乙型肝炎病毒(hepatitis B virus，HBV)的包膜包含大(large，L)、中(middle，M)、小(small，S)包膜蛋白(envelope protein)，其中S蛋白含量最高。当前慢性乙型肝炎的治疗目标是功能性治愈，其关键指标是外周血中的乙肝表面抗原(hepatitis B surface antigen, HBsAg)持续清除。但目前HBsAg降解的机制尚未明确。本研究在前期工作中发现，靶向HBV包膜蛋白的嵌合抗原受体(chimeric antigen receptor, CAR)G12-CAR可抑制HBsAg的分泌，并促进其在胞内降解。为阐明这一机制，本研究采用多种表达载体转染Huh-7人肝癌细胞表达S蛋白，再用巴佛洛霉素A1(bafilomycin A1)和硼替佐米(bortezomib)分别抑制细胞内的溶酶体自噬和蛋白酶体降解途径，并通过蛋白免疫印迹和酶联免疫吸附试验检测细胞内和细胞外的S蛋白。结果显示，S蛋白可通过溶酶体自噬和蛋白酶体途径降解；L蛋白可干扰S蛋白在细胞内的降解；编码S蛋白的mRNA的3′-非翻译区(3′-untranslated region, 3′-UTR)序列也影响了S蛋白在细胞内的降解。以上研究结果为研制以蛋白降解相关分子为靶点的抗乙肝病毒药物提供了一定的依据。

The envelope proteins of hepatitis B virus (HBV) include large (L), middle (M) and small (S) envelope proteins, with S protein being the most abundant viral transmembrane protein. Functional cure is currently regarded as the therapeutic goal for chronic hepatitis B. One of the key indicators of functional cure is the sustained clearance of circulating hepatitis B surface antigen (HBsAg). However, the mechanisms whereby intracellular S protein is degraded remain elusive. The previous study of this research group demonstrated that G12-CAR, a chimeric antigen receptor (CAR) specifically targeting hepatitis B virus envelope proteins, inhibited S protein secretion and promoted its intracellular degradation. In this study, Huh-7 hepatoma cells transfected by different vectors expressing S protein were treated with bafilomycin A1 and bortezomib, respectively inhibiting lysosomal autophagy and proteasomal degradation pathways. Subsequently, intracellular and extracellular S proteins were detected by Western blot and enzyme-linked immunosorbent assay (ELISA). The results showed that S protein can be degraded via both lysosomal and proteasomal pathways, while other HBV components also can regulate S degradation. Both L protein and 3′-untranslated region (3′-UTR) sequences of the viral S mRNA were identified to affect intracellular S degradation. Hence, these findings may offer novel therapeutic targets for promoting HBsAg clearance.