Hope for escape from a prison of bone: Cellular and molecular targets for fibrodysplasia ossificans progressiva

Frederick S Kaplan

doi:10.4103/HMJ.HMJ_78_18

REVIEW ARTICLE

Year : 2018 | Volume : 11 | Issue : 4 | Page : 144-150

Hope for escape from a prison of bone: Cellular and molecular targets for fibrodysplasia ossificans progressiva

Frederick S Kaplan
Department of Orthopaedic Surgery, Medicine, The Center for Research in FOP and Related Disorders, The Perelman School of Medicine, The University of Pennsylvania, Philadelphia, PA, USA

Date of Web Publication

9-Nov-2018

Correspondence Address:
Prof. Frederick S Kaplan
Division of Orthopaedic Molecular Medicine, Department of Orthopaedic Surgery, Penn Musculoskeletal Center, The Perelman School of Medicine, The University of Pennsylvania, Suite 600, 3737 Market Street, Philadelphia, PA 19104
USA

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/HMJ.HMJ_78_18

Abstract

The progressive pathological metamorphosis of one normal organ system into another is unique to fibrodysplasia ossificans progressiva and unveils a pathophysiologic process that can be exploited for therapy of disabling extraskeletal ossification. Here, we review the recent remarkable insight from medical research and the potential of targeted therapy for this rare condition as well as for many common forms of heterotopic ossification.

Keywords: Activin A, activin receptor A Type I, bone morphogenetic protein, bone morphogenetic protein signalling, fibrodysplasia ossificans progressiva, heterotopic ossification

How to cite this article:
Kaplan FS. Hope for escape from a prison of bone: Cellular and molecular targets for fibrodysplasia ossificans progressiva. Hamdan Med J 2018;11:144-50

How to cite this URL:
Kaplan FS. Hope for escape from a prison of bone: Cellular and molecular targets for fibrodysplasia ossificans progressiva. Hamdan Med J [serial online] 2018 [cited 2023 Mar 29];11:144-50. Available from: http://www.hamdanjournal.org/text.asp?2018/11/4/144/245133

‘Nature is nowhere accustomed more openly to display her secret mysteries than in cases where she shows traces of her workings apart from the beaten path; nor is there any better way to advance the proper practice of medicine than to give our minds to the discovery of the usual law’ of nature, by the careful investigation of cases of rare forms of disease. For it has been found in almost all things, that what they contain of useful or applicable nature, is hardly perceived unless we are deprived of them, or they become deranged in some way.’

William Harvey, London, 1657.^[1]

Introduction

During embryogenesis, undifferentiated cells are programmed to develop into mature tissues and organs. Once formed, tissue and organ identities remain stable throughout life. For example, the heart does not turn into lung. The liver does not turn into pancreas. The kidney does not turn into brain. However, in the ultra-rare and disabling condition known as fibrodysplasia ossificans progressiva (FOP), muscles, tendons, ligaments and fascia turn into mature bone – progressively forming a second skeleton that locks the joints into permanent immobility, rendering movement impossible. The extraskeletal (or heterotopic) bone of the second skeleton is perfectly normal in every way – except, it should not be there.

The ability of mature organisms to stabilise phenotypes has enormous selective advantage across all phyla, but the mechanisms have been largely unexplored.^[2] Individuals with FOP undergo a pathological metamorphosis in which one normal tissue is transformed into another through a highly regulated process of tissue destruction and phenotype reassignment. This disabling metamorphosis is mediated by the FOP metamorphogene, which encodes a mutant bone morphogenetic protein (BMP) Type I receptor (BMP1) that exhibits mild constitutive activity during development and severe episodic dysregulation post-natally due to the action of promiscuous ligands. The discovery of the FOP metamorphogene reveals a highly conserved target for drug development and identifies a fundamental defect in the BMP signalling pathway – which when triggered by injury and inflammation transforms one tissue into another.

Although ultra-rare, focussed research efforts over the past 25 years have begun to decipher the genetic, molecular, cellular and immunologic basis of this medical mystery and have identified multiple targets for therapy of this disabling disorder – and by example – into more common disorders of non-genetic heterotopic ossification (HO) that plague millions of people worldwide – HO that forms from total hip replacement, muscle trauma, war wounds, civilian injuries, trauma to the brain and spinal cord, atherosclerosis and valvular heart disease.^[3] Conversely, the riddles of FOP unveil new opportunities for genetic engineering of new bone for those who desperately need new bone – such as for healing of non-unions, traumatic skeletal injuries, osteoporotic fractures, spinal fusions and congenital skeletal anomalies. In this article, we will briefly review FOP and the progress that has been made to shed light on therapy for this rare condition and vastly more common problems.

Fibrodysplasia Ossificans Progressiva

FOP (MIM135100) is a progressively disabling genetic disorder of extraskeletal ossification, which leads to the formation of a second skeleton of heterotopic bone. Individuals with FOP appear normal at birth, except for characteristic malformations of the great toes that are present in all classically affected individuals.^[4],[5] During the first decade of life, episodic soft tissue swellings (or flare-ups) can arise in the neck and back. While some flare-ups regress spontaneously, most undergo pathological metamorphosis into mature heterotopic bone through an endochondral pathway.^[6] Minor trauma such as intramuscular immunisations, mandibular blocks for dental work, muscle fatigue, blunt muscle trauma, bumps, bruises, falls or influenza-like viral illnesses can trigger new flare-ups of FOP leading to progressive HO.^[4],[5]

Bone formation in FOP is episodic, but disability is cumulative. Most patients are confined to a wheelchair or immobilised in a standing position by the third decade of life and require lifelong assistance for activities of daily living [Figure 1]. The median estimated lifespan is 56 years; death often results from complications of thoracic insufficiency syndrome.^[7]

Figure 1: Clinical appearance and skeleton of a man with fibrodysplasia ossificans progressiva. The rigid posture in this 25-year-old man with fibrodysplasia ossificans progressiva was due to ankylosis of the spine, shoulders, elbows, hips and knees. He died of pneumonia at the age of 40 years. Plates and ribbons of ectopic bone contour the skin over the back and arms (a) and can be seen directly on the skeleton (b). (Courtesy Frederick Kaplan; The New England Journal of Medicine)

Click here to view

Cellular and Molecular Features of Fibrodysplasia Ossificans Progressiva

A recurrent heterozygous missense mutation in activin receptor A Type I (ACVR1), a BMP1, was identified in all classically affected individuals with sporadic or familial FOP – approximately 97% of all known FOP patients worldwide.^[8] Phenotypic and genotypic variants of FOP occur in 3% of all FOP patients worldwide.^[9] DNA sequence analysis of ACVR1 in patients who have classic FOP reveals the identical single nucleotide mutation in ACVR1 (c.617G>A), which results in the substitution of arginine by histidine at codon 206 (p.R206H) in the intracellular GS domain of the receptor, upstream of the kinase domain.^[8]

Most cases of FOP are sporadic, but inheritance can occur by autosomal dominant transmission.^[4] The mutation causes loss of autoinhibition of ACVR1^[10],[11] and renders it susceptible to dysregulated BMP and activin A (Act A) signalling through the SMAD 1/5/8 branch of the BMP signalling pathway.^[12],[13] The pathophysiology of the disorder is multifactorial with immune activation, abnormal cellular response to hypoxia and dysregulation of multiple signalling pathways.^[14]

All of the ACVR1 mutations identified in individuals with classic or variant FOP occur in highly conserved amino acids, indicating their functional importance.^[8],[9] Protein structure homology modelling of the resulting ACVR1 proteins predicted that these mutant receptors activate ACVR1 and enhance receptor signalling – now a well-confirmed finding.^[11] Many studies have demonstrated that signal transduction through the BMP pathway is altered in specific cells from individuals with FOP,^[15],[16] with increased BMP expression,^[17] decreased expression of BMP antagonists, increased phosphorylation of BMP pathway signalling mediators (BMP-specific SMAD proteins and p38MAPK), dysregulated trafficking of BMP receptors and increased expression of BMP transcriptional targets in the absence of exogenous BMP ligand.^{[10],[14],[18]}

Although mutant ACVR1 (mACVR1) exhibits dysregulated basal activity and increased ligand sensitivity, binding of the Type II receptor is necessary for signalling. In fact, a key determinant for ACVR1 hyperactivity is a functional Type II receptor.^[19]

The interaction of mACVR1 with FKBP12, a glycine/serine domain-binding protein that prevents leaky BMP1 activation in the absence of ligand, has been investigated. mACVR1 exhibits reduced binding to FKBP12 in in vitro assays, suggesting that increased BMP pathway activity in cells with mACVR1 is due, at least in part, to decreased binding of this inhibitory factor.^[20]

In addition to numerous in vitro studies, the BMP signalling pathway has been studied in several highly informative animal models of FOP including Drosophila melanogaster and the zebrafish, Danio rerio providing important insight into the cellular and molecular mechanisms of BMP signalling and the activities of the evolutionarily conserved ACVR1 receptor, its orthologs and ligands in vivo.^{[10],[19],[21]} As in vertebrates, elevated basal BMP pathway signalling associated with mACVR1 in Drosophila is BMP ligand-independent.^[19] Wild-type (wt) ACVR1 can antagonise as well as promote BMP pathway signalling while mACVR1 can promote signalling with or without ligand.^[12],[13]

The development of mACVR1 knock-in mouse models of FOP has propelled in vivo research on FOP.^[21],[22] Although germline transmission of mACVR1 in the mouse leads to perinatal lethality, mice that are 70%–90% chimeric for mACVR1 cells exhibit clinical features of FOP, including embryonic skeletal malformations and post-natal HO that is identical to that seen in the human condition.^[22] Importantly, knock-in mACVR1 mice also develop spontaneous and injury-induced FOP-like lesions that progress to mature heterotopic bone through a cellular cascade identical to that seen in individuals with FOP. These mice validate that mACVR1 is a direct genetic cause of FOP and that mild basal activity and increased ligand sensitivity of mACVR1 reproduce the HO seen in humans.^[22]

Recently, mACVR1 was shown to directly regulate early chondrogenic fate in FOP. Importantly, chondrogenic differentiation was accelerated in mACVR1 cells due, in part, to enhanced sensitivity to BMP4 and Act A ligands.^[23]

Targeted Therapy for Fibrodysplasia Ossificans Progressiva: Hope for Escape

At present, standard-of-care medical management is supportive.^[24],[25] High-dose glucocorticoids have limited use in the management of the early inflammatory flare-ups. Surgical removal of FOP lesions is strongly discouraged as it is uniformly followed by recurrence. Surgical release of joint contractures is unsuccessful and risks new, trauma-induced HO. Intramuscular injections must be avoided. Prevention of falls, influenza, recurrent pulmonary infections and complications of restrictive chest wall disease is vital.^[24],[25]

Definitive treatments and cures are not yet available for FOP, and there is a critical unmet need for the development of effective therapies. The discovery of the FOP gene, emerging insights into its mechanism of action and identification of associated pathway abnormalities reveal at least five plausible approaches to the treatment and/or prevention of FOP:^[26]

Diminishing activity of mACVR1
Blocking inflammatory triggers of FOP flare-ups
Redirecting FOP progenitor cells to an alternate tissue fate other than cartilage or bone
Blocking the body's response to microenvironmental signals (such as hypoxia) that promote the development of FOP lesions
Blocking multiple nodes in the network of physiologic targets.

Worldwide interest in FOP skyrocketed in the wake of the historic discovery of the FOP gene in 2006 – the first hard target for FOP therapy. Soon after the FOP gene discovery, The New York Times published an article entitled, ‘Finally, With Gene Discovery, Hope for Escape from a Prison of Bone’.^[27] The FOP gene discovery launched an industry that catapulted the field to clinical trials. FOP research is now a worldwide enterprise. This new enterprise is leading to massive research investment in FOP on the part of academia and industry worldwide – all propelled by the discovery of the FOP gene and by the identification of its robust therapeutic target the ACVR1 and its interacting molecules and pathways.

Target: The Mutant Activin Receptor a Type I Receptor

The discovery of the FOP gene identified ACVR1, the mutated BMP receptor, as the prime pharmacologic target for the treatment of FOP. Multiple therapeutic strategies for inhibiting dysregulated BMP signalling in FOP are being pursued [Figure 2].

Figure 2: Potential therapies for identified targets in fibrodysplasia ossificans progressiva. mAbs = monoclonal antibodies; STIs = signal transduction inhibitors; siRNA = small inhibitory RNA

Click here to view

Signal transduction inhibitors

Signal transduction inhibitors (STIs) are important molecular tools for studying BMP pathway signalling in FOP and have great potential for the development into powerful therapeutic drugs for FOP.^{[26],[28],[29]} STIs are small molecules, generally well-absorbed into the bloodstream through the gastrointestinal tract that work by blocking the ‘kinase mouth’ of ACVR1 so that it cannot transmit metamorphic signals. Selective STIs for FOP will inhibit ACVR1 (also known as ALK2) rather than ALK1, ALK3, or ALK6 (bone-forming receptors in the same family). Such highly selective molecules are being developed by pharmaceutical companies for use in FOP clinical trials. Broad spectrum STIs that are currently available and that target ACVR1 (among other receptors) are also being considered for repurposing in FOP clinical trials. Recent investigations are exploring novel allosteric destabilisers as therapeutics for FOP.^[29]

Blocking antibodies against activin receptor A Type I receptor

mACVR1 (in FOP) signals at low levels all the time (even when it should be inactivated). mACVR1 is also hyper-responsive to locally produced hormone-like molecules (also known as ligands) such as BMP4 and Act A, thus the rationale for using blocking antibodies to ACVR1 in the prevention and treatment of FOP.^[26],[29]

Therapeutic monoclonal antibodies that target ACVR1 are under development by several pharmaceutical companies.

Blocking antibodies against activin A

Act A potently stimulates the mutant FOP receptor with little or no activation of wt ACVR1.^[12],[13] The molecular, physiologic and structural basis for the sensitivity of mtACVR1 to Act A is still unknown. Act A has previously been recognised as a potent hormone-like molecule that is a key regulator of the immune system.

Importantly, Act A induces HO in FOP mice.^[12] Inhibition of Act A with a fully humanised monoclonal antibody blocks spontaneous and trauma-induced HO in FOP mice.^[12] The unexpected discovery of Act A in the pathogenesis of FOP identifies a therapeutic target for FOP and excavates a foundation for clinical development.^{[12],[13],[26],[29]}

Ligand traps

Ligand traps, magnet-like molecules that attract molecules such as BMP4 and Act A and prevent them from activating ACVR1, have been proposed as a therapeutic strategy for FOP. Such molecules have been produced and been shown experimentally to inhibit dysregulated BMP signalling in FOP cells and suppress cartilage and bone formation in mutant allele-specific inhibitory RNA.^{[26],[29],[30]}

Inhibitory RNA capable of suppressing the expression of ACVR1 in connective tissue progenitor cells from FOP patients restores dysregulated BMP signalling to levels observed in control cells and blocks cartilage and bone formation in vitro. While providing proof-of-principle for allele-specific inhibition of ACVR1 in the prevention of HO in FOP, the in vivo utility of this approach must be confirmed in mouse models of FOP. A hurdle to human application is safe, effective and durable delivery of small inhibitory RNA molecules to relevant progenitor cells. While small molecule inhibitors or biologics may dominate near-term therapeutic options, opportunities on the distant horizon using inhibitory RNA are appreciable.

Target: Inflammatory and Immune Triggers

Loss of autoinhibition of mACVR1 results in dysregulated BMP pathway signalling and is necessary for the myriad developmental features of FOP but does not appear sufficient to induce the episodic flare-ups that lead to disabling post-natal HO.^[31] Post-natal FOP flare-ups strongly implicate an underlying immunological trigger involving inflammation and the innate immune system. Recent studies implicate canonical and non-canonical/BMP family ligands in the amplification of mACVR1 signalling leading to the formation of FOP lesions and resultant HO. BMP and activin ligands that stimulate mACVR1 signalling also have critical regulatory functions in the immune system. Cross-talk between the morphogenetic and immunological pathways that regulate tissue maintenance and wound healing identifies potential robust therapeutic targets for FOP.^{[12],[13],[31]}

Despite the occurrence of germline activating mutations of ACVR1 in FOP patients, and the presence of mild ligand-independent elevation of basal BMP signalling, individuals with FOP do not form bone continuously, but rather episodically and often following trivial injury – a finding that suggests that innate immune-related triggers induce tissue metamorphosis in the setting of altered microenvironmental thresholds. Many clinical and pathologic features of FOP strongly point to an underlying immunological component to HO. These myriad epidemiologic, clinical, pathological and molecular features of FOP support that the innate immune system plays a prominent and provocative role in the pathophysiology of FOP.^[31]

Immune cells of hematopoietic origin have been implicated in the HO of FOP.^[32] Clinical observation and in vivo murine transplantation studies precisely determined the contribution of hematopoietic cells to HO. A patient with FOP who had undergone bone marrow transplantation for the treatment of intercurrent aplastic anaemia 25 years earlier was evaluated to determine whether the clinical course of his FOP had been influenced by bone marrow replacement or immunosuppression or both. In complementary studies, haematopoietic stem cells were transplanted from constitutively expressing lacZ transgenic mice to identify the contribution of hematopoietic cells to BMP4-inducedHO, an early histopathologic model of FOP.^[32]

Replacement of haematopoietic cells was not sufficient to prevent HO in the patient with FOP, but pharmacologic suppression of the normal donor immune system following transplantation in the new host modulated the activity of the post-natal FOP phenotype and diminished the formation of HO. In complementary murine transplantation studies, cells of haematopoietic origin contributed to the early inflammatory and late marrow-repopulating stages of BMP4-induced HO but were not represented in the fibroproliferative, chondrogenic or osteogenic stages of HO. Interestingly, both the recombinant human BMP4 induction of HO in an animal model and the dysregulated BMP signalling pathway in a patient with FOP were sufficient to recruit at least two populations of cells, one of the haematopoietic origin and at least one of the non-haematopoietic origin, that contribute to the formation of an ectopic skeleton.^{[15],[32],[33]}

Taken together, these findings demonstrated that bone marrow transplantation did not cure FOP in the patient, most likely because the haematopoietic cells were not the population, or at least not the dominant target population, of the intrinsic dysregulation of the BMP signalling pathway in FOP. However, following transplantation of bone marrow from a presumably normal donor, immunosuppression of the immune system appeared to ameliorate HO in a genetically susceptible host. Thus, cells of haematopoietic origin may initiate the formation of an ectopic skeleton although they may not be sufficient to complete the process alone. Furthermore, while cells of haematopoietic origin may contribute robustly to FOP flare-ups, wt haematopoietic cells may be sufficient to stimulate the process in resident FOP osteoprogenitor cells.^[32]

Episodic flare-ups of FOP are characterised clinically by severe, often posttraumatic, connective tissue swelling and intramuscular oedema, followed histologically by an intense and highly angiogenic fibroproliferative reaction. This early inflammatory and angiogenic fibroproliferative response is accompanied by the presence of abundant mast cells far in excess of other reported myopathies.^[34] Using an injury-induced, constitutively active transgenic mouse model of FOP, investigators showed that mast cell inhibition by cromolyn, a mast cell stabiliser, results in a dramatic reduction of HO. Cromolyn significantly decreases the total number of mast cells in FOP lesions. Furthermore, cromolyn specifically diminishes the number of degranulating mast cells in pre-osseous lesions. This work demonstrated that consideration of FOP as a type of localised mastocytosis may offer new therapeutic interventions for treatment of this devastating condition.^[35]

In a recent study using the classic FOP knock-in mouse model, investigators found that the ACVR1 R206H mutation caused increased BMP signalling in post-traumatic FOP lesions and early divergence from the normal skeletal muscle repair program with elevated and prolonged immune cell infiltration.^[36] The pro-inflammatory cytokine response of tumour necrosis factor a, IL-1β and IL-6 was elevated and prolonged in FOP lesions and in FOP mast cells. Importantly, depletion of mast cells and macrophages significantly impaired injury-induced HO in FOP mice, reducing injury-induced HO volume by = 50% with depletion of each cell population independently and = 75% with combined depletion of both cell populations. These data show that the immune system contributes to the initiation and development of HO in FOP and unveils novel therapeutic targets for treatment of FOP and non-genetic forms of HO.^[36]

Target: The Pre-Cartilage Scaffold

In FOP, heterotopic bone forms by a mechanism called endochondral ossification – in other words, through an obligate cartilage scaffold. Palovarotene, a drug now in clinical trials for FOP, selectively inhibits cartilage formation by blocking the BMP signalling pathway downstream of mACVR1 in pre-chondrogenic cells involved in HO. Investigators showed in 2011 that palovarotene inhibited HO in mice genetically engineered to form heterotopic bone.^[37] The HO in FOP flare-ups progresses through a cartilage stage before replacement with mature bone. If the cartilage scaffold can be inhibited, bone will not form. Thus, the pre-cartilage scaffold is a target for FOP.

In September 2016, scientists announced a major breakthrough in understanding the role of palovarotene in FOP. In their paper, ‘palovarotene inhibits HO and maintains limb mobility and growth in mice with the human ACVR1(R206H) FOP FOP mutation’, investigators tested palovarotene in a classic FOP mouse model.^[38] They reported that palovarotene prevented spontaneous HO in FOP mice. In addition, palovarotene protected growing newborn FOP mice when given to lactating mothers. Importantly, palovarotene maintained joint, limb and body motion, providing clear evidence for its encompassing therapeutic potential as a treatment for FOP. In addition, palovarotene not only inhibited spontaneous HO but also prevented it when mice were experimentally injured. This finding also indicates potential benefits for treating or preventing HO that results from trauma, head injuries, war wounds or joint replacement.^[3],[38]

Target: Microenvironmental Amplifiers of Dysregulated Bone Morphogenetic Protein Pathways Signalling

Hypoxia and inflammation are implicated in the episodic induction of HO; however, until recently, the molecular mechanisms were unknown. Hypoxia-inducible factor HIF-1α integrates the cellular response to both hypoxia and inflammation and is a prime candidate for regulating HO.^[14] A recent study investigated the role of hypoxia and HIF-1α in FOP and found that HIF-1α increased the intensity and duration of BMP signalling through Rabaptin 5 (RABEP1)-mediated retention of ACVR1, in the endosomal compartment of hypoxic connective tissue progenitor cells from patients with FOP. The study further showed that early inflammatory FOP lesions in humans and in a mouse model were markedly hypoxic. Importantly, inhibition of HIF-1α by genetic or pharmacologic means restored canonical BMP signalling to normoxic levels in human FOP cells and profoundly reduced HO in a mouse model of FOP. Thus, an inflammation and cellular oxygen-sensing mechanism that modulates intracellular retention of a mutant BMP receptor determine, in part, its pathologic activity in FOP.^[14] The study provided critical insight into a previously unrecognised role of HIF-1α in the ligand-independent and hypoxic amplification of BMP signalling and in the episodic induction of HO in FOP and further identified HIF-1α as a therapeutic target for FOP and perhaps non-genetic forms of HO.^[14]

Multifactorial Targets in Fibrodysplasia Ossificans Progressiva Pathophysiology

Research studies have identified multiple potential targets for therapy in FOP, and novel drug candidates are being developed for testing in clinical trials. A complementary approach seeks to identify approved drugs that could be re-purposed for off-label use against defined targets in FOP. One such drug is imatinib mesylate, a tyrosine kinase inhibitor originally developed for use in patients with chronic myeloid leukaemia.

Imatinib has the desirable effect of attacking multiple targets involved in the early hypoxic and inflammatory stages of FOP flare-ups, including HIF-1α, PDGFRα, c-KIT and multiple MAP kinases.^[39] Based on compelling biologic rationale, strong preclinical data and a favourable safety profile, imatinib has been prescribed on an off-label basis in a non-trial setting in seven children with continuous FOP flare-ups, predominantly in the axial regions, and which were not responsive to standard-of-care regimens. Anecdotal reports in these seven isolated cases documented that the medication was well tolerated with a ubiquitous reported decrease in the intensity of flare-ups in the six children who took the medication. These early clinical observations support the implementation of clinical trials in children with uncontrolled FOP flare-ups to determine if imatinib may ameliorate symptoms or alter the natural history of this debilitating and life-threatening disease.^[39]

Perspectives and Outstanding Questions

BMP ligand-independent basal activation of the BMP signalling pathway in FOP is a well-noted molecular signature of the disease and is plausibly responsible for the myriad developmental features of FOP such as the congenital malformations of the great toes, thumbs, cervical facet, costovertebral and hip joints; osteochondromas and hearing loss, as well as numerous variant and congenital phenotypic features. While the molecular basis of the congenital features of FOP is intriguing and will likely elucidate additional therapeutic targets, it is the post-natal, episodic, progressive and disabling formation of HO that commands greatest attention and is most relevant to the successful treatment of FOP and more common forms of HO.

Clinical observations strongly suggest that inflammatory triggers, initiated through the innate immune system, are responsible for trauma-induced flare-ups and subsequent HO in FOP. It is intriguing to speculate that perhaps all flare-ups, even those that appear spontaneous, are activated by the innate immune system.^[31] Clearly, the innate immune system is ubiquitously active and functional post-natally in vertebrates even in the absence of overt injury. On-going studies will elucidate the role of down-stream and re-entrant activation of cognate and orphan ligands such as BMP4 and Act A in this process, their cross-talk with the inflammatory pathways and their impingement on resident connective tissue progenitor cells capable of orchestrating the requisite pathologic cascade of HO.

These and other questions are the focus of intense on-going investigation. As Tom Maeder wrote in The Atlantic Monthly, ‘FOP and its problems lie at the crossroads of several seemingly unrelated disciplines. Answers to questions that FOP poses will also address grander issues of how the body first creates its shape and knows where to stop; how tissues decide to become what they are, and why they don't be turn into something else’.^[40]

Acknowledgements

The authors thank Mr Robert Caron and Mrs. Kamlesh Rai for their invaluable technical and administrative assistance.

Financial support and sponsorship

This work was supported in part by the International Fibrodysplasia Ossificans Progressiva Association, The Center for Research in FOP and Related Disorders, the Ian Cali Endowment for FOP Research, the Whitney Weldon Endowment for FOP Research, the Ashley Martucci Research Fund the Isaac and Rose Nassau Professorship of Orthopaedic Molecular Medicine, the Penn Center for Musculoskeletal Disorders, and the National Institutes of Health (NIH R01-AR41916).

Conflicts of interest

There are no conflicts of interest.

References

1.	Willis R, editor. The Works of William Harvey. Philadelphia: University of Pennsylvania Press; 1989.
2.	Kaplan FS, Pignolo RJ, Shore EM. The FOP metamorphogene encodes a novel type I receptor that dysregulates BMP signaling. Cytokine Growth Factor Rev 2009;20:399-407.
3.	Pignolo RJ, Foley KL. Nonhereditary heterotopic ossification. Clin Rev Bone Miner Metab 2005;3:261-6.
4.	Shore EM, Kaplan FS. Inherited human diseases of heterotopic bone formation. Nat Rev Rheumatol 2010;6:518-27.
5.	Pignolo RJ, Shore EM, Kaplan FS. Fibrodysplasia ossificans progressiva: Clinical and genetic aspects. Orphanet J Rare Dis 2011;6:80.
6.	Pignolo RJ, Bedford-Gay C, Liljesthrom M, Durbin-Johnson BP, Shore EM, Rocke DM, et al. The natural history of flare-ups in fibrodysplasia ossificans progressiva: A comprehensive global assessment. J Bone Miner Res 2016;31:650-6.
7.	Kaplan FS, Zasloff MA, Kitterman JA, Shore EM, Hong CC, Rocke DM, et al. Early mortality and cardiorespiratory failure in patients with fibrodysplasia ossificans progressiva. J Bone Joint Surg Am 2010;92:686-91.
8.	Shore EM, Xu M, Feldman GJ, Fenstermacher DA, Cho TJ, Choi IH, et al. Arecurrent mutation in the BMP type I receptor ACVR1 causes inherited and sporadic fibrodysplasia ossificans progressiva. Nat Genet 2006;38:525-7.
9.	Kaplan FS, Xu M, Seemann P, Connor JM, Glaser DL, Carroll L, et al. Classic and atypical fibrodysplasia ossificans progressiva (FOP) phenotypes are caused by mutations in the bone morphogenetic protein (BMP) type I receptor ACVR1. Hum Mutat 2009;30:379-90.
10.	Shen Q, Little SC, Xu M, Haupt J, Ast C, Katagiri T, et al. The fibrodysplasia ossificans progressiva R206H ACVR1 mutation activates BMP-independent chondrogenesis and zebrafish embryo ventralization. J Clin Invest 2009;119:3462-72.
11.	Chaikuad A, Alfano I, Kerr G, Sanvitale CE, Boergermann JH, Triffitt JT, et al. Structure of the bone morphogenetic protein receptor ALK2 and implications for fibrodysplasia ossificans progressiva. J Biol Chem 2012;287:36990-8.
12.	Hatsell SJ, Idone V, Wolken DM, Huang L, Kim HJ, Wang L, et al. ACVR1R206H receptor mutation causes fibrodysplasia ossificans progressiva by imparting responsiveness to activin A. Sci Transl Med 2015;7:303ra137.
13.	Hino K, Ikeya M, Horigome K, Matsumoto Y, Ebise H, Nishio M, et al. Neofunction of ACVR1 in fibrodysplasia ossificans progressiva. Proc Natl Acad Sci U S A 2015;112:15438-43.
14.	Wang H, Lindborg C, Lounev V, Kim JH, McCarrick-Walmsley R, Xu M, et al. Cellular hypoxia promotes heterotopic ossification by amplifying BMP signaling. J Bone Miner Res 2016;31:1652-65.
15.	Lounev VY, Ramachandran R, Wosczyna MN, Yamamoto M, Maidment AD, Shore EM, et al. Identification of progenitor cells that contribute to heterotopic skeletogenesis. J Bone Joint Surg Am 2009;91:652-63.
16.	Wosczyna MN, Biswas AA, Cogswell CA, Goldhamer DJ. Multipotent progenitors resident in the skeletal muscle interstitium exhibit robust BMP-dependent osteogenic activity and mediate heterotopic ossification. J Bone Miner Res 2012;27:1004-17.
17.	Shafritz AB, Shore EM, Gannon FH, Zasloff MA, Taub R, Muenke M, et al. Overexpression of an osteogenic morphogen in fibrodysplasia ossificans progressiva. N Engl J Med 1996;335:555-61.
18.	Ahn J, Serrano de la Pena L, Shore EM, Kaplan FS. Paresis of a bone morphogenetic protein-antagonist response in a genetic disorder of heterotopic skeletogenesis. J Bone Joint Surg Am 2003;85-A: 667-74.
19.	Le VQ, Wharton KA. Hyperactive BMP signaling induced by ALK2(R206H) requires type II receptor function in a drosophila model for classic fibrodysplasia ossificans progressiva. Dev Dyn 2012;241:200-14.
20.	Groppe JC, Wu J, Shore EM, Kaplan FS. in vitro analyses of the dysregulated R206H ALK2 kinase-FKBP12 interaction associated with heterotopic ossification in FOP. Cells Tissues Organs 2011;194:291-5.
21.	Kaplan FS, Chakkalakal SA, Shore EM. Fibrodysplasia ossificans progressiva: Mechanisms and models of skeletal metamorphosis. Dis Model Mech 2012;5:756-62.
22.	Chakkalakal SA, Zhang D, Culbert AL, Convente MR, Caron RJ, Wright AC, et al. An Acvr1 R206H knock-in mouse has fibrodysplasia ossificans progressiva. J Bone Miner Res 2012;27:1746-56.
23.	Culbert AL, Chakkalakal SA, Theosmy EG, Brennan TA, Kaplan FS, Shore EM, et al. Alk2 regulates early chondrogenic fate in fibrodysplasia ossificans progressiva heterotopic endochondral ossification. Stem Cells 2014;32:1289-300.
24.	Kaplan FS, Le Merrer M, Glaser DL, Pignolo RJ, Goldsby RE, Kitterman JA, et al. Fibrodysplasia ossificans progressiva. Best Pract Res Clin Rheumatol 2008;22:191-205.
25.	Kaplan FS, Shore EM, Pignolo RJ, The International Clinical Consortium on FOP. The medical management of fibrodysplasia ossificans progressiva: Current treatment considerations. Clin Proc Int Clin Consort FOP 2011;4:1-100.
26.	Kaplan FS, Pignolo RJ, Al Mukaddam MM, Shore EM. Hard targets for a second skeleton: Therapeutic horizons for fibrodysplasia ossificans progressiva (FOP). Expert Opin Orphan Drugs 2017;5:291-4.
27.	Mason M. Finally, with Genetic Discovery, Hope for Escape from a Prison of Bone. New York Times; 09 May, 2006.
28.	Hong CC, Yu PB. Applications of small molecule BMP inhibitors in physiology and disease. Cytokine Growth Factor Rev 2009;20:409-18.
29.	Kaplan FS, Pignolo RJ, AI Mukaddam M, Shore EM. Twenty-Sixth Annual Report of the Fibrodysplasia Ossificans Progressiva (FOP) Collaborative Research Project. Annual Report of the Center for Research in FOP and Related Disorders. Summer; 2017.
30.	Kaplan J, Kaplan FS, Shore EM. Restoration of normal BMP signaling levels and osteogenic differentiation in FOP mesenchymal progenitor cells by mutant allele-specific targeting. Gene Ther 2012;19:786-90.
31.	Kaplan FS, Pignolo RJ, Shore EM. Granting immunity to FOP and catching heterotopic ossification in the act. Semin Cell Dev Biol 2016;49:30-6.
32.	Kaplan FS, Glaser DL, Shore EM, Pignolo RJ, Xu M, Zhang Y, et al. Hematopoietic stem-cell contribution to ectopic skeletogenesis. J Bone Joint Surg Am 2007;89:347-57.
33.	Billings PC, Fiori JL, Bentwood JL, O'Connell MP, Jiao X, Nussbaum B, et al. Dysregulated BMP signaling and enhanced osteogenic differentiation of connective tissue progenitor cells from patients with fibrodysplasia ossificans progressiva (FOP). J Bone Miner Res 2008;23:305-13.
34.	Gannon FH, Glaser D, Caron R, Thompson LD, Shore EM, Kaplan FS, et al. Mast cell involvement in fibrodysplasia ossificans progressiva. Hum Pathol 2001;32:842-8.
35.	Brennan TA, Lindborg CM, Bergbauer CR, Wang H, Kaplan FS, Pignolo RJ, et al. Mast cell inhibition as a therapeutic approach in fibrodysplasia ossificans progressiva (FOP). Bone 2018;109:259-66.
36.	Convente MR, Chakkalakal SA, Yang E, Caron RJ, Zhang D, Kambayashi T, et al. Depletion of mast cells and macrophages impairs heterotopic ossification in an Acvr1^R206H mouse model of fibrodysplasia ossificans progressiva. J Bone Miner Res 2018;33:269-82.
37.	Shimono K, Tung WE, Macolino C, Chi AH, Didizian JH, Mundy C, et al. Potent inhibition of heterotopic ossification by nuclear retinoic acid receptor-γ agonists. Nat Med 2011;17:454-60.
38.	Chakkalakal SA, Uchibe K, Convente MR, Zhang D, Economides AN, Kaplan FS, et al. Palovarotene inhibits heterotopic ossification and maintains limb mobility and growth in mice with the human ACVR1(R206H) fibrodysplasia ossificans progressiva (FOP) mutation. J Bone Miner Res 2016;31:1666-75.
39.	Kaplan FS, Andolina JR, Adamson PC, Teachey DT, Finklestein JZ, Ebb DH, et al. Early clinical observations on the use of imatinib mesylate in FOP: A report of seven cases. Bone 2018;109:276-80.
40.	Maeder T. A few hundred people turned to bone. Atl Mon 1998;281:81-9.