IF lysosome-targeted recombinant DNase II nanoparticles (lyso-DNase II; formulated as mannose-6-phosphate receptor-targeted lipid nanoparticles encapsulating recombinant human DNase II optimized for activity at pH 4.5–5.0, administered intravenously at 3 mg/kg twice weekly for 8 weeks) are administered to aged female C57BL/6J mice (22–24 months),

THEN a significant reduction (≥40%) in cytoplasmic mitochondrial DNA (mtDNA) burden within Lin⁻Sca1⁺cKit⁺ (LSK) HSCs, suppression of cGAS-STING-NFκB signaling, restoration of HSC long-term repopulation capacity to ≥60% of young controls in competitive transplantation assays, and reduction of myeloid-biased output will be observed,

BECAUSE the following causal chain operates:

1. Aged HSCs exhibit lysosomal hyperactivation that is paradoxically dysfunctional: despite elevated degradative enzyme activity, lysosomal membrane permeabilization and impaired substrate processing result in failure to degrade mitochondrial DNA delivered via mitophagy, causing cytoplasmic mtDNA accumulation (Mount Sinai group work: https://scholars.mssm.edu/en/publications/reversing-lysosomal-dysfunction-restores-youthful-state-in-aged-h/).

2. Aged cells accumulate extranuclear DNA that engages the autophagy-lysosome system for clearance; when this clearance fails, cytoplasmic DNA persists and activates innate immune sensing (extranuclear DNA accumulation drives senescence and inflammation via autophagy-lysosome processing failure)[https://doi.org/10.1111/acel.12901].

3. Leaked cytoplasmic mtDNA activates cGAS, producing cGAMP, which engages STING, triggering IRF3 and NFκB transcription factor cascades. NFκB activation within HSCs is directly associated with increased quiescence, impaired symmetric self-renewal, and durable long-term reconstitution failure following transplantation (NFκB cell-intrinsic activation drives HSC quiescence and long-term reconstitution impairment)[https://doi.org/10.1101/2025.01.14.632900].

4. DNase II is the principal lysosomal endonuclease responsible for degrading DNA delivered to lysosomes via autophagy; it degrades single-stranded and double-stranded DNA at acidic pH, and its deficiency is the canonical cause of sterile type I interferon inflammation driven by undigested nuclear/mitochondrial DNA. Lysosome-targeted supplementation with exogenous DNase II via mannose-6-phosphate receptor-mediated delivery directly repairs the deficit in mtDNA degradative capacity—removing already-accumulated cytoplasmic mtDNA rather than preventing new mitochondrial damage [SPECULATIVE: M6P-targeted nanoparticle delivery efficiency to HSC lysosomes in vivo has not been directly demonstrated; tropism will require empirical validation].

5. By enzymatically degrading the cytoplasmic and intra-lysosomal mtDNA pool, lyso-DNase II removes the cGAS-STING ligand, collapsing downstream NFκB signaling and resolving the chronic inflammatory state within aged HSCs, analogous to the mechanism by which DNASE2 knockout mice ...

SENS category: LysoSENS

Key references: • doi.org/10.1111/acel.12901]. • doi.org/10.1101/2025.01.14.632900]. • doi.org/10.1371/journal.pbio.2003389]. • doi.org/10.1126/science.aag3048]. • doi.org/10.1101/2025.04.11.648417].