RESUMO
Oncogenic Kras activates a plethora of signaling pathways, but our understanding of critical Ras effectors is still very limited. We show that cell-autonomous phosphoinositide 3-kinase (PI3K) and 3-phosphoinositide-dependent protein kinase 1 (PDK1), but not Craf, are key effectors of oncogenic Kras in the pancreas, mediating cell plasticity, acinar-to-ductal metaplasia (ADM), and pancreatic ductal adenocarcinoma (PDAC) formation. This contrasts with Kras-driven non-small cell lung cancer, where signaling via Craf, but not PDK1, is an essential tumor-initiating event. These in vivo genetic studies together with pharmacologic treatment studies in models of human ADM and PDAC demonstrate tissue-specific differences of oncogenic Kras signaling and define PI3K/PDK1 as a suitable target for therapeutic intervention specifically in PDAC.
Assuntos
Neoplasias Pancreáticas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Fator 3 Associado a Receptor de TNF/metabolismo , Proteínas ras/metabolismo , Proteínas Quinases Dependentes de 3-Fosfoinositídeo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Proliferação de Células , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Ativação Enzimática , Humanos , Indazóis/farmacologia , Litostatina/metabolismo , Metaplasia , Camundongos , Camundongos Endogâmicos NOD , Transplante de Neoplasias , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Proteínas Proto-Oncogênicas p21(ras) , Transdução de Sinais , Sulfonamidas/farmacologia , Transplante Heterólogo , Células Tumorais CultivadasRESUMO
Four new single-chain Fv antibody fragments (scFvs) specific for the human leucocyte surface antigen CD123 (interleukin-3 receptor alpha) were generated to achieve preferential targeting of leukaemia stem cells (LSCs) in acute myeloid leukaemia (AML). The scFvs were isolated from a phage display library generated with spleen RNA from mice, immunized with a fusion protein consisting of the extracellular domain of CD123 and the Fc domain of a human immunoglobulin G1. The scFvs displayed CD123-specific binding on tumour cells (binding constants (K(D)) 4.5-101 nmol/l). The scFv with the highest affinity was used to design two cell death-inducing molecules. First, an immunotoxin, a fusion protein with truncated Pseudomonas Exotoxin A, induced potent apoptosis of AML-derived MOLM-13 and SKNO-1 cells at nanomolar concentrations. Second, the fusion to another scFv, specific for the low affinity Fcgamma-receptor III (CD16), created a bispecific single chain Fv (bsscFv). This bsscFv [123 x ds16] mediated potent lysis of AML-derived MOLM-13, THP-1 and SKNO-1 cells in antibody-dependent cellular cytotoxicity (ADCC) reactions at picomolar concentrations. The recruitment of CD16-positive effector cells for the lysis of AML cells via CD123 represents a novel combination with attractive prospects for future clinical testing.
