title: Engineering tumor cell targeting in nanoscale amyloidal materials.
abstract: Bacterial inclusion bodies are non-toxic, mechanically stable and functional protein amyloids within the nanoscale size range that are able to naturally penetrate into mammalian cells, where they deliver the embedded protein in a functional form. The potential use of inclusion bodies in protein delivery or protein replacement therapies is strongly impaired by the absence of specificity in cell binding and penetration, thus preventing targeting. To address this issue, we have here explored whether the genetic fusion of two tumor-homing peptides, the CXCR4 ligands R9 and T22, to an inclusion body-forming green fluorescent protein (GFP), would keep the interaction potential and the functionality of the fused peptides and then confer CXCR4 specificity in cell binding and further uptake of the materials. The fusion proteins have been well produced in Escherichia coli in their full-length form, keeping the potential for fluorescence emission of the partner GFP. By using specific inhibitors of CXCR4 binding, we have demonstrated that the engineered protein particles are able to penetrate CXCR4+cells, in a receptor-mediated way, without toxicity or visible cytopathic effects, proving the availability of the peptide ligands on the surface of inclusion bodies. Since no further modification is required upon their purification, the biological production of genetically targeted inclusion bodies opens a plethora of cost-effective possibilities in the tissue-specific intracellular transfer of functional proteins through the use of structurally and functionally tailored soft materials.
authors.affiliation: Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193 Barcelona, Spain. Department de Genètica i de Microbiologia, Universitat Autònoma de Barcelona, Bellaterra, 08193 Barcelona, Spain. CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Bellaterra, 08193 Barcelona, Spain.
authors.forename: Ugutz
authors.initials: U
authors.lastname: Unzueta
authors.forename: Joaquin
authors.initials: J
authors.lastname: Seras-Franzoso
authors.forename: María Virtudes
authors.initials: MV
authors.lastname: Céspedes
authors.forename: Paolo
authors.initials: P
authors.lastname: Saccardo
authors.forename: Francisco
authors.initials: F
authors.lastname: Cortés
authors.forename: Fabián
authors.initials: F
authors.lastname: Rueda
authors.forename: Elena
authors.initials: E
authors.lastname: Garcia-Fruitós
authors.forename: Neus
authors.initials: N
authors.lastname: Ferrer-Miralles
authors.forename: Ramon
authors.initials: R
authors.lastname: Mangues
authors.forename: Esther
authors.initials: E
authors.lastname: Vázquez
authors.forename: Antonio
authors.initials: A
authors.lastname: Villaverde
mesh_terms: D000682:Amyloid / Q000008:administration & dosage* / Q000737:chemistry* / Q000378:metabolism; D045744:Cell Line, Tumor; D016503:Drug Delivery Systems / Q000379:methods; D004926:Escherichia coli / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D002479:Inclusion Bodies / Q000737:chemistry* / Q000378:metabolism; D049329:Nanostructures / Q000008:administration & dosage* / Q000737:chemistry*; D009369:Neoplasms / Q000188:drug therapy*; D010455:Peptides / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism; D019718:Receptors, CXCR4 / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism
chemical_list: D000682:Amyloid; D010455:Peptides; D019718:Receptors, CXCR4; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins