We treated HL-60 and ATRA-resistant HL-60 (HL-60R) cells that express mutated RARalpha and very low levels of RARbeta, RARgamma and RXRalpha with 4-HPR (2 microM) for 3 days.
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.
A monoclonal or oligoclonal pattern could be obtained by digestion with Hha I in 18/22 (82%) patients with acute myeloid leukaemia who had previously shown hypermethylation of both alleles using Hpa II, although in six of these patients differences in methylation could still be demonstrated between leukaemic and remission samples.