TY - JOUR
T1 - Class I β-tubulin mutations in 2-methoxyestradiol-resistant acute lymphoblastic leukemia cells
T2 - Implications for drug-target interactions
AU - Liaw, Tracy Y E
AU - Salam, Noeris K.
AU - McKay, Matthew J.
AU - Cunningham, Anne M.
AU - Hibbs, David E.
AU - Kavallaris, Maria
PY - 2008/10/1
Y1 - 2008/10/1
N2 - 2-Methoxyestradiol (2ME2) is a naturally occurring derivative of estradiol that has been shown to be an active small molecule that has antitumor and antiangiogenic properties. 2ME2 binds to β-tubulin near the colchicine-binding site, inhibits microtubule polymerization, and induces mitotic arrest. To improve understanding of the mechanisms of action and resistance to 2ME2, we selected leukemia cells, CCRF-CEM, that display increasing resistance to 2ME2, and three of the highly resistant sublines were chosen for detailed analysis. The 2ME2 cells selected in 7.2 to 28.8 μmol/L were found to be 47- to 107-fold resistant to 2ME2 and exhibited low levels of cross-resistance to vinblastine. Two of the lowest 2ME2-resistant sublines were significantlyh ypersensitive to colchicine and epothilone B, but the hypersensitive effects were lost in the highest 2ME2-resistant subline. Moreover, 2ME2-resistant cells require 10-fold higher concentrations of 2ME2 to induce G2-M cell cycle arrest and have higher amounts of tubulin polymer compared with parental cells. Gene and protein sequencing revealed four class I B-tubulin mutations, S25N, D197N, A248T, and K350N, in the 2ME2-resistant cells. The S25N mutation is within the paclitaxel-binding site, whereas A248T and K350N are within the colchicine-binding site on β-tubulin, yet the resistant cells were not cross-resistant to paclitaxel or colchicine. This strongly suggests that the mutations have induced conformational changes to the binding site that resulted in 2ME2 resistance. The 2ME2-resistant leukemia cells provide novel insights into microtubule stability and drug-target interactions.
AB - 2-Methoxyestradiol (2ME2) is a naturally occurring derivative of estradiol that has been shown to be an active small molecule that has antitumor and antiangiogenic properties. 2ME2 binds to β-tubulin near the colchicine-binding site, inhibits microtubule polymerization, and induces mitotic arrest. To improve understanding of the mechanisms of action and resistance to 2ME2, we selected leukemia cells, CCRF-CEM, that display increasing resistance to 2ME2, and three of the highly resistant sublines were chosen for detailed analysis. The 2ME2 cells selected in 7.2 to 28.8 μmol/L were found to be 47- to 107-fold resistant to 2ME2 and exhibited low levels of cross-resistance to vinblastine. Two of the lowest 2ME2-resistant sublines were significantlyh ypersensitive to colchicine and epothilone B, but the hypersensitive effects were lost in the highest 2ME2-resistant subline. Moreover, 2ME2-resistant cells require 10-fold higher concentrations of 2ME2 to induce G2-M cell cycle arrest and have higher amounts of tubulin polymer compared with parental cells. Gene and protein sequencing revealed four class I B-tubulin mutations, S25N, D197N, A248T, and K350N, in the 2ME2-resistant cells. The S25N mutation is within the paclitaxel-binding site, whereas A248T and K350N are within the colchicine-binding site on β-tubulin, yet the resistant cells were not cross-resistant to paclitaxel or colchicine. This strongly suggests that the mutations have induced conformational changes to the binding site that resulted in 2ME2 resistance. The 2ME2-resistant leukemia cells provide novel insights into microtubule stability and drug-target interactions.
UR - http://www.scopus.com/inward/record.url?scp=55749087503&partnerID=8YFLogxK
U2 - 10.1158/1535-7163.MCT-08-0442
DO - 10.1158/1535-7163.MCT-08-0442
M3 - Article
C2 - 18852118
AN - SCOPUS:55749087503
SN - 1535-7163
VL - 7
SP - 3150
EP - 3159
JO - Molecular Cancer Therapeutics
JF - Molecular Cancer Therapeutics
IS - 10
ER -