TY - JOUR
T1 - Reduced folate carrier mutations are not the mechanism underlying methotrexate resistance in childhood acute lymphoblastic leukemia
AU - Kaufman, Yotam
AU - Drori, Stavit
AU - Cole, Peter D
AU - Kamen, Barton A
AU - Sirota, Jenny
AU - Ifergan, Ilan
AU - Arush, Myriam Weyl Ben
AU - Elhasid, Ronit
AU - Sahar, Dvora
AU - Kaspers, Gert Jan L
AU - Jansen, Gerrit
AU - Matherly, Larry H
AU - Rechavi, Gideon
AU - Toren, Amos
AU - Assaraf, Yehuda G
N1 - Copyright 2003 American Cancer Society.
PY - 2004/2/15
Y1 - 2004/2/15
N2 - BACKGROUND: Although the majority of children with acute lymphoblastic leukemia (ALL) are cured with combination chemotherapy containing methotrexate (MTX), drug resistance contributes to treatment failure for a substantial fraction of patients. The primary transporter for folates and MTX is the reduced folate carrier (RFC). Impaired drug transport is a documented mechanism of MTX resistance in patients with ALL; however, to the authors' knowledge it is not known whether inactivating RFC mutations are a contributing factor.METHODS: The authors devised a genomic polymerase chain reaction-single strand conformational polymorphism assay followed by sequencing and screened the entire RFC coding region for sequence alterations in DNA from 246 leukemia specimens from patients with diverse ethnic variation, 24 at the time of recurrence and the rest at the time of diagnosis. This cohort was comprised of 203 B-precursor ALL specimens (82.5%), 32 T-lineage ALL specimens (13%), and 11 acute myeloblastic leukemia specimens (4.5%).RESULTS: Of 246 DNA samples, only 3 diagnosis B-precursor ALL specimens (1.2%) were found to harbor alterations in the RFC gene, including heterozygous single nucleotide changes resulting in D56H and D522N substitutions in the first extracellular loop and the C-terminus of this transporter, respectively. The third sample had a sequence alteration in exon 3 that could not be identified because of the lack of availability of DNA.CONCLUSIONS: Whereas inactivating RFC mutations are a frequent mechanism of MTX resistance in human leukemia cell lines and in patients with osteosarcoma, they are not common and do not appear to play any significant role in intrinsic or acquired resistance to MTX in childhood leukemia. This is the first study of RFC mutations in multiple pediatric leukemia specimens.
AB - BACKGROUND: Although the majority of children with acute lymphoblastic leukemia (ALL) are cured with combination chemotherapy containing methotrexate (MTX), drug resistance contributes to treatment failure for a substantial fraction of patients. The primary transporter for folates and MTX is the reduced folate carrier (RFC). Impaired drug transport is a documented mechanism of MTX resistance in patients with ALL; however, to the authors' knowledge it is not known whether inactivating RFC mutations are a contributing factor.METHODS: The authors devised a genomic polymerase chain reaction-single strand conformational polymorphism assay followed by sequencing and screened the entire RFC coding region for sequence alterations in DNA from 246 leukemia specimens from patients with diverse ethnic variation, 24 at the time of recurrence and the rest at the time of diagnosis. This cohort was comprised of 203 B-precursor ALL specimens (82.5%), 32 T-lineage ALL specimens (13%), and 11 acute myeloblastic leukemia specimens (4.5%).RESULTS: Of 246 DNA samples, only 3 diagnosis B-precursor ALL specimens (1.2%) were found to harbor alterations in the RFC gene, including heterozygous single nucleotide changes resulting in D56H and D522N substitutions in the first extracellular loop and the C-terminus of this transporter, respectively. The third sample had a sequence alteration in exon 3 that could not be identified because of the lack of availability of DNA.CONCLUSIONS: Whereas inactivating RFC mutations are a frequent mechanism of MTX resistance in human leukemia cell lines and in patients with osteosarcoma, they are not common and do not appear to play any significant role in intrinsic or acquired resistance to MTX in childhood leukemia. This is the first study of RFC mutations in multiple pediatric leukemia specimens.
KW - Antimetabolites, Antineoplastic/pharmacology
KW - Blotting, Northern
KW - Carrier Proteins/genetics
KW - Child
KW - DNA Mutational Analysis
KW - Drug Resistance, Neoplasm
KW - Humans
KW - Membrane Transport Proteins
KW - Methotrexate/pharmacology
KW - Osteosarcoma/drug therapy
KW - Polymerase Chain Reaction
KW - Polymorphism, Genetic
KW - Polymorphism, Single-Stranded Conformational
KW - Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy
KW - Reduced Folate Carrier Protein
KW - Tumor Cells, Cultured
U2 - 10.1002/cncr.20018
DO - 10.1002/cncr.20018
M3 - Article
C2 - 14770434
VL - 100
SP - 773
EP - 782
JO - Cancer
JF - Cancer
SN - 0008-543X
IS - 4
ER -