慢性骨髓性白血病病人以Imatinib治療其JunB與BCR-ABL之基因表現及臨床反應

Evaluating the effect of imatinib mesylate in chronic myeloid leukemia patients in the expression of JunB and BCR-ABL gene

劉益昌(Yi-Chang Liu)

指導教授 ： 林勝豐教授

高雄醫學大學/醫學院/醫學研究所/碩士(2003年)

摘要

Imatinib mesylate是一種有效的Bcr-Abl酪胺酸激酶〔tyrosine kinase〕抑制劑，這個激酶是慢性骨髓性白血病的病理發生學上一相當關鍵的蛋白質，此藥物是最近對癌症治療研究最熱門的所謂標靶治療〔target therapy〕的藥物之一，它已顯示出相當有效的臨床治療反應，包括血液學上的緩解及細胞遺傳學上的緩解，特別是對慢性期的病人，其療效也已得到包括phase III在內的各種臨床試驗證實。JunB是activating protein-1 轉錄因子的Jun 家族之一員，調節細胞的成長分化及增生轉變。最近的研究發現缺少JunB表現的基因轉殖的老鼠，會表現出骨髓增殖性疾病，類似於人類慢性骨髓性白血病的變化，其在骨髓性細胞生成上，扮演類似腫瘤抑制基因的角色。另外的研究也發現在慢性骨髓性白血病的病人其JunB的表現缺乏，是由於JunB基因的promoter區域被甲基化而導致其不活化，而其疾病不同的表現和被甲基化的比例有關。在此我們利用即時定量RT-PCR的方法，定期追蹤慢性骨髓性白血病病人在接受imatinib治療後，其JunB和Bcr-Abl在週邊血液的表現。總共有22位病人接受imatinib的治療，19位接受規則追蹤的病人中，9位男性，10位女性，平均年齡37歲，平均追蹤期為6個月，14位病人為慢性期患者，5位為加速期患者。14位慢性期的病人〔100%〕均達到完全血液學緩解，達到主要細胞遺傳學反應有10位〔71.4%〕，完全細胞遺傳學反應有9位〔64.3%〕，平均時間為16及18星期，而分子學上反應則有8位〔57.1%〕，平均時間為21星期。在其餘5位加速期的病人，有1位〔20%〕達完全細胞遺傳學及分子學上反應，有2位〔40%〕仍維持在加速期，有2位〔40%〕疾病進展至芽細胞危象期。副作用方面，全部病人因治療導致的grade 3和grade 4的白血球低下和血小板低下分別有8位〔36.4%〕及6位〔27.3%〕，其餘非血液學上副作用較少發生。在JunB和Bcr-Abl的表現上，隨治療而JunB表現增加而Bcr-Abl下降或偵測不出的病人有5位，臨床上都能達完全細胞遺傳學反應〔100%〕及分子學上反應〔100%〕。而JunB表現越來越少甚至很低且Bcr-Abl一直存在的病人有2位，均為加速期進展至blast crisis而導致mortality，且合併clonal evolution的病人。無規則的JunB表現且Bcr-Abl沒偵測到有3位，臨床上均達完全細胞遺傳學反應，而Bcr-Abl逐漸減少甚至消失有3位，其中2位達完全細胞遺傳學反應，1位達主要細胞遺傳學反應，其平均診斷至開始治療期間及平均達主要細胞遺傳學反應之時間和JunB增加組無明顯之差異。另無規則的JunB和Bcr-Abl表現共6位，4位慢性期而2位加速期病人，臨床上均維持於原疾病分期，並無細胞遺傳學反應，治療反應未如預期。而JunB表現增加與否，和主要細胞遺傳學反應，完全細胞遺傳學反應及分子學上反應之有無，有顯著性差異存在。因此我們認為，雖然所收集的病人及檢體數不夠多，但臨床上對於治療反應的評估，包括主要細胞遺傳學反應和完全細胞遺傳學反應的比例，和其他研究結果相類似，也證實了imatinib的臨床療效，惟可能有較多grade 3-4的白血球低下和血小板低下的情況。而JunB表現的變化，特別是那些隨著治療而增加表現的族群和良好臨床反應的關係，暗示著JunB的表現增加，可能可以當作一個預測良好治療效果的指標，及發展新治療方法的依據。而由於其他能達到完全細胞遺傳學反應的病人也有部分呈現不一致的JunB表現，也暗示著可能另有其他的機制，能夠共同調節JunB的表現。而是否imatinib的治療，能夠導致更多的JunB基因的promoter area的demethylation或經由其他機制使JunB的表現增加，是需再探討之處。而另外衍生出的問題，包括對於Bcr-Abl的其他下游路徑及不同的受質，是否會因imatinib的治療同時改變其表現，還有對於那些JunB和Bcr-Abl表現均起伏不定的病患，臨床上可能產生抗藥性的原因，以及對於這些病人更長期的追蹤變化，包括能否持續維持良好反應，也需更長時間及更進一步的研究來證實。

關鍵字

imatinib ； JunB ； Bcr-Abl ； 即時定量RT-PCR ； 慢性骨髓性白血病

並列摘要

Imatinib mesylate, a potent inhibitor of the Bcr-Abl tyrosine kinase that is central to the pathogenesis of CML, has demonstrated remarkable hematologic and cytogenetic response especially in chronic phase. This was well documented in various studies including phase III study. JunB is a component of the Jun family genes of the activating protein-1 transcription factors that are important in the control of cell growth, differentiation and neoplastic transformation. It was demonstrated that transgenic mice lacking JunB expression developed myeloproliferative disease and eventually progressing to blast crisis that resembles human CML. Recently, it was demonstrated that down-regulated JunB expression in CML was due to the inactivation of JunB gene by methylation in the promoter area, and the differential expression was correlated to the ratio of cells being methylated. Here we used real-time quantitative RT-PCR to monitor the serial changes of both JunB and Bcr-Abl expression in peripheral blood in our patients receiving imatinib. Total 22 patients were treated with imatinib, and in 19 of the 22 patients (9 male and 10 female, median age 37 years-old) who received regular follow-up, 14 patients were in chronic phase and 5 patients were in accelerating phase. All of the 14 chronic phase patients achieved complete hematologic response (100%), and major cytogenetic response was found in 10 of 14 (71.4%) patients and complete cytogenetic response in 9 of 14 (64.3%) patients, with median time 16 and 18 weeks respectively. Molecular response was also noted in 8 of 14 (57.1%) patients with median time 21 weeks. Among the 5 patients in
accelerating phase, one of them (20%) achieved complete cytogenetic response and molecular response, two of them (40%) remained in accelerating phase and two of them (40%) with disease progression to blast crisis with clonal evolution. Grade III to IV leukopenia and thrombocytopenia caused by imatinib was found in 8 of 19 (36.4%) and 6 of 19 (27.3%) patients, and other side effects were not so obvious compared with other studies. Gradual increase in JunB expression with decrease or undetectable Bcr-Abl expression during imatinib treatment was found in 5 patients, all of them achieved complete cytogenetic response (100%) and even molecular response (100%). Gradual decrease to even very low level of JunB expression with variable Bcr-Abl expression was found in two patients who possessed disease progression to blast crisis with clonal evolution and leaded to mortality. Variable JunB expression with undetectable Bcr-Abl expression was found in 3 patients, all of them achieved complete cytogenetic response. Variable JunB expression with gradual decrease Bcr-Abl expression was found in other 3 patients, 2 of them achieved complete cytogenetic response and one of them achieved major cytogenetic response. No significance existed on the median time to complete cytogenetic response and median time to initiation of imatinib between these two groups. The other 6 patients (4 in chronic phase and 2 in accelerating phase) with variable JunB and Bcr-Abl expression revealed no cytogenetic response and remained in stable disease. Significance existed between increased JunB expression and good clinical response such as complete cytogenetic response or molecular response. Conclusively, similar clinical response rate including major and complete cytogenetic response during chronic and accelerating phase was found in our results compared with previous studies, indicated the clinical efficacy of imatinib in the treatment of CML. It seemed that more grades III to IV leukopenia and thrombocytopenia were found during treatment in our study. The correlation between those who possessed gradual increase in JunB expression and good clinical response implied that the increase in JunB expression during imatinib treatment might be used as a predictor for good clinical response and a basis for developing new treatment tragedy. The fact that variable JunB expression could be found in those who possessed complete cytogenetic response indicated that there were still other mechanisms responsible for regulation of JunB expression. The possible mechanism accounting for increasing JunB expression is caused by more demethylation in promoter area of JunB gene or by other mechanism needs to be further investigated. Furthermore, the changes in specific genes expression other than JunB that is down-streamed to Bcr-Abl protein during imatinib treatment, the possible mechanisms for those who revealed drug resistance, and the long-term effects of imatinib, still needed to be further investigated.

並列關鍵字

imatinib ； JunB ； Bcr-Abl ； real-time quantitative RT-PCR ； chronic myeloid leukemia

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