CRISPR-Based Screening Implicates Mitochondrial Phosphatase Gene in Hypoxic Survival of Liver Cancer Cells

Hypoxia is irrefutably important in the development of solid tumors. The hypoxic environment within the tumor results from low blood supply and the tumor limiting the opportunities for the organism’s immune system to attack it. This condition also applies to solid tumors resulting from Hepatocellular carcinoma (HCC).

Primary liver cancer due to HCC is the 4th most common and 2nd most fatal cancer globally. HCC is hard to fight due to difficulties in early detection. Current treatments are restricted to tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors like Nivolumab for advanced HCC patients.

Solid cancers, like HCC, are frequently oxygen deprived due to the lack of blood supply. In the case of these cancers, a transcription factor, hypoxia-inducible factor (HIF), allows cells to adapt to hypoxia. Contrary to the traditional idea that mitochondria are “useless” in hypoxia, a new study investigated whether mitochondrial integrity is essential to cell survival in hypoxia.

Study Results

By performing a genome-wide CRISPR-Cas9 knock out library (GeCKO v2A library) screening, researchers identified PTPMT1 (protein-tyrosine phosphatase mitochondrial 1) as the third most significant gene for hypoxic adaptation, ranking right after HIF-1α and HIF-1β.

PTPMT1 is a phosphatase that is necessary for cardiolipin (CL) synthesis. CL is a major component of the mitochondrial membranes and is critical in forming mitochondrial cristae and stable complexes in the electron transport chain (ETC). In the event of CL not functioning, the mitochondrial cristae and ETC experience issues that allow ROS to build up and increase oxidative stress on the cell.

When HCC cell lines were treated with PTPMT1 inhibitor, Alexidine dihydrochloride, the authors observed that PTPMT1 was more effective in killing hypoxic vs. normoxic cells. The results from this experiment lead the researchers to the conclusion that the hypoxic cells were more vulnerable to oxidative stress than the normoxic cells. Although a more specific PTPMT1 inhibitor would be preferred, AD was a good proof of concept to demonstrate the translational implications of the library screening.

By using the human GeCKO v2A library, comprising 65,386 unique single-guide RNAs (sgRNAs) targeting 19,052 protein-coding genes and 1,864 microRNAs, the researchers identified HIF-1α and HIF-1β as the first and second most essential genes for hypoxic adaptation with PTPMT1 as the third.

Implications

This study revealed the metabolic advantages of PTPMT1 in hypoxic cells as well as highlight its clinical relevance in solid cancers. This has strong possibilities in developing new treatments that can be selected for hard tumor cancer cells and not have the same negative effects of chemotherapy. The selectivity created by PTPMT1 is incredibly valuable and is part of the next generation of high specificity cancer drugs. This knowledge will help in the development of therapeutic strategies for hypoxic HCC cells and other cancer cells.

Related Article: China’s Regulatory Body Accepts Application for First Selective Internal Radiation Therapy for HCC

References

1. https://pubmed.ncbi.nlm.nih.gov/33503428/

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Author

Eduardo Longoria