Oweira Hani, Lahdou Imad, Mehrle Stefan, Khajeh Elias, Nikbakhsh Rajan, Ghamarnejad Omid, Terness Peter, Reißfelder Christoph, Sadeghi Mahmoud, Ramouz Ali
Department of Surgery, Medical Faculty Mannheim, University of Heidelberg, 68167 Mannheim, Germany.
Department of Transplantation Immunology, University of Heidelberg, 69120 Heidelberg, Germany.
J Clin Med. 2022 Aug 16;11(16):4794. doi: 10.3390/jcm11164794.
There are two main enzymes that convert tryptophan (Trp) to kynurenine (Kyn): tryptophan-2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO). Kyn accumulation can promote immunosuppression in certain cancers. In this study, we investigated Trp degradation to Kyn by IDO and TDO in primary human hepatocytes (PHH) and tumoral HepG2 cells. To quantify Trp-degradation and Kyn-accumulation, using reversed-phase high-pressure liquid chromatography, the levels of Trp and Kyn were determined in the culture media of PHH and HepG2 cells. The role of IDO in Trp metabolism was investigated by activating IDO with IFN-γ and inhibiting IDO with 1-methyl-tryptophan (1-DL-MT). The role of TDO was investigated using one of two TDO inhibitors: 680C91 or LM10. Real-time PCR was used to measure TDO and IDO expression. Trp was degraded in both PHH and HepG2 cells, but degradation was higher in PHH cells. However, Kyn accumulation was higher in the supernatants of HepG2 cells. Stimulating IDO with IFN-γ did not significantly affect Trp degradation and Kyn accumulation, even though it strongly upregulated IDO expression. Inhibiting IDO with 1-DL-MT also had no effect on Trp degradation. In contrast, inhibiting TDO with 680C91 or LM10 significantly reduced Trp degradation. The expression of TDO but not of IDO correlated positively with Kyn accumulation in the HepG2 cell culture media. Furthermore, TDO degraded L-Trp but not D-Trp in HepG2 cells. Kyn is the main metabolite of Trp degradation by TDO in HepG2 cells. The accumulation of Kyn in HepG2 cells could be a key mechanism for tumor immune resistance. Two TDO inhibitors, 680C91 and LM10, could be useful in immunotherapy for liver cancers.
有两种主要的酶可将色氨酸（Trp）转化为犬尿氨酸（Kyn）：色氨酸-2,3-双加氧酶（TDO）和吲哚胺2,3-双加氧酶（IDO）。Kyn的积累可促进某些癌症中的免疫抑制。在本研究中，我们调查了原代人肝细胞（PHH）和肿瘤性HepG2细胞中IDO和TDO介导的Trp向Kyn的降解情况。为了量化Trp降解和Kyn积累，使用反相高压液相色谱法测定了PHH和HepG2细胞培养基中Trp和Kyn的水平。通过用γ干扰素激活IDO并用1-甲基色氨酸（1-DL-MT）抑制IDO来研究IDO在Trp代谢中的作用。使用两种TDO抑制剂之一680C91或LM10来研究TDO的作用。采用实时PCR检测TDO和IDO的表达。Trp在PHH和HepG2细胞中均有降解，但在PHH细胞中的降解更高。然而，HepG2细胞上清液中的Kyn积累更高。用γ干扰素刺激IDO对Trp降解和Kyn积累没有显著影响，尽管它强烈上调了IDO的表达。用1-DL-MT抑制IDO对Trp降解也没有影响。相反，用680C91或LM10抑制TDO可显著降低Trp降解。在HepG2细胞培养基中，TDO而非IDO的表达与Kyn积累呈正相关。此外，TDO在HepG2细胞中可降解L-Trp但不能降解D-Trp。Kyn是HepG2细胞中TDO介导的Trp降解的主要代谢产物。HepG2细胞中Kyn的积累可能是肿瘤免疫抵抗的关键机制。两种TDO抑制剂680C91和LM10可能对肝癌免疫治疗有用。
