Effect of a poly(ADP-ribose) polymerase-1 inhibitor against esophageal squamous cell carcinoma cell lines
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Tomomitsu Nasuno1, Sachiyo Mimaki, Makito Okamoto
1Division of Translational Research, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan; Department of Otorhinolaryngology, University of Kitasato Hospital, Minami-ku, Sagamihara, Kanagawa, Japan.
Abstract:
Effective molecular target drugs that improve therapeutic efficacy with fewer adverse effects for esophageal cancer are highly anticipated. Poly(ADP-ribose) polymerase (PARP) inhibitors have been proposed as low-toxicity agents to treat double strand break (DSB)-repair defective tumors. Several findings imply the potential relevance of DSB repair defects in the tumorigenesis of esophageal squamous cell carcinoma (ESCC). We evaluated the effect of a PARP Inhibitor (AZD2281) on the TE-series ESCC cell lines. Of these eight cell lines, the clonogenic survival of one (TE-6) was reduced by AZD2281 to the level of DSB repair-defective Capan-1 and HCC1937 cells. AZD2281-induced DNA damage was implied by increases in γ-H2AX and cell cycle arrest at G2/M phase. The impairment of DSB repair in TE-6 cells was suggested by a sustained increase in γ-H2AX levels and the tail moment calculated from a neutral comet assay after X-ray irradiation. Because the formation of nuclear DSB repair protein foci was impaired in TE-6 cells, whole-exome sequencing of these cells was performed to explore the gene mutations that might be responsible. A novel mutation in RNF8, an E3 ligase targeting γ-H2AX was identified. Consistent with this, polyubiquitination of γ-H2AX after irradiation was impaired in TE-6 cells. Thus, AZD2281 induced growth retardation of the DSB repair-impaired TE-6 cells. Interestingly, a strong correlation between basal expression levels of γ-H2AX and sensitivity to AZD2281was observed in the TE-series cells (R(2) = 0.5345). Because the assessment of basal DSB status could serve as a biomarker for selecting PARP inhibitor-tractable tumors, further investigation is warranted.
Insights
Poly(ADP-ribose) polymerase (PARP) inhibitors show promise for esophageal cancer. AZD2281 effectively inhibited growth in esophageal squamous cell carcinoma cells with DNA repair defects, identified by a novel RNF8 mutation.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Esophageal cancer lacks effective molecularly targeted therapies with reduced side effects.
- Poly(ADP-ribose) polymerase (PARP) inhibitors are emerging as low-toxicity agents for tumors with DNA double-strand break (DSB) repair defects.
- DSB repair defects are implicated in esophageal squamous cell carcinoma (ESCC) tumorigenesis.
Purpose of the Study:
- To evaluate the efficacy of the PARP inhibitor AZD2281 in ESCC cell lines.
- To investigate the underlying mechanisms of PARP inhibitor sensitivity in ESCC.
- To identify potential biomarkers for predicting treatment response.
Main Methods:
- Screening of eight ESCC cell lines (TE-series) for sensitivity to AZD2281.
- Assessing DNA damage (γ-H2AX), cell cycle arrest, and DSB repair capacity (neutral comet assay) after AZD2281 treatment.
- Performing whole-exome sequencing to identify genetic mutations in sensitive cell lines.
- Analyzing the correlation between basal γ-H2AX expression and AZD2281 sensitivity.
Main Results:
- One ESCC cell line (TE-6) exhibited reduced clonogenic survival upon AZD2281 treatment, similar to known DSB repair-defective cells.
- AZD2281 induced DNA damage and G2/M cell cycle arrest in TE-6 cells.
- TE-6 cells showed impaired DSB repair, evidenced by sustained γ-H2AX, increased tail moment in comet assays, and deficient DSB repair protein foci formation.
- Whole-exome sequencing revealed a novel mutation in RNF8, an E3 ligase involved in γ-H2AX ubiquitination, which was impaired in TE-6 cells.
- A significant correlation (R² = 0.5345) was observed between basal γ-H2AX expression levels and AZD2281 sensitivity across the TE-series cell lines.
Conclusions:
- AZD2281 effectively inhibits the growth of ESCC cells with impaired DSB repair, linked to a novel RNF8 mutation.
- Basal expression of γ-H2AX may serve as a predictive biomarker for identifying PARP inhibitor-tractable esophageal tumors.
- Further research is warranted to validate γ-H2AX as a biomarker for guiding PARP inhibitor therapy in ESCC.
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