Department of Oncology, Shanghai Xuhui District Center Hospital, Shanghai 200031, China.
World J Gastroenterol. 2012 Jul 28;18(28):3745-51. doi: 10.3748/wjg.v18.i28.3745.
To assess the significance of phosphatidylinositol 3-kinase (PI3K) in colorectal cancer (CRC) and toxicity of LY294002 in CRC cells with different metastatic abilities.
Sixty formalin-fixed and paraffin-embedded CRC tumor specimens were investigated. Adjacent normal colonic mucosa specimens from 10 of these cases were selected as controls. PI3K protein was detected by immunohistochemistry and PIK3CA mutations were investigated by gene sequencing analysis. A flow-cytometry-based apoptosis detection kit was used to determine PI3K inhibitor-induced apoptosis in CRC cell lines SW480 and SW620. Expression of phosphorylated protein kinase B in CRC cell lines was detected by Western blotting.
There was a significant difference in the proportion of primary lesions (30%, 18/60) vs metastatic lesions (46.7%, 28/60) that were positive for PI3K (P < 0.05). Mutations were detected in exon 9 (13.3%) and exon 20 (8.3%). Out of 60 cases, seven mutations were identified: two hotspot mutations, C.1633G>A resulting in E545A, and C.3140A>G resulting in H1047R; two novel missense mutations C.1624G>A and C.3079G>A; and three synonymous mutations (C.1641G>A, C.1581C>T and C.3027T>A). Exposure of SW480 cells to PI3K inhibitor for 48 h resulted in a significant increase of apoptotic cells in a dose-dependent manner [3.2% apoptotic cells in 0 μmol/L, 4.3% in 5 μmol/L, 6.3% in 10 μmol/L (P < 0.05), and 6.7% in 20 μmol/L (P < 0.05)]. Moreover, PI3K inhibitor induced a similar significant increase of apoptotic cells in the SW620 cell line for 48 h [3.3% apoptotic cells in 0 μmol/L, 13.3% in 5 μmol/L (P < 0.01), 19.2% in 10 μmol/L (P < 0.01), and 21.3% in 20 μmol/L (P < 0.01)].
High PI3K expression is associated with CRC metastasis. PI3K inhibitor induced apoptosis in CRC cells and displayed strong cytotoxicity for highly metastatic cells. PI3K inhibition may be an effective treatment for CRC.
评估磷脂酰肌醇 3-激酶（PI3K）在结直肠癌（CRC）中的意义和 LY294002 对不同转移能力的 CRC 细胞的毒性。
检测了 60 例福尔马林固定石蜡包埋的 CRC 肿瘤标本。从其中 10 例中选择相邻的正常结肠黏膜标本作为对照。采用免疫组织化学法检测 PI3K 蛋白，基因测序分析检测 PIK3CA 突变。使用基于流式细胞术的凋亡检测试剂盒检测 CRC 细胞系 SW480 和 SW620 中 PI3K 抑制剂诱导的凋亡。用 Western blot 检测 CRC 细胞系中磷酸化蛋白激酶 B 的表达。
PI3K 阳性的原发灶（30%，18/60）与转移灶（46.7%，28/60）之间存在显著差异（P<0.05）。在外显子 9（13.3%）和外显子 20（8.3%）中检测到突变。在 60 例中发现了 7 种突变：两种热点突变，C.1633G>A 导致 E545A 和 C.3140A>G 导致 H1047R；两种新的错义突变 C.1624G>A 和 C.3079G>A；以及三种同义突变 C.1641G>A、C.1581C>T 和 C.3027T>A。PI3K 抑制剂暴露于 SW480 细胞 48 小时后，细胞凋亡呈剂量依赖性显著增加[0μmol/L 时 3.2%凋亡细胞，5μmol/L 时 4.3%（P<0.05），10μmol/L 时 6.3%（P<0.05），20μmol/L 时 6.7%（P<0.05）]。此外，PI3K 抑制剂在 SW620 细胞系中也诱导了类似的 48 小时细胞凋亡显著增加[0μmol/L 时 3.3%凋亡细胞，5μmol/L 时 13.3%（P<0.01），10μmol/L 时 19.2%（P<0.01），20μmol/L 时 21.3%（P<0.01）]。
高 PI3K 表达与 CRC 转移有关。PI3K 抑制剂诱导 CRC 细胞凋亡，并对高转移性细胞表现出强烈的细胞毒性。PI3K 抑制可能是 CRC 的有效治疗方法。
