Pathological analysis of GIST using transgenic or knock-in-mouse

首先，我们试图产生具有c-kit基因突变的转基因小鼠，这种突变在家族性和多发性GIST(胃肠道间质瘤)患者中可见。获得了两种类型的转基因小鼠，一种是膜旁结构域突变，另一种是酪氨酸激酶n结构域突变。然而，这些小鼠既没有表现出Cajal间质细胞(IGCs)的增殖，也没有表现出多发性GIST。我们正在产生敲击小鼠，作为人类家族GIST的更具生理学的模型。敲击小鼠在酪氨酸激酶II结构域有c-kit基因突变。我们将在不久的将来研究敲击小鼠。在上述过程中，我们检测了家族性GIST患者ICCs弥漫性增殖的克隆性。增生性病变呈多克隆性，而每一种GIST均为单克隆性。我们还表明，选择性酪氨酸激酶抑制剂伊马替尼不能有效地抑制由外显子17突变引起的KIT激活。Imatinib也不能完全抑制KIT信号转导的下游分子。我们探讨了GIST无c-KIT基因突变的原因。在无c-kit基因突变的GIST中，约有一半存在PDGFRα基因突变。我们发现了两种类型的PDGFRα基因突变，Imatinib有效地抑制了膜旁区域的突变，但酪氨酸激酶n区域的突变没有。我们证明了在Imatinib治疗期间GIST的再生长(产生耐药克隆)是由于在原有c-kit基因突变的基础上增加了c-kit基因突变所致。此外，我们还证明了来自1型神经母细胞瘤患者的GIST没有任何c-kit基因突变。

First, we tried to generate transgenic mice possessing c-kit gene mutation which was seen in patients with familial and multiple GISTs (gastrointestinal stromal tumors). Two types of transgenic mice was gained; one had juxtamembrane domain mutation and the other had tyrosine kinase n domain mutation. However, these mice showed neither hyperplasia of interstitial cells of Cajal (IGCs) nor multiple GISTs. We are generating knock-in-mouse as a more physiological model of human familial GISTs. The knock-in-mouse has c-kit gene mutation at tyrosine kinase II domain. We will investigate the knock-in-mouse in near future.During the above process, we examined the clonality of diffuse proliferation of ICCs in familial GIST patients. The proliferate lesion showed polyclonal nature while each GIST demonstrated to be monoclonal. We also showed that KIT activation by exon 17 mutation was not effectively inhibited by a selective tyrosine kinase inhibitor, Imatinib. The downstream molecules of KIT signal transduction were not also fully inhibited by Imatinib.We investigated the cause of GISTs without c-kit gene mutation. Approximately half of GISTs without c-kit gene mutation had PDGFR alpha gene mutation. Two types of PDGFR alpha gene mutation were seen, and the juxtamembrane domain mutation was effectively inhibited by Imatinib but the tyrosine kinase n domain mutation was not We demonstrated that regrowth of GISTs during the Imatinib treatment (development of resistant clone) was caused by an additional c-kit gene mutation to original c-kit gene mutatioa Moreover, we showed that GISTs from neuroflbromatosis type1 patients did not have any c-kit gene mutation.