Growth arrest specific 6 (Gas6) is a vitamin-K dependent protein that is involved in the regulation of a wide array of cellular activities, including adhesion, migration, immune evasion, and apoptosis. Gas6 as well as its receptors Axl, Mer, and Tyro3 are highly expressed in multiple malignancies suggesting that signaling through this axis may be tumor promoting in human disease. A strategy was devised to generate fully human neutralizing antibodies against Gas6 using XenoMouse technology. Hybridoma supernatants were first screened for their ability to bind Gas6, then for their ability to inhibit Gas6 binding to the Axl receptor and block Gas6-induced Axl phosphorylation in human cells. Two purified antibodies isolated from the screened hybridomas, 1D9.1 and 1G9.1, maintained potency in all 3 assays and also inhibited Gas6-induced cellular migration and proliferation. In the scratch wound migration assay, the antibodies inhibited both basal and Gas6-induced wound closure to a greater extent than the soluble extracellular domain of the receptor Axl (Axl- Fc). In the proliferation assay, the antibody 1D9.1 completely inhibited Gas6-induced proliferation with a half maximal effective concentration (EC50) of 0.15nM. In vivo characterization of one of the antibodies, 1D9.1, was conducted using a pharmacodynamic (PD) assay that measured the ability of the antibody to inhibit Gas6-induced Akt activation in the mouse spleen. Treatment of mice with a single dose (100-1000 μg) of 1D9.1 led to greater than 90% inhibition of Gas6-induced phosphorylated Akt (pAkt) for up to 72 hours. Based on the target coverage observed in the PD assay, we tested the efficacy of 1D9.1 against Panc-1 human pancreatic adenocarcinoma xenografts implanted in athymic nu/nu female mice. At doses of 50 and 150 μg, twice weekly, the Gas6 neutralizing antibody 1D9.1 was able to inhibit 55% and 76% of tumor growth, respectively. (p<0.001 for both treatments vs. control Ig). When combined with gemcitabine, 1D9.1 was able to inhibit tumor growth to a greater extent than either agent alone (p<0.001 vs. either monotherapy). Together, the data suggest that Gas6 neutralization should be further explored as a potential strategy for the treatment of pancreatic cancer.

Citation Format: Gordon E. Moody, Brian Belmontes, Chadwick King, Shuying Liu, Stephanie Masterman, Chris Murawsky, Trace Tsuruda, Wei Wang, Robert Radinsky, Pedro J. Beltran. Generation of a fully human Gas6 neutralizing antibody with anti-tumor activity in vivo. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5158. doi:10.1158/1538-7445.AM2013-5158