Loss of imprinting of the insulin-like growth factor 2 and the H19 gene in testicular seminomas detected by real-time PCR approach.
IGF2 and H19 are imprinted genes in normal human tissue, but many studies have observed a loss of imprinting (LOI) of these genes in tumors as an epigenetic alteration of the DNA, that leads to a biallelic expression predisposing cells to carcinogenesis and tumor growth. The aim of this study was to test the reliability of LightCycler-assisted Real-time PCR in detecting LOI of IGF2 and H19 in 39 patients with testicular germ cell tumors by comparing these results with the analysis generated by the golden standard restriction fragment length polymorphism (RFLP). With LightCycler-assisted Real-time PCR for IGF2 44% and for H19 49% of the patients were found to be heterozygous. This was consistent with the results obtained by RFLP, but surprisingly RFLP failed in more than 7% of the patients. In detecting LOI (for IGF2 in 41% and for H19 in 68% of the informative patients) the approach by RFLP was superior, since the results derived from LightCycler-assisted Real-time PCR showed reliable results in 76 and 10% of the samples concerning IGF2 and H19, respectively. Again, no discrepancy between the results obtained by the two methods occurred. In sum, LightCycler-assisted Real-time PCR is a sufficiently working approach for the rapid and reliable detection of heterozygosity of IGF2 or H19 gene and identification of LOI of IGF2 and thus may be helpful in conducting large epidemiological studies. However, for the identification of LOI of the H19 gene in this cohort it possesses only restrictive use.
Stier
Sebastian
S
Medizinische Universitäts-Poliklinik Bonn, Wilhelmstrasse 35-37, 53111 Bonn, Germany.
Neuhaus
Thomas
T
Albers
Peter
P
Wernert
Nicolas
N
Grünewald
Elisabeth
E
Forkert
Randolf
R
Vetter
Hans
H
Ko
Yon
Y
D015331:Cohort Studies; D015972:Gene Expression Regulation, Neoplastic; D018392:Genomic Imprinting; D006579:Heterozygote; D006801:Humans; D007335:Insulin-Like Growth Factor II; D008297:Male; D012150:Polymorphism, Restriction Fragment Length; D011237:Predictive Value of Tests; D011506:Proteins / Q000032:analysis / Q000235:genetics*; D062085:RNA, Long Noncoding; D012333:RNA, Messenger / Q000032:analysis; D022661:RNA, Untranslated / Q000032:analysis / Q000235:genetics*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D018239:Seminoma / Q000737:chemistry / Q000235:genetics*; D013736:Testicular Neoplasms / Q000737:chemistry / Q000235:genetics*
C094335:H19 long non-coding RNA; C035170:IGF2 protein, human; D011506:Proteins; D062085:RNA, Long Noncoding; D012333:RNA, Messenger; D022661:RNA, Untranslated; D007335:Insulin-Like Growth Factor II