长链非编码 RNA ZNF649-AS1 通过调控 EXOC7 的可变剪接促进乳腺癌对曲妥珠单抗的耐药性和 TAM 依赖性 PD-L1 的表达。

LncRNA ZNF649-AS1 promotes trastuzumab resistance and TAM-dependent PD-L1 expression in breast cancer by regulating EXOC7 alternative splicing.

机构信息

Department of General Surgery, Hainan General Hospital, Hainan Affiliated Hospital of Hainan Medical University, Haikou, 570311, Hainan Province, PR China.

Department of Breast Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, Henan Province, PR China.

出版信息

Arch Biochem Biophys. 2024 Nov;761:110128. doi: 10.1016/j.abb.2024.110128. Epub 2024 Aug 17.

DOI:10.1016/j.abb.2024.110128

PMID:39159899

Abstract

BACKGROUND

Trastuzumab resistance is a serious clinical problem in the treatment of HER2-positive breast cancer (BC). The lncRNA ZNF649-AS1 was previously found to promote HER2-positive BC trastuzumab resistance. The study aims to explore the molecular mechanism of ZNF649-AS1 in HER2-positive BC trastuzumab resistance.

METHODS

Tumor tissue and peripheral blood samples were collected from 20 HER2-positive BC patients with trastuzumab-resistant and non-resistant, respectively. Trastuzumab-resistant BC cell lines SKBR-3-TR and BT474-TR were established. RIP was employed to confirm the binding of ZNF649-AS1, PRPF8 and exocyst complex component 7 (EXOC7). RNA expression of EXOC7-L (Full length of EXOC7) and EXOC7-S (Spliceosome of EXOC7) were detected using agarose gel electrophoresis. Expressions of macrophage markers CD68 CD206 were measured by flow cytometry.

RESULTS

ZNF649-AS1 expression was upregulated in HER2-positive BC trastuzumab resistance. ZNF649-AS1 downregulation inhibited trastuzumab resistance in HER2-positive BC. ZNF649-AS1 regulated EXOC7 alternative splicing by binding with PRPF8. EXOC7-S knockdown suppressed trastuzumab resistance and TAM-dependent PD-L1 expression in HER2-positive BC. EXOC7-S overexpression abolished the effects of ZNF649-AS1 knockdown on trastuzumab resistance and TAM-dependent PD-L1 expression in HER2-positive BC.

CONCLUSION

ZNF649-AS1 promoted trastuzumab resistance and TAM-dependent PD-L1 expression in HER2-positive BC via promoting alternative splicing of EXOC7 by PRPF8.

摘要

背景

曲妥珠单抗耐药是 HER2 阳性乳腺癌（BC）治疗中的一个严重临床问题。先前发现长链非编码 RNA ZNF649-AS1 可促进 HER2 阳性 BC 曲妥珠单抗耐药。本研究旨在探讨 ZNF649-AS1 在 HER2 阳性 BC 曲妥珠单抗耐药中的分子机制。

方法

分别收集 20 例曲妥珠单抗耐药和非耐药的 HER2 阳性 BC 患者的肿瘤组织和外周血样本。建立曲妥珠单抗耐药 BC 细胞系 SKBR-3-TR 和 BT474-TR。采用 RIP 实验证实 ZNF649-AS1 与 PRPF8 和外泌体复合物成分 7（EXOC7）的结合。琼脂糖凝胶电泳检测 EXOC7-L（EXOC7 的全长）和 EXOC7-S（EXOC7 的剪接体）的 RNA 表达。采用流式细胞术检测巨噬细胞标志物 CD68、CD206 的表达。

结果

ZNF649-AS1 在 HER2 阳性 BC 曲妥珠单抗耐药中表达上调。下调 ZNF649-AS1 抑制了 HER2 阳性 BC 的曲妥珠单抗耐药。ZNF649-AS1 通过与 PRPF8 结合调节 EXOC7 的可变剪接。EXOC7-S 敲低抑制了 HER2 阳性 BC 中的曲妥珠单抗耐药和 TAM 依赖性 PD-L1 表达。EXOC7-S 过表达消除了 ZNF649-AS1 敲低对 HER2 阳性 BC 中曲妥珠单抗耐药和 TAM 依赖性 PD-L1 表达的影响。