Pemetrexed (PMX, Alimta®) is a novel multi-targeted antifolate approved for the treatment of malignant pleural mesothelioma and non-small cell lung cancer (NSCLC). Given the high incidence of brain metastasis in NSCLC patients and wide use of the classic antifolate, methotrexate (MTX), in primary CNS lymphoma, the clinical use of PMX may eventually include the treatment of CNS tumors. However, previous studies in our laboratory indicated that, like MTX, PMX has difficulty in penetrating across the blood-brain barrier (BBB). Factors limiting the CNS distribution of PMX remain unidentified. One important determinant of CNS distribution is efflux transport by BBB transporters. The overall objective of this study was to characterize the brain-to-blood efflux transport of PMX and to examine role of BBB efflux transporters such as BCRP, MRP2 and other organic anion transporters in brain distribution of PMX. The interaction of PMX with BCRP was examined in vitro and in vivo. In vitro results revealed that PMX is a substrate for BCRP-mediated transport. In vivo examination indicated that deletion of Bcrpl has little influence on brain penetration of PMX. Using the brain efflux index method, the mechanism responsible for the brain efflux of PMX and MTX was investigated. The results revealed that brain elimination half-life of PMX and MTX were 48 and 32 minutes, respectively and both PMX and MTX undergo saturable efflux transport across the BBB. MRP2 does not play a role in the brain distribution of either antifolate. However, BCRP makes a significant contribution to brain elimination of MTX, but not PMX. In addition, it was observed that brain-to-blood transport of PMX and MTX was markedly inhibited by probenecid and benzylpenicillin, suggesting the involvement of organic anion transporters, possibly OAT3. Given the knowledge that solid tumors and some physiological barriers have an acidic extracellular environment, the effect of pH on transport activity of BCRP for PMX was examined. In addition, the molecular basis of the observed pH-dependency in BCRP transport activity was explored based on the recent homology model of BCRP. This study has important implications in the handling of PMX and other chemotherapy drug molecules in the acidic environment of tumors and in the distribution and elimination of the drug molecules.
Studies presented in this dissertation provide useful information about the specific mechanisms involved in limiting the brain penetration of PMX and MTX. This new knowledge will help in formulating strategies to improve CNS delivery of these antifolates and maylead to more successful treatment of primary and secondary CNS tumors.