Pyrrolizidine alkaloids (PAs) are among the most common phytotoxins with emerging evidence to contaminate soil, water, and nearby plants. Humans are frequently exposed to PA-contaminated food products or PA-containing herbal remedies. The reactive metabolites of PAs rapidly alkylate DNA to form pyrrole–DNA adducts (PDAs) and seed a population of mutations that initiate tumorigenesis in experimental models. However, the clinical evidence of PA-induced initial DNA damage is absent. This study developed a liquid biopsy approach for detecting PDAs in blood to diagnose PA-induced genotoxicity. Blood DNA samples, extracted from PA-exposed rats, were reacted with silver nitrate to cleave the N-linkage between conjugated DNA bases and pyrrole. The released pyrrole was chemically derivatized to a stable product which was measured by liquid chromatography-tandem mass spectrometry. Using this method, PDAs were, for the first time, detected in blood samples from 18 patients with PA-induced liver injury. Notably, PDAs were detectable in patients’ blood samples collected one month after hospital admission, indicating adequate persistence of PDAs for use as a clinical biomarker of PA-induced genotoxicity. Our findings provide clinical indication of PA-induced genotoxicity, which warrants early detection of PDAs as an actionable approach for prevention of PA-associated carcinogenesis.

中文翻译：

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血吡咯-DNA 加合物定义了吡咯里西啶生物碱诱导的肝损伤患者的早期致瘤风险

吡咯里西啶生物碱 (PA) 是最常见的植物毒素之一，新出现的证据表明它会污染土壤、水和附近的植物。人类经常接触受 PA 污染的食品或含 PA 的草药。PA 的反应性代谢物迅速将 DNA 烷基化以形成吡咯-DNA 加合物 (PDA)，并在实验模型中引发一系列突变，从而引发肿瘤发生。然而，缺乏 PA 诱导的初始 DNA 损伤的临床证据。本研究开发了一种液体活检方法，用于检测血液中的 PDA，以诊断 PA 诱导的基因毒性。从暴露于 PA 的大鼠中提取的血液 DNA 样本与硝酸银反应以裂解N- 共轭 DNA 碱基与吡咯之间的连接。释放的吡咯化学衍生为稳定的产物，通过液相色谱-串联质谱法进行测量。使用这种方法，首次在 18 名 PA 引起的肝损伤患者的血液样本中检测到 PDA。值得注意的是，在入院一个月后收集的患者血液样本中可检测到 PDA，表明 PDA 具有足够的持久性，可用作 PA 诱导的基因毒性的临床生物标志物。我们的研究结果提供了 PA 诱导的基因毒性的临床迹象，这保证了早期检测 PDAs 作为预防 PA 相关致癌作用的可行方法。