Hypothesis

Liver‑targeted maintenance of mTORC1 activity combined with muscle‑ and brain‑specific mTORC1 inhibition will extend mammalian lifespan more effectively than global mTOR suppression, by preserving hepatic anabolic output while enhancing autophagy and proteostasis in tissues where damage accumulates.

Mechanistic Rationale

mTORC1 functions as a metabolic rheostat that balances biosynthesis with cellular cleanup [1]. Systemic inhibition with rapamycin extends lifespan but imposes trade‑offs such as glucose intolerance and reduced muscle mass because it forces all tissues into a survival mode [3]. Human genetics shows that natural long‑lived variants attenuate rather than abolish mTOR signaling, suggesting that optimal aging involves tissue‑specific tuning [2][4]. The liver requires mTORC1 for IGF‑1 production, albumin synthesis, and lipid homeostasis [1]; suppressing it compromises these functions and can aggravate metabolic disease. In contrast, skeletal muscle, cardiac tissue, and the brain benefit from lowered mTORC1 activity through increased autophagy, reduced SASP, and enhanced Klotho secretion [1][6]. Recent AAV engineering enables precise spatial control: liver‑detargeted capsids achieve >1000‑fold CNS enrichment versus AAV9 [5] and liver‑detargeted myotropic vectors deliver uniform skeletal and cardiac transduction at low doses [6]. By pairing an AAV that overexpresses Rheb (a potent mTORC1 activator) under a liver‑specific promoter with an AAV that expresses a dominant‑negative Raptor under a muscle‑ or brain‑specific promoter, we can create a push‑pull system that keeps the liver in a growth‑permissive state while shifting peripheral tissues toward maintenance.

Experimental Design

1. Vector construction – AAV8‑TBG‑Rheb (liver‑specific) and AAV‑PHP.eB‑CKM‑DN‑Raptor (muscle) or AAV‑PHP.eB‑Syn‑DN‑Raptor (brain).
2. Animal cohorts (n=30 per group, both sexes):
   • Global mTORC1 inhibition (systemic rapamycin)
   • Liver‑only mTORC1 activation (AAV‑TBG‑Rheb)
   • Muscle‑only mTORC1 inhibition (AAV‑CKM‑DN‑Raptor)
   • Brain‑only mTORC1 inhibition (AAV‑Syn‑DN‑Raptor)
   • Combined liver activation + muscle inhibition
   • Combined liver activation + brain inhibition
   • Triple combination (liver activation + muscle + brain inhibition)
   • Empty vector controls.
3. Readouts (longitudinal): survival, frailty index, glucose tolerance, insulin sensitivity, body composition (MRI), hepatic IGF‑1 and serum albumin levels, muscle fiber cross‑sectional area, cardiac ejection fraction, hippocampal neurogenesis, markers of autophagy (LC3‑II/p62) and SASP (IL‑6, TNF‑α) in each tissue.
4. Mechanistic probes – hyperinsulinemic‑euglycemic clamps, polysome profiling for protein synthesis flux, and secretory proteomics to assess Klotho and FGF21 levels.

Predicted Outcomes

• Groups with liver‑specific mTORC1 activation will maintain or increase circulating IGF‑1 and albumin relative to rapamycin‑treated mice, preserving hepatic metabolic function.
• Muscle‑ and brain‑specific mTORC1 inhibition will show elevated autophagy flux, reduced p62 accumulation, and lower SASP compared with controls.
• The combined liver‑activation + peripheral‑inhibition groups are expected to exhibit the longest median lifespan, improved glucose homeostasis, and better preservation of lean mass and cognitive performance than global rapamycin treatment.
• If hepatic mTORC1 activation exacerbates tumorigenesis or insulin resistance despite peripheral benefits, the hypothesis would be refuted.

Falsifiability

A clear falsifiable outcome is that the triple‑combination (liver activation + muscle + brain inhibition) does not surpass the lifespan of the global rapamycin group, or that hepatic IGF‑1 elevation correlates with increased tumor burden or worsened glucose tolerance without compensatory longevity benefits. Conversely, a significant lifespan extension coupled with preserved hepatic anabolic markers and enhanced peripheral autophagy would support the hypothesis.