WD40重复蛋白43在肺腺癌中的表达及其对细胞紫杉醇耐药的影响

doi:10.3969/j.issn.1674-8115.2022.12.002

上海交通大学学报（医学版） ›› 2022, Vol. 42 ›› Issue (12): 1656-1665.doi: 10.3969/j.issn.1674-8115.2022.12.002

• 论著 · 基础研究 • 上一篇

WD40重复蛋白43在肺腺癌中的表达及其对细胞紫杉醇耐药的影响

赵雪¹^,²(), 董春燕³()

^1.同济大学医学院，上海 200092
^2.上海市普陀区长寿街道社区卫生服务中心，上海 200060
^3.同济大学附属东方医院肿瘤科，上海 200120

收稿日期:2022-08-16 接受日期:2022-12-11 出版日期:2022-12-28 发布日期:2022-12-28
通讯作者: 董春燕 E-mail:2031143@tongji.edu.cn;cy_dong@tongji.edu.cn
作者简介:赵雪（1989—），女，主治医师，硕士；电子信箱：2031143@tongji.edu.cn。
基金资助:
国家自然科学基金(82073387)

Correlation of WD 40 repeat 43 expression in lung adenocarcinoma and its effect on paclitaxel-resistance of cells

ZHAO Xue¹^,²(), DONG Chunyan³()

^1.Tongji University School of Medicine, Shanghai 200092, China
^2.Changshou Community Health Service Center of Putuo District, Shanghai 200060, China
^3.Department of Oncology, East Hospital, Tongji University, Shanghai 200120, China

Received:2022-08-16 Accepted:2022-12-11 Online:2022-12-28 Published:2022-12-28
Contact: DONG Chunyan E-mail:2031143@tongji.edu.cn;cy_dong@tongji.edu.cn
Supported by:
National Natural Science Foundation of China(82073387)

摘要/Abstract

摘要：

目的·分析WD40重复蛋白43（WD 40 repeat protein 43，WDR43）mRNA在肺腺癌（lung adenocarcinoma，LUAD）细胞中的表达及其与LUAD患者预后的相关性，并探讨WDR43对LUAD细胞发生紫杉醇耐药的影响。方法·通过癌症治疗反应门户网站（Cancer Therapeutics Response Portal，CTRP），下载LUAD细胞的紫杉醇敏感性与其基因表达的相关性系数，筛选紫杉醇耐药相关基因，行基因本体数据库（Gene Ontology，GO）功能分析和京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genome，KEGG）通路分析，并以相关基因WDR43作为研究对象行后续分析及验证。检索人类蛋白图谱（Human Protein Atlas，HPA）数据库，分析WDR43蛋白在LUAD组织中的表达和定位。检索癌症基因组图谱（The Cancer Genome Atlas，TCGA）数据库，分析WDR43 mRNA在LUAD组织和正常肺组织中的表达差异、LUAD患者的WDR43突变情况以及WDR43 mRNA与不同临床病理因素的相关性。检索CaArray数据库，分析WDR43 mRNA与LUAD患者预后之间的相关性。检索肿瘤浸润性免疫细胞分析（Tumor Immune Estimation Resource，TIMER）数据库，分析WDR43表达与LUAD组织中免疫细胞浸润水平的相关性。通过癌症细胞系百科全书（Cancer Cell Line Encyclopedia，CCLE）数据库比较不同LUAD细胞中WDR43基因表达，构建差异表达WDR43的LUAD细胞，并采用MTT实验和平板克隆形成实验验证WDR43表达对紫杉醇耐药性的影响。结果·通过CTRP筛选出了LUAD细胞紫杉醇敏感性相关基因WDR43。GO功能分析和KEGG通路分析的结果显示，WDR43等基因与rRNA合成、细胞形态调控等相关，且主要富集在单纯疱疹病毒1感染信号通路和胞浆DNA感受器信号通路。HPA数据库的分析结果显示，WDR43蛋白主要表达于LUAD细胞的细胞核、细胞质及细胞膜中。TCGA数据库的分析结果显示，WDR43 mRNA在LUAD组织中的表达明显高于正常肺组织（P=0.000），在LUAD吸烟患者的癌组织中的表达明显高于非吸烟者（P=0.000），在TP53突变型患者中的表达亦高于TP53野生型患者（P=0.000）。CaArray数据库的分析结果显示，WDR43 mRNA高表达的LUAD患者的预后较差（均P=0.000）。TIMER数据库的分析结果显示，WDR43表达可促进LUAD组织中的中性粒细胞、CD8⁺T细胞、骨髓来源的抑制性细胞、巨噬细胞的浸润，抑制CD4⁺T细胞的浸润。体外实验证实，过表达WDR43可增强LUAD细胞对紫杉醇的耐药能力。结论·WDR43在LUAD组织中的表达显著增加且与患者预后呈负相关，WDR43的过表达可增强LUAD细胞的紫杉醇耐药能力。

关键词: WD40重复蛋白43, 肺腺癌, 紫杉醇, 化疗耐药, 预后

Abstract:

Objective ·To investigate the expression of WD 40 repeat protein 43 (WDR43) mRNA in lung adenocarcinoma (LUAD) cells and its correlation with the prognosis of LUAD patients, and analyze the effect of WDR43 on the paclitaxel-resistance of LUAD cells. Methods ·Cancer Therapeutics Response Portal (CTRP) was used to download the correlation coefficient between gene expression and paclitaxel sensitivity of LUAD cells, and the paclitaxel resistance-associated genes were screened. Gene Ontology (GO) function analysis and Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis were used to analyze the paclitaxel resistance-associated genes, and WDR43 gene was chosen to perform further study. Human Protein Atlas (HPA) database was used to explore the expression and location of WDR43 protein in LUAD tissues. The cancer genome atlas (TCGA) database was retrieved to analyze the difference of WDR43 mRNA expression in LUAD tissues and normal lung tissues, the mutation of WDR43 in LUAD patients, and the correlation between WDR43 mRNA and clinicopathological factors. CaArray database was used to analyze the relationship between WDR43 mRNA and the prognosis of LUAD patients. Tumor Immune Estimation Resource (TIMER) was used to analyze the relationship between WDR43 expression and the infiltration level of immune cells in LUAD tissues. The expressions of WDR43 gene in LUAD cell lines were analyzed by Cancer Cell Line Encyclopedia (CCLE) database, the WDR43 differential-expressed LUAD cells were constructed and the effect of WDR43 expression on paclitaxel resistance was validated by MTT assay and plate colony formation assay. Results ·WDR43, a paclitaxel sensitivity-related gene, was selected by CTRP. The results of GO function analysis and KEGG pathway analysis showed that WDR43 and other genes were associated with rRNA processing and regulation of cell shape, and were enriched on the pathway such as herpes simplex virus 1 infection and cytosolic DNA-sensing pathway. The analysis results of HPA database showed that WDR43 protein was mainly expressed in the cellular nucleus, cellular cytoplasm and cellular membrane of LUAD tissues. The analysis results of TCGA database showed that the expression of WDR43 mRNA in LUAD tissues was higher than that in normal lung tissues (P=0.000), the expression in cancer tissues of LUAD smokers was higher than that of non-smokers (P=0.000), and the expression in TP53 mutant patients was also higher than that in TP53 wild-type patients (P=0.000). The analysis results of CaArray database showed that LUAD patients with WDR43 mRNA-high expression had poor prognosis (both P=0.000). The analysis results of TIMER database showed that the expression of WDR43 could promote the infiltration level of neutrophils, CD8⁺T cells, myeloid-derived suppressor cells and macrophages, and inhibit the infiltration level of CD4⁺T cells in LUAD tissues. In vitro experiments confirmed that overexpression of WDR43 could enhance the drug resistance of LUAD cells to paclitaxel. Conclusion ·The expression of WDR43 in LUAD tissues is significantly increased and negatively correlated with the prognosis of patients. Overexpression of WDR43 could enhance the paclitaxel resistance of LUAD cells.

Key words: WD40 repeat 43 (WDR43), lung adenocarcinoma, paclitaxel, chemoresistance, prognosis

中图分类号:

R734.2

赵雪, 董春燕. WD40重复蛋白43在肺腺癌中的表达及其对细胞紫杉醇耐药的影响[J]. 上海交通大学学报（医学版）, 2022, 42(12): 1656-1665.

ZHAO Xue, DONG Chunyan. Correlation of WD 40 repeat 43 expression in lung adenocarcinoma and its effect on paclitaxel-resistance of cells[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2022, 42(12): 1656-1665.

图/表 9

表 1 qPCR引物序列

Tab 1 Primer sequences for qPCR

Primer	Forward （5’→3’）	Reverse （5’→3’）
WDR43	CCTACTTCGCTTTGGCCTCTA	GAAGGCACGTACTCCTGGTG
GAPDH	AATGGGCAGCCGTTAGGAAA	GCCCAATACGACCAAATCAGAG

图1 WDR43 表达与LUAD细胞紫杉醇敏感性的相关性分析Note： Red dot—paclitaxel sensitivity-associated genes of LUAD cells.

Fig 1 Correlation between the expression of WDR43 and the sensitivity of paclitaxel in LUAD cells

图2 紫杉醇敏感性相关基因的GO功能分析Note： A. Biological process. B. Cellular component. C. Molecular function. ERK1—extracellular signal-regulated kinases 1; ERK2—extracellular signal-regulated kinases 2.

Fig 2 GO function analysis of paclitaxel sensitivity-related genes

图3 紫杉醇敏感性相关基因的KEGG通路分析

Fig 3 KEGG pathway analysis of paclitaxel sensitivity-related genes

图4 WDR43 在LUAD组织中的表达水平和突变情况Note：A. Immunohistochemistry images of WDR43 expression in LUAD tissues by HPA database. Scale bar from left to right is 100, 50 and 25 μm. B. Expression level of WDR43 mRNA in tumor tissues and normal lung tissues of LUAD patients. C. Expression level of WDR43 mRNA in different AJCC (American Joint Committee on Cancer) stages in tumor tissues and normal lung tissues of LUAD patients. D. Expression level of WDR43 mRNA in tumor tissues and normal lung tissues of LUAD based on patients' smoking habits. E. Expression level of WDR43 mRNA in tumor tissues and normal lung tissues of TP53 wild-type and TP53 mutation LUAD patients. F. Alteration frequency of WDR43. G. Mutation spectrum and count of WDR43 and TMB nonsynonymous in the TCGA-LUAD cohort by OncoPrint. TMB—tumor mutation burden. H. Mutation frequency and site of WDR43 in LUAD patients. WD40 zoom (left) is WD domain, G-β repeat (119?154); WD40 zoom (right) is WD domain, G-β repeat (159?193).

Fig 4 Expression level and mutation of WDR43 in LUAD tissues

表2 WDR43 mRNA高表达组和 WDR43 mRNA低表达组患者的临床病理因素分析

Tab 2 Analysis of clinicopathological factors of patients in WDR43 mRNA-high expression and WDR43 mRNA-low expression

Clinicopathological factor	WDR43 mRNA-low expression group （n=267）	WDR43 mRNA-high expression group （n=268）	P value
Age/year	66 （59， 73）	66 （59， 72）	0.614
Gender/n（%）			0.010
Female	158 （59.18）	128 （47.76）
Male	109 （40.82）	140 （52.24）
T stage/n（%）			0.003
T1	106 （39.70）	69 （25.75）
T2	129 （48.31）	160 （59.70）
T3	24 （8.99）	25 （9.33）
T4	6 （2.25）	13 （4.85）
N stage/n（%）			0.023
N0	181 （67.79）	167 （62.31）
N1	49 （18.35）	46 （17.16）
N2	25 （9.36）	49 （18.28）
N3	1 （0.37）	1 （0.37）
M stage/n（%）			0.142
M0	178 （66.67）	183 （68.28）
M1	8 （3.00）	17 （6.34）
Tumor stage/n（%）			0.033
Stage 1	157 （58.80）	137 （51.12）
Stage 2	64 （23.97）	59 （22.01）
Stage 3	32 （11.99）	52 （19.40）
Stage 4	9 （3.37）	17 （6.34）
Smoking/n（%）			0.000
No	53 （19.85）	22 （8.21）
Yes	203 （76.03）	243 （90.67）
Number of smoking/n（%）			0.988
<40 packets per year	84 （41.38）	104 （42.80）
≥40 packets per year	82 （40.39）	99 （40.74）

图5 基于CaArray数据库的 WDR43 mRNA高表达组和低表达组患者的总体生存期（A）、无进展生存期（B）分析

Fig 5 Analysis of OS (A) and PFS (B) of patients with WDR43 mRNA-high expression and WDR43 mRNA-low expression based on CaArray database

图6 WDR43 表达与LUAD组织中不同免疫细胞浸润水平的相关性

Fig 6 Correlation between WDR43 expression and infiltration levels of different immune cells in LUAD tissues

图7 体外实验验证 WDR43 表达与紫杉醇敏感性的相关性Note： A. Expression level of WDR43 in LUAD cell lines by CCLE database. B Detection of relative expression of WDR43 in HCC827-WDR43 cells and HCC827-Vector cells by qPCR. C. Detection of proliferation activity of HCC827-WDR43 cells and HCC827-Vector cells by MTT assay. D/E. Colony formation (D) and statistical analysis (E) of HCC827-WDR43 cells and HCC827-Vector cells.

Fig 7 Correlation between WDR43 expression and the sensitivity of paclitaxel in vitro

参考文献 20

1	SUNG H, FERLAY J, SIEGEL R L, et al. Global cancer statistics 2020: globocan estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2021, 71(3): 209-249.
2	O'KEEFFE L M, TAYLOR G, HUXLEY R R, et al. Smoking as a risk factor for lung cancer in women and men: a systematic review and meta-analysis[J]. BMJ Open, 2018, 8(10): e021611.
3	SHEN D J, WANG J F, WU J, et al. Neoadjuvant pembrolizumab with chemotherapy for the treatment of stage ⅡB-ⅢB resectable lung squamous cell carcinoma[J]. J Thorac Dis, 2021, 13(3): 1760-1768.
4	CHENG Z A, YU C T, CUI S H, et al. circTP63 functions as a ceRNA to promote lung squamous cell carcinoma progression by upregulating FOXM1[J]. Nat Commun, 2019, 10(1): 3200.
5	WEST H, MCCLEOD M, HUSSEIN M, et al. Atezolizumab in combination with carboplatin plus nab-paclitaxel chemotherapy compared with chemotherapy alone as first-line treatment for metastatic non-squamous non-small-cell lung cancer (IMpower130): a multicentre, randomised, open-label, phase 3 trial[J]. Lancet Oncol, 2019, 20(7): 924-937.
6	MUELLER H S, FOWLER C E, DALIN S, et al. Acquired resistance to PRMT5 inhibition induces concomitant collateral sensitivity to paclitaxel[J]. Proc Natl Acad Sci USA, 2021, 118(34): e2024055118.
7	CAI X Y, CHEN Z H, DENG M L, et al. Unique genomic features and prognostic value of COSMIC mutational signature 4 in lung adenocarcinoma and lung squamous cell carcinoma[J]. Ann Transl Med, 2020, 8(18): 1176.
8	SUGAWARA S, LEE J S, KANG J H, et al. Nivolumab with carboplatin, paclitaxel, and bevacizumab for first-line treatment of advanced nonsquamous non-small-cell lung cancer[J]. Ann Oncol, 2021, 32(9): 1137-1147.
9	SONDALLE S B, BASERGA S J, YELICK P C. The contributions of the ribosome biogenesis protein Utp5/WDR43 to craniofacial development[J]. J Dent Res, 2016, 95(11): 1214-1220.
10	PEÑA C, HURT E, PANSE V G. Eukaryotic ribosome assembly, transport and quality control[J]. Nat Struct Mol Biol, 2017, 24(9): 689-699.
11	BI X J, XU Y H, LI T, et al. RNA targets ribogenesis factor WDR43 to chromatin for transcription and pluripotency control[J]. Mol Cell, 2019, 75(1): 102-116.e9.
12	PARANT J M, GEORGE S A, PRYOR R, et al. A rapid and efficient method of genotyping zebrafish mutants[J]. Dev Dyn, 2009, 238(12): 3168-3174.
13	COUCH F J, KUCHENBAECKER K B, MICHAILIDOU K, et al. Identification of four novel susceptibility loci for oestrogen receptor negative breast cancer[J]. Nat Commun, 2016, 7: 11375.
14	MENDOZA-FANDIÑO G, LYRA P C M, NEPOMUCENO T C, et al. Two distinct mechanisms underlie estrogen-receptor-negative breast cancer susceptibility at the 2p23.2 locus[J]. Eur J Hum Genet, 2022, 30(4): 465-473.
15	LI Z J, FENG M, ZHANG J, et al. WD40 repeat 43 mediates cell survival, proliferation, migration and invasion via vimentin in colorectal cancer[J]. Cancer Cell Int, 2021, 21(1): 418.
16	HUANG Z Y, LI F, LI Q C. Expression profile of RNA binding protein in cervical cancer using bioinformatics approach[J]. Cancer Cell Int, 2021, 21(1): 647.
17	SCHAPIRA M, TYERS M, TORRENT M, et al. WD40 repeat domain proteins: a novel target class?[J]. Nat Rev Drug Discov, 2017, 16(11): 773-786.
18	UHLÉN M, FAGERBERG L, HALLSTRÖM B M, et al. Proteomics. Tissue-based map of the human proteome[J]. Science, 2015, 347(6220): 1260419.
19	SUN H J, SUN Q, QIU X, et al. WD repeat domain 43 promotes malignant progression of non-small cell lung cancer by regulating CDK2[J]. Int J Biochem Cell Biol, 2022, 151: 106293.
20	史清文. 天然药物化学史话: 紫杉醇[J]. 中草药, 2011, 42(10): 1878-1884.
	SHI Q W. Historical story on natural medicinal chemistry of Taxol[J]. Chinese Traditional and Herbal Drugs, 2011, 42(10): 1878-1884.

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WD40重复蛋白43在肺腺癌中的表达及其对细胞紫杉醇耐药的影响

Correlation of WD 40 repeat 43 expression in lung adenocarcinoma and its effect on paclitaxel-resistance of cells

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