Plasma proteome profiling reveals dynamic of cholesterol marker after dual blocker therapy.

Dual blocker therapy (DBT) has the enhanced antitumor benefits than the monotherapy. Yet, few effective biomarkers are developed to monitor the therapy response. Herein, we investigate the DBT longitudinal plasma proteome profiling including 113 longitudinal samples from 22 patients who received anti-PD1 and anti-CTLA4 DBT therapy. The results show the immune response and cholesterol metabolism are upregulated after the first DBT cycle. Notably, the cholesterol metabolism is activated in the disease non-progressive group (DNP) during the therapy. Correspondingly, the clinical indicator prealbumin (PA), free triiodothyronine (FT3) and triiodothyronine (T3) show significantly positive association with the cholesterol metabolism. Furthermore, by integrating proteome and radiology approach, we observe the high-density lipoprotein partial remodeling are activated in DNP group and identify a candidate biomarker APOC3 that can reflect DBT response. Above, we establish a machine learning model to predict the DBT response and the model performance is validated by an independent cohort with balanced accuracy is 0.96. Thus, the plasma proteome profiling strategy evaluates the alteration of cholesterol metabolism and identifies a panel of biomarkers in DBT.

PD-1 and LAG-3 positive T cells are related with the prognosis of patients with chronic kidney disease.

OBJECTIVE: Our objective was to study the frequency of circulating LAG-3+ and PD-1+ T cells in chronic kidney disease (CKD) patients and their correlation with cytokines and patient prognosis. METHODS: A total of 83 patients with CKD between June 2020 and June 2022 were enrolled. We measured serum levels of IL-6, CRP, IL-1ß, and TNF-α by ELISA. The frequency of PD-1+ and LAG-3+ T cells was measured using flow cytometry. All patients were followed up for 1 year, and the occurrence of any of the following conditions during the follow-up period was considered as major adverse cardiac events (MACE) indicating poor prognosis. RESULTS: The frequencies of LAG-3+PD-1+, LAG-3+ and PD-1+ cells were significantly increased in CKD group compared to healthy volunteers. Additionally, CKD patients had remarkably enhanced levels of cytokines. Compared to the non-MACE group, MACE group had significantly higher frequencies of LAG-3PD-1, LAG-3 and PD-1 expression on CD8 and CD4. Positive correlations were observed between IL-1ß, IL-6 and frequencies of PD-1+LAG-3+. CD4+LAG-3+PD-1+ frequency displayed the highest diagnostic value for CKD patients with MACE. Moreover, CD8+LAG-3+, CD4+LAG-3+PD-1+, CD4+PD-1+, IL-1ß and IL-6 were identified as risk factors for the occurrence of MACE in patients with CKD. CONCLUSION: In summary, the present research showed that the frequencies of LAG-3+ and PD-1+ T cells were remarkably enhanced in CKD patients. These findings offer novel insights and potential therapeutic targets for the management of CKD.

The diversity of peripheral blood T cells and myeloid-derived suppressor cells in patients with idiopathic membranous nephropathy.

OBJECTIVE: Membranous nephropathy is a leading cause of adult-onset nephrotic syndrome. Peripheral T cells and myeloid-derived suppressor cells (MDSCs) are closely associated with autoimmune diseases, while their exact roles and interaction in these processes are unclear. Here, we studied the roles of T cells, MDSCs, and their subsets in patients with idiopathic membranous nephropathy (IMN). MATERIALS AND METHODS: 35 IMN patients and 30 healthy controls were included in this retrospective study. Flow cytometry was performed to determine the phenotype of human T cells and MDSCs in peripheral blood mononuclear cells (PBMCs). Anti-PLA2R was measured by ELISA. Values ≥ 20 RU/mL were defined as positive and < 14 RU/mL as negative. RESULTS: A higher ratio of CD4/CD8 T cells with a lower proportion of Tregs, a remarkably lower proportion of G-MDSCs (but not M-MDSCs), lower frequency of PD-L2+G-MDSCs, and higher frequency of PD-L1+M-MDSCs were found in IMN patients compared to healthy controls. The ratio of CD4/CD8 T cells was higher, and the frequencies of PD-1+CD4+ T cells, CTLA-4+CD4+ T cells, PD-1+Tregs, and CTLA-4+Tregs were lower in PBMCs of PLA2R-positive IMN patients compared to PLA2R-negative IMN patients. CONCLUSION: Tregs and G-MDSCs were reduced in the circulation of the IMN patients, which may promote understanding of the crucial functions that are mediated by these cells in the pathogenesis of IMN.

Missing prognostic value of soluble PD-1, PD-L1 and PD-L2 in lung cancer patients undergoing chemotherapy - A CEPAC-TDM biomarker substudy.

BACKGROUND: Programmed cell death receptors and ligands in cancer tissue samples are established companion diagnostics for immune checkpoint inhibitor (ICI) therapies. OBJECTIVE: To investigate the relevance of soluble PD-1, PD-L1 and PD-L2 for estimating therapy response and prognosis in non-small cell lung cancer patients (NSCLC) undergoing platin-based combination chemotherapies. METHODS: In a biomarker substudy of a prospective, multicentric clinical trial (CEPAC-TDM) on advanced NSCLC patients, soluble PD-1, PD-L1 and PD-L2 were assessed in serial serum samples by highly sensitive enzyme-linked immunosorbent assays and correlated with radiological response after two cycles of chemotherapy and with overall survival (OS). RESULTS: Among 243 NSCLC patients, 185 achieved response (partial remission and stable disease) and 58 non-response (progression). The distribution of PD-1, PD-L1 and PD-L2 at baseline (C1), prior to staging (C3) and the relative changes (C3/C1) greatly overlapped between the patient groups with response and non-response, thus hindering the discrimination between the two groups. None of the PD markers had prognostic value regarding OS. CONCLUSIONS: Neither soluble PD-1, PD-L1 nor PD-L2 did provide clinical utility for predicting response to chemotherapy and prognosis. Studies on the relevance of PD markers in ICI therapies are warranted.

The combination of soluble forms of PD-1 and PD-L1 as a predictive marker of PD-1 blockade in patients with advanced cancers: a multicenter retrospective study.

Introduction: The clinical relevance of soluble forms of programmed cell death-1 (sPD-1) and programmed cell death-ligand 1 (sPD-L1) remains unclear. We here investigated the relation between the efficacy of PD-1 blockade and pretreatment plasma levels of sPD-1 and sPD-L1 across a broad range of cancer types. Methods: We retrospectively analyzed clinical data from 171 patients with advanced solid tumors who received nivolumab or pembrolizumab monotherapy regardless of treatment line. The concentrations of sPD-1 and sPD-L1 were measured with a fully automated immunoassay (HISCL system). Results: The study subjects comprised patients with head and neck cancer (n = 50), urothelial cancer (n = 42), renal cell cancer (n = 37), gastric cancer (n = 20), esophageal cancer (n = 10), malignant pleural mesothelioma (n = 6), or microsatellite instability-high tumors (n = 6). High or low levels of sPD-1 or sPD-L1 were not significantly associated with progression-free survival (PFS) or overall survival (OS) for PD-1 blockade in the entire study population. Comparison of treatment outcomes according to combinations of high or low sPD-1 and sPD-L1 levels, however, revealed that patients with low sPD-1 and high sPD-L1 concentrations had a significantly poorer PFS (HR of 1.79 [95% CI, 1.13-2.83], p = 0.01) and a tendency toward poorer OS (HR of 1.70 [95% CI, 0.99-2.91], p = 0.05) compared with all other patients. Conclusion: Our findings suggest that the combination of low sPD-1 and high sPD-L1 levels is a potential negative biomarker for PD-1 blockade therapy.

Soluble forms of PD-1/PD-L immune checkpoint receptor and ligand in blood serum of breast cancer patients: association with clinical pathologic factors and molecular type of the tumor.

Results of enzyme-linked immunosorbent assay of the soluble forms of PD-1/PD-L immune checkpoint receptor and ligand (sPD-1 and sPD-L1) in pretreatment blood serum of 88 breast cancer patients at various disease stages aged 30-83 years are presented. The control group included 55 practically healthy women aged 19-82 years. Serum sPD-1 and sPD-L1 levels in breast cancer patients highly significantly (p<0.0001) differ from control and these changes are opposite: soluble receptor level is more than 6-fold decreased, while soluble ligand concentration - 5.5 fold increased. Both markers separately, as well as their ratio demonstrate very high sensitivity (94-100%) and specificity (95-100%) in relation to healthy control. No statistically significant associations of sPD-1 and sPD-L1 levels with clinical stage, individual TNM system criteria, tumor histological structure, grade, receptor status, and molecular type were established. In particular, no significant peculiarities of the markers' levels in triple negative breast cancer successfully treated with anti-PD-1/PD-L1 preparations were revealed. Long-term follow-up and dynamic studies of sPD-1 and sPD-L1serum levels in the course of treatment are required for evaluation of their independent from clinical and morphological factors prognostic significance and the possibility of application as low invasive tests for prediction and monitoring of corresponding targeted therapy efficiency.

Soluble programmed cell death-1 predicts hepatocellular carcinoma development during nucleoside analogue treatment.

Soluble immune checkpoint molecules are emerging novel mediators of immune regulation. However, it is unclear whether soluble immune checkpoint proteins affect the development of hepatocellular carcinoma (HCC) during nucleos(t)ide analogue (NA) treatment in patients with chronic hepatitis B virus infection. This study included 122 NA-naïve patients who received NA therapy. We assessed the associations of clinical factors, including soluble immune checkpoint proteins, with HCC development during NA treatment. The baseline serum concentrations of 16 soluble immune checkpoint proteins were measured using multiplexed fluorescent bead-based immunoassay. In total, 13 patients developed HCC during the follow-up period (median duration, 4.3 years). Of the 16 proteins, soluble inducible T-cell co-stimulator (≥ 164.71 pg/mL; p = 0.014), soluble programmed cell death-1 (sPD-1) (≤ 447.27 pg/mL; p = 0.031), soluble CD40 (≤ 493.68 pg/mL; p = 0.032), and soluble herpes virus entry mediator (≤ 2470.83 pg/mL; p = 0.038) were significantly associated with HCC development (log-rank test). In multivariate analysis, an sPD-1 level ≤ 447.27 pg/mL (p = 0.014; hazard ratio [HR], 4.537) and α-fetoprotein level ≥ 6.4 ng/mL (p = 0.040; HR, 5.524) were independently and significantly associated with HCC development. Pre-treatment sPD-1 is a novel predictive biomarker for HCC development during NA treatment.

The role of PD-1/PD-Ls in the pathogenesis of IgG4-related disease.

OBJECTIVE: To investigate the role of programmed cell death protein 1 (PD-1) and its two ligands, PD-L1 and PD-L2, in the pathogenesis of IgG4-related disease (IgG4-RD). METHODS: Patients with IgG4-RD (n = 43) and healthy controls (n = 34) were recruited. Expression levels of PD-1, PD-L1 and PD-L2 in plasma, submandibular gland and T cell subsets were determined by ELISA, immunohistochemistry and flow cytometry. Naïve T cells were stimulated with or without PD-L1/PD-L2 or anti-PD-L1/anti-PD-L2 for 7 days and the proportion of CD4+CD25+ Treg cells was detected by flow cytometry. RESULTS: The expression of PD-1, PD-L1 and PD-L2 in the plasma, submandibular gland and on the surface of Treg cells was increased in IgG4-RD patients. Plasma soluble (s)PD-1 was positively correlated with serum IgG, IgG1, IgG3, IgG4, IgG4-RD responder index and numbers of organs involved, and negatively correlated with serum IgM, IgA, C3 and C4. Plasma sPD-L2 was positively correlated with serum IgG1, and plasma sPD-L1 was positively correlated with sPD-L2 and negatively correlated with C3. Stimulation of PD-L1 but not PD-L2 promoted the differentiation of naïve T cells from IgG4-RD patients into CD4+CD25+ Treg cells. CONCLUSION: Plasma concentrations of sPD-1, sPD-L1 and sPD-L2 were significantly increased in patients with IgG4-RD, and the expression of PD-1 and PD-L2 on Treg cells was upregulated. PD-1-PD-L1 can promote the differentiation of naïve T cells into Treg cells and thus participate in the pathogenesis of IgG4-RD.

Blood PD-1+TFh and CTLA-4+CD4+ T cells predict remission after CTLA-4Ig treatment in early rheumatoid arthritis.

OBJECTIVE: Treatment with CTLA-4Ig blocks T-cell activation and is clinically effective in RA. However, it is unknown if specific CD4+ T-cell subsets in blood at baseline predict remission after CTLA-4Ig, or other biological treatments with different modes of action, and how treatment affects CD4+ T cells in patients with untreated early RA (eRA). METHODS: This study included 60 patients with untreated eRA from a larger randomized trial. They were treated with methotrexate combined with CTLA-4Ig (abatacept, n = 17), anti-IL6 receptor (tocilizumab, n = 21) or anti-TNF (certolizumab-pegol, n = 22). Disease activity was assessed by clinical disease activity index (CDAI), DAS28, swollen joint counts, tender joint counts, CRP and ESR. The primary outcome was CDAI remission (CDAI ≤ 2.8) at week 24. Proportions of 12 CD4+ T-cell subsets were measured by flow cytometry at baseline and after 4, 12 and 24 weeks of treatment. RESULTS: In patients treated with CTLA-4Ig, the proportions of PD-1+TFh and CTLA-4+ conventional CD4+ T cells at baseline predicted CDAI remission at week 24. CD4+ T-cell subset proportions could not predict remission after treatment with anti-IL6R or anti-TNF. The percentage of regulatory T cells (Tregs) expressing CTLA-4 decreased in all treatment arms by 24 weeks, but only CTLA-4Ig treatment significantly reduced the proportions of Tregs and PD-1+T follicular helper (TFh) cells. CONCLUSION: These findings indicate that circulating proportions PD-1+TFh and CTLA-4+ conventional CD4+ T cells at baseline may serve as predictive biomarkers for remission in early RA after CTLA-4Ig treatment.

Relationship between the absolute lymphocyte count/absolute monocyte count ratio, soluble interleukin 2 receptor level, serum programmed cell death 1 level, and the prognosis of patients with diffuse large B-cell lymphoma.

BACKGROUND: To analyze the relationship between the peripheral blood absolute lymphocyte count (ALC)/absolute monocyte count (AMC) ratio, soluble interleukin 2 receptor (sIL-2R) level, serum programmed cell death 1 (PD-1) level, and the prognosis of patients with diffuse large B-cell lymphoma (DLBCL). METHODS: A total of 78 patients with DLBCL admitted to hospital and 30 healthy controls were enrolled as the case group and control group between August 2019 and June 2020, respectively. The ALC/AMC ratio and the levels of sIL-2R and serum PD-1 between the 2 groups and among patients with different prognoses were compared. The evaluation efficiency of these 3 factors for the prognosis of DLBCL patients was analyzed by receiver operating characteristic (ROC) curves. The risk factors affecting the 1-year survival rate were analyzed by the Cox hazard model. RESULTS: The levels of sIL-2R, AMC, and PD-1 in the case group were significantly higher than those in the control group, while the ALC/AMC ratio was lower than that in the control group (P<0.05). The levels of sIL-2R and PD-1 in the poor prognosis group were significantly higher than those in the good prognosis group, while the ALC/AMC ratio was lower than that in the good prognosis group (P<0.05). The areas under the ROC curve (AUCs) of sIL-2R level, serum PD-1 level, and the ALC/AMC ratio in evaluating the prognosis of DLBCL patients were 0.805 (95% CI: 0.700-0.886), 0.825 (95% CI: 0.722-0.902), 0.792 (95% CI: 0.685-0.876), respectively. The critical values were 474.80 µg/L, 206.85 pg/mL and 3.01, respectively. The differences in the 1-year survival rate among DLBCL patients with different tumor sizes, B symptoms, sIL-2R levels, and ALC/AMC ratios were statistically significant (P<0.05). B symptoms (RR =1.721) and ALC/AMC ratio lower than 3.01 (RR =1.484) were independent influencing factors of the 1-year survival rate in DLBCL patients (P<0.05). CONCLUSIONS: The ALC/AMC ratio, sIL-2R level, and serum PD-1 level can effectively assess the prognosis of DLBCL patients. B symptoms and ALC/AMC ratio lower than 3.01 are risk factors affecting the 1-year survival rate of patients.

Increased Circulating CD4+CXCR5+ Cells and IgG4 Levels in Patients with Myelodysplastic Syndrome with Autoimmune Diseases.

OBJECTIVES: Immune abnormalities play an important role in the pathogenesis and progression of myelodysplastic syndrome (MDS). Some patients with MDS have autoimmune diseases (AI). Follicular helper T (Tfh) cells help B cells produce antibodies. The role of Tfh in MDS with AI has not been studied. METHODS: We enrolled 21 patients with MDS with AI and 21 patients with MDS without AI. The proportion of peripheral blood CD4+CXCR5+ cells and the PD1 expression on CD4+CXCR5+ cells were detected by flow cytometry. Serum levels of immunoglobulin G (IgG) and IgG4 were measured. The survival and progression of MDS to acute myeloid leukemia (AML) in MDS patients with or without AI were compared. RESULTS: MDS with AI accounted for 19.6% of all MDS cases in our study. The overall response rate was 81% (17/21) in MDS patients with AI for the first-line treatment. The proportion of circulating CD4+CXCR5+ cells was increased, but the expression of PD1 was decreased in MDS patients with AI. Serum IgG4 levels were also increased in MDS patients with AI. The proportion of peripheral blood CD4+CXCR5+ cells and the level of serum IgG4 decreased after therapy, but the expression of PD1 increased. There were no differences in overall survival and progress to acute myeloid leukemia between MDS with AI and without AI groups. CONCLUSION: CD4+CXCR5+ cells and IgG4 levels increased in patients with MDS and AI.

Soluble programmed death molecule 1 (sPD-1) as a predictor of interstitial lung disease in rheumatoid arthritis.

BACKGROUND: Previous studies have indicated that the programmed death molecule 1 (PD-1) signaling pathway may play a key role in rheumatoid arthritis (RA). However, the pathogenesis of rheumatoid arthritis-related interstitial lung disease (RA-ILD) is not clear. We examined the serum levels of soluble PD-1 in patients with RA and its relationship with RA-ILD. METHODS: Blood samples were obtained from 87 patients with RA (58 with ILD and 29 without ILD) and 45 healthy controls. Serum sPD-1 was measured by Enzyme-Linked Immunosorbent Assay. The pulmonary interstitial disease score was completed by a pulmonary physician and a radiologist through chest high-resolution computed tomography. Patients with RA-ILD were tested for lung function [e.g., forced vital capacity (FVC%), diffusing capacity of lungs for carbon monoxide (DLCO%)]. Associations between ILD and various markers, including sPD-1 and confounding factors, were investigated by logistic regression analysis. Diagnostic values of sPD-1 for the presence of ILD were investigated using receiver operating characteristic curve analysis. RESULTS: Serum sPD-1 levels were higher in RA patients with ILD than in RA patients without ILD and healthy controls (185.1 ± 109.0 pg/ml vs. 119.1 ± 77.5 pg/ml vs. 52.1 ± 21.7 pg/ml, P < 0.05). Serum sPD-1 levels were positively correlated with RF titer (P = 0.02, r = 0.249), anti-cyclic citrullinated peptide antibody status (P = 0.02, r = 0.243), and serum IgG levels (P < 0.001, r = 0.368), negatively associated with FVC% (P = 0.02, r = - 0.344), forced expiratory volume (FEV1%) (P = 0.01, r = - 0.354), total lung capacity (TLC%) (P = 0.046, r = - 0.302), and was independently associated with the presence of ILD in RA patients by multivariate logistic regression analysis. The sensitivity and specificity of sPD-1 levels for the detection of ILD in RA patients were 58.6% and 75.9%, respectively. The area under the curve was 0.689. CONCLUSION: Serum sPD-1 levels were increased in RA patients with ILD. Increased sPD-1 may be a valuable biomarker to predict the presence of ILD in patients with RA.

Low Soluble Programmed Cell Death Protein 1 Levels After Allogeneic Stem Cell Transplantation Predict Moderate or Severe Chronic GvHD and Inferior Overall Survival.

Programmed cell death protein-1 (PD-1) is an inhibitory co-receptor required for regulating immune responsiveness and maintaining immune homeostasis. As PD-1 can be released as bioactive soluble molecule, we investigated the clinical significance of soluble PD-1 (sPD-1) after allogeneic hematopoietic stem cell transplantation (HSCT) regarding graft-versus-host disease (GvHD), relapse, and overall survival (OS) in a mono-centric cohort of 82 patients. Compared to pre-HSCT and to healthy controls, post-HSCT sPD-1 plasma levels were significantly increased during an observation time of three months. Univariate analysis revealed that low sPD-1 plasma levels at month one, two or three post HSCT were associated with acute GvHD grade III-IV, the onset of moderate/severe chronic GvHD (cGvHD) and inferior OS, DFS, and TRM, respectively. No relationship was detected to relapse rates. sPD-1 plasma levels were significantly increased in ATG-treated patients compared to ATG-untreated patients. Multivariate analysis revealed that a low sPD-1 plasma levels status at one or two month(s) after HSCT is an independent indicator for inferior OS, DFS, or TRM. A low sPD-1 plasma levels status at month three post HSCT is predictive for the onset of moderate/severe cGvHD. Thus, our study pinpoints the soluble inhibitory co-receptor PD-1 as a promising candidate molecule for the prediction of clinical HSCT outcome.

Evaluation of circulating PD-1 and PD-L1 as diagnostic biomarkers in dogs with tumors.

BACKGROUND: Programmed cell death protein-1 (PD-1) and programmed cell death ligand-1 (PD-L1) have important roles in tumor evasion of the immune system. OBJECTIVES: This study aimed to assess the diagnostic utility of circulating PD-1 and PD-L1 levels in healthy dogs and dogs with tumors. METHODS: Circulating PD-1 and PD-L1 levels in the serum of 71 dogs with tumors were compared with those of 52 healthy dogs by performing enzyme-linked immunosorbent assay (ELISA). RESULTS: The ELISA results revealed higher circulating PD-1 and PD-L1 levels in dogs with tumors (2.9 [2.2-3.7] ng/mL; median [IQR] and 2.4 [1.4-4.4] ng/mL, respectively) than in healthy dogs (2.4 [1.9-3.0] ng/mL; p = 0.012 and 1.4 [0.9-2.1] ng/mL; p < 0.001, respectively). Especially, there was a significant difference in circulating PD-1 levels between healthy dogs and dogs with malignant epithelial tumors (2.4 [1.9-3.0] ng/mL and 3.1 [2.6-4.4] ng/mL, respectively; p < 0.01). In addition, there was a significant difference in circulating PD-L1 levels between healthy dogs and dogs with lymphomas (1.4 [0.9-2.1] ng/mL and 2.7 [1.6-5.8] ng/mL, respectively; p < 0.001). CONCLUSION: This study indicates that circulating PD-1 and PD-L1 have potential as tumor diagnostic biomarkers in dogs with tumors.

T cell exhaustion is associated with the risk of papillary thyroid carcinoma and can be a predictive and sensitive biomarker for diagnosis.

BACKGROUND: The incidence of papillary thyroid carcinoma (PTC) has been steadily increasing over the past decades. Hashimoto's thyroiditis (HT) is the most common autoimmune disease, and is related to the pathogenesis of PTC. Programmed death-1 (PD-1) is currently used for the treatment of PTC, but there are very few studies on the clinical value of PD-1 in the diagnosis and targeted therapy of PTC. METHODS: The expression of T, B, NK cells and PD-1 in the peripheral blood of 132 patients with PTC (PTC group), 48 patients with nodular goiter (NG group) and 63 healthy subjects (HP group) were detected by flow cytometry. The expression of plasma T3, T4, FT3, FT4, TSH, TGAb and TPO was detected by chemiluminescence immunoassay. Among 132 PTC, 49 PTC&HT and 83 PTC&noHT were included. Among 48 NG, 10 NG&HT and 38 NG&noHT were included. The expressions of programmed death- ligand1(PD-L1) in tumor tissues of PTC group and thyroid tissues of NG group, PD-1 and CD3 in tumor infiltration lymphocyte (TIL) were detected by immunohistochemistry. RESULTS: The expression of FT3, TGAb, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC and NG was significantly higher than that in the HP group. Moreover, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ expression had significant differences between the PTC group and the NG group. In addition, the expression of TGAb, TPO, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC&HT group was significantly higher than that in the PTC&noHT group. While, the expression of B cells, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC&HT group was higher than that in NG&HT group. PD-1 showed a significant correlation with PTC lymph node metastasis. CD3+PD-1+ and CD3+CD4+PD-1+ was higher in N1 stage than in N0 stage. Immunohistochemical results showed that the expression of PD-1, CD3 and PD-L1 in PTC was significantly higher than that in NG. CONCLUSIONS: T cell exhaustion might act as a biomarker for the differential diagnosis of PTC and NG. Patients with PTC&HT have obvious T cell exhaustion and increased expression of PD-1, PD-L1.Targeting the PD-1/PD-L1 pathway could be a new approach to prevent malignant transformation from HT to PTC&HT in the future.

Circulating CD4+ FoxP3- CXCR5- CXCR3+ PD-1hi cells are elevated in active rheumatoid arthritis and reflect the severity of the disease.

OBJECTIVE: To examine the expression and clinical significance of circulating CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells in rheumatoid arthritis (RA). METHODS: CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells in peripheral blood of 35 patients with active RA, 17 with RA in stable remission, and 24 healthy controls were analyzed by flow cytometry. Serum IgG and circulating plasmablast percentages were measured and correlations with CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells were systematically analyzed. Disease Activity Scale 28 (DAS28) scores were also calculated and correlation analysis with CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells was conducted. The levels of CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells were compared before and after disease-modifying anti-rheumatic drug treatment. Cytokine levels in plasma and cytokine secretion in CD4 cells were measured and their correlations with CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells were further analyzed. RESULTS: The levels of CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells in the peripheral blood of patients with active RA were significantly increased compared with healthy controls. CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells in patients with active RA were positively correlated with serum IgG and DAS28 scores. CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells were significantly decreased in patients after treatment. Plasma interleukin-10 concentrations and interleukin-10-positive CD4 cell percentages were significantly positively correlated with CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cell levels. CONCLUSION: Circulating CD4+  FoxP3-  CXCR5-  CXCR3+  PD-1hi cells in patients with active RA are increased and could reflect the severity of the disease, which may play a potential role in the pathogenesis of RA.

Unbalanced expression of membrane-bound and soluble programmed cell death 1 and programmed cell death ligand 1 in systemic juvenile idiopathic arthritis.

The study aimed to investigate the soluble programmed death-1 (sPD-1) and its ligand (sPD-L1) levels in systemic juvenile idiopathic arthritis (sJIA) patients and elucidate its underlying immunomodulatory mechanisms. Plasma levels of sPD-1, sPD-L1 and related cytokines and proteins were detected using an enzyme-linked immunosorbent assay (ELISA) and Luminex. The effects of PD-1/PD-L1 signal on mDC (myeloid dendritic cell) and IL-6 secretion were measured using flow cytometry. The results revealed decreased levels of sPD-1 in sJIA patients negatively correlated with JADAS-27, PGA, PtGA and CRP. sJIA patients had lower CD86 and MHC-II expression on mDC. When blocking PD-1/PD-L1 signal, IL-6 secretion of DC were increased. Our finding displayed downregulated sPD-1 was related with clinical indicators and could be a new biomarker for sJIA diagnosis. The reduced membrane and soluble forms of PD-1/PD-L1 might take part in sJIA pathogenesis by enhancing mDC proliferation and IL-6 secretion.

Serum HBV pregenomic RNA exhibited opposite associations with NKdim and NKbright cell immunity in treatment-naïve chronic hepatitis B patients.

Hepatitis B virus (HBV) pregenomic RNA (pgRNA) is a new biomarker that reflects HBV replication, but its relationship with natural killer (NK) cell immunity in chronic hepatitis B (CHB) is unknown. We assessed serum HBV pgRNA levels in 323 CHB patients by reverse transcription-polymerase chain reaction, assessed cytokine production and activation and inhibitory markers of NK cells by flow cytometry, and measured serum cytokines by enzyme-linked immunosorbent assays (ELISAs). Among the different CHB phases, the serum HBV pgRNA level was highest in the immune-tolerant (IT) and immune-active (IA) phases. Regarding NK and NKdim cells, HBV pgRNA was negatively associated with frequencies, but positively associated with NKp44 and NKp46 expression (activation markers). Regarding NKbright cells, serum HBV pgRNA was positively associated with frequency and programmed cell death protein 1 (PD1) expression (inhibitory marker), but negatively associated with NKp44 and NKp46. Serum HBV pgRNA was not associated with NKp30 (activation marker) on NK cells or subsets. Lastly, serum HBV pgRNA was positively correlated with the levels of serum IL-7 and IL-12P40 (NK cell-promoting cytokines) and negatively correlated with serum prostaglandin E2 (PGE2) level (which negatively regulates NK cells). In conclusion, we found varied relationships between serum HBV pgRNA and NK cells and subsets, indicating that HBV pgRNA may play a complicated role in NK cell-related immunity, providing new information on HBV and host immunity.

Bloodborne Cytokines for Predicting Clinical Benefits and Immune-Related Adverse Events in Advanced Non-Small Cell Lung Cancer Treated With Anti-Programmed Cell Death 1 Inhibitors.

BACKGROUND: Programmed cell death ligand 1 is a biomarker of immune checkpoint inhibitors (ICIs) for treating advanced non-small-cell lung cancer (NSCLC). Here, we evaluated serum proteins from patients with advanced NSCLC treated with ICIs to determine their potential as noninvasive predictive biomarkers for efficacy and immune-related adverse events (irAEs). PATIENTS AND METHODS: Patients with advanced NSCLC who received nivolumab or pembrolizumab monotherapy until disease progression or unacceptable toxicity were integrated with previously reported nivolumab-treated patients. Blood samples were collected serially from baseline until the disease progressed. Serum protein levels were quantified using the Luminex assay. Associations of clinical benefit (CB) and onset of irAEs with serum protein levels were evaluated. RESULTS: Sixty-three patients with advanced NSCLC were studied, and we used 63 and 47 paired serum samples at baseline and the second sampling point, respectively, for efficacy analysis. Baseline growth-regulated oncogene 1 (GRO-1) levels were significantly lower in durable CB (DCB) patients than in non-DCB patients (P < .05). Changes in monocyte chemoattractant protein 1 (MCP-1) levels significantly decreased between baseline and the second sampling point (P < .05). Patients with the low GRO-1/decreased MCP-1 subtype showed significantly longer progression-free survival (PFS) and overall survival (OS) than the high GRO-1/increased MCP-1 subgroup did (median PFS, not reached vs. 47 days, P < .0001; median OS, 985 days vs. 148 days, P = .0002, respectively). Elevated GRO-1 levels were associated with immune-related adverse event onset. CONCLUSIONS: Serum GRO-1 and MCP-1 levels can identify patients with advanced NSCLC who are likely to benefit from ICI treatment. Time-course tracing of these protein levels might be valuable in ICI treatment.

Molecular Mechanism of Expression Changes of Immunological Indexes of PD-1/sPD-L1 after Radiotherapy in Nonsmall Cell Lung Cancer.

It is aimed at investigating the changes of serum soluble programmed death-ligand 1 (sPD-L1) expression level in nonsmall cell lung cancer (NSCLC) before and after radiotherapy, the correlation of PD-L1, PD-1, and proteins of Akt (protein kinase B), mTOR, and HIF-1α, and the molecular mechanism of the PD-1/PD-L1 pathway in the development of NSCLS. A total of 126 NSCLC patients receiving radiotherapy in Liaoning Cancer Hospital from September 2018 to September 2019 were selected as the observation group, and another 58 healthy volunteers were selected as the control group. NSCLC patients were divided into group A (stage I-II, stereotactic radiotherapy) and group B (stage III, intensity-modulated radiation therapy) according to the cancer stage. The efficacy of radiotherapy was evaluated, and sPD-L1 expression was detected by ELISA. The immunohistochemical staining was adopted to detect protein expressions of Akt, mTOR, and HIF-1α in NSCLC tissues. The correlation between their expression and expression of PD-L1 and PD-1 was analyzed. The results showed that the overall response rate (ORR) of group A was 89.29%, the clinical benefit response (CBR) was 96.43%, the median survival time (MST) was 25 months, and the survival rate within three years was 72.56%. In group B, the ORR was 70.41%, the CBR was 97.96%, the MST was 18 months, and the survival rate within three years was 34.67%. Comparison of overall serum sPD-L1 expression in the control group, group A, and group B and between groups before radiotherapy was statistically significant (P < 0.01). After radiotherapy, serum sPD-L1 expression in group A and group B decreased compared with that before radiotherapy (P < 0.01). Among NSCLC patients, the positive expression rate of Akt, mTOR, and HIF-1α was 71.32%, 41.26%, and 80.65%, respectively. PD-L1 expression and Akt, mTOR, and HIF-1α expression showed a significant correlation. PD1 expression and Akt, mTOR, and HIF-1α expression also showed a significant correlation. It indicated that the expression level of sPD-L1 in NSCLC patients was higher than that in normal subjects, but the expression level of sPD-L1 was decreased after radiotherapy. PD-1/PD-L1 may play important roles in NSCLC procession through the Akt/mTOR and HIF-1α pathway.