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2013 122: 2770-2771 doi:10.1182/blood-2013-09-522441

Targeting IDH: the next big thing in AML Mark Levis

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Interferon-regulatory factor 4 (IRF4) is a critical transcription factor for hematopoietic development and the immune system. It has been identified as an oncogene in multiple myeloma6 but a tumor suppressor in CLL.7 In this study, Shukla and colleagues backcrossed Vh11 mice which have a dramatically expanded B1 cell population into IRF4deficiency mice and found that 100% (n 5 12) of IRF42/2Vh11 mice developed CLL within 10 months (see figure). Among those IRF42/2Vh11 mice, 70% resemble indolent CLL and 30% exhibit aggressive CLL. Even after just 5 months, 7 of 12 (58%) IRF42/2Vh11 mice developed CLL and the rest developed monoclonal B-cell lymphocytosis (MBL). In contrast, no CLL or MBL was developed in the IRF41/1Vh11 control mice within 12 months. The authors also reported that IgM1CD51 CLL cells started to increase in the blood of IRF42/2Vh11 mice at 2 to 4 months of age and occupied ;69% of peripheral blood mononuclear cells at 8 months of age (see figure). IRF42/2Vh11 mice exhibited splenomegaly and lymph node enlargement and those with aggressive CLL had enlarged livers. The authors then identified the surface phenotype of IRF42/2Vh11 CLL cells as CD191, B220low/2, CD232, CD212, IgDlow, and CD1dint, and demonstrated that IRF42/2Vh11 CLL cells were transplantable in immunodeficient host mice. To further characterize the IRF42/2Vh11 CLL cells, Shukla and coworkers studied proliferation, survival, and molecular signatures of these cells and found that IRF42/2Vh11 CLL cells mainly proliferated in spleen while not in blood nor in lymph node, and these cells were resistant to apoptosis. Consistent with these findings, reexpression of IRF4 in IRF42/2Vh11 CLL cells in vitro promoted apoptosis. Very interestingly, the authors found that the expression of Mcl-1 which is a critical prosurvival factor for CLL cells8 was significantly increased in all 5 IRF42/2Vh11 CLL samples compared with controls, while the expression of TCL1 and miR15a/16-1 was not deregulated. Collectively, the findings in this study1 strongly indicate an important role of IRF4 in the initiation and progression of CLL and potential applications of this novel IRF42/2Vh11 mouse model in understanding CLL etiology and testing preclinical drugs.

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There are also several questions that need to be further addressed. First, in order to verify the role of IRF4 in the initiation of CLL, the transplantable ability of untransformed IRF42/2Vh11 cells should be further investigated. Second, IRF4 deficiency affects other lymphocyte subsets9; therefore, other potential abnormalities in IRF42/2Vh11 mice need to be characterized. Third, as the authors discussed in the manuscript, more generations of backcrossing are preferred to get a pure genetic background in IRF42/2Vh11 mice to further facilitate reproducibility of experiments. Overall, this study represents a significant step forward in our understanding of CLL, and the CLL field is looking forward to the applications of this new mouse model with respect to immunology, experimental therapeutics, and biology of this disease. Conflict-of-interest disclosure: The authors declare no competing financial interests. n REFERENCES 1. Shukla V, Ma S, Hardy RR, Joshi SS, Lu R. A role for IRF4 in the development of CLL. Blood. 2013;122(16):2848-2855.

2. Yan XJ, Albesiano E, Zanesi N, et al. B cell receptors in TCL1 transgenic mice resemble those of aggressive, treatment-resistant human chronic lymphocytic leukemia. Proc Natl Acad Sci USA. 2006;103(31):11713-11718. 3. Klein U, Lia M, Crespo M, et al. The DLEU2/miR15a/16-1 cluster controls B cell proliferation and its deletion leads to chronic lymphocytic leukemia. Cancer Cell. 2010;17(1):28-40. 4. Santanam U, Zanesi N, Efanov A, et al. Chronic lymphocytic leukemia modeled in mouse by targeted miR29 expression. Proc Natl Acad Sci USA. 2010;107(27): 12210-12215. 5. Zapata JM, Krajewska M, Morse HC III, Choi Y, Reed JC. TNF receptor-associated factor (TRAF) domain and Bcl-2 cooperate to induce small B cell lymphoma/chronic lymphocytic leukemia in transgenic mice. Proc Natl Acad Sci USA. 2004;101(47):16600-16605. 6. Shaffer AL, Emre NC, Lamy L, et al. IRF4 addiction in multiple myeloma. Nature. 2008;454(7201):226-231. 7. Chang CC, Lorek J, Sabath DE, et al. Expression of MUM1/IRF4 correlates with clinical outcome in patients with B-cell chronic lymphocytic leukemia. Blood. 2002; 100(13):4671-4675. 8. Pepper C, Lin TT, Pratt G, et al. Mcl-1 expression has in vitro and in vivo significance in chronic lymphocytic leukemia and is associated with other poor prognostic markers. Blood. 2008;112(9):3807-3817. 9. Mittru¨ cker HW, Matsuyama T, Grossman A, et al. Requirement for the transcription factor LSIRF/IRF4 for mature B and T lymphocyte function. Science. 1997; 275(5299):540-543. © 2013 by The American Society of Hematology

l l l MYELOID NEOPLASIA

Comment on Chaturvedi et al, page 2877

Targeting IDH: the next big thing in AML ----------------------------------------------------------------------------------------------------Mark Levis1

1

JOHNS HOPKINS UNIVERSITY

In this issue of Blood, Chaturvedi and coworkers use a small molecule inhibitor of mutant isocitrate dehydrogenase 1 (IDH1) to reverse the myeloproliferative effects induced by the “oncometabolite” 2-hydroxyglutarate (2-HG).1

J

ust occasionally, things in biomedical research actually work out the way they are supposed to. A few years back, groups of researchers all around the world began using next-generation sequencing techniques to sequence whole exomes and whole genomes from acute myeloid leukemia (AML) samples, a grand scheme that was supposed to identify new mutations that could be targeted with new therapeutics. One set of mutations emerging from these screens was found in genes encoding for the 2 isoforms IDH1 and IDH2. The mutations, first identified in gliomas,2 were noted to occur in 15% to 20%

of newly diagnosed AML patients, particularly in those with normal cytogenetics.3,4 After dusting off our biochemistry textbooks and reviewing the intermediates in the Krebs cycle, many of us in the field immediately wondered how such mutations could promote malignant transformation. Investigators who were not intimidated by a little biochemistry took up the challenge, and a clearer picture of how these new mutations promote transformation is now emerging.5-8 The IDH enzymes, as homodimers, convert isocitrate into a-ketoglutarate (a-KG), which turns out to be not only an intermediate in the

BLOOD, 17 OCTOBER 2013 x VOLUME 122, NUMBER 16

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Mutant IDH enzymes convert a Krebs cycle intermediate, a-KG, into 2-HG. This alters the energy balance of the cell and interferes with a host of metabolic functions, including the epigenetic milieu.

formation in methylcellulose of IDH1-mutant human primary bone marrow cells relative to control, thus establishing an important proof of principle that these mutant enzymes can be successfully targeted and their metabolic effects are to some degree reversible. So, it seems that the next big target in AML has been identified. We can anticipate that the pharmaceutical–industrial complex will swing into action and launch an IDH inhibitor arms race of sorts. We can also anticipate limited single-agent activity of such drugs, given what we have learned about this disease over the past decade—namely, that AML emerges from what is at least a 2-step process and that targeting single defects (eg, as with FLT3 inhibitors) is only going to get us part way to our goal of cure. Nonetheless, we can only be encouraged by the progress we are seeing, at how chipping away at these different molecular defects will eventually cause this disease to crumble at last. Conflict-of-interest disclosure: The author declares no competing financial interests. n REFERENCES

Krebs cycle, but also an essential cosubstrate for a host of cellular metabolic functions. Several different dioxygenases, including TET2 and histone demethylases (enzymes that clearly influence the epigenetic milieu of the cell), depend on a-KG for their normal function. The IDH mutations occur at critical arginine residues in the active site of the enzyme (R132 in IDH1 and R140 and R172 in IDH2). The resulting amino acid substitution not only prevents the normal catalytic function of the enzyme, it also results in a new (or neomorphic) function—the conversion of a-KG to 2-HG, which (in the case of the R-enantiomer) is a competitive inhibitor of dioxygenases. For this to work most efficiently, the mutation must occur in heterozygous form, with the wild-type enzyme generating a-KG and the mutant enzyme promptly converting it to 2-HG (see figure). The activity of the mutant heterodimer thus affects the energy landscape of the cell, and, with the generation of the 2-HG “oncometabolite,” leads to epigenetic reprogramming of a cell, blocking differentiation and contributing to a transformed phenotype. Thus, gene

sequencing studies gave us the mutations, whereas the diligent work of the biochemists unraveled the mechanisms that contribute to the transformed phenotype (and, as a bonus, gave us 2 exciting new terms to bandy about: neomorphic and oncometabolite). As has been found with most other putative oncogenes in AML, IDH mutations by themselves do not appear sufficient to induce transformation. Chaturvedi and colleagues take an important next step in this story. They and others had noted increased Hox family gene expression in association with IDH mutations. Using a retrovirally transduced murine bone marrow model, they induced a myeloproliferative phenotype when IDH1 was combined with HoxA9. The double-mutant cells contained increased levels of the inhibitory R-enantiomer of 2-HG, were cycling more, and had decreased cdk gene expression and increased mitogenactivated protein kinase activity. A small molecule inhibitor of mutant IDH1 caused decreased cell-cycling and increased apoptosis in the double-mutant cells. The same inhibitor induced apoptosis and decreased colony

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1. Chaturvedi A, Araujo Cruz MM, Jyotsana N, et al. Mutant IDH1 promotes leukemogenesis in vivo and can be specifically targeted in human AML. Blood. 2013;122(16):2877-2887. 2. Yan H, Parsons DW, Jin G, et al. IDH1 and IDH2 mutations in gliomas. N Engl J Med. 2009;360(8): 765-773. 3. Mardis ER, Ding L, Dooling DJ, et al. Recurring mutations found by sequencing an acute myeloid leukemia genome. N Engl J Med. 2009;361(11): 1058-1066. 4. Abbas S, Lugthart S, Kavelaars FG, et al. Acquired mutations in the genes encoding IDH1 and IDH2 both are recurrent aberrations in acute myeloid leukemia: prevalence and prognostic value. Blood. 2010;116(12): 2122-2126. 5. Dang L, White DW, Gross S, et al. Cancer-associated IDH1 mutations produce 2-hydroxyglutarate. Nature. 2009;462(7274):739-744. 6. Xu W, Yang H, Liu Y, et al. Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of a-ketoglutarate-dependent dioxygenases. Cancer Cell. 2011;19(1):17-30. 7. Pietrak B, Zhao H, Qi H, et al. A tale of two subunits: how the neomorphic R132H IDH1 mutation enhances production of aHG. Biochemistry. 2011;50(21): 4804-4812. 8. Losman JA, Looper RE, Koivunen P, et al. (R)-2hydroxyglutarate is sufficient to promote leukemogenesis and its effects are reversible. Science. 2013;339(6127): 1621-1625. © 2013 by The American Society of Hematology

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