IF oral administration of a USP30 inhibitor (MTX652, dose scaled to 10–30 mg/kg/day based on preclinical pharmacokinetic data described in the Evidence Set, given by oral gavage) is initiated 2 weeks after completing a 3-month course of dasatinib (5 mg/kg) plus quercetin (50 mg/kg, monthly oral gavage) in 18–24 month old male and female C57BL/6J mice, and continued for 12 weeks (weeks 14–26 of the study),

THEN the sequential D+Q → USP30i cohort will exhibit ≥40% lower p16^INK4a^+ senescent cell density in liver and gastrocnemius muscle compared to either monotherapy arm at week 26, alongside a ≥1.5-fold increase in the mt-Keima red:green ratiometric index (indicating enhanced mitophagy flux), improved maximal respiratory capacity in isolated mitochondria, and reduced tissue IL-6, IL-1β, and MMP-3 concentrations,

BECAUSE the following step-by-step causal chain operates:

1. Accumulated dysfunctional mitochondria in aged hepatocytes and myocytes are the proximate trigger for de novo senescence. Mitochondrial damage in aged cells generates elevated ROS and cytosolic mitochondrial DNA, which activates cGAS-STING and p53/p21 signaling, ultimately stabilizing p16^INK4a and initiating SASP. This source of ongoing senescence induction cannot be addressed by D+Q alone because D+Q clears already-senescent cells but does not remove the upstream mitochondrial damage in sub-threshold, non-yet-senescent cells. (Evidence Set: "The accumulation of damaged mitochondria leads to increased production of reactive oxygen species (ROS), disrupted cellular bioenergetics, and the release of mitochondrial DNA into the cytosol. These factors collectively trigger DNA damage responses and activate senescence pathways, culminating in the expression of senescence markers such as p16^INK4a and the secretion of pro-inflammatory SASP cytokines.")

2. USP30 is the primary molecular brake on PINK1/Parkin-mediated mitophagy at the outer mitochondrial membrane (OMM). USP30 constitutively removes phospho-ubiquitin chains from OMM substrates ubiquitinated by Parkin downstream of PINK1 stabilization, thereby preventing autophagy receptor (OPTN, NDP52) recruitment and maintaining a high threshold for mitophagy activation. In aged muscle and liver, where basal PINK1/Parkin pathway activity is insufficient to clear the accumulated burden of damaged organelles, USP30 renders the system effectively locked in a state of mitophagy insufficiency. (Evidence Set: "USP30 is a mitochondrial-localized DUB that directly opposes Parkin activity. By continuously stripping ubiquitin chains from OMM proteins, USP30 acts as a fundamental brake on the mitophagy process, preventing the clearance of mitochondria under basal conditions and setting a high threshold for mitophagy activation.")

3. **SASP cytokines from residual senescent cells in aged tissue actively suppress mitophagy in neighboring non-senescent cells via paracrine mTORC1 activation — the key mechanistic link that makes sequential,...

SENS category: GlycoSENS