探討CBP磷酸化缺失對AOM/DSS誘發結腸癌之作用

Abstract

大腸直腸癌(CRC)是致死率全球第二大的惡性腫瘤。2018年，全球約有90萬人死於該疾病。而inflammatory bowel disease (IBD)病人得到CRC的機率會高出2.4倍。CREB-binding protein (CBP)具有histone acetyltransferase的功能，使它可以乙烯化(acetylate) histone tail，促進基因表達。在我們先前的研究發現，IKKα磷酸化CBP Ser1382/1386會抑制p53-mediated gene的表達，同時促進NF-κB signaling。CBP Ser1383/1387Ala knockin mice(AA-mice) ( Ser1383/1387無法被IKK α磷酸化) (對應人類CBP Ser1382/1386)會有自發性的腸發炎；給予colitogenic chemical (Dextran sulfate sodium, DSS)後，會引起更嚴重的發炎反應。但對於colitis-associated cancer (CAC)的影響仍然未知。 AOM/DSS實驗中， AA小鼠大腸發炎比WT嚴重，且AA小鼠大腸內adenomas的生長較WT嚴重。在cohoused AOM/DSS的實驗中，WT與AA小鼠發炎沒有差異；AA大腸組織的dysplasia較WT嚴重，但AA小鼠的adenoma卻占了較少的比例，表示AA Colorectal cancers (CRC)的進展大多停留在早期(dysplasia)階段。腸組織western blot與IHC的結果中， AA小鼠的大腸組織含有較多beta-catenin與beta-catenin下游基因(MMP9 and c-Myc)，有利於adenomas的生成。但AA也表達較高的apoptosis相關蛋白(caspase3 and Bax)。在in vitro的實驗中，給予TNF-α後，transfected AA-CBP plasmid的HCT116 (AA-HCT116)會大幅表現apoptosis相關蛋白(caspase3 and Bax)與p53下游基因(p21, Bax)，但AA-CBP抑制腫瘤生長的原因仍需進一步的探討。綜合以上結論，當小鼠的CBP Ser1383/1387無法被磷酸化，會提高發炎和Wnt/beta-catenin signaling，促進adenomas的生成；但阻礙adenomas進展的原因還有待研究。

Colorectal cancer (CRC) is the second leading cause of death globally, accounting for almost 9 hundred thousand deaths annually. It is associated with a 2.4-fold higher risk in patients with inflammatory bowel disease (IBD), especially ulcerative colitis patients. CREB-binding protein (CBP) belongs to the family of histone acetyltransferases regulating the accessibility of DNA for the transcription factors. Phosphorylation of human CBP by IKKa at Ser1382/1386 plays a critical role in the regulation of cell fate by suppressing p53-mediated gene expression. Our previous study demonstrated that CBP-AA-knockin mice(AA-mice) replacing CBP Ser1383/1387 with alanines exhibit a spontaneous colitis phenotype instigated by impaired phosphorylation of CBP in non-hematopoietic cells origin. In addition, AA mice caused exacerbated colon inflammation upon colitogenic chemical (DSS) induction. Whether initiation and progression of inflammation in AA mice drives colitis-associated cancer (CAC) was investigated by AOM/DSS model. Both WT and AA mice showed dysplasia and adenoma after AOM/DSS treatment, while two types of results were shown. The first one was higher extent of inflammation in AA mice with higher adenomas burden. The second one was no difference in the extent of inflammation between both mice. H E staining revealed AA-mice showed more dysplasia (early stage of adenoma) but fewer adenomas than WT-mice. Higher expression of Wnt/beta-catenin and its target genes in AA-mice was found in both non-polyps and polyps (adenoma), indicating the initiation of dysplasia by beta-catenin. HCT116 cells overexpressing WT CBP (WT-HCT116) and S1382A/S1386A mutant CBP (AA-HCT116) treatment with TNF-α showed more caspase3, Bax, and p21 in AA HCT116cells, indicating over activation of apoptosis and p53 signaling in AA-HCT116. The underlying mechanism will be discussed.