Introduction
Pediatric acute lymphoblastic leukemia and lymphoma (ALL) is the most common childhood malignancy, accounting for approximately 25% of all pediatric cancer diagnosis. Chemotherapy inhibiting folate metabolism, particularly methotrexate (MTX), has remained a core component of ALL therapy since the first published remissions as a monotherapy in 1948. Currently within Children’s Oncology Group (COG)-based protocols, MTX is administered in by multiple modes of delivery including intravenous, intrathecal, and oral administration during all phases of ALL treatment. One of the major contributions of MTX is to combat leukemia in sanctuary sites, namely the CNS and testicles, thereby preventing local relapse.
The antileukemic activity of MTX is attributed to its ability to disrupt folate metabolism. MTX enters the cell through the reduced folate carrier and then polyglutamated which helps retain the drug intracellularly. It is also believed that the polyglutamated form is more bioactive, inhibiting MTX’s main targets, Dihydrofolate Reductase and Thymidylate Synthase, which are important for DNA synthesis. Through the disruption of folate metabolism, there is also reduction in the conversion of homocysteine to methionine resulting in decreased protein synthesis and methylation. Methylenetetrahydrofolate reductase (MTHFR) is an enzyme that also exists in this pathway and serves as the irreversible, rate limiting step of the conversion of 5,10-methylentetrahydrofolate (5,10-CH2-THF) to 5-methyltetrahydrofolate (5-CH-THF). Two important single nucleotide polymorphisms (SNPs), C677T and A1298C, are frequently identified in the general population and thought to be clinically significant based upon relative enzymatic function. The C677T SNP is thought to reduce MTHFR activity to 60-70% and 30% in heterozygous and homozygous individuals respectively. This SNP has also been associated with elevated homocysteine, which may lead to a prothrombotic state. The A1298C SNP is more active with 60 and 80% in its homozygous and heterozygous form, respectively. However, for those with one copy of the A1298C SNP, activity is estimated to be slightly decreased at 50-60% when coexisting with a single C677T SNP, otherwise known as compound heterozygosity.
Patients treated with MTX can experience a variety of multisystem toxicities including, but not limited to, myelosuppression, transaminitis, subacute leukoencephalopathy, mucositis, and renal toxicity. The influence of MTHFR SNPs on toxicity has been examined previously and results have remained controversial. It should be noted that most studies to date have either focused on high dose MTX (HDMTX) with typical doses of 5 g/m2/dose given intravenously, examined ethnically homogenous populations, or have not differentiated between differing modalities of MTX treatment. In a retrospective analysis of CCG-1891 after analyzing for common polymorphic genes involved in drug metabolism, the MTHFR C677T genotype was determined to have no significant effect on cumulative oral MTX received during maintenance, although a 25-30% reduction of cumulative dose was observed across the population. This method of analysis, however, can be misleading as it fails to account for several confounders, including medication holds, average dose received, and compliance.
The aim of our study was to directly examine the influence of MTHFR SNPs on methotrexate tolerance at various phases of treatment, with particular attention given to oral methotrexate which is given during the maintenance phase of treatment, the last and longest phase of treatment on COG-based ALL treatment protocols. Dosing of MTX in maintenance along with oral 6-mercaptopurine (6-MP), is regulated by both degree of myelosuppression (goal absolute neutrophil count 500-1500/ul and Platelet count > 50,000 cell/ul) and to a lesser extent liver toxicity. There is extreme variability between patients likely from a multitude of factors including individual pharmacogenomics. As part of the folate metabolic pathway, MTHFR has the potential to influence MTX efficacy, as well as toxicity, and those with certain MTHFR genotypes may be at increased risk for toxicity.
Our overarching goal was to utilize our local ALL patient population to identify clinically significant genotypes that would identify individuals who need reduced MTX dose adjustment upfront and therefore provide guidance to providers taking care of these patients. Furthermore, we also examined the influence of MTHFR genotypes on other phases of treatment as well as risk for developing additional toxicities such as subacute leukoencephalopathy and thrombosis.