Antibiotic resistance is an ever-increasing problem for modern chemotherapy of bacterial infectious diseases. In light of the limited pipeline of new antibacterials, drug-resistant pathogens are a clear and urgent danger to public health and national biodefense. Recent evidence suggests that the bacterial RecA protein's functions, which are carried out through two distinct protein conformation states (termed A and P), allow bacteria to overcome or adapt to antibacterial actions. Because of the inherent complexity of these processes involving interconnected networks of genes and gene products, recA mutants are pleiotropic. We hypothesize that functionally selective small-molecule probes capable of modulating predetermined RecA functions would be useful in teasing apart the relative contributions of its myriad activities to key biological processes involved in bacterial survival or adaptation to antibacterial therapies. Moreover, small-molecule inhibitors of RecA may serve as leads for the development of combination chemotherapies for bacterial infectious diseases. A prototypical RecA inhibitor was previously discovered from a screen of a multi- thousand compound library. This represents the first small molecule with demonstrated inhibitory activity in live Escherichia coli. Furthermore, the prototype demonstrated functional selectivity by only inhibiting RecA P-state activities. The purpose of the proposed research is to initiate a program that utilizes the prototypical inhibitor as a platform for the design, synthesis, and biological evaluation of analogous RecA inhibitors that selectively modulate recombinogenic processes in pathogenic bacteria. Specifically, we aim to (1) evaluate the abilities of a first-generation pilot library of prototype analogues to inhibit genetic recombination processes in pathogenic bacteria;(2) synthesize and biochemically evaluate second-generation analogues to identify improved RecA inhibitors;(3) characterize second-generation RecA inhibitors against E. coli utilizing robust microbiological and biochemical assays;and (4) characterize second-generation RecA inhibitors against Neisseria gonnorhoea and Streptococcus pneumonia. As functionally selective P-state inhibitors, these small-molecule probes will allow us to identify and characterize specific roles that are ascribed to the A- and P-state conformations of RecA in the development and transmission of antibiotic resistance. Additionally, we expect the research program to serve as platform enabling the exploitation of RecA as a therapeutic target. Realization of the goals outlined in this proposal will have a significant impact on the research areas of bacterial infections (how bacteria adapt to antibacterial actions), sexually transmitted diseases (study of genetic recombination in N. gonnorhoea), and overall global health (study of various disease states from S. pneumonia infections). Furthermore, the research should make important contributions to the fields of drug development (identification of non-classical targets, potentiation of known antibacterials) and biodefense development.
Antibiotic resistance is an ever-increasing problem for modern chemotherapy of bacterial infectious diseases. In light of the limited pipeline of new antibacterials, drug-resistant pathogens are a clear and urgent danger to public health and national biodefense. The bacterial RecA protein is an emerging target for adjuvants for antibiotic chemotherapy that attenuate the development and transmission of antibiotic resistance genes and increase the antibiotic therapeutic index. It is envisioned that the proposed plan will lead to small-molecule probes that selectively inhibit predetermined functions of the RecA protein in live bacteria, thus affording greater understanding of the protein's roles in various aspects of bacterial pathogenicity - including the de novo development and transmission of antibiotic resistance genes. This new knowledge will significantly impact approaches to sexually transmitted disease, biodefense, and overall global public health related to infectious disease. The research platform will also increase the efficiency of the discovery of small- molecule effectors that address the urgent unmet need to overcome antibiotic resistance in biothreat and other pathogenic bacteria.
