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Keywords:

  • RANKL;
  • osteoprotegerin;
  • osteoporosis;
  • bone resorption;
  • AMG 162

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

The safety and bone antiresorptive effect of a single subcutaneous dose of AMG 162, a human monoclonal antibody to RANKL, was investigated in 49 postmenopausal women. AMG 162 is a potent antiresorptive agent for diseases such as osteoporosis.

Introduction: RANKL is an essential osteoclastic differentiation and activation factor.

Materials and Methods: The bone antiresorptive activity and safety of AMG 162, a fully human monoclonal antibody to RANKL, were evaluated in postmenopausal women in this randomized, double-blind, placebo-controlled, single-dose, dose escalation study. Six cohorts of eight to nine women were randomly assigned to receive a single subcutaneous injection of either AMG 162 or placebo (3:1 ratio). AMG 162 doses were 0.01, 0.03, 0.1, 0.3, 1.0, and 3.0 mg/kg. Subjects were followed up to 6 months in all cohorts and 9 months in the three highest dose cohorts. Second morning void urinary N-telopeptide/creatinine (NTX; Osteomark), serum NTX, and serum bone-specific alkaline phosphatase (BALP, Ostase) were assessed as bone turnover markers.

Results and Conclusions: Forty-nine women were enrolled. A single subcutaneous dose of AMG 162 resulted in a dose-dependent, rapid (within 12 h), profound (up to 84%), and sustained (up to 6 months) decrease in urinary NTX. At 6 months, there was a mean change from baseline of −81% in the 3.0 mg/kg AMG 162 group compared with −10% in the placebo group; serum NTX changes were −56% and 2%, respectively. BALP levels did not decrease remarkably until after 1 month, indicating that the effect of AMG 162 is primarily antiresorptive. Intact parathyroid hormone (PTH) levels increased up to ∼3-fold after 4 days in the 3.0 mg/kg dose group, but returned toward baseline with follow-up. Albumin-adjusted serum calcium did not decrease >10% on average in any group, and no subject had values below 2 mmol/liter. AMG 162 was well tolerated. No related serious adverse events occurred. No clinically meaningful laboratory changes, other than those described above, were observed. In summary, a single subcutaneous dose of AMG 162 resulted in a dose-dependent rapid and sustained decrease from baseline in bone turnover and could be an effective and convenient treatment for osteoporosis.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

RANKL, originally identified in dendritic cells,(1) has been well documented as a critical factor in the terminal differentiation and activation of osteoclasts.(2–4) Genetic experiments have shown that mice with a disrupted RANKL gene have severe osteopetrosis and absence of osteoclasts.(5) As a soluble member of the TNF receptor family, osteoprotegerin (OPG) blocks bone resorption by binding to RANKL and preventing its interaction with RANK, a cell surface receptor on pre-osteoclasts and osteoclasts.(6,7) We previously showed that an Fc-OPG fusion protein is able to effectively block bone resorption in postmenopausal women(8) and patients with breast carcinoma or multiple myeloma.(9) Since these discoveries, RANKL signaling has been implicated in many bone diseases associated with increased bone resorption, such as osteoporosis(8,10,11), multiple myeloma,(9,12–14) bone metastasis associated with breast carcinoma(9,15–18) and prostate carcinoma,(19,20) bone pain associated with bone metastasis,(21–24) rheumatoid arthritis,(25–29) psoriatic arthritis,(30) familial expansile osteolysis,(31) Paget's disease,(32) juvenile Paget's disease,(33) giant cell tumors,(34) and periprosthetic bone loss.(35)

AMG 162, a fully human monoclonal antibody to RANKL, blocks binding of RANKL to RANK. This report describes the findings from a single-dose, placebo-controlled study with AMG 162 in postmenopausal women to determine its safety and bone antiresorptive effect.

Results from this study have been presented previously in abstract form.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Study design

This is a single-dose, placebo-controlled, dose escalation study in healthy postmenopausal women conducted at two centers. The main objectives were to assess safety and tolerability, the effect of AMG 162 on bone turnover as measured by biochemical markers, and the pharmacokinetics of AMG 162. Subjects (eight to nine per dose cohort) were randomized in a 3:1 ratio to receive either AMG 162 or matching placebo by subcutaneous abdominal injection. AMG 162 is a fully human monoclonal antibody (IgG2) with high affinity and specificity for human RANKL. Placebo consisted of vehicle only and was indistinguishable from AMG 162 preparations. The AMG 162 doses tested were 0.01, 0.03, 0.1, 0.3, 1.0, and 3.0 mg/kg. After dosing, the three lower dose cohorts were followed for 6 months and the three higher dose cohorts for 9 months because of prolonged suppression of bone turnover.

Study subjects

Subjects were at least 1 year postmenopausal and were enrolled at two study centers (Lenexa, Kansas and Miami, FL, USA). They were healthy and were not allowed to be on any therapies that might significantly affect bone turnover, e.g., bisphosphonates or fluoride within 12 months, and estrogens, selective estrogen receptor modulators, calcitonin, parathyroid hormone, high doses of Vitamin D (> 1,000 IU daily), anabolic steroids, systemic glucocorticosteroids, or calcitriol within 6 months of enrollment. Subjects were also excluded if they had evidence of a disease that might influence the results, e.g., hyperparathyroidism, hyperthyroidism, hypothyroidism, rheumatoid arthritis, Paget's disease, osteomalacia, or recent fracture (within 6 months).

Study procedures

Subjects received a single dose of study drug in the morning and were followed for 6 or 9 months. The following procedures were performed during the study: medical and medication history (prestudy) and physical examinations, vital signs, hematology, serum chemistry, coagulation parameters, immunoglobulins, urinalysis, T- and B-cell enumeration (CD3, CD4, CD8, CD20, and CD56), anti-AMG 162 antibodies, and ECG (prestudy and periodically during the study). The following parameters were measured to assess the effect of AMG 162 on bone metabolism: urinary N-telopeptide/creatinine (NTX, Osteomark, Seattle, WA, USA), serum NTX (Osteomark), serum bone-specific alkaline phosphatase (BALP; Tandem-R Ostase; Hybritech, San Diego, CA, USA), serum intact parathyroid hormone (iPTH; Nichols assay), serum albumin-adjusted calcium, serum phosphorus, urinary calcium/creatinine, and urinary phosphorus/creatinine. A second morning void sample was used for the urinary parameters. Predose urine and blood samples were collected on 2 separate days before dosing to establish the baseline level for urine and serum NTX and BALP, and the average of these two assays was used as the baseline value.

Adverse events and concomitant medications were assessed at all study visits after dosing. This study was conducted in accordance with ICH guidelines and was approved by the local Institutional Review Boards.

Data analysis

The data from the subjects who received placebo in all six dose cohorts were pooled. Serum albumin-adjusted calcium (mmol/liter) was calculated as serum calcium (mmol/liter) − [0.02 × albumin (g/liter)] + 0.8. Change and percent change from baseline was calculated for all subjects for the bone metabolism parameters, and the mean change and/or percent change from baseline was compared across dose groups. The mean percent changes from baseline in urinary NTX/creatinine, serum NTX, and BALP were compared at each visit between each AMG 162 dose and placebo using a linear model controlling for dose and baseline value. Comparisons were adjusted for multiple testing using Dunnett's method.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Demographics and baseline characteristics

In Table 1, the baseline demographics and characteristics of the study population are summarized. No marked imbalances were noted across the dose groups. The mean age ranged from 54 to 63 years, and subjects were 7-15 years postmenopause. The majority of subjects (81%) were white, 16% were Hispanic, and 3% were black. The mean body mass index (BMI) ranged from 23.6 to 29.5 kg/m2 across groups. The mean baseline urinary NTX/creatinine ranged from 36 to 66 nmol/mmol, serum NTX from 12 to 17 nmol/liter, BALP from 13 to 17 μg/liter, iPTH from 4.2 to 5.6 pM, and albumin-adjusted serum calcium from 2.26 to 2.40 mmol/liter. The bone density of study subjects at baseline is unknown, because bone densitometry was not a required study procedure.

Table Table 1.. Demographics and Baseline Characteristics of the Study Population
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Table Table 2.. Change from Baseline in Urinary NTX/Creatinine (%)
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Table Table 3.. Change From Baseline in Serum NTX (%)
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Bone metabolism

After a single subcutaneous dose of AMG 162, there was a dose-dependent decrease in bone turnover as reflected by the changes observed in urinary NTX/creatinine (Fig. 1; Table 2) and serum NTX (Table 3). At the higher doses of AMG 162, decreases were observed as early as 12 h after dosing in urine NTX/creatinine (−46% in placebo and −77% in the 3.0 mg/kg AMG 162 group; see Table 2). Note that the placebo group also showed a decrease at 12 h. This was expected because of the diurnal variation of this marker(36): the predose urine sample was taken in the morning, when the urine NTX/creatinine level is relatively high, and the 12-h sample was taken in the early evening, when the urine NTX/creatinine level is relatively low. Twenty-four hours after dosing, there was a mean decrease from baseline in urinary NTX/creatinine of 73% in the 3.0 mg/kg AMG 162 group compared with 10% in the placebo group. The maximum urinary NTX/creatinine reduction was observed at 2 weeks in the 0.01, 0.03, 0.3, and 1.0 mg/kg AMG 162 groups, 1 month in the 0.1 mg/kg AMG 162 group, and 3 months in the 3.0 mg/kg AMG 162 group (84%). The pharmacodynamic response to AMG 162, as measured by urinary NTX/creatinine, was consistently observed across all subjects in the higher dose groups. For example, at 6 months, the range of percent changes observed was −37% to −79% in the 1.0 mg/kg group and −56% to −94% in the 3.0 mg/kg group compared with −48 to +62% in the placebo group. There was a mean change of −9% (median, −10%) in the placebo group at 9 months compared with −51% in the 3.0 mg/kg AMG 162 group (not different statistically). The treatment effect of AMG 162 was reversible, as indicated by a return toward baseline levels at ∼2 months in the 0.01 and 0.03 mg/kg AMG 162 groups, 4 months in the 0.1 mg/kg group, 6 months in the 0.3 mg/kg AMG 162 group, and 9 months in the 1.0 and 3.0 mg/kg dose groups.

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Figure FIG. 1.. The effect of AMG 162 treatment on bone resorption as reflected by changes in a second morning void urinary NTX/creatinine (nmol BCE/mmol creatinine) over time. Data are presented as mean and SEM. Placebo, no symbol; 0.01 mg/kg AMG 162, ○; 0.03 mg/kg AMG 162, □; 0.1 mg/kg AMG 162, ▵; 0.3 mg/kg AMG 162, ▿; 1.0 mg/kg AMG 162, [Hollow Diamond] 3.0 mg/kg AMG 162, *.

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The serum NTX data (Table 3) confirmed the findings with urinary NTX/creatinine. The magnitude of the decreases in serum NTX was not as large as with urinary NTX. This has been observed in other studies.(37,38) The maximum mean decrease observed in any treatment group was 65% at 1 month in the 0.1 mg/kg AMG 162 group. As with urine NTX, the reversibility of the treatment effect was confirmed. At 6 months, the mean change from baseline was 2%, −43%, and −56% in the placebo, 1.0 mg/kg AMG 162, and 3.0 mg/kg AMG 162 groups, respectively, and at 9 months, −11%, −13%, and −40%, respectively, again confirming the prolonged antiresorptive effect at the higher AMG 162 doses. The change at 9 months in the 3.0 mg/kg AMG 162 group (−40%) was statistically significantly (p < 0.05) different from placebo (−11%).

The BALP levels remained close to baseline levels in all groups up to ∼2 weeks before showing a dose-dependent decrease (Table 4). The maximum mean change was −53% in the 3.0 mg/kg group at 5 months (compared with 8% in the placebo group; data not shown in Table 4), and there was still a mean change of −37% at 9 months (compared with 33% in the placebo group).

Table Table 4.. Change From Baseline in BALP (%)
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The albumin-adjusted serum calcium levels in the AMG 162-treated subjects show early but modest decreases from baseline, which were especially evident in the higher dose groups (Table 5). The maximum mean decrease at any time point was 10% (in the 0.3 mg/kg AMG 162 group at 14 days). None of the subjects had values below 2 mmol/liter. Note that subjects were instructed not to take calcium or vitamin D supplements during the study to ensure that hypocalcemic events would not be masked. The serum phosphorus levels also decreased in a similar manner to the serum calcium levels because of the antiresorptive effect of AMG 162 (data not shown). There were dose-dependent early increases in iPTH levels in the AMG 162 groups (Table 6). The maximum mean increase was ∼3-fold in the 3.0 mg/kg group 4 days after dosing, but the mean levels decreased over time. At 6 months, the mean change from baseline was 8% and 67% in the 1.0 and 3.0 mg/kg groups, respectively, compared with −3% in the placebo group. There were decreases in urinary calcium/creatinine levels in subjects receiving AMG 162, but urinary phosphorus/creatinine levels remained relatively stable compared with the placebo control group (data not shown).

Table Table 5.. Albumin-Adjusted Serum Calcium (mmol/liter)
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Table Table 6.. Serum Intact PTH (pmol/liter)
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Safety and tolerability

No related serious adverse events were reported. Two unrelated events required hospitalization: a subject with moderately severe abdominal pain with no identified underlying pathology in the 0.01 mg/kg AMG 162 group, and another with cholecystitis in the 0.1 mg/kg AMG 162 group. None of the subjects were discontinued from the study because of an adverse event. The incidence of reported infectious events was similar across groups (33% in placebo and 38% in the AMG 162 group overall, with no apparent dose-dependent increase). The subcutaneous injections were well tolerated; one subject who received 1.0 mg/kg reported injection site pain and another who received 3.0 mg/kg had injection site rash and burning. No clinically significant changes in any other laboratory variables were noted. The mean white blood cell (WBC) count remained stable (within 10% of the mean baseline value) across all AMG 162 groups, and there were no changes associated with AMG 162 treatment in T- and B-cell counts (CD3, CD4, CD8, CD20, and CD56), coagulation parameters, and immunoglobulins. Serum from all subjects was assessed on a weekly basis through month 1, biweekly through month 3, and monthly through month 9 for anti-AMG 162 antibodies, and all tests were negative.

Pharmacokinetics

The SC pharmacokinetics of AMG 162 in postmenopausal women were nonlinear with dose. The serum profiles were characterized by three distinct phases (Fig. 2): (1) a prolonged absorption phase, which resulted in maximum serum concentrations that increased disproportionately greater (2.6-fold) than the increase in dose and were observed between 5 and 21 days after administration; (2) a prolonged β-phase, characterized by half-lives that increased with dose to a maximum of 32 days; and (3) a more rapid terminal phase observed at concentrations <1000 ng/ml with a half-life that increased from 5 to 10 days as dose increased from 0.01 to 3.0 mg/kg. Because of the nonlinear pharmacokinetics, the mean serum residence time (MRT) increased with dose from 12 to 46 days.

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Figure FIG. 2.. The serum concentration profile of AMG 162 (ng/ml) over time. Data are presented as mean and SEM.

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Because RANKL plays such a pivotal role in osteoclast development and bone resorptive activity, the ability to control its interaction with RANK and thereby manage bone diseases characterized by increased osteoclastic activity, such as osteoporosis, bone metastasis, rheumatoid arthritis, Paget's disease, and others, is compelling. AMG 162, a specific fully human monoclonal antibody to RANKL, which prevents RANKL binding to RANK, has been developed and tested in this first-in-human clinical study. The results show that AMG 162 seems to specifically and profoundly inhibit osteoclastic bone resorption, as indicated by the changes observed in the biochemical markers, urinary NTX/creatinine, and serum NTX. The effect is evident within 12 h of dosing, which indicates that mature and active osteoclastic activity is inhibited almost immediately. The effect is also profound as indicated by the low NTX levels observed during the period of maximum effect, which increases as the dose was increased. Therefore, little if any osteoclastic activity remains while AMG 162 is in circulation, which is for a prolonged period based on the pharmacokinetic profile. However, the effect is reversible as indicated by a return of NTX levels when AMG 162 is cleared from the circulation.

BALP levels do not show such a rapid decrease as NTX. This was anticipated, because AMG 162 does not primarily interfere with osteoblastic activity. By interrupting terminal osteoclastic development, it reduces the activation frequency (or birth rate) of basic multicellular units, the cellular units responsible for bone turnover. Osteoclastic bone resorption on dormant bone surfaces normally initiates the bone turnover cycle, which is followed by osteoblastic bone formation.(39,40) Therefore, when the activation frequency is decreased, the bone formation rate also decreases as a result. This decrease in BALP has been observed with bisphosphonates(41–43) and raloxifene.(44)

The degree of bone turnover suppression (up to 81% in the 3.0 mg/kg dose at month 6) is at least comparable with the most potent antiresorptive agents. Alendronate, at the marketed dose of 10 mg daily, is associated with a mean decrease in urinary NTX/creatinine of 64-70%,(41,45) and at 70 mg weekly, the mean decrease reported was 56%.(41) Risedronate (5 mg daily) resulted in a mean decrease of 40-60%,(46,47) and with 35 mg weekly, 61%.(47) A mean decrease of 54-65% has been reported at month 12 with 4 mg intravenous zoledronate.(43) Raloxifene (60 mg daily) showed a mean decrease of 17%.(44)

Transient decreases in serum albumin-adjusted calcium, phosphorus, and urinary calcium/creatinine are consistent with the antiresorptive effect of AMG 162, as is the compensatory increase in PTH secretion.

Because AMG 162 is effectively acting similarly to OPG, it is of interest to compare them. The safety and efficacy of Fc-OPG fusion molecules were studied in postmenopausal women(8) and patients with breast carcinoma-associated bone metastases or multiple myeloma(9) and indicated that these molecules are effective antiresorptive agents. However, data from the study reported here indicate that AMG 162 has superior characteristics: it is more potent, showing greater decreases in bone turnover markers at lower doses, and the duration of antiresorptive effect is also longer at equivalent doses.

AMG 162 seemed to be well tolerated, and no significant safety issues have been identified. Although AMG 162 is a potent antiresorptive agent, no significant degree of hypocalcemia has been observed. It is of note that the nadir of the mean serum albumin-adjusted calcium was observed 2-8 weeks after doses of 0.3 mg/kg or higher of AMG 162, which is later than observed with the Fc-OPG fusion molecules (2-8 days after dosing(8); data on file). This may be related to differences in the pharmacokinetics of the compounds.

Partial inhibition of early T- and B-lymphocyte development has been observed in RANKL-deficient mice.(5) There was no clinically significant effect on lymphocyte counts overall (CD3), T-cells (CD4, CD8, CD56), or B-cells (CD20) in this study, and no meaningful differences were observed among treatment groups regarding incidence of infectious events.

A potential risk with an OPG molecule is the generation of anti-OPG antibodies, which might cross-react with endogenous OPG, neutralizing its activity. Anti-OPG antibodies were observed in one subject in a phase 1 study with an Fc-OPG fusion molecule (P Bekker, unpublished observations, 2001), and although there was no apparent negative effect clinically, safety concerns could arise with chronic dosing. This prospect is avoided with AMG 162, because it does not resemble OPG structurally, and even if anti-AMG 162 antibodies were elicited, they would not be expected to cross-react with endogenous OPG. No evidence of anti-AMG 162 antibodies was observed in this study.

Another potential concern is binding of OPG to TNF-related apoptosis-inducing ligand (TRAIL,(48) a survival factor for tumor cells) and interference with a natural defense mechanism against tumorigenesis. Binding of OPG to a soluble form of TRAIL has been reported.(49) Although OPG has a low affinity for membrane-associated TRAIL (B Boyle, unpublished observations, 2001), this could nevertheless be a potential concern at high doses of OPG. Because AMG 162 is specific for RANKL, it does not bind to TRAIL. In an in vitro competition assay with RANKL-expressing Chinese hamster ovary cells, TRAIL, TNFα, TNFβ, and CD40L (tested at doses up to 1 μg/ml) were unable to compete with anti-RANKL antibody binding to RANKL, whereas this binding was competed by exogenously added human RANKL (P Kostenuik, unpublished observations, 2004). Therefore, the concern regarding AMG 162 binding to TRAIL was alleviated.

The effect of AMG 162 treatment at the bone histological level has not been assessed in this study, and conclusions regarding its safety in this regard cannot be drawn. This will require larger, long-term studies.

The results from this study indicate that AMG 162 could potentially have therapeutic applications to several bone disorders, including osteoporosis, cancer-related bone disease, rheumatoid arthritis, psoriatic arthritis, Paget's disease, osteoclastoma, prosthesis loosening, and periodontal disease. Longer-term, multiple dose studies are ongoing to assess the use of AMG 162 in modifying the course of bone disease.

Although this study was conducted at only two study centers in a small number of healthy postmenopausal subjects, it is likely that, based on results from this study, a similar bone antiresorptive effect would be observed in osteoporotic patients and those with other bone diseases. The effects of AMG 162 on bone density were not assessed in this study, but it is likely, based on the data from this study and the established link between bone resorption and bone density, that a positive effect on bone density would be observed in patients treated with AMG 162. Ongoing studies are assessing the effects of this treatment on bone density.

In summary, this randomized, single-dose, double-blind, placebo-controlled study showed that AMG 162, a fully human, high-affinity, specific anti-RANKL monoclonal antibody, is a potent, long-acting, well-tolerated bone antiresorptive agent with potential broad application in the treatment of bone disorders.

Acknowledgements

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

This study was supported with funding from Amgen, Inc.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES
  • 1
    Anderson DM, Maraskovsky E, Billingsley WL, Dougall WC, Tometsko ME, Roux ER, Teepe MC, DuBose RF, Cosman D, Galibert L 1997 A homologue of the TNF receptor and its ligand enhance T-cell growth and dendritic-cell function. Nature 390: 175179.
  • 2
    Lacey DL, Timms E, Tan HL, Kelley MJ, Dunstan CR, Burgess T, Elliott R, Colombero A, Elliott G, Scully S, Hsu H, Sullivan J, Hawkins N, Davy E, Capparelli C, Eli A, Qian YX, Kaufman S, Sarosi I, Shalhoub V, Senaldi G, Guo J, Delaney J, Boyle WJ 1998 Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell 93: 165176.
  • 3
    Yasuda H, Shima N, Nakagawa N, Yamaguchi K, Kinosaki M, Mochizuki S, Tomoyasu A, Yano K, Goto M, Murakami A, Tsuda E, Morinaga T, Higashio K, Udagawa N, Takahashi N, Suda T 1998 Osteoclast differentiation factor is a ligand for osteoprotegerin/osteoclastogenesis-inhibitory factor and is identical to TRANCE/RANKL. Proc Natl Acad Sci USA 95: 35973602.
  • 4
    Burgess TL, Qian YX, Kaufman S, Ring BD, Van G, Capparelli C, Kelley M, Hsu HL, Boyle WJ, Dunstan CR, Hu S, Lacey DL 1999 The ligand for osteoprotegerin (OPGL) directly activates mature osteoclasts. J Cell Biol 145: 527538.
  • 5
    Kong YY, Yoshida H, Sarosi I, Tan HL, Timms E, Capparelli C, Morony S, Oliveira-dos SA, Van G, Itie A, Khoo W, Wakeham A, Dunstan CR, Lacey DL, Mak TW, Boyle WJ, Penninger JM 1999 OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis. Nature 397: 315323.
  • 6
    Simonet WS, Lacey DL, Dunstan CR, Kelley M, Chang MS, Luethy R, Nguyen HQ, Wooden S, Bennett L, Boone T, Shimamoto G, DeRose M, Elliott R, Colombero A, Tan HL, Trail G, Sullivan J, Davy E, Bucay N, Renshaw GL, Hughes TM, Hill D, Pattison W, Campbell P, Boyle WJ 1997 Osteoprotegerin: A novel secreted protein involved in the regulation of bone density. Cell 89: 309319.
  • 7
    Tsuda E, Goto M, Mochizuki SI, Yano K, Kobayashi F, Morinaga T, Higashio K 1997 Isolation of a novel cytokine from human fibroblasts that specifically inhibits osteoclastogenesis. Biochim Biophys Res Commun 234: 137142.
  • 8
    Bekker PJ, Holloway D, Nakanishi A, Arrighi M, Leese PT, Dunstan CR 2001 The effect of a single dose of osteoprotegerin in postmenopausal women. J Bone Miner Res 16: 348360.
  • 9
    Body JJ, Greipp P, Coleman RE, Facon T, Geurs F, Fermand JP, Harousseau JL, Lipton A, Mariette X, Williams CD, Nakanishi A, Holloway D, Martin SW, Dunstan CR, Bekker PJ 2003 A phase I study of AMGN-0007, a recombinant osteoprotegerin construct, in patients with multiple myeloma or breast carcinoma related bone metastases. Cancer 97: 887892.
  • 10
    Bucay N, Sarosi I, Dunstan CR, Morony S, Tarpley J, Capparelli C, Scully S, Tan HL, Xu WL, Lacey DL, Boyle WJ, Simonet WS 1998 Osteoprotegerin-deficient mice develop early onset osteoporosis and arterial calcification. Genes Dev 12: 12601268.
  • 11
    Kostenuik PJ, Capparelli C, Morony S, Adamu S, Shimamoto G, Shen V, Lacey DL, Dunstan CR 2001 OPG and PTH-(1–34) have additive effects on bone density and mechanical strength in osteopenic ovariectomized rats. Endocrine 142: 42954304.
  • 12
    Croucher PI, Shipman CM, Lippitt J, Perry M, Asosingh K, Hijzen A, Brabbs AC, Van Beek EJR, Holen I, Skerry TM, Dunstan CR, Russell GR, Van Camp B, Vanderkerken K 2001 Osteoprotegerin inhibits the development of osteolytic bone disease in multiple myeloma. Blood 98: 35343540.
  • 13
    Sezer O, Heider U, Zavrski I, Kuhne CA, Hofbauer LC 2003 RANK ligand and osteoprotegerin in myeloma bone disease. Blood 101: 20942098.
  • 14
    Roux S, Meignin V, Quillard J, Meduri G, Guiochon-Mantel A, Fermand JP, Milgrom E, Mariette X 2002 RANK (receptor activator of nuclear factor-kappaB) and RANKL expression in multiple myeloma. Brit J Haematol 117: 8692.
  • 15
    Thomas RJ, Guise TA, Yin JJ, Elliott J, Horwood NJ, Martin TJ, Gillespie MT 1999 Breast cancer cells interact with osteoblasts to support osteoclast formation. Endocrine 140: 44514458.
  • 16
    Mancino AT, Klimberg VS, Yamamoto M, Manolagas SC, Abe E 2001 Breast cancer increases osteoclastogenesis by secreting M-CSF and upregulating RANKL in stromal cells. J Surg Res 100: 1824.
  • 17
    Morony S, Capparelli C, Lee R, Shimamoto G, Boone T, Lacey DL, Dunstan CR 1999 A chimeric form of osteoprotegerin inhibits hypercalcemia and bone resorption induced by IL-1 beta, TNF-alpha, PTH, PTHrP, and 1,25(OH)(2)D-3. J Bone Miner Res 14: 14781485.
  • 18
    Kitazawa S, Kitazawa R 2002 RANK ligand is a prerequisite for cancer-associated osteolytic lesions. J Pathol 198: 228236.
  • 19
    Brown JM, Corey E, Lee ZD, True LD, Yun TJ, Tondravi M, Vessella RL 2001 Osteoprotegerin and RANK ligand expression in prostate cancer. Urology 57: 611616.
  • 20
    Zhang J, Dai J, Qi Y, Lin DL, Smith P, Strayhorn C, Mizokami A, Fu Z, Westman J, Keller ET 2001 Osteoprotegerin inhibits prostate cancer-induced osteoclastogenesis and prevents prostate tumor growth in the bone. J Clin Invest 107: 12351244.
  • 21
    Honore P, Luger NM, Sabino MA, Schwei MJ, Rogers SD, Mach DB, O'Keefe PF, Ramnaraine ML, Clohisy DR, Mantyh PW 2000 Osteoprotegerin blocks bone cancer-induced skeletal destruction, skeletal pain and pain-related neurochemical reorganization of the spinal cord. Nat Med 6: 521528.
  • 22
    Mantyh PW, Clohisy DR, Koltzenburg M, Hunt SP 2002 Molecular mechanisms of cancer pain. Nature Rev Cancer 2: 201209.
  • 23
    Clohisy DR, Ramnaraine ML, Scully S, Qi M, Van G, Hong LT, Lacey DL 2000 Osteoprotegerin inhibits tumor-induced osteoclastogenesis and bone tumor growth in osteopetrotic mice. J Orthop Res 18: 967976.
  • 24
    Luger NM, Honore P, Sabino MA, Schwei MJ, Rogers SD, Mach DB, Clohisy DR, Mantyh PW 2001 Osteoprotegerin diminishes advanced bone cancer pain. Cancer Res 61: 40384047.
  • 25
    Kong YY, Feige U, Sarosi I, Bolon B, Tafuri A, Morony S, Capparelli C, Li J, Elliott R, McCabe S, Wong T, Campagnuolo G, Moran E, Bogoch ER, Van G, Nguyen LT, Ohashi PS, Lacey DL, Fish E, Boyle WJ, Penninger JM 1999 Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand. Nature 402: 304309.
  • 26
    Romas E, Sims NA, Hards DK, Lindsay M, Quinn JWM, Ryan PFJ, Dunstan CR, Martin TJ, Gillespie MT 2002 Osteoprotegerin reduces osteoclast numbers and prevents bone erosion in collagen-induced arthritis. Am J Pathol 161: 14191427.
  • 27
    Haynes DR, Crotti TN, Loric M, Bain GI, Atkins GJ, Findlay DM 2001 Osteoprotegerin and receptor activator of nuclear factor kappaB ligand (RANKL) regulate osteoclast formation by cells in the human rheumatoid arthritic joint. Rheumatology 40: 623630.
  • 28
    Crotti TN, Smith MD, Weedon H, Ahern MJ, Findlay DM, Kraan M, Tak PP, Haynes DR 2002 Receptor activator NF-kappa B ligand (RANKL) expression in synovial tissue from patients with rheumatoid arthritis, spondyloarthropathy, osteoarthritis, and from normal patients: Semiquantitative and quantitative analysis. Ann Rheum Dis 61: 10471054.
  • 29
    Gravallese EM, Manning C, Tsay A, Naito A, Pan C, Amento E, Goldring SR 2000 Synovial tissue in rheumatoid arthritis is a source of osteoclast differentiation factor. Arthritis Rheum 43: 250258.
  • 30
    Ritchlin CT, Haas-Smith SA, Li P, Hicks DG, Schwarz EM 2003 Mechanisms of TNF-alpha- and RANKL-mediated osteoclastogenesis and bone resorption in psoriatic arthritis. J Clin Invest 111: 821831.
  • 31
    Sparks AB, Peterson SN, Bell C, Loftus BJ, Hocking L, Cahill DP, Frassica FJ, Streeten EA, Levine MA, Fraser CM, Adams MD, Broder S, Venter JC, Kinzler KW, Vogelstein B, Ralston SH 2001 Mutation screening of the TNFRSF11A gene encoding receptor activator of NF kappa B (RANK) in familial and sporadic Paget's disease of bone and osteosarcoma. Calcif Tissue Int 68: 151155.
  • 32
    Hughes AE, Ralston SH, Marken J, Bell C, MacPherson H, Wallace RGH, van-Hul W, Whyte MP, Nakatsuka K, Hovy L, Anderson DM 2000 Mutations in TNFRSF11A, affecting the signal peptide of RANK, cause familial expansile osteolysis. Nat Genet 24: 4548.
  • 33
    Whyte MP, Obrecht SE, Finnegan PM, Jones JL, Podgornik MN, McAlister WH, Mumm S 2002 Osteoprotegerin deficiency and juvenile Paget's disease. N Engl J Med 347: 175184.
  • 34
    Atkins GJ, Bouralexis S, Haynes DR, Graves SE, Geary SM, Evdokiou A, Zannettino AC, Hay S, Findlay DM 2001 Osteoprotegerin inhibits osteoclast formation and bone resorbing activity in giant cell tumors of bone. Bone 28: 370377.
  • 35
    Haynes DR, Crotti TN, Potter AE, Loric M, Atkins GJ, Howie DW, Findlay DM 2001 The osteoclastogenic molecules RANKL and RANK are associated with periprosthetic osteolysis. J Bone Joint Surg Br 83: 902911.
  • 36
    Ju HSJ, Leong S, Brown B, Stringer MA, Leigh S, Scherrer C, Shepard K, Jenkins D, Knudsen J, Cannon R 1997 Comparison of analytical performance and biological variability of three bone resorption assays. Clin Chem 43: 15701576.
  • 37
    Chesnut CH III, Bell NH, Clark GS, Drinkwater BL, English SC, Johnson CC Jr, Notelovitz M, Rosen C, Cain DF, Flessland KA, Mallinak NJ 1997 Hormone replacement therapy in postmenopausal women: Urinary N-telopeptide of type I collagen monitors therapeutic effect and predicts response of bone mineral density. Am J Med 102: 2937.
  • 38
    Eastell R, Mallinak N, Weiss S, Ettinger M, Pettinger M, Cain D, Fressland K, Chesnut C III 2000 Biological variability of serum and urinary N-telopeptides of type I collagen in postmenopausal women. J Bone Miner Res 15: 594598.
  • 39
    Frost HM 1969 Tetracycline-based histological analysis of bone remodeling. Calcif Tissue Res 3: 211237.
  • 40
    Eriksen EF 1986 Normal and pathological remodeling of human trabecular bone: Three dimensional reconstruction of the remodeling sequence in normals and in metabolic bone disease. Endocr Rev 7: 379408.
  • 41
    Schnitzer T, Bone HG, Crepaldi G, Adami S, McClung M, Kiel D, Felsenberg D, Recker RR, Tonino RP, Roux C, Pinchera A, Foldes AJ, Greenspan SL, Levine MA, Emkey R, Santora AC, Kaur A, Thompson DE, Yates J, Orloff JJ, Alendronate-Once WS-G 2000 Therapeutic equivalence of alendronate 70 mg once-weekly and alendronate 10 mg daily in the treatment of osteoporosis. Aging Clin Exp Res 12: 112.
  • 42
    Harris ST, Watts NB, Genant HK, McKeever CD, Hangartner T, Keller M, Chesnut CH III, Brown J, Eriksen EF, Hoseyni MS, Axelrod DW, Miller PD 1999 Effects of risedronate treatment on vertebral and nonvertebral fractures in women with postmenopausal osteoporosis: A randomized controlled trial. Vertebral Efficacy with Risedronate Therapy (VERT) Study Group. JAMA 282: 13441352.
  • 43
    Reid IR, Brown JP, Burckhardt P, Horowitz Z, Richardson P, Trechsel U, Widmer A, Devogelaer JP, Kaufman JM, Jaeger P, Body JJ, Brandi ML, Broell J, Di Micco R, Genazzani AR, Felsenberg D, Happ J, Hooper MJ, Ittner J, Leb G, Mallmin H, Murray T, Ortolani S, Rubinacci A, Saaf M, Samsioe G, Verbruggen L, Meunier PJ 2002 Intravenous zoledronic acid in postmenopausal women with low bone mineral density. N Engl J Med 346: 653661.
  • 44
    Johnston CC Jr, Bjarnason NH, Cohen FJ, Shah A, Lindsay R, Mitlak BH, Huster W, Draper MW, Harper KD, Heath H III, Gennari C, Christiansen C, Arnaud CD, Delmas PD 2000 Long-term effects of raloxifene on bone mineral density, bone turnover, and serum lipid levels in early postmenopausal women: Three-year data from 2 double-blind, randomized, placebo-controlled trials. Arch Intern Med 160: 34443450.
  • 45
    Tonino RP, Meunier PJ, Emkey R, Rodriguez-Portales JA, Menkes CJ, Wasnich RD, Bone HG, Santora AC, Wu M, Desai R, Ross PD 2000 Skeletal benefits of alendronate: 7-year treatment of postmenopausal osteoporotic women. Phase III Osteoporosis Treatment Study Group. J Clin Endocrinol Metab 85: 31093115.
  • 46
    Fogelman I, Ribot C, Smith R, Ethgen D, Sod E, Reginster JY 2000 Risedronate reverses bone loss in postmenopausal women with low bone mass: Results from a multinational, double-blind, placebo-controlled trial. BMD-MN Study Group. J Clin Endocrinol Metab 85: 18951900.
  • 47
    Brown JP, Kendler DL, Mcclung MR, Emkey RD, Adachi JD, Bolognese MA, Li Z, Balske A, Lindsay R 2002 The efficacy and tolerability of risedronate once a week for the treatment of postmenopausal osteoporosis. Calcif Tissue Int 71: 103111.
  • 48
    Wiley SR, Schooley K, Smolak PJ, Din WS, Huang CP, Nicholl JK, Sutherland GR, Smith TD, Rauch C, Smith CA 1995 Identification and characterization of a new member of the TNF family that induces apoptosis. Immunity 3: 673682.
  • 49
    Emery JG, McDonnell P, Burke MB, Deen KC, Lyn S, Silverman C, Dul E, Appelbaum ER, Eichman C, DiPrinzio R, Dodds RA, James IE, Rosenberg M, Lee JC, Young PR 1998 Osteoprotegerin is a receptor for the cytotoxic ligand TRAIL. J Biol Chem 273: 1436314367.