Inhibition Of Osteoclastic Bone Resorption Research Articles

Einfluss von Bisphosphonaten auf die entzündliche Gelenkdestruktion bei rheumatoider Arthritis und in Arthritismodellen

In context with the increasing evidence for the significance of osteoclastic bone resorption mediated by the RANKL-RANK-OPG system in the pathogenesis of postmenopausal, glucocorticoid-induced and inflammation-associated osteoporosis as well as in joint destruction in rheumatoid arthritis (RA), there is an increasing interest in the combination of anti-inflammatory and anti-osteoclastic therapies in RA. Because of their suppressive effects on both osteoclastic bone resorption and inflammation due to inhibitory effects on the secretion of pro-inflammatory cytokines and matrix metalloproteinases, bisphosphonates are implicated to be a useful adjuvant therapy in RA. Furthermore, these substances are relatively cheap and have only few side effects. The effects of various bisphosphonates on inflammation, joint destruction and periarticular bone resorption were investigated in different animal models of RA and also in some small studies in RA patients. In various animal models, a suppressive effect of different non-amino- and aminobisphosphonates on periarticular bone resorption was found. But the results with respect to the inhibition of joint and cartilage destruction and inflammation are inconsistent. Newly developed, highly potent aminobisphosphonates such as zoledronate have been shown to inhibit articular bone erosion not only in animal models but also in RA. The assessment of data from animal models is difficult because various bisphosphonates were administered in different doses in heterogeneous animal models with a partly different pathogenesis. Furthermore, the effects of bisphophonates on bone and joint destruction were investigated using different methods or combinations of these methods. Data from animal models and from RA patients have shown that the doses of bisphosphonates necessary for the suppression of inflammation and joint destruction are significantly higher than those needed for the inhibition of osteoclastic bone resorption. It is not clear whether or not these relatively high bisphosphonate doses may result in an oversuppression of bone turnover. The effects of various bisphosphonates on joint destruction and inflammation in RA and animal models of RA are reviewed systematically and discussed in this contribution.

Recent developments in the management of postmenopausal osteoporosis with bisphosphonates: enhanced efficacy by enhanced compliance

Bisphosphonates are the current mainstay of treatment for postmenopausal osteoporosis. Although daily oral dosing is effective, it is associated with poor compliance, partly because of the pre and postdose fasting and posture requirements. This negatively impacts treatment outcomes, leading to a reduced clinical benefit. Improved, yet still suboptimal adherence has been noticed with less frequent bisphosphonate dosing e.g. once-weekly and once-monthly oral regimens. The recently approved quarterly intravenous (i.v.) injection regimen of ibandronate and yearly i.v. infusion of zoledronic acid are attractive options in the management of postmenopausal osteoporosis. These regimens may assure quarterly and year long compliance.

Serum osteoprotegerin concentrations are decreased in women with the polycystic ovary syndrome

Osteoprotegerin (OPG), an inhibitor of osteoclastic bone resorption, has a variety of functions including anti-inflammatory effects and a possible cardiovascular protective role. Both low-grade chronic inflammation and cardiovascular risk are increased in women with the polycystic ovary syndrome (PCOS). We aimed to study serum OPG concentrations in PCOS patients. Case-control study including 40 PCOS patients matched with 40 non-hyperandrogenic women for age and body mass index. Basal serum sampling and standard oral glucose tolerance test, and measurement of serum OPG concentrations by commercial ELISA. Serum OPG concentrations were lower in women with PCOS compared with those of controls (304+/-120 vs 363+/-105 pg/ml respectively; F=7.641, P=0.007) independently of obesity. No differences were observed in serum receptor activator of nuclear factor-kappaB ligand (RANKL) levels and in the RANKL/OPG molar ratio. A multivariate linear regression model (R(2)=0.208, F=6.579, P=0.001) showed that PCOS (beta=-0.281, P=0.008), obesity (beta=-0.245, P=0.022) and age (beta=0.296, P=0.006) were predictive of serum OPG concentrations. Serum OPG concentrations are reduced in PCOS patients independently of obesity. Considering the anti-inflammatory effects of OPG, its reduced serum concentrations might contribute to the proinflammatory state and cardiovascular risk of PCOS patients.

Bisphosphonates in the management of postmenopausal osteoporosis – optimizing efficacy in clinical practice

Nitrogen-containing bisphosphonates are potent inhibitors of osteoclastic bone resorption. With their individually proven efficacy to significantly reduce the incidence of vertebral and/or non-vertebral fractures and with their overall beneficial safety profile, alendronate, ibandronate, risedronate, and zoledronate are considered today a treatment of first choice in postmenopausal osteoporosis. However, treatment effects in an individual patient and cost-effectiveness in public health perspective are vitally dependent on the long-term patient adherence as well as on compliance and persistence. As compliance and persistence with daily oral bisphosphonates are shown to be suboptimal in many patients, leading to an increased fracture incidence in non-compliant patients, there is a need to improve overall adherence for bisphosphonate treatment in order to achieve maximum treatment effects. One option is to extend dosing intervals to weekly (alendronate, risedronate) or monthly (ibandronate) oral regimens. Less frequent oral regimens are generally preferred by majority of patients. Another alternative is intravenous, instead of oral application (ibandronate, zoledronate). Treatment acceptance could be further improved by IV bisphosphonates with their benefit of only quarterly, or even once-yearly, application. Treatment decisions should be based on anti-fracture efficacy data first. In addition, to ensure best possible patient adherence and maximum treatment benefits, physicians should consider individual patient conditions affecting compliance and persistence as well as patient preferences.

Extracellular calcium increases bisphosphonate-induced growth inhibition of breast cancer cells

IntroductionBisphosphonates have become standard therapy for the treatment of skeletal complications related to breast cancer. Although their therapeutic effects mainly result from an inhibition of osteoclastic bone resorption, in vitro data indicate that they also act directly on breast cancer cells, inhibiting proliferation and inducing apoptosis.MethodsThe present study examined the effects of calcium (from 0.6 to 2.0 mmol/l) on the antitumour activity of the bisphosphonate ibandronate (1 to 1,000 nmol/l) on MDA-MB-231 and MCF-7 breast cancer cells. Cell culture densities were determined using crystal violet staining assay. Apoptotic cell death was assessed by annexin V-phycoerythrin and 7-amino-actinomycin double staining.ResultsAt low calcium concentration, 30 μmol/l ibandronate had no effect on MDA-MB-231 cells growth and only slightly inhibited MCF-7 cells growth. Higher calcium levels significantly increased growth inhibition as well as cell apoptosis induced by ibandronate. We observed similar effects with zoledronic acid. Of note, enhancement of ibandronate-induced growth inhibition was also observed in other breast cancer cell lines (T-47D, ZR-75, Hs-578T and BT-549 cells). The growth inhibitory effect of ibandronate in the presence of high concentrations of calcium was partly suppressed by the calcium chelator EGTA (ethylene glycol tetra-acetic acid). In addition, in the presence of calcium at high concentrations, cells accumulated more [14C]ibandronate than at low calcium concentrations. We obtained further evidence of enhancement of cellular ibandronate accumulation by calcium by demonstrating that high calcium levels increased the inhibition of protein prenylation induced by the bisphosphonate.ConclusionAltogether, our data suggest that extracellular calcium, probably through its binding to ibandronate, markedly increased its cellular accumulation and its inhibitory activity on breast tumour cells. Thus, calcium released during the process of tumour-induced osteolysis might enhance the antitumour effects of bisphosphonates and contribute to their therapeutic efficacy.

Bisphosphonates and osteomyelitis of the jaw: a pathogenic puzzle

The maxillary and mandibular bones undergo high-turnover remodeling to maintain mechanical competence. Common dental or periodontal diseases can increase local bone turnover. Bisphosphonates (BPs) accumulate almost exclusively in skeletal sites that have active bone remodeling. The maxillary and mandibular bones are preferential sites for accumulation of BPs, which become buried under new layers of bone and remain biologically inactive for a long time. Surgical odontostomatological procedures create open bony wounds that heal quickly and without infection, as a result of activation of osteoclasts and subsequently osteoblasts. Once BPs are removed from the bone via activation of osteoclasts after a tooth extraction or a periodontal procedure, they induce osteoclast apoptosis. This inhibition of osteoclast bone resorption impairs bone wound healing because of decreased production of cytokines derived from the bone matrix, and the bone is exposed to the risk of osteomyelitis and necrosis. The pathogenic relationship between BPs and osteonecrosis of the jaw is unclear, but there is evidence to indicate an association between high-dose BP treatment and exposure to dental infections or oral surgical procedures. A better knowledge of the interactions between BPs and jaw and maxillary bone biology will improve clinical and therapeutic approaches.

Diphyllin, a Novel and Naturally Potent V-ATPase Inhibitor, Abrogates Acidification of the Osteoclastic Resorption Lacunae and Bone Resorption

Dissolution of the inorganic phase of bone by the osteoclasts mediated by V-ATPase and ClC-7 is a prerequisite for bone resorption. Inhibitors of osteoclastic V-ATPase or ClC-7 are novel approaches for inhibition of osteoclastic bone resorption. By testing natural compounds in acidification assays, diphyllin was identified. We characterized diphyllin with respect to the pharmacological effects on osteoclasts. Osteoclastic acidification of the resorption lacuna and bone resorption requires activity of both V-ATPase and the chloride channel ClC-7. Inhibition of these processes represents a novel approach for treatment of bone metabolic disorders. We identified diphyllin, a novel inhibitor of V-ATPase, and characterized this natural compound with respect to activity in human osteoclasts. Diphyllin was tested in the acid influx assay and V-ATPase assay using bovine chromaffin granules. Human osteoclasts were generated from CD14+ monocytes cultured with macrophage-colony stimulating factor (M-CSF) and RANKL. The effect of diphyllin on lysosomal acidification in human osteoclasts was studied using acridine orange. The effect of diphyllin on bone resorption by osteoclasts was measured as release of C-terminal cross-linked telopeptide of type I collagen (CTX-I) and calcium into the supernatants and by scoring pit area. Osteoclast number, TRACP activity, and cell viability were measured. Furthermore, the effect of diphyllin on bone nodule formation was tested using the mouse osteoblast cell line MC3T3-E1. In the acid influx assay, diphyllin potently inhibited the acid influx (IC50 = 0.6 nM). We found that diphyllin inhibited V-ATPase with an IC50 value of 17 nM, compared with 4 nM for bafilomycin A1. Moreover, diphyllin dose-dependently inhibited lysosomal acidification in human osteoclasts. Furthermore, we found that diphyllin inhibited human osteoclastic bone resorption measured by CTX-I (IC50 = 14 nM), calcium release, and pit area, despite increasing TRACP activity, numbers of osteoclasts, and cell viability. Finally, diphyllin showed no effect on bone formation in vitro, whereas bafilomycin A1 was toxic. We identified a natural compound that potently inhibits V-ATPase and thereby lysosomal acidification in osteoclasts, which leads to abrogation of bone resorption. Because recent studies indicate that inhibition of the osteoclastic acidification leads to inhibition of resorption without inhibiting formation, we speculate that diphyllin is a potential novel treatment for bone disorders involving excessive resorption.

Bisphosphonate Therapy for Reduced Bone Mineral Density During Treatment of Acute Lymphoblastic Leukemia in Childhood and Adolescence

Osteopenia is a common consequence of the treatment of acute lymphoblastic leukemia (ALL) in children and adolescents, due predominantly to glucocorticosteroid therapy. The pathogenesis relates to an imbalance of resorption over formation of bone. Alendronate (Fosamax), an inhibitor of osteoclastic bone resorption, was administered for at least 6 months to 15 children with ALL during maintenance chemotherapy, after the diagnosis of osteopenia/osteoporosis by dual energy x-ray absorptiometry. The height velocity was also measured during the administration of alendronate and again 2 years later. Areal bone mineral density Z scores of the lumbar spine had a median value of -1.32 before administration of alendronate and a median gain of +0.64, with 14/15 children showing improvement. There was no adverse effect of alendronate on height velocity, and the drug was well tolerated with no short-term toxicity. This preliminary experience suggests a potential value in the use of alendronate for the treatment of osteopenia/osteoporosis in children with ALL and points to the need for a randomized controlled trial of this intervention.

Heparin enhances osteoclastic bone resorption by inhibiting osteoprotegerin activity

Heparin is a highly sulfated glycosaminoglycan and has been shown to activate osteoclastic bone resorption though how is not yet clear. Here we investigate the molecule involved in heparin-induced activation of osteoclasts using an in vitro osteoclast culture assay. The formation and activation of osteoclasts are induced by receptor activator of NFκB ligand (RANKL) on osteoblasts, and inhibited by osteoprotegerin (OPG), a decoy receptor of RANKL, which is secreted from osteoblasts. In a coculture of mouse bone marrow cells and osteoblasts treated with 1,25-dihydroxyvitamin D 3 and prostaglandin E 2 on dentin slices, the bone marrow cells differentiate into osteoclasts, and resorption pits are formed on the dentin slices. Addition of heparin, various glycosaminoglycans, and chemically modified heparins to the coculture reveals that heparin enhances the pit-forming activity of osteoclasts, and this effect of heparin on the activation of osteoclasts is dependent on its sugar chain structure. By contrast, mRNA expression levels of RANKL, RANK, and OPG in the coculture are not altered by heparin treatment. Furthermore, neither RANK nor RANKL binds to heparin, suggesting that heparin does not directly interact with these proteins. Instead, heparin specifically binds to OPG and prevents OPG-mediated inhibition of osteoclastic bone resorption in the coculture. Heparin treatment does not enhance osteoclastic bone resorption in a monoculture of osteoclasts derived from bone marrow cells, and in the coculture using osteoblasts from OPG-deficient mice. A 125I-OPG binding assay showed that OPG binds to osteoblasts and that this binding is inhibited by the addition of heparin, suggesting that OPG binds to RANKL on the osteoblast membrane and that heparin blocks this interaction. These results demonstrate that heparin enhances osteoclastic bone resorption by inhibiting OPG activity.

Inhibition of tooth movement by osteoprotegerin vs. pamidronate under conditions of constant orthodontic force

The undesired movement of anchor teeth, and relapse of previously moved teeth, are major clinical problems in orthodontics. Dental implants are increasingly used to preserve anchorage, but these are costly and require invasive surgical procedures. An alternative strategy for maintaining anchorage may be the use of biological inhibitors of osteoclastic bone resorption. In the present study, we investigated the relative efficacy of pamidronate vs. osteoprotegerin (OPG) in inhibiting bone resorption and tooth movement, using a new orthodontic model in mice in which maxillary molars are moved for prolonged periods by near-constant, clinically relevant forces. Osteoclast influx to compression sites was initiated on day 3, was maximal on day 4, and persisted until at least day 12 after force application. Tooth movement paralleled osteoclast numbers. Minimal osteoclast apoptosis was observed, suggesting that recruitment, rather than programmed cell death, is a critical regulatory mechanism under conditions of constant force. Osteoclasts were reduced at compression sites by both OPG (95%) and pamidronate (70%); tooth movement was more dramatically inhibited by OPG (77% vs. 34%). Our findings indicate that constant orthodontic force regulates the recruitment, activation, and viability of osteoclasts, and that OPG could have clinical utility in preventing undesired tooth movement.

Expression and Regulation of Osteoprotegerin in Adipose Tissue

PurposeOsteoprotegerin (OPG), a potent inhibitor of osteoclastic bone resorption, has a variety of biological functions that include anti-inflammatory effects. Adipocytes and osteoblasts share a common origin, and the formation of new blood vessels often precedes adipogenesis in developing adipose tissue microvasculature. We examined whether OPG is secreted from adipocytes, therefore contributing to the prevention of neovascularization and protecting the vessels from intimal inflammation and medial calcification.Materials and MethodsThe mRNA expression of OPG and receptor activator of NF-κB ligand (RANKL) was measured in differentiated 3T3L1 adipocytes and adipose tissues.ResultsOPG mRNA expression increased with the differentiation of 3T3L1 adipocytes, while RANKL expression was not significantly altered. OPG mRNA was expressed at higher levels in white adipose tissue than in brown adipose tissue and was most abundant in the epididymal portion. In differentiated 3T3L1 adipocytes, Rosiglitazone and insulin reduced the OPG/RANKL expression ratio in a dose- and time-dependent manner. In contrast, tumor necrosis factor-α (TNF-α) increased the expression of both OPG and RANKL in a time-dependent manner. The OPG/RANKL ratio was at a maximum two hours after TNF-α treatment and then returned to control levels. Furthermore, OPG was abundantly secreted into the media after transfection of OPG cDNA with Phi C31 integrase into 3T3L1 cells.ConclusionOur results indicate that OPG mRNA is expressed and regulated in the adipose tissue. Considering the role of OPG in obesity-associated inflammatory changes in adipose tissue and vessels, we speculate that OPG may have both a protective function against inflammation and anti-angiogenic effects on adipose tissue.

The effect of zoledronic acid incorporated in a poly(D,L‐lactide) implant coating on osteoblasts in vitro

Bisphosphonates such as zoledronic acid (ZOL) are used in diseases associated with osteoclast-mediated bone loss. However, their antiresorptive activity is partly due to their effect on osteoblasts. Local application might increase the therapeutical fence and their local efficiency and reduce systemic side effects. Aim of the study was to investigate the effect of ZOL on human osteoblasts like cells in vitro with special focus on the synthesis of factors mediating osteoclast differentiation (RANKL, OPG). ZOL was incorporated in an implant coating based on poly(D,L-lactide) (PDLLA) in different concentrations (10-150 microM). Control groups were treated with uncoated implants, PDLLA-coated implants, and ZOL pure substance in corresponding concentrations. After an experimental period of 144 h, primary human osteoblasts were stained with alamar blue and cell viability was measured. Procollagen I synthesis, osteoprotegerin (OPG) secretion, and soluble receptor activator of nuclear factor-kappaB ligand (sRANKL) were analyzed. Results showed that cell viability was not affected when treated with doses equivalent up to 100 microM ZOL-coated implants (ZOL-CI). Procollagen I synthesis was highest when treated with 50 microM ZOL-CI. OPG increased significantly in the 10 microM ZOL-CI group, whereas sRANKL decreased significantly with different concentrations of ZOL-CI. Higher concentrations or exposure to the pure substance showed a decrease in cell viability, collagen I, OPG, and sRANKL synthesis. In conclusion, exposure to specific concentrations of ZOL-CI showed a beneficial effect on osteoblast differentiation and protein synthesis without influencing their proliferation. Changes in sRANKL and OPG production may contribute to the inhibition of osteoclastic bone resorption. This local antiresorptive effect might be clinically useful in osseous implant integration and fracture healing.

Antitumor and Antimetastatic Activities of Docetaxel Are Enhanced by Genistein through Regulation of Osteoprotegerin/Receptor Activator of Nuclear Factor-κB (RANK)/RANK Ligand/MMP-9 Signaling in Prostate Cancer

Bone metastasis is very common in advanced prostate cancer. Docetaxel has been shown to improve survival in patients with metastatic prostate cancer. However, treatment with docetaxel is associated with a certain degree of toxicity. Genistein, derived from soybeans, has been found to inhibit cancer cell growth without toxicity. We have recently reported that genistein could potentiate the antitumor activity of chemotherapeutic agents both in vitro and in vivo. However, the molecular mechanism of this novel effect of genistein has not been fully elucidated. In this study, we found that genistein significantly potentiated the antitumor, anti-invasive, and antimetastatic activities of docetaxel both in culture and in severe combined immunodeficient (SCID)-human model of experimental prostate cancer bone metastasis. We further conducted microarray analysis, real-time reverse transcription-PCR, Western blot analysis, small interfering RNA and cDNA transfection, matrix metalloproteinase-9 (MMP-9) activity assay, and invasion assay. We found that the expression of osteoprotegerin (OPG) was induced by genistein and inhibited by docetaxel, whereas genistein significantly down-regulated the expression and secretion of receptor activator of nuclear factor-kappaB (RANK) ligand (RANKL) and inhibited osteoclast formation. Moreover, genistein down-regulated the expression and activity of MMP-9, which was induced by docetaxel treatment, and inhibited invasion of PC-3 cells. These results suggest that the observed potentiation of antitumor activity of docetaxel by genistein in the SCID-human model of experimental bone metastasis could be mediated by regulation of OPG/RANK/RANKL/MMP-9 signaling, resulting in the inhibition of osteoclastic bone resorption and prostate cancer bone metastasis. From these results, we conclude that genistein could be a promising nontoxic agent to improve the treatment outcome of metastatic prostate cancer with docetaxel.

Role of osteoprotegerin (OPG) in cancer

OPG (osteoprotegerin), a secreted member of the TNF (tumour necrosis factor) receptor superfamily, has a variety of biological functions which include the regulation of bone turnover. OPG is a potent inhibitor of osteoclastic bone resorption and has been investigated as a potential therapeutic for the treatment of both osteoporosis and tumour-induced bone disease. Indeed, in murine models of cancer-induced bone disease, inhibition of osteoclastic activity by OPG was also associated with a reduction in tumour burden. The discovery that OPG can bind to and inhibit the activity of TRAIL (TNF-related apoptosis-inducing ligand) triggered extensive research into the potential role of OPG in the regulation of tumour cell survival. A number of reports from studies using in vitro models have shown that OPG protects tumour cells from the effects of TRAIL, thereby possibly providing tumour cells that produce OPG with a survival advantage. However, the ability of OPG to act as a tumour cell survival factor remains to be verified using appropriate in vivo systems. A third area of interest has been the use of OPG as a prognostic marker in various cancer types, including myeloma, breast and prostate cancer. This review provides an overview of the role of OPG in cancer, both in cancer-induced bone disease and in tumour growth and survival.

Application of bisphosphonates for periodontitis

Bisphosphonates are known as highly effective inhibitors of osteoclastic bone resorption that selectively affect osteoclasts. They are world-wildly used for the treatment of metabolic bone diseases with excessive bone resorption such as osteoporosis and Paget's disease. Alveolar bone loss in periodontitis results from local inflammatory reactions by periodontopathic bacteria. It has been reported that bisphosphonates efficiently prevent experimentally induced alveolar bone resorption in animals with periodontitis, and that bisphosphonate therapy improves the outcome of periodontal treatment in human being. Administration of bisphosphonates may be an appropriate adjunctive treatment to preserve periodontal bone mass. Further studies are needed regarding topical drug delivery system, effective dose and frequency of administration, possible side effects, and which bisphosphonate is suitable for periodontitis.

644. A Dual-Action Armed Replicating Adenovirus for the Treatment of Bone Metastases of Breast Cancer

The majority of patients with advanced breast cancer develop osteolytic metastases in the skeleton, which lead to such complications as pain, fractures, hypercalcemia and a general decrease in the quality of life. Current therapies for bone metastases are not curative in nature. Thus, new treatments are needed for osteolytic bone metastasis of breast cancer. Conditionally replicating adenoviruses (CRADs) are a new class of anti-cancer agents that offer the potential ability to infect and lyse all the cells of a tumor. In clinical trials, replicating adenoviruses have exhibited safety but not the desired level of efficacy, demonstrating a need for improvement. One approach to enhance the efficacy of a CRAD is to arm it with a therapeutic transgene. Osteoprotegerin (OPG), the recently described inhibitor of osteoclastic bone resorption, represents a promising candidate with which to arm a replicating adenovirus designed to treat osteolytic bone metastasis. We hypothesize that a replicating adenovirus armed with a gene for OPG will be able to inhibit breast cancer bone metastasis and reduce the tumor burden in the bone through two actions: direct lysis of the tumor cells by replication of the virus and inhibition of bone resorption by the production of OPG.

Advances in the management of myeloma bone disease

Multiple myeloma is the second most common adult haematological malignancy. One of the major clinical features is the development of a unique osteolytic bone disease, characterised by progressive and devastating bone destruction, bone pain, pathological fractures and hypercalcaemia. Bisphosphonates, inhibitors of osteoclastic bone resorption, are the standard therapy for myeloma-induced bone disease. However, as our understanding of the molecular mechanisms involved in the development of myeloma bone disease increases, new molecular targets have been identified for the treatment of this devastating bone disease.

Effects of osteoprotegerin administration on osteoclast differentiation and trabecular bone structure in osteoprotegerin-deficient mice

Osteoprotegerin (OPG)-deficient mice exhibit severe bone loss including the destruction of growth plate cartilage. Using OPG-deficient mice, we attempted to clarify the differentiation and ultrastructure of osteoclasts located on the destroyed growth plate cartilage and trabecular bone matrix in long bones. In (-/-) homozygous OPG knockout mice, adjacent to the growth plate cartilage, the formation of bone trabeculae without a calcified cartilaginous core resulted in an irregular chondrocyte distribution in the growth plate cartilage. At the metaphyseal ossification center, TRAP-positive osteoclasts showed unusual localization on both type-II collagen-positive cartilage and type-I collagen-positive bone matrix. Osteoclasts located on cartilage matrix lacked a typical ruffled border structure, but formed resorption lacunae. During growth plate cartilage destruction, osteoclasts formed ruffled border structures on bone matrix deposited on the remaining cartilage surfaces. These findings suggest that, in OPG (-/-) mice, osteoclast structure differs, depending on the matrix of either cartilage or bone. Then, we examined the effects of OPG administration on the internal trabecular bone structure and osteoclast differentiation in OPG (-/-) mice. OPG administration to OPG (-/-) mice significantly inhibited trabecular bone loss and maintained the internal trabecular bone structure, but did not reduce the osteoclast number on bone trabeculae. For most osteoclasts, OPG administration caused disappearance or reduction of the ruffled border, but induced neither necrotic nor apoptotic damages. These results suggest that OPG administration is an effective means of maintaining the internal structure and volume of trabecular bone in metabolic bone diseases by inhibition of osteoclastic bone resorption.

Pamidronate as adjuvant treatment for prevention of bone metastasis in breast cancer

888 Background: Bone is the most common site of distant recurrence in breast cancer. The primary mechanism of metastatic bone disease is stimulation of osteoclastic bone resorption by factors produced by the cancer cells. Bisphosphonates are potent inhibitors of osteoclastic bone resorption with proven efficacy in reducing tumor-associated skeletal complications. Several studies have investigated the adjuvant, or preventive, use of these drugs in breast cancer. Laboratory experiments have shown that the development of bone metastases can be inhibited by Bisphosphonates. Pamidronate is an easy to use potent inhibitor of osteolysis, given in conjunction with standard anticancer therapies. We evaluated the efficacy of adjuvant pamidronate therapy for prevention of bone metastasis in women with breast cancer. Methods: We analyzed the data of 429 patients with primary operable stage I-III breast cancer that were treated at KNUH during the period January 1999-December 2002. All patients had radical local therapy and systemic therapy performed according to our guidelines at that time. 258 patients were received adjuvant pamidronate therapy(parmidronate 15mg iv every 4 weeks and oral parmidronate 100mg daily during adjuvant chemotherapy). 171 patients had no bisphosphonate therapy. The patients were perimenopausal women. The mean age of patients was 53 years and the median follow-up period was 3.5 years. The clinicopathological characteristics of the patients were well balanced. Results: The incidence of bone metastasis was 2.3% in the parmidronate group and 8.7% in control group. The incidence of other sites metastasis in pamidronate group was lower than control group but no statistically significant. Overall survival and disease-free survival rate were equal in the both groups. Bone metastasis free survival was significantly higher in the pamidronate group compared to the control group. Conclusions: In the parmidronate group, the incidence of bone metastases was significantly reduced and bone metastasis free survival was significantly higher. We conclude that adjuvant parmidronate therapy significantly reduced the development of bone metastases in perimenopausal women with primary breast cancer. No significant financial relationships to disclose.

Suppression by incadronate of invasion and growth of A-375 human melanoma in mandible in nude mice.

Frequent invasion of oral cancers into the neighboring jaw bones is a major cause of increased mortality. Moreover, majority of patients with these oral cancers suffer from functional inconveniences and esthetic disadvantages during clinical course and after surgical treatments. Understanding of the mechanisms underlying oral cancer invasion into jaw bones is, therefore, critical to the better management of oral cancer patients. To study the pathophysiology of mandibular invasion of oral cancers, we established an animal model by inoculating the A-375 human melanoma cells onto mandible through masseter muscle in nude mice. Histological study showed that A-375 tumor cells aggressively destroyed the mandibular cortical bones and invaded into the bone marrow cavities. Histochemical examination revealed that there were numerous tartrate-resistant acid phosphatase-positive osteoclasts on the residual bone surfaces facing A-375 tumors. To examine the role of osteoclasts in mandibular bone invasion of A-375 tumors, we next studied the effects of the bisphosphonate incadronate (INC), a potent inhibitor of osteoclastic bone resorption, using this model. Daily subcutaneous injections of INC (1 mg/kg) markedly decreased A-375-increased osteoclasts, increased numbers of apoptotic osteoclasts and inhibited mandibular cortical bone destruction. Furthermore, INC significantly suppressed the growth of A-375 tumors, increased numbers of apoptotic A-375 cells and decreased the tumor-associated blood vessel density. These results suggest that invasion and growth of oral cancers in mandibular bone is dependent on osteoclastic bone resorption and that INC can inhibit the aggressive behavior of oral cancer through inhibiting osteoclastic bone resorption.