CORE--MINIMAL DISEASE DETECTION BY PCR

慢性粒细胞白血病(CML)是一种多能干细胞疾病， 它与一种被称为费城的异常染色体有关 染色体(Ph)。Ph融合了部分bcr和abl基因，而 产生的bcr-abl癌蛋白在血液和骨髓细胞中表达。 在慢性粒细胞白血病患者中。有两种治疗模式非常有效， 慢性粒细胞白血病慢性期患者诱导长期缓解：骨髓 来自匹配供者的移植和干扰素-α治疗。 目前有几种检测方法用于监测慢性粒细胞白血病的治疗情况，但只有 逆转录酶(RT)聚合酶链式反应(PCR)方法是 足够灵敏，可以检测到微小的残留病。RT-PCR是已知的 只检测到一个细胞表达bcr-abl 10万个细胞。然而，标准的RT-PCR方法作为NOT被广泛使用 量化的。竞争性定量聚合酶链式反应方法已在 最近几年。我们正在采用克罗斯和高盛开发的方法。 该方法包括添加已知水平的BCR-ABL连接序列， 包含插入以将其与天然连接序列区分开来， 一个未知的样本，并比较放大器的强度。这个 通过对连续病人样本的分析，提高了方法的可靠性 从一段时间来确定bcr-abl-RNA水平是否 递减的或递增的例如，一种递减水平的模式 Bcr-abl核糖核酸可提示病情进一步缓解。这笔赠款的目的是 是采用克罗斯和戈德曼的方法，并使用测试来监测骨骼 骨髓移植治疗慢性粒细胞白血病和慢性粒细胞白血病患者 干扰素。这种方法也将用于Ph+ALL患者。这个 将使用已知的bcr-Abl阳性和阴性细胞建立方法 接下来将对来自血液和骨髓的细胞进行研究 强调最小残留疾病；其效用将通过 对病人样本进行盲法研究。最后，一个核心实验室 将建立用于分析CML患者血液和骨髓样本的方法 通过竞争性聚合酶链式反应在P01资助范围内研究的样本。

Chronic myelogenous leukemia (CML) is a pluripotent stem cell disease, which is associated with an abnormal chromosome, termed the Philadelphia chromosome (Ph). The Ph fuses parts of the BCR and ABL genes, and the resulting Bcr-Abl oncoprotein is expressed in blood and bone marrow cells of CML patients. Two modes of therapy have been unusually effective at inducing long term remission in chronic phase CML patients: bone marrow transplantation from a matched donor and interferon-alpha treatment. Several tests are currently used to monitor treatment of CML but only the reverse transcriptase (RT) polymerase chain reaction (PCR) method is sufficiently sensitive to detect minimal residual disease. RT-PCR is known to detect as few as one cell expressing BCR-ABL in a total of about 100,000 cells. However, the standard RT-PCR method is widely used as not quantitative. Competitive quantitative PCR methods have been developed in recent years. We are adapting an approach developed by Cross and Goldman. This method involves adding known levels of BCR-ABL junction sequences, containing an insert to distinguish it from the native junction sequences, to an unknown sample and comparing the intensity of the amplimers. The reliability of the method is increased by analyzing serial patient samples from a period of time to determine whether the level of BCR-ABL-RNA is decreasing or increasing. For example, a pattern of decreasing level of BCR-ABL RNA would indicate a deepening remission. The aims of this grant are to adapt the Cross and Goldman method and use the test to monitor bone marrow transplant CML patients and CML patients being treated with interferon alpha. This approach will be also used in Ph+ ALL patients. The method will be established using known Bcr-Abl positive and negative cell lines; next studies will be done on cells from blood and marrow with emphasis on minimal residual disease; and the utility will be evaluated by performing a blind study with patient samples. Lastly, a core laboratory will be established for analysis blood and marrow samples of CML patient samples under study within the P01 grant by competitive PCR.