Introduction: After stroke, penumbral cells express neo-epitopes that are recognized by natural IgM antibodies (NIgM) leading to complement activation and propagation of inflammation and injury. We previously identified NIgMs, namely B4IgM, that only recognizes ischemic cells, and developed a site-targeted complement inhibitor by fusing a single chain antibody derived from B4mAb to the complement inhibitor Crry. We hypothesize that the fusion construct B4Crry inhibits neurodegenerative effects of IgM and complement and improves outcome after stroke.
Methods: Stroke was induced in adult and aged mice by 60 min right MCA occlusion. B4Crry was administered intravenously at 2-24 hours after stroke. Animals were tested for infarct volume and performance on motor and cognitive tasks. Brains were extracted for histological analyses.
Results: A single dose of B4Crry 6-24 hours post-ischemia specifically targeted to the ischemic hemisphere and inhibited complement and IgM deposition in the penumbra, yielding significant reduction in infarct volumes, neuronal loss and neurological deficits (p<0.01) at 24hrs in adult males, females and aged mice. B4Crry-mediated neuroprotection persisted throughout 15 days of recovery yielding reduced forelimb laterality (p<0.01), improved spatial learning and memory (Barnes maze, p<0.05), and improved skilled handling (Pasta task, p<0.01) compared to vehicle. Vehicle treated animals showed a sustained inflammatory response with continuous IgM and complement deposition and robust proinflammatory microglial activation detected 15 days post-stroke. Acute B4Crry therapy interrupted the chronic neuroinflammatory cycle by significantly inhibiting complement and IgM deposition, and shifted microglial polarity to anti-inflammatory phenotypes resulting in preserved penumbral neuronal density. Simultaneously, B4Crry resulted in pronounced increase in neurogenesis and neuronal migration. Finally, we show that B4Crry bound specifically to the ischemic penumbra of postmortem brain samples obtained from acute stroke patients, but not to normal brain tissue from the same patient.
Conclusions: B4Crry is a novel translational therapeutic agent that interrupts neurodegenerative inflammatory cascades and improves outcomes after stroke.
Patient Care: Work presented here describes a novel anti-inflammatory therapeutic construct (B4Crry) that can target specifically to the human ischemic brain after stroke resulting in inhibition of inflammation and improved outcomes as well as a novel approach to deliver therapeutics locally to the ischemic brain avoiding systemic complications. The construct, B4Crry, is currently under preclinical development for potential translation as a new treatment for ischemic stroke patients especially given its wide therapeutic window of at least 24 hrs. after stroke.
Learning Objectives: By the conclusion of this session, participants should be able to:
1) Identify the role of natural IgM antibodies and complement activation in propagating secondary injury after ischemic stroke
2) Describe the dual effects of limiting neuroinflammatory cascades by reducing injury and damage and removing the brakes off regenerative processes after stroke
3) Identify the utility of innate self-recognition systems that are conserved between human and mice to deliver therapeutics locally to sites of ischemia and injury