Abstract
Recently, it has been reported that anti-viral drugs, such as indinavir and lopinavir (originally targeted for HIV), also inhibit E6-mediated proteasomal degradation of mutant p53 in E6-transfected C33A cells. In order to understand more about the mode-of-action(s) of these drugs the metabolome of HPV16 E6 expressing cervical carcinoma cell lines was investigated using mass spectrometry (MS)-based metabolic profiling. The metabolite profiling of C33A parent and E6-transfected cells exposed to these two anti-viral drugs was performed by ultra performance liquid chromatography (UPLC)-MS and gas chromatography (GC)-time of flight (TOF)-MS. Using a combination of univariate and multivariate analyses, these metabolic profiles were investigated for analytical and biological reproducibility and to discover key metabolite differences elicited during anti-viral drug challenge. This approach revealed both distinct and common effects of these two drugs on the metabolome of two different cell lines. Finally, intracellular drug levels were quantified, which suggested in the case of lopinavir that increased activity of membrane transporters may contribute to the drug sensitivity of HPV infected cells.
Highlights
It has been reported that anti-viral drugs, such as indinavir and lopinavir, inhibit E6-mediated proteasomal degradation of mutant p53 in E6-transfected C33A cells
Intracellular metabolite changes caused by indinavir and lopinavir in the presence and absence of HPV16 E6 against an isogenic background (C33A cervical cancer cells) were investigated using mass spectrometry (MS)-based metabolomics
We confirm that the change in the levels of phenylalanine, 2-octenedioic acid,butyrylcarnitine and deoxyguanosinemonophosphate in drug challenged C33AP and E6 cells could be thought of as general drug effects on both cell lines whereas a reduction of the levels of proline, indoline and methionine in only lopinavir exposed C33AE6 cells represents E6 oncogene specified effects of the drug
Summary
It has been reported that anti-viral drugs, such as indinavir and lopinavir (originally targeted for HIV), inhibit E6-mediated proteasomal degradation of mutant p53 in E6-transfected C33A cells. Using a combination of univariate and multivariate analyses, these metabolic profiles were investigated for analytical and biological reproducibility and to discover key metabolite differences elicited during anti-viral drug challenge. This approach revealed both distinct and common effects of these two drugs on the metabolome of two different cell lines. Whilst GC-MS can detect a range of volatile compounds, non-polar fatty acids, or primary metabolites (when derivatised), UPLC-MS is more suited to the analysis of secondary metabolites and lipids when C18 reversed-phase LC is applied Since both MS techniques are fully automated they are amenable to high-throughput metabolomics analysis. UPLCMS and GC-MS have been used widely to investigate metabolic changes in biological processes, to discover new biomarkers and drugs, and diagnose diseases.[13,14,15,16,17,18,19,20,21]
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