Type I receptor tyrosine kinases, including the epidermal growth factor receptor (EGFR) and erbB2, have been implicated in mammary carcinoma growth and metastasis. Recent evidence suggests that type I receptor signaling may be mediated by the CD44 family of transmembrane glycoproteins that also have been implicated in mammary tumor progression. Here, the authors tested whether CD44, EGFR, and erbB2 interacted and colocalized with one another in four mammary carcinoma cell lines (MCF-7, MDA-MB-231, MDA-MB-435, and MDA-MB-436) and in cytology samples obtained from patients with metastatic breast cancer. CD44 constitutively colocalized and coimmunoprecipitated with erbB2 and EGFR in all four mammary carcinoma cell lines. CD44 also colocalized with erbB2 and EGFR in all cytology samples expressing erbB2. CD44 colocalized with EGFR in cells from only 1 of 16 erbB2-negative cytology samples. These data indicate that CD44-EGFR-erbB2 protein complexes occur in a high proportion of metastatic mammary carcinomas and suggest that CD44-type I receptor colocalization may be a novel prognostic marker for aggressive mammary cancers.