oral) of the antibiotic rifampicin and 250 mg / kg / day (oral) of the composition of the present invention for 10 consecutive days (250 mg refers to the mass of the lyophilized product in Table 1 contained in the amount of mixed solution administered), or co-treated with 10 mg / kg / day (oral) of the antibiotic rifampicin and 500 mg / kg / day (oral) of the composition of the present invention for 10 consecutive days. Survival after 10 days was recorded. In the untreated group, no animals survived after 10 days. In the group treated with antibiotics alone, 2 out of 10 animals survived. In the group treated simultaneously with 250 mg / kg / day of the composition of the present invention, 7 out of 10 animals survived. In the group treated simultaneously with 500 mg / kg / day of the composition of the present invention, all animals survived.

[0016] In vivo Animal Study No. 2 Male Wistar rats weighing 200-250 g were infected with a resistant strain of Escherichia coli (O157.H7) using the granulation pouch model. Specifically, a pouch was created by injecting 30 ml of sterile filtered air, followed by a deep injection of 0.5 ml of 0.5% croton oil dissolved in sesame oil into the dorsal loose connective tissue. Two days later, approximately 10 ml of air was aspirated, and two more days later, 2 ml of a saline solution of 1% peptone and 0.35% agar was injected into the pouch. On day 10, the rats were inoculated with 10 ml of E. coli suspended in 1.5 ml of THB containing 2.5% gastric mucin and 0.35% agar. 6 CFU were infected. The rats were divided into groups of five and administered the following drugs, either alone or simultaneously, every 12 hours for three consecutive days starting 24 hours after infection, for a total of six doses: cotrimoxazole (Septrin, GlaxoSmithKline) at 30 mg / kg (intraperitoneally) and the composition of the present invention at 500 mg / kg (orally) (500 mg refers to the mass of the lyophilized product in Table 1 contained in the amount of mixed solution administered). A control group of five untreated control animals received six oral doses of placebo and intraperitoneal injections of saline at the same intervals.

[0017] Bursal fluid samples (0.5 mL) were aspirated from each animal immediately prior to treatment (time 0), 3 hours after the first dose, immediately prior to the final dose, and 3 and 24 hours after the final dose. Bursal fluid was diluted 10-fold six times with 1% peptone in saline, and 10 μl of each diluted and undiluted bursal fluid was spread in duplicate on the surface of blood agar plates and incubated at 37°C for 24 hours. Results are the mean values ​​obtained at the same intervals from all animals in the same group and expressed as log10 CFU / mL. Following standard analysis of variance, one-way analysis of variance and Bonferroni's multiple comparison test were used to determine significance at the 0.05 level. The efficacy of treatment based on bacterial counts is shown in the figure. There were no significant differences between the control group and the groups treated with the composition of the present invention alone or cotrimoxazole alone at any time interval. A 0.93-log reduction (p≦0.01) in bacterial counts was observed in the groups tested with the composition of the present invention and cotrimoxazole as early as 3 hours after the first dose. This reduction increased with successive doses, reaching a 3.6-log reduction (p≦0.01) before the final dose (6th dose) and sustaining a 4.3-log reduction (p≦0.01) compared to the control group 24 hours after the final dose. When the difference from initial counts was used as the response measure, the groups treated with cotrimoxazole and the composition of the present invention were still significantly lower than the control group at all time points.

[0018] Clinical trials The planned Phase II clinical trial design will include several patients with bacterial urinary tract infections. Patients will be treated for 10 days with a combination of the composition of the present invention (20 ml of the combined solution administered twice daily according to Table 3 above) and the appropriate antibiotic. Results will be evaluated and benchmarked against treatment with antibiotics alone. In detail, the study design is as follows. Phase II Indications: Recurrent urinary tract infections the purpose: Primary Objective: To evaluate the safety of the compositions of the present invention and to evaluate the effect of the compositions of the present invention in combination with antimicrobial treatment on urine cultures (microbiological cure, no microbial growth in 24-48 hour cultures). Secondary objective: To evaluate the effect of the compositions of the present invention in combination with antibacterial treatment on disease-related symptoms (clinical cure, disappearance of symptoms and signs).

[0019] Design: Randomized, double-blind, placebo-controlled, multicenter trial. Patients with recurrent urinary tract infections will receive a blinded oral dose of the composition of the present invention or a matching placebo twice daily for 10 days in addition to an empirical antibiotic. Population: Male and female patients diagnosed with recurrent urinary tract infections over the age of 18. Sample size: 100 patients from each center. Dosage regimen: All patients will receive the following: 1. Trimethoprim-sulfamethoxazole 160 / 800 mg orally twice daily, every 12 hours, for 10 days. 2. A composition of the invention or placebo (according to randomization) twice daily, every 12 hours, orally, for 10 days.

[0020] Efficacy Data: - Complete response is defined as clinical cure (disappearance of symptoms and signs) and microbiological cure (negative urine culture) 5-7 days after the end of treatment. - Incomplete response is defined as clinical recovery (disappearance of symptoms and signs) accompanied by a positive bacterial culture 5-7 days after the end of treatment. - Failure is defined as the absence of a clinical response within 5 days of initiating treatment. - Relapse is the appearance of a new symptom of urinary tract infection 60 ± 10 days after day 1 of study drug administration in a patient who was previously clinically and microbiologically cured with a positive urine culture. Safety Data: Adverse events reported by the subject or observed by the observer.

[0021] statistics: The primary validity analysis will use reliability procedures. Two-sided confidence intervals will be calculated for response frequencies (e.g., Clopper-Pearson method). Paired t-tests will be applied as secondary efficacy analyses. Adverse events will be summarized in frequency tables and categorized by body system. Frequency tables by treatment group will show all treatment-emergent adverse events (TEAEs), all possible related TEAEs, and all adverse events during treatment. These will be analyzed by the intensity of the adverse event, and pre- and post-treatment adverse events will be presented in the subject list. P values ​​will be used to identify potentially significant adverse events. Treatment group comparisons of adverse event frequencies will be examined using Fisher's exact test. Test period: The study is expected to last 18 months. Treatment will last for 10 days, after which each patient will be followed for 8 weeks.

Claims

1. 1. A composition for use in a method of treating a bacterial urinary tract infection in a patient, the composition comprising ribose and the amino acids glycine, alanine, and glutamine in an aqueous solution, the method comprising orally administering the composition to the patient who is being concomitantly treated with an antibiotic or a sulfonamide; the amount of ribose administered to the patient per day is between 1200 and 2400 mg; the amount of glycine administered to the patient per day is between 60 and 150 mg; the amount of alanine administered to the patient per day is between 600 and 1500 mg; and the amount of glutamine administered to the patient per day is between 800 and 1800 mg; the relative amounts of the amino acids administered to the patient are 1:8-12:10-15 (glycine:alanine:glutamine); The composition wherein the relative amounts of the ribose and the amino acids glycine, alanine, and glutamine administered to the patient are 1:0.8-2.0 (ribose:amino acids).

2. 2. The composition for use according to claim 1, wherein the ribose is D-ribose, and / or the alanine is racemic (D / L)-alanine, and / or the glutamine is L-glutamine.

3. the concentration of ribose in the aqueous solution of said composition is between 30 and 40 g / l, and / or the concentration of glycine in the aqueous solution of said composition is between 1.5 and 2.5 g / l, and / or the concentration of alanine in the aqueous solution of said composition is between 15 and 25 g / l, and / or 3. The composition for use according to claim 1 or 2, wherein the concentration of glutamine in the aqueous solution of said composition is between 20 and 30 g / l.

4. 3. The composition for use according to claim 1, wherein the relative concentrations of the amino acids in the aqueous solution of the composition are 1:8-12:10-15 (glycine:alanine:glutamine).

5. 3. The composition for use according to claim 1 or 2, wherein the relative concentrations of the ribose and the amino acids glycine, alanine, and glutamine in the aqueous solution of the composition are 1:0.8-2.0 (ribose:amino acids).

6. The composition for use according to claim 1 or 2, wherein the volume of the aqueous solution of the composition is between 10 and 30 ml.

7. 3. The composition for use according to claim 1 or 2, which is administered to the patient two or three times a day and / or administered to the patient daily for 1 to 3 weeks.

8. 3. The composition for use according to claim 1 or 2, wherein the antibiotic co-administered to the patient is a penicillin, in particular ampicillin, a cephalosporin antibiotic such as cefazolin, or a fluoroquinolone antibiotic such as ciprofloxacin.

9. A composition for use according to claim 1 or 2, wherein the sulfonamide co-administered to the patient is sulfamethoxazole.

10. A composition for use according to claim 1 or 2, wherein the sulfonamide co-administered to the patient is co-administered to the patient in combination with trimethoprim.

11. 3. The composition for use according to claim 1 or 2, wherein the urinary tract infection is caused by a bacterial strain with multiple resistance to antibiotics, in particular Escherichia coli or Klebsiella pneumoniae with multiple resistance to antibiotics.

12. 3. The composition for use according to claim 1 or 2, which is co-administered with every dose or every other dose of an antibiotic.

13. 3. The composition for use according to claim 1 or 2, which is administered at least 30 minutes and at most 5 hours before each or every other dose of the antibiotic.

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