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We recently found that overexpression of p185c-erbB2 in c-erbB2 transfected MDA-MB-435 breast cancer cells (435.eB transfectants) confers a 5–9-fold increase in Taxol resistance. To examine whether Taxol resistance is a common phenomenon in other c-erbB2 overexpressing breast cancer cell lines, we tested a panel of human breast cancer cell lines established from different patients and expressing p185c-erbB2 at different levels for their sensitivity to Taxol and Taxotere, a synthetic taxoid. Higher expression of p185c-erbB2 in these breast cancer cell lines indeed correlated well with resistance to Taxol and Taxotere, and the degree of resistance was about 100-fold that in c-erbB2-overexpressing 435.eB transfectants, demonstrating that these breast cancer cells are highly resistant to Taxol. Since mdr-1-encoded p-glycoprotein (p170mdr-1) has been implicated in Taxol resistance, we next examined the p170mdr-1 levels in these breast cancer cell lines that are highly resistant to Taxol. Higher levels of p170mdr-1 expression were found in several breast cancer cell lines that are highly resistant to Taxol. Since these same breast cancer cell lines also expressed higher levels of p185c-erbB2, we sought to determine the relative contribution of p185c-erbB2 and p170mdr-1 overexpression to Taxol resistance. We first specifically down-regulated cell surface p185c-erbB2 using anti-p185c-erbB2 monoclonal antibodies and assayed sensitivity of these cells to Taxol. We next specifically inactivated p170mdr-1 function using p170mdr-1 blockers (thioridazine or verapamil) and again assayed Taxol sensitivity. Both p185c-erbB2 down-regulation and p170mdr-1 blockade significantly sensitized the breast cancer cell lines to Taxol. The results indicate that overexpression of either p185c-erbB2 or... [ABSTRACT FROM AUTHOR]

Background: Triple negative breast cancer (TNBC) represents a significant clinical challenge. Chemotherapy remains the mainstay for a large part of TNBC patients, whereas drug resistance and tumor recurrence frequently occur. It is in urgent need to identify novel molecular targets for TNBC and develop effective therapy against the aggressive disease. Methods: Immunohistochemistry was performed to examine the expression of HER3 in TNBC samples. Western blots were used to assess protein expression and activation. Cell proliferation and viability were determined by cell growth (MTS) assays. TCGA databases were analyzed to correlate HER3 mRNA expression with the clinical outcomes of TNBC patients. Specific shRNA was used to knockdown HER3 expression. IncuCyte system was utilized to monitor cell growth and migration. LIVE/DEAD Cell Imaging was to detect live and dead cells. HER3 recognition by our anti-HER3 monoclonal antibody (mAb) 4A7 was verified by ELISA, flow cytometry, and co-immunoprecipitation assays. Orthotopic tumor models were established in nude mice to determine the capability of TNBC cells forming tumors and to test if our mAb 4A7 could potentiate the antitumor activity of paclitaxel in vivo. Results: Elevated expression of HER3 was observed in approximately half of the TNBC specimens and cell lines tested. Analyses of TCGA databases found that the TNBC patients with high HER3 mRNA expression in the tumors showed significantly worse overall survival (OS) and relapse-free survival (RFS) than those with low HER3 expression. Specific knockdown of HER3 markedly inhibited TNBC cell proliferation and mammosphere formation in vitro and tumor growth in vivo. Our mAb 4A7 abrogated heregulin (a ligand for HER3), but not SDF-1 (a ligand for CXCR4)-induced enhancement of TNBC cell migration. Combinations of 4A7 and the EGFR-tyrosine kinase inhibitor (TKI) gefitinib dramatically decreased the levels of phosphorylated HER3, EGFR, Akt, and ERK1/2 in TNBC cells and potently induced growth inhibition and cell death. Moreover, 4A7 in combination with paclitaxel exerted significant antitumor activity against TNBC in vitro and in vivo. Conclusions: Our data demonstrate that increased HER3 is an effective therapeutic target for TNBC and our anti-HER3 mAb (4A7) may enhance the efficacy of gefitinib or paclitaxel in TNBC. [ABSTRACT FROM AUTHOR]

The impact of the cyclin dependent kinase (CDK) inhibitors p21Waf1 and p27Kip1 on paclitaxel-mediated cytotoxicity was investigated in RKO human colon adenocarcinoma cells with the ecdysone-inducible expression of p21Waf1 or p27Kip1. Ectopic expression of p27Kip1 arrested cells at G1 phase, whereas p21Waf1 expression arrested cells at G1 and G2. Expression of p21Waf1 after paclitaxel treatment produced much greater resistance to paclitaxel than did expression of p27Kip1. We attributed this difference to the additional block at G2 induced by p21Waf1, which prevented cells from entering M phase and becoming paclitaxel susceptible. Expression of p21Waf1 inhibited p34cdc2 activity and markedly reduced paclitaxel-mediated mitotic arrest, from 87.5 to 23%. In contrast, p27Kip1 expression also inhibited p34cdc2 but reduced mitotic arrest only slightly, from 87.4 to 74.5%. We concluded that the G2 block produced by p21Waf1, but not by p27Kip1, contributed to their unequal modulation of sensitivity to paclitaxel-mediated apoptosis in RKO cells, and there is no causal relationship between paclitaxel-mediated cytotoxicity and elevation of p34cdc2 activity. Oncogene (2000) 19, 2423–2429 [ABSTRACT FROM AUTHOR]