PD-1 Dynamically Regulates Inflammation and Development of Brain-Resident Memory CD8 T Cells During Persistent Viral Encephalitis

Programmed cell death-1 (PD-1) receptor signaling dampens the functionality of T cells faced with repetitive antigenic stimulation from chronic infections or tumors. Using intracerebral (i.c.) inoculation with mouse polyomavirus (MuPyV), we have shown that CD8 T cells establish a PD-1hi, tissue-resident memory population in the brains (bTRM) of mice with a low-level persistent infection. In MuPyV encephalitis, PD-L1 was expressed on infiltrating myeloid cells, microglia and astrocytes, but not on oligodendrocytes. Engagement of PD-1 on anti-MuPyV CD8 T cells limited their effector activity. NanoString gene expression analysis showed that neuroinflammation was higher in PD-L1−/− than wild type mice at day 8 post-infection, the peak of the MuPyV-specific CD8 response. During the persistent phase of infection, however, the absence of PD-1 signaling was found to be associated with a lower inflammatory response than in wild type mice. Genetic disruption and intracerebroventricular blockade of PD-1 signaling resulted in an increase in number of MuPyV-specific CD8 bTRM and the fraction of these cells expressing CD103, the αE integrin commonly used to define tissue-resident T cells. However, PD-L1−/− mice persistently infected with MuPyV showed impaired virus control upon i.c. re-infection with MuPyV. Collectively, these data reveal a temporal duality in PD-1-mediated regulation of MuPyV-associated neuroinflammation. PD-1 signaling limited the severity of neuroinflammation during acute infection but sustained a level of inflammation during persistent infection for maintaining control of virus re-infection

Regulation of transcription factor IRF8 in myeloid progenitors is a critical checkpoint for formation of defective myeloid cells in cancer.

Abstract Defectivemyeloid cells are commonly observed across a large range of solid cancers. These defects contribute to immune suppression and tumor progression tometastatic disease. Early studies focused on characterizing such myeloid-derivedsuppressor cells (MDSC) in peripheral tissues and the tumor microenvironment, but molecular processes underlying their development remained unclear. Bone marrowis the major site for normal myeloid production and is known to respond to signalsfrom the periphery. Thus, we hypothesizedthat tumors perturb bone marrow myelopoiesis, resulting in production of MDSC. During myelopoiesis, transcription factor IRF8 expression within granulocyte/monocyteprogenitors (GMP) determines differentiation such that high IRF8 favorsmonocytes while low IRF8 favors granulocytes. We previously revealed that IRF8 suppressionby tumor-derived factors leads to accumulation of MDSC, but exactly when andwhere IRF8 is compromised was unknown. Here we utilize a novel mouse model expressingan IRF8-EGFP fusion protein to investigate specific changes in IRF8 during myelopoiesisin both orthotopic and spontaneous mammary tumor progression. Our results show that:1) GMP can be divided into IRF8hi and IRF8lo expressingpopulations corresponding to monocytic and granulocytic progenitor phenotypes,2) IRF8lo GMP preferentially expand with increasing tumor size andincreasing G-CSF serum concentration, and 3) enforced IRF8 expression restrainsthis expansion and reduces the frequency of myeloid suppressors in theperiphery. Altogether, these data identify modulation of IRF8 in myeloidprogenitors as a consequence of tumor development and potential target fortherapeutic intervention.</jats:p

Abstract 3249: Regulation of interferon regulatory factor-8 (IRF8) during myelopoiesis is a critical checkpoint for the formation of defective myeloid cells in cancer

Abstract Aberrant myelopoiesis is commonly observed in patients with solid cancers. This results in the production of myeloid-derived suppressive cells (MDSCs) which promote tumor growth and metastasis. However, the molecular mechanisms underlying MDSC development have remained poorly understood. The transcription factor IRF8 is integral for overseeing myelopoietic fate; high IRF8 expression during normal myelopoiesis favors monocytic differentiation and directly inhibits granulocytic differentiation. We recently showed that tumor-derived STAT3/5-activating cytokines downregulate the expression of IRF8 in myeloid cells leading to the accumulation of MDSCs in the periphery, which are primarily granulocytic. However it is unclear if MDSC production is a consequence of IRF8 downregulation in the periphery or upstream in the bone marrow during myelopoiesis. To that end, we utilized a novel mouse model expressing an IRF8-EGFP fusion protein which allowed us to investigate changes in IRF8 expression during both orthotopic and spontaneous mammary tumor growth and progression. Our results showed that: 1) the total granulocyte-monocyte progenitor (GMP) fraction, as with the peripheral MDSC population, was responsive to tumor burden and greatly expanded with increasing tumor size, and 2) IRF8 expression within the total GMPs significantly decreased during tumor growth, reflecting the expansion of a newly defined primarily IRF8lo granulocyte progenitor (GP) population, and 3) tumor-induced GPs were hyper-responsive not only to G-CSF, but also M-CSF to generate a ‘biased’ granulocytic MDSC-like phenotype. Expression of IRF8 in other progenitor populations was not affected, suggesting that IRF8lo GMPs/GPs are a key source of the peripheral MDSC response. Lastly, genetically enforced IRF8 expression in myeloid progenitors of the bone marrow constrained aberrant myelopoiesis during tumor growth, resulting in delayed autochthonous tumor growth and reduced spontaneous lung metastasis. Altogether, these data reinforce the notion that modulation of IRF8 in myeloid progenitors is an early consequence of the neoplastic process and a potential target for therapeutic intervention. NIH R01CA140622 NIH T32CA085183 Citation Format: Colleen S. Netherby, Michelle N. Messmer, Scott I. Abrams. Regulation of interferon regulatory factor-8 (IRF8) during myelopoiesis is a critical checkpoint for the formation of defective myeloid cells in cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3249.</jats:p

Balancing Inflammation and Central Nervous System Homeostasis: T Cell Receptor Signaling in Antiviral Brain TRM Formation and Function

Tissue-resident memory (TRM) CD8 T cells provide early frontline defense against regional pathogen reencounter. CD8 TRM are predominantly parked in nonlymphoid tissues and do not circulate. In addition to this anatomic difference, TRM are transcriptionally and phenotypically distinct from central-memory T cells (TCM) and effector-memory T cells (TEM). Moreover, TRM differ phenotypically, functionally, and transcriptionally across barrier tissues (e.g., gastrointestinal tract, respiratory tract, urogenital tract, and skin) and in non-barrier organs (e.g., brain, liver, kidney). In the brain, TRM are governed by a contextual milieu that balances TRM activation and preservation of essential post-mitotic neurons. Factors contributing to the development and maintenance of brain TRM, of which T cell receptor (TCR) signal strength and duration is a central determinant, vary depending on the infectious agent and modulation of TCR signaling by inhibitory markers that quell potentially pathogenic inflammation. This review will explore our current understanding of the context-dependent factors that drive the acquisition of brain (b)TRM phenotype and function, and discuss the contribution of TRM to promoting protective immune responses in situ while maintaining tissue homeostasis.</jats:p

Tumor-induced myeloid dysfunction and its implications for cancer immunotherapy

Immune function relies on an appropriate balance of the lymphoid and myeloid responses. In the case of neoplasia, this balance is readily perturbed by the dramatic expansion of immature or dysfunctional myeloid cells accompanied by a reciprocal decline in the quantity/quality of the lymphoid response. In this review, we seek to: 1) define the nature of the atypical myelopoiesis observed in cancer patients and the impact of this perturbation on clinical outcomes; 2) examine the potential mechanisms underlying these clinical manifestations; and 3) explore potential strategies to restore normal myeloid cell differentiation to improve activation of the host antitumor immune response. We posit that fundamental alterations in myeloid homeostasis triggered by the neoplastic process represent critical checkpoints that govern therapeutic efficacy, as well as offer novel cellular-based biomarkers for tracking changes in disease status or relapse

CD8 T Cells and STAT1 Signaling Are Essential Codeterminants in Protection from Polyomavirus Encephalopathy

A comprehensive understanding of JCPyV-induced PML pathogenesis is needed to define determinants that predispose patients to PML, a goal whose urgency is heightened by the lack of anti-JCPyV agents. A handicap to achieving this goal is the lack of a tractable animal model to study PML pathogenesis. Using intracerebral inoculation with MuPyV, we found that MuPyV encephalitis in wild-type mice causes an encephalopathy, which is markedly exacerbated in mice deficient in STAT1, a molecule involved in transducing signals from type I, II, and III IFN receptors. CD8 T cell deficiency compounded the severity of MuPyV neuropathology and resulted in dramatically elevated virus levels in the CNS. These findings demonstrate that STAT1 signaling and CD8 T cells concomitantly act to mitigate MuPyV-encephalopathy and control viral infection.</jats:p

IL-21 from high-affinity CD4 T cells drives differentiation of brain-resident CD8 T cells during persistent viral infection

High-affinity CD4 T cells produce IL-21 to help CD8 T cells differentiate into tissue-resident memory cells in the brain.</jats:p

The Granulocyte Progenitor Stage Is a Key Target of IRF8-Mediated Regulation of Myeloid-Derived Suppressor Cell Production

Abstract Alterations in myelopoiesis are common across various tumor types, resulting in immature populations termed myeloid-derived suppressor cells (MDSCs). MDSC burden correlates with poorer clinical outcomes, credited to their ability to suppress antitumor immunity. MDSCs consist of two major subsets, monocytic and polymorphonuclear (PMN). Intriguingly, the latter subset predominates in many patients and tumor models, although the mechanisms favoring PMN-MDSC responses remain poorly understood. Ordinarily, lineage-restricted transcription factors regulate myelopoiesis that collectively dictate cell fate. One integral player is IFN regulatory factor (IRF)–8, which promotes monocyte/dendritic cell differentiation while limiting granulocyte development. We recently showed that IRF8 inversely controls MDSC burden in tumor models, particularly the PMN-MDSC subset. However, where IRF8 acts in the pathway of myeloid differentiation to influence PMN-MDSC production has remained unknown. In this study, we showed that: 1) tumor growth was associated with a selective expansion of newly defined IRF8lo granulocyte progenitors (GPs); 2) tumor-derived GPs had an increased ability to form PMN-MDSCs; 3) tumor-derived GPs shared gene expression patterns with IRF8−/− GPs, suggesting that IRF8 loss underlies GP expansion; and 4) enforced IRF8 overexpression in vivo selectively constrained tumor-induced GP expansion. These findings support the hypothesis that PMN-MDSCs result from selective expansion of IRF8lo GPs, and that strategies targeting IRF8 expression may limit their load to improve immunotherapy efficacy.</jats:p