Human Cancer Biology Gene Expression Profiling of Bone Marrow Endothelial Cells in Patients with Multiple Myeloma

Purpose: To determine a “gene/molecular fingerprint” of multiple myeloma endothelial cells and identify vascular mechanisms governing the malignant progression from quiescent monoclonal gammopathy of undetermined significance. Experimental Design: Comparative gene expression profiling of multiple myeloma endothelial cells and monoclonal gammopathy of undetermined significance endothelial cells with the Affymetrix U133A Arrays was carried out in patients at diagnosis; expression and function of selective vascular markers was validated by real-time reverse transcriptase-PCR, Western blot, and small interfering RNA analyses. Results: Twenty-two genes were found differentially expressed (14 down-regulated and eight up-regulated) at relatively high stringency in multiple myeloma endothelial cells compared with monoclonal gammopathy of undetermined significance endothelial cells. Functional annotation revealed a role of these genes in the regulation of extracellular matrix formation and bone remodeling, cell adhesion, chemotaxis, angiogenesis, resistance to apoptosis, and cell-cycle regulation. Validation was focused on six genes (DIRAS3, SERPINF1, SRPX, BNIP3, IER3, and SEPW1) not previously found to be functionally correlated to the overangiogenic phenotype of multiple myeloma endothelial cells in active disease. The small interfering RNA knockdown of BNIP3, IER3, and SEPW1 genes affected critical multiple myeloma endothelial cell functions correlated with the overangiogenic phenotype. Conclusions: The distinct endothelial cell gene expression profiles and vascular phenotypes detected in this study may influence remodeling of the bone marrow microenvironment in patients with active multiple myeloma. A better understanding of the linkage between plasma cells and endothelial cells in multiple myeloma could contribute to the molecular classification of the disease and thus pinpoint selective gene targets for more effective antiangiogenic treatments. (Clin Cancer Res 2009;15(17):5369–78) The unique markers expressed by tumor vasculature distinguish it from normal endothelium. These abnormalities reflect the pathologic nature of its induction and attempts to discover tumor endothelial cell markers have always been hampered by technical difficulties in isolating functionally intact and phenotypically stable endothelial cells from patient samples. St. Croix et al. (1) were the first to show that colorectal cancer endothelial cells overexpress specific transcripts as a result of qualitative Authors' Affiliations: Departments of Internal Medicine and Clinical Oncology and Human Anatomy and Histology, University of Bari Medical School; Hematology Unit, Institute of Oncology “Giovanni Paolo II,” Bari, Italy; Clinical Proteomics Unit, “Vito Fazzi” Hospital, University of Salento, Lecce, Italy; Hematology Unit, Hospital of Cosenza, Cosenza, Italy; Department of Medical Sciences, University of Milan, and Hematology 1, Fondazione IRCCS Policlinico MaRe, Milan, Italy; and Division of Hematology, Department of Cellular Biotechnology and Hematology, University “La Sapienza” Medical School, Rome, Italy Received 1/8/09; revised 5/20/09; accepted 5/27/09; published OnlineFirst 8/18/09. Grant support: Associazione Italiana per la Ricerca sul Cancro, Milan (A. Vacca and A. Neri); the Ministry of Education, University and Research (Progetti di Ricerca di Interesse Nazionale Projects 2008); the Ministry of Health; Progetto Oncologia 2006, Humanitas Mirasole Società per Azioni, Rome; and Amelia Scorza Onlus Foundation, Cosenza, Italy; and a fellowship of the Fondazione Italiana per la Ricerca sul Cancro, Milan (K. Todoerti). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). R. Ria and K. Todoerti contributed equally to the work. Current address for M. Mattioli: Department of Biomedical Sciences and Technologies, University of Milan, Milan, Italy. The sponsors of this study are public or nonprofit organizations that support science in general; they had no role in gathering, analyzing, or interpreting the data. Requests for reprints: Angelo Vacca, Department of Internal Medicine and ClinicalOncology,Policlinico–PiazzaGiulioCesare, 11, I-70124Bari, Italy. Phone: 39-080-559-34-44; Fax: 39-080-559-21-89; E-mail: [email protected] and Antonino Neri, Department of Medical Sciences, University of Milan, Istituto di Ricovero e Cura a Carattere Scientifico PoliclinicoMangiagalli e Regina Elena, Via Francesco Sforza 35, 20122, I-20122 Milan, Italy. Phone: 39-0255033328; Fax: 39-02-50320403; E-mail: [email protected]. F 2009 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-09-0040 5369 Clin Cancer Res 2009;15(17) September 1, 2009 www.aacrjournals.org Cancer Research. on October 15, 2017. © 2009 American Association for clincancerres.aacrjournals.org Downloaded from Published OnlineFirst August 18, 2009; DOI: 10.1158/1078-0432.CCR-09-0040