在小鼠模式中探討結合中和HBV表面抗原的抗體和TLR7刺激劑兩種藥物對清除B型肝炎病毒的影響

Abstract

B型肝炎(Hepatitis B)是藉由B型肝炎病毒(Hepatitis B virus, HBV)感染肝臟引起的疾病，根據世界衛生組織估計有二億五千七百萬人有感染HBV，為全球性的共同議題。以目前的治療方式無法完全治療的原因為HBV的covalently closed circular DNA (cccDNA)無法根除以及表面抗原(HBV surface antigen, HBsAg)無法清除，所以需要新的HBV治療策略。而在感染HBV後，HBV可能藉由抑制宿主的先天性免疫反應，而不會誘發conventional innate immunity產生IFN, IL-1β, TNF等等的cytokine，以及interferon related gene的induction量很有限，在後天性免疫方面，因為高表現HBV的抗原使得T 細胞耗竭，失去了產生反應的功能，B細胞的反應也不足夠，所以未來的治療趨勢可能可以藉由改善先天性或後天性免疫的方式，達到治療效果。在之前有研究指出在餵食TLR7 agonist GS-9620給慢性感染HBV的動物後有達到很好的抑制HBV的功效，目前已知的功能為促使TLR7訊息傳遞路徑進行，產生type I interferon和cytokine，而這type I interferon會藉由receptor送至受HBV感染的肝細胞中，產生下游interferon stimulated gene。雖然在慢性感染的動物model中可以看到serum中的HBsAg和DNA表現量減少，以及肝臟中的DNA量減少，但是在慢性感染HBV的病人給予GS-9620後仍測得與未給藥相似的HBsAg含量。但是在另一篇報導指出GS-9620和會結合於SHIV表面抗原的抗體PGT121給予於感染SHIV的猴子中，可以看出和單獨給各自的藥比起來病毒會復發的猴子數量變少，並且所需要的復發時間會延長許多。而在實驗室先前與廈門大學合作中所使用的抗體E6F6使用於hydrodynamic injection HBV mice中可以有效的抑制HBsAg，但是也達不到清除的效果。因此假設E6F6將大量的HBsAg減少後且使T cell immune restoration，再藉由GS9620活化和NK cell的方式，而這活化的NK cell是否可以像在SHIV的模式中一樣，接上E6F6後，E6F6接上受HBV感染肝細胞表面的HBsAg，直接殺死受HBV感染肝細胞，增加受HBV感染肝細胞的清除率。結果顯示，每周測量血清中HBsAg的結果，在有給予E6F6的三個組別的小鼠都呈現低至個位數字的程度直至第十七周，而在GS-9620組和控制組都沒有看到這樣的變化，但是在第十七周後，反而在給E6F6加上GS-9620的兩組比單獨給E6F6來的先回升。相對的，在停藥後HBV DNA都看不到每組間有太多的差異。本研究說明了兩種藥物比單獨給藥來的先復發，希望未來可以了解這其間的機制，進而去改善 免疫反應。

Hepatitis B is a disease caused by the infection of liver by hepatitis B virus (HBV). It is estimated that 257 million people have HBV infections as a common global problem. The reasons for the current cure are not fully cured for the inability of cccDNA of HBV to be eradicated and for the inability of HBsAg to be cleared, so new HBV cure strategies are needed. After infecting HBV, it may inhibit the conventional innate immunity of the host. And in the adaptive immunity, T cells exhaustion and not enough function of the productive antibody response are result from high expression of HBsAg. so the uncoming cure may be effective by improving innate or adaptive immunity. There have been studies that point to an eclipse of TLR7 agonist GS-9620 has been able to suppress HBV in chronic HBV-infected animals, and now known functions are to promote TLR7, the production of type I interferon and cytokines. And then, type I Interferon is delivered by the receptor to the hepatocytes, which is infected with HBV, and produces a downstream interferon stimulated gene. Although the HBsAg and DNA in serum are reduced in chronically infected animal model, and the amount of DNA in the liver is low, the HBsAg titer is still not given similar to that of HBsAg after the patient who is chronically infected with HBV is given GS-9620. But in another paper, GS-9620 together with the antibody PGT121, which is bound to the SHIV surface antigen, are shown to be giving less monkeys than only giving GS-9620 or PGT121, and the days to viral rebound are longer. The antibody E6F6 can be effective in suppressing HBsAg in hydrodynamic injection HBV mice, but it also does not achieve the effect of clearance. Therefore, the hypothesis is E6F6 will deplete large amounts of HBsAg and leave T cell restoration, and then GS-9620 will activate NK cell just like in SHIV mode. NK cells is linked by E6F6 which is attached to HBsAg that is on the surface of hepatocytes. So NK cells can directly kill HBV-infected liver cells, and increase the rate of removal of HBV-infected hepatocytes. The results show that HBsAg in the weekly serum are shown to be low in three groups that have been given E6F6 until the seventeenth week, but not seen in GS-9620 group and control group. However, after the seventeenth week, the HBsAg titer of the E6F6 plus the GS-9620 groups are lifted even faster than of the only E6F6 group. In contrast to the HBsAg titer, HBV DNA did not see too much difference in each group after drug discontinuation. This study showed that HBsAg rebound in groups received E6F6 and GS-9620 combined are more faster than the E6F6 only. In the future, we hope to understand the mechanism in E6F6 and GS-9620 to regulate the immune response.