Abstract
Standard methods to assess tumor progression and treatment response for leukemia patients rely on repeat sampling of bone marrow. These methods evaluate overall disease status, but are not sensitive to early biochemical or cellular responses to therapies. Identification and validation of additional pharmacodynamic endpoints that are useful for accurate prediction of therapeutic efficacy would allow for earlier assessment of clinical response and thereby facilitate the development of effective individualized treatment regimens. Ideally, these markers would be assessed using non-invasive methods. Toward this end, we used in vivo magnetic resonance imaging (MRI) and ex vivo magnetic resonance spectroscopy (MRS) to investigate changes in the bone marrow, spleen, and/or blood of leukemic MLL-AF9 transgenic (Tg) mice relative to wild-type littermates.
MLL-AF9 Tg mice with leukemia exhibited a statistically significant 1.5-fold increase in bone marrow T1-weighted MRI signal intensity. Increased signal intensity preceded development of leukemia and is therefore likely to be due, at least in part, to increased bone marrow cellularity, initially as a result of pre-neoplastic myeloproliferation and later as a result of marrow infiltration with leukemic blasts. We are currently determining microvessel density in the bone marrow of leukemic MLL-AF9 Tg mice and littermate controls to determine whether an increase in blood supply may also contribute to changes in MRI signal intensity in leukemic mice. These studies suggest that T1-weighted MRI signal intensity may be useful as an indicator of bone marrow tumor burden.
Leukemic MLL-AF9 Tg mice also exhibited statistically significant changes in metabolite levels in spleen, blood, and bone marrow. The Warburg effect, whereby cancer cells utilize aerobic glycolysis to meet their increased energy demands, was evident in all tissues examined as indicated by increased glycolysis rates, increased glucose utilization, and increased levels of lactate and alanine, the end-products of glycolysis. Additional changes in metabolite levels were observed in the bone marrow and/or spleen of leukemic mice. Absolute levels of glutathione were increased. Glutathione reduces reactive oxygen species that are generated as a result of increased glycolysis and high levels of glutathione are associated with chemoresistance and poor prognosis in patients with acute leukemia (Maung et al., 1994, Leukemia 8:1487–91; Kearns et al., 2001, Blood 97:393–8). Increased glycine levels were also observed and may be associated with increased pyridine and DNA synthesis in leukemia cells. Decreased glutamate and glutamine levels may reflect a decrease in utilization of the mitochondrial Krebs cycle in tumor cells. Decreased levels of myo-inositol and taurine, which function as osmoregulators, may occur as a result of osmotic stress due to increased cellularity in leukemic organs. Levels of aromatic acids, lysine and arginine, and creatine and phosphocreatine were also decreased. Many of these changes in metabolism are recapitulated in immunocompetent mice orthotopically transplanted with MLL-AF9 Tg leukemias.
This study represents the first description of a comprehensive analysis of the “metabolomic” profile associated with development of acute leukemia and is, to our knowledge, the first description of changes in metabolism in an animal model of acute leukemia. The data presented here suggest novel metabolic targets for therapeutic intervention. In addition, because metabolic changes often precede detectable changes in tumor burden, they may be particularly useful as early indicators of therapeutic efficacy and may thereby allow for more rapid determination of clinical response, decreased exposure to toxic therapies in resistant patients, and more expedient conversion to effective therapies. Because changes in glucose metabolism are a central feature of tumorigenesis and changes in glutathione levels have been associated with clinical outcome in patients with acute leukemia, these metabolites are of particular interest. Future studies will use MLL-AF9 transgenic mice to investigate the roles of metabolic changes during de novo development of leukemia. Mice transplanted with MLL-AF9 Tg leukemias will also be used for studies investigating the utility of metabolic changes as indicators of therapeutic efficacy.
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