The CD44+/CD24- phenotype is enriched in basal-like breast tumors

Abstract

Introduction: Human breast tumors are heterogeneous and consist of phenotypically diverse cells. Breast cancer cells with a CD44+/CD24- phenotype have been suggested to have tumor-initiating properties with stem cell-like and invasive features, although it is unclear whether their presence within a tumor has clinical implications. There is also a large heterogeneity between tumors, illustrated by reproducible stratification into various subtypes based on gene expression profiles or histopathological features. We have explored the prevalence of cells with different CD44/CD24 phenotypes within breast cancer subtypes.

Methods: Double-staining immunohistochemistry was used to quantify CD44 and CD24 expression in 240 human breast tumors for which information on other tumor markers and clinical characteristics was available. Gene expression data were also accessible for a cohort of the material.

Results: A considerable heterogeneity in CD44 and CD24 expression was seen both between and within tumors. A complete lack of both proteins was evident in 35% of the tumors, while 13% contained cells of more than one of the CD44+/CD24-, CD44-/CD24+ and CD44+/CD24+ phenotypes. CD44+/CD24- cells were detected in 31% of the tumors, ranging in proportion from only a few to close to 100% of tumor cells. The CD44+/CD24- phenotype was most common in the basal-like subgroup--characterized as negative for the estrogen and progesterone receptors as well as for HER2, and as positive for cytokeratin 5/14 and/or epidermal growth factor receptor, and particularly common in BRCA1 hereditary tumors, of which 94% contained CD44+/CD24- cells. The CD44+/CD24- phenotype was surprisingly scarce in HER2+ tumors, which had a predominantly CD24+ status. A CD44+/CD24- gene expression signature was generated, which included CD44 and alpha6-integrin (CD49f) among the top-ranked overexpressed genes.

Conclusion: We demonstrate an association between basal-like and particularly BRCA1 hereditary breast cancer and the presence of CD44+/CD24- cells. Not all basal-like tumors and very few HER2+ tumors, however, contain CD44+/CD24- cells, emphasizing that a putative tumorigenic ability may not be confined to cells of this phenotype and that other breast cancer stem cell markers remain to be identified.

Figure 1

Immunohistochemical double-staining of human breast tumors for CD44 and CD24. CD44 is stained with Permanent Red and CD24 with diaminobenzidene (DAB). Magnification × 20. (a) A tumor positive for both CD44⁺/CD24^- (white arrow) and CD44^-/CD24⁺ (black arrow) cancer cells, although the predominant phenotype is CD44^-/CD24^-. (b) Almost all cells in this tumor are CD44⁺/CD24^-. No CD24 staining is seen. (c) A tumor with predominantly CD44⁺/CD24^- cells. A few CD44⁺/CD24⁺ cells are also present (black arrow). (d) A tumor positive for the CD44^-/CD24⁺ phenotype. No CD44 staining is present.

Figure 2

Hierarchical clustering of 69 tumor samples with available gene expression data. Clustering was based on 364 genes from the intrinsic gene list published by Sørlie and colleagues [9] that matched our cDNA clones. Colored boxes indicate classification of each tumor into subtypes/subgroups. Filled or open boxes indicate the percentage of cells in each tumor positive for the CD44⁺/CD24^- and CD44^-/CD24⁺ phenotypes as determined by immunohistochemistry. SR, steroid receptor. Hu classification, Hu and colleagues [10].