Laboratorio de Biología Molecular e Histocompatibilidad, Hospital Privado Universitario de Córdoba , Córdoba, Argentina.
Instituto Universitario de Ciencias Biomédicas de Córdoba , Córdoba, Argentina.
Br J Biomed Sci. 2020 Jul;77(3):105-111. doi: 10.1080/09674845.2020.1730625. Epub 2020 Mar 13.
CAT25 (Tmononucleotide repeat of the Caspase 2 gene), is a promising DNA marker for detecting microsatellite instability (MSI) in colorectal cancer. CAT25 has the potential to be incorporated into the Bethesda panel, a commonly used panel of DNA microsatellites, or replace it in its entirety. We aimed to develop and validate a high-resolution melting-PCR (HRM-PCR) method for CAT25 instability detection in clinical samples.
The instability of CAT25, BAT25 (a poly(A) tract occurring in ) and BAT26 (a poly(A) tract localized in ) microsatellites were assessed in DNA from tumour and peripheral blood obtained from 110 patients with colorectal cancer using HRM-PCR and capillary electrophoresis. Immunohistochemistry (IHC) staining for MSH2, MSH6, MLH1, and PMS2 enzymes was performed on tumours with jigj MSI. Allelic size variation of CAT25 was analysed on peripheral blood DNA from 208 healthy volunteers.
The HRM-PCR for CAT25 was validated in clinical samples. CAT25 showed a tight range of 64-66 base pairs. Of 110 tumours, 11 had High MSI, later confirmed by IHC. CAT25 defines MSI alone as well as when used together with BAT25 and BAT26. CAT25 results provided 100% predictive values and p < 0.0001 to classify a tumour as having high MSI.
We developed and validated a new HRM-PCR assay to detect CAT25 instability. Our findings showed a limited allelic size variation of CAT25 and highlighted to CAT25 as a promising marker for MSI analysis.
CAT25（Caspase 2 基因的单核苷酸重复）是一种有前途的检测结直肠癌微卫星不稳定性（MSI）的 DNA 标志物。CAT25 有可能被纳入 Bethesda 面板，即一种常用的 DNA 微卫星面板，或完全替代它。我们旨在开发和验证一种用于临床样本中 CAT25 不稳定性检测的高分辨率熔解-PCR（HRM-PCR）方法。
采用 HRM-PCR 和毛细管电泳法，对 110 例结直肠癌患者肿瘤和外周血 DNA 中的 CAT25、BAT25（位于）和 BAT26（位于）微卫星不稳定性进行评估。对 MSI 结直肠癌肿瘤进行 MSH2、MSH6、MLH1 和 PMS2 酶免疫组化（IHC）染色。对 208 例健康志愿者的外周血 DNA 进行 CAT25 等位基因大小变异分析。
该 HRM-PCR 方法在临床样本中得到验证。CAT25 显示出 64-66 个碱基的紧密范围。在 110 个肿瘤中，有 11 个具有高 MSI，随后通过 IHC 得到证实。CAT25 单独定义为 MSI，也可与 BAT25 和 BAT26 一起使用。CAT25 结果提供了 100%的预测值（p<0.0001），用于将肿瘤分类为高 MSI。
我们开发并验证了一种新的 HRM-PCR 检测方法来检测 CAT25 不稳定性。我们的研究结果显示 CAT25 的等位基因大小变异有限，并强调 CAT25 是 MSI 分析的一个有前途的标志物。
