"植化素抑制大腸直腸癌中indoleamine-2,3-dioxygenase活性之探討"

Abstract

癌細胞經常利用許多方式來抑制病人的免疫系統以躲避免疫系統的清除，其中一個免疫抑制的機制即是透過抑制indoleamine-2,3-dioxygenase (IDO)此酵素之活性。IDO會將tryptophan降解成kynurenine，而造成微環境中tryptophan缺乏的現象，又因tryptophan是T細胞的必需胺基酸，且其下游代謝產物kynurenines會誘導T細胞走向凋亡，因此是一種具抑制免疫能力的酵素。過去文獻指出，除腫瘤組織浸潤的dendritic cells可能表現較高的IDO活性外，癌細胞本身也會表現IDO，使得腫瘤組織逃過免疫系統的監控及毒殺。文獻指出大腸直腸癌病人中高IDO表現可能與低T細胞數量有關，不利於預後，因此本研究選擇目前已知具抗癌能力的植化素(phytochemical)，以大腸直腸癌細胞模式來篩選可能抑制大腸直腸癌中IDO活性的植化素，並探討其可能的作用機制。研究結果顯示，大腸直腸癌細胞株HCT116及LoVo皆因interferon-γ (IFN-γ)之誘導而表現IDO之mRNA，且可於細胞培養液中偵測到tryptophan的減少及kynurenine的增加，而LoVo還表現tryptophan-2,3-dioxygenase (TDO)此與IDO作用類似的酵素的mRNA。而人體息肉ex vivo的實驗中可以發現該樣本亦因IFN-γ之誘導而表現IDO之mRNA。第二部份使用此兩株細胞株針對9種植化素：Epigallocatechin gallate (EGCG)、curcumin、 resveratrol、quercetin、sulforaphane、6-shogaol、diallyl trisulfide (DATS)、brassinin及其化學衍生物5-br-brassinin，進行抑制IDO活性的實驗，結果發現EGCG、6-shogaol、brassinin、5-br-brassinin、sulforaphane、DATS可顯著使細胞培養液中kynurenine減少。第三部份探討EGCG及DATS之抑制機制，實驗結果顯示EGCG具抑制IDO及TDO mRNA表現的活性，然而DATS並不具抑制IDO或TDO mRNA的效果，因此DATS產生抑制活性之機制仍待深入探討。

Tumor cells display multiple immunosuppressive mechanisms against the host immune system. One of the tumor immunosuppression mechanisms involves the immunoregulatory enzyme indoleamine-2,3-dioxygenase (IDO). IDO degrades tryptophan which results tryptophan starvation of local environment. Tryptophan starvation and the proapoptotic tryptophan catabolites, kynurenine, consequently inhibit T cell response and proliferation. Recent studies indicated that many tumor cells constantly express IDO which enable tumor cells to avoid immune attack of T cells. In several colorectal cancer case studies, it has been proposed that high-IDO expression was associated with significantly low T cells, which may correlate to poor prognosis and liver metastasis. Phytochemicals are chemical compounds that occur naturally in plants and have been reported to have antitumor activity. According to recent studies, some of the phytochemicals have ability to inhibit IDO activity in cancer cells. The aim of this research was to use colonrectal cancer cell lines to identify the phytochemicals which have anti-IDO effects on colorectal cancer. We first prepared two colorectal cancer cell lines, LoVo and HCT116, and used them as IDO-expressing cell models. We found that these two cell lines express IDO mRNA by the induction of interferon-γ (IFN-γ). In addition, IFN-γ decreased tryptophan and increased kynurenine concentratins in the culture medium.We further found that LoVo cells expressed tryptophan-2,3-dioxygenase (TDO) mRNA. In our human colon polyp ex vivo study, it showed that IDO mRNA was also induced by IFN-γ stimulation. In the second part of our research, we tested 9 phytochemicals including EGCG, curcumin, resveratrol, quercetin, sulforaphane, 6-shogaol, diallyl trisulfide (DATS), brassinin andits derivate 5-br-brassinin, to see if they were able to inhibit colorectal cancer cells to convert tryptophan to kynurenine. Results showed that EGCG, 6-shogaol, brassinin, 5-br-brassinin, sulforaphane, DATS were able to inhibit both LoVo and HCT116 cells to release kynurenine into cell medium. In the third part, we investigated the inhibition mechanism of EGCG and DATS. Results indicated that EGCG showed IDO and TDO mRNA inhibiting activity. However, DATS did not show inhibition effect on IDO or TDO gene expression. The mechanism of how DATS inhibits IDO activity in colorectal cancer cells is still unclear and needs further study.