Real-Time Methylomic Aberrations During Initiation and Progression of Induced Human Mammary Epithelial Cell Tumorigenesis

Abstract

Aim: Neoplastic transformation provides one of the few existing opportunities to analyze molecular changes in real time during the initiation and progression of breast cancer. Materials & methods: Human mammary epithelial cells underwent neoplastic reprogramming, generating one line of semitransformed, premalignant cells and two separate, temporal lines of fully transformed human mammary epithelial cells (THMECs). An Illumina Infinium HumanMethylation27 BeadChip was used to analyze DNA methylation alterations in 27,578 CpG loci at three consecutive time points over an 80-day (d) transformation period. Results: The mean β value for semitransformed human mammary epithelial cells CpG loci (0.245) was much greater than for either THMEC-40d (0.055) or THMEC-80d (0.066), indicating a large loss of methylation after neoplastic induction. In addition, 54% of CpG loci were hypermethylated during the THMEC-40d to THMEC-80d transition. We observed that the CpG loci exhibiting DNA methylation changes during early oncogenesis were enriched for biological functions like cellular movement; this was distinctly different than in the later, more progressive stages of the transformation process enriched for processes involving differentiation. Conclusion: The timing of major methylomic changes may be important in directing the cell toward a more cancerous phenotype. In addition, gene-specific hypermethylation appears to silence developmentally related genes, leading to dedifferentiation.

KEYWORDS::

• breast cancer
• DNA methylation
• epigenetics
• Illumina Infinium HumanMethylation27 BeadChip
• neoplastic reprogramming

Acknowledgements

The authors appreciate the help of T Hardy in proofreading the manuscript. The authors thank JT DeAngelis for providing the cell lines.

Financial & competing interests disclosure

This work was funded by grants to TO Tollefsbol from the American Institute for Cancer Research and the National Institutes of Health (CA 129415). The authors acknowledge the support of the UAB Comprehensive Cancer Center (P30-CA013148) Heflin Center for Genomic Sciences Genomics Core and its personnel for assisting with all portions of the Illumina Infinium HumanMethylation27 BeadChip experiments. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Ethical conduct of research

The authors state that they have obtained appropriate institutional review board approval or have followed the principles outlined in the Declaration of Helsinki for all human or animal experimental investigations. In addition, for investigations involving human subjects, informed consent has been obtained from the participants involved.

Additional information

Funding

This work was funded by grants to TO Tollefsbol from the American Institute for Cancer Research and the National Institutes of Health (CA 129415). The authors acknowledge the support of the UAB Comprehensive Cancer Center (P30-CA013148) Heflin Center for Genomic Sciences Genomics Core and its personnel for assisting with all portions of the Illumina Infinium HumanMethylation27 BeadChip experiments. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.