Peptide microarrays enable rapid mimotope optimization for pharmacokinetic analysis of the novel therapeutic antibody IMAB362

Abstract

As membrane proteins play an important role in a variety of life-threatening diseases, the development of therapeutic monoclonal antibodies against membrane proteins is of significant interest. Among many other requirements, the process of antibody drug development requires a set of tailor-made assays for the characterization of the antibodies and for monitoring their activity. Designing assays to characterize antibodies directed to membrane proteins is challenging, because the natural targets are often not available in a format that is compatible with a biochemical assay setup. Thus, alternatives that mimic the targeted membrane proteins are needed. In this study, we developed optimal peptidic mimotopes for the ELISA-based detection of the novel therapeutic antibody IMAB362 in biological samples. Initial hits were identified using phage display and these hits were optimized with the help of structure-activity relationship analysis on peptide microarrays. The optimized peptides showed binding constants in the low nanomolar to picomolar range, an improvement by a factor of up to 30 compared to the initial hits. The best mimotope (apparent KD = 0.15 nM) was successfully used for the ELISA-based quantification of IMAB362 in samples from a mouse pharmacokinetic study. The process described allows the rapid discovery of mimotopes for target proteins that are difficult to produce or handle, which can then be used in pre-clinical and clinical assays or for the purification of biological products.

Claudin 18.2 as a New Biomarker in Gastric Cancer—What Should We Know?

“…Initial trials aimed at monoclonal antibodies against CLDN18.2 and zolbetuximab (claudiximab and previously named IMAB362), which were some of the first agents to be studied. Zolbetuximab is a mouse chimeric monoclonal antibody with human IgG1 constant regions designed to bind specifically to CLDN18.2 (Figure 1) [66] with high affinity without cross-binding to other claudin family members [67,68]. Pre-clinical data demonstrate that zolbetuximab elicited antibody-dependent cell-mediated cytotoxicity in CLDN18.2-expressing GC cell lines and also induced complement-dependent cytotoxicity in CLDN18.2-positive GC cells [37].…”

The role of claudin 18.2 and HER-2 in pancreatic cancer outcomes

“…[ 8 ] The monoclonal antibody zolbetuximab, discovered by Schnatbaum, binds to cancer cells that are CLDN18.2 positive and leads to cancer cell death by antibody-dependent cellular toxicity and complement-dependent cytotoxicity. [ 9 ] In the phase 2a MONO study, conducted by Türeci, [ 10 ] and phase 2 FAST study conducted by Sahin, [ 11 ] zolbetuximab was effective in patients with metastatic gastric and esophageal adenocarcinoma. Clinical trials of zolbetuximab in the first-line setting in combination with chemotherapy and immunotherapy, are ongoing worldwide for metastatic gastric and gastroesophageal junction cancer (NCT03505320, NCT03504397, NCT03653507).…”

Alanine Scanning of the Unstructured Region of Ara h 2 and of a Related Mimotope Reveals Critical Amino Acids for IgE Binding

“…[22] Measuring IgE binding to the DPYSP motif of Ara h 2 enhances in vitro diagnositics. [25] To determine the core sequence of epitope 3 (epitope 3-core) and the relative importance of amino acids throughout its sequence and throughout the sequence of a mimotope that contains the central DPY_P motif of epitope 3, we performed standard ELISAs and then conducted alanine scanning [26][27][28][29] using microarrays. The relevance of these studies was enhanced by changing the "P" of the DPYSP motif in epitope 3 and in the mimotope to "h" prior to alanine scanning.…”