Thiolactomycin-Based Inhibitors of Bacterial β-Ketoacyl-ACP Synthases with in Vivo Activity

Abstract

β-Ketoacyl-ACP synthases (KAS) are key enzymes involved in the type II bacterial fatty acid biosynthesis (FASII) pathway and are putative targets for antibacterial discovery. Several natural product KAS inhibitors have previously been reported, including thiolactomycin (TLM), which is produced by Nocardia spp. Here we describe the synthesis and characterization of optically pure 5R-thiolactomycin (TLM) analogues that show improved whole cell activity against bacterial strains including methicillin-resistant Staphylococcus aureus (MRSA) and priority pathogens such as Francisella tularensis and Burkholderia pseudomallei. In addition, we identify TLM analogues with in vivo efficacy against MRSA and Klebsiella pneumoniae in animal models of infection.

Figure 1

Fatty Acid Biosynthesis pathway in E. coli. β-Ketoacyl-ACP synthases (FabB, FabF and FabH) catalyze the decarboxylative Claisen condensation of malonyl-ACP with the acyl primer. While FabB and FabF are responsible for the elongation step of FASII, FabH initiates FASII by synthesizing acetoacetyl-ACP. Thiolactomycin (TLM), a natural product thiolactone, preferentially inhibits the FabB and FabF β-ketoacyl-ACP synthases.

Figure 2

Average survival time of mice infected with, A) Methicillin resistant S. aureus BAA1762 and treated with vehicle (●, n=7), TLM (■, n=7), TLM 5 (▲, n=6), TLM 6 (▼, n=6), TLM 11 (◆, n=6) and vancomycin (○, n=6); B) K. pneumoniae KPC2ST258 and treated with vehicle (●, n=10), TLM (■, n=8), TLM 3 (▲, n=8), TLM 5 (▼, n=8), TLM 6 (◆, n=8) and Gentamicin (▽, n=10). The infected animals were monitored twice a day for 5 days post infection for the MRSA infection model and 7 days for the K. pneumoniae infection model. The average survival time was calculated at the end of each study. The one-tailed t test was used to calculate the p value of each data set with respect to the vehicle control.

Scheme 1

Synthesis of building blocks 32 and 1.

Scheme 2

Synthesis of C3 analogs of 5R-(+)-Thiolactomycin (TLM 2 – 4, 12 – 16, 19)

Scheme 3

Synthesis of C4 analogs of 5R-(+)-Thiolactomycin (TLM 22 – 25)

Scheme 4

Synthesis of C3 analogs of 5R-(+)-Thiolactomycin (TLM 17, 18, 20, 21)

Scheme 5

Synthesis of C3 analogs of 5R-(+)-Thiolactomycin (TLM 5–11, 26)