Quantitative evaluation of berberine subcellular distribution and cellular accumulation in non-small cell lung cancer
Zhong-Wen Yuan1, Elaine Lai-Han Leung1, Xing-Xing Fan1
1State Key Laboratory of Quality Research in Chinese Medicines, Macau Institute for Applied Research in Medicine and Health, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macau.
Abstract:
Berberine, an isoquinoline alkaloid, has been demonstrated to be a safe anti-cancer agent with multiple effects on mitochondria. Intracellular concentration and distribution around the targeting sites are determinants of efficacy, but subcellular distribution of berberine has not been fully elucidated yet, which relies on the sensitive and robustness assay. In this study, a sensitive and robust UPLC-MS/MS method has been developed and validated with optimized extraction solvents and detection conditions. Key factors such as the purity and integrity of isolated organelle fractions, and the effects of isolation procedures on the subcellular concentration of berberine were systemically evaluated. With the developed assay, we found that the intracellular accumulations of berberine in two gefitinib resistant NSCLC cell lines H1650 and H1975 were 2-3 folds higher than that of normal epithelial cells BEAS-2B. Moreover, significantly different subcellular distribution profiles in NSCLC cancer cells from that of BEAS-2B cells with a striking increase in content in most organelles may contribute to its selective cytotoxicity to cancer cells. Furthermore, a predominant accumulation of berberine was observed for the first time in microsomal fraction for all three cell lines. Therefore, this method could be used for quantitative evaluation of subcellular distribution and cellular accumulation of berberine and for further evaluation of the concentration-effects relationship.
Insights
Berberine accumulates more in cancer cells than normal cells, with higher concentrations found in specific cell parts. This finding aids in understanding berberine
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Berberine, an isoquinoline alkaloid, shows anti-cancer properties and affects mitochondria.
- Understanding berberine's intracellular concentration and distribution is crucial for its efficacy.
- Subcellular distribution of berberine remains largely unelucidated, necessitating sensitive assays.
Purpose of the Study:
- To develop and validate a sensitive and robust UPLC-MS/MS method for quantifying berberine.
- To investigate the subcellular distribution of berberine in normal and non-small cell lung cancer (NSCLC) cells.
- To evaluate the impact of isolation procedures on berberine's subcellular concentration.
Main Methods:
- Development and validation of a UPLC-MS/MS assay for berberine quantification.
- Optimization of extraction solvents and detection conditions.
- Systematic evaluation of organelle fraction purity and isolation procedure effects on berberine concentration.
Main Results:
- Intracellular berberine accumulation was 2-3 times higher in gefitinib-resistant NSCLC cell lines (H1650, H1975) compared to normal BEAS-2B cells.
- Significantly different subcellular distribution profiles were observed in NSCLC cells versus BEAS-2B cells.
- A predominant accumulation of berberine was identified in the microsomal fraction across all tested cell lines.
Conclusions:
- The developed UPLC-MS/MS method is suitable for quantitative evaluation of berberine's subcellular distribution and cellular accumulation.
- Differential subcellular distribution, particularly increased organelle content in NSCLC cells, may explain berberine's selective cytotoxicity.
- Findings provide a basis for further studies on the concentration-effect relationship of berberine in cancer therapy.
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