Christophe de Romeuf

E23196 Background: Expression of anti-Müllerian hormone receptor (AMHRII) is described in gynecologic cancers, tumors which are largely infiltrated by immune myeloid cells. Based on these observations, a humanized glyco-engineered monoclonal antibody (GM102) able to increase effector cells engagement has been developed displaying a high affinity towards AMHR2 (subnanomolar) and CD16 (nanomolar), and triggering enhanced ADCC and phagocytosis by M1 and M2 macrophages. Methods: In vivo antitumor activity of GM102 alone and in combination with first line combo (Paclitaxel + Carboplatin) has been evaluated in a panel of AMHRII+ ovarian PDXs. Safety profile of GM102 was documented during a regulatory toxicology study in cynomolgus with repeated IV infusions up to 300 mg/kg/day every two weeks for eight weeks, associated with ancillary pharmacokinetics (PK) studies. Human and monkey tissues were tested for tissue cross reactivity (TCR) studies. Results: A meta-analysis of the fivePDXs studied showed a significant effect on response rate (p = 0.018) in favor of combo + GM102 versus combo alone. The overall CR rates reached 32% with GM102 versus 12% with combo alone. In cynomolgus, GM102 did not elicit any signs of toxicity with a NOAEL ≥ 300 mg/kg. Very low levels of Anti-Drug Antibodies were found in two females indicating no immunogenic potential. TCR studies showed GM102 positive binding in reproductive organs. Few off-target binding (myocardium, lens) were observed and shown of no toxicological significance. The PK of GM102 consisted of low clearance and distribution volume, prolonged half-life (177 hours), and dose-proportional increase in exposure. Conclusions: Altogether, this preclinical profile supports the current clinical development (Phase I) of GM102 in gynecological malignancies expressing AMHRII, a novel target not yet addressed by cancer therapeutics.

While glyco-engineered monoclonal antibodies (mAbs) with improved antibody-dependent cell-mediated cytotoxicity (ADCC) are reaching the market, extensive efforts have also been made to improve their pharmacokinetic properties to generate biologically superior molecules. Most therapeutic mAbs are human or humanized IgG molecules whose half-life is dependent on the neonatal Fc receptor FcRn. FcRn reduces IgG catabolism by binding to the Fc domain of endocytosed IgG in acidic lysosomal compartments, allowing them to be recycled into the blood. Fc-engineered mAbs with increased FcRn affinity resulted in longer in vivo half-life in animal models, but also in healthy humans. These Fc-engineered mAbs were obtained by alanine scanning, directed mutagenesis or in silico approach of the FcRn binding site. In our approach, we applied a random mutagenesis technology (MutaGenTM) to generate mutations evenly distributed over the whole Fc sequence of human IgG1. IgG variants with improved FcRn-binding were then isolated from these Fc-libraries using a pH-dependent phage display selection process. Two successive rounds of mutagenesis and selection were performed to identify several mutations that dramatically improve FcRn binding. Notably, many of these mutations were unpredictable by rational design as they were located distantly from the FcRn binding site, validating our random molecular approach. When produced on the EMABling® platform allowing effector function increase, our IgG variants retained both higher ADCC and higher FcRn binding. Moreover, these IgG variants exhibited longer half-life in human FcRn transgenic mice. These results clearly demonstrate that glyco-engineering to improve cytotoxicity and protein-engineering to increase half-life can be combined to further optimize therapeutic mAbs.

Summary: Patients with chronic lymphocytic leukaemia (CLL) treated with a combination of fludarabine, cyclophosphamide and rituximab show a high response rate. However, only a poor response is observed following rituximab monotherapy. The use of chemo‐immunotherapy is often associated with haematological and infectious complications. Thus, an antibody with an enhanced ability to kill CLL cells could lead to better clinical responses to antibody monotherapy and the possibility of lowering drug doses during chemo‐immunotherapy. We generated a chimeric anti‐CD20 monoclonal antibody (mAb), EMAB‐6, which has a low fucose content. Apoptosis and complement activities for EMAB‐6 were similar to those seen for rituximab. By contrast, EMAB‐6 mAb showed improved Fcγ receptor IIIA (FcγRIIIA)/CD16 binding and FcγRIIIA‐dependent effector functions. It induced a higher in vitro antibody‐dependent cellular cytotoxicity against CLL cells and a higher FcγRIIIA‐mediated interleukin‐2 production by FcγRIIIA⁺ Jurkat cells in the presence of CLL cells at both low and maximally saturating concentrations. Comparative studies between CLL and lymphoma cells coated with EMAB‐6 or rituximab indicated that the difference of efficacy was more pronounced at low doses and when target cells expressed fewer CD20 molecules. Thus, EMAB‐6 mAb represents a promising drug candidate for the treatment of CLL by inducing a strong cytotoxicity against tumour cells that express low CD20 levels.