P2-01-14: High Level of CD44 Expression and Soluble CD44 Secretion Mediate Trastuzumab Resistance in HER2 Positive Breast Cancer.

Abstract

Abstract Background: Overexpression of HER2 occurs in 15–25% of invasive breast cancer. Trastuzumab, a humanized monoclonal antibody targeting the extracellular domain of HER2, markedly improves disease-free and overall survival in patients with HER2−positive breast cancer. However, resistance to trastuzumab develops in most patients with HER2−overexpressing metastatic breast cancer. Breast cancer stem cells have been shown to express HER2, although its role in the development or maintenance of stem cells is not well characterized. CD44 is one of the proposed markers for breast cancer stem cells. We investigated the role of CD44 in acquired trastuzumab resistance in HER2 positive breast cancer. Methods: CD44 mRNA expression was measured by quantitative real-time PCR. The proportion of CD44 positive cells was measured by fluorescence-activated cell sorting (FACS). Soluble CD44 level was detected by enzyme-linked immunosorbent assay (ELISA). In vitro tumorigenicity was evaluated by soft agar colony formation assay. Cell invasion was tested by using a matrigel invasion assay. In vivo tumor growth was evaluated using a mammary fat pad xenograft mouse model. Results: CD44 mRNA expression, CD44 positive cell population by FACS, and CD44 secretion were significantly increased in both SKBR3 derived trastuzumab resistant SKBR3 clone 3 and BT474 derived trastuzumab resistant HR20 cells compared to parental, trastuzumab sensitive cells. Consistent with the increase in CD44, trastuzumab resistant cells showed an increase in soft agar colony formation and cell invasion, which were blocked by siRNA-induced downregulation of CD44 expression. Tumor growth and incidence were enhanced in mice injected with trastuzumab resistant HR20 cells compared to the mice injected with trastuzumab sensitive parental BT474 cells. Knock-down of CD44 expression by the transfection with CD44-specific siRNA suppressed in vivo tumor growth in the mice injected with trastuzumab resistant HR20 cells. Finally, we found high serum CD44 levels correlated with advanced tumor progression in HER2 positive breast cancer. Conclusion: Our findings indicate that high levels of CD44 mediate trastuzumab resistance in HER2 positive breast cancer cells as well as tumor progression in mice. Circulating CD44 is prognostic in patients with HER2 positive breast cancer. CD44 may be a therapeutic target for HER2 positive breast cancer. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P2-01-14.