Interleukin-2 Gene Research Articles

Pre-treatment with IL2 gene therapy alleviates Staphylococcus aureus arthritis in mice

BackgroundStaphylococcus aureus (S. aureus) arthritis is one of the most detrimental joint diseases known and leads to severe joint destruction within days. We hypothesized that the provision of auxiliary immunoregulation via an expanded compartment of T regulatory cells (Tregs) could dampen detrimental aspects of the host immune response whilst preserving its protective nature. Administration of low-dose interleukin 2 (IL2) preferentially expands Tregs, and is being studied as a treatment choice in several autoimmune conditions. We aimed to evaluate the role of IL2 and Tregs in septic arthritis using a well-established mouse model of haematogenously spred S. aureus arthritis.MethodsC57BL/6 or NMRI mice we intravenously (iv) injected with a defined dose of S. aureus LS-1 or Newman and the role of IL2 and Tregs were assessed by the following approaches: IL2 was endogenously delivered by intraperitoneal injection of a recombinant adeno-associated virus vector (rAAV) before iv S. aureus inoculation; Tregs were depleted before and during S. aureus arthritis using antiCD25 antibodies; Tregs were adoptively transferred before induction of S. aureus arthritis and finally, recombinant IL2 was used as a treatment starting day 3 after S. aureus injection. Studied outcomes included survival, weight change, bacterial clearance, and joint damage.ResultsExpansion of Tregs induced by IL2 gene therapy prior to disease onset does not compromise host resistance to S. aureus infection, as the increased proportions of Tregs reduced the arthritis severity as well as the systemic inflammatory response, while simultaneously preserving the host’s ability to clear the infection.ConclusionsPre-treatment with IL2 gene therapy dampens detrimental immune responses but preserves appropriate host defense, which alleviates S. aureus septic arthritis in a mouse model.

Non-significant association between \u2212\u2009330\u2009T/G polymorphism in interleukin-2 gene and chronic periodontitis: findings from a meta-analysis

BackgroundChronic periodontitis (CP) is an immune-inflammatory disease that promotes tissue damage around the teeth. Among the several inflammatory mediators that orchestrate the periodontitis, there is the interleukin (IL)-2. Genetic variations in IL2 gene may be associated with the risk and severity of the disease. Contrary results are available in the literature with inconclusive findings and none meta-analysis to gather these data.MethodsA literature search was performed for studies published before June 11, 2019 in diverse scientific and educational databases. The data was extracted by two investigators and the statistical evaluation was performed by Review Manager statistical program with heterogeneity (I2) and Odds Ratio (OR) with 95% of Confidence Intervals (CI) calculations and a sensitive analysis to assess the accuracy of the obtained results. The publication bias was evaluated by Begg’ and Egger’s test with Comprehensive meta-analysis software. The value of P < 0.05 was considered as significant.ResultsFive studies were identified in diverse ethnical groups with 1425 participants. The − 330 T/G polymorphism in IL2 gene was not significantly associated with CP in allelic evaluation (P > 0.05) as well as in the genotypic comparisons (P = 0.15). The Begg’s test and the linear regression Egger’s test did not show any evidence of publication bias risk (P > 0.05) which was corroborated by the absence of obvious asymmetry in Funnel plot graphic.ConclusionsThis meta-analysis showed a non-significant association between − 330 T/G polymorphism in IL2 gene and CP in any allelic evaluation.

Health-related quality of life in dogs treated with electrochemotherapy and/or interleukin-12 gene electrotransfer.

The aim of this study was to evaluate the owners' perception of health‐related quality of life (HRQoL) of dogs after treatment with electrochemotherapy (ECT) alone or combined with interleukin‐12 gene electrotransfer (IL‐12 GET) and/or surgery. The owners of 44 dogs with histologically different tumours were offered the »Cancer Treatment Form« at least one month after treatment. The owners assessed their dogs’ quality of life (QoL) after treatment as good (mean 7.4) (from 1–very poor to 10–excellent) and the general health compared with the initial diagnosis of cancer as improving (mean 3.9) (from 1–worse to 5–better). The assessment of the current QoL was better within the group of dogs treated with non‐invasive treatment (ECT and/or IL‐12 GET only), compared with those that received invasive treatment, where, in addition to ECT and/or IL‐12 GET, surgery was performed (p < .05). The owners of dogs that achieved an objective response (OR) to the treatment assessed the QoL as significantly better compared with those whose dogs did not respond to the treatment (p < .05). The majority of the owners (86.4%) would opt for the therapy again, regardless of the financial costs. In conclusion, the results of this study demonstrate that the majority of the owners of dogs assessed their dogs’ QoL as good and felt that it improved after the treatment, especially in dogs, treated with non‐invasive treatment and in those that responded to the treatment. This supports further use of ECT and IL‐12 GET as suitable methods for the treatment of selected tumours in veterinary medicine.

Systematic review and meta-analysis of the association between C-746T and C-703T polymorphisms of the interleukin-5 gene and asthma

Background: Asthma is a common disease with a complex risk architecture including both genetic and environmental factors. A number of studies have identified the association between interleukin (IL)-5 C-746T and C-703T polymorphisms and asthma risk, however, the results still remain inconclusive. The objective of the present study was to identify the effect of IL-5 polymorphisms in asthma susceptibility. Methods: PubMed and Google Scholar databases were searched. The odds ratio (OR) with its 95% confidence interval (CI) was employed to calculate the effect and strength of association in the random-effects model or fixed-effects model. Results: A total of 4 case–control and 2 cross-sectional studies were screened out, including 1499 asthma patients and 3766 controls. Two single-nucleotide polymorphisms of the IL-5 gene were identified. Our results detected a no significant association between IL-5 C-746T polymorphisms and asthma risk in the total population (CC genotype showed OR = 0.82, 95% CI = 0.67–1.00, P = 0.05, I2 = 0%; CT genotype showed OR = 0.94, 95% CI = 0.78–1.13, P = 0.50, I2 = 24%; and TT genotype showed OR = 0.85, 95% CI = 0.64–1.13, P = 0.27, I2 = 0%) while a significant association between IL-5 C-703T polymorphisms and asthma in children according to review results. Conclusion: The meta-analysis findings suggest a lack of direct association between the IL-5 C-746T polymorphism and asthma and found that C-703T polymorphisms of the IL-5 gene might contribute to asthma risk. Future well-designed case–control studies with a large population and more ethnicities are still needed to estimate the association.

Effects of interleukin-12 gene polymorphism on response to hepatitis B vaccination among hemodialysis Egyptian patients

BackgroundHepatitis B virus (HBV) infection is a major health problem among hemodialysis (HD) patients. Interleukin (IL)-12 gene polymorphisms may be associated with immune response variability to recombinant HBV vaccines. The aim was to determine the correlation between IL-12 gene polymorphism and hepatitis B surface-antibody (HBs-Ab) titer in response to HBV vaccine among HD Egyptian patients.Patients and methodsSeventy patients receiving long-term HD and 20 age-matched and sex-matched healthy controls were enrolled. All participants were non-HBV vaccinated and seronegative for HBV and HIV. Recombinant HBV vaccine was given (three-dose scheduled). Thereafter, HBs-Ab titer and IL-12 gene polymorphism were evaluated 8 weeks after the last vaccination dose.ResultsThere was no response (HBs-Ab<10 μIU/ml) in 20% of HD patients and 10% of the controls. HBs-Ab titers showed no significant correlation with duration of HD, BMI, serum albumin, hemoglobin, leucocytic count, parathyroid hormone level, or IL-12 gene polymorphism. Responders to vaccination had significantly lower transferrin saturation and significantly higher levels of urea reduction ratio, Kt/V and lymphopenia. IL-12B genotype frequency was as follows: AA (58.3 vs. 55.6%), AC (37.5%) and CC (4.2 vs. 0%) in responders of either HD or control participants, respectively (P>0.05 for all).ConclusionThere was no significant association between IL-12B gene polymorphism and HBs-Ab response in Egyptian HD patients. In HD patients, lymphocytopnea, diabetes mellitus (DM), high transferrin saturation and inefficient HD were associated with HBV vaccine hyporesponsiveness.

Optimization of Enhanced Expression of Interferon-γ and Interleukin-2 in Chicken Peripheral Blood Mononuclear Cells by Polysaccharide Immunomodulators Using Historical-Data Response Surface Methodology

Background: Natural polysaccharides have been found to act as potent immunomodulatory agents. Objectives: The present study aimed to assess and optimize the concentration of polysaccharides driven from Chlorella by-product (CBP) for interferon-γ (IFN-γ) and interleukin-2 (IL-2) gene expression by chicken peripheral blood mononuclear cells (PBMCs). Methods: In this study, a crude polysaccharide (CP) was extracted from CBP and fractionated to F1 and F2 using anion-exchange chromatography. Extracted polysaccharides were used at the concentrations of 200 to 1000 μg/mL in the culture medium for PBMCs. The data were analyzed by historical-data response surface methodology (RSM) approach. The analysis of variance test was applied to check the significance of RSM results. Results: The effects of immunomodulator type and concentration were statistically significant (P < 0.0001) on both cytokines. In addition, the interaction effect was also significant for IFN-γ expression (P = 0.03). The regression models of IFN-γ and IL-2 expression were confirmed by the non-significant lack of fit (P = 0.056 and P = 0.066, respectively), the determination coefficient (R2 = 0.98 and R2 = 0.96, respectively), and adjusted determination coefficient (Adj. R2 = 0.97 and Adj. R2 = 0.94, respectively). The predicted determination coefficients (Pred. R2 = 0.96 and Pred. R2 = 0.92 for IFN-γ and IL-2 expression, respectively) were in reasonable agreement with the adjusted R2 values and confirmed that the models had a good predictive ability. The adequate precision values of above 4 (37.51 and 24.45 for IFN-γ and IL-2, respectively) also proved the established models could navigate the design space. Furthermore, the optimization results by the historical-data design of RSM revealed that F1 at 786.68 μg/mL with the desirability of 0.87 could be selected to maximize the gene expression of IFN-γ and IL-2 by chicken PBMCs. Conclusions: It is concluded that the immunostimulatory effect was more prominent for F1 than for CP or F2 polysaccharides. In addition, historical-data RSM showed to be a valuable technique to optimize the factors and reach maximum gene expression in chicken PBMCs. Further investigations are now in progress to determine the interrelationship between the molecular structure and bioactivities of CP and fractionated polysaccharides.

TAC Chemotherapy and Pembrolizumab Plus Interleukin-12 Gene Therapy and L-NMMA in Triple Negative Breast Cancer

TAC Chemotherapy and Pembrolizumab Plus Interleukin-12 Gene Therapy and L-NMMA in Triple Negative Breast Cancer

Associations of IL-12, IL12R polymorphisms and serum IL-12 levels with high-risk human papillomavirus susceptibility in rural women from Luohe, Henan, China.

Background:Interleukin 12 (IL-12) and interleukin 12 receptor (IL12R), key inflammatory cytokines in the immune system, participate in bridging the innate immunity and adaptive immunity. No previous work has reported the role of IL-12 and IL12R in high-risk human papillomavirus (hrHPV) susceptibility. The purpose of this study was to investigate the association of IL-12, IL12R polymorphisms, and serum IL-12 levels with hrHPV susceptibility in rural women from Luohe, Henan, China.Methods:Two hundred sixty cases with hrHPV infection and 260 healthy controls were selected. Enzyme-linked immunosorbent assays were used to detect the serum IL-12 levels, and the polymorphisms of IL12B rs3212227, IL12RB1 rs393548, and IL12RB1 rs436857 were determined using DNA sequencing.Results:The serum IL-12 levels were significantly lower in cases with hrHPV infection compared with those in healthy controls (P < .01).There was no significant difference in IL12 rs3212227, IL12RB1rs436857, and IL12RB1rs393548 genotype and allele frequencies between cases and controls (P > .05). Furthermore, with respect to the IL12 rs3212227 polymorphism with serum IL-12 levels, although serum IL-12 levels were lower in cases than in controls, we did not find any differences between serum IL-12 levels and genotypes in cases(P > .05).Conclusions:Our data demonstrates that low serum IL-12 levels may be associated with hrHPV susceptibility but are not associated with IL-12 gene polymorphisms; furthermore, IL-12 and IL12R gene polymorphisms may not contribute susceptibility to hrHPV in rural women from Luohe, Henan, China.

Genetic Variants of the IL2 Gene Related to Risk and Survival in Patients With Colorectal Cancer.

Interleukin 2 (IL2) is a significant factor activating T-cell-mediated immune response by stimulation of natural killer cells, T-cells and in development of regulatory T (Treg) cells. Recent studies have that IL2 participates in cancer development by modifying the local immune response. Based on the suggested role of the single nucleotide polymorphisms (SNPs) rs2069762, rs6822844 and rs11938795 of IL2 in the pathogenesis of certain diseases, the relationship of these SNPs with clinicopathological variables and their possible implication for prognosis and disease outcome were evaluated in a cohort of Swedish patients with colorectal cancer (CRC). TaqMan SNP genotype assays based on polymerase chain reaction were used for analysis of the IL2 SNPs in 467 patients with CRC and 467 healthy controls. Expression analysis of IL2 in plasma and CRC tissue was also performed. The allelic variants T in rs11938795 and G in rs6822844 were significantly associated with a higher risk of CRC. Kaplan-Meier analysis showed that cancer-specific survival was worse for individuals with C allele for rs2069762 with stage II CRC and with T allele for rs6822844 with stage III CRC. SNPs rs2069762, rs6822844 and rs11938795 of the IL2 gene may be helpful as prognostic biomarkers in the follow-up and management of the patients.

Influence of introduction of interleukin-2 and interleukin-12 into experimental marker DNA-vaccine

Aim. To determine the effects of the combined administration of recombinant expression vectors containing genes of murine interleukin-2 and interleukin-12 on humoral immune response, elicited by the experimental marker DNA-vaccine against classical swine fever. Methods. Expression of chimeric proteins in vitro and in vivo was determined by western-blot analysis. The antibodies specific to target antigens in blood serum have been detected by ELISA. Results. A series of recombinant plasmid vectors containing the genes of murine interleukin-2 and chimeric murine interleukin-12 have been developed. It has been shown that target chimeric proteins were expressed from the developed vectors both in vitro in HEK293 and in vivo in murine muscle tissue. The use of combined administration of murine interleukin-2 and interleukin-12 genes resulted in significant enhancement of titer of the anti-E2 and anti-β-galactosidase IgG, induced by vaccination with experimental marker DNA-vaccine against CSF, and model DNA-vaccine respectively. Conclusions. The data obtained show that the introduction of recombinant expressing vectors containing genes of murine interleukin-2 and interleukin-12 into vaccine preparations enhances humoral immune response elicited by the experimental marker DNA-vaccine against CSF and modelDNA-vaccine. Keywords: DNA-vaccine, humoral immune response, interleukin-2, interleukin-12, classical swine fever.

Generation and functional evaluation of a DNA vaccine co-expressing Vibrio anguillarum VAA protein and flounder interleukin-2

In our previous study, a DNA plasmid encoding the VAA gene of Vibrio anguillarum was constructed and demonstrated to confer moderated protection against V. anguillarum challenge. Here, a bicistronic DNA vaccine (pVAA-IRES-IL2), co-expressing the VAA gene of V. anguillarum and Interleukin-2 (IL2) gene of flounder, was constructed to increase the protective efficacy of VAA DNA vaccine. The potential of pVAA-IRES-IL2 to express both VAA and IL2 in transfected HINAE cell lines was confirmed by immunofluorescence assay. Further, the variation of sIgM+, CD4-1+, CD4-2+ lymphocytes and production of VAA-specific antibodies in flounder, which was intramuscularly immunized with three DNA plasmids (pIRES, pVAA-IRES, pVAA-IRES-IL2), were investigated, respectively. The bacterial burden and relative percentage survival (RPS) of flounder exposed to V. anguillarum infection were both analyzed to evaluate the efficacy of bicistronic DNA plasmid. Our results revealed that the percentages of sIgM+, CD4-1+, CD4-2+ lymphocytes and antibodies specific to VAA were remarkably increased in pVAA-IRES or pVAA-IRES-IL2 immunized fish. Moreover, the co-expression of IL2 enhanced the immune response in response to VAA DNA vaccination, as shown by the higher percentages of sIgM+, CD4-1+, CD4-2+ lymphocytes and production of specific antibody. Importantly, the RPS in pVAA-IRES-IL2 and pVAA-IRES groups reached 64.1% and 51.3%, respectively, when compared with the 97.5% cumulative mortality in pIRES group. Furthermore, the number of V. anguillarum in liver, spleen and kidney of pVAA-IRES or pVAA-IRES-IL2 immunized flounder after V. anguillarum challenge was significantly reduced, as compared to that in pIRES group. These suggest that the bicistronic DNA vaccine can be an effective immunization strategy in inducing immune response against V. anguillarum infection and IL2 has the potential as the adjuvant for VAA DNA vaccine.

Bone marrow–derived mesenchymal stromal cells promote resistance to tyrosine kinase inhibitors in chronic myeloid leukemia via the IL-7/JAK1/STAT5 pathway

Chronic myeloid leukemia (CML) is caused by the fusion of the BCR activator of RhoGEF and GTPase activating protein (BCR) and ABL proto-oncogene, the nonreceptor tyrosine kinase (ABL) genes. Although the tyrosine kinase inhibitors (TKIs) imatinib (IM) and nilotinib (NI) have remarkable efficacy in managing CML, the malignancies in some patients become TKI-resistant. Here, we isolated bone marrow (BM)-derived mesenchymal stem cells (MSCs) from several CML patients by Ficoll-Hypaque density-gradient centrifugation for coculture with K562 and BV173 cells with or without TKIs. We used real-time quantitative PCR to assess the level of interleukin 7 (IL-7) expression in the MSCs and employed immunoblotting to monitor protein expression in the BCR/ABL, phosphatidylinositol 3-kinase (PI3K)/AKT, and JAK/STAT signaling pathways. We also used a xenograft tumor model to examine the in vivo effect of different MSCs on CML cells. MSCs from patients with IM-resistant CML protected K562 and BV173 cells against IM- or NI-induced cell death, and this protection was due to increased IL-7 secretion from the MSCs. Moreover, IL-7 levels in the BM of patients with IM-resistant CML were significantly higher than in healthy donors or IM-sensitive CML patients. IL-7 elicited IM and NI resistance via BCR/ABL-independent activation of JAK1/STAT5 signaling, but not of JAK3/STAT5 or PI3K/AKT signaling. IL-7 or JAK1 gene knockdown abrogated IL-7-mediated STAT5 phosphorylation and IM resistance in vitro and in vivo Because high IL-7 levels in the BM mediate TKI resistance via BCR/ABL-independent activation of JAK1/STAT5 signaling, combining TKIs with IL-7/JAK1/STAT5 inhibition may have significant utility for managing CML.

Growth Promotion and Immune Stimulation in Nile Tilapia, Oreochromis niloticus, Fingerlings Following Dietary Administration of a Novel Marine Probiotic, Psychrobacter maritimus S.

A 50-day feeding trial was conducted to evaluate the effects of dietary supplementation of a novel marine psychrotrophic bacterium, Psychrobacter maritimus S, on growth performance and immune responses of Nile tilapia (Oreochromis niloticus) fingerlings. Triplicate groups of Nile tilapia (10 fish each) were fed for 50days with three different diets: T0 (without probiotic, negative control), T0.5 (supplemented with 3.3 × 108CFUg-1P. maritimus S), and T1 (supplemented with 6.6 × 108CFUg-1P. maritimus S). The test diets were fed to fish at a daily rate of 3% of their live weights, three times per day. On the 50th day, fish were weighed and blood samples were collected to determine the main nonspecific humoral and cellular immune responses and digestive enzymes (protease, amylase, and lipase). Fish performance (weight gain, specific growth rate, feed conversion ratio, protein efficiency ratio, and protein productive value) was also calculated. Growth rates and digestive enzyme activity were significantly increased (P < 0.05) with increasing dietary P. maritimus S up to 0.5%, and leveled off with further increase in bacterial concentrations. Phagocytic activity, lysozyme activity, alternative complement hemolysis, and hematological parameters were also significantly increased (P < 0.05) with increasing P. maritimus S concentration to 0.5%, followed by a slight decrease (P > 0.05) at 1% level. The expression of interleukin-4 and interleukin-12 genes was significantly upregulated, while heat shock protein gene was downregulated, with dietary supplementation of P. maritimus S up to 0.5% level. These results suggest that 0.5% of dietary Psychrobacter maritimus S supplementation could be considered as a novel probiont for optimum growth performance and immune response of Nile tilapia fingerlings.

Effects of cyclosporine and dexamethasone on canine T cell expression of interleukin-2 and interferon-gamma

Cyclosporine and glucocorticoids are powerful immunosuppressive agents used to treat many inflammatory diseases in dogs. Cyclosporine inhibits calcineurin-dependent pathways of T cell activation and resultant T cell cytokine production, and glucocorticoids directly inhibit genes coding for cytokines. Little work has been done comparing the effects of these agents on T cell cytokine production in dogs. Our study measured T cell interleukin-2 (IL-2) and interferon-gamma (IFN-γ) production using flow cytometry and T cell IL-2 and IFN-γ gene expression using quantitative reverse transcription polymerase chain reaction (qRT-PCR) in activated canine T cells incubated with cyclosporine and dexamethasone in vitro. For flow cytometric assays, diluted whole blood was cultured for 7 h in the presence of cyclosporine (10, 100, 500, and 1000 ng/mL) or dexamethasone (10 ng/mL, 100 ng/mL, 1 μg/mL, and 10 μg/mL). For qRT-PCR, whole blood was cultured for 5 h with the same drugs at the same concentrations, and RNA was then extracted from leukocytes. Flow cytometry and qRT-PCR both demonstrated inhibition of IL-2 and IFN-γ that was concentration-dependent in response to cyclosporine, and was more variable for dexamethasone. Quantitative RT-PCR but not flow cytometry documented significant reduction of IL-2 expression after dexamethasone treatment, while both methods showed concentration-dependent suppression of IFN-γ. Quantitative RT-PCR also revealed additional cytokine suppression at higher cyclosporine concentrations, an effect not found using flow cytometry, and may therefore be the preferred method for cytokine determination in dogs. Suppression of IL-2 and IFN-γ in activated T cells may have potential as an indicator of the efficacy of cyclosporine and glucocorticoids in suppressing canine T cell function in vivo, and may therefore be of value for characterizing the immunosuppression induced by these drugs in clinical patients.

Mode of Action of Aspirin in Experimental Autoimmune Encephalomyelitis.

Multiple sclerosis (MS) is a chronic and debilitating autoimmune disorder of the central nervous system in which the autoimmune T cells destroy myelin, thus causing lesion, damage, and neuronal dysfunction. Experimental autoimmune encephalomyelitis (EAE) is an animal model of MS that is particularly useful for testing new therapeutic approaches against MS. Aspirin (acetyl salicylic acid) is one of the oldest and widely used medicines in the world, and recently it has been shown that low-dose aspirin is capable of suppressing the disease process of EAE in mice. One of the root causes of this autoimmune disease process is the decrease and/or suppression of Foxp3-expressing anti-autoimmune regulatory T cells (Tregs) and associated increase in autoimmune T-helper 1 (Th1) and Th17 cells. Aspirin upregulates Tregs and decreases Th1 and Th17 responses. Accordingly, the suppression of Tregs abrogates the protective effect of aspirin on EAE, indicating that aspirin protects EAE via Tregs. While there are several mechanisms for the maintenance of Tregs under immune insults, aspirin increases the level of interleukin-11 (IL-11), an immunomodulatory cytokine, and IL-11 alone is sufficient to protect Tregs. Being a multifunctional molecule, aspirin stimulates the activation of cAMP-response element-binding (CREB) to promote the recruitment of CREB to the IL-11 gene promoter and stimulate the transcription of IL-11 in splenocytes. Therefore, it appears that low-dose aspirin protects EAE via CREB-mediated stimulation of IL-11-Treg pathway and that aspirin may have therapeutic importance in MS.

THOR-707: Using synthetic biology to reprogram the therapeutic activity of interleukin-2 (IL-2).

2603 Background: Recombinant interleukin-2 (rIL-2; aldesleukin) is an approved immunotherapy in melanoma and renal cell carcinoma based on complete durable remissions. The anti-neoplastic properties of IL-2 are mediated by interactions with the beta-gamma chain (IL-2Rβγ) on naïve CD8+ T cells, which lead to their expansion and differentiation into T effector and T memory cells directed against the tumor. However, the widespread use of IL-2 in oncology is limited by interaction with the high affinity IL-2 receptor alpha chain (IL-2R⍺) on regulatory CD4+ T cells (Tregs), which leads to immunosuppression, and on innate lymphoid cells in the vascular endothelium, which leads to eosinophilic recruitment and activation, and the sometimes fatal complication of vascular leak syndrome (VLS). A rIL-2 biased toward IL-2βγ affinity with no IL-2R⍺interaction could fill unmet needs in oncology. Methods: Using a synthetic biology platform, we have engineered THOR-707, a rIL-2 that contains a novel amino acid encoded in the IL-2 gene via a new DNA base pair (X-Y). The novel amino acid serves as a hook for site specific pegylation that extends half-life, blocks IL-2R⍺ engagement and binds to the IL-2Rβγ. Results: In non-human primates, THOR-707 can be dosed to maximize the level of cytotoxic CD8+ T lymphocytes without elevation of VLS-inducing eosinophils. In murine tumor models, THOR-707 induced the expansion of peripheral and intratumoral CD8+ T cells without expansion of suppressive Tregs. Single-agent dose-dependent anti-tumor efficacy was observed in two syngeneic mouse models. In combination with a PD-1 inhibitor, survival of tumor-bearing mice was longer than either agent as monotherapy. Efficacy in tumor models was durable, suggesting activation of CD8+ memory T cell populations. Conclusions: THOR-707 is a reprogrammed, site-directed, singly-pegylated rIL-2 that changes the pharmacologic profile of IL-2, potentially providing a favorable risk-benefit profile. First-in-human studies are expected to begin this year evaluating THOR-707 as monotherapy and in combination with a PD-1 inhibitor. Based on preclinical evidence to-date, THOR-707 may potentially address existing and emerging unmet needs across multiple solid tumors.

Enhanced antitumor effects by combining IL-12 gene electrotransfer with anti-PD1 in preclinical metastatic cancer model

Abstract The survival and expansion of effector cytotoxic T lymphocytes (CTLs) during an immunological response are critical for the successful elimination of cancer cells. Among the factors that regulate the immune response, interleukin-12 (IL-12), is known to stimulate growth and survival of T cells. Major differences exist between these lymphokines in their capacity to act on various T-cell types such as CD4+ or CD8+ and effector versus memory T lymphocytes. PD1 expression can have a negative effect on effector or memory T-cell populations causing anergy or cell death. Limited information is available regarding the function of combining IL-12 gene electrotransfer (GET) with anti-PD1 in preclinical metastatic cancer models. Here, we report that IL-12 (GET) induced expansion of effector CD8 CTLs. Combining IL-12 GET with anti-PD1 i.p. injection resulted in long-term survival of CD8+PD1− T cells. Most significantly, the effector cytolytic activity of CTLs were expanded and maintained for up to 60 days, indicating that these cells do not differentiate into a memory functional phenotype. In a two site metastatic model, B16F10 melanoma cells expressing luciferase were injected via intraperitoneal route and B16.F10 cells (no luciferase) were injected subcutaneously. Results on day 60 using an In Vivo Imaging System (IVIS) illuminated a lower flux signal in intraperitoneal metastases. Simultaneously, the primary subcutaneous tumors completely regressed in 80% of these mice. The expression of CD44+ CD62L− increased with time, which was detected on CD8 CTLs, suggesting that the memory response was still active at 120 days. The present findings have practical implications for the combination of IL-12 gene electrotransfer with anti-PD1.

Immunization of mice by the co-administration of codon-optimized HPV16 E7 and lL12 genes against HPV16-associated cervical cancer

BackgroundVarious promising procedures have been used to improve the potency of DNA vaccines for the treatment of human papillomavirus type 16 (HPV16) infections. Interleukin-12 (IL12) is a powerful adjuvant that can contribute to T cell-mediated protection against many pathogens, specifically viruses. Considering the important role of T cell-mediated immunity in tumor clearance, the induction of these responses can help control the progression of tumors in animal models. We have demonstrated that the co-administration of codon-optimized E7 (uE7) gene of HPV16 with interleukin-12 is effective in the development of antitumor responses. ObjectivesThe present study examined the co-administration of codon-optimized HPV16 E7 gene with murine interleukin-12 gene (mIL-12) as a vaccine adjuvant in tumor mice model. Materials and methodsC57BL/6 mice were studied for tumor progression after injection of recombinant DNA vaccines. Lactate dehydrogenase (LDH) and IFN-γ were measured to evaluate the activity of cytotoxic T lymphocytes (CTLs). Measurements of tumor volume and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay were used for assessment of therapeutic antitumor effects of the vaccines. ResultsResults showed that DNA vaccines, specifically codon-optimized E7/murine interleukin-12 (mIL-12), elicited significant differences in levels of IFN-γ and cytotoxic T lymphocyte (CTLs) responses compared to control groups. Furthermore, higher antitumor response and lower tumor size in the vaccine group was significantly evident compared to control group. ConclusionThe co-administration of codon-optimized HPV16 E7 gene with IL12 significantly enhances the DNA vaccine potency against HPV16-associated cervical cancer.

Effect of dietary supplementation of mannanoligosaccharides on hepatic gene expressions and humoral and cellular immune responses in aflatoxin-contaminated broiler chicks

The present study was conducted to investigate the effects of dietary supplementation of mannanoligosaccharides (MOS) on expression of hepatic immunological genes and immune responses in aflatoxin-contaminated broiler chicks. A total of 336 seven-day-old Ross broiler chicks were randomly allotted to 7 experimental treatments with 4 replicates and 12 birds per replicate. Experimental treatments consisted of 2 aflatoxin levels (0.5 and 2 ppm) and 3 supplemental MOS levels (0, 1 and 2 g/kg) as a 2 × 3 factorial arrangement in comparison with a control group (unchallenged group). The chicks were challenged with a mix of aflatoxins during 7–28 d of age. Results showed that aflatoxin challenge resulted in the lower antibody titers against infectious bronchitis (IBV) and bursal (IBD) diseases viruses. In addition, aflatoxin-contaminated birds had a lower (P < 0.0001) lymphocyte percentage and a decline in (P < 0.01) interleukin-2 (IL-2) mRNA abundance. Likewise, heterophil proportion, heterophil to lymphocyte ratio and gene expressions of hepatic interleukin-6 (IL-6) and C reactive protein (CRP) were raised (P < 0.001) by increasing dietary aflatoxin level. Dietary inclusion of MOS increased (P < 0.05) antibody titers against IBV, IBD and Newcastle disease virus. Lymphocyte proportion and hepatic IL-2 gene expression were greater (P < 0.0001) in MOS-supplemented birds. Furthermore, supplemental MOS decreased hepatic IL-6 and CRP abundances. Additionally, inclusion of 2 g/kg MOS resulted in the upregulation (P < 0.01) of hepatic IL-2 gene expression in birds contaminated with 0.5 ppm aflatoxin. The present results indicate that supplemental MOS could improve cellular immunity via the upregulation of hepatic IL-2 gene expression in birds challenged with aflatoxins.

Association of expression of inflammatory response genes and DNA repair genes in colorectal carcinoma.

Inflammation is an important etiological factor of colorectal carcinoma and may be related to colorectal carcinoma growth and proliferation. This study aimed to verify whether the presence of chronic inflammation represented by tumor necrosis factor-α, interleukin-2, interleukin-6, and interleukin-10 gene expression is related to hMLH1, hMSH2, hMSH6, and PMS2 gene expression and the corresponding protein levels of these genes from the DNA repair system. A total of 83 patients were operated on for curative or palliative colorectal carcinoma. Expression of the inflammatory response genes tumor necrosis factor-α, interleukin-2, interleukin-6, and interleukin-10 as well as expression of the hMLH1, hMSH2, hMSH6, and PMS2 genes of the DNA repair system (mismatch repair) and the expression levels of the corresponding mismatch repair proteins were measured in neoplastic tissue by reverse transcription polymerase chain reaction and immunohistochemistry, respectively. Associations were observed between hMSH6 mRNA expression and interleukin-2 mRNA expression (p = 0.026) as well as between hMLH1 and hMSH2 gene expression and tumor necrosis factor-α gene expression (p = 0.042). Higher tissue levels of interleukin-2 and tumor necrosis factor-α gene expression were associated with lower hMSH6, hMLH1, and hMSH2 gene expression.