IF a dual-vector allotopic rescue system — comprising (i) AAV2 (intravitreal, ~1×10¹⁰ vg/eye) delivering a codon-recoded, mitochondrial targeting sequence (MTS)-fused MT-ND4 allotope modeled on the lenadogene nolparvovec/GS010 construct, and (ii) AAV9 (systemic intravenous, ~2×10¹³ vg/kg) delivering the identical nuclear-encoded ND4 cassette — is administered to aged (24-month-old), male and female C57BL/6J mice harboring a mosaic burden of somatic ND4/Complex I mtDNA mutations,

THEN a ≥30% rescue of mitochondrial Complex I–linked oxygen consumption rate (OCR) will be observed in both retinal ganglion cells (RGCs) and skeletal muscle fibers within 16 weeks post-injection, accompanied by ≥20% preservation of surviving RGC density (OCT/RBPMS immunostaining), ≥15% improvement in rotarod endurance, and a measurable reduction in the fraction of cells exhibiting Complete I enzyme histochemistry (COX/SDH) dropout to levels approaching those of 6-month-old reference animals,

BECAUSE the following causal chain connects intervention to outcome:

1. Aged post-mitotic cells accumulate somatic ND4 point mutations and large-scale mtDNA deletions through clonal expansion, creating heteroplasmic mosaic dysfunction within individual RGCs and skeletal muscle fibers where Complex I assembly and electron flux are compromised once heteroplasmy crosses the tissue-specific biochemical threshold for OXPHOS failure. (Evidence Set: Literature Task Output — "mtDNA is indispensable for the generation of cellular ATP… close physical proximity to the electron transport chain — the primary intracellular source of reactive oxygen species")

2. The allotopic expression strategy (nuclear-encoded, MTS-targeted ND4) demonstrated by Yu-Wai-Man and GenSight Biologics in the LHON/GS010 clinical program has established proof-of-concept that a recoded hydrophobic ND4 subunit can be imported into the mitochondrial inner membrane with sufficient efficiency to rescue Complex I function in human RGCs carrying the pathogenic G11778A ND4 mutation, providing validated construct architecture (MTS selection, codon optimization for nuclear codon usage, AAV2 tropism for RGCs via intravitreal delivery). (Evidence Set: Research Context — "proof-of-concept established by Yu-Wai-Man for Leber hereditary optic neuropathy"; Literature Task Output — "lenadogene nolparvovec / GS010… MTS, codon optimization, AAV2 vector")

3. Critically, the GS010 allotope is mutation-agnostic at the functional level: the nuclear-encoded wild-type ND4 protein, once imported, contributes to Complex I assembly independent of which specific somatic mutation has corrupted the mtDNA-encoded copy. This distinguishes age-directed allotopic repair from LHON therapy — rather than compensating for one germline variant, the same construct provides a permanent nuclear backup that rescues ANY ND4-disrupting somatic event in a given cell, regardless of heteroplasmic mutation identity. [SPECULATIVE — direct evidence for mut...

SENS category: GlycoSENS