Mechanisms of methotrexate resistance in osteosarcoma.
High-dose methotrexate is a major component of current protocols for the treatment of osteosarcoma, but some tumors seem to be resistant. Potential mechanisms of resistance include decreased transport through the reduced folate carrier (RFC) and increased expression of dihydrofolate reductase (DHFR). To investigate methotrexate resistance, tumors were obtained from 42 patients with high-grade osteosarcoma. RFC and DHFR mRNA expression were studied by semiquantitative reverse transcription-PCR. The RFC and DHFR genes were studied for deletions and amplification by Southern blot. Thirteen of 20 (65%) osteosarcoma samples were found to have decreased RFC expression at the time of initial biopsy. At definitive surgery and relapse, 10 of 22 (45%) were found to have decreased RFC expression. Seventeen of 26 (65%) samples with a poor response to chemotherapy had decreased RFC expression, whereas 5 of 14 (36%) samples with a good response had a decrease (P = 0.03). None of the samples had an RFC gene deletion. Two of 20 samples (10%) showed increased DHFR expression at initial biopsy. The frequency of increased DHFR expression was significantly higher in metastatic or recurrent tumors (62%, P = 0.014). None of the samples showed evidence of DHFR gene amplification. The high frequency of decreased RFC expression in the biopsy material suggests that impaired transport of methotrexate is a common mechanism of intrinsic resistance in osteosarcoma. Increased DHFR expression in the pulmonary metastases may be a mechanism of acquired methotrexate resistance or a difference between primary and metastatic lesions.
Guo
W
W
Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Healey
J H
JH
Meyers
P A
PA
Ladanyi
M
M
Huvos
A G
AG
Bertino
J R
JR
Gorlick
R
R
D000293:Adolescent; D000328:Adult; D000368:Aged; D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology* / Q000627:therapeutic use; D001692:Biological Transport; D015139:Blotting, Southern; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics; D002648:Child; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005260:Female; D005784:Gene Amplification; D017353:Gene Deletion; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D008297:Male; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D008727:Methotrexate / Q000494:pharmacology* / Q000627:therapeutic use; D008875:Middle Aged; D012516:Osteosarcoma / Q000175:diagnosis / Q000188:drug therapy / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D058978:Reduced Folate Carrier Protein; D013762:Tetrahydrofolate Dehydrogenase / Q000096:biosynthesis / Q000235:genetics
D000964:Antimetabolites, Antineoplastic; D002352:Carrier Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D012333:RNA, Messenger; D058978:Reduced Folate Carrier Protein; C548699:SLC19A1 protein, human; C120043:SLC19A2 protein, human; D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate