2008 Summer Meeting - Leeds - The Pathological Society of Great ...

P131Immunophenotype of Ductal Carcinoma in Situ in BRCAGermline Mutation CarriersP Van Der Groep* 1 , PJ Van Diest* 1 , E Van Der Wall# 11 University Medical Centre Utrecht, *Dept of Pathology and #Division ofInternal Medicine and Dermatology,The NetherlandsBackground: Germline BRCA1 related breast cancers have a distinctbasal/triple negative immunophenotype, and show EGFR and HIF1 expression.Little is known about the immunophenotype of precursor lesions in BRCA1/2germline mutation carriers. The aim of this study was to examine whether thischaracteristic phenotype is already present in the pre- invasive stage.Material and Methods: DCIS of 6 proven BRCA1 and 4 BRCA2 germlinemutation carriers were stained by immunohistochemistry for ER, PR, HER-2/neu, Ck5/6, Ck14, EGFR and Ki67.Results: 4/11 cases (36%) were ER positive, 0/7 (0%) were PR positive, 0/10(0%) were HER2 positive, 5/10 (50%) were CK5/6 positive, 1/9 (11%) wereCK14 positive, and 6/10 (60%) were EGFR positive. Mean percentage Ki67nuclear staining was 30% (range 0-100). These percentages are similar to thosethat have been reported for invasive cancers in BRCA1/2 mutation carriers,except for ER that is generally even lower in BRCA1/2 related cancers.Discussion: DCIS in BRCA1/2 germline mutation carriers shows a so calledbasal immunophenotype with high proliferation and EGFR positivity similar tothat of invasive cancers in such patients. This may be useful to identify“BRCA-ness” in cases of DCIS in diagnostic pathology, and opens up newways for targeted therapy against EGFR to prevent development of invasivecancer in case of a germline mutation.P133Perinecrotic HIF-1 Expression and Necrosis PredictPrognosis in Patients with Endometrioid EndometrialCarcinomaLMS Seeber# 1 , N Horrée# 1 , P Van Der Groep* 1 ,RHM Verheijen# 1 , PJ Van Diest* 1 .1 University Medical Centre Utrecht, Departments of #SurgicalGynecology and Oncology, and *Pathology, The NetherlandsBackground. Hypoxia-inducible factor 1 (HIF-1) plays an essential role inthe adaptive response of cells to hypoxia, triggering biologic events associatedwith aggressive tumour behaviour. Hypoxia and its key regulator HIF-1 playan important role in endometrial carcinogenesis, but contradictory results havebeen published as to the prognostic value of HIF-1 expression in endometrialcarcinoma. We therefore re-evaluated the prognostic value of HIF-1expression in a large representative group of endometrioid endometrial cancerusing well-established methodology.Methods. In 98 patients with endometrioid endometrial cancer, expressionlevels of HIF-1 and p27 were analyzed by immunohistochemistry. Presence ofnecrosis, and type of HIF-1 expression (perinecrotic, diffuse, or mixed) werenoted.Results. Stage, grade and depth of invasion showed prognostic value asexpected. Indicators of poor prognosis were presence of necrosis (p=0.05) andperinecrotic type of HIF-1 expression (p=0.03). In patients with perinecrotictype of HIF-1 expression, high p27 expression was an additional prognosticfactor. In Cox regression, HIF-1 was an additional prognostic factor to stage.Conclusion. In patients with endometrioid endometrial cancer, necrosis andnecrosis related expression of HIF-1 are important prognostic factors. In viewof the proposed role of hypoxia and HIF-1 in endometrial cancer, HIF-1 isthereby an attractive therapeutic target.P132Hypoxia-Inducible Factor 1a is Essential for Hypoxic p27Induction in Endometrioid Endometrial CarcinomaN Horrée# 1 ,EH Gort* 1 , P Van Der Groep* 1 , APM Heintz# 1 ,M Vooijs* 1 , PJ Van Diest* 11 University Medical Centre Utrecht, *Departments of Pathology and#Surgical Gynaecology and Oncology,The NetherlandsHypoxia-inducible factor 1 (HIF-1) plays an essential role in the cellularadaptive hypoxia response. The cyclin-dependent kinase inhibitor p27(Kip1) ishighly expressed in the normal endometrium but is lost during endometrialcarcinogenesis. However, in high-grade cancers, p27 re-expression is observed.We analysed the role of HIF-1 in hypoxia-induced expression of p27 inendometrial cancer. Paraffin-embedded specimens from 39 endometrioidendometrial carcinomas were immunohistochemically stained for HIF-1, p27,and Ki67. HEC1B, an endometrial carcinoma cell line, was cultured undernormoxic or hypoxic conditions in the presence or absence of transientlyexpressed shRNAs targeting HIF-1. Protein expression of p27 and HIF-1 wasassessed by western blotting.Immunohistochemical staining revealed perinecrotic HIF-1 expression in 67%of the cases and p27 staining centrally in the tumour islands, mostly aroundnecrosis, in 46% of the cases. In 50% of the tumours with perinecrotic HIF-1expression, p27 and HIF-1 perinecrotic/central co-localization was observed.Hypoxia-associated p27 expression showed less proliferation around necrosis.In HEC1B, p27 protein expression was induced by hypoxia. This induction wasabrogated by transient knockdown of HIF-1 using RNAi. Furthermore,hypoxia induced cell cycle arrest in HEC1B cells. We conclude that, inendometrioid endometrial carcinoma, p27 re-expression by hypoxia is HIF-1dependentand leads to cell cycle arrest. This may contribute to the survival ofcancer cells in hypoxic parts of the tumour.P134Nitric oxide down-regulates expression of the haemoglobinhaptoglobinscavenger receptor (CD163) on human monocyte /macrophagesG Hutchins 1 ,PM Guyre 2 , NJ Goulding 31 Dept. Histopathology, Leeds Teaching Hospitals NHS Trust, Leeds, UK2 Dept. Microbiology and Immunology, Darmouth Medical School,Lebanon, NH, USA, 3 William Harvey Research Institute, Barts and TheLondon, Queen Mary's School of Medicine and Dentistry, LondonNitric oxide (NO) mediates many effects on immune system function. Althoughexaggerated NO production is well-characterised in many pathological states,it’s effect on the expression and function of the haemoglobin-haptoglobinscavenger receptor (CD163) is unknown.Human monocytes were isolated by density centrifugation andsubsequently exposed in 18-24 hour cultures to the NO generator DETA-NONOate. Co-incubation with factors known to promote CD163 expressionwas also performed. Metalloproteinase inhibitors were utilised to assessshedding as possible regulatory mechanism. CD163 expression was quantifiedby flow cytometry with supernatant soluble CD163 concentrations determinedby ELISA. Post-incubatory cell viability was confirmed by metabolic capacityand CD14 expression. CD163 expression was also evaluated followingexposure to the guanylate cyclase activator 8-Br-cGMP.Nitric oxide downregulated monocyte CD163 expression by upto 70% atmaximal concentrations. Similar attenuation was observed following coexposureto both NO and interleukin-10 or dexamethasone. CD163 expressionwas downregulated by 24% through NO exposure following super-induction ofCD163 expression by co-incubation with IL-10 and dexamethasone. Nitricoxide had no effect on cell viability but did induce a reduction in solubleCD163 detected in the culture supernatants relative to controls, thus excludingshedding as a downregulatory mechanism. The guanylate cyclase activator, 8-Br-cGMP, also induced downregulation of CD163, indicating a possible role ofguanylyl cyclase in the downregulatory process.This study has established a role of nitric oxide in regulating expression ofCD163, possibly through activation of guanylate cyclase.Summer Meeting (194 th ) 1–4 July 2008 Scientific Programme65