RESUMO
BACKGROUND: TGFß signaling has typically been associated with suppression of tumor initiation while the role it plays in metastasis is generally associated with progression of malignancy. However, we present evidence here for an anti-metastatic role of TGFß signaling. METHODS: To test the importance of TGFß signaling to cell survival and metastasis we compared human colon carcinoma cell lines that are either non-tumorigenic with TGFß response (FET), or tumorigenic with TGFß response (FETα) or tumorigenic with abrogated TGFß response via introduction of dominant negative TGFßRII (FETα/DN) and their ability to metastasize. Metastatic competency was assessed by orthotopic transplantation. Metastatic colony formation was assessed histologically and by imaging. RESULTS: Abrogation of TGFß signaling through introduction of a dominant negative TGFß receptor II (TGFßRII) in non-metastatic FETα human colon cancer cells permits metastasis to distal organs, but importantly does not reduce invasive behavior at the primary site. Loss of TGFß signaling in FETα-DN cells generated enhanced cell survival capabilities in response to cellular stress in vitro. We show that enhanced cellular survival is associated with increased AKT phosphorylation and cytoplasmic expression of inhibitor of apoptosis (IAP) family members (survivin and XIAP) that elicit a cytoprotective effect through inhibition of caspases in response to stress. To confirm that TGFß signaling is a metastasis suppressor, we rescued TGFß signaling in CBS metastatic colon cancer cells that had lost TGFß receptor expression due to epigenetic repression. Restoration of TGFß signaling resulted in the inhibition of metastatic colony formation in distal organs by these cells. These results indicate that TGFß signaling has an important role in the suppression of metastatic potential in tumors that have already progressed to the stage of an invasive carcinoma. CONCLUSIONS: The observations presented here indicate a metastasis suppressor role for TGFß signaling in human colon cancer cells. This raises the concern that therapies targeting inhibition of TGFß signaling may be imprudent in some patient populations with residual TGFß tumor suppressor activity.
Assuntos
Neoplasias do Colo/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Células HEK293 , Humanos , Immunoblotting , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microscopia de Fluorescência , Microscopia de Vídeo , Invasividade Neoplásica , Metástase Neoplásica , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo II , Receptores de Fatores de Crescimento Transformadores beta/genética , Transdução de Sinais/genética , Proteínas Smad/genética , Proteínas Smad/metabolismo , Transplante Heterólogo , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/metabolismoRESUMO
The alpha isoform of the phosphatidylinositol-3-kinases (PI3Kα) is often mutated, amplified and overexpressed in human tumors. In an effort to develop new inhibitors targeting this enzyme, we carried out a pharmacophore model study based on six PI3Kα-selective compounds. The pharmacophore searching identified three structurally novel inhibitors of PI3Kα and its H1047R mutant. Our biological studies show that two of our hit molecules suppressed the formation of pAKT, a downstream effector of PI3Kα, and induced apoptosis in the HCT116 colon cancer cell line. QPLD-based docking showed that residues Asp933, Glu849, Val851, and Gln859 appeared to be key binding residues for active inhibitors.
Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Inibidores de Proteínas Quinases/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Estrutura Molecular , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
This work describes our efforts to optimize the lead PI3Kα inhibitor N-benzyl 4-hydroxy-2-quinolone-3-carboxamide using structure-based design and molecular docking. We identified a series of N-phenyl 4-hydroxy-2-quinolone-3-carboxamides as selective inhibitors of mutant H1047R versus wild-type PI3Kα and we also showed that the cell growth inhibition by these compounds likely occurs by inhibiting the formation of pAKT and induction of apoptosis.