Development of Herceptin resistance in breast cancer cells

Abstract

Background Herceptin, a humanized antibody to HER‐2, is now utilized in the clinic for metastatic breast cancer treatment. The response rate for HER‐2+ patients is only 30% and little is known as to mechanisms of resistance. The mechanism of Herceptin action is also unknown but has been related to cell cycle inhibition. Methods The effects of Herceptin and other antibody treatments were determined by cell counting and cell cycle analysis. HER‐2 and p27 expression levels were analyzed by flow cytometry and levels of activated AKT were compared by Western blot analysis. Cellular HER‐2 and p27 expression was measured by immunofluorescence. Results Herceptin treatment of BT‐474 cells results in inhibition of cell growth and arrest in the G1 phase. The efficacy of growth arrest was not directly correlated to the binding affinity of antibodies to Her‐2. Our laboratory has developed cell lines that are resistant to Herceptin treatment. In resistant cell lines, binding of antibodies is not hindered. However, Herceptin has completely lost the ability to inhibit cell proliferation. Yet, the mouse isotype 4D5 maintains significant inhibitory activity upon Herceptin‐resistant clones. Conclusions Herceptin binds effectively to Her‐2 on the cell surface of Herceptin‐resistant cell lines and the level of Her‐2 expression on the cell surface is not downregulated. Herceptin resistance is not due to downregulation of levels of AKT protein expression, although, phosphorylation of AKT is enhanced in resistant lines and could have a role in resistance. Resistance appears to correlate with the loss of nuclear expression of the cyclin‐dependent kinase inhibitor, p27, as defined by immunofluorescence and flow cytometry studies and cdk‐2 binding studies. © 2004 Wiley‐Liss, Inc.

Stimulated γδ T Cells Increase the In Vivo Efficacy of Trastuzumab in HER-2+ Breast Cancer

“…MDA-MB-231 was included as an HER-2/neu negative tumor cell line control, and SK-BR-3 was used to represent HER-2/neu positive tumor cells. Specific lysis by Vg9 T cells was measured by standard 4-h 51 Crrelease assays. The lysis rates were obtained by the following equation: (experimental release -spontaneous release)/(maximum releasespontaneous release) 3 100.…”

“…The lysis rates were obtained by the following equation: (experimental release -spontaneous release)/(maximum releasespontaneous release) 3 100. Maximum and spontaneous releases were determined, respectively, by adding 0.1% Triton X-100 or complete medium to 51 Cr-labeled tumor target cells in the absence of gd T cells. Data are presented as the mean of triplicate samples.…”

Trastuzumab causes antibody-dependent cellular cytotoxicity–mediated growth inhibition of submacroscopic JIMT-1 breast cancer xenografts despite intrinsic drug resistance

“…Animals were euthanized, and the tumors were removed and cut into small pieces with a sterile blade, washed twice with sterile PBS, and placed in culture dishes in medium containing F-12/ DMEM (1:1) supplemented with 20% FCS, 60 units/L insulin, and antibiotics. The medium of JIMT-1 X+ cells also contained 10 Ag/mL trastuzumab (28). After 3 days, debris and dead cells were removed, and the medium was refreshed.…”

Trastuzumab-Resistant Luminal B Breast Cancer Cells Show Basal-Like Cell Growth Features Through NF-κB-Activation

“…(19,(22)(23)(24)(25) To rule out loss of epitope or epitopemasking effects, first, we validated the overexpression of HER2 receptor and function in BT-474 cells. The HER2 expression remained unaltered between the parental cells (BT-474) and the resistant cells (BT-474-R), both at the mRNA (Fig.…”

“…Their results indicate that the resistant clones have not lost HER2 expression, but overexpress epidermal growth factor receptor (EGFR) and HER3. (41,42) On the other hand, Kute and colleagues (23) developed resistant cells by treating with the antibody over 2 weeks and found that PI3K and overexpression of EGFR mediate resistance. Chan and colleagues (43) cultures) and thus provide a snapshot of the evolution of TZR (Fig.…”