title: Integrin αVβ3 silencing sensitizes malignant glioma cells to temozolomide by suppression of homologous recombination repair.
abstract: Integrins have been suggested as possible targets in anticancer therapy. Here we show that knockdown of integrins αVβ3, αVβ5, α3β1 and α4β1 and pharmacological inhibition using a cyclo-RGD integrin αVβ3/αVβ5 antagonist sensitized multiple high-grade glioma cell lines to temozolomide (TMZ)-induced cytotoxicity. The greatest effect was observed in LN229 cells upon integrin β3 silencing, which led to inhibition of the FAK/Src/Akt/NFκB signaling pathway and increased formation of γH2AX foci. The integrin β3 knockdown led to the proteasomal degradation of Rad51, reduction of Rad51 foci and reduced repair of TMZ-induced DNA double-strand breaks by impairing homologous recombination efficiency. The down-regulation of β3 in Rad51 knockdown (LN229-Rad51kd) cells neither further sensitized them to TMZ nor increased the number of γH2AX foci, confirming causality between β3 silencing and Rad51 reduction. RIP1 was found cleaved and IκBα significantly less degraded in β3-silenced/TMZ-exposed cells, indicating inactivation of NFκB signaling. The anti-apoptotic proteins Bcl-xL, survivin and XIAP were proteasomally degraded and caspase-3/-2 cleaved. Increased H2AX phosphorylation, caspase-3 cleavage, reduced Rad51 and RIP1 expression, as well as sustained IκBα expression were also observed in mouse glioma xenografts treated with the cyclo-RGD inhibitor and TMZ, confirming the molecular mechanism in vivo. Our data indicates that β3 silencing in glioma cells represents a promising strategy to sensitize high-grade gliomas to TMZ therapy.
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Christmann
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Diesler
authors.affiliation: Laboratory for Cell Biology and Signaling, Division of Molecular Biology, Ruđer Bošković Institute, HR-10000 Zagreb, Croatia.
authors.forename: Dragomira
authors.initials: D
authors.lastname: Majhen
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Steigerwald
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Nancy
authors.initials: N
authors.lastname: Berte
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Halima
authors.initials: H
authors.lastname: Freund
authors.affiliation: Laboratory for Cell Biology and Signaling, Division of Molecular Biology, Ruđer Bošković Institute, HR-10000 Zagreb, Croatia.
authors.forename: Nikolina
authors.initials: N
authors.lastname: Stojanović
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Kaina
authors.affiliation: Laboratory for Cell Biology and Signaling, Division of Molecular Biology, Ruđer Bošković Institute, HR-10000 Zagreb, Croatia.
authors.forename: Maja
authors.initials: M
authors.lastname: Osmak
authors.affiliation: Laboratory for Cell Biology and Signaling, Division of Molecular Biology, Ruđer Bošković Institute, HR-10000 Zagreb, Croatia.
authors.forename: Andreja
authors.initials: A
authors.lastname: Ambriović-Ristov
authors.affiliation: Department of Toxicology, University Medical Center Mainz, D-55131 Mainz, Germany.
authors.forename: Maja T
authors.initials: MT
authors.lastname: Tomicic
mesh_terms: D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001932:Brain Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D003606:Dacarbazine / Q000031:analogs & derivatives / Q000494:pharmacology / Q000627:therapeutic use; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D005909:Glioblastoma / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D039661:Integrin beta3 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D000072000:NF-KappaB Inhibitor alpha / Q000378:metabolism; D060787:Neoplasm Grading; D010456:Peptides, Cyclic / Q000494:pharmacology; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000378:metabolism; D019030:Receptors, Vitronectin / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D059767:Recombinational DNA Repair; D015398:Signal Transduction; D000077204:Temozolomide; D023041:Xenograft Model Antitumor Assays
chemical_list: D018906:Antineoplastic Agents, Alkylating; D051017:Apoptosis Regulatory Proteins; C061248:ITGB3 protein, human; D039302:Integrin alphaVbeta3; D039661:Integrin beta3; C000606204:NFKBIA protein, human; D010456:Peptides, Cyclic; D034741:RNA, Small Interfering; D019030:Receptors, Vitronectin; C507169:cyclic arginine-glycine-aspartic acid peptide; C064660:integrin alphaVbeta5; D000072000:NF-KappaB Inhibitor alpha; D003606:Dacarbazine; D051057:Proto-Oncogene Proteins c-akt; C506249:RIPK1 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C495153:RAD51 protein, human; D051135:Rad51 Recombinase; D000077204:Temozolomide
keywords: Rad51; homologous recombination repair; integrin αVβ3 silencing; malignant gliomas; temozolomide