44 research outputs found
A Plug-and-Play Approach for the De Novo Generation of Dually Functionalized Bispecifics
Diseases are multifactorial, with redundancies and synergies between various pathways. However, most of the antibody-based therapeutics on the market interact with only one target, thus limiting their efficacy. The targeting of multiple epitopes could improve the therapeutic index of treatment and counteract mechanisms of resistance. To this effect, a new class of therapeutics has emerged: bispecific antibodies. Bispecific formation using chemical methods is rare and low-yielding and/or requires a large excess of one of the two proteins to avoid homodimerization and heterogeneity. In order for chemically prepared bispecifics to deliver their full potential, high-yielding, modular, and reliable cross-linking technologies are required. Herein, we describe a novel approach not only for the rapid and high-yielding chemical generation of bispecific antibodies from native antibody fragments, but also for the site-specific dual functionalization of the resulting bioconjugates. Based on orthogonal clickable functional groups, this strategy enables the assembly of functionalized bispecifics with controlled loading in a modular and convergent manner
Employing defined bioconjugates to generate chemically functionalised gold nanoparticles for in vitro diagnostic applications
Novel methods for introducing chemical and biological functionality to the surface of gold nanoparticles serve to increase the utility of this class of nanomaterials across a range of applications. To date, methods for functionalising gold surfaces have relied upon uncontrollable non-specific adsorption, bespoke chemical linkers, or non-generalisable proteinâprotein interactions. Herein we report a versatile method for introducing functionality to gold nanoparticles by exploiting the strong interaction between chemically functionalised bovine serum albumin (f-BSA) and citrate-capped gold nanoparticles (AuNPs). We establish the generalisability of the method by introducing a variety of functionalities to gold nanoparticles using cheap, commercially available chemical linkers. The utility of this approach is further demonstrated through the conjugation of the monoclonal antibody Ontruzant to f-BSAâAuNPs using inverse electron-demand DielsâAlder (iEDDA) click chemistry, a hitherto unexplored chemistry for AuNPâIgG conjugation. Finally, we show that the AuNPâOntruzant particles generated via f-BSAâAuNPs have a greater affinity for their target in a lateral flow format when compared to conventional physisorption, highlighting the potential of this technology for producing sensitive diagnostic tests
Modular Chemical Construction of IgG-like Mono- and Bispecific Synthetic Antibodies (SynAbs)
In recent years there has been rising interest in the
field of proteinâprotein conjugation, especially related to bispecific
antibodies (bsAbs) and their therapeutic applications. These
constructs contain two paratopes capable of binding two distinct
epitopes on target molecules and are thus able to perform complex
biological functions (mechanisms of action) not available to
monospecific mAbs. Traditionally these bsAbs have been
constructed through protein engineering, but recently chemical
methods for their construction have started to (re)emerge. While
these have been shown to offer increased modularity, speed, and for
some methods even the inherent capacity for further functionalization (e.g., with small molecule cargo), most of these approaches lacked the ability to include a fragment crystallizable (Fc) modality.
The Fc component of IgG antibodies offers effector function and increased half-life. Here we report a first-in-class disulfide
rebridging and click-chemistry-based method for the generation of Fc-containing, IgG-like mono- and bispecific antibodies. These
are in the FcZ-(FabX)-FabY format, i.e., two distinct Fabs and an Fc, potentially all from different antibodies, attached in a
homogeneous and covalent manner. We have dubbed these molecules synthetic antibodies (SynAbs). We have constructed a T cellengager (TCE) SynAb, FcCD20-(FabHER2)-FabCD3, and have confirmed that it exhibits the expected biological functions, including the
ability to kill HER2+ target cells in a coculture assay with T cells