The work described in the application is necessary for furthering the development of a new therapeutic for treating Gram-positive infections. Preliminary in vitro and in vivo activity studies of novel analogs of mutacin 1140 suggest that they hold great promise as an antimicrobial product for clinical use. Mutacin 1140 is produced by the bacterium Streptococcus mutans JH1140 and the bacterium has been engineered to synthesize novel analogs of the native compound. Our studies have revealed that select mutacin 1140 analogs have a rapid bactericidal activity and that they are superior to native compound in terms of inhibitory activity, serum stability, and performance in an in vivo infection study. Further, the analogs have been shown to be more effective than vancomycin in treating a systemic MRSA infection. Mutacin 1140 analogs have no observable toxicity in mice at a 50 mg/kg intravenous dose. Furthermore, the analogs were shown to significantly reduce bacterial load of MRSA in the kidneys and liver of infected mice in an acute infection study. All these studies point to the need to further the preclinical development of one or more of the novel analogs of mutacin 1140. The major impediment to furthering investigational studies on these unique antibacterial compounds is the poor yield from culture liquor. The goal of this application is to enhance the yield and purity of mutacin analogs following fermentation. Three approaches will be explored to increase the yield of the antibacterial compound. First, media components will be screened to determine their effect on the production levels of the antibacterial compound. Second, a molecular based approach will be used to determine whether an increase in production of the analogs is possible through genetic manipulation of S. mutans. Our third approach is transposon or ethyl-methanesulfonate (EMS) random mutagenesis coupled with next-generation DNA sequencing to identify mutations that lead to enhanced production. The last component of the application is to determine the most effective means of extracting and purifying the antibacterial compound from the fermentation liquor. These experiments will enable exploratory studies for defining a novel drug product for investigational new drug (IND) studies.
Public Health Relevance Statement: Project Narrative The proposal addresses the need for the development of novel treatments for Gram- positive infections. We will investigate means of enhancing the production and purification of lead analogs of the antibacterial compound mutacin 1140. This work is necessary for promoting the production of sufficient amounts of the analogs for the completion of the required preclinical studies that are needed for filing an Investigational New Drug (IND) application.
Project Terms: acute infection; Address; Anabolism; analog; Anti-Bacterial Agents; Antibiotics; antimicrobial; Bacteria; Bacterial Infections; bactericide; base; Biological; Biomedical Engineering; Bioreactors; Carbon; Cell Density; Centers for Disease Control and Prevention (U.S.); Cessation of life; Child; Clinical; Community Healthcare; Coupled; Culture Media; Development; distilled alcoholic beverage; DNA sequencing; Dose; Drug Kinetics; Drug resistance; Elderly; Engineering; Ethyl Methanesulfonate; experience; experimental study; Fermentation; Financial Hardship; Gene Cluster; Genetic; genetic manipulation; genomic locus; Genomics; Goals; Gram-Positive Bacteria; Gram-Positive Bacterial Infections; Growth; Healthcare Systems; Human; improved; In Vitro; in vivo; Incidence; Individual; Infection; Intravenous; Investigation; Investigational Drugs; Investigational New Drug Application; Kidney; Knowledge; Lead; Liver; Measures; methicillin resistant Staphylococcus aureus; Methods; Molecular; Morbidity - disease rate; mortality; Multiple Bacterial Drug Resistance; Mus; Mutagenesis; mutant; Mutation; Natural Products; next generation; Nitrogen; novel; novel therapeutics; Oxygen; pathogen; Patients; peptide analog; Peptides; Performance; Pharmaceutical Preparations; preclinical development; preclinical study; Predisposition; Production; promoter; Regulatory Element; Research; Resistance; Ribosomes; Sepsis; Serum; Sodium Chloride; Source; Staphylococcus aureus; Staphylococcus aureus infection; Streptococcus mutans; Streptococcus pneumoniae; System; Systemic infection; Techniques; Temperature; Therapeutic Agents; Time; Toxic effect; United States; Vancomycin; Work
