Proliferation Of Peripheral Blood Research Articles

Phase 1/2 study of monalizumab plus durvalumab in patients with advanced solid tumors

BackgroundThe combination of monalizumab (anti-NKG2A/CD94) and durvalumab (anti-programmed death ligand-1) may promote antitumor immunity by targeting innate and adaptive immunity. This phase 1/2 study of monalizumab and durvalumab evaluated safety,...

Nivolumab Reduces PD1 Expression and Alters Density and Proliferation of Tumor Infiltrating Immune Cells in a Tissue Slice Culture Model of Renal Cell Carcinoma.

Simple SummaryImmune checkpoint inhibitors (ICIs) have become a first-choice therapy option in the treatment of clear cell renal cell carcinoma (ccRCC). A predictive biomarker is urgently needed since not all patients respond and adverse events occur. Therefore, an ex vivo tissue slice culture (TSC) model was tested to investigate the effects of nivolumab on tumor infiltrating immune cells (TIIC). A decrease in programmed death receptor 1 expression, as well as effects on density and proliferation of TIIC, were observed. Thus, the TSC model could serve as a test platform for response prediction to ICIs.Background: In the treatment of clear cell renal cell carcinoma (ccRCC), nivolumab is an established component of the first-line therapy with a favorable impact on progression free survival and overall survival. However, treatment-related adverse effects occur and, to date, there is no approved predictive biomarker for patient stratification. Thus, the aim of this study was to establish an ex vivo tissue slice culture model of ccRCC and to elucidate the impact of nivolumab on tumor infiltrating immune cells. Methods: Fresh tumor tissue of ccRCC was treated with the immune checkpoint inhibitor nivolumab using ex vivo tissue slice culture (TSC). After cultivation, tissue slices were formalin-fixed, immunohistochemically stained and analyzed via digital image analysis. Results: The TSC model was shown to be suitable for ex vivo pharmacological experiments on intratumoral immune cells in ccRCC. PD1 expression on tumor infiltrating immune cells was dose-dependently reduced after nivolumab treatment (p < 0.01), whereas density and proliferation of tumor infiltrating T-cells and cytotoxic T-cells were inter-individually altered with a remarkable variability. Tumor cell proliferation was not affected by nivolumab. Conclusions: This study could demonstrate nivolumab-dependent effects on PD1 expression and tumor infiltrating T-cells in TSC of ccRCC. This is in line with results from other scientific studies about changes in immune cell proliferation in peripheral blood in response to nivolumab. Thus, TSC of ccRCC could be a further step to personalized medicine in terms of testing the response of individual patients to nivolumab.

Anti-tumor activity and immunogenicity of a succinoglycan riclin

Natural polysaccharides have attracted considerable interests due to diverse biological activities. Succinoglycan is an extracellular polysaccharide produced by most Agrobacterium strains. Here, we confirmed riclin was a typical succinoglycan by NMR and methylation analysis, and investigated the antitumor effects of riclin in sarcoma 180 tumor-bearing mice. The results showed that riclin inhibited the tumor growth significantly as well as cyclophosphamide (CTX). While CTX caused serious damage to spleen structure, riclin increased the spleen index and promoted lymphocytes proliferation in peripheral blood, spleen and lymph nodes. Riclin decreased splenocytes apoptosis as evidenced by alterations of B-cell lymphoma-2 family proteins and Cleaved Caspase-3 protein. Moreover, 1H nuclear magnetic resonance (NMR)-based metabolomics analysis revealed that riclin partially altered the metabolic profiles of splenocytes. In conclusion, riclin is a succinoglycan that performed strong immunogenicity and suppressed sarcoma growth in mice. Succinoglycan riclin could be a potential antitumor agent for functional food and pharmaceutical purpose.

Preliminary safety and efficacy of GX-I7, a long-acting interleukin-7, in combination with pembrolizumab in patients with refractory or recurrent metastatic triple negative breast cancer (mTNBC): Dose escalation period of Phase Ib/II study (KEYNOTE-899).

1072 Background: Pembrolizumab monotherapy did not significantly improve OS as 2nd and 3rd-lines treatment for mTNBC compared to standard chemotherapy in phase III study (KEYNOTE-119) leading to high unmet needs of effective treatment. Recent studies showed that higher lymphocyte count is an independent factor which correlates with better response to checkpoint blockade in cancer patients. GX-I7, a long-acting interleukin-7, could potentially provide synergistic anti-tumor efficacy with pembrolizumab by increasing number of T cells both in tumor microenvironment (TME) and peripheral blood (PB). Methods: This is an open-label, phase Ib/II study in patients with refractory or recurrent TNBC who failed from standard chemotherapy in the metastatic setting, with ≤3rd-lines of previous chemotherapy. The dose escalation phase adopted the 3+3 design. The GX-I7 doses were administered IM q9w or q12w, with or without cyclophosphamide pre-conditioning depending on the allocation, and in combination with pembrolizumab 200 mg IV q3w. The objectives were dose limiting toxicities (DLTs), safety, pharmacodynamic markers including lymphocyte increase and RP2D. Results: As of January 30, 2020, GX-I7 and pembrolizumab were exposed to 24 patients (median age 46.0 years [29-75], ECOG PS 1 [58.3%], median cycle no. 3 [1-9]). Treatment was discontinued in 13 (54.2%), majority due to PD and 11 patients are ongoing. No DLTs were reported in all dose groups. Treatment related AEs occurred in 91.7% of patients with grade 1-2 and 9.1% with grade 3 (no grade 4). Common AEs were injection site reaction (39.0%) and fever (13.0%), which were easily managed. Grade 3 toxicity were AST/ALT elevation and infusion related reaction, reported from 1 patient each (4.2%). GX-I7 induced dose-dependent lymphocyte proliferation in PB, with approximately 4-folds increase in high doses. 17 patients were evaluable; confirmed objective responses from ongoing patients included one partial response (5.9%), 2 stable disease (11.8%) and 1 durable unconfirmed PD (5.9%). Conclusions: GX-I7 in combination with pembrolizumab was well tolerated, with no DLTs reported. There was no apparent increase of immune-related AEs with the addition of GX-I7. Pharmacodynamics data support proof of mechanism and it warrants further clinical studies. Clinical trial information: NCT03752723 .

Anti-CD19/CD22 Dual CAR-T Therapy for Refractory and Relapsed B-Cell Acute Lymphoblastic Leukemia

Introduction Chimeric antigen receptor (CAR) T cell therapy targeting CD19 has demonstrated high success for B-cell acute lymphoblastic leukemia (B-ALL). Despite the initial high complete remission (CR) rate, about half of patients (pts) relapse at 1 year. CD19 antigen loss was observed in a significant number of relapsed patients. CD22 is another leukemic marker that often expressed on the surface of CD19- relapsed B-ALL blasts. We have developed a bispecific CAR construct targeting CD19 and CD22. Here we report results from a phase Ⅰ clinical trial of CD19/CD22 (GC022) dual CAR-T to evaluate the safety and feasibility of treating patients with relapsed/refractory B-ALL. Methods The CD19/CD22 dual CAR-T cells were manufactured in a cGMP facility. Patients' peripheral blood (PB) mononuclear cells were first collected, and CD3+ T cells were separated. The cells were then transfected by lentivirus encoded with CD19 CD22 bispecific scFv sequences. The CAR-T cells produced in this way contained a 4-1BB co-stimulatory signal domain. The CAR-T cells were then cultured for 8-14 days until sufficient cells were harvested for infusion. All pts received conditioning regimen of fludarabine and cyclophosphamide intravenously for 3 consecutive days with doses of 30 mg/m2/day and 250 mg/m2/day, respectively before a single infusion of CAR-T cells. The level of infused CAR-T cell proliferation in PB was analyzed by qPCR and flow cytometry. The primary end points were to evaluate feasibility and toxicity, and the secondary end points included disease response and engraftment/persistence of infused CD19/CD22 dual CAR-T cells. Results From Feb. 2019 to 23 July. 2019, 17 patients (pts) with relapsed/refractory B-ALL including 4 pts who previously treated with CD19 CAR-T cells were enrolled and pts were treated with CD19/CD22 dual CAR-T GC022 (US NIH Clinical#: NCT03825731). Four were adults, 13 pediatrics (age 1-45, Table 1). The median bone marrow (BM) blasts was 19.09 (0.36-87.82) %. Four patients received a low-dose (2.5-5×105/kg) dual CAR-T, 7 received a medium-dose (1-2.5×106/kg) and 5, a high-dose (3-5×106/kg). One patient withdrew immediately before CAR-T infusion due to his personal issue. Anti-leukemic efficacy was evaluated in 11/16 pts (5 pts have not yet reached D15). The 3/4 pts received low dose of GC022 had no response to treatment and 1 had MRD-positive CR. Seven patients who received medium dose achieved 100% CR on D15, highlighting the dose-dependent anti-leukemic activity. Six out of seven pts had MRD negative CR in this medium dose group. Five pts in high dose group have not reached the time for evaluation. No one relapsed with a median observation time of 60 (7-139) days. Cellular kinetic data was analyzed. Median peak of CAR-T copies was 1.09 (0.0022-4.98) x105 copy number/µg PB genomic DNA (Fig.1). The proliferation of medium or high dose groups was significantly better than the low dose group 3.47(0.43-4.98) x105 vs. 0.023(0.0022-0.81) x105(P=0.02) and 2.02(1.89-2.16) x105 vs. 0.023(0.0022-0.81) x105(P=0.004), (Fig.2). The peaks of IL-6, IFN-γ, IL-10, and CD25 were observed around day 7-10. Sixteen out of seventeen pts had grade 0-1 cytokine release syndrome (CRS) and only 1 patient experienced grade 2 CRS. None developed neurotoxicity. Conclusion Our study demonstrates safety and technical feasibility of CD19 and CD22 dual CAR-T in treating patients with CD19+CD22+ relapsed/refractory B-ALL. A low toxicity with dose-dependent high CR rate including pts who previously treated with CD19 CAR-T cells were observed. Longer observation time and more patients are needed to evaluate a beneficial advantage of the CD19/CD22 dual CAR-T over CD19 CAR-T product. Disclosures No relevant conflicts of interest to declare.

Abstract CT082: Phase (Ph) I/Ib study of NIZ985 with and without spartalizumab (PDR001) in patients (pts) with metastatic/unresectable solid tumors

Abstract Background: NIZ985 is a soluble IL-15/IL-15 receptor α heterodimer (hetIL-15) that binds IL-2/IL-15 receptor β/γ and promotes effector T-cell and NK cell expansion and activation. Spartalizumab is a humanized IgG4 mAb that blocks binding of programmed death 1 (PD-1) to programmed death ligand 1/2 (PD-L1/2). In preclinical studies, IL-15 and anti-PD-1/PD-L1 combinations demonstrated synergistic antitumor activity. Here we report dose escalation results from a Ph I/Ib study of NIZ985 as a single agent (SA) and in combination with spartalizumab (NCT02452268). Methods: Pts with metastatic/unresectable solid tumors received subcutaneous NIZ985 TIW (2 weeks on/2 weeks off); 0.25-4 μg/kg NIZ985 SA or 1 μg/kg NIZ985 with IV spartalizumab (400 mg, Q4W). Ph I endpoints: DLTs, safety, PK, pharmacodynamics (PD), preliminary efficacy, and recommended dose for expansion (RDE), statistically guided by a 3+3 study design. Results: As of Oct 9, 2018, 14 pts (median age 56.5 y [range 42.0-73.0]) received NIZ985 SA and 11 pts (median age 61 y [range 34.0-75.0]) received NIZ985 + spartalizumab. Treatment was discontinued in 13 (93%) and 10 (91%) pts receiving NIZ985 SA and NIZ985 + spartalizumab, respectively, mainly due to disease progression (57% and 82%); 1 pt in each group is ongoing. No DLTs were reported in the first cycle. AEs suspected to be drug related were reported in 100% of pts (36% grade 3/4) for NIZ985 SA and 100% of pts treated with the combination (27% grade 3/4). Common AEs [all grades; grade 3/4], defined as ≥50% of pts for any grade, were injection site reaction [100%; 7%], chills [64%; 0%], and fatigue [57%; 0%] in NIZ985 SA; and injection site reaction [100%; 9%], fatigue [82%; 0%], diarrhea [64%; 9%], and nausea [55%; 0%] in NIZ985 + spartalizumab. Median duration of NIZ985 exposure (range) across doses was 7.6 weeks (2.6-15.6) in NIZ985 SA and 21.6 weeks (5.6-53.6) in NIZ985 + spartalizumab. In 13 evaluable NIZ985 SA pts, confirmed best overall responses (BORs) were stable disease (SD; n=3, 23%), progressive disease (n=8, 62%), and unknown (n=2, 15%); SD is ongoing in 1 pt with spindle cell ocular melanoma. Confirmed BORs in 11 evaluable NIZ985 + spartalizumab pts were SD (n=5, 45%) and progressive disease (n=6, 55%); SD is ongoing in 1 pt with gastric adenocarcinoma. RDE was declared as 1 μg/kg NIZ985 (TIW; 2 weeks on/2 weeks off) for SA and with spartalizumab (400 mg, Q4W). PD analyses showed NIZ985 induces lymphocyte proliferation in peripheral blood. Preliminary antitumor activity, PK, and PD data from tumors and peripheral blood will be presented. Conclusions: NIZ985 SA and with spartalizumab was well tolerated in pts with metastatic/unresectable solid tumors, with no DLTs reported. RDEs were NIZ985 1 μg/kg (TIW, 2 weeks on/2 weeks off) for SA and with spartalizumab (400 mg, Q4W). PD data support proof of mechanism. Dose escalation with an alternate NIZ985 schedule is ongoing. Citation Format: Kevin Conlon, Rom Leidner, Douglas McNeel, Sumati Gupta, Andrea Wang-Gillam, Thomas A. Waldmann, George Pavlakis, Mirek Dostalek, Sema Kurtulus, Niladri Roy Chowdhury, Jessica O'Keeffe, Nehal S. Parikh, John A. Thompson. Phase (Ph) I/Ib study of NIZ985 with and without spartalizumab (PDR001) in patients (pts) with metastatic/unresectable solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT082.

Apoptosis and proliferation ofperipheral blood T-cells as alternative processes in pathogenesis of diabetes mellitus type 1

Apoptosis, along with proliferation, is a form of lymphocyte response to activating stimuli. In the early stages of cell differentiation, the apoptotic response prevails and it results to the formation of tolerance to inductor antigen. Mature lymphocytes proliferate in response to stimulation and it means the initial stage in the development of the immune response. Since in this case apoptosis and proliferation acts as alternative processes, their ratio can serve as a measure of the effectiveness of the cellular response to activating signals. The resistance of autoreactive T-cells to apoptosis is the main key point in the development of type 1 diabetes mellitus (T1DM). Autoreactive T-cells migrates from the bloodstream to the islet tissue of the pancreas and take an active part in b cells destruction. The resistance of autoreactive effector T-cells to apoptosis may suggest their high proliferative potential. Therefore, the comparative evaluation of apoptosis and proliferation of peripheral blood lymphocytes can give a more complete picture of their functional state and thus will help to reveal the causes of ineffective peripheral blood T-ceiis apoptosis in patients with T1DM and will help to understand more deeply the pathogenesis of the disease. in this article, the features of proliferative response of peripheral blood T-cells in patients with T1DM and in individuals with high risk of developing T1DM have been studied. Apoptosis of T-cell subpopulations has been investigated. The correlation between the apoptotic markers and the intensity of spontaneous and activation- induced in vitro T-cells proliferation of was revealed. it was determined, that autoreactive peripheral blood T-cells were resistant to apoptosis and demonstrated the increased proliferative potential in patients with T1DM and in individuals with high risk of developing T1DM.

Chronic Myeloid Leukemia with Cryptic Philadelphia Translocation Presenting Initially As Extramedullary Lymphoid Blast Phase

Abstract We report an adult male diagnosed as chronic myeloid leukemia (CML) - chronic phase when he was shortly presented with treatment-naive extramedullary lymphoid blast crises involving multiple lymph nodes, with no features of acceleration or blast crises in the peripheral blood (PB) and bone marrow (BM). In addition the patient has Philadelphia negative (Ph -ve) BCR/ABL-1 - positive CML. Up to our knowledge; this is the first report of CML with extramedullary lymphoid blast phase (as initial presentation) that shows cryptic Ph translocation. This is a 45 year- old male who presented with one month history of swelling of medial canthus of his left eye, and left sided neck swelling for three days. His initial physical examination showed bilateral cervical lymphadenopathy and splenomegaly. CBC showed marked leukocytosis (WBC: 159x10^3/ul (4.0-10.0), with Hb of 12.2gm/dl (13.0-17.0) and thrombocytopenia (123x10^3/ul (150-400)). BM biopsy was markedly hypercellular with granulocytic hyperplasia and prominent eosinophils. Reticulin stain showed focal increase reticulin fibrosis (grade 1-2 out of 3). FISH analysis revealed BCR/ABL1 rearrangement, t(9;22) in 97% of the cells analyzed with deletion of ABL1 as confirmed by metaphase FISH analysis: der(9)t(9;22),del(9)(q34q34)(ABL1+,BCR+),der(22)t(9;22)(BCR+,ABL1-)[10]. (fig 1 B). The fusion signal was formed on chromosome 9 instead of 22. Conventional cytogenetics confirmed the presence of cryptic BCR/ABL1 rearrangement. and normal karyotype: 46,XY. (fig1 c). BCR/ABL1 transcript confirmed by PCR.The overall findings consistent with CML- chronic phase. Immunohistochemical studies showed positivity for CD45, CD20, CD79a and BCL2 with strong and diffuse positive staining for Tdt. Ki67 shows high expression of about 90% of cellular nuclei. T-cell markers including CD3, CD5, CD4 and CD8 are negative in tumor cells. The morphological and immunohistochemical features are consistent with nodal involvement by precursor B-cell lymphoblastic lymphoma. FISH analysis performed on nodal tissue and confirmed the presence of BCR/ABL-1 rearrangement confirming that nodal involvement is lymphoid blast phase (BP) of CML and excluding the possibility of denovo ALL as second pathology. The patient started on hyper CVAD chemotherapy and Dasatinib .Still under treatment . In the great majority of CML; BCR/ABL is cytogenetically visualized as t(9;22); giving rise to the Ph chromosome, harboring the chimeric gene. Cryptic or masked translocation occurs in 2-10% patients with no cytogenetic evidence for the BCR/ABL rearrangement but are positive by FISH and/or (RT-PCR. Cryptic BCR/ABL rearrangements can be found in cases with a normal karyotype and in cases with complex karyotype in which the t(9;22) is not detected by conventional cytogenetics. These latter patients are described as CML Ph-ve/ BCR/ABL1 +ve with the chimeric gene present on the derivative chromosome 22, as in most CML cases, or alternatively on the rivative 9 as in our case. The prognostic significance of cryptic Ph translocations has been discussed in previous reports, some authors reported a worse prognosis associated with location of BCR/ABL on the der (9). Data regarding the outcome of Ph-neg CML patients treated with TKIs are limited, only 10 Ph-neg CML cases treated with imatinib have been described in 6 previous reports . The prevalence of extramedullary blast phase (BP) has been reported to be 7-17% in patients with BP . Extramedullary BP in the newly diagnosed patients is extremely rare as the prevalence of an accelerated phase and BP as initial presentations has been reported to be only 5-10% in CML patients . Conclusion: This is a report of a case of (CML) with a combination of a couple of unusual events including cryptic Philadelphia (Ph) translocation together with extramedullary lymphoid blast crisis as an initial presentation We aim in this report to highlight this case with unusual features, to describe clinical features and disease out come and to focus that the absence of hematologic blast proliferation in PB or BM in cases of CML doesn't by all means exclude the possibility of concomitant extramedullary blast crisis as an initial event. The combined cytogenetic, molecular and clinical studies in CML cases can bring to light the frequency of this event and disease out come. Although rare; this possibility should be raised and thoroughly investigated whenever clinically suspected. Download : Download high-res image (336KB) Download : Download full-size image Disclosures No relevant conflicts of interest to declare.

ID:2049 Effect of human umbilical cord-derived MSCs on the proliferation of peripheral blood T lymphocytes with different level HLA matching

Human umbilical cord-derived MSCs are very attractive sources of stem cells for the non-autologous cell transplantation therapy to treat many kinds of diseases based on their efficiency in treating graft versus host disease (GVHD), a condition that patient might suffer after receive cell-transplantation. However, the immune properties of MSCs are also critical for their efficiency in regenerative medicine applications. Thus, the mechanism of their action is now under intense investigation. A number of researches showed that MSCs exert strong anti-inflammatory and immunosuppressive effects on the main immune cell subsets through their production of various soluble factors. As a result, scientists all over the globe are changing their attention to the immunomodulatory property of MSCs, hence we are not an exception. Our research was carried out in order to strengthen our knowledge about this spectacular aspect of MSCs, especially the effect of cord-derived MSCs on the proliferation of peripheral blood T lymphocytes with different level HLA matching. Our results indicated that at passage 4 of culture, human umbilical cord-derived MSCs had no effect on the proliferation of T cells with all blood sample tested.

NKG2D Gene Polymorphism Is Associated with Disease Control of Chronic Myeloid Leukemia By Dasatinib

Dasatinib (DA) is a tyrosine kinase inhibitor (TKI), and is approved for the treatment of naïve and relapsed/refractory Philadelphia chromosome positive leukemia. DA is a potent and multi-target TKI, and a part of patients treated with DA are reported to exhibit lymphocytosis and that is caused by a proliferation of NK cells or CD8 T cells. Nowadays, a lot of reports about the stop TKI trials suggested that treatment free remission of the CML after TKI discontinuation was significantly associated with NK cell proliferation in peripheral blood of the patients. So far, little is known about the individual differences in NK cell response among patients treated with Dasatinib.NKG2D is an activating receptor expressed on NK cells. Recently, functional SNP were identified between LNK1 and HNK1 haplotypes of the NKG2D gene. We analyzed the association between patient's NKG2D polymorphism and achievement of the deep molecular response by Dasatinib. We retrospectively investigated thirty-one patients who were treated with Dasatinib for the first-line treatment of CML at Tokai university school of medicine between 2011 and 2015. Patients with HNK1 haplotype tended to achieve MR4.5 more frequently (not significant), and these patients accomplished MR4.5significantly faster than others.Next, we harvested NK cells from healthy volunteers, and investigated NK cell responses by Dasatinib in vitro. There were differences in the degree of NK cell proliferation capacity among subjects after DA treatment in vitro. We found that NK cells with NKG2D LNK1/LNK1 haplotype exhibited significantly lower proliferation capacity than others in the population. NK cells of higher proliferation capacity exhibited Vav-1 Tyr174 phosphorylation, but they didn't show any differences in cytotoxicity capacity.In this study, our data suggest that DA may stimulate NK cells with NKG2D HNK1 haplotype more strongly than LNK1 haplotype, and therefore patients with HNK1 haplotype can achieve deep molecular response faster than others. So, personalized medicine according to their NKG2D haplotype may be useful to the treatment of CML, especially when they are considered to discontinue DA in stop TKI trials. DisclosuresNo relevant conflicts of interest to declare.

The effects of telmisartan on the nuclear factor of activated T lymphocytes signalling pathway in hypertensive patients.

Hypothesis:Previous studies provide links between the nuclear factor of activated T lymphocytes (NFAT) signalling pathway and the development of hypertension. Our preliminary studies indicate that telmisartan can block Kv1.3 potassium channels and effectively inhibit potassium current densities, along with Kv1.3 mRNA and protein expression levels. This paper aims to investigate whether telmisartan has an inhibitory effect on the NFAT signalling pathway after activation and proliferation of peripheral blood T lymphocytes in Kazakh patients with essential hypertension (EH) from Xinjiang, China.Materials and methods:T lymphocytes were isolated using the immunomagnetic cell sorting method (MACS). The mRNA expression of NFATc1, IL-6 and TNF-α was measured by quantitative polymerase chain reaction (qRT-PCR) and relative protein levels were evaluated by Western blot. T cell samples from 50 hypertensive Kazakh patients from Xinjiang were randomly divided into control, telmisartan, cyclosporin A (CsA), VIVIT, and 4-aminopytidine (4-AP) groups. Peripheral blood T lymphocytes were first activated and proliferated in vitro, then incubated for 48 h under different treatment conditions before determination of protein and mRNA expression of NFATc1, IL-6, and TNF-α by Western blot and qRT-PCR analyses, respectively.Results:There were no significant differences in cardiovascular risk factors among the patients with samples assigned to the five groups (p > 0.05). Expression of NFATc1, IL-6, and TNF-α mRNA and protein was significantly reduced in T lymphocytes in all treatment groups (telmisartan, CsA, VIVIT, and 4-AP) compared with controls.Conclusions:Antihypertensive function and inhibitory effects of telmisartan on the T lymphocyte NFAT signalling pathway are unlikely to affect the normal immune function of hypertensive patients. Telmisartan may exert anti-inflammatory effects by inhibition of the NFAT signalling pathway in the T lymphocytes of hypertensive patients.

IMCT-10A PHASE I DOSE ESCALATION STUDY TO TEST THE SAFETY OF INTRATUMORAL ADOPTIVE IMMUNE THERAPY WITH AlloCTL IN RECURRENT GLIOMA PATIENTS

In a Phase 1 dose escalation study we evaluated the safety, feasibility, toxicity, and survival of patients treated with allogeneic cytotoxic T lymphocytes and recombinant IL-2 (alloCTL, rIL-2) infusion administered to the resection cavity of patients with recurrent glioma (WHO grade III or IV). AlloCTL are generated when gamma-irradiated lymphoblasts from the brain tumor patient stimulate the proliferation of peripheral blood mononuclear cells (PBMC) isolated from an HLA-disparate healthy donor in ex vivo culture via one-way mixed lymphocyte reaction. Nine patients were treated on the protocol and received a range of 0.5-5 cycles of therapy. Patients and donors were mismatched at a minimum of 2 HLA A, B, DR loci. Infusates were at least 30% CD3+. Cytotoxicity of alloCTL against patient PBMC ranged from 21.0-84.4% at a 30:1 effector to target ratio. The ratio of CD4/CD8 T cells averaged 1.12 (range 0.14-2.95). In vitro restimulation of alloCTL with patient lymphoblasts allowed for quantitation of immune cell subset proliferation and production of IFN-γ. Adverse events were noted at all three dose levels tested, and were primarily limited to Grade 1-2. Grade 3-4 events were noted only at dose levels 2 and 3. One patient experienced a dose limiting toxicity characterized by Grade 3 fatigue, arthralgia, headache, and non-infectious meningitis. Patient serum samples were collected during the course of therapy to determine Th1/Th2 cytokine profiles using Luminex multiplex analysis. Tumor progression was evident in 8/9 patients resulting in removal from the clinical trial. Median overall survival from the time of inclusion into the study was 8.8 months (range- 3.6-35.7 months). Two patients are still alive at greater than 35.7 and 16.4 months. The results suggest that alloCTL infusion is safe, feasible, and may warrant further study based on the clinical results.

Intramuscular delivery of heterodimeric IL-15 DNA in macaques produces systemic levels of bioactive cytokine inducing proliferation of NK and T cells.

Interleukin-15 (IL-15) is a common γ-chain cytokine that has a significant role in the activation and proliferation of T and NK cells and holds great potential in fighting infection and cancer. We have previously shown that bioactive IL-15 in vivo comprises a complex of the IL-15 chain with the soluble or cell-associated IL-15 receptor alpha (IL-15Rα) chain, which together form the IL-15 heterodimer. We have generated DNA vectors expressing the heterodimeric IL-15 by optimizing mRNA expression and protein trafficking. Repeated administration of these DNA plasmids by intramuscular injection followed by in vivo electroporation in rhesus macaques resulted in sustained high levels of IL-15 in plasma, with no significant toxicity. Administration of DNAs expressing heterodimeric IL-15 also resulted in an increased frequency of NK and T cells undergoing proliferation in peripheral blood. Heterodimeric IL-15 led to preferential expansion of CD8+NK cells, all memory CD8+ T-cell subsets and effector memory CD4+ T cells. Expression of heterodimeric IL-15 by DNA delivery to the muscle is an efficient procedure to obtain high systemic levels of bioactive cytokine, without the toxicity linked to the high transient cytokine peak associated with protein injection.

Effect of a combination of green and blue monochromatic light on broiler immune response

Our previous study suggested that green light or blue light would enhance the broiler immune response; this study was conducted to evaluate whether a combination of green and blue monochromatic light would result in improved immune response. A total of 192 Arbor Acre male broilers were exposed to white light, red light, green light, and blue light from 0 to 26days. From 27 to 49days, half of the broilers in green light and blue light were switched to blue light (G–B) and green light (B–G), respectively. The levels of anti-Newcastle disease virus (NDV) and anti-bovine serum albumin (BSA) IgG in G–B group were elevated by 11.9–40.3% and 17.4–48.7%, respectively, compared to single monochromatic lights (P<0.05). Moreover, the proliferation of peripheral blood T and B lymphocytes and the IL-2 concentration in the G–B groups increased by 10.4–36.2%, 10.0–50.0% and 24.7–60.3% (P<0.05), respectively, compared with the single monochromatic light groups. However, the serum TNF-α concentration in the G–B group was reduced by 3.64–40.5% compared to other groups, and no significant difference was found between the G–B and B–G groups in any type of detection index at the end of the experiment. These results suggested that the combination of G–B and B–G monochromatic light could effectively enhance the antibody titer, the proliferation index of lymphocytes and alleviate the stress response in broilers. Therefore, the combination of green and blue monochromatic light can improve the immune function of broilers.

Changes in the immune cell population and cell proliferation in peripheral blood after gemcitabine-based chemotherapy for pancreatic cancer

Regulatory T cells (Tregs) play a role in the immunosuppressive state in pancreatic cancer patients. We aimed to evaluate the changes of immune cells population including Tregs caused by gemcitabine (GEM)-based chemotherapy. Fifty-three patients with pancreatic cancer were enrolled in this study, of which 32 received GEM- based chemotherapy. Blood samples were collected before and at least 2weeks after the last dose of chemotherapy. The peripheral blood mononuclear cells (PBMCs) were subjected to flow cytometry analysis after labeling with anti-CD4, anti-CD25, and anti-Foxp3 antibodies. Other lymphocytes and NK cell markers were also measured. The proliferative capacity of PBMCs stimulated with anti-CD3 was analyzed using H(3) thymidine. The percentage and number of Tregs were significantly decreased after chemotherapy (p=0.032, p=0.003, respectively). The other immune cells and the proliferative capacity did not change. This study showed that GEM-based chemotherapy produced an immunomodulatory effect via the depletion of Tregs.

English

To investigate the effects of various monochromatic lights on T lymphocytes proliferation and serum nitric oxide production in chicken vaccinated with infectious bursal disease and newcastle disease vaccines, a total of 60 one-day-old broilers were exposed to red, green, blue, white and yellow light by using a light-emitting diode system for 6 weeks. The results indicated that the proliferation of peripheral blood T lymphocytes in the chickens exposed to white and green lights significantly increased when compared with other groups at 37 days of age (P < 0.05). In the early days, the results were different. The enhancement of T lymphocytes proliferation with green and yellow lights occurred at 18 days, whereas the enhancement with green light was less than other lights at 30 days. Red light promoted NO (nitrix oxide) production at maximum level in the chickens, while green light suppressed it at minimum level after 37 days. These results suggested that green and white lights had strong effects on immunity, especially at the last days of rearing. Key words: Monochromatic lights, vaccination, nitric oxide, broiler chicken.

Normalised EBV Quantity in Patients After Allogeneic Haematopoietic Stem Cell Transplantation – Benign Reactivation, Localised and Generalised EBV Lymphoproliferative Disease

EBV remains important pathogen in patients after haematopoietic stem cell transplantation (HSCT). Beside benign reactivation, EBV can cause localised or generalised form of EBV associated lymphoproliferative disease (EBV-LPD) with clinical symptoms such as fever, hepathopathy, lymphoma-like symptoms and generalised B cell proliferation in peripheral blood.

Effect of Vanadium on the Subset and Proliferation of Peripheral Blood T Cells, and Serum Interleukin-2 Content in Broilers

The purpose of this 42-day study was to investigate the effects of dietary excess vanadium on immune function by determining changes of the subsets and proliferation function of peripheral blood T cells. Four hundred twenty 1-day-old avian broilers were divided into six groups and fed on a corn-soybean basal diet as control diet or the same diet amended to contain 5, 15, 30, 45, and 60ppm vanadium supplied as ammonium metavanadate. In comparison with those of the control group, the percentages of CD (3) (+) , CD (3) (+) CD (4) (+) , and CD (3) (+) CD (8) (+) were decreased in 45 and 60ppm groups from 14 to 42days of age, and the percentages of CD (3) (+) and CD (3) (+) CD (4) (+) were increased in 5ppm group at 42days of age. The CD (4) (+) /CD (8) (+) ratio was increased in 45 and 60ppm groups at 28days of age. Meanwhile, the proliferation function of peripheral blood T cell were decreased in 30, 45, and 60ppm groups from 14 to 42days of age. Also, the serum interleukin-2 contents were decreased in 45 and 60ppm groups from 14 to 42days of age and increased in 5ppm group at 28days of age. Histopathologically, hypocellularity appeared in the thymus in 45 and 60ppm groups. It was concluded that dietary vanadium in excess of 30ppm reduced the percentages of peripheral blood T-cell subsets and the proliferation function and serum interleukin-2 contents. The cellular immune function was finally impaired in broilers.

Systemic immunization with CCL27/CTACK modulates immune responses at mucosal sites in mice and macaques

Plasmid DNA is a promising vaccine platform that has been shown to be safe and able to be administered repeatedly without vector interference. Enhancing the potency of DNA vaccination through co-delivery of molecular adjuvants is one strategy currently under investigation. Here we describe the use of the novel chemokine adjuvant CCL27/CTACK to enhance immune responses to an HIV-1 or SIV antigen in mice and rhesus macaques. CCL27 has been shown to play a role in inflammatory responses through chemotaxis of CCR10+ cells, and we hypothesized that CCL27 may modulate adaptive immune responses. Immunizations in mice with HIV-1gag/CCL27 enhanced immune responses both at peripheral and, surprisingly, at mucosal sites. To confirm these findings in a large-animal model, we created optimized CCL27 and SIV antigenic plasmid constructs for rhesus macaques. 10 macaques ( n = 5/group) were immunized intramuscularly with 1 mg/construct of antigenic plasmids ± CCL27 with electroporation. We observed significant IFN-γ secretion and CD8+ T-cell proliferation in peripheral blood. Interestingly, CCL27 co-immunized macaques exhibited a trend toward greater effector CD4+ T cells in the bronchiolar lavage (BAL). CCL27 co-delivery also elicited greater antigen-specific IgA at unique sites including BAL and fecal samples but not in the periphery. Future studies incorporating CCL27 as an adjuvant in vaccine or therapy models where eliciting immune responses in the lung are warranted.

Self-complementary AAV Virus (scAAV) Safe and Long-term Gene Transfer in the Trabecular Meshwork of Living Rats and Monkeys

AAV vectors produce stable transgene expression and elicit low immune response in many tissues. AAVs have been the vectors of choice for gene therapy for the eye, in particular the retina. scAAVs are modified AAVs that bypass the required second-strand DNA synthesis to achieve transcription of the transgene. The goal was to investigate the ability of AAV vectors to induce long-term, safe delivery of transgenes to the trabecular meshwork of living animals. Single doses of AAV2.GFP and AAV2.RGD.GFP/Ad5.LacZ were injected intracamerally (IC) into rats (n = 28 eyes). A single dose of scAAV.GFP was IC-injected into rats (n = 72 eyes) and cynomolgus monkeys (n = 3). GFP expression was evaluated by fluorescence, immunohistochemistry, and noninvasive gonioscopy. Intraocular pressure (IOP) was measured with calibrated tonometer (rats) and Goldmann tonometer (monkeys). Differential expression of scAAV-infected human trabecular meshwork cells (HTM) was determined by microarrays. Humoral and cell-mediated immune responses were evaluated by ELISA and peripheral blood proliferation assays. No GFP transduction was observed on the anterior segment tissues of AAV-injected rats up to 27 days after injection. In contrast, scAAV2 transduced the trabecular meshwork very efficiently, with a fast onset (4 days). Eyes remained clear and no adverse effects were observed. Transgene expression lasted >3.5 months in rats and >2.35 years in monkeys. The scAAV viral vector provides prolonged and safe transduction in the trabecular meshwork of rats and monkeys. The stable expression and safe properties of this vector could facilitate the development of trabecular meshwork drugs for gene therapy for glaucoma.