Plasma and CNS pharmacokinetics of O⁴-benzylfolic acid (O ⁴BF) and metabolite in a non-human primate model

Purpose: O⁶-alkylguanine-DNA alkyltransferase (AGT) repairs DNA damage from alkylating agents by transferring the alkyl adducts from the O ⁶-position of guanine in DNA to AGT. The folate analog O ⁴-benzylfolic acid (O⁴BF) is an inhibitor of AGT with reported selectivity of the alpha-folate receptor in tumors. We studied plasma and cerebrospinal fluid (CSF) pharmacokinetics and CSF penetration of O ⁴BF in a non-human primate model. Methods: Rhesus monkeys (Macaca mulatta) received O⁴BF (10-50 mg/kg) intravenously, and serial blood and CSF samples were obtained. Analyte concentrations in plasma were measured by HPLC/photo diode array, and an HPLC/MS/MS assay was used for CSF samples. Results: A putative metabolite of O⁴BF was detected in plasma and CSF. O⁴BF and the metabolite inactivated purified AGT with ED ₅₀ of 0.04 mcM. The median clearance of O⁴BF was 8 ml/min/kg and half-life was 1.1 h. The metabolite had a substantially longer half-life (>20 h) and greater AUC than O⁴BF. The AUC of the metabolite increased disproportionately to the dose of O⁴BF, suggesting saturable elimination. CSF penetration of O⁴BF and its metabolite was < 1%. At the 50 mg/kg dose level, the C_max in CSF for O⁴BF was less than 0.09 mcM and for the metabolite the C _max ranged from 0.02 to 0.04 mcM (O⁴BF equivalents). Conclusions: Concentrations of O⁴BF and the metabolite in CSF exceeded the ED₅₀ of AGT; however, recently reported lack of receptor specificity and pharmacokinetic data suggesting saturable elimination of both O⁴BF and its metabolite may limit dose-escalation and future clinical development of this agent.