Impact of heme iron from red meat on colorectal carcinogenesis and relevance of DNA repair

众所周知，饮食因素会增加患结直肠癌（CRC）的风险，结直肠癌是德国第三大常见的肿瘤实体。各种研究表明，血红素铁作为红肉的相关成分，在结直肠癌的发生中起着重要作用。然而，潜在的机制仅被部分理解，可能涉及a)与微生物生态失调相关的结肠隐窝增殖增加，b)结直肠中遗传毒性n -亚硝基化合物（NOC）的产生以及c)脂质过氧化的诱导。NOC是一种烷基化剂，它能产生致突变前DNA加合物，如o6 -甲基鸟嘌呤（O6-MeG）和o6 -羧甲基鸟嘌呤（O6-CMG）。反过来，脂质过氧化产物如丙二醛和羟基壬烯醛（HNE）引起突变前DNA损伤，如嘧啶嘌呤酮（M1G）和HNE- g加合物。这就提出了一个问题，即红肉中的血红素铁是否在结直肠癌中具有引发肿瘤和/或促进肿瘤的潜力，以及哪种DNA修复途径参与了防止血红素铁相关结直肠癌形成的保护。O6-MeG和O6-CMG是o6 -甲基鸟嘌呤- dna甲基转移酶（MGMT）直接逆转损伤的底物。M1G和HNE-G加合物进行核苷酸切除修复，主要涉及着色性干皮病补体组A （XPA）蛋白。在我们的小组中，有MGMT和XPA基因缺失的转基因小鼠模型，这将允许研究这些DNA加合物在血红素铁引发的结直肠癌发生中的因果作用。首先，将在野生型动物中分析膳食血红素铁的肿瘤启动和促肿瘤特性，并与铁平衡对照饮食进行比较。为此，将使用非侵入性微型内窥镜装置，通过定量方式监测肿瘤的形成和进展，并辅以整个结直肠的组织病理学分析。此外，血红素铁引发的结直肠癌形成的剂量-反应关系和DNA修复缺陷的影响将被评估。接下来，我们将分析血红素铁在结直肠组织中诱导的关键DNA损伤，以及其他终点，如细胞死亡诱导、细胞增殖和MGMT启动子甲基化。最后，我们将分析肠道炎症在血红素相关结直肠癌发生中的作用。

Dietary factors are well known to increase the risk to develop colorectal cancer (CRC), which is the third most common tumor entity in Germany. Various studies suggest that the intake of heme iron, as relevant constituent of red meat, plays an important role in colorectal carcinogenesis. However, the underlying mechanisms are only partially understood and may involve a) an increased proliferation of the colon crypts linked to microbial dysbiosis, b) the generation of genotoxic N-nitroso compounds (NOC) in the colorectum and c) the induction of lipid peroxidation. NOC are alkylating agents, which cause premutagenic DNA adducts such as O6-methylguanine (O6-MeG) and O6-carboxymethylguanine (O6-CMG). In turn, lipid peroxidation products such as malondialdehyde and hydroxynonenal (HNE) give rise to premutagenic DNA lesions such as pyrimidopurinone (M1G) and HNE-G adducts. This raises the issue whether heme iron from red meat exhibits tumor initiating and/or tumor promoting potential in the colorectum and which DNA repair pathway is involved in the protection against heme iron-associated CRC formation. O6-MeG and O6-CMG are substrates for direct damage reversal by O6-methylguanine-DNA methyltransferase (MGMT). M1G and HNE-G adducts undergo nucleotide excision repair, which essentially involves xeroderma pigmentosum complementation group A (XPA) protein. Transgenic mouse models with deficiencies in MGMT and XPA are available in our group, which will allow studying the causal role of these DNA adducts in heme iron-triggered colorectal carcinogenesis. First, the tumor initiating and tumor promoting properties of dietary heme iron will be analyzed in wildtype animals in comparison to an iron balanced control diet. To this end, a non-invasive mini-endoscopy unit will be used, by which tumor formation and progression is monitored over time in a quantitative manner, which is complemented by histopathological analysis of the whole colorectum. Furthermore, the dose-response relationship in heme iron-triggered CRC formation and the impact of DNA repair deficiencies will be assessed. Next, we will analyze critical heme iron-induced DNA damage in colorectal tissue as well as other endpoints such as cell death induction, cell proliferation and MGMT promoter methylation. Finally, the contribution of intestinal inflammation in heme-associated colorectal carcinogenesis will be analyzed.