Study of Lewis y (Ley) expression and anti Le y immune response through circulating immune complexes (Ley/CIC) detection in breast cancer patients.

ISLA LARRAIN M; CRESPO M; LACUNZA E; BARBERA A

Lyon, France

Congreso; 20th Meeting of the European Association for Cancer Research; 2008

EACR

The aims of this study were: 1-to detect Lewisy (Ley) antigen in breast tumour samples; 2- to determine the role of MUC1 as a carrier of Ley and 3- to investigate the induction of humoral immune response through the detection of circulating immune complexes (CIC) carrying Ley in breast sera. Materials and methods: 137 breast tissue and serum samples: 72 malignant tumours, 30 benign diseases and 35 normals. The expression of Ley was determined by standard immunohistochemistry (IHC); percentage of stained cells, intensity and pattern of the reaction were analyzed. An immunoprecipitation was performed in order to determine if MUC1 may be a possible carrier for Ley. HMFG1, an anti MUC1 monoclonal antibody (MAb) was used to precipitate MUC1 from breast cancer serum samples. Immunoprecipitates (IP) obtained were run in SDS-PAGE and Western blot (WB) assays. Sheets were incubated with C14 (anti Ley) and HMFG1 (anti. MUC1). An ELISA was developed to study the presence of CIC carrying Ley. Briefly C14 was adsorbed at multiwell microplates and incubated overnight at 4ºC. After washing, 1% bovine serum albumin/PBS was added for 3 hours at 37ºC. Serum samples were incubated overnight at 4ºC and 1:2000 anti human IgM or 1:3000 IgG reacted with the complexes and revealed with ABTS and 30% H2O2 in sodium citrate buffer, pH 5.0; OD was measured at 405 nm. Results: By IHC with C14, positive results were found in 34% malignant tumours; 33% benign diseases and 35% normals. Any statistical difference was found. The pattern of expression differed between malignant and non malignant samples: cancer specimens showed more frequently a cytoplasmic and membrane non apical reaction while non malignant samples showed an apical membrane reaction. By WB, IP displayed a band at >200KDa with both C14 and HMFG1. By ELISA, mean OD for IgMCIC in breast cancer sera, benign and normal samples were: 0.538, 0.949 and 0.942, respectively. Significant statistical differences between breast cancer and normal and benign samples were found by ANOVA. Mean OD for IgGCIC were: 0.414, 0.438 and 0.492, respectively. Conclusions: 1- By IHC, non statistical differences among breast cancer, benign and normal samples were found although a different pattern of expression was observed; 2- Immunoprecipitation and WB indicate that MUC1 may behave as a possible carrier for Ley in breast cancer; and 3- a significant statistical difference between breast cancer and normal and benign samples were found in IgMCIC.