Upregulated heme biosynthesis, an exploitable vulnerability in MYCN-driven leukemogenesis
Yu Fukuda, Yao Wang, Shangli Lian, John Lynch, Shinjiro Nagai, Bruce Fanshawe, Ayten Kandilci, Laura J. Janke, Geoffrey Neale, Yiping Fan, Brian P. Sorrentino, Martine F. Roussel, Gerard Grosveld, John D. Schuetz
Yu Fukuda, Yao Wang, Shangli Lian, John Lynch, Shinjiro Nagai, Bruce Fanshawe, Ayten Kandilci, Laura J. Janke, Geoffrey Neale, Yiping Fan, Brian P. Sorrentino, Martine F. Roussel, Gerard Grosveld, John D. Schuetz
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Research Article Metabolism

Upregulated heme biosynthesis, an exploitable vulnerability in MYCN-driven leukemogenesis

Abstract

The increased heme biosynthesis long observed in leukemia was previously of unknown significance. Heme, synthesized from porphyrin precursors, plays a central role in oxygen metabolism and mitochondrial function, yet little is known about its role in leukemogenesis. Here, we show increased expression of heme biosynthetic genes, including UROD, only in pediatric AML samples that have high MYCN expression. High expression of both UROD and MYCN predicts poor overall survival and unfavorable outcomes in adult AML. Murine leukemic progenitors derived from hematopoietic progenitor cells (HPCs) overexpressing a MYCN cDNA (MYCN-HPCs) require heme/porphyrin biosynthesis, accompanied by increased oxygen consumption, to fully engage in self-renewal and oncogenic transformation. Blocking heme biosynthesis reduced mitochondrial oxygen consumption and markedly suppressed self-renewal. Leukemic progenitors rely on balanced production of heme and heme intermediates, the porphyrins. Porphyrin homeostasis is required because absence of the porphyrin exporter, ABCG2, increased death of leukemic progenitors in vitro and prolonged the survival of mice transplanted with Abcg2-KO MYCN-HPCs. Pediatric AML patients with elevated MYCN mRNA display strong activation of TP53 target genes. Abcg2-KO MYCN-HPCs were rescued from porphyrin toxicity by p53 loss. This vulnerability was exploited to show that treatment with a porphyrin precursor, coupled with the absence of ABCG2, blocked MYCN-driven leukemogenesis in vivo, thereby demonstrating that porphyrin homeostasis is a pathway crucial to MYCN leukemogenesis.

Authors

Yu Fukuda, Yao Wang, Shangli Lian, John Lynch, Shinjiro Nagai, Bruce Fanshawe, Ayten Kandilci, Laura J. Janke, Geoffrey Neale, Yiping Fan, Brian P. Sorrentino, Martine F. Roussel, Gerard Grosveld, John D. Schuetz

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Figure 4

Porphyrin cytotoxicity is abrogated by p53 absence, and AML is cured by enhancing porphyrins in the absence of ABCG2.

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Porphyrin cytotoxicity is abrogated by p53 absence, and AML is cured by ...
WT-MYCN HPCs were incubated with increasing doses of ALA for 72 hours in the absence or presence of FTC. (A) Cell viability and (B) PPIX levels were measured (n = 3, mean ± SEM). (C) Abcg2-KO MYCN-HPCs accumulate high levels of porphyrins compared with WT-MYCN HPCs (n = 3, mean ± SEM). (D) High porphyrin levels are especially detrimental to colony formation of Abcg2-KO MYCN-HPCs (n = 4, mean ± SEM). (E) AML patients with MYCN levels in the upper quartile display strong activation of a p53 gene set. (F) Abcg2-KO MYCN-HPCs show activation of p53 target genes. (G) CFU-c assays were performed using MYCN-transduced HPCs isolated from mice of the indicated genotypes. Cells were incubated with ALA and colonies were counted. Loss of p53 renders Abcg2-KO MYCN cells resistant to porphyrin toxicity (2 independent experiments, with 2 replicates per condition per experiment, and values shown as an aggregate mean ± SD; 1-way ANOVA). (H) WT or Abcg2-KO MYCN-expressing HPCs were treated with ALA for 24 hours prior to transplantation into lethally irradiated recipient mice. Survival curve showing that Abcg2-KO cells treated with ALA did not produce leukemia. Median survival for WT vehicle, Abcg2-KO vehicle, and WT ALA were 61 days, 97 days, and 91 days, respectively. The Abcg2-KO and WT leukemia were myeloperoxidase positive, as shown in Supplemental Figure 6. Median survival for Abcg2-KO ALA was not determined (n = 10 for vehicle-treated and n = 14 for ALA-treated samples; P = 0.05 between WT vehicle and Abcg2-KO vehicle, log-rank Mantel-Cox test). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Two-way ANOVA was used for A–D.