Final Report

1. Project objective

RNA-polymerase is a validated target for the treatment of tuberculosis represented by Rifampicin.  There are, however, some Mycobacterium tuberculosis (Mtb) strains resistant to Rifampicin.  In addition, Rifampicin is not the easiest agent to use in drug combination therapies because it causes induction of CYP enzymes. This project was based on a novel natural product recently reported in the literature.  Our plan was to preselect microorganisms that would contain a certain biosynthetic gene sequence of the producing microorganism of this natural product.

2. Project design

Operationally, this was a four-way collaboration involving, Daiichi Sankyo RD Novare, University of Illinois at Chicago (UIC), Texas A&M University, and TB Alliance.  Selection of the producing organisms and preparation of the extracts was carried out by Daiichi Sankyo RD Novare.  Our objective for the first year was to demonstrate its feasibility and to identify at least one fraction of interest for an additional investigation involving further scale-up fermentation and characterization of the active components.

3. Results, lessons learned

Approximately 300 extracts were screened using the target enzyme assays and a phenotypic screening against Mtb.  We selected 4 producing organisms at the end based on our predetermined decision tree.  Fractionation of the extracts from these organisms has eliminated known natural product inhibitors of RNA-polymerase and left one fraction that could contain a novel RNA-polymerase inhibitor.  Completion of this work would require scaling up of the fermentation and isolation of the active components in the second year, but this one-year investigation has established the feasibility of this screening paradigm.  This approach can enhance the productivity of screening natural products based on the combination of genome information of the producing organisms, the enzymatic assays, and phenotypic screening against Mtb.