In vitro study of radiosensitivity in colorectal cancer cell lines associated with Lynch syndrome
File(s)
Published version
Author(s)
Type
Journal Article
Abstract
Introduction: Lynch syndrome patients have an inherited predisposition to cancer due to a deficiency in DNA mismatch repair (MMR) genes which could lead to a higher risk of developing cancer if exposed to ionizing radiation. This pilot study aims to reveal the association between MMR deficiency and radiosensitivity at both a CT relevant low dose (20 mGy) and a therapeutic higher dose (2 Gy).
Methods: Human colorectal cancer cell lines with (dMMR) or without MMR deficiency (pMMR) were analyzed before and after exposure to radiation using cellular and cytogenetic analyses i.e., clonogenic assay to determine cell reproductive death; sister chromatid exchange (SCE) assay to detect the exchange of DNA between sister chromatids; γH2AX assay to analyze DNA damage repair; and apoptosis analysis to compare cell death response. The advantages and limitations of these assays were assessed in vitro, and their applicability and feasibility investigated for their potential to be used for further studies using clinical samples.
Results: Results from the clonogenic assay indicated that the pMMR cell line (HT29) was significantly more radio-resistant than the dMMR cell lines (HCT116, SW48, and LoVo) after 2 Gy X-irradiation. Both cell type and radiation dose had a significant effect on the yield of SCEs/chromosome. When the yield of SCEs/chromosome for the irradiated samples (2 Gy) was normalized against the controls, no significant difference was observed between the cell lines. For the γH2AX assay, 0, 20 mGy and 2 Gy were examined at post-exposure time points of 30 min (min), 4 and 24 h (h). Statistical analysis revealed that HT29 was only significantly more radio-resistant than the MLH1-deficient cells lines, but not the MSH2-deficient cell line. Apoptosis analysis (4 Gy) revealed that HT29 was significantly more radio-resistant than HCT116 albeit with very few apoptotic cells observed.
Discussion: Overall, this study showed radio-resistance of the MMR proficient cell line in some assays, but not in the others. All methods used within this study have been validated; however, due to the limitations associated with cancer cell lines, the next step will be to use these assays in clinical samples in an effort to understand the biological and mechanistic effects of radiation in Lynch patients as well as the health implications.
Methods: Human colorectal cancer cell lines with (dMMR) or without MMR deficiency (pMMR) were analyzed before and after exposure to radiation using cellular and cytogenetic analyses i.e., clonogenic assay to determine cell reproductive death; sister chromatid exchange (SCE) assay to detect the exchange of DNA between sister chromatids; γH2AX assay to analyze DNA damage repair; and apoptosis analysis to compare cell death response. The advantages and limitations of these assays were assessed in vitro, and their applicability and feasibility investigated for their potential to be used for further studies using clinical samples.
Results: Results from the clonogenic assay indicated that the pMMR cell line (HT29) was significantly more radio-resistant than the dMMR cell lines (HCT116, SW48, and LoVo) after 2 Gy X-irradiation. Both cell type and radiation dose had a significant effect on the yield of SCEs/chromosome. When the yield of SCEs/chromosome for the irradiated samples (2 Gy) was normalized against the controls, no significant difference was observed between the cell lines. For the γH2AX assay, 0, 20 mGy and 2 Gy were examined at post-exposure time points of 30 min (min), 4 and 24 h (h). Statistical analysis revealed that HT29 was only significantly more radio-resistant than the MLH1-deficient cells lines, but not the MSH2-deficient cell line. Apoptosis analysis (4 Gy) revealed that HT29 was significantly more radio-resistant than HCT116 albeit with very few apoptotic cells observed.
Discussion: Overall, this study showed radio-resistance of the MMR proficient cell line in some assays, but not in the others. All methods used within this study have been validated; however, due to the limitations associated with cancer cell lines, the next step will be to use these assays in clinical samples in an effort to understand the biological and mechanistic effects of radiation in Lynch patients as well as the health implications.
Date Issued
2024-04-04
Date Acceptance
2024-03-18
Citation
Frontiers in Public Health, 2024, 12
ISSN
2296-2565
Publisher
Frontiers Media S.A.
Journal / Book Title
Frontiers in Public Health
Volume
12
Copyright Statement
© 2024 Sun, Moquet, Barnard, Mancey,
Burling, Baldwin-Cleland, Monahan,
Latchford, Lloyd, Bou er, Badie, Anyamene
and Ainsbury. This is an open-access article
distributed under the terms of the Creative
Commons Attribution License (CC BY). The
use, distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication in
this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms.
Burling, Baldwin-Cleland, Monahan,
Latchford, Lloyd, Bou er, Badie, Anyamene
and Ainsbury. This is an open-access article
distributed under the terms of the Creative
Commons Attribution License (CC BY). The
use, distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication in
this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms.
License URL
Identifier
https://www.frontiersin.org/journals/public-health/articles/10.3389/fpubh.2024.1369201/full
Subjects
ASSAY
BIODOSIMETRY
BIOMARKER
colorectal cancer cell lines
CT
DEFECTS
DNA-DAMAGE
DOUBLE-STRAND BREAKS
HOMOLOGOUS RECOMBINATION
Life Sciences & Biomedicine
Lynch syndrome
mismatch repair (MMR) deficiency
MISMATCH-REPAIR
Public, Environmental & Occupational Health
radiation effects
radiosensitivity
RISK
Science & Technology
Publication Status
Published
Article Number
1369201
Date Publish Online
2024-04-04