c-Kit Mutations and Their Role in Tumor Biology

描述(申请人提供)：肥大细胞肿瘤(MCT)是最常见的被诊断为狗的恶性肿瘤。我们先前已经证明，至少30%的犬MCT具有原癌基因c-kit的突变，该突变由编码负调控膜旁结构域的第11/12外显子的串联复制组成。这些突变导致Kit在没有配体结合的情况下发生结构性磷酸化。Kit的异常表达也被发现存在于许多人类肿瘤中，包括异常表达(小细胞肺癌、泌尿生殖系肿瘤)和细胞质区域的突变导致结构性激活(肥大细胞紊乱、胃肠道间质瘤)。最近对各种实验性激酶抑制剂的研究表明，抑制Kit信号转导可能对这些恶性肿瘤患者有显著好处。然而，为了它们的成功应用，更清楚地确定c-kit突变在肿瘤疾病的发生和发展中的作用是至关重要的。这一假设的基础是，c-kit的激活突变通过促进细胞存活和诱导与侵袭和转移相关的金属蛋白酶基因表达，与恶性肥大细胞疾病的发生和发展有关。本研究的具体目的是：1)对犬MCT中c-kit突变进行细致的分析，包括它们对MCT生物学行为的影响，并确定与其发育相关的潜在风险因素；2)通过建立在高度调控的可诱导启动子下表达各种形式的c-kit突变的转基因小鼠，研究Kit调控失调对体内正常细胞群体的影响；以及3)评估吲哚酮激酶抑制剂在体外和Kit突变小鼠模型中对Kit调控失调的影响。结合对小鼠Kit失调的详细研究和对狗c-Kit自发突变模型的全面研究，将有助于澄清此类突变的生物学和生化后果。此外，将激酶抑制剂纳入这些研究提供了一个独特的机会来评估这些药物在治疗Kit功能障碍明显的肿瘤疾病中的潜在有用性和有效性。

DESCRIPTION (provided by applicant): Mast cell tumors (MCTs) are the most frequently diagnosed malignant tumor of the dog. We have previously demonstrated that at least 30% of canine MCTs possess mutations in the proto-oncogene c-kit consisting of tandem duplications in exons 11/12 encoding the negative regulatory juxtamembrane domain. These mutations result in constitutive phosphorylation of Kit in the absence of ligand binding. Dysregulation of Kit has also been found to occur in many human tumors including aberrant expression (small cell lung carcinoma, genitourinary cancers) and mutation in the cytoplasmic domain leading to constitutive activation (mast cell disorders, gastrointestinal stromal tumors). Recent studies with various experimental kinase inhibitors suggest that inhibition of Kit signaling may be of significant benefit to patients with these malignancies. However, for their application to be successful, it is critical that the role of c-kit mutations in the initiation and progression of neoplastic disorders be more clearly defined. The hypothesis underlying this proposal is that activating mutations of c-kit are associated with the development and progression of malignant mast cell disease through the promotion of cell survival and induction of metalloproteinase gene expression linked to invasion and metastasis. The specific aims of this study are: 1) to undertake a meticulous characterization of c-kit mutations in dog MCTs including their impact on the biologic behavior of MCTs, and identification of potential risk factors associated with their development; 2) to study the effect of Kit dysregulation on normal cell populations in vivo through the generation of transgenic mice expressing various forms of mutant c-kit under a highly regulated inducible promoter; and 3) to evaluate the effects of indolinone kinase inhibitors on dysregulated Kit both in vitro and in mouse models of Kit mutation. The integration of detailed investigations of Kit dysregulation in the mouse with comprehensive studies of a spontaneous model of c-kit mutation in the dog will help to clarify the biological and biochemical consequences of such mutations. Moreover, the incorporation of kinase inhibitors into these studies offers a unique opportunity to evaluate the potential usefulness and efficacy of such agents in the treatment of neoplastic diseases in which Kit dysfunction is evident.