The ultimate goal of this proposal and any subsequent phase II proposal is to develop a unique protein agent to prevent Clostridium difficile associated diseases in those patients at high risk rather than wait to treat their dangerous and costly infections. We aim in this SBIR phase I project to determine the feasibility of oral delivery of a lead, specifically targeted bactericidal protein to eliminate C. difficile cariage without untended collateral damage to the intestinal microbiota. The lead candidate, termed a "diffocin", kills 27 of a collection of 28 BI/NAP1/027 strains; and Lawley et al (2010) have described a mouse model of C. difficile carriage. We have recently shown that another engineered R-type bacteriocin administered orally to rabbits can transit the GI tract to act as a specific bactericidal agent killing another bacterial pathogen in the terminal ileum and colon. Thus, after improving the lab scale production of this lead recombinant protein in B. subtilis and conducting preliminary pharmacology and pharmacodynamic studies to guide dosing, we shall evaluate the efficacy of diffocins in C57Bl/6 mice that are asymptomatic carriers and shedders of a sensitive strain of C. difficile. If diffocins eliminate C. difficile from carrier mice, the efect of oral diffocins on the mouse intestinal microbiota will be determined by ribotyping the fecal microbiome of normal and C. difficile carrier mice after diffocin administration.
Public Health Relevance: Clostridium difficile is a bacterium that can reside in human intestines and not cause disease until the healthy bacteria sharing the intestinal space are damaged by antibiotics. Because Clostridium difficile bacteria are usually resistant to antibiotics they can then proliferate, make potent toxins, and cause severe diarrhea and potentially lethal inflammation of the colon. Such infections are now more common in hospitals than MRSA. We have discovered, cloned and made a protein that specifically kills the most toxic form of Clostridium difficile bacteria. We intend to determine whether this agent, a "diffocin", when administered orally is capable of eliminating Clostridium difficile bacteria residing innocuously i the intestine of a mouse model of the human infection. If so, we plan to develop this bacteria-killing protein as an agent to kill Clostridium difficile present in patients before they receive antibiotics and thereby prevent the severe, recurring infections without damaging the healthy bacteria of the gut.
Public Health Relevance Statement: Clostridium difficile is a bacterium that can reside in human intestines and not cause disease until the healthy bacteria sharing the intestinal space are damaged by antibiotics. Because Clostridium difficile bacteria are usually resistant to antibiotics they can then proliferate, make potent toxins, and cause severe diarrhea and potentially lethal inflammation of the colon. Such infections are now more common in hospitals than MRSA. We have discovered, cloned and made a protein that specifically kills the most toxic form of Clostridium difficile bacteria. We intend to determine whether this agent, a "diffocin", when administered orally is capable of eliminating Clostridium difficile bacteria residing innocuously i the intestine of a mouse model of the human infection. If so, we plan to develop this bacteria-killing protein as an agent to kill Clostridium difficile present in patients before they receive antibiotics and thereby prevent the severe, recurring infections without damaging the healthy bacteria of the gut.
NIH Spending Category: Biodefense; Digestive Diseases; Emerging Infectious Diseases; Infectious Diseases
Project Terms: Antibiotic Resistance; Antibiotics; Antibodies; Bacteria; bactericide; Bacteriocin Typing; Biological Assay; Clinical; Clostridium difficile; Collection; Colon; Diarrhea; Disease; Distal part of ileum; Dose; Drug Kinetics; Engineering; Gastrointestinal tract structure; Goals; Health; high risk; Hospitals; Human; improved; in vivo; Incidence; Infection; Inflammation; innovation; Intestines; Killings; Lead; methicillin resistant Staphylococcus aureus (organism); microbiome; mouse model; Mus; North America; Nosocomial Infections; Oral; Oryctolagus cuniculus; pathogen; Patients; Pharmacodynamics; Pharmacology; Phase; pre-clinical; prevent; Process; Production; Proliferating; Proteins; Recombinant Proteins; Ribotypes; Ribotyping; Rifles; Role; Shotguns; Small Business Innovation Research Grant; Source; Specificity; System; Toxicology; Toxin
