Molecular mechanism of the toxicity of the antifungal antibiotic amphotericin B
Research project objectives/Hypothesis
Interdisciplinary research, at the interface of medicinal biology and molecular biophysics will be carried out, aimed to unveil molecular mechanisms responsible for the severe toxic side effects of antifungal antibiotic amphotericin B (AmB). This antibiotic belongs to the group of life saving pharmaceuticals and despite of its nephrotoxicity and hepatotoxicity is used for treatment of deep seated mycotic infections due to its effectiveness. The main objective of the project is unveiling and understanding of molecular mechanisms responsible for AmB toxicity, in particular, verification of our working hypothesis based on the results of our pilot studies. According to this hypothesis:
Amphotericin B molecules, in water environment, self-organize spontaneously into the forms of dimers which can further associate into tetramers. The surface of lipid membranes can act as a catalytic platform for the tetramer formation. After incorporation into lipid bilayers, tetrameric structures of AmB function as transmembrane ion channels affecting electrophysiological homeostasis of living cells. Such a mechanism is one of the major causes responsible for severe side effects for patients.
The research planned will be carried out with the use of the molecular spectroscopy and molecular imaging techniques applied to single model lipid membrane objects (liposomes) and single transport protein molecules (“single molecule” approach). In particular, the research planned will be based on FLIM (Fluorescence Lifetime Imaging Microscopy), FCS (Fluorescence Correlation Spectroscopy), Raman imaging microscopy coupled with AFM (Atomic Force Microscopy), CD (Circular Dichroism) spectroscopy and FTIR (Fourier-Transform Infrared) absorption spectroscopy. The detailed research tasks are planned to unveil and understand: (i) molecular mechanisms responsible for self-organization of AmB to dimers and tetramers, in the water phase and in the environments of transport proteins and lipid membranes, (ii) impact of individual AmB molecular organization forms on lipid membrane structure and transmembrane ion transfer, (iii) possibilities of developing of new pharmacological formulas of AmB, based on protein and lipid structures, to prevent formation of tetrameric structures. In parallel, the research will be conducted (iv), based on molecular dynamics calculations of the process of AmB tetramer formation in the lipid membrane environment.
Research project impact
The planned research will result in obtaining precise information on molecular mechanisms responsible for formation of supramolecular structures of AmB, which can be responsible for toxic side effects of the antibiotic. This knowledge will allow designing of chemical modifications of the drug and engineering of new pharmaceutical formulations which retain therapeutic efficacy with substantially reduced toxicity for patients.
Pioneering nature of the research project
The technically highly advanced “single molecule” approach will be applied to gain precise knowledge on AmB self-organization and on impact of individual molecular structures of the drug on membrane properties and physiological functionality. The hypothesis on AmB toxicity based on the transmembrane-localized specific tetrameric structures is highly innovative and consistent with both the current knowledge and all the results of the pilot studies.