RESUMEN
Nucleolin (NCL) is a nucleocytoplasmic protein involved in many biological processes, such as ribosomal assembly, rRNA processing, and mRNA stabilization. NCL also regulates the biogenesis of specific microRNAs (miRNAs) involved in tumor development and aggressiveness. Interestingly, NCL is expressed on the surface of actively proliferating cancer cells, but not on their normal counterparts. Therefore, NCL is an attractive target for antineoplastic treatments. Taking advantage of phage-display technology, we engineered a fully human single-chain fragment variable, named 4LB5. This immunoagent binds NCL on the cell surface, it is translocated into the cytoplasm of target cells, and it abrogates the biogenesis of NCL-dependent miRNAs. Binding of 4LB5 to NCL on the cell surface of a variety of breast cancer and hepatocellular carcinoma cell lines, but not to normal-like MCF-10a breast cells, dramatically reduces cancer cell viability and proliferation. Finally, in orthotopic breast cancer mouse models, 4LB5 administration results in a significant reduction of the tumor volume without evident side effects. In summary, here we describe, to our knowledge, the first anti-NCL single-chain fragment variable displaying antineoplastic activity against established solid tumors, which could represent the prototype of novel immune-based NCL-targeting drugs with clinical potential as diagnostic and therapeutic tools in a wide variety of human cancers.
Asunto(s)
Antineoplásicos/química , Neoplasias/inmunología , Neoplasias/terapia , Fosfoproteínas/química , Proteínas de Unión al ARN/química , Anticuerpos de Cadena Única/química , Animales , Apoptosis , Carcinoma Hepatocelular/metabolismo , Línea Celular Tumoral , Membrana Celular/metabolismo , Movimiento Celular , Proliferación Celular , Supervivencia Celular , Citoplasma/metabolismo , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Neoplasias Hepáticas/metabolismo , Ratones , Ratones SCID , Trasplante de Neoplasias , Neoplasias/metabolismo , Biblioteca de Péptidos , Ingeniería de Proteínas , Proteínas Recombinantes/química , NucleolinaRESUMEN
Zinc half sites are present in all human lactogenic hormones: human prolactin (hPRL), growth hormone (hGH), placental lactogens (hPL) and the hPRL receptor (hPRLr). The influence of divalent zinc (Zn(2+)) as measured by intrinsic fluorescence or FRET in each of these hormones is unique and is affected by the presence of varying stoichiometries of hPRLr. These data show that both Zn(2+) and hPRLr binding influence hPRL conformers in an interdependent fashion. Although each of these three lactogenic hormones bind hPRLr and induce a biological response that is sensitive to the presence of increasing concentrations of Zn(2+), each hormone is unique in the mechanistic details of this process.
