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Comparison of Cellular Death Pathways after mTHPC-mediated Photodynamic Therapy (PDT) in Five Human Cancer Cell Lines
Carsten Lange1, Christiane Lehmann2, Martin Mahler3
1Department of Pharmaceutical and Medicinal Chemistry, Institute of Pharmacy, University of Greifswald, Friedrich-Ludwig-Jahn-Straße 17, 17489 Greifswald, Germany. langec@uni-greifswald.de.
Abstract:
One of the most promising photosensitizers (PS) used in photodynamic therapy (PDT) is the porphyrin derivative 5,10,15,20-tetra(m-hydroxyphenyl)chlorin (mTHPC, temoporfin), marketed in Europe under the trade name Foscan®. A set of five human cancer cell lines from head and neck and other PDT-relevant tissues was used to investigate oxidative stress and underlying cell death mechanisms of mTHPC-mediated PDT in vitro. Cells were treated with mTHPC in equitoxic concentrations and illuminated with light doses of 1.8-7.0 J/cm2 and harvested immediately, 6, 24, or 48 h post illumination for analyses. Our results confirm the induction of oxidative stress after mTHPC-based PDT by detecting a total loss of mitochondrial membrane potential (Δψm) and increased formation of ROS. However, lipid peroxidation (LPO) and loss of cell membrane integrity play only a minor role in cell death in most cell lines. Based on our results, apoptosis is the predominant death mechanism following mTHPC-mediated PDT. Autophagy can occur in parallel to apoptosis or the former can be dominant first, yet ultimately leading to autophagy-associated apoptosis. The death of the cells is in some cases accompanied by DNA fragmentation and a G2/M phase arrest. In general, the overall phototoxic effects and the concentrations as well as the time to establish these effects varies between cell lines, suggesting that the cancer cells are not all dying by one defined mechanism, but rather succumb to an individual interplay of different cell death mechanisms. Besides the evaluation of the underlying cell death mechanisms, we focused on the comparison of results in a set of five identically treated cell lines in this study. Although cells were treated under equitoxic conditions and PDT acts via a rather unspecific ROS formation, very heterogeneous results were obtained with different cell lines. This study shows that general conclusions after PDT in vitro require testing on several cell lines to be reliable, which has too often been ignored in the past.
Insights
Photodynamic therapy using mTHPC (temoporfin) induces oxidative stress and apoptosis in cancer cells. However, cell death mechanisms vary significantly across different cell lines, highlighting the need for multi-cell line testing in PDT research.
Area of Science:
- Biochemistry
- Cell Biology
- Oncology
Background:
- 5,10,15,20-tetra(m-hydroxyphenyl)chlorin (mTHPC, temoporfin) is a promising photosensitizer for photodynamic therapy (PDT).
- Understanding the precise mechanisms of cell death induced by mTHPC-mediated PDT is crucial for optimizing its clinical application.
- Investigating oxidative stress and cell death pathways in various cancer cell lines provides insights into treatment efficacy.
Purpose of the Study:
- To investigate the oxidative stress and cell death mechanisms induced by mTHPC-mediated photodynamic therapy (PDT) in vitro.
- To compare the cellular responses across a panel of five human cancer cell lines.
- To determine the predominant cell death pathways and the role of oxidative stress in mTHPC-based PDT.
Main Methods:
- Treatment of five human cancer cell lines with mTHPC in equitoxic concentrations.
- Illumination with specific light doses (1.8-7.0 J/cm²).
- Analysis of mitochondrial membrane potential, reactive oxygen species (ROS) formation, lipid peroxidation (LPO), cell membrane integrity, apoptosis, autophagy, and cell cycle progression at various time points post-illumination.
Main Results:
- mTHPC-PDT confirmed induction of oxidative stress, evidenced by loss of mitochondrial membrane potential and increased ROS formation.
- Apoptosis was identified as the predominant cell death mechanism, with autophagy occurring in parallel or preceding apoptosis.
- Significant heterogeneity in phototoxic effects and cell death mechanisms was observed across the five cell lines, despite equitoxic treatment conditions.
Conclusions:
- mTHPC-mediated PDT primarily induces apoptosis, often in conjunction with autophagy, and triggers significant oxidative stress.
- Cancer cell lines exhibit diverse responses to mTHPC-PDT, indicating that cell-specific interplay of death mechanisms dictates outcomes.
- Reliable conclusions regarding PDT efficacy necessitate testing across multiple cell lines to account for inherent cellular variability.
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