Novel antifungal activity of broxyquinoline against ITC-Resistant mutants via inhibition of methionine aminopeptidase in Aspergillus fumigatus.

BACKGROUND

A rising public health threat of invasive aspergillosis causing by azole-resistant Aspergillus species necessitates the development of novel antifungal agents. We performed a screening of antifungal compounds against azole-resistant fungi and broxyquinoline was identified. Our aim was to investigate its antifungal activities and antifungal mechanism.

METHODS

Fungal susceptibility testing for broxyquinoline carried out by the broth-based microdilution methods. Cell viability treated by broxyquinoline was tested by resazurin dye testing. The synergistic effect of broxyquinoline and itraconazole was evaluated using checkboard assay. Enzyme activity was investigated with spectrophotometric methods. The key amino acid residues in the binding of broxyquinoline with A. fumigatus methionine aminopeptidase (AfMetAP) were indicated by structural analyses, site-directed mutagenesis, and microscale thermophoresis (MST) assays.

RESULTS

Broxyquinoline has fungicidal activity against azole-resistant A. fumigatus and synergistic effects with itraconazole. Moreover, we discovered that broxyquinoline is strong inhibitor of AfMetAP through interacting with key residues T279A/W280A of AfMetAP1 and Y527A of AfMetAP2, and in vivo, the strains with high expression of AfMetAP are resistant to BQ.

CONCLUSIONS

Broxyquinoline has antifungal activities against azole-resistant Aspergillus spp. and is also a potentiator of ITC in vitro. AfMetAP may biologically targets to BQ in vivo regulating antifungal activity. T279A/W280A of AfMetAP1 and Y527A of AfMetAP2 are key amino acid sites to maintain stable interactions between broxyquinoline and target enzymes AfMetAP.