Construction of a pIX-modified Adenovirus Vector Able to Effectively Bind to Nanoantibodies for Targeting

Abstract

Current targeting strategies for genetic vectors imply the creation of a specific vector for every targeted receptor, which is time-consuming and expensive. Therefore, the development of a universal vector system whose surface can specifically bind molecules to provide efficient targeting is of particular interest. In this study, we propose a new approach in creating targeted vectors based on the genome of human adenovirus serotype 5 carrying the modified gene of the capsid protein pIX (Ad5-EGFP-pIX-ER): recombinant pseudoadenoviral nanoparticles (RPANs). The surfaces of such RPANs are able to bind properly modified chimeric nanoantibodies that specifically recognize a particular target antigen (carcinoembryonic antigen (CEA)) with high affinity. The efficient binding of nanoantibodies (aCEA-RE) to the RPAN capsid surfaces has been demonstrated by ELISA. The ability of the constructed vector to deliver target genes has been confirmed by experiments with the tumor cell lines A549 and Lim1215 expressing CEA. It has been shown that Ad5-EGFP-pIX-ER carrying aCEA-RE on its surface penetrates into the tumor cell lines A549 and Lim1215 via the CAR-independent pathway three times more efficiently than unmodified RPAN and Ad5-EGFP-pIX-ER without nanoantibodies on the capsid surface. Thus, RPAN Ad5-EGFP-pIX-ER is a universal platform that may be useful for targeted gene delivery in specific cells due to “nanoantibody–modified RPAN” binding.

Nanobodies—Useful Tools for Allergy Treatment?

“…They include increasing the hydrodynamic radius of a protein by attaching highly flexible and hydrophilic molecules such as polyethylene glycol (PEG) and carbohydrates or by genetic fusion with polypeptide chains mimicking the biochemical properties of PEG, fusion of V H H to the Fc region of IgG, fusion or non-covalent binding to albumin ( 104 ) ( Figure 2D ). Nanobodies can also be used as modules to engineer larger molecules with several valencies and/or specificities, such as multivalent ( 105 – 108 ), bispecific ( 105 , 109 ), and other ( 110 , 111 ) constructs that may acquire considerably higher specificity, binding efficiency and biological activity ( 106 , 107 , 111 ). Nanobodies were also considered as possible ligands to design new highly specific immunosorbents ( 112 – 114 ).…”

Oncolytic Adenoviruses: Strategies for Improved Targeting and Specificity

“…However, expressing these retargeting molecules separately and then assembling them with virus particles into a functional complex before delivery has not been easy either. Synthetic leucine-zipper-based dimers [146,147] and bispecific T-cell engagers (BiTEs) are examples of retargeting molecules that may need extensive maturation. Several studies have tried to use them to increase tumor specificity, however the overall outcomes have not been significant [147][148][149][150][151][152][153][154][155].…”

“…Antibody-antibody-based bridging molecules Anti-AdV protein conjugated to VEGFR, TIE-2, integrins, EPCAM, EGFR, HER2, Endoglin, HMWMAA [127][128][129][130][131][132] Antibody-ligand-based bridging molecules anti-AdV protein conjugated to Folate, TNFa, IGF1, EGF [133,134] Peptide-antibody-based bridging molecules p75 neurotropin receptor on hepatic stellate cells [135] Soluble receptors conjugated to ligands (sCAR, FX) EGF, anti-Cd40 ScFv, ApoE ligand, anti-ErbB2, anti-CEA, Polysialyc-acid (PSA), CXCL12 [136][137][138][139][140][141][142][143][144][145] BiTE, Leucine-zipper-based linkers CD44v6, anti-B-cell maturation antigen, FR-α, EGFR, EpCAM, carcinoembryonic antigen, CD40 [147][148][149][150][151][152][153][154] b. Selective Expression of Effectors (Inhibitors/Enhancers) in Specific Cells.…”

A novel capsid-modified oncolytic recombinant adenovirus type 5 for tumor-targeting gene therapy by intravenous route

“…We hypothesized that a tumor-targeted Ad vector can be achieved via highly specific association with secreted bioactive TRAIL proteins by employing synthetic leucine zipper-like dimerization domains (zippers) that have been optimized for structural compatibility between the Ad capsid and TRAIL. The feasibility and effectiveness of such strategy has been confirmed by M. N. Garas' study recently [26]. In this report, based on a 24-base-pair deletion mutant E1A oncolytic Ad (Δ24E1A) [27–29], we outlined the biochemical analysis, functional validation and anti-tumor activity of a novel TRAIL-modified Ad vector and demonstrated that this engineered Ad virion with TRAIL on the surface could target cancer tissues administered by IV injection in vivo .…”