Amphotericin has remained the powerful but highly toxic last line of defense in treating life-threatening fungal infections in humans for over 50 years with minimal development of microbial resistance. Understanding how this small molecule kills yeast is thus critical for guiding development of derivatives with an improved therapeutic index and other resistance-refractory antimicrobial agents. In the widely accepted ion channel model for its mechanism of cytocidal action, amphotericin forms aggregates inside lipid bilayers that permeabilize and kill cells. In contrast, we report that amphotericin exists primarily in the form of large, extramembranous aggregates that kill yeast by extracting ergosterol from lipid bilayers. These findings reveal that extraction of a polyfunctional lipid underlies the resistance-refractory antimicrobial action of amphotericin and suggests a roadmap for separating its cytocidal and membrane-permeabilizing activities. This new mechanistic understanding is also guiding development of the first derivatives of amphotericin that kill yeast but not human cells.

Multiple human diseases ensue from a hereditary or acquired deficiency of iron-transporting protein function that diminishes transmembrane iron flux in distinct sites and directions. Because other iron-transport proteins remain active, labile iron gradients build up across the corresponding protein-deficient membranes. Here we report that a small molecule natural product, hinokitiol, can harness such gradients to restore iron transport into, within, and/or out of cells. The same compound promotes gut iron absorption in DMT1-deficient rats and ferroportin-deficient mice, as well as hemoglobinization in DMT1- and mitoferrin-deficient zebrafish. These findings illuminate a general mechanistic framework for small molecule-mediated site- and direction-selective restoration of iron transport. They also suggest small molecules that partially mimic the function of missing protein transporters of iron, and possibly other ions, may have potential in treating human diseases.

Loss-of-function mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) compromise epithelial HCO 3 − and Cl − secretion, reduce airway surface liquid (ASL) pH, and impair respiratory host defenses in people with cystic fibrosis (CF) 1 – 3 . Here we report that apical addition of an unselective ion channel-forming small molecule, amphotericin B (AmB), restored HCO 3 − secretion and increased ASL pH in cultured human CF airway epithelia. These effects required the basolateral Na + /K + ATPase, indicating that apical AmB channels functionally interfaced with this driver of anion secretion. AmB also restored ASL pH, viscosity, and antibacterial activity in primary cultures of airway epithelia from people with CF caused by different mutations, including ones that yield no CFTR, and increased ASL pH in CFTR -null pigs in vivo . Thus, unselective small molecule ion channels can restore CF airway host defenses via a mechanism that is CFTR-independent and therefore genotype-independent.

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Deficiencies of protein ion channels underlie many currently incurable human diseases. Robust networks of pumps and channels are usually responsible for the directional movement of specific ions in organisms ranging from microbes to humans. We thus questioned whether minimally selective small molecule mimics of missing protein channels might be capable of collaborating with the corresponding protein ion pumps to restore physiology. Here we report vigorous and sustainable restoration of yeast cell growth by replacing missing protein ion channels with imperfect small molecule mimics. We further provide evidence that this tolerance for imperfect mimicry is attributable to collaboration between the channel-forming small molecule and protein ion pumps. These results illuminate a mechanistic framework for pursuing small molecule replacements for deficient protein ion channels that underlie a range of challenging human diseases.