Cancer Research and Mitochondria

Cancer, Cell Death, and Mitochondria

Apoptosis is a protective defense mechanism that effectively suppresses tumor growth and eliminates tumor cells.
One of the main mechanisms that trigger apoptosis is the increase in mitochondrial metabolic activity, which leads to elevated ROS levels in cancer cells. Excessive ROS damage mitochondrial function, causing mitochondrial membrane depolarization, which subsequently activates the intrinsic apoptosis pathway. Tumor cell immune evasion is a key feature of tumor pathophysiology, and mitochondria play a central role in both inhibiting and promoting immune evasion within the  complex mechanism    s involved in immune responses.1)

The Potential of Ferroptosis in Cancer Therapy: Many studies have found that ferroptosis sensitivity can be used to target tumors resistant to conventional therapies (such as triple-negative breast cancer and glioblastoma).2)

Ferroptosis and the Immune Microenvironment: Neutrophils in the tumour microenvironment die spontaneously by ferotosis and the lipid peroxide released suppresses T-cell activity, thereby suppressing tumour immunity.3)

Reference
1) Gao, J., Cancer Gene Therapy, 2024, 31, 970-983
2) Yang, F., Cell Metabolism, 2023, 35(1), 84-100
3) Kim, R., Nature, 2022, 612, 338-346


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Cancer and Metabolism

Cancer cells undergo significant metabolic reprogramming to support their rapid growth and survival. This altered metabolism is a hallmark of cancer and is essential for tumor development, progression, and therapy resistance. Key metabolic changes in cancer include:

1、Warburg Effect: Cancer cells often rely on glycolysis for energy production, even in the presence of oxygen, a phenomenon known as the Warburg effect. This shift allows for faster ATP production and the generation of metabolic intermediates necessary for biosynthesis, despite being less efficient than oxidative phosphorylation.

2、Mitochondrial Function and Metabolic Flexibility: Mitochondria in cancer cells are highly dynamic, adapting to various metabolic stresses. These adaptations enable cancer cells to switch between different energy sources (such as fatty acids, glucose, and glutamine) depending on the environment, ensuring continued growth and survival.

3、Altered Lipid Metabolism: Cancer cells frequently modify their lipid metabolism to meet the demands for membrane synthesis and energy storage. Increased lipid synthesis and uptake of fatty acids are common in tumors and support cancer cell proliferation and survival.

4、Amino Acid Metabolism: Cancer cells often exhibit altered amino acid metabolism, including increased consumption of glutamine and other amino acids, which supports protein synthesis and energy production.

Despite a century of research, the exact mechanisms by which cells integrate glycolysis and oxidative phosphorylation remain unclear, and this issue is even more complex at the individual cell level. Studies have shown that in certain tissues, one specific cell type tends to preferentially perform glucose glycolysis, while an adjacent cell type prefers to utilize lactate through mitochondrial respiration to produce ATP.


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Experimental Example: Changes in various indicators of cell death induced by drugs

HepG2 cells treated with the apoptosis-inducing agent staurosporine or the ferroptosis-inducing agents Erastin and RSL3. After treatment, extracellular LDH, phosphatidylserine, cell viability, intracellular Fe2+ and lipid peroxidation were determined.

The results showed that apoptosis-induced cells treated with staurosporine showed an increase in phosphatidylserine, a decrease in cell viability and an increase in extracellular LDH, indicating that cell death had occurred. On the other hand, intracellular Fe2+, an indicator of ferroptosis, remained unchanged. In cells treated with Erastin, a ferroptosis inducer, intracellular Fe2+ increased and cell viability decreased, but extracellular LDH and lipid peroxidation (lipid peroxidation: decrease in red fluorescence and increase in green fluorescence) did not increase. In cells in which ferroptosis was more strongly induced by co-treatment with RSL3 in addition to Erastin, increased intracellular Fe2+ and lipid peroxidation were observed. Moreover, decreased cell viability and increased dead cells were detected. Meanwhile, phosphatidylserine showed a lower rate of increase during ferroptosis induction compared to apoptosis-induced cells. These results suggest that cell death can be distinguished by evaluating a combination of cell death indicators.

Experimental Example: Comparison of metabolic pathway dependence in different cell lines

Products:
Glycolysys/OXPHOS - G270 Glycolysis/OXPHOS Assay Kit
OCR - E297 Extracellular OCR Plate Assay Kit

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

*Please refer to the "Supplementary information on technology and products used" section on the right for additional information on the results.

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Experimental Example：Analysis of Apoptosis-Induced Changes in OCR and Mitochondrial Membrane Potential

Products:
Dead Cell Detection - C555 Dead Cell Makeup Blue - Higher Retention than PI
Mitochondrial Membrane Potential Detection - MT09 JC-1 MitoMP Detection Kit
OCR - E297 Extracellular OCR Plate Assay Kit

1. HepG2 and Jurkat cells were treated with Staurosporine (4µM) to induce apoptosis.

2. Using OCR measurement reagents, mitochondrial membrane potential detection reagent (JC-1), and a dead cell detection reagent,we analyzed the time-dependent changes in OCR and mitochondrial membrane potential associated with apoptosis induction.

3. In Jurkat cells, cell death was observed in a time-dependent manner after Staurosporine treatment, and mitochondrial activity (membrane potential and OCR) decreased with increasing cell death. On the other hand, 4 µM Staurosporine treatment of HepG2 cells showed no cell death at any treatment time and a decrease in basal mitochondrial respiration, but rather an increase in maximal respiratory activity with shorter Staurosporine treatment.

 

 

 

 

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