Author ORCID Identifier

Defense Date


Document Type


Degree Name

Doctor of Philosophy


Oral Health Research

First Advisor

Anthony Faber


High risk neuroblastoma (NB) is responsible for nearly 15% of all cancer related deaths in the pediatric population. The transcription factor MYCN is the most studied oncogene in NB, automatically conferring high-risk and has been found to be difficult to target. One way that MYCN amplification in NB functions to support tumorigenicity is through altered metabolism, satisfying the demand for rapid proliferation by increasing the nutrient flux through biosynthetic pathways. We performed an unbiased screen on select metabolic targeted therapy combinations and correlated sensitivity with over 20 subsets of cancer. We found that MYCN-amplified NB is hypersensitive to the combination of an inhibitor of the lactate transporter monocarboxylate transporter 1 (MCT1), AZD3965, and complex I of the mitochondrion, phenformin. Our data demonstrate that monocarboxylate transporter 4 (MCT4) is highly correlated with resistance to the combination in the screen and lowly expressed in MYCN-amplified NB. Low MCT4 combines with high expression of the monocarboxylate transporter 2 (MCT2) and MCT1 chaperone cluster of differentiation 147 (CD147) in MYCN-amplified NB, altogether conferring sensitivity to the AZD3965 and phenformin combination. The result is simultaneous disruption of glycolysis and oxidative phosphorylation, resulting in dramatic disruption of adenosine triphosphate (ATP) production, endoplasmic reticulum (ER) stress, and cell death. In mouse models of MYCN-amplified NB, the combination was tolerable at concentrations where it shrank tumors and did not increase white-blood-cell toxicity compared to single drugs. Therefore, we demonstrate that a metabolic combination screen can identify vulnerabilities in subsets of cancer and put forth a metabolic combination therapy tailored for MYCN-amplified NB that demonstrates efficacy and tolerability in vivo.

Epigenetics and metabolism are closely connected, with oncogene-driven metabolic rewiring modifying the epigenetic landscape through increases of metabolites that are critical for the activities of epigenetic enzymes, and conversely the enhanced epigenetic activity further regulating the activity of metabolic enzymes. Deoxyribonucleic acid (DNA) methyltransferases (DNMTs) play an important role in this, as methylation patterns lead to changes in gene expression that play a role in metabolic reprogramming. NB has recently emerged as an epigenomic disease, with MYCN amplification in NB driving an epigenetically distinct cancer. We have screened epigenetic targets and have found evidence that DNA methyltransferase 1 (DNMT1) represents a druggable vulnerability in these high-risk MYCN-amplified NB. We have found that MYCN-amplified NB has increased expression of DNMT1 correlating with higher methylation levels, which is often associated with poor prognosis. Interestingly, we have found a feed forward loop between MYCN and DNMT1, where MYCN induces DNMT1 expression by binding to a non-canonical E-box near the transcriptional start site (TSS) of DNMT1, and DNMT1 in turn regulates MYCN expression. We found that MYCN expression led to increased levels of methionine and S-adenosyl-L-methionine (SAM), the universal methyl donor for methyltransferase reactions, further enhancing its reliance on DNMT1. We also found that DNMT1 inhibition through treatment with decitabine (DAC) led to mitochondrial dysfunction in MYCN-amplified NB including loss of oxidative phosphorylation. Finally, we found that DNMT1 inhibitors are effective in MYCN-amplified NB patient derived xenograft (PDX) in vivo mouse models, as well as multiple rational combination therapies to enhance DACs anti-NB activity. Overall, we have found that MYCN drives aberrant DNMT1 activity to feed-forward enhance MYCN expression, which in turn dysregulates key metabolic pathways. We propose that pharmacological inhibition of DNMT1 with in-clinic DNMT1 inhibitors can downregulate these pathways and targeting DNMT1 is lethal in MYCN-amplified NB.


© Krista M. Dalton

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Available for download on Tuesday, July 30, 2024