|Clinical characteristics of patients and healthy subjects|
|Healthy control||ESCC patients||mESCC patients|
|No. of subjects||28||38||38|
Tryptophan；3-Hydroxyanthranilic acid；Kynurenine；Kynurenic acid；5-Hydroxytryptophan；5-Hydroxyindole-3-acetic acid；Tryptamine；N-acetylserotonin；5-Hydroxytryptamine.
后期，最终选择其中五个代谢物 （即色氨酸 (TRP)、 犬尿氨酸 （KYN）、 5-羟基色氨酸 (5HTP)、 5-羟基吲哚-3-乙酸 (5HIAA)、 5-羟色胺 (5HT)） 进行定量比较，健康人群、ESCC患者、mESCC患者血样中这五种代谢物的平均含量如下图：
通过热图可以看到，相对于正常人群样本来说，ESCC和 mESCC 患者的色氨酸（TRP）含量下调了，而5-羟色胺 (5HT)上调，而其他三个代谢物则出现动态变化。为了阐明这些差异，研究团队奖下游代谢产物 （5HT、 5HTP、 5HIAA和KYN） 与上游的前体 (TRP) 的计算比率，进过数据预处理之后，研究团队使用其他四种代谢物与色氨酸的比值作为指标进行分析。相对于正常人群样本，研究团队发现这些比值，ESCC和 mESCC 患者的这些比值都呈现了上调的趋势。再以此建立两种诊断模型。研究团队发现这两种模式能有效的预测和诊断ESCC和 mESCC。
In mammals, tryptophan metabolism is a physiological means of preserving immune homeostasis, which avoids acute and chronic hyper-inﬂammatory reactions. Recently, some studies report that immune homeostasis is important in cancer progression and the disturbed tryptophan metabolism is correlated to tumorigenesis and metastasis. In this study, a targeted metabolomics approach was conducted to explore variations of tryptophan metabolism in ESCC patients.
In procedure of tryptophan metabolism, tryptophan can be degraded in different pathways , in which some physiologically important metabolites are produced, including 5HTP, 5HT, TRA and KYN: (1)TRP is catalyzed by the 5-monooxygenase and converted in o 5HTP, which is subsequently converted in o 5HT through decarboxylation reaction, and 5HT is converted in o 5HIAA ﬁnally; (2)TRP can be degraded in o KYN through the kynurenine pathway, and KYN can be transformed to 3HAA and KYA; (3) TRP can be transformed to TRA by the aromatic-L-amino-acid decarboxylase.
TRP is the precursor of 5HTP, 5HT, and 5HIAA. In this study, concentration of TRP was decreased signiﬁcantly both in ESCC and mESCC patients, which was in consistent with some previous cancer studies. Decreased tryptophan leads to biostatic starvation and thus limits lymphocyte expansion, which has a close correlation with tumorigenesis. In this study, serum 5HTP was down-regulated in ESCC patients due to decreased concentration of TRP and enhanced activity of aromatic-L-aminoacid decarboxylase. 5HT, degraded from the 5HTP by decarboxylation reaction, was upregulated both in non-metastatic and metastatic patients. It has been reported that 5HT is a mitogenic factor and high concentration of 5HT promotes progress of caners. On the other hand, TRP can be converted in o N-formylkynurenine and then hydrolyzed to KYN through the kynurenine pathway, in which the downstream metabolites, KYA and 3HAA, are produced.
The ratio of KYN to TRP is a widely accepted indicator presenting the enzyme activity involved in the kynurenine pathway. Variation of this ratio in health controls, ESCC, and mESCC patients was showed in. We found that the ratios of KYN/TRP, 5HTP/ TRP, 5HIAA/TRP and 5HT/TRP were increased when compared the ESCC and mESCC patients to healthy controls. Moreover, the increased trends of these ratios of metabolite to tryptophan were in accordance with malignant transformation of ESCC. These results implied that there was a higher enzyme activity in ESCC and mESCC patients compared to healthy controls. The higher enzyme activity has been observed in several types of human solid tumors, including colorectal, breast, ovarian, lung cancers and melanoma. It is worth noting that high enzyme activity of kynurenine pathway is associated with immune escape, which is a trigger factor of tumor progression and migration. The incensement of KYN can inhibit T-cell proliferation and induce T-cell apoptosis, which leads to immune tolerance, thus providing a microenvironment for tumor progression and migration.
在色氨酸代谢的过程中，色氨酸可以在不同的路径被降解，其中会产生一些具有重要生理学作用的代谢物，包括 5HTP，5HT，TRA和KYN: (1) TRP是由 5-单加氧酶催化和转换成 5HTP，随后通过脱羧反应被转化为 5HT，最终5HT 转换成 5HIAA;(2)TRP可以通过犬尿氨酸通路，降解成KYN，KYN可以再转化为 3HAA 和KYA;（3） TRP可以被芳香-L-氨基酸脱羧酶转化成 TRA .
TRP是 5HTP、 5HT 和 5HIAA 的先导。在此研究中，在ESCC和 mESCC 患者中，色氨酸浓度显著下降，符合一些以前癌症研究。色氨酸降低可以饿死肿瘤细胞从而限制了淋巴细胞扩张，这与肿瘤转移密切相关。在本研究中， ESCC患者血清中5HTP 下调是由于色氨酸浓度降低，芳香-L-氨基酸脱羧酶的活性增强。5HTP通过脱羧反应降解成5HT，而5HT在非转移和远处转移的患者中表达上调。据报 5HT 是一种有丝分裂因子，高浓度的 5HT 促进了癌症的发展。另一方面，TRP可以转化为 N-甲酰犬尿氨酸，然后通过犬尿氨酸途径，水解成KYN，再产生的下游的代谢产物KYA和 3HAA.
对于呈现酶参与的犬尿氨酸途径，KYN/TRP的值是一个被广泛接受的指标。这一比率在健康组、 ESCC和 mESCC 患者的变化如图 3 。我们发现，与健康对照组相比，在ESCC和 mESCC 患者中，KYN/TRP，5HTP / TRP，5HIAA/TRP和 5HT/TRP的比例都增加了。此外，这些色氨酸代谢产物的比值的增加的趋势与ESCC恶性转化吻合。这些结果暗示ESCC和 mESCC 患者与健康对照组相比，具有更高的酶活性。包括结直肠癌、 乳腺癌、 卵巢癌、 肺癌和黑色素瘤，过几种类型的人类肿瘤患者中都观测到较高的酶活性。值得注意的是犬尿氨酸途径的高酶活性与免疫逃逸有关，免疫逃逸是肿瘤的生长和迁移触发因素。KYN的刺激可以抑制 T 细胞增殖并诱导 T 细胞凋亡，从而导致免疫耐受，从而为肿瘤的生长和迁移提供一个微环境。
Title:Disturbed tryptophan metabolism correlating to progression and metastasis of esophageal squamous cell carcinoma.
Author：Jing Cheng , Hai Jin , Xiaobei Hou , Jie Lv , Xianfu Gao *, Guangyong Zheng ** .
Journal: Biochemical and Biophysical Research Communications.
Keywords: Tryptophan metabolism,Esophageal squamous cell carcinoma Lymph node metastasis,Liquid chromatography Tandem mass spectrometry.
Esophageal squamous cell carcinoma (ESCC) is one of the most frequent malignancies worldwide. Lymph node metastasis is the leading cause of death in ESCC patients. To identify early diagnostic and prognostic biomarkers of ESCC and elucidate underlying pathogenesis of the disease, a targeted metabolomics strategy based on liquid chromatography combined with tandem mass spectrometry was applied to explore tryptophan metabolism between ESCC patients, metastatic ESCC patients (mESCC), and healthy controls. Statistical analysis on metabolite expression abundance and compound concentration ratio was conducted to discriminate patients from healthy controls. The concentration ratio of kynurenine, 5-hydroxytryptophan, 5-hydroxyindole-3-acetic acid, 5-hydroxytryptamine to their precursor tryptophan were identiﬁed as potential biomarkers, presenting high diagnostic capacity for distinguishing ESCC and mESCC patients from healthy controls. Moreover, a prognostic prediction model was also built on these ratios to distinguish metastasis patients from non-metastasis patients successfully. The high performance of ESCC prediction models suggest that concentration ratios of compounds may be used as biomarkers for early diagnosis and prognosis of the disease. In addition, concentration ratios of compounds show a progressively increased trend from non-metastasis to metastasis patients compared with healthy controls, which is in accordance with process of malignant transformation of ESCC. This interested ﬁnding suggests that disturbed tryptophan metabolism is correlated to progression and metastasis of ESCC since concentration ratios of compounds reﬂect activity of enzymes involved in tryptophan metabolism. This study reveals the impact of tryptophan metabolism to tumorigenesis and metastasis of ESCC, which help biologists investigate mechanism of the disease.