title: A single capsule formulation of RHB-104 demonstrates higher anti-microbial growth potency for effective treatment of Crohn's disease associated withMycobacterium aviumsubspeciesparatuberculosis.
abstract: BACKGROUND
  Most recently we reported that RHB‑104 triple antibiotics combination in culture is bactericidal and should be effective for treatment of Crohn's disease (CD)-associated withMycobacterium aviumsubspeciesparatuberculosis(MAP) (Alcedo et al. in Gut Pathog 14:32, 2016). The combination exhibited unique synergistic antimicrobial growth activity. The proprietary RHB-104 capsule formulation contains active ingredients (63.3 % Clarithromycin (CLA), 6.7 % Clofazimine (CLO) and 30 % Rifabutin (RIF)). In our earlier study, we could not dissolve the proprietary RHB-104 capsule formulation in one compatible solvent. Consequently, we re-created RHB-104 analog by adding appropriate concentrations of each of the three antibiotics into the cultures. The Minimum inhibitory concentration (MIC) for RHB-104 analog, CLA, CLO, RIF, CLA-CLO, CLA-RIF, CLO-RIF and their individual solvents were reported earlier (Alcedo et al. in Gut Pathog 14:32, 2016). In this study, we succeeded in dissolving the proprietary RHB-104 capsule formulation in a single proprietary solvent. This study is designed to compare of the MIC the proprietary RHB-104 capsule formulation to RHB-104 analog against MAP and other microorganisms.
  
  
  METHODS
  BD Bactec™ MGIT™ Para-TB medium (Sparks, MD) system was used to determine the MIC of the proprietary RHB-104 capsule formulation and RHB-104 analog and their solvents against MAP and several other microorganisms. The final concentration of solvents used to dissolve all the drugs were ≤0.5 % (v/v).
  
  
  RESULTS
  The MIC for the RHB-104 proprietary solvent against MAP was consistent against all microorganisms tested in the study at 12.5 % (v/v). The MIC for the proprietary RHB-104 capsule formulation was similar to RHB-104 analog against several MAP clinical strains with MIC ≤ 0.2 μg/mL. The MIC for the proprietary RHB-104 capsule formulation was at 2.0 μg/mL against MAP strain MS 137 andM. aviumstrain JF7 compared to 4.0 ug/mL for RHB-104 analog. Similarly, the MIC of RHB-104 formulation capsule was significantly lower than RHB-104 analog againstM. tuberculosisHR237,M. fortuitismsubspeciesfortuitum, M. smegmatisATCC 27199,Staphylococcus aureusATCC 25923 andListeria monocytogenesATCC 19112.
  
  
  CONCLUSIONS
  The data demonstrated that the proprietary RHB-104 capsule formulation is more potent in culture against Mycobacteria and other microorganisms especially those with MIC >0.2. Formulation of multi-drugs in a single capsule results in potent synergistic anti-microbial activity far exceeds treatment the culture with multi-individually dissolved drugs. RHB-104 capsule formulation should be more effective to eradicate MAP infection in patients with CD. The study provides evidence that combining weak antibiotics in one formulation might be the new silver bullet to combat bacteria.
authors.affiliation: Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, 4110 Libra Drive, Orlando, FL USA.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Qasem
authors.affiliation: Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, 4110 Libra Drive, Orlando, FL USA.
authors.forename: Mitra
authors.initials: M
authors.lastname: Safavikhasraghi
authors.affiliation: Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, 4110 Libra Drive, Orlando, FL USA.
authors.forename: Saleh A
authors.initials: SA
authors.lastname: Naser
keywords: Antibiotics; Crohn’s disease; IBD; MIC; Mycobacterium paratuberculosis (MAP); RHB-104