Abstract 4437: The relevance of aberrant FPGS splicing for ex vivo MTX resistance and clinical outcome in childhood acute lymphoblastic leukemia

Methotrexate (MTX) is one of the key components of current ALL treatment protocols. This antifolate inhibits de novo nucleotide biosynthesis, hindering DNA replication, hence leading to cell death. However, despite the overall good cure rates in ALL, drug resistance continues to hamper the efficacy of MTX, contributing to relapse of leukemia. Multiple molecular mechanisms underlying drug resistance have been characterized, however, their clinical relevance often remains unclear. We recently identified impaired FPGS splicing, in particular intron 8 partial retention (intron 8 PR), as a possible contributor to decreased FPGS activity in both MTX-resistant leukemia cell lines and primary ALL specimens.In the current study we explored the association between FPGS splicing alterations and MTX resistance as well as the clinical outcome of 91 pediatric ALL patients. With respect to MTX resistance, we determined the levels of MTX polyglutamates in leukemic cells, as well as FPGS activity along with the expression of several enzymes and transporters involved in (anti)folate metabolism and transport, including FPGS, FPGH, RFC, DHFR and TS. The sensitivity of leukemic cells to MTX was determined using the thymidylate synthase inhibition assay (TSIA). Moreover, the sensitivity to other chemotherapeutics as measured in the MTT assay was determined for these patient specimens. The levels of FPGS splicing alterations were semi-quantified using PCR and fragment analysis. We found that intron...
Source: Cancer Research - Category: Cancer & Oncology Authors: Tags: Experimental and Molecular Therapeutics Source Type: research