Monitoring of methylation changes in 9p21 region in patients with myelodysplastic syndromes and acute myeloid leukemia.

Epigenetic de novo methylation of CpG islands is an important event in malignant transformation. Two genes are frequently methylated: cyclin-dependent kinase inhibitor 2B (CDKN2B) and cyclin-dependent kinase inhibitor 2A (CDKN2A). In our study methylation of these genes was studied in 63 patients with myelodysplastic syndromes (MDS), 2 with myelodysplastic/myeloproliferative neoplasms (MDS/MPN) and 13 with acute myeloid leukemia (AML). Five patients were monitored during 5-azacytidine treatment. Twenty-six healthy donors were tested in a control group. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) method with all associated techniques was used for detection. Aberrant methylation was present in the CDKN2A gene in 38% and in the CDKN2B gene in 77% of the patients in MDS group. The level of methylation was higher in the group of AML patients - 77% in CDKN2A gene and 100% in CDKN2B gene. In MDS patients, an aberrant methylation was associated with a tendency to disease progression towards more advanced forms according to the World Health Organization (WHO) classification and the International Prognostic Scoring System (IPSS). Significant differences in methylation level were observed between early and advanced forms of MDS in CDKN2B gene (P value < 0.05) but not for CDKN2A gene. The trend of methylation in patients treated with azacitidine was analyzed in CDKN2B gene and correlated with the course of the disease. Increased methylation was connected with disease progression. We concluded that the methylation level of CDKN2B gene might be used as a marker of leukemic transformation in MDS. Our study indicates the role of hypermethylation as an important event in the progression of MDS to AML.

[Familial haemophagocytic lymphohistiocytosis caused by perforin deficit can be successfully treated by haematopoietic stem cell transplantation--the first diagnosed case in the Czech Republic]. / Familiární hemofagocytující lymfohistiocytóza na podklade deficitu perforinu uspesne lécená transplantací hematopoetických kmenových bunek--první diagnostikovaný prípad v Ceské republice.

Familial haemophagocytic lymphohistiocytosis (FHL) is an inherited disorder characterized by an impaired cytotoxicity of T lymphocytes and NK cells typically manifesting within first few months after birth. If not treated adequately, it is inevitably fatal within several months. The incidence in Caucasians has been estimated to 1: 50 000 births. Haematopoietic stem cell transplantation represents the only curative treatment for FHL. Recently, several genetic defects underlying molecular defects in FHL have been identified. In approximately 30% of patients FHL is caused by mutations in PRF1 gene coding for perforin. Further 30% of patients were found to have mutations in UNC13D coding for hMunc13-4 protein. Very recent report has identified another cause of FHL, mutations in STX11 gene on chromosome 6, coding for syntaxin 11. Absence of any of those proteins severely impairs the process of exocytosis of cytotoxic granules. We describe patient with clinical symptoms of FHL. Immunological and molecular biology methods led to the identification of perforin mutation as a cause of the disease. Patient received an allogeneic SCT from HLA-matched unrelated donor. SCT was followed by rapid normalization of clinical symptoms and laboratory findings. In patient described in this study, FHL manifested with typical clinical and laboratory symptoms. Adequate immunosuppressive treatment and subsequent SCT led to the sustained remission of FHL and correction of molecular defect. This is the first case of FHL in Czech Republic where perforin mutation was identified as a molecular cause both at cellular and molecular level.