Significance The global problem of TB has worsened in recent years with the emergence of drug-resistant organisms, and new drugs are clearly needed. In a cell-based high-throughput screen, a small molecule, TCA1, was discovered that has activity against replicating and nonreplicating Mycobacterium tuberculosis . It is also efficacious in acute and chronic rodent models of TB alone or combined with frontline TB drugs. TCA1 functions by a unique mechanism, inhibiting enzymes involved in cell wall and molybdenum cofactor biosynthesis. This discovery represents a significant advance in the search for new agents to treat persistent and drug-resistant TB.

Genotoxic chemotherapy is the most common cancer treatment strategy. However, its untargeted generic DNA-damaging nature and associated systemic cytotoxicity greatly limit the therapeutic applications. Here, we employed a haploid genetic screen in human cells to discover an absolute dependency of the clinically evaluated anti-cancer compound YM155 on SLC35F2, an uncharacterized member of the solute carrier protein family that is highly expressed in a variety of human cancers. YM155 generated DNA damage through intercalation, which was contingent on the expression of SLC35F2 and its drug importing activity. SLC35F2 expression and YM155 sensitivity correlated across a panel of cancer cell lines and targeted genome editing verified SLC35F2 as the main determinant of YM155-mediated DNA damage toxicity in vitro and in vivo.These findings suggest a novel route to targeted DNA damage by exploiting tumor and patientspecific import of YM155.

Benzothiazinones (BTZs) form a new class of potent antimycobacterial agents. Although the target of BTZs has been identified as decaprenylphosphoryl-β-D-ribose 2'-epimerase (DprE1), their detailed mechanism of action remains obscure. Here we demonstrate that BTZs are activated in the bacterium by reduction of an essential nitro group to a nitroso derivative, which then specifically reacts with a cysteine residue in the active site of DprE1.

Benzothiazinones (BTZs) are antituberculosis drug candidates with nanomolar bactericidal activity against tubercle bacilli. Here we demonstrate that BTZs are suicide substrates of the FAD-dependent decaprenylphosphoryl-β-D-ribofuranose 2′-oxidase DprE1, an enzyme involved in cell-wall biogenesis. BTZs are reduced by DprE1 to an electrophile, which then reacts in a nearquantitative manner with an active-site cysteine of DprE1, thus providing a rationale for the extraordinary potency of BTZs. Mutant DprE1 enzymes from BTZ-resistant strains reduce BTZs to inert metabolites while avoiding covalent inactivation. Our results explain the basis for drug sensitivity and resistance to an exceptionally potent class of antituberculosis agents.T he increasing number of drug-resistant Mycobacterium tuberculosis strains that fail to respond to first-and secondline drug treatment demands the development of new antituberculosis drugs. 1−3 Benzothiazinones (BTZs) such as BTZ043 ( Figure 1A) are a promising class of new compounds that kill M. tuberculosis in vitro, ex vivo, and in mouse models of tuberculosis. 4 The minimal inhibitory concentration (MIC) of BTZ043 against M. tuberculosis is 1 ng/mL, which is significantly lower than the MICs of all currently used tuberculosis drugs and drug candidates. Decaprenylphosphoryl-β-D-ribofuranose 2′-epimerase was identified as a BTZ target ( Figure 1B). 4 The enzyme is constituted of DprE1 and DprE2 that together catalyze the epimerization of decaprenylphosphoryl-β-D-ribofuranose (DPR) to decaprenylphosphoryl-β-Darabinofuranose (DPA), the arabinosyl donor for the biosynthesis of mycobacterial cell wall arabinan polymers. 4−6 The reaction is believed to proceed via the keto intermediate decaprenylphosphoryl-D-2′-keto-erythro-pentofuranose (DPX) ( Figure 1B). 5 The Cys387Gly and Cys387Ser point mutations in DprE1 result in 250-and 10 000-fold increases in the MIC, respectively. 4 We previously isolated a covalent adduct of DprE1 and BTZ043 from mycobacteria incubated with BTZ043 and proposed a mechanism of action involving reduction of the essential nitro group of BTZ043 to a nitroso group that then reacts with Cys387 of DprE1 to form a stable semimercaptal ( Figure 1A). 7 However, the mechanism of action of BTZs still poses numerous questions, as it is unclear how BTZs are activated and what the basis of the specificity of the proposed nitroso derivative for DprE1 is. Furthermore, it has not been elucidated whether the observed modification of DprE1 indeed affects the activity of DprE1 or functions by inhibiting the activity of DprE2 (or both); neither have the exact roles of DprE1 and DprE2 in the epimerization reaction been validated.A more detailed characterization of the mechanism of action of BTZs requires the availability of pure DprE1 and DprE2. As our previous attempts to purify recombinant DprE1 and DprE2 of M. tuberculosis H37Rv in their active form were unsuccessful,