Th17 Pathway Research Articles

Th Pathways in Immune-Mediated Skin Disorders: A Guide for Strategic Treatment Decisions.

In recent years, there have been significant breakthroughs in the identification of immunological components of skin diseases and in the development of immunomodulatory drugs. Novel therapies create exciting prospects for personalized care. This article provides an overview of the role played by Th1, Th2, Th17, and follicular Th pathways in the most common skin diseases. Additionally, it elucidates the impact of current and upcoming treatments on each of these signaling cascades. Skin diseases predominantly influenced by a single dominant Th pathway such as psoriasis and atopic dermatitis are well-suited for biologics. However, in many other disorders a complex interplay between different immune pathways exists. This can lead to inconsistent efficacy of biologics based on individual patient profiles. In case of activation of several Th pathways, it may be more suitable to consider conventional therapies or JAK inhibitors. Increasing immunological insights have transitioned from laboratory research to practical applications, a trend that is expected to continue growing in the future.

Comparison of inflammatory molecular mechanisms between osteoarthritis and rheumatoid arthritis via gene microarrays.

Osteoarthritis (OA) and rheumatoid arthritis (RA) treatment requires exact arthritis type diagnosis. We compared inflammatory molecular mechanisms between OA and RA to introduce reliable molecular biomarkers. The GSE55235 and GSE100786 microarray datasets were acquired from the GEO. Data preprocessing and differential expression analysis were conducted in OA and RA groups and their control groups applying GEO2R. Differentially expressed genes (DEGs) with a |LogFC|>1 and adj. p<0.05 were determined. Gene ontology (GO) and signaling pathway analysis were done utilizing PANTHER and Enrichr. The suitability of gene expression alterations as biomarkers was tested using the receiver operating characteristic (ROC) curve analysis. We found 2129 DEGs between the OA and control groups and 2494 DEGs between the RA and control groups. GO on the DEGs showed enrichment in binding, cellular processes, and cellular anatomical entities in molecular functions, biological processes, and cellular components, respectively. Enrichr found the cell differentiation pathways of Th1 and Th2 only in RA. The ROC curve analysis indicated HLA-DQA1 downregulation and MAPK8IP3 upregulation as reliable biomarkers to discriminate RA from OA in peripheral blood and bone marrow samples, respectively. We found more DEGs in patients with OA than those with RA and determined inflammatory pathways and genes unique to RA as reliable biomarkers to discriminate RA from OA. Gene expression alterations associated with Th1 and Th2 cell differentiation pathways, including HLA-DQA1 downregulation and MAPK8IP3 upregulation, could be novel molecular biomarkers to diagnose RA.

The IL-23/Th17 pathway inhibitors in the treatment of psoriasis and the risk of skin malignancies: a review

The IL-23/Th17 pathway inhibitors in the treatment of psoriasis and the risk of skin malignancies: a review

Biomarkers in rosacea: A systematic review.

Rosacea is a chronic and psychologically ladened disease affecting 1%-3% of people worldwide. The identification and validation of biomarkers in rosacea patients has the potential to improve disease progression, support diagnosis, provide objective measures for clinical trials and aid in management. The objective of this review is to systematically identify all rosacea biomarkers, categorize them by type and identify trends to improve disease expression. Eligibility criteria for this review (PROSPERO CRD42023397510) include randomized controlled trials, case-control studies, cohort studies and other observational studies. No restrictions were placed on patient demographics (age, sex, ethnicity) or language of publication until February 2023. Quality of studies was assessed using the National Institute of Health quality assessment tool. The literature search was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines. A total of 805 unique articles were screened based on the applied inclusion and exclusion criteria. After the articles were screened based on title/abstract and full-text, a total of 38 studies were included, reporting on a total of 119 unique biomarkers. The results of this review and current rosacea pathogenic mechanisms provide the greatest support for the innate cathelicidin and inflammasome, Th1 and Th17 pathways. The most commonly reported biomarkers include IL-1β, TNF-α, IL-37, IFN-γ and MMP-9. Biomarkers identified in this study support current theories of rosacea pathogenesis and provide direction for research to further our knowledge. However, more research is needed to identify biomarkers panels that can provide diagnostic utility. This may be difficult due to the heterogeneity of the disease and potential differences between rosacea subtypes.

Identification of Fibroinflammatory and Fibrotic Transcriptomic Subsets of Human Cutaneous Sclerotic Chronic Graft-Versus-Host Disease

Cutaneous sclerotic chronic graft-versus-host disease (cGVHD) is a common and highly morbid complication of allogeneic hematopoietic stem cell transplantation (HSCT). Our goals were to identify signals active in the skin of sclerotic cGVHD patients in an effort to better understand how to treat this manifestation and to explore the heterogeneity of the disease. We identified genes that are significantly upregulated in the skin of sclerotic cGVHD patients (n = 17) compared to HSCT patients without cutaneous cGVHD (n = 9) by bulk RNA sequencing. Sclerotic cGVHD was most associated with Th1, phagocytic, and fibrotic pathways. Additionally, different transcriptomic groups of affected patients were discovered: those with fibrotic and inflammatory/Th1 gene expression (the fibroinflammatory group) and those with predominantly fibrotic/TGFβ-associated expression (the fibrotic group). Further study will help elucidate if these gene expression findings can be used to tailor treatment decisions. Multiple proteins encoded by highly-induced genes in the skin (SFRP4, SERPINE2, COMP) were also highly induced in the plasma of sclerotic cGVHD patients (n = 16) compared to control HSCT patients without sclerotic cGVHD (n = 17), suggesting these TGFβ and Wnt pathway mediators as candidate blood biomarkers of disease.

Tape strips detect molecular alterations and cutaneous biomarkers in skin of patients with hidradenitis suppurativa

Hidradenitis suppurativa (HS) has a high unmet need for better treatments. Biopsies are considered the gold standard for studying molecular alterations in skin. A reproducible, minimally invasive approach is needed for longitudinal monitoring in trials and in pediatric populations. To determine whether skin tape strips can detect molecular alterations in HS and identify biomarkers of disease activity. We performed RNA sequencing on tape strips collected from lesional and healthy-appearing (nonlesional) HS skin (n=22) and healthy controls (n=21). We correlated the expression of skin biomarkers between tape strips and a previously published gene-signature of HS biopsies. Tape strips detected upregulation of known HS biomarkers (eg, Interleukin[IL]-17A) in nonlesional and/or lesional skin and also identified novel clinically actionable targets, including OX40 and JAK3. The expression of Th17 and tumor necrosis factor-α pathways were highly correlated between tape strips and biopsies. HS clinical severity was significantly associated with expression of biomarkers (eg tumor necrosis factor-α , IL-17A/F, OX40, JAK1-3, IL-4R) in HS lesional and/or nonlesional skin. Sample size. Tape stripping is limited in depth. This study validates tape strips as a minimally-invasive approach to identify cutaneous biomarkers in HS. This provides a novel avenue for monitoring treatment efficacy and a potential step toward individualized therapy in HS.

Mechanism of Yixishu lotion in the treatment of vaginitis based on network pharmacology combined with experimental validation: an experimental research study

Objective: Yixishu lotion (YXSL) originates from the summary of traditional Chinese medicine clinical experience and constantly improves in practice in clinical validation of the exact efficacy of traditional Chinese medicine prescription. To explore the mechanism of YXSL in treating vaginitis and the potential mechanisms based on network pharmacology and experimental verification. Methods: The active components and drug-related targets of YXSL were retrieved from the TCMSP (Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform) database, and the target was predicted by the UniProt database. Searching for genes related to ‘vaginitis’ disease in the GeneCards database, a total of 2581 drug targets were obtained. The interaction between proteins (PPI – protein–protein interaction) relationship was obtained by STRING database and visualized by Cytoscape software. Finally, the ‘Bioconductor’ installation package in R software was used to analyze the GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways of the target. Results: In this study, by the method of network pharmacology, the key active components of YXSL were flavonoids such as quercetin, apigenin, kaempferol, luteolin, β-sitosterol; the main core proteins included MAPK14, TP53, FGF2, ESR1, MAPK3, MAPK1, VEGFA, JUN, IL-6, and the KEGG pathway was mainly involved in MAPK pathway, Th17 pathway, Malaria, TNF pathway, and other signaling pathways. Animal experiments showed that the clinical symptoms and vaginal tissue lesions of the YXSL group and the fluconazole group were improved, and the levels of TNF-α (tumor necrosis factor alpha), IL-6 (interleukin-6), MDA (malondialdehyde), SOD (superoxide dismutase), IL-4, and IFN-γ (interferon-γ) in vaginal tissue and serum were better than the model group. Conclusion: YXSL may achieve its therapeutic effect on vaginitis by reducing the inflammatory response, improving oxidative stress response, and improving body immunity, and it provides a theoretical basis for further research on its pharmacodynamic material basis and mechanism of action.

Immunoinformatics Prediction and Protective Efficacy of Vaccine Candidate PiuA-PlyD4 Against Streptococcus Pneumoniae.

This study was designed to evaluate the immune protective efficacy of the novel Streptococcus pneumoniae (S. pneumoniae) protein vaccine PiuA-PlyD4 through immunoinformatics prediction and in vitro and in vivo experiments. In this study, we conducted immunoinformatics prediction and protection analysis on the fusion protein PiuA-PlyD4. The epitope composition of the vaccine was analyzed based on the prediction of B-cell and helper T-cell epitopes. Meanwhile, the molecular docking of PiuA and TLR2/4 was simulated. After immunizing C57BL/6 mice with the prepared vaccine, the biological safety, immunogenicity and conservation were evaluated. By constructing different infection models and from the aspects of adhesion inhibition and cytokines, the protective effect of the fusion protein vaccine PiuA-PlyD4 on S. pneumoniae infection was explored. PiuA-PlyD4 has abundant B-cell and helper T-cell epitopes and shows a high antigenicity score and structural stability. Molecular docking analysis suggested the potential interaction between PiuA and TLR2/4. The specific antibody titer of fusion protein antiserum was as high as (7.81±2.32) ×105. The protective effect of the immunized mice on nasal and lung colonization was significantly better than that of the control group, and the survival rate against S. pneumoniae infection of serotype 3 reached 50%. Cytokine detection showed that the humoral immune response, Th1, Th2 and Th17 cellular immune pathways were all involved in the process. The study indicates that PiuA-PlyD4, whether the results are predicted by immunoinformatics or experimentally validated in vivo and in vitro, has good immunogenicity and immunoreactivity and can provide effective protection against S. pneumoniae infection. Therefore, it can be considered a promising prophylactic vaccine candidate for S. pneumoniae.

Abstract B004: Neoantigen adenoviral cancer vaccine generates improved CD8+ T-cell responses compared to adjuvanted peptide vaccine

Abstract Development of large, functional CD8+ T cell responses is crucial for successful cancer vaccine therapeutics. Neoantigens have demonstrated their potential as cancer vaccine immunogens, but improvements in delivery method are still necessary. In this work, we characterized five candidate Adenovirus Serotype 26 (Ad26) cancer vaccines for prophylactic efficacy in the MC-38 cancer model. All five single shot vaccines generated cellular immune responses at a higher level than prime boost peptide vaccination. The vaccines generated varying degrees of protection against tumor growth. The best Ad26 vaccine candidates showed improved protection compared to long synthetic peptide vaccines containing the same immunogens, the current standard in the field. Ad26 expressing seven neoantigens conjugated to the Herpes Viral Protein 22 (VP22) (Ad26.VP22.7Epi) was found to reduce tumor growth the most. Ad26.VP227Epi was compared to peptide vaccination in immune recall post challenge studies, in both the spleen and the tumor. Both Ad26.VP22.7Epi and Peptide showed a clear recall response after challenge in the spleen. This response was primarily focused on the Adpgk neoantigen but Ad26.VP22.7Epi additionally induced responses to the Irgq neoantigen while the peptide vaccine had stronger responses against the Reps1 neoantigen. The Ad26.VP227Epi vaccine generated increased numbers of infiltrating CD8+ T-cells, IFN+ CD8+ T-cells, and CD107a+ CD8+ T-cells, which all correlated with reduced tumor growth. We used single cell RNA-seq (scRNA-seq) to characterize tumor infiltrating lymphocyte populations (defined as CD45+) generated by the two platforms. We found that both Ad26.VP22.7Epi and Peptide induced higher numbers of infiltrating CD8+ T-cells within the tumor as compared to Sham. Single-cell analysis also showed higher numbers of tumor infiltrating Tregs in peptide vaccinated mice. Higher numbers of CD8+ T-cells correlated with protection as expected but interestingly increased numbers of Tregs positively correlated with tumor volume only among vaccinated mice, which could further explain differences between Ad vector and peptide vaccination. Differential gene expression analysis of CD8+ T-cells showed an upregulation of Th1 pathways and general T cell activation when comparing Ad vector vaccinated mice to peptide vaccinated. Peptide vaccinated CD8+ T-cells showed upregulation in the IL-10 pathway, potentially caused by an increased Tregs. Further investigation of these differences among CD8+ T cell and Treg populations could further elucidate potential mechanisms explaining the differences in platform efficacy. Citation Format: Gabriel Dagotto, Alessandro Colarusso, Robert Patio, David Li, Tochi Anioke, Victoria Giffin, Malika Aid, Dan Barouch. Neoantigen adenoviral cancer vaccine generates improved CD8+ T-cell responses compared to adjuvanted peptide vaccine [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr B004.

Variants of IL6, IL10, FCN2, RNASE3, IL12B and IL17B loci are associated with Schistosoma mansoni worm burden in the Albert Nile region of Uganda.

Individuals genetically susceptible to high schistosomiasis worm burden may contribute disproportionately to transmission and could be prioritized for control. Identifying genes involved may guide development of therapy. A cohort of 606 children aged 10-15 years were recruited in the Albert Nile region of Uganda and assessed for Schistosoma mansoni worm burden using the Up-Converting Particle Lateral Flow (UCP-LF) test detecting circulating anodic antigen (CAA), point-of-care Circulating Cathodic Antigen (POC-CCA) and Kato-Katz tests. Whole genome genotyping was conducted on 326 children comprising the top and bottom 25% of worm burden. Linear models were fitted to identify variants associated with worm burden in preselected candidate genes. Expression quantitative trait locus (eQTL) analysis was conducted for candidate genes with UCP-LF worm burden included as a covariate. Single Nucleotide Polymorphism loci associated with UCP-LF CAA included IL6 rs2066992 (OR = 0.43, p = 0.0006) and rs7793163 (OR = 2.0, p = 0.0007); IL21 SNP kgp513476 (OR 1.79, p = 0.0025) and IL17B SNP kgp708159 (OR = 0.35, p = 0.0028). A haplotype in the IL10 locus was associated with lower worm burden (OR = 0.53, p = 0.015) and overlapped SNPs rs1800896, rs1800871 and rs1800872. Significant haplotypes (p<0.05, overlapping significant SNP) associated with worm burden were observed in IL6 and the Th17 pathway IL12B and IL17B genes. There were significant eQTL in the IL6, IL5, IL21, IL25 and IFNG regions. Variants associated with S. mansoni worm burden were in IL6, FCN2, RNASE3, IL10, IL12B and IL17B gene loci. However only eQTL associations remained significant after Bonferroni correction. In summary, immune balance, pathogen recognition and Th17 pathways may play a role in modulating Schistosoma worm burden. Individuals carrying risk variants may be targeted first in allocation of control efforts to reduce the burden of schistosomiasis in the community.

Case report: Interleukin-17 targeted biological therapy in netherton syndrome

Netherton syndrome (NS) is rare and multisystemic congenital skin disorder classically distinguied as a triad of congenital ichthyosiform erythroderma, trichorrhexis invaginata (TI), and an atopic diathesis. Recent advances in pathogenesis have explored the role of IL-23/Th17 pathway in NS. Herein, we present a 17 years old girl harbouring homozygous four base pair deletion in exon 26 of the SPINK5 gene, presented with pruritus, scaling, dry skin and generalized eczematous lesions. She was administered anti IL17A (subcutaneous secukinumab) therapy. The treatment was well tolerated and resulted in a favourable clinical response, reduction of the IL17A gene expression and CD4 + Th17 cell population after 6 months which revealed an abrogation of Th17-skewing during therapy.

Defining the Drivers of Idelalisib-Related Early Autoimmune Toxicity in Chronic Lymphocytic Leukemia

Chronic lymphocytic leukemia (CLL) patients with disease resistant to BTK and BCL-2 inhibitors can potentially be treated with PI3K inhibitors to overcome resistance and improve outcomes. PI3K inhibitors such as idelalisib and duvelisib have shown promising efficacy in CLL patients, but severe toxicities remain a significant hurdle to their implementation in the clinic (PMID: 24615777). By investigating the molecular mechanisms that lead to severe early autoimmune toxicity with idelalisib, we hope to predict or prevent toxicity, thus expanding the use of this efficacious PI3K inhibitor class of drugs. To this end, the Brown lab showed that idelalisib-induced hepatitis is autoimmune. CyTOF analysis in CLL patients with early severe toxicity in a front-line idelalisib clinical trial cohort showed a significant decrease in CD4+ T regulatory cells (Treg) with a relative shift in favor of T effectors in peripheral blood. In addition, the Brown lab showed that the Th17 pathway is more active at baseline, with a further increase in patients with toxicity (PMID: 27247136, 36732326). Since the CYTOF data is restricted to fewer mostly T-cell markers in our studies, an unbiased single cell RNA sequencing (scRNA-seq) approach is warranted to explore the full effects of idelalisib on immune cells and CLL cells in patients with and without autoimmune toxicity. The question remains what molecular mechanisms drive the idelalisib-induced decrease in the Treg population, its effect on natural killer (NK) and CD8 T-cell function, and the activation of the Th17 pathway in CLL patients with early toxicity. We therefore carried out the first systematic effort to understand the mechanisms of idelalisib toxicity in molecular detail using combined scRNA and scTCR-seq. In this study, we profile T-cell enriched PBMCs of matched pre-treatment and on-idelalisib treatment samples from CLL patients with early severe toxicity and without any toxicity (n=4 CLL patients from each group, a total of 16 samples) using 10X single cell transcriptomic and TCR profiling. Raw sequencing data files were processed using the Cell Ranger suite (version 7.1.0) from 10x Genomics. Sample identification based on SNP information was done using the Popscle suite. Seurat package (version 4.9.9.9) was used for the single cell analysis. After filtering cells with fewer than 600 features, less than 1000 UMIs, mitochondrial content higher than 15%, 51,487 cells were left for the downstream analysis. To explore transcriptional heterogeneity and perform initial cell clustering, principal component analysis and nonlinear dimensional reduction using UMAP were applied using 20 dimensions. Leiden algorithm was used for cluster identification. Cell types were identified using Azimuth with the PBMC reference dataset and confirmed with expression plots. The initial observations of the cell proportions show that baseline samples from patients with early idelalisib toxicity have fewer Tregs, NK, and CD4+ cytotoxic T (CTL) cells compared to patients without toxicity. In addition, we observed a decline in Tregs, gamma-delta (γδ) T, and CD4+ CTL cells and a concurrent increase in proliferative CD4+ and CD8+ T-cells and proliferative NK on idelalisib treatment in the early toxicity group. On the contrary, CD4+ CTL and CD4+ T effector memory (TEM) cells are enriched with idelalisib therapy in patients without toxicities. At the same time, more CD4+ T-Cell Memory (TCM), γδ T, CD8+ naive T, CD8+ TCM, and Mucosal-associated invariant T (MAIT) cells are present in pretreatment samples from patients with toxicity compared to patients without toxicity. Furthermore, proliferative T-cells are depleted with idelalisib therapy in patients without toxicity whereas they are enriched in patients with early toxicity. To further characterize idelalisib-induced depletion of Tregs, we reclustered them to identify distinct subgroups and found particular clusters of Treg were depleted in patients with early toxicity. In the non-toxicity group, more diversity of Tregs was identified with depletion of some clusters but also an increase in a cluster. At present, we are integrating the scRNA &amp; scTCR-seq data to characterize the cellular diversity and heterogeneous phenotypic states of the T-cell populations and updated results unraveling molecular mechanisms that drive early severe toxicity in CLL patients with idelalisib treatment will be presented at the meeting.

New Enhancing MRI Lesions Associate with IL-17, Neutrophil Degranulation and Integrin Microparticles: Multi-Omics Combined with Frequent MRI in Multiple Sclerosis.

Blood-barrier (BBB) breakdown and active inflammation are hallmarks of relapsing multiple sclerosis (RMS), but the molecular events contributing to the development of new lesions are not well explored. Leaky endothelial junctions are associated with increased production of endothelial-derived extracellular microvesicles (EVs) and result in the entry of circulating immune cells into the brain. MRI with intravenous gadolinium (Gd) can visualize acute blood-barrier disruption as the initial event of the evolution of new lesions. Here, weekly MRI with Gd was combined with proteomics, multiplex immunoassay, and endothelial stress-optimized EV array to identify early markers related to BBB disruption. Five patients with RMS with no disease-modifying treatment were monitored weekly using high-resolution 3T MRI scanning with intravenous gadolinium (Gd) for 8 weeks. Patients were then divided into three groups (low, medium, or high MRI activity) defined by the number of new, total, and maximally enhancing Gd-enhancing lesions and the number of new FLAIR lesions. Plasma samples taken at each MRI were analyzed for protein biomarkers of inflammation by quantitative proteomics, and cytokines using multiplex immunoassays. EVs were characterized with an optimized endothelial stress EV array based on exosome surface protein markers for the detection of soluble secreted EVs. Proteomics analysis of plasma yielded quantitative information on 208 proteins at each patient time point (n = 40). We observed the highest number of unique dysregulated proteins (DEPs) and the highest functional enrichment in the low vs. high MRI activity comparison. Complement activation and complement/coagulation cascade were also strongly overrepresented in the low vs. high MRI activity comparison. Activation of the alternative complement pathway, pathways of blood coagulation, extracellular matrix organization, and the regulation of TLR and IGF transport were unique for the low vs. high MRI activity comparison as well, with these pathways being overrepresented in the patient with high MRI activity. Principal component analysis indicated the individuality of plasma profiles in patients. IL-17 was upregulated at all time points during 8 weeks in patients with high vs. low MRI activity. Hierarchical clustering of soluble markers in the plasma indicated that all four MRI outcomes clustered together with IL-17, IL-12p70, and IL-1β. MRI outcomes also showed clustering with EV markers CD62E/P, MIC A/B, ICAM-1, and CD42A. The combined cluster of these cytokines, EV markers, and MRI outcomes clustered also with IL-12p40 and IL-7. All four MRI outcomes correlated positively with levels of IL-17 (p < 0.001, respectively), and EV-ICAM-1 (p < 0.0003, respectively). IL-1β levels positively correlated with the number of new Gd-enhancing lesions (p < 0.01), new FLAIR lesions (p < 0.001), and total number of Gd-enhancing lesions (p < 0.05). IL-6 levels positively correlated with the number of new FLAIR lesions (p < 0.05). Random Forests and linear mixed models identified IL-17, CCL17/TARC, CCL3/MIP-1α, and TNF-α as composite biomarkers predicting new lesion evolution. Combination of serial frequent MRI with proteome, neuroinflammation markers, and protein array data of EVs enabled assessment of temporal changes in inflammation and endothelial dysfunction in RMS related to the evolution of new and enhancing lesions. Particularly, the Th17 pathway and IL-1β clustered and correlated with new lesions and Gd enhancement, indicating their importance in BBB disruption and initiating acute brain inflammation in MS. In addition to the Th17 pathway, abundant protein changes between MRI activity groups suggested the role of EVs and the coagulation system along with innate immune responses including acute phase proteins, complement components, and neutrophil degranulation.

Chemokine CCL19 promotes type 2 T-cell differentiation and allergic airway inflammation

Allergic asthma is driven largely by allergen-specific TH2 cells, which develop in regional lymph nodes on the interaction of naive CD4+ T cells with allergen-bearing dendritic cells that migrate from the lung. This migration event is dependent on CCR7 and its chemokine ligand, CCL21. However, is has been unclear whether the other CCR7 ligand, CCL19, has a role in allergic airway disease. This study sought to define the role of CCL19 in TH2 differentiation and allergic airway disease. Ccl19-deficient mice were studied in an animal model of allergic asthma. Dendritic cells or fibroblastic reticular cells from wild-type and Ccl19-deficient mice were cultured with naive CD4+ T cells, and cytokine production was measured by ELISA. Recombinant CCL19 was added to CD4+ T-cell cultures, and gene expression was assessed by RNA-sequencing and quantitative PCR. Transcription factor activation was assessed by flow cytometry. Lungs of Ccl19-deficient mice had less allergic airway inflammation, reduced airway hyperresponsiveness, and less IL-4 and IL-13 production compared with lungs of Ccl19-sufficient animals. Naive CD4+ T cells cocultured with Ccl19-deficient dendritic cells or fibroblastic reticular cells produced lower amounts of type 2 cytokines than did T cells cocultured with their wild-type counterparts. Recombinant CCL19 increased phosphorylation of STAT5 and induced expression of genes associated with TH2 cell and IL-2 signaling pathways. These results reveal a novel, TH2 cell-inducing function of CCL19 in allergic airway disease and suggest that strategies to block this pathway might help to reduce the incidence or severity of allergic asthma.

High expression of interleukin-17A in cutaneous sarcoidosis.

Increasing evidence of Th17 pathways is implicated in sarcoidosis granulomatous inflammation and maintenance. Our study evaluated the presence of IL-17A, a downstream proinflammatory cytokine, in cutaneous sarcoidosis biopsies. We found that all 26 biopsy samples had increased expression of IL-17A, predominantly located in macrophages within granulomas, in the cytoplasm of multinucleated histiocytes surrounding the granulomas and within lymphocytic infiltrates surrounding the inflammation.

Beyond CLL Progression: Unveiling Th17 and Th2 T Cell Subsets As Predictors of Pembrolizumab Response in Richter's Transformation

Despite notable advancements in treating chronic lymphocytic leukemia (CLL) with targeted therapies, a significant clinical challenge arises as a subset of CLL patients progress and develop Richter's transformation (RT), an aggressive form of lymphoma. While immune cell dysfunction and T cell defects are well-known characteristics of CLL, the alterations in immune cell subsets during CLL progression to RT remain largely unexplored. While CLL patients typically fail to respond to anti-PD1 pembrolizumab, a positive response has been observed in 20-40% of RT patients. Previously, our study involving three RT patients (1 responder and 2 progressors) revealed enriched regulatory T cells (Tregs) in the responders compared to progressors after pembrolizumab treatment (Iyer et al, ASH2020). Hence, we propose that dynamic changes in T cell subsets at baseline may be associated with response to pembrolizumab treatment. To this end, we performed single-cell RNA sequencing on sorted T cells (CD3 +) from PBMCs derived from 20 patients (responders (RES), n=5; stable disease (SD), n=5; progressive disease/progressors (PD), n=10). A total of 408,627 cells were captured, and an average of 1,393 genes were detected per cell per patient. Using a graph-based clustering approach (Seurat V4.3), we identified 19 clusters based on marker genes and transcriptional signatures and identified enriched pathways using Enrichr analysis. Our results showed that RES had an increased Th17 subset at baseline compared to PD (15% vs. 5%, p=0.01) and reduced Th2 T cells compared to SD (p=0.004) and PD (p=0.05). The Th17/Th2 ratio was higher in RES at baseline (Th17/Th2 R: 1.24, PD: 0.21, SD: 0.25, p0.05) and post-pembrolizumab (Th17/Th2 RES: 1.31, PD: 0.34, SD: 0.30, p0.05). Furthermore, RES had an increased fraction of proliferating CD4 T cells at baseline (4% RES vs. 0.4% SD and 0.4% PD, p=0.02) and post-treatment (1.9% RES vs. 0.8% PD and 0.34% SD, p=0.02). Although we observed an increasing trend in the fraction of Tregs in the RES at baseline [(7.6% RES vs.6.1% PD and 6.5% SD, p=0.4; ns) and post-treatment (10.3% RES vs. 8.3% PD and 5.8% SD, p= 0.2; ns] compared to PD and SD, the difference was not significant. Similarly, among CD8+ T cells, we observed an increasing trend in CD8+ effector T cell fraction in the responders at baseline (29.8% RES vs. 21% PD and 15.6% SD, p=0.9, ns) and post-treatment (30.1% RES vs. 21.2% PD and 19.9% SD, p=0.44, ns). However, the difference was not statistically significant. These findings highlight the dynamic changes in T cell subsets associated with pembrolizumab response. In terms of transcriptional changes in Tregs following pembrolizumab treatment among RES, PD, and SD, we observed dynamic changes on a few Treg marker genes, including TIGIT and TOX. TIGIT and TOX expression drastically decreased after treatment in RES (p &amp;lt; 0.0001), while expression did not change in PD and SD. There were no significant changes in the expression of FOXP3 or CD27 in RES, PD, and SD before and after the treatment. In CD8 effector T cells, GZMK, GZMA (granzymes), and PRF1 (perforin) are marker genes. We observed higher expression of GZMK and GZMA in RES (p &amp;lt;0.0001) at the baseline compared to SD and PD. In the proliferating CD4 T cells, we observed increased expression of IDH2 (Isocitrate dehydrogenase 2-key enzyme of TCA cycle) in RES at the baseline, as compared to PD and SD, indicating increased TCA cycle activity in the proliferating T cells. Pathway analysis of T cell clusters using the Enrichr database showed TNFA-IFNG-MYC targets are enriched in CD8 + cells, IL2-STAT5 signaling is upregulated in Th17, TREGs, and E2F cell cycle associated pathways enriched in CD4 + proliferation cluster. In conclusion, our study unveils significant baseline differences in CD4 subsets between RES and PD/SD subjects and dynamic changes in T cell subsets and transcriptional signatures. These findings offer potential predictors of pembrolizumab response in RT, such as increased Th17/Th2 ratio, enriched proliferating CD4 + T cells, and elevated TIGIT, TOX gene expression in Tregs, GZMK, and GZMA in CD8 effector cells. This deeper understanding of the immunological landscape in RT patients sheds light on the interplay between T cell subsets and treatment response in RT, paving the way for personalized therapeutic strategies. Further validation and exploration can lead to improved outcomes and tailored interventions for RT.

Investigation of Immunostimulatory Effects of IFN-γ Cytokine and CD40 Ligand Costimulatory Molecule for Development of HIV-1 Therapeutic Vaccine Candidate.

The most crucial disadvantage of DNA-based vaccines is their low immunogenicity; therefore, finding an effectual adjuvant is essential for their development. Herein, immunostimulatory effects of IFNγ cytokine and a CD40 ligand (CD40L) costimulatory molecule are evaluated as combined with an antigen, and also linked to an antigen in mice. For this purpose, after preparation of the HIV-1 Nef, IFNγ, and CD40L DNA constructs, and also their recombinant protein in an Escherichia coli expression system, nine groups of female BALB/c mice are immunized with different regimens of DNA constructs. About 3 weeks and also 3 months after the last injection, humoral and cellular immune responses are assessed in mice sera and splenocytes. Additionally, mice splenocytes are exposed to single-cycle replicable (SCR) HIV-1 virions for evaluating their potency in the secretion of cytokines in vitro. The data indicate that the linkage of IFNγ and CD40L to Nef antigen can significantly induce the Th-1 pathway and activate cytotoxic T lymphocytes compared to other regimens. Moreover, groups receiving the IFNγ-Nef and CD40L-Nef fusion DNA constructs show higher secretion of IFNγ and TNF-α from virion-infected lymphocytes than other groups. Therefore, the IFNγ-Nef and CD40L-Nef fusion DNA constructs are suggested to be a potential option for development of an efficient HIV-1 vaccine.

Donor-derived regulatory dendritic cell infusion modulates effector CD8+ T cell and NK cell responses after liver transplantation.

Immune cell-based therapies are promising strategies to facilitate immunosuppression withdrawal after organ transplantation. Regulatory dendritic cells (DCreg) are innate immune cells that down-regulate alloimmune responses in preclinical models. Here, we performed clinical monitoring and comprehensive assessment of peripheral and allograft tissue immune cell populations in DCreg-infused live-donor liver transplant (LDLT) recipients up to 12 months (M) after transplant. Thirteen patients were given a single infusion of donor-derived DCreg 1 week before transplant (STUDY) and were compared with 40 propensity-matched standard-of-care (SOC) patients. Donor-derived DCreg infusion was well tolerated in all STUDY patients. There were no differences in postoperative complications or biopsy-confirmed acute rejection compared with SOC patients up to 12M. DCreg administration was associated with lower frequencies of effector T-bet+Eomes+CD8+ T cells and CD16bright natural killer (NK) cells and an increase in putative tolerogenic CD141+CD163+ DCs compared with SOC at 12M. Antidonor proliferative capacity of interferon-γ+ (IFN-γ+) CD4+ and CD8+ T cells was lower compared with antithird party responses in STUDY participants, but not in SOC patients, at 12M. In addition, lower circulating concentrations of interleukin-12p40 (IL-12p40), IFN-γ, and CXCL10 were detected in STUDY participants compared with SOC patients at 12M. Analysis of 12M allograft biopsies revealed lower frequencies of graft-infiltrating CD8+ T cells, as well as attenuation of cytolytic TH1 effector genes and pathways among intragraft CD8+ T cells and NK cells, in DCreg-infused patients. These reductions may be conducive to reduced dependence on immunosuppressive drug therapy or immunosuppression withdrawal.

The selection of the initial drug in the treatment of severe psoriasis

Psoriasis is an immune-mediated skin disease associated with an increased risk of comorbidities and a significant negative impact on the quality of life of patients. In moderate and severe forms of psoriasis it is necessary to assign systemic therapies. The newest paradigm of treatment has become possible as a result of constant deepening of knowledge of pathophysiology of the disease. A clear mechanism is finally known down to the molecular level as to which cytokines are involved in the pathogenesis of psoriatic disease. Interleukin (IL)-23 mediates the activation of the Th17 pathway, which is hypothesised to be a major contributor to he inflammation observed in psoriasis, as proven, among other things, by the high efficacy of IL-23 inhibitor biological agents. It is obvious that great progress has been made in the field of genetically engineered biological therapy for psoriasis, both in terms of safety and efficacy. However, the issue of selecting a biologic drug individually in each patient is pressing, including in the case of initiation of the first genetically engineered biological drug in bionaive patients. The article provides an overview of the key points in the process of biological drug selection depending on the present comorbidities, and also describes a clinical case of successful therapy of a bionaive patient with concomitant depressive disorder in the anamnesis against the backdrop of a severe course of psoriasis. Successful use of IL-23 inhibitor (Guselkumab) allowed to achieve persistent remission and improve the quality of life, which in turn had a positive effect on the patient’s comorbid profile. This observation allows us to conclude that the use of guselkumab as the first biological agent is a rather effective, safe and promising option in the treatment of severe psoriasis.

Sclerotic-Type Cutaneous Chronic Graft-Versus-Host Disease Exhibits Activation of T Helper 1 and OX40 Cytokines

Sclerotic-type cutaneous chronic graft-versus-host disease is a severe complication of allogeneic hematopoietic stem cell transplantation, with profound morbidity. A dearth of effective, targeted treatment options necessitates further investigation into the molecular mechanisms underlying this T-cell-mediated disease. In this study, we compared the transcriptome in skin biopsies from pediatric and young adult (aged <25 years) patients with sclerotic-type cutaneous chronic graft-versus-host disease (n= 7) with that in demographically matched healthy controls (n= 8) and patients with atopic dermatitis (n= 10) using RNA sequencing with RT-PCR and immunohistochemistry validation. Differential expression was defined as fold change > 1.5 and false discovery rate < 0.05. Sclerotic-type cutaneous chronic graft-versus-host disease exhibited strong and significant T helper (Th)1 skewing through key related cytokines and chemokines (CXCL9/10/11, IFNG/IFN-γ, STAT1/signal transducer and activator of transcription 1). Several markers related to the TSLP-OX40 axis were significantly upregulated relative to those in both controls and lesional atopic dermatitis, including TNFSF4/OX40L, TSLP, and IL33, as well as fibroinflammatory signatures characterized in a prior study in systemic sclerosis. Gene set variation analysis reflected marker-level findings, showing the greatest enrichment of the Th1 and fibroinflammatory pathways, with no global activation identified in Th2 or Th17/Th22. Cell-type deconvolution revealed a significant representation of macrophages and vascular endothelial cells. Sclerotic-type cutaneous chronic graft-versus-host disease in young patients may therefore be characterized by strong Th1-related upregulation with a unique TSLP-OX40 signature, suggesting new therapeutic avenues for this devastating disease.