Th2 Cytokine Secretion Research Articles

Human endoglin-CD3 bispecific T cell engager antibody induces anti-tumor effect in vivo.

Rationale: Endoglin, also known as CD105, is a homo-dimeric membrane glycoprotein required for angiogenesis and serves as a marker for cancer vasculature. In this study, we constructed a bispecific T-cell engager (BiTE) antibody that targets human endoglin and CD3 (hEND-CD3/BiTE). We examined BiTE binding to endoglin-expressing cells and its effects on the cytolytic activity of T cells and cancer development.Methods: The in vitro effects of hEND-CD3/BiTE, including binding to target cells, T-cell activation, proliferation, and cytotoxicity, were examined in endoglin-expressing 293T cells, human umbilical vascular endothelial cells, tumor-derived endothelial cells, and CD3+ T cells. An in vivo xenograft tumor model was established using A549 human lung cancer cells. The therapeutic efficacy of hEND-CD3/BiTE was assessed by monitoring tumor growth, angiogenesis, and mouse survival.Results: hEND-CD3/BiTE specifically bound to endoglin-expressing cells and CD3+ T cells in vitro and stimulated T-cell activation, proliferation, and Th1 cytokine secretion, and promoted T-cell-mediated cytolysis of endoglin-expressing cells. The hEND-CD3/BiTE in vivo caused minimal toxicity to major organs, reduced tumor neoangiogenesis, inhibited tumor growth, and significantly improved mouse survival.Conclusions: Our study demonstrated the therapeutic potential of hEND-CD3/BiTE and provided a novel approach to clinical cancer treatment.

Experimental autoimmune encephalomyelitis inhibited by huangqi guizhi wuwu decoction via th2 cytokine enhancement

Background: Huangqi Guizhi Wuwu decoction (HQGZWW) exhibits good effects when administered to treat multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Understanding the precise mechanism of this decoction is thus important. Based on the findings of our previous study, the aim of the present study was to understand the role of antigen-specific CD8+ T-cells on the pathogenesis of MS/EAE when HQGZWW is administered as treatment. Methods: Myelin oligodendrocyte glycoprotein (MOG) 35-55-induced mice were administered distilled water, prednisone, and high dose or low dose HQGZWW. After purified CD4+ and CD8+ T-cells were stimulated with the MOG35-55 peptide, proliferation and cytokine secretion assays were performed. To establish the adoptive transfer EAE model, naive mice were injected with MOG35-55 -CD8+ or CD4+ T-cells. Results: Significant improvements in EAE score and pathology were observed in the high dose HQGZWW and prednisone groups. Compared to the low dose HQGZWW and distilled water groups, lower antigen-specific responses, lower levels of interferon-gamma, and higher levels of interleukin (IL)-4 and IL-10 from CD8+ and CD4+ T cells were observed in the high dose HQGZWW and prednisone groups. Finally, the EAE score was observed to be similar between the high dose HQGZWW group and prednisone group; however, this finding was not observed in the low dose HQGZWW group. Conclusion: Our findings suggest that high dose HQGZWW has similar effects on cell proliferation, cytokine secretion, and EAE score to prednisone, while low dose HQGZWW does not have such effect. The protective role of HQGZWW against EAE might thus depend on the Th2 cytokine secretion profile induced by either MOG35-55 specific CD8+ or CD4+ T-cells.

T-bet is a critical determinant in the instability of the IL-17-secreting T-helper phenotype

IL-23, an IL-12-related cytokine, induces an IL-17-secreting T-helper phenotype that is involved in autoimmune diseases and host defense against certain pathogens. Although the transcription factors required for development of IL-23-stimulated cells are unknown, we show that T-bet is a critical negative regulator of the IL-23-primed T-cell phenotype, which we term Th1beta. Th1 or Th1beta Tbx21-/- cultures secrete higher than WT levels of IL-17 in response to T-cell receptor (TCR) or IL-23 + IL-18 stimulation. Ectopic T-bet expression in Th1beta cells promotes IFN-gamma secretion but decreases IL-17 production. Although antigen-receptor stimulation of Th1beta cells stimulates IL-17 production, it also induces the IFN-gamma-independent expression of T-bet and progression to a Th1 cytokine secretion pattern. T-bet is required for the progression to the Th1 phenotype, because Tbx21-/- Th1beta cultures maintain the IL-17-secreting phenotype after 2 weeks of culture. Addition of IFN-gamma to Tbx21-/- Th1beta cultures cannot recover the progression to the Th1 phenotype, suggesting T-bet, rather than IFN-gamma, mediates Th1beta to Th1 progression. The transient nature of the Th1beta phenotype suggests that these cells are a component of type I immunity and that T-bet expression is a critical determinant of Th1 versus Th1beta cell fate.

Robust Production of Merkel Cell Polyomavirus Oncogene Specific T Cells From Healthy Donors for Adoptive Transfer.

Virus positive Merkel cell carcinoma (VP-MCC) is an aggressive but immunogenic skin malignancy driven by Merkel cell polyomavirus (MCPyV) T antigen (TAg). Since adoptive T cell transfer (ACT) can be effective against virus-driven malignancies, we set out to develop a methodology for generating MCPyV TAg specific T cells. MCPyV is a common, asymptomatic infection and virus-exposed healthy donors represent a potential source of MCPyV TAg specific T cells for ACT. Virus specific T cells were generated using monocyte-derived dendritic cells (moDCs) pulsed with MCPyV TAg peptide libraries and co-cultured with autologous T cells in supplemented with pro-inflammatory and homeostatic cytokines for 14 days. Specific reactivity was observed predominantly within the CD4+ T cell compartment in the cultures generated from 21/46 random healthy donors. Notably, responses were more often seen in donors aged 50 years and older. TAg specific CD4+ T cells specifically secreted Th1 cytokines and upregulated CD137 upon challenge with MCPyV TAg peptide libraries and autologous transduced antigen presenting cells. Expanded T cells from healthy donors recognized epitopes of both TAg splice variants found in VP-MCC tumors, and minimally expressed exhaustion markers. Our data show that MCPyV specific T cells can be expanded from healthy donors using methods appropriate for the manufacture of clinical grade ACT products.

Callicarpa japonica Thunb. ameliorates allergic airway inflammation by suppressing NF-κB activation and upregulating HO-1 expression

Ethnopharmacological relevanceCallicarpa japonica Thunb., as an herbal medicine has been used for the treatment of inflammatory diseases in China and Korea. Materials and methodsUltra performance liquid chromatography–photodiode array–quadrupole time–of–flight mass spectrometer (UPLC–PDA–QTof MS) was used to detect the major phenylethanoid glycosides in the C. japonica extract. BALB/c mice were intraperitoneally sensitized by ovalbumin (OVA) (on days 0 and 7) and challenged by OVA aerosol (on days 11–13) to induce airway inflammatory response. The mice were also administered with C. japonica Thunb. (CJT) (20 and 40 mg/kg Per oral) on days 9–13. CJT pretreatment was conducted in lipopolysaccharide (LPS)-stimulated RAW264.7 or phorbol 12-myristate 13-acetate (PMA)-stimulated A549 cells. ResultsCJT administration significantly reduced the secretion of Th2 cytokines, TNF-α, IL-6, immunoglobulin E (IgE) and histamine, and the recruitment of eosinophils in an OVA-exposed mice. In histological analyses, the amelioration of inflammatory cell influx and mucus secretion were observed with CJT. The OVA-induced airway hyperresponsiveness (AHR), iNOS expression and NF-κB activation were effectively suppressed by CJT administration. In addition, CJT led to the upregulation of HO-1 expression. In an in vitro study, CJT pretreatment suppressed the LPS-induced TNF-α secretion in RAW264.7 cells and attenuated the PMA-induced IL-6, IL-8 and MCP-1 secretion in A549 cells. These effects were accompanied by downregulated NF-κB phosphorylation and by upregulated HO-1 expression. ConclusionThese results suggested that CJT has protective activity against OVA-induced airway inflammation via downregulation of NF-κB activation and upregulation of HO-1, suggesting that CJT has preventive potential for the development of allergic asthma.

In vitro effects of 1-(4-dimethylaminobenzylidene)-2-(2-hydroxybenzylidene) hydrazine and its copper complex on lymphocyte proliferation and function in the presence of free radical generator.

In this study, the in vitro effects of 1-(4-dimethylaminobenzylidene)-2-(2-hydroxybenzylidene) hydrazone (L1) and its corresponding copper complex [Cu(L1)], synthesized in our laboratory, were investigated on the proliferative responses, Th1 (interleukin-2 (IL-2), interferon-γ (INFγ)) and Th2 (IL-4) cytokine secretion, adenosine triphosphate (ATP) levels and intracellular redox status of T lymphocytes submitted to H2O2/FeSO4-mediated oxidative stress. T cells were isolated on histopaque density gradient by differential centrifugation, and were cultured with the mitogen concanavalin A (Con A), free radical generator (H2O2/FeSO4) and with different concentrations of L1 and [Cu(L1)] (1 - 100 μM). Proliferation (MTT assay), cytokines (Elisa kits), ATP levels, cytotoxic effect (micronucleus test) and oxidative markers (glutathione, catalase, superoxide dismutase, hydroperoxide and carbonyl protein contents) were investigated after 48-h incubation. Our results showed that H2O2/FeSO4 treatment induced a reduction in T lymphocyte proliferation, cytokine secretion and ATP levels associated to an evident intracellular oxidative stress, inflammatory profile and DNA damage. Addition of L1 at 100 μM was able to increase cell proliferation, IL-2, IL-4 and INFγ secretion and ATP contents and to reduce hydroperoxide and carbonyl protein contents, catalase activity and micronuclei number in lymphocytes under oxidative stress, with a partial protection. The [Cu(L1)] exhibited protective effects in T lymphocytes by inhibiting H2O2/FeSO4 - induced cell proliferation suppression, inflammatory status, ATP loss and oxidative stress generation, whatever the concentration used. In conclusion, in the situation of excessive oxidative stress, [Cu(L1)] treatment improved T lymphocyte proliferation, cytokine production, ATP contents and oxidant/antioxidant status. [Cu(L1)] could be effective at improving oxidative stress and T cell abnormalities.

Elucidating the Pivotal Immunomodulatory and Anti-Inflammatory Potentials of Chloroquine and Hydroxychloroquine.

Chloroquine (CQ) and hydroxychloroquine (HCQ) are derivatives of 4-aminoquinoline compounds with over 60 years of safe clinical usage. CQ and HCQ are able to inhibit the production of cytokines such as interleukin- (IL-) 1, IL-2, IL-6, IL-17, and IL-22. Also, CQ and HCQ inhibit the production of interferon- (IFN-) α and IFN-γ and/or tumor necrotizing factor- (TNF-) α. Furthermore, CQ blocks the production of prostaglandins (PGs) in the intact cell by inhibiting substrate accessibility of arachidonic acid necessary for the production of PGs. Moreover, CQ affects the stability between T-helper cell (Th) 1 and Th2 cytokine secretion by augmenting IL-10 production in peripheral blood mononuclear cells (PBMCs). Additionally, CQ is capable of blocking lipopolysaccharide- (LPS-) triggered stimulation of extracellular signal-modulated extracellular signal-regulated kinases 1/2 in human PBMCs. HCQ at clinical levels effectively blocks CpG-triggered class-switched memory B-cells from differentiating into plasmablasts as well as producing IgG. Also, HCQ inhibits cytokine generation from all the B-cell subsets. IgM memory B-cells exhibits the utmost cytokine production. Nevertheless, CQ triggers the production of reactive oxygen species. A rare, but serious, side effect of CQ or HCQ in nondiabetic patients is hypoglycaemia. Thus, in critically ill patients, CQ and HCQ are most likely to deplete all the energy stores of the body leaving the patient very weak and sicker. We advocate that, during clinical usage of CQ and HCQ in critically ill patients, it is very essential to strengthen the CQ or HCQ with glucose infusion. CQ and HCQ are thus potential inhibitors of the COVID-19 cytokine storm.

Mycobacterium tuberculosis–Specific Antigen Rv3619c Effectively Alleviates Allergic Asthma in Mice

Despite significant advances, asthma remains a cause of premature death, and current treatments are suboptimal. Antigen-specific Th2 cells and their cytokines are primary mediators of the pathophysiological changes seen in asthma. Studies in animal models have shown that mycobacteria can suppress the asthma phenotype by alteration of the Th1/Th2 cytokines ratio. In this study, utilizing a Th1 delivery system to modulate the allergic airway inflammation in a Th2-driven model of asthma, we evaluated the efficacy of immunization with Mycobacterium tuberculosis-specific antigen Rv3619c, either alone or in combination with low dose dexamethasone. The rv3619c gene was cloned in an expression plasmid pGES-TH-1, expressed in Escherichia coli, and the recombinant protein Rv3619c was purified to homogeneity using affinity chromatography. Mice were immunized with the recombinant protein emulsified in Freund’s Incomplete Adjuvant (IFA) alone and in combination with low dose dexamethasone, and then challenged with ovalbumin (OVA). Airway inflammation was assessed by quantifying airway cytology, histological changes and Th2 cytokine (IL-5) secretion from splenocytes. OVA-specific IgE, IgG and IgG1 from sera was assessed, as well as pERK1/2 expression in the lung tissue. Immunization with recombinant Rv3619c alone inhibited the OVA-induced increase in total cell counts, eosinophil airway cell infiltration in BAL fluid, perivascular and peribronchial inflammation and fibrosis, and goblet cell hyper/metaplasia. In addition, Rv3619c/IFA inhibited the OVA-induced IL-5 in spleen cells, OVA-specific IgE, IgG, and IgG1 levels in sera, and pERK1/2 expression in lung tissue. Immunization with Rv3619c/IFA in combination with low dose dexamethasone resulted in an enhanced effect on some but not all the asthma features. Taken together, this study demonstrates that immunization with Rv3619c/IFA, alone or in combination with dexamethasone, may be an effective treatment strategy for the prevention of asthma.

Food allergy promotes a Th2/Th17 response that drives house dust mite-induced allergic airway inflammation in humanized mice.

Food allergy is related to increasing risk of the development of allergic asthma, but the precise interplay between sensitization to different allergens in different compartments of the body is not fully understood. The aim of this study was to develop a novel humanized murine model of mixed food and respiratory allergy that recapitulates the human anaphylactic response and to more clearly understand the impact of food allergies on asthma. Immunodeficient mice transferred with peripheral blood mononuclear cells (PBMCs) from donors with peanut and house dust mite (HDM) allergy were exposed and challenged to peanut. Between peanut exposure and challenge, mice were intranasally treated to HDM. Allergic parameters were analyzed. Allergen-specific immunoglobulin (Ig)E in sera could only be measured in mice treated with peripheral blood mononuclear cells (PBMCs) plus allergen. A preceding peanut exposure increased IgE levels, histamine release, bronchial hyper-responsiveness and lung inflammation. Recruitment of inflammatory cells to the airways was aggravated associated with an enhanced T helper type 2 (Th2)/Th17 cytokine secretion when the two allergies were present. A preceding peanut exposure amplifies allergic asthma in this humanized model, which may contribute to the understanding of underlying immunological mechanism of polysensitization occurring in allergic individuals and evaluation of therapeutic interventions.

Association between IL-33 Gene Polymorphism (Rs7044343) and Risk of Allergic Rhinitis

ABSTRACT Background Allergic rhinitis (AR) is a T helper type 2 (Th2)-mediated upper airways disease in which genetics factors including cytokine genes play a prominent role. Interleukin-33 (IL-33) is a major cytokine for naive T cells polarization into Th2 phenotype as well as enhances the secretion of Th2 cytokines. The aim of the present study was to investigate the relationship between IL-33 single nucleotide polymorphisms (SNPs) and IL-33 serum level with Allergic rhinitis. Methods Blood samples were collected from 130 AR patients and 130 healthy individuals. SNPs (rs7044343 C > T, rs1929992 A > G, rs12551256 A > G) of IL-33 gene were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Serum level of IL-33 was measured by enzyme-linked immunosorbent assay (ELISA). Results Statistical analysis showed that the TT genotype (OR = 1.996, CI: 1.168–3.412, P = .01), as well as the T allele (OR = 0.675, CI: 0.476–0.957, P = .02) of rs7044343 C > T were significantly associated with reduced risk of AR. In addition, individuals carrying the TT genotype were associated with lower levels of IL-33 compared to subjects with CC and CT genotypes; however, these differences were not statistically significant. No association was found between rs1929992 and rs12551256 variants and risk of AR, but the GG genotype from rs1929992 A > G was associated with increased serum levels of IL-33 in control group (p = .01). Furthermore, serum IL-33 levels were not significantly different between AR patients and healthy controls (p > .05). Conclusion Our results suggest that the TT genotype of rs7044343 C > T may act as a protective agent against allergic rhinitis.

The establishment of humanized IL-4/IL-4RA mouse model by gene editing and efficacy evaluation

Asthma is a common respiratory immune disease in children and adults, and interleukin-4 (IL-4) is one of the key factors for the onset of asthma. Therefore, targeting human IL-4 and IL-4 receptor alpha (IL-4RA) has become one of the strategies for targeted therapy of cytokines. Herein, we established an animal model of asthmatic airway inflammation using double humanized IL-4/IL-4RA (hIL-4/hIL-4RA) mice, where human IL-4 and IL-4RA replaced their murine counterparts, respectively. We successfully identified the phenotype by Southern blotting, ELISA, and flow cytometry. The hIL-4/hIL-4RA mice induced by ovalbumin (OVA) exhibited several important features of asthma, such as inflammatory cell infiltration, IgE release, goblet cell hyperplasia, and Th2 cytokine secretion. Furthermore, treatment of these humanized mice with anti-human IL-4RA antibodies significantly inhibited level of these pathological indicators. Thus, hIL-4/hIL-4RA mice provide a validated preclinical mouse model to interrogate new therapeutic agents targeting this specific cytokine pathway in asthma.

B lymphocytes contribute to indirect pathway T cell sensitization via acquisition of extracellular vesicles.

B cells have been implicated in transplant rejection via antibody-mediated mechanisms and more recently by presenting donor antigens to T cells. We have shown in patients with chronic antibody-mediated rejection that B cells control the indirect T cell alloresponses. To understand more about the role of B cells as antigen-presenting cells for CD4+ T cell with indirect allospecificity, B cells were depleted in C57BL/6 mice, using an anti-CD20 antibody, prior to receiving MHC class I-mismatched (Kd ) skin. The absence of B cells at the time of transplantation prolonged skin graft survival. To study the mechanisms behind this observation, T cells with indirect allospecificity were transferred in mice receiving a Kd skin transplant. T cell proliferation was markedly inhibited in the absence of recipient B cells, suggesting that B cells contribute to indirect pathway sensitization. Furthermore, we have shown that a possible way in which B cells present alloantigens is via acquisition of MHC-peptide complexes. Finally, we demonstrate that the addition of B cell depletion to the transfer of regulatory T cells (Tregs) with indirect alloresponse further prolonged skin graft survival. This study supports an important role for B cells in indirect T cell priming and further emphasizes the advantage of combination therapies in prolonging transplant survival.

Enhanced immunogenicity of leukemia-derived exosomes via transfection with lentiviral vectors encoding costimulatory molecules

Background: Tumor cell-derived exosomes (TEXs) have been widely used to induce antitumor immune responses in animal models and clinical trials. Similarly, leukemia cell-derived exosomes (LEXs) can induce antileukemia immune responses in animal models. However, the antileukemia immunity induced by LEXs is less effective, which may be due to an inadequate costimulatory capacity.Methods: In this study, we transduced L1210 leukemia cells with a lentiviral vector encoding two B7 costimulatory molecules (CD80, CD86) and obtained LEXs that highly expressed CD80 and CD86. The antileukemia immune response derived from these LEXs was examined in vitro and in vivo in animal models.Results: We found that B7 gene-modified LEXs, including LEX-CD80, LEX-CD86, and LEX-8086, could significantly boost the expression of CD80 and CD86 in dendritic cells (DCs) and promote the secretion of functional cytokines such as TNF-α and IL-12. Moreover, these B7 gene-modified LEXs, particularly LEX-CD8086, could effectively induce CD4+ T cell proliferation, Th1 cytokine secretion, and an antigen-specific anti-leukemia cytotoxic T lymphocyte (CTL) response. Additional animal studies indicated that immunization with B7 gene-modified LEXs, in particular LEX-CD8086, could significantly retard tumor growth compared to the control LEXnull group.Conclusions: This study sheds light on the feasibility of obtaining LEXs that overexpress costimulatory molecules via genetically modified leukemia cells, thereby enhancing their anti-leukemia immunity and providing a potential therapeutic strategy that contributes to leukemia immunotherapy.

589 Effects of CCR4 antagonists in cutaneous T-cell lymphoma cells

CC chemokine receptor 4 (CCR4) is responsible for T-cell skin homing. It is overexpressed in type 2 helper T-cells (Th2) as well as cutaneous T-cell lymphoma (CTCL) cells. Mycosis fungoides (MF) and Sézary syndrome (SS) are two most common types of CTCL. Currently, Mogamulizumab, a humanized anti-CCR4 antibody, has been FDA-approved to treat MF and SS. However, the clinical response rates for SS patients was 30% and for MF patients was 22% and severe drug eruptions following treatment were reported. Thus, a more effective and less toxic strategy is needed for CCR4-targeted therapy. CCR4 antagonists have been studied in diseases where Th2 cells participate, such as asthma and atopic dermatitis, but little has been done for CTCL. Here, we tested two CCR4 antagonists, C021 (class I antagonist) and AZD2098 (class II antagonist), in MF derived cell line (MJ) and SS derived cell line (Hut78). The effects of two compounds on cell chemotaxis, proliferation, apoptosis, and Th2 cytokine secretion were assessed. As expected, CCR4 was highly expressed on MJ and Hut78 cells. MJ and Hut78 cells showed chemotactic responses to TARC/CCL17 and MDC/CCL22. Both C021 and AZD2098 inhibited chemotactic responses to TARC/CCL17 and MDC/CCL22 in MJ and Hut78 cells. Of note, C021 inhibited chemotaxis to CCL17 at a much lower concentration (IC50: 0.186 μM) than to CCL22 (1.3 μM) in MJ cells. AZD2098 inhibited chemotaxis to CCL17 at a much lower concentration (IC50: 0. 12 μM) than to CCL22 (0.866 μM) in Hut78 cells. Interestingly, only C021 downregulated CCR4 expression on cell surfaces. Only C021 inhibited cell proliferation, at a higher concentration (IC50: 3.21 μM in MJ cells; 5.98 μM in Hut78 cells), and induced cell apoptosis. AZD2098 had no such effect at any concentrations tested. In addition, the expressions of IL-5 and IL-13 proteins were decreased after treatment with C021 but not with AZD2098. Our results suggest that both CO21 and AZD2098 have inhibitory effects on chemotaxis of CTCL cells, but only C021 exerts other effects on CTCL cells. The in vivo study is on the way.

356 IL-1β induces rapid secretion of the antimicrobial protein IL-26 from Th17 cells

Th17 cells play a fundamental role in both immunity and autoimmunity at mucosal surfaces, including skin. Recent work has implicated the Th17 cytokine IL-26 as directly antimicrobial to extracellular and intracellular bacteria, including Mycobacterium leprae, the causative agent of leprosy. IL-26 protein expression was greater in skin lesions from patients with the resistant vs. progressive form of the disease. To characterize the mechanism of IL-26 induction, we examined the kinetics of IL-26 secretion by PBMCs exposed to Mycobacterium leprae sonicate, which revealed significant release at 3 hours, reminiscent of an innate response. Pattern recognition receptor ligands induced IL-26 secretion from PBMC, which was mediated by IL-1β. Recombinant IL-1β alone was sufficient to stimulate IL-26 release from PBMC and memory CD4+ T cells in the absence of T cell receptor (TCR) activation. IL-1β stimulated IL-26 secretion more rapidly from memory CD4+ T cells as compared to TCR activation with crosslinking antibodies. Further investigation revealed that IL-1RI expression was necessary for IL-1β induced IL-26 from memory Th17 cells. IL-1β stimulation did not lead to secretion of other Th17 cytokines, unlike TCR activation. RNA sequencing of IL-1β treated IL-1RI+ Th17 cells revealed enrichment for NF-κB regulated genes. Inhibition of NF-κB signaling with Bay 11-7082 abrogated IL-26 production in response to either stimulus. Finally, supernatants from IL-1β treated memory T cells showed antimicrobial activity against E. coli in an IL-26 dependent manner. Together, these results identify IL-1RI+ Th17 cells as an antimicrobial Th17 subpopulation with the ability to rapidly respond to IL-1β and induce IL-26 to kill extracellular bacteria.

Targeting the interleukin-7 receptor alpha by an anti-CD127 monoclonal antibody improves allergic airway inflammation in mice.

Interleukin-7 (IL-7) is the most important cytokine for T-cell homeostasis. IL-7 signals through the IL-7 receptor (IL-7R) which is composed of an alpha chain (IL-7Rα), also called CD127 and a common gamma chain. T lymphocytes, especially T helper type 2, play a crucial role in the pathobiology of allergic asthma. To study the effects of an anti-CD127 monoclonal antibody (mAb) in a murine model of allergic airway inflammation induced by house dust mite (HDM). Allergic airway inflammation was induced in mice using a protocol comprising 4 weekly percutaneous sensitizations followed by 2 weekly intranasal challenges with total HDM extracts and treated by intraperitoneal injections of an anti-CD127 mAb. Because CD127 is shared by both IL-7R and the receptor for thymic stromal lymphopoietin (TSLP), a group of mice was also treated with an anti-IL-7 mAb to block only the IL-7 signalling pathway. Anti-CD127 mAb-treated mice showed significantly lower airway resistance in response to methacholine and improvement in lung histology compared with isotype mAb-treated animals. Anti-CD127 mAb treatment significantly decreased the mRNA expression of Th2 cytokines (IL-4, IL-5, and IL-13) and chemokines (CCL5/RANTES) in lung tissue, decreased the secretion of Th2 cytokines (IL-4, IL-5, and IL-13) and chemokines (CXCL1 and CCL11/eotaxin) in bronchoalveolar lavage fluid (BALF), decreased serum HDM-specific IgE, and reduced the number of total leucocytes and leucocyte subpopulations such as eosinophils, macrophages, lymphocytes, T lymphocytes, and ILC2 in BALF and lung tissue. Mice treated with anti-IL-7 mAb also showed less allergic airway inflammation as evidenced by significantly lower airway resistance and fewer leucocytes in BALF and lung tissue compared to mice treated with the corresponding isotype control mAb. Targeting the IL-7Rα by an anti-CD127 mAb improves allergic airway inflammation in mice and presents as a potential therapeutic approach for allergic asthma.

CD2 Regulates Pathogenesis of Asthma Induced by House Dust Mice Extract

Characteristic of allergic asthma, CD4+Th2 lymphocytes secrete Th2 cytokines, interleukin (IL)-4, IL-13, and IL-5 that mediate the inflammatory immune response. Surface expression of CD2 and its ligand, CD58, is increased on the monocytes and eosinophils of asthma patients, which correlate with elevated serum IgE levels, suggesting that CD2 may contribute to allergic airway inflammation. Using a murine model of asthma, we observed that house dust mice extract (HDME)-exposed Balb/c mice have increased airway hyperresponsiveness (AHR), lung inflammation, goblet cell hyperplasia, and elevated levels of Th2 cytokines in the lungs, as well as increased serum IgE levels as compared to the control mice. In contrast, with the exception of serum IgE levels, all the other parameters were significantly reduced in HDME-treated Cd2−/− mice. Interestingly, Il13 but not Il4 or Il5 gene expression in the lungs was dramatically decreased in HDME-exposed Cd2−/− mice. Of note, the gene expression of IL-13 downstream targets (Muc5b and Muc5ac) and high affinity IL-13Rα2 were upregulated in the lungs of HDME-exposed Balb/c mice but were significantly reduced in HDME-exposed Cd2−/− mice. Consistently, gene expression of microRNAs regulating mucin production, inflammation, airway smooth muscle cell proliferation and IL-13 transcripts were increased in the lungs of HDME-exposed Cd2−/− mice. Given the established role of IL-13 in promoting goblet cell hyperplasia, lung inflammation and AHR in allergic asthma, our studies reveal a unique role for CD2 in the regulation of Th2-associated allergic asthma.

Potent Bidirectional Cross-Talk Between Plasmacytoid Dendritic Cells and γδT Cells Through BTN3A, Type I/II IFNs and Immune Checkpoints.

Plasmacytoid DCs (pDCs) and γδT cells are both critical players in immunosurveillance against pathogens and cancer due to their ability to sense microbes and cell stress through recognition of pathogen-associated molecular patterns or altered metabolism [phosphoantigens (PAgs)]. Their unique features, high functional plasticity and ability to interact with many immune cell types allow them to bridge innate and adaptive immunity, initiating and orientating widely immune responses, hence contributing to protective and pathogenic immune responses. Yet, despite strategic and closed missions, potential interactions between pDCs and γδT cells are still unknown. Here we investigated whether there is interplay between pDCs and γδT cells and the underlying molecular mechanisms. Purified human pDCs and γδT cells were cocultured in presence of TLR-L, PAg, and zoledronate (Zol) to mimic both infectious and tumor settings. We demonstrated that TLR7/9L- or Zol-stimulated pDCs drive potent γδT-cell activation, Th1 cytokine secretion and cytotoxic activity. Conversely PAg-activated γδT cells trigger pDC phenotypic changes and functional activities. We provided evidence that pDCs and γδT cells cross-regulate each other through soluble factors and cell-cell contacts, especially type I/II IFNs and BTN3A. Such interplay could be modulated by blocking selective immune checkpoints. Our study highlighted crucial bidirectional interactions between these key potent immune players. The exploitation of pDC-γδT cells interplay represents a promising opportunity to design novel immunotherapeutic strategies and restore appropriate immune responses in cancers, infections and autoimmune diseases.

Anti-CS1 × Anti-CD3 Bispecific Antibody (BiAb)-Armed Anti-CD3 Activated T Cells (CS1-BATs) Kill CS1+ Myeloma Cells and Release Type-1 Cytokines.

Background: Multiple myeloma (MM) remains incurable despite significant advances in chemotherapy, targeted therapies, and immunotherapy. Bispecific antibody (BiAb)-armed activated T cells (BATs) have been developed for targeting and treatment of solid and hematologic malignancies. BATs are serial killers of tumor cells, secrete Th1 cytokines, and induce adaptive cellular and humoral immune responses in patients (pts). This study provides preclinical data using bispecific anti-CS1 (elotuzumab) × anti-CD3 (OKT3) antibody (CS1Bi)-armed activated T cells (CS1- BATs) that provide a strong rationale for applying CS1-BATs to pts with MM.Methods: CS1-BATs and unarmed activated T cells (ATC) were incubated with MM cell targets at various effector to target ratios (E:T) in a quantitative flow cytometry-based assay to determine the degree of cell loss relative to target cells incubated without ATC. ATC from up to 8 normal donors were armed with various concentrations of CS1 BiAb and tested against 5 myeloma cells lines for CS1-BATs-mediated killing and release of Th1 cytokines, chemokines and granzyme B.Results: CS1-BATs from normal donors killed each of 5 MM cell lines proportional to E:T ratios ranging between 1:1 and 10:1 and arming concentrations of 12.5 to 50 ng/million ATC, which was accompanied by release of Th1 cytokines, chemokines and granzyme B. CS1-BATs prepared from MM pts' peripheral blood mononuclear cells (PBMC) showed increasing cytotoxicity and T cell expansion over time against ARH77 MM cells. The optimal arming dose of CS1Bi is 50 ng/106 ATC.Conclusions: These data demonstrate the therapeutic potential of CS1-BATs-mediated cytotoxicity and Th1 cytokines release at low E:T and support advancing their clinical development in pts with MM.

MicroRNAs: future biomarkers and targets of therapy in asthma?

MicroRNAs (miRNAs) are small noncoding RNA molecules that are considered one of the fundamental regulatory mechanisms of gene expression. They are involved in many biologic processes, such as signal transduction, cell proliferation and differentiation, apoptosis and stress responses. The purpose of this review is to present recent knowledge about the role of miRNAs in asthma and outline possible applications of miRNAs. A core set of miRNAs involved in asthma includes downregulated let-7 family, miR-193b, miR-375 as well as upregulated miR-21, miR-223, miR-146a, miR-142-5p, miR-142-3p, miR-146b and miR-155. Recently it has been shown that most of the involved miRNAs increase secretion of Th2 cytokines, decrease secretion of Th1 cytokines, promote differentiation of T cells towards Th2 or play a role in hyperplasia and hypertrophy of bronchial smooth muscle cells. The profiles of miRNAs correlate with clinical characteristics, including lung function, phenotype and severity of asthma. Recent publications confirmed crucial regulatory role of miRNAs in the pathomechanism of asthma. Some single miRNAs or their sets hold the promise for their use as asthma biomarkers facilitating diagnosis or prediction of treatment outcomes. They are also possible target of future therapies. The studies in this field are lacking though.