Probes for INS

γδ T cells are a subset of nonconventional T cells that play a critical role in bridging the innate and adaptive arms of the immune system. γδ T cells are particularly abundant in ruminant species and may constitute up to 60% of the circulating lymphocyte pool in young cattle. The frequency of circulating γδ T cells is highest in neonatal calves and declines as the animal ages, suggesting these cells may be particularly important in the immune system of the very young. Bovine respiratory syncytial virus (BRSV) is a significant cause of respiratory infection in calves, and is most severe in animals under one year of age. BRSV is also a significant factor in the development of bovine respiratory disease complex (BRDC), the leading cause of morbidity and mortality in feedlot cattle. Human respiratory syncytial virus (RSV) is closely related to BRSV and a leading cause of lower respiratory tract infection in infants and children worldwide. BRSV infection in calves shares striking similarities with RSV infection in human infants. To date, there have been few studies defining the role of γδ T cells in the immune response to BRSV or RSV infection in animals or humans, respectively. However, emerging evidence suggests that γδ T cells may play a critical role in the early recognition of infection and in shaping the development of the adaptive immune response through inflammatory chemokine and cytokine production. Further, while it is clear that γδ T cells accumulate in the lungs during BRSV and RSV infection, their role in protection vs. immunopathology remains unclear. This review will summarize what is currently known about the role of γδ T cells in the immune response to BRSV and BRDC in cattle, and where appropriate, draw parallels to the role of γδ T cells in the human response to RSV infection.

Abstract

BACKGROUND:

Craniopharyngiomas are neoplasms of the sellar/parasellar region that are classified into adamantinomatous (ACP) and papillary (PCP) subtypes. Surgical resection of craniopharyngiomas is challenging, and recurrence is common, frequently leading to profound morbidity. BRAF V600E mutations render PCP susceptible to BRAF/MEK inhibitors, but effective targeted therapies are needed for ACP. We explored the feasibility of targeting the PD-1/PD-L1 immune checkpoint pathway in ACP and PCP.

METHODS:

We mapped and quantified PD-L1 and PD-1 expression in ACP and PCP resections using immunohistochemistry, immunofluorescence, and RNA in situ hybridization. We used tissue-based cyclic immunofluorescence (t-CyCIF) to map the spatial distribution of immune cells and characterize cell cycle and signaling pathways in ACP tumor cells which intrinsically express PD-1.

RESULTS:

All ACP (15±14% of cells, n=23, average±S.D.) and PCP (35±22% of cells, n=18) resections expressed PD-L1. In ACP, PD-L1 was predominantly expressed by tumor cells comprising the cyst-lining. In PCP, PD-L1 was highly-expressed by tumor cells surrounding the stromal fibrovascular cores. ACP also exhibited tumor cell-intrinsic PD-1 expression in whorled epithelial cells with nuclear-localized beta-catenin. These cells exhibited evidence of elevated mTOR and MAPK signaling. Profiling of immune populations in ACP and PCP showed a modest density of CD8+ T-cells.

CONCLUSIONS:

ACP exhibit PD-L1 expression in the tumor cyst-lining and intrinsic PD-1 expression in cells proposed to comprise an oncogenic stem-like population. In PCP, proliferative tumor cells express PD-L1 in a continuous band at the stromal-epithelial interface. Targeting PD-L1 and/or PD-1 in both subtypes of craniopharyngioma might therefore be an effective therapeutic strategy.

Astrocytes are the most abundant glial cells of the central nervous system and have recently been recognized as crucial in the regulation of brain immunity. In most neuropathological conditions, astrocytes are prone to a radical phenotypical change called reactivity, which plays a key role in astrocyte contribution to neuroinflammation. However, how astrocytes regulate brain immunity in healthy conditions is an understudied question. One of the astroglial molecule involved in these regulations might be Connexin 43 (Cx43), a gap junction protein highly enriched in astrocyte perivascular endfeet-terminated processes forming the glia limitans. Indeed, Cx43 deletion in astrocytes (Cx43KO) promotes a continuous immune recruitment and an autoimmune response against an astrocyte protein, without inducing any brain lesion. To investigate the molecular basis of this unique immune response, we characterized the polysomal transcriptome of hippocampal astrocytes deleted for Cx43. Our results demonstrate that, in the absence of Cx43, astrocytes adopt an atypical reactive status with no change in most canonical astrogliosis markers, but with an upregulation of molecules promoting immune recruitment, complement activation as well as anti-inflammatory processes. Intriguingly, while several of these upregulated transcriptional events suggested an activation of the γ-interferon pathway, no increase in this cytokine or activation of related signaling pathways were found in Cx43KO. Finally, deletion of astroglial Cx43 was associated with the upregulation of several angiogenic factors, consistent with an increase in microvascular density in Cx43KO brains. Collectively, these results strongly suggest that Cx43 controls immunoregulatory and angiogenic properties of astrocytes.

PD-L1 expression may be a predictive marker for anti-PD-1 therapeutic efficacy. No standard detection method of PD-L1 expression was available for advanced gastric cancer (AGC), which would be investigated in this study using RNA in situ hybridization and immunohistochemistry. Patients (N = 165) with AGC treated at Peking University Cancer Hospital from October 2008 to February 2013 were retrospectively studied. Tissue samples prior to chemotherapy were assessed for PD-L1 expression using RNA in situ hybridization (an RNAscope assay) and immunohistochemistry (IHC). The correlations of PD-L1 expression to patient characteristics and clinical outcomes were statistically analyzed. PD-L1 mRNA signals were located in tumor compartments or the mesenchyme in a brown dotted or clustered pattern, and PD-L1 mRNA expression in gastric cancer was heterogeneous. PD-L1-positive expressions were observed in 33.9% (56/165) and 35.1% (46/131) patients in mRNA level and protein level, respectively. A positive relationship was found between PD-L1 mRNA and PD-L1 protein, and compared to IHC, RNAscope assay could provide an intuitional and quantitative data with potential clinical application. No statistically significant differences occurred between PD-L1 expression and clinical response to chemotherapy, or survival. However, we found that PD-L1 expression was higher in intestinal type than in diffuse type. These findings suggested that the RNAscope assay may be a promising method for patient assessment in gastric cancer clinical trials, which would be illustrated in further study.

ABSTRACT

Triple-negative breast cancer (TNBC) represents a more aggressive and difficult subtype of breast cancer where responses to chemotherapy occur, but toxicity is significant and resistance often follows. Immunotherapy has shown promising results in various types of cancer, including breast cancer. Here, we investigated a new combination strategy where histone deacetylase inhibitors (HDACi) are applied with immune checkpoint inhibitors to improve immunotherapy responses in TNBC.

Testing different epigenetic modifiers, we focused on the mechanisms underlying HDACi as priming modulators of immunotherapy. Tumor cells were co-cultured with human peripheral blood mononuclear cells (PBMCs) and flow cytometric immunophenotyping was performed to define the role of epigenetic priming in promoting tumor antigen presentation and immune cell activation. We found that HDACi up-regulate PD-L1 mRNA and protein expression in a time-dependent manner in TNBC cells, but not in hormone responsive cells. Focusing on TNBC, HDACi up-regulated PD-L1 and HLA-DR on tumor cells when co-cultured with PBMCs and down-regulated CD4+ Foxp3+ Treg in vitro. HDACi significantly enhanced the in vivo response to PD-1/CTLA-4 blockade in the triple-negative 4T1 breast cancer mouse model, the only currently available experimental system with functional resemblance to human TNBC. This resulted in a significant decrease in tumor growth and increased survival, associated with increased T cell tumor infiltration and a reduction in CD4+Foxp3+ T cells in the tumor microenvironment. Overall, our results suggest a novel role for HDAC inhibition in combination with immune checkpoint inhibitors and identify a promising therapeutic strategy, supporting its further clinical evaluation for TNBC treatment.