Masato Maekawa

Extensive but hemiallelic methylation of the hMLH1 promoter region in early-onset sporadic colon cancers with microsatellite instability

BACKGROUND AND AIMSnMethylation of the hMLH1 promoter region is frequently observed in microsatellite instability (MSI)-positive sporadic colorectal carcinomas. We studied hMLH1 promoter methylation in peripheral blood lymphocytes of 87 index patients representing 29 cases of hereditary nonpolyposis colorectal cancers (HNPCCs), 28 cases of atypical HNPCCs, and 30 sporadic cases of the development of early-onset colorectal carcinomas or multiple primary cancers.nnnMETHODSnMethylation of the hMLH1 promoter region was analyzed by Na-bisulfite polymerase chain reaction/single-strand conformation polymorphism analysis or methylation-specific polymerase chain reaction. MSI, allelic status of the hMLH1 locus, and loss of hMLH1 protein expression were examined in cases for which tumor tissues were available.nnnRESULTSnExtensive methylation of the hMLH1 promoter was detected in peripheral blood lymphocytes of 4 of 30 patients with sporadic early-onset colon cancer, among whom multiple primary cancers (1 colon and 1 endometrial cancer) developed in 2 cases. This methylation was not detected in analyses of HNPCC or atypical HNPCC groups or healthy control subjects. MSI was positive, and extensive methylation was detected in both cancers (colon and endometrial cancer) and normal tissues (colon, gastric mucosa, endometrium, and bone marrow) in all of the examined cases (3 of 3). Analysis of a polymorphic site in the hMLH1 promoter in 2 informative cases showed that methylation was hemiallelic. In 1 case, the unmethylated allele was lost in the colon cancer but not in the metachronous endometrial cancer.nnnCONCLUSIONSnConstitutive, hemiallelic methylation of the hMLH1 promoter region was shown to be associated with carcinogenesis in sporadic, early-onset MSI-positive colon cancers.

Efficacy of gemtuzumab ozogamicin on ATRA- and arsenic-resistant acute promyelocytic leukemia (APL) cells

Acute promyelocytic leukemia (APL) cells express a considerable level of CD33, which is a target of gemtuzumab ozogamicin (GO), and a significantly lower level of P-glycoprotein (P-gp). In this study, we examined whether GO was effective on all-trans retinoic acid (ATRA)- or arsenic trioxide (ATO)-resistant APL cells. Cells used were an APL cell line in which P-gp was undetectable (NB4), ATRA-resistant NB4 (NB4/RA), NB4 and NB4/RA that had been transfected with MDR-1 cDNA (NB4/MDR and NB4/RA/MDR, respectively), ATO-resistant NB4 (NB4/As) and blast cells from eight patients with clinically ATRA-resistant APL including two patients with ATRA- and ATO-resistant APL. The efficacy of GO was analyzed by 3H-thymidine incorporation, the dye exclusion test and cell cycle distribution. GO suppressed the growth of NB4, NB4/RA and NB4/As cells in a dose-dependent manner. GO increased the percentage of hypodiploid cells significantly in NB4, NB4/RA and NB4/As cells, and by a limited degree in NB4/MDR and NB4/RA/MDR cells. Similar results were obtained using blast cells from the patients with APL. GO is effective against ATRA- or ATO-resistant APL cells that do not express P-gp, and the mechanism of resistance to GO is not related to the mechanism of resistance to ATRA or ATO in APL cells.

Epigenetics: relations to disease and laboratory findings.

Epigenetics is a postmeiotic modification of gene expression that is independent of the primary DNA sequence. DNA methylation, methylated DNA binding proteins, and histone modification-related enzymes are associated with epigenetics. Abnormalities in DNA methylation of CpG islands which are important for gene expression control, affect gene expression, which may influence carcinogenesis, aging, and other diseases. Aberrant DNA methylation occurs with aging, inflammation, viral infection, and carcinogenesis. DNA methylation can be evaluated for molecular analysis for diagnosis of early cancer. It is also important for laboratory diagnosis by using classic and authentic laboratory tests because the tests can be affected by epigenetics-controlled gene expression. It is also related to the effectiveness of therapeutic agents affecting DNA methylation and histone deacetylation, and the strategy in search of genetic abnormality for epigenetic as well as genetic error.