Targeted BRD4 protein degradation by dBET1 ameliorates acute ischemic brain injury and improves functional outcomes associated with reduced neuroinflammation and oxidative stress and preservation of blood–brain barrier integrity

Abstract

Bromodomain-containing protein 4 (BRD4), a member of the bromodomain and extra-terminal domain (BET) protein family, plays a crucial role in regulating inflammation and oxidative stress that are tightly related to stroke development and progression. Consequently, BRD4 blockade has attracted increasing interest for associated neurological diseases, including stroke. dBET1 is a novel and effective BRD4 degrader through the proteolysis-targeting chimera (PROTAC) strategy. We hypothesized that dBET1 protects against brain damage and neurological deficits in a transient focal ischemic stroke mouse model by reducing inflammation and oxidative stress and preserving the blood–brain barrier (BBB) integrity. Post-ischemic dBET1 treatment starting 4 h after stroke onset significantly ameliorated severe neurological deficits and reduced infarct volume 48 h after stroke. dBET1 markedly reduced inflammation and oxidative stress after stroke, indicated by multiple pro-inflammatory cytokines and chemokines including IL-1β, IL-6, TNF-α, CCL2, CXCL1 and CXCL10, and oxidative damage markers 4-hydroxynonenal (4-HNE) and gp91phox and antioxidative proteins SOD2 and GPx1. Meanwhile, stroke-induced BBB disruption, increased MMP-9 levels, neutrophil infiltration, and increased ICAM-1 were significantly attenuated by dBET1 treatment. Post-ischemic dBET1 administration also attenuated ischemia-induced reactive gliosis in microglia and astrocytes. Overall, these findings demonstrate that BRD4 degradation by dBET1 improves acute stroke outcomes, which is associated with reduced neuroinflammation and oxidative stress and preservation of BBB integrity. This study identifies a novel role of BET proteins in the mechanisms resulting in ischemic brain damage, which can be leveraged to develop novel therapies.

Bromodomain and Extra-Terminal Proteins in Brain Physiology and Pathology: BET-ing on Epigenetic Regulation

“…Recent studies employed models of permanent and transient cerebral ischemia to evaluate the effects of BET inhibition on the inflammatory response mediated by NF-kB [ 131 , 132 , 133 , 134 ]. In rats, transient cerebral ischemia obtained by middle cerebral artery occlusion (MCAO) resulted in a significant increase in BRD4 expression in the MCAO group compared to the control group, which was prevented by JQ1 treatment.…”

“…In fact, evaluation of oxidative stress in MCAO mice revealed an increase in the oxidative damage marker 4-hydroxy-2-nonenal (4-HNE), a concurrent buildup in the protein levels of GP91phox (NOX2) subunit of the pro-oxidant NADPH oxidase complex, and a decrease in anti-oxidant enzymes such as superoxide dismutase 2 (SOD2) and glutathione peroxidase 1 (GPX1). On the contrary, dBET1 significantly attenuated the levels of 4-HNE and NOX2 and simultaneously increased SOD2 and GPX1 expression [ 133 ]. These effects could be attributable to the ability of BET proteins to modulate Nrf2-dependent transcription of anti-oxidant genes.…”

Neuroprotective Mechanism of Icariin on Hypoxic Ischemic Brain Damage in Neonatal Mice

“…Injection. Pups were anesthetized by continuous inhalation of isoflurane (3% for induction and 1.5% for maintenance [57]). The top of the head of each pup was sterilized with alcohol, and the head skin (approximately 0.3 cm) was incised along the midline of the head with a sterile blade to expose the bregma.…”

Trilobatin attenuates cerebral ischemia/reperfusion-induced blood-brain-barrier dysfunction by targeting MMP9: The legend of a food additive

“…Brain microvascular endothelial cells (BMVECs) guarantee the tightness of BBB and own multiple unique elements such as specialized TJs proteins including claudin-5, occludin and zonula occludens-1(ZO-1) (Ng et al, 2022). Owing to the crucial structures of BBB, BMVECs are rapidly activated after ischemia, along with TJs loss, allowing the inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6 into the brain and eventually leading to BBB integrity damage (L. Liu et al, 2022). Emerging evidence demonstrates that apolipoprotein E (APOE) 4, cyclophilin A (CypA), nuclear factor kappa B (NF-κB) and matrix metalloproteinase (MMP) 9 have momentous and necessary roles in BBB disruption after ischemic stroke (Montagne, Nation, & Zlokovic, 2020;Palomino-Antolin et al, 2022).…”