InflaMab: A novel NLRP3 Inflammasome inhibitor provides neuroprotection in a mouse model of glaucoma

Purpose : Recent evidence shows an essential role for NLRP3 inflammasome activation in the pathogenesis of glaucoma and we previously reported that NLRP3 knockout mice were protected from axon degeneration and death of RGCs in an inducible mouse model of glaucoma. These data support pharmacologic inhibition of NLRP3 as a potential neuroprotective therapy in glaucoma. However, the short half-life of the currently available small molecule inhibitors of NLRP3 make them poor candidates for local intravitreal administration, the ideal route of administration for glaucoma. The aim of this study is to characterize the anti-inflammatory effects of a novel biologic (InflaMab) that targets the NLRP3 inflammasome and to determine its’ neuroprotective potential in an inducible mouse model of glaucoma. Methods : InflaMab is a bi-specific antibody that was developed to gain access to the intracellular compartment and target NLRP3. The anti-inflammatory potential of InflaMab was tested in vitro in a THP1 macrophage cell line and murine bone marrow-derived macrophages (BMDMs). IL-1β and TNFα secretion was measured using ELISA and Caspase-1 activation was assessed by staining cells with a Fluorescent Inhibitor of Caspase-1 (FLICA). In vivo, the neuroprotective effect of InflaMab was assessed in a microbead-induced mouse model of glaucoma. WT C57BL/6J mice received one intravitreal injection of InflaMab (250 ng/ml) on Day 0 (just prior to microbead injection). IOP was monitored by rebound tonometry and at Day 28 post microbead injection pattern electroretinogram (pERG) was performed to assess RGC function and retinal whole mounts and optic nerves were prepared for RGC and axon quantification. Results : In vitro, InflaMab significantly inhibited IL-1β secretion in THP1 cells and BMDMs (p<0.05) and had no inhibitory effect on TNFα production, demonstrating specificity. Confocal microscopy revealed a significant reduction of caspase-1 activation in THP1 cells treated with InflaMab as opposed to non-treated cells. In vivo, a single intravitreal injection of Inflamab provided significant neuroprotection in terms of RGC function (pERG amplitude) and RGC and axon survival when compared to untreated controls Conclusions : These data demonstrate that (i) InflaMab inhibits inflammasome activation and (ii) intravitreal administration of InflaMab provides sustained neuroprotection in an inducible mouse model of glaucoma.