Abstract B34: CD56+ immune cell infiltration is decreased in benign breast lobules with fibrocystic changes
Authors: Brahmbhatt Rushin D. , Kerekes Daniel , Hoskin Tanya L. , Pena Alvaro , Visscher Daniel W. , Stallings-Mann Melody L. , Radisky Derek C. , Murphy Linda M. , Pankratz Vernon Shane , Knutson Keith L. , Frost Marlene , Degnim Amy C. 
Published: October 2015

Abstract
Introduction: CD56+ lymphocytes (a defining marker of natural killer cells) have a predominantly cytotoxic phenotype and a hypothetical role in breast tumor immunosurveillance. Here we investigate whether CD56+ staining density varies in nonmalignant breast lobules according to confirmed breast cancer risk factors- age, and histologic features of fibrocystic change and lobular involution.
Methods: Archived breast tissue samples were obtained from 94 women with benign breast disease (BBD). The sample set was selected as a nested case-control study (47 cases, 47 controls) from within a large BBD cohort; cases were those who developed breast cancer subsequent to their benign breast biopsy; controls were matched to cases on age at biopsy, year of biopsy, and length of breast cancer free follow-up at least as long as the time-to-cancer in the matched case. Up to 10 representative lobules in each sample were characterized by H&E for fibrocystic changes and the degree of epithelial proliferation (normal, nonproliferative, proliferative, atypia) and for degree of lobular involution (none, partial, complete). Consecutive sections were immunostained for CD56 and hematoxylin. Using digital image analysis, CD56 staining was quantified on a per lobule basis by pixel ratio (PR) (positive stain:total lobule). Statistical analysis was performed using linear mixed effects regression on the rank transformed density to account for correlations among multiple lobules from the same patient in assessing the association of density with lobule characteristics and using conditional logistic regression for association with case-control status.
Results: Among 94 women (median age 52, range 35-73), 876 lobules were evaluated: 431 in BBD cases and 445 in BBD controls. Overall, 391 lobules (45%) were normal and the remaining 485 (55%) demonstrated fibrocystic changes. Among fibrocystic lobules, 247 (51%) had nonproliferative changes, 218 (45%) had epithelial proliferation, and 20 (4%) had atypical hyperplasia. Among normal lobules, 61 (16%) had no involution, 139 (36%) had partial involution, and 191 (49%) had complete involution. The vast majority of lobules (861/876, 98%) demonstrated presence of CD56+ cells, while only 15 of 876 lobules in 11 patients had zero CD56+ cells [7 cases (15%) versus 4 controls (8.5%) OR 1.75, p = 0.37].
The median CD56+ PR was 0.55% overall. Fibrocystic lobules as a group showed significantly lower median CD56+ PR compared to normal lobules (0.37% vs 0.85%, p &lt; 0.0001). Among subcategories of fibrocystic lobules, the median CD56+ PR was lower in proliferative lobules (0.29%) compared to non-proliferative (0.42%), but this difference was not statistically significant (p = 0.55). Older women (age &gt; 55) showed the highest CD56+ PR (median of 0.92%), as compared to 0.38% in women 45-55 (p=0.02), and 0.52% in women age &lt;45 (p = 0.26). Among normal lobules, median CD56+ PR was similar between lobules with no involution (0.55%) and partial involution (0.52%) but was considerably higher among lobules with complete involution (1.3%, p = 0.09 and p = 0.05, respectively). In a multivariable model, CD56+ PR differences between fibrocystic and normal lobules remained significant (p = 0.0002) and also remained significant between age groups of &gt; 55 and 45-55 (p = 0.02), while involution was non-significant. Median CD56+ PR was lower in cases (0.43%) compared to controls (0.72%), but this was not statistically significant (p=0.19) in our sample of 47 case-control pairs.
Conclusions: Breast lobules with fibrocystic changes show less CD56+ cells, raising a question of possible immune suppression induced by early epithelial abnormalities. These findings encourage further investigation to improve understanding of the immune microenvironment in premalignant breast tissues.
Citation Format: Rushin D. Brahmbhatt, Daniel Kerekes, Tanya L. Hoskin, Alvaro Pena, Daniel W. Visscher, Melody L. Stallings-Mann, Derek C. Radisky, Linda M. Murphy, Vernon Shane Pankratz, Keith L. Knutson, Marlene Frost, Amy C. Degnim. CD56+ immune cell infiltration is decreased in benign breast lobules with fibrocystic changes. [abstract]. In: Proceedings of the Thirteenth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2014 Sep 27-Oct 1; New Orleans, LA. Philadelphia (PA): AACR; Can Prev Res 2015;8(10 Suppl): Abstract nr B34.