Tumor Long Interspersed Nucleotide Element-1 (LINE-1) Hypomethylation in Relation to Age of Colorectal Cancer Diagnosis and Prognosis

Naohiko Akimoto^{1

2}, Melissa Zhao¹, Tomotaka Ugai^{1

3}, Rong Zhong^{1

3

4}, Mai Chan Lau¹, Kenji Fujiyoshi¹, Junko Kishikawa¹, Koichiro Haruki¹, Kota Arima¹, Tyler S Twombly¹, Xuehong Zhang^{5

6}, Edward L Giovannucci^{3

5

6}, Kana Wu^{3

5

6}, Mingyang Song^{6

7

8}, Andrew T Chan^{5

7

8

9}, Yin Cao^{10

11

12}, Jeffrey A Meyerhardt¹³, Kimmie Ng¹³, Marios Giannakis^{13

14

15}, Juha P Väyrynen^{1

13

16}, Jonathan A Nowak¹, Shuji Ogino^{1

3

14

17}

Affiliations

¹ Program in MPE Molecular Pathological Epidemiology, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
² Department of Gastroenterology, Nippon Medical School, Graduate School of Medicine, Tokyo 1138602, Japan.
³ Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston 02115, MA, USA.
⁴ Department of Epidemiology and Biostatistics, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
⁵ Channing Division of Network Medicine, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA.
⁶ Department of Nutrition, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
⁷ Clinical and Translational Epidemiology Unit, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.
⁸ Division of Gastroenterology, Massachusetts General Hospital, Boston, MA 02114, USA.
⁹ Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
¹⁰ Division of Public Health Sciences, Department of Surgery, Washington University in St. Louis, St. Louis, MO 63110, USA.
¹¹ Alvin J. Siteman Cancer Center, Washington University School of Medicine, St. Louis, MO 63110, USA.
¹² Division of Gastroenterology, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
¹³ Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, MA 02215, USA.
¹⁴ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
¹⁵ Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
¹⁶ Cancer and Translational Medicine Research Unit, Medical Research Center Oulu, Oulu University Hospital, and University of Oulu, 90220 Oulu, Finland.
¹⁷ Cancer Immunology and Cancer Epidemiology Programs, Dana-Farber Harvard Cancer Center, Boston, MA 02215, USA.

PMID: 33922024
PMCID: PMC8122644
DOI: 10.3390/cancers13092016

Tumor Long Interspersed Nucleotide Element-1 (LINE-1) Hypomethylation in Relation to Age of Colorectal Cancer Diagnosis and Prognosis

Naohiko Akimoto et al. Cancers (Basel). 2021.

. 2021 Apr 22;13(9):2016.

doi: 10.3390/cancers13092016.

Authors

Affiliations

¹ Program in MPE Molecular Pathological Epidemiology, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
² Department of Gastroenterology, Nippon Medical School, Graduate School of Medicine, Tokyo 1138602, Japan.
³ Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston 02115, MA, USA.
⁴ Department of Epidemiology and Biostatistics, School of Public Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
⁵ Channing Division of Network Medicine, Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA.
⁶ Department of Nutrition, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
⁷ Clinical and Translational Epidemiology Unit, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.
⁸ Division of Gastroenterology, Massachusetts General Hospital, Boston, MA 02114, USA.
⁹ Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, MA 02115, USA.
¹⁰ Division of Public Health Sciences, Department of Surgery, Washington University in St. Louis, St. Louis, MO 63110, USA.
¹¹ Alvin J. Siteman Cancer Center, Washington University School of Medicine, St. Louis, MO 63110, USA.
¹² Division of Gastroenterology, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
¹³ Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, MA 02215, USA.
¹⁴ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
¹⁵ Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
¹⁶ Cancer and Translational Medicine Research Unit, Medical Research Center Oulu, Oulu University Hospital, and University of Oulu, 90220 Oulu, Finland.
¹⁷ Cancer Immunology and Cancer Epidemiology Programs, Dana-Farber Harvard Cancer Center, Boston, MA 02215, USA.

PMID: 33922024
PMCID: PMC8122644
DOI: 10.3390/cancers13092016

Abstract

Evidence indicates the pathogenic role of epigenetic alterations in early-onset colorectal cancers diagnosed before age 50. However, features of colorectal cancers diagnosed at age 50-54 (hereafter referred to as "intermediate-onset") remain less known. We hypothesized that tumor long interspersed nucleotide element-1 (LINE-1) hypomethylation might be increasingly more common with decreasing age of colorectal cancer diagnosis. In 1356 colorectal cancers, including 28 early-onset and 66 intermediate-onset cases, the tumor LINE-1 methylation level measured by bisulfite-PCR-pyrosequencing (scaled 0 to 100) showed a mean of 63.6 (standard deviation (SD) 10.1). The mean tumor LINE-1 methylation level decreased with decreasing age (mean 64.7 (SD 10.4) in age ≥70, 62.8 (SD 9.4) in age 55-69, 61.0 (SD 10.2) in age 50-54, and 58.9 (SD 12.0) in age <50; p < 0.0001). In linear regression analysis, the multivariable-adjusted β coefficient (95% confidence interval (CI)) (vs. age ≥70) was -1.38 (-2.47 to -0.30) for age 55-69, -2.82 (-5.29 to -0.34) for age 50-54, and -4.54 (-8.24 to -0.85) for age <50 (P_trend = 0.0003). Multivariable-adjusted hazard ratios (95% CI) for LINE-1 methylation levels of ≤45, 45-55, and 55-65 (vs. >65) were 2.33 (1.49-3.64), 1.39 (1.05-1.85), and 1.29 (1.02-1.63), respectively (P_trend = 0.0005). In conclusion, tumor LINE-1 hypomethylation is increasingly more common with decreasing age of colorectal cancer diagnosis, suggesting a role of global DNA hypomethylation in colorectal cancer arising in younger adults.

Keywords: carcinogenesis; colorectal neoplasms; epigenomics; genomic instability; long interspersed nuclear element; molecular pathology; retrotransposon; screening; transposable element; young-onset cancer.

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Conflict of interest statement

J.A.M. has received institutional research funding from Boston Biomedical, has served as an advisor/consultant to COTA Healthcare, and has served on a grant review panel for the National Comprehensive Cancer Network funded by Taiho Pharmaceutical. M.G. received research funding from Bristol-Myers Squibb, Merck, and Servier. This study was not funded by any of these commercial entities. A.T.C. previously served as a consultant for Bayer Healthcare and Pfizer Inc. This study was not funded by Bayer Healthcare or Pfizer Inc. K.N. has received institutional research funding from Evergrande Group, Genentech, Gilead Sciences, Pharmavite, Revolution Medicines, Tarrex Biopharma, and Trovagene and has served on advisory boards for Array Biopharma, Bayer, and Seattle Genetics. The other authors declare that they have no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Figures

**Figure 1**
Flow chart of case selection in the Nurses’ Health Study and the Health Professionals Follow-up Study. Abbreviations: HPFS, Health Professionals Follow-up Study; LINE-1, long interspersed nucleotide element-1; NHS, Nurses’ Health Study.

**Figure 2**
Pyrosequencing assay to measure LINE-1 methylation. (A) Tumor with LINE-1 hypomethylation (methylation level of 46.7). (B) Tumor with LINE-1 methylation level of 58.9. (C) Tumor with LINE-1 methylation level of 76.1. In panels A–C, the % numbers (in blue shade) are proportions of T and C nucleotides at each CpG site after bisulfite conversion. The proportion of C nucleotides (%) can be interpreted as the methylation level of each CpG site. The first, third, and fourth CpG sites follow mononucleotide T repeats, resulting in higher T peaks (in yellow shade) than the second CpG site. Accordingly, the proportion of C nucleotides (%) has been adjusted. No residual C nucleotides at the non-CpG site are indicated by the arrows, providing evidence for a complete bisulfite conversion reaction. We used the average of the proportions of C nucleotides at the 4 CpG sites as the LINE-1 methylation level (a scale of 0 to 100) of each tumor. To avoid confusion with other % numbers, “%” is not used for this LINE-1 methylation level to keep consistency between the text, tables, and figures. Abbreviation: LINE-1, long interspersed nucleotide element-1.

**Figure 3**
Validation procedure to assess precision of bisulfite conversion and PCR-pyrosequencing. Bisulfite conversion was performed on seven aliquots (B1 to B7) from each specimen. PCR-pyrosequencing was performed for seven bisulfite-treated specimens (B1 to B7) and was repeated seven times on two specimens (B1 and B2) on seven different days (P1 to P7). Abbreviation: PCR, polymerase chain reaction.

**Figure 4**
Distribution of tumor LINE-1 methylation levels in different age groups. In the scatter dot plot, the blue horizontal bar marks the mean and the red horizontal bar indicates the standard deviation of tumor LINE-1 methylation levels in each age group. A significant difference was observed between four age groups (≥70, 55–69, 50–54, and <50) (P_trend <0.0001). Abbreviation: LINE-1, long interspersed nucleotide element-1.

**Figure 5**
Kaplan–Meier survival curves of colorectal cancer-specific survival (A) and overall survival (B) according to the LINE-1 methylation level. The p values were calculated using the log-rank test for trend (two-sided).

See this image and copyright information in PMC

References

1. Siegel R.L., Torre L.A., Soerjomataram I., Hayes R.B., Bray F., Weber T.K., Jemal A. Global patterns and trends in colorectal cancer incidence in young adults. Gut. 2019;68:2179–2185. doi: 10.1136/gutjnl-2019-319511. - DOI - PubMed
1. Saad El Din K., Loree J.M., Sayre E.C., Gill S., Brown C.J., Dau H., De Vera M.A. Trends in the epidemiology of young-onset colorectal cancer: A worldwide systematic review. BMC Cancer. 2020;20:288. doi: 10.1186/s12885-020-06766-9. - DOI - PMC - PubMed
1. Chung R.Y., Tsoi K.K.F., Kyaw M.H., Lui A.R., Lai F.T.T., Sung J.J. A population-based age-period-cohort study of colorectal cancer incidence comparing Asia against the West. Cancer Epidemiol. 2019;59:29–36. doi: 10.1016/j.canep.2019.01.007. - DOI - PubMed
1. Sung H., Siegel R.L., Rosenberg P.S., Jemal A. Emerging cancer trends among young adults in the USA: Analysis of a population-based cancer registry. Lancet Public Health. 2019;4:e137–e147. doi: 10.1016/S2468-2667(18)30267-6. - DOI - PubMed
1. Hofseth L.J., Hebert J.R., Chanda A., Chen H., Love B.L., Pena M.M., Murphy E.A., Sajish M., Sheth A., Buckhaults P.J., et al. Early-onset colorectal cancer: Initial clues and current views. Nat. Rev. Gastroenterol. Hepatol. 2020;17:352–364. doi: 10.1038/s41575-019-0253-4. - DOI - PMC - PubMed

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Tumor Long Interspersed Nucleotide Element-1 (LINE-1) Hypomethylation in Relation to Age of Colorectal Cancer Diagnosis and Prognosis

Affiliations

Tumor Long Interspersed Nucleotide Element-1 (LINE-1) Hypomethylation in Relation to Age of Colorectal Cancer Diagnosis and Prognosis

Authors

Affiliations

Abstract

Conflict of interest statement

Figures

References

Grants and funding

LinkOut - more resources

Full Text Sources