RANKL inhibition with denosumab improves fibrous dysplasia by decreasing lesional cell proliferation and increasing osteogenesis

Abstract

BACKGROUND: Fibrous dysplasia (FD) is a rare, disabling disease with no established treatments. Growing evidence supports inhibiting the pro-osteoclastic factor receptor activator of nuclear Kappa-B ligand (RANKL) as a potential treatment strategy. We conducted a phase 2 trial evaluating the anti-RANKL drug denosumab in adults with FD, with an emphasis on investigating post-discontinuation bone turnover rebound, and cellular mechanisms underlying anti-RANKL effects on FD osteoprogenitors. METHODS: Eight subjects received denosumab for 6-months and were observed for 8-months post-discontinuation. Efficacy and safety were evaluated using bone turnover markers, 18F-NaF PET/CT, and lesion biopsies. RANKL neutralization effects were assessed by histology, RNASeq, and an FD mouse model. Interplay between osteoclasts and FD osteoprogenitors was assessed in an ex vivo lesion model. RESULTS: Denosumab markedly reduced bone turnover and radiographic lesional activity in all subjects. Denosumab was well-tolerated during the treatment period, however post-discontinuation turnover reached or exceeded pre-treatment in six subjects, associated with severe hypercalcemia in one. Histology and whole-exome RNA sequencing showed reduced FD cell proliferation and increased osteogenic maturation, with increased lesional bone formation. The ex vivo model supported the dependence of FD cell proliferation on osteoclast activation. CONCLUSIONS: Osteoclast inhibition by anti-RANKL decreased FD cell proliferation and lesional activity, enabling osteogenic maturation and bone formation. These findings provide new understanding of FD pathogenesis as driven by crosstalk between osteoclasts and pre-osteoblast/osteoblasts, and support denosumab as a mechanistically-driven treatment strategy. Marked bone turnover rebound with post-discontinuation hypercalcemia occurs in a subset of patients, particularly younger individuals with high disease burden. TRIAL REGISTRATION: ClinicalTrials.govNCT03571191FUNDING: This work was supported by the Intramural Research Program of the NIDCR, NICHD, and Clinical Center, National Institutes of Health. Clinical trialNCT03571191was conducted as an investigator-sponsored study supported by Amgen, Inc. This research was supported in part by the NIDCR Genomics and Computational Biology Core: ZIC DC000086 and Veterinary Resources Core: ZIC DE000740-05. Work in MTC lab and LVC labs were supported by the of Research on Women's Health (ORWH) through the Bench to Bedside Program award #884515.

Transcriptomic signature and pro-osteoclastic secreted factors of abnormal bone marrow stromal cells in fibrous dysplasia

“…Although our understanding of the lesional cell population dynamics in FD pathogenesis has advanced significantly in recent years, we still have limited knowledge of the transcriptional effects of hyperactive Gαs and cAMP excess in BMSCs, the underlying cause of the lesions. Previous differential gene expression analyses of bulk FD tissue fail to capture FD BMSCs-specific transcriptomic changes (9,14,15) , and attempts seeking to characterize the effects of Gαs R201C expression in human BMSCs by lentiviral transduction of wildtype BMSCs involved high cell manipulation, limiting the model's validity to emulate the transcriptomic profile of FD BMSCs (17,18) . In the present study, to analyze the cell-intrinsic effects of GNAS gain-of-function in FD BMSCs unaffected by the lesional microenvironment, we performed a comprehensive exploration of the transcriptome and secretome of FD BMSCs cultured in the absence of other cell types.…”

“…This resulted in a better mineralized and organized bone in FD lesions and a decrease in co-morbidities associated with the skeletal lesions. Based on these findings, our current understanding of FD pathophysiology is that of a positive feedback loop between altered BMSCs and osteoclasts, through which FD BMSCs release proosteoclastic factors inducing osteoclastogenesis, and osteoclasts respond increasing BMSC proliferation and contributing to their altered differentiation through intercellular signals like RANK-containing extracellular vesicles (8,9) . Despite this advancement of our understanding of the cellular dynamics in FD tissue, little is known about the transcriptional effects of Gαs activation/cAMP excess in BMSCs and how this translates into their modification of the FD lesion microenvironment through the modulation of released signaling factors.…”

“…On the other hand, we and others carried out efforts to characterize the transcriptomic profile of FD, but our approach involved the isolation of mRNA in bulk FD tissue, which does not allow for determination of the contribution of BMSCs bearing gain-of-function variants of GNAS to the differential expression genes identified (9,(14)(15)(16) . In addition, previous attempts were done to characterize the transcriptomic changes caused by GNAS activation in FD BMSCs using cultured human BMSCs transduced with Gαs R201C , though the cellular manipulation of this technique may limit its reliability to capture the transcriptomic effects of Gαs gain of function in lesional BMSCs (17,18) .…”

An inducible explant model for dissecting osteoclast-osteoblast coordination in health and disease

“…We believe our culture system will be useful for addressing mechanisms underpinning the basic formation and coordination of osteoclasts and osteoblasts, as an acute model of the excessive osteoclast formation that accompanies many resorptive bone diseases and can help us better address how osteoblast-osteoclast coordination is accomplished in health and disease. We have utilized this novel model in two recent manuscripts to identify a novel regulator of osteoclast fusion and assess the molecular mechanism of Denosumab treatment of fibrous dysplasia (39, 40). It is now our hope to disseminate this tool and partner with our colleagues to implement its use in answering new, vital questions concerning how the BMU and bone remodeling are managed in health and disease.…”

Press release from the American Society for Bone and Mineral Metabolism 2022 Annual Meeting and Vitamin D Workshop 2022

“…Ряд докладов был посвящен исследованию таких новых препаратов в лечении редких заболеваний, как мезенхимальные стволовые клетки при несовершенном остеогенезе [18], высокие дозы деносумаба при фиброзной дисплазии [19], буросумаб при X-сцепленной гипофосфатемии [20], энкалерет при аутосомно-доминантной гипокальциемии 1 типа [21]. Также представлена динамика МПК и маркеров костного обмена в исследовании второй фазы препарата TransCon PTH -пролекарства паратиреоидного гормона -у пациентов с гипопаратиреозом [22].…”