A chemical genetic screen for modulators of asymmetrical 2,2'-dimeric naphthoquinones cytotoxicity in yeast.
BACKGROUND
Dimeric naphthoquinones (BiQ) were originally synthesized as a new class of HIV integrase inhibitors but have shown integrase-independent cytotoxicity in acute lymphoblastic leukemia cell lines suggesting their use as potential anti-neoplastic agents. The mechanism of this cytotoxicity is unknown. In order to gain insight into the mode of action of binaphthoquinones we performed a systematic high-throughput screen in a yeast isogenic deletion mutant array for enhanced or suppressed growth in the presence of binaphthoquinones.


RESULTS
Exposure of wild type yeast strains to various BiQs demonstrated inhibition of yeast growth with IC(50)s in the microM range. Drug sensitivity and resistance screens were performed by exposing arrays of a haploid yeast deletion mutant library to BiQs at concentrations near their IC(50). Sensitivity screens identified yeast with deletions affecting mitochondrial function and cellular respiration as having increased sensitivity to BiQs. Corresponding to this, wild type yeast grown in the absence of a fermentable carbon source were particularly sensitive to BiQs, and treatment with BiQs was shown to disrupt the mitochondrial membrane potential and lead to the generation of reactive oxygen species (ROS). Furthermore, baseline ROS production in BiQ sensitive mutant strains was increased compared to wild type and could be further augmented by the presence of BiQ. Screens for resistance to BiQ action identified the mitochondrial external NAD(P)H dehydrogenase, NDE1, as critical to BiQ toxicity and over-expression of this gene resulted in increased ROS production and increased sensitivity of wild type yeast to BiQ.


CONCLUSIONS
In yeast, binaphthoquinone cytotoxicity is likely mediated through NAD(P)H:quonine oxidoreductases leading to ROS production and dysfunctional mitochondria. Further studies are required to validate this mechanism in mammalian cells.
Department of Internal Medicine and Oncology, Johns Hopkins School of Medicine, Baltimore, Maryland, USA.
Ashkan
A
Emadi
Ashley E
AE
Ross
Kathleen M
KM
Cowan
Yolanda M
YM
Fortenberry
Milena
M
Vuica-Ross
D000332:Aerobiosis / Q000187:drug effects; D003470:Culture Media; D005285:Fermentation / Q000187:drug effects; D005820:Genetic Testing; D016681:Genome, Fungal / Q000235:genetics; D007536:Isomerism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D050296:Microbial Viability / Q000187:drug effects*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009285:Naphthoquinones / Q000737:chemistry / Q000633:toxicity*; D017382:Reactive Oxygen Species / Q000378:metabolism; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
D003470:Culture Media; D009285:Naphthoquinones; D017382:Reactive Oxygen Species; D029701:Saccharomyces cerevisiae Proteins