Research output: Contribution to journal › Article › peer-review
Size and Methylation Index of Cell-Free and Cell-Surface-Bound DNA in Blood of Breast Cancer Patients in the Contest of Liquid Biopsy. / Tamkovich, Svetlana; Tupikin, Alexey; Kozyakov, Anton et al.
In: International Journal of Molecular Sciences, Vol. 23, No. 16, 8919, 10.08.2022.Research output: Contribution to journal › Article › peer-review
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TY - JOUR
T1 - Size and Methylation Index of Cell-Free and Cell-Surface-Bound DNA in Blood of Breast Cancer Patients in the Contest of Liquid Biopsy
AU - Tamkovich, Svetlana
AU - Tupikin, Alexey
AU - Kozyakov, Anton
AU - Laktionov, Pavel
N1 - Publisher Copyright: © 2022 by the authors.
PY - 2022/8/10
Y1 - 2022/8/10
N2 - Aberrantly methylated circulating DNA (cirDNA) has proven to be a good cancer marker, but its detection is limited by low concentrations, fragmentation, and insufficiency. Since the methylated cirDNA was shown to be more stable in circulation than the unmethylated one and was shown to bind with the blood cell surface, we studied the concentration, representation, and fragmentation of tumor-derived methylated DNA in cell-free and cell-surface-associated DNA. We found that long DNA fragments (more than 10 kb) are mainly associated with the surface of blood cells. However, in plasma short DNA fragments (100-1000 bp) were also found along with long DNA fragments. Isolation of short fragments after separation of cirDNA in 6% PAGE followed by quantitative PCR (L1 element) has shown that short DNA fragments in healthy females represent 22% versus 0.5-4.4% in breast cancer patients. The methylated form of the RARβ2 gene was detected only in long DNA fragments by Real-time TaqMan PCR of bisulfite-converted DNA. The methylation index of cirDNA from healthy women was estimated at 0%, 9%, and 7% in plasma, PBS-EDTA, and trypsin eluates from the surface of blood cells, respectively. The methylation index of breast cancer patients' DNA was found to be 33%, 15%, and 61% in the same fractions confirming the overrepresentation of methylated DNA in csbDNA.
AB - Aberrantly methylated circulating DNA (cirDNA) has proven to be a good cancer marker, but its detection is limited by low concentrations, fragmentation, and insufficiency. Since the methylated cirDNA was shown to be more stable in circulation than the unmethylated one and was shown to bind with the blood cell surface, we studied the concentration, representation, and fragmentation of tumor-derived methylated DNA in cell-free and cell-surface-associated DNA. We found that long DNA fragments (more than 10 kb) are mainly associated with the surface of blood cells. However, in plasma short DNA fragments (100-1000 bp) were also found along with long DNA fragments. Isolation of short fragments after separation of cirDNA in 6% PAGE followed by quantitative PCR (L1 element) has shown that short DNA fragments in healthy females represent 22% versus 0.5-4.4% in breast cancer patients. The methylated form of the RARβ2 gene was detected only in long DNA fragments by Real-time TaqMan PCR of bisulfite-converted DNA. The methylation index of cirDNA from healthy women was estimated at 0%, 9%, and 7% in plasma, PBS-EDTA, and trypsin eluates from the surface of blood cells, respectively. The methylation index of breast cancer patients' DNA was found to be 33%, 15%, and 61% in the same fractions confirming the overrepresentation of methylated DNA in csbDNA.
KW - breast cancer
KW - cell surface bound DNA
KW - cell-free DNA
KW - circulating DNA
KW - fragment sizes
KW - liquid biopsy
KW - methylation
UR - http://www.scopus.com/inward/record.url?scp=85137127128&partnerID=8YFLogxK
U2 - 10.3390/ijms23168919
DO - 10.3390/ijms23168919
M3 - Article
C2 - 36012175
AN - SCOPUS:85137127128
VL - 23
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
SN - 1661-6596
IS - 16
M1 - 8919
ER -
ID: 37098149