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27.03.2017 | Preclinical study

ΔNp63 activates EGFR signaling to induce loss of adhesion in triple-negative basal-like breast cancer cells

verfasst von: Jitka Holcakova, Marta Nekulova, Paulina Orzol, Rudolf Nenutil, Jan Podhorec, Marek Svoboda, Petra Dvorakova, Mariana Pjechova, Lenka Hernychova, Borivoj Vojtesek, Philip J. Coates

Erschienen in: Breast Cancer Research and Treatment | Ausgabe 3/2017

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Abstract

Purpose

The basal-A subtype of triple-negative breast cancer is characterized by high levels of ΔNp63. Various functions have been proposed for p63 in breast cancer initiation and growth, and p63 mediates chemotherapeutic response in a subset of triple-negative breast cancers. We investigated the signaling pathways that are controlled by ΔNp63 in basal-A triple-negative breast cancer.

Methods

Human basal-A triple-negative breast cancer cell lines with ΔNp63α induction or inhibition were studied, along with primary human triple-negative breast cancer tissues. Proteomic, phospho-kinase array, mRNA measurements, and immunohistochemistry were employed.

Results

Global phosphoproteomics identified increased EGFR phosphorylation in MDA-MB-468 cells expressing ΔNp63α. ΔNp63α expression increased EGFR mRNA, total EGFR protein, and phospho-EGFR(Y1086), whereas silencing endogenous ΔNp63 in HCC1806 cells reduced both total and phospho-EGFR levels and inhibited the ability of EGF to activate EGFR. EGFR pathway gene expression analysis indicated that ΔNp63 alters EGFR-regulated genes involved in cell adhesion, migration, and angiogenesis. Addition of EGF or neutralizing EGFR antibodies demonstrated that EGFR activation is responsible for ΔNp63-mediated loss of cellular adhesion. Finally, immunohistochemical staining showed that p63-positive triple-negative breast cancers were more likely to express high levels of EGFR than p63-negative cancers, corroborated by in silico analysis of gene expression profiling data.

Conclusions

These data identify EGFR as a major target for ΔNp63 regulation that influences cancer cell adhesion in basal-like triple-negative breast cancer.

Nur mit Berechtigung zugänglich

Crum CP, McKeon FD (2010) p63 in epithelial survival, germ cell surveillance, and neoplasia. Annu Rev Pathol 5:349–371. doi:10.1146/annurev-pathol-121808-102117 CrossRefPubMed

Nekulova M, Holcakova J, Coates P, Vojtesek B (2011) The role of p63 in cancer, stem cells and cancer stem cells. Cell Mol Biol Lett 16(2):296–327. doi:10.2478/s11658-011-0009-9 CrossRefPubMed

Orzol P, Holcakova J, Nekulova M, Nenutil R, Vojtesek B, Coates PJ (2015) The diverse oncogenic and tumour suppressor roles of p63 and p73 in cancer: a review by cancer site. Histol Histopathol 30(5):503–521. doi:10.14670/HH-30.503 PubMed

Nylander K, Vojtesek B, Nenutil R, Lindgren B, Roos G, Zhanxiang W, Sjostrom B, Dahlqvist A, Coates PJ (2002) Differential expression of p63 isoforms in normal tissues and neoplastic cells. J Pathol 198(4):417–427. doi:10.1002/path.1231 CrossRefPubMed

Lehmann BD, Pietenpol JA (2014) Identification and use of biomarkers in treatment strategies for triple-negative breast cancer subtypes. J Pathol 232(2):142–150. doi:10.1002/path.4280 CrossRefPubMedPubMedCentral

Leong CO, Vidnovic N, DeYoung MP, Sgroi D, Ellisen LW (2007) The p63/p73 network mediates chemosensitivity to cisplatin in a biologically defined subset of primary breast cancers. J Clin Invest 117(5):1370–1380. doi:10.1172/JCI30866 CrossRefPubMedPubMedCentral

Rakha EA, Elsheikh SE, Aleskandarany MA et al (2009) Triple-negative breast cancer: distinguishing between basal and nonbasal subtypes. Clin Cancer Res 15(7):2302–2310. doi:10.1158/1078-0432.CCR-08-2132 CrossRefPubMed

Thike AA, Cheok PY, Jara-Lazaro AR, Tan B, Tan P, Tan PH (2010) Triple-negative breast cancer: clinicopathological characteristics and relationship with basal-like breast cancer. Mod Pathol 23(1):123–133. doi:10.1038/modpathol.2009.145 CrossRefPubMed

Neve RM, Chin K, Fridlyand J et al (2006) A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell 10(6):515–527. doi:10.1016/j.ccr.2006.10.008 CrossRefPubMedPubMedCentral

10.

Carroll DK, Carroll JS, Leong CO, Cheng F, Brown M, Mills AA, Brugge JS, Ellisen LW (2006) p63 regulates an adhesion programme and cell survival in epithelial cells. Nat Cell Biol 8(6):551–561. doi:10.1038/ncb1420 CrossRefPubMed

11.

Nekulova M, Holcakova J, Gu X, Hrabal V, Galtsidis S, Orzol P, Liu Y, Logotheti S, Zoumpourlis V, Nylander K, Coates PJ, Vojtesek B (2016) ΔNp63α expression induces loss of cell adhesion in triple-negative breast cancer cells. BMC Cancer 16(1):782. doi:10.1186/s12885-016-2808-x CrossRefPubMedPubMedCentral

12.

Buckley NE, Conlon SJ, Jirstrom K et al (2011) The {Delta}Np63 proteins are key allies of BRCA1 in the prevention of basal-like breast cancer. Cancer Res 71(5):1933–1944. doi:10.1158/0008-5472.CAN-10-2717 CrossRefPubMed

13.

Li N, Singh S, Cherukuri P, Li H, Yuan Z, Ellisen LW, Wang B, Robbins D, DiRenzo J (2008) Reciprocal intraepithelial interactions between TP63 and hedgehog signaling regulate quiescence and activation of progenitor elaboration by mammary stem cells. Stem Cells 26(5):1253–1264. doi:10.1634/stemcells.2007-0691 CrossRefPubMedPubMedCentral

14.

Chakrabarti R, Wei Y, Hwang J, et al (2014) DeltaNp63 promotes stem cell activity in mammary gland development and basal-like breast cancer by enhancing Fzd7 expression and Wnt signalling. Nat Cell Biol 16(10): 1004–1015, 1001–1013. doi: 10.1038/ncb3040

15.

Memmi EM, Sanarico AG, Giacobbe A et al (2015) p63 sustains self-renewal of mammary cancer stem cells through regulation of Sonic Hedgehog signaling. Proc Natl Acad Sci USA 112(11):3499–3504. doi:10.1073/pnas.1500762112 CrossRefPubMedPubMedCentral

16.

Rocca A, Viale G, Gelber RD, Bottiglieri L, Gelber S, Pruneri G, Ghisini R, Balduzzi A, Pietri E, D’Alessandro C, Goldhirsch A, Colleoni M (2008) Pathologic complete remission rate after cisplatin-based primary chemotherapy in breast cancer: correlation with p63 expression. Cancer Chemother Pharmacol 61(6):965–971. doi:10.1007/s00280-007-0551-3 CrossRefPubMed

17.

Isakoff SJ, Mayer EL, He L et al (2015) TBCRC009: a multicenter phase II clinical trial of platinum monotherapy with biomarker assessment in metastatic triple-negative breast cancer. J Clin Oncol 33(17):1902–1909. doi:10.1200/JCO.2014.57.6660 CrossRefPubMedPubMedCentral

18.

Badve S, Dabbs DJ, Schnitt SJ et al (2011) Basal-like and triple-negative breast cancers: a critical review with an emphasis on the implications for pathologists and oncologists. Mod Pathol 24(2):157–167. doi:10.1038/modpathol.2010.200 CrossRefPubMed

19.

Turner NC, Reis-Filho JS (2013) Tackling the diversity of triple-negative breast cancer. Clin Cancer Res 19(23):6380–6388. doi:10.1158/1078-0432.CCR-13-0915 CrossRefPubMed

20.

Masuda H, Zhang D, Bartholomeusz C, Doihara H, Hortobagyi GN, Ueno NT (2012) Role of epidermal growth factor receptor in breast cancer. Breast Cancer Res Treat 136(2):331–345. doi:10.1007/s10549-012-2289-9 CrossRefPubMed

21.

Hochgrafe F, Zhang L, O’Toole SA et al (2010) Tyrosine phosphorylation profiling reveals the signaling network characteristics of Basal breast cancer cells. Cancer Res 70(22):9391–9401. doi:10.1158/0008-5472.CAN-10-0911 CrossRefPubMed

22.

Dvorakova P, Nekulova M, Holcakova J, Vojtesek B, Hernychova L (2015) Analysis of phosphoproteome changes in MDA MB 468 cancer cell line in response to expression of p63 isoforms using mass spectrometry. Klin Onkol 28(Suppl 2):2S11–2S19. doi:10.14375/amko20152S11 CrossRefPubMed

23.

Boersema PJ, Mohammed S, Heck AJ (2009) Phosphopeptide fragmentation and analysis by mass spectrometry. J Mass Spectrom 44(6):861–878. doi:10.1002/jms.1599 CrossRefPubMed

24.

Wisniewski JR, Zougman A, Nagaraj N, Mann M (2009) Universal sample preparation method for proteome analysis. Nat Methods 6(5):359–362. doi:10.1038/nmeth.1322 CrossRefPubMed

25.

Taus T, Kocher T, Pichler P, Paschke C, Schmidt A, Henrich C, Mechtler K (2011) Universal and confident phosphorylation site localization using phosphoRS. J Proteome Res 10(12):5354–5362. doi:10.1021/pr200611n CrossRefPubMed

26.

Li C, Wong WH (2001) Model-based analysis of oligonucleotide arrays: expression index computation and outlier detection. Proc Natl Acad Sci U S A 98(1):31–36. doi:10.1073/pnas.011404098 CrossRefPubMed

27.

Orzol P, Nekulova M, Holcakova J, Muller P, Votesek B, Coates PJ (2016) ΔNp63 regulates cell proliferation, differentiation, adhesion, and migration in the BL2 subtype of basal-like breast cancer. Tumour Biol 37(8):10133–10140. doi:10.1007/s13277-016-4880-x CrossRefPubMed

28.

Huang Y, Jeong JS, Okamura J, Sook-Kim M, Zhu H, Guerrero-Preston R, Ratovitski EA (2012) Global tumor protein p53/p63 interactome: making a case for cisplatin chemoresistance. Cell Cycle 11(12):2367–2379. doi:10.4161/cc.20863 CrossRefPubMedPubMedCentral

29.

Daub H, Olsen JV, Bairlein M, Gnad F, Oppermann FS, Korner R, Greff Z, Keri G, Stemmann O, Mann M (2008) Kinase-selective enrichment enables quantitative phosphoproteomics of the kinome across the cell cycle. Mol Cell 31(3):438–448. doi:10.1016/j.molcel.2008.07.007 CrossRef

30.

Dephoure N, Zhou C, Villen J, Beausoleil SA, Bakalarski CE, Elledge SJ, Gygi SP (2008) A quantitative atlas of mitotic phosphorylation. Proc Natl Acad Sci U S A 105(31):10762–10767. doi:10.1073/pnas.0805139105 CrossRefPubMedPubMedCentral

31.

Heisermann GJ, Gill GN (1988) Epidermal growth factor receptor threonine and serine residues phosphorylated in vivo. J Biol Chem 263(26):13152–13158PubMed

32.

Decker SJ (1993) Transmembrane signaling by epidermal growth factor receptors lacking autophosphorylation sites. J Biol Chem 268(13):9176–9179PubMed

33.

Sorkin A, Waters C, Overholser KA, Carpenter G (1991) Multiple autophosphorylation site mutations of the epidermal growth factor receptor. Analysis of kinase activity and endocytosis. J Biol Chem 266(13):8355–8362PubMed

34.

Van Schaeybroeck S, Karaiskou-McCaul A, Kelly D, Longley D, Galligan L, Van Cutsem E, Johnston P (2005) Epidermal growth factor receptor activity determines response of colorectal cancer cells to gefitinib alone and in combination with chemotherapy. Clin Cancer Res 11(20):7480–7489. doi:10.1158/1078-0432.CCR-05-0328 CrossRefPubMed

35.

Sorkin A, Goh LK (2008) Endocytosis and intracellular trafficking of ErbBs. Exp Cell Res 314(17):3093–3106. doi:10.1016/j.yexcr.2008.08.013 CrossRefPubMedPubMedCentral

36.

Barbieri CE, Tang LJ, Brown KA, Pietenpol JA (2006) Loss of p63 leads to increased cell migration and up-regulation of genes involved in invasion and metastasis. Cancer Res 66(15):7589–7597. doi:10.1158/0008-5472.CAN-06-2020 CrossRefPubMed

37.

Boldrup L, Coates PJ, Gu X, Nylander K (2009) DeltaNp63 isoforms differentially regulate gene expression in squamous cell carcinoma: identification of Cox-2 as a novel p63 target. J Pathol 218(4):428–436. doi:10.1002/path.2560 CrossRefPubMed

38.

Gu X, Coates PJ, Boldrup L, Nylander K (2008) p63 contributes to cell invasion and migration in squamous cell carcinoma of the head and neck. Cancer Lett 263(1):26–34. doi:10.1016/j.canlet.2007.12.011 CrossRefPubMed

39.

Long W, Yi P, Amazit L, LaMarca HL, Ashcroft F, Kumar R, Mancini MA, Tsai SY, Tsai MJ, O’Malley BW (2010) SRC-3Delta4 mediates the interaction of EGFR with FAK to promote cell migration. Mol Cell 37(3):321–332. doi:10.1016/j.molcel.2010.01.004 CrossRefPubMedPubMedCentral

40.

Lu Z, Jiang G, Blume-Jensen P, Hunter T (2001) Epidermal growth factor-induced tumor cell invasion and metastasis initiated by dephosphorylation and downregulation of focal adhesion kinase. Mol Cell Biol 21(12):4016–4031. doi:10.1128/MCB.21.12.4016-4031.2001 CrossRefPubMedPubMedCentral

41.

Craig AL, Holcakova J, Finlan LE, Nekulova M, Hrstka R, Gueven N, DiRenzo J, Smith G, Hupp TR, Vojtesek B (2010) DeltaNp63 transcriptionally regulates ATM to control p53 Serine-15 phosphorylation. Mol Cancer 9:195. doi:10.1186/1476-4598-9-195 CrossRefPubMedPubMedCentral

42.

Lin YL, Sengupta S, Gurdziel K, Bell GW, Jacks T, Flores ER (2009) p63 and p73 transcriptionally regulate genes involved in DNA repair. PLoS Genet 5(10):e1000680. doi:10.1371/journal.pgen.1000680 CrossRefPubMedPubMedCentral

43.

Danilov AV, Neupane D, Nagaraja AS, Feofanova EV, Humphries LA, DiRenzo J, Korc M (2011) DeltaNp63alpha-mediated induction of epidermal growth factor receptor promotes pancreatic cancer cell growth and chemoresistance. PLoS ONE 6(10):e26815. doi:10.1371/journal.pone.0026815 CrossRefPubMedPubMedCentral

44.

Bos PD, Zhang XH, Nadal C, Shu W, Gomis RR, Nguyen DX, Minn AJ, van de Vijver MJ, Gerald WL, Foekens JA, Massague J (2009) Genes that mediate breast cancer metastasis to the brain. Nature 459(7249):1005–1009. doi:10.1038/nature08021 CrossRefPubMedPubMedCentral

45.

Reis-Filho JS, Pinheiro C, Lambros MB et al (2006) EGFR amplification and lack of activating mutations in metaplastic breast carcinomas. J Pathol 209(4):445–453. doi:10.1002/path.2004 CrossRefPubMed

46.

Mouneimne G, Brugge JS (2007) Tensins: a new switch in cell migration. Dev Cell 13(3):317–319. doi:10.1016/j.devcel.2007.08.010 CrossRefPubMed

47.

Dang TT, Westcott JM, Maine EA, Kanchwala M, Xing C, Pearson GW (2016) DeltaNp63alpha induces the expression of FAT2 and Slug to promote tumor invasion. Oncotarget. doi:10.18632/oncotarget.8696

48.

Giacobbe A, Compagnone M, Bongiorno-Borbone L, Antonov A, Markert EK, Zhou JH, Annicchiarico-Petruzzelli M, Melino G, Peschiaroli A (2016) p63 controls cell migration and invasion by transcriptional regulation of MTSS1. Oncogene 35(12):1602–1608. doi:10.1038/onc.2015.230 CrossRefPubMed

49.

Matheny KE, Barbieri CE, Sniezek JC, Arteaga CL, Pietenpol JA (2003) Inhibition of epidermal growth factor receptor signaling decreases p63 expression in head and neck squamous carcinoma cells. Laryngoscope 113(6):936–939. doi:10.1097/00005537-200306000-00004 CrossRefPubMed

50.

Ripamonti F, Albano L, Rossini A, Borrelli S, Fabris S, Mantovani R, Neri A, Balsari A, Magnifico A, Tagliabue E (2013) EGFR through STAT3 modulates DeltaN63alpha expression to sustain tumor-initiating cell proliferation in squamous cell carcinomas. J Cell Physiol 228(4):871–878. doi:10.1002/jcp.24238 CrossRefPubMed

51.

Amit I, Citri A, Shay T et al (2007) A module of negative feedback regulators defines growth factor signaling. Nat Genet 39(4):503–512. doi:10.1038/ng1987 CrossRefPubMed

52.

Forster N, Saladi SV, van Bragt M, Sfondouris ME, Jones FE, Li Z, Ellisen LW (2014) Basal cell signaling by p63 controls luminal progenitor function and lactation via NRG1. Dev Cell 28(2):147–160. doi:10.1016/j.devcel.2013.11.019 CrossRefPubMedPubMedCentral

53.

Assefnia S, Kang K, Groeneveld S, Yamaji D, Dabydeen S, Alamri A, Liu X, Hennighausen L, Furth PA (2014) Trp63 is regulated by STAT5 in mammary tissue and subject to differentiation in cancer. Endocr Relat Cancer 21(3):443–457. doi:10.1530/ERC-14-0032 CrossRefPubMedPubMedCentral

54.

Walker SR, Xiang M, Frank DA (2014) Distinct roles of STAT3 and STAT5 in the pathogenesis and targeted therapy of breast cancer. Mol Cell Endocrinol 382(1):616–621. doi:10.1016/j.mce.2013.03.010 CrossRefPubMed

Titel: ΔNp63 activates EGFR signaling to induce loss of adhesion in triple-negative basal-like breast cancer cells
verfasst von: Jitka Holcakova
Marta Nekulova
Paulina Orzol
Rudolf Nenutil
Jan Podhorec
Marek Svoboda
Petra Dvorakova
Mariana Pjechova
Lenka Hernychova
Borivoj Vojtesek
Philip J. Coates
Publikationsdatum: 27.03.2017
Verlag: Springer US
Erschienen in: Breast Cancer Research and Treatment / Ausgabe 3/2017
Print ISSN: 0167-6806
Elektronische ISSN: 1573-7217
DOI: https://doi.org/10.1007/s10549-017-4216-6

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ΔNp63 activates EGFR signaling to induce loss of adhesion in triple-negative basal-like breast cancer cells