Functional analysis of PIK3CA gene mutations in human colorectal cancer
Functional analysis of PIK3CA gene mutations in human colorectal cancer
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DOI：
10.1158/0008-5472.can-04-4114
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发表时间：
2005-06-01
期刊：
影响因子：
11.2
通讯作者：
Omata, M
中科院分区：
文献类型：
--
作者：
Ikenoue, T;Kanai, F;Omata, M
PIK3CA基因编码磷脂酰肌醇3-激酶（PI3K）的p110 α催化亚基，其突变已在包括结直肠癌在内的人类癌症中被报道。大多数突变集中在螺旋结构域和激酶结构域的热点上。据报道，热点突变体之一14104711具有升高的脂激酶活性，而其他突变的功能后果尚未得到研究。在本研究中，我们检测了结肠癌相关PIK3CA突变对体外脂激酶活性、体内下游靶点Akt和p70S6K的激活以及NIH 3t3转化能力的影响。在检测的8个突变中，与野生型p110 α相比，所有突变都显示出脂激酶活性增加。与野生型p110 α相比，所有突变体都能强烈激活Akt和p70S6K，通过磷酸化特异性抗体进行免疫印迹检测。这些突变体还诱导了NIH 3T3细胞的形态改变、接触抑制丧失和不依赖于锚定的生长。本研究检测的热点突变E542K、E545K和H1047R都具有较高的酶和转化活性。这些结果表明，几乎所有结肠癌相关的P1K3CA突变都具有功能活性，因此它们很可能参与了癌变。
Mutations in the PIK3CA gene, which encodes the p110 alpha catalytic subunit of phosphatidylinositol 3-kinase (PI3K), have been reported in human cancers, including colorectal cancer. Most of the mutations cluster at hotspots within the helical and kinase domains. Whereas 14104711, one of the hotspot mutants, is reported to have elevated lipid kinase activity, the functional consequences of other mutations have not been examined. In this study, we examined the effects of colon cancer-associated PIK3CA mutations on the lipid kinase activity in vitro, activation of the downstream targets Akt and p70S6K in vivo and NIH 3T3-transforming ability. Of eight mutations examined, all showed increased lipid kinase activity compared with wild-type p110 alpha. All the mutants strongly activated Akt and p70S6K compared with wild-type p110 alpha as determined by immunoblotting using phospho-specific antibodies. These mutants also induced morphologic changes, loss of contact inhibition, and anchorage-independent growth of NIH 3T3 cells. The hotspot mutations examined in this study, E542K, E545K, and H1047R, all had high enzymatic and transforming activities. These results show that almost all the colon cancer-associated P1K3CA mutations are functionally active so that they are likely to be involved in carcinogenesis.