Publications by authors named "Sarah Pagan"

13 Publications

  • Page 1 of 1

Differential IRF8 Transcription Factor Requirement Defines Two Pathways of Dendritic Cell Development in Humans.

Immunity 2020 08 30;53(2):353-370.e8. Epub 2020 Jul 30.

Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne NE2 4HH, UK; Northern Centre for Cancer Care, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne NE7 7DN, UK. Electronic address:

The formation of mammalian dendritic cells (DCs) is controlled by multiple hematopoietic transcription factors, including IRF8. Loss of IRF8 exerts a differential effect on DC subsets, including plasmacytoid DCs (pDCs) and the classical DC lineages cDC1 and cDC2. In humans, cDC2-related subsets have been described including AXLSIGLEC6 pre-DC, DC2 and DC3. The origin of this heterogeneity is unknown. Using high-dimensional analysis, in vitro differentiation, and an allelic series of human IRF8 deficiency, we demonstrated that cDC2 (CD1cDC) heterogeneity originates from two distinct pathways of development. The lymphoid-primed IRF8 pathway, marked by CD123 and BTLA, carried pDC, cDC1, and DC2 trajectories, while the common myeloid IRF8 pathway, expressing SIRPA, formed DC3s and monocytes. We traced distinct trajectories through the granulocyte-macrophage progenitor (GMP) compartment showing that AXLSIGLEC6 pre-DCs mapped exclusively to the DC2 pathway. In keeping with their lower requirement for IRF8, DC3s expand to replace DC2s in human partial IRF8 deficiency.
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http://dx.doi.org/10.1016/j.immuni.2020.07.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7447982PMC
August 2020

Donor monocyte-derived macrophages promote human acute graft-versus-host disease.

J Clin Invest 2020 09;130(9):4574-4586

Human Dendritic Cell Laboratory, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom.

Myelopoiesis is invariably present and contributes to pathology in animal models of graft-versus-host disease (GVHD). In humans, a rich inflammatory infiltrate bearing macrophage markers has also been described in histological studies. In order to determine the origin, functional properties, and role in pathogenesis of these cells, we isolated single-cell suspensions from acute cutaneous GVHD and subjected them to genotype, transcriptome, and in vitro functional analysis. A donor-derived population of CD11c+CD14+ cells was the dominant population of all leukocytes in GVHD. Surface phenotype and NanoString gene expression profiling indicated the closest steady-state counterpart of these cells to be monocyte-derived macrophages. In GVHD, however, there was upregulation of monocyte antigens SIRPα and S100A8/9 transcripts associated with leukocyte trafficking, pattern recognition, antigen presentation, and costimulation. Isolated GVHD macrophages stimulated greater proliferation and activation of allogeneic T cells and secreted higher levels of inflammatory cytokines than their steady-state counterparts. In HLA-matched mixed leukocyte reactions, we also observed differentiation of activated macrophages with a similar phenotype. These exhibited cytopathicity to a keratinocyte cell line and mediated pathological damage to skin explants independently of T cells. Together, these results define the origin, functional properties, and potential pathogenic roles of human GVHD macrophages.
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http://dx.doi.org/10.1172/JCI133909DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7456218PMC
September 2020

Serum Flt3 ligand is a biomarker of progenitor cell mass and prognosis in acute myeloid leukemia.

Blood Adv 2019 10;3(20):3052-3061

Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom.

Fms-like tyrosine kinase 3 (Flt3) is expressed on progenitor cells and acute myeloid leukemia (AML) blasts. Fms-like tyrosine kinase 3 ligand (Flt3L) is detectable during homeostasis and increases in hypoplasia due to genetic defects or treatment with cytoreductive agents. Conversely, Flt3+ AML is associated with depletion of Flt3L to undetectable levels. After induction chemotherapy, Flt3L is restored in patients entering complete remission (CR) but remains depressed in those with refractory disease. Weekly sampling reveals marked differences in the kinetics of Flt3L response during the first 6 weeks of treatment, proportionate to the clearance of blasts and cellularity of the bone marrow. In the UK NCRI AML17 trial, Flt3L was measured at day 26 in a subgroup of 140 patients with Flt3 mutation randomized to the tyrosine kinase inhibitor lestaurtinib or placebo. In these patients, attainment of CR was associated with higher Flt3L at day 26 (Mann-Whitney UP < .0001). Day 26 Flt3L was also associated with survival; Flt3L ≤291 pg/mL was associated with inferior event-free survival (EFS), and Flt3L >1185 pg/mL was associated with higher overall survival (OS; P = .0119). The separation of EFS and OS curves increased when minimal residual disease (MRD) status was combined with Flt3L measurement, and Flt3L retained a near-significant association with survival after adjusting for MRD in a proportional hazards model. Serial measurement of Flt3L in patients who had received a hematopoietic stem cell transplant for AML illustrates the potential value of monitoring Flt3L to identify relapse. Measurement of Flt3L is a noninvasive test with the potential to inform clinical decisions in patients with AML.
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http://dx.doi.org/10.1182/bloodadvances.2019000197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6849950PMC
October 2019

Early Molecular Stratification of High-risk Primary Biliary Cholangitis.

EBioMedicine 2016 Dec 21;14:65-73. Epub 2016 Nov 21.

Institute of Cellular Medicine, Medical School, Newcastle University, Newcastle upon Tyne, UK. Electronic address:

High-risk primary biliary cholangitis (PBC), defined by inadequate response at one year to Ursodeoxycholic acid (UDCA), is associated with disease progression and liver transplantation. Stratifying high-risk patients early would facilitate improved approaches to care. Using long-term follow-up data to define risk at presentation, 6 high-risk PBC patients and 8 low-risk patients were identified from biopsy, transplant and biochemical archival records. Formalin-fixed paraffin-embedded (FFPE) liver biopsies taken at presentation were graded (Scheuer and Nakanuma scoring) and gene expression analysed using the NanoString® nCounter PanCancer Immunity 770-gene panel. Principle component analysis (PCA) demonstrated discrete gene expression clustering between controls and high- and low-risk PBC. High-risk PBC was characterised by up-regulation of genes linked to T-cell activation and apoptosis, INF-γ signalling and leukocyte migration and down-regulation of those linked to the complement pathway. CDKN1a, up-regulated in high-risk PBC, correlated with significantly increased expression of its gene product, the senescence marker p21, by biliary epithelial cells. Our findings suggest high- and low-risk PBC are biologically different from disease outset and senescence an early feature in high-risk disease. Identification of a high-risk 'signal' early from standard FFPE tissue sections has clear clinical utility allowing for patient stratification and second-line therapeutic intervention.
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http://dx.doi.org/10.1016/j.ebiom.2016.11.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5161439PMC
December 2016

Langerin-expressing dendritic cells in human tissues are related to CD1c+ dendritic cells and distinct from Langerhans cells and CD141high XCR1+ dendritic cells.

J Leukoc Biol 2015 Apr 16;97(4):627-34. Epub 2014 Dec 16.

Human Dendritic Cell Laboratory, Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom

Langerin is a C-type lectin expressed at high level by LCs of the epidermis. Langerin is also expressed by CD8(+)/CD103(+) XCR1(+) cross-presenting DCs of mice but is not found on the homologous human CD141(high) XCR1(+) myeloid DC. Here, we show that langerin is expressed at a low level on DCs isolated from dermis, lung, liver, and lymphoid tissue and that langerin(+) DCs are closely related to CD1c(+) myeloid DCs. They are distinguishable from LCs by the level of expression of CD1a, EpCAM, CD11b, CD11c, CD13, and CD33 and are found in tissues and tissue-draining LNs devoid of LCs. They are unrelated to CD141(high) XCR1(+) myeloid DCs, lacking the characteristic expression profile of cross-presenting DCs, conserved between mammalian species. Stem cell transplantation and DC deficiency models confirm that dermal langerin(+) DCs have an independent homeostasis to LCs. Langerin is not expressed by freshly isolated CD1c(+) blood DCs but is rapidly induced on CD1c(+) DCs by serum or TGF-β via an ALK-3-dependent pathway. These results show that langerin is expressed outside of the LC compartment of humans and highlight a species difference: langerin is expressed by the XCR1(+) "DC1" population of mice but is restricted to the CD1c(+) "DC2" population of humans (homologous to CD11b(+) DCs in the mouse).
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http://dx.doi.org/10.1189/jlb.1HI0714-351RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370053PMC
April 2015

Human dermal CD14⁺ cells are a transient population of monocyte-derived macrophages.

Immunity 2014 Sep 4;41(3):465-477. Epub 2014 Sep 4.

Institute of Cellular Medicine, The Medical School, Newcastle University, Framlington Place, Newcastle upon Tyne NE2 4HH, UK. Electronic address:

Dendritic cells (DCs), monocytes, and macrophages are leukocytes with critical roles in immunity and tolerance. The DC network is evolutionarily conserved; the homologs of human tissue CD141(hi)XCR1⁺ CLEC9A⁺ DCs and CD1c⁺ DCs are murine CD103⁺ DCs and CD64⁻ CD11b⁺ DCs. In addition, human tissues also contain CD14⁺ cells, currently designated as DCs, with an as-yet unknown murine counterpart. Here we have demonstrated that human dermal CD14⁺ cells are a tissue-resident population of monocyte-derived macrophages with a short half-life of <6 days. The decline and reconstitution kinetics of human blood CD14⁺ monocytes and dermal CD14⁺ cells in vivo supported their precursor-progeny relationship. The murine homologs of human dermal CD14⁺ cells are CD11b⁺ CD64⁺ monocyte-derived macrophages. Human and mouse monocytes and macrophages were defined by highly conserved gene transcripts, which were distinct from DCs. The demonstration of monocyte-derived macrophages in the steady state in human tissue supports a conserved organization of human and mouse mononuclear phagocyte system.
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http://dx.doi.org/10.1016/j.immuni.2014.08.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4175180PMC
September 2014

Rapid detection of dendritic cell and monocyte disorders using CD4 as a lineage marker of the human peripheral blood antigen-presenting cell compartment.

Front Immunol 2013 27;4:495. Epub 2013 Dec 27.

Human Dendritic Cell Laboratory, Institute of Cellular Medicine, Newcastle University , Newcastle upon Tyne , UK.

Dendritic cells (DCs) and monocytes are critical regulators and effectors of innate and adaptive immune responses. Monocyte expansion has been described in many pathological states while monocyte and DC deficiency syndromes are relatively recent additions to the catalog of human primary immunodeficiency disorders. Clinically applicable screening tests to diagnose and monitor these conditions are lacking. Conventional strategies for identifying human DCs and monocytes have been based on the use of a lineage gate to exclude lymphocytes, thus preventing simultaneous detection of DCs, monocytes, and lymphocyte subsets. Here we demonstrate that CD4 is a reliable lineage marker for the human peripheral blood antigen-presenting cell compartment that can be used to identify DCs and monocytes in parallel with lymphocytes. Based on this principle, simple modification of a standard lymphocyte phenotyping assay permits simultaneous enumeration of four lymphocyte and five DC/monocyte populations from a single sample. This approach is applicable to clinical samples and facilitates the diagnosis of DC and monocyte disorders in a wide range of clinical settings, including genetic deficiency, neoplasia, and inflammation.
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http://dx.doi.org/10.3389/fimmu.2013.00495DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3873601PMC
January 2014

The evolution of cellular deficiency in GATA2 mutation.

Blood 2014 Feb 17;123(6):863-74. Epub 2013 Dec 17.

Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom;

Constitutive heterozygous GATA2 mutation is associated with deafness, lymphedema, mononuclear cytopenias, infection, myelodysplasia (MDS), and acute myeloid leukemia. In this study, we describe a cross-sectional analysis of 24 patients and 6 relatives with 14 different frameshift or substitution mutations of GATA2. A pattern of dendritic cell, monocyte, B, and natural killer (NK) lymphoid deficiency (DCML deficiency) with elevated Fms-like tyrosine kinase 3 ligand (Flt3L) was observed in all 20 patients phenotyped, including patients with Emberger syndrome, monocytopenia with Mycobacterium avium complex (MonoMAC), and MDS. Four unaffected relatives had a normal phenotype indicating that cellular deficiency may evolve over time or is incompletely penetrant, while 2 developed subclinical cytopenias or elevated Flt3L. Patients with GATA2 mutation maintained higher hemoglobin, neutrophils, and platelets and were younger than controls with acquired MDS and wild-type GATA2. Frameshift mutations were associated with earlier age of clinical presentation than substitution mutations. Elevated Flt3L, loss of bone marrow progenitors, and clonal myelopoiesis were early signs of disease evolution. Clinical progression was associated with increasingly elevated Flt3L, depletion of transitional B cells, CD56(bright) NK cells, naïve T cells, and accumulation of terminally differentiated NK and CD8(+) memory T cells. These studies provide a framework for clinical and laboratory monitoring of patients with GATA2 mutation and may inform therapeutic decision-making.
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http://dx.doi.org/10.1182/blood-2013-07-517151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3916878PMC
February 2014

Human tissues contain CD141hi cross-presenting dendritic cells with functional homology to mouse CD103+ nonlymphoid dendritic cells.

Immunity 2012 Jul 12;37(1):60-73. Epub 2012 Jul 12.

Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK.

Dendritic cell (DC)-mediated cross-presentation of exogenous antigens acquired in the periphery is critical for the initiation of CD8(+) T cell responses. Several DC subsets are described in human tissues but migratory cross-presenting DCs have not been isolated, despite their potential importance in immunity to pathogens, vaccines, and tumors and tolerance to self. Here, we identified a CD141(hi) DC present in human interstitial dermis, liver, and lung that was distinct from the majority of CD1c(+) and CD14(+) tissue DCs and superior at cross-presenting soluble antigens. Cutaneous CD141(hi) DCs were closely related to blood CD141(+) DCs, and migratory counterparts were found among skin-draining lymph node DCs. Comparative transcriptomic analysis with mouse showed tissue DC subsets to be conserved between species and permitted close alignment of human and mouse DC subsets. These studies inform the rational design of targeted immunotherapies and facilitate translation of mouse functional DC biology to the human setting.
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http://dx.doi.org/10.1016/j.immuni.2012.04.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3476529PMC
July 2012

Sensitizing primary acute lymphoblastic leukemia to natural killer cell recognition by induction of NKG2D ligands.

Leuk Lymphoma 2013 Jan 8;54(1):167-73. Epub 2012 Sep 8.

Institute of Cellular Medicine, Newcastle University, Newcastle upon Tyne, UK.

Natural killer (NK) cell immunosurveillance may be impaired by malignant disease, resulting in tumor escape and disease progression. Therapies that enhance NK cytotoxicity may therefore prove valuable in remission-induction and maintenance treatment regimens. Acute lymphoblastic leukemia (ALL) has previously been considered resistant to NK cell lysis and not tractable to this approach. Our study demonstrates that bortezomib, valproate and troglitazone can up-regulate NK activating ligands on a B-ALL cell line and on a proportion but not all adult primary B-ALL samples. Drug-treated ALL cells trigger higher levels of NK degranulation, as measured by CD107a expression, and this effect is dependent on signaling through the NK activating receptor NKG2D. These results suggest that bortezomib, valproate and troglitazone may have clinical utility in sensitizing ALL to NK mediated lysis in vivo.
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http://dx.doi.org/10.3109/10428194.2012.708026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6588533PMC
January 2013

Exome sequencing identifies GATA-2 mutation as the cause of dendritic cell, monocyte, B and NK lymphoid deficiency.

Blood 2011 Sep 15;118(10):2656-8. Epub 2011 Jul 15.

Institute of Cellular Medicine, International Centre for Life, Newcastle University, Newcastle upon Tyne, United Kingdom.

The human syndrome of dendritic cell, monocyte, B and natural killer lymphoid deficiency presents as a sporadic or autosomal dominant trait causing susceptibility to mycobacterial and other infections, predisposition to myelodysplasia and leukemia, and, in some cases, pulmonary alveolar proteinosis. Seeking a genetic cause, we sequenced the exomes of 4 unrelated persons, 3 with sporadic disease, looking for novel, heterozygous, and probably deleterious variants. A number of genes harbored novel variants in person, but only one gene, GATA2, was mutated in all 4 persons. Each person harbored a different mutation, but all were predicted to be highly deleterious and to cause loss or mutation of the C-terminal zinc finger domain. Because GATA2 is the only common mutated gene in 4 unrelated persons, it is highly probable to be the cause of dendritic cell, monocyte, B, and natural killer lymphoid deficiency. This disorder therefore constitutes a new genetic form of heritable immunodeficiency and leukemic transformation.
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http://dx.doi.org/10.1182/blood-2011-06-360313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5137783PMC
September 2011

The human syndrome of dendritic cell, monocyte, B and NK lymphoid deficiency.

J Exp Med 2011 Feb 17;208(2):227-34. Epub 2011 Jan 17.

Institute of Cellular Medicine, Newcastle University, Framlington Place, Newcastle upon Tyne, England, UK.

Congenital or acquired cellular deficiencies in humans have the potential to reveal much about normal hematopoiesis and immune function. We show that a recently described syndrome of monocytopenia, B and NK lymphoid deficiency additionally includes the near absence of dendritic cells. Four subjects showed severe depletion of the peripheral blood HLA-DR(+) lineage(-) compartment, with virtually no CD123(+) or CD11c(+) dendritic cells (DCs) and very few CD14(+) or CD16(+) monocytes. The only remaining HLA-DR(+) lineage(-) cells were circulating CD34(+) progenitor cells. Dermal CD14(+) and CD1a(+) DC were also absent, consistent with their dependence on blood-derived precursors. In contrast, epidermal Langerhans cells and tissue macrophages were largely preserved. Combined loss of peripheral DCs, monocytes, and B and NK lymphocytes was mirrored in the bone marrow by complete absence of multilymphoid progenitors and depletion of granulocyte-macrophage progenitors. Depletion of the HLA-DR(+) peripheral blood compartment was associated with elevated serum fms-like tyrosine kinase ligand and reduced circulating CD4(+)CD25(hi)FoxP3(+) T cells, supporting a role for DC in T reg cell homeostasis.
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http://dx.doi.org/10.1084/jem.20101459DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3039861PMC
February 2011
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