title: Breast cancer humoral immune response: involvement of Lewis y through the detection of circulating immune complexes and association with Mucin 1 (MUC1).
abstract: BACKGROUND
  In cancer patients, MUC1 glycoprotein may carry Lewis y which could be involved in immune response.
  
  
  OBJECTIVE
  1- to evaluate the presence of Lewis y and MUC1 in circulating immune complexes (Lewis y/CIC and MUC1/CIC, respectively) and their correlation; 2- to analyze the possible presence of Lewis y in carbohydrate chains of tumoral MUC1 glycoprotein and 3- to correlate serum and tissue parameters considered.
  
  
  METHODS
  Pretreatment serum and tissue breast samples from 76 adenocarcinoma, 34 benign and 36 normal specimens were analyzed. Anti-MUC1 and anti-Lewis y MAbs were employed. To detect Lewis y/CIC and MUC1/CIC, ELISA tests were developed; serum samples containing MUC1 were previously selected by Cancer Associated Serum Antigen (CASA). Immunoprecipitation (IP) was performed in 9 malignant, benign and normal samples and analyzed by SDS-PAGE and Western blot. Lewis y and MUC1 expression was studied by immunohistochemistry (IHC). Statistical analysis was performed employing principal component analysis (PCA), ANOVA, Tukey HSD, Chi square test and classical correlation (p < 0.05).
  
  
  RESULTS
  By ELISA, Lewis y/IgM/CIC levels showed statistically significant differences between breast cancer versus benign and normal samples; mean +/- SD values expressed in OD units were: 0.525 +/- 0.304; 0.968 +/- 0.482 and 0.928 +/- 0.447, for breast cancer, benign disease and normal samples, respectively, p < 0.05. Lewis y/IgG/CIC did not show any statistically significant difference. MUC1/IgM/CIC correlated with Lewis y/IgM/CIC. By CASA, 9 samples with MUC1 values above the cut off were selected and IP was performed, followed by SDS-PAGE and Western blot; bands at 200 kDa were obtained with each MAb in all the samples. By IHC, with C14 MAb, 47.5%, 31% and 35% of malignant, benign and normal samples, respectively, showed positive reaction while all the samples were positive with anti-MUC1 MAb; in both cases, with a different pattern of expression between malignant and non malignant samples.
  
  
  CONCLUSIONS
  Our findings support that in breast cancer there was a limited humoral immune response through Lewis y/IgM/CIC levels detection which correlated with MUC1/IgM/CIC. We also found that Lewis y might be part of circulating MUC1 glycoform structure and also that Lewis y/CIC did not correlate with Lewis y expression.
authors.affiliation: Centre for Basic and Applied Immunological Research, Faculty of Medical Sciences, National University of La Plata, Buenos Aires, Argentina. marinaislalarrain@hotmail.com
authors.forename: Marina
authors.initials: M
authors.lastname: Isla Larrain
authors.forename: Sandra
authors.initials: S
authors.lastname: Demichelis
authors.forename: Marina
authors.initials: M
authors.lastname: Crespo
authors.forename: Ezequiel
authors.initials: E
authors.lastname: Lacunza
authors.forename: Alberto
authors.initials: A
authors.lastname: Barbera
authors.forename: Aldo
authors.initials: A
authors.lastname: Cretón
authors.forename: Francisco
authors.initials: F
authors.lastname: Terrier
authors.forename: Amada
authors.initials: A
authors.lastname: Segal-Eiras
authors.forename: María Virginia
authors.initials: MV
authors.lastname: Croce
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000936:Antigen-Antibody Complex / Q000097:blood* / Q000276:immunology; D014408:Biomarkers, Tumor / Q000097:blood / Q000276:immunology; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000097:blood* / Q000276:immunology* / Q000473:pathology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D056724:Immunity, Humoral; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D007983:Lewis Blood Group Antigens / Q000097:blood* / Q000276:immunology; D008875:Middle Aged; D018396:Mucin-1 / Q000097:blood* / Q000276:immunology; D009367:Neoplasm Staging
chemical_list: D000936:Antigen-Antibody Complex; D014408:Biomarkers, Tumor; D007983:Lewis Blood Group Antigens; C094162:Lewis Y antigen; D018396:Mucin-1