FOXO transcription factors throughout T cell biology

Abstract | The outcome of an infection with any given pathogen varies according to the dosage and route of infection, but, in addition, the physiological state of the host can determine the efficacy of clearance, the severity of infection and the extent of immunopathology. Here we propose that the forkhead box O (FOXO) transcription factor family -which is central to the integration of growth factor signalling, oxidative stress and inflammation -provides connections between physical well-being and the form and magnitude of an immune response. We present a case that FOXO transcription factors guide T cell differentiation and function in a context-driven manner, and might provide a link between metabolism and immunity

Identification of pre-leukaemic haematopoietic stem cells in acute leukaemia.

In acute myeloid leukaemia (AML), the cell of origin, nature and biological consequences of initiating lesions, and order of subsequent mutations remain poorly understood, as AML is typically diagnosed without observation of a pre-leukaemic phase. Here, highly purified haematopoietic stem cells (HSCs), progenitor and mature cell fractions from the blood of AML patients were found to contain recurrent DNMT3A mutations (DNMT3A(mut)) at high allele frequency, but without coincident NPM1 mutations (NPM1c) present in AML blasts. DNMT3A(mut)-bearing HSCs showed a multilineage repopulation advantage over non-mutated HSCs in xenografts, establishing their identity as pre-leukaemic HSCs. Pre-leukaemic HSCs were found in remission samples, indicating that they survive chemotherapy. Therefore DNMT3A(mut) arises early in AML evolution, probably in HSCs, leading to a clonally expanded pool of pre-leukaemic HSCs from which AML evolves. Our findings provide a paradigm for the detection and treatment of pre-leukaemic clones before the acquisition of additional genetic lesions engenders greater therapeutic resistance

S-2-hydroxyglutarate regulates CD8+ T-lymphocyte fate.

R-2-hydroxyglutarate accumulates to millimolar levels in cancer cells with gain-of-function isocitrate dehydrogenase 1/2 mutations. These levels of R-2-hydroxyglutarate affect 2-oxoglutarate-dependent dioxygenases. Both metabolite enantiomers, R- and S-2-hydroxyglutarate, are detectible in healthy individuals, yet their physiological function remains elusive. Here we show that 2-hydroxyglutarate accumulates in mouse CD8+ T cells in response to T-cell receptor triggering, and accumulates to millimolar levels in physiological oxygen conditions through a hypoxia-inducible factor 1-alpha (HIF-1α)-dependent mechanism. S-2-hydroxyglutarate predominates over R-2-hydroxyglutarate in activated T cells, and we demonstrate alterations in markers of CD8+ T-cell differentiation in response to this metabolite. Modulation of histone and DNA demethylation, as well as HIF-1α stability, mediate these effects. S-2-hydroxyglutarate treatment greatly enhances the in vivo proliferation, persistence and anti-tumour capacity of adoptively transferred CD8+ T cells. Thus, S-2-hydroxyglutarate acts as an immunometabolite that links environmental context, through a metabolic-epigenetic axis, to immune fate and function

G-CSF/anti-G-CSF antibody complexes drive the potent recovery and expansion of CD11b+Gr-1+ myeloid cells without compromising CD8+ T cell immune responses

BACKGROUND: Administration of recombinant G-CSF following cytoreductive therapy enhances the recovery of myeloid cells, minimizing the risk of opportunistic infection. Free G-CSF, however, is expensive, exhibits a short half-life, and has poor biological activity in vivo. METHODS: We evaluated whether the biological activity of G-CSF could be improved by pre-association with anti-G-CSF mAb prior to injection into mice. RESULTS: We find that the efficacy of G-CSF therapy can be enhanced more than 100-fold by pre-association of G-CSF with an anti-G-CSF monoclonal antibody (mAb). Compared with G-CSF alone, administration of G-CSF/anti-G-CSF mAb complexes induced the potent expansion of CD11b(+)Gr-1(+) myeloid cells in mice with or without concomitant cytoreductive treatment including radiation or chemotherapy. Despite driving the dramatic expansion of myeloid cells, in vivo antigen-specific CD8(+) T cell immune responses were not compromised. Furthermore, injection of G-CSF/anti-G-CSF mAb complexes heightened protective immunity to bacterial infection. As a measure of clinical value, we also found that antibody complexes improved G-CSF biological activity much more significantly than pegylation. CONCLUSIONS: Our findings provide the first evidence that antibody cytokine complexes can effectively expand myeloid cells, and furthermore, that G-CSF/anti-G-CSF mAb complexes may provide an improved method for the administration of recombinant G-CSF

Identification of regulatory variants associated with genetic susceptibility to meningococcal disease.

Non-coding genetic variants play an important role in driving susceptibility to complex diseases but their characterization remains challenging. Here, we employed a novel approach to interrogate the genetic risk of such polymorphisms in a more systematic way by targeting specific regulatory regions relevant for the phenotype studied. We applied this method to meningococcal disease susceptibility, using the DNA binding pattern of RELA - a NF-kB subunit, master regulator of the response to infection - under bacterial stimuli in nasopharyngeal epithelial cells. We designed a custom panel to cover these RELA binding sites and used it for targeted sequencing in cases and controls. Variant calling and association analysis were performed followed by validation of candidate polymorphisms by genotyping in three independent cohorts. We identified two new polymorphisms, rs4823231 and rs11913168, showing signs of association with meningococcal disease susceptibility. In addition, using our genomic data as well as publicly available resources, we found evidences for these SNPs to have potential regulatory effects on ATXN10 and LIF genes respectively. The variants and related candidate genes are relevant for infectious diseases and may have important contribution for meningococcal disease pathology. Finally, we described a novel genetic association approach that could be applied to other phenotypes

Plasma lipid profiles discriminate bacterial from viral infection in febrile children

Fever is the most common reason that children present to Emergency Departments. Clinical signs and symptoms suggestive of bacterial infection ar

Role of myeloid Hypoxia-Inducible Factor-1alpha in the tumor microenvironment

Solid tumors are frequently infiltrated by large numbers of non-cancerous hematopoietic cells, including macrophages. The pro-tumor or anti-tumor role of macrophages in the tumor microenvironment is unclear. Tumors are also characterized by regions of low oxygen tension. Mammalian cells respond transcriptionally to low oxygen tension via the Hypoxia-Inducible Factor-1alpha, which upregulates genes involved in glycolysis, angiogenesis, and cell survival. Previous work has shown that myeloid cells deficient for HIF-1alpha are impaired in their inflammatory responses. To genetically test the role of the myeloid hypoxic response mediated by HIF- 1alpha in the tumor microenvironment, we used the loxP/ tissue specifc cre recombianse to generate murine myeloid specific deletion of HIF-1alpha in rodents with an established transgenic model of breast cancer, MMTV-PyMT. Lack of HIF-1alpha in the tumor microenvironment resulted in tumors with less mass, but with increased cell death. Enzyme activities of iNOS and ArgI were reduced in whole tumor lysates. In vitro experiments demonstrated HIF-1 alpha, not HIF-2alpha, control of L-arginine degrading enzymes iNOS and arginase I. Further characterization of the relationship between macrophages and tumor cells using co-culture strategies revealed that tumor cells induce ArgI in a HIF-1alpha and hypoxia dependent fashion at the protein level. iNOS was detected at the RNA level after co -culture with MECs, but was scarcely detectable at the protein level. ArgI and iNOS have been implicated in T cell immunosuppression. PyMT tumor bearing mice displayed evidence of T cell activation, yet T cells isolated from myeloid HIF WT tumors were less responsive than those from myeloid HIF KO tumors after stimulation with CD3/28 ex vivo. We propose myeloid HIF-1alpha contributes to local tumor immunosuppression of T cell function. This suggests inhibition of HIF-1 may have beneficial effects not only by blocking tumor growth and survival under hypoxia, but also by relieving myeloid cell mediated immunosuppression in the tumor microenvironmen

Role of myeloid Hypoxia-Inducible Factor-1alpha in the tumor microenvironment

Solid tumors are frequently infiltrated by large numbers of non-cancerous hematopoietic cells, including macrophages. The pro-tumor or anti-tumor role of macrophages in the tumor microenvironment is unclear. Tumors are also characterized by regions of low oxygen tension. Mammalian cells respond transcriptionally to low oxygen tension via the Hypoxia-Inducible Factor-1alpha, which upregulates genes involved in glycolysis, angiogenesis, and cell survival. Previous work has shown that myeloid cells deficient for HIF-1alpha are impaired in their inflammatory responses. To genetically test the role of the myeloid hypoxic response mediated by HIF- 1alpha in the tumor microenvironment, we used the loxP/ tissue specifc cre recombianse to generate murine myeloid specific deletion of HIF-1alpha in rodents with an established transgenic model of breast cancer, MMTV-PyMT. Lack of HIF-1alpha in the tumor microenvironment resulted in tumors with less mass, but with increased cell death. Enzyme activities of iNOS and ArgI were reduced in whole tumor lysates. In vitro experiments demonstrated HIF-1 alpha, not HIF-2alpha, control of L-arginine degrading enzymes iNOS and arginase I. Further characterization of the relationship between macrophages and tumor cells using co-culture strategies revealed that tumor cells induce ArgI in a HIF-1alpha and hypoxia dependent fashion at the protein level. iNOS was detected at the RNA level after co -culture with MECs, but was scarcely detectable at the protein level. ArgI and iNOS have been implicated in T cell immunosuppression. PyMT tumor bearing mice displayed evidence of T cell activation, yet T cells isolated from myeloid HIF WT tumors were less responsive than those from myeloid HIF KO tumors after stimulation with CD3/28 ex vivo. We propose myeloid HIF-1alpha contributes to local tumor immunosuppression of T cell function. This suggests inhibition of HIF-1 may have beneficial effects not only by blocking tumor growth and survival under hypoxia, but also by relieving myeloid cell mediated immunosuppression in the tumor microenvironmen

FOXO1 opposition of CD8+ T cell effector programming confers early memory properties and phenotypic diversity.

The factors and steps controlling postinfection CD8+ T cell terminal effector versus memory differentiation are incompletely understood. Whereas we found that naive TCF7 (alias "Tcf-1") expression is FOXO1 independent, early postinfection we report bimodal, FOXO1-dependent expression of the memory-essential transcription factor TCF7 in pathogen-specific CD8+ T cells. We determined the early postinfection TCF7high population is marked by low TIM3 expression and bears memory signature hallmarks before the appearance of established memory precursor marker CD127 (IL-7R). These cells exhibit diminished TBET, GZMB, mTOR signaling, and cell cycle progression. Day 5 postinfection, TCF7high cells express higher memory-associated BCL2 and EOMES, as well as increased accumulation potential and capacity to differentiate into memory phenotype cells. TCF7 retroviral transduction opposes GZMB expression and the formation of KLRG1pos phenotype cells, demonstrating an active role for TCF7 in extinguishing the effector program and forestalling terminal differentiation. Past the peak of the cellular immune response, we report a gradient of FOXO1 and TCF7 expression, which functions to oppose TBET and orchestrate a continuum of effector-to-memory phenotypes