RESUMEN
Translation initiation factor eIF4E mediates normal cell proliferation, yet induces tumorigenesis when overexpressed. The mechanisms by which eIF4E directs such distinct biologic outputs remain unknown. We found that mouse mammary morphogenesis during pregnancy and lactation is accompanied by increased cap-binding capability of eIF4E and activation of the eIF4E-dependent translational apparatus, but only subtle oscillations in eIF4E abundance. Using a transgenic mouse model engineered so that lactogenic hormones stimulate a sustained increase in eIF4E abundance in stem/progenitor cells of lactogenic mammary epithelium during successive pregnancy/lactation cycles, eIF4E overexpression increased self-renewal, triggered DNA replication stress, and induced formation of premalignant and malignant lesions. Using complementary in vivo and ex vivo approaches, we found that increasing eIF4E levels rescued cells harboring oncogenic c-Myc or H-RasV12 from DNA replication stress and oncogene-induced replication catastrophe. Our findings indicate that distinct threshold levels of eIF4E govern its biologic output in lactating mammary glands and that eIF4E overexpression in the context of stem/progenitor cell population expansion can initiate malignant transformation by enabling cells to evade DNA damage checkpoints activated by oncogenic stimuli. Maintaining eIF4E levels below its proneoplastic threshold is an important anticancer defense in normal cells, with important implications for understanding pregnancy-associated breast cancer.
Asunto(s)
Neoplasias de la Mama/genética , Carcinogénesis/genética , Factor 4E Eucariótico de Iniciación/biosíntesis , Glándulas Mamarias Humanas/metabolismo , Animales , Neoplasias de la Mama/patología , Proliferación Celular/genética , Replicación del ADN/genética , Factor 4E Eucariótico de Iniciación/genética , Femenino , Humanos , Glándulas Mamarias Animales/metabolismo , Glándulas Mamarias Animales/patología , Glándulas Mamarias Humanas/patología , Ratones , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Embarazo , Biosíntesis de Proteínas , Proteínas Proto-Oncogénicas c-myc/biosíntesis , Proteínas ras/biosíntesisRESUMEN
Fibroblasts have a central role in the maintenance of tissue homeostasis and repair after injury. Currently, there are no tractable, cost-effective model systems for studying the biology of human fibroblasts in vivo. Here we demonstrate that primary human fibroblasts survive transplantation into zebrafish embryos. Transplanted cells migrate and proliferate, but do not integrate into host tissues. We used this system to study the intrinsic motility of lung fibroblasts from a prototype fibrotic lung disease, idiopathic pulmonary fibrosis (IPF). IPF fibroblasts displayed a significantly higher level of motility than did fibroblasts from nonfibrotic lungs. This is the first in vivo examination of primary human lung fibroblast motility in health and disease using zebrafish models.
Asunto(s)
Movimiento Celular , Fibroblastos/fisiología , Modelos Animales , Pez Cebra/embriología , Animales , Técnicas de Cultivo de Célula , Línea Celular , Fibroblastos/citología , Fibroblastos/trasplante , Humanos , Trasplante HeterólogoRESUMEN
Normal growth and development depends upon high fidelity regulation of cap-dependent translation initiation, a process that is usurped and redirected in cancer to mediate acquisition of malignant properties. The epithelial-to-mesenchymal transition (EMT) is a key translationally regulated step in the development of epithelial cancers and pathological tissue fibrosis. To date, no compounds targeting EMT have been developed. Here we report the synthesis of a novel class of histidine triad nucleotide binding protein (HINT)-dependent pronucleotides that interdict EMT by negatively regulating the association of eIF4E with the mRNA cap. Compound eIF4E inhibitor-1 potently inhibited cap-dependent translation in a dose-dependent manner in zebrafish embryos without causing developmental abnormalities and prevented eIF4E from triggering EMT in zebrafish ectoderm explants without toxicity. Metabolism studies with whole cell lysates demonstrated that the prodrug was rapidly converted into 7-BnGMP. Thus we have successfully developed the first nontoxic small molecule able to inhibit EMT, a key process in the development of epithelial cancer and tissue fibrosis, by targeting the interaction of eIF4E with the mRNA cap and demonstrated the tractability of zebrafish as a model organism for studying agents that modulate EMT. Our work provides strong motivation for the continued development of compounds designed to normalize cap-dependent translation as novel chemo-preventive agents and therapeutics for cancer and fibrosis.
Asunto(s)
Sistemas de Liberación de Medicamentos , Epitelio/embriología , Mesodermo/embriología , Complejo Proteico Nuclear de Unión a la Caperuza/antagonistas & inhibidores , Complejo Proteico Nuclear de Unión a la Caperuza/metabolismo , Pez Cebra/embriología , Amidas/síntesis química , Amidas/química , Amidas/farmacología , Animales , Secuencia de Bases , Embrión no Mamífero/embriología , Factor 4E Eucariótico de Iniciación/antagonistas & inhibidores , Humanos , Concentración 50 Inhibidora , Datos de Secuencia Molecular , Neoplasias/metabolismo , Ácidos Fosfóricos/síntesis química , Ácidos Fosfóricos/química , Ácidos Fosfóricos/farmacología , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The anti-proliferative effects of a novel bromomethoxylated arylphosphate derivative of zidovudine (compound DDE46, CAS 213982-96-8) were first examined in a zebra fish embryo model. DDE46 blocked the cell division at the 2-cell stage of the embryonic development followed by total cell fusion. DDE46 also inhibited the proliferation of the leukemic cell lines NALM-6 and MOLT-3. DDE46 enhanced the activity of the pro-apoptotic enzymes Caspase-3, Caspase-6, Caspase-8, and Caspase-9 leading to the apoptotic death of the leukemic cell line Jurkat. These results justify the further development of this agent as a new anti-leukemic drug candidate.