Evading persistent drug resistance in cancer and bacteria is quintessential to restore health in humans, and impels intervention strategies. A distinct property of the cancer phenotype is enhanced glucose metabolism and oxidative stress. Reactive oxygen species (ROS) are metabolic byproducts of aerobic respiration and are responsible for maintaining redox homeostasis in cells. Redox balance and oxidative stress are orchestrated by antioxidant enzymes, reduced thiols and NADP(H) cofactors, which is critical for cancer cells survival and progression. Similarly, Escherichia coli (E. coli) and life-threatening infectious pathogens such as Staphylococcus aureus (SA) and Mycobacterium tuberculosis (Mtb) are appreciably sensitive to changes in the intracellular oxidative environment. Thus, small molecules that modulate antioxidant levels and/or enhance intracellular ROS could disturb the cellular oxidative environment and induce cell death, and hence could serve as novel therapeutics. Presented here are a collection of approaches that involve ROS modulation in cells as a strategy to target cancer and bacteria.

Regeneration of myelin is mediated by oligodendrocyte progenitor cells-an abundant stem cell population in the central nervous system (CNS) and the principal source of new myelinating oligodendrocytes. Loss of myelin-producing oligodendrocytes in the CNS underlies a number of neurological diseases, including multiple sclerosis and diverse genetic diseases. High-throughput chemical screening approaches have been used to identify small molecules that stimulate the formation of oligodendrocytes from oligodendrocyte progenitor cells and functionally enhance remyelination in vivo. Here we show that a wide range of these pro-myelinating small molecules function not through their canonical targets but by directly inhibiting CYP51, TM7SF2, or EBP, a narrow range of enzymes within the cholesterol biosynthesis pathway. Subsequent accumulation of the 8,9-unsaturated sterol substrates of these enzymes is a key mechanistic node that promotes oligodendrocyte formation, as 8,9-unsaturated sterols are effective when supplied to oligodendrocyte progenitor cells in purified form whereas analogous sterols that lack this structural feature have no effect. Collectively, our results define a unifying sterol-based mechanism of action for most known small-molecule enhancers of oligodendrocyte formation and highlight specific targets to propel the development of optimal remyelinating therapeutics.

Mycobacterium tuberculosis (Mtb) has evolved protective and detoxification mechanisms to maintain cytoplasmic redox balance in response to exogenous oxidative stress encountered inside host phagocytes. In contrast, little is known about the dynamic response of this pathogen to endogenous oxidative stress generated within Mtb. Using a noninvasive and specific biosensor of cytoplasmic redox state of Mtb, we for first time discovered a surprisingly high sensitivity of this pathogen to perturbation in redox homeostasis induced by elevated endogenous reactive oxygen species (ROS). We synthesized a series of hydroquinone-based small molecule ROS generators and found that ATD-3169 permeated mycobacteria to reliably enhance endogenous ROS including superoxide radicals. When Mtb strains including multidrug-resistant (MDR) and extensively drug-resistant (XDR) patient isolates were exposed to this compound, a dose-dependent, long-lasting, and irreversible oxidative shift in intramycobacterial redox potential was detected. Dynamic redox potential measurements revealed that Mtb had diminished capacity to restore cytoplasmic redox balance in comparison with Mycobacterium smegmatis (Msm), a fast growing nonpathogenic mycobacterial species. Accordingly, Mtb strains were extremely susceptible to inhibition by ATD-3169 but not Msm, suggesting a functional linkage between dynamic redox changes and survival. Microarray analysis showed major realignment of pathways involved in redox homeostasis, central metabolism, DNA repair, and cell wall lipid biosynthesis in response to ATD-3169, all consistent with enhanced endogenous ROS contributing to lethality induced by this compound. This work provides empirical evidence that the cytoplasmic redox poise of Mtb is uniquely sensitive to manipulation in steady-state endogenous ROS levels, thus revealing the importance of targeting intramycobacterial redox metabolism for controlling TB infection.

Highlights d A small-molecule screen identified novel enhancers of oligodendrocyte formation d Most top compounds target cholesterol biosynthesis enzymes CYP51, TM7SF2, or EBP d Evaluation of analogs identified CW3388 as a potent, selective EBP inhibitor d CW3388 is poised for further optimization toward EBPtargeting remyelinating drugs