Comment on 'Palovarotene reduces heterotopic ossification in juvenile FOP mice but exhibits pronounced skeletal toxicity'
- The Children's Hospital of Philadelphia, United States
- University of Pennsylvania, United States
Abstract
We are writing to communicate our concerns regarding the recently published study by Lees-Shepard et al. (2018).
https://doi.org/10.7554/eLife.43173.001Introduction
Lees-Shepard et al. reported that palovarotene (a retinoid agonist) had severe side effects on growth plate and limb joints in an Acvr1R206H knock-in mouse model of fibrodysplasia ossificans progressiva (FOP; Lees-Shepard et al., 2018). We have a number of concerns about this publication. Lees-Shepard et al. write that “it is important for FOP therapeutics to exhibit an acceptable safety profile in juvenile patients”: however, they used a retinoid drug delivery method that is never used in the treatment of patients, thus undermining their goal. Moreover, the mouse model used does not sufficiently mimic FOP patients.
Discussion
Route of palovarotene administration
Palovarotene was originally designed for oral administration (Hind and Stinchcombe, 2009), allowing for low and steady drug adsorption by the digestive system and an overall absorption efficiency of 50% to 65% of administered dose, typical of retinoids (Blaner and Olson, 1994). Instead, Lees-Shepard et al. injected the drug intraperitoneally (IP) every day for 4 weeks. This IP route of administration is known to invariably cause a sharp and sudden increase in circulating drug levels, resulting in a higher and faster Cmax compared to oral delivery (Eaton and Gilbert, 2013). Indeed, palovarotene Cmax levels were found to be about 6-fold higher after IP injection compared to oral administration in mice (Clementia Pharmaceuticals, internal data). Circulating retinoids are quickly taken up by tissues and cells (Cullum and Zile, 1985), and cells accumulate retinoids in large amounts intracellularly (Kurlandsky et al., 1995; Randolph and Simon, 1997) where their action on cell function and phenotype can last hours to days (Chen and Gudas, 1996). Thus, daily IP injections are expected to result in ever increasing and possibly confounding drug effects over time. Lees-Shepard et al. stated that they used “palovarotene doses….that correspond to approximate adult human equivalent doses of 3.6 mg and 7.2 mg” under testing in ongoing efficacy and safety study of palovarotene for the treatment of FOP being conducted by Clementia Pharmaceuticals (https://clinicaltrials.gov/ct2/show/NCT03312634. This trial has recently entered phase 3). However, the IP administration changed the effective dose and undermined the goal of testing a clinically relevant treatment.
FOP mouse model
Lees-Shepard et al. utilized a conditional mouse model in which the FOP causative gene was activated in Pdgfrα-expressing cells by mating floxed Acvr1R206H mice with Pdgfrα-Cre mice. Those cells were described as a subpopulation of progenitor cells particularly sensitive to effects of the mutation. This mouse model is at sharp variance with FOP patients in which the mutant ACVR1R206H gene characterizes the majority of, if not all, cells. Thus, data obtained with the Pdgfrα-Cre mouse model may be skewed by targeting a specific subpopulation of cells that could also be affected by wild-type cells, with the wild-type cells likely outnumbering the cells in the specific Acvr1R206H subpopulation and interacting with them. Thus, the model does not faithfully reproduce the overall genetic makeup and cell and tissue responses in FOP patients. While Lees-Shepard et al. observed that Pdgfrα+ cells contributed to heterotopic ossification (HO) in their model, it is not known whether this specific cell population is clinically relevant in FOP patients. Further, Lees-Shepard et al. presented no data showing that targeted Pdgfrα+ cells were present in the growing skeleton.
Effects on growth plates and joints
These latter issues are particularly relevant to the severe effects on growth plates and joints reported by Lees-Shepard et al. Given that the Pdgfrα gene is not known to be expressed in chondrocytes (Hamilton et al., 2003), it is nearly certain that growth plate and articular chondrocytes were not targeted by Pdgfrα-Cre and remained largely, if not totally, wild-type. Lees-Shepard et al. included no data to demonstrate that Pdgfrα-Cre had targeted growth plate and joint chondrocytes. Studies dating back decades have established that cartilage is sensitive to systemic retinoid levels (Wolbach and Hegsted, 1953). In our studies (Chakkalakal et al., 2016; Shimono et al., 2011), we did observe a slight growth retardation of about 10% in control wild type mice after oral palovarotene administration, although their growth plates remained open and there were no obvious effects on their joints. Notably, there were no obvious growth retardation and growth plate effects after oral palovarotene administration in our Prrx1-Cre; Acvr1R206H mutant mice in which the entire limb mesenchymal cell population was targeted, including skeletal cells. The higher tolerance of mutant growth plates was likely due to higher basal levels of canonical BMP signaling activity. As we pointed out previously (Chakkalakal et al., 2016), this provides support to the notion that palovarotene may be quite safe for – and well tolerated by – FOP patients, as supported by the announcement of Phase 2 Part B data from the ongoing efficacy and safety study being conducted by Clementia (Clementia Pharmaceuticals, 2018).
Conclusions
In sum, the severe side effects on growth plates and joints observed by Lees-Shepard et al. are likely the results of: (i) the route of drug administration; (ii) excessive palovarotene levels and effects likely caused by the daily IP administration; and (iii) reliance on a mouse model that does not mimic FOP patients, including apparent lack of mutation expression in cartilage. We strongly recommend that data and conclusions in that study should be considered and interpreted with caution with regard to drug effects and action in skeletal tissues and for relevance to ongoing clinical studies.
References
-
BookRetinol and retinoic acid metabolismIn: Sporn MB, Roberts AB, Goodman DS, editors. The Retinoids: Biology, Chemistry, and Medicine (Second Edition). New York: Raven Press. pp. 229–255.
-
An analysis of retinoic acid-induced gene expression and metabolism in AB1 embryonic stem cellsJournal of Biological Chemistry 271:14971–14980.https://doi.org/10.1074/jbc.271.25.14971
-
Metabolism of all-trans-retinoic acid and all-trans-retinyl acetate. Demonstration of common physiological metabolites in rat small intestinal mucosa and circulationThe Journal of Biological Chemistry 260:10590–10596.
-
Casarett and Doull's Toxicology: The Basic Science of PoisonsPrinciples of toxicology, Casarett and Doull's Toxicology: The Basic Science of Poisons, Eighth Edition, London, McGraw-Hill.
-
Evolutionary divergence of platelet-derived growth factor alpha receptor signaling mechanismsMolecular and Cellular Biology 23:4013–4025.https://doi.org/10.1128/MCB.23.11.4013-4025.2003
-
Palovarotene, a novel retinoic acid receptor gamma agonist for the treatment of emphysemaCurrent Opinion in Investigational Drugs 10:1243–1250.
-
Plasma delivery of retinoic acid to tissues in the ratJournal of Biological Chemistry 270:17850–17857.https://doi.org/10.1074/jbc.270.30.17850
-
Metabolism of all-trans-retinoic acid by cultured human epidermal keratinocytesJournal of Lipid Research 38:1374–1383.
-
Hypervitaminosis A in young ducks: The epiphyseal cartilageArchives of Pathology 55:47–54.
Article and author information
Author details
Senior Editor
- Mark I McCarthy, University of Oxford, United Kingdom
Reviewing Editor
- Clifford J Rosen, Maine Medical Center Research Institute, United States
Version history
- Received: October 29, 2018
- Accepted: January 9, 2019
- Version of Record published: January 30, 2019 (version 1)
Copyright
© 2019, Pacifici and Shore
This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
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Comment on 'Palovarotene reduces heterotopic ossification in juvenile FOP mice but exhibits pronounced skeletal toxicity'
eLife 8:e43173.
https://doi.org/10.7554/eLife.43173
Further reading
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- Stem Cells and Regenerative Medicine
Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disorder characterized by debilitating heterotopic ossification (HO). The retinoic acid receptor gamma agonist, palovarotene, and antibody-mediated activin A blockade have entered human clinical trials, but how these therapeutic modalities affect the behavior of pathogenic fibro/adipogenic progenitors (FAPs) is unclear. Using live-animal luminescence imaging, we show that transplanted pathogenic FAPs undergo rapid initial expansion, with peak number strongly correlating with HO severity. Palovarotene significantly reduced expansion of pathogenic FAPs, but was less effective than activin A inhibition, which restored wild-type population growth dynamics to FAPs. Palovarotene pretreatment did not reduce FAPs’ skeletogenic potential, indicating that efficacy requires chronic administration. Although palovarotene inhibited chondrogenic differentiation in vitro and reduced HO in juvenile FOP mice, daily dosing resulted in aggressive synovial joint overgrowth and long bone growth plate ablation. These results highlight the challenge of inhibiting pathological bone formation prior to skeletal maturation.
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Retinitis pigmentosa (RP), a heterogenous group of inherited retinal disorder, causes slow progressive vision loss with no effective treatments available. Mutations in the rhodopsin gene (RHO) account for ~25% cases of autosomal dominant RP (adRP). In this study, we describe the disease characteristics of the first-ever reported mono-allelic copy number variation (CNV) in RHO as a novel cause of adRP. We (a) show advanced retinal degeneration in a male patient (68 years of age) harboring four transcriptionally active intact copies of rhodopsin, (b) recapitulated the clinical phenotypes using retinal organoids, and (c) assessed the utilization of a small molecule, Photoregulin3 (PR3), as a clinically viable strategy to target and modify disease progression in RP patients associated with RHO-CNV. Patient retinal organoids showed photoreceptors dysgenesis, with rod photoreceptors displaying stunted outer segments with occasional elongated cilia-like projections (microscopy); increased RHO mRNA expression (quantitative real-time PCR [qRT-PCR] and bulk RNA sequencing); and elevated levels and mislocalization of rhodopsin protein (RHO) within the cell body of rod photoreceptors (western blotting and immunohistochemistry) over the extended (300 days) culture time period when compared against control organoids. Lastly, we utilized PR3 to target NR2E3, an upstream regulator of RHO, to alter RHO expression and observed a partial rescue of RHO protein localization from the cell body to the inner/outer segments of rod photoreceptors in patient organoids. These results provide a proof-of-principle for personalized medicine and suggest that RHO expression requires precise control. Taken together, this study supports the clinical data indicating that RHO-CNV associated adRPdevelops as a result of protein overexpression, thereby overloading the photoreceptor post-translational modification machinery.
