Lymphocyte Survival Research Articles

Effects of Perioperative Inflammatory Response in Cervical Cancer: Laparoscopic versus Open Surgery.

There are few studies between postoperative neutrophil to lymphocyte ratio (NLR) and survival in cervical cancer. We compared postoperative changes in NLR according to surgical methods and analyzed the effect of these changes on 5-year mortality of cervical cancer patients. A total of 929 patients were assigned to either the laparoscopic radical hysterectomy (LRH) (n = 721) or open radical hysterectomy (ORH) (n = 208) group. Propensity score matching analysis compared the postoperative NLR changes between the two groups, and multivariate logistic regression analysis evaluated the association between NLR changes and 5-year mortality. Surgical outcomes between the two groups were also compared. In the LRH group, NLR changes at postoperative day (POD) 0 and POD 1 were significantly lower than in the ORH group after matching (NLR change at POD 0, 10.4 vs. 14.3, p < 0.001; NLR change at POD 1, 3.5 vs. 5.4, p < 0.001). In multivariate logistic regression analysis, postoperative NLR change was not associated with 5-year mortality (2nd quartile: OR 1.55, 95% CI 0.56–4.29, p = 0.401; 3rd quartile: OR 0.90, 95% CI 0.29–2.82, p = 0.869; 4th quartile: OR 1.40, 95% CI 0.48–3.61, p = 0.598), whereas preoperative NLR was associated with 5-year mortality (OR 1.23, 95% CI 1.06–1.43, p = 0.005). After matching, there were no significant differences in surgical outcomes between the two groups. There were significantly fewer postoperative changes of NLR in the LRH group. However, the extent of these NLR changes was not associated with 5-year mortality. By contrast, preoperative NLR was associated with 5-year mortality.

Brain-derived neurotrophic factor in autoimmune inflammatory diseases (Review).

Numerous recent studies reported that brain-derived neurotrophic factor (BDNF) also exists in the peripheral blood to regulate the proliferation, differentiation and survival of lymphocytes. Besides the role of BDNF in neuron repair, circulatory BDNF also enhances the proliferation and reduces apoptosis of lymphocytes. Peripheral lymphocytes express both BDNF and its receptors. Increasing evidence has indicated that altered BDNF serum levels significantly affect patients with autoimmune inflammatory diseases and may also be linked to the pathogenesis of diseases. For instance, systemic lupus erythematosus, an autoimmune inflammatory disease involving multiple organs, is frequently linked to altered B lymphocyte function, imbalance of T-cell subpopulations and loss of immune tolerance, which dysregulates the immune regulatory network with excessive secretion of inflammatory cytokines. The present review summarized studies that suggest a potential link between circulatory BDNF and autoimmune inflammatory diseases.

Next-generation immunotherapy for pancreatic ductal adenocarcinoma: navigating pathways of immune resistance.

To date, the use of immune checkpoint inhibitors has proven largely ineffective in patients with advanced pancreatic ductal adenocarcinoma. A combination of low tumor antigenicity, deficits in immune activation along with an exclusive and suppressive tumor microenvironment result in resistance to host defensives. However, a deepening understanding of these immune escape and suppressive mechanisms has led to the discovery of novel molecular targets and treatment strategies that may hold the key to a long-awaited therapeutic breakthrough. In this review, we describe the tumor-intrinsic and microenvironmental barriers to modern immunotherapy, examine novel immune-based and targeted modalities, summarize relevant pre-clinical findings and human experience, and, finally, discuss novel synergistic approaches to overcome immune-resistance in pancreatic cancer. Beyond checkpoint inhibition, immune agonists and anti-tumor vaccines represent promising strategies to stimulate host response via activation and expansion of anti-tumor immune effectors. Off-the-shelf natural killer cell therapies may offer an effective method for bypassing downregulated tumor antigen presentation. In parallel with this, sophisticated targeting of crosstalk between tumor and tumor-associated immune cells may lead to enhanced immune infiltration and survival of anti-tumor lymphocytes. A future multimodal treatment strategy involving immune priming/activation, tumor microenvironment reprogramming, and immune checkpoint blockade may help transform pancreatic cancer into an immunogenic tumor.

Peripheral Blood Lymphocyte Percentage May Predict Chemotolerance and Survival in Patients with Advanced Pancreatic Cancer. Association between Adaptive Immunity and Nutritional State.

Simple SummaryAlterations of the immune system that consist of induced inflammation and reduction in blood lymphocytes are a major factor contributing to cancer progression in patients with pancreatic carcinoma (PC). Identification of the percentage of lymphocytes in the Total number of White Blood Cells (L% TWBC) that could be associated with chemotolerance and survival time may be important for predicting treatment efficacy. The aim of this retrospective study was to highlight the best value of L% for predicting chemotolerance (n° of cycles tolerated) and survival beyond 12 months from diagnosis. The study found that L ≥ 29.7% TWBC compared to L < 29.7% predicted chemotolerance (p < 0.0001) and survival at every time point of follow-up: 6 (p = 0.04), 12 (p = 0.0003) and 18 months (p = 0.004) after diagnosis. Simple, rapid, routine laboratory data can be useful to predict treatment tolerance and efficacy in PC patients.Pancreatic Carcinoma (PC) cells have the ability to induce patient immunosuppression and to escape immunosurveillance. Low circulating lymphocytes are associated with an advanced stage of PC and reduced survival. Blood lymphocytes expressed as a percentage of Total White Blood Cells (L% TWBC) could predict chemotolerance (n° of tolerated cycles), survival time and Body Weight (BW) more effectively than lymphocytes expressed as an absolute value (LAB > 1500 n°/mm3) or lymphocytes >22%, which is the lowest limit of normal values in our laboratory. Forty-one patients with advanced PC, treated with chemotherapy, were selected for this observational retrospective study. Patients were evaluated at baseline (pre-chemotherapy), and at 6, 12 and 18 months, respectively, after diagnosis of PC. The study found L ≥ 29.7% to be a better predictor of survival (COX model, using age, sex, BW, serum creatinine, bilirubin and lymphocytes as covariates), chemotolerance (r = +0.50, p = 0.001) and BW (r = +0.35, p = 0.027) than LAB > 1500 or L > 22%. BW did not significantly correlate with chemotolerance or survival. The preliminary results of this study suggest that L ≥ 29.7% is more effective than LAB > 1500 or L > 22% at predicting chemotolerance, survival time and nutritional status. A possible impact of nutritional status on chemotherapy and survival seems to be lymphocyte-mediated given the association between BW and L%. This study may serve as the basis for future research to explore whether nutritional interventions can improve lymphopenia, and if so, how this may be possible.

Gene Expression Pattern of Peyer's Patch Lymphocytes Exposed to Kagocel Suggests Pattern-Recognition Receptors Mediate Its Action.

Kagocel is a synthetic carboxymethylcellulose derivative copolymerized with gossypol. Clinical data evidence its safety and efficiency for the treatment of flu and other viral infections via enhancement of interferon production. The gut-associated lymphoid tissue seems a likely site of kagocel action. The study was aimed to investigate the molecular mechanisms of its action using murine Peyer’s patches lymphocytes as a test system and the cytokines production and gene expression patterns as the primary outcomes. The Peyer’s patches lymphocytes isolated from BALB/c mice were stimulated with concanavalin A, or, to mimic viral infection, with a combination of concanavalin A and TLR3 ligand poly I:C. After 24 h of stimulation the cells were treated with saline, 30, 100, or 300 μg/ml of kagocel, or, as positive controls, 300 μg/ml oats b-D-glucan or 300 μg/ml lentinan. After 24 and 72 h of incubation with these drugs cytokines production was analyzed with ELISA and gene expression pattern was investigated using nCounter Inflammation panel chips followed by bioinformatics analysis. Expression of genes involved in the inflammatory response, antiviral defense, lymphocytes survival and proliferation (C1qa, C2, C3, Ccl21a, Il11, Il1b, Il23a, Il5, Ltb4r2, Alox15, Pla2g4a, Ptger1, Mapkapk5, Hras, Ifna1, Tlr2, Mrc1, Mx2) was upregulated in kagocel-treated Peyer’s patches lymphocytes. A list of plausible transcription factors (CEBPs, IRF, NFκB, RXR, Stat, Tead4, and ZSCAN) and master-regulators has been identified (cIAP, CIKS, dock9, MEKK1, FXR, IKK, IRAK, TRAF, dsRNA:TLR3:TRIF). The changes in gene expression pattern and the outcomes of bioinformatics analysis suggest that pattern recognition receptors, TLRs and dectin-1, are the key mediators of kagocel immunomodulatory action, with the possible involvement of interferon autocrine loop. The genes upregulated with kagocel include diverse components of the innate immune defense system.

Impact of effective dose to immune cells (EDIC) on lymphocyte nadir and survival in limited-stage SCLC

BackgroundEffective dose to immune cell (EDIC), an estimated radiation dose to the circulating lymphocytes, is of significance for overall survival (OS) in non-small cell lung cancer. This study aimed to validate the EDIC’s OS effect on limited-stage small cell lung cancer (LS-SCLC). Method and materialsThis study included LS-SCLC patients received definitive chemo-radiation in one single center from 2012 to 2017. All patients had multiple complete-blood-count tests including lymphocyte count at pre-, during- and end- radiotherapy. EDIC, computed according to doses of the lung, heart, and the total body, was assessed for its correlation with lymphocyte nadir, OS and progression free survival (PFS). ResultsOf 503 eligible patients, the mean EDIC was 7.34 Gy. The mean lymphocyte nadir was 0.48 × 109 cells/L, significantly lower than 1.65 × 109 cells/L at pre-radiotherapy (p < 0.001). EDIC was significantly correlated with lymphocyte nadir under both univariate (p < 0.001) and multivariable linear regression (p < 0.001). Multivariable analysis showed EDIC (HR = 0.1072, p = 0.005) and lymphocyte nadir (HR = 0.345, p = 0.003) were both significant for OS. EDIC was also significant for PFS (HR = 1.046, p = 0.026). The C-indexes of OS prediction were 0.593, 0.617, 0.676, and 0.684, for lymphocyte nadir alone, EDIC alone, combined lymphocyte nadir model, and combined EDIC model, respectively. ConclusionsThis study demonstrated that EDIC is an independent predictor for lymphocyte nadir, PFS and OS. EDIC may serve as a predictor for lymphocyte nadir and a surrogate marker for OS in LS-SCLC. More attention should be paid to EDIC to decease the lymphocyte toxicity and improve survival.

The Fractalkine Receptor CX3CR1 Links Lymphocyte Kinetics in CMV-Seropositive Patients and Acute Myocardial Infarction With Adverse Left Ventricular Remodeling.

AimsLatent cytomegalovirus (CMV) infection is associated with adverse cardiovascular outcomes. Virus-specific CX3CR1+ effector memory T-cells may be instrumental in this process due to their pro-inflammatory properties. We investigated the role of CX3CR1 (fractalkine receptor) in CMV-related lymphocyte kinetics and cardiac remodeling in patients with ST-elevation myocardial infarction (STEMI) undergoing primary percutaneous coronary intervention (pPCI).Methods and ResultsWe retrospectively analysed lymphocyte count, troponin, and survival in 4874 STEMI/pPCI patients, evaluated lymphocyte kinetics during reperfusion in a prospective cohort, and obtained sequential cardiac MRI (cMRI) to assess remodeling. Pre-reperfusion lymphopenia independently predicted mortality at 7.5 years. Prior to reperfusion, CCR7+ T-lymphocytes appeared to be depleted. After reperfusion, T-lymphocytes expressing CX3CR1 were depleted predominantly in CMV-seropositive patients. During ischaemia/reperfusion, a drop in CX3CR1+ T-lymphocytes was significantly linked with microvascular obstruction in CMV+ patients, suggesting increased fractalkine-receptor interaction. At 12 weeks, CMV+ patients displayed adverse LV remodeling.ConclusionWe show that lymphopenia occurs before and after reperfusion in STEMI by different mechanisms and predicts long-term outcome. In CMV+ patients, increased fractalkine induction and sequestration of CX3CR1+ T-cells may contribute to adverse remodeling, suggesting a pro-inflammatory pathomechanism which presents a novel therapeutic target.

Altered expression profile of BAFF receptors on peripheral blood B lymphocytes in Graves\u2019 disease

BackgroundB lymphocyte activating factor (BAFF) is a growth factor regulating B lymphocytes survival and maturation. Serum BAFF levels were elevated in patients affected with autoimmune thyroid diseases (AITD), including Graves’ disease (GD) and Hashimoto’s thyroiditis (HT). The aim of this study is to explore the association of expression levels of BAFF and its receptors with AITD.MethodsFifty-two GD patients, 39 Hashimoto’s thyroiditis (HT) patients and 23 healthy controls (HC) were recruited in this study. Serum BAFF levels were measured by ELISA. Expression of BAFF receptors, including BAFF receptor 3 (BR3) and transmembrane activator and calcium-modulating and cyclophilin ligand interactor (TACI), on B lymphocytes were analyzed by flowcytometry. Effects of steroids on serum BAFF levels and expression of BR3 and TACI were also observed in 10 patients with Graves’ orbitopathy (GO) receiving steroids therapy.ResultsSerum BAFF levels were significantly elevated from 0.93 ± 0.24 ng/ml in HC to 1.18 ± 0.33 ng/ml in GD (P = 0.0027) and 1.02 ± 0.24 ng/ml in HT (P = 0.0331). BR3 expression on peripheral B lymphocytes were elevated in GD (mean MFI: 4.52 ± 2.06 in GD vs. 3.00 ± 0.87 in HC, P = 0.0015), while TACI expression on peripheral B lymphocytes were decreased in GD without significance (mean MFI: 7.96 ± 4.06 in GD vs. 9.10 ± 3.37 in HC, P = 0.1285). Expression of BR3 and TACI was not changed significantly in HT patients. Steroids significantly suppressed serum BAFF concentrations (from 1.18 ± 0.27 ng/ml to 0.97 ± 0.10 ng/ml, P = 0.0364) and BR3 expression in GO patients (mean MFI from 6.26 ± 4.91 to 4.05 ± 1.58, P = 0.0083).ConclusionsAltered expression of BAFF and its receptor may mediate the autoimmunity in GD. Restoring the normal expression profile of receptors for BAFF could be a new strategy to treat GD.

SASH3 variants cause a novel form of X-linked combined immunodeficiency with immune dysregulation

AbstractSterile alpha motif (SAM) and Src homology-3 (SH3) domain-containing 3 (SASH3), also called SH3-containing lymphocyte protein (SLY1), is a putative adaptor protein that is postulated to play an important role in the organization of signaling complexes and propagation of signal transduction cascades in lymphocytes. The SASH3 gene is located on the X-chromosome. Here, we identified 3 novel SASH3 deleterious variants in 4 unrelated male patients with a history of combined immunodeficiency and immune dysregulation that manifested as recurrent sinopulmonary, cutaneous, and mucosal infections and refractory autoimmune cytopenias. Patients exhibited CD4+ T-cell lymphopenia, decreased T-cell proliferation, cell cycle progression, and increased T-cell apoptosis in response to mitogens. In vitro T-cell differentiation of CD34+ cells and molecular signatures of rearrangements at the T-cell receptor α (TRA) locus were indicative of impaired thymocyte survival. These patients also manifested neutropenia and B-cell and natural killer (NK)-cell lymphopenia. Lentivirus-mediated transfer of the SASH3 complementary DNA–corrected protein expression, in vitro proliferation, and signaling in SASH3-deficient Jurkat and patient-derived T cells. These findings define a new type of X-linked combined immunodeficiency in humans that recapitulates many of the abnormalities reported in mice with Sly1–/– and Sly1Δ/Δ mutations, highlighting an important role of SASH3 in human lymphocyte function and survival.

Radiation-Induced Lymphopenia Risks of Photon Versus Proton Therapy for Esophageal Cancer Patients.

PurposeTo assess possible differences in radiation-induced lymphocyte depletion for esophageal cancer patients being treated with the following 3 treatment modalities: intensity-modulated radiation therapy (IMRT), passive scattering proton therapy (PSPT), and intensity-modulated proton therapy (IMPT).Methods and MaterialsWe used 2 prediction models to estimate lymphocyte depletion based on dose distributions. Model I used a piecewise linear relationship between lymphocyte survival and voxel-by-voxel dose. Model II assumes that lymphocytes deplete exponentially as a function of total delivered dose. The models can be fitted using the weekly absolute lymphocyte counts measurements collected throughout treatment. We randomly selected 45 esophageal cancer patients treated with IMRT, PSPT, or IMPT at our institution (15 per modality) to demonstrate the fitness of the 2 models. A different group of 10 esophageal cancer patients who had received PSPT were included in this study of in silico simulations of multiple modalities. One IMRT and one IMPT plan were created, using our standards of practice for each modality, as competing plans to the existing PSPT plan for each patient. We fitted the models by PSPT plans used in treatment and predicted absolute lymphocyte counts for IMRT and IMPT plans.ResultsModel validation on each modality group of patients showed good agreement between measured and predicted absolute lymphocyte counts nadirs with mean squared errors from 0.003 to 0.023 among the modalities and models. In the simulation study of IMRT and IMPT on the 10 PSPT patients, the average predicted absolute lymphocyte count (ALC) nadirs were 0.27, 0.35, and 0.37 K/μL after IMRT, PSPT, and IMPT treatments using Model I, respectively, and 0.14, 0.22, and 0.33 K/μL using Model II.ConclusionsProton plans carried a lower predicted risk of lymphopenia after the treatment course than did photon plans. Moreover, IMPT plans outperformed PSPT in terms of predicted lymphocyte preservation.

Prognostic value of pretreatment circulating basophils in patients with glioblastoma.

Accumulating evidence demonstrated that atopic diseases were inversely related to glioma susceptibility and associated with improved prognosis of patients with glioma. This study aimed to elucidate the impacts of basophils, one of the important effector cells in the pathobiology of atopic disease, on prognosis of patients with glioblastoma (GBM). A total of 268 patients were newly diagnosed with GBM and treated with operation at our institution from January 2010 to December 2017. The association between pre-operation circulating eosinophil, basophil, neutrophil, lymphocyte, monocyte count and GBM progression free survival (PFS) was investigated. Moreover, based on the results of multivariate analysis, a prognostic nomogram was established and evaluated. Kaplan-Meier method showed that basophils ≥0.015 × 109/L (p = 0.015) and lymphocytes ≥1.555 × 109/L (p = 0.005) were correlated with better PFS. Cox regression model showed that basophils ≥0.015 × 109/L were an independent prognostic factor for PFS. Prognostic nomogram was established and the concordance index (C-index) for PFS prediction was 0.629. The calibration plots for the probability of 0.5-, 1- and 3-year PFS showed optimal consistency between the prediction by nomogram and actual observation. Increased pre-operation circulating basophils portend better PFS, which might be a useful and novel marker for the prognosis of GBM patients.

Bortezomib Sustains T Cell Function by Inducing miR-155-Mediated Downregulation of SOCS1 and SHIP1.

Suppressive mechanisms operating within T cells are linked to immune dysfunction in the tumor microenvironment. We have previously reported using adoptive T cell immunotherapy models that tumor–bearing mice treated with a regimen of proteasome inhibitor, bortezomib - a dipeptidyl boronate, show increased antitumor lymphocyte effector function and survival. Here, we identify a mechanism for the improved antitumor CD8+ T cell function following bortezomib treatment. Intravenous administration of bortezomib at a low dose (1 mg/kg body weight) in wild-type or tumor-bearing mice altered the expression of a number of miRNAs in CD8+ T cells. Specifically, the effect of bortezomib was prominent on miR-155 - a key cellular miRNA involved in T cell function. Importantly, bortezomib–induced upregulation of miR-155 was associated with the downregulation of its targets, the suppressor of cytokine signaling 1 (SOCS1) and inositol polyphosphate-5-phosphatase (SHIP1). Genetic and biochemical analysis confirmed a functional link between miR-155 and these targets. Moreover, activated CD8+ T cells treated with bortezomib exhibited a significant reduction in programmed cell death-1 (PD-1) expressing SHIP1+ phenotype. These data underscore a mechanism of action by which bortezomib induces miR-155–dependent downregulation of SOCS1 and SHIP1 negative regulatory proteins, leading to a suppressed PD-1–mediated T cell exhaustion. Collectively, data provide novel molecular insights into bortezomib–mediated lymphocyte–stimulatory effects that could overcome immunosuppressive actions of tumor on antitumor T cell functions. The findings support the approach that bortezomib combined with other immunotherapies would lead to improved therapeutic outcomes by overcoming T cell exhaustion in the tumor microenvironment.

B lymphocyte stimulating factor of Trachinotus ovatus (TroBLyS) promotes antibacterial immune defense

B lymphocyte stimulating factor (BLyS) belongs to the tumor necrosis family (TNF), which is a key cytokine in promoting the survival, proliferation, differentiation and apoptosis of B cells, and regulating the functions of T cells. However, functions of BLyS in teleosts are not well known. In the present study, the functions of BLyS in golden pompano (Trachinotus ovatus) (TroBLyS) were conducted through in vivo and in vitro experiments. Gene expression analysis suggested that TroBLyS was ubiquitously expressed in different tissues. Particularly, TroBLyS was most expressed in head kidney, followed by spleen, and liver. After Vibrio harveyi stimulation, the expressions of TroBLyS in head kidney, spleen, and liver were significantly upregulated. Recombinant TroBLyS (rTroBLyS) in vitro could promote lymphocytes proliferation and survival. Moreover, overexpression of TroBLyS in vivo in fish significantly promoted the activation of macrophages, increased the secretion of cytokines and chemokines, and reduced V. harveyi colonization in tissues. When the expression of TroBLyS was inhibited in T. ovatus, the bacterial count of V. harveyi was significantly increased than that in the control fish. Furthermore, the mortality of fish which was administrated of pTroBLyS for two weeks, was significantly reduced after V. harveyi infection, suggesting that TroBLyS had the potential as an immune enhancer. Our study indicated that TroBLyS might be involved in the antibacterial immune responses and has the potential as an immune system enhancer in T. ovatus.

Therapeutic efficacy of TACE 125I seed implantation and its combination with intra-tumor injection of cisplatin for the treatment of hepatocellular carcinoma

This study aimed to compare the therapeutic efficacy of transcatheter arterial chemoembolization (TACE) combined with either 125I seed implantation or 125I seed implantation and intra-tumor injection of cisplatin in treating hepatocellular carcinoma (HCC).A total of 100 patients with HCC were analyzed. The control group (n = 50) received TACE combined with 125I seed implantation therapy. The therapy group (n = 50) was treated with an intra-tumor injection of cisplatin along with TACE and 125I seed implantation therapy. After treatment, routine blood, liver and kidney function, tumor volume, T lymphocyte subset count (CD3, CD4, and CD8), implanted metastases, and survival were studied.The tumor volume decreased by 27.4% on average in the control group, and by 38.6% in the therapy group. Alpha fetoprotein (AFP) level decreased in all cases, and it was significantly lower in the therapy group than in the control group. Remote metastasis was observed in both groups (7 in the control group and 3 in the therapy group). No significant difference in routine blood, liver and kidney function, and T-lymphocyte subset counts were found between the two groups. Eight patients died of metastases in the control group and 2 in the therapy group at 1-year follow-up (P < 0.05).TACE combined with either 125I seed implantation or 125I seed implantation and intra-tumor injection of cisplatin was effective for the treatment of HCC. Of the 2 combination therapies, TACE combined with 125I seed implantation and intra-tumor injection of cisplatin was more effective for the treatment of HCC.

Organ-specific effects on glycolysis by the dioxin-activated aryl hydrocarbon receptor.

Activation of the aryl hydrocarbon receptor (AHR) by the environmental toxin dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD) causes diverse toxicities, including thymus atrophy and hepatosteatosis. The mechanisms by which AHR activation by TCDD leads to these toxicities are not fully understood. Here we studied the effects of TCDD on a major energy pathway, glycolysis, using the chick embryo close to hatching, a well-established model for studying dioxin toxicity. We showed that 24 hr of TCDD treatment causes changes in glycolysis in both thymus and liver. In thymus glands, TCDD decreased mRNAs for glycolytic genes and glucose transporters, glycolytic indices and levels of IL7 mRNA, phosphorylated AKT (pAKT) and HIF1A, stimulators of glycolysis and promoters of survival and proliferation of thymic lymphocytes. In contrast, in liver, TCDD increased mRNA levels for glycolytic genes and glucose transporters, glycolytic endpoints and pAKT levels. Similarly, increases by TCDD in mRNA levels for glycolytic genes and glucose transporters in human primary hepatocytes showed that effects in chick embryo liver pertain also to human cells. Treatment with the glycolytic inhibitor 2-deoxy-d-glucose exacerbated the effects on thymus atrophy by TCDD, supporting a role for decreased glycolysis in thymus atrophy by TCDD, but did not prevent hepatosteatosis. NAD+ precursors abolished TCDD effects on glycolytic endpoints in both thymus and liver. In summary, we report here that dioxin disrupts glycolysis mediated energy metabolism in both thymus and liver, and that it does so in opposite ways, decreasing it in the thymus and increasing it in the liver. Further, the findings support NAD+ boosting as a strategy against metabolic effects of environmental pollutants such as dioxins.

Abstract PR08: IL4 signaling increases acetyl-CoA metabolism and histone acetylation to promote breast cancer metastasis

Abstract Interleukin-4 (IL-4) has a number of well-established functions in immune cells including proliferation, and survival of lymphocytes, as well as polarization of macrophages to the pro-tumor M2 tumor associated macrophage phenotype. Many of the signaling effects of IL4 in immune cells involve regulation of metabolism such as increased glycolysis in B-lymphocytes and increased fatty acid oxidation in IL4 polarized macrophages. Despite understanding of the effects of IL4 signaling in immune cells not much is known about the consequences of IL4 signaling in tumor cells that express IL-4 Receptor (IL-4R). Previous work from our lab has shown that loss of IL-4R significantly abrogates metastasis in experimental models in mice. Thus, we wondered whether changes in metabolism downstream of IL-4 receptor activation can contribute to the metastatic phenotypes observed. Since altered metabolism has been shown to regulate the epigenetic state of cancer cells, we are examining histone modifications as a mechanism for the connections between metabolic regulation and increased metastatic capability in IL-4R expressing breast cancer cells. MDA-MB-231 and BT549 human triple negative breast cancer cell lines were treated with IL4 for various times. Samples of media from these cells were assayed for glucose and lactate. IL-4 treatment increased the consumption of glucose, and the release of lactate in IL-4R expressing breast cancer cell lines. This finding was confirmed using NBDG uptake assays. In addition, breast cancer cell lines were treated with IL-4 before undergoing biochemical fractionation to obtain nuclear lysates. These nuclear lysates were assessed for histone acetyltransferase (HAT) activity, and we observed that IL-4 treated lysates showed increased HAT activity. Western blotting was used to assess abundance of specific histone acetylation modifications and found that IL-4 increases acetylation of histone 3 lysine 9 as well as histone 3 lysine 27. In addition, we have also shown that IL-4 increases phosphorylation at serine 455 of ATP-Citrate-lyase (ACLY), an enzyme that plays a vital role in the production of nuclear and cytosolic pools of acetyl-CoA. This phosphorylation is a putative positive regulator of ACLY and its enzymatic activity. We’ve shown that treatment of IL-4R expressing cancer cells with inhibitors of HAT enzymes decreases proliferation, and clonogenic capabilities of those cells. However, IL-4 treatment is able to rescue the effects of HAT inhibition. Taken together these data suggest that the metastatic phenotype associated with IL-4 signaling in IL-4R expressing breast cancer is mediated at least partially by epigenetic modification, specifically histone acetylation. Citation Format: Demond Williams, Chilesi Uririr, Barbara Fingleton. IL4 signaling increases acetyl-CoA metabolism and histone acetylation to promote breast cancer metastasis [abstract]. In: Abstracts: AACR Special Virtual Conference on Epigenetics and Metabolism; October 15-16, 2020; 2020 Oct 15-16. Philadelphia (PA): AACR; Cancer Res 2020;80(23 Suppl):Abstract nr PR08.

Targeting TSLP-Induced Tyrosine Kinase Signaling Pathways in CRLF2-Rearranged Ph-like ALL.

Philadelphia (Ph)-like acute lymphoblastic leukemia (ALL) is characterized by aberrant activation of signaling pathways and high risk of relapse. Approximately 50% of Ph-like ALL cases overexpress cytokine receptor-like factor 2 (CRLF2) associated with gene rearrangement. Activated by its ligand thymic stromal lymphopoietin (TSLP), CRLF2 signaling is critical for the development, proliferation, and survival of normal lymphocytes. To examine activation of tyrosine kinases regulated by TSLP/CRLF2, phosphotyrosine (P-Tyr) profiling coupled with stable isotope labeling of amino acids in cell culture (SILAC) was conducted using two CRLF2-rearranged (CRLF2r) Ph-like ALL cell lines stimulated with TSLP. As a result, increased P-Tyr was detected in previously reported TSLP-activated tyrosine kinases and substrates, including JAK1, JAK2, STAT5, and ERK1/2. Interestingly, TSLP also increased P-Tyr of insulin growth factor 1 receptor (IGF1R) and fibroblast growth factor receptor 1 (FGFR1), both of which can be targeted with small-molecule inhibitors. Fixed-ratio combination cytotoxicity assays using the tyrosine kinase inhibitors BMS-754807 and ponatinib that target IGF1R and FGFR1, respectively, revealed strong synergy against both cell line and patient-derived xenograft (PDX) models of CRLF2r Ph-like ALL. Further analyses also indicated off-target effects of ponatinib in the synergy, and novel association of the Ras-associated protein-1 (Rap1) signaling pathway with TSLP signaling in CRLF2r Ph-like ALL. When tested in vivo, the BMS-754807/ponatinib combination exerted minimal efficacy against 2 Ph-like ALL PDXs, associated with low achievable plasma drug concentrations. Although this study identified potential new targets in CRLF2r Ph-like ALL, it also highlights that in vivo validation of synergistic drug interactions is essential. IMPLICATION: Quantitative phosphotyrosine profiling identified potential therapeutic targets for high-risk CRLF2-rearranged Ph-like ALL.

The relationship between 18F-FDG-PETCT-derived tumour metabolic activity, nutritional risk, body composition, systemic inflammation and survival in patients with lung cancer

The aim of this study was to examine the relationship between PET-CT derived tumour glucose uptake as measured by maximum standard glucose uptake (SUVmax) and total lesion glycolysis (TLG), nutritional risk as measured by the malnutrition universal screening tool (MUST), CT derived body composition as measured by skeletal muscle index (SMI) and skeletal muscle radiodensity (SMD), the systemic inflammatory response as measured by the modified Glasgow prognostic score (mGPS) and the neutrophil to lymphocyte ratio (NLR) and survival in patients with lung cancer, treated with radiotherapy. In a retrospective cohort study, 119 patients were included in final analyses. The majority of patients were over 65 (86%), female (52%), had a performance status (ECOG-PS) of 0 or 1 (57%), were at nutritional risk (57%), were overweight (53%), had visceral obesity (62%), had a normal SMI (51%), had a low SMD (62%) and were systemically inflammed (mGPS 1/2, 51%). An elevated TLG was associated with sex (p < 0.05), TNM stage (p < 0.001), MUST (p < 0.01) and mGPS (p < 0.01). An elevated mGPS was associated with age (p < 0.05), NLR (p < 0.01), MUST (p < 0.01), and TLG (p < 0.01). On univariate survival analysis, TNM stage (p < 0.01), mGPS (p < 0.05), NLR (p < 0.01), MUST (p ≤ 0.001), Low SMD (p < 0.05), SUVmax (p ≤ 0.001) and TLG (p < 0.001) were associated with overall survival. On multivariate survival analysis MUST (HR: 1.49 95%CI 1.12–01.98 p < 0.01) and TLG (HR: 2.02 95%CI 1.34–3.04 p = 0.001) remained independently associated with survival. In conclusion, elevated tumour metabolic activity was associated with more advanced stage, greater nutritional risk, the systemic inflammatory response and poorer survival but not body composition analysis in patients with lung cancer. These results suggest that detrimental body composition is not directly determined by tumour metabolic activity but rather an ongoing systemic inflammatory response.

137 Randel Lecture: Genetic control of thermotolerance in cattle at the whole-animal and cellular level

Abstract Research by Randel and others has revealed the extensive genetic diversity in thermotolerance existing in cattle. Genetic differences in resistance to heat stress involve genes that affect body temperature regulation and that stabilize cellular function during hyperthermia. There are large differences between breeds in ability to regulate body temperature during heat stress. There is also genetic variation in thermoregulatory ability within breeds that arose in cold climates – the estimate of heritability of rectal temperature during heat stress in Holsteins is 0.17. Efforts are underway to identify genes conferring increased thermoregulatory ability in thermotolerant breeds and transfer these to thermosensitive breeds. One such gene is the prolactin receptor gene (PRLR). Several mutations in PRLR exist in criollo-type B. taurus that result in formation of a truncated receptor protein. Cattle with this mutation, which is dominant, have a short sleek hair coat and are referred to as “slick”. The slick mutation has been introduced into Holsteins by crossbreeding with Senepol cattle and in Red Angus by gene editing. Slick Holsteins are less affected by heat stress than wild-type Holsteins with respect to rectal temperature, milk yield and reproduction. Genetic variation also exists in cellular responses to elevated temperature (i.e., heat shock). Exposure of preimplantation embryos to heat shock, for example, causes a greater reduction in subsequent development of Angus, Holstein, Angus x Holstein or Jersey embryos than embryos from Brahman or Nelore (B. indicus) or Romosinuano (criollo). One gene with alleles affecting cellular resistance is the heat shock protein 70 gene HSPA1A. A mutation in the promoter region of HSPA1Ais associated with increased transcription of the encoded protein and enhanced survival of lymphocytes exposed to heat shock. Identification of other genes conferring thermotolerance will result in the increased ability to produce thermotolerant lines of cattle via genetic selection or gene transfer.

Deep sequencing reveals a DAP1 regulatory haplotype that potentiates autoimmunity in systemic lupus erythematosus

BackgroundSystemic lupus erythematosus (SLE) is a clinically heterogeneous autoimmune disease characterized by the development of anti-nuclear antibodies. Susceptibility to SLE is multifactorial, with a combination of genetic and environmental risk factors contributing to disease development. Like other polygenic diseases, a significant proportion of estimated SLE heritability is not accounted for by common disease alleles analyzed by SNP array-based GWASs. Death-associated protein 1 (DAP1) was implicated as a candidate gene in a previous familial linkage study of SLE and rheumatoid arthritis, but the association has not been explored further.ResultsWe perform deep sequencing across the DAP1 genomic segment in 2032 SLE patients, and healthy controls, and discover a low-frequency functional haplotype strongly associated with SLE risk in multiple ethnicities. We find multiple cis-eQTLs embedded in a risk haplotype that progressively downregulates DAP1 transcription in immune cells. Decreased DAP1 transcription results in reduced DAP1 protein in peripheral blood mononuclear cells, monocytes, and lymphoblastoid cell lines, leading to enhanced autophagic flux in immune cells expressing the DAP1 risk haplotype. Patients with DAP1 risk allele exhibit significantly higher autoantibody titers and altered expression of the immune system, autophagy, and apoptosis pathway transcripts, indicating that the DAP1 risk allele mediates enhanced autophagy, leading to the survival of autoreactive lymphocytes and increased autoantibody.ConclusionsWe demonstrate how targeted sequencing captures low-frequency functional risk alleles that are missed by SNP array-based studies. SLE patients with the DAP1 genotype have distinct autoantibody and transcription profiles, supporting the dissection of SLE heterogeneity by genetic analysis.