Peripheral Counts Research Articles

Blood T cell phenotypes correlate with fatigue severity in post-acute sequelae of COVID-19.

Post-acute sequelae of COVID-19 (PASC) affect approximately 10% of convalescent patients. The spectrum of symptoms is broad and heterogeneous with fatigue being the most often reported sequela. Easily accessible blood biomarkers to determine PASC severity are lacking. Thus, our study aimed to correlate immune phenotypes with PASC across the severity spectrum of COVID-19. A total of 176 originally immunonaïve, convalescent COVID-19 patients from a prospective cohort during the first pandemic phase were stratified by initial disease severity and underwent clinical, psychosocial, and immune phenotyping around 10weeks after first COVID-19 symptoms. COVID-19-associated fatigue dynamics were assessed and related to clinical and immune phenotypes. Fatigue and severe fatigue were commonly reported irrespective of initial COVID-19 severity or organ-specific PASC. A clinically relevant increase in fatigue severity after COVID-19 was detected in all groups. Neutralizing antibody titers were higher in patients with severe acute disease, but no association was found between antibody titers and PASC. While absolute peripheral blood immune cell counts in originally immunonaïve PASC patients did not differ from unexposed controls, peripheral CD3+CD4+ T cell counts were independently correlated with fatigue severity across all strata in multivariable analysis. Patients were at similar risk of self-reported PASC irrespective of initial disease severity. The independent correlation between fatigue severity and blood T cell phenotypes indicates a possible role of CD4+ T cells in the pathogenesis of post-COVID-19 fatigue, which might serve as a blood biomarker.

Relative Monocytosis Is Predictive of Disease Progression and Confirmatory of Response Assessment in Langerhans Cell Histiocytosis

Introduction Langerhans cell histiocytosis (LCH) is a clonal histiocytic neoplasm, which arises when myeloid precursors expressing CD207 (Langerin) and CD1a expand and circulate as mature dendritic cells, clinically manifesting in multisystemic eosinophilic granulomas at osseous sites (notably the cranium), lungs, cutaneous tissue, and visceral structures (pituitary gland, liver, spleen). Molecular alterations in MAP kinase signaling are common, specifically in BRAFV600E, which is identified in greater than 50% of patients. Limited LCH is commonly managed with local therapies, such as surgery and radiation, while disseminated disease is managed with systemic therapies, including multi-agent chemotherapy or targeted agents, such as BRAF inhibitors. While diagnostic and management paradigms are well-established in LCH, clinical studies evaluating indicators of disease progression are lacking. Methods Clinical, laboratory, radiographic and treatment data for 49 patients with a histopathologic diagnosis of Langerhans Cell Histiocytosis treated at a single institution over a 20-year period was reviewed by multiple study investigators. Specific parameters reviewed included biological sex, age at and time to diagnosis, molecular diagnostic data by immunohistochemistry and/or next generation sequencing, diagnostic imaging utilized, treatment rendered, clinical and/or radiographic response. Radiography in all but 1/49 patients was used to assess treatment responses, by RECIST 1.1 where MRI, or CT was utilized and PERCIST for serial PET imaging. The peripheral blood absolute monocyte count (in K/µL) was also assessed in each patient at various timepoints, including at initial diagnosis and the most recent value. We also performed a more focused assessment of absolute monocytes at the time of disease progression (when applicable) and again following reinstitution of therapy. Statistical analyses to compare sets of data were conducted using unpaired two-sided t-tests. Results Of the 49 patients included in this analysis, 28 were female and 21 male, with an average age at diagnosis of approximately 38 years; time between symptom onset and diagnosis varied widely across the cohort but was generally less than 12 months. Molecular diagnostics were employed in only 8/49 patients, 3 by IHC, 4 by NGS and 1 by both; BRAFV600E was assessed in 6 patients and was positive in 2. Approach to imaging at diagnosis and throughout treatment varied but most commonly included conventional CT (25), followed by MRI (17) and PET (16). LCH was observed as a primary pulmonary disease in 18 patients (36.7%), as involvement of skull/CNS/ocular structures in 10 and as localized cutaneous involvement in 5. Complete responses were observed in 17 of 49 patients, 11/17 of whom underwent local therapy (radiation and/or surgery) at ≥1 point in his or her treatment course. On the other end of the spectrum, 5 patients never achieved any response and always had progressive/persistent disease, all of whom had been diagnosed with pulmonary LCH in the setting of chronic tobacco use. Peripheral blood absolute monocyte count between the earliest-assessed timepoint and latest value, lastly, did not differ. The mean value on progression (0.94 K/µL), however, was significantly higher than that following re-institution of therapy (0.31, p=0.0005) as depicted in Figure 1. Discussion The identification of relative monocytosis at the time of LCH progression was an unexpected finding in this study. LCH pathogenesis has been felt to berelated to dysregulated hematopoiesis and acquisition of somatic mutations during monocyte differentiation, ultimately driving clonal expansion of CD1a+/CD207+ histiocytes. Within this working model of disruption to monocyte differentiation, one explanation of our observed findings is a simple expansion in circulating monocytes as a manifestation of disease progressing throughout various tissues. Another possibility is that circulating monocytes are upregulated and later differentiate into mature dendritic cells for MHC Class I presentation. Similar observations have been made at our institution in cohorts of non-LCH histiocytosis, specifically Rosai Dorfman and Erdheim Chester Disease. Expanded integrated genomic-transcriptomic sequencing analyses to evaluate the molecular character of LCH and ultimately clarify the origin of this monocyte trend are currently underway.

Factors Influencing Lymphocyte Collection Efficiency for the Manufacture of CAR-T Cells in Adults with B Cell Malignancies , a Single Center Experience

Aim: During leukapheresis, accurate estimation of the proportion of T cells that are removed from peripheral blood (collection efficiency, CE) is essential to minimise apheresis time, reduce lab storage requirements and prevent excess product collection. Based on historical precedent from autologous HPC donors, many centres estimate a CE of 40% for autologous T cell donors, although data are lacking. We aimed to identify patient characteristics and collection parameters that might influence leukapheresis CE in order to maximise efficiency and minimize waste during CAR T manufacture. Methods: We reviewed data from 33 consecutive patients who underwent leukapheresis for CAR-T cell treatment at our centre from March 2020 to October 2022. Apheresis was performed on the Spectra Optia, using the CMNC collection protocol. Apheresis were performed on the Spectra Optia, using the CMNC collection protocol. peripheral collection was used if patients had adequate venous access, otherwise central venous access was used. Peripheral blood flow cytometry was done prior to apheresis. Results: 33 apheresis procedures were documented from patients with 3% acute lymphocytic leukemia (ALL) (n=1), 82% non-Hodgkin lymphoma (NHL) (n=27) and 15% Multiple myeloma (n=5). Median age was 62 (18 -76 ) , Male to Female ratio was 1.38 , the median weight was 75 kg (45-136) the median of prior lines of therapy was 2 (1-7) with 33% of pts receiving prior autologous HCT and 3% Prior allogeneic HCT. The median time from diagnosis to collection was 15 months (3-109) The median WBC value was 6× 10 9/L (2-24), median Hematocrit was 0,34% (0,25-0,45), Median Neutrophil was 4,43×10 9/L (1-21), Monocytes 0,62 ×10 9/L (0,36-1,43), Platelets 185× 10 9/L (94-385), and median absolute Lymphocytes count was 0.8 (0.21-2,4). With median CD3 value was 560×10 9/L and a median percentage of ALC of 76%. CD4 and CD8 absolute median values were 276 and 309 respectively. The median CRP, LDH and Ferritinemia were 29,238 and 252 respectively. The collection was successful in all patients with 32 (97%) having a high CE of >40%. The median CD3 lymphocyte count in the apheresis product was 4,3 × 10 9 with an overall viability of 99% . The median collection efficiency was 71% (31-96) . Median Run time was 210 minutes, total blood volume and apheresis product volume were 9750 and 278ml. In the apheresis product, median CD3 value was 15415 ×10 6 and the median CD3 value per kg was 58×10 6 /kg. In multivariate analysis, baseline characteristics did not significantly impact the collection efficiency. ALC and CD3 pre-apheresis strongly predicted the CD3 yield after apheresis (p< 0,001). No major adverse events were recorded. Conclusion: High efficiency leukapheresis is safe and feasible in autologous T-cell donors. Our data also confirms that high ALC and peripheral blood CD3 counts pre-apheresis are strongly correlated to the CD3 yield in the product. Our centre now uses a CE estimate of 60% for collection of CAR T starting material.

Poor Hematopoietic Reserve at the Time of CD19 CAR T-Cell Infusion Is Associated with Long Term B-Cell Aplasia

Background: On-target off-tumor toxicities of anti-CD19 CAR T-cell therapy include B-cell aplasia and hypogammaglobulinemia. Long term B-cell aplasia is hypothesized to represent functional CAR T-cell engraftment (Melenhorst, Nature 2022), while peripheral blood B-cell recovery represents the loss of functional CAR-T cells against target over time. Patients with large B-cell lymphoma (LBCL) who are clinical responders have a 50-70% chance of B-cell recovery in the first year, with the remainder experiencing longer term B-cell aplasia (Logue, Haematologica 2021). Here, we sought to identify factors associated with long term B-cell aplasia using a cohort of responding patients who received CAR T-cell therapy for LBCL. Methods:This retrospective cohort study included 57 patients with LBCL treated with CD19-targeted CAR T-cell therapy between June 2016 and August 2020 at Moffitt Cancer Center who exhibited complete or partial response at six months after CAR T-cell infusion. Patients were treated with with axicabtagene ciloleucel (axi-cel; n=50) or tisagenlecleucel (tisa-cel; n=7), either as standard-of-care therapy (n=47) or as part of a previously published clinical trial (n=10; NCT02348216, NCT03391466, NCT03153462). Peripheral blood B-cells were measured by flow cytometry using CD19 expression as part of routine clinical care at baseline and periodically after infusion. Prolonged B-cell aplasia was defined as a CD19+ B cell count comprising less than 1% of peripheral blood mononuclear cells during a minimum of two separate time points beyond 6 months of follow-up. Patients were defined as having B cell recovery if >1% B-cells were observed in peripheral blood at any time after CAR T-cell infusion. Results: At a median follow up of 33 months (95% CI: 28.7, 37.3 months), 29 (51%) patients had persistent B-cell aplasia and 28 (49%) had recovery of peripheral B-cell counts. Median B-cell count at 12 months in the recovery group was 84/µL (IQR 2.5-197) compared to 0/µL (IQR 0-1) in the aplasia group. Patients with long-term B-cell aplasia had lower CD4 T-cell counts prior to CAR T-cell infusion [median 200/µL (IQR 136-278) vs. 330/µL (IQR 224-451); P= 0.008] and over time compared to patients with B-cell recovery (Fig. A). Patients with long-term B cell aplasia also had lower pre-infusion absolute neutrophil counts (2.3K/µL (IQR 1.4-4.15) vs. 3.5K/µL (IQR 2.8-4.2); P<0.001) and higher baseline CAR-HEMATOTOX scores [2 (IQR 1-3.25) vs 1 (IQR 0-1)], consistent with poor baseline hematopoietic reserve (Rejeski, Blood 2021). While disease-related mortality did not significantly differ between groups, non-relapse mortality was markedly higher in patients with long-term B-cell aplasia due to infection: all 8 patient deaths in the aplasia group were due to infections occurring beyond day 30 post-infusion, with 6 of the 8 deaths due to COVID-19. However, no patients in the B-cell recovery group died of late infection (Fig. B). Conclusions: Long term B-cell aplasia is hypothesized to represent ongoing CAR T-cell activity against normal B-cells. However, our results demonstrate that B-cell aplasia is associated with low baseline hematopoietic reserve and subsequent poor immune reconstitution of multiple lineages including B-cells, T-cells, and myeloid cells. Further studies are warranted to assess whether poor immune reconstitution and B-cell aplasia are related to long term CAR T-cell activity or other processes. These patients are at higher risk for non-relapse mortality from infection, and mitigation strategies are needed.

Immune Responses after One Versus Two Influenza A/B Vaccinations in Patients with Multiple Myeloma

Background: Multiple Myeloma (MM) is a hematological cancer affecting plasma cells, suppressing the immune system and increasing the risk of severe Influenza A/B infection. International guidelines recommend Influenza A/B vaccination on an annual basis. Data from the pre-lenalidomide era highlighted poor vaccination responses, with sufficient seroconversion in 39% of patients with MM after single vaccination and an increase to 55% after additional boosting. Lenalidomide is now widely established in different (maintenance) therapy protocols and other novel agents (e.g. anti-CD38-agents) are incorporated into the therapy lines. Hence, it is of high relevance to characterize the sufficient immune response to Influenza A/B vaccination in this context, evaluate the necessity for a prime and boosting approach, and identify patients with high risk of insufficient response and subsequent hazard of severe disease. Methods: In this retrospective single-center study, we analyzed the serological response from 71 patients with MM treated at our institution who received the annual Influenza A/B vaccination (Influvac Tetra, Mylan, Troisdorf, Germany) in the 2019/20 season. All patients received at least one vaccination with 52 patients receiving a second vaccination. Hemagglutination-inhibition (HI) assay titers for the five most prevalent virus strains were measured before, 21-28 days after the first and 21-28 days after the second vaccination or at observation (42-56 days after first vaccination in case of single vaccination). Peripheral immune cell counts were measured at the initial vaccination event by flow cytometry. Disease, therapy, and patient characteristics were retrieved from the electronical health care system at our institution. Primary endpoint was the sufficient responder status as indicated by ≥1:40 HI titer or an at least four-fold increase in the median HI titers of all strains. All patients declared written informed consent and the study was approved by the local ethics committee. Results: 71 patients with MM were included in the present study who mostly had IgG MM (46.5%), the minority displayed genetic high-risk MM (25.4%), 64.8% were in their first line of therapy and 69% were in good remission (CR/VGPR). Overall, only 5.6% (n=4) showed sufficient titers before vaccination. 63.3% (n=45) achieved a sufficient serological response after one or two Influenza A/B vaccinations. A strong increase in HI titers against all strains was observed after already the first vaccination. In patients receiving a prime and boost approach, significantly higher titers but no significant increase in responder rates were observed after the boost vaccination. Responders were more frequently without current therapy or under maintenance therapy, showed significantly higher rates of CR/VGPR and significantly less immunoparesis. Interestingly, they trended to lower rates of previous anti-CD38 therapy (24.4% versus 50%, p=0.053). Non-responders displayed significantly lower median CD19 + B-cell counts (20.0/µl versus 79.0/µl p=0.001) and reduced median CD4 + T-cell counts (167.5/µl versus 274.0/µl, p=0.033). Multivariate regression analysis highlighted current CR/VGPR (OR 4.1 [95% CI 1.09-16.57], p=0.04) and no immunoparesis (OR 5.37 [95% CI 1.13-33.21], p=0.046) at timepoint of vaccination as independent predictors for sufficient serological response. Those patients achieving a sufficient responder status only after the boost vaccination among all responders displayed significantly shorter time (5.0 versus 26.5 months, p=0.046) since the last high-dose chemotherapy and autologous stem cell transplantation (autoHSCT). Conclusions: Together, 63.3% of patients with MM achieved a sufficient serological response after tetravalent Influenza A/B vaccination without a significant general benefit for a prime and boost approach. Current therapy, uncontrolled MM, present immunoparesis and low CD19 + B-cell counts were associated with inferior response rates. Those patients exceptionally benefitting from a boost vaccination were characterized by recent autoHSCT. Hence, annual one-time Influenza A/B vaccination is applicable for patients with MM and patients directly after autoHSCT can be considered for a prime and boost approach. Further studies are warranted to investigate this in a prospective and randomized fashion.

Phase 1/2 First-in-Human Study of the Menin-MLL Inhibitor DSP-5336 in Patients with Relapsed or Refractory Acute Leukemia

Introduction: Menin inhibitors are an exciting new class of agents in development for patients (pts) with acute leukemia who overexpress HOXA9 and MEIS1 genes. Although there is a growing list of fusion genes in acute leukemia that appear to be dependent on homeobox-related gene activity, menin inhibition has been shown to be particularly active in pts with NPM1 mutations (NPM1c) and KMT2A rearrangements (MLLr). DSP-5336 is an investigational, oral small molecule designed to inhibit the menin and MLL protein interaction. Methods: A Phase 1/2 study of DSP-5336 is being conducted in pts with relapsed or refractory (R/R) acute leukemia. The dose escalation portion of the study consists of two parallel arms (Arm A: without concomitant anti-fungal azole therapy; Arm B: with concomitant azole therapy). Pts were eligible with R/R AML, ALL or acute leukemia of ambiguous lineage without a limit on number of prior therapies, with a focus on those with MLLr and NPM1c. Results: Accrual is ongoing with 24 pts enrolled as of April 2023; 14 pts in Arm A at doses of 40 mg BID (n=2), 80 mg BID (n=4), 100 mg BID (n=2) and 120 mg BID (n=6), and 10 pts in Arm B at doses of 40 mg BID (n=4) and 60 mg BID (n=6). The median age was 67.0 years (range 20 - 85) and 62.5% were male. All pts had AML except for 1 pt with acute leukemia of ambiguous lineage. Pts had received a median of 3 (range 1 to 9) prior treatments, and 6 (25%) had received prior alloSCT. MLLr or NPM1c were present in 6 (25%) and 4 (17%) patients, respectively. No dose-limiting toxicities have been observed thus far. Most treatment-emergent adverse events (TEAEs) were grade 1/2. TEAEs assessed as related to DSP-5336 in ≥ 3 pts were nausea (Grade 1 or 2, n = 3 pts) and vomiting (Grade 1 or 2, n = 4 pts). Grade >3 TEAEs occurring in ≥ 3 pts included anemia (6 pts), pneumonia (5 pts), sepsis (3 pts) and hypokalemia (3 pts), all unrelated to study drug. A possible Grade 4 differentiation syndrome (DS) was observed in 1 pt with a TP53 mutation in Arm A receiving 80 mg BID although concurrent pneumonia and sepsis made a clear attribution difficult. No other DS events of any grade have been reported to date. No cardiac toxicities, and no QTc prolongation of any grade related to DSP-5336 have been observed. Out of the 6 pts enrolled with MLLr, 4 were treated at dose levels projected to be active based on preclinical modeling (≥ 60 mg BID with azoles, or ≥ 120 mg BID without azoles). Of these 4 pts, all had received prior intensive induction chemotherapy as well as a venetoclax-based regimen, and 3 had received prior allo-transplant. Of these 4 pts, 1 achieved CRi with a duration of therapy of 5.1 months, 1 achieved MLFS with a duration of therapy of 6.2 months (ongoing), and 1 achieved SD with clearance of peripheral blasts, recovery of peripheral counts, resolution of leukemic gingival infiltration, and reduction in bone marrow (BM) blasts from 85% to 31%. Of the 4 enrolled pts with NPM1c, 2 were treated at doses projected to be effective. Stable disease with complete clearance of peripheral blasts were observed in both pts, and BM blasts were reduced by 66% and 83% respectively. Preliminary pharmacokinetic (PK) analysis has demonstrated generally higher mean exposures at higher dose levels. Across all the dose levels, plasma t 1/2 ranged from 2-6 hours in Arm A and from 2-10 hours in Arm B, with T max reached within 2 hours. Little to no drug accumulation was observed with repeat dosing in both arms. Data to date suggests that azoles may not have a significant effect on DSP-5336 exposure, but enrollment and comparative analyses are ongoing. Target pharmacodynamic changes have been achieved in pts who have MLLr or NPM1m, including rapid decreases in HOXA9, MEIS1, and PBX3 with treatment and, conversely, increases in CD11b compared to pretreatment. Conclusion: DSP-5336 has been well tolerated with no DLTs to date in heavily pretreated R/R AML patients with NPM1c and MLLr AML. Importantly, no cardiac signals (including no QTcF prolongation) have been observed. PK studies have not identified a significant drug-drug interaction with azoles. Although there are early signs of clinical activity and pharmacodynamic changes during administration of DSP-5336, the study is ongoing to determine a recommended Phase 2 dose for single-agent expansion and potential combination regimens. Updated safety and efficacy data will be presented at the meeting.

Cell Count and Cell Density Decrease as Age Increases in Cadaveric Pediatric Medial Menisci

To examine the histologic changes in terms of cellularity, cell density, and nuclear shape in medial meniscal cellularity during skeletal development using pediatric cadaver specimens. Medial menisci from 26 pediatric cadavers, 11 female and 15 male (total 36 menisci), were obtained from tissue bank. Mean age of female donors was 34 months (1-108 months) and of male donors was 52 months (1-132 months). Menisci were processed and embedded in paraffin blocks. Each tissue block containing 6 representative areas of meniscus (anterior root, anterior horn, body [n= 2], posterior horn, and posterior root) was sectioned at 4 microns and stained with hematoxylin and eosin for evaluation of chondrocyte nuclei. Each of the 6 representative areas was imaged at 10×; one image on peripheral one-third of section, the second image on central two-thirds of the section. FIJI imaging software was used to measure cell count, cell density, and nuclear morphology (1= perfect circle). Data analysis included linear mixed models, Type II analysis of variance tests, and pairwise tests with the Tukey correction to assess statistical significance. Peripheral meniscus was more cellular than central meniscus. The cell count was found to decrease by 14% per year of age. Peripheral cell count decreased at a rate similar to the cell count in the central meniscus. Meniscal cell density was 2× higher peripherally than centrally. Overall average cell density in all locations in the menisci decreased by an average of 14% per year of age. The results of this study reveal decreases in cell count, cell density, and circularity as age increases in cadaveric pediatric medial menisci. To better understand the development of pediatric menisci at a cellular level and use this knowledge in the future on how to maintain the menisci in a younger, healthier state.

A Mendelian randomization-based approach to explore the relationship between leukocyte counts and breast cancer risk in European ethnic groups

Exploring the potential association between peripheral blood leukocyte counts and breast cancer risk by Mendelian randomization (MR) analysis methods. Genetic data related to peripheral blood sorting counts of leukocytes were collected from a genome-wide association study by Blood Cell Consortium (BCX). Single nucleotide polymorphic loci predicting peripheral blood sorting counts of these leukocytes were selected as instrumental variables according to the correlation assumption, independence assumption and exclusivity assumption of MR. The data on breast cancer and its subtypes were obtained from Breast Cancer Association Consortium (BCAC) and FinnGen Consortium. In this study, the Inverse-Variance Weighted (IVW), Weighted Median, MR-Egger, Maximum Likelihood (ML), MR-PRESSO and Constrained Maximum Likelihood and Model Averaging (cML-MA) methods of random effects models were used for MR analysis. Cochran’s Q analysis, and MR-Egger intercept analysis were applied for sensitivity analysis. IVW and cML-MA were considered the primary analytical tools, and the results of the other 4 MRs were used as complementary and validation. The results suggest that there is no significant causal relationship between leukocyte count and breast cancer risk (IVW OR = 0.98 [95% CI: 0.93–1.03], p-value = 0.35; CML-MA OR = 1.01 [95% CI: 0.98–1.05], p-value = 0.51). In addition, we analyzed whether there was a potential correlation between the five main types of categorized leukocyte counts and different breast cancer subtypes. We did not find significant evidence to support a significant correlation between leukocyte counts and breast cancer subtypes.

Shenqi Wenfei Formula alleviates chronic obstructive pulmonary disease in rats with pulmonary qi deficiency syndrome by regulating NLRP3/GSDMD pyroptosis pathway

To investigate the effects of Shenqi Wenfei (SQWF) Formula on the NLRP3/GSDMD signaling pathway in pulmonary qi deficiency syndrome rats with chronic obstructive pulmonary diseases (COPD). Rat models of COPD and lung qi deficiency syndrome induced by exposure to cigarette smoke and lipopolysaccharide (LPS) injection were randomized (n=8) for treatment with low-, medium- and high-dose SQWF or Yu Ping Feng (YPF). The changes in body weight, grip strength, lung function, pulmonary pathology, peripheral blood inflammatory cell counts, and levels of inflammatory factors in the bronchoalveolar lavage fluid (BALF) were examined. The expressions of proteins associated with the NLRP3/GSDMD signaling pathway in the lung tissues were determined with RT-qPCR and immunohistochemistry. The rat models of COPD and lung qi deficiency syndrome showed significantly reduced body weight, grip strength and lung function (P<0.01) with severe lung pathologies, increased levels of WBC, NEU% and MON% (P<0.01), decreased LYM% (P<0.01), and increased levels of TNF-α, IL-6, IL-1β and IL-18 in the BALF (P<0.01); the mRNA and protein expression levels of NLRP3, ASC, GSDMD and IL-1β all increased significantly in the lung tissue (P<0.01). Treatment with SQWF and YPF obviously increased body weight and improved grip strength, pulmonary function and lung pathology of the COPD rats (P<0.05). The treatments obviously improved the changes in peripheral blood inflammatory cell counts and inflammatory factors in the BALF of the rat models (P<0.01) and lowered the expressions of NLRP3, ASC, GSDMD and IL-1β in the lung tissues (P<0.05) without significantly affecting the mRNA levels of caspase-1 and IL-18 (P>0.05), and the effects of SQWF were dose-dependent. SQWF Formula relieves inflammatory response and injury in the lung tissue of COPD rats with pulmonary qi deficiency syndrome possibly by inhibiting pyroptosis through regulating the NLRP3/GSDMD pathway.

Serum interleukin-33 combined with FEF75% z-score and FeNO improves the diagnostic accuracy of asthma in children

ObjectiveTo investigate the diagnostic efficacy of serum IL-33 single indicator and combined indicators for asthma in children. Methods132 children were initially diagnosed with asthma during acute exacerbation and 100 healthy children were included. Serum IL-33 concentration differences were compared between asthmatic and normal children. Correlations between IL-33 with pulmonary function parameters, FeNO, peripheral blood EOS counts and serum total IgE were analyzed in asthmatic children. ROC curves were used to assess IL-33 diagnostic efficacy and its combined indicators. To prevent overfitting of the predictive model, the hold-out cross-validation method was used. Results(1) Serum IL-33 concentrations were significantly higher in children with asthma than in normal children (p < 0.001). (2) IL-33 concentration was negatively correlated with FVC z-score, FEV1 z-score and FEF75% z-score in asthmatic children (p < 0.05). (3) The area under the ROC curve of IL-33 was 0.821, which was higher than those of FeNO, FVC z-score, and FEV1 z-score. (4) Cross-validation of the combined indicators showed that IL-33 significantly improved asthma diagnostic efficacy. The combination of IL-33, FEF75% z-score, and FeNO showed the highest diagnostic efficacy, with the AUC, sensitivity, and specificity of the combined indicator being 0.954, 90.1%, and 89. 0%, respectively, and good extrapolation of the predictive model. ConclusionSerum IL-33 is higher in children with asthma and increases with the severity of pulmonary ventilation obstruction. A single indicator of serum IL-33 demonstrates moderate diagnostic accuracy, and its combination with FEF75% z-score and FeNO significantly improves the diagnostic accuracy in childhood asthma.

Gene Expressions and High Lymphocyte Count May Predict Durable Clinical Benefits in Patients with Advanced Non-Small-Cell Lung Cancer Treated with Immune Checkpoint Inhibitors.

Not all patients with advanced non-small cell lung cancer (NSCLC) benefit from immune checkpoint inhibitors (ICIs). Therefore, we aimed to assess the predictive potential of gene expression profiling (GEP), peripheral immune cell counts, and clinical characteristics. The primary endpoint of this prospective, observational study was a durable clinical benefit (DCB) defined as progression-free survival >6 months. In a subgroup with histological biopsies of sufficient quality (n = 25), GEP was performed using the nCounter® PanCancer IO 360 panel. DCB was observed in 49% of 123 included patients. High absolute lymphocyte count (ALC) and absence of liver metastases were associated with DCB (OR = 1.95, p = 0.038 and OR = 0.36, p = 0.046, respectively). GEP showed clustering of differentially expressed genes according to DCB, and a strong association between PD-L1 assessed by GEP (CD274) and immunohistochemistry (IHC) was observed (p = 0.00013). The TGF-β, dendritic cell, and myeloid signature scores were higher for patients without DCB, whereas the JAK/STAT loss signature scores were higher for patients with DCB (unadjusted p-values < 0.05). ALC above 1.01 × 109/L and absence of liver metastases were significantly associated with DCB in ICI-treated patients with NSCLC. GEP was only feasible in 20% of the patients. GEP-derived signatures may be associated with clinical outcomes, and PD-L1 could be assessed by GEP rather than IHC.

Development and validation of a peripheral cell ratio and lactate score for differentiating status epilepticus from prolonged psychogenic nonepileptic seizures.

Differentiating status epilepticus (SE) from prolonged psychogenic nonepileptic seizures (pPNES) can be difficult clinically. We aimed to define the utility of peripheral cell counts, cell ratios, and lactate levels in distinguishing SE from pPNES. Retrospective two-center study investigating the sensitivity and specificity of acute (≤12 h of event offset) peripheral cell counts, cell ratios (neutrophil-lymphocyte ratio, neutrophil-monocyte ratio, monocyte-lymphocyte ratio, platelet-lymphocyte ratio, systemic immune-inflammatory index [SII], systemic inflammatory response index [SIRI]), and lactate levels in differentiating SE from pPNES. Patients were identified from two tertiary hospitals, with one forming the development cohort and the other the validation cohort. Using generalized additive models to generate biomarker vs time curves, optimal blood collection times were defined for set parameters. Three diagnostic scores combining neutrophil count, SII, or SIRI with lactate levels were developed and validated in separate cohorts. For the development cohort, 1262 seizure-like events were reviewed and 79 SE and 44 pPNES events were included. For the validation cohort, 241 events were reviewed and 20 SE and 11 pPNES events were included. Individually, the biomarkers generally had low sensitivity and reasonable specificity for differentiating SE from pPNES, with the neutrophil count, SIRI, and SII performing best with sensitivities of 0.65-0.84, specificities of 0.64-0.89, and ROC AUCs of 0.78-0.79. Lactate levels peaked at 60 min, while cell counts and ratios peaked after 240 min. Combining early peaking lactate levels and later peaking neutrophil count, SIRI or SII resulted in three scores that improved predictive potential with sensitivities of between 0.75 and 0.79, specificities between 0.93 and 1.00, and ROC AUCs of 0.89-0.91. Lactate levels peak early post-SE, whereas cell counts and ratios do so later. The differing post-event time profiles of lactate levels vs neutrophil count, SIRI, and SII allow incorporation into three separate scores which can assist in differentiating SE from pPNES.

The Value of Neutrophil-to-Monocyte-Plus-Lymphocyte Ratio as a Marker for Discriminating Pulmonary Tuberculosis from Community Acquired Pneumonia

Abstract Background Tuberculosis (TB) is an infectious disease usually caused by Mycobacterium tuberculosis (MTB) bacteria. On the other hand, pneumonia is usually caused by viral or bacteria and less commonly by other microorganisms. Objectives To evaluate the value of Neutrophil to Monocyte plus lymphocyte ratio (NMLR) as a marker for discriminating pulmonary tuberculosis from community acquired pneumonia. Patients and Methods A case control study, was conducted at Abbassia Chest Hospital and included 60 patients with pulmonary infection (TB and community acquired pneumonia, 30 patients in each group) and 30 healthy controlled individuals. All patients subjected to full history, physical examination, Chest X-ray and 2 cc of blood sample was collected for total and differential peripheral leucocytic count before starting antibiotic therapy, Neutrophil-toMonocyte-Plus-Lymphocyte Ratio (NMLR) was calculated as the quotient of the absolute neutrophils, Monocyte and lymphocyte counts and Neutrophil-to-Lymphocyte Ratio (NLR) was calculated). Result The study involved 90 subjects; the mean of age of TB patients was (29±8.17) years, which significantly lower than other with community acquired pneumonia (40.4±16.81) years. Moreover, male sex was significantly associated with TB group. By blood analysis, NLR and NMLR were significantly higher in pneumonia than TB or control group, P value&amp;lt; 0.001 for both. Also, NLR &amp;gt; 2.96 and NMLR &amp;gt;2.385 cutoff values have sensitivity and specificity of 90% and 100% respectively for both. Conclusion We concluded that Cutoff value of NLR &amp;gt; 2.96 and NMLR &amp;gt;2.385 may use for prediction of pneumonia and T.B. the modified blood indices as NLR and NMLR had the capability to discriminate between inflammatory condition as pneumonia and TB and healthy condition as same as ordinary marker like CRP and ESR did.

Blood eosinophils and clinical features of pneumoconiosis complicated with chronic obstructive pulmonary disease: A cross-sectional study

Objective: To analyze the correlation between peripheral blood eosinophil (EOS) level and clinical characteristics of patients with pneumoconiosis complicated with chronic obstructive pulmonary disease (COPD) . Methods: From January 2007 to November 2020, newly diagnosed patients with pneumoconiosis complicated with COPD in Beijing Chaoyang Hospital, were retrospectively analyzed. These patients were stratified into EOS<100 cells/μl group and EOS≥100 cells/μl group, taking 100 cells/μl as the cut-off value. Demographic characteristics, clinical symptoms, lung function and laboratory indexes were compared between the two groups. Results: The median EOS count of patients with pneumoconiosis complicated with COPD was 100 (40, 180) cells/μl. 50.2% (160/319) had blood eosinophil counts ≥100 cells/μl, and 11.0% (35/319) had blood eosinophil counts ≥300 cells/μl. In comparison with EOS<100 cells/μl group, EOS≥100 cells/μl group were older (P=0.035), had higher body mass index (P=0.008), and had lower forced respiratory volume in the first second (P=0.017), had higher the ratio of residual volume to total lung volume (P=0.010), and had lower diffusing capacity of the lung for carbon monoxide (P=0.008). Arterial partial pressure of oxygen was significantly reduced in EOS≥100 cells/μl group (P=0.039). The peripheral blood EOS count was negatively correlated with forced vital capacity, forced breathing volume in the first second, carbon monoxide diffusion, peak expiratory flow, and maximum mid expiratory flow as a percentage of expected values (r(s)=-0.22, -0.18, -0.19, -0.19, -0.19, P=0.000, 0.001, 0.003, 0.008, 0.002), and positively correlated with the ratio of residual air volume to total lung volume (r(s)=0.17, P=0.002) . Conclusion: There was a correlation between blood EOS count and pulmonary function parameters, can proide reference for the diagnosis and treatment of chnoric obstuctive pulmmory disease in clinical practice.

Increased serum soluble interleukin-2 receptor levels in dermatomyositis are associated with Th17/Treg immune imbalance.

Dermatomyositis (DM) represents a multifaceted chronic inflammatory myopathy, primarily manifesting as progressive deterioration of muscular and cutaneous tissues. Despite an incomplete comprehension of DM's etiology and pathogenesis, current evidence implicates the involvement of T lymphocyte infiltration, extensive cytokine release, myositis-specific antibodies, and myositis-associated antibodies in disease development. Serum soluble interleukin-2 receptor (sIL-2R) frequently serves as a marker for T cell activation; however, its role remains elusive. Consequently, this investigation sought to elucidate the association between sIL-2R levels, peripheral blood lymphocyte subset counts, and related cytokines in DM patients, with the aim of uncovering the intricate mechanisms underlying DM and establishing a theoretical foundation for the implementation of precise, targeted, individualized immunomodulatory therapy. In this study, a cohort of 60 dermatomyositis (DM) patients, comprising 32 with inactive DM and 28 with active DM, was enrolled and stratified into inactive and active groups based on the Myositis Disease Activity Visual Analogue Scale (MYOACT). Flow cytometry was employed to quantify the absolute counts of peripheral lymphocyte subsets and CD4+T cell subsets in each group, while a flow cytometry bead array was utilized to measure serum cytokine levels. In a comparative analysis between healthy individuals and patients diagnosed with DM, we observed a marked elevation in serum sIL-2R concentrations (P < 0.001) and T-helper 17 cell/regulatory T cell (Th17/Treg) ratios (P < 0.01) within the latter group. A positive correlation was identified between serum sIL-2R levels and various parameters, including ESR, CRP, VAS, AST, CKMB, LDH, HBDH, PT, APTT, DDi, IL-6, IL-10, and IFN-γlevels (P < 0.05). In contrast, serum sIL-2R levels demonstrated a negative correlation with LY, HGB, ALB, Th17 cell populations, and Th17/Treg cell ratios (P < 0.05). Employing multivariate logistic regression, we identified serum sIL-2R concentrations as an independent risk factor for both disease activity and hepatic involvement in DM patients. Moreover, receiver operating characteristic (ROC) curve analyses revealed that serum sIL-2R levels significantly contributed to the differentiation of disease activity and the detection of liver involvement in DM patients, with areas under the ROC curve (AUC) of 0.757 (95% CI 0.630-0.884, P = 0.001) and 0.826 (95% CI 0.717-0.935, P < 0.001), respectively. This study highlights the potential utility of serum sIL-2R levels as a valuable biomarker for assessing disease activity and liver involvement in dermatomyositis. Elevated serum concentrations of sIL-2R were observed in patients with DM, exhibiting significant associations with Th17 cell populations and Th17/ Treg ratios. These findings indicate that sIL-2R may be implicated in the immunopathogenesis of DM, thereby warranting further investigation to elucidate its role in the disease process.

Identification of Plasma Thrombopoietin Level and Its Significance in Patients with Aplastic Anemia and Myelodysplastic Syndrome.

Objective Our objective was to investigate the concentration of plasma thrombopoietin (TPO) in patients with aplastic anemia (AA) and myelodysplastic syndrome (MDS), as well as its relationship with patients' responses to recombined human TPO (rhTPO) therapy. Methods We detected the concentration of plasma TPO in 31 patients with AA, 27 patients with MDS, and 11 normal controls using enzyme-linked immunosorbent assay. Results The median concentration of plasma TPO in patients with AA, MDS, and controls was (841.08 ± 768.64), (212.41 ± 338.93), and (35.09 ± 18.21) pg/mL, respectively. The TPO concentration in patients with AA and MDS was significantly higher than that in controls ( p < 0.05). The median platelet (PLT) counts were (184 ± 34) ×10 9 /L in the control group and (24 ± 19) ×10 9 /L and (80 ± 71) ×10 9 /L in AA and MDS patients, respectively. Negative correlations were found between plasma TPO concentration and PLT counts as well as megakaryocytes in bone marrow ( p < 0.05). In AA patients treated with rhTPO, a negative correlation was observed between increased PLT counts and pretreatment TPO levels ( p < 0.05). Conclusion Plasma TPO concentration in AA and MDS was significantly higher than that in normal controls. Plasma TPO was negatively correlated with peripheral blood PLT counts and bone marrow megakaryocyte counts. The pretreatment TPO level may serve as a prognostic indicator for the therapeutic effect of rhTPO in AA patients.

EVALUATION OF THE ACTIVITY OF PERIPHERAL BLOOD NATURAL KILLER CELLS IN HEALTHY SUBJECTS AND BREAST CANCER PATIENTS

Objectives: To study Natural Killer (NK) cell activity (NK-IFNg) and some indicators related to immunological characteristics of peripheral blood NK cells in healthy people and breast cancer patients. Methods: We evaluated NK cell activity through its secretory characteristics (NK-IFNg) using ATGen's commercial NK VUE TestÒ (ATGen kit) on 35 healthy subjects (medical staff) and 132 cancer patients treated at Vietnam National Cancer Hospital from August 2020 to February 2023. At the same time, we collected peripheral blood samples to conduct immunological characterization of peripheral blood NK cells based on the flow cytometer system (number, expression of the activation and inhibitory receptors such as NKG2A, NKG2D), and some other biomarkers (CEA, CA 15.3). Results: The secretory activity of peripheral blood NK cells (NKA-IFNγ) in breast cancer patients (1,013.46 ± 1,115.87 pg/mL) was statistically significantly lower than that in the healthy group ( 2,571.38 ± 827.52 pg/mL) ( ± SD) (p &lt; 0.001). The proportion of breast cancer patients with NKA-IFNγ activity of a very low level (≤ 200 pg/mL) was 40.9% and that of the healthy subjects was 0%. There was no relationship between NKA-IFNγ and peripheral blood NK cell count, and the expression levels of the activating receptor NKG2D and the inhibitory receptor NKG2A. Patients with CEA and CA 15.3 serum levels above the normal limits accounted for relatively low percentages (10.6% and 6.1%). Conclusion: Breast cancer patients with very low NK cell activity accounted for a significant proportion (40.9%). NKA-IFNγ activity could be used as a potential tool for monitoring the immune system health status of breast cancer patients before and after receiving treatment interventions.

Association between immune cells in peripheral blood and psychiatric symptoms.

There are bidirectional associations between immunological dysfunction and psychiatric symptoms. However, the associations between the levels of immune cells in the peripheral blood and psychiatric symptoms remain unclear. The present study aimed to evaluate levels of immune cells in peripheral blood in people with positive psychiatric symptoms. This retrospective study analyzed data from routine blood tests and psychopathology and sleep quality assessments. Data were compared between a group of 45 patients with de novo psychological symptoms and 225 matched controls. Patients with psychiatric symptoms had higher white blood cell and neutrophil counts compared with controls. However, in a subgroup analysis, neutrophil counts were significantly higher than in controls only in patients with multiple psychiatric symptoms. Furthermore, monocyte counts were significantly higher in patients with multiple psychiatric symptoms than in controls. Further, sleep quality was lower in patients with psychiatric symptoms than in controls. White blood cell and neutrophil counts in the peripheral blood of patients with psychiatric symptoms were significantly higher and sleep quality was significantly lower than in controls. Participants with multiple psychiatric symptoms showed more significant differences in peripheral blood immune cell counts than other subgroups. These results provided evidence for the relationship between psychiatric symptoms, immunity, and sleep.

#4709 PERSONALISED INDUCTION THERAPY IN ANCA GLOMERULONEPHRITIS

Abstract Background and Aims Patient and kidney outcome in ANCA glomerulonephritis (GN) remains unsatisfactory. Rituximab use failed to reduce the rate of infectious complications and demonstrates a significant rate of hypogammaglobulinaemia in ANCA vasculitis. Adjusting the rituximab dosage to the individual patient's need promises a reduction in treatment toxicity. Method Retrospective analysis of a single centre cohort of ANCA GN patients with disease flares between 2019 – 2022 using a stratified induction immunosuppression with rituximab and cyclophosphamide. Treatment intensity was adjusted to resolution of systemic inflammation and microscopic haematuria, peripheral CD19 cell count depletion and immunoglobulin G (IgG) levels. Results 45 patients (median age 57 years, 55.7% female, median eGFR 27 mls/min) were treated with a median dose of 1.5 grams Rituximab during induction (interquartile range, IQR 1 – 2 grams). 18 patients received concomitant intravenous cyclophosphamide pulses to control the acute inflammation (median cumulative dose of 2 grams, IQR 1.65 – 3 grams). Peripheral B cell depletion was 91.4% at 3 months and 90.3% at 6 months (&amp;lt;3 CD19 cells/ul). During the initial 12 months treatment, IgG level fell below 7g/l in 20 patients, below 5g/l in nine patients and below 3g/l in one patient. Seven patients required renal replacement therapy at diagnosis (15.6%). Five patients recovered kidney function and three patients developed end stage kidney disease (6.7%). Twelve infections were reported requiring antibiotic treatment (26.7%), seven severe infections were documented requiring hospitalisation (13.3%) and one patient died (2.2%). Conclusion An early assessment of treatment response assists adjusting vasculitis therapy. Adjusting management to the patient's individual immunological and treatment response may result in better outcome with less treatment toxicity.

The Potential of Fractional Exhaled Nitric Oxide as a Biomarker in Predicting and Optimizing Use of Treatment in Asthma

Asthma affects the respiratory system and causes airway inflammation. The indication of asthma includes a triad of airway inflammation, hyperresponsiveness, and obstruction. Nitric Oxide (NO) is a gas that is exhaled and is a sign of airway inflammation. NO levels in the exhaled breath of patients with type 2 asthma are elevated, and fractional exhaled nitric oxide (FeNO) is an objective biomarker of airway inflammation. Measurements of FeNO are noninvasive, require minimal patient effort, and are easy to collect in clinical settings. The current review is a systematic review performed using PubMed, Science Direct, and Google Scholar according to The Preferred Reporting Items for Systematic Reviews and Meta-Analysis Protocol (PRISMA-P) guidelines. This review discusses the entanglement of understanding FeNO measurement and supplementing existing diagnostic and assessment tools for inflammatory lung diseases. Monitoring FeNO can also help identify different asthma phenotypes within the asthma syndrome and suggest the optimal administration of inhaled corticosteroids (ICS) as elevated FeNO levels indicate ICS response. Non-adherence to ICS is a significant contributor to the failure of asthma treatment. A FeNO suppression test can be done to determine non-adherence. FeNO levels should be used with a careful history, conventional spirometric testing with bronchodilator reversibility, measures of bronchial hyperreactivity using methacholine, and other measures of eosinophilic inflammation, such as a peripheral blood eosinophil cell count. FeNO is more sensitive and specific when paired with other lung function tests.