New potent orally bioavailable nitro-containing compounds to treat mycobacterial infections - Diseases caused by mycobacteria, including tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) and non-tuberculous mycobacterial (NTM) infections caused by M. abscessus (Mab), are challenging to treat due to intrinsic drug tolerance and acquired multi-drug resistance. Mtb requires treatment of up to 6 months, while NTM requires up to 18 months and is often treated as a chronic disease with recurrence for multiple years. Some NTM infections are fatal with no effective cure. Both diseases are treated with multiple antibiotics with significant side effects highlighting a tremendous need for both new TB and NTM drugs. We discovered a new series of nitrofurans, parent compound HC2210, that are potent inhibitors of both Mtb and Mab. When HC2210 was administered orally in a chronic murine model of TB infection, HC2210 reduced Mtb burden by >10-fold in the lungs and spleens of infected mice. These proof-of-concept data show HC2210 is orally bioavailable, tolerable and efficacious in a chronic mouse model of Mtb infection, attaining a critical milestone in the development of a new antimicrobial. Initial structure-activity-relationship (SAR) studies, including the synthesis of ~110 HC2210 analogs, has defined even more potent analogs, with whole cell EC50 of 2 nM and 24 nM against Mtb and Mab, respectively. Compared to the potencies of pretomanid against Mtb (~620 nM EC50) and amikacin against Mab (~5000 nM EC50), these new nitro-containing compounds are >300-fold and >200-fold more potent, respectively. In Aim 1, we will define SAR and optimize activity of HC2210 analogs (Aim 1.1); conduct integrated SAR and pharmacokinetic studies to prioritize compounds for in vivo efficacy studies (Aim 1.2); and, demonstrate in vivo efficacy of HC2210 analogs against Mtb and Mab infections (Aim 1.3). Resistant mutants isolated in both Mtb and Mab support that HC2210 is a prodrug, dependent on Ddn, an F420-dependent nitroreductase, for its activation. Upon metabolism by Ddn, intracellular nitric oxide (NO) is released which poisons the electron transport chain. Mammalian cells, and most other bacteria, do not carry the activating nitroreductases, ensuring that the analogs do not release NO until reaching the infecting mycobacteria. Interestingly, mutations in ddn provide full resistance to pretomanid, but only moderate resistance to HC2210, suggesting that other nitro reductases may activate HC2210. Indeed, several new analogs only have activity against Mtb or Mab and we hypothesize the differential activity may be driven by differential interactions with Mtb vs. Mab nitroreductases. In Aim 2, we will define mechanisms of resistance and drug penetration (Aim 2.1), examine combination drug therapies (Aim 2.2) and post-antibiotic effect (Aim 2.3), which will allow the design of analogs that are more durable to resistance evolution and have improved activity specifically for Mtb or Mab. OVERALL IMPACT: This focused, 3-year project will optimize novel Mtb and Mab inhibitors, define their function in vivo and identify combinations of compounds or new analogs that will provide key data required for a go-no-go decision for further preclinical development of this series.
