Publications by authors named "Stephanie K Dougan"

67 Publications

Radiation combines with immune checkpoint blockade to enhance T cell priming in a murine model of poorly immunogenic pancreatic cancer.

Open Biol 2021 11 17;11(11):210245. Epub 2021 Nov 17.

Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA 02215, USA.

Radiation has been a pillar of cancer therapy for decades. The effects of radiation on the anti-tumour immune response are variable across studies and have not been explicitly defined in poorly immunogenic tumour types. Here, we employed combination checkpoint blockade immunotherapy with stereotactic body radiation therapy and examined the effect on tumour growth and immune infiltrates in subcutaneous and orthotopic mouse models of pancreatic cancer. Although immune checkpoint blockade and radiation were ineffective alone, their combination produced a modest growth delay in both irradiated and non-irradiated tumours that corresponded with significant increases in CD8+ T cells, CD4+ T cells and tumour-specific T cells as identified by IFNγ ELISpot. We conclude that radiation enhances priming of tumour-specific T cells in poorly immunogenic tumours and that the frequency of these T cells can be further increased by combination with immune checkpoint blockade.
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http://dx.doi.org/10.1098/rsob.210245DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8595997PMC
November 2021

Eosinophils in Health and Disease: A State-of-the-Art Review.

Mayo Clin Proc 2021 10 15;96(10):2694-2707. Epub 2021 Sep 15.

Division of Rheumatology, Department of Medicine, and Division of Clinical Epidemiology, Department of Biostatistics, Epidemiology, and Informatics, University of Pennsylvania, Philadelphia.

Eosinophils play a homeostatic role in the body's immune responses. These cells are involved in combating some parasitic, bacterial, and viral infections and certain cancers and have pathologic roles in diseases including asthma, chronic rhinosinusitis with nasal polyps, eosinophilic gastrointestinal disorders, and hypereosinophilic syndromes. Treatment of eosinophilic diseases has traditionally been through nonspecific eosinophil attenuation by use of glucocorticoids. However, several novel biologic therapies targeting eosinophil maturation factors, such as interleukin (IL)-5 and the IL-5 receptor or IL-4/IL-13, have recently been approved for clinical use. Despite the success of biologic therapies, some patients with eosinophilic inflammatory disease may not achieve adequate symptom control, underlining the need to further investigate the contribution of patient characteristics, such as comorbidities and other processes, in driving ongoing disease activity. New research has shown that eosinophils are also involved in several homeostatic processes, including metabolism, tissue remodeling and development, neuronal regulation, epithelial and microbiome regulation, and immunoregulation, indicating that these cells may play a crucial role in metabolic regulation and organ function in healthy humans. Consequently, further investigation is needed into the homeostatic roles of eosinophils and eosinophil-mediated processes across different tissues and their varied microenvironments. Such work may provide important insights into the role of eosinophils not only under disease conditions but also in health. This narrative review synthesizes relevant publications retrieved from PubMed informed by author expertise to provide new insights into the diverse roles of eosinophils in health and disease, with particular emphasis on the implications for current and future development of eosinophil-targeted therapies.
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http://dx.doi.org/10.1016/j.mayocp.2021.04.025DOI Listing
October 2021

Type 2 immunity is maintained during cancer-associated adipose tissue wasting.

Immunother Adv 2021 Jan 2;1(1):ltab011. Epub 2021 Jun 2.

Department of Cancer Immunology and Virology, Dana Farber Cancer Institute, Boston, MA, USA.

Objectives: Cachexia is a systemic metabolic disorder characterized by loss of fat and muscle mass, which disproportionately impacts patients with gastrointestinal malignancies such as pancreatic cancer. While the immunologic shifts contributing to the development of other adipose tissue (AT) pathologies such as obesity have been well described, the immune microenvironment has not been studied in the context of cachexia.

Methods: We performed bulk RNA-sequencing, cytokine arrays, and flow cytometry to characterize the immune landscape of visceral AT (VAT) in the setting of pancreatic and colorectal cancers.

Results: The cachexia inducing factor IL-6 is strongly elevated in the wasting VAT of cancer bearing mice, but the regulatory type 2 immune landscape which characterizes healthy VAT is maintained. Pathologic skewing toward Th1 and Th17 inflammation is absent. Similarly, the VAT of patients with colorectal cancer is characterized by a Th2 signature with abundant IL-33 and eotaxin-2, albeit also with high levels of IL-6.

Conclusions: Wasting AT during the development of cachexia may not undergo drastic changes in immune composition like those seen in obese AT. Our approach provides a framework for future immunologic analyses of cancer associated cachexia.
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http://dx.doi.org/10.1093/immadv/ltab011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8286632PMC
January 2021

Opportunities for Utilization of DNA Repair Inhibitors in Homologous Recombination Repair-Deficient and Proficient Pancreatic Adenocarcinoma.

Clin Cancer Res 2021 12 20;27(24):6622-6637. Epub 2021 Jul 20.

Dana-Farber Brigham and Women's Cancer Center, Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts.

Pancreatic cancer is rapidly progressive and notoriously difficult to treat with cytotoxic chemotherapy and targeted agents. Recent demonstration of the efficacy of maintenance PARP inhibition in germline mutated pancreatic cancer has raised hopes that increased understanding of the DNA damage response pathway will lead to new therapies in both homologous recombination (HR) repair-deficient and proficient pancreatic cancer. Here, we review the potential mechanisms of exploiting HR deficiency, replicative stress, and DNA damage-mediated immune activation through targeted inhibition of DNA repair regulatory proteins.
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http://dx.doi.org/10.1158/1078-0432.CCR-21-1367DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8678153PMC
December 2021

cIAP1/2 antagonism eliminates MHC class I-negative tumors through T cell-dependent reprogramming of mononuclear phagocytes.

Sci Transl Med 2021 05;13(594)

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA 02215, USA.

Loss of major histocompatibility complex (MHC) class I and interferon-γ (IFN-γ) sensing are major causes of primary and acquired resistance to checkpoint blockade immunotherapy. Thus, additional treatment options are needed for tumors that lose expression of MHC class I. The cellular inhibitor of apoptosis proteins 1 and 2 (cIAP1/2) regulate classical and alternative nuclear factor κB (NF-κB) signaling. Induction of noncanonical NF-κB signaling with cIAP1/2 antagonists mimics costimulatory signaling, augmenting antitumor immunity. We show that induction of noncanonical NF-κB signaling induces T cell-dependent immune responses, even in β-microglobulin (βM)-deficient tumors, demonstrating that direct CD8 T cell recognition of tumor cell-expressed MHC class I is not required. Instead, T cell-produced lymphotoxin reprograms both mouse and human macrophages to be tumoricidal. In wild-type mice, but not mice incapable of antigen-specific T cell responses, cIAP1/2 antagonism reduces tumor burden by increasing phagocytosis of live tumor cells. Efficacy is augmented by combination with CD47 blockade. Thus, activation of noncanonical NF-κB stimulates a T cell-macrophage axis that curtails growth of tumors that are resistant to checkpoint blockade because of loss of MHC class I or IFN-γ sensing. These findings provide a potential mechanism for controlling checkpoint blockade refractory tumors.
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http://dx.doi.org/10.1126/scitranslmed.abf5058DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8406785PMC
May 2021

Inhibition of CDK4/6 Promotes CD8 T-cell Memory Formation.

Cancer Discov 2021 Oct 3;11(10):2564-2581. Epub 2021 May 3.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts.

CDK4/6 inhibitors are approved to treat breast cancer and are in trials for other malignancies. We examined CDK4/6 inhibition in mouse and human CD8 T cells during early stages of activation. Mice receiving tumor-specific CD8 T cells treated with CDK4/6 inhibitors displayed increased T-cell persistence and immunologic memory. CDK4/6 inhibition upregulated MXD4, a negative regulator of MYC, in both mouse and human CD8 T cells. Silencing of in mouse CD8 T cells demonstrated the importance of this axis for memory formation. We used single-cell transcriptional profiling and T-cell receptor clonotype tracking to evaluate recently activated human CD8 T cells in patients with breast cancer before and during treatment with either palbociclib or abemaciclib. CDK4/6 inhibitor therapy in humans increases the frequency of CD8 memory precursors and downregulates their expression of MYC target genes, suggesting that CDK4/6 inhibitors in patients with cancer may augment long-term protective immunity. SIGNIFICANCE: CDK4/6 inhibition skews newly activated CD8 T cells toward a memory phenotype in mice and humans with breast cancer. CDK4/6 inhibitors may have broad utility outside breast cancer, particularly in the neoadjuvant setting to augment CD8 T-cell priming to tumor antigens prior to dosing with checkpoint blockade..
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http://dx.doi.org/10.1158/2159-8290.CD-20-1540DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8487897PMC
October 2021

EZH2 inhibition activates a dsRNA-STING-interferon stress axis that potentiates response to PD-1 checkpoint blockade in prostate cancer.

Nat Cancer 2021 Apr 22;2(4):444-456. Epub 2021 Mar 22.

Laboratory of Genitourinary Cancer Pathogenesis, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.

Prostate cancers are considered to be immunologically 'cold' tumors given the very few patients who respond to checkpoint inhibitor (CPI) therapy. Recently, enrichment of interferon-stimulated genes (ISGs) predicted a favorable response to CPI across various disease sites. The enhancer of zeste homolog-2 (EZH2) is overexpressed in prostate cancer and known to negatively regulate ISGs. In the present study, we demonstrate that EZH2 inhibition in prostate cancer models activates a double-stranded RNA-STING-ISG stress response upregulating genes involved in antigen presentation, Th1 chemokine signaling and interferon response, including programmed cell death protein 1 (PD-L1) that is dependent on STING activation. EZH2 inhibition substantially increased intratumoral trafficking of activated CD8 T cells and increased M1 tumor-associated macrophages, overall reversing resistance to PD-1 CPI. Our study identifies EZH2 as a potent inhibitor of antitumor immunity and responsiveness to CPI. These data suggest EZH2 inhibition as a therapeutic direction to enhance prostate cancer response to PD-1 CPI.
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http://dx.doi.org/10.1038/s43018-021-00185-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8061902PMC
April 2021

Understanding and treating the inflammatory adverse events of cancer immunotherapy.

Cell 2021 03 5;184(6):1575-1588. Epub 2021 Mar 5.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, MA 02215, USA; Department of Immunology, Harvard Medical School, 25 Shattuck Street, Boston, MA 02115, USA; Department of Neurology, Brigham & Women's Hospital and Harvard Medical School, 25 Shattuck Street, Boston, MA 02115, USA. Electronic address:

During the past decade, immunotherapies have made a major impact on the treatment of diverse types of cancer. Inflammatory toxicities are not only a major concern for Food and Drug Administration (FDA)-approved checkpoint blockade and chimeric antigen receptor (CAR) T cell therapies, but also limit the development and use of combination therapies. Fundamentally, these adverse events highlight the intricate balance of pro- and anti-inflammatory pathways that regulate protective immune responses. Here, we discuss the cellular and molecular mechanisms of inflammatory adverse events, current approaches to treatment, as well as opportunities for the design of immunotherapies that limit such inflammatory toxicities while preserving anti-tumor efficacy.
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http://dx.doi.org/10.1016/j.cell.2021.02.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979511PMC
March 2021

TGFβ: Protecting PD-1 from mRNA Decay.

Cancer Immunol Res 2020 12;8(12):1464

Dana-Farber Cancer Institute, Boston, Massachusetts.

Coordinated regulation of genes is key to determining cell fate. Although this is best understood for master regulator transcription factors, posttranscriptional regulation of mRNA stability and nuclear export can be equally effective at altering gene expression. Indeed, the heterogeneity of RNA-binding proteins and miRNAs suggests a deep complexity to posttranscriptional regulatory processes. In this issue, Wu and colleagues report a new mechanism for TGFβ-mediated immune suppression via regulation of mRNA-binding proteins in CD8 T cells..
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http://dx.doi.org/10.1158/2326-6066.CIR-20-0881DOI Listing
December 2020

Molecular Pathways of Colon Inflammation Induced by Cancer Immunotherapy.

Cell 2020 08 29;182(3):655-671.e22. Epub 2020 Jun 29.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA 02215, USA; Department of Immunology, Harvard Medical School, Boston, MA 02115, USA; Department of Neurology, Brigham & Women's Hospital and Harvard Medical School, Boston, MA 02115, USA. Electronic address:

Checkpoint blockade with antibodies specific for the PD-1 and CTLA-4 inhibitory receptors can induce durable responses in a wide range of human cancers. However, the immunological mechanisms responsible for severe inflammatory side effects remain poorly understood. Here we report a comprehensive single-cell analysis of immune cell populations in colitis, a common and severe side effect of checkpoint blockade. We observed a striking accumulation of CD8 T cells with highly cytotoxic and proliferative states and no evidence of regulatory T cell depletion. T cell receptor (TCR) sequence analysis demonstrated that a substantial fraction of colitis-associated CD8 T cells originated from tissue-resident populations, explaining the frequently early onset of colitis symptoms following treatment initiation. Our analysis also identified cytokines, chemokines, and surface receptors that could serve as therapeutic targets for colitis and potentially other inflammatory side effects of checkpoint blockade.
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http://dx.doi.org/10.1016/j.cell.2020.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415717PMC
August 2020

Neoleukin-2 enhances anti-tumour immunity downstream of peptide vaccination targeted by an anti-MHC class II VHH.

Open Biol 2020 02 5;10(2):190235. Epub 2020 Feb 5.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA.

Cancer-specific mutations can lead to peptides of unique sequence presented on MHC class I to CD8 T cells. These neoantigens can be potent tumour-rejection antigens, appear to be the driving force behind responsiveness to anti-CTLA-4 and anti-PD1/L1-based therapies and have been used to develop personalized vaccines. The platform for delivering neoantigen-based vaccines has varied, and further optimization of both platform and adjuvant will be necessary to achieve scalable vaccine products that are therapeutically effective at a reasonable cost. Here, we developed a platform for testing potential CD8 T cell tumour vaccine candidates. We used a high-affinity alpaca-derived VHH against MHC class II to deliver peptides to professional antigen-presenting cells. We show and that peptides derived from the model antigen ovalbumin are better able to activate naive ovalbumin-specific CD8 T cells when conjugated to an MHC class II-specific VHH when compared with an irrelevant control VHH. We then used the VHH-peptide platform to evaluate a panel of candidate neoantigens in a mouse model of pancreatic cancer. None of the candidate neoantigens tested led to protection from tumour challenge; however, we were able to show vaccine-induced CD8 T cell responses to a melanoma self-antigen that was augmented by combination therapy with the synthetic cytokine mimetic Neo2/15.
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http://dx.doi.org/10.1098/rsob.190235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7058936PMC
February 2020

SMAC mimetics throw a molecular switch to control T17 responses.

Sci Signal 2019 08 27;12(596). Epub 2019 Aug 27.

Division of Gastroenterology, Department of Medicine, Massachusetts General Hospital, Boston, MA 02114, USA.

IL-17 produced by T17 cells plays a central role in numerous protective and pathologic immune responses. In this issue of , Rizk define a molecular switch controlled by the cellular inhibitor of apoptosis proteins that regulates T17 immunity.
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http://dx.doi.org/10.1126/scisignal.aay3986DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7525609PMC
August 2019

Transnuclear mice reveal Peyer's patch iNKT cells that regulate B-cell class switching to IgG1.

EMBO J 2019 07 31;38(14):e101260. Epub 2019 May 31.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA.

Tissue-resident iNKT cells maintain tissue homeostasis and peripheral surveillance against pathogens; however, studying these cells is challenging due to their low abundance and poor recovery from tissues. We here show that iNKT transnuclear mice, generated by somatic cell nuclear transfer, have increased tissue resident iNKT cells. We examined expression of PLZF, T-bet, and RORγt, as well as cytokine/chemokine profiles, and found that both monoclonal and polyclonal iNKT cells differentiated into functional subsets that faithfully replicated those seen in wild-type mice. We detected iNKT cells from tissues in which they are rare, including adipose, lung, skin-draining lymph nodes, and a previously undescribed population in Peyer's patches (PP). PP-NKT cells produce the majority of the IL-4 in Peyer's patches and provide indirect help for B-cell class switching to IgG1 in both transnuclear and wild-type mice. Oral vaccination with α-galactosylceramide shows enhanced fecal IgG1 titers in iNKT cell-sufficient mice. Transcriptional profiling reveals a unique signature of PP-NKT cells, characterized by tissue residency. We thus define PP-NKT as potentially important for surveillance for mucosal pathogens.
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http://dx.doi.org/10.15252/embj.2018101260DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6627243PMC
July 2019

Programmable bacteria as cancer therapy.

Nat Med 2019 07;25(7):1030-1031

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA.

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http://dx.doi.org/10.1038/s41591-019-0513-4DOI Listing
July 2019

GM-CSF, IL-3, and IL-5 Family of Cytokines: Regulators of Inflammation.

Immunity 2019 04;50(4):796-811

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA, USA; Department of Immunology, Harvard Medical School, Boston, MA, USA. Electronic address:

The β common chain cytokines GM-CSF, IL-3, and IL-5 regulate varied inflammatory responses that promote the rapid clearance of pathogens but also contribute to pathology in chronic inflammation. Therapeutic interventions manipulating these cytokines are approved for use in some cancers as well as allergic and autoimmune disease, and others show promising early clinical activity. These approaches are based on our understanding of the inflammatory roles of these cytokines; however, GM-CSF also participates in the resolution of inflammation, and IL-3 and IL-5 may also have such properties. Here, we review the functions of the β common cytokines in health and disease. We discuss preclinical and clinical data, highlighting the potential inherent in targeting these cytokine pathways, the limitations, and the important gaps in understanding of the basic biology of this cytokine family.
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http://dx.doi.org/10.1016/j.immuni.2019.03.022DOI Listing
April 2019

Broadening the Impact of Immunotherapy to Pancreatic Cancer: Challenges and Opportunities.

Gastroenterology 2019 05 18;156(7):2056-2072. Epub 2019 Jan 18.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, and Department of Immunology, Harvard Medical School, Boston, Massachusetts. Electronic address:

Pancreatic ductal adenocarcinoma (PDAC) is projected to become the second deadliest cancer in the United States by 2025, with 5-year survival at less than 10%. In other recalcitrant cancers, immunotherapy has shown unprecedented response rates, including durable remissions after drug discontinuation. However, responses to immunotherapy in PDAC are rare. Accumulating evidence in mice and humans suggests that this remarkable resistance is linked to the complex, dueling role of the immune system in simultaneously promoting and restraining PDAC. In this review, we highlight the rationale that supports pursuing immunotherapy in PDAC, outline the key barriers that limit immunotherapy efficacy, and summarize the primary preclinical and clinical efforts to sensitize PDAC to immunotherapy.
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http://dx.doi.org/10.1053/j.gastro.2018.12.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6486864PMC
May 2019

Altered Binding of Tumor Antigenic Peptides to MHC Class I Affects CD8 T Cell-Effector Responses.

Cancer Immunol Res 2018 12 23;6(12):1524-1536. Epub 2018 Oct 23.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts.

T-cell priming occurs when a naïve T cell recognizes cognate peptide-MHC complexes on an activated antigen-presenting cell. The circumstances of this initial priming have ramifications on the fate of the newly primed T cell. Newly primed CD8 T cells can embark onto different trajectories, with some becoming short-lived effector cells and others adopting a tissue resident or memory cell fate. To determine whether T-cell priming influences the quality of the effector T-cell response to tumors, we used transnuclear CD8 T cells that recognize the melanoma antigen TRP1 using TRP1 or TRP1 TCRs that differ in both affinity and fine specificity. From a series of altered peptide ligands, we identified a point mutation (K8) in a nonanchor residue that, when analyzed crystallographically and biophysically, destabilized the peptide interaction with the MHC binding groove. , the K8 peptide induced robust proliferation of both TRP1 and TRP1 CD8 T cells but did not induce expression of PD-1. Cytokine production from K8-stimulated TRP1 cells was minimal, whereas cytotoxicity was increased. Upon transfer into B16 tumor-bearing mice, the reference peptide (TRP1-M9)- and K8-stimulated TRP1 cells were equally effective at controlling tumor growth but accomplished this through different mechanisms. TRP1-M9-stimulated cells produced more IFNγ, whereas K8-stimulated cells accumulated to higher numbers and were more cytotoxic. We, therefore, conclude that TCR recognition of weakly binding peptides during priming can skew the effector function of tumor-specific CD8 T cells.
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http://dx.doi.org/10.1158/2326-6066.CIR-18-0348DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6290996PMC
December 2018

Radiation and Local Anti-CD40 Generate an Effective Vaccine in Preclinical Models of Pancreatic Cancer.

Front Immunol 2018 7;9:2030. Epub 2018 Sep 7.

Department of Radiation Oncology, Brigham and Women's Hospital, Boston, MA, United States.

Radiation therapy induces immunogenic cell death, which can theoretically stimulate T cell priming and induction of tumor-specific memory T cell responses, serving as an vaccine. In practice, this abscopal effect is rarely observed. We use two mouse models of pancreatic cancer to show that a single dose of stereotactic body radiation therapy (SBRT) synergizes with intratumoral injection of agonistic anti-CD40, resulting in regression of non-treated contralateral tumors and formation of long-term immunologic memory. Long-term survival was not observed when mice received multiple fractions of SBRT, or when TGFβ blockade was combined with SBRT. SBRT and anti-CD40 was so effective at augmenting T cell priming, that memory CD8 T cell responses to both tumor and self-antigens were induced, resulting in vitiligo in long-term survivors.
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http://dx.doi.org/10.3389/fimmu.2018.02030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6137176PMC
October 2019

Rapid CLIP dissociation from MHC II promotes an unusual antigen presentation pathway in autoimmunity.

J Exp Med 2018 10 5;215(10):2617-2635. Epub 2018 Sep 5.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, MA

A number of autoimmunity-associated MHC class II proteins interact only weakly with the invariant chain-derived class II-associated invariant chain peptide (CLIP). CLIP dissociates rapidly from I-A even in the absence of DM, and this property is related to the type 1 diabetes-associated β57 polymorphism. We generated knock-in non-obese diabetic (NOD) mice with a single amino acid change in the CLIP segment of the invariant chain in order to moderately slow CLIP dissociation from I-A These knock-in mice had a significantly reduced incidence of spontaneous type 1 diabetes and diminished islet infiltration by CD4 T cells, in particular T cells specific for fusion peptides generated by covalent linkage of proteolytic fragments within β cell secretory granules. Rapid CLIP dissociation enhanced the presentation of such extracellular peptides, thus bypassing the conventional MHC class II antigen-processing pathway. Autoimmunity-associated MHC class II polymorphisms therefore not only modify binding of self-peptides, but also alter the biochemistry of peptide acquisition.
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http://dx.doi.org/10.1084/jem.20180300DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6170167PMC
October 2018

One-step CRISPR/Cas9 method for the rapid generation of human antibody heavy chain knock-in mice.

EMBO J 2018 09 7;37(18). Epub 2018 Aug 7.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA

Here, we describe a one-step, CRISPR/Cas9 nuclease-mediated strategy to generate knock-in mice. We produced knock-in (KI) mice wherein a 1.9-kb DNA fragment bearing a pre-arranged human B-cell receptor heavy chain was recombined into the native murine immunoglobulin locus. Our methodology relies on Cas9 nuclease-induced double-stranded breaks directed by two sgRNAs to occur within the specific target locus of fertilized oocytes. These double-stranded breaks are subsequently repaired via homology-directed repair by a plasmid-borne template containing the pre-arranged human immunoglobulin heavy chain. To validate our knock-in mouse model, we examined the expression of the KI immunoglobulin heavy chains by following B-cell development and performing single B-cell receptor sequencing. We optimized this strategy to generate immunoglobulin KI mice in a short amount of time with a high frequency of homologous recombination (30-50%). In the future, we envision that such knock-in mice will provide much needed vaccination models to evaluate immunoresponses against immunogens specific for various infectious diseases.
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http://dx.doi.org/10.15252/embj.201899243DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6138433PMC
September 2018

Unmasking Pancreatic Cancer: Epitope Spreading After Single Antigen Chimeric Antigen Receptor T-Cell Therapy in a Human Phase I Trial.

Gastroenterology 2018 07 6;155(1):11-14. Epub 2018 Jun 6.

Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts. Electronic address:

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http://dx.doi.org/10.1053/j.gastro.2018.06.023DOI Listing
July 2018

Regulation of innate and adaptive antitumor immunity by IAP antagonists.

Immunotherapy 2018 07 29;10(9):787-796. Epub 2018 May 29.

Harvard Medical School, Boston, MA 02115, USA.

Inhibition of the T-cell co-inhibitory checkpoint receptors or their ligands CTLA-4, PD-1 and PD-L1 using monoclonal antibodies has proven to be highly effective against many cancers. Yet many cancers remain resistant to checkpoint blockade, and durable remissions occur in only a minority of patients. Novel approaches to enhancing antitumor responses are thus necessary in order to expand the reach of these treatments. The inhibitor of apoptosis (IAP) protein family comprises a diverse group of proteins, many of which have immunoregulatory roles. Small molecule IAP antagonists have been developed and are undergoing early phase clinical testing. These drugs were initially developed to promote tumor cell apoptosis; however, a considerable body of work now indicates that IAP antagonists induce antitumor activity through modulation of innate and adaptive immunity. Primarily through inhibition of cellular (c)-IAP1 and c-IAP2, IAP antagonists can activate alternative NF-κB signaling, promoting B-cell survival, activation of dendritic cells and delivering a broad co-stimulatory signal to T cells. At the same time, IAP antagonists can promote tumor cell intrinsic sensitization to innate immune signals, and enhance tumor cell killing by inflammatory cytokines and phagocytic macrophages. These drugs thus represent an attractive investigational approach to immunotherapy, providing a positive signaling counterpart to the relief of signal inhibition conferred by checkpoint blockade.
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http://dx.doi.org/10.2217/imt-2017-0185DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6462841PMC
July 2018

The systemic response to surgery triggers the outgrowth of distant immune-controlled tumors in mouse models of dormancy.

Sci Transl Med 2018 04;10(436)

Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.

Patients undergoing surgical resection of primary breast tumors confront a risk for metastatic recurrence that peaks sharply 12 to 18 months after surgery. The cause of early metastatic relapse in breast cancer has long been debated, with many ascribing these relapses to the natural progression of the disease. Others have proposed that some aspect of surgical tumor resection triggers the outgrowth of otherwise-dormant metastases, leading to the synchronous pattern of relapse. Clinical data cannot distinguish between these hypotheses, and previous experimental approaches have not provided clear answers. Such uncertainty hinders the development and application of therapeutic approaches that could potentially reduce early metastatic relapse. We describe an experimental model system that definitively links surgery and the subsequent wound-healing response to the outgrowth of tumor cells at distant anatomical sites. Specifically, we find that the systemic inflammatory response induced after surgery promotes the emergence of tumors whose growth was otherwise restricted by a tumor-specific T cell response. Furthermore, we demonstrate that perioperative anti-inflammatory treatment markedly reduces tumor outgrowth in this model, suggesting that similar approaches might substantially reduce early metastatic recurrence in breast cancer patients.
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http://dx.doi.org/10.1126/scitranslmed.aan3464DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6364295PMC
April 2018

Anti-CTLA-4 therapy requires an Fc domain for efficacy.

Proc Natl Acad Sci U S A 2018 04 26;115(15):3912-3917. Epub 2018 Mar 26.

Division of Gastroenterology, Massachusetts General Hospital, Boston, MA 02114;

Ipilimumab, a monoclonal antibody that recognizes cytotoxic T lymphocyte antigen (CTLA)-4, was the first approved "checkpoint"-blocking anticancer therapy. In mouse tumor models, the response to antibodies against CTLA-4 depends entirely on expression of the Fcγ receptor (FcγR), which may facilitate antibody-dependent cellular phagocytosis, but the contribution of simple CTLA-4 blockade remains unknown. To understand the role of CTLA-4 blockade in the complete absence of Fc-dependent functions, we developed H11, a high-affinity alpaca heavy chain-only antibody fragment (VHH) against CTLA-4. The VHH H11 lacks an Fc portion, binds monovalently to CTLA-4, and inhibits interactions between CTLA-4 and its ligand by occluding the ligand-binding motif on CTLA-4 as shown crystallographically. We used H11 to visualize CTLA-4 expression in vivo using whole-animal immuno-PET, finding that surface-accessible CTLA-4 is largely confined to the tumor microenvironment. Despite this, H11-mediated CTLA-4 blockade has minimal effects on antitumor responses. Installation of the murine IgG2a constant region on H11 dramatically enhances its antitumor response. Coadministration of the monovalent H11 VHH blocks the efficacy of a full-sized therapeutic antibody. We were thus able to demonstrate that CTLA-4-binding antibodies require an Fc domain for antitumor effect.
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http://dx.doi.org/10.1073/pnas.1801524115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5899492PMC
April 2018

Targeting Cytokine Therapy to the Pancreatic Tumor Microenvironment Using PD-L1-Specific VHHs.

Cancer Immunol Res 2018 04 19;6(4):389-401. Epub 2018 Feb 19.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts.

Cytokine-based therapies for cancer have not achieved widespread clinical success because of inherent toxicities. Treatment for pancreatic cancer is limited by the dense stroma that surrounds tumors and by an immunosuppressive tumor microenvironment. To overcome these barriers, we developed constructs of single-domain antibodies (VHHs) against PD-L1 fused with IL-2 and IFNγ. Targeting cytokine delivery in this manner reduced pancreatic tumor burden by 50%, whereas cytokines fused to an irrelevant VHH, or blockade of PD-L1 alone, showed little effect. Targeted delivery of IL-2 increased the number of intratumoral CD8 T cells, whereas IFNγ reduced the number of CD11b cells and skewed intratumoral macrophages toward the display of M1-like characteristics. Imaging of fluorescent VHH-IFNγ constructs, as well as transcriptional profiling, demonstrated targeting of IFNγ to the tumor microenvironment. Many tumors and tumor-infiltrating myeloid cells express PD-L1, rendering them potentially susceptible to this form of targeted immunotherapy. .
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http://dx.doi.org/10.1158/2326-6066.CIR-17-0495DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6079513PMC
April 2018

Generation of Ca2+-independent sortase A mutants with enhanced activity for protein and cell surface labeling.

PLoS One 2017 4;12(12):e0189068. Epub 2017 Dec 4.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts, United States of America.

Sortase A, a calcium-dependent transpeptidase derived from Staphylococcus aureus, is used in a broad range of applications, such as the conjugation of fluorescent dyes and other moieties to proteins or to the surface of eukaryotic cells. In vivo and cell-based applications of sortase have been somewhat limited by the large range of calcium concentrations, as well as by the often transient nature of protein-protein interactions in living systems. In order to use sortase A for cell labeling applications, we generated a new sortase A variant by combining multiple mutations to yield an enzyme that was both calcium-independent and highly active. This variant has enhanced activity for both N- and C-terminal labeling, as well as for cell surface modification under physiological conditions.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0189068PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5714338PMC
December 2017

The Pancreatic Cancer Microenvironment.

Cancer J 2017 Nov/Dec;23(6):321-325

From the Dana-Farber Cancer Institute, Boston, MA.

Pancreatic ductal adenocarcinoma (PDAC) is composed of a minority of malignant cells within a microenvironment of extracellular matrix, fibroblasts, endothelial cells, and immune cells. Therapeutic failures of chemotherapy, targeted therapy, and immunotherapy have all been attributed to the PDAC microenvironment. In this review, we dissect the components of the microenvironment and explain how each cell type contributes to form a highly immunosuppressive, hypoxic, and desmoplastic cancer. New efforts in single-cell profiling will enable a better understanding of the composition of the microenvironment in primary and metastatic PDAC, as well as an understanding of how the microenvironment may respond to novel therapeutic approaches.
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http://dx.doi.org/10.1097/PPO.0000000000000288DOI Listing
July 2018

IAP Antagonists Enhance Cytokine Production from Mouse and Human iNKT Cells.

Cancer Immunol Res 2018 01 29;6(1):25-35. Epub 2017 Nov 29.

Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Boston, Massachusetts.

Inhibitor of apoptosis protein (IAP) antagonists are in clinical trials for a variety of cancers, and mouse models show synergism between IAP antagonists and anti-PD-1 immunotherapy. Although IAP antagonists affect the intrinsic signaling of tumor cells, their most pronounced effects are on immune cells and the generation of antitumor immunity. Here, we examined the effects of IAP antagonism on T-cell development using mouse fetal thymic organ culture and observed a selective loss of iNKT cells, an effector cell type of potential importance for cancer immunotherapy. Thymic iNKT-cell development probably failed due to increased strength of TCR signal leading to negative selection, given that mature iNKT cells treated with IAP antagonists were not depleted, but had enhanced cytokine production in both mouse and human cultures. Consistent with this, mature mouse primary iNKT cells and iNKT hybridomas increased production of effector cytokines in the presence of IAP antagonists. administration of IAP antagonists and α-GalCer resulted in increased IFNγ and IL-2 production from iNKT cells and decreased tumor burden in a mouse model of melanoma lung metastasis. Human iNKT cells also proliferated and increased IFNγ production dramatically in the presence of IAP antagonists, demonstrating the utility of these compounds in adoptive therapy of iNKT cells. .
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http://dx.doi.org/10.1158/2326-6066.CIR-17-0490DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5754232PMC
January 2018

PD-L1 is an activation-independent marker of brown adipocytes.

Nat Commun 2017 09 21;8(1):647. Epub 2017 Sep 21.

Whitehead Institute for Biomedical Research, Cambridge, MA, 02142, USA.

Programmed death ligand 1 (PD-L1) is expressed on a number of immune and cancer cells, where it can downregulate antitumor immune responses. Its expression has been linked to metabolic changes in these cells. Here we develop a radiolabeled camelid single-domain antibody (anti-PD-L1 VHH) to track PD-L1 expression by immuno-positron emission tomography (PET). PET-CT imaging shows a robust and specific PD-L1 signal in brown adipose tissue (BAT). We confirm expression of PD-L1 on brown adipocytes and demonstrate that signal intensity does not change in response to cold exposure or β-adrenergic activation. This is the first robust method of visualizing murine brown fat independent of its activation state.Current approaches to visualise brown adipose tissue (BAT) rely primarily on markers that reflect its metabolic activity. Here, the authors show that PD-L1 is expressed on brown adipocytes, does not change upon BAT activation, and that BAT volume in mice can be measured by PET-CT with a radiolabeled anti-PD-L1 antibody.
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http://dx.doi.org/10.1038/s41467-017-00799-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5608754PMC
September 2017
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