Detection of specific chromosome translocation by interphase FISH method and its clinical appication

人白血病细胞具有疾病特异性染色体易位，慢性髓细胞白血病为t(9; 22)，急性髓细胞白血病为t（8; 21）、t（15; 17）、inv（16）、t（3; 21）和t（12; 21）。我们利用荧光原位杂交（FISH）方法建立了鉴定间期核易位的检测方法。在每个染色体易位的断点区域上使用多个cosmid和YAC探针来检测每个易位。我们将FISH方法应用于临床诊断，以及化疗和骨髓移植（BMT）后残留白血病细胞的检测。检测水平约为3%至5%，但在性别不匹配的BMT病例中，使用易位和Y染色体混合探针的FISH检测水平提高到10^<-3>水平。通过对FISH和常规g带分析检出率的比较研究发现，g带法不能识别出占细胞总数不到20%的异常细胞。t（15; 17）和单体7异常尤其难以检测。间期FISH方法有助于检测这些被掩盖的染色体畸变。此外，我们开发了同步FISH分析来检测基因改变和蛋白质表达。利用该方法揭示BCR-ABL、p53、IRF-1和t（9; 22）蛋白表达与5号和17号染色体缺失的关系。这也有助于了解TRF-1端粒蛋白与端粒酶活性与染色体改变、细胞凋亡和基因改变的关系。定性FISH分析将更适用于白血病的病理研究和临床应用。

Human leukemic cells had disease-specific chromosome translocations which are t (9 ; 22) in chronic myelocytic leukemia, t (8 ; 21), t (15 ; 17), inv (16), t (3 ; 21) and t (12 ; 21) in acute myelocytic leukemia. We established detection method to identify these translocations on interphase nuclei using fluorescence in situ hybridization (FISH) method. Several cosmid and YAC probes spanning on breakpoint region of each chromosome translocation were used for detecting each translocation. We applied this FISH method for clinical use to make diagnosis and to detect remaining residual leukemic cell after chemotherapy and bone marrow transplantation (BMT). The detection levels were about three to five percent, but in sex mismatched BMT cases the levels were increased to 10^<-3> levels by FISH using mixture probes of translocation and Y chromosome. Comparative study on the detection rates between FISH and conventional G-banding analyzes were revealed that G-banding method could not identify the aberrant cells containing less than 20 % of the cell population. The t (15 ; 17) and monosomy 7 abnormalities were specially difficult to detect. Interphase FISH method was helpful detect these masked chromosome aberrations. Futhermore we developed simultaneous FISH analysis for detecting gene alteration and protein expression. The method was applied to reveal the relationship between protein expressions of BCR-ABL,p53, IRF-1 and t (9 ; 22) and deletions of chromosomes 5 and 17. It was also helpful to know the relationship between TRF-1 telomere protein and telomerase activities and chromosome alterations, apoptosis and gene alteration. The qualitative FISH analysis will be more applicable for pathological study and clinical use in leukemia.