Impact of aging on the pharmacokinetic profile of everolimus in male mice

Age-related changes in drug-metabolizing enzymes and transporters can alter pharmacokinetics, yet data on everolimus, a widely used mTOR inhibitor, remain limited. This study investigated the pharmacokinetic profile of everolimus in male C57BL/6J mice at three life stages—young (≈ 9 weeks), adult (≈ 23 weeks), and old (≈ 56 weeks)—and examined whether age-related changes in hepatic and intestinal Cyp3a11 and Abcb1a/b (P-gp; P-glycoprotein) expression parallel these alterations. Mice received a single oral dose of everolimus (5 mg/kg, ZT1), and plasma and hepatic concentrations were quantified over 24 h by validated HPLC-UV, with non-compartmental analysis performed (n = 5 per age group per time point). In drug-naïve, age-matched cohorts, gene and protein expression were measured by qPCR and western blot (n = 5 per age group). The area under the concentration–time curve (AUCtotal) was 129% greater in young mice and 86% more in mature mice compared to old mice (young vs. old, p < 0.05). Peak plasma concentration (Cmax) also decreased with age (young + 95%, adult + 72% compared to elderly; p < 0.01 and p < 0.05). Oral clearance (CL/F) increased by 68% in adults and 230% in older individuals compared to young animals, whereas the apparent volume of distribution (Vd/F) climbed by 47% and 187%, respectively. In liver, Abcb1a remained unchanged across age groups, whereas Abcb1b transcripts rose ~ 1.5-fold in old vs. young mice (p < 0.001). Hepatic Cyp3a11 declined stepwise, falling by ~ 55% in old animals (p < 0.01). At the protein level, total hepatic P-gp dropped in adult and old mice (− 45% vs. young, p < 0.05) and Cyp3a protein showed a non-significant downward trend. In ileum, Abcb1a increased sharply with age (adult ≈ 5-fold, old ≈ 7-fold vs. young), translating to a ~ 5-fold rise in P-gp protein in adult mice (p < 0.01); Abcb1b transcripts were unchanged. Ileal Cyp3a11 mRNA peaked in adults (≈ 3-fold vs. young, p < 0.05), although Cyp3a protein differences did not reach significance. These findings suggest that reduced everolimus bioavailability in older mice is primarily driven by increased intestinal P-gp–mediated efflux rather than hepatic metabolism, supporting the need for age-adjusted dosing to optimize therapeutic outcomes across the lifespan.