Receptores estrogênicos regulam a migração e a invasão das células de câncer prostático independentes de andrógenos Pc-3

• Autor(a) principal: Lombardi, Ana Paola Giometti [UNIFESP]
• Data de Publicação: 2018
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da UNIFESP
• Texto Completo: https://sucupira.capes.gov.br/sucupira/public/consultas/coleta/trabalhoConclusao/viewTrabalhoConclusao.jsf?popup=true&id_trabalho=7033372; https://repositorio.unifesp.br/handle/11600/52238
• topic: Estrogen receptors Esr1 (Er") And Esr2 (Er"); Prostate cancer; Cancer models; Receptores estrogênicos Esr1 (Er") E Esr2 (Er"; Células de câncer; Câncer Prostático

Recent studies from our laboratory have shown the expression of estrogen receptors ESR1 (ERα) and ESR2 (ERβ) in androgenindependent prostate cancer cells (PC3), used as castrationresistant prostate cancer models. Furthermore, estrogen plays a role in PC3 cell proliferation through a novel pathway, involving ESR2mediated activation of βcatenin. It is important to emphasize that nonphosphorylated βcatenin associate with transcription factor TCF/LEF1 in the nucleus, and, together with coactivators, is implicated in cell proliferation, migration, and invasion in several cancer types. This study was performed to investigate the effects of estrogen receptors activation and the proteins involved with intracellular signaling pathways, such as SRC, PI3K, AKT and βcatenin, on migration, invasion and colony formation of the androgenindependent prostate cancer cells PC3. It showed that: 1. The treatment of PC3 cells with 17βestradiol (E2), ESR1selective agonist PPT and ESR2selective agonist DPN induced a rapid increase in the phosphorylation state of SRC. These effects were blocked by ESR2selective antagonist (PHTPP), ESR1selective antagonist (MPP) and selective inhibitor of the SRC family of protein tyrosine kinases (PP2). 2. A diffuse immunostaining for nonphosphorylated βcatenin was detected in the cytoplasm of PC3 cells. Low levels of nonphosphorylated βcatenin immunostaining were also detected near the plasma membrane. Treatment of PC3 cells with DPN for 2 hour markedly increased nonphosphorylated βcatenin expression in cytoplasm, near the plasma membrane and nuclei. These effects were blocked by pretreatment with PP2, suggesting the involvement of SRC on βcatenin expression induced by DPN. 3. Treatment with E2 and ESR2selective agonists (DPN and ERB041) caused an enhancement on migration and invasion of PC3 cells. The pretreatment with ESR2selective antagonist (PHTPP) decreased the effect of E2 and blunted the effect induced by DPN on migration and invasion of PC3 cells, indicating that ESR2 is involved in these processes. Previous study from our laboratory showed that ESR2selective agonist DPN increased ESR1 levels. In fact, treatment with ESR1selective antagonist (MPP) decreased 30% the migration and 25% the invasion of PC3 cells induced by DPN. These results suggest that ESR1 also plays a role in the regulation of DPNESR2 effects. On the other hand, ESR1selective agonist (PPT) did not have any effect on cell migration, but increased cell invasion (27%). The pretreatment with MPP blunted the effect induced by PPT on cell invasion, indicating that ESR1 is also involved in this process. 4. The upregulation induced by DPN on migration and invasion of PC3 cells was blocked by pretreatment with PKF 118310, a compound that disrupts the complex βcateninTCF/ LEF transcription factor, suggesting the involvement of ESR2βcatenin on migration and invasion of PC3 cells. The pretreatment with PPT did not have any effect on cell migration, but increased cell invasion (27%). This effect was partially reduced (59%) by PKF 118310, suggesting also the involvement of ESR1βcatenin on invasion of PC3 cells. 5. The pretreatment with SRC inhibitor (PP2), PI3K inhibitor (Wortmannin), AKT inhibitor (MK2206) and metalloproteases (MMPs) inhibitor (GM6001) blunted the effect induced by DPN and reduced the effect induced by PPT on invasion of PC3 cells. VEGF inhibitor (Bevacizumab) decreased the effect of both agonists DPN and PPT on invasion of PC3 cells. Taken together, these results suggest the involvement of these estrogen receptors and these intracellular proteins on invasion of PC3 cells. It is important to emphasize the upregulation of VEGF expression by E2, DPN and PPT. This effect was blunted by PKF 118310, indicating the involvement of βcatenin. 6. To further analyze the tumorigenic potential of estrogen receptors in PC3 cells, we performed the colony formation, anchorageindependent growth assay. E2, PPT and DPN increased the number and the size of colony formed by PC3 cells (rounds spheroids strutures) compared with control (absence of agonists). Furthermore, E2, PPT and DPN formed stellate or invasive structures. These effects were blocked by MPP and PHTTP, indicating that ESR1 and ESR2 are upstream receptors regulating these processes. The proteins involved in the intracellular signaling pathways of these receptors, such as βcatenin, SRC, PI3K and AKT also was involved the tumorigenic potential of estrogen receptors in PC3 cells. In summary, the activation of ESR2 (ERβ1 isoform) by the ESR2selective agonist (DPN) increases βcatenin expression and induces migration, cell invasion and colony formation. Activation of ESR1 by ESR1selective agonist (PPT) also enhances βcatenin expression, cell invasion, and colony formation. In addition to βcatenin expression, stimulation of ESR2 and ESR1 active SRC, PI3K, AKT, metalloproteinases and VEGF and these proteins are also involved in cell invasion and colony formation of PC3 cells. It is important to mention that the effects of ESR2 activation are predominant in these cells when compared to the activation of ESR1. Future studies in animal, xenographic, and large tissue models of prostate cancer patients could improve our understanding of the role of ESR2 and ESR1 in this cancer. The identification of new signaling pathways is important in the development of therapeutic targets for the treatment of CRPC. Combining therapies targeting more than one pathway is the logical approach coming from these new molecular evidences in order to obtain longer lasting clinical responses and increase the survival of patients with prostate cancer.