William L. Trepicchio

Interleukin-11 therapy selectively downregulates type I cytokine proinflammatory pathways in psoriasis lesions

Psoriasis is a chronic inflammatory skin disease in which epidermal hyperplasia results from skin infiltration by type I T lymphocytes and release of associated cytokines. A multifunctional cytokine, rhIL-11, modulates macrophage and type I T-lymphocyte function in cell culture and shows anti-inflammatory activity in animal models. We are testing subcutaneous delivery of rhIL-11 to patients with psoriasis in a phase 1 open-label dose-escalation clinical trial. Tissue was obtained from lesional and uninvolved skin before and during treatment with rhIL-11 and was examined by histology/immunohistochemistry and quantitative RT-PCR. Expression of over 35 genes was examined in all patients, and multiple genetic markers of psoriasis were identified. Expression of numerous proinflammatory genes was elevated in psoriatic tissue compared with nonlesional skin. Seven of 12 patients responded well to rhIL-11 treatment. Amelioration of disease by rhIL-11, as shown by reduced keratinocyte proliferation and cutaneous inflammation, was associated with decreased expression of products of disease-related genes, including K16, iNOS, IFN-gamma, IL-8, IL-12, TNF-alpha, IL-1beta, and CD8, and with increased expression of endogenous IL-11. We believe that this is the first study in humans to indicate that type I cytokines can be selectively suppressed by an exogenous immune-modifying therapy. The study highlights the utility of pharmacogenomic monitoring to track patient responsiveness and to elucidate anti-inflammatory mechanisms.

Molecular classification of psoriasis disease-associated genes through pharmacogenomic expression profiling

Psoriasis is recognized as the most common T cell-mediated inflammatory disease in humans. Genetic linkage to as many as six distinct disease loci has been established but the molecular etiology and genetics remain unknown. To begin to identify psoriasis disease-related genes and construct in vivo pathways of the inflammatory process, a genome-wide expression screen of multiple psoriasis patients was undertaken. A comprehensive list of 159 genes that define psoriasis in molecular terms was generated; numerous genes in this set mapped to six different disease-associated loci. To further interpret the functional role of this gene set in the disease process, a longitudinal pharmacogenomic study was initiated to understand how expression levels of these transcripts are altered following patient treatment with therapeutic agents that antagonize calcineurin or NF-κB pathways. Transcript levels for a subset of these 159 genes changed significantly in those patients who responded to therapy and many of the changes preceded clinical improvement. The disease-related gene map provides new insights into the pathogenesis of psoriasis, wound healing and cellular-immune reactions occurring in human skin as well as other T cell-mediated autoimmune diseases. In addition, it provides a set of candidate genes that may serve as novel therapeutic intervention points as well as surrogate and predictive markers of treatment outcome.

Hematopoietic, immunomodulatory and epithelial effects of interleukin-11

Interleukin 11 (IL-11) is a pleiotropic cytokine with biological activities on many different cell types. Recombinant human IL-11 (rhIL-11) is produced by recombinant DNA technology in Escherichia coli. Both in vitro and in vivo, rhIL-11 has shown effects on multiple hematopoietic cell types. Its predominant in vivo hematopoietic activity is the stimulation of peripheral platelet counts in both normal and myelosuppressed animals. This activity is mediated through effects on both early and late progenitor cells to stimulate megakaryocyte differentiation and maturation. rhIL-11 has been approved for the treatment of chemotherapy-induced thrombocytopenia. The hematopoietic effects of rhIL-11 are most likely direct effects on progenitor cells and megakaryocytes in combination with other cytokines or growth factors. rhIL-11 also induces secretion of acute phase proteins (ferritin, haptoglobin, C-reactive protein, and fibrinogen) from the liver. The induction of heme oxidase and inhibition of several P450 oxidases have been reported from in vitro studies. In vivo, rhIL-11 treatment decreases sodium excretion by the kidney by an unknown mechanism and induces hemodilution. rhIL-11 also exhibits anti-inflammatory effects in a variety of animal models of acute and chronic inflammation, including inflammatory bowel disease, inflammatory skin disease, autoimmune joint disease, and various infection-endotoxemia syndromes. rhIL-11 has trophic effects on non-transformed intestinal epithelium under conditions of mucosal damage. The mechanism of the anti-inflammatory activity of rhIL-11 has been extensively studied. rhIL-11 directly affects macrophage and T cell effector function. rhIL-11 inhibits tumor necrosis factor-alpha (TNFα), interleukin 1β (IL-1β), interleukin 12 (IL-12), interleukin 6 (IL-6), and nitric oxide (NO) production from activated macrophages in vitro. The inhibition of cytokine production was associated with inhibition of nuclear translocation of the transcription factor, nuclear factor kappa B (NF-κB). The block to NF-κB nuclear translocation correlates with the ability of rhIL-11 to maintain or enhance production of the inhibitors of NF-κB, IκB-α and IκB-β. In addition to effects on macrophages, rhIL-11 also reduces CD4+ T cell production of Th1 cytokines, such as IFNγ induced by IL-12, while enhancing Th2 cytokine production. rhIL-11 also blocks IFNγ production in vivo. The molecular effects of rhIL-11 have also been studied in a clinical trial. Molecular analysis of skin biopsies of patients with psoriasis before and during rhIL-11 treatment demonstrates a decrease in mRNA levels of TNFα, IFNγ and iNOS. These activities suggest that in addition to its thrombopoietic clinical use, rhIL-11 may also be valuable in the treatment of inflammatory diseases. The clinical utility of the anti-inflammatory properties of rhIL-11 is being investigated in patients with Crohn’s disease, psoriasis and rheumatoid arthritis. These diseases are believed to be initiated and maintained by activated CD4+ Th1 cells in conjunction with activated macrophages.

Interleukin-11 modulates Th1/Th2 cytokine production from activated CD4+ T cells.

Recombinant human interleukin-11 (rHuIL-11) is a pleiotropic cytokine with effects on multiple cell types. rHuIL-11 reduces activated macrophage activity and downregulates production of proinflammatory mediators, such as tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO). In vitro and in vivo, rHuIL-11 inhibits production of key immunostimulatory cytokines, including IL-12 and interferon-gamma (IFN-gamma). rHuIL-11 has recently demonstrated immunomodulatory activity to downregulate IFN-gamma production, increase IL-4 production, and reduce inflammatory tissue injury in a human psoriasis clinical trial. The cellular mechanisms of these effects are not fully elucidated. We demonstrate here that expression of gp130 and IL-11 receptor (IL-11R) alpha mRNA, components of the IL-11R complex, are detected in human and murine CD4(+) and CD8(+) lymphocytes, suggesting that rHuIL-11 can directly interact with T cells. In a cell culture model of murine T cell differentiation, rHuIL-11 acts to inhibit IL-2 production as well as IL-12-induced IFN-gamma production and enhances IL-4 and IL-10 production. rHuIL-11 had no effect on T cell proliferation. The ability of rHuIL-11 to modulate cytokine production from activated CD4(+) T cells provides a mechanism through which rHuIL-11 may ameliorate such inflammatory diseases as psoriasis.

Protective Effect of rhIL-11 in a Murine Model of Acetaminophen-Induced Hepatotoxicity

Acetaminophen intoxication results in hepatotoxicity associated with increased serum concentrations of hepatocellular leakage enzymes such as aspartate aminotransferase, lactate dehydrogenase, and alanine aminotransferase, centrilobular degeneration and necrosis, and activation of Kupffer cells. Recombinant human Interleukin-11 (rhIL-11) downregulates the production of proinfl ammatory mediators from activated macrophages and has direct effects on hepatocyte gene expression. Based on these biological activities of rhIL-11, the effect of pretreatment with rhIL-11 in a murine model of acetaminophen-induce d hepatotoxicity was examined. Administration of 500 ug/kg acetaminophen to B6C3F1 mice resulted in progressive hepatotoxicity as demonstrated by elevated serum concentration s of hepatocellular leakage enzymes and TNFα and histopathology. Pretreatment with 250 or 500 ug/kg of subcutaneously administered rhIL-11 2 hours before acetaminophen administration reduced serum concentrations of hepatocellular leakage enzyme s and TNFα by 40—50%. This was associated with a statistically significant decreas e in mean severity score for centrilobular hemorrhag e and necrosis from grade 3 to grade 2 for rhIL-11-treated animals compared to vehicle. These results indicate that treatment with rhIL-11 has a protective effect in a model of acetaminophen-induce d liver damage.

The therapeutic utility of Interleukin-11 in the treatment of inflammatory disease

Interleukin-11 (IL-11) is a pleiotropic cytokine that exhibits anti-inflammatory and mucosal protective effects in a variety of animal models of acute and chronic inflammation, such as mucositis, inflammatory bowel disease and autoimmune joint disease. This reduction in inflammation and epithelial damage is mediated in part through effects of recombinant human (rh) IL-11 on macrophage effector function and epithelial cell growth. In vitro studies indicate that rhIL-11 inhibits tumour necrosis factor (TNF)-alpha, IL-1beta, IL-12, IL-6, and nitric oxide production from activated macrophages. Analysis of the effects of rhIL-11 on transcription factors that activate pro-inflammatory cytokines demonstrate that the level of induced nuclear factor kappa B (NF-kappaB) binding activity in the nucleus of rhIL-11-treated peritoneal macrophages is significantly reduced. Studies of normal intestinal epithelial cells indicate that rhIL-11 reduces the rate of cellular proliferation. Analysis of cell-cycle progression demonstrates that growth inhibition of epithelial cells by rhIL-11 correlates with delayed entry into S phase and suppression of pRB phosphorylation. IL-11 also protects intestinal crypt stem cells from radiation- or chemotherapy-induced insults. Such immunomodulatory and epithelial activities may contribute to the protective effects of this cytokine and support the clinical utility of rhIL-11 in the treatment of mucositis, as well as a variety of chronic inflammatory diseases, such as Crohns disease and rheumatoid arthritis.

A single administration of recombinant human interleukin-12 is associated with increased expression levels of interferon-gamma and signal transducer and activator of transcription in healthy subjects

The objectives of this study were to assess the safety and tolerability of single doses of 1, 4, and 8 μg of recombinant human interleukin‐12 (rhIL‐12) administered subcutaneously to healthy subjects. The pharmacokinetics, pharmacodynamics, and pharmacogenomics of rhIL‐12 were evaluated. Recombinant human IL‐12 was well tolerated in these healthy male and female subjects. The most frequently reported adverse events were flu‐like symptoms, which exhibited a dose‐response relationship. Pharmacokinetic analysis suggested that serum IL‐12 levels increased with dose. Analysis of serum levels indicated that interferon‐γ increased with the dose of rhIL‐12, whereas IL‐6 levels showed no changes with rhIL‐12 treatment. The messenger ribonucleic acid expression of signal transducer and activator of transcription was significantly increased 24 hours after the administration of rhIL‐12 for all dose groups versus placebo, and results indicated that the magnitude of increase may be dose dependent. This study suggests that interferon‐γ and signal transducer and activator of transcription are biomarkers of rhIL‐12 activity.