title: Small synthetic ligands for the enrichment of viral particles pseudotyped with amphotropic murine leukemia virus envelope.
abstract: Retroviral vectors gained popularity toward other viral vectors as they integrate their genome into hosts' genome, a characteristic required for the modification of stem cells. However, the production of viable particles for gene therapy is hampered by the low ratio of infectious to non-infectious viral particles after purification, low titers and limited number of competent viral receptors. We have developed de novo two fully synthetic triazine-based ligands that can selectively bind retroviral particles pseudotyped with amphotropic murine leukemia virus envelope (AMPHO4070A). A 78-membered library of triazine-based ligands was designed in silico and was virtually screened against the modeled structure of the AMPHO4070A protein. Ligands displaying the highest energy of binding were synthesized on cross-linked agarose and experimentally tested. Adsorbents containing ligands A5A10 and A10A11 showed selectivity toward viral particles containing the target protein (VLP-AMPHO), binding 19 ± 5 μg/g support and 47 ± 13 μg/g support, respectively. The elution conditions for both ligands were mild and with high recovery yields (80-100%), in comparison with common purification practices. These results were based on a lab-scale experimental setting with VLP integrity being confirmed through TEM. In particular, the elution buffer containing 12 mM imidazole allowed the recovery of intact amphotropic viral particles.
authors.affiliation: UCIBIO, REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal.
authors.forename: Cláudia S M
authors.initials: CS
authors.lastname: Fernandes
authors.affiliation: iBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, Portugal.
authors.forename: Rute
authors.initials: R
authors.lastname: Castro
authors.affiliation: iBET, Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2780-901 Oeiras, Portugal.
authors.forename: Ana Sofia
authors.initials: AS
authors.lastname: Coroadinha
authors.affiliation: UCIBIO, REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal. Electronic address: cecilia.roque@fct.unl.pt.
authors.forename: A Cecília A
authors.initials: AC
authors.lastname: Roque
mesh_terms: D000818:Animals; D005822:Genetic Vectors / Q000302:isolation & purification; D009052:Leukemia Virus, Murine / Q000737:chemistry*; D008024:Ligands; D051379:Mice; D011991:Receptors, Virus; D012190:Retroviridae / Q000302:isolation & purification / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism*; D014771:Virion / Q000145:classification / Q000302:isolation & purification*; D014773:Virology / Q000379:methods*
chemical_list: D008024:Ligands; D011991:Receptors, Virus; D014759:Viral Envelope Proteins
keywords: Purification; Triazine scaffold; Viral particles; Virtual screening