Background & Aims

Strategies to develop virus-specific T cells against hepatic viral infections have been hindered by safety concerns. We engineered nonlytic human T cells to suppress replication of hepatitis B virus (HBV) and hepatitis C virus (HCV) without overt hepatotoxicity and investigated their antiviral activity.

Methods

We electroporated resting T cells or T cells activated by anti-CD3 with mRNAs encoding HBV or HCV-specific T-cell receptors (TCRs) to create 2 populations of TCR-reprogrammed T cells. We tested their ability to suppress HBV or HCV replication without lysis in 2-dimensional and 3-dimensional cultures of HepG2.2.15 cells and HBV-infected HepG2-hNTCP cells. We also injected TCR-reprogrammed resting and activated T cells into HBV-infected urokinase-type plasminogen activator/severe combined immunodeficiency disease/interleukin 2γ mice with humanized livers and measured levels of intrahepatic and serological viral parameters and serum alanine aminotransferase. Livers were collected for analysis of gene expression patterns to determine effects of the TCR-reprogrammed T cells.

Results

TCR-reprogrammed resting T cells produced comparable levels of interferon gamma but lower levels of perforin and granzyme than activated T cells and did not lyse HCV- or HBV-infected hepatoma cells. Although T-cell secretion of interferon gamma was required to inhibit HCV replication, the HBV-specific TCR-reprogrammed resting T cells reduced HBV replication also through intracellular activation of apolipoprotein B mRNA editing enzyme, catalytic polypeptide 3 (APOBEC3). The mechanism of APOBEC3 intracellular activation involved temporal expression of lymphotoxin-β receptor ligands on resting T cells after TCR-mediated antigen recognition and activation of lymphotoxin-β receptor in infected cells.

Conclusions

We developed TCR-reprogrammed nonlytic T cells capable of activating APOBEC3 in hepatoma cells and in HBV-infected human hepatocytes in mice, limiting viral infection. These cells with limited hepatotoxicity might be developed for treatment of chronic HBV infection.

中文翻译：

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能够表达病毒特异性T细胞受体的非溶解淋巴细胞可通过激活APOBEC3限制HBV感染

背景与目标

安全问题阻碍了开发针对肝病毒感染的病毒特异性T细胞的策略。我们设计了非溶解性人类T细胞，以抑制乙型肝炎病毒（HBV）和丙型肝炎病毒（HCV）的复制而没有明显的肝毒性，并研究了它们的抗病毒活性。

方法

我们用编码HBV或HCV特异性T细胞受体（TCR）的mRNA电穿孔静息T细胞或被抗CD3激活的T细胞，以创建2个TCR重新编程的T细胞群体。我们测试了它们在不裂解HepG2.2.15细胞和HBV感染的HepG2-hNTCP细胞的2维和3维培养物中不裂解的情况下抑制HBV或HCV复制的能力。我们还将TCR重新编程的静息和活化T细胞注入人源化肝脏的HBV感染的尿激酶型纤溶酶原激活剂/重度联合免疫缺陷疾病/白介素2γ小鼠中，并测量了肝内和血清学病毒参数水平以及血清丙氨酸氨基转移酶。收集肝脏用于基因表达模式分析，以确定TCR重新编程的T细胞的作用。

结果

经TCR重新编程的静息T细胞产生的干扰素γ水平与活化的T细胞相当，但穿孔素和颗粒酶的水平却低于活化的T细胞，并且不裂解被HCV或HBV感染的肝癌细胞。尽管需要干扰素γ的T细胞分泌来抑制HCV复制，但HBV特异性TCR重编程的静止T细胞也通过载脂蛋白B mRNA编辑酶催化性多肽3（APOBEC3）的细胞内激活而降低了HBV复制。APOBEC3细胞内激活的机制涉及在TCR介导的抗原识别和感染细胞中淋巴毒素β受体的活化后，静止的T细胞上淋巴毒素β受体配体的瞬时表达。

结论

我们开发了能够激活小鼠肝癌细胞和HBV感染人类肝细胞中APOBEC3的TCR重新编程的非溶解性T细胞，可限制病毒感染。这些肝毒性有限的细胞可能被开发用于治疗慢性HBV感染。