Advances in Osteoporosis Therapy: Focus on Osteoanabolic Agents, Secondary Fracture Prevention, and Perioperative Bone Health.

PURPOSE OF REVIEW: This review summarizes recently published data and other developments around osteoanabolic osteoporosis therapies in patients with very high fracture risk, including those undergoing bone-related surgery. RECENT FINDINGS: Two osteoanabolic agents, abaloparatide and romosozumab, were recently approved for treatment of patients with osteoporosis at high fracture risk. These agents, along with teriparatide, are valuable for primary and secondary fracture prevention. Orthopedic surgeons are well positioned to facilitate secondary fracture prevention via referrals to fracture liaison services or other bone health specialist colleagues. This review aims to help surgeons understand how to identify patients with sufficiently high fracture risk to warrant consideration of osteoanabolic therapy. Recent evidence around the perioperative use and potential benefits of osteoanabolic agents in fracture healing and other orthopedic settings (e.g., spinal fusion and arthroplasty) in individuals with osteoporosis is also discussed. Osteoanabolic agents should be considered for patients with osteoporosis at very high fracture risk, including those with prior osteoporotic fractures and those with poor bone health who are undergoing bone-related surgery.

Dual Inhibition of the Wnt Inhibitors DKK1 and Sclerostin Promotes Fracture Healing and Increases the Density and Strength of Uninjured Bone: An Experimental Study in Nonhuman Primates.

BACKGROUND: Fracture repair involves the reactivation of developmental signaling cascades, including Wnt signaling that stimulates bone formation and bone regeneration. Rodent data indicate that dual inhibition of the Wnt signaling antagonists sclerostin and Dickkopf-1 (DKK1) increases callus bone volume and strength while increasing bone mass systemically. METHODS: We evaluated the effects of 16 weeks of subcutaneously administered carrier solution (vehicle, VEH), anti-sclerostin antibody (Scl-Ab), anti-DKK1 antibody (DKK1-Ab), or Scl-Ab plus DKK1-Ab combination therapy (COMBO) on ulnar osteotomy healing in nonhuman primates (cynomolgus monkeys; 20 to 22 per group). RESULTS: Scl-Ab and COMBO therapy increased systemic markers of bone formation versus VEH, with COMBO leading to synergistic increases versus Scl-Ab or DKK1-Ab monotherapies. The COMBO and Scl-Ab groups showed reduced serum markers of bone resorption versus VEH. The COMBO and DKK1-Ab groups exhibited greater callus bone mineral density (BMD), torsional stiffness, and torsional rigidity versus VEH. Lumbar vertebrae from the Scl-Ab and COMBO groups showed greater BMD and bone formation rate versus VEH, and the femoral mid-diaphysis of the Scl-Ab and COMBO groups showed greater periosteal and endocortical bone formation rates versus VEH. CONCLUSIONS: DKK1-Ab increased BMD and strength at the ulnar osteotomy site, Scl-Ab increased bone formation and BMD at uninjured skeletal sites, and Scl-Ab plus DKK1-Ab combination therapy induced all of these effects, in some cases to a greater degree versus 1 or both monotherapies. These results in nonhuman primates suggest that DKK1 preferentially regulates bone healing while sclerostin preferentially regulates systemic bone mass. CLINICAL RELEVANCE: Combination therapy with antibodies against sclerostin and DKK1 may offer a promising therapeutic strategy for both fracture treatment and fracture prevention.

Twelve Months of Denosumab and/or Alendronate Is Associated With Improved Bone Fatigue Life, Microarchitecture, and Density in Ovariectomized Cynomolgus Monkeys.

Prolonged use of antiresorptives such as the bisphosphonate alendronate (ALN) and the RANKL inhibitor denosumab (DMAb) are associated with rare cases of atypical femoral fracture (AFF). The etiology of AFF is unclear, but it has been hypothesized that potent osteoclast inhibitors may reduce bone fatigue resistance. The purpose of this study was to quantify the relationship between antiresorptive treatment and fatigue life (cycles to failure) in bone from ovariectomized cynomolgus monkeys. We analyzed humeral bone from 30 animals across five treatment groups. Animals were treated for 12 months with subcutaneous (sc) vehicle (VEH), sc DMAb (25 mg/kg/month), or intravenous (iv) ALN (50 µg/kg/month). Another group received 6 months VEH followed by 6 months DMAb (VEH-DMAb), and the final group received 6 months ALN followed by 6 months DMAb (ALN-DMAb). A total of 240 cortical beam samples were cyclically tested in four-point bending at 80, 100, 120, or 140 MPa peak stress. High-resolution imaging and density measurements were performed to evaluate bone microstructure and composition. Samples from the ALN (p = 0.014), ALN-DMAb (p = 0.008), and DMAb (p < 0.001) groups illustrated higher fatigue-life measurements than VEH. For example, at 140 MPa the VEH group demonstrated a median ± interquartile range (IQR) fatigue life of 1987 ± 10593 cycles, while animals in the ALN, ALN-DMAb, and DMAb groups survived 9850 ± 13648 (+395% versus VEH), 10493 ± 16796 (+428%), and 14495 ± 49299 (+629%) cycles, respectively. All antiresorptive treatment groups demonstrated lower porosity, smaller pore size, greater pore spacing, and lower number of canals versus VEH (p < 0.001). Antiresorptive treatment was also associated with greater apparent density, dry density, and ash density (p ≤ 0.03). We did not detect detrimental changes following antiresorptive treatments that would explain their association with AFF. In contrast, 12 months of treatment may have a protective effect against fatigue fractures. © 2022 American Society for Bone and Mineral Research (ASBMR).

Denosumab treatment is associated with decreased cortical porosity and increased bone density and strength at the proximal humerus of ovariectomized cynomolgus monkeys.

Upper extremity fractures, including those at the humerus, are common among women with postmenopausal osteoporosis. Denosumab was shown to reduce humeral fractures in this population; however, no clinical or preclinical studies have quantified the effects of denosumab on humerus bone mineral density or bone microarchitecture changes. This study used micro-computed tomography (µCT) and computed tomography (CT), alongside image-based finite element (FE) models derived from both modalities, to quantify the effects of denosomab (DMAb) and alendronate (ALN) on humeral bone from acutely ovariectomized (OVX) cynomolgus monkeys. Animals were treated with 12 monthly injections of s.c. vehicle (VEH; n = 10), s.c. denosumab (DMAb; 25 mg/kg, n = 9), or i.v. alendronate (ALN; 50 µg/kg, n = 10). Two more groups received 6 months of VEH followed by 6 months of DMAb (VEH-DMAb; n = 7) or 6 months of ALN followed by 6 months of DMAb (ALN-DMAb; n = 9). After treatment, humeri were harvested and µCT was used to quantify tissue mineral density, trabecular morphology, and cortical porosity at the humeral head. Clinical CT imaging was also used to quantify trabecular and cortical bone mineral density (BMD) at the ultra-proximal, proximal, 1/5 proximal and midshaft of the bone. Finally, µCT-based FE models in compression, and CT-based FE models in compression, torsion, and bending, were developed to estimate differences in strength. Compared to VEH, groups that received DMAb at any time demonstrated lower cortical porosity and/or higher tissue mineral density via µCT; no effects on trabecular morphology were observed. FE estimated strength based on µCT was higher after 12-months DMAb (p = 0.020) and ALN-DMAb (p = 0.024) vs. VEH; respectively, FE predicted mean (SD) strength was 4649.88 (710.58) N, and 4621.10 (1050.16) N vs. 3309.4 (876.09) N. All antiresorptive treatments were associated with higher cortical BMD via CT at the 1/5 proximal and midshaft of the humerus; however, no differences in CT-based FE predicted strength were observed. Overall, these results help to explain the observed reductions in humeral fracture rate following DMAb treatment in women with postmenopausal osteoporosis.

Sclerostin antibody stimulates periodontal regeneration in large alveolar bone defects.

Destruction of the alveolar bone in the jaws can occur due to periodontitis, trauma or following tumor resection. Common reconstructive therapy can include the use of bone grafts with limited predictability and efficacy. Romosozumab, approved by the FDA in 2019, is a humanized sclerostin-neutralizing antibody (Scl-Ab) indicated in postmenopausal women with osteoporosis at high risk for fracture. Preclinical models show that Scl-Ab administration preserves bone volume during periodontal disease, repairs bone defects surrounding dental implants, and reverses alveolar bone loss following extraction socket remodeling. To date, there are no studies evaluating Scl-Ab to repair osseous defects around teeth or to identify the efficacy of locally-delivered Scl-Ab for targeted drug delivery. In this investigation, the use of systemically-delivered versus low dose locally-delivered Scl-Ab via poly(lactic-co-glycolic) acid (PLGA) microspheres (MSs) was compared at experimentally-created alveolar bone defects in rats. Systemic Scl-Ab administration improved bone regeneration and tended to increase cementogenesis measured by histology and microcomputed tomography, while Scl-Ab delivered by MSs did not result in enhancements in bone or cemental repair compared to MSs alone or control. In conclusion, systemic administration of Scl-Ab promotes bone and cemental regeneration while local, low dose delivery did not heal periodontal osseous defects in this study.

Immobilization of Denosumab on Titanium Affects Osteoclastogenesis of Human Peripheral Blood Monocytes.

Immobilization of proteins has been examined to improve implant surfaces. In this study, titanium surfaces were modified with nanofunctionalized denosumab (cDMAB), a human monoclonal anti-RANKL IgG. Noncoding DNA oligonucleotides (ODN) served as linker molecules between titanium and DMAB. Binding and release experiments demonstrated a high binding capacity of cDMAB and continuous release. Human peripheral mononuclear blood cells (PBMCs) were cultured in the presence of RANKL/MCSF for 28 days and differentiated into osteoclasts. Adding soluble DMAB to the medium inhibited osteoclast differentiation. On nanofunctionalized titanium specimens, the osteoclast-specific TRAP5b protein was monitored and showed a significantly decreased amount on cDMAB-titanium in PBMCs + RANKL/MCSF. PBMCs on cDMAB-titanium also changed SEM cell morphology. In conclusion, the results indicate that cDMAB reduces osteoclast formation and has the potential to reduce osteoclastogenesis on titanium surfaces.

Serum calcium concentration is maintained when bone resorption is suppressed by osteoprotegerin in young growing male rats.

Serum calcium (Ca) is maintained in a narrow range through regulation of Ca metabolism in the intestine, kidney, and bone. Calcium is incorporated and resorbed from bone during bone remodeling via cellular processes as well as by exchange. Both routes contribute to calcium homeostasis. To assess the magnitude of bone turnover contribution to calcium homeostasis we labeled bone with a Ca tracer and measured Ca release following stimulation or suppression of bone resorption. Young growing male rats (n = 162) were dosed with 45Ca to label skeletal Ca. After a one-month period to allow the label to incorporate into the skeleton, rats were treated with a bone resorption antagonist (OPG), a bone resorption agonist (RANKL), or vehicle control (PBS). Serum and urine 45Ca and total Ca, and serum TRACP5b (a bone resorption biomarker), were monitored for 45 days following treatment. Tracer data were analyzed by a compartmental model using WinSAAM to quantify dynamic changes in Ca metabolism and identify sites of change following treatment. In RANKL treated rats, both serum 45Ca and serum TRACP5b were increased by >70% due to a 25-fold increase in bone resorption. In OPG treated rats, both serum 45Ca and serum TRACP5b were suppressed by >70% due to a 75% decrease in bone resorption, a 3-fold increase in bone formation, and a 50% increase in absorption. Because TRACP5b and 45Ca responded similarly, we conclude that Ca release from bone into serum occurs mostly via osteoclast-mediated bone resorption. However, because serum Ca concentration did not change with altered resorption in response to either RANKL or OPG treatment, we also conclude that serum Ca concentration under normal dietary conditions in young growing male rats is maintained by processes in addition to cellular bone resorption.

Selective inhibition of receptor activator of NF-κB ligand (RANKL) in hematopoietic cells improves outcome after experimental myocardial infarction.

The RANK (receptor activator of nuclear factor κB)/RANKL (RANK ligand)/OPG (osteoprotegerin) axis is activated after myocardial infarction (MI), but its pathophysiological role is not well understood. Here, we investigated how global and cell compartment-selective inhibition of RANKL affects cardiac function and remodeling after MI in mice. Global RANKL inhibition was achieved by treatment of human RANKL knock-in (huRANKL-KI) mice with the monoclonal antibody AMG161. huRANKL-KI mice express a chimeric RANKL protein wherein part of the RANKL molecule is humanized. AMG161 inhibits human and chimeric but not murine RANKL. To dissect the pathophysiological role of RANKL derived from hematopoietic and mesenchymal cells, we selectively exchanged the hematopoietic cell compartment by lethal irradiation and across-genotype bone marrow transplantation between wild-type and huRANKL-KI mice, exploiting the specificity of AMG161. After permanent coronary artery ligation, mice were injected with AMG161 or an isotype control antibody over 4 weeks post-MI. MI increased RANKL expression mainly in cardiomyocytes and scar-infiltrating cells 4 weeks after MI. Only inhibition of RANKL derived from hematopoietic cellular sources, but not global or mesenchymal RANKL inhibition, improved post-infarct survival and cardiac function. Mechanistically, hematopoietic RANKL inhibition reduced expression of the pro-inflammatory cytokine IL-1ß in the cardiac cellular infiltrate. In conclusion, inhibition of RANKL derived from hematopoietic cellular sources is beneficial to maintain post-ischemic cardiac function by reduction of pro-inflammatory cytokine production. KEY MESSAGES: Experimental myocardial infarction (MI) augments cardiac RANKL expression in mice. RANKL expression is increased in cardiomyocytes and scar-infiltrating cells after MI. Global or mesenchymal cell RANKL inhibition has no influence on cardiac function after MI. Inhibition of RANKL derived from hematopoietic cells improves heart function post-MI. Hematopoietic RANKL inhibition reduces pro-inflammatory cytokines in scar-infiltrating cells.

Estrogen Regulates Bone Turnover by Targeting RANKL Expression in Bone Lining Cells.

Estrogen is critical for skeletal homeostasis and regulates bone remodeling, in part, by modulating the expression of receptor activator of NF-κB ligand (RANKL), an essential cytokine for bone resorption by osteoclasts. RANKL can be produced by a variety of hematopoietic (e.g. T and B-cell) and mesenchymal (osteoblast lineage, chondrocyte) cell types. The cellular mechanisms by which estrogen acts on bone are still a matter of controversy. By using murine reconstitution models that allow for selective deletion of estrogen receptor-alpha (ERα) or selective inhibition of RANKL in hematopoietic vs. mesenchymal cells, in conjunction with in situ expression profiling in bone cells, we identified bone lining cells as important gatekeepers of estrogen-controlled bone resorption. Our data indicate that the increase in bone resorption observed in states of estrogen deficiency in mice is mainly caused by lack of ERα-mediated suppression of RANKL expression in bone lining cells.

One year of abaloparatide, a selective peptide activator of the PTH1 receptor, increased bone mass and strength in ovariectomized rats.

Abaloparatide is a novel 34 amino acid peptide selected to be a potent and selective activator of the parathyroid hormone receptor 1 (PTHR1) signaling pathway. The effects of 12months of abaloparatide treatment on bone mass, bone strength and bone quality was assessed in osteopenic ovariectomized (OVX) rats. SD rats were subjected to OVX or sham surgery at 6months of age and left untreated for 3months to allow OVX-induced bone loss. Eighteen OVX rats were sacrificed after this bone depletion period, and the remaining OVX rats received daily s.c. injections of vehicle (n=18) or abaloparatide at 1, 5 or 25µg/kg/d (n=18/dose level) for 12months. Sham controls (n=18) received vehicle daily. Bone changes were assessed by DXA and pQCT after 0, 3, 6 or 12months of treatment, and destructive biomechanical testing was conducted at month 12 to assess bone strength and bone quality. Abaloparatide dose-dependently increased bone mass at the lumbar spine and at the proximal and diaphyseal regions of the tibia and femur. pQCT revealed that increased cortical bone volume at the tibia was a result of periosteal expansion and endocortical bone apposition. Abaloparatide dose-dependently increased structural strength of L4-L5 vertebral bodies, the femur diaphysis, and the femur neck. Increments in peak load for lumbar spine and the femur diaphysis of abaloparatide-treated rats persisted even after adjusting for treatment-related increments in BMC, and estimated material properties were maintained or increased at the femur diaphysis with abaloparatide. The abaloparatide groups also exhibited significant and positive correlations between bone mass and bone strength at these sites. These data indicate that gains in cortical and trabecular bone mass with abaloparatide are accompanied by and correlated with improvements in bone strength, resulting in maintenance or improvement in bone quality. Thus, this study demonstrated that long-term daily administration of abaloparatide to osteopenic OVX rats led to dose-dependent improvements in bone mass, geometry and strength.

Finite Element Analysis of Denosumab Treatment Effects on Vertebral Strength in Ovariectomized Cynomolgus Monkeys.

Finite element analysis has not yet been validated for measuring changes in whole-bone strength at the hip or spine in people after treatment with an osteoporosis agent. Toward that end, we assessed the ability of a clinically approved implementation of finite element analysis to correctly quantify treatment effects on vertebral strength, comparing against direct mechanical testing, in cynomolgus monkeys randomly assigned to one of three 16-month-long treatments: sham surgery with vehicle (Sham-Vehicle), ovariectomy with vehicle (OVX-Vehicle), or ovariectomy with denosumab (OVX-DMAb). After treatment, T12 vertebrae were retrieved, scanned with micro-CT, and mechanically tested to measure compressive strength. Blinded to the strength data and treatment codes, the micro-CT images were coarsened and homogenized to create continuum-type finite element models, without explicit porosity. With clinical translation in mind, these models were then analyzed for strength using the U.S. Food and Drug Administration (FDA)-cleared VirtuOst software application (O.N. Diagnostics, Berkeley, CA, USA), developed for analysis of human bones. We found that vertebral strength by finite element analysis was highly correlated (R(2) = 0.97; n = 52) with mechanical testing, independent of treatment (p = 0.12). Further, the size of the treatment effect on strength (ratio of mean OVX-DMAb to mean OVX-Vehicle, as a percentage) was large and did not differ (p = 0.79) between mechanical testing (+57%; 95% CI [26%, 95%]) and finite element analysis (+51% [20%, 88%]). The micro-CT analysis revealed increases in cortical thickness (+45% [19%, 73%]) and trabecular bone volume fraction (+24% [8%, 42%]). These results show that a preestablished clinical finite element analysis implementation-developed for human bone and clinically validated in fracture-outcome studies-correctly quantified the observed treatment effects of denosumab on vertebral strength in cynomolgus monkeys. One implication is that the treatment effects in this study are well explained by the features contained within these finite element models, namely, the bone geometry and mass and the spatial distribution of bone mass. © 2016 American Society for Bone and Mineral Research.

Influence of Fatigue Loading and Bone Turnover on Bone Strength and Pattern of Experimental Fractures of the Tibia in Mice.

Bone fragility depends on bone mass, structure, and material properties, including damage. The relationship between bone turnover, fatigue damage, and the pattern and location of fractures, however, remains poorly understood. We examined these factors and their integrated effects on fracture strength and patterns in tibia. Adult male mice received RANKL (2 mg/kg/day), OPG-Fc (5 mg/kg 2×/week), or vehicle (Veh) 2 days prior to fatigue loading of one tibia by in vivo axial compression, with treatments continuing up to 28 more days. One day post fatigue, crack density was similarly increased in fatigued tibiae from all treatment groups. After 28 days, the RANKL group exhibited reduced bone mass and increased crack density, resulting in reduced bone strength, while the OPG-Fc group had greater bone mass and bone strength. Injury repair altered the pattern and location of fractures created by ex vivo destructive testing, with fractures occurring more proximally and obliquely relative to non-fatigued tibia. A similar pattern was observed in both non-fatigued and fatigued tibia of RANKL. In contrast, OPG-Fc prevented this fatigue-related shift in fracture pattern by maintaining fractures more distal and transverse. Correlation analysis showed that bone strength was predominantly determined by aBMD with minor contributions from structure and intrinsic strength as measured by nanoindentation and cracks density. In contrast, fracture location was predicted equally by aBMD, crack density and intrinsic modulus. The data suggest that not only bone strength but also the fracture pattern depends on previous damage and the effects of bone turnover on bone mass and structure. These observations may be relevant to further understand the mechanisms contributing to fracture pattern in long bone with different levels of bone remodeling, including atypical femur fracture.

Combination therapy with BMP-2 and a systemic RANKL inhibitor enhances bone healing in a mouse critical-sized femoral defect.

Recombinant human BMP-2 (rhBMP-2) is a potent osteoinductive agent, but has been associated not only with bone formation, but also osteoclastogenesis and bone resorption. Osteoprotegerin (OPG) is a RANKL inhibitor that blocks differentiation and function of osteoclasts. We hypothesized that the combination of local BMP-2 (recombinant protein or a product of gene therapy) plus systemic OPG-Fc is more effective than BMP-2 alone in promoting bone repair. To test this hypothesis we used a mouse critical-sized femoral defect model. Col2.3eGFP (osteoblastic marker) male mice were treated with rhBMP-2 (group I), rhBMP-2 and systemic OPG (group II), rhBMP-2 and delayed administration of OPG (group III), mouse BM cells transduced with a lentiviral vector containing the BMP-2 gene (LV-BMP-2; group IV), LV-BMP-2 and systemic OPG (group V), a carrier alone (group VI) and administration of OPG alone (group VII). All bone defects treated with BMP-2 (alone or combined with OPG) healed, whereas minimal bone formation was noted in animals treated with the carrier alone or OPG alone. MicroCT analysis showed that bone volume (BV) in rhBMP-2+OPG and LV-BMP-2+OPG groups was significantly higher compared to rhBMP-2 alone (p<0.01) and LV-BMP-2 alone (p<0.001). Similar results were observed in histomorphometry, with rhBMP-2 alone defects exhibiting significantly lower bone area (B.Ar) compared to rhBMP-2+OPG defects (p<0.005) and LV-BMP-2 defects having a significantly lower B.Ar compared to all BMP-2+OPG treated groups (p≤0.01). TRAP staining demonstrated a major osteoclast response in the groups that did not receive OPG (rhBMP-2, LV-BMP-2 and sponge alone) beginning as early as 7days post-operatively. In conclusion, we demonstrated that locally delivered BMP-2 (recombinant protein or gene therapy) in combination with systemically administered OPG improved bone healing compared to BMP-2 alone in a mouse critical-sized bone defect. These data indicate that osteoclasts can diminish healing responses to BMP-2 and that RANKL inhibition may thus accentuate BMP-2 efficacy.

Osteoprotegerin is an effective countermeasure for spaceflight-induced bone loss in mice.

Bone loss associated with microgravity exposure poses a significant barrier to long-duration spaceflight. Osteoprotegerin-Fc (OPG-Fc) is a receptor activator of nuclear factor kappa-B ligand (RANKL) inhibitor that causes sustained inhibition of bone resorption after a single subcutaneous injection. We tested the ability of OPG-Fc to preserve bone mass during 12 days of spaceflight (SF). 64-day-old female C57BL/6J mice (n=12/group) were injected subcutaneously with OPG-Fc (20mg/kg) or an inert vehicle (VEH), 24h prior to launch. Ground control (GC) mice (VEH or OPG-Fc) were maintained under environmental conditions that mimicked those in the space shuttle middeck. Age-matched baseline (BL) controls were sacrificed at launch. GC/VEH, but not SF/VEH mice, gained tibia BMD and trabecular volume fraction (BV/TV) during the mission (P<0.05 vs. BL). SF/VEH mice had lower BV/TV vs. GC/VEH mice, while SF/OPG-Fc mice had greater BV/TV than SF/VEH or GC/VEH. SF reduced femur elastic and maximum strength in VEH mice, with OPG-Fc increasing elastic strength in SF mice. Serum TRAP5b was elevated in SF/VEH mice vs. GC/VEH mice. Conversely, SF/OPG-Fc mice had lower TRAP5b levels, suggesting that OPG-Fc preserved bone during spaceflight via inhibition of osteoclast-mediated bone resorption. Decreased bone formation also contributed to the observed osteopenia, based on the reduced femur periosteal bone formation rate and serum osteocalcin level. Overall, these observations suggest that the beneficial effects of OPG-Fc during SF are primarily due to dramatic and sustained suppression of bone resorption. In growing mice, this effect appears to compensate for the SF-related inhibition of bone formation, while preventing any SF-related increase in bone resorption. We have demonstrated that the young mouse is an appropriate new model for SF-induced osteopenia, and that a single pre-flight treatment with OPG-Fc can effectively prevent the deleterious effects of SF on mouse bone.

Hypocalcaemia in patients with metastatic bone disease treated with denosumab.

BACKGROUND: This analysis was performed to further characterise treatment-emergent hypocalcaemia in patients with bone metastases receiving denosumab. METHODS: Laboratory abnormalities and adverse events of hypocalcaemia in patients with metastatic bone disease were analysed using data from three identically designed phase 3 trials of subcutaneous denosumab 120 mg (n = 2841) versus intravenous zoledronic acid 4 mg (n = 2836). RESULTS: The overall incidence of laboratory events of hypocalcaemia grade ⩾ 2 was higher with denosumab (12.4%) than with zoledronic acid (5.3%). Hypocalcaemia events were primarily grade 2 in severity and usually occurred within the first 6 months of treatment. Patients who reported taking calcium and/or vitamin D supplements had a lower incidence of hypocalcaemia. Prostate cancer or small-cell lung cancer, reduced creatinine clearance and higher baseline bone turnover markers of urinary N-telopeptide of type I collagen (uNTx; > 50 versus ⩽ 50 nmol/mmol) and bone-specific alkaline phosphatase (BSAP; > 20.77 µg/L [median] versus ⩽ 20.77 µg/L) values were important risk factors for developing hypocalcaemia. The risk associated with increased baseline BSAP levels was greater among patients who had > 2 bone metastases at baseline versus those with ⩽ 2 bone metastases at baseline. CONCLUSION: Hypocalcaemia was more frequent with denosumab versus zoledronic acid, consistent with denosumab's greater antiresorptive effect. Low serum calcium levels and potential vitamin D deficiency should be corrected before initiating treatment with a potent osteoclast inhibitor, and corrected serum calcium levels should be monitored during treatment. Adequate calcium and vitamin D intake appears to substantially reduce the risk of hypocalcaemia.

Sustained Modeling-Based Bone Formation During Adulthood in Cynomolgus Monkeys May Contribute to Continuous BMD Gains With Denosumab.

Denosumab (DMAb) administration to postmenopausal women with osteoporosis is associated with continued bone mineral density (BMD) increases and low fracture incidence through 8 years, despite persistently reduced bone turnover markers and limited fluorochrome labeling in iliac crest bone biopsies. BMD increases were hypothesized to result from additional accrual of bone matrix via modeling-based bone formation-a hypothesis that was tested by examining fluorochrome labeling patterns in sections from ovariectomized (OVX) cynomolgus monkeys (cynos) treated with DMAb for 16 months. Mature OVX or Sham cynos were treated monthly with vehicle for 16 months, whereas other OVX cynos received monthly 25 or 50 mg/kg DMAb. DMAb groups exhibited very low serum bone resorption and formation biomarkers and near-absent fluorochrome labeling in proximal femur cancellous bone. Despite these reductions, femoral neck dual-energy X-ray absorptiometry (DXA) BMD continued to rise in DMAb-treated cynos, from a 4.6% increase at month 6 to 9.8% above baseline at month 16. Further examination of cortical bone in the proximal femur demonstrated consistent and prominent labeling on the superior endocortex and the inferior periosteal surface, typically containing multiple superimposed labels from month 6 to 16 over smooth cement lines, consistent with continuous modeling-based bone formation. These findings were evident in all groups. Quantitative analysis at another modeling site, the ninth rib, demonstrated that DMAb did not alter the surface extent of modeling-based labels, or the cortical area bound by them, relative to OVX controls, while significantly reducing remodeling-based bone formation and eroded surface. This conservation of modeling-based formation occurred concomitantly with increased femoral neck strength and, when coupled with a reduction in remodeling-based bone loss, is likely to contribute to increases in bone mass with DMAb treatment. Thus, this study provides preclinical evidence for a potential mechanism that could contribute to the clinical observations of continued BMD increases and low fracture rates with long-term DMAb administration.

Effects of denosumab, alendronate, or denosumab following alendronate on bone turnover, calcium homeostasis, bone mass and bone strength in ovariectomized cynomolgus monkeys.

Postmenopausal osteoporosis is a chronic disease wherein increased bone remodeling reduces bone mass and bone strength. Antiresorptive agents including bisphosphonates are commonly used to mitigate bone loss and fracture risk. Osteoclast inhibition via denosumab (DMAb), a RANKL inhibitor, is a newer approach for reducing fracture risk in patients at increased risk for fracture. The safety of transitioning from bisphosphonate therapy (alendronate; ALN) to DMAb was examined in mature ovariectomized (OVX) cynomolgus monkeys (cynos). One day after OVX, cynos (7-10/group) were treated with vehicle (VEH, s.c.), ALN (50 µg/kg, i.v., twice monthly) or DMAb (25 mg/kg/month, s.c.) for 12 months. Other animals received VEH or ALN for 6 months and then transitioned to 6 months of DMAb. DMAb caused significantly greater reductions in serum CTx than ALN, and transition from ALN to DMAb caused further reductions relative to continued ALN. DMAb and ALN decreased serum calcium (Ca), and transition from ALN to DMAb resulted in a lesser decline in Ca relative to DMAb or to VEH-DMAb transition. Bone histomorphometry indicated significantly reduced trabecular and cortical remodeling with DMAb or ALN. Compared with ALN, DMAb caused greater reductions in osteoclast surface, eroded surface, cortical porosity and fluorochrome labeling, and transition from ALN to DMAb reduced these parameters relative to continued ALN. Bone mineral density increased in all active treatment groups relative to VEH controls. Destructive biomechanical testing revealed significantly greater vertebral strength in all three groups receiving DMAb, including those receiving DMAb after ALN, relative to VEH controls. Bone mass and strength remained highly correlated in all groups at all tested skeletal sites, consistent with normal bone quality. These data indicate that cynos transitioned from ALN to DMAb exhibited reduced bone resorption and cortical porosity, and increased BMD and bone strength, without deleterious effects on Ca homeostasis or bone quality.

Progressive increases in bone mass and bone strength in an ovariectomized rat model of osteoporosis after 26 weeks of treatment with a sclerostin antibody.

The effects of up to 26 weeks of sclerostin antibody (Scl-Ab) treatment were investigated in ovariectomized (OVX) rats. Two months after surgery, 6-month-old osteopenic OVX rats were treated with vehicle or Scl-Ab (25 mg/kg, sc, one time per week) for 6, 12, or 26 weeks. In vivo dual-energy x-ray absorptiometry analysis demonstrated that the bone mineral density of lumbar vertebrae and femur-tibia increased progressively through 26 weeks of Scl-Ab treatment along with progressive increases in trabecular and cortical bone mass and bone strength at multiple sites. There was a strong correlation between bone mass and maximum load at lumbar vertebra, femoral neck, and diaphysis at weeks 6 and 26. Dynamic histomorphometric analysis showed that lumbar trabecular and tibial shaft endocortical and periosteal bone formation rates (BFR/BS) increased and peaked at week 6 with Scl-Ab-treatment; thereafter trabecular and endocortical BFR/BS gradually declined but remained significantly greater than OVX controls at week 26, whereas periosteal BFR/BS returned to OVX control levels at week 26. In the tibia metaphysis, trabecular BFR/BS in the Scl-Ab treated group remained elevated from week 6 to week 26. The osteoclast surface and eroded surface were significantly lower in Scl-Ab-treated rats than in OVX controls at all times. In summary, bone mass and strength increased progressively over 26 weeks of Scl-Ab treatment in adult OVX rats. The early gains were accompanied by increased cortical and trabecular bone formation and reduced osteoclast activity, whereas later gains were attributed to residual endocortical and trabecular osteoblast stimulation and persistently low osteoclast activity.