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A p21/WAF1 mutation favors the appearance of drug resistance to paclitaxel in human noncancerous epithelial mammary cells
Author(s) -
Galmarini Carlos María,
Bouchet Benjamin Pierre,
Audoynaud Carole,
Lamblot Christelle,
Falette Nicole,
Bertholon Jacques,
Wang Qing,
Beghin Anne,
Dumontet Charles,
Puisieux Alain
Publication year - 2006
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.21770
Subject(s) - paclitaxel , microbiology and biotechnology , mutation , exon , biology , mutant , cancer research , point mutation , western blot , drug resistance , gene , cancer , genetics
We investigated the mechanisms responsible for paclitaxel resistance in HME‐1 cells (human mammary epithelial cells immortalized with hTERT). These cells were exposed to paclitaxel (10 pM for 7 days) and 20 cellular surviving populations (PSP) were obtained. PSP demonstrated high levels of resistance to paclitaxel cytotoxicity as compared with HME‐1 cells. Activation of mdr‐1 gene expression was observed in 2 PSP. Protein expression analysis using a C‐terminal targeted antibody showed that 13 PSP were negative for p21/WAF1 expression after ionizing radiation (6 Gy) or doxorubicin (100 nM) treatment. Sequencing of the 3 exons of the CDKN1A gene revealed that 13 PSP contained a point mutation in exon 2. This mutation consisted in a T insertion at codon 104 leading to a premature STOP codon appearance. Mismatch amplification mutation assay and RFLP‐PCR confirmed the presence of the mutation in 16 PSP. Western blot using an N‐terminal targeted antibody demonstrated that the C‐terminal‐truncated p21/WAF1 protein (14 kDa) was indeed expressed in the 13 PSP. Our data suggest that p21/WAF1 inactivation may confer a strong resistance to paclitaxel in noncancerous breast epithelial cells harboring a p21/WAF1 mutant. © 2006 Wiley‐Liss, Inc.