Immune Profile Of Patients Research Articles

The interactive association between Immune system and Kidney function

The Immune System (IS) and Kidney function are closely and interactively connected. IS dysfunction happening in autoimmune diseases, infections, malignancies, etc. is implicated in the pathogenesis of kidney involvement, leading to a great spectrum of glomerular and interstitial injury through several immunological pathways, such as immune complex deposition, activation of complement and signaling pathways. Complications of chronic kidney disease (CKD), including accumulation of uremic toxins, O2 free radicals, advanced glycation end products, increased expression of Toll Like Receptors on monocytes, cytokine overproduction, result in a situation characterized by chronic inflammation combined with premature ageing, and usually designated as “inflamm-aging”, followed by detrimental clinical consequences in CKD patients, such as increased cardiovascular risk, susceptibility to infections and malignancies. Kidney transplant can potentially restore the immune profile of patients, albeit immunosuppression treatment may be followed by further complications. The presence of certain T cell subsets at time of renal transplantation may affect response to immunosuppression and acute or chronic rejection, suggesting that patients’ immune profile at time of transplantation may have substantial impact in short and long term graft function.

Immune effects of CDK4/6 inhibitors in patients with HR+/HER2- metastatic breast cancer: Relief from immunosuppression is associated with clinical response.

SummaryBackgroundCyclin-dependent kinase 4/6 inhibitors (CDK4/6i) are innovative small target molecules that, in combination with endocrine therapy, have recently been employed in the treatment of patients with HR+/HER2− metastatic breast cancer (mBC). In this prospective study, we investigate the impact of CDK4/6i on the immune profile of patients with HR+/HER2− mBC.MethodsImmune cell subsets were analysed using flow cytometry of peripheral blood mononuclear cells (PBMCs) isolated from patients with HR+/HER2− mBC, both before and during treatment. Regulatory T cells (Tregs) were identified using the markers CD4, CD25, CTLA4, CD45RA, and intracellular FOXP3. Monocytic and polymorphonuclear myeloid-derived suppressor cells (M-MDSCs and PMN-MDSCs) and other immune populations were analysed using CD45, CD14, CD66b, CD11c, HLA-DR, CD3, CD8, CD28, CD137, PD1, CD45RA, CCR7, and Ki67.FindingsThe percentage of circulating Tregs and M/PMN-MDSCs was significantly downregulated from baseline during CDK4/6i-treatment (p<0.0001 and p<0.05, respectively). In particular, the effector Treg subset (CD4+CD25+FOXP3highCD45RA−) was strongly reduced (p<0.0001). The decrease in Treg levels was significantly greater in responder patients than in non-responder patients. Conversely, CDK4/6i treatment was associated with increased levels of CD4+ T cells and anti-tumour CD137+CD8+ T cells (p<0.05).InterpretationCDK4/6i treatment results in downregulation of Tregs, M-MDSCs, and PMN-MDSCs, thus weakening tumour immunosuppression. This decrease is associated with response to treatment, highlighting the importance of unleashing immunity in cancer treatment efficacy. These results suggest a novel mechanism of immunomodulation in mBC and provide valuable information for the future design of novel treatments combining CDK4/6i with immunotherapy in other cancer settings.FundingSapienza University of Rome.

Systemic cytokines, chemokines and growth factors reveal specific and shared immunological characteristics in infectious cardiomyopathies

Heart disease is a major cause of death worldwide. Chronic Chagas cardiomyopathy (CCC) caused by infection with Trypanosoma cruzi leading to high mortality in adults, and rheumatic heart disease (RHD), resulting from infection by Streptococcus pyogenes affecting mainly children and young adults, are amongst the deadliest heart diseases in low-middle income countries. Despite distinct etiology, the pathology associated with both diseases is a consequence of inflammation. Here we compare systemic immune profile in patients with these cardiopathies, to identify particular and common characteristics in these infectious heart diseases. We evaluated the expression of 27 soluble factors, employing single and multivariate analysis combined with machine-learning approaches. We observed that, while RHD and CCC display higher levels of circulating mediators than healthy individuals, CCC is associated with stronger immune activation as compared to RHD. Despite distinct etiologies, univariate analysis showed that expression of TNF, IL-17, IFN-gamma, IL-4, CCL4, CCL3, CXCL8, CCL11, CCL2, PDGF-BB were similar between CCC and RHD, consistent with their inflammatory nature. Network analysis revealed common inflammatory pathways between CCC and RHD, while highlighting the broader reach of the inflammatory response in CCC. The final multivariate model showed a 100% discrimination power for the combination of the cytokines IL-12p70, IL-1Ra, IL-4, and IL-7 between CCC and RHD groups. Thus, while clear immunological distinctions were identified between CCC and RHD, similarities indicate shared inflammatory pathways in these infectious heart diseases. These results contribute to understanding the pathogenesis of CCC and RHD and may impact the design of immune-based therapies for these and other inflammatory cardiopathies that may also share immunological characteristics.

Deep dissection of the antiviral immune profile of patients with COVID-19

In light of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) variants potentially undermining humoral immunity, it is important to understand the fine specificity of the antiviral antibodies. We screened 20 COVID-19 patients for antibodies against 9 different SARS-CoV-2 proteins observing responses against the spike (S) proteins, the receptor-binding domain (RBD), and the nucleocapsid (N) protein which were of the IgG1 and IgG3 subtypes. Importantly, mutations which typically occur in the B.1.351 “South African” variant, significantly reduced the binding of anti-RBD antibodies. Nine of 20 patients were critically ill and were considered high-risk (HR). These patients showed significantly higher levels of transforming growth factor beta (TGF-β) and myeloid-derived suppressor cells (MDSC), and lower levels of CD4+ T cells expressing LAG-3 compared to standard-risk (SR) patients. HR patients evidenced significantly higher anti-S1/RBD IgG antibody levels and an increased neutralizing activity. Importantly, a large proportion of S protein-specific antibodies were glycosylation-dependent and we identified a number of immunodominant linear epitopes within the S1 and N proteins. Findings derived from this study will not only help us to identify the most relevant component of the anti-SARS-CoV-2 humoral immune response but will also enable us to design more meaningful immunomonitoring methods for anti-COVID-19 vaccines.

Immunomodulation with β-glucan from Pleurotus ostreatus in patients with endocrine-dependent breast cancer.

Aim: To evaluate the effect of pleuran (β-glucan from Pleurotus ostreatus) administration on the immune profile of patients with endocrine-dependent breast cancer (clinical stages I-II) in clinical and imaging remission. Methodology: Antitumor cellular immunity (CD19+, CD3+, CD4+ and CD8+ T lymphocytes and natural killercells) of 195 patients (49 in the pleuran group and 146 in the control group) was measured by flow cytometry. Results: We observed a significant increase in the absolute number of CD3+, CD19+, CD4+ and CD8+ T lymphocytes in the pleuran group compared with the control group. Conclusion: Our results suggest potential benefit of continuous pleuran administration on immune rehabilitation of cellular antitumor immunity and better prognosis in breast cancer patients in remission.

Alteration of immune profiles is associated with pulmonary function and symptoms in patients with chronic obstructive pulmonary disease

Inflammation serves a key role in chronic obstructive pulmonary disease (COPD). However, changes in the immune profiles of patients with COPD remain unclear. The present prospective observational study aimed to determine the expression profiles of immune cells and inflammatory factors of patients with COPD and healthy controls, and to analyze the relationship between immune profiles and smoking history. A total of 140 subjects were enrolled in the present study between September 2018 and April 2019 at West China Hospital of Sichuan University (Chengdu, China). These included 87 patients with stable COPD and 24 patients with acute exacerbated COPD, as well as 29 healthy controls. Baseline characteristics were recorded during the screening period, and levels of immune cells were examined using flow cytometry. In addition, levels of inflammatory factors were measured using ELISAs. The results revealed increased expression of the CD64+/CD14+ mononuclear phagocyte system (MPS) and CD16+CD66+ neutrophils, and decreased expression of CD3+CD4+ T cells and CD3+ CD8+ T cells (all P<0.05) in the peripheral blood of patients with COPD and smokers relative to non-smoking controls. In addition, significant differences were observed in protein levels of IL-6, IL-1β, TNF-α, TGF-α, IFN-γ, IL-8, IL-17A and CRP among the three groups (all P<0.05). Furthermore, the IL-17A, TNF and NF-κB signaling pathways were found to serve a key role in the inflammatory network of COPD. Pearson's correlation analysis revealed a positive correlation between CD3+T lymphocyte counts and pulmonary function, and a negative correlation with smoking and exacerbations. Furthermore, neutrophils and MPS were negatively associated with pulmonary function, and IL-8 was positively associated with cough. There was also a negative association between CRP and IL-17A with pulmonary function but a positive correlation with symptoms and exacerbation. Club cell secretory protein was also negatively associated with emphysema parameters. In conclusion, the present findings revealed significant differences in profiles of immune factors among patients with COPD, smokers and non-smoking controls and their association with clinical characteristics. The clinical trial registration number of the present study is: ChiCTR1800015700 (registered with http://www.chictr.org.cn/index.aspx, 2018/04/16).

Cereblon pathway biomarkers and immune profiles in patients with myeloma receiving post-ASCT lenalidomide maintenance (LEOPARD)

LEOPARD was a single arm, phase II study of lenalidomide (LEN) and alternate day prednisolone maintenance in patients with newly diagnosed multiple myeloma (MM) following autologous stem cell transplantation (ASCT). Sixty patients were enrolled. Estimated median potential follow-up was 44 m, median PFS was 38.3 m, median OS was not reached (landmark 36 m OS: 71.4%). Correlative immunohistochemistry performed on pre-ASCT trephines demonstrated high MM tumor cereblon (total/cytoplasmic) was associated with superior OS (p = .045, p = .031, respectively), whereas high c-Myc was associated with inferior PFS (p = .04). Patients with high cereblon (total/nuclear) were more likely to improve depth of response, whereas patients with high c-Myc were less likely, suggesting alternative/more effective post-ASCT strategies for patients with high c-Myc need identification. Peripheral blood immune profiling (mass cytometry) informed a more sustained response to LEN maintenance, demonstrating enrichment of activated/cytotoxic NK cells and cytotoxic T cells in patients with durable responses, contrasting with enrichment of B-regs in early relapsers.

Immune Profile of Patients (pts) with Relapsed and/or Refractory Multiple Myeloma (RRMM) Treated with Pomalidomide (POM) + Low-Dose Dexamethasone (LoDEX) + Daratumumab (DARA) in the Second Line Immediately after Lenalidomide (LEN)

Immune Profile of Patients (pts) with Relapsed and/or Refractory Multiple Myeloma (RRMM) Treated with Pomalidomide (POM) + Low-Dose Dexamethasone (LoDEX) + Daratumumab (DARA) in the Second Line Immediately after Lenalidomide (LEN)

60-LB: Dendritic Cells Engineered with Tailored Multiepitopes-Encoding mRNA as Precision Medicine for Immunotherapy of Type 1 Diabetes

Type 1 diabetes (T1D) is a complex autoimmune disease characterized by heterogeneity in genetic risk factors and immune profiles of patients and for which a cure or approved treatment is not yet available. Antigen-specific immunotherapy (ASIT) offers a targeted treatment of T1D that selectively inhibits autoreactive T cells. So far, ASIT trials showed limited efficacy, as only a subset of patients is responsive. Thus, approaches that consider the genetic and immunological heterogeneity of T1D patients to provide more tailored treatments are needed. Using mRNA encoding multiple epitopes tailored to the NOD mouse (model of T1D), we engineered bone marrow-derived dendritic cells (DCs) by mRNA electroporation (mRNA-DCs). The mRNA is designed to enable optimal presentation of epitopes to both CD4 and CD8 T cells. Antigen presentation persisted at least three days in vivo. Markers of anergy as well as IL-10 expression were found to be upregulated in antigen-specific CD4 and CD8 T cells after treating NOD mice with mRNA-DCs. T1D development was significantly suppressed with 3 biweekly treatments with 106 DCs containing 2 µg of antigen mRNA. Moreover, efficacy of the mRNA-DCs was enhanced when mRNAs encoding antigens and IL-27 were co-delivered into DCs by electroporation or when DCs were treated with rapamycin during differentiation. Endogenous expression of antigens may enable post-translational modifications and more prolonged presentation. Thus, we developed a promising and customizable ASIT platform to deliver multiple epitopes to exogenous tolerogenic DCs (or to endogenous APCs using a nanoparticle-based platform not covered here) that will consider the patient’s immune profile for a precision medicine treatment of T1D. Disclosure R. Fite: None. R. J. Creusot: None. Funding American Diabetes Association (1-19-PMF-022 to R.F.F.)

Cross-sectional analysis of peripheral blood mononuclear cells in lymphopenic and non-lymphopenic relapsing-remitting multiple sclerosis patients treated with dimethyl fumarate.

Background: Relapsing-remitting multiple sclerosis (RRMS) is an autoimmune disorder of the central nervous system. Dimethyl Fumarate is a disease-modifying medication used to treat RRMS patients that can induce lymphopenia. We aimed to immunophenotype peripheral blood mononuclear cells (PBMC) in RRMS patients cross-sectionally and examine the characteristics and modifications of lymphopenia over time. Methods: Characterization of PBMC was done by multiparametric flow cytometry. Patients had been on treatment for up to 4 years and were grouped into lymphopenic (DMF-L) and non-lymphopenic (DMF-N) patients. Results: Lymphopenia affected the cell population changes over time, with other patient characteristics (gender, age, and previous treatment status) also having significant effects. In both lymphopenic and non-lymphopenic patients, PBMC percentages were reduced over time. While overall T and B cells frequencies were not affected, males, older patients and untreated patients had significant changes in B cell subpopulations over time. CD4+ to CD8+T cell ratio increased significantly in lymphopenic patients over time. CD4−CD8−T cell population was similarly reduced in both lymphopenic and non-lymphopenic patients, over time. While the monocyte and NK overall populations were not changed, non-classical monocyte subpopulation decreased over time in lymphopenic patients. We also found CD56-CD16+ and CD56-CD16- NK cells frequencies changed over time in lymphopenic patients. Immune populations showed correlations with clinical outcomes measured by EDSS and relapse rate. Analysis of the overall immunophenotype showed that, while groups divided by other patient characteristics showed differences, the lymphopenia status overrode these differences, resulting in similar immunophenotype within DMF-L. Conclusions: Our data provide evidence that under the same therapy, lymphopenia affects how the immunophenotype changes over time and can override the differences associated with other patient characteristics and possibly mask other significant changes in the immune profile of patients.

Immune Profile in Patients With COVID-19: Lymphocytes Exhaustion Markers in Relationship to Clinical Outcome.

The velocity of the COVID-19 pandemic spread and the variable severity of the disease course has forced scientists to search for potential predictors of the disease outcome. We examined various immune parameters including the markers of immune cells exhaustion and activation in 21 patients with COVID-19 disease hospitalised in our hospital during the first wave of the COVID-19 pandemic in Slovakia. The results showed significant progressive lymphopenia and depletion of lymphocyte subsets (CD3+, CD4+, CD8+ and CD19+) in correlation to the disease severity. Clinical recovery was associated with significant increase in CD3+ and CD3+CD4+ T-cells. Most of our patients had eosinopenia on admission, although no significant differences were seen among groups with different disease severity. Non-survivors, when compared to survivors, had significantly increased expression of PD-1 on CD4+ and CD8+ cells, but no significant difference in Tim-3 expression was observed, what suggests possible reversibility of immune paralysis in the most severe group of patients. During recovery, the expression of Tim-3 on both CD3+CD4+ and CD3+CD8+ cells significantly decreased. Moreover, patients with fatal outcome had significantly higher proportion of CD38+CD8+ cells and lower proportion of CD38+HLA-DR+CD8+ cells on admission. Clinical recovery was associated with significant decrease of proportion of CD38+CD8+ cells. The highest AUC values within univariate and multivariate logistic regression were achieved for expression of CD38 on CD8+ cells and expression of PD1 on CD4+ cells alone or combined, what suggests, that these parameters could be used as potential biomarkers of poor outcome. The assessment of immune markers could help in predicting outcome and disease severity in COVID-19 patients. Our observations suggest, that apart from the degree of depletion of total lymphocytes and lymphocytes subsets, increased expression of CD38 on CD3+CD8+ cells alone or combined with increased expression of PD-1 on CD3+CD4+ cells, should be regarded as a risk factor of an unfavourable outcome in COVID-19 patients. Increased expression of PD-1 in the absence of an increased expression of Tim-3 on CD3+CD4+ and CD3+CD8+ cells suggests potential reversibility of ongoing immune paralysis in patients with the most severe course of COVID-19.

Changes in Leptin, CCL16 and sTNF-RII as a Distinctive Plasma Immune Profile in Patients with Fast Progressing ALS (1276)

Changes in Leptin, CCL16 and sTNF-RII as a Distinctive Plasma Immune Profile in Patients with Fast Progressing ALS (1276)

Gene coexpression network approach to develop an immune prognostic model for pancreatic adenocarcinoma

BackgroundPancreatic adenocarcinoma (PAAD) is a nonimmunogenic tumor, and very little is known about the relationship between the host immune response and patient survival. We aimed to develop an immune prognostic model (IPM) and analyze its relevance to the tumor immune profiles of patients with PAAD.MethodsWe investigated differentially expressed genes between tumor and normal tissues in the TCGA PAAD cohort. Immune-related genes were screened from highly variably expressed genes with weighted gene correlation network analysis (WGCNA) to construct an IPM. Then, the influence of IPM on the PAAD immune profile was comprehensively analyzed.ResultsA total of 4902 genes highly variably expressed among primary tumors were used to construct a weighted gene coexpression network. One hundred seventy-five hub genes in the immune-related module were used for machine learning. Then, we established an IPM with four core genes (FCGR2B, IL10RA, and HLA-DRA) to evaluate the prognosis. The risk score predicted by IPM was an independent prognostic factor and had a high predictive value for the prognosis of patients with PAAD. Moreover, we found that the patients in the low-risk group had higher cytolytic activity and lower innate anti-PD-1 resistance (IPRES) signatures than patients in the high-risk group.ConclusionsUnlike the traditional methods that use immune-related genes listed in public databases to screen prognostic genes, we constructed an IPM through WGCNA to predict the prognosis of PAAD patients. In addition, an IPM prediction of low risk indicated enhanced immune activity and a decreased anti-PD-1 therapeutic response.

ACPA Status Correlates with Differential Immune Profile in Patients with Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a progressive erosive autoimmune disease that affects 1% of the world population. Anti-citrullinated protein autoantibodies (ACPA) are routinely used for the diagnosis of RA, however 20–30% of patients are ACPA negative. ACPA status is a delineator of RA disease endotypes with similar clinical manifestation but potentially different pathophysiology. Profiling of key peripheral blood and synovial tissue immune populations including B cells, T follicular helper (Tfh) cells and CD4 T cell proinflammatory cytokine responses could elucidate the underlying immunological mechanisms involved and inform a treat to target approach for both ACPA-positive and ACPA-negative RA. Detailed high dimensionality flow cytometric analysis with supervised and unsupervised algorithm analysis revealed unique RA patient peripheral blood B cell and Tfh cell profiles. Synovial tissue single cell analysis of B cell subpopulation distribution was similar between ACPA− and ACPA+ RA patients, highlighting a key role for specific B cell subsets in both disease endotypes. Interestingly, synovial tissue single cell analysis of CD4 T cell proinflammatory cytokine production was markedly different between ACPA− and APCA+ RA patients. RNAseq analysis of RA patient synovial tissue highlighted disease endotype specific gene signatures. ACPA status associates with unique immune profile signatures that reinforce the need for a treat to target approach for both endotypes of RA.

Effect of Obstructive Sleep Apnea on Immunity in Cases of Chronic Rhinosinusitis With Nasal Polyps.

Objectives.Chronic rhinosinusitis with nasal polyps (CRSwNP) is a more severe inflammatory form of CRS that often coexists with obstructive sleep apnea (OSA). However, little is known about the relationship between OSA and the immune profile in patients with CRSwNP. We aimed to investigate the immune profile of patients with CRSwNP according to OSA severity.Methods.This study included 63 patients with CRSwNP and nine control subjects. Protein levels of inflammatory mediators were determined using multiplex immunoassays. All patients underwent standard polysomnography.Results.In patients with eosinophilic CRSwNP (ECRSwNP), interleukin (IL)-6 and chemokine [C-X-C motif] ligand (CXCL)-1 (type 1 immune-related markers) were upregulated in cases of moderate-to-severe OSA. Additionally, IL-4, IL-13, C-C motif chemokine (CCL)-11, CCL-24 (type 2 immune-related markers), and IL-17A (a type 3 immune-related marker) were present at elevated levels in patients with moderate-to-severe OSA. Although there were no significant differences in type 1, 2, or 3 immune-related markers among patients with non-eosinophilic CRSwNP (NECRSwNP) according to the severity of OSA, transforming growth factor-beta expression was higher in those with moderate-to-severe OSA. Furthermore, in ECRSwNP with moderate-to-severe OSA, associations were detected between serum markers and some upregulated inflammatory markers.Conclusion.OSA may increase the heterogeneity of the immune profile (types 1, 2, and 3) in patients with ECRSwNP, but not in those with NECRSwNP.

Peripheral Blood Immune Profiling of Convalescent Plasma Donors Reveals Alterations in Specific Immune Subpopulations Even at 2 Months Post SARS-CoV-2 Infection.

Immune profiling of patients with COVID-19 has shown that SARS-CoV-2 causes severe lymphocyte deficiencies (e.g., lymphopenia, decreased numbers, and exhaustion of T cells) and increased levels of pro-inflammatory monocytes. Peripheral blood (PB) samples from convalescent plasma (CP) donors, COVID-19 patients, and control subjects were analyzed by multiparametric flow cytometry, allowing the identification of a wide panel of immune cells, comprising lymphocytes (T, B, natural killer (NK) and NKT cells), monocytes, granulocytes, and their subsets. Compared to active COVID-19 patients, our results revealed that the immune profile of recovered donors was restored for most subpopulations. Nevertheless, even 2 months after recovery, CP donors still had reduced levels of CD4+ T and B cells, as well as granulocytes. CP donors with non-detectable levels of anti-SARS-CoV-2-specific antibodies in their serum were characterized by higher Th9 and Th17 cells, which were possibly expanded at the expense of Th2 humoral immunity. The most noticeable alterations were identified in previously hospitalized CP donors, who presented the lowest levels of CD8+ regulatory T cells, the highest levels of CD56+CD16− NKT cells, and a promotion of a Th17-type phenotype, which might be associated with a prolonged pro-inflammatory response. A longer follow-up of CP donors will eventually reveal the time needed for full recovery of their immune system competence.

Immune Cell Profiles in Patients Treated with Lenalidomide and Alternate Day Prednisolone Maintenance Post Upfront ASCT for Multiple Myeloma (LEOPARD Trial)

The LEOPARD trial evaluated lenalidomide and alternate day prednisolone (RAP) as post ASCT maintenance in newly diagnosed transplant eligible MM patients (TE NDMM). 60 patients were recruited. Estimated median potential follow-up was 44 months (IQR 26m - 52m). Median PFS from time of commencing RAP was 38.3m (95% CI, 25.8 to 54.8); median OS was not reached (71.4% of patients were alive at 36 months). Here we present the findings from correlative immune studies of this trial. Aims: To undertake mass cytometry (CyTOF) based immune profiling in patients with TE NDMM treated with RAP maintenance post ASCT. Methods: The LEOPARD trial was a phase II, multi centre, open label, single arm study of RAP maintenance after a single melphalan conditioned (200mg/m2) ASCT as part of up-front therapy. Patients were restaged at D+42 ASCT, and if eligible, were commenced on RAP maintenance (LEN 10mg daily increasing to 15mg daily after 8 weeks and alternate day prednisolone 50mg) within 8 weeks of D+0 of the ASCT. Therapy continued until toxicity/progression. CyTOF was performed in sequential samples in two selected groups of patients: long runners (LR, n=7), defined as those with PFS &amp;gt; 36 months (median) and early relapsers (ER, n=8), defined as those who progressed/died before reaching the lower quartile of PFS. [All patients had peripheral blood collected at baseline (pre-ASCT), 6w post-ASCT and weeks 4, 8, 12, 20, 28 and 40 of RAP]. Cells were barcoded using the Cell-ID 20-Plex Pd barcoding kit (Fluidigm) followed by staining with sub-set/function defining antibodies (targeting myeloid, B, T and NK cells: CD45, CD3, CD19, CD5, CD1c, CD226. CD8, CD11c, CD16, CD127, CD138, CD123, NKG2A, TIGIT, TIM3, CD45RA. CD274, CD27, CD197, CD28, Ki67, CD66b, CD183, KLRG1, CD43, NKG2D, CD38, CD278/ICOS, CD25, HLA-DR, CD4, CD57, GramB, PD-1, CD14, CD56, CD11b, Tbet, CD33). Samples were acquired on the Helios instrument. Supervised analysis was performed to determine differences in canonical immune cell populations. Unsupervised analysis was then performed: data were clustered in the VORTEX package. Significant differences in cluster frequency were assessed by Mann-Whitney test for statistical significance. Cluster phenotypes were determined and validated via multiple visualisation approaches. Results: Median age was 56yrs for LR versus 63yrs for ER. Median PFS for LR was 46.3m (38.4 - 51.5m) versus 10.2 m (2.1 - 21.3m) for ER. Supervised analysis was performed on all samples, dichotomized into baseline and last time point sampled for each patient. At baseline, Ki67+CD8+ T cells, ICOS+CD8+ T cells, HLA-DR+CD4+ T cells and CD11c+ myeloid cells were enriched in LR compared to ER. At the last timepoint sampled, Ki67+CD8+ T cells and ICOS+CD8+ T cells were again enriched in LR compared to ER. Conversely, B-reg-like cells (CD19+CD5+CD43-) were enriched in ER compared to LR at the last timepoint sampled. Unsupervised analysis was performed on all samples (all timepoints were pooled). Five clusters were significantly enriched in LR compared to ER. Four of these clusters represented activated/cytotoxic NK cells: CD56 dim, CD16-, NKG2A(CD159a)+, NKG2D(CD314)+, Granzyme B+ and CD38+, and additional expression of CD57 on one cluster; one cluster represented a mature myeloid population, with high expression of HLA-DR, CD11b and CD11c and low expression of CD33. One cluster was significantly enriched in ER compared to LR, representing activated neutrophils, with high expression of CD66b, CD11b and CD16. The clusters that were enriched were then assessed longitudinally over all time points. There was no difference in the kinetics of these populations between groups. Conclusions Significant differences in both T-cell and NK cell populations were demonstrable at baseline in LR versus ER patients. Subsequently, durable responses to post-ASCT lenalidomide maintenance were associated with a cytotoxic, controlled immune response whereas early relapse was characterised by a more uncontrolled inflammatory response and the emergence of B-reg-like cells prior to relapse. We conclude that immune profiling at baseline and after initiation of therapy may help to predict a more sustained response to lenalidomide maintenance enabling pre-emptive tailored treatment decisions. Disclosures Kalff: Roche: Honoraria; Janssen: Honoraria; Amgen: Honoraria; CSL: Honoraria; Celgene: Honoraria. Young:Bristol Meyers Squibb: Current Employment, Current equity holder in publicly-traded company. Pierceall:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company. Thakurta:Bristol Myers Squibb: Current Employment, Current equity holder in publicly-traded company; Oxford University: Other: visiting professor. Oppermann:Bristol Meyers Squibb: Research Funding. Guo:Bristol Meyers Squibb: Research Funding. Reynolds:Novartis AG: Current equity holder in publicly-traded company. Spencer:AbbVie, Celgene, Haemalogix, Janssen, Sanofi, SecuraBio, Specialised Therapeutics Australia, Servier and Takeda: Consultancy; AbbVie, Amgen, Celgene, Haemalogix, Janssen, Sanofi, SecuraBio, Specialised Therapeutics Australia, Servier and Takeda: Honoraria; Amgen, Celgene, Haemalogix, Janssen, Servier and Takeda: Research Funding; Celgene, Janssen and Takeda: Speakers Bureau.

Alteration of systemic and uterine endometrial immune populations in patients with endometriosis.

Endometriosis is defined as growth of endometrial tissue in ectopic locations; it is associated with infertility and chronic pain and affects ~12% of reproductive-aged women. Although inflammation is known to play a key role in endometriosis, knowledge related to immune phenotypes associated with this disease is lacking. This study aimed to characterize immune profiles in patients with endometriosis, to assess inflammatory mediators, to and determine if surgical and/or hormonal therapies restore immune homeostasis. Samples from nine controls and 20 histologically confirmed endometriosis patients were collected upon surgery and ~1-3weeks post-surgical intervention. Subjects were either not utilizing hormonal suppression or were currently on monophasic hormonal therapy. Tolerant regulatory T cells (Tregs=natural [nTregs] +inducible [iTregs]) and inflammatory T helper 17 (Th17) cells were identified in peripheral blood via flow cytometry and within the eutopic/ectopic endometrial tissues via immunohistochemistry and real-time-qPCR. Cytokines were assessed via 10-plex-ELISA. Patients with endometriosis not utilizing hormonal therapy exhibited lower iTregs (tolerant), greater Th17 (inflammatory), and a reduction in Treg/Th17 ratio (P<.05), indicative of systemic inflammation. Treg and Th17 localizations were enhanced within the ectopic endometrial implant, which promotes lesion development. Hormonal therapy decreased systemic and local inflammation (eutopic/ectopic endometrium) via decreased iTregs and Th17 cells in patients with endometriosis (P<.05). Thus, imbalance within immune populations correlated with increased inflammation in patients with endometriosis, which was mitigated by hormonal therapy. Patients with endometriosis exhibited systemic and localized inflammation within ectopic and endometrial tissues. Hormonal therapy dampened inflammation caused by disease.

The Metabolic Changes and Immune Profiles in Patients With COVID-19.

To explore the metabolic changes and immune profiles in patients with COVID-19, we analyzed the data of patients with mild and severe COVID-19 as well as young children with COVID-19. Of the leukocytes, 47% (IQR, 33–59) were lymphocytes [2.5 × 109/L (IQR, 2.2–3.3)], and monocytes were 0.51 × 109/L (IQR, 0.45–0.57) in young children with COVID-19. In 32 mild COVID-19 patients, circulating monocytes were 0.45 × 109/L (IQR, 0.36–0.64). Twenty-one severe patients had low PO2 [57 mmHg (IQR, 50–73)] and SO2 [90% (IQR, 86–93)] and high lactate dehydrogenase [580 U/L (IQR, 447–696)], cardiac troponin I [0.07 ng/mL (IQR, 0.02–0.30)], and pro-BNP [498 pg/mL (IQR, 241–1,726)]. Serum D-dimer and FDP were 9.89 mg/L (IQR, 3.62–22.85) and 32.7 mg/L (IQR, 12.8–81.9), and a large number of RBC (46/μL (IQR, 4–242) was presented in urine, a cue of disseminated intravascular coagulation (DIC) in severe patients. Three patients had comorbidity with diabetes, and 18 patients without diabetes also presented high blood glucose [7.4 mmol/L (IQR, 5.9–10.1)]. Fifteen of 21 (71%) severe cases had urine glucose +, and nine of 21 (43%) had urine ketone body +. The increased glucose was partially caused by reduced glucose consumption of cells. Severe cases had extraordinarily low serum uric acid [176 μmol/L (IQR, 131–256)]. In the late stage of COVID-19, severe cases had extremely low CD4+ T cells and CD8+ T cells, but unusually high neutrophils [6.5 × 109/L (IQR, 4.8–9.6)], procalcitonin [0.27 ng/mL (IQR, 0.14–1.94)], C-reactive protein [66 mg/L (IQR, 25–114)] and an extremely high level of interleukin-6. Four of 21 (19%) severe cases had co-infection with fungi, and two of 21 (9%) severe cases had bacterial infection. Our findings suggest that, severe cases had acute respiratory distress syndrome (ARDS) I–III, and metabolic disorders of glucose, lipid, uric acid, etc., even multiple organ dysfunction (MODS) and DIC. Increased neutrophils and severe inflammatory responses were involved in ARDS, MODS, and DIC. With the dramatical decrease of T-lymphocytes, severe cases were susceptible to co-infect with bacteria and fungi in the late stage of COVID-19. In young children, extremely high lymphocytes and monocytes might be associated with the low morbidity of COVID-19. The significantly increased monocytes might play an important role in the recovery of patients with mild COVID-19.

Abstract 3171: Biomarkers of responses to immunotherapy in advanced hepatocellular carcinoma

Abstract Biomarkers from the peripheral blood may serve as a useful tool to identify who will likely benefit from immunotherapy in patients with advanced hepatocellular carcinoma; hence, oncologists might be able to monitor treatment responses regularly via blood tests in addition to routine radiological assessment. To investigate immune profiles of patients with advanced HCC after exposure to immunotherapy by analysing circulating T cells from peripheral blood, our study included stage IV HCC patients (n = 14) seen at the Department of Medical Oncology, Queen Mary Hospitals. Nine patients received PD-1/PD-L1 inhibitor immunotherapy at the discretion of their clinicians, and five treatment-naïve patients were included as a control group. A control group of healthy subjects was also included in the study. Samples from patients who received PD-1/PD-L1 inhibitor therapy were collected after the treatment response was confirmed by radiological assessment. T cell profiles and exhaustion markers were compared along with treatment response as determined by radiological assessment. A total of 14 HCC patients were enrolled in the study: 4 patients showed a partial response to PD-1/PD-L1 inhibitor treatment, 5 patients progressed during PD-1/PD-L1 inhibitor treatment, and 5 patients were not treated with PD-1/PD-L1 inhibitors. Five buffy coat samples from healthy subjects were included as controls. Decreases in the percentage of regulatory T cells (0.8% of T cells in responders, p&amp;lt;0.0001) and in the ratio of effector memory T cells to regulatory T cells (EMT/Treg) were noted in patients who responded to PD-1/PD-L1 inhibitor therapy compared to those who did not, and thus may serve as biomarkers for immunotherapy responses in HCC. Similarly, we observed decreases in Tim-3 (2.09%, p&amp;lt;0.0001) and OX-40 (6.12%, p&amp;lt;0.0001) expression between responders and non-responders, and thus expression levels of these factors could potentially serve as response predictors. An increase in the proportion of CD4 CD8 double-positive (DP) T cells in peripheral blood was observed in immunotherapy responders (16.63%) versus non-responders (4.03%, p&amp;lt;0.05). Biomarkers in circulating T cells allow the investigation of immune profiles after treatment with immunotherapy using non-invasive approach from peripheral blood. Coupled with conventional surveillance strategies, we have identified biomarkers may serve as predictors for HCC patients who will likely benefit from immunotherapy. Citation Format: Vox Z. Ting, Valerie Chew, Carmen C. Wong, Thomas C. Yau. Biomarkers of responses to immunotherapy in advanced hepatocellular carcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3171.