Kyriaki Dunussi-Joannopoulos

Interleukin (IL)-22 and IL-17 are coexpressed by Th17 cells and cooperatively enhance expression of antimicrobial peptides

Th17 cells are a distinct lineage of effector CD4+ T cells characterized by their production of interleukin (IL)-17. We demonstrate that Th17 cells also expressed IL-22, an IL-10 family member, at substantially higher amounts than T helper (Th)1 or Th2 cells. Similar to IL-17A, IL-22 expression was initiated by transforming growth factor β signaling in the context of IL-6 and other proinflammatory cytokines. The subsequent expansion of IL-22–producing cells was dependent on IL-23. We further demonstrate that IL-22 was coexpressed in vitro and in vivo with both IL-17A and IL-17F. To study a functional relationship among these cytokines, we examined the expression of antimicrobial peptides by primary keratinocytes treated with combinations of IL-22, IL-17A, and IL-17F. IL-22 in conjunction with IL-17A or IL-17F synergistically induced the expression of β-defensin 2 and S100A9 and additively enhanced the expression of S100A7 and S100A8. Collectively, we have identified IL-22 as a new cytokine expressed by Th17 cells that synergizes with IL-17A or IL-17F to regulate genes associated with skin innate immunity.

IL-22 is required for Th17 cell–mediated pathology in a mouse model of psoriasis-like skin inflammation

Psoriasis is a chronic skin disease resulting from the dysregulated interplay between keratinocytes and infiltrating immune cells. We report on a psoriasis-like disease model, which is induced by the transfer of CD4(+)CD45RB(hi)CD25(-) cells to pathogen-free scid/scid mice. Psoriasis-like lesions had elevated levels of antimicrobial peptide and proinflammatory cytokine mRNA. Also, similar to psoriasis, disease progression in this model was dependent on the p40 common to IL-12 and IL-23. To investigate the role of IL-22, a Th17 cytokine, in disease progression, mice were treated with IL-22-neutralizing antibodies. Neutralization of IL-22 prevented the development of disease, reducing acanthosis (thickening of the skin), inflammatory infiltrates, and expression of Th17 cytokines. Direct administration of IL-22 into the skin of normal mice induced both antimicrobial peptide and proinflammatory cytokine gene expression. Our data suggest that IL-22, which acts on keratinocytes and other nonhematopoietic cells, is required for development of the autoreactive Th17 cell-dependent disease in this model of skin inflammation. We propose that IL-22 antagonism might be a promising therapy for the treatment of human psoriasis.

An IL-17F/A Heterodimer Protein Is Produced by Mouse Th17 Cells and Induces Airway Neutrophil Recruitment

IL-17A and IL-17F are related homodimeric proteins of the IL-17 family produced by Th17 cells. In this study, we show that mouse Th17 cells also produce an IL-17F/A heterodimeric protein. Whereas naive CD4+ T cells differentiating toward the Th17 cell lineage expressed IL-17F/A in higher amounts than IL-17A/A homodimer and in lower amounts than IL-17F/F homodimer, differentiated Th17 cells expressed IL-17F/A in higher amounts than either homodimer. In vitro, IL-17F/A was more potent than IL-17F/F and less potent than IL-17A/A in regulating CXCL1 expression. Neutralization of IL-17F/A with an IL-17A-specific Ab, and not with an IL-17F-specific Ab, reduced the majority of IL-17F/A-induced CXCL1 expression. To study these cytokines in vivo, we established a Th17 cell adoptive transfer model characterized by increased neutrophilia in the airways. An IL-17A-specific Ab completely prevented Th17 cell-induced neutrophilia and CXCL5 expression, whereas Abs specific for IL-17F or IL-22, a cytokine also produced by Th17 cells, had no effects. Direct administration of mouse IL-17A/A or IL-17F/A, and not IL-17F/F or IL-22, into the airways significantly increased neutrophil and chemokine expression. Taken together, our data elucidate the regulation of IL-17F/A heterodimer expression by Th17 cells and demonstrate an in vivo function for this cytokine in airway neutrophilia.

Th17 cytokines and their emerging roles in inflammation and autoimmunity

Summary: T‐helper 17 (Th17) cells are a new lineage of CD4+ T cells that are characterized by their production of interleukin‐17A (IL‐17A). Recent studies show that these cells can also express IL‐17F, IL‐22, and IL‐21. IL‐17A and IL‐17F can form a heterodimeric cytokine, which mediates biological activities, at least in part, through shared receptors with IL‐17A and IL‐17F homodimers. The cytokines made by Th17 cells represent three distinct gene families, highlighting the unique biology of these cells. Accumulating data support a role for Th17 cells and these cytokines in inflammatory processes and in animal models of autoimmunity or inflammation. Emerging data in clinical trials support our understanding of the importance of Th17 cells in inflammatory disease. Future clinical studies will allow us to evaluate the role of each cytokine independently in contributing to human diseases with immune‐mediated pathologies and to design optimal cytokine‐targeted therapies for these diseases.

IL-21 Activates Both Innate and Adaptive Immunity to Generate Potent Antitumor Responses that Require Perforin but Are Independent of IFN-γ

IL-21 is a key factor in the transition between innate and adaptive immune responses. We have used the cytokine gene therapy approach to study the antitumor responses mediated by IL-21 in the B16F1 melanoma and MethA fibrosarcoma tumor models in mice. Retrovirally transduced tumor cells secreting biologically functional IL-21 have growth patterns in vitro similar to that of control green fluorescent protein-transduced cells, but are completely rejected in vivo. We show that IL-21 activates NK and CD8+ T cells in vivo, thus mediating complete rejection of poorly immunogenic tumors. Rejection of IL-21-secreting tumors requires the presence of cognate IL-21R and does not depend on CD4+ T cell help. Interestingly, perforin, but not IFN-γ or other major Th1 and Th2 cytokines (IL-12, IL-4, or IL-10), is required for the IL-21-mediated antitumor response. Moreover, IL-21 results in 50% protection and 70% cure of nonimmunogenic tumors when given before and after tumor challenge, respectively, in C57BL/6 mice. We conclude that IL-21 immunotherapy warrants clinical evaluation as a potential treatment for cancer.

IL-22 Induces an Acute-Phase Response

IL-22 is made by a unique set of innate and adaptive immune cells, including the recently identified noncytolytic NK, lymphoid tissue-inducer, Th17, and Th22 cells. The direct effects of IL-22 are restricted to nonhematopoietic cells, its receptor expressed on the surface of only epithelial cells and some fibroblasts in various organs, including parenchymal tissue of the gut, lung, skin, and liver. Despite this cellular restriction on IL-22 activity, we demonstrate that IL-22 induces effects on systemic biochemical, cellular, and physiological parameters. By utilizing adenoviral-mediated delivery of IL-22 and systemic administration of IL-22 protein, we observed that IL-22 modulates factors involved in coagulation, including fibrinogen levels and platelet numbers, and cellular constituents of blood, such as neutrophil and RBC counts. Furthermore, we observed that IL-22 induces thymic atrophy, body weight loss, and renal proximal tubule metabolic activity. These cellular and physiological parameters are indicative of a systemic inflammatory state. We observed that IL-22 induces biochemical changes in the liver including induction of fibrinogen, CXCL1, and serum amyloid A that likely contribute to the reported cellular and physiological effects of IL-22. Based on these findings, we propose that downstream of its expression and impact in local tissue inflammation, circulating IL-22 can further induce changes in systemic physiology that is indicative of an acute-phase response.

Mitoxantrone, Etoposide, and Cyclosporine Therapy in Pediatric Patients With Recurrent or Refractory Acute Myeloid Leukemia

PURPOSE To determine the remission rate and toxicity of mitoxantrone, etoposide, and cyclosporine (MEC) therapy, multidrug resistance-1 (MDR1) status, and steady-state cyclosporine (CSA) levels in children with relapsed and/or refractory acute myeloid leukemia. PATIENTS AND METHODS MEC therapy consisted of mitoxantrone 6 mg/m(2)/d for 5 days, etoposide 60 mg/m(2)/d for 5 days, and CSA 10 mg/kg for 2 hours followed by 30 mg/kg/d as a continuous infusion for 98 hours. Because of pharmacokinetic interactions, drug doses were decreased to 60% of those found to be effective without coadministration of CSA. MDR1 expression was evaluated by reverse transcriptase polymerase chain reaction, flow cytometry, and the ability of CSA at 2.5 micromol/L to increase intracellular accumulation of (3)H-daunomycin in blasts from bone marrow specimens. RESULTS The remission rate was 35% (n = 23 of 66). Overall, 35% of patients (n = 23) achieved complete remission (CR), 12% of patients (n = 8) achieved partial remission, and 9% of patients (n = 6) died of infection. Exposure to CSA levels of greater than 2,400 ng/mL was achieved in 95% of patients (n = 56 of 59). Toxicities included infection, cardiotoxicity, myelosuppression, stomatitis, and reversible increases in serum creatinine and bilirubin. In most who had relapsed while receiving therapy or whose induction therapy had failed, response was not significantly different for MDR1-positive and MDR1-negative patients. CONCLUSION Serum levels of CSA capable of reversing multidrug resistance are achievable in children with acceptable toxicity. The CR rate of 35% achieved in this study is comparable to previously reported results using standard doses of mitoxantrone and etoposide. The use of CSA may have improved the response rate for the MDR1-positive patients so that it was not different from that for the MDR1-negative patients.

IL-22R, IL-10R2, and IL-22BP binding sites are topologically juxtaposed on adjacent and overlapping surfaces of IL-22.

Interleukin (IL) 22 is a type II cytokine that is produced by immune cells and acts on nonimmune cells to regulate local tissue inflammation. As a product of the recently identified T helper 17 lineage of CD4(+) effector lymphocytes, IL-22 plays a critical role in mucosal immunity as well as in dysregulated inflammation observed in autoimmune diseases. We used comprehensive mutagenesis combined with mammalian cell expression, ELISA cell-based, and structural methods to evaluate how IL-22 interacts with its cell surface receptor, IL-22R/IL-10R2, and with secreted IL-22 binding protein. This study identifies those amino acid side chains of IL-22 that are individually important for optimal binding to IL-22R, considerably expands the definition of IL-22 surface required for binding to IL-10R2, and demonstrates how IL-22 binding protein prevents IL-22R from binding to IL-22. The IL-22R and IL-10R2 binding sites are juxtaposed on adjacent IL-22 surfaces contributed mostly by helices A, D, and F and loop AB. Our results also provide a model for how IL-19, IL-20, IL-24, and IL-26 which are other IL-10-like cytokines, interact with their respective cell surface receptors.

Gene therapy with B7.1 and GM-CSF vaccines in a murine AML model.

Purpose Characterization of B7.1 and GM-CSF vaccines on the induction of anti-tumor immunity in a murine AML model. Materials and Methods Primary AML cells were retrovirally transduced with the murine costimulatory molecule B7.1, a natural ligand for the T-cell receptors CD28 and CTLA-4, or the cytokine GM-CSF. Mice were vaccinated with irradiated AML cells expressing B7.1 or GM-CSF before or after inoculation of wild type AML cells. Results Intravenous injection of irradiated B7.1 or GM-CSF expressing AML cells can provide long lasting systemic immunity against a subsequent challenge of wild type AML cells. Vaccination with irradiated B7.1 or GM-CSF expressing AML cells results in rejection of established leukemia when the vaccination occurs in the early stages of the disease. However, when the vaccines are administered >2 weeks after leukemic inoculation, only mice which receive the GM-CSF vaccine are cured of leukemia. Conclusions: These results suggest that tumor burden and vaccine efficiency are most likely to be the limiting factors in the curative potential of tumor vaccines. Novel approaches such as this experiment could provide improved therapeutic outcomes in patients with AML and other cancers.

Induction of Tumor Regression by Administration of B7-Ig Fusion Proteins: Mediation by Type 2 CD8+ T Cells and Dependence on IL-4 Production

CD28 signals contribute to either type 1 or type 2 T cell differentiation. Here, we show that administration of B7.2-Ig fusion proteins to tumor-bearing mice induces tumor regression by promoting the differentiation of antitumor type 2 CD8+ effector T cells along with IL-4 production. B7.2-Ig-mediated regression was not induced in IL-4−/− and STAT6−/− mice. However, it was elicited in IFN-γ−/− and STAT4−/− mice. By contrast, IL-12-induced tumor regression occurred in IL-4−/− and STAT6−/− mice, but not in IFN-γ−/− and STAT4−/− mice. Moreover, B7.2-Ig treatment was effective in a tumor model not responsive to IL-12. B7.2-Ig administration elicited elevated levels of IL-4 production. Tumor regression was predominantly mediated by CD8+ T cells, although the induction of these effector cells required CD4+ T cells. Tumor regression induced by CD8+ T cells alone was inhibited by neutralizing the IL-4 produced during B7.2-Ig treatment. Thus, these results indicate that stimulation in vivo of CD28 with B7.2-Ig in tumor-bearing mice results in enhanced induction of antitumor type 2 CD8+ T cells (Tc2) leading to Tc2-mediated tumor regression.