[369] Proteasome inhibitor bortezomib (Velcade) and trastuzumab in HER-2 overexpressing breast cancer cell lines.
[Abstract from the San Antonio Breast Cancer Symposium]
Laes J-F, Cardoso F, Durbecq V, Lagneaux L, Hennuy B, Lallemand F, Gonze I, Di Leo A, Kern F, Ross J, Piccart MJ, Sotiriou C JBI, Brussels, Belgium; BioVallee A.S.B.L., Gosselies, Belgium; Southern Research Institute, AL; Millennium Pharmaceutics, Cambridge, MA
Note: * co-first authors
Background: Even in the subset of breast cancer (BC) patients with HER-2 highly positive tumours (3+ score by IHC or FISH+) only 40% of responses to trastuzumab are obtained and the median duration of response is 9 months. Resistance to trastuzumab, both primary and acquired, is an important problem. The proteasome inhibitor bortezomib, formerly known as PS-341, is a new promising anticancer agent. The proteasome has been implicated in the degradation of several proteins, such as HER-2 and Ikb. We hypothesized that the combination of both drugs might result in a synergistic or at least additive effect since both agents prevent the NFkB activation and increase p27 levels. Additionally, the inhibition of HER-2 degradation by bortezomib may also influence the response to trastuzumab. Methods: Four BC cell lines were used: MDA-MB-361 (HER-2+/ER+), MDA-MB-453 (HER-2+/ER-), HER-2-transfected-MCF-7 (HER-2+++/ER+) and MCF-7 cells (HER-2-/ER+) as control. Each cell line was incubated with 0.01M of bortezomib for injection, for 1, 2 or 3 days. After washing, the cells were treated with 20g/ml of trastuzumab for 1, 2 or 3 days. Techniques: Kit Annexin-V FITC (Biosource) for apoptosis measurements; analysis of DNA content after PI staining and cytomorphological analysis with May-Grunwald-Giemsa (MGG); Affymetrix cDNA microarray chips for genomic profiling. Results: 1) Trastuzumab and bortezomib antiproliferative effects were directly dependent on HER-2 status but not on ER status: in MCF-7 cell lines, apoptosis was seen 48h after treatment with bortezomib and no apoptosis was seen with trastuzumab; in HER-2+ cells lines apoptosis was seen 48h after treatment with bortezomib and only 72h after trastuzumab administration; in HER-2 ++ or +++ cells, apoptosis was seen only with a ten-fold increase in bortezomib dose, and with trastuzumab no apoptosis was detected but a cell cycle arrest in G1 was observed. 2) According to these results, different combination schedules of trastuzumab and bortezomib were tested in HER-2 ++ and +++ cells. A very strong synergistic effect between both drugs was observed in vitro, which was stronger in HER-2 +++ cells than in HER-2 ++. Conclusions: 1) Response to bortezomib and trastuzumab appears to be linked with HER-2 status and independent of ER status and of cell growth rate. 2) The combination bortezomib + trastuzumab seems to be synergistic in vitro. 3) This synergy is more important in cells with high levels of HER-2. 4) Genomic profiling of all cell lines and evaluation of p27 and NFkB levels, before and after treatment, are ongoing.
Work supported by a research grant from the Belgian non-profit organization Fondation Lambeau-Marteau and by a research grant from Millennium Pharmaceuticals.
Thursday, December 4, 2003 4:30 PM
Poster Session: Tumor Cell Biology: Drug Resistance (4:30 PM-7:00 PM)
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