Baharudin Rashidah, Ab Mutalib Nurul-Syakima, Othman Sri N, Sagap Ismail, Rose Isa M, Mohd Mokhtar Norfilza, Jamal Rahman
UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia Kuala Lumpur, Malaysia.
Department of Surgery, Faculty of Medicine, Universiti Kebangsaan Malaysia Kuala Lumpur, Malaysia.
Front Pharmacol. 2017 Feb 13;8:47. doi: 10.3389/fphar.2017.00047. eCollection 2017.
Resistance to 5-Fluorouracil (5-FU) is a major obstacle to the successful treatment of colorectal cancer (CRC) and posed an increased risk of recurrence. DNA methylation has been suggested as one of the underlying mechanisms for recurrent disease and its contribution to the development of drug resistance remains to be clarified. This study aimed to determine the methylation phenotype in CRC for identification of predictive markers for chemotherapy response. We performed DNA methylation profiling on 43 non-recurrent and five recurrent CRC patients using the Illumina Infinium HumanMethylation450 Beadchip assay. In addition, CRC cells with different genetic backgrounds, response to 5-FU and global methylation levels (HT29 and SW48) were treated with 5-FU and DNA methylation inhibitor 5-aza-2'-deoxycytidine (5-azadC). The singular and combined effects of these two drug classes on cell viability and global methylation profiles were investigated. Our genome-wide methylation study on the clinical specimens showed that recurrent CRCs exhibited higher methylation levels compared to non-recurrent CRCs. We identified 4787 significantly differentially methylated genes ( < 0.05); 3112 genes were hyper- while 1675 genes were hypomethylated in the recurrent group compared to the non-recurrent. Fifty eight and 47 of the significantly hypermethylated and hypomethylated genes have an absolute recurrent/non-recurrent methylation difference of ≥20%. Most of the hypermethylated genes were involved in the MAPK signaling pathway which is a key regulator for apoptosis while the hypomethylated genes were involved in the PI3K-AKT signaling pathway and proliferation process. We also demonstrate that 5-azadC treatment enhanced response to 5-FU which resulted in significant growth inhibition compared to 5-FU alone in hypermethylated cell lines SW48. In conclusion, we found the evidence of five potentially biologically important genes in recurrent CRCs that could possibly serve as a new potential therapeutic targets for patients with chemoresistance. We postulate that aberrant methylation of , and in CRC might be associated with the recurrence of CRC and 5-azadC-mediated restoration of 5-FU sensitivity is mediated at least in part by MAPK signaling pathway.
对5-氟尿嘧啶（5-FU）耐药是结直肠癌（CRC）成功治疗的主要障碍，并增加了复发风险。DNA甲基化被认为是复发性疾病的潜在机制之一，其对耐药性发展的作用仍有待阐明。本研究旨在确定CRC中的甲基化表型，以鉴定化疗反应的预测标志物。我们使用Illumina Infinium HumanMethylation450 Beadchip检测法对43例非复发性和5例复发性CRC患者进行了DNA甲基化分析。此外，用5-FU和DNA甲基化抑制剂5-氮杂-2'-脱氧胞苷（5-azadC）处理具有不同遗传背景、对5-FU的反应和整体甲基化水平的CRC细胞（HT29和SW48）。研究了这两类药物对细胞活力和整体甲基化谱的单一和联合作用。我们对临床标本进行的全基因组甲基化研究表明，与非复发性CRC相比，复发性CRC表现出更高的甲基化水平。我们鉴定出4787个显著差异甲基化基因（<0.05）；与非复发性组相比，复发性组中有3112个基因高甲基化，1675个基因低甲基化。58个和47个显著高甲基化和低甲基化基因的复发/非复发甲基化绝对差异≥20%。大多数高甲基化基因参与MAPK信号通路，这是细胞凋亡的关键调节因子，而低甲基化基因参与PI3K-AKT信号通路和增殖过程。我们还证明，在高甲基化细胞系SW48中，5-azadC处理增强了对5-FU的反应，与单独使用5-FU相比，导致显著的生长抑制。总之，我们在复发性CRC中发现了五个潜在生物学重要基因的证据，这些基因可能成为化疗耐药患者的新潜在治疗靶点。我们推测，CRC中、和的异常甲基化可能与CRC的复发有关，5-azadC介导的5-FU敏感性恢复至少部分由MAPK信号通路介导。
