The use of antibody engineering combined with subcellular trafficking analyses to design therapeutic antibodies in two areas will be discussed: First, the development of engineered antibodies that clear pathogenic antibodies. Second, the design of antibody-drug conjugates (ADCs) that deliver their cytotoxic payload more efficiently to lysosomes within cells, resulting in a strategy to generate ADCs that are effective at lower doses.

Oncology targets Claudin 6 (CLDN6) and 18.2 (CLDN18.2) are overexpressed in select cancers but are absent in most adult healthy tissues. Using our MPS Antibody Discovery platform, we discovered and humanized lead candidate antibodies that bind unique residues on CLDN6 but not closely related receptors CLDN9, CLDN3, and CLDN4, or members of the human proteome. We also present a panel of humanized CLDN18.2 MAbs with picomolar affinities that are superior to the clinical-stage benchmark.

We discovered and developed the concept that auto reactivity in arthritis is against neoepitopes that is formed as a result of oxidative post translational modification (oxPTM) collagen type II (oxPTM-CII) that are formed specifically in the arthritic joints. Hence, autoantibodies to oxPTM-CII are biomarkers that can be utilised for early diagnosis and for stratification of patients with RA. We showed that specific targeting of a payload drug by anti-oxPTM-CII antibody to inflamed arthritic joints (Etanercept or viral IL-10) and hence nanoparticles loaded with multiple drugs. This resulted in significantly enhanced therapeutic effecacy compared with non-targeted systemic treatments. We also showed that anti-oxPTM-CII antibodies can detect very early OA before evident cartilage erosion.

We used an antibody specific to oxidative post-translationally modified type II collagen (oxPTM-CII) to stain rat cartilage in two different chronic osteoarthritis models. Several time points were evaluated as early as one day post operation. We could observe that oxPTM-CII staining is mainly localized in the deep zone and is detectable before the appearance of cartilage lesions. It indicates that oxPTM-CII could serve to detect early osteoarthritis.

We found that mast cell tryptase levels are elevated in severe asthma patients independent of type 2 inflammation which correlates with active ß-tryptase allele count. We discovered an inhibitory antibody against human ß-tryptase, that dissociates active tryptase tetramers into inactive monomers. A co-crystal structure along with biochemical studies reveal the molecular basis for inhibition. This antibody inhibits tryptase activity in two in vivo models, providing a foundation for clinical development.

Activating Natural Killer (NK) cell receptors represent promising target structures to elicit potent anti-tumor immune responses. Here, novel immunoligands were generated that bridge the activating NK cell receptor NKp30 on NK cells with epidermal growth factor receptor (EGFR) on tumor cells in a bispecific IgG-like format based on affinity-optimized versions of B7-H6 and the Fab arm derived from Cetuximab. To enhance NKp30 binding, the solitary N‑terminal IgV domain of B7-H6 (DB7-H6) was affinity matured by an evolutionary library approach combined with yeast surface display. Biochemical and functional characterization of 36 of these novel DB7‑H6-derived NK cell engagers revealed an up to 45-fold enhanced affinity for NKp30 and significantly improved NK cell-mediated, EGFR-dependent killing of tumor cells compared to the NK cell engagers based on the wild-type DB7-H6 domain. In this regard, potencies (EC₅₀ killing) of the best immunoligands were substantially improved by up to 87‑fold. Moreover, release of interferon-γ (IFN-g) and tumor necrosis factor-α (TNF-α) was significantly increased. Importantly, equipment of the DB7-H6-based NK cell engagers with a human IgG1 Fc part competent in Fc receptor binding resulted in an almost 10-fold superior killing of EGFR-overexpressing tumor cells compared to molecules either triggering FcgRIIIa or NKp30. Additionally, INF-g and TNF-α release was increased compared to molecules solely triggering FcgRIIIa including the clinically approved antibody Cetuximab. Thus, incorporating affinity-matured ligands for NK cell activating receptors might represent an effective strategy for the generation of potent novel therapeutic agents with unique effector functions in cancer immunotherapy.

Complex cellular targets such as GPCRs, ion channels, and other multi-transmembrane proteins represent a significant challenge for therapeutic antibody discovery, primarily because of poor stability of the target protein upon extraction from cell membranes.  Here we have used different membrane-bound antigen formats to identify and optimise anti CCR-1 functional antibodies using an in vitro yeast-based antibody discovery platform (Adimab^TM). These data exemplify a methodology to generate potent fully-human mAbs for challenging targets rapidly using whole cells as antigen and also define a route to the identification of affinity-matured variants.

The antibody drug development field suffers from a crowded target space and an approach for discovery of both novel antibodies and targets is phenotypic screening, using phage display and selection on whole cells, followed by functional testing and target deconvolution. In this strategy, the generated pool of antibodies will be very complex and by applying various mining technologies antibodies against a broad range of cell surface receptors are discovered.

We will present Icosagen's latest breakthrough in developing highly potent virus neutralizing antibodies against SARS-CoV-2 based on multiple antibody formats. We demonstrate the implementation of the Hybrifree technology in rapidly developing antibodies with drug like properties and highly potent virus neutralizing efficacy. 

The in vitro affinity and/or functional maturation of naïve antibodies is common practice. In most cases, targeted introduction of sequence diversity into a limited number of complementarity determining region (CDR) loops coupled with selection for improved variants through phage or ribosome display is sufficient to deliver the required affinity or functional improvements. Occasionally, ‘hard to mature’ clones are seen that are inherently intractable to optimisation, necessitating a more heuristic, unbiased approach to achieve the desired improvements. In this talk, I will describe the use of ribosome display to optimise these ‘hard to mature’ clones, using the affinity optimisation of a inhibitory antibody to human Arginase 2 as a case study. This work exemplifies the application of novel Shuffle and Shuffle/StEP libraries as well as pool maturation and error-prone libraries to deliver significant improvements in potency, affinity and mode of binding, that would not be achievable through more conventional methods.

Developing human neutralizing antibodies (nAbs) against emerging viral pathogens can be seriously delayed due to limited access of blood specimens from acute/convalescent infected individuals due to privacy, public safety, regulatory and national security concerns. However, seminal discoveries of nAbs for emerging coronaviruses have also been made the old-fashioned way through antibody phage display using non-immune libraries. I will discuss the lessons that we have learned for nAb discovery during these outbreaks.

We have built a platform to rapidly isolate anti-influenza antibodies in as little as 10 days. We then optimized parallel manufacturing workflows to deliver one of these antibodies by plasmid DNA, mRNA and AAV platforms that afforded mice complete protection from lethal challenge. The complete process was completed in approximately 60 days and could form the basis for a rapid therapeutic response to upcoming pandemics.