Oncostatin M Regulation of Inflammatory Responses By

Regulation of Inflammatory Responses by Oncostatin M Philip M. Wallace,1,2* John F. MacMaster,† Katherine A. Rouleau,† T. Joseph Brown,* James K. Loy,† Karen L. Donaldson,3* and Alan F. Wahl3* Oncostatin M (OM) is a pleiotropic cytokine produced late in the activation cycle of T cells and macrophages. In vitro it shares properties with related proteins of the IL-6 family of cytokines; however, its in vivo properties and physiological function are as yet ill defined. We show that administration of OM inhibited bacterial LPS-induced production of TNF-a and lethality in a dose-dependent manner. Consistent with these findings, OM potently suppressed inflammation and tissue destruction in murine models of rheumatoid arthritis and multiple sclerosis. T cell function and Ab production were not impaired by OM treatment. Taken together these data indicate the activities of this cytokine in vivo are antiinflammatory without concordant immunosuppression. The Journal of Immunology, 1999, 162: 5547–5555. he normal development of an inflammatory response must from the inflammatory effector phase back to homeostasis also are be rapidly followed by the engagement of a feedback sys- being evaluated for their clinical potential as drugs. The cytokines T tem to minimize adventitious tissue damage and regulate IL-10 and IL-11 both appear to accelerate this process and their the eventual return to homeostasis. This system involves a multi- administration have proven effective in resolving several animal tude of regulators including cytokines, adhesion molecules, pro- models of chronic inflammatory disease (10). teases, corticosteroids, and subsequent regulators of each of these Oncostatin M (OM)4 is a pleiotropic cytokine that is produced agents. A normal response to infection or other insult is a self- by activated T cells and macrophages and has shown in vitro prop- limiting process that by way of temporal expression of both reg- erties that would be expected to influence the course of inflamma- ulators, and effector molecules, causes the resolution of the initi- tory responses (11, 12). The protein is structurally and functionally ating event. The failure to resolve the causative insult or to redress related to IL-6, leukemia inhibitory factor (LIF), and IL-11, pro- the balance of pro- and antiinflammatory agents results in tissue teins that also influence immune and inflammatory function (13). injury and destruction that characterize the pathology of various Despite each protein signaling via a family of related receptors and chronic inflammatory diseases (1, 2). sharing various common properties, each is endowed with a unique The exact participation of each cytokine in the inflammatory disease process is poorly understood in part due to their complex array of biological functions (13). Numerous activities have been interplay. However, the ability of a variety of cytokine and cyto- ascribed to OM in vitro, including the differentiation of kine agonists to alter the severity or course of various inflamma- megakaryocytes, inhibition of tumor cell growth, induction of neu- tory diseases is an impressive testament to the clinical value of rotrophic peptides, regulation of cholesterol metabolism, and ef- cytokines as a target for therapeutic intervention (2, 3). Such data fects on bone-derived cells (7, 14, 15). Recently a collective pic- has been accrued using animal models of disease, transgenic ani- ture of OM has emerged that strongly suggests a natural role of the mals and, more recently, clinical trials of cytokine inhibitors (4, 5). cytokine in the wound healing process and attenuation of the in- A variety of approaches are currently being studied to alter cyto- flammatory response. We have previously found that OM can kine function to bring about the regulation of aberrant inflamma- modulate the expression of IL-6, an important regulator of various tory responses (6). Inhibitors of proinflammatory cytokines, most aspects of the host defense system (16). OM has been shown to notably TNF-a inhibitors, have been successful in moderating un- regulate the expression by human cells of acute phase proteins and toward inflammatory responses (2). Abs to TNF-a and soluble protease inhibitors that have been implicated in modulating cyto- receptors are currently in clinical trials against a variety of diseases kine function and limiting tissue damage at sites of inflammation. including rheumatoid arthritis, multiple sclerosis, and Crohn’s dis- Recently many of these in vitro effects have been found to occur ease (7–9). Their efficacy has helped establish a set of common in rodents and nonhuman primates following OM administration effectors in these apparently disparate diseases. Alternatively, the (17). Here we have extended these in vivo findings to further un- cytokines whose normal physiological role is to usher a response derstand the role of OM in regulating cytokine networks following inflammatory stimuli. We have also examined the effects of OM *Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, WA 98121; and treatment in two murine models of disease in which common † Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, NJ 08543 proinflammatory cytokines have been previously shown to play Received for publication November 19, 1998. Accepted for publication February key roles. 16, 1999. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Address correspondence and reprint requests to Dr. Philip M. Wallace, Xcyte Ther- apies, 2203 Airport Way South, Suite 300, Seattle, WA 98134. E-mail address: 4 Abbreviations used in this paper: OM, oncostatin M; LIF, leukemia inhibitory fac- [email protected] tor; a1-Pi, a1-proteinase inhibitor; EAE, experimental autoimmune encephalomyeli- tis; KLH, keyhole limpet hemocyanin; MMP, matrix metalloproteinase; PLP, prote- 2 Current address: Xcyte Therapies, 2203 Airport Way South, Suite 300, Seattle, WA olipid protein; SAA, serum amyloid A; TIMP-1, tissue inhibitor of 98134. metalloproteinase-1; BMS-PRI, Bristol-Myers Squibb Pharmaceutical Research 3 Current address: Seattle Genetics, 22215, 26th Avenue SE, Bothell, WA 98021. Institute. Copyright © 1999 by The American Association of Immunologists 0022-1767/99/$02.00 5548 OM ATTENUATES INFLAMMATION Materials and Methods decalcified in HCl, processed by routine methods, and embedded into par- m Animals affin. The specimens were sectioned at 4–6 m, stained with hematoxylin and eosin, and examined by light microscopy. Sections were graded with- Studies used female mice (;8 wk old) that were held in quarantine for out prior knowledge of the treatment group. Tibiotarsal (hock) joints were 2 wk before admission to any study, during which time serological graded as to the severity of inflammation, pannus formation, cartilage dam- examination was performed. BALB/c and C57BL/6 mice were obtained age, and osseuos changes. Each parameter was examined separately and from Taconic (Germantown, NY), and B10.S-H2(S)SgMcdJ mice were graded as follows: grade 0, unremarkable; grade 1, minimal change; grade obtained from The Jackson Laboratory (Bar Harbor, ME). Animals 2, mild; grade 3, moderate; and grade 4, severe. The inflammation score were housed according to the American Association for the Accredita- was derived from evaluation of soft tissue inflammation, synovitis, and tion of Laboratory Animal Care and institutional guidelines. Experi- angiogenesis. Pannus formation was defined as hypertrophic synovial tis- ments shown are representative of at least three independent studies. sue composed of intraarticular inflammatory exudate accompanied by sy- Statistical analyses were performed using a Wilcoxon rank test (Primer novial cell hyperplasia. Cartilage destruction and loss of matrix were eval- for Biostatistics, McGraw-Hill, NY). uated on the articular surfaces of the distal tibia, the talus, the calcaneus, and the tarsal bones to yield the cartilage damage score. The depth of Reagents erosion of the subchondral bone and the amount of periosteal exocytosis in the distal tibia, the talus, the calcaneus, and the tarsal bones were evaluated Recombinant human OM was expressed in Chinese hamster ovary cells to yield the osseous changes score. The above parameters were then eval- and purified as described (18). OM was administered via various routes of uated as to the percent of tissue involved in the disease process: 1, 0–25%; injection in PBS. Escherichia coli LPS (#L3012) and IFA were purchased a 2, 26–50%; 3, 51–75%; 4, 76–100%. The severity and extent of involve- from Sigma (St. Louis, MO). TNF- and IL-6 ELISA was obtained from ment were then combined to yield the global arthritis score for each joint Endogen (Woburn, MA), and no cross reactivity was found with OM (data (maximum possible score, 32). not shown). Anti-collagen II monoclonal hybridomas were purchased from Chondrex (Redmond, WA). Abs were produced from hybridoma superna- Peptide synthesis tant and purified by protein A Sepharose chromatography. Mycobacterium tuberculosis was purchased from Difco (Detroit, MI), and pertussis toxin The peptide 139–151 from proteolipid peptide (PLP) was assembled on a was obtained from List Biological Laboratories (Campbell, CA). SRBC Gilson multiple peptide synthesizer (Middleton, WI) using F-moc amino were purchased from PML Microbiologicals (Tualatin, OR), and keyhole acids. The peptide resin was treated

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