sPD-1 Levels Research Articles

Decreased serum soluble programmed cell death ligand-1 level as a potential biomarker for missed miscarriage.

Can maternal serum levels of soluble programmed cell death-1 (sPD-1) and its ligand (sPD-L1) serve as biomarkers for missed miscarriage (MM)? Serum sPD-L1 levels are significantly decreased in MM patients and may serve as a potential predictive biomarker for miscarriage. Programmed cell death-1 (PD-1) and its ligand (PD-L1) comprise important immune inhibitory checkpoint signaling to maintain pregnancy. Their soluble forms are detectable in human circulation and are associated with immunosuppression. Three independent cohorts attending tertiary referral hospitals were studied. The first (discovery) cohort was cross-sectional and included MM patients and healthy pregnant (HP) women matched on BMI. The second validation cohort contained MM patients and women with legally induced abortion (IA). The third prospective observational study recruited subjects requiring IVF treatment. In the discovery cohort, we enrolled 108 MM patients and 115 HP women who had a full-term pregnancy at 6-14 weeks of gestation. In the validation cohort, we recruited 25 MM patients and 25 women with IA. Blood samples were collected at the first prenatal visit for HP women or on the day of dilatation and curettage surgery (D&C) for MM and IA subjects to determine serum sPD-1 and sPD-L1 levels. Placenta samples were harvested during the D&C within the validation cohort to measure gene and protein expression. The prospective cohort collected serial blood samples weekly from 75 volunteers with embryo transfer (ET) after IVF. Circulating sPD-L1 levels were reduced by 50% in patients with MM (55.7 ± 16.04 pg/ml) compared to HP controls (106.7 ± 58.46 pg/ml, P < 0.001) and the difference remained significant after adjusting for maternal age and gestational age, whereas no significant differences in sPD-1 level were observed. Likewise, serum sPD-L1 was lower in MM patients than in IA subjects and accompanied by downregulated PD-L1-related gene expression levels in the placenta. In the IVF cohort, applying the changing rate of sPD-L1 level after ET achieved a predictive performance for miscarriage with receiver operating characteristics = 0.73 (95% CI: 0.57-0.88, P < 0.01). The study was mainly confined to East Asian pregnant women. Further large prospective pregnancy cohorts are required to validate the predictive performance of sPD-L1 on miscarriage. Reduced circulating sPD-L1 level and downregulated placental PD-L1 expression in miscarriage indicate that dysfunction in PD-L1 signals is a potential underlying mechanism for pregnancy loss. Our findings further extend the importance of the PD-L1 axis in pregnancy maintenance in early pregnancy. This study was financially supported by grants from the Subject Innovation Team of Shaanxi University of Chinese Medicine (2019-Y502), General Research Fund (14122021), and Key Laboratory of Model Animal Phenotyping and Basic Research in Metabolic Diseases (2018KSYS003). The authors declare that they have no competing interests to be disclosed. N/A.

The Possible Clinical Significance of a Decreased Serum Level of Soluble PD-L1 in Discoid Lupus Erythematosus, but Not in Subacute Cutaneous Lupus Erythematosus-A Pilot Study.

Cutaneous lupus erythematosus (CLE) is an autoimmune skin disease with various clinical forms, including the subtypes of discoid lupus erythematosus (DLE) and subacute cutaneous lupus erythematosus (SCLE). The altered function of the programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1) axis in CLE pathogenesis has been suggested. Here, the soluble forms of PD-1 (sPD-1) and PD-L1 (sPD-L1) were explored in untreated DLE and SCLE. Levels of sPD-1 and sPD-L1 were determined by enzyme-linked immunosorbent assay in serums of 21 DLE, 18 SCLE, 13 systemic lupus erythematosus (SLE) patients and 20 healthy controls (HCs). Differences between patient groups and HCs, and the association between clinical activity of skin symptoms and sPD-1/sPD-L1 levels were analyzed with Mann-Whitney U-test and Spearmann's correlation. Regarding sPD-1 levels, no statistically significant differences were found between DLE and SCLE groups, nor compared to HCs. As for sPD-L1, a significantly lower level was found in the DLE group compared to the SCLE and HC groups (p = 0.027 and p = 0.009, respectively). In SLE, significantly higher sPD-1 was found compared to HCs (p = 0.002). No association between skin symptom activity and sPD-1/sPD-L1 levels was found in CLE. Alterations of the inhibitory effect of sPD-L1 on T-cell activity might elucidate the differences between DLE and SCLE.

Elevated programmed cell death-1 protein/ligand (PD-1/PD-L1) and variants are associated with susceptibility to multiple myeloma: a case-control study in the Chinese cohort

Multiple myeloma (MM) is a malignant disorder characterised by progressive immune dysregulation. The importance of programmed cell death protein 1 (PD-1)/programmed cell death ligand 1 (PD-L1) in MM has been documented in various populations, but studies have been limited to the Chinese cohort. In the present study, we examined the role of PD-1/PDL-1 in large cohorts of Chinese patients with MM and healthy controls to reveal a possible association with MM. Three hundred thirty-four MM patients and 202 healthy age-sex-matched subjects were enrolled in the present study. Serum levels of PD-1 and PD-L1 were quantified by ELISA. Percentages of T cells (CD4+ and CD8+ T cells) expressing PD-1 receptor were assessed by flow cytometry. Variants in PD-L1 (rs4143815) and PD-1 gene (rs2227981, rs2227982, rs7421861 and rs11568821) were genotyped by PCR-RFLP method. Patients with multiple myeloma had higher levels of PD-1 and PDL-1 than healthy controls, indicating an important role for programmed cell death protein-1 and its ligand in the pathogenesis of MM. T cells expressing PD-1 receptors were also significantly higher in MM patients than in controls. Mutants for PD-L1 (rs4143815) and PD-1 (rs2227982 and rs7421861) polymorphisms were significantly more common in MM than in HC. Interestingly, PD-L1 (rs4143815) and PD-1 (rs2227982 and rs7421861) variants were linked to higher sPD-L1 and sPD-1 levels, respectively. PD-1/PD-L1 levels are significantly higher in MM patients and could be a promising biomarker for the disease. Variants of PD-L1 and PD-1 are linked to serum-soluble proteins and are associated with the development of MM.

Dynamics of Soluble Forms of the Immune Checkpoint Components PD-1/PD-L1/B7-H3, CD314/ULBP1, and HLA-G in Peripheral Blood of Melanoma Patients Receiving Blockers of Programmed Cell Death Protein PD-1.

The content of the soluble forms of immune checkpoint components sPD-1, sPD-L1 in blood serum, and sB7-H3, sCD314, sULBP1, sHLA-G in blood plasma of 30 melanoma patients receiving immunotherapy with anti-PD-1 antibodies (nivolumab, pembrolisumab) was measured before and in 4 and 8 weeks after the start of immunotherapy. The control group comprised 70 practically healthy donors. Standard immunoassay kits were used. In melanoma patients, the levels of sPD-L1 and sB7-H3 were significantly higher than in the control group (p<0001), sPD-1 level did not differ from the control, while sCD314 and sHLA-G levels were insignificantly decreased. During therapy, opposite changes in the levels of markers in individual patients were observed, and frequently after the initial increase (or decrease) after the first 4 weeks normalization did occur in the further 4 weeks. No statistically significant associations between the initial levels of markers and direction of their changes during treatment were found, but some trends indicating to the potential benefits from assessment of soluble forms of immune checkpoint proteins for evaluation and monitoring of the efficiency of the therapy with immune checkpoint blockers were revealed: significant decrease of sB7-H3 and sPD-1 levels in the course of treatment, higher initial sPD-1 level in patients with future progression than in those with stabilization or partial effect, and lower progression frequency in patients with increasing sPD-1 and sPD-L1 levels than in those with decreasing markers levels.

P-478 The decreases in serum soluble programmed cell death ligand −1 level are associated with missed miscarriage in humans

Abstract Study question The perturbations in serum levels of soluble forms of programmed cell death-1 (sPD-1) and its ligand (sPD-L1) in missed miscarriage (MM) are unknown. Summary answer Dysfunction in PD-L1 signal has been demonstrated in MM as evidenced by a lower circulating sPD-L1 level and declined expressions of PD-L1 in placenta. What is known already Programmed cell death-1 (PD-1) and its ligand (PD-L1) comprise important immune inhibitory checkpoint signaling to maintain pregnancy. During pregnancy, PD-L1 is highly expressed in the trophoblasts of the placenta. The reduced placental PD-L1 expression is associated with the risk of miscarriage. Therefore, disturbance of PD-1/PD-L1 axis in the fetomaternal interface is susceptible to be an important pathogenesis for pregnancy loss. Interestingly, the soluble form of these immune checkpoint molecules is detectable and associated with immunosuppression, it raises the diagnostic potential to monitor their level to monitor pregnancy health. Study design, size, duration In the discovery cohort, we enrolled 108 patients with MM and 115 body mass index (BMI)-matched, healthy women with a full-term pregnancy at 6 to 13 weeks of gestation. Another cohort of 25 MM patients and 25 subjects with induced abortion was recruited for validation. Participants/materials, setting, methods Blood samples were collected at the first prenatal visit for HP women or on the day of dilatation and curettage surgery (D&amp;C) for subjects with MM or IA to measure serum sPD-1 and sPD-L1 levels. Placenta samples were harvested during the D&amp;C within cohort 2 to examine related gene and protein expression. Main results and the role of chance Our results showed that circulating sPD-L1 levels were reduced by 50% in patients with MM (55.71 ± 1.54 pg/mL) compared to HP controls (107.27 ± 5.48 pg/mL, P&amp;lt;0.001) and remained significant after adjusting for maternal age and gestational age, whereas no significant differences in sPD-1 were observed between the two groups. Applying serum sPD-L1 levels achieved an area under the receiver operating characteristics curve (AUROC) of 0.83 (95% CI: 0.77 to 0.88, P&amp;lt;0.001) with the optimal cut-off value of 81.52 pg/ml to detect MM. Our data in the validation cohort further demonstrated that sPD-L1 was lower in MM patients relative to IA subjects. Likewise, placental PD-L1-related gene expressions were downregulated at both mRNA and protein levels in miscarriage samples relative to samples in IA groups and were positively associated with sPD-L1 levels in maternal circulation Limitations, reasons for caution This study was conducted in East Asian women, further studies are required for the evaluation of the serum PD-1 and sPD-L1 levels in different ethnic groups. Secondly, due to the cross-sectional nature of the study design, whether the sPD-L1 levels could utilize as a predictive biomarker is questioned. Wider implications of the findings The maternal serum sPD-L1 level has the potential to be a diagnostic aid for pregnancy maintenance. Moreover, insufficient PD-L1 expression in the feto-maternal interface may act as one of the potential pathogenic events to trigger early pregnancy loss. Trial registration number not applicable

#3789 SERUM AND URINARY SOLUBLE PD-1, PD-L1 AND PD-L2 AS BIOMARKERS FOR CHECKPOINT INHIBITOR RELATED ACUTE INTERSTITIAL NEPHRITIS

Abstract Background and Aims Acute interstitial nephritis (AIN) related to Immune Checkpoint Inhibitors (ICI) is a frequently described adverse effect, although the mechanisms are not fully understood. The gold standard for the diagnosis is kidney biopsy. Although it is invasive, it is necessary as AIN can be confounded with other causes of acute kidney injury (AKI) in oncologic patients. The diagnosis of AIN has therapeutical consequences such as the need to withdraw the drug if confirmed, possibly diminishing patients’ life expectancy. Few biomarkers have been proposed for non-invasive diagnosis or the follow-up of these patients, which could be useful when the biopsy is contraindicated or not available. Our aim was to study the Immune Checkpoint pathway during the event, as well as analyse potential biomarkers directly related to the pathway. Method We performed an observational study. We recruited patients with incident diagnosis of AIN related to ICI in our institution from 2018 to 2022. We obtained serum and urine at the time of diagnosis after signing informed consent. We recruited n = 18 patients with the diagnosis of AIN related to ICI. For comparison, we recruited patients with non ICI-related AIN (n = 18), patients with active ANCA-vasculitis (n = 35), patients with acute tubular necrosis (ATN) (n = 22), and controls (n = 36) (patients with diagnosis of malignancy without AKI (n = 22), and healthy controls (n = 14)). We determined soluble PD-1 (sPD-1), sPD-L1 and sPD-L2 in serum and urine using a multiplex bead-based Luminex assay. Besides, we performed PD-L1 immunohistochemistry and PD-L2 immunofluorescence staining of kidney biopsies from patients with ICI-related AIN (n = 14) and compared to patients with non ICI-related AIN (n = 10). Results Serum sPD-1 was higher in patients with AIN compared to controls (p = 0.0004) and patients with ATN (p = 0.021). There were no differences between controls and ATN (p = 0.76). Urinary sPD-1 was lower in subjects with AKI compared to controls (control vs AIN (p = 0.019), control vs active vasculitis (p&amp;lt;0.0001) and control vs ATN (p = 0.0304)). Patients with ATN exhibited the lowest urinary sPD-1 levels, and there were statistically significant differences when compared to AIN (p = 0.0304). Focusing on oncologic patients, urinary sPD-1 concentration &amp;lt;129.3 pg/ml had a 71.43% sensitivity and 94.12% specificity to differentiate ATN from AIN, with a likelihood ratio of 12.14. Serum and urinary sPD-L1 and sPD-L2 did not show statistically significant differences between AIN and ATN. In kidney biopsies, patients with ICI related AIN showed higher density of PD-L1 positive tubules than patients with non-ICI related AIN (p = 0.037). Proportion of patients having more than 2.64/mm2 PD-L2 positive tubules was higher among patients with ICI related AIN compared to non ICI-related AIN (p = 0.034). There was a positive correlation (p = 0.009, r = 0.72) between urinary sPD-1 and the density of PD-L1 positive tubules (tubules/mm2). As for PD-L2, serum PD-L2 was higher in patients with positive tubular PD-L2 expression (p = 0.009). Conclusion Serum sPD-1 increases in AIN, contrary to patients with ATN and controls. Urinary sPD-1 decreases in patients with AKI. This decrease is more intense in the case of patients with ATN, thus being useful in the differentiation between ATN and AIN, especially in patients with neoplasms. Patients with ICI-related AIN exhibit higher positivity for PD-L1 and PD-L2 staining compared to other causes of AIN. Altogether, our findings suggest a role for these soluble molecules in the ICI-related AIN pathophysiology.

Association of circulating levels of soluble PD-1, PD-1 gene polymorphisms with HBV infection and HBV infection-associated hepatocellular carcinoma

Objective: To investigate the association of circulating sPD-1 level and PD-1 gene polymorphisms with HBV infection and HBV infection-associated hepatocellular carcinoma. Methods: A case-control study was conducted. A total of 237 chronic HBV infection cases and 138 HBV infection-associated hepatocellular carcinoma in the Department of Infectious Diseases of the First Hospital of Shanxi Medical University from 2018 to 2021 were selected as the case group. About 250 individuals who visited a hospital physical examination center for routine physical examination during the same period were selected as the control group. Plasma sPD-1 levels were measured by using an ELISA kit and genotyping was performed by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. The association of sPD-1 levels and PD-1 polymorphisms with HBV infection as well as HBV infection-associated hepatocellular carcinoma was analyzed by using logistic regression models after adjusting for age, sex, alcohol consumption, smoking, ALT and AST levels. The sPD-1 level and PD-1 polymorphisms were independent variables, and HBV infection was the dependent variable. Results: The age of 237 chronic HBV infections, 138 HBV infection-related liver cancer case subjects and 250 control subjects in the study was (49.1±10.8), (51.9±12.7) and (50.7±11.9) years, respectively. Multivariate logistic regression model analysis showed that with a 1 pg/ml increase in sPD-1 level, the OR (95%CI) values for the risk of incident HBV infection cases and HBV hepatocellular carcinoma cases were 1.92 (1.68-2.19) and 2.02 (1.69-2.40). For rs2227981, compared with the CC genotype, the TT genotype had a lower risk of HBV infection and liver cancer associated with HBV infection, with OR (95%CI) values of 0.45 (0.22-0.91) and 0.35 (0.14-0.91). For rs2227982, compared with the CC genotype, the CT and TT genotypes also had a lower risk of HBV infection [OR (95%CI) values of 0.72 (0.53-0.97) and 0.57 (0.35-0.93)] and HBV infection-related liver cancer [OR (95%CI) values of 0.64 (0.45-0.92) and 0.52 (0.29-0.93)]. Conclusions: Plasma sPD-1 levels and PD-1 gene polymorphisms are associated with HBV infection and HBV infection-associated hepatocellular carcinoma.

Clinical roles of soluble PD-1 and PD-L1 in plasma of NSCLC patients treated with immune checkpoint inhibitors.

Immune checkpoint inhibitors (ICIs) have significantly improved the prognosis of non-small cell lung cancer (NSCLC). However, only a limited proportion of patients can benefit from this therapy, and clinically useful predictive biomarkers remain to be elucidated. Blood was collected from 189 patients with NSCLC before and six weeks after the initiation of ICI treatment (anti-PD-1 or anti-PD-L1 antibody). Soluble PD-1 (sPD-1) and PD-L1 (sPD-L1) in plasma before and after treatment were analyzed to evaluate their clinical significance. Cox regression analysis demonstrated that higher sPD-L1 levels before treatment significantly predicted unfavorable progression-free survival (PFS; HR 15.4, 95% CI 1.10-86.7, P = 0.009) and overall survival (OS; HR 11.4, 95% CI 1.19-52.3, P = 0.007) in NSCLC patients treated with ICI monotherapy (n = 122) but not in those treated with ICIs combined with chemotherapy (n = 67: P = 0.729 and P = 0.155, respectively). In addition, higher sPD-1 levels after treatment were significantly associated with better OS (HR 0.24, 95% CI 0.06-0.91, P = 0.037) in patients treated with anti-PD-1 monotherapy, whereas higher sPD-L1 levels after treatment were significantly associated with worse PFS (HR 6.09, 95% CI 1.42-21.0, P = 0.008) and OS (HR 42.6, 95% CI 6.83-226, P < 0.001). The levels of sPD-L1 at baseline closely correlated with those of other soluble factors, such as sCD30, IL-2Ra, sTNF-R1, and sTNF-R2, which are known to be released from the cell surface by zinc-binding proteases ADAM10/17. These findings suggest the clinical significance of pretreatment sPD-L1 as well as posttreatment sPD-1 and sPD-L1 in NSCLC patients treated with ICI monotherapy.

CD4+ T cell counts and soluble programmed death-1 at baseline correlated with hepatitis B surface antigen decline in HIV/HBV coinfection during combined antiretroviral therapy.

Several studies have described the rapid decline and clearance of hepatitis B surface antigen (HBsAg) in human immunodeficiency virus (HIV)/hepatitis B virus (HBV) coinfection after initiating combined antiretroviral therapy (cART). Early decline of HBsAg levels is associated with HBsAg seroclearance in the treatment of chronic HBV infection. This study aims to evaluate the HBsAg kinetics and the determinants of early HBsAg decline in patients with HIV/HBV coinfection during cART. A total of 51 patients with HIV/HBV coinfection were enrolled from a previously established HIV/AIDS cohort and followed for a median of 59.5 months after cART initiation. Biochemical tests, virology and immunology assessments were measured longitudinally. The kinetics of HBsAg during cART were analyzed. Soluble programmed death-1 (sPD-1) levels and immune activation markers (CD38 and HLA-DR) were measured at baseline, 1-year and 3-year during treatment. HBsAg response was defined as a decline of more than 0.5 log10 IU/ml at 6 months from the baseline after initiation of cART. HBsAg declined faster (0.47 log10 IU/mL) in the first six months and attained a decrease of 1.39 log10 IU/mL after 5-year therapy. Seventeen (33.3%) participants achieved a decline of more than 0.5 log10 IU/ml at the first 6 months of cART(HBsAg response) of which five patients achieved HBsAg clearance at a median of 11 months (range: 6-51 months). Multivariate logistic analysis showed the lower baseline CD4+ T cell levels (OR=6.633, P=0.012) and sPD-1 level (OR=5.389, P=0.038) were independently associated with HBsAg response after cART initiation. The alanine aminotransferase abnormality rate and HLA-DR expression were significantly higher in patients who achieved HBsAg response than in those who did not achieve HBsAg response after cART initiation. Lower CD4 + T cells, sPD-1, and immune activation were related to a rapid HBsAg decline in patients with HIV/HBV-coinfection after the initiation of cART. These findings imply that immune disorders induced by HIV infection may disrupt immune tolerance to HBV, leading to a faster decline in HBsAg levels during coinfection.

Soluble forms of PD-1 and sPD-L1/2 in serum and urine of patients with head and neck cancer and their clinical significance

ABSTRACT This study explored the soluble forms of PD-1 and sPD-L1/2 in serum and urine of patients with head and neck cancer (HNCs) and associated the data with clinical state and 5-year survival. The sPD-1 and sPD-L1/2 levels were evaluated by ELISA in sufferers (N=110) and normal controls (N=82). Patients in the case group were more likely to be male smokers or former smokers. Compared with the normal control group, the serum levels of sPD-1, sPD-L1 and sPD-L2 and the urine level of sPD-L1 in patients with HNCs were increased. Furthermore, sPD-1 and sPD-L1 serum levels existed a positive connection, and sPD-1 and sPD-L2 serum levels positively correlated in HNCs sufferers. The urine sPD-1 and sPD-L1 had a positive relationship. sPD-1 serum levels had a positive connection with urine sPD-1, sPD-L1 urine levels had a positive relationship with sPD-L1, and sPD-L2 serum levels positively connected to urine sPD-L2. Lower serum sPD-1 and sPD-L1/L2 were associated with disease progression and survival at the examination time. sPD-1 and sPD-L1/L2 serum levels above median were markedly related to a decreased probability of 5-years OS in patients with HNCs. The sPD-1 and sPD-L1/2 were complementary markers representing clinical condition and illness outcomes for HNCs patients. The sPD-L1 might accelerate the characterization of high-risk patients with disapproving illness outcomes. sPD-1 and sPD-L1/2 could be easily accessed through liquid biopsy. The incorporation of them as indicators for risk evaluation throughout treatment scheduling and follow-up seems to be an appreciated method.

Abstract 3013: The immunogenetic basis of hepatitis B virus-related hepatocellular carcinoma

Abstract Background: The reported associations of genetic variants within the human leukocyte antigen (HLA) region with hepatis B virus (HBV)-related hepatocellular carcinoma (HCC) are not consistent, and the structure-function patterns involved in this disease remain unclear. Methods: Utilizing genotype data from 706 HBV-related HCC cases and 6197 chronic HBV carriers without HCC collected from three resources in Taiwan, we imputed the classical HLA alleles at 2-field resolution and analyzed the molecular landscape of HLA complexes by deeply dissecting their genotypic configurations, functional divergence, and HBV-antigen binding capabilities in associations with the HCC risk. Logistic regression models were used, and all models were adjusted for sex, age, and population structure using the top 10 principal components. We also tested the underlying biological implications of our findings by investigating the HLA binding affinities and HLA effect on control of viral replication (i.e, HBV DNA load), viral population diversity (i.e, quasispecies), and inflammation (i.e, chemokines, cytokines, and soluble programmed cell death 1 [sPD-1]-associated T cell exhaustion). Results: We found high accuracy (≥95% concordance) of HLA imputation at all HLA class I and class II alleles. The strongest evidence for an association with HCC was observed for HLA-DQB1*03:01 (adjusted OR= 1.35 in an additive genetic model, P=1.3×10-6). Three amino acids at specific positions within or near the peptide-binding groove of DQB1 molecules might explain the observed allele association, which leads to loss of binding HBV nucleocapsid protein (i.e., low binding affinity predicted by NetMHCIIpan 4.0 for HLA-DQB1*03:01). HLA-DQB1*03:01 was significantly associated with a higher HBV DNA load (P= 4.3×10-4) and an increased level of serum sPD-1 (P=6.8×10-4), but not with increased levels of chemokines, cytokines, and viral complexity (i.e., Shannon entropy for HBV quasispecies, all P values&amp;gt;0.05). Analysis of HLA zygosity confirmed that HLA-DQB1 heterozygosity was associated with a decreased risk of HCC (adjusted OR=0.77, P= 0.015). Analysis of HLA evolutionary divergence (HED) showed that high HED at HLA-DRB1 loci was marginally associated with a decreased risk of HCC among HLA-DQB1*03:01 non-carriers (P=0.05). Conclusion: Our data shed light on the immunogenetic risk associated with HBV-related HCC. The effect of HLA-DQB1*03:01 could be partly explained by its low binding affinity with HBV nucleocapsid antigen, and/or its effect on viral load control and sPD-1, but not by its control on other inflammation markers (chemokines and cytokines) and viral population diversity. The association between HLA-DQB1 heterozygosity and HCC risk could be independent of the HLA-DQB1*03:01 effect, which indicates underlying mechanisms related to increased immunosurveillance in clearing potential viral antigens or neoantigens. Citation Format: Zhiwei Liu, Chih-Jen Huang, Yu-Han Huang, Mei-Hung Pan, Mei-Hsuan Lee, Mathias Viard, Allan Hildesheim, Ruth M. Pfeiffer, Mary Carrington, Chien-Jen Chen, Tobias L. Lenz, Hwai-I Yang. The immunogenetic basis of hepatitis B virus-related hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3013.

Soluble immune checkpoints as correlates for HIV persistence and T cell function in people with HIV on antiretroviral therapy.

In people with HIV (PWH) both off and on antiretroviral therapy (ART), the expression of immune checkpoint (IC) proteins is elevated on the surface of total and HIV-specific T-cells, indicating T-cell exhaustion. Soluble IC proteins and their ligands can also be detected in plasma, but have not been systematically examined in PWH. Since T-cell exhaustion is associated with HIV persistence on ART, we aimed to determine if soluble IC proteins and their ligands also correlated with the size of the HIV reservoir and HIV-specific T-cell function. We used multiplex bead-based immunoassay to quantify soluble programmed cell death protein 1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin domain and mucin domain 3 (TIM-3), PD-1 Ligand 1 (PD-L1) and PD-1 Ligand 2 (PD-L2) in plasma from PWH off ART (n=20), on suppressive ART (n=75) and uninfected controls (n=20). We also quantified expression of membrane-bound IC and frequencies of functional T-cells to Gag and Nef peptide stimulation on CD4+ and CD8+ T-cells using flow cytometry. The HIV reservoir was quantified in circulating CD4+ T-cells using qPCR for total and integrated HIV DNA, cell-associated unspliced HIV RNA and 2LTR circles. Soluble (s) PD-L2 level was higher in PWH off and on ART compared to uninfected controls. Higher levels of sPD-L2 correlated with lower levels of HIV total DNA and higher frequencies of gag-specific CD8+ T-cells expressing CD107a, IFNγ or TNFα. In contrast, the concentration of sLAG-3 was similar in uninfected individuals and PWH on ART, but was significantly elevated in PWH off ART. Higher levels of sLAG-3 correlated with higher levels of HIV total and integrated DNA, and lower frequency of gag-specific CD4+ T cells expressing CD107a. Similar to sLAG-3, levels of sPD-1 were elevated in PWH off ART and normalized in PWH on ART. sPD-1 was positively correlated with the frequency of gag-specific CD4+ T cells expressing TNF-a and the expression of membrane-bound PD-1 on total CD8+ T-cells in PWH on ART. Plasma soluble IC proteins and their ligands correlate with markers of the HIV reservoir and HIV-specific T-cell function and should be investigated further in in large population-based studies of the HIV reservoir or cure interventions in PWH on ART.

Soluble programmed cell death ligand 1 predicts prognosis for gastric cancer patients treated with nivolumab: Blood-based biomarker analysis for the DELIVER trial

BackgroundThe clinical value of soluble forms of programmed cell death–1 (sPD-1), PD ligand 1 (sPD-L1) and cytotoxic T lymphocyte–associated protein–4 (sCTLA-4) for gastric cancer (GC) patients treated with nivolumab monotherapy has remained unknown. MethodsBlood samples collected before nivolumab treatment from 439 GC patients enrolled in the DELIVER (Japan Clinical Cancer Research Organisation GC-08) trial were analysed for sPD-1, sPD-L1 and sCTLA-4. Corresponding baseline clinical data were also retrieved. ResultsHigher plasma levels of sPD-1 (hazard ratio [HR] = 1.27, p = 0.020), sPD-L1 (HR = 1.86, p < 0.001) and sCTLA-4 (HR = 1.33, p = 0.008) were significantly associated with shorter overall survival (OS), whereas only higher sPD-L1 levels was significantly associated with shorter progression-free survival (HR = 1.30, p = 0.008). The sPD-L1 concentration was significantly associated with the Glasgow prognostic score (GPS) (p < 0.001), but both sPD-L1 (HR = 1.67, p < 0.001) and GPS (HR = 1.39, p = 0.009 for GPS 0 versus 1; HR = 1.95, p < 0.001 for GPS 0 versus 2) were independently associated with OS. Patients with a GPS of 0 and low sPD-L1 thus showed the longest OS (median, 12.0 months) and those with a GPS of 2 and high sPD-L1 showed the shortest OS (median, 3.1 months), yielding a HR of 3.69 (p < 0.001). ConclusionBaseline sPD-L1 levels have the potential to predict survival for advanced GC patients treated with nivolumab, with the prognostic accuracy of sPD-L1 being improved by its combination with GPS.

Role of soluble forms of follicular helper T-cell membrane molecules in the pathogenesis of myasthenia gravis

This study examined the role of Tfh and Treg associated molecules also known as checkpoint molecules, their ligands, and IL-21 in myasthenia gravis (MG) pathogenesis. Serum levels of sPD-1, sPD-L1, sICOS, sICOSLG, sCTLA4, and IL-21 were measured in 39 patients with acetylcholine receptor (AChR) antibody-positive generalized MG and 27 controls. sPD-1 and IL-21 levels were higher in MG patients than in controls. Additionally, sPD-1 levels correlated positively with the levels of IL-21, sICOSLG, sCTLA4, and AChR antibody titers. sICOS are correlated with MGADL and AChR antibody titers. These Tfh associated molecules could be used as biomarkers of MG disease activity.

Prognostic role of soluble PD-1 and BTN2A1 in overweight melanoma patients treated with nivolumab or pembrolizumab: finding the missing links in the symbiotic immune-metabolic interplay.

Individual response to immune checkpoint inhibitors (ICIs) is currently unpredictable in patients with melanoma. Recent findings highlight a striking improvement in the clinical outcomes of overweight/obese patients treated with ICIs, which seems driven, at least in part, by programmed cell death protein 1 (PD-1)-mediated T-cell dysfunction. A putative role of butyrophilins (BTNs) is under investigation as a novel mechanism of cancer immune evasion and obesity-associated inflammation. This study investigates the role of baseline plasma levels of soluble PD-1 (sPD-1), soluble programmed cell death ligand 1 (sPD-L1), BTN2A1 (sBTN2A1), BTN3A1 (sBTN3A1), along with body mass index (BMI), as predictive biomarkers of immunotherapy response in metastatic melanoma patients treated with nivolumab or pembrolizumab as first-line treatment. In all, 41 patients were included in the study. The baseline plasma level of sPD-1 was significantly lower, and the sBTN2A1 was significantly higher, in long-responder patients to nivolumab or pembrolizumab (median sPD-1: 10.3 ng/ml versus 16.6 ng/ml, p = 0.001; median sBTN2A1: 4.4 ng/ml versus 3.77 ng/ml, p = 0.004). Lower levels of sPD-1 and higher levels of sBTN2A1 were also significantly associated with better overall response rate. Notably, when we further stratified the study cohort using BMI along with sPD-1, patients with BMI ⩾ 25 and sPD-1 < 11.24 ng/ml had longer time to treatment failure after PD-1 inhibitor than other subgroups of patients (p < 0.001). Circulating sPD-1 and sBTN2A1 detection, along with BMI, could give more insights into the immune-metabolic interactions underlying the benefit observed in overweight/obese patients, improving the use of dynamic, noninvasive, biomarkers for patient selection.

Soluble PD-L1 Predict Poor Outcome and Disease Progression in Patients with De Novo Myelodysplastic Syndrome

Background: Myelodysplastic syndromes (MDS) are a group of myeloid neoplasms characterized by clonal proliferation of hematopoietic stem cells, recurrent genetic abnormalities, myelodysplasia, ineffective hematopoiesis, peripheral-blood cytopenia, and a high risk of evolution to acute myeloid leukemia (AML). Mounting evidence indicates that dysregulating immune checkpoint pathways are implicated in the pathogenesis of MDS, which thus promoting a number of trials employing immune checkpoints inhibitors (ICIs) in different settings. Soluble ICs (sICs) are soluble forms of ICs, which can be measured from circulation. In solid tumors, sICs have been identified with roles in predicting prognosis and mediating resistance to ICIs. However, in MDS, it is still unclear whether sICs levels are dysregulating in this disease and already has a prognostic influence at the time of diagnosis. Method: Seventy-nine adult patients with untreated de novo MDS were enrolled in this prospective study and managed upon clinical prognostic risk assessment by Revised International Prognostic Scoring System (IPSS-R), 10 healthy donors (HD) were enrolled as control. Profile of sICs (sPD-1, sPD-L1, sCTLA-4, sLAG-3, sTIM-3, sGITR, sOX40, s4-1BB, sST-2) and cytokines (IFN-γ, IFN-α2, IL-2, IL-2α, IL-6, IL-7, IL-10, IL-15, IL-17, TNF-α) were tested by ELISA, and the expression levels of PD-1 on T cells (CD3+, CD4+, CD8+, DPT, DNT）were measured using flow cytometry (FCM). Follow-up time ended on June 21, 2022. Result: MDS patients showed significant higher levels of sPD-1 and sPD-L1 (P＜0.01). Patients with high pretreatment sPD-L1 (>73.79 pg/mL) and high percent of PD-1+CD8+T cells (>7.3%) had worse OS (P < 0.05). Using a multivariate cox regression model adjusting established prognostic factors, including IPSS-R and transfusion dependency, high pretreatment sPD-L1 level was retained as a negative independent prognostic factor for OS (HR 3.151, 95%CI 1.061-9.364, P = 0.039). Moreover, to explore the roles of the PD-1/PD-L1 pathway in the pathogenesis of MDS, we examined the correlations between sPD-1, sPD-L1 and percent of PD-1+CD8+T cells and parameters associating with disease progression (Table. 1). Among clinico-pathological features, male patients showed higher levels of sPD-L1 and percent of PD-1+CD8+T cells, lower hemoglobin (＜80 g/L) significantly correlated to high sPD-L1 levels. In addition, in low- and very low- risk group according to IPSS-R, levels of sPD-L1 were significantly lower than intermediate-, high- and very high- risk group. Furthermore, in previous studies, it was reported that mPD-1/mPD-L1 can be induced by multiple inflammatory cytokines in cancer, including IFN-γ, IFN-α2, IL-2, IL-2α, IL-6, IL-7, IL-10, IL-15, IL-17, TNF-α, and we further tested them in our study. Correlation analysis showed that significant correlations were observed among different sICs as well as cytokines (Figure. 1). In addition, a positive correlation was found between TNF-α and sICs, including sPD-1, sPD-L1 and sST2 (P < 0.05). Conclusions: In conclusion, sPD-L1 was a negative independent prognostic factor for MDS patients at the time of diagnosis. High levels of sPD-L1 were presented in patients with lower hemoglobin and higher risk group, associating with disease progressing. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Sex difference in the expression of PD-1 of non-small cell lung cancer.

Evidence increasingly indicated that lung cancer incidence in female individuals continue to rise, and women have a higher risk to develop adenocarcinoma than men. Male and female individuals differ in their innate and adaptive immune responses, and there are sex differences in response to the PD-1/PD-L1-dependent blocking immunotherapy. Whether the differential expression of PD-1 between genders affect the response to blocking treatment is currently unknown. In this study, we examined sex differences in serum sPD-1, mPD-1 expression on T cells, and sex hormone levels in non-small cell lung cancer (NSCLC) patients. Our results revealed a higher level of sPD-1 and expression of PD-1 on CD4+T cell in female patients than in male patients; we identified that serum sPD-1 level and the expression of mPD-1 on T cells were significantly reduced in NSCLC; we also found that serum testosterone level increased in female patients compared with control subjects and that increased testosterone downregulated the expression of mPD-1 on T cell. These findings provide a better understanding of the differences in PD-1 expression between genders in NSCLC patients and the effect of sex hormones on PD-1 expression and supply evidence for early lung cancer diagnosis and responsiveness to immune checkpoint inhibitors.

Effect of tacrolimus on soluble costimulatory molecules in patients with refractory myasthenia gravis

ObjectivesTo investigate the expression and possible role of soluble costimulatory molecules in the treatment of refractory myasthenia gravis. MethodsThirty-two patients with refractory myasthenia gravis were enrolled into this study and given tacrolimus 3 mg/day. At the beginning of treatment and 12 months follow-up period, clinical data were collected and recorded. The clinical classification of myasthenia gravis Foundation (MGFA) was performed. The MGFA-quantitative myasthenia gravis score (MGFA-QMGS), manual muscle test (MMT), MG activity of daily living (MG-ADL) and the activity of daily living (MG-ADL), the 15-item myasthenia gravis quality of life (MG QOL-15) and the dose change of prednisone were used to evaluate the efficacy. The expression levels of soluble costimulatory molecules and their ligands (sPD-1/sPD-L1, sICOS/sICOSL, sCD40/sCD40L), soluble CD25 and IL-2 in serum were determined by enzyme-linked immunosorbent assay (ELISA). ResultsWe observed that oral administration of 3 mg tacrolimus daily for 1 year can significantly improve the clinical symptoms of patients with refractory myasthenia gravis, which is characterized by a significant reduction in clinical scores, such as QMG, MMT, ADL, MGQOL-15, and a reduction daily oral prednisolone (PSL) dose (P < 0.0001).We also found that the levels of plasma sPD-1, sCD40, IL-2 in refractory MG patients increased significantly, and those decreased significantly 12 months after tacrolimus treatment (P < 0.05). The level of sCD25 was negatively correlated with clinical severity scores (P < 0.05). After tacrolimus treatment, the level of sPD-L1 increased although there was no significant difference. ConclusionTacrolimus could relieve the symptoms of refractory MG and significantly decrease the levels of plasma sPD-1, sICOSL, sCD40, sCD25 and IL-2. Soluble costimulatory molecules might be potential biomarkers for MG and tacrolimus treatment.

Soluble PD-L1 in blood correlates positively with neutrophil and negatively with lymphocyte mRNA markers and implies adverse sepsis outcome

Sepsis causes a myriad of immunological reactions that result in life-threatening alterations in the human body. Immunosuppression in sepsis is partly attributed to the programmed death receptor (PD-1) and its associated ligand (PD-L1) via the regulation of lymphocytes and neutrophils. Although the soluble forms of these proteins (i.e., sPD-1 and sPD-L1, respectively) are recognized as possible sepsis biomarkers, their functional implications are yet to be elucidated. Our research assessed the correlation between sPD-1 and sPD-L1 and blood mRNA markers and sepsis outcome. Blood samples of septic patients of urogenital origin versus control patients (both groups: n = 18) were analyzed. Blood serum sPD-1 and sPD-L1 levels were determined using the enzyme-linked immunosorbent assay (ELISA). The whole blood mRNA concentrations of PD-1, PD-L1, neutrophil markers (CEACAM8 and MPO), and T-lymphocyte markers (TCRβ, CD4 and CD8) were determined via reverse transcriptase quantitative PCR (RT-qPCR). sPD-L1 levels were significantly increased in septic patients when compared to the controls, whereas sPD-1 levels were unaltered. Patients with high sPD-L1 levels, as dichotomized to the median, had a significantly shorter survival rate than those with low sPD-L1 levels. The sensitivity/specificity characteristics of sPD-L1 proved significant for sepsis detection. Furthermore, sPD-L1 correlated with the mRNA concentrations of PD-L1, CEACAM, and MPO, as well as major inflammatory markers (C-reactive protein and procalcitonin). However, sPD-L1 negatively correlated with TCRβ, CD4, and CD8 mRNAs. sPD-L1 was found to be significantly increased in septic patients. Notably, sPD-L1 correlated with PD-L1 mRNA and neutrophil markers and was indicative of adverse outcomes.

Characterization of multiple soluble immune checkpoints in individuals with different Mycobacterium tuberculosis infection status and dynamic changes during anti-tuberculosis treatment

BackgroundImmune checkpoints are crucial for the maintenance of subtle balance between self-tolerance and effector immune responses, but the role of soluble immune checkpoints (sICs) in Mycobacterium tuberculosis (M. tb) infection remains unknown. We assessed the levels of multiple sICs in individuals with distinct M. tb infection status, and their dynamic changes during anti-tuberculosis treatment.MethodsWe enrolled 24 patients with pulmonary tuberculosis, among which 10 patients were diagnosed with tuberculous pleurisy (TBP), 10 individuals with latent tuberculosis infection (LTBI), and 10 healthy volunteers from Wuxi Fifth People’s Hospital and Huashan Hospital between February 2019 and May 2021. Plasma concentrations of thirteen sICs were measured at enrollment and during anti-tuberculosis treatment using luminex-based multiplex assay. sICs levels in tuberculous pleural effusion (TPE) and their relations to laboratory test markers of TPE were also assessed in TBP patients.ResultsThe circulating levels of sPD-1, sPD-L1, sCTLA-4, sBTLA, sGITR, sIDO, sCD28, sCD27 and s4-1BB were upregulated in tuberculosis patients than in healthy controls. A lower sPD-L1 level was found in LTBI individuals than in tuberculosis patients. In TBP patients, the levels of sPD-1, sPD-L2, sCD28, sCD80, sCD27, sTIM-3, sLAG-3, sBTLA, s4-1BB and sIDO increased significantly in TPE than in plasma. In TPE, sBTLA and sLAG-3 correlated positively with the adenosine deaminase level. sIDO and sCD80 correlated positively with the lactate dehydrogenase level and the percentage of lymphocytes in TPE, respectively. Meanwhile, sCD27 correlated negatively with the specific gravity and protein level in TPE. In tuberculosis patients, the circulating levels of sBTLA and sPD-L1 gradually declined during anti-tuberculosis treatment.ConclusionsWe characterized the changing balance of sICs in M. tb infection. And our results revealed the relations of sICs to laboratory test markers and treatment responses in tuberculosis patients, indicating that certain sICs may serve as potential biomarkers for disease surveillance and prognosis of tuberculosis.