O1FABP7 Expression Predicts Better Outcome in Basal-LikeBreast CancerHZhang 1 ,EA Rakha 1 , GR Ball 2 , EC Paish 1 , C Caldas 3 , IO Ellis 1 ,AR Green 11 Division <strong>of</strong> Pathology, School <strong>of</strong> Molecular Medical Sciences,Nottingham University Hospitals and University <strong>of</strong> Nottingham,Nottingham, 2 School <strong>of</strong> Biomedical and Natural Sciences, NottinghamTrent University, Nottingham, 3 Cancer Research UK, CambridgeResearch Institute, Li Ka-Shing Centre, Cambridge<strong>The</strong> recently identified basal-like cancers (BP) have attracted attention as a poorprognostic class <strong>of</strong> breast cancer. However, BP appear to encompassbiologically and clinically heterogeneous tumours; resulting in the lack <strong>of</strong> aconsensus definition <strong>of</strong> BP.In order to refine BP, we analysed expression <strong>of</strong> 48,000 gene transcripts in 132breast carcinomas using Artificial Neural Network analysis and identified twogenes (OATP2 and FABP7) associated with BP (defined by cytokeratin(CK)5/6 and/or CK14 positivity). Using a large invasive breast carcinomacohort (n=900), prepared as TMA, we assessed the immunohistochemicalexpression <strong>of</strong> OATP2 and FABP7 and investigated their association withclinicopathological variables, patients’ outcome and ability to refine BPclassification.A total <strong>of</strong> 8.9% and 15.6% cases were OATP2 and FABP7 positiverespectively. OATP2 was associated with high grade tumours (p
O5An Altered Myoepithelial Cell Phenotype in DCIS InfluencesTumour-Associated AngiogenesisSJL Payne 1 ,MAllen 1 ,IHart 1 , JL Jones 11 Queen Mary University London<strong>The</strong> process <strong>of</strong> angiogenesis is a critical factor in breast tumour progression andhas also been demonstrated in preneoplastic breast lesions.Limited published data suggests that myoepithelial cells are anti-angiogenic.This observation is based on ‘normal’ myoepithelial cells. It is recognised,however, that myoepithelial cells are altered in DCIS. We have previouslydemonstrated a difference characterised by the up-regulation v6. This studyaimed to investigate whether expression <strong>of</strong> v6 in DCIS is associated withangiogenesis, and to establish if v6 expression could contribute to enhancedangiogenesis.Immunohistochemistry for v6 and the vascular marker CD31 was carried outon tissue microarrays (TMAs) <strong>of</strong> 148 cases <strong>of</strong> pure DCIS, and on wholesections from 36 <strong>of</strong> these cases. TMAs were found to be <strong>of</strong> limited use inanalysis. In whole sections, 47% expressed v6 with 53% negative for v6.<strong>The</strong> angiogenic pattern was scored as rim pattern, where vessels are adjacent tothe DCIS ducts and diffuse pattern, where vessels are present in the surroundingstroma. A rim pattern <strong>of</strong> staining was only seen in the v6 positive group.Using an immortalised myoepithelial cell line (1089) and an v6 overexpressingcounterpart (6-1089), we demonstrated that 6-1089 cells releasehigher levels <strong>of</strong> VEGF than 1089 cells and also stimulated more vascularsprouting in a mouse aortic ring assay. This supports the hypothesis that upregulation<strong>of</strong> v6 on myoepithelial cells in DCIS is <strong>of</strong> functional significanceand may contribute to invasive progression.O7Neural networks identify gene pr<strong>of</strong>iles that predict clinicaloutcome with high accuracy in breast cancer: CA IX isidentified as the principal prognostic indicatorLJ Lancashire 1 , DG Powe 2 ,ERakha 2 , AR Green 2 ,RMukta 2 ,EC Paish 2 , RC Rees 3 , IO Ellis 2 , GR Ball 31 Clinical and Experimental Pharmacology, Paterson Institute for CancerResearch, 2 Department <strong>of</strong> Histopathology, Nottingham UniversityHospitals Trust and University <strong>of</strong> Nottingham, 3 <strong>The</strong> Nottingham TrentUniversity, School <strong>of</strong> Science and TechnologyGene expression microarrays allow for the high throughput analysis <strong>of</strong>hundreds <strong>of</strong> thousands <strong>of</strong> gene transcripts. This technology has been applied tothe molecular classification <strong>of</strong> cancer patients. Subsequently, it is recognisedthat different patient subgroups show distinct molecular signaturescorresponding to cancer-associated disease pathways. Importantly, these genepr<strong>of</strong>iles provide information on clinical outcome. A potential handicap <strong>of</strong> suchhigh throughput technologies is the unprecedented amount <strong>of</strong> data generated,<strong>of</strong>ten <strong>of</strong> high complexity. This has created a demand for novel data analysismethodologies. Here, we present an Artificial Neural Network basedmethodology that has identified a prognostic gene expression signature in breastcancer. In applying this approach to a previously published seminal data set wehave identified nine key genes whose expression pr<strong>of</strong>iles are capable <strong>of</strong>predicting development <strong>of</strong> distant metastases to accuracies <strong>of</strong> 98%. <strong>The</strong>principal prognostic indicator identified was carbonic anhydrase IX (CA IX), acomponent <strong>of</strong> the hypoxia-pathway. We subsequently tested the suitability <strong>of</strong>CA IX for predicting survival outcome using semi-quantitativeimmunohistochemical morphometry on a large number <strong>of</strong> unselected breastcancers. Interestingly, our results showed that membranous CA IX expression ispredictive <strong>of</strong> shortened overall survival, reduced disease free interval, andlymph node involvement.O6Genomic and Immunophenotypical Characterisation <strong>of</strong> PureMicropapillary Carcinomas <strong>of</strong> the BreastCMarchio 1 ,MIravani 1 , R Natrajan 1 , MB Lambros 1 , K Savage 1 ,S Di Palma 2 , FC Schmitt 3 , G Bussolati 4 , IO Ellis 5 , A Ashworth 1 ,ASapino 4 , JS Reis-Filho 1 .1 <strong>The</strong> Breakthrough Breast Cancer Research Centre – Institute <strong>of</strong> CancerResearch, London, 2 RSCH, Guildford, 3 IPATIMUP, Porto, Portugal4 Department <strong>of</strong> Biomedical Sciences and Human Oncology, University <strong>of</strong>Turin, Torino, Italy, 5 Department <strong>of</strong> Histopathology, Nottingham CityHospital NHS Trust and University <strong>of</strong> Nottingham, NottinghamPure invasive micropapillary carcinoma (MPC) is a special type <strong>of</strong> breastcancer characterised by a distinctive growth pattern and a more aggressiveclinical behaviour than invasive ductal carcinomas <strong>of</strong> no special type (IDC-NSTs). To define the molecular characteristics <strong>of</strong> MPCs, we pr<strong>of</strong>iled 12 MPCsand 24 grade and oestrogen receptor (ER)-matched IDC-NSTs using microarraycomparative genomic hybridisation (aCGH). In addition, we generated a tissuemicroarray containing a series <strong>of</strong> 24 MPCs and performedimmunohistochemistical analysis with ER, PR, Ki67, HER2, CK5/6, CK14,CK17, EGFR, topoisomerase-II, cyclin D1, caveolin-1, E-cadherin and -catenin antibodies. In situ hybridisation probes were employed to evaluate theprevalence <strong>of</strong> amplification <strong>of</strong> HER2, TOP2A, EGFR, CCND1, MYC, ESR1and FGFR1 genes. aCGH analysis demonstrated that MPCs significantlydiffered from IDC-NSTs at the genomic level. Gains <strong>of</strong> 1q, 2q, 4p, 6p, 6q23,7p, 7q, 8p, 8q, 9p, 10p, 11q, 12p, 12q, 16p, 17p, 17q, 19p, 20p, 20q and 21qand losses <strong>of</strong> 1p, 2p, 6q11, 6q21, 9p, 11p, 15q and 19q were more prevalent inMPCs. Amplifications <strong>of</strong> 8p12, 8q12, 8q13, 8q21, 8q23, 8q24, 17q21, 17q23and 20q13 were significantly associated with MPCs. A comparison between 24MPCs and a series <strong>of</strong> 48 grade and ER matched IDC-NSTs revealed that highcyclin D1 expression, high proliferation rates and MYC amplification weresignificantly associated with MPCs. Our results demonstrate that MPCs havedistinctive histological features and molecular genetic pr<strong>of</strong>iles supporting thecontention that they constitute a distinct pathological entity.O8Clear Cells <strong>of</strong> the Nipple Epidermis: a Reserve <strong>of</strong> MotileMammary Precursor Cells?JGoing 1 ,MCurrie 11 University <strong>of</strong> GlasgowBackground: Clear cells <strong>of</strong> the nipple epidermis [Toker, Cancer 1970] have noknown function but, with a relationship to mammary duct ostia andultrastructural features in common with light cells <strong>of</strong> the mammary epithelium,it has been proposed that they migrate into nipple epidermis from subjacentducts [Marucci, Virchows Archiv 2002].Aims and methods: We examined clear cell morphology and relationships inthe nipple in 30 mastectomy breasts. Paraffin sections were immunostained forcytokeratins 7+8, 14 and progesterone receptor (PR).Results: Ck 7/8 and PR positive clear cells clustered around duct ostia wereabundant in 6, fairly numerous in 7, scanty in 10 and absent in 7 cases. <strong>The</strong>ymay be single, clumped or form hollow spheroids. Some have a 'foot' onbasement membrane. A subset show structural features implying motility. Athin protrusion inserted with the aid <strong>of</strong> a leading filopodium (microspike)between adjacent keratinocytes is anchored by a rounded, club-like termination.<strong>The</strong> protrusion thickens, most strikingly at its far end, and adopts a flattened,lamellipodium-like contact pr<strong>of</strong>ile with adjacent keratinocytes beforeshortening to draw the cell body in the direction <strong>of</strong> travel.Conclusions: Clear cells <strong>of</strong> the nipple epidermis are associated with ductopenings and combine a lack <strong>of</strong> differentiated features with apparent motility.We propose that they represent a reserve <strong>of</strong> mammary progenitor cells, and thattrafficking <strong>of</strong> precursor cells between separate mammary lobes via the nippleepidermis is a possible mode <strong>of</strong> mammary gland maintenance over repeatedcycles <strong>of</strong> pregnancy and lactation.<strong>Summer</strong> <strong>Meeting</strong> (194 th ) 1–4 July <strong>2008</strong> Scientific Programme79
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