MYCN amplification confers enhanced folate dependence and methotrexate sensitivity in neuroblastoma.

Diana T Lau,Giovanni Perini,Chengyuan Xue,Benedikt Brors,Samuele Gherardi,Jamie I Fletcher,Matthias Fischer,Dilafruz Juraeva,Glenn M Marshall,Claudia L Flemming,Michelle Haber,Lesley J Ashton,Murray D Norris,André Oberthuer

doi:10.18632/oncotarget.3732

Abstract

MYCN amplification occurs in 20% of neuroblastomas and is strongly related to poor clinical outcome. We have identified folate-mediated one-carbon metabolism as highly upregulated in neuroblastoma tumors with MYCN amplification and have validated this finding experimentally by showing that MYCN amplified neuroblastoma cell lines have a higher requirement for folate and are significantly more sensitive to the antifolate methotrexate than cell lines without MYCN amplification. We have demonstrated that methotrexate uptake in neuroblastoma cells is mediated principally by the reduced folate carrier (RFC; SLC19A1), that SLC19A1 and MYCN expression are highly correlated in both patient tumors and cell lines, and that SLC19A1 is a direct transcriptional target of N-Myc. Finally, we assessed the relationship between SLC19A1 expression and patient survival in two independent primary tumor cohorts and found that SLC19A1 expression was associated with increased risk of relapse or death, and that SLC19A1 expression retained prognostic significance independent of age, disease stage and MYCN amplification. This study adds upregulation of folate-mediated one-carbon metabolism to the known consequences of MYCN amplification, and suggests that this pathway might be targeted in poor outcome tumors with MYCN amplification and high SLC19A1 expression.

Highlights

Folate is essential for DNA synthesis and cell growth in rapidly dividing cells
To test whether upregulation of the folate-mediated one-carbon metabolism pathway corresponds to an increased reliance on folate in MYCN-amplified cells, we assessed the colony forming ability of two MYCNamplified cell lines (CHP-134 and Kelly) and two nonamplified lines (SH-EP and NB69) in folate-free medium supplemented with increasing concentrations of folinic acid
Activation of the Myc oncogene in particular reprograms metabolic pathways, providing increased biosynthetic and bioenergetic capacity [25] through upregulation of protein synthesis [26], glycolysis [27], glutaminolysis [28], nucleotide synthesis [29] and polyamine synthesis [21], all of which contribute to rapid proliferation

Summary

Introduction

Folate is essential for DNA synthesis and cell growth in rapidly dividing cells. In cancer, folate uptake and metabolism are frequently up-regulated to meet the elevated need for nucleotides. Inhibition of folate metabolism is the basis for several widely used cancer drugs, including methotrexate [1,2,3], which exerts its toxicity by inhibiting dihydrofolate reductase (DHFR) thereby disrupting purine and thymidylate biosynthesis, inhibiting DNA replication and promoting cell death. This folate antagonist has been used as a chemotherapeutic agent to treat a variety of hematological malignancies and solid tumors including acute lymphoblastic leukemia, www.impactjournals.com/oncotarget non-Hodgkin’s lymphoma, osteosarcoma, head and neck cancer, choriocarcinoma, small cell lung cancer, and breast cancer [4]. Little is known about the relationship between folate metabolism and neuroblastoma biology, folate-mediated one-carbon metabolism has previously been identified as correlating with neuroblastoma aggressiveness in bioinformatics analyses [13], suggesting that inhibition of folate metabolism may be beneficial in high-risk disease

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Conclusion