Creative Biolabs conducts a panel of well-designed in vitro assays to elaborate the mechanism of action of potential antifungal compounds, especially in respect of protein synthesis inhibition.
Fungal cell envelope (plasma membrane and cell wall) has long been taken as the primary target for developing most antifungal therapies, for their well-characterized structure, easy access, and adequate pathogenicity. However, the increasing impact of fungal infections along with drug resistance has greatly driven development and discovery of novel targets. Since the 1980s, thanks to the emerging technology of whole-cell sequencing and high throughput screening of natural products, a new class of agents was found to inhibit protein synthesis in fungi. These chemicals include sodarins (blocking the function of fungal translation elongation factor 2), myristate analogues (N-myristylation of fungal proteins), azoxybacilin (inhibiting gene expression of sulfite reductase), and cispentacin (perturbing transport and metabolism of amino acid).
In the effort to discover novel antifungal compounds by uncovering the MOA of protein synthesis inhibition potential, Creative Biolabs has launched an exclusive technical system, based on profound knowledge of fungi physiology and a comprehensive range of assays.
Two key factors must be taken into consideration during MOA verification. First, potential cross-reactivity between fungi and mammalian species must be validated in each step. Owing to the eukaryotic nature of fungi, therapeutic targets shall be restricted with high selectivity toward fungal components, which do not overlap with mammalian counterparts. Cross-reactivity can cause severe toxic side effects. Second, since many similar antifungal chemicals exert species-specific effects, the inhibitory ability and sensitivity should be evaluated among different fungi species. This can not only better characterize the effective spectra of susceptible species but also provide additional mechanical insight.
We offer integrated investigation service including but not limited to following steps:
Growth Inhibition
Overall growth inhibition and toxicity rate can be generated using growth inhibition assay of mammalian cell model and fungi species of interest. IC50 values can generally assess the whole-cell toxicity and sensitivity.
Whole-cell Protein Synthesis Labeling
By radiolabeling of substrate amino acids, interference with whole-cell protein synthesis will be monitored and examined with high accuracy.
Cell-free Transcription/Translation Test
Our well-established in vitro transcription/translation platform can validate the specific inhibiting mode of action, using cell-free lysate capable of protein synthesis and luciferase reporter system. This enables in-depth verification of MOA of test articles in subcellular or even molecular level.
Expression Profile
In case your compounds display varied sensitivity among different species, further information can be provided by analyzing the expression levels and structural variants of the target components. One example is that ER2, which poses as the primary inhibition target for sodarins, varies in certain amino acid residues and thereby exhibits species-specific sensitivity.
To fully illustrate the mechanism of action of your compound, Creative Biolabs is always a trustworthy and reliable partner, working closely with you in every step of your project. For more detailed information, please feel free to contact us or directly sent an inquiry.
Reference
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