Comprehensive Metabolic Profiling of MYC-Amplified Medulloblastoma Tumors Reveals Key Dependencies on Amino Acid, Tricarboxylic Acid and Hexosamine Pathways.

Khoa Pham,Brad A Poore,Allison R Hanaford,Jesse Alt,Charles G Eberhart,Micah J Maxwell,Heather Sweeney,Barbara S Slusher,Rana Rais,Akhila Parthasarathy,Eric H Raabe

doi:10.3390/cancers14051311

Abstract

Simple SummaryThe oncogene MYC alters cellular metabolism. Medulloblastoma is the most common malignant pediatric brain tumor. MYC-amplified medulloblastoma has a poor prognosis, and the metabolism of MYC-amplified medulloblastoma is poorly understood. We performed comprehensive metabolic profiling of MYC-amplified medulloblastoma and found increased reliance on potentially targetable pathways. We also found that the metabolism of MYC-amplified cell lines differed from orthotopic brain tumors in vitro and in flank tumors, suggesting that analyses conducted in vitro or in flank tumors may miss key vulnerabilities.Reprograming of cellular metabolism is a hallmark of cancer. Altering metabolism allows cancer cells to overcome unfavorable microenvironment conditions and to proliferate and invade. Medulloblastoma is the most common malignant brain tumor of children. Genomic amplification of MYC defines a subset of poor-prognosis medulloblastoma. We performed comprehensive metabolic studies of human MYC-amplified medulloblastoma by comparing the metabolic profiles of tumor cells in three different conditions—in vitro, in flank xenografts and in orthotopic xenografts in the cerebellum. Principal component analysis showed that the metabolic profiles of brain and flank high-MYC medulloblastoma tumors clustered closely together and separated away from normal brain and in vitro MYC-amplified cells. Compared to normal brain, MYC-amplified medulloblastoma orthotopic xenograft tumors showed upregulation of the TCA cycle as well as the synthesis of nucleotides, hexosamines, amino acids and glutathione. There was significantly higher glucose uptake and usage in orthotopic xenograft tumors compared to flank xenograft tumors and cells in culture. In orthotopic tumors, glucose was the main carbon source for the de novo synthesis of glutamate, glutamine and glutathione through the TCA cycle. In vivo, the glutaminase II pathway was the main pathway utilizing glutamine. Glutathione was the most abundant upregulated metabolite in orthotopic tumors compared to normal brain. Glutamine-derived glutathione was synthesized through the glutamine transaminase K (GTK) enzyme in vivo. In conclusion, high MYC medulloblastoma cells have different metabolic profiles in vitro compared to in vivo, and key vulnerabilities may be missed by not performing in vivo metabolic analyses.

Highlights

Malignant transformation is a process that drives normal cells to become cancerous through the accumulation of alterations in proto-oncogenes and tumor suppressors [1–4].Among different types of tumors, genetic alterations in PI3/mTOR, RAS/BRAF, MYC and TP53 reprogram metabolic pathways, allowing cancer cells to overcome unfavorable conditions and enabling them to proliferate at a pathologic rate and metastasize [5–11].Identifying and interrupting the abnormal metabolic pathways that benefit cancer cells could yield a therapeutic index in which cancer cells are targeted while normal cells are not harmed [12–14].In vitro metabolic studies and flux analysis using stable isotopes can provide a picture of the intracellular metabolite levels and how those metabolites change in response to therapy
hematoxylin and eosin (HE) stained sections demonstrate that the orthotopic xenograft tumors used in our study grew in the native microenvironment, with histology similar to that of human primary
In one of the orthotopic MED211 tumors, we found that the tumor and normal brain metabolomes data were distinct from the other samples in the PCA, likely due to technical issues

Summary

Introduction

Malignant transformation is a process that drives normal cells to become cancerous through the accumulation of alterations in proto-oncogenes and tumor suppressors [1–4].Among different types of tumors, genetic alterations in PI3/mTOR, RAS/BRAF, MYC and TP53 reprogram metabolic pathways, allowing cancer cells to overcome unfavorable conditions and enabling them to proliferate at a pathologic rate and metastasize [5–11].Identifying and interrupting the abnormal metabolic pathways that benefit cancer cells could yield a therapeutic index in which cancer cells are targeted while normal cells are not harmed [12–14].In vitro metabolic studies and flux analysis using stable isotopes can provide a picture of the intracellular metabolite levels and how those metabolites change in response to therapy. In vitro models miss the influence from the native microenvironment, such as physiologic or hypoxic oxygen tension and pH and limited availability of nutrients as well as interaction with stromal cells and tumor-associated macrophages, all of which could have a significant impact on the intracellular metabolites of cancer cells [15–18]. These differences could potentially confound the applicability of cell culture metabolic and therapeutic findings to in vivo tumors, as shown in some type of cancers [19–22]. We sought to assess how different types of in vitro and in vivo microenvironments affected the metabolic profiles of medulloblastoma

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