Preclinical Validation For Treatment With Rg7787 In Ovarian Cancer

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PARG7787 is composed of a Fab fragment from an anti-MSLN antibody fused to a de-immunized and truncated pseudomonas endotoxin A variant. Once RG7787 is bound and internalized by MSLN positive cells, the toxin is transported to the cytosol, where it inhibits protein synthesis, eventually causing tumor cell death.Mesothelin (MSLN) is a tumor specific differentiation antigen. On normal tissue, its expression is restricted to differentiated mesothelial cells that line as single cell layer body cavities and major organs (e.g. pleura, pericardium, and peritoneum). In cancer, MSLN is highly expressed not only on mesotheliomas but also on a number of other types of solid tumors like ovarian and pancreatic cancer. In both indications patients frequently also have significant serum levels of the cancer antigen-125 (CA-125), sometimes even u003e1000 U/ml.MSLN has been described to bind CA-125 and this interaction has been suggested to play a role for the ability of cancer cells to metastasize e.g. to the peritoneum. The region in MSLN that is responsible for its interaction with CA-125 has been reported to overlap with the binding epitope of RG7787 potentially resulting in competition between CA-125 and RG7787 for binding to MSLN. The high percentage of MSLN positive cases as well as a clear unmet medical need makes ovarian and pancreatic cancer promising indications for clinical development of RG7787. We investigated, whether abundance of CA-125 can negatively affect ability of RG7787 to bind and be taken up by tumor cells thereby antagonizing RG7787 treatment.We found that indeed RG7787 competes with CA-125 for binding to mesothelin. However, in SPR experiments, the interaction of MSLN with CA-125 was not strong enough to block binding of the anti-MSLN Fab moiety that targets RG7787 to tumor cells. At 20°C the absolute affinity (KD) of RG7787 for human MSLN was determined to be 12.5 pM using two orthogonal “affinity in solution” methods, ELISA and SPR. In agreement with such high affinity binding, we found that adding soluble CA-125 to cell viability assays did not reduce the cytotoxic potency of RG7787. In one set of assays, ascites fluid containing a 10 fold excess of CA-125 compared to the average levels observed in sera of ovarian patients was used for competition. In another set of assays, we used a 100 fold excess of a truncated recombinant CA-125 fragment, containing the domain that interacts with MSLN. Neither the ascites fluid, nor the recombinant CA-125 fragment had any effect on the in-vitro potency of RG7787, indicating that soluble CA-125 levels in patients will not antagonize RG7787 treatment. Finally we demonstrated in cell-cell interaction assays that RG7787 can efficiently block the attachment of MSLN positive cells to OVCAR3 cells that express high levels of CA-125 on their surface. Based on these encouraging data we are currently evaluating in-vivo efficacy of RG7787 in a patient-derived ovarian cancer model.Citation Format: Gwendlyn Kollmorgen, Klara Palme, Annette Seidl, Stefan Scheiblich, Christian Clemens, Edgar Voss, Martin Kaufmann, Klaus Hirzel, Pamela Wilfert, Moritz Marcinowski, Bernd Satzinger, Frank Herting, Gerhard Niederfellner. Preclinical validation for treatment with RG7787 in ovarian cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5321. doi:10.1158/1538-7445.AM2015-5321