Discussion
It has long been known that individual treatment responses and tolerance
in pediatric ALL are influenced by genetics. Certain SNPs in the TPMT
gene were previously identified as a predictor of 6-MP and 6-TG toxicity
in ALL treatment and assessment of TPMT genotype is now standard of care
in the upfront setting. The role of MTHFR SNPs in ALL pathogenesis and
treatment has been extensively studied yet remains controversial. Many
studies are small, contain a multitude of patients on different
treatment protocols, contain genetically and ethnically homogenous
populations, or have been limited by their retrospective nature. Here,
our study examines a large number of patients sourced from a genetically
heterogenous population treated with similar COG-based protocols and
suggests two MTHFR genotypes, homozygous 677TT and compound heterozygous
677CT/1298AC, are associated with increased MTX toxicity and require
decreased oral MTX dosing during ALL maintenance therapy. Based upon
this data, one could surmise that appropriate dosing of oral MTX for
patients with either homozygous 677TT or compound heterozygous
677CT/1298AC would benefit from initial dose reductions as final average
dosing was around 50% of the recommended starting dose for both groups.
With both homozygous 677TT and compound heterozygous 677CT/1298AC
genotypes having known reduced MTHFR enzymatic activity (30% and
50-60% respectively) relative to heterozygous 677CT and heterozygous
1298AC genotypes (60-70% and >80% respectively), our
findings correlate well with known enzymatic function. Surprisingly, our
results did not completely correlate with toxicity to IV methotrexate
formulations, although could be a limitation of our study given the
limited sample size of patients receiving either C-MTX or HDMTX. As far
as predicting MTHFR genotype, myelosuppression was the only clinical
factor significantly associated with detection of a clinically
significant variant. No other toxicities demonstrated statistical
significance and were not as common. We also assessed the association of
specific MTHFR genotypes and other MTX toxicities, namely methotrexate
leukoencephalopathy and thrombosis. Here we found that no MTHFR genotype
was shown to increase risk of either toxicity. It should be noted that
thrombosis risk in relation to MTHFR polymorphisms is attributed to high
homocysteine levels and previously literature suggests that these levels
may be mildly elevated at most. Transaminitis and mucositis are also
major toxicities associated with methotrexate as well as other
chemotherapeutic agents received by ALL patients. We found a large
proportion of patients regardless of MTHFR status developed mucositis at
some point during treatment although were not able to elaborate on the
severity or causal relationship with MTX versus other therapies given
limitations of information in the EHR.
There is also evidence that MTHFR status can influence outcomes, namely
the likelihood of relapse or eventual death from ALL disease. Within our
cohort of patients tested for MTHFR SNP, only one had refractory disease
(1 patient with heterozygous 677CT SNP) and ultimately achieved
remission after CAR T cell therapy and bone marrow transplant. Compared
to most recent statistics showing greater 90% survival among ALL
patients, this certainly does not demonstrate worse outcome for patients
with clinically meaningful MTHFR genotypes, although would not speculate
with regards to a clinical benefit given the small sample size, large
variation of time since diagnosis and treatment, as well as significant
population of patients within cohort that never received oral or IV
methotrexate or had MTHFR genetic testing evaluated given early
relapsed/refractory disease.
Being retrospective in nature, there are significant limitations to our
study, mainly that information extracted is reliant on accurate medical
documentation which is sometimes incomplete. We also combined patients
whose MTHFR genotyping was not tested with those who tested wildtype,
which was an assumption that may not have been true. In addition, it is
likely certain biases played a role in the decisions to adjust oral MTX
dosing. By knowing MTHFR status, providers may have preferentially
decreased oral MTX dosing in response to toxicity whether it was a
clinically significant genotype or not. Given the frequency of MTHFR
SNPs in the population combined with targeted testing and the fact this
was a single institutional cohort, our sample sizes were limited,
especially among different genotype groups. It is very likely additional
patients with MTHFR variants, both clinically significant and
insignificant, exist within the control population and were never
tested. Although this would be expected to further reinforce the
significance of our results, it does limit our ability to extrapolate
information such as targeted oral MTX dosing and it also may limit some
of our other comparisons that did not achieve statistical significance.
Nonetheless, our study was able to demonstrate significantly decreased
tolerance to oral MTX among patients with homozygous 677TT and compound
heterozygous 677CT/1298AC genotypes as demonstrated by level of
myelosuppression. These results do warrant further investigations.
Ideally, this would be done through a prospective study. However, a more
practical approach could involve a multi-institutional effort where
genetic data can be harvested through existing biobanks and MTX dose and
toxicity could retrospectively be reviewed for each sample. As was
demonstrated with TPMT related to 6-MP toxicity, MTHFR may also
influence MTX toxicity and therefore, identification of MTHFR genotypes
upfront may lead to reduced ALL treatment complications in the future.
Conflicts of Interest: The authors declare no conflict of interest.
Acknowledgments: CH is supported by the National Institutes of Health
(T32 CA060441) and Hartwell Foundation.