Cell Subsets In Peripheral Blood Research Articles

Circulatory age-associated B cells: Their distinct transcriptomic characteristics and clinical significance in drug-naïve patients with rheumatoid arthritis.

Age-associated B cells (ABCs) have been implicated in the pathogenesis of autoimmune diseases. However, the global gene expression and clinical significance of circulatory ABCs in rheumatoid arthritis (RA) remain poorly understood. Here, single-cell RNA sequencing identified nine B cell subsets in peripheral blood of RA patients, including ABCs. Increased phagocytosis and antigen presentation were functionally enriched by the genes expressed differentially in ABCs. Network analysis and in vitro experiments demonstrated SYK as a key regulator defining the myeloid-like phenotypes in ABCs. Flow cytometry showed that the proportion of ABCs correlated with RA activity and serum tumor necrosis factor-alpha level. Notably, ABCs above a cutoff threshold specifically distinguished RA from healthy controls and indicated higher disease activity. This study highlights the myeloid characteristics of circulatory ABCs regulated by SYK in RA. Increased ABCs may reflect disease activity and could serve as a potential biomarker in RA.

T-cell immune status in patients with acute exacerbation of chronic obstructive pulmonary disease: a case-control study

IntroductionImmune inflammatory response plays an important role in chronic obstructive pulmonary disease (COPD). However, the cellular immune status of patients with COPD at different phases is unclear. Herein, we aim to investigate the distribution and functional status of T cell subsets in different phases of COPD (acute exacerbation of COPD [AECOPD] and stable COPD [SCOPD]).MethodsThis is an observational case-control study undertaken in West China Hospital. The distribution of T cell subsets in peripheral blood of AECOPD, SCOPD, and healthy controls (HCs) was measured using multi-color flow cytometry, and the functional status was analyzed by additional staining of activation markers.ResultsA total of 43 HCs, 43 SCOPD patients, and 64 AECOPD patients were evaluated. The total number and percentage of lymphocytes and the CD4+/CD8+ T cells ratio were significantly lower in AECOPD patients when compared to HCs. HLA-DR expression in CD3+, CD4+, CD8+, CD8+ TCR aβ, and CD4+ TCR aβ T cells was upregulated in the AECOPD group. Similarly, the expressions of HLA-DR, CD57, and PD-1 were higher in T cell subsets in the AECOPD group. Compared with the SCOPD and HC groups, the AECOPD had a significantly lower proportion of CD4+CD27+CD28+ T cells, but opposite results were found for CD4+CD27-CD28- T cells. In addition, the proportion of CD4+CD39+ T cells and CD4+CD25+FoxP3+ T cells was significantly higher in the AECOPD and SCOPD groups when compared to the HC group (P < 0.05).DiscussionThe distribution of nearly half the T cell subsets in AECOPD patients was significantly different from that in SCOPD patients and HCs. AECOPD patients may have cellular immune suppression, immune dysfunction, abnormal activation, and higher senescence depletion of T cells.

Changes in and Potential Mechanisms of Circulating IgA+CD27-Class-Switched Memory B Cells in Patients With Allergic Rhinitis.

The role of memory B cells and their subgroups in allergic rhinitis (AR) and allergen immunotherapy (AIT) remains unclear. This study aimed to investigate the characteristics of memory B cells in the circulation of patients with AR and those undergoing AIT, as well as their clinical significance. This study involved a cohort comprising 32healthy control subjects, 39 individuals diagnosed with AR, and 31 AR patients who had received AIT for over one year. Visual analog scale (VAS) scores were used for symptom assessment, and the serum concentrations of immunoglobulins and cytokines were quantified. This study evaluated alterations in the proportions of peripheral blood memory B cells and their subpopulations, plasma cells, and various T-cell subsets across the three participant groups. The proportion of IgA+CD27- class-switched memory B cells in the AR group significantly decreased compared to the control group, but significantly increased following AIT (P < 0.05). In AR patients, circulating IgA+CD27- class-switched memory B cells were significantly positively correlated with Treg cells, IL-10, and IL-4 and significantly negatively correlated with IFN-γ, total IgE, sIgE, and VAS scores (P < 0.05). After AIT, the number of circulating IgA+CD27- class-switched memory B cells in AR patients was significantly positively correlated with the number of Treg cells and IL-10 and significantly negatively correlated with the VAS score (P < 0.05). The IgA+CD27- class-switched memory cell subset in human peripheral blood may serve as a potential biomarker for evaluating AR symptoms and treatment efficacy. Its mechanism may be associated with interactions between T and B cells.

Analysis of frequency changes in CD8+ regulatory T cell subsets in peripheral blood of individuals with type 1 diabetes

BackgroundThis study aims to examine the alterations and clinical significance of CD8+ regulatory T cell subsets in the peripheral blood of individuals with type 1 diabetes mellitus (T1DM).MethodsFrom January 2020 to December 2023, a study was conducted involving 40 individuals with T1DM, who visited the Department of Endocrinology at the First Affiliated Hospital of Nanjing Medical University (T1DM group). For comparison, 40 healthy individuals who underwent routine physical examinations at the same hospital during this period were selected as the control group. Peripheral blood mononuclear cells were isolated, and CD8+ T cells were labeled with CD3, CD25 and FoxP3 to analyze their subset frequencies using flow cytometry. The study examined differences in subset frequencies between the two groups and explored correlations between subset frequency, disease duration, and age of onset.ResultsThe frequencies of CD8+ CD25+, CD8+ FoxP3+, CD8+ CD25+ CD3+ and CD8+ FoxP3+ CD3+ subsets in peripheral blood mononuclear cells did not significantly differ between the healthy control group and the T1DM group (P > 0.05). In the T1DM group, the expression level of CD25 on CD8+ T cells showed no correlation with the age of onset or disease duration, and FoxP3 levels were also unrelated to the age of onset, with no statistical differences (P > 0.05). However, within the T1DM group, FoxP3 levels progressively decreased with longer disease duration, demonstrating a statistically significant negative correlation (Pearson r = -0.331, P < 0.05). In the T1DM group, the level of CD3+ CD8+ T cells expressed CD25, and there was no correlation between Foxp 3 level and age of onset, not statistically significant (P > 0.05), but the level of Foxp 3 in the T1DM group decreased with the duration of the disease, (Pearson r= − 0.363, P < 0.05).ConclusionThe levels of CD8+FoxP3+ regulatory T cells in peripheral blood mononuclear cells of patients with T1DM show a significant correlation with disease duration, suggesting that these cells may play a critical role in the progression of T1DM.

Alterations in peripheral T-cell and B-cell subsets in the ankylosing spondylitis patients with gut inflammation.

This study investigates changes in immune cell subsets in peripheral blood of ankylosing spondylitis (AS) patients with colitis or terminal ileitis. It aims to explore the connection between changes in lymphocyte subsets and gut inflammation, providing insights for early detection. Overall, 50 AS patients undergoing colonoscopy were enrolled. Flow cytometry was employed to analyze lymphocyte subsets, including T and B cells, in peripheral blood. Disease activity was assessed using CRP, ESR, BASDAI, ASDAS-CRP, and ASDAS-ESR. Compared to AS patients without gut inflammation, those with colorectal inflammation showed a significant increase in total T cells (p < .05), an increase in exhausted CD4+ T cells (p < .05), and a decrease in Th2 cells and total Tc cells (p < .05). Notably, in AS patients with terminal ileitis, there was an increase in total B cells and classic switched B cells (p < .05), with a decrease in double-positive T cells (p < .05). However, no significant differences were observed in the distribution of Tfh-cell subpopulations (Tfh1, Tfh2, Tfh17) and Tc-cell subpopulations (Tc1, Tc2, Tc17) between AS patients with either colorectal inflammation or terminal ileitis (p > .05). We explored the relationship between disease activity scores, ESR, CRP, and lymphocyte subsets, but found no statistically significant correlation between them. Distinct immune patterns may exist in AS with different types of intestinal inflammation. Colitis in AS is primarily characterized by a significant increase in exhausted CD4+ T cells, along with a decrease in Th2 cells. In contrast, terminal ileum inflammation in AS is marked by an increase in total B cells and classic switched B cells. These findings offer new insights for early detection and therapeutic intervention.

OMIP-069 version 2: Update to the 40-color full Spectrum flow cytometry panel for deep immunophenotyping of major cell subsets in human peripheral blood.

OMIP-069 version 2: Update to the 40-color full Spectrum flow cytometry panel for deep immunophenotyping of major cell subsets in human peripheral blood.

Non-Skewed X-inactivation Results in NF-κB Essential Modulator (NEMO) Δ-exon 5-autoinflammatory Syndrome (NEMO-NDAS) in a Female with Incontinentia Pigmenti

PurposeGenetic hypomorphic defects in X chromosomal IKBKG coding for the NF-κB essential modulator (NEMO) lead to ectodermal dysplasia and immunodeficiency in males and the skin disorder incontinentia pigmenti (IP) in females, respectively. NF-κB essential modulator (NEMO) Δ-exon 5-autoinflammatory syndrome (NEMO-NDAS) is a systemic autoinflammatory disease caused by alternative splicing and increased proportion of NEMO-Δex5. We investigated a female carrier presenting with IP and NEMO-NDAS due to non-skewed X-inactivation.MethodsIKBKG transcripts were quantified in peripheral blood mononuclear cells isolated from the patient, her mother, and healthy controls using RT-PCR and nanopore sequencing. Corresponding proteins were analyzed by western blotting and flow cytometry. Besides toll-like receptor (TLR) and tumor necrosis factor (TNF) signaling, the interferon signature, cytokine production and X-inactivation status were investigated.ResultsIP and autoinflammation with recurrent fever, oral ulcers, hepatitis, and neutropenia, but no immunodeficiency was observed in a female patient. Besides moderately reduced NEMO signaling function, type I interferonopathy, and elevated IL-18 and CXCL10 were found. She and her mother both carried the heterozygous variant c.613 C > T p.(Gln205*) in exon 5 of IKBKG previously reported in NEMO-deficient patients. However, X-inactivation was skewed in the mother, but not in the patient. Alternative splicing led to increased ratios of NEMO-Dex5 over full-length protein in peripheral blood cell subsets causing autoinflammation. Clinical symptoms partially resolved under treatment with TNF inhibitors.ConclusionNon-skewed X-inactivation can lead to NEMO-NDAS in females with IP carrying hypomorphic IKBKG variants due to alternative splicing and increased proportions of NEMO-∆ex5.

Reduced TRIM expression correlates with anomalous CD4 T cell activation in systemic lupus Erythematosus and its clinical diagnostic potential

ObjectiveThis study seeks to elucidate the expression, function, and clinical relevance of the T cell receptor interacting molecule (TRIM) within circulating CD4+T cell subsets in systemic lupus erythematosus (SLE) patients. MethodsWe assessed TRIM expression across distinct subpopulations of human peripheral blood mononuclear cells (PBMCs) through the analysis of publicly available single-cell RNA sequencing data. In addition, TRIM expression was investigated within CD4+T cell subsets of peripheral blood and spleens in mice. PBMCs were isolated from both SLE patients, healthy controls (HCs) and rheumatoid arthritis (RA) patients with subsequent measurement and comparative analysis of TRIM expression and functional molecules using flow cytometry. To gauge the clinical relevance of TRIM in SLE, correlation and ROC curve analyses were performed. ResultsIn both healthy humans and mice, TRIM was higher expressed within CD4+T cell subsets, especially in naive CD4+T cells. TRIM+ Tregs exhibited lower Helios+ cells and CD45RA-FoxP3hi cells percentages compared to TRIM- Treg cells. TRIM+T cells demonstrated reduced granzyme B and perforin secretion and increased IFN-γ secretion in comparison to TRIM- T cells. Notably, the proportion of TRIM+CD4+T cells was diminished in SLE patients. The downregulation of TRIM+ in CD4+T cells positively correlated with diminished complement C3 and C1q levels and inversely correlated with CRP. The identification of TRIM-associated CD4 T cell subsets aids in distinguishing SLE patients from HCs and those with RA. ConclusionsReduced TRIM expression is linked to abnormal CD4+T cell activation in SLE. TRIM-associated CD4+T cells may be implicated in the pathogenesis of SLE and hold potential for clinical diagnostic purposes.

148P Clinical significance of CD4+ T cell subsets in peripheral blood for anti-PD-1/PD-L1 therapy

148P Clinical significance of CD4+ T cell subsets in peripheral blood for anti-PD-1/PD-L1 therapy

The effects of postoperative targeted immunotherapy on peripheral blood cytokines and immune cell profile in lung cancer patients.

Cytokines and cell subsets are important components of the tumor microenvironment. Previous research has revealed that there are differences in cytokines and cell subsets in the peripheral blood of lung cancer (LCA) patients before and after eradication. The purpose of this study is to explore the monitoring value of cytokines and cellular subpopulations as biomarkers in post-immunotherapy monitoring of patients with LCA after surgery. We conducted a case-control study using double-antibody sandwich magnetic microsphere flow cytometry with immunofluorescence technology and fluorescent monoclonal antibody multiparameter flow cytometry to detect differences in peripheral blood cytokines and cell subsets between LCA patients after immunotherapy and healthy controls. Our research results show that there are differences in the levels of IL-4, IL-6, IL-10, IL-17, IFN-γ, TNF-α in the peripheral blood of LCA patients (n=70) after immunotherapy compared to the healthy controls (n=55) (P<0.05), and there are differences in 10 cell subgroups including DP T Cells, AT cells, and NLR in the peripheral blood compared to the healthy controls (n=35) (P<0.05). Further analysis revealed significant differences in the detection data of IL-6, IL-10, IFN-γ, CD56dim NK cells, Total B cells, Total NE cells, CD15+M cells, and NLR between LCA deceased patients (n=25) and LCA surviving patients (n=27) during the same period (P<0.05). The continuous monitoring of cytokines and cell subsets is far more valuable than a single-time test, as abnormal fluctuations in the data of cytokines and cell subsets are often associated with poor prognosis. In addition, IL-6 and NLR showed the strongest discriminative ability between postoperative immunotherapy-treated LCA patients and healthy controls, with AUC values of 0.840 and 0.822, respectively. There was a significant association between IFN-γ and distant metastasis in LCA (P<0.05), as well as between CD56dim NK cells and lymph node infiltration (P<0.05). This research results support peripheral blood cytokines and cell subsets as biomarkers for monitoring the postoperative immune status and predicting the prognosis of LCA patients after immunotherapy. The continuous monitoring of cytokines and cell subsets is far more valuable than a single-time detection.

Roginolisib, an oral, highly selective and allosteric modulator of phosphoinositide 3-kinase inhibitor delta (PI3Kδ) in patients with uveal melanoma and advanced cancers.

9597 Background: The highly selective, oral allosteric modulator of PI3Kδ, roginolisib (IOA-244), blocks the activity of PI3Kδ-dependent signaling, in both tumor cells and Tregs. Methods: Continuous daily dosing of roginolisib at 10, 20, 40 and 80 mg in patients (pts) with pretreated solid tumours and Follicular Lymphoma (FL) were evaluated (Part A). Part B consists of a dose confirmation cohort in uveal melanoma (UM) pts of 2 parts, B.1 and B.2. Primary objective: safety of the anticipated biologically effective dose (BED). Secondary objectives: PK; PD (e.g., inhibition of CD63 expression on basophils, changes in immune cell subsets in peripheral blood); RECIST 1.1 or Lugano-based responses; PFS and OS. Exploratory studies: phenotypic changes in circulating immune cells by mass cytometry (i.e., CyTOF); response assessments by radiomics; analysis of circulating protein signatures via Olink proteomics. Results: Part A Solid Tumour (completed): Sixteen pts were treated in 4 cohorts of 4 pts each. Pts characteristics: uveal melanoma (9/16; 56%), cutaneous melanoma (5/16; 31%) and pleural mesothelioma (2/16; 13%). Only Grade 1 and 2 AEs by CTCAE v.5 were observed, including 2 cases of transient (lasting &lt; 24 hrs) diarrhoea and 2 of AST/ALT elevation. Part A FL Cohort (completed): 8 pts were treated at 20 mg (n=4) and 80 mg (n=4). Grade 3 toxicities were transient and observed in 2/8 (25%) pts. 2/4 pts (50%) at the 80 mg had PR as per Lugano criteria. Treatment-emergent adverse events (TEAEs) did not result in study drug discontinuation, immune-related toxicity, or dose-limiting toxicity in either the solid tumor or hematological patients. UM pts (Part A and Part B)&gt;: 29 pts (Part A: 9 pts; Part B.1: 7 pts, Part B.2: 13 pts), of which 23 were treated at 80 mg QD (RP2D). Mean time on treatment: 10.7 mo (range: 1.5-39.6 mo). ORR (RECIST 1.1): PR: 1/29 (3%); SD: 21/29 (72%). Median OS for Part A: 20.8 mo (5/9 with 4 pts alive); Part B.1 20.1 (4/7 with 3 pts alive); Part B.2 not determined; 1-year OS rate: Part A 66.7%; Part B.1 71%; Part B.2 immature. Exploratory Investigations: At 3.7 mo (Cycle 5), there were 2 groups: one with SD (13/29; 45%) and the other with PD (16/29; 55%). At 6 mo, this split continued with 38% (11/29) showing SD and 62% having PD (18/29). Volumetric changes matched durable SDs while having higher sensitivity in the 15 analyzed patients. Interestingly, tumor burden reduction in mediastinal lymph nodes was indicative of SD and 70% of pts had volumetric spleen size reduction. Volumetric and radiomic changes were matched with peripheral changes in plasma proteins and immune cell changes. Conclusions: Roginolisib is well tolerated at 80 mg, demonstrates efficacy, and induces phenotypic changes in circulating immune cells. At the RP2D, the most robust anti-tumour activity was observed in FL (50% PR) and UM pts (3% PR; 72% SD). Clinical trial information: NCT04328844 .

Analysis of Immune Cell Subsets in Peripheral Blood from Patients with Engineered Stone Silica-Induced Lung Inflammation.

Silicosis caused by engineered stone (ES-silicosis) is an emerging worldwide issue characterized by inflammation and fibrosis in the lungs. To our knowledge, only a few reports have investigated leukocyte/lymphocyte subsets in ES-silicosis patients. The present study was designed to explore the proportions of the main lymphocyte subsets in ES-silicosis patients stratified into two groups, one with simple silicosis (SS) and the other with a more advanced state of the disease, defined as progressive massive fibrosis (PMF). The proportions of B (memory and plasmablasts) cells, T (helper, cytotoxic, regulatory) cells, and natural killer (NK) (regulatory and cytotoxic) cells were investigated by multiparameter flow cytometry in 91 ES-silicosis patients (53 SS patients and 38 PMF patients) and 22 healthy controls (HC). Although the total number of leukocytes did not differ between the groups studied, lymphopenia was observed in patients compared to healthy controls. Compared with those in healthy controls, the proportions of memory B cells, naïve helper T cells, and the CD4+/CD8+ T cells' ratio in the peripheral blood of patients with silicosis were significantly decreased, while the percentages of plasma cells, memory helper T cells, and regulatory T cells were significantly increased. For the NK cell subsets, no significant differences were found between the groups studied. These results revealed altered cellular immune processes in the peripheral blood of patients with ES-silicosis and provided further insight into silicosis pathogenesis.

Pan‑immune‑inflammation value as a novel prognostic biomarker in nasopharyngeal carcinoma.

The pan-immune-inflammation-value (PIV) is a comprehensive biomarker that integrates different peripheral blood cell subsets. The present study aimed to evaluate the prognostic ability of PIV in patients with nasopharyngeal carcinoma (NPC) undergoing chemoradiotherapy. PIV was assessed using the following equation: (Neutrophil count × platelet count × monocyte count)/lymphocyte count. The Kaplan-Meier method and Cox hazards regression models were used for survival analyses. The optimal cut-off values for PIV and systemic immune-inflammation index (SII) were determined using receiver operating characteristic analysis to be 428.0 and 1032.7, respectively. A total of 319 patients were recruited. Patients with a low baseline PIV (≤428.0) accounted for 69.9% (n=223) and patients with a high baseline PIV (>428.0) accounted for 30.1% (n=96). Compared with patients with low PIV, patients with a high PIV had significantly worse 5-year progression-free survival [PFS; 66.8 vs. 77.1%; hazard ratio (HR), 1.97; 95% confidence interval (CI), 1.22-3.23); P=0.005] and 5-year overall survival (OS; 68.7 vs. 86.9%, HR, 2.71; 95% CI, 1.45-5.03; P=0.001). PIV was also a significant independent prognostic indicator for OS (HR, 2.19; 95% CI, 1.16-4.12; P=0.016) and PFS (HR, 1.86; 95% CI, 1.14-3.04; P=0.013) and outperformed the SII in multivariate analysis. In conclusion, the PIV was a powerful predictor of survival outcomes and outperformed the SII in patients with NPC treated with chemoradiotherapy. Prospective validation of the PIV should be performed to better stratify radical treatment of patients with NPC.

Immune cell phenotype and function patterns across the life course in individuals from rural Uganda.

To determine the pattern of immune cell subsets across the life span in rural sub-Saharan Africa (SSA), and to set a reference standard for cell subsets amongst Africans, we characterised the major immune cell subsets in peripheral blood including T cells, B cells, monocytes, NK cells, neutrophils and eosinophils, in individuals aged 3 to 89 years from Uganda. Immune phenotypes were measured using both conventional flow cytometry in 72 individuals, and full spectrum flow cytometry in 80 individuals. Epstein-Barr virus (EBV) IFN-γ T cell responses were quantified in 332 individuals using an ELISpot assay. Full blood counts of all study participants were also obtained. The percentages of central memory (TCM) and senescent CD4+ and CD8+ T cell subsets, effector memory (TEM) CD8+ T cells and neutrophils increased with increasing age. On the other hand, the percentages of naïve T (TN) and B (BN) cells, atypical B cells (BA), total lymphocytes, eosinophils and basophils decreased with increasing age. There was no change in CD4+ or CD8+ T effector memory RA (TEMRA) cells, exhausted T cells, NK cells and monocytes with age. Higher eosinophil and basophil percentages were observed in males compared to females. T cell function as measured by IFN-γ responses to EBV increased with increasing age, peaking at 31-55 years. The percentages of cell subsets differ between individuals from SSA compared to those elsewhere, perhaps reflecting a different antigenic milieu. These results serve as a reference for normal values in this population.

Imbalance of Tfollicular helper cell subsets trigger the differentiation of pathogenic B cells in idiopathic membranous nephropathy.

This study aims to elucidate the role of T follicular helper (Tfh) cells and their subsets in idiopathic membranous nephropathy (IMN). The frequencies of Tfh cell subsets and B cell subsets in peripheral blood (PB) were detected in both IMN patients and healthy controls (HCs). The involvement of Tfh cells in the disease pathogenesis was examined by coculturing human Tfh cells with B cells. The dynamic changes of Tfh cells in PB or spleen were monitored in passive Heymann nephritis (PHN) rats. The frequencies of circulating Tfh (cTfh) cells, cTfh2 cells, and plasmablasts were enriched in the PB of patients with IMN. cTfh cells expressed higher ICOS, and lower BTLA than healthy counterparts. The frequency of ICOS + cTfh2 was associated with the severity of IMN, including 24h urine protein, IgG4 concentration and the IgG4: IgG ratio. Positive correlations were also observed between the frequency of cTfh2 cells with plasmablasts, serum IL-21 and IL-4 levels. Importantly, cTfh cells isolated from IMN patients were able to induce the differentiation of B cells to memory B cells (MBC) and plasmablasts, this process could be substantially attenuated by blocking the IL-21. Similar increases of ICOS + cTfh cells were also detected in spleen of PHN rats, concomitant with elevated urine protein levels. Collectively, our results demonstrate that the imbalance of cTfh cell subsets play a crucial pathogenic role in IMN by inducing the differentiation of B cells through IL-21, and cTfh2 cells might serve as useful markers to evaluate the progression of IMN.

Sofosbuvir (+) daclatasvir (+) ribavirin in Egyptian patients with hepatitis C virus: Therapeutic outcomes and the prognostic role of natural killer cells

BackgroundOne of the prominent causes of chronic liver disease worldwide is the hepatitis C virus (HCV). HCV believed that innate immunity contributes to a sustained virological response (SVR) to the treatment of Sofosbuvir (SOF) (+) Daclatasvir (DCV) (+) Ribavirin (RBV). This study aimed to evaluate the impact of SOF (+) DCV (+) RBV therapy and persistent HCV infection on the subset of natural killer cells (NK) in HCV genotype four patients from Egypt. Materials and MethodsOne hundred and ten patients with persistent HCV infections requiring SOF (+) DCV (+) RBV therapy were grouped, and a flow cytometry (FCM) study of the NK cell subset in peripheral blood was performed. The assessment was performed before and after three and/or six months of the cessation of viral suppression therapy when a patient had a long-term viral response (SVR). One hundred and ten volunteers from the National Liver Institute’s (NLI) blood bank were selected as controls. ResultsPatients with chronic HCV infection before therapy had considerably lower CD16+ and CD3− CD56+ cells than controls. Their levels increase during SOF (+) DCV (+) RBV therapy. In patients with SVR during treatment, CD16+ and CD3– CD56+ cells increased significantly compared to those who did not get SVR. Furthermore, CD56+ cells were significantly higher in patients with persistent infection before treatment than controls but diminished with the response to treatment. Conclusion: NK cell activation following SOF (+) DCV (+) RBV therapy and polarization to cytotoxicity occurred early in HCV antiviral therapy and was elevated in the respondents. Our data illustrated that establishing an inhibitory cytotoxic NK profile is related to therapeutic outcomes.

Repeated iv anti-CD20 treatment in multiple sclerosis: Long-term effects on peripheral immune cell subsets.

Repeated intravenous administration of anti-CD20 depleting monoclonal antibodies 6 months apart is among the highly effective treatment options in multiple sclerosis (MS). Here, we aimed to investigate peripheral immune cell subset depletion kinetics following either rituximab (RTX) or ocrelizumab (OCR) infusions in people with MS (pwMS). We studied pwMS treated de-novo with either RTX (n = 7) or OCR (n = 8). The examinations were scheduled before the initiation of anti-CD20 therapy and every 12 weeks for up to 15 months. Immunophenotyping of immune cell subsets in peripheral blood was performed by multiparametric fluorescence cytometry. A significant, persistent decrease of CD19+ B cells was observed already with the first anti-CD20 infusion (p < 0.0001). A significant proportional reduction of memory B cells within the B-cell pool was achieved only after two treatment cycles (p = 0.005). We observed a proportional increase of immature (p = 0.04) and naive B cells (p = 0.004), again only after the second treatment cycle. As for the peripheral T-cell pool, we observed a continuous proportional increase of memory T helper (TH) cells/central memory TH cells (p = 0.02/p = 0.008), while the number of regulatory T cells (Treg) decreased (p = 0.007). The percentage of B-cell dependent TH17.1 central memory cells dropped after the second treatment cycle (p = 0.02). No significant differences in the depletion kinetics between RTX and OCR were found. Peripheral immune cell profiling revealed more differentiated insights into the prompt and delayed immunological effects of repeated intravenous anti-CD20 treatment. The observation of proportional changes of some pathogenetically relevant immune cell subsets only after two infusion cycles deserves further attention.

Characterization of the immune cell profile in metastatic nasopharyngeal carcinoma treated with chemotherapy and immune checkpoint inhibitors.

Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus (EBV)-associated cancer, and immune checkpoint inhibitors (ICIs) have shown efficacy in its treatment. The combination of chemotherapy and ICIs represents a new trend in the standard care for metastatic NPC. In this study, we aim to clarify the immune cell profile and related prognostic factors in the ICI-based treatment of metastatic NPC. Programmed cell death ligand 1 (PD-L1) expression was measured in 81 metastatic tissue samples that had not received prior ICI treatment. The combined positive score (CPS) was positive in 58.0% of the samples, with a statistically significant correlation to median overall survival (OS) (CPS ≥ 1 vs. CPS < 1: 28 vs. 16 months, P = 0.004). For the combination treatment of metastatic NPC, 62 patients were enrolled in a retrospective analysis, yielding a median OS of 39.3 months. The objective response rate for this combination therapy was 71%, with a complete response rate of 45.2%. With a cutoff value of 4.8 for the pre-treated neutrophil-lymphocyte ratio (NLR) in peripheral blood (PB), the difference in median OS was statistically significant (P = 0.021). Thirty-seven patients received local treatment following the combination therapy of ICIs and chemotherapy, which provided additional survival benefits. Most hyper-responders exhibited a prolonged low NLR (< 3), a high total lymphocyte count, and an undetectable or stable EBV DNA load in PB during treatment. Peripheral blood mononuclear cells (PBMCs) from most patients receiving the combination treatment were rich in PD-1+CD8+ lymphocytes, which showed high expression of both IFN-γ and Granzyme B, demonstrating the ability to kill the EBV-positive NPC cell line and xenografts in vitro and in vivo. Responders also displayed increased levels of CD4+CD45RA-CCR7-CD28+CD57- cells (effector memory cell subset) in peripheral blood. These results indicate that in the context of combined chemotherapy and ICIs, high PD-L1 expression in pre-treated metastatic tumor tissue, a low NLR before treatment, a decrease in NLR after treatment, and local treatment can provide significant benefits for patients with metastatic NPC.

Roginolisib a Highly Selective Allosteric Modulator of the Phosphoinositide 3-Kinase Delta (PI3Kδ) in Patients with Refractory/Relapsed Follicular Lymphoma

Introduction PI3Kδ inhibitors are active in patients (pts) with lymphomas, but their clinical development has been hampered by toxicity (Brown et al, Lancet Oncol 2022). Roginolisib (IOA-244) exerts an allosteric modulation of the PI3Kδ C-terminus and functions as a non-ATP competitive PI3Kδ inhibitor, delivering a selective inhibition without off-target effects typically seen with other PI3K inhibitors (Johnson et al, Cancer Res Commun 2023). Using the recommended phase 2 dose (RP2D) of 80 mg QD established in solid tumors patients, patients with refractory/relapsed (r/r) follicular lymphoma (FL) were treated to investigate its safety profile in pts with hematologic malignancies (NCT04328844). Methods In Part A of the FiH dose escalation study, roginolisib was investigated at 20 and 80 mg QD in pts with r/r FL. Primary objective: safety of the RP2D established in solid tumor pts. Secondary objectives: PK; PD (e.g., changes in immune cell subsets in peripheral blood); Lugano-based responses; PFS and OS. Results In Part A for pts with r/r FL, there were two cohorts: (a) Cohort 1: 20 mg QD daily (4/4; 2 female, 2 male); (b) Cohort 2: 80 mg QD (4/4; 3 female, 1 male). There were no DLT observed and no dose modifications were needed. The mean time on treatment was 1.9 months (20 mg QD) and 4.3 months (80 mg QD), with one patient at the 80 mg receiving Roginolisib for 9 mo. More than 4 prior lines of treatment were in 3/8 pts. Transient (lasting less than 48 hrs) platelet reduction (G3) and AST/ALT elevation (G3) were observed in 2/8 pts, which improved while pts continued without a treatment intervention, such as dose modification or medical treatment with steroids. Lugano-based PRs were observed in 2/4 pts at the 80 mg QD dose and none at the 20 mg QD. At the 80 mg QD dose a third pt had a clinical response manifested by improvement in night sweats and fever, reduction in cervical palpable lymph nodes and relief from chronic pleural effusion. This clinical improvement lasted less than 5 weeks. While Overall Survival (OS) is still ongoing, the median has not been reached in this study and is currently at 14.3 mo. Subsequent to ending Roginolisib treatment, patients received at least one additional therapy (6/8), one patient was able to undergo autologous transplant with a complete metabolic response (CMR). Biomarker evaluation using flow cytometry showed low counts of T regulatory cells (Treg) in peripheral blood in all 8 r/r FL patients compared to solid tumor patients at baseline and throughout the treatment. Myeloid derived suppressor cells in peripheral blood remained unchanged during the course of treatment with Roginolisib. Conclusions The safety profile of roginolisib in pts with r/r FL appears to match that in pts with solid malignancies. In contrast to other PI3Kδ inhibitors, roginolisib is highly selective for binding to PI3Kδ, favoring an inactive confirmation of PI3Kd and functions as a non-ATP competitive inhibitor.

Baseline CD57 + and CD16 + NK-Cells Predict Treatment Outcome in Non-Transplant Eligible Newly Diagnosed Multiple Myeloma Patients Treated with Daratumumab-Ixazomib-Dexamethasone

Background Non-fit multiple myeloma (MM) patients have inferior survival, increased toxicity, and higher rates of treatment discontinuation compared with their fit peers. Effective and less toxic treatment strategies are therefore urgently needed for this vulnerable population. The HOVON-143 trial (EudraCT: 2016-002600-90) was specifically designed to assess the efficacy and safety of daratumumab-ixazomib-dexamethasone in non-transplant eligible (NTE), intermediate fit and frail newly diagnosed (ND)MM patients. Patients were treated with nine 28-day induction cycles comprising daratumumab 16 mg/kg intravenously (cycles 1-2: days 1, 8, 15, and 22; cycles 3-6: days 1 and 15; cycles 7-9: day 1), ixazomib 4 mg orally (days 1, 8, and 15) and dexamethasone orally (only on days that daratumumab was given; cycle 1-2: 20 mg; cycle 3-9: 10 mg), followed by 8-week maintenance cycles consisting of daratumumab (day 1) and ixazomib (days 1, 8, 15, 29, 36, 43) for two years or until disease progression. Aims Since daratumumab has well-known NK-cell-mediated effector mechanisms and T-cell-stimulating immunomodulatory effects, we monitored these cell subsets in bone marrow (BM) and peripheral blood (PB) samples obtained before and during treatment. We specifically aimed to better understand these cell subsets related to frailty status, their predictive value for treatment outcome, and their dynamics during treatment. Methods 126 NDMM patients, among which 64 intermediate fit and 62 frail, were monitored. BM was collected at study-entry (baseline) and at disease progression; PB was collected at baseline, before cycle 4, before maintenance, and at disease progression. Absolute numbers of total T-cells and NK-cells (in PB) or percentages of T-cell and NK-cell subsets (in PB and BM) were profiled by flow cytometry. Data were analyzed using FlowSOM and UMAP. Total cells and cell subsets at baseline were compared between intermediate fit and frail patients, between responders (≥ partial response) and non-responders (&amp;lt; partial response), and associated with progression-free survival (PFS) and overall survival (OS). Results At baseline, we found no difference in absolute numbers of total T-cells and NK-cells between intermediate fit and frail patients. Besides, NK-cell phenotype was similar between both groups. However, within the T-cell compartment frail patients had a decreased CD4 +:CD8 + T-cell ratio, reduced naïve CD4 + and CD8 + T cells, and increased PD1 + CD4 + T-cells compared with intermediate fit patients in both BM and PB, independent of age. Interestingly, differences in T-cell phenotype were more pronounced in female than in male patients. Response data were available for 124 patients, of which 92 (74%) responded to treatment. In both BM and PB, preliminary results showed no clear association between response and absolute numbers of total T-cells or certain T-cell phenotypes at baseline. However, responders had higher absolute numbers of total NK-cells and higher percentages of NK-cells positive for maturation marker CD57 or fc-gamma receptor CD16 than non-responders, while higher absolute numbers of total NK-cells and higher percentages of CD57 + NK-cells were also associated with superior PFS and OS (Figure 1). Conversely, higher proportions of baseline NK-cells positive for inhibitory markers TIGIT and PD1 were associated with non-response, while higher percentages of CD56 bright and PD1 + NK-cells were negatively associated with PFS and OS. As expected, absolute numbers of total NK-cells markedly declined during the treatment course. Interestingly, percentages of CD57 + and CD16 + NK-cells decreased during treatment and remained low at disease progression, while percentages of CD56 bright and NKG2A + NK-cells substantially increased compared with baseline. Conclusions Frail patients had a distinct T-cell profile compared with intermediate fit patients, while NK-cell profiles were similar. Treatment outcome of NTE-NDMM patients treated with daratumumab-ixazomib-dexamethasone was positively associated with higher absolute numbers of total NK-cells and higher percentages of CD57 + and CD16 + NK-cells at baseline, corresponding to a mature and cytotoxic phenotype capable of mediating potent antibody-dependent cellular cytotoxicity. This study highlights the importance of NK-cell phenotype for the effectivity of daratumumab-based combination therapies in NTE-NDMM.