Clusterin (CLU) is a heterodimeric secreted glycoprotein implicated in several physiological and pathological processes including cancer. Although recent data showed that overexpression of CLU is closely associated with disease progression in patients with breast tumor, the functional role of CLU expression in this tumor hystotype remains to be determined. The objectives in this study were to evaluate CLU expression levels after treatment with Trastuzumab, a HER2-targeted monoclonal antibody used in the clinical management of advanced breast cancer patients, and to test the usefulness of combined treatment with OGX-011, the second generation 2′-methoxyethyl gapmer oligonucleotides targeting the CLU gene, and Trastuzumab in this tumor hystotype. By using the HER-2 gene amplified-BT474 human breast cancer cells, we found Trastuzumab decreased HER-2 expression and inhibited cell proliferation without affecting apoptosis. Interestingly, Trastuzumab treatment up-regulated CLU protein expression in a dose-dependent fashion. We therefore hypothesized that the treatment with OGX-011, by blocking Trastuzumab-induced CLU expression, might potentiate the growth-inhibitory effect of Trastuzumab alone. Although OGX-011 had no effect on the behavior of the BT474 cells when used alone, it significantly enhanced the sensitivity of cells to Trastuzumab. A significant increase in the percentage of apoptotic cells, analyzed in terms of annexin V positivity and cleavage of poly(ADP-ribose) polymerase, was observed after combined treatment with OGX-011 plus Trastuzumab but not with either agent alone. Altogether our findings suggest that combined targeting of HER-2 and CLU may represent a novel, rational approach to breast cancer therapy. © 2005 Wiley-Liss, Inc.