探討天然物Piperlongumine對TNF-α誘導HeLa細胞的發炎反應中NF-κB p65的影響

Abstract

天然物是一類從生物體內製造並且被純化出來的小分子。許多從中草藥萃取出來的天然物，都有抗發炎和抗癌的功效，而且從結構上看，其中有許多可能是共價藥物。天然物piperlongumine是從胡椒屬的植物長胡椒(Piper longum)中，所萃取出來的一種小分子。目前研究發現，piperlongumine可以殺死某些癌細胞，但是不影響正常細胞；除此之外，還有抑制血小板活化的功效。另外，長胡椒的萃取物可以抗發炎，而piperlongumine也是萃取物的成分之一；但它在抗發炎的機制依舊不明。NF-κB是一種轉錄因子，可活化許多和細胞生長相關的基因表現，包括使白血球分泌細胞激素去促進細胞增生，因此NF-κB為影響細胞生長的關鍵調控者。在NF-κB過度活化的情況下，會造成慢性發炎，進而造成其他疾病，包括提高癌症風險。Piperlongumine從結構上分析，我們預測它可能是一種共價藥物。藉由麥可加成反應的方式，piperlongumine可以被半胱胺酸殘基攻擊可進行麥可加成反應的位置，進而和蛋白質形成共價鍵，對蛋白質進行修飾。我們的研究，以新的方法學：抗piperlongumine多株抗體的方式，發現以piperlongumine處理細胞之後，NF-κB會被piperlongumine所修飾。在子宮頸癌細胞，以腫瘤壞死因子-α (tumor necrosis factor-α) 去誘導NF-κB的活化的模式中，我們發現先以piperlongumine處理的細胞，NF-κB從細胞質到細胞核的移動會被抑制。因此，我們推測NF-κB上的某些半胱胺酸殘基被piperlongumine修飾後，可能會影響NF-κB移動至細胞核和NF-κB的活化；甚至有其他piperlongumine的目標蛋白也可能一起影響NF-κB的活化。

Natural compounds are small molecules extracted from organisms. Many natural compounds from herbals have the properties of anti-inflammation and anti-cancer and they may be covalent drugs according to their structures. The natural compound, piperlongumine, is a small molecule from the Piper species plant, Piper longum. Piperlongumine has been shown to kill certain cancer cells but not the normal cells; furthermore, piperlongumine has the activities for anti-platelet. The extracts of Piper longum also have the ability for anti-inflammation, and piperlongumine is a component of the extracts. However, the anti-inflammatory mechanism of piperlongumine is still unknown. NF-κB is a transcription factor that can activate many genes required for the cell proliferation including inducing secretion of interleukins from leukocytes to increase the activities of cell growth. Therefore, NF-κB is a key regulator of the cell proliferation. The NF-κB hyper-activation can lead to chronic inflammation and diseases such as the cancers. We analyzed the structure of piperlongumine and predicted that piperlongumine may be a covalent drug. Piperlongumine can be attacked by the cysteine residues of some proteins through Michael addition to form the covalent bonds. We used a new methodology to generate anti-piperlongumine polyclonal antibody for target identification, and found that the NF-κB can be modified by piperlongumine after piperlongumine treatment in the cells. In the TNF-α (tumor necrosis factor-α) induced NF-κB activation model, we found that the NF-κB translocation from the cytoplasm to the nucleus can be inhibited in cervical cancer cells. Therefore, we predicted that modifications of some cysteine residues on NF-κB can affect the NF-κB translocation and activation.