Prominent Microvascular Proliferationin Clinically Aggressive Neuroblastoma RadhikaPeddinti, 1 RanaZeine, 2 DragosLuca, 5 RoopaSeshadri, 1 AlexandreChlenski, 2 KristinaCole, 4 BrucePawel, 4 HelenR.Salwen, 2 JohnM.Maris, 4 andSusanL.Cohn 3 Abstract Purpose: Tumor vasculature is disorganizedand glomeruloid microvascular proliferation (MVP) hasbeenidentifiedasapoorprognosticatorinsomeadultcancers.Todeterminetheclinicalsig- nificance of MVP, including glomeruloid MVP inneuroblastoma, we initially examined vessel architecture in tumor sections from 51children diagnosed at Children’s Memorial Hospital (CMH) and subsequently evaluated154 neuroblastoma tumors on a tissue microarray con- structedatChildren’sHospitalofPhiladelphia(CHOP). Experimental Design: H&Esectionswereexaminedforthepresenceofstructurallyabnormal vesselsandfurthercharacterizedbyimmunostainingforCD31andvonWillebrandfactortohigh- lightendothelialcellsand a-smoothmuscleactinforpericytes.Tumorswiththickenedwallscon- taining a complete layer of hypertrophic endothelial cells plus additional layers of vascular mural cellswereclassifiedasMVPpositive.AssociationsbetweenMVPandestablishedclinicopatho- logicfeaturesandoutcomewereassessed. Results: Inbothseries,MVPwassignificantlyassociatedwithSchwannianstroma-poorhistol- ogy (CMH, P = 0.008; CHOP, P < 0.001) and decreased survivalprobability (CMH, P = 0.017; CHOP, P = 0.014). In the CHOP series, MVP was associated withhigh-risk group classification (P < 0.001),althoughthisassociationwasnotseeninthesmallerCMHcohort. Conclusions: TheassociationbetweenMVPandpooroutcomeprovidesfurthersupportforthe conceptthatangiogenesisplaysanimportantroleindeterminingthebiologicalbehaviorofneu- roblastomatumors.OurresultsalsoindicatethatangiogenesisisregulateddifferentlyinSchwan- nian stroma-rich versus stroma-poor neuroblastoma tumors. Further studies investigating the activityofangiogenicinhibitorsinchildrenwithclinicallyaggressivestroma-poorneuroblastoma arewarranted. Neuroblastoma, the most common extracranial solid tumor in children, is unique among pediatric cancers for its broad spectrum of clinical behavior (1). This tumor is composed of two main cell types, neuroblastic/ganglion cells and Schwann cells, and the amount of Schwannian stroma strongly affects prognosis (2). Favorable outcome is associated with tumors with abundant Schwannian stroma. In addition to tumor histology, various genetic and biological factors have been shown to correlate with outcome. However, the cellular mechanisms that underlie the clinical variability observed in neuroblastoma remain largely unknown. It is widely accepted that malignant solid tumors must acquire a new blood supply for their progressive growth to a clinically relevant size and for metastasis (3, 4). In contrast to normal vessels, tumor vasculature tends to be disorganized with vessels that are dilated and tortuous and have uneven diameters and excessive branching (5). Microvascular proliferation (MVP) ranges from slight to extreme degrees. Focal proliferative buddings of endothelial cells resembling a renal glomerulus [glomeruloid MVPs (GMP)] have been described in some types of adult cancers, and several studies have shown an association between the presence of GMP and shorter survival (6–8). Wesseling et al. (9) have shown by electron microscopy that the process of MVP involves not only endothelial cell proliferation but also a significant contribution from pericytes. In the WHO grading of astrocytic neoplasms, GMP is a criterion for increasing the tumor grade (10). However, the clinical significance of MVP has not been evaluated previously in neuroblastoma or other pediatric cancers. Human Cancer Biology Authors’Affiliations: 1 DepartmentofPediatrics, Children’s Memorial Hospital; 2 RobertH.LurieComprehensiveCancerCenter,NorthwesternUniversity; 3 Institute forMolecularPediatricSciences,UniversityofChicago,Chicago,Illinois; 4 Department ofPediatrics,Children’sHospitalofPhiladelphiaandtheUniversityofPennsylvania SchoolofMedicine, Philadelphia,Pennsylvania; and 5 Departmentof Pathology, Children’sHospitalsandClinicsofMinnesota,Minneapolis,Minnesota Received2/2/07;revised3/14/07;accepted3/29/07. Grant support: Clinical Oncology ResearchTraining grantT32 CA079447, NIH grants R01NS049814 and P01CA97323, the Neuroblastoma Children’s Cancer Society, Friends for Steven Pediatric Cancer Research Fund, the EliseAnderson Neuroblastoma Research Fund, Neuroblastoma Kids, Alex’s Lemonade Stand Foundation, andThe Robert H. Lurie Comprehensive Cancer Center, NIH, National CancerInstituteCoregrant5P30CA60553. Thecostsofpublicationofthisarticleweredefrayedinpartbythepaymentofpage charges.Thisarticlemustthereforebeherebymarked advertisement inaccordance with18U.S.C.Section1734solelytoindicatethisfact. Requests for reprints: Susan L. Cohn, Clinical Sciences, Institute for Molecular Pediatric Sciences, University of Chicago, 5841Maryland Avenue, MC 4060, Room N114, Chicago, IL 60637. Phone: 773-702-2571; Fax: 773-834-1329; E-mail:scohn@peds.bsd.uchicago.edu. F 2007AmericanAssociationforCancerResearch. doi:10.1158/1078-0432.CCR-07-0237 www.aacrjournals.org Clin Cancer Res 2007;13(12) June15, 2007 3499