RESUMO
PURPOSE: To examine the involvement of specific components of the PI3K/AKT pathway in urinary bladder cancer development. METHODS: Samples from 65 tumors and 13 normal bladder tissues were collected. Genomic DNA isolation from snap-frozen and paraffin-embedded laser-microdissected tissues was followed by Sanger sequencing, whereas total RNA was purified for use in RT-PCR analyses. Immunohistochemistry was carried out on sections of paraffin-embedded biopsy material. RESULTS: Three pathogenic mutations (two missense and one frameshift) were identified in exon 20 of PIK3CA {c.3140A>G (p.His1047Arg), c.[3172A>T(;)3174C>T] (p.lle1058Phe), c.3203dupA (p.Asn1068Lysfs*5)} after laser capture microdissection, whereas PTEN mRNA expression was found to be downregulated in bladder cancer tissues compared to normal bladder urothelium. Upregulation of cytoplasmic and nuclear p-AKT expression was detected in low grade tumors, whereas in infiltrating carcinomas p-AKT was shown to be downregulated and confined to the cytoplasm. PTEN expression was weak and mainly cytoplasmic in superficial tumors, but stronger and nuclear in the infiltrating tumors. CONCLUSIONS: PI3K/AKT pathway activation is crucial for bladder cancer initiation and progression. In this context, PIK3CA, p-AKT and nuclear PTEN could be used along with other biomarkers for prognosis and selection of appropriate therapy in the clinical management of bladder cancer.
Assuntos
Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias da Bexiga Urinária/genética , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Feminino , Humanos , Masculino , PTEN Fosfo-Hidrolase/metabolismo , Transdução de Sinais/genética , Neoplasias da Bexiga Urinária/enzimologia , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologiaRESUMO
Doxorubicin is an important component of combination therapy for muscle-invasive urinary bladder cancer. Treatment with this topoisomerase II poison is able to interfere with cell cycle progression and lead to cancer cell death. Using FACS analysis, Western immunoblotting and semi-quantitative RT-PCR, we studied the effects of doxorubicin on cell cycle progression and apoptosis, and also explored the possibility of using groups of genes as biomarkers of prognosis and/or response to doxorubicin treatment in human urinary bladder cancer cells. Doxorubicin induced dose-dependent G2/M and/or G1/S cell cycle arrest, followed by grade- and dose-dependent reduction in the amount of the cytosolic trimeric form of FasL, activation of Caspase-8, Caspase-9, Caspase-3, cleavage of PARP, Lamin A/C, Bcl-XL/S and interestingly Hsp90, and finally cell death. Data presented here also suggest the use of the expression patterns of Cyclin-E2, Cyclin-F, p63, p73, FasL, TRAIL, Tweak, Tweak-R, XAF-1, OPG and Bok genes for identification of the differentiation grade, and Cyclin-B2, GADD45A, p73, FasL, Bik, Bim, TRAIL, Fas, Tweak-R, XAF-1, Bcl-2, Survivin, OPG, DcR2 and Bcl-XL genes for the detection of response to doxorubicin in human bladder cancer cells.
