Endogenous Zinc Protoporphyrin in Intracerebral Hemorrhage

Project Summary Intracerebral hemorrhage (ICH) is a leading cause of mortality. Despite extensive efforts, current therapeutic interventions targeting recognized causes of ICH-induced brain damage have been ineffective. Our recent study revealed an intriguing finding: endogenously formed zinc protoporphyrin (ZnPP) significantly contributes to ICH brain damage. Our preliminary results show that ICH-generated ZnPP accumulated in brain tissue surrounding the hematoma, which was dependent on ferrochelatase (FECH) catalysis. Notably, inhibiting FECH markedly reduced ZnPP generation, decreased brain damage, and improved long-term outcomes. However, little is known about the factors that are required for ZnPP formation, nor the mechanisms by which ZnPP exerts its neurotoxicity in ICH. Building upon our published and preliminary findings, our overarching hypothesis is that ICH causes a hypoxic rim around hemorrhagic area, triggering ROS generation, free Zn2+ release and iron oxidation, and promoting FECH-mediated formation of ZnPP; subsequently, ZnPP contributes to brain damage via HO-1 inhibition, HIF-1a upregulation, Zn toxicity enhancement and ROS generation. In Aim 1, we will determine conditions for ZnPP generation and how ZnPP contributes to brain damage following ICH. We will test the hypothesis that ICH-induced local tissue hypoxia increases Zn2+ release and Fe3+/Fe2+ ratio, promoting the Zn2+ insertion by FECH to generate ZnPP, ultimately leading to brain damage through HO- 1 inhibition and further increased Zn2+ release ROS generation. In Aim 2, we will determine why ZnPP is toxic in hypoxia and whether oxygen treatment could mitigate ZnPP-induced brain damage following ICH. We will investigate the hypothesis that ICH-induced tissue hypoxia enhances ZnPP formation by promoting oxidative stress and zinc release and exacerbates brain injury by upregulation of HIF-1a. We will also evaluate the effect of normobaric oxygen on reversing brain damage and improving outcomes. In Aim 3, we will determine whether off-target FECH inhibition by protein kinase inhibitors reduces peri-hematoma ZnPP level and alleviates brain damage in ICH. We will investigate FECH requirement for ZnPP generation and associated brain damage. Additionally, we will evaluate the efficacy of FDA approved off-target FECH inhibition drugs in mitigating ICH-induced brain damage and improving neurological outcomes. The findings form this project will clearly elucidate the intricate relationship between ZnPP, ROS, Zn2+ and Fe3+/Fe2+, providing an unprecedented mechanistic picture underlying ICH brain injury. Furthermore, the results will reveal the relationship between brain pO2 and ZnPP toxicity, demonstrating that the differential toxic vs. neuroprotective properties of ZnPP is dependent on hypoxia, resolving the current controversy on the role of ZnPP in various biological systems. Most importantly, our approach to pharmacologically inhibit ZnPP-induced exacerbation of brain damage may pave the way for the development of new therapies for ICH, either alone or in combination with other treatments.