Department of General Surgery, The 305 hospital of People's Liberation Army, Beijing, 100017, China.
Department of Anesthesiology, The 305 hospital of People's Liberation Army, Beijing, 100017, China.
Cell Biochem Biophys. 2023 Sep;81(3):569-576. doi: 10.1007/s12013-023-01158-9. Epub 2023 Aug 12.
Colorectal cancer is a malignant tumor with higher morbidity and mortality. The purpose of this study is to investigate whether inhibition of Protein Kinase, Membrane Associated Tyrosine/Threonine 1 (PKMYT1) affects tumor cell proliferation, survival and migration in colon tumors with high Cyclin E1 (CCNE1) expression. PcDNA3.1-CCNE1 vector and si-PKMYT1 were transfected in SW480 cells by Lipofectamine 2000. Q-PCR and western blot assay were processed to detect the expression. Transwell assay and Edu assay were undertaken to verify the migration and proliferation. CCNE1 promotes the proliferation and migration of SW480. Silencing of PKMYT1 inhibited the proliferation of tumor cells. Silencing the expression of PKMYT1 under the premise of overexpression of CCNE1, the level of Cyclin Dependent Kinase 1 (CDK1)-PT14 was reduced, indicating that the cell cycle was blocked. The expression of γH2AX increased significantly, indicating that the DDR pathway of tumor cells was activated and DNA damage accumulated. The results of immunofluorescence microscopy showed significantly increased expression of DNA damage-associated marker (γH2AX: H2AX Variant Histone). In CCNE1 amplificated colorectal tumor cells, knockdown of PKMYT1 reduced cells in S phase, inhibited cell proliferation and promoted cell apoptosis, confirming that PKMYT1 was a potential therapeutic target for colorectal tumor. This study may verify a potential therapeutic target and provide a new idea for the treatment of colorectal cancer in the future.
结直肠癌是一种发病率和死亡率较高的恶性肿瘤。本研究旨在探讨抑制蛋白激酶，膜相关酪氨酸/苏氨酸 1（PKMYT1）是否会影响高细胞周期蛋白 E1（CCNE1）表达的结肠肿瘤中的肿瘤细胞增殖、存活和迁移。通过 Lipofectamine 2000 将 PcDNA3.1-CCNE1 载体和 si-PKMYT1 转染到 SW480 细胞中。通过 Q-PCR 和 Western blot 检测来检测表达情况。通过 Transwell 检测和 Edu 检测来验证迁移和增殖情况。CCNE1 促进了 SW480 的增殖和迁移。沉默 PKMYT1 抑制了肿瘤细胞的增殖。在过表达 CCNE1 的前提下沉默 PKMYT1 的表达，细胞周期蛋白依赖性激酶 1（CDK1）-PT14 的水平降低，表明细胞周期被阻断。γH2AX 的表达显著增加，表明肿瘤细胞的 DDR 途径被激活，DNA 损伤累积。免疫荧光显微镜的结果显示，与 DNA 损伤相关的标记物（γH2AX：H2AX 变体组蛋白）的表达显著增加。在 CCNE1 扩增的结直肠肿瘤细胞中，PKMYT1 的敲低减少了 S 期细胞，抑制了细胞增殖并促进了细胞凋亡，证实 PKMYT1 是结直肠肿瘤的潜在治疗靶点。本研究可能验证了一个潜在的治疗靶点，并为未来结直肠癌的治疗提供了新的思路。
