Checkpoint Inhibitor Sensitizes Human Tumor Cells
Exposing a cell to ionizing radiation can result in the formation of DNA double strand breaks. These breaks destabilize the cell’s genomic integrity and must be repaired for cell survival. In response to the damage, proteins become modified (green flags) to inhibit the cell cycle and to direct repair factors to the break site. Cells with normal p53 in the presence or absence of a Chk1 inhibitor are able to repair the damaged DNA. Cells expressing mutated p53, however, fail to properly repair the double strand breaks when treated with the inhibitor.
One unfortunate and detrimental side effect of ionizing radiation as a treatment for cancer is the damage it imparts to normal tissue near the targeted tumor. Technology has improved radiation delivery, minimizing the volume of normal tissue in the radiation field, but has not eliminated it completely. Thus, the identification of drugs that increase the sensitivity of cancer cells to radiation while sparing normal cells would go a long way toward improving patient quality of life and outcome.
When actively growing cells are exposed to radiation, two important checkpoints are activated to block further progression through the cell cycle, allowing the cell to repair any damaged DNA. Drugs inhibiting these checkpoints could prevent DNA repair pathways and eventually lead to cell death, making them ideal targets for altering a cell’s susceptibility to radiation. Unlike normal cells, half of human tumors have altered p53, an important protein that controls the first checkpoint. James Mitchell, Ph.D. and researchers from the Radiation Biology and Radiation Oncology Branches decided to focus their efforts on a novel inhibitor of the protein Chk1, which regulates the second checkpoint.
Their study, published in Clinical Cancer Research, showed that treating various cancer cell lines expressing p53 mutations with the Chk1 inhibitor significantly increased cell death following exposure to radiation. Interestingly, cells with normal p53 showed the same survival rate after radiation with or without the drug. This suggests that cells with altered p53 become sensitized to radiation in the presence of the drug while normal cells do not.
Mitchell and colleagues then wanted to understand how the inhibitor sensitized the p53 mutant cells. They first investigated whether the drug affected the cell cycle arrest triggered by radiation exposure. Measuring the amount of DNA in p53 normal and mutant cells treated with radiation and the inhibitor together demonstrated that the drug prevented cell cycle arrest in both cell types.
Since the Chk1 inhibitor caused universal cell cycle progression, the researchers next determined the ability of the cells to repair radiation-induced DNA damage in the presence of the agent. In p53 normal cells, immediately following radiation and drug treatment, there was an increase in a protein modification indicative of DNA damage. After 24 hours the modified protein returned to pre-radiation levels demonstrating the completion of DNA repair. In the mutant p53 cells, however, there was an extended up-regulation of the modified protein, even at 24 hours, suggesting a block in DNA damage repair. This inhibition of repair resulted in increased nuclear condensation, a marker of cell death.
Finally, the researchers tested the effectiveness of the drug in an animal tumor model. Radiation treatment of tumors that were established by injecting p53 mutant cells into immune compromised mice somewhat slowed tumor growth. The added injection of the inhibitor just after radiation exposure and 8 hours later significantly decreased tumor growth. Treatment with the inhibitor also prolonged the modification of the DNA damage response protein in the tumor similar to that observed in the cell lines.
These studies successfully demonstrated the efficacy of blocking Chk1, a regulator of the second cell cycle checkpoint, as a means of sensitizing p53 mutant cancer cells to radiation in cell lines and in the mouse. This novel inhibitor could now be tested in patients to see whether tumors expressing mutant p53 can also be sensitized to radiation by slowing DNA damage repair, and if this can improve patient outcomes.
Summary Posted: 06/2010
Clin Cancer Res. 2010 16(7):2076-84. PubMed link
Reviewed by Jennifer Crawford, PhD
- Understanding Papillary Renal Cell Carcinoma
- Insight into Prostate Cancer Stem Cells
- Naïve-Derived T Cell Adoptive Immunotherapy is Impaired by Memory T Cells
- Identifying Stem-like Cells Using Mitochondrial Membrane Potential
- Noncoding RNA Shows Context-Dependent Function
- T Cells that Recognize HPV Protein Can Target Virus-Infected Cells
- Pancreatic Cancer Sensitive to Selective p38 Pathway Inhibition
- Mutations Allow JC Polyomaviruses to Elude Antibody Recognition
- Mitochondrial DNA Unwinding Enzyme Required for Liver Regeneration
- Regulatory RNA Key Player in p53-Mediated Apoptosis in Embryonic Stem Cells
- New Method for Producing Significant Amounts of RNA Labeled at Specific Sites
- Mutant HABP2 Causes Non-Medullary Thyroid Cancer
- Paradigm Shift in Thyroid Hormone Mechanism of Action
- Changes in Bacteria Induce Inflammatory Skin Diseases