Semi-quantitative study of calcitonin gene methylation in myelodysplastic syndrome

OBJECTIVE: To evaluate whether hypermethylation of calcitonin (CT) gene could serve as a transforming signal of myelodysplastic syndrome (MDS) to leukemia. METHODS: Bone marrow aspirates from 35 MDS patients, including 25 refractory anemia (RA), 10 refractory anemia with excess of blasts (RAEB) or refractory anemia with excess of blasts in transformation (RAEBt) and 7 cases of acute myeloid leukemia (AML) transformed from MDS, were studied on methylation rate in 5' end of CT gene by polymerase chain reaction (PCR) technique using methylation-sensitive endonuclease Hpa II with external references of undigested DNA and Msp I digested DNA and internal reference of 112 bp fragment containing codon 61 of N-ras oncogene. The results were expressed as calcitonin gene methylation rate (CTMR) calculated from the densitometer-analyzed integral calculus of PCR products of 566 bp CT(a1), 112 bp N-ras(b1) by using Hpa II-digested DNA and PCR products of 566 bp CT (a0), 112 bp N-ras(b0) by using undigested DNA according to the formula, CTMR = (a1/b1)/(a0/b0) x 100%. RESULTS: The CTMRs in total 35 MDS, 25 RA, 10 RAEBt and 7 cases of AML transformed from MDS were 36.87% +/- 25.10%, 28.12% +/- 24.01%, 58.74% +/- 16.49%, and 54.03% +/- 7.06% respectively, significantly higher than that in control group (P < 0.001, P < 0.05, P < 0.001 and P < 0.001, respectively). CONCLUSION: The results suggest that hypermethylation of CT gene occurs in early stage of leukemic transformation and CTMR might be a useful marker in predicting the transformation of MDS to AML.