Evaluation of the molecular mechanisms involved in the gain of function of a Li-Fraumeni TP53 mutation.
The TP53 tumor suppressor gene is the most frequent target for genetic alterations in human cancer. TP53 gene alterations may result in the gain of oncogenic functions such as neoangiogenesis and resistance to therapy. The TP53 germ line mutation c.659A>C (p.Y220S) was identified in stored DNA from related patients with Li-Fraumeni syndrome (LFS) who died after developing clinically aggressive tumors. All of the patients were treated with protocols that included doxorubicin hydrochloride (DX) as a pivotal drug. To define the in vitro mutational phenotype of this germ line mutation, we used murine fibroblasts explanted from wild-type (wt) and p53 knockout (KO) mice from the same littermate. p53Y220S and p53R175H fibroblasts, obtained from p53KO fibroblasts transfected with expression vectors encoding the human Y220S and R175H p53 mutants, respectively, exhibited resistance to DX treatment. Moreover, p53Y220S fibroblasts exhibited angiogenetic properties, and after DX treatment, p53Y220S failed to translocate into the nucleus and showed an increase in its cytosolic levels. DX treatment does not influence p53 distribution within the nuclear and cytosolic compartments in p53R175H fibroblasts. Peroxiredoxin II (Prx II), a protein that is involved in eliminating reactive oxygen species (ROS), showed increased expression intensity in p53Y220S fibroblasts after DX treatment, as observed by two-dimensional electrophoresis analysis. Moreover, Thioredoxin (Trx), a protein that cooperates with Prx II, is overexpressed in p53Y220S mutants under basal conditions. These data suggest a relationship between the presence of the p53Y220S mutation and enhanced levels of Prx II and Trx in mutant fibroblasts. Since one of the mechanisms of the DX antitumor effect has been ascribed to production of ROS, future studies will evaluate the involvement of PrxII and Trx in the chemoresistance of p53Y220S fibroblasts to DX.
Genetics Unit, Laboratory of Oncology Research, Rizzoli Orthopedic Institute, Bologna, Italy.
Silvia
S
Capponcelli
Elena
E
Pedrini
Maria Antonietta
MA
Cerone
Valeria
V
Corti
Silvia
S
Fontanesi
Massimo
M
Alessio
Angela
A
Bachi
Silvia
S
Soddu
Domenico
D
Ribatti
Piero
P
Picci
Lee J
LJ
Helman
Giorgio
G
Cantelli-Forti
Luca
L
Sangiorgi
D000293:Adolescent; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D002648:Child; D002675:Child, Preschool; D049033:Chorioallantoic Membrane / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology; D005260:Female; D005347:Fibroblasts / Q000378:metabolism; D016158:Genes, p53; D006801:Humans; D016864:Li-Fraumeni Syndrome / Q000235:genetics*; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*
D000903:Antibiotics, Antineoplastic; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin