title: Annexin-A1 regulates microRNA-26b* and microRNA-562 to directly target NF-κB and angiogenesis in breast cancer cells.
abstract: Annexin 1 (ANXA1) is an endogenous anti-inflammatory protein implicated in cancer. ANXA1 was previously shown to be regulated by hsa-miR-196a. However, whether ANXA1 itself regulates microRNA (miR) expression is unknown. Therefore, we investigated the regulation of miR by ANXA1 in MCF7 breast cancer cells. MCF7-EV (Empty vector) and MCF7-V5 (ANXA1-V5 expressing cells) were subjected to a miR microarray. Microarray analysis revealed a number of miRNAs which were dysregulated in MCF7-V5 cells. 2 novel miRNAs (miR562 and miR26b*) were validated, cloned and functionally characterized. As ANXA1 constitutively activates NF-κB activity to modulate breast cancer metastasis, we found that miR26b* and miR562 directly targeted the canonical NF-κB pathway by targeting the 3' UTR and inhibiting expression of Rel A (p65) and NF-κB1 (p105) respectively. MiR562 inhibited wound healing, which was reversed when ANXA1 was overexpressed. Overexpression of either miR562 or miR26b* in MCF-7 cells enhanced endothelial tube formation when cocultured with human umbilical cord endothelial cells while conversely, treatment of MCF7 cells with either anti-miR562 or anti-miR26b* inhibited endothelial tube formation after co-culture. Further analysis of miR562 revealed that miR562-transfected cell conditioned media enhances endothelial cell tube formation, indicating that miR562 increased angiogenic secreted factors from MCF-7 breast tumor cells. TNFα was increased upon overexpression of miR562, which was reversed when ANXA1 was co-transfected In conclusion, this data suggests that ANXA1-regulated miR26b* and miR562 may play a role in wound healing and tumor-induced endothelial cell tube formation by targeting NF-κB expression and point towards a potential therapeutic target for breast cancer.
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Durkeshwari
authors.initials: D
authors.lastname: Anbalagan
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Gracemary
authors.initials: G
authors.lastname: Yap
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Yi
authors.initials: Y
authors.lastname: Yuan
authors.affiliation: Cancer Science Institute, 14 Medical Drive, #12-01, Centre for Translational Medicine, MD6 Singapore, 117599, Singapore.
authors.forename: Vijay K
authors.initials: VK
authors.lastname: Pandey
authors.affiliation: Cancer Science Institute, 14 Medical Drive, #12-01, Centre for Translational Medicine, MD6 Singapore, 117599, Singapore.
authors.forename: Wai Hoe
authors.initials: WH
authors.lastname: Lau
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Suruchi
authors.initials: S
authors.lastname: Arora
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Pradeep
authors.initials: P
authors.lastname: Bist
authors.affiliation: Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; NUS Immunology Program, Life Sciences Institute, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore.
authors.forename: Justin S B
authors.initials: JS
authors.lastname: Wong
authors.affiliation: Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, MD11, 10 Medical Drive, Singapore, 117597, Singapore.
authors.forename: Gautam
authors.initials: G
authors.lastname: Sethi
authors.affiliation: Astar-Bioprocessing Technology Institute, 20 Biopolis Way, 138668, Singapore, Singapore.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Nissom
authors.affiliation: Cancer Science Institute, 14 Medical Drive, #12-01, Centre for Translational Medicine, MD6 Singapore, 117599, Singapore.
authors.forename: Peter E
authors.initials: PE
authors.lastname: Lobie
authors.affiliation: Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Centre for Life Sciences, 28 Medical Drive, Singapore, 117456, Singapore; Astar-Bioprocessing Technology Institute, 20 Biopolis Way, 138668, Singapore, Singapore.
authors.forename: Lina H K
authors.initials: LH
authors.lastname: Lim
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D017305:Annexin A1 / Q000378:metabolism*; D001483:Base Sequence; D001943:Breast Neoplasms / Q000098:blood supply* / Q000235:genetics* / Q000473:pathology; D003001:Cloning, Molecular; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006801:Humans; D061986:MCF-7 Cells; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D009389:Neovascularization, Pathologic / Q000235:genetics* / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D015203:Reproducibility of Results; D051996:Transcription Factor RelA / Q000378:metabolism; D014945:Wound Healing
chemical_list: D020413:3' Untranslated Regions; D017305:Annexin A1; D035683:MicroRNAs; D016328:NF-kappa B; D051996:Transcription Factor RelA; C528163:MIRN26 microRNA, human; C545728:MIRN562 microRNA, human