胰腺癌吉西他滨化疗耐药生物标志物—INPP4B的探索性研究

任加强; 武帅; 苏童; 李杰; 韩亮; 仵正

doi:10.19401/j.cnki.1007-3639.2024.12.003

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胰腺癌吉西他滨化疗耐药生物标志物—INPP4B的探索性研究

论著 | 更新时间：2025-12-31

- 胰腺癌吉西他滨化疗耐药生物标志物—INPP4B的探索性研究
- An exploratory study of INPP4B, a biomarker of gemcitabine chemoresistance in pancreatic cancer
- 中国癌症杂志 2024年34卷第12期页码：1090-1099
- 作者机构：
  
  1. 西安交通大学第一附属医院肝胆外科，陕西西安 710061
  2. 西安交通大学医学部公共卫生学院，陕西西安 710061
- 作者简介：
  
  [ "任加强（ORCID: 0000-0002-4957-7789），博士研究生" ]
  仵正（ORCID: 0000-0002-2441-0229），教授，主任医师，博士生导师。
- 基金信息：
- DOI：10.19401/j.cnki.1007-3639.2024.12.003
  中图分类号：
- 收稿：2024-08-30，
  
  纸质出版：2024-12-30
- 稿件说明：
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任加强, 武帅, 苏童, 等. 胰腺癌吉西他滨化疗耐药生物标志物—INPP4B的探索性研究[J]. 中国癌症杂志, 2024,34(12):1090-1099.

Jiaqiang REN, Shuai WU, Tong SU, et al. An exploratory study of INPP4B, a biomarker of gemcitabine chemoresistance in pancreatic cancer[J]. China Oncology, 2024, 34(12): 1090-1099.
任加强, 武帅, 苏童, 等. 胰腺癌吉西他滨化疗耐药生物标志物—INPP4B的探索性研究[J]. 中国癌症杂志, 2024,34(12):1090-1099. DOI： 10.19401/j.cnki.1007-3639.2024.12.003.

Jiaqiang REN, Shuai WU, Tong SU, et al. An exploratory study of INPP4B, a biomarker of gemcitabine chemoresistance in pancreatic cancer[J]. China Oncology, 2024, 34(12): 1090-1099. DOI： 10.19401/j.cnki.1007-3639.2024.12.003.

摘要

背景与目的：

胰腺癌是具有高侵袭性的消化系统实体肿瘤，因病程初期缺乏特异性临床表现，约80%的患者无法行根治性切除术。吉西他滨作为胰腺癌化疗的一线用药，由于化疗耐药性，其使用并未显著改善患者预后。迄今，胰腺癌对吉西他滨治疗发生耐药的具体机制仍不清楚。本研究借助胰腺癌吉西他滨耐药细胞系及胰腺癌病理组织标本，结合在线数据库，探索胰腺癌吉西他滨化疗耐药的潜在生物标志物。并通过收集与分析胰腺癌患者的随访资料，明确相关靶点对胰腺癌患者预后的影响。

方法：

本研究采用吉西他滨浓度递增法逐步诱导构建胰腺癌耐药细胞株;对耐药细胞行二代高通量RNA-seq测序，并联合在线网络数据库高通量基因表达数据库（Gene Expression Omnibus，GEO）的4个胰腺癌吉西他滨耐药相关GEO数据集（GSE106336、GSE110580、GSE35141和GSE140077），用生物信息学分析筛选共同差异表达基因;采用实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction，RTFQ-PCR）、蛋白质印迹法（Western blot）及免疫组织化学验证目标分子的表达水平;最后选取自2018年6月—2021年6月就诊于西安交通大学第一附属医院并经手术后病理学检查确诊为胰腺腺癌的部分患者，从西安交通大学第一附属医院生物样本信息资源中心获取行胰腺癌根治性切除术的部分患者手术标本共70例（其中30例对吉西他滨治疗耐药，16例未耐药，同时取24例癌旁胰腺组织作为对照;伦理批件号：2021伦审科字第（457）号，No.XJTU1AF2021LSK-457），并收集其临床和预后相关信息，用log-rank检验评价目标分子的表达与患者预后的关联性。

结果：

检测胰腺癌吉西他滨耐药细胞株（MiaPaCa-2 gemcitabine resistant，Mia GR）及同种野生型细胞系（MiaPaCa-2 wild type，Mia WT）针对吉西他滨杀伤的半数抑制浓度（half maximal inhibitory concentration，IC

），结果表明Mia GR组的IC

显著高于Mia WT组（258.10 μmol/L

0.18 μmol/L），耐药指数（resistence index，RI）高达1 443.9。对耐药株与野生细胞系的二代转录组测序结果分析得到3 985个差异表达基因，再与4个胰腺癌吉西他滨耐药的在线GEO数据集（GSE106336、GSE110580、GSE35141和GSE140077）中的数据作横向比较，发现有25个共同差异表达基因。结合文献回顾和对基因的功能注释和通路富集分析，进一步筛选发现INPP4B在其中发挥关键作用。接下来通过细胞学的RTFQ-PCR及Western blot检测，证实INPP4B在耐药细胞中的mRNA表达和蛋白水平均显著高于野生型（

0.05）。最后对30例耐药胰腺癌组织、16例未耐药胰腺癌组织以及24例癌旁胰腺组织进行免疫组织化学分析。结果显示，与未耐药组相比，耐药组胰腺癌患者的INPP4B表达水平明显更高，而正常组织中INPP4B的表达水平明显低于这两种类型的癌组织。将胰腺癌组织中的INPP4B表达水平与患者生存信息相对应，绘制Kaplan-Meier曲线，结果表明，INPP4B低表达组患者的无进展生存期（progression-free survival，PFS）显著优于INPP4B高表达组（HR=2.874，95% CI：1.262~6.544，

=0.013）。同时，低INPP4B表达组患者的总生存期（overall survival，OS）相比INPP4B高表达组也有明显优势，虽然其间差异无统计学意义（HR=1.484，95% CI：0.518~4.250，

=0.465）。

结论：

INPP4B可能是一种潜在的胰腺癌吉

西他滨化疗耐药生物标志物，且与耐药胰腺癌患者的不良预后相关。针对此靶点设计相应的检测及治疗手段，将有望早期识别对吉西他滨治疗潜在耐药的胰腺癌患者，极大地改善其预后。

Abstract

Background and purpose:

Pancreatic cancer is a highly aggressive solid tumor of the digestive system

with radical resection being unfeasible in approximately 80% of patients due to the absence of specific clinical manifestations in the early stages. The use of gemcitabine as a first-line chemotherapeutic agent has not significantly improved patient prognosis

primarily due to the development of chemoresistance. The precise mechanisms underlying gemcitabine resistance in pancreatic cancer remain unclear. This study aimed to explore potential biomarkers associated with gemcitabine chemoresistance in pancreatic cancer by utilizing gemcitabine-resistant cell lines and pathological pancreatic cancer tissues

in conjunction with data from online databases. Additionally

we analyzed follow-up data from pancreatic cancer patients to assess the impact of relevant targets on patient prognosis.

Methods:

In this study

gemcitabine-resistant cell lines were developed through stepwise induction using a gemcitabine concentration gradient. Second-generation high-throughput RNA-seq sequencing was conducted on these resistant cells

and bioinformatics analysis was employed to identify four pancreatic cancer genes from the Gene Expression Omnibus (GEO) datasets (GSE106336

GSE110580

GSE35141

and GSE140077). Co-expressed genes were screened using real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR)

Western blot and immunohistochemistry to verify the expression levels of target molecules. Surgical specimens from 70 patients diagnosed with pancreatic adenocarcinoma at the First Affiliated Hospital of Xi'an Jiao Tong University between June 2018 and June 2021 were analyzed. These included 30 specimens resistant to gemcitabine

16 non-resistant specimens

and 24 normal pancreatic tissues as controls. Ethical appro

val was obtained (Ethical approval: 2021 LunxuanKeZi No. 457

No. XJTU1AF2021LSK-457). Clinical and prognostic information was collected

and the log-rank test was used to evaluate the relationship between target molecule expression and patient prognosis.

Results:

The half maximal inhibitory concentration (IC

) for gemcitabine was significantly higher in the gemcitabine-resistant cell strain (Mia GR) than in the wild-type cell line (Mia WT) (258.10 μmol/L

0.18 μmol/L)

with a resistance index (RI) of 1 443.9. Transcriptome sequencing identified 3 985 differentially expressed genes

of which 25 were shared with the GEO datasets. Further analysis highlighted

INPP

as a key gene. RTFQ-PCR and Western blot confirmed that INPP4B mRNA and protein levels were significantly elevated in drug resistant cells compared to wild-type cells (

0.05). Immunohistochemical analysis revealed that INPP4B expression was significantly higher in drug resistant pancreatic cancer tissues compared to non-drug resistant tissues

and lower in normal tissues than in both cancerous tissue types. Kaplan-Meier curves demonstrated that patients with low INPP4B expression had significantly better progression-free survival (PFS) than those with high expression (HR=2.874

95% CI: 1.262-6.544

=0.013). Although patients with low INPP4B expression also showed better overall survival (OS)

the difference was not statistically significant (HR=1.484

95% CI: 0.518-4.250

=0.465).

Conclusion:

INPP4B may serve as a potential biomarker for gemcitabine chemoresistance in pancreatic cancer and is associated with poor prognosis in drug resistant patients. Developing targeted assays and treatments for INPP4B could facilitate early identification of patients likely to exhibit resistance to gemcitabine therapy

thereby improving their prognosis.

关键词

Keywords

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