AAV-based temporal APOE4-to-APOE2 replacement reveals rebound adaptation and RAB24-mediated Aβ and cholesterol dysregulation

APOE-targeted gene therapy offers a promising strategy for modifying Alzheimers disease (AD) risk, yet the temporal dynamics and context-dependent effects of APOE isoform modulation remain poorly defined. Here, we developed a rapid AAV-based platform enabling inducible in vivo replacement of APOE4 with APOE2. In 5xFAD mice, sustained APOE4 expression exacerbated cognitive decline, A{beta} deposition (parenchymal and vascular), and glial activation, whereas long-term APOE2 expression--with concurrent APOE4 silencing--significantly reversed these pathological features and rescued cognitive function. In contrast, short-term APOE2 replacement conferred no benefit and unexpectedly worsened behavioral and pathological outcomes. Transcriptomic profiling revealed that APOE4-associated gene signatures were broadly reversed by long-term APOE2 expression, but paradoxically aggravated by short-term replacement. Among these, RAB24--a regulator of autophagic trafficking--was upregulated by APOE4 and short-term APOE2 but suppressed by long-term APOE2. RAB24 elevation impaired A{beta} clearance and cholesterol homeostasis via lysosomal retention in primary astrocytes and neurons. Together, these findings uncover a rebound-adaptation mechanism that shapes APOE2 therapeutic outcomes, identify RAB24 as a modifiable node in A{beta} and cholesterol metabolism, and establish a temporally controlled gene therapy platform to inform the design of future APOE-targeted interventions in AD.