Kortüm Benedikt, Campregher Christoph, Lang Michaela, Khare Vineeta, Pinter Matthias, Evstatiev Rayko, Schmid Gerald, Mittlböck Martina, Scharl Theresa, Kucherlapati Melanie H, Edelmann Winfried, Gasche Christoph
Christian Doppler Laboratory for Molecular Cancer Chemoprevention, Medical University of Vienna, Vienna, Austria.
Division of Gastroenterology and Hepatology, Department of Medicine 3, Medical University of Vienna, Vienna, Austria.
Gut. 2015 Dec;64(12):1905-12. doi: 10.1136/gutjnl-2014-307663. Epub 2014 Nov 26.
Lynch syndrome is caused by germline mutations in DNA mismatch repair genes leading to microsatellite instability (MSI) and colorectal cancer. Mesalazine, commonly used for the treatment of UC, reduces MSI in vitro. Here, we tested natural compounds for such activity and applied mesalazine and thymoquinone in a Msh2(loxP/loxP) Villin-Cre mouse model for Lynch syndrome.
Flow cytometry was used for quantitation of mutation rates at a CA13 microsatellite in human colon cancer (HCT116) cells that had been stably transfected with pIREShyg2-enhanced green fluorescent protein/CA13, a reporter for frameshift mutations. Mice were treated for 43 weeks with mesalazine, thymoquinone or control chow. Intestines were analysed for tumour incidence, tumour multiplicity and size. MSI testing was performed from microdissected normal intestinal or tumour tissue, compared with mouse tails and quantified by the number of mutations per marker (NMPM).
Besides mesalazine, thymoquinone significantly improved replication fidelity at 1.25 and 2.5 µM in HCT116 cells. In Msh2(loxP/loxP) Villin-Cre mice, tumour incidence was reduced by mesalazine from 94% to 69% (p=0.04) and to 56% (p=0.003) by thymoquinone. The mean number of tumours was reduced from 3.1 to 1.4 by mesalazine (p=0.004) and to 1.1 by thymoquinone (p<0.001). Interestingly, MSI was reduced in normal intestinal tissue from 1.5 to 1.2 NMPM (p=0.006) and to 1.1 NMPM (p=0.01) by mesalazine and thymoquinone, respectively. Thymoquinone, but not mesalazine, reduced MSI in tumours.
Mesalazine and thymoquinone reduce tumour incidence and multiplicity in Msh2(loxP/loxP) Villin-Cre mice by reduction of MSI independent of a functional mismatch repair system. Both substances are candidate compounds for chemoprevention in Lynch syndrome mutation carriers.
林奇综合征由DNA错配修复基因的种系突变引起，导致微卫星不稳定性（MSI）和结直肠癌。常用于治疗溃疡性结肠炎的美沙拉嗪可在体外降低MSI。在此，我们测试了天然化合物的这种活性，并在Msh2(loxP/loxP)Villin-Cre林奇综合征小鼠模型中应用了美沙拉嗪和百里醌。
使用流式细胞术对稳定转染了pIREShyg2-增强型绿色荧光蛋白/CA13（一种移码突变报告基因）的人结肠癌细胞（HCT116）中CA13微卫星处的突变率进行定量。用美沙拉嗪、百里醌或对照饲料对小鼠治疗43周。分析肠道的肿瘤发生率、肿瘤多发性和大小。从显微切割的正常肠道或肿瘤组织进行MSI检测，与小鼠尾巴进行比较，并通过每个标记的突变数（NMPM）进行定量。
除美沙拉嗪外，百里醌在1.25和2.5µM时可显著提高HCT116细胞的复制保真度。在Msh2(loxP/loxP)Villin-Cre小鼠中，美沙拉嗪使肿瘤发生率从94%降至69%（p=0.04），百里醌使其降至56%（p=0.003）。美沙拉嗪使肿瘤平均数量从3.1降至1.4（p=0.004），百里醌使其降至1.1（p<0.001）。有趣的是，美沙拉嗪和百里醌分别使正常肠道组织中的MSI从1.5 NMPM降至1.2 NMPM（p=0.006）和1.1 NMPM（p=0.01）。百里醌而非美沙拉嗪降低了肿瘤中的MSI。
美沙拉嗪和百里醌通过降低MSI降低了Msh2(loxP/loxP)Villin-Cre小鼠的肿瘤发生率和多发性，且不依赖于功能性错配修复系统。这两种物质都是林奇综合征突变携带者化学预防的候选化合物。
