|Table of Contents|

Transcriptome sequencing to explore the mechanism of EGFR-TKI damage to HaCaT cells

Journal Of Modern Oncology[ISSN:1672-4992/CN:61-1415/R]

Issue:
2023 19
Page:
3539-3547
Research Field:
Publishing date:

Info

Title:
Transcriptome sequencing to explore the mechanism of EGFR-TKI damage to HaCaT cells
Author(s):
ZHONG Di1; 2; XIAO Bingying1; 2; LIN Xiaohui2; CHEN Xin1; 2; WANG Yujia2; NIE Xuekun2; LIN Minhua2
1.Fujian Medical University,Fujian Fuzhou 350100,China;2.Department of Pharmacy,Ningde Municipal Hospital of Ningde Normal University,Fujian Ningde 352100,China.
Keywords:
transcriptome sequencing; molecular docking; EGFR-TKI; epidermal toxicity
PACS:
R730
DOI:
10.3969/j.issn.1672-4992.2023.19.003
Abstract:
Objective:To investigate the mechanism of epidermal growth factor receptor-tyrosine kinase inhibitors(EGFR-TKIs) causing damage to human immortalized keratinocytes(HaCaT) cells,and thus to explore the preliminary mechanism of epidermal adverse reactions induced by this drug.Methods:MTS and Hoechst staining were used to model TKI drug damage to HaCaT cells.Transcriptome sequencing was used to screen for differential genes.The database was used to collect targets for epidermal adverse reactions.The intersection of differential genes and targets for epidermal adverse reactions was taken to obtain potential targets.The potential targets were analyzed by the Kyoto encyclopedia of genes and genomes(KEGG) and gene ontology(GO) using DAVID and R software.Protein protein interaction(PPI) analysis was performed using the STRING database,and Cytoscape software was used to visualisation and topological parameters were analysed using Cytoscape software.Molecular docking of key genes to EGFR-TKI drugs was performed using Autodock software.Real-time fluorescence PCR was used to detect the lowest binding energy targets in molecular docking and Wesrtern Blot was used to detect pathway-related protein expression.Results:MTS and Hoechst staining showed that ositinib and afatinib had a significant inhibitory effect on cell survival.176 potential targets of action were obtained after taking intersection.PPI showed that IL6,HSPA5,FN1,VEGFA,XBP1,TLR4,MMP1,CCL2,PTGS2 and VWF may be key targets of EGFR-TKI epidermal toxicity.KEGG signaling pathway includes IL-17,PI3K-Akt pathway.GO function involves endoplasmic reticulum stress,oxidative stress,lipopolysaccharide response.Molecular docking showed that oxitinib and afatinib have good binding activity with PTGS2,HSPA5,MMP1,VEGFA and FN1.Real-time fluorescence quantitative PCR showed that FN1,HSPA5 and PTGS2 expression were up-regulated and VEGFA and MMP1 expression were down-regulated after treatment of HaCaT cells with axitinib and afatinib.Immunoblotting experiments showed that FN1 protein expression increased and VEGFA protein expression decreased.Conclusion:EGFR-TKI drugs may produce epidermal toxicity by inhibiting MMP1 and VEGFA production in PI3K-Akt and IL-17 signaling pathways,promoting PTGS2,FN1 and HSPA5 expression and damaging HaCaT cells.

References:

[1]XIA C,DONG X,LI H,et al.Cancer statistics in China and United States,2022:profiles,trends,and determinants［J］.Chin Med J(Engl),2022,135(5):584-590.
[2]MULLER-HERMELINK HK,STROBEL P,ZETTL A,et al.Pathology and genetics of tumours of the lung,pleura,thymus and heart(WHO classification of tumours series)［M］.Lyon:IARC Press,2004:26-67.
[3]MOLINA JR,YANG P,CASSIVI SD,et al.Non-small cell lung cancer:epidemiology,risk factors,treatment,and survivorship［J］.Mayo Clin Proc,2008,83(5):584-594.
[4]LEE CC,SHIAO HY,WANG WC,et al.Small-molecule EGFR tyrosine kinase inhibitors for the treatment of cancer［J］.Expert Opin Investig Drugs,2014,23(10):1333-1348.
[5]MOORE MJ,GOLDSTEIN D,HAMM J,et al.Erlotinib plus gemcitabine compared with gemcitabine alone in patients with advanced pancreatic cancer:a phase III trial of the National Cancer Institute of Canada Clinical Trials Group［J］.J Clin Oncol,2007,25(15):1960-1966.
[6]BOULOUGOURIS P,ELDER J.Epidermal growth factor receptor structure,regulation,mitogenic signalling and effects of activation［J］.Anticancer Res,2001,21(4A):2769-2775.
[7]CALIFANO R,TARIQ N,COMPTON S,et al.Expert consensus on the management of adverse events from EGFR tyrosine kinase inhibitors in the UK［J］.Drugs,2015,75(12):1335-1348.
[8]TROTT O,OLSON AJ.AutoDock Vina:improving the speed and accuracy of docking with a new scoring function,efficient optimization,and multithreading［J］.J Comput Chem,2010,31(2):455-461.
[9]徐森楠,庄莉,翟园园,等.基于网络药理学研究二至丸防治骨质疏松症的物质基础与作用机制［J］.中国药学杂志,2018,53(22):1913-1920. XU SN,ZHUANG L,ZHAI YY,et al.Network-based pharmacological study of the material basis and mechanism of action of Erzhi Pills in the prevention and treatment of osteoporosis［J］.Chinese Journal of Pharmacy,2018,53(22):1913-1920.
[10]CHO JH,LIM SH,AN HJ,et al.Osimertinib for patients with non-small-cell lung cancer harboring uncommon EGFR mutations:A multicenter,open-label,phase II trial(KCSG-LU15-09)［J］.J Clin Oncol,2020,38(5):488-495.
[11]WAHLER G,HECK DE,HEINDEL ND,et al.Antioxidant/stress response in mouse epidermis following exposure to nitrogen mustard［J］.Exp Mol Pathol,2020,114:104410.
[12]LICHTENBERGER BM,GERBER PA,HOLCMANN M,et al.Epidermal EGFR controls cutaneous host defense and prevents inflammation［J］.Sci Transl Med,2013,5(199):199ra111.
[13]叶丹,谢长生.表皮生长因子受体酪氨酸激酶抑制剂相关皮疹的中西医治疗进展［J］.肿瘤学杂志,2019,25(03):256-260. YE D,XIE CS.Advances in Chinese and Western medicine for the treatment of epidermal growth factor receptor tyrosine kinase inhibitor-associated skin rashes［J］.Journal of Oncology,2019,25(03):256-260.
[14]LI Y,FU R,JIANG T,et al.Mechanism of lethal skin toxicities induced by epidermal growth factor receptor inhibitors and related treatment strategies［J］.Front Oncol,2022,12:804212.
[15]WILGUS TA,ROSS MS,PARRETT ML,et al.Topical application of a selective cyclooxygenase inhibitor suppresses UVB mediated cutaneous inflammation［J］.Prostaglandins Other Lipid Mediat,2000,62(4):367-384.
[16]WANG Y,ZHANG Y,DING C,et al.Exploration of the potential mechanism of Qi Yin San Liang San Decoction in the treatment of EGFRI-related adverse skin reactions using network pharmacology and in vitro experiments［J］.Front Oncol,2022,12:790713.
[17]FAOUR WH,MANCINI A,HE QW,et al.T-cell-derived interleukin-17 regulates the level and stability of cyclooxygenase-2(COX-2) mRNA through restricted activation of the p38 mitogen-activated protein kinase cascade:role of distal sequences in the 3'-untranslated region of COX-2 mRNA［J］.J Biol Chem,2003,278(29):26897-26907.
[18]LINDNER D,ZIETSCH C,BECHER PM,et al.Differential expression of matrix metalloproteases in human fibroblasts with different origins［J］.Biochem Res Int,2012,2012:875742.
[19]SHIH MF,CHERNG JY.Potential protective effect of fresh grown unicellular green algae component(resilient factor) against PMA- and UVB-induced MMP1 expression in skin fibroblasts［J］.Eur J Dermatol,2008,18(3):303-307.
[20]BREMBILLA NC,MONTANARI E,TRUCHETET ME,et al.Th17 cells favor inflammatory responses while inhibiting type I collagen deposition by dermal fibroblasts:differential effects in healthy and systemic sclerosis fibroblasts［J］.Arthritis Res Ther,2013,15(5):R151.
[21]MHHP A,SH B,MA A.Wound healing potential of pomegranate peel extract in human dermal fibroblasts through regulating the expression of FN1 gene［J］.South African Journal of Botany,2022,146:222-229.
[22]KNAUP J,VERWANGER T,GRUBER C,et al.Epidermolysis bullosa - a group of skin diseases with different causes but commonalities in gene expression［J］.Exp Dermatol,2012,21(7):526-530.
[23]MOON SJ,BAE JM,PARK KS,et al.Compendium of skin molecular signatures identifies key pathological features associated with fibrosis in systemic sclerosis［J］.Ann Rheum Dis,2019,78(6):817-825.
[24]JEON HH,YU Q,LU Y,et al.FOXO1 regulates VEGFA expression and promotes angiogenesis in healing wounds［J］.J Pathol,2018,245(3):258-264.
[25]CHEN J,GU Z,WU M,et al.C-reactive protein can upregulate VEGF expression to promote ADSC-induced angiogenesis by activating HIF-1α via CD64/PI3k/Akt and MAPK/ERK signaling pathways［J］.Stem Cell Res Ther,2016,7(1):114.
[26]ZHAO M,LUO J,XIAO B,et al.Endoplasmic reticulum stress links psoriasis vulgaris with keratinocyte inflammation［J］.Postepy Dermatol Alergol,2020,37(1):34-40.
[27]YANG GH,LI S,PESTKA JJ.Down-regulation of the endoplasmic reticulum chaperone GRP78/BiP by vomitoxin(Deoxynivalenol)［J］.Toxicol Appl Pharmacol,2000,162(3):207-217.

Memo

Memo:
福建省自然科学基金面上项目（编号：2020J011341）;福建省宁德市科技局指导性项目（编号：20190045，20170104）
Last Update: 2023-08-31