Lipid metabolism disorders, characterized by abnormal blood lipid levels, are central to the pathogenesis of obesity, nonalcoholic fatty liver disease (NAFLD), and atherosclerosis. These conditions increase the risk of type 2 diabetes, cardiovascular diseases (CVD), and stroke, highlighting the need for novel therapeutic approaches. Emerging evidence suggests a complex interplay between bone and lipid metabolism, with RANKL playing a key role. This review explores the potential of denosumab, a RANKL-targeting monoclonal antibody, in modulating lipid metabolism and its broader metabolic implications.

We conducted a comprehensive literature review to analyze the molecular mechanisms by which denosumab influences lipid metabolism, with a focus on the OPG/RANKL/RANK signaling pathway. Additionally, we examined the roles of immune modulation, bone marrow adipose tissue, and gut microbiota in metabolic diseases.

Denosumab, primarily known for its anti-resorptive effects in osteoporosis, may also exert beneficial effects on lipid metabolism. Preclinical and clinical studies suggest its potential in ameliorating obesity, NAFLD, and atherosclerosis. The OPG/RANKL/RANK axis appears to mediate crosstalk between bone and metabolic pathways, while immune regulation and gut microbiota may further contribute to these effects.

Denosumab shows promise as a therapeutic agent for lipid metabolism disorders, though long-term metabolic effects remain unclear. Further research is needed to validate its efficacy and elucidate underlying mechanisms, which could pave the way for novel treatments targeting metabolic diseases.

Osteoarthritis (OA) is a multifactorial degenerative disease involved subchondral bone remodeling, cartilage destruction and synovium inflammation. While receptor activator of nuclear factor-κB ligand (RANKL), a tumor necrosis factor (TNF) superfamily protein, is the critical regulator in bone metabolism associated with subchondral bone resorption, TNF-α is also an important inflammatory factor involved in the OA inflammation and cartilage destruction. Based on previous compound Y1599, we identified a novel tetrahydro-β-carboline derivative Y2641 with both RANKL and TNF-α inhibition in this study. Y2641 exhibited potent RANKL-induced osteoclastogenic inhibition (IC50 = 109.1 nM), and had anti-inflammatory and cartilage destruction inhibiting effects at 10 μM with low cytotoxicity. SPR assays demonstrated the binding affinity of Y2641 to RANKL (Kd = 3.984 μM) and TNF-α (Kd = 18.59 μM). In vivo assay further revealed the disease-modifying effects of Y2641 in OA rats, establishing Y2641 as a promising lead compound for the development of disease-modifying osteoarthritis drugs.

This review aimed to evaluate the effects of bisphosphonates (BP) and denosumab on dental implants, including implant failure (IF), marginal bone loss (MBL), and medication-related osteonecrosis of the jaw (MRONJ).

A literature search was conducted in four databases and OpenGrey. Studies examining IF/MBL/MBRONJ associated with BP or denosumab were included. ROBINS-I was used to assess the risk of bias. Trial sequential analysis and the GRADE approach were used to examine the certainty of evidence. Statistical analyses were conducted using R version 4.3.1.

Twenty-one studies were included. BP was associated with IF only at the implant level (RR 1.74; 95% CI: 1.10-2.75) but not at the patient level (RR 1.01; 95% CI: 0.35-2.91). The analysis of two studies indicated no significant correlation between BP and MBL (MD 0.05; 95% CI: -0.12 to 0.21). BP was associated with MRONJ (RR 3.45; 95% CI: 2.56-4.65), whereas denosumab showed no significant statistical correlation with MRONJ (RR 25.98; 95% CI: 0.31-2165.63).

In patients with dental implants, the existing very low certainty level evidence suggests that BP intake may be associated with greater risks of IF and MRONJ but not with MBL, whereas it is currently unknown whether denosumab is associated with MRONJ.

Denosumab, a RANK ligand inhibitor, is primarily used to prevent osteoclastogenesis in the treatment of conditions such as osteoporosis, bone metastasis, and giant cell tumor of bone (GCTB). RANK ligand also plays an important role in immunity by activating NF-κB and its target genes, including the osteopontin-coding gene SPP1 (also known as OPN), which is linked to CXCL9:SPP1 macrophage polarization and prognosis. In this study, we explored an additional role of denosumab in enhancing antitumor immunity in patients. Single-cell RNA sequencing was performed on nine human GCTB samples, including six untreated and three treated only with denosumab, to exclude confounding treatment factors linked with bone metastasis samples. We further analyzed paired samples collected before and after denosumab treatment from a cohort of nine patients with GCTB and conducted a pan-cancer analysis of 34 distinct types of cancers. Our single-cell analysis of GCTB resulted in a comprehensive cell atlas revealing an antitumor role of denosumab in inhibiting SPP1 expression and augmenting active cytotoxic T-cell abundance. Furthermore, we validated this immunomodulatory role of denosumab using the paired GCTB samples. Finally, the pan-cancer analysis supported a negative correlation between SPP1 and CD8A levels, with the CD8A:SPP1 ratio correlating with overall survival in 14 cancer types, which was superior to either CD8A or SPP1 alone. Our research provides clinical evidence that denosumab improves antitumor immunity by decreasing SPP1 expression and enhancing cytotoxic T-cell activity, serving as a milestone in the development of innovative use of denosumab and offering potential benefits to patients with elevated levels of SPP1.

Osteoporosis and osteopenia are common among patients undergoing posterior spine fusion surgery, presenting challenges such as pseudarthrosis, screw loosening, and poor patient outcomes. While pharmacological interventions are available, no consensus exists regarding the optimal perioperative treatment for these patients. Furthermore, the effectiveness of various treatment options in improving fusion rates and minimizing complications remains uncertain.

To compare the effects of teriparatide, bisphosphonates, denosumab, and romosozumab in patients with posterior spine fusion with low bone mineral density (BMD).

Systematic review and meta-analysis PATIENT SAMPLE: Adult patients with low BMD receiving osteoporosis medications and undergoing posterior spine fusion surgery OUTCOME MEASURES: Fusion rate, subsequent vertebral fracture (VF), screw loosening, cage subsidence, proximal junctional kyphosis(PJK), and patient-reported outcomes (PROs), particularly the Visual Analogue Scale (VAS) and Oswestry Disability Index (ODI).

A systematic search was conducted using PubMed, EMBASE, and Cochrane Library. Two reviewers independently selected and assessed relevant studies. Four groups were analyzed to evaluate the comparative effectiveness of antiosteoporosis medication on the outcome measures: Bisphosphonate versus Control; Teriparatide versus Control; Teriparatide versus Bisphosphonate; and Denosumab versus Control.

Bisphosphonate showed reduced subsequent VFs (odds ratio [OR]=0.27, 95% confidence interval [CI]=0.09-0.81) and cage subsidence (OR=0.29, 95% CI=0.11-0.75) and improved ODI scores at 12 months (standardized mean difference [SMD] [95% CI]=-0.75 [-1.42, -0.08]) compared to the control. Teriparatide showed a higher fusion rate (OR=3.52, 95% CI=1.84-6.75), lower screw loosening (OR=0.23, 95% CI=0.09-0.60), and improved ODI scores at 24 months (SMD [95% CI]=-0.57 [-0.99, -0.15]) compared to the control. Moreover, teriparatide showed a higher fusion rate (OR=2.28, 95% CI=1.67-3.11), lower subsequent VF (OR=0.22, 95% CI=0.09-0.51), and improved VAS score for back pain (VASB) (mean difference [MD] [95% CI]=-0.30 [-0.54, -0.07]) and ODI (SMD [95% CI]=-0.38[-0.64, -0.12]) scores at 12 months compared to bisphosphonate. Denosumab showed no significant difference in fusion rate or other complications compared to control.

Our results indicated that teriparatide should be used as the first-line perioperative treatment for patients with poor bone quality scheduled for posterior spine fusion. Teriparatide exhibited better fusion rates and reduced complications than controls and bisphosphonates, resulting in improved PROs. Moreover, bisphosphonates can be utilized in patients with contraindications to teriparatide since the former prevents osteoporosis-related complications compared to controls, resulting in improved PROs. Further studies are warranted to evaluate the potential effects of denosumab and romosozumab.

Both diabetes and osteoporosis are serious chronic conditions. Evidence is mounting that several bone-derived hormones play a role in glucose metabolism in patients with diabetes. Notably, novel biotargeted anti-osteoporotic agents have been recently found to reduce the risk of diabetes. This review explores the correlation of osteokines, including the receptor activator of nuclear factor-κB ligand (RANKL), sclerostin, and Dickkopf-1 (DKK1) with glycemic indicators in patients with diabetes, as well as the effects of their respective monoclonal antibodies on glucose metabolism and their possible mechanisms. Denosumab, the monoclonal antibody against RANKL, has been shown to reduce glycated hemoglobin (HbA1c) and the risk of diabetes, possibly by enhancing pancreatic β-cell survival and glucagon-like peptide-1 secretion. Sclerostin was positively correlated with HbA1c and may induce insulin resistance via endoplasmic reticulum stress. The association of DKK1 with fasting plasma glucose and HbA1c is still unclear, though decreasing DKK1 levels may correlate with β-cell survival. However, few studies have investigated the effects of antibodies against sclerostin or DKK1 on glucose metabolism. Further research is required to elucidate the influence of novel anti-osteoporotic biotargeted agents on glucose homeostasis in patients with diabetes and their underlying mechanisms.

Osteoporosis is a common inflammation-related disease in which the release of proinflammatory cytokines promotes bone loss. Oleandrin is a monomer compound extracted from the leaves of the Nerium oleander plant, has been shown to exert an anti-inflammatory effect on a variety of inflammation-related diseases. However, its role in osteoporosis and the underlying mechanisms remain unclear. In this study, Oleandrin was shown to reduce bone loss in ovariectomy-induced osteoporotic mice in vivo. Additionally, Oleandrin inhibited RANKL-induced osteoclast differentiation in a concentration-dependent manner in vitro. Signalling pathway studies showed that Oleandrin could inhibit osteoclast differentiation by targeting MAPK and NF-κB signalling pathways. Further mechanistic studies showed that Oleandrin binds to low-density lipoprotein receptor-related protein 4 in osteoclast, thereby exerting inhibitory effects on osteoclast differentiation. In conclusion, this study lays the foundation for further research on the anti-inflammatory and anti-osteoporotic effects of Oleandrin on osteoporosis and its underlying mechanism and provides new possibilities for the treatment of osteoporosis.

Musculoskeletal disorders, affecting as many as 1.3 billion people worldwide, are the leading cause of disability and impose a substantial health and socioeconomic burden. Despite the high prevalence of these conditions, translational research in this field is far from optimal, highlighting the need for stronger collaboration between basic and clinical scientists. This paper, authored by members of the basic and clinical action groups of the European Calcified Tissue Society (ECTS) and endorsed by the Board of the ECTS, examines the key barriers to effective translational research in musculoskeletal diseases, including clinician workload, differences in professional language and culture, physical distance between research sites, and insufficient interdisciplinary funding. Through interviews with eight institutional managers across five European countries, we observed that in some institutions, the collaboration between basic scientists and clinicians was regarded as no concern (but with room for improvement), and in most institutions it was recognised as a serious issue. We found consensus on the importance of collaboration yet identified discrepancies in the provision of structural and financial support. Based on these findings, we propose strategic initiatives to bridge the gap between basic and clinical research. Suggested measures include dedicated translational funding, integrated research facilities, collaborative scientific forums, strategic collaborations, establishment of physician-scientists, and, finally, bringing basic and clinical researchers together in the same building or even in a combined department. Notable successes, such as the development of the anti-osteoporotic drugs, romosozumab and denosumab, underscore the value of a coordinated approach and exemplify how shared insights between laboratory research and clinical practice can lead to impactful therapeutic advances. Moving forward, we advocate for institutional commitments to foster a robust translational research environment, as well as tailored funding initiatives to support such efforts. This paper serves as a call for discussion and action to enhance interdisciplinary cooperation to advance musculoskeletal medicine and improve outcomes for patients with debilitating musculoskeletal diseases.

Bone remodeling is regulated by the interaction between receptor activator of nuclear factor kappa-B ligand (RANKL) and its receptor RANK on osteoblasts and osteoclasts, respectively. Osteoprotegerin (OPG) is secreted from osteoblasts and inhibits osteoclast differentiation by acting as a decoy receptor for RANKL. Despite its importance, the mechanism underlying the secretion of OPG remains poorly understood. Here, we applied a method of video-rate bioluminescence imaging using a fusion protein with Gaussia luciferase (GLase) and visualized the secretion of OPG from living mouse osteoblastic MC3T3-E1 cells. The bioluminescence imaging revealed that the secretion of OPG fused to GLase (OPG-GLase) occurred frequently and widely across the cell surface. Notably, co-expression of RANKL significantly reduced the secretion of OPG-GLase, indicating an inhibitory role of RANKL on OPG secretion within cells. Further imaging and biochemical analyses using deletion mutants of OPG and RANKL, as well as RANKL mutants that cause autosomal recessive osteopetrosis, demonstrated the essential role of protein-protein interaction between OPG and RANKL in the inhibition of OPG secretion. Treatment with proteasome inhibitors resulted in increased levels of OPG in both culture medium and cell lysates. However, the fold-increase of OPG was similar regardless of the presence or absence of RANKL, suggesting that the regulation of OPG secretion by RANKL is independent of proteasome activity. This report visualized the secretion of OPG from living cells and provided evidence for a novel intracellular inhibitory effect of RANKL on OPG secretion.

The teleost scale is a unique calcified tissue that contains osteoclasts, osteoblasts, osteocytes and the bone matrix, similar to mammalian bone. Here, the effects of low-intensity pulsed ultrasound (LIPUS) on osteoblasts and osteoclasts in goldfish scales were investigated. Scales were treated with LIPUS, which is equivalent to use under clinical conditions (30 mW/cm2 for 20 min), then cultured at 15˚C. Alkaline phosphatase activity, a marker of osteoblasts, or tartrate-resistant acid phosphatase (TRAP) activity, a marker of osteoclasts was measured. The gene expression profile was examined using RNA-sequencing. Gene network and biological function analyses were performed using the Ingenuity® Pathways Knowledge Base. A single exposure of LIPUS significantly increased ALP activity but did not affect TRAP activity. These data indicated that LIPUS induced osteoblastic activation in goldfish scales. Using RNA-sequencing, numerous genes that were significantly and differentially expressed 3, 6, and 24 h after LIPUS exposure were observed. Ingenuity® pathway analysis demonstrated that three gene networks, GN-3h, GN-6h, and GN-24h, were obtained from upregulated genes at 3, 6 and 24 h culture, respectively, and included several genes associated with osteoblast differentiation, such as protein kinase D1, prostaglandin-endoperoxide synthase 2, TNFRSF11B (tumor necrosis factor receptor superfamily, member 11b) and WNT3A (Wnt family member 3A). A significant upregulation of expression levels of these genes in scales treated with LIPUS was confirmed by reverse transcription-quantitative polymerase chain reaction. These results contribute to elucidating the molecular mechanisms of osteoblast activation induced by LIPUS.

RANK pathway has attracted increasing interest as a promising target in breast cancer, given the availability of denosumab, an anti-RANKL drug. RANK signaling mediates progesterone-driven regulation of mammary gland development and favors breast cancer initiation by controlling mammary cell proliferation and stem cell fate. RANK activation promotes luminal mammary epithelial cell senescence, acting as an initial barrier to tumorigenesis but ultimately facilitating tumor progression and metastasis. Comprehensive analyses have demonstrated that RANK protein expression is an independent biomarker of poor prognosis in postmenopausal and estrogen receptor-negative breast cancer patients. RANK pathway also has multiple roles in immunity and inflammation, regulating innate and adaptive responses. In the tumor microenvironment, RANK and RANKL are expressed by different immune cell populations and contribute to the regulation of tumor immune surveillance, mainly driving immunosuppressive effects.Herein, we discuss the preventive and therapeutic potential of targeting RANK signaling in breast cancer given its tumor cell intrinsic and extrinsic effects. RANKL inhibition has been shown to induce mammary tumor cell differentiation and an antitumor immune response. Moreover, loss of RANK signaling increases sensitivity of breast cancer cells to chemotherapy, targeted therapies such as HER2 and CDK4/6 inhibitors, and immunotherapy. Finally, we describe clinical trials of denosumab for breast cancer prevention, such as those ongoing in women with high risk of developing breast cancer, large phase III clinical trials where the impact of adjuvant denosumab on disease-free survival has been assessed, and window trials to evaluate the immunomodulatory effects of denosumab in breast cancer and other solid tumors.

The 24th Annual Santa Fe Bone Symposium (SFBS) was held in Santa Fe, New Mexico, USA, on August 2-3, 2024. This was a "hybrid" meeting, with in-person and real-time remote participants representing a broad range of geographical locations and medical disciplines. The focus was on new developments in the care of patients with osteoporosis, other metabolic bone diseases, and inherited skeletal disorders. The most current medical evidence was presented and discussed with consideration of implications for patient management. Topics included an update on clinical uses of osteoanabolic agents, management of patients discontinuing denosumab, bone health optimization for orthopedic surgery, estrogen and testosterone in the management of osteoporosis, osteoporosis treatment in the very old, overview of rare bone diseases, treat-to-target for osteoporosis, and a progress report on global activities of Bone Health ECHO. There were two highly interactive faculty panel discussions - one with case presentations by attendees and another with open microphone for all topics of interest. Endocrinology fellows, selected from attendees of the Santa Fe Fellows Workshop on Metabolic Bone Diseases, held the two days preceding the SFBS, participated with presentations of oral abstracts. Ancillary events addressed modern approaches to menopause and bone health, case studies of management of patients at very high fracture risk, and management of patients with rare bone diseases, such as hypophosphatasia, fibrodysplasia ossificans progressiva, X-linked hypophosphatemia, and hypoparathyroidism. These proceedings of the SFBS present the clinical highlights of the plenary sessions and the discussions that followed.

Osteosarcopenia (OS) is a geriatric syndrome characterized by the concurrent presence of osteoporosis and sarcopenia, predominantly affecting the elderly population. Osteoporosis (OP) is a systemic skeletal disorder characterized by decreased bone mass, compromised bone microarchitecture, and heightened bone fragility, substantially elevating fracture risk. Sarcopenia (SP) is defined by decreased muscle mass, strength, and/or functional capacity. Both conditions are age-related degenerative diseases with overlapping pathophysiological mechanisms, commonly co-occurring in elderly individuals and substantially increasing fracture risk. Denosumab, a targeted anti-osteoporotic agent, mediates therapeutic effects by inhibiting bone resorption through the RANK-RANKL-OPG (RRO) pathway, consequently enhancing bone mineral density. International studies indicate that Denosumab not only treats osteoporosis but also improves sarcopenia-related metrics, suggesting its potential as a sarcopenia treatment. However, research focusing on the Chinese population remains limited. Additionally, the pathophysiological mechanisms of sarcopenia and the pathways through which Denosumab ameliorates sarcopenia are not yet fully understood, warranting further experimental investigation. In summary, Denosumab's therapeutic efficacy in osteoporosis treatment and its potential impact on sarcopenia are of substantial research interest. However, research and literature on these topics in China remain notably scarce. This article aims to offer a systematic review and critical analysis of these topics.

Osteoclasts are the sole bone-resorbing cells and are formed by the fusion of osteoclast precursor cells (OCPs) derived from myeloid lineage cells. Animal studies reveal that circulating OCPs (cOCPs) in blood travel to bone and fuse with bone-resident osteoclasts. However, the characteristics of human cOCPs and their association with bone diseases remain elusive. We have identified and characterized human cOCPs and found a positive association between cOCPs and osteoclast activity. Sorted cOCPs have a higher osteoclastogenic potential than other myeloid cells and effectively differentiate into osteoclasts. cOCPs exhibit distinct morphology and transcriptomic signatures. The frequency of cOCPs in the blood varies among treatment-naive postmenopausal women and has an inverse correlation with lumbar spine bone density and a positive correlation with serum CTX, a bone resorption marker. The increased cOCPs in treatment-naive patients with osteoporosis were significantly diminished by denosumab, a widely used antiresorptive therapy. Our study reveals the distinctive identity of human cOCPs and the potential link between the dynamic regulation of cOCPs and osteoporosis and its treatment. Taken together, our study enhances our understanding of human cOCPs and highlights a potential opportunity to measure cOCPs through a simple blood test, which could potentially identify high-risk individuals.

Osteoporosis and aortic valve calcification, prevalent in the elderly, have unclear common mechanisms. This study aims to uncover them through bioinformatics analysis.

Microarray data from GEO was analyzed for osteoporosis and aortic valve calcification. Differential expression analysis identified co-expressed genes. SVM-RFE and random forest selected key genes. GO and KEGG enrichment analyses were performed. Immunoinfiltration and GSEA analyses were subsequently performed. NetworkAnalyst analyzed microRNAs/TFs. HERB predicted drugs, and molecular docking assessed targeting potential.

Thirteen genes linked to osteoporosis and aortic valve calcification were identified. TNFSF11, KYNU, and HLA-DMB emerged as key genes. miRNAs, TFs, and drug predictions offered therapeutic insights. Molecular docking suggested 17-beta-estradiol and vitamin D3 as potential treatments.

The study clarifies shared mechanisms of osteoporosis and aortic valve calcification, identifies biomarkers, and highlights TNFSF11, KYNU, and HLA-DMB. It also suggests 17-beta-estradiol and vitamin D3 as potential effective treatments.

Denosumab is a potent antiresorptive medication, commonly used in the treatment of osteoporosis, as well as in a variety of other diseases. Potential adverse rebound effects after its cessation include a loss in bone mineral density and an increased risk of osteoporotic fractures. Hypercalcemia is a less frequently reported rebound phenomenon after denosumab discontinuation, that may pose a diagnostic challenge to physicians as a rare non-parathyroid hormone (PTH) dependent cause of hypercalcemia. In our case, a 47-year-old male presented with rebound hypercalcemia after denosumab cessation during follow-up after surgical treatment for parathyroid carcinoma. This non-PTH-dependent hypercalcemia resolved after re-initiation of denosumab. We performed a systematic literature review on rebound hypercalcemia after denosumab cessation and identified 52 individual patient cases. Children appear to be more prone to developing rebound hypercalcemia, which could be attributed to their higher baseline bone turnover, underlying conditions, or denosumab dosage regimens. In most cases, patients initially presented with acute and often severe symptoms of hypercalcemia that occur from 1.75 to 9 months after denosumab cessation (4 to 9 months in adults). Most effective treatment approaches to sufficiently decrease serum calcium levels were bisphosphonates or re-administration of denosumab. A watch and wait strategy may be sufficient in asymptomatic cases, which are less common and probably underdiagnosed. Subsequent antiresorptive treatment after denosumab cessation, which is a common practice in osteoporosis treatment, may reduce the risk of rebound hypercalcemia. As denosumab is a frequently used drug in patients with advanced malignant diseases and rebound hypercalcemia with low PTH levels may raise the suspicion for skeletal metastases, awareness of this rebound effect may be for particular relevance in such settings.

Chikungunya fever (CHIKF), caused by the Chikungunya virus (CHIKV), manifests as acute febrile illness often associated with polyarthritis and polyarthralgia. Although the acute symptoms resolve within two weeks, many patients experience prolonged joint pain and inflammation, resembling rheumatoid arthritis (RA). This study aimed to identify molecular markers related to joint pain and chronicity in CHIKV-infected individuals by analyzing blood transcriptomes using bulk RNA sequencing. B- and T-cell receptor (BCR and TCR) diversity was assessed through computational analysis of RNA-seq data, revealing a significant reduction in CDR3 diversity in CHIKV-infected individuals compared to healthy controls. This reduced diversity was associated with the upregulation of genes involved in osteoclast differentiation and activation, particularly through the RANK/RANKL signaling pathway. These findings suggest a potential link between immune dysregulation and enhanced osteoclast activity, which may contribute to the persistence of joint pain in chronic CHIKF. Targeting osteoclast-related pathways could offer therapeutic strategies for managing chronic symptoms in CHIKF patients.

Receptor activator of nuclear factor-κB ligand (RANKL) is considered the principal regulator of osteoclast differentiation. Therefore, strategies interfering with the RANKL-RANK signaling pathway may effectively inhibit osteoclast differentiation and mitigate bone resorption. Consequently, RANKL has become a promising target for new drug design strategies. Despite extensive research on specific drugs and antibodies, only a few have shown efficacy in treating osteoporosis. To address this challenge, we aimed to explore new approaches for designing drugs for osteoporosis. In this study, a 3D quantitative structure-activity relationship (QSAR) pharmacophore model was built for RANKL with reference to known inhibitor IC50 values. The optimal pharmacophore model was then employed as a 3D query to screen databases for novel lead compounds. The obtained compounds were subjected to ADMET and TOPKAT analyses to predict drug pharmacokinetics and toxicity. Molecular docking and de novo evolution approaches were applied to verify the docking binding affinities of the compounds. Five candidate compounds were subjected to further in vitro analyses to assess their anti-osteoporotic effects, among which compound 4 demonstrated significant inhibitory activity, achieving an inhibitory rate of 92.6 % on osteoclastogenesis at a concentration of 10 μM. Subsequent molecular dynamics (MD) simulations to assess the stability and behavior of compound 4 and its evolved variant, ZINC00059014397_Evo, within the RANKL binding site revealed that the variant is a potential therapeutic agent for targeting osteoclasts. This study offers valuable insights for developing next generation RANKL inhibitors for osteoporosis treatments.

After denosumab (Dmab) discontinuation C-terminal telopeptide (CTX) levels increase, bone mineral density (BMD) decreases and multiple vertebral fractures (FX) may occur with relevant impacts on women's health. A sequential therapy with bisphosphonates is recommended, and the European Calcified Tissue Society (ECTS) proposed repeated zoledronate (ZOL) administrations in patients with persistently high CTX levels, although the efficacy of this schedule is unknown. In this retrospective study, we describe BMD changes and FX rate in 52 patients managed according to the ECTS recommendations.

We measured CTX levels and administered ZOL after 1 month from Dmab withdrawal (t0). After 6 months (t1), we administered a second ZOL infusion, if CTX levels were ≥280 ng/L. BMD changes and FX rate were assessed on average after 17 months from Dmab withdrawal.

Seventy-five percent of patients repeated ZOL infusion. In this group, spine BMD declined significantly (-5.5 ± 5.6%), while it remained stable in the group with CTX levels <280 ng/L (-0.1 ± 5.5%, P = 0.008). All fractured patients (9.6%) had received >5 Dmab injections and 2 ZOL infusions. The BMD worsening after Dmab withdrawal was associated with CTX t1 [odds ratio (OR) 2.9, interquartile range (IQR) 1.3-6.6, P = .009] and spine BMD gain during Dmab therapy corrected for the number of Dmab injections (OR 3.0, IQR 1.2-7.2, P = .014). A CTX level at t1 > 212 ng/L had 100% sensitivity in predicting the BMD loss.

In patients with uncontrolled CTX levels after Dmab withdrawal, 2 ZOL infusions 6 months apart do not prevent BMD loss and FX.

Osteoprotegerin (OPG) plays an important role in the inhibition of osteoclast formation and bone resorption. Studies have reported lower OPG levels among women with a pathogenic variant (mutation) in the BRCA1 gene, and thus, may be at greater risk for skeletal bone loss. Thus, we investigated the association between circulating OPG and two validated markers of bone health: 1) bone fracture risk score (FRAX) and 2) bone mineral density (BMD), among BRCA mutation carriers.

Women with a blood sample and clinical data were included in this analysis. An enzyme-linked immunosorbent assay (ELISA) was used to quantify serum OPG (pg/mL) and the 10-year risk of major osteoporotic fracture (FRAXmajor) and hip fracture (FRAXhip) (%) was estimated using a web-based algorithm. For a subset of women, lumbar spine BMD was previously assessed by dual x-ray absorptiometry (DXA)(T-score). A Mann-Whitney U test was used to evaluate the association between OPG and FRAX score, while linear regression was used to assess the association of OPG and BMD.

Among 701 women with a BRCA1 mutation, there was a significant (and unexpected) positive association between OPG levels and FRAX score (FRAXmajor: 2.12 (low OPG) vs. 2.53 (high OPG) P < 0.0001; FRAXhip: 0.27 (low OPG) vs. 0.44 (high OPG) P < 0.0001). In a subset with BMD measurement (n = 50), low serum OPG was associated with a significantly lower BMD T-score (-1.069 vs. -0.318; P = 0.04).

Our findings suggest that women with inherently lower OPG may be at risk of lower BMD, the gold standard marker of bone disease. Due to the young age of our cohort, on-going studies are warranted to re-evaluate the association between OPG and FRAX in BRCA mutation carriers.

During the progression of proliferative vitreoretinopathy (PVR) following ocular trauma, previously quiescent retinal pigment epithelial (RPE) cells transition into a state of rapid proliferation, migration, and secretion. The elusive molecular mechanisms behind these changes have hindered the development of effective pharmacological treatments, presenting a pressing clinical challenge. In this study, by monitoring the dynamic changes in chromatin accessibility and various histone modifications, we chart the comprehensive epigenetic landscape of RPE cells in male mice subjected to traumatic PVR. Coupled with transcriptomic analysis, we reveal a robust correlation between enhancer activation and the upregulation of the PVR-associated gene programs. Furthermore, by constructing transcription factor regulatory networks, we identify the aberrant activation of enhancer-driven RANK-NFATc1 pathway as PVR advanced. Importantly, we demonstrate that intraocular interventions, including nanomedicines inhibiting enhancer activity, gene therapies targeting NFATc1 and antibody therapeutics against RANK pathway, effectively mitigate PVR progression. Together, our findings elucidate the epigenetic basis underlying the activation of PVR-associated genes during RPE cell fate transitions and offer promising therapeutic avenues targeting epigenetic modulation and the RANK-NFATc1 axis for PVR management.

Breast cancer (BC) is the most prevalent cancer and the leading cause of death among women worldwide. The osteoprotegerin (OPG) cytokine, a decoy receptor for RANKL and a key player in bone homeostasis, has pro-and anti-carcinogenic effects in various types of cancer, including breast neoplasms. In the present study, we have shown that ectopic expression of OPG in breast epithelial/cancer cells promotes the pro-metastatic processes epithelial-to-mesenchymal transition (EMT), stemness, angiogenesis as well as the activation of breast stromal fibroblasts. Furthermore, proteomics analysis, which allows the identification and quantification of a plethora of known and unknown proteins, has shown a strong and significant correlation between OPG upregulation and the expression of proteins with functions in EMT and stemness. On the other hand, OPG knockdown in triple-negative breast cancer (TNBC) cells inhibited the formation of cancer stem cells. Importantly, while OPG upregulation significantly enhanced the resistance of luminal BC cells to cisplatin and docetaxel, OPG downregulation sensitized TNBC cells to these chemotherapeutic drugs. We have also shown that OPG negatively controls estrogen receptor α (ERα), and OPG upregulation correlated well with the expression of genes related to ER-negative claudin low cells. Collectively, these results show that OPG promotes stemness and the consequent chemoresistance of breast cancer cells.

T-2 toxin, the most toxic type A trichothecene, is widely present in grain and animal feed, causing growth retardation and tissue damage in poultry. Geese are more sensitive to T-2 toxin than chickens and ducks. Although T-2 toxin has been reported to cause tibial growth plate (TGP) chondrodysplasia in chickens, tibial damage caused by T-2 toxin in geese has not been fully demonstrated. This study aims to investigate the adverse effects of T-2 toxin on tibial bone development, bone quality, chondrocyte differentiation, and bone metabolism. Here, forty-eight one-day-old male Yangzhou goslings were randomly divided into four groups and daily gavaged with T-2 toxin at concentrations of 0, 0.5, 1.0, and 2.0 mg/kg body weight for 21 days, respectively. The development of gosling body weight and size was determined by weighing and taking body measurements after exposure to different concentrations of T-2 toxin. Changes in tibial development and bone characteristics were determined by radiographic examination, phenotypic measurements, and bone quality and composition analyses. Chondrocyte differentiation in TGP and bone metabolism was characterized by cell morphology, tissue gene-specific expression, and serum marker levels. Results showed that T-2 toxin treatment resulted in a lower weight, volume, length, middle width, and middle circumference of the tibia in a dose-dependent manner (p < 0.05). Moreover, decreased bone-breaking strength, bone mineral density, and contents of ash, Ca, and P in the tibia were observed in T-2 toxin-challenged goslings (p < 0.05). In addition, T-2 toxin not only reduced TGP height (p < 0.05) but also induced TGP chondrocytes to be disorganized with reduced numbers and indistinct borders. As expected, the apoptosis-related genes (CASP9 and CASP3) were significantly up-regulated in chondrocytes challenged by T-2 toxin with a dose dependence, while cell differentiation and maturation-related genes (BMP6, BMP7, SOX9, and RUNX2) were down-regulated (p < 0.05). Considering bone metabolism, T-2 toxin dose-dependently and significantly induced a decreased number of osteoblasts and an increased number of osteoclasts in the tibia, with inhibited patterns of osteogenesis-related genes and enzymes and increased patterns of osteoclast-related genes and enzymes (p < 0.05). Similarly, the serum Ca and P concentrations and parathyroid hormone, calcitonin, and 1, 25-dihydroxycholecalciferol levels decreased under T-2 toxin exposure (p < 0.05). In summary, 2.0 mg/kg T-2 toxin significantly inhibited tibia weight, length, width, and circumference, as well as decreased bone-breaking strength, density, and composition (ash, calcium, and phosphorus) in 21-day-old goslings compared to the control and lower dose groups. Chondrocyte differentiation in TGP was delayed by 2.0 mg/kg T-2 toxin owing to cell apoptosis. In addition, 2.0 mg/kg T-2 toxin promoted bone resorption and inhibited osteogenesis in cellular morphology, gene expression, and hormonal modulation patterns. Thus, T-2 toxin significantly inhibited tibial growth and development with a dose dependence, accompanied by decreased bone geometry parameters and properties, hindered chondrocyte differentiation, and imbalanced bone metabolism.

Alzheimer's disease (AD) and osteoporosis are two diseases that mainly affect elderly people, with increases in the occurrence of cases due to a longer life expectancy. Several epidemiological studies have shown a reciprocal association between both diseases, finding an increase in incidence of osteoporosis in patients with AD, and a higher burden of AD in osteoporotic patients. This epidemiological relationship has motivated the search for molecules, genes, signaling pathways and mechanisms that are related to both pathologies. The mechanisms found in these studies can serve to improve treatments and establish better patient care protocols.

Denosumab and bisphosphonates for primary osteoporosis are generally well-tolerated, but their comparative safety remains unclear. We aimed to explore the comparative safety of denosumab and bisphosphonates in primary osteoporosis. Databases such as PubMed and Google Scholar were searched for relevant peer-reviewed randomized controlled trials published in English (as of December 2023). Trials comparing adverse events (AE) between denosumab and bisphosphonates in patients with primary osteoporosis were investigated. Data were pooled using a fixed- or random-effects model to determine the risk ratios (RR) and 95% confidence intervals (CIs) for various AEs in patients treated with denosumab in comparison to patients treated with bisphosphonates. Eleven trials (5,545 patients; follow-up period: 12-24 months) were included in this meta-analysis. All trials had a risk of bias (e.g., reporting bias linked to secondary endpoints and selection bias linked to random allocation). In comparison to bisphosphonates, denosumab was significantly associated with less withdrawal due to AEs (RR = 0.49; 95% CI 0.34-0.71), more five-point major adverse cardiovascular events (RR = 2.05; 95% CI 1.03-4.09), more cardiovascular AEs (RR = 1.61; 95% CI 1.07-2.41), more infections (RR = 1.14; 95% CI 1.02-1.27), more upper respiratory tract infections (RR = 1.56; 95% CI 1.08-2.25), less vertebral fractures (RR = 0.54; 95% CI 0.31-0.93), and less abdominal pain (RR = 0.44;95% CI 0.22-0.87). We explored the comparative safety of denosumab and bisphosphonates for primary osteoporosis, some of which could be attributed to their beneficial effects. However, all trials had a risk of bias. Further investigations are required to confirm our results.

Antiresorptive medications do not negatively affect fracture healing in humans. Teriparatide may decrease time to fracture healing. Romosozumab has not shown a beneficial effect on human fracture healing.

Fracture healing is a complex process. Uncertainty exists over the influence of osteoporosis and the medications used to treat it on fracture healing.

Narrative review authored by the members of the Fracture Working Group of the Committee of Scientific Advisors of the International Osteoporosis Foundation (IOF), on behalf of the IOF and the Société Internationale de Chirurgie Orthopédique et de Traumatologie (SICOT).

Fracture healing is a multistep process. Most fractures heal through a combination of intramembranous and endochondral ossification. Radiographic imaging is important for evaluating fracture healing and for detecting delayed or non-union. The presence of callus formation, bridging trabeculae, and a decrease in the size of the fracture line over time are indicative of healing. Imaging must be combined with clinical parameters and patient-reported outcomes. Animal data support a negative effect of osteoporosis on fracture healing; however, clinical data do not appear to corroborate with this. Evidence does not support a delay in the initiation of antiresorptive therapy following acute fragility fractures. There is no reason for suspension of osteoporosis medication at the time of fracture if the person is already on treatment. Teriparatide treatment may shorten fracture healing time at certain sites such as distal radius; however, it does not prevent non-union or influence union rate. The positive effect on fracture healing that romosozumab has demonstrated in animals has not been observed in humans.

Overall, there appears to be no deleterious effect of osteoporosis medications on fracture healing. The benefit of treating osteoporosis and the urgent necessity to mitigate imminent refracture risk after a fracture should be given prime consideration. It is imperative that new radiological and biological markers of fracture healing be identified. It is also important to synthesize clinical and basic science methodologies to assess fracture healing, so that a convergence of the two frameworks can be achieved.

Adipogenesis, the process of white adipose tissue expansion, plays a critical role in the development of obesity. Osteoprotegerin (OPG), known for its role in bone metabolism regulation, emerges as a potential regulator in mediating adipogenesis during obesity onset.

This study aims to elucidate the involvement of OPG in adipogenesis during the early phases of diet-induced obesity and explore its therapeutic potential in obesity management.

Using a diet-induced obesity model, we investigated OPG expression patterns in adipocytes and explored the mechanisms underlying its involvement in adipogenesis. We also assessed the effects of targeted silencing of OPG and recombinant OPG administration on obesity progression and insulin resistance. Additionally, the impact of electroacupuncture treatment on OPG levels and obesity management was evaluated in both animal models and human participants.

OPG expression was prominently activated in adipocytes of white adipose tissues during the early phase of diet-induced obesity. Hyperlipidemia induced Cbfa1-dependent OPG transcription, initiating and promoting adipogenesis, leading to cell-size expansion and lipid storage. Intracellular OPG physically bound to RAR and released the PPARɤ/RXR complex, activating adipogenesis-associated gene expression. Targeted silencing of OPG suppressed obesity development, while recombinant OPG administration promoted disease progression and insulin resistance in obese mice. Electroacupuncture treatment suppressed obesity development in an OPG-dependent manner and improved obesity parameters in obese human participants.

OPG emerges as a key regulator in mediating adipogenesis during obesity development. Targeting OPG holds promise for the prevention and treatment of obesity, as evidenced by the efficacy of electroacupuncture treatment in modulating OPG levels and managing obesity-related outcomes.

As a highly dynamic tissue, bone is continuously rebuilt throughout life. Both bone formation by osteoblasts and bone resorption by osteoclasts constitute bone reconstruction homeostasis. The equilibrium of bone homeostasis is governed by many complicated signaling pathways that weave together to form an intricate network. These pathways coordinate the meticulous processes of bone formation and resorption, ensuring the structural integrity and dynamic vitality of the skeletal system. Dysregulation of the bone homeostatic regulatory signaling network contributes to the development and progression of many skeletal diseases. Significantly, imbalanced bone homeostasis further disrupts the signaling network and triggers a cascade reaction that exacerbates disease progression and engenders a deleterious cycle. Here, we summarize the influence of signaling pathways on bone homeostasis, elucidating the interplay and crosstalk among them. Additionally, we review the mechanisms underpinning bone homeostatic imbalances across diverse disease landscapes, highlighting current and prospective therapeutic targets and clinical drugs. We hope that this review will contribute to a holistic understanding of the signaling pathways and molecular mechanisms sustaining bone homeostasis, which are promising to contribute to further research on bone homeostasis and shed light on the development of targeted drugs.

Osteomyelitis (OM) is a major challenge in orthopedic surgery. The diagnosis of OM is based on imaging and laboratory tests, but it still presents some limitations. Therefore, a deeper comprehension of the pathogenetic mechanisms could enhance diagnostic and treatment approaches. OM pathogenesis is based on an inflammatory response to pathogen infection, leading to bone loss. The present study aims to investigate the potential diagnostic role of a panel of osteoimmunological serum biomarkers in the clinical approach to OM. The focus is on the emerging infection biomarker sCD14-ST, along with osteoimmunological and inflammatory serum biomarkers, to define a comprehensive biomarker panel for a multifaced approach to OM. The results, to our knowledge, demonstrate for the first time the diagnostic and early prognostic role of sCD14-ST in OM patients, suggesting that this biomarker could address the limitations of current laboratory tests, such as traditional inflammatory markers, in diagnosing OM. In addition, the study highlights a relevant diagnostic role of SuPAR, the chemokine CCL2, the anti-inflammatory cytokine IL-10, the Wnt inhibitors DKK-1 and Sclerostin, and the RANKL/OPG ratio. Moreover, CCL2 and SuPAR also exhibited early prognostic value.

Preclinical models (typically ovariectomized rats and genetically altered mice) have underpinned much of what we know about skeletal biology. They have been pivotal for developing therapies for osteoporosis and monogenic skeletal conditions, including osteogenesis imperfecta, achondroplasia, hypophosphatasia, and craniodysplasias. Further therapeutic advances, particularly to improve cortical strength, require improved understanding and more rigorous use and reporting. We describe here how trabecular and cortical bone structure develop, are maintained, and degenerate with aging in mice, rats, and humans, and how cortical bone structure is changed in some preclinical models of endocrine conditions (eg, postmenopausal osteoporosis, chronic kidney disease, hyperparathyroidism, diabetes). We provide examples of preclinical models used to identify and test current therapies for osteoporosis, and discuss common concerns raised when comparing rodent preclinical models to the human skeleton. We focus especially on cortical bone, because it differs between small and larger mammals in its organizational structure. We discuss mechanisms common to mouse and human controlling cortical bone strength and structure, including recent examples revealing genetic contributors to cortical porosity and osteocyte network configurations during growth, maturity, and aging. We conclude with guidelines for clear reporting on mouse models with a goal for better consistency in the use and interpretation of these models.

Cortactin (CTTN), a cytoskeletal protein and substrate of Src kinase, is implicated in tumor aggressiveness. However, its role in bone cell differentiation remains unknown. The current study revealed that CTTN was upregulated during osteoblast and adipocyte differentiation. Functional experiments demonstrated that CTTN promoted the in vitro differentiation of mesenchymal stem/progenitor cells into osteogenic and adipogenic lineages. Mechanistically, CTTN was able to stabilize the protein level of mechanistic target of rapamycin kinase (mTOR), leading to the activation of mTOR signaling. In-depth investigation revealed that CTTN could bind with casitas B lineage lymphoma-c (c-CBL) and counteract the function of c-CBL, a known E3 ubiquitin ligase responsible for the proteasomal degradation of mTOR. Silencing c-Cbl alleviated the impaired differentiation of osteoblasts and adipocytes caused by CTTN siRNA, while silencing mTOR mitigated the stimulation of osteoblast and adipocyte differentiation induced by CTTN overexpression. Notably, transplantation of CTTN-silenced bone marrow stromal cells (BMSCs) into the marrow of mice led to a reduction in trabecular bone mass, accompanied by a decrease in osteoblasts and an increase in osteoclasts. Furthermore, CTTN-silenced BMSCs expressed higher levels of receptor activator of nuclear factor κB ligand (RANKL) than control BMSCs did and promoted osteoclast differentiation when cocultured with bone marrow-derived osteoclast precursor cells. This study provides evidence that CTTN favors osteoblast differentiation by counteracting the c-CBL-induced degradation of mTOR and inhibits osteoclast differentiation by downregulating the expression of RANKL. It also suggests that maintaining an appropriate level of CTTN expression may be advantageous for maintaining bone homeostasis.

Denosumab is a potential therapeutic agent for osteogenesis imperfecta (OI), but its efficacy and safety remain unclear in children with OI.

We aimed to investigate the effects of denosumab on bone mineral density (BMD), spinal morphometry, and safety in children with OI compared with zoledronic acid.

In this prospective study, 84 children or adolescents with OI were randomized to receive denosumab subcutaneous injection every 6 months or zoledronic acid intravenous infusion once. Changes of BMD and its Z-score, vertebral shape, serum levels of calcium and bone turnover biomarkers were assessed during the 1-year treatment.

After 12 months of treatment, BMD at the lumbar spine, femoral neck, and total hip significantly increased by 29.3%, 27.8%, and 30.2% in the denosumab group, and by 32.2%, 47.1%, and 41.1% in the zoledronic acid group (all P < .001 vs baseline). Vertebral height and projection area significantly increased after denosumab and zoledronic acid treatment. Rebound hypercalcemia was found to be a common and serious side effect of denosumab, of which 14.3% reached hypercalcemic crisis. Rebound hypercalcemia could be alleviated by switching to zoledronic acid treatment.

Treatment with denosumab or zoledronic acid is beneficial in increasing BMD and improving the spinal morphometry of children with OI. However, denosumab should be used with caution in pediatric patients with OI because of its common and dangerous side effect of rebound hypercalcemia. The appropriate dosage and dosing interval of denosumab need to be further explored in children with OI.

By 2060, an estimated one in four Americans will be elderly. Consequently, the prevalence of osteoporosis and fragility fractures will also increase. Presently, no available intervention definitively prevents or manages osteoporosis. This study explores whether Pool 7 Compound 3 (P7C3) reduces progressive bone loss and fragility following the onset of ovariectomy (OVX)-induced osteoporosis. Results confirm OVX-induced weakened, osteoporotic bone together with a significant gain in adipogenic body weight. Treatment with P7C3 significantly reduced osteoclastic activity, bone marrow adiposity, whole-body weight gain, and preserved bone area, architecture, and mechanical strength. Analyses reveal significantly upregulated platelet derived growth factor-BB and leukemia inhibitory factor, with downregulation of interleukin-1 R6, and receptor activator of nuclear factor kappa-B (RANK). Together, proteomic data suggest the targeting of several key regulators of inflammation, bone, and adipose turnover, via transforming growth factor-beta/SMAD, and Wingless-related integration site/be-catenin signaling pathways. To the best of the knowledge, this is first evidence of an intervention that drives against bone loss via RANK. Metatranscriptomic analyses of the gut microbiota show P7C3 increased Porphyromonadaceae bacterium, Candidatus Melainabacteria, and Ruminococcaceae bacterium abundance, potentially contributing to the favorable inflammatory, and adipo-osteogenic metabolic regulation observed. The results reveal an undiscovered, and multifunctional therapeutic strategy to prevent the pathological progression of OVX-induced bone loss.

Skeletal disorders can seriously threaten the health and the performance of poultry, such as tibial dyschondroplasia (TD) and osteoporosis (OP). Oligomeric proanthocyanidins (OPC) are naturally occurring polyphenolic flavonoid compounds that can be used as potential substances to improve the bone health and the growth performance of poultry. Eighty 7-day-old green-eggshell yellow feather layer chickens were randomly divided into 4 groups: basal diet and basal diet supplementation with 25, 50, and 100 mg/kg OPC. The results have indicated that the growth performance and bone parameters of chickens were significantly improved supplementation with OPC in vivo, including the bone volume (BV), the bone mineral density (BMD) and the activities of antioxidative enzymes, but ratio of osteoprotegerin (OPG)/receptor activator of NF-κB (RANK) ligand (RANKL) was decreased. Furthermore, primary bone marrow mesenchymal stem cells (BMSCs) and bone marrow monocytes/macrophages (BMMs) were successfully isolated from femur and tibia of chickens, and co-cultured to differentiate into osteoclasts in vitro. The osteogenic differentiation derived from BMSCs was promoted treatment with high concentrations of OPC (10, 20, and 40 µmol/L) groups in vitro, but emerging the inhibition of osteoclastogenesis by increasing the ratio of OPG/RANKL. In contrary, the osteogenic differentiation was also promoted treatment with low concentrations of OPC (2.5, 5, and 10 µmol/L) groups, but osteoclastogenesis was enhanced by decreasing the ratio of OPG/RANKL in vitro. In addition, OPG inhibits the differentiation and activity of osteoclasts by increasing the autophagy in vitro. Dietary supplementation of OPC can improve the growth performance of bone and alter the balance of osteoblasts and osteoclasts, thereby improving the bone health of chickens.

Recent evidence suggests additional immunomodulatory properties of RANKL inhibition possibly boosting the clinical efficacy of immune checkpoint inhibitors (ICI).

We conducted a prospective, multicentre clinical trial in unresectable stage IV melanoma patients with bone metastases who received denosumab in parallel with dual ICI (BONEMET) and performed comprehensive immune monitoring at baseline and 4, 12, and 24 weeks after initiation of therapy. Secondary endpoints included tolerability and efficacy. For comparison, biospecimens from melanoma patients treated with dual ICI without denosumab were analyzed accordingly and served as retrospective reference cohort.

In both the BONEMET (n = 16) and the reference cohort (n = 18) serum levels of 17 cytokines, including IFNγ were significantly increased after 4 weeks of treatment. Patients who received ICI and denosumab showed a significantly higher increase in serum CXCL-13 and a significant decrease in VEGFc compared with the reference cohort. While no changes in T cell composition were observed at 4 weeks, patients in the BONEMET cohort showed a significant decrease in the peripheral naïve T-cell population and an increase in CD8+ effector cells after 12 weeks. Treatment-related adverse events occurred with comparable frequency (93.8% in the BONEMET cohort versus 83.3% in the reference cohort). 7/16 patients in the BONEMET cohort and 8/18 patients in the reference cohort achieved disease control.

Denosumab in combination with dual ICI modulates cytokine expression and T-cell composition in peripheral blood. The upregulation of CXCL-13, a key factor for initiating tertiary lymphoid structures, strengthens the hypothesis that denosumab indeed boost immunological effects.