From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
equivalent CR rates; however, no superiority in OS has been shown.8 Concerns about toxicity of high-dose (90 mg/m2) daunorubicin, the wide use of the 60-mg/m2 dose as a newer “standard,” and similar remission rates of the 60-mg/m2 dose led the UK NCRI AML Study Group to compare the 60- vs 90-mg/m2 doses in induction in a prospective randomized trial.1 At first glance, the conclusions from the study were that there is no benefit to the higher daunorubicin dosing, nor was there a subgroup where the higher dose benefited patients. However, one needs to look beyond the simple calculation of anthracycline dose in the first induction and consider the total dose of anthracycline given over several courses. Compared with the ECOG, the Dutch-Belgian Cooperative Trial Group for Hematology Oncology (HOVON), and Korean trials, where a total of 270 mg/m2 of daunorubicin was given to a majority of patients,6,9,10 in this trial, the total dose of daunorubicin was 330 mg/m2 for the 60-mg/m2 group and 420 mg/m2 for the 90-mg/m2 group. Therefore, the total anthracycline in the lower-dose arm was slightly higher than the high-dose arms in the 3 randomized trials. There were also differences in the dose schedules of anthracycline, where pharmacokinetics and pharmacodynamics could have impacted leukemic cell exposure and outcomes. The UK NCRI AML17 trial brings to light that the true benefit of anthracycline in AML may be in a defined therapeutic window. A threshold level is required for benefit in CR rates, but further dose escalation of the drug may not significantly improve, and may even worsen, outcomes. CR rates were similar in both arms, with no difference in mortality at 30 days. However, in addition to the lack of benefit of the higher dose, the trial was stopped prematurely because of the higher 60-day mortality noted in the 90-mg/m2 arm. Perhaps this study defines the point of diminishing returns where the benefit of increasing anthracycline begins to erode. The final factor in this trial is the short follow-up of the UK NCRI AML17 study group. The other published comparative trials had a much longer follow-up. Moreover, with time, certain interactions have become more evident. For example, with an 80-month follow-up in the ECOG trial, FLT3ITD–positive AML patients did benefit
BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
from a higher dose of anthracycline. In the UK NCRI AML17 trial, there was a trend to a higher-dose daunorubicin benefit in the FLT3-positive patients. Longer follow-up is required to determine whether it becomes significant. The appealing factor here for the UK NCRI AML17 study is that the lower dose of daunorubicin may allow for a third targeting drug to be more easily inserted into the regimen to treat FLT3-positive, core binding factor-positive, or other subgroups of AML patients. The foundation of therapy for this disease has been available for 40 years. Collectively, we are optimizing the appropriate dosing in induction. In this and future studies, these variations on the theme need to be perused and not just given a quick glance. Conflict-of-interest disclosure: The author declares no competing financial interests. n REFERENCES 1. Burnett AK, Russell NH, Hills RK, et al, on behalf of the UK NCRI AML Study Group. A randomized comparison of daunorubicin 90 mg/m2 vs 60 mg/m2 in AML induction: results from the UK NCRI AML17 trial in 1206 patients. Blood. 2015;125(25):3878-3885. 2. Yates JW, Wallace HJ Jr, Ellison RR, Holland JF. Cytosine arabinoside (NSC-63878) and daunorubicin (NSC-83142) therapy in acute nonlymphocytic leukemia. Cancer Chemother Rep. 1973;57(4):485-488. 3. Bishop JF, Matthews JP, Young GA, et al. A randomized study of high-dose cytarabine in induction in acute myeloid leukemia. Blood. 1996;87(5): 1710-1717.
4. Weick JK, Kopecky KJ, Appelbaum FR, et al. A randomized investigation of high-dose versus standarddose cytosine arabinoside with daunorubicin in patients with previously untreated acute myeloid leukemia: a Southwest Oncology Group study. Blood. 1996;88(8): 2841-2851. 5. Hann IM, Stevens RF, Goldstone AH, et al; Adult and Childhood Leukaemia Working Parties of the Medical Research Council. Randomized comparison of DAT versus ADE as induction chemotherapy in children and younger adults with acute myeloid leukemia. Results of the Medical Research Council’s 10th AML trial (MRC AML10). Blood. 1997;89(7):2311-2318. 6. Fernandez HF, Sun Z, Yao X, et al. Anthracycline dose intensification in acute myeloid leukemia. N Engl J Med. 2009;361(13):1249-1259. 7. Luskin MR, Lee J-W, Fernandez HF, et al. High dose daunorubicin improves survival in AML up to age 60, across all cytogenetic risk groups including patients with unfavorable cytogenetic risk, and FLT3-ITD mutant AML: updated analyses from Eastern Cooperative Oncology Trial E1900 [abstract]. Blood. 2014;124(21). Abstract 373. 8. Mandelli F, Vignetti M, Suciu S, et al. Daunorubicin versus mitoxantrone versus idarubicin as induction and consolidation chemotherapy for adults with acute myeloid leukemia: the EORTC and GIMEMA Groups Study AML-10. J Clin Oncol. 2009; 27(32):5397-5403. 9. Lee JH, Joo YD, Kim H, et al; Cooperative Study Group A for Hematology. A randomized trial comparing standard versus high-dose daunorubicin induction in patients with acute myeloid leukemia. Blood. 2011;118(14): 3832-3841. 10. L¨owenberg B, Ossenkoppele GJ, van Putten W, et al; Dutch-Belgian Cooperative Trial Group for HematoOncology (HOVON); German AML Study Group (AMLSG); Swiss Group for Clinical Cancer Research (SAKK) Collaborative Group. High-dose daunorubicin in older patients with acute myeloid leukemia. N Engl J Med. 2009;361(13):1235-1248. © 2015 by The American Society of Hematology
l l l IMMUNOBIOLOGY
Comment on Goettel et al, page 3886
Daring to learn from humanized mice ----------------------------------------------------------------------------------------------------Silke Paust and Matthew Bettini
BAYLOR COLLEGE OF MEDICINE
In this issue of Blood, Goettel and colleagues introduce a novel humanized mouse model of immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome, whereby mice lacking murine major histocompatibility complex class II (MHC II) and expressing human HLA-DR1 (NOD. PrkdcscidIl2rg 2/2H2-Ab12/2; NSGAb°DR1) are reconstituted with hematopoietic stem cells (HSCs) from a patient with IPEX syndrome to generate a humanized model for primary immune deficiency presenting as fatal autoimmunity.1
“H
umanized mice” allow the study of human immune cells in the context of human diseases for the evaluation of organ-specific pathologies for which human
samples may not be available or accessible, and for the assessment of experimental approaches that may be harmful when performed in patients or human volunteers. Thus,
3829
From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
Overview of the generation of the HLA-DR1-NSG humanized IPEX syndrome mouse models, appropriate controls, and disease phenotypes. TCR, T-cell receptor.
humanized mice have become a valuable translational research tool for the study of pathogen transmission, immunopathogenesis, viral latency, pathogen evolution, immune escape, prevention therapy assessments, and vaccine development.2 Humanization of tissues in mice is usually achieved by the transplant of human tissues and/or HSCs into lymphopenic hosts. The resulting cohort of humanized mice is genetically identical (individual members within a mouse strain are genetically identical due to a century of sibling inbreeding),3 and several dozen humanized mice can be generated from 1 human donor. Now, adoptive transfers of isolated human lymphocytes can be performed without rejection by the recipient immune system, and antibody-mediated in vivo lymphocyte depletion enables the assessment of the importance of individual immune cell types or pathways. Several sophisticated humanized mouse models that differ in host genetics and/or their reconstitution methods have been established, including (1) mice that allow the evaluation of functional human innate and adaptive immune responses to infection
3830
and/or vaccination2; (2) mice that enable the humanization of hepatocytes and the study of hepatotropic pathogens4 such as malaria parasites or hepatitis viruses; and (3) germfree mice, in which a human microbiome has been established by oral fecal transplant.5 However, translational autoimmune disease models are rare, with the exception of graft-versus-host disease models, and currently, no humanized mouse models exist to study primary immune deficiency disorders, which present as autoimmune diseases. In a particularly elegant study in the current issue of Blood, Goettel and colleagues describe the development and disease phenotype of a novel humanized mouse model of a primary immunodeficiency called IPEX syndrome.1 IPEX syndrome is a heritable X-linked recessive disorder caused by a number of mutations, mostly missense, found in the DNA-binding domain of the forkhead box P3 (FOXP3) protein.6 The FOXP3 protein is required for the development and function of CD41CD251 regulatory T cells (Tregs) derived from both the thymus Tregs and periphery Tregs. FOXP3-expressing Tregs are critical in the establishment and maintenance of peripheral self-tolerance.
Therefore, the immunopathology of patients with IPEX syndrome is swift, often presenting between 1 and 3 months of age, and includes autoimmune enteropathy; endocrinopathy; eczematous dermatitis, which manifests as excessive cytokine production; elevated immunoglobulin (Ig)E levels; and chronic inflammation, leading to death. The various mutations within FOXP3 can lead to multiple functional fates, including a loss of transcriptional repression, messenger RNA stability, DNA binding, and FOXP3 dimerization. Goettel et al have significantly advanced the study of IPEX syndrome in vivo and, potentially, any hematopoiesis-derived disease, in their newly generated NSGAb°DR1 mouse model. Mice that carry the scurfy mutation have defective FOXP3 that closely resembles IPEX syndrome, including widespread autoimmunity and high quantities of lymphocyte infiltrates within the lung, pancreas, stomach, skin, liver, and gut.7 However, the recapitulation of IPEX within mice using HSCs from IPEX patients has not been reported. In this issue, Goettel and colleagues successfully transplanted CD341 cells into an NOD.PrkdcscidIl2rg2/2 (NSG) host that expresses the human MHC II HLA-DR1 (NSGAb°DR1), in the absence of murine MHC II (HLA-DR1-NSG; see figure). The lack of murine MHC II presumably allows for the full maturation of T and B cells and promotes T-cell help to B cells and full immune function. Indeed, Goettel and colleagues demonstrate that when CD341 cells are engrafted into the HLA-DR1–expressing NSG mice, there is a significant improvement in both mature B- and T-lymphocyte development and function. Importantly, the HLA-DR1–expressing mice are able to develop delayed-type hypersensitivity, exhibiting increased IgG and IgE levels. To test whether this improved adaptive immune response could recapitulate a spontaneous and defined immunologic disorder, Goettel and colleagues transplanted HSCs from an IPEX patient. Remarkably, nearly all of the IPEX (DR1) (NSGAb°DR1) mice succumbed to multiorgan inflammation due to autoantibody production and the lack of a functional Treg compartment (see figure). It is important to note that IPEX(NSG) mice without the HLA-DR1–expressing allele displayed an increase in serum antibodies and an overall absolute increase in T-cell numbers;
BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
however, these mice did not develop widespread autoimmunity. This result is particularly significant because it highlights the importance of an intact mature lymphocyte compartment in the establishment of a fully adaptive immune response. There are many diseases that will benefit from the advancement of immune disease–specific humanized mice, including monogenic immune disorders in which there is reliance on the adaptive immune arm with an intact mature T- and B-cell compartment. The newly developed NSGAb°DR1 mouse emphasizes just how invaluable a mouse model can be to dissect complex pathological disorders, including IPEX syndrome and other rare monogenic immune disorders, diabetes, and inflammatory bowel disease.8 By utilizing CD341 HSCs, Goettel and colleagues are able to select T cells in the thymus on HLA-DR1, although the donor population was HLA mismatched. The ability to study the development of a spontaneous disease in-depth, before onset and during the development of pathogenesis, will move science forward immensely. In particular, the NSGAb°DR1 mouse will provide novel information about the initiation of IPEX due to particular point mutations in FOXP3, including direct and indirect involvement of Tregs in B-cell maturation and autoantibody production, and interactions with other immune populations including natural killer cells and macrophages. It will be interesting to learn in future studies how closely NSGAb°DR1-derived IPEX Tregs functionally behave in comparison with human-derived IPEX Tregs. Can NSGAb°DR1 Tregs express similar amounts of inhibitory cytokines (interleukin [IL]-35, IL-10, and transforming growth factor b) and inhibitory molecules (programmed death ligand 1 and cytotoxic T-lymphocyte–associated protein 4)?9,10 Thus, NSGAb°DR1 mice, alone or in combination with additional HLA molecules and/or other human cytokines, will enable in-depth analyses of developmental and functional defects that underlie immunodeficiencies and autoimmune disorders, and much-needed evaluations of emerging immunotherapies in a preclinical translational animal model. Conflict-of-interest disclosure: The authors declare no competing financial interests. n BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
REFERENCES
6. Verbsky JW, Chatila TA. Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) and IPEX-related disorders: an evolving web of heritable autoimmune diseases. Curr Opin Pediatr. 2013;25(6): 708-714.
1. Goettel JA, Biswas S, Lexmond WS, et al. Fatal autoimmunity in mice reconstituted with human hematopoietic stem cells encoding defective FOXP3. Blood. 2015;125(25):3886-3895. 2. Brehm MA, Jouvet N, Greiner DL, Shultz LD. Humanized mice for the study of infectious diseases. Curr Opin Immunol. 2013;25(4):428-435. 3. Keane TM, Goodstadt L, Danecek P, et al. Mouse genomic variation and its effect on phenotypes and gene regulation. Nature. 2011;477(7364):289-294. 4. Bissig KD, Wieland SF, Tran P, et al. Human liver chimeric mice provide a model for hepatitis B and C virus infection and treatment. J Clin Invest. 2010;120(3): 924-930. 5. Turnbaugh PJ, Ridaura VK, Faith JJ, et al. The effect of diet on the human gut microbiome: a metagenomic analysis in humanized gnotobiotic mice. Sci Transl Med. 2009;1(6):6ra14.
7. Ramsdell F, Ziegler SF. FOXP3 and scurfy: how it all began. Nat Rev Immunol. 2014;14(5):343-349. 8. Touitou I, Galeotti C, Rossi-Semerano L, Hentgen V, Piram M, Kon´e-Paut I; CeR´eMAI, French reference center for autoinflammatory diseases. The expanding spectrum of rare monogenic autoinflammatory diseases. Orphanet J Rare Dis. 2013;8:162. 9. Nguyen LT, Ohashi PS. Clinical blockade of PD1 and LAG3—potential mechanisms of action. Nat Rev Immunol. 2015;15(1):45-56. 10. Vignali DA, Collison LW, Workman CJ. How regulatory T cells work. Nat Rev Immunol. 2008;8(7):523-532. © 2015 by The American Society of Hematology
l l l IMMUNOBIOLOGY
Comment on Schwartz et al, page 3896
Mechanism of enhanced eosinophil survival in inflammation ----------------------------------------------------------------------------------------------------Nives Zimmermann and Marc E. Rothenberg
CINCINNATI CHILDREN’S HOSPITAL MEDICAL CENTER
In this issue of Blood, Schwartz et al identify the nuclear factor (NF)-kB/Bcl-xL pathway as critical for regulation of eosinophil survival in inflammatory conditions.1
C
ell death is essential for many physiologic processes, and its dysregulation characterizes numerous human diseases. This is especially true for eosinophils, because these cells do not undergo substantial extramedullary hematopoiesis, yet their tissue levels can be markedly and selectively increased by a combination of recruitment from the blood and regulation of their cell death within tissues. The field of eosinophil survival has come a long way since the original finding that human blood–derived eosinophils had prolonged survival (.14 days) when cocultured with endothelial cells and with the subsequent identification of interleukin (IL)-5, granulocyte macrophage–colony-stimulating factor (GM-CSF), and IL-3 as key eosinophil survival factors.2 It is now appreciated that these eosinophil-directed hematopoietins inhibit eosinophil apoptosis and that IL-5 has a special capacity to promote eosinophil development, which has led to several therapeutics that lower eosinophils by
blocking IL-5, such as mepolizumab and reslizumab, which are at advanced stages of development.3 However, whether eosinophils are better “dead or alive” is still an unresolved question. Simplistically, both living and dead eosinophils can lead to positive and negative outcomes (see figure). For instance, eosinophils are homeostatically present in some tissues with no ill effect (figure panel A, upper left)4 and have been shown to contribute to a variety of homeostatic functions, including production of secretory immunoglobulin A in the intestine.5 Yet during many eosinophilic inflammatory diseases, eosinophils display an activated phenotype and likely induce tissue damage (figure panel A, upper right).6 Eosinophil cell death was previously viewed dichotomously, as either noninflammatory apoptotic (figure panel A, lower left) or proinflammatory necrotic (figure panel A, lower right) cell death, but recent studies have identified additional cell death pathways, including regulated necrosis,
3831
From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
2015 125: 3829-3831 doi:10.1182/blood-2015-04-639435
Daring to learn from humanized mice Silke Paust and Matthew Bettini
Updated information and services can be found at: http://www.bloodjournal.org/content/125/25/3829.full.html Articles on similar topics can be found in the following Blood collections Free Research Articles (4041 articles) Information about reproducing this article in parts or in its entirety may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#repub_requests Information about ordering reprints may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#reprints Information about subscriptions and ASH membership may be found online at: http://www.bloodjournal.org/site/subscriptions/index.xhtml
Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly by the American Society of Hematology, 2021 L St, NW, Suite 900, Washington DC 20036. Copyright 2011 by The American Society of Hematology; all rights reserved.
equivalent CR rates; however, no superiority in OS has been shown.8 Concerns about toxicity of high-dose (90 mg/m2) daunorubicin, the wide use of the 60-mg/m2 dose as a newer “standard,” and similar remission rates of the 60-mg/m2 dose led the UK NCRI AML Study Group to compare the 60- vs 90-mg/m2 doses in induction in a prospective randomized trial.1 At first glance, the conclusions from the study were that there is no benefit to the higher daunorubicin dosing, nor was there a subgroup where the higher dose benefited patients. However, one needs to look beyond the simple calculation of anthracycline dose in the first induction and consider the total dose of anthracycline given over several courses. Compared with the ECOG, the Dutch-Belgian Cooperative Trial Group for Hematology Oncology (HOVON), and Korean trials, where a total of 270 mg/m2 of daunorubicin was given to a majority of patients,6,9,10 in this trial, the total dose of daunorubicin was 330 mg/m2 for the 60-mg/m2 group and 420 mg/m2 for the 90-mg/m2 group. Therefore, the total anthracycline in the lower-dose arm was slightly higher than the high-dose arms in the 3 randomized trials. There were also differences in the dose schedules of anthracycline, where pharmacokinetics and pharmacodynamics could have impacted leukemic cell exposure and outcomes. The UK NCRI AML17 trial brings to light that the true benefit of anthracycline in AML may be in a defined therapeutic window. A threshold level is required for benefit in CR rates, but further dose escalation of the drug may not significantly improve, and may even worsen, outcomes. CR rates were similar in both arms, with no difference in mortality at 30 days. However, in addition to the lack of benefit of the higher dose, the trial was stopped prematurely because of the higher 60-day mortality noted in the 90-mg/m2 arm. Perhaps this study defines the point of diminishing returns where the benefit of increasing anthracycline begins to erode. The final factor in this trial is the short follow-up of the UK NCRI AML17 study group. The other published comparative trials had a much longer follow-up. Moreover, with time, certain interactions have become more evident. For example, with an 80-month follow-up in the ECOG trial, FLT3ITD–positive AML patients did benefit
BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
from a higher dose of anthracycline. In the UK NCRI AML17 trial, there was a trend to a higher-dose daunorubicin benefit in the FLT3-positive patients. Longer follow-up is required to determine whether it becomes significant. The appealing factor here for the UK NCRI AML17 study is that the lower dose of daunorubicin may allow for a third targeting drug to be more easily inserted into the regimen to treat FLT3-positive, core binding factor-positive, or other subgroups of AML patients. The foundation of therapy for this disease has been available for 40 years. Collectively, we are optimizing the appropriate dosing in induction. In this and future studies, these variations on the theme need to be perused and not just given a quick glance. Conflict-of-interest disclosure: The author declares no competing financial interests. n REFERENCES 1. Burnett AK, Russell NH, Hills RK, et al, on behalf of the UK NCRI AML Study Group. A randomized comparison of daunorubicin 90 mg/m2 vs 60 mg/m2 in AML induction: results from the UK NCRI AML17 trial in 1206 patients. Blood. 2015;125(25):3878-3885. 2. Yates JW, Wallace HJ Jr, Ellison RR, Holland JF. Cytosine arabinoside (NSC-63878) and daunorubicin (NSC-83142) therapy in acute nonlymphocytic leukemia. Cancer Chemother Rep. 1973;57(4):485-488. 3. Bishop JF, Matthews JP, Young GA, et al. A randomized study of high-dose cytarabine in induction in acute myeloid leukemia. Blood. 1996;87(5): 1710-1717.
4. Weick JK, Kopecky KJ, Appelbaum FR, et al. A randomized investigation of high-dose versus standarddose cytosine arabinoside with daunorubicin in patients with previously untreated acute myeloid leukemia: a Southwest Oncology Group study. Blood. 1996;88(8): 2841-2851. 5. Hann IM, Stevens RF, Goldstone AH, et al; Adult and Childhood Leukaemia Working Parties of the Medical Research Council. Randomized comparison of DAT versus ADE as induction chemotherapy in children and younger adults with acute myeloid leukemia. Results of the Medical Research Council’s 10th AML trial (MRC AML10). Blood. 1997;89(7):2311-2318. 6. Fernandez HF, Sun Z, Yao X, et al. Anthracycline dose intensification in acute myeloid leukemia. N Engl J Med. 2009;361(13):1249-1259. 7. Luskin MR, Lee J-W, Fernandez HF, et al. High dose daunorubicin improves survival in AML up to age 60, across all cytogenetic risk groups including patients with unfavorable cytogenetic risk, and FLT3-ITD mutant AML: updated analyses from Eastern Cooperative Oncology Trial E1900 [abstract]. Blood. 2014;124(21). Abstract 373. 8. Mandelli F, Vignetti M, Suciu S, et al. Daunorubicin versus mitoxantrone versus idarubicin as induction and consolidation chemotherapy for adults with acute myeloid leukemia: the EORTC and GIMEMA Groups Study AML-10. J Clin Oncol. 2009; 27(32):5397-5403. 9. Lee JH, Joo YD, Kim H, et al; Cooperative Study Group A for Hematology. A randomized trial comparing standard versus high-dose daunorubicin induction in patients with acute myeloid leukemia. Blood. 2011;118(14): 3832-3841. 10. L¨owenberg B, Ossenkoppele GJ, van Putten W, et al; Dutch-Belgian Cooperative Trial Group for HematoOncology (HOVON); German AML Study Group (AMLSG); Swiss Group for Clinical Cancer Research (SAKK) Collaborative Group. High-dose daunorubicin in older patients with acute myeloid leukemia. N Engl J Med. 2009;361(13):1235-1248. © 2015 by The American Society of Hematology
l l l IMMUNOBIOLOGY
Comment on Goettel et al, page 3886
Daring to learn from humanized mice ----------------------------------------------------------------------------------------------------Silke Paust and Matthew Bettini
BAYLOR COLLEGE OF MEDICINE
In this issue of Blood, Goettel and colleagues introduce a novel humanized mouse model of immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome, whereby mice lacking murine major histocompatibility complex class II (MHC II) and expressing human HLA-DR1 (NOD. PrkdcscidIl2rg 2/2H2-Ab12/2; NSGAb°DR1) are reconstituted with hematopoietic stem cells (HSCs) from a patient with IPEX syndrome to generate a humanized model for primary immune deficiency presenting as fatal autoimmunity.1
“H
umanized mice” allow the study of human immune cells in the context of human diseases for the evaluation of organ-specific pathologies for which human
samples may not be available or accessible, and for the assessment of experimental approaches that may be harmful when performed in patients or human volunteers. Thus,
3829
From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
Overview of the generation of the HLA-DR1-NSG humanized IPEX syndrome mouse models, appropriate controls, and disease phenotypes. TCR, T-cell receptor.
humanized mice have become a valuable translational research tool for the study of pathogen transmission, immunopathogenesis, viral latency, pathogen evolution, immune escape, prevention therapy assessments, and vaccine development.2 Humanization of tissues in mice is usually achieved by the transplant of human tissues and/or HSCs into lymphopenic hosts. The resulting cohort of humanized mice is genetically identical (individual members within a mouse strain are genetically identical due to a century of sibling inbreeding),3 and several dozen humanized mice can be generated from 1 human donor. Now, adoptive transfers of isolated human lymphocytes can be performed without rejection by the recipient immune system, and antibody-mediated in vivo lymphocyte depletion enables the assessment of the importance of individual immune cell types or pathways. Several sophisticated humanized mouse models that differ in host genetics and/or their reconstitution methods have been established, including (1) mice that allow the evaluation of functional human innate and adaptive immune responses to infection
3830
and/or vaccination2; (2) mice that enable the humanization of hepatocytes and the study of hepatotropic pathogens4 such as malaria parasites or hepatitis viruses; and (3) germfree mice, in which a human microbiome has been established by oral fecal transplant.5 However, translational autoimmune disease models are rare, with the exception of graft-versus-host disease models, and currently, no humanized mouse models exist to study primary immune deficiency disorders, which present as autoimmune diseases. In a particularly elegant study in the current issue of Blood, Goettel and colleagues describe the development and disease phenotype of a novel humanized mouse model of a primary immunodeficiency called IPEX syndrome.1 IPEX syndrome is a heritable X-linked recessive disorder caused by a number of mutations, mostly missense, found in the DNA-binding domain of the forkhead box P3 (FOXP3) protein.6 The FOXP3 protein is required for the development and function of CD41CD251 regulatory T cells (Tregs) derived from both the thymus Tregs and periphery Tregs. FOXP3-expressing Tregs are critical in the establishment and maintenance of peripheral self-tolerance.
Therefore, the immunopathology of patients with IPEX syndrome is swift, often presenting between 1 and 3 months of age, and includes autoimmune enteropathy; endocrinopathy; eczematous dermatitis, which manifests as excessive cytokine production; elevated immunoglobulin (Ig)E levels; and chronic inflammation, leading to death. The various mutations within FOXP3 can lead to multiple functional fates, including a loss of transcriptional repression, messenger RNA stability, DNA binding, and FOXP3 dimerization. Goettel et al have significantly advanced the study of IPEX syndrome in vivo and, potentially, any hematopoiesis-derived disease, in their newly generated NSGAb°DR1 mouse model. Mice that carry the scurfy mutation have defective FOXP3 that closely resembles IPEX syndrome, including widespread autoimmunity and high quantities of lymphocyte infiltrates within the lung, pancreas, stomach, skin, liver, and gut.7 However, the recapitulation of IPEX within mice using HSCs from IPEX patients has not been reported. In this issue, Goettel and colleagues successfully transplanted CD341 cells into an NOD.PrkdcscidIl2rg2/2 (NSG) host that expresses the human MHC II HLA-DR1 (NSGAb°DR1), in the absence of murine MHC II (HLA-DR1-NSG; see figure). The lack of murine MHC II presumably allows for the full maturation of T and B cells and promotes T-cell help to B cells and full immune function. Indeed, Goettel and colleagues demonstrate that when CD341 cells are engrafted into the HLA-DR1–expressing NSG mice, there is a significant improvement in both mature B- and T-lymphocyte development and function. Importantly, the HLA-DR1–expressing mice are able to develop delayed-type hypersensitivity, exhibiting increased IgG and IgE levels. To test whether this improved adaptive immune response could recapitulate a spontaneous and defined immunologic disorder, Goettel and colleagues transplanted HSCs from an IPEX patient. Remarkably, nearly all of the IPEX (DR1) (NSGAb°DR1) mice succumbed to multiorgan inflammation due to autoantibody production and the lack of a functional Treg compartment (see figure). It is important to note that IPEX(NSG) mice without the HLA-DR1–expressing allele displayed an increase in serum antibodies and an overall absolute increase in T-cell numbers;
BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
however, these mice did not develop widespread autoimmunity. This result is particularly significant because it highlights the importance of an intact mature lymphocyte compartment in the establishment of a fully adaptive immune response. There are many diseases that will benefit from the advancement of immune disease–specific humanized mice, including monogenic immune disorders in which there is reliance on the adaptive immune arm with an intact mature T- and B-cell compartment. The newly developed NSGAb°DR1 mouse emphasizes just how invaluable a mouse model can be to dissect complex pathological disorders, including IPEX syndrome and other rare monogenic immune disorders, diabetes, and inflammatory bowel disease.8 By utilizing CD341 HSCs, Goettel and colleagues are able to select T cells in the thymus on HLA-DR1, although the donor population was HLA mismatched. The ability to study the development of a spontaneous disease in-depth, before onset and during the development of pathogenesis, will move science forward immensely. In particular, the NSGAb°DR1 mouse will provide novel information about the initiation of IPEX due to particular point mutations in FOXP3, including direct and indirect involvement of Tregs in B-cell maturation and autoantibody production, and interactions with other immune populations including natural killer cells and macrophages. It will be interesting to learn in future studies how closely NSGAb°DR1-derived IPEX Tregs functionally behave in comparison with human-derived IPEX Tregs. Can NSGAb°DR1 Tregs express similar amounts of inhibitory cytokines (interleukin [IL]-35, IL-10, and transforming growth factor b) and inhibitory molecules (programmed death ligand 1 and cytotoxic T-lymphocyte–associated protein 4)?9,10 Thus, NSGAb°DR1 mice, alone or in combination with additional HLA molecules and/or other human cytokines, will enable in-depth analyses of developmental and functional defects that underlie immunodeficiencies and autoimmune disorders, and much-needed evaluations of emerging immunotherapies in a preclinical translational animal model. Conflict-of-interest disclosure: The authors declare no competing financial interests. n BLOOD, 18 JUNE 2015 x VOLUME 125, NUMBER 25
REFERENCES
6. Verbsky JW, Chatila TA. Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) and IPEX-related disorders: an evolving web of heritable autoimmune diseases. Curr Opin Pediatr. 2013;25(6): 708-714.
1. Goettel JA, Biswas S, Lexmond WS, et al. Fatal autoimmunity in mice reconstituted with human hematopoietic stem cells encoding defective FOXP3. Blood. 2015;125(25):3886-3895. 2. Brehm MA, Jouvet N, Greiner DL, Shultz LD. Humanized mice for the study of infectious diseases. Curr Opin Immunol. 2013;25(4):428-435. 3. Keane TM, Goodstadt L, Danecek P, et al. Mouse genomic variation and its effect on phenotypes and gene regulation. Nature. 2011;477(7364):289-294. 4. Bissig KD, Wieland SF, Tran P, et al. Human liver chimeric mice provide a model for hepatitis B and C virus infection and treatment. J Clin Invest. 2010;120(3): 924-930. 5. Turnbaugh PJ, Ridaura VK, Faith JJ, et al. The effect of diet on the human gut microbiome: a metagenomic analysis in humanized gnotobiotic mice. Sci Transl Med. 2009;1(6):6ra14.
7. Ramsdell F, Ziegler SF. FOXP3 and scurfy: how it all began. Nat Rev Immunol. 2014;14(5):343-349. 8. Touitou I, Galeotti C, Rossi-Semerano L, Hentgen V, Piram M, Kon´e-Paut I; CeR´eMAI, French reference center for autoinflammatory diseases. The expanding spectrum of rare monogenic autoinflammatory diseases. Orphanet J Rare Dis. 2013;8:162. 9. Nguyen LT, Ohashi PS. Clinical blockade of PD1 and LAG3—potential mechanisms of action. Nat Rev Immunol. 2015;15(1):45-56. 10. Vignali DA, Collison LW, Workman CJ. How regulatory T cells work. Nat Rev Immunol. 2008;8(7):523-532. © 2015 by The American Society of Hematology
l l l IMMUNOBIOLOGY
Comment on Schwartz et al, page 3896
Mechanism of enhanced eosinophil survival in inflammation ----------------------------------------------------------------------------------------------------Nives Zimmermann and Marc E. Rothenberg
CINCINNATI CHILDREN’S HOSPITAL MEDICAL CENTER
In this issue of Blood, Schwartz et al identify the nuclear factor (NF)-kB/Bcl-xL pathway as critical for regulation of eosinophil survival in inflammatory conditions.1
C
ell death is essential for many physiologic processes, and its dysregulation characterizes numerous human diseases. This is especially true for eosinophils, because these cells do not undergo substantial extramedullary hematopoiesis, yet their tissue levels can be markedly and selectively increased by a combination of recruitment from the blood and regulation of their cell death within tissues. The field of eosinophil survival has come a long way since the original finding that human blood–derived eosinophils had prolonged survival (.14 days) when cocultured with endothelial cells and with the subsequent identification of interleukin (IL)-5, granulocyte macrophage–colony-stimulating factor (GM-CSF), and IL-3 as key eosinophil survival factors.2 It is now appreciated that these eosinophil-directed hematopoietins inhibit eosinophil apoptosis and that IL-5 has a special capacity to promote eosinophil development, which has led to several therapeutics that lower eosinophils by
blocking IL-5, such as mepolizumab and reslizumab, which are at advanced stages of development.3 However, whether eosinophils are better “dead or alive” is still an unresolved question. Simplistically, both living and dead eosinophils can lead to positive and negative outcomes (see figure). For instance, eosinophils are homeostatically present in some tissues with no ill effect (figure panel A, upper left)4 and have been shown to contribute to a variety of homeostatic functions, including production of secretory immunoglobulin A in the intestine.5 Yet during many eosinophilic inflammatory diseases, eosinophils display an activated phenotype and likely induce tissue damage (figure panel A, upper right).6 Eosinophil cell death was previously viewed dichotomously, as either noninflammatory apoptotic (figure panel A, lower left) or proinflammatory necrotic (figure panel A, lower right) cell death, but recent studies have identified additional cell death pathways, including regulated necrosis,
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From www.bloodjournal.org by guest on September 11, 2016. For personal use only.
2015 125: 3829-3831 doi:10.1182/blood-2015-04-639435
Daring to learn from humanized mice Silke Paust and Matthew Bettini
Updated information and services can be found at: http://www.bloodjournal.org/content/125/25/3829.full.html Articles on similar topics can be found in the following Blood collections Free Research Articles (4041 articles) Information about reproducing this article in parts or in its entirety may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#repub_requests Information about ordering reprints may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#reprints Information about subscriptions and ASH membership may be found online at: http://www.bloodjournal.org/site/subscriptions/index.xhtml
Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly by the American Society of Hematology, 2021 L St, NW, Suite 900, Washington DC 20036. Copyright 2011 by The American Society of Hematology; all rights reserved.
