Development of Inhibitors Against Mycobacterium Abscessus tRNA (m1G37) Methyltransferase (TrmD) Using Fragment-Based Approaches

Abstract

Mycobacterium abscessus (Mab) is a rapidly growing species of multidrug-resistant nontuberculous mycobacteria (NTM) that has emerged as a growing threat to individuals with cystic fibrosis, and other pre-existing chronic lung diseases. Mab pulmonary infections are difficult, or sometimes impossible, to treat and result in accelerated lung function decline and premature death. There is therefore an urgent need to develop novel antibiotics with improved efficacy. tRNA (m1G37) methyltransferase (TrmD) is a promising target for novel antibiotics. It is essential in Mab and other mycobacteria, improving reading frame maintenance on the ribosome to prevent frameshift errors. In this work a fragment-based approach was employed with the merging of two fragments bound to the active site, followed by structure-guided elaboration to design potent nanomolar inhibitors against Mab TrmD. Several of these compounds exhibit promising activity against mycobacterial species, including Mycobacterium tuberculosis and Mycobacterium leprae in addition to Mab, supporting the use of TrmD as a target for the development of antimycobacterial compounds.

Fragment-based discovery of a new class of inhibitors targeting mycobacterial tRNA modification

“…Structure-guided elaboration of the merged compound AW1 ( K d 0.11 mM, LE 0.36, IC 50 0.23 mM) was performed, initially utilizing the indole nitrogen as a vector for growth. While the detailed medicinal chemistry strategy of all the compounds and intermediates are described separately ( 37 ) and in supporting information (Schemes 1–4 and Supplementary Table S3 ), in this section we illustrate the key features in the fragment hit to lead optimization of TrmD inhibitors. The addition of a 2-picolyl moiety successfully increased the affinity of AW1 by an order of magnitude in compound AW2 ( K d 12 μM, LE 0.30, IC 50 33 μM) (Table 1 and Supplementary Figures S5 and S6 ).…”

“…ITC experiments to quantify binding of ligands to TrmD were done as described in ( 37 ) using Malvern MicroCal iTC200 or Auto-iTC200 systems at 25°C. Titrations consisted of an initial injection (0.2 μl), discarded during data processing, followed by either 19 (2 μl) or 39 (1 μl) injections separated by intervals of 60–150 s duration.…”

“…A more in-depth discussion around the medicinal chemistry strategy for fragment merging and lead compound development is described in a corresponding publication by Whitehouse et al. ( 37 ). The compounds AW1-7 are listed in this publication as follows: AW1 (Compound 23 ), AW2 (Compound 24f ), AW3 (Compound 26f ), AW4 (Compound 28 ), AW5 (Compound 29a ), AW6 (Compound 31a ) and AW7 (Compound 29d ).…”

“…Briefly, mycobacteria were grown to optical density ( A 600 nm ) of 0.2–0.3 in liquid culture and 1 × 10 5 bacteria were added to each well of 96-well plates containing serial dilutions of compound (400, 200, 100, 50, 25, 12.5, 6.3, 3.1, 1.6, 0.8, 0.4, 0 μM), in triplicate wells per condition, and incubated at 37°C until growth was seen in the control wells. MIC measurements using M. tuberculosis H37Rv were performed as reported in ( 37 ). M. tuberculosis H37Rv was grown in Middlebrook 7H9 base containing 14 mg/l dipalmitoyl phosphatidylcholine (DPPC), 0.81 g/l NaCl, 0.3 g/l casitone, and 0.05% Tyloxapol.…”

Evolutionary repair reveals an unexpected role of the tRNA modification m1G37 in aminoacylation

“…TrmD is a high-priority antibiotic target and development of TrmD inhibitors is currently an active area of research (15)(16)(17)(18)(19)(20)(21). Several factors have established TrmD as a promising antibiotic target.…”