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Normal p53 status and function despite the development of drug resistance in human breast cancer cells
K Wosikowski1, J T Regis, R W Robey
1Medicine Branch, National Cancer Institute, Bethesda, Maryland 20892, USA.
Abstract:
Loss of or mutations in p53 protein have been shown to decrease both radio- and chemosensitivity. The present study assessed the p53 gene status, ability to arrest in G1 of the cell cycle, the functionality of the p53 transduction pathway, and apoptosis following treatment with radiation in a series of drug-resistant human breast cancer cells to determine whether p53 alterations occur during the development of drug resistance. We used 13 sublines derived from MCF-7, ZR75B, and T47D cells, which were resistant to doxorubicin, paclitaxel, vinblastine, cisplatin, etoposide, and amsacrine. Eleven of 12 drug-resistant sublines retained the parental p53 gene status, as determined by sequence analysis and functional yeast assay; only one subline was found to have acquired a mutation in the p53 gene. The MCF-7 TH subline was found to both acquire mutated p53 and to have major changes in p53 protein expression and function. In 12 other drug-resistant sublines, the G1 checkpoint was conserved or only slightly impaired. A normal accumulation of p53, p21Cip1/Waf1, and Mdm2 proteins and hypophosphorylation of Rb protein occurred in response to radiation with only small differences noted in the kinetics of p53 and p21Cip1/Waf1 induction. Increased susceptibility to apoptosis was found in the ZR75B drug-resistant sublines, whereas no evidence for apoptosis was observed in the ZR75B, MCF-7, and T47D parentals and the MCF-7 and T47D drug-resistant sublines. This effect could not be explained by alterations in bcl-2 or bax expression. Our results demonstrate that alterations in: (a) p53 gene status; (b) ability to arrest in G1; (c) induction of p53 protein and p53-dependent genes; and (d) decreased activation of apoptosis is not a requirement for the onset of drug resistance. The function of p53 appears to be dissociated from drug resistance in our model system.
Insights
p53 gene alterations are not required for drug resistance in human breast cancer cells. Most drug-resistant cells maintained normal p53 function, indicating p53 status is dissociated from drug resistance development.
Area of Science:
- * Oncology
- * Molecular Biology
- * Genetics
Background:
- * Loss of or mutations in the p53 protein can reduce sensitivity to radiation and chemotherapy.
- * The p53 tumor suppressor protein plays a critical role in cell cycle arrest, DNA repair, and apoptosis.
- * Understanding the role of p53 in drug resistance is crucial for developing effective cancer therapies.
Purpose of the Study:
- * To investigate whether alterations in the p53 gene occur during the development of drug resistance in human breast cancer cells.
- * To assess the p53 gene status, cell cycle arrest, p53 pathway functionality, and apoptosis following radiation treatment in drug-resistant breast cancer sublines.
- * To determine the relationship between p53 alterations and the onset of drug resistance.
Main Methods:
- * Generation of 13 drug-resistant human breast cancer sublines from MCF-7, ZR75B, and T47D cells resistant to various chemotherapeutic agents.
- * Sequence analysis and functional yeast assays to determine p53 gene status.
- * Assessment of G1 cell cycle arrest, p53 protein accumulation, p21Cip1/Waf1, Mdm2, and Rb protein phosphorylation in response to radiation.
- * Analysis of apoptosis and expression of bcl-2 and bax.
Main Results:
- * Eleven of 12 drug-resistant sublines retained the parental p53 gene status; only one acquired a p53 mutation.
- * The G1 checkpoint was largely conserved in most drug-resistant sublines.
- * Radiation treatment induced normal p53, p21Cip1/Waf1, and Mdm2 accumulation in most sublines.
- * Increased apoptosis was observed in ZR75B drug-resistant sublines, but not explained by bcl-2 or bax alterations.
Conclusions:
- * Alterations in p53 gene status, cell cycle arrest, p53 pathway induction, or apoptosis are not required for the development of drug resistance.
- * The function of the p53 protein appears to be dissociated from drug resistance in this model system.
- * These findings suggest that targeting p53 may not be a universally effective strategy for overcoming drug resistance in breast cancer.