Epigenetic Profile in Chronic Lymphocytic Leukemia using Methylation-Specific Multiplex Ligation-Dependent Probe Amplification

Abstract

Aim: To analyze the methylation status of 35 tumor suppressor genes using methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) in chronic lymphocytic leukemia (CLL). Materials & methods: The DNA of 37 samples from patients with CLL, six healthy donors, and Jurkat and Ramos cell lines was analyzed by MS-MLPA. Results: Our results confirm that hypermethylation is a common and not randomly distributed event in CLL, and some genes, such as WT1, CDH13, IGSF4/TSLC1, GATA5, DAPK1 and RARB, are hypermethylated in more than 25% of the analyzed samples. Importantly, MS-MLPA also detected hypermethylation of some genes not reported previously in CLL, and their methylation status was confirmed by bisulfite sequencing. Conclusion: These results indicate that MS-MLPA is a useful technique for the detection of methylation in CLL samples. Selecting CLL-specific methylation targets in order to generate a CLL-specific MS-MLPA probe set could enhance its usefulness as a tool in studies of risk stratification and guiding the best therapeutic decision.

KEYWORDS::

• CLL
• DNA methylation
• epigenetic profiling
• MS-MLPA

Acknowledgements

The authors would like to thank the Scientific-Technical Services (CCiTUB) of the Unitat de Bellvitge at the Universitat de Barcelona (Barcelona, Spain) for helpful discussions and suggestions. Furthermore, the authors thank the Unitat de Genòmica from CCiTUB at the Universitat de Barcelona for their technical support. Finally, the authors thank MA Peinado‘s research group at the Institut de Medicina Predictiva i Personalitzada del Càncer (Barcelona, Spain) for technical assistance in bisulfite sequencing and reagents.

Financial & competing interests disclosure

This study was supported by grants from the Ministerio de Ciencia e Innovación and FEDER (SAF2010-20519), the Instituto de Salud Carlos III (RTICC RD06/0020/0097 and RD06/0020/1040) and the AGAUR-Generalitat de Catalunya (AGAUR-2009SGR395). D Iglesias-Serret, DM González-Gironès, A Pérez-Perarnau and C Rubio-Patiño are recipients of research fellowships from the Ministerio de Economía y Competitividad. 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 study was supported by grants from the Ministerio de Ciencia e Innovación and FEDER (SAF2010-20519), the Instituto de Salud Carlos III (RTICC RD06/0020/0097 and RD06/0020/1040) and the AGAUR-Generalitat de Catalunya (AGAUR-2009SGR395). D Iglesias-Serret, DM González-Gironès, A Pérez-Perarnau and C Rubio-Patiño are recipients of research fellowships from the Ministerio de Economía y Competitividad. 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.