The Clinical Significance Of Lysine Methylation Of Histone H3 In Colorectal Cancer

ObjectiveWe aimed to explore the expressions of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 protein in colorectal cancer tissues,and their relationship with clinicopathological features and prognosis.Nextly,we purposed to demonstrate the effect of WNT/β-catenin signaling pathway on H3K4me2,H3K4me3,H3K9me3 and H3K27me3 proteins.Methods1.We collected specimens and clinical data of colorectal cancer surgery patients and follow-up,and then used tissue microarray and immunohistochemical techniques to detect the protein expressions of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 in colorectal cancer tissues.Furthermore,we analyzed the relationships between H3K4me2,H3K4me3,H3K9me3,H3K27me3 and clinicopathological features,and investigated which influential factor would be a predictor of prognosis for patients with colorectal cancer.2.We treated colon cancer cells(SW480 and SW620)with Wnt/β-catenin signaling pathway small-molecule regulators XAV939 and BIO,and observed the protein expressions of β-catenin,H3K4me2,H3K4me3,H3K9me3 and H3K27me3 using Western blot.ResultsClinical significance of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 protein expression in colorectal cancer1.The positive protein expression levels of H3K4me2,H3K4me3,H3K9me3,H3K27me3 were significantly higher in colorectal cancer tissues,which was 43.5%,13.0%,78.2%,3.7% respectively,comparing with adjacent normal tissues(p<0.001).We found the positive protein expression rates of H3K4me3,H3K9me3 were significantly higher in primary colorectal cancer,which was 18.5%,82.1% respectively,comparing with lymph node metastatic cancer tissues(p<0.05).We did not find any significant difference of the positive protein expression rates of H3K4me2 and H3K27me3 in primary colorectal cancer and lymph node metastatic cancer tissues(p>0.05).2.There were significant higher of H3K4me2 protein staining in male(48.2%),high histological grade(55.2%)patients than those female(37.6%),low histological grade(42.1%)patients(p<0.05).The positive expression of H3K4me3,H3K9me3 and H3K27me3 proteins were not correlated with gender,histological grade(p>0.05).H3K4me2,H3K4me3,H3K9me3 and H3K27me3 proteins staining in colorectal cancer patients with lymph node metastasis(56.2%,17.8%,83.0% and 5.1%).It was significantly higher than that in those without lymph node metastasis(32.0%,8.6%,73.8%and 2.4%)(p<0.05).H3K4me2,H3K4me3 and H3K9me3 proteins staining in colorectal cancer patients with TNM stage III-IV(54.5%,16.5% and 81.9%),which were significantly higher than those in patients with TNM stage I-II(29.9%,8.6% and73.6%)(p<0.05).However,the positive expression of H3K27me3 protein were not correlated with TNM stage(p>0.05).The positive expression of H3K4me2,H3K4me3,H3K9me3,and H3K27me3 proteins were not correlated with age,tumor site and histological type(p>0.05).3.Spearman correlation analysis showed that H3K4me2 and H3K4me3,H3K4me2 and H3K9me3,H3K4me2 and H3K27me3,H3K9me3 and H3K27me3 protein expression were statistically significant(r=0.201,p<0.001;r= 0.382,p <0.001;r=0.197,p<0.001;r=0.087,p<0.05).4.Survival analysis showed that the 5-year survival rate of patients with colorectal cancer after surgery was 64.9%.Univariate analysis showed that age,histological type,histological grade,lymph node metastasis,TNM stage,H3K4me2 and H3K9me3 proteins expression had significantly associated with survival(p<0.05);Multivariate Cox regression demonstrated that age(HR=1.039,p<0.001)and TNM stage(HR=4.564,p<0.001)were independent prognostic factors in colorectal cancer patients.5.The colorectal cancer patients with TNM III-IV stage,who accepted chemotherapy,had significantly longer overall survival than who without.Furthermore,the patients with expressions of H3K4me2,H3K9me3 positive and H3K4me3,H3K27me3 negative had significantly longer overall survival after chemotherapy than those without chemotherapy(p<0.05),while the patients with protein expressions of H3K4me2,H3K9me3 negative and H3K4me3,H3K27me3 positive regardless of chemotherapy whether or not,there was no statistical difference in their overall survival time(p>0.05).The effects of Wnt/β-catenin signaling pathway on the expressions of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 protein in colon cancer cells1.BIO treatment of SW480 and SW620 cells upregulated β-catenin protein expression,and H3K4me3 protein expressions was also upregulated,while H3K4me2,H3K9me3 and H3K27me3 protein expressions was decreased.2.XAV939 decreased β-catenin and H3K4me3 protein expression markedly in SW480 and SW620 cells,while H3K4me2,H3K9me3 and H3K27me3 protein expressions was increased.Conclusion1.The expression rates of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 protein in colorectal cancer tissues were higher than adjacent normal tissues,suggesting that the above four histone methylation may participate in colorectal carcinogenesis.2.The positive expression rates of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 proteins in colorectal cancer patients with lymph node metastasis was higher than that in the group without lymph node metastasis.The positive expression rates of H3K4me2,H3K4me3 and H3K9me3 proteins in colorectal cancer stage TNM III-IV was significantly higher than that of TNM stage I-II,suggesting that the above four histone methylation may promote the progression of colorectal cancer.3.The patients who suffered from TNM stage III or IV colorectal cancer,with positive or negative expression of H3K4me2,H3K4me3,H3K9me3 and H3K27me3,had different reaction on chemotherapy.H3K4me2,H3K4me3,H3K9me3 and H3K27me3 would be recognized as biomarkers,and guided postoperative chemotherapy in colorectal cancer patients with TNM III-IV stage.4.The protein expression of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 was related to the expression of β-catenin in colorectal cancer tissues.It was verified that the activity of Wnt/β-catenin pathway affected the protein expressions of H3K4me2,H3K4me3,H3K9me3 and H3K27me3 in vitro.