Role of Vav and Rac in KIT oncogenesis

描述（由申请方提供）：KIT是一种独特的受体，在黑素细胞、生殖细胞、Cajal间质细胞、肥大细胞和造血干细胞中具有重要功能作用。与KIT在这些定义的组织中的重要性一致，在生殖细胞肿瘤、胃肠道间质瘤（GIST）、鼻窦淋巴瘤、急性髓性白血病（AML）和系统性肥大细胞增多症（SM）中描述了KIT的激活突变。这些疾病中的一个显著比例通常携带KIT激活环突变KITD 816 V。在核心结合因子-急性髓性白血病（CBF-AML）中也观察到KIT的激活环突变，这些白血病携带t（8;21）或inv（16）细胞遗传学异常，分别产生融合基因AML 1- ETO或CBF 2-MYH 11。对成人和儿童AML的研究表明，携带t（8;21）的CBF-AML中KITD 816 V突变的存在会影响基于几个临床指标的预后。尽管在GIST中常见的内膜区域内的KIT突变对酪氨酸激酶抑制剂甲磺酸伊马替尼（Gleevec）的抑制敏感;但酪氨酸激酶结构域的羧基末端叶内的KIT突变（如KITD 816 V）使KIT活化环构象稳定在其活性形式，从而阻止了伊马替尼与酪氨酸激酶抑制剂的充分结合。因此，与成功使用Gleevec治疗GIST相反，Gleevec对于治疗携带KIT活化环突变体（即KITD 816 V）的血液疾病（包括SM和CBF-AML）无效。重要的是，KIT（KITD 816 V）中激活环突变诱导原代造血细胞转化的受体近端和/或下游信号的性质定义不清。我们有证据证明KITD 816 V（小鼠中的KITD 814 V）诱导的原代造血干细胞和祖细胞转化导致GEF Vav/Rho GT3 Rac途径的组成型激活，并且小鼠中造血特异性Vav 1和/或Rac 2的遗传破坏通过KITD 814 V消除了配体非依赖性生长，这使我们假设参与KITD 814 V诱导的转化的信号可能部分地通过该途径的超活化来介导。此外，我们有证据表明突变KITD 814 V细胞内结构域内的酪氨酸残基导致KITD 814 V诱导的配体非依赖性生长的完全丧失，这使我们假设细胞膜内的酪氨酸和KITD 814 V的激酶插入区域可能有助于KITD 814 V诱导的转化。基于这些发现，本申请的中心假设是Vav/Rac通路的过度活化促成与系统性肥大细胞增多症、AML以及涉及KITD 814 V突变的其他疾病相关的疾病的病因。我们提出的研究将提供对Vav/Rac途径的生理意义以及质膜和激酶插入序列参与调节KITD 814 V诱导的转化的机制性见解，目前还没有药物存在。公共卫生相关性：现在明确的是，激活突变的KIT有助于生殖细胞肿瘤，胃肠道间质瘤（GIST），鼻窦淋巴瘤，急性髓性白血病（AML）和系统性肥大细胞增多症（SM）。我们提出的研究将提供机制的洞察信号通路，通过激活KIT突变，目前没有药物存在的调节转化。我们的研究结果有望为AML和SM等疾病的分子治疗提供新的靶点。

DESCRIPTION (provided by applicant): KIT is a unique receptor with important functional roles in melanocytes, germ cells, interstitial cells of Cajal, mast cells, and hematopoietic stem cells. Consistent with the importance of KIT in these defined tissues, activating mutations of KIT have been described in germ cell tumors, gastrointestinal stromal tumors (GISTs), sinonasal lymphomas, acute myeloid leukemia (AML), and systemic mastocytosis (SM). A significant proportion of these diseases commonly bear the KIT activation loop mutation KITD816V. Activation loop mutations of KIT have also been observed in core binding factor-acute myeloid leukemia (CBF-AML), leukemias that bear either the t(8;21) or inv(16) cytogenetic abnormality, generating the fusion genes AML1- ETO or CBF2-MYH11, respectively. Studies examining both adult and pediatric AML have indicated that the presence of the KITD816V mutation in CBF-AML carrying t(8;21) worsens the prognosis based on several clinical indices. Although KIT mutations within the juxtamembrane region that are commonly found in GISTs are sensitive to inhibition by the tyrosine kinase inhibitor, imatinib mesylate (Gleevec); KIT mutations within the carboxy-terminal lobe of the tyrosine kinase domain, such as KITD816V, stabilizes the KIT activation loop conformation in its active form, which precludes sufficient imatinib binding for tyrosine kinase inhibition. Therefore, in contrast to successful use of Gleevec to treat GISTs, Gleevec is ineffective for the treatment of the hematologic diseases harboring the KIT activation loop mutants (i.e. KITD816V), including SM and CBF- AML. Importantly, nature of the receptor proximal and/or downstream signals by which activation loop mutations in KIT (KITD816V) induce transformation in primary hematopoietic cells are poorly defined. We have evidence to demonstrate that KITD816V (KITD814V in mice) induced transformation in primary hematopoietic stem and progenitor cells results in constitutive activation of GEF Vav/Rho GTPase Rac pathway and that genetic disruption of hematopoietic specific Vav1 and/or Rac2 in mice abrogates ligand independent growth via KITD814V, leading us to hypothesize that signals involved in KITD814V induced transformation may in part be mediated via the hyperactivation of this pathway. Furthermore, we have evidence demonstrating that mutating the tyrosine residues within the intracellular domain of KITD814V results in complete loss of KITD814V induced ligand independent growth, leading us to hypothesize that the intracellular tyrosines within the juxtamembrane and the kinase insert region of KITD814V are likely to contribute to KITD814V induced transformation. Based on these findings, the central hypothesis of this application is that hyperactivation of the Vav/Rac pathway contributes to the etiology of diseases associated with systemic mastocytosis, AML as well as other diseases involving the KITD814V mutation. Our proposed studies will provide mechanistic insight into the physiologic significance of the Vav/Rac pathway as well as the involvement of the juxtamembrane and the kinase insert sequences in regulating KITD814V induced transformation for which currently no drugs exist. PUBLIC HEALTH RELEVANCE: It is now unequivocal that activating mutations of KIT contribute to germ cell tumors, gastrointestinal stromal tumors (GISTs), sinonasal lymphomas, acute myeloid leukemia (AML), and systemic mastocytosis (SM). Our proposed studies will provide mechanistic insight into the signaling pathways that regulate transformation via an activating KIT mutation for which currently no drugs exist. Our results are expected to provide new targets for molecular therapies for the treatment of diseases such as AML and SM.