Institute of Pathology, University of Bern, Murtenstrasse 31, Room L310, 3008, Bern, Switzerland.
Graduate School for Cellular and Biomedical Sciences, University of Bern, Freiestrasse 1, 3012, Bern, Switzerland.
Clin Epigenetics. 2018 Sep 26;10(1):120. doi: 10.1186/s13148-018-0548-2.
In colorectal cancer, CDX2 expression is lost in approximately 20% of cases and associated with poor outcome. Here, we aim to validate the clinical impact of CDX2 and investigate the role of promoter methylation and histone deacetylation in CDX2 repression and restoration.
CDX2 immunohistochemistry was performed on multi-punch tissue microarrays (n = 637 patients). Promoter methylation and protein expression investigated on 11 colorectal cancer cell lines identified two CDX2 low expressors (SW620, COLO205) for treatment with decitabine (DNA methyltransferase inhibitor), trichostatin A (TSA) (general HDAC inhibitor), and LMK-235 (specific HDAC4 and HDAC5 inhibitor). RNA and protein levels were assessed. HDAC5 recruitment to the CDX2 gene promoter region was tested by chromatin immunoprecipitation.
Sixty percent of tumors showed focal CDX2 loss; 5% were negative. Reduced CDX2 was associated with lymph node metastasis (p = 0.0167), distant metastasis (p = 0.0123), and unfavorable survival (multivariate analysis: p = 0.0008; HR (95%CI) 0.922 (0.988-0.997)) as well as BRAF, mismatch repair deficiency, and CpG island methylator phenotype. Decitabine treatment alone induced CDX2 RNA and protein with values from 2- to 25-fold. TSA treatment ± decitabine also led to successful restoration of RNA and/or protein. Treatment with LMK-235 alone had marked effects on RNA and protein levels, mainly in COLO205 cells that responded less to decitabine. Lastly, decitabine co-treatment was more effective than LMK-235 alone at restoring CDX2.
CDX2 loss is an adverse prognostic factor and linked to molecular features of the serrated pathway. RNA/protein expression is restored in CDX2 low-expressing CRC cell lines by demethylation and HDAC inhibition. Importantly, our data underline HDAC4 and HDAC5 as new epigenetic CDX2 regulators that warrant further investigation.
在结直肠癌中，约 20%的病例中 CDX2 表达缺失，并与不良预后相关。在此，我们旨在验证 CDX2 的临床影响，并研究启动子甲基化和组蛋白去乙酰化在 CDX2 抑制和恢复中的作用。
对多针组织微阵列（n=637 例患者）进行 CDX2 免疫组织化学染色。在 11 种结直肠癌细胞系中检测启动子甲基化和蛋白表达，确定了两种 CDX2 低表达细胞系（SW620、COLO205），用于使用地西他滨（DNA 甲基转移酶抑制剂）、曲古抑菌素 A（TSA）（通用组蛋白去乙酰化酶抑制剂）和 LMK-235（特异性组蛋白去乙酰化酶 4 和 5 抑制剂）进行治疗。评估 RNA 和蛋白水平。通过染色质免疫沉淀检测 HDAC5 募集到 CDX2 基因启动子区域。
60%的肿瘤表现为局灶性 CDX2 缺失；5%为阴性。CDX2 表达降低与淋巴结转移（p=0.0167）、远处转移（p=0.0123）和不良生存相关（多变量分析：p=0.0008；HR（95%CI）0.922（0.988-0.997））以及 BRAF、错配修复缺陷和 CpG 岛甲基化表型。单独使用地西他滨治疗可诱导 CDX2 RNA 和蛋白表达增加 2-25 倍。TSA 治疗±地西他滨也能成功恢复 RNA 和/或蛋白。单独使用 LMK-235 治疗对 RNA 和蛋白水平有显著影响，主要在 COLO205 细胞中，对地西他滨的反应较小。最后，与单独使用 LMK-235 相比，地西他滨联合治疗在恢复 CDX2 方面更有效。
CDX2 缺失是一个不良的预后因素，并与锯齿状途径的分子特征相关。在 CDX2 低表达的结直肠癌细胞系中，通过去甲基化和组蛋白去乙酰化抑制可恢复 RNA/蛋白表达。重要的是，我们的数据强调了组蛋白去乙酰化酶 4 和 5 作为新的 CDX2 表观遗传调控因子，值得进一步研究。
