Soluble Decoy Receptor Research Articles

Antiangiogenic Therapies in Epithelial Ovarian Cancer

Angiogenesis is a critical component of tumor development and proliferation, and increased angiogenesis has been associated with a worse clinical outcome in a number of solid tumors, including ovarian cancer. Therefore, agents that target the angiogenic process are of considerable interest in the treatment of ovarian cancer. Studies evaluating the efficacy of antiangiogenic agents in ovarian cancer are reported. Antiangiogenic agents examined include vascular endothelial growth factor (VEGF) pathway inhibitors, including monoclonal antibodies, tyrosine kinase inhibitors (TKIs), and a soluble receptor decoy, as well as inhibitors of other angiogenic factors and vascular disrupting agents. The VEGF inhibitor bevacizumab has been shown to have efficacy in ovarian cancer in phase II trials and a progression-free survival advantage in one phase III trial. TKIs block the VEGF receptors and secondary angiogenic pathways and have shown activity in phase I and II trials. Alternative angiogenesis inhibitors include EphA2 inhibitors and a selective angiopoietin 1/2-neutralizing peptibody. Another strategy is to destroy the existing tumor vasculature, and a number of vascular disrupting agents are being studied in preclinical and phase I trials. Antiangiogenic agents have a unique side effect profile, likely due to inhibition of normal physiologic angiogenesis. Phase II and early phase III trials have demonstrated that antiangiogenic therapies have significant activity in ovarian cancer. The results of phase III trials in the front-line and recurrent settings will determine the extent of clinical benefit of antiangiogenic therapies in combination with chemotherapy. Antiangiogenic agents have a distinct side effect profile, and further studies are necessary to evaluate how to minimize the incidence of these events and to identify women most likely to benefit from these therapies.

Angiogenesis-Related Cytokines, RANKL, and Osteoprotegerin in Multiple Myeloma Patients in relation to Clinical Features and Response to Treatment

An essential cytokine system for the osteoclast biology in multiple myeloma (MM) consists of the receptor of activator of NF-κB ligand (RANKL), its receptor (RANK), and the soluble decoy receptor, osteoprotegerin (OPG). Myeloma cells cause imbalance in OPG/RANKL interactions. We measured serum levels of OPG, soluble (s) RANKL, sRANKL/OPG ratio, markers of disease activity [LDH, CRP, interleukin-6 (IL-6), β2-microglobulin (B2M)], and angiogenic factors [hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF)], in 54 newly diagnosed MM patients and in 25 of them in plateau phase. All the above values were higher in MM patients compared to controls and decreased in plateau phase. sRANKL and RANKL/OPG were higher with advancing disease stage and skeletal grade. Significant correlations were found among RANKL and RANKL/OPG with HGF, LDH, VEGF, IL-6, and B2M. In conclusion, RANKL and OPG play significant roles in MM pathophysiology, as regulators of bone turnover and mediators of angiogenesis.

VEGF receptor signaling links inflammation and tumorigenesis in colitis-associated cancer

Whereas the inhibition of vascular endothelial growth factor (VEGF) has shown promising results in sporadic colon cancer, the role of VEGF signaling in colitis-associated cancer (CAC) has not been addressed. We found that, unlike sporadic colorectal cancer and control patients, patients with CAC show activated VEGFR2 on intestinal epithelial cells (IECs). We then explored the function of VEGFR2 in a murine model of colitis-associated colon cancer characterized by increased VEGFR2 expression. Epithelial cells in tumor tissue expressed VEGFR2 and responded to VEGF stimulation with augmented VEGFR2-mediated proliferation. Blockade of VEGF function via soluble decoy receptors suppressed tumor development, inhibited tumor angiogenesis, and blocked tumor cell proliferation. Functional studies revealed that chronic inflammation leads to an up-regulation of VEGFR2 on IECs. Studies in conditional STAT3 mutant mice showed that VEGFR signaling requires STAT3 to promote epithelial cell proliferation and tumor growth in vivo. Thus, VEGFR-signaling acts as a direct growth factor for tumor cells in CAC, providing a molecular link between inflammation and the development of colon cancer.

Comparison of Platelet-Activating Potential of Bevacizumab+VEGF and Aflibercept+VEGF Complexes

AbstractAbstract 3200Anti-angiogenic therapy with the monoclonal antibody (mAb) bevacizumab (bev; Genentech) is associated with venous and arterial thrombosis in cancer patients. We recently showed that bev immune complexes (ICs) activate platelets in vitro and cause thrombotic thrombocytopenia in mice transgenic for the human platelet IgG receptor, FcγRIIa (FCGR2A mice; Jax Labs). Previous studies conducted in other labs using wild type mice injected with bevacizumab failed to identify thrombotic side effects; however, wild type mice do not have platelet FcγRIIa IgG receptors. Platelets from human donors or from FCGR2A mice are directly activated when FcγRIIa is clustered by the Fc domains of mAbs that have been assembled into higher order ICs by multivalent antigens, such as was shown for bev (Meyer, et al, J Thromb Haemost, 2009;7:171) and for anti-CD154 ICs (Robles-Carrillo, et al, J Immunol, 2010;185:1577). Aflibercept (aflib, Regeneron/sanofi-aventis, also known as VEGF Trap) is a soluble decoy receptor protein that binds all isoforms of VEGF-A including VEGF165 and inhibits VEGF165 activity. Aflib also binds the related angiogenic molecule PlGF, and like bev has anti-angiogenic activity. The aflib molecule contains the Fc domain of the IgG isotype antibody; however, although VEGF165 efficiently clusters bev into higher order ICs, VEGF165 does not cluster aflib into higher order complexes (Rudge JS, et al, PNAS, 2007;104:18363). This suggests that aflib+VEGF165 should not be able to trigger platelet FcγRIIa activation (because FcγRIIa requires clustering for activation). We therefore hypothesized that aflib+VEGF165 complexes would fail to activate platelet FcγRIIa in vitro or in FCGR2A transgenic mice. For in vitro studies, we chose the washed platelet aggregation method as a measure of IC-induced platelet activation. To assess platelet activation in vivo, we measured IC-induced reductions in the number of circulating platelets (thrombocytopenia) following intravenous injection of ICs into FCGR2A mice. Aflib+VEGF165 or bev+VEGF165 complexes were pre-formed at room temperature in the presence or absence of heparin. (Heparin can enhance bev IC-induced platelet activation. Many cancer patients are routinely exposed to heparin to maintain patency of chemotherapy ports.) In 5 donors, bev+VEGF165 complexes (500nM) rapidly induced full aggregation of human platelets in an FcγRIIa-dependent manner, whereas 500nM aflib+VEGF165 failed to induce aggregation under all conditions tested. Within 5 minutes following intravenous injection of bev+VEGF165, FCGR2A mice (n=10) experienced signs of distress consistent with thrombotic shock, including rapid shallow breathing, hunched posture, and decreased or dysfunctional locomotor activity. Animals receiving aflib complexes (n=10) did not exhibit these symptoms. Platelet counts 10 minutes following IC injection revealed thrombocytopenia in animals receiving bev+VEGF165 but not aflib+VEGF165 complexes. Specifically, animals injected with PBS (vehicle; n=5) had baseline platelet counts of (1173±179;mean±SD), whereas animals receiving bev+VEGF165 had mean platelet counts of 331±217 (with heparin; P < 0.001) and 725±303 (without heparin; P < 0.003). Animals receiving aflib+VEGF165 had mean platelet counts of 966±168 (with heparin) and 878±250 (without heparin), which were not statistically different from baseline animal counts. These results indicate that aflibercept lacks that platelet activating potential observed with bevacizumab, and also suggest that anti-angiogenic therapy with aflibercept may lack the capacity to induce the platelet associated thrombotic side effects observed in patients receiving bevacizumab. Disclosures:Meyer:Regeneron Pharmaceuticals, Inc.: Florida Hospital received research funding from Regeneron Pharmaceuticals in support of some of this work.

Structure-based development of a receptor activator of nuclear factor-kappaB ligand (RANKL) inhibitor peptide and molecular basis for osteopetrosis.

The receptor activator of nuclear factor-κB (RANK) and its ligand RANKL, which belong to the tumor necrosis factor (TNF) receptor-ligand family, mediate osteoclastogenesis. The crystal structure of the RANKL ectodomain (eRANKL) in complex with the RANK ectodomain (eRANK) combined with biochemical assays of RANK mutants indicated that three RANK loops (Loop1, Loop2, and Loop3) bind to the interface of a trimeric eRANKL. Loop3 is particularly notable in that it is structurally distinctive from other TNF-family receptors and forms extensive contacts with RANKL. The disulfide bond (C125-C127) at the tip of Loop3 is important for determining the unique topology of Loop3, and docking E126 close to RANKL, which was supported by the inability of C127A or E126A mutants of RANK to bind to RANKL. Inhibitory activity of RANK mutants, which contain loops of osteoprotegerin (OPG), a soluble decoy receptor to RANKL, confirmed that OPG shares the similar binding mode with RANK and OPG. Loop3 plays a key role in RANKL binding. Peptide inhibitors designed to mimic Loop3 blocked the RANKL-induced differentiation of osteoclast precursors, suggesting that they could be developed as therapeutic agents for the treatment of osteoporosis and bone-related diseases. Furthermore, some of the RANK mutations associated with autosomal recessive osteopetrosis (ARO) resulted in reduced RANKL-binding activity and failure to induce osteoclastogenesis. These results, together with structural interpretation of eRANK-eRANKL interaction, provided molecular understanding for pathogenesis of ARO.

Cytokines in bone diseases. Osteoprotegerin and juvenile Paget's disease

Juvenile Paget's disease (JPD) is a rare autosomal-recessive hereditary disease and is typically diagnosed in infants or young children. JPD causes bone pain, bone expansion and deformity, and severity generally increases during adolescence. Recently, differentiation and maturation of osteoclasts, controlled by RANK, which is expressed in osteoclast precursors and its ligand, RANKL, which is expressed in osteoblasts or marrow stromal cells, have been clarified. Recent studies elucidated that JPD was caused by mutation of TNFRSF11B, which encodes osteoprotegerin, a soluble decoy receptor of RANKL. We summarize the outline and etiology of JPD.

PKA-induced Receptor Activator of NF-κB Ligand (RANKL) Expression in Vascular Cells Mediates Osteoclastogenesis but Not Matrix Calcification

Vascular calcification is a predictor of cardiovascular mortality and is prevalent in patients with atherosclerosis and chronic renal disease. It resembles skeletal osteogenesis, and many bone cells as well as bone-related factors involved in both formation and resorption have been localized in calcified arteries. Previously, we showed that aortic medial cells undergo osteoblastic differentiation and matrix calcification both spontaneously and in response to PKA agonists. The PKA signaling pathway is also involved in regulating bone resorption in skeletal tissue by stimulating osteoblast-production of osteoclast regulating cytokines, including receptor-activator of nuclear κB ligand (RANKL) and interleukins. Therefore, we investigated whether PKA activators regulate osteoclastogenesis in aortic smooth muscle cells (SMC). Treatment of murine SMC with the PKA agonist forskolin stimulated RANKL expression at both mRNA and protein levels. Forskolin also stimulated expression of interleukin-6 but not osteoprotegerin (OPG), an inhibitor of RANKL. Consistent with these results, osteoclastic differentiation was induced when monocytic preosteoclasts (RAW264.7) were cocultured with forskolin-treated aortic SMC. Oxidized phospholipids also slightly induced RANKL expression in T lymphocytes, another potential source of RANKL in the vasculature. Because previous studies have shown that RANKL treatment alone induces matrix calcification of valvular and vascular cells, we next examined whether RANKL mediates forskolin-induced matrix calcification by aortic SMC. RANKL inhibition with OPG had little or no effect on osteoblastic differentiation and matrix calcification of aortic SMC. These findings suggest that, as in skeletal tissues, PKA activation induces bone resorptive factors in the vasculature and that aortic SMC calcification specifically induced by PKA, is not mediated by RANKL.

Shedding of RANKL by Tumor-Associated MT1-MMP Activates Src-Dependent Prostate Cancer Cell Migration

Membrane type 1 matrix metalloproteinase (MT1-MMP) plays an essential role in protease-mediated extracellular matrix (ECM) degradation, but it also functions as a sheddase releasing non-ECM substrates such as receptor activator of NF-kappaB ligand (RANKL), an osteoclastogenic factor typically confined to the surface of osteoblasts. We previously found high expression of MT1-MMP in skeletal metastasis of prostate cancer patients, in a pattern similar to RANKL expression. We also showed that overexpression of MT1-MMP in prostate cancer cells increases tumor growth and osteolysis in an intratibial mouse model of bone metastasis, and that soluble factor(s) shed by tumor-derived MT1-MMP enhance osteoclast differentiation in a RANKL-dependent manner. Recent evidence indicates that the cognate receptor for RANKL, RANK, is expressed in prostate cancer cells, suggesting the presence of an autocrine pathway. In this study, we show that MT1-MMP-expressing LNCaP prostate cancer cells display enhanced migration. Moreover, conditioned medium from LNCaP cells expressing both RANKL and MT1-MMP stimulates the migration of MT1-MMP-deficient C42b prostate cancer cells. This enhanced chemotaxis can be abrogated by osteoprotegerin (soluble decoy receptor of RANKL), MIK-G2 (a selective inhibitor for MT1-MMP), and PP2 (a Src inhibitor). These findings indicate that tumor-derived MT1-MMP enhances tumor cell migration through initiation of an autocrine loop requiring ectodomain shedding of membrane-bound RANKL in prostate cancer cells, and that Src is a key downstream mediator of RANKL-induced migration of prostate cancer cells.

Advances in understanding and treating cardiac cachexia: Highlights from the 5th Cachexia Conference

Cardiac cachexia as a terminal stage of chronic heart failure carries a devastating prognosis. This article focuses on novel insights into the pathophysiology and on new treatment approaches to cardiac cachexia. Drugs that have been used in preclinical and clinical studies that may also confer beneficial effects in cardiac cachexia include but are not limited to the type 4 melanocortin receptor antagonist SNT 207979, the appetite promoting synthetic ghrelin SUN11031, the soluble myostatin decoy receptor ActRIIB-Fc, the fast skeletal muscle troponin activating substance CK-2017357, the anti-catabolic/anabolic transforming agent MT-102, the anti-inflammatory agent celecoxib, and testosterone supplementation.

Requirement for BAFF and APRIL during B Cell Development in GALT

The effects of B cell-activating factor belonging to the TNF family (BAFF) on B cell maturation and survival in the mouse are relatively well understood. In contrast, little is known about the role of BAFF in B cell development in other mammals, such as rabbits, that use GALT to develop and maintain the B cell compartment. We examined the expression and requirement of BAFF and a proliferation-inducing ligand (APRIL) during peripheral B cell development in young rabbits. By neutralizing BAFF and APRIL in neonates with a soluble decoy receptor, transmembrane activator calcium modulator and cyclophilin ligand interactor-Fc, we found a marked reduction in the number of peripheral B cells, but found no change in the bone marrow (BM) compartment. In the appendix, the size and number of proliferating B cell follicles were greatly reduced, demonstrating that although BAFF/APRIL is dispensable for B cell development in BM, it is required for B cell development in GALT. We found that all rabbit B cells expressed BAFF receptor 3, but did not bind rBAFF, suggesting that the BAFF-binding receptors (BBRs) are bound by endogenous soluble BAFF. Further, we found that B cells themselves express BAFF, suggesting that the soluble BAFF bound to BBRs may be endogenously produced and stimulate B cells in an autocrine fashion. Additionally, we propose that this chronic occupancy of BBRs on B cells may provide a tonic and/or survival signal for the maintenance of peripheral B cells in adults after B lymphopoiesis is arrested in BM.

Porphyromonas gingivalis Induces RANKL in T-cells

Porphyromonas gingivalis is an oral pathogen highly implicated in chronic periodontitis, a disease characterized by inflammatory destruction of the tooth-supporting alveolar bone and eventually, tooth loss. T-cell innate immune responses are actively involved in this pathological process. Receptor activator of NF-κB Ligand (RANKL) is a cytokine that stimulates bone resorption, while its soluble decoy receptor osteoprotegerin (OPG) blocks its action. This study aimed to investigate in Jurkat T-cells the effects of P. gingivalis on the RANKL-OPG system and the major inflammatory mediator of bone resorption prostaglandin E(2) (PGE(2)). P. gingivalis caused concentration-dependent up-regulation of RANKL gene expression and protein production, assessed by quantitative PCR and ELISA, respectively. PGE(2) production was also enhanced. However, OPG was not detected. In conclusion, P. gingivalis induces RANKL and PGE(2) in T-cells, potentially favoring bone resorption. These T-cell responses to P. gingivalis may contribute to the pathogenesis of inflammatory alveolar bone destruction occurring in chronic periodontitis.

Epithelial-derived IL-33 and its receptor ST2 are dysregulated in ulcerative colitis and in experimental Th1/Th2 driven enteritis

IL-33 is a novel member of the IL-1 family and ligand for the IL-1 receptor-related protein, ST2. Recent evidence suggests that the IL-33/ST2 axis plays a critical role in several autoimmune and inflammatory disorders; however, its role in inflammatory bowel disease (IBD) has not been clearly defined. We characterized IL-33 and ST2 expression and modulation after conventional anti-TNF therapy in Crohn's disease and ulcerative colitis (UC) patients and investigated the role of IL-33 in SAMP1/YitFc (SAMP) mice, a mixed Th1/Th2 model of IBD. Our results showed a specific increase of mucosal IL-33 in active UC, localized primarily to intestinal epithelial cells (IEC) and colonic inflammatory infiltrates. Importantly, increased expression of full-length IL-33, representing the most bioactive form, was detected in UC epithelium, whereas elevated levels of cleaved IL-33 were present in IBD serum. ST2 isoforms were differentially modulated in UC epithelium, and sST2, a soluble decoy receptor with anti-inflammatory properties, was also elevated in IBD serum. Infliximab (anti-TNF) treatment of UC decreased circulating IL-33 and increased sST2, whereas stimulation of HT-29 IEC confirmed IL-33 and sST2 regulation by TNF. Similarly, IL-33 significantly increased and correlated with disease severity, and potently induced IL-5, IL-6, and IL-17 from mucosal immune cells in SAMP mice. Taken together, the IL-33/ST2 system plays an important role in IBD and experimental colitis, is modulated by anti-TNF therapy, and may represent a specific biomarker for active UC.

The formation of osteoclasts in multiple myeloma bone disease patients involves the secretion of soluble decoy receptor 3

Soluble decoy receptor 3 (DcR3), a member of the tumor necrosis factor receptor superfamily, has recently been reported to increase osteoclast (OC) differentiation. Its impact on the skeleton was reinforced by a study on DcR3 transgenic mice showing a decreased bone mass through the elevation of OC number, providing some initial evidence of DcR3 involvement in bone diseases. In this study we show that malignant plasma cells and T lymphocytes from myeloma patients directly produce DcR3, and this molecule supports the elevated formation of OCs in both peripheral blood and bone marrow from the patients. We also show that serum DcR3 levels in myeloma patients are significantly higher compared to controls.

Inhibition of RANKL increases the anti-tumor effect of the EGFR inhibitor panitumumab in a murine model of bone metastasis

Bone metastases cause severe skeletal complications and are associated with osteoclast-mediated bone destruction. RANKL is essential for osteoclast formation, function, and survival, and is the primary effector of tumor-induced osteoclastogenesis and osteolysis. RANKL inhibition by its soluble decoy receptor osteoprotegerin (OPG) prevents tumor-induced osteolysis and decreases skeletal tumor burden. Because osteoclast-mediated bone resorption releases growth factors from the bone matrix, the host bone microenvironment induces a vicious cycle of bone destruction and tumor proliferation and survival. A prediction of this vicious cycle hypothesis is that targeting the host bone microenvironment by osteoclast inhibition would reduce tumor growth and survival and may enhance the anti-tumor effects of targeted therapies. The epidermal growth factor receptor (EGFR) pathway regulates critical processes such as cell growth and survival, and anti-EGFR therapies can cause tumor cell arrest and apoptosis. We evaluated whether reduction of osteolysis by RANKL inhibition could enhance the anti-tumor effects of an anti-EGFR antibody (panitumumab) in a novel murine model of human A431 epidermoid carcinoma bone metastasis. Skeletal tumor progression was assessed longitudinally by bioluminescence imaging. RANKL inhibition by OPG-Fc treatment resulted in a reduction in tumor progression in bony sites. OPG-Fc treatment also caused a dose-dependent reduction in tumor-induced osteolysis, supporting the essential role of RANKL in this process. In combination, RANKL inhibition increased the anti-tumor efficacy of an anti-EGFR antibody, and completely blocked tumor-induced bone breakdown, demonstrating that addition of the indirect anti-tumor effect of RANKL inhibition increases the anti-tumor efficacy of panitumumab, a targeted anti-EGFR antibody.

Activin A promotes multiple myeloma-induced osteolysis and is a promising target for myeloma bone disease

Understanding the pathogenesis of cancer-related bone disease is crucial to the discovery of new therapies. Here we identify activin A, a TGF-beta family member, as a therapeutically amenable target exploited by multiple myeloma (MM) to alter its microenvironmental niche favoring osteolysis. Increased bone marrow plasma activin A levels were found in MM patients with osteolytic disease. MM cell engagement of marrow stromal cells enhanced activin A secretion via adhesion-mediated JNK activation. Activin A, in turn, inhibited osteoblast differentiation via SMAD2-dependent distal-less homeobox-5 down-regulation. Targeting activin A by a soluble decoy receptor reversed osteoblast inhibition, ameliorated MM bone disease, and inhibited tumor growth in an in vivo humanized MM model, setting the stage for testing in human clinical trials.

Creation of a lysine-deficient LIGHT mutant with the capacity for site-specific PEGylation and low affinity for a decoy receptor

The cytokine LIGHT is a promising candidate for cancer therapy. However, the therapeutic effect of LIGHT as a systemic anticancer agent is currently insufficient because of its instability and its binding to nonfunctional soluble decoy receptor 3 (DcR3), which is overexpressed in various tumors. Modification of proteins with polyethylene glycol (PEGylation) can improve their in vivo stability, but PEGylation may occur randomly at all lysine residues and the NH 2-terminus; therefore, PEGylated proteins are generally heterogeneous and have decreased bioactivity. In this study, we attempted to create a lysine-deficient LIGHT mutant that could be PEGylated site-specifically and would have lower affinity for DcR3. We prepared phage libraries expressing LIGHT mutants in which all the lysine residues were replaced with other amino acids. A lysine-deficient LIGHT mutant [mLIGHT-Lys(−)] was isolated by panning against lymphotoxin β receptor (LTβR). mLIGHT-Lys(−) could be site-specifically PEGylated at its NH 2-terminus, yielding molecular uniformity and in vitro bioactivity equal to that of non-PEGylated, wild-type LIGHT. Furthermore, mLIGHT-Lys(−) was not trapped by the nonfunctional DcR3, despite binding to its functional receptors. These results suggest that mLIGHT-Lys(−) might be a useful candidate for cancer therapy.

Prolonged effect of BAFF on chicken B cell development revealed by RCAS retroviral gene transfer in vivo

Chickens provide an important model for developmental biology as well as phylogenetic studies of the immune system. In many species cytokines are important regulators of immune functions and recently we identified the chicken homologue of BAFF (B cell activating factor of the TNF family), shown to be an essential survival factor for B cells in mammals. Characterisation of BAFF function in the phylogenetically distant chicken requires in vivo studies, but despite considerable progress to date no efficient transgene or knockout technology for chickens is available. Thus the retroviral vector system RCAS (replication-competent ASLV long terminal repeat (LTR) with a Splice acceptor) was used to generate chickens stably expressing chBAFF or a soluble chBAFF neutralizing decoy receptor. Expression of RCAS based proteins was demonstrated in ovo and for the complete observation period of 2 month after hatch. In contrast to injections with recombinant proteins the prolonged RCAS based cytokine expression allowed assessment of BAFF function not only in newly hatched chicks but also in more mature birds. We demonstrate that RCAS based secreted products were biologically active, with chBAFF overexpression leading to a drastic and continuous increase in B cell numbers and neutralisation of chBAFF by expression of a soluble decoy receptor leading to severely reduced B cell numbers in transduced birds. This increase in B cell numbers was matched by increased immunoglobulin levels. Using RCASBP transduced birds we provide evidence that the RCAS system provides a fast and highly efficient gene transfer system for analysis of secreted protein function in the chicken and we show that in the chicken BAFF is necessary for B cell survival but in contrast to mammals, not only mature B cells but also immature B cells in the bursa of Fabricius are maintained by BAFF.

Markers of bone remodeling and skeletal morbidity in patients with solid tumors metastatic to the skeleton receiving the biphosphonate zoledronic acid

The molecular triad, which includes the receptor activator of nuclear factor kappa-B ligand (RANKL), its receptor RANK, and the endogenous soluble RANKL decoy receptor osteoprotegerin (OPG), has emerged as an important determinant of bone metabolism. We aimed to evaluate the effect of treatment with the biphosphonate zoledronic acid (ZA) on biochemical markers of bone remodeling and to detect possible correlations of markerlevel changes with skeletal morbidity and clinical outcomes in patients with solid tumors and osseous metastases. The following serum markers were measured at the onset of skeletal metastases and after 6 months of treatment with ZA (4 mg intravenously monthly) in 70 patients with breast (n = 30), lung (n = 18), or prostate (n = 22) cancer: RANKL, OPG, C-terminal cross-linking telopeptide of type I collagen (CTX), tartrate-resistant acid phosphatase isoform 5b (TRACP-5b), bone-specific alkaline phosphatase (bALP), and osteopontin (OPN). Logistic regression models were applied to assess the correlation between marker-level changes and skeletal related events (SRE, primary endpoint), recurrence or progression, and death. Within a median follow-up of 32 months, 34 patients (48.6%) presented with at least 1 SRE and 48 patients (68.6%) relapsed. The RANKL/OPG ratio was upregulated in patients with breast and lung cancer, and it tended to decline after treatment with ZA, whereas prostate cancer patients presented with profound elevation of OPG only that persisted after treatment. CTX levels were significantly reduced after treatment in the whole study population (P = 0.003). None of the markers was able to predict skeletal morbidity or clinical outcomes independently of well-established prognostic clinical parameters.

EFFECTS OF THE TREM 1 PATHWAY MODULATION DURING HEMORRHAGIC SHOCK IN RATS

The triggering receptor expressed on myeloid cells (TREM) 1, a receptor expressed on the surface of neutrophils and monocytes/macrophages, synergizes with the Toll-like receptors in amplifying the inflammatory response mediated by microbial components. Because the pathogenesis of severe blood loss-induced excessive inflammation and multiple organ failure implies leukocyte activation and bacterial translocation, we hypothesized that the TREM-1 pathway modulation would prove beneficial in this setting. Wistar rats were subjected to a 1-h period of hemorrhagic shock and then reperfused with shed blood and ringer lactate for 1 h. At the time of reperfusion, animals were administered with LP17 (a synthetic soluble TREM-1 decoy receptor), a control peptide, or a vehicle (isotonic sodium chloride solution). Plasma concentration of TNF-alpha, IL-6, and soluble TREM-1 were measured by enzyme-linked immunosorbent assay. Lung permeability was assessed by the weight-dry ratio and fluorescein isothiocyanate-labeled albumin lung-blood ratio. Organ dysfunction was appreciated by measuring plasma aspartate aminotransferase and urea concentrations. Bacterial translocation was estimated by blood, mesenteric lymph nodes, and spleens culture. Hemorrhagic shock associated with cardiovascular collapse, lactic acidosis, systemic inflammatory response, and organ dysfunction that was partly prevented by LP17 administration. Hemorrhagic shock induced a marked increase in lung permeability that was also prevented by TREM-1 modulation. Finally, LP17 improved survival. Thus, the early modulation of the TREM-1 pathway by means of a synthetic peptide may be useful during severe hemorrhagic shock in rats in preventing organ dysfunction and improving survival.

The Effect of Bortezomib On Bone Metabolism of Patients with Multiple Myeloma: a Korean Multicenter Prospective Study.

Abstract 4871 BACKGROUNDBortezomib is increasingly being employed as an important treatment for relapsed/refractory patients with multiple myeloma (MM) who have received at least one prior therapy. Osteolytic bone disease is a major problem in the management of MM. Myeloma bone disease is a result of excessive osteoclast activation and impaired osteoblast function. METHODSThis study aimed to evaluate the effect of bortezomib on bone remodeling in the patients with MM who received one prior therapy. Total 104 patients were enrolled and serum samples were collected from 81 patients at baseline and after 4 cycles of bortezomib treatment, respectively. An enzyme linked immunosorbent assay (ELISA) was used for the detection of the following serum markers; (1) the osteoblastic markers including serum osteocalcin (OC) and bone-specific alkaline phosphatase (BAP), (2) dickkopf-1 (DKK-1), an osteoblastic inhibitor and (3) a main inducer of osteoclast activity, receptor activator nuclear factor κB ligand (RANKL) and its soluble decoy receptor, osteoprotegerin (OPG). RESULTSThere was no correlation between bone disease status and the concentrations of the bone markers; patients with myeloma bone disease at baseline did not have increased values of DKK-1, RANKL and RANKL/OPG ratio as well as decreased levels of OC and BAP when compared with those without osteolytic bone lesions. To our surprise, there was also a strong tendency towards a negative correlation with serum levels of OC (P=.009) and BAP (P=.049) and disease duration (≥ vs. < 6 months). Moreover, DKK-1 concentrations were positively associated with disease duration (P=.047). The objective response rate (≥ partial response) after four cycles of therapy was 58%. Bortezomib administration significantly reduced serum DKK-1 (P=.011) and sRANKL/OPG ratio (P=.034) after 4 cycles irrespective of treatment response. OPG levels were significantly different according to the type of previous therapy (stem cell transplantation vs. alkylator-based, P=.029) CONCLUSIONSThis study suggests that bortezomib can inhibit the down-regulation of osteoblastic response and bone resorption by reducing circulating levels of DKK-1 and sRANKL/OPG ratio. Serum concentrations of the bone-remodeling markers can be different according to the disease duration rather than the status of bone disease. Bortezomib treatment early after diagnosis may benefit myeloma patients with bone disease considering that the change of bone markers was bound up with disease duration. DisclosuresMin:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Lee:Janssen Korea: Research Funding. Suh:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Yoon:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Kang:Janssen Korea: Research Funding. Choi:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Kwak:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding. Joo:Janssen Korea: Research Funding. Mun:Janssen Korea: Research Funding. Jo:Janssen Korea: Research Funding. Park:Janssen Korea: Research Funding. Park:Janssen Korea: Research Funding. Kim:Janssen Korea: Research Funding.