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Project Article PNAS (IF = 9.1) | Nano‑Immunomodulation of the Aβ‑Activated Feedback Mechanism in Microglia for Alzheimer’s Disease Therapy

　　Alzheimer’s disease (AD) is a common neurodegenerative disorder characterized by an insidious onset, slow progression, and the presence of behavioral and cognitive impairments, making it one of the most challenging conditions to treat worldwide. Amyloid‑β plaque deposition and neuroinflammation in the brain are key pathological hallmarks of AD; however, the molecular‑biological links between these two processes and the mechanisms by which they jointly influence the disease’s pathogenesis remain poorly understood.

　　A research team led by Professor Li Suxin at China Pharmaceutical University has discovered that, under pathological conditions, excess Aβ accumulated in the brain activates the cGAS–STING pathway within microglia via mitochondrial damage, triggering a cascade of inflammatory responses. In turn, neuroinflammation further disrupts the balance between Aβ uptake and clearance in the brain, exacerbating Aβ deposition and establishing a vicious pathological cycle. The findings were published in PNAS (Proceedings of the National Academy of Sciences of the United States of America) in an article titled “Nano‑immunomodulation of the Aβ–STING feedback machinery in microglia for Alzheimer’s disease treatment.”

　　

 

　　This study elucidates the molecular feedback mechanism by which Aβ induces mitochondrial damage, triggers mtDNA leakage, activates the cGAS/STING axis to drive neuroinflammation, and disrupts the balance between Aβ uptake and clearance. Based on these findings, we propose an Aβ–STING synergistic immunosilencing therapy, termed ASSIST (Aβ–STING Synergistic ImmunoSilencing Therapy). This is a nano‑micellar formulation capable of crossing the blood–brain barrier; it competitively disaggregates Aβ plaques and prevents their re‑aggregation, thereby enabling Aβ in its oligomeric form to be selectively internalized by microglia rather than neurons. Once inside microglia, the formulation releases a STING inhibitor, blocking inflammatory signaling, restoring Aβ clearance, and reducing neuronal damage, thus collectively ameliorating cognitive deficits.

　　Research findings

　　| A multivalent nanomicelle, G‑NP, was prepared, which can effectively disaggregate Aβ aggregates, inhibit monomer aggregation, and promote the uptake and clearance of Aβ by microglial cells.

　　

 

　　Using a variety of experimental approaches—including TEM, ThT fluorescence, FRET, and CD spectroscopy—our studies have demonstrated that G‑NPs not only efficiently disaggregate Aβ aggregates but also inhibit the further aggregation of Aβ monomers, thereby reducing Aβ plaque formation at its source. Specifically, the GKLVFF peptide segment within G‑NPs can specifically recognize and bind to Aβ aggregates, disrupting their β‑sheet structure and converting them into more readily cleared monomeric forms. At the same time, G‑NPs can block the aberrant aggregation of Aβ monomers, establishing a dual regulatory mechanism. Confocal microscopy and flow cytometry assays in both single‑cell and multicellular systems confirmed that, following disaggregation of Aβ aggregates, G‑NPs significantly enhance microglial uptake of Aβ. The internalized Aβ exhibits strong colocalization with lysosomes and is almost completely degraded within 24 hours, indicating that G‑NPs hold substantial promise for promoting Aβ clearance and degradation.

　　| G-NP can cross the BBB and exert therapeutic effects in the brain, but its efficacy is limited.

　　

 

　　In vitro Transwell assays and in vivo fluorescence imaging demonstrate that G‑NPs exhibit favorable brain penetration. Intravenous administration of G‑NPs to AD mice revealed therapeutic efficacy, though with certain limitations: G‑NPs did not significantly ameliorate cerebral hemorrhage in the brains of these mice, likely because they failed to effectively modulate the inflammatory milieu in the brain, thereby restricting their overall therapeutic impact.

　　| Aβ activates the cGAS-STING pathway, triggering an inflammatory response.

　　

 

　　Bioinformatics analysis of transcriptomic data from Alzheimer’s disease (AD) patients revealed that key downstream pathways of the STING signaling pathway are significantly upregulated in brain tissue, suggesting that this pathway may contribute to the pathogenesis of AD. To validate this finding, we treated BV2 microglial cells with Aβ; the results demonstrated that high concentrations of Aβ robustly activate the STING signaling pathway. Further mechanistic studies showed that Aβ induces mitochondrial damage, leading to the release of mitochondrial DNA into the cytoplasm, where it binds to cGAS and triggers a cascade of STING‑mediated signaling events. In the hippocampus of 5×FAD mice, phosphorylation levels of p‑STING, p‑IRF3, and p‑TBK1 were markedly elevated, confirming aberrant activation of the STING pathway in an AD animal model.

　　Furthermore, in the cerebral cortex and hippocampus of Aβ‑induced AD mouse models, the expression of the downstream effector molecules IFN‑β, as well as the chemokines CXCL9 and CXCL10, was significantly upregulated, thereby providing additional evidence for the widespread activation of the STING pathway in Alzheimer’s disease. This study not only highlights the critical role of the STING pathway in AD pathogenesis but also elucidates the molecular mechanism by which Aβ drives neuroinflammation via a mitochondrial damage–mtDNA release–cGAS/STING signaling axis, offering new theoretical insights and potential therapeutic targets for precision medicine in Alzheimer’s disease.

　　| Preparation of ASSIST and its in vitro anti-inflammatory activity

　　

 

　　To enhance the therapeutic efficacy of G‑NPs, we incorporated the STING inhibitor H151 as a combination therapy agent. By encapsulating H151 within G‑NPs, we successfully developed a nanomedicinal formulation termed Aβ‑STING Synergistic ImmunoSilencing Therapy (ASSIST). ASSIST not only preserves the G‑NP–mediated abilities to disaggregate Aβ aggregates, inhibit Aβ monomer aggregation, and promote microglial phagocytosis of Aβ, but also enables the controlled release of H151, thereby effectively suppressing STING pathway activation triggered by excessive Aβ uptake and further mitigating the inflammatory brain microenvironment.

　　| ASSIST inhibits the STING pathway in the brains of AD mice, alleviating inflammation.

　　

 

　　To investigate the therapeutic efficacy of ASSIST in Alzheimer’s disease (AD) mice, this study administered ASSIST intravenously to Aβ‑induced AD mice and 5×FAD mice. Using qPCR and ELISA, we found that ASSIST significantly suppressed the activation of the STING pathway in the brain and reduced the secretion of pro‑inflammatory cytokines, thereby ameliorating the inflammatory microenvironment. Nissl staining revealed that ASSIST treatment markedly attenuated neuronal damage in the hippocampus and cortical regions of AD mice, further confirming its neuroprotective effects.

　　| ASSIST improves cognitive impairment in AD mice

　　

 

　　This study systematically evaluated the effects of ASSIST on cognitive function and brain pathology in 5xFAD model mice. Behavioral assays, including the Morris water maze, the Y-maze, and the nest-building test, demonstrated that ASSIST significantly improved long-term memory, short-term memory, and overall cognitive performance, while enhancing the animals’ everyday behavioral capabilities. Furthermore, ASSIST markedly reduced Aβ plaque deposition in the brain, attenuated pathological hallmarks associated with cerebral amyloid angiopathy (CAA), and ameliorated symptoms of cerebral hemorrhage.

　　Summary

　　In summary, excessive Aβ impairs microglia, leading to the leakage of mitochondrial DNA (mtDNA) into the cytoplasm. mtDNA then activates the cGAS–STING signaling cascade, prompting microglia to secrete large amounts of pro-inflammatory cytokines. This severe inflammatory milieu not only exacerbates neuroinflammation but also hinders microglial clearance of Aβ, thereby establishing a negative feedback loop. Based on this mechanism, our study has designed the ASSIST nanomedicine.

　　The GKLVFF moiety carried on the ASSIST surface can effectively destabilize Aβ plaques and inhibit monomer aggregation, thereby promoting the phagocytosis of Aβ by microglia rather than neurons and enhancing Aβ clearance. Moreover, ASSIST blocks the STING signaling pathway triggered by excessive Aβ uptake by microglia, attenuating brain inflammation, restoring the microglial capacity to clear Aβ, and reducing multiple forms of neuronal damage. ASSIST ameliorates cognitive deficits in AD‑model mice and improves their memory performance, offering a novel therapeutic target and a promising nanomedicinal strategy for the precision treatment of Alzheimer’s disease.

　　The 5-month-old 5×FAD transgenic mice and the 2-month-old wild-type C57BL/6 mice used in this study were both provided by Aniphe BioLab.

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