A high-throughput and sensitive method to measure global DNA methylation: application in lung cancer.
BACKGROUND
Genome-wide changes in DNA methylation are an epigenetic phenomenon that can lead to the development of disease. The study of global DNA methylation utilizes technology that requires both expensive equipment and highly specialized skill sets.


METHODS
We have designed and developed an assay, CpGlobal, which is easy-to-use, does not utilize PCR, radioactivity and expensive equipment. CpGlobal utilizes methyl-sensitive restriction enzymes, HRP Neutravidin to detect the biotinylated nucleotides incorporated in an end-fill reaction and a luminometer to measure the chemiluminescence. The assay shows high accuracy and reproducibility in measuring global DNA methylation. Furthermore, CpGlobal correlates significantly with High Performance Capillary Electrophoresis (HPCE), a gold standard technology. We have applied the technology to understand the role of global DNA methylation in the natural history of lung cancer. World-wide, it is the leading cause of death attributed to any cancer. The survival rate is 15% over 5 years due to the lack of any clinical symptoms until the disease has progressed to a stage where cure is limited.


RESULTS
Through the use of cell lines and paired normal/tumor samples from patients with non-small cell lung cancer (NSCLC) we show that global DNA hypomethylation is highly associated with the progression of the tumor. In addition, the results provide the first indication that the normal part of the lung from a cancer patient has already experienced a loss of methylation compared to a normal individual.


CONCLUSIONS
By detecting these changes in global DNA methylation, CpGlobal may have a role as a barometer for the onset and development of lung cancer.
Anisowicz
Anthony
A
Molecular Therapeutics Division, AmberGen Incorporated, Waltham, Massachusetts, USA.
Huang
Hui
H
Braunschweiger
Karen I
KI
Liu
Ziying
Z
Giese
Heidi
H
Wang
Huajun
H
Mamaev
Sergey
S
Olejnik
Jerzy
J
Massion
Pierre P
PP
Del Mastro
Richard G
RG
D000368:Aged; D000369:Aged, 80 and over; D001360:Avidin / Q000378:metabolism; D019921:Biotinylation; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis* / Q000235:genetics*; D045744:Cell Line, Tumor; D018899:CpG Islands; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D008297:Male; D008875:Middle Aged; D012044:Regression Analysis
C093483:neutravidin; D001360:Avidin