Experimental and Molecular Pathology, Institute of Pathology, Ludwig-Maximilians-University Munich, Munich, Germany.
Institute of Pathology, Ludwig-Maximilians-University Munich, Munich, Germany; Institute of Pathology, Charité-Universitätsmedizin Berlin, Berlin, Germany; German Cancer Consortium, Heidelberg, Germany; German Cancer Research Center, Heidelberg, Germany.
Gastroenterology. 2018 Dec;155(6):1868-1882. doi: 10.1053/j.gastro.2018.08.011. Epub 2018 Aug 10.
BACKGROUND & AIMS: Combined inactivation of the microRNA 34a gene (MIR34A, by methylation) and the TP53 gene (by mutation or deletion) is observed in 50% of colorectal tumors that progress to distant metastases. We studied mice with intestinal disruption of Mir34a and Tp53 to investigate mechanisms of colorectal carcinogenesis and identify strategies to block these processes.
Mice with disruption of Mir34a and/or Tp53 specifically in intestinal epithelial cells (IECs) (Mir34a mice, Tp53 mice, and Mir34a/Tp53 mice) and controls (Mir34a/Tp53) were given azoxymethane to induce colorectal carcinogenesis. Some mice were given intraperitoneal injections of an antibody against mouse interleukin 6 receptor (IL6R), or received an inhibitor of PAI1 (tiplaxtinin) in their chow. Intestinal tissues were collected and analyzed by immunohistochemistry; gene expression profiles were analyzed by RNA sequencing. We determined the expression and localization of PAI1 in 61 human primary colon cancers and compared them to MIR34A methylation and inactivating mutations in TP53. Data on mRNA levels, methylation, and clinical features of 628 colon and rectal adenocarcinomas were obtained from The Cancer Genome Atlas portal.
Mir34a/Tp53 mice developed larger and more colorectal tumors, with increased invasion of surrounding tissue and metastasis to lymph nodes, than control mice or mice with disruption of either gene alone. Cells in tumors from the Mir34a/Tp53 mice had decreased apoptosis and increased proliferation compared to tumor cells from control mice, and expressed higher levels of genes, that regulate inflammation (including Il6r and Stat3) and epithelial-mesenchymal transition. The gene expression pattern of the tumors from Mir34a/Tp53 mice was similar to that of human colorectal tumor consensus molecular subtype 4 (mesenchymal, invasive). We identified the Pai1 messenger RNA as a target of Mir34a; levels of PAI1 protein were increased in primary colon cancer samples, that displayed methylation of MIR34A and mutational inactivation of TP53. Administration of tiplaxtinin or anti-IL6R antibody to Mir34a/Tp53 mice decreased proliferation of cancer cells, and reduced colorectal tumor invasion and metastasis.
In mice, we demonstrated that combined inactivation of Mir34a and Tp53 promotes azoxymethane-induced colorectal carcinogenesis and tumor progression and metastasis by increasing levels of IL6R and PAI1. Strategies to inhibit these processes might be developed to slow progression of colorectal cancer.
在进展为远处转移的 50%结直肠肿瘤中观察到 microRNA 34a 基因（MIR34A，通过甲基化）和 TP53 基因（通过突变或缺失）的联合失活。我们研究了肠道 Mir34a 和 Tp53 破坏的小鼠，以研究结直肠癌变的机制，并确定阻断这些过程的策略。
Mir34a 和/或 Tp53 特异性在肠上皮细胞（IEC）中破坏的小鼠（Mir34a 小鼠、Tp53 小鼠和 Mir34a/Tp53 小鼠）和对照小鼠（Mir34a/Tp53）接受氧化偶氮甲烷诱导结直肠癌变。一些小鼠接受抗小鼠白细胞介素 6 受体（IL6R）的腹腔注射，或在饲料中接受 PAI1 抑制剂（tiplaxtinin）。通过免疫组织化学收集和分析肠组织；通过 RNA 测序分析基因表达谱。我们在 61 个人类原发性结肠癌中确定了 PAI1 的表达和定位，并将其与 MIR34A 甲基化和 TP53 失活突变进行了比较。从癌症基因组图谱门户获得了 628 例结肠和直肠腺癌的 mRNA 水平、甲基化和临床特征数据。
Mir34a/Tp53 小鼠比对照小鼠或单独破坏任一基因的小鼠发展出更大和更多的结直肠肿瘤，并且周围组织的侵袭和淋巴结转移增加。与对照小鼠的肿瘤细胞相比，Mir34a/Tp53 小鼠的肿瘤细胞凋亡减少，增殖增加，并且表达更高水平的调节炎症（包括 Il6r 和 Stat3）和上皮-间充质转化的基因。Mir34a/Tp53 小鼠肿瘤的基因表达模式与人类结直肠癌共识分子亚型 4（间质，侵袭）相似。我们确定了 Pai1 信使 RNA 是 Mir34a 的靶标；在显示 MIR34A 甲基化和 TP53 失活突变的原发性结肠癌样本中，PAI1 蛋白水平升高。给予 Mir34a/Tp53 小鼠 tiplaxtinin 或抗 IL6R 抗体可减少癌细胞的增殖，并降低结直肠肿瘤的侵袭和转移。
在小鼠中，我们证明 Mir34a 和 Tp53 的联合失活通过增加 IL6R 和 PAI1 的水平促进氧化偶氮甲烷诱导的结直肠癌发生和肿瘤进展和转移。抑制这些过程的策略可能会被开发出来以减缓结直肠癌的进展。
