A Translational Strategy Targeting Type I BMP Receptors to Prevent Heterotopic Ossification
Shailesh Agarwal, MD, Shawn Loder, BS, David Cholok, BS, Christopher Breuler, BS, Michael Chung, MD, Cameron Brownley, BS, Jonathon Peterson, MD, John Li, MD, Hsiao Hsieh Hsung, DDS, Kavitha Ranganathan, MD, Joseph Habbouche, BS, Arminder Kaura, Caitlin Priest, BS, Shuli Li, PhD, Yuji Mishina, PhD, Benjamin Levi, MD.
University of Michigan, Ann Arbor, MI, USA.
PURPOSE: Trauma-induced heterotopic ossification (tHO) is the aberrant growth of ectopic bone in soft tissue, which develops in patients following severe musculoskeletal trauma. Much of HO literature focuses on a related pathology known as fibrodysplasia ossificans progressiva (FOP), which is caused by a hyperactivating mutation in the type I bone morphogenetic protein receptor (T1-BMPR) ACVR1 (ACVR1 R206H). Consequently, emphasis has been placed on developing inhibitors with improved specificity for ACVR1. However, patients who develop tHO do not harbor known ACVR1 mutations, and it is unclear whether emphasis on ACVR1-specific inhibition is beneficial for the treatment of tHO. Here investigate whether any single T1-BMPR is required for tHO, or whether these receptors perform overlapping roles during tHO development. We further evaluate the efficacy of the BMP ligand trap, Alk3-Fc, as a broad-spectrum inhibitor of T1-BMP receptors in the treatment and prevention tHO.
METHODS: Wild-type, tamoxifen-inducible Acvr1 knockout (Acvr1 tmKO: Ub.creERT/Acvr1fl/fl), tamoxifen-inducible Bmpr1a knockout (Bmpr1a tmKO: Ub.creERT/Bmpr1afl/fl), and Bmpr1b knockout (Bmpr1b-/-) mice receivedhindlimb Achille’s tendon transection and dorsal 30% total body surface area partial-thickness burn. A second cohort of wild-type mice received daily i.p. injections of Alk3-fc (2 mg/kg) or saline for 6-weeks post-injury or during weeks 0-2 (inflammation), weeks2-4 (chondrogenesis), or weeks 4-6 (ossification) after injury. Mice receiving abbreviated treatment with Alk3-fc received intraperitoneal saline injections for the remainder of their 6-week treatment period. HO was quantified by microCT 9-weeks post-injury, and cartilage formation was assessed via pentachrome staining.
RESULTS: Genetic loss of Acvr1 (p>0.05), Bmpr1a (p>0.05), or Bmpr1b (p>0.05) alone was unable to significantly or substantially reduce tHO after injury (Fig. 1A). However, genetic loss of both Acvr1 and Bmpr1a (Acvr1;Bmpr1a tmKO: Ub.creERT /Acvr1fl/fl;Bmpr1afl/fl) led to a substantial and significant reduction in tHO volume (16.7-fold decrease vs. wild-type; p<0.05; Fig. 1A). Based on these findings we studied the effect of Alk3-Fc, a BMP ligand trap which inhibits BMP receptor signaling capable of broadly T1-BMPR inhibition at different points during tHO development. Mice treated daily with Alk3-Fc for 6-weeks post-injury demonstrated a significant and substantial reduction in tHO volume (2.9-fold; p<0.05; Fig. 1B). Mice treated daily with Alk3-Fc from 0-2 weeks demonstrated similar reductions in tHO volume (2.3-fold; p<0.05; Fig. 1B). This was confirmed histologically, with diminished cartilage content in 6-week and 0-2 week treated groups when compared with controls.
CONCLUSIONS: Here we demonstrate that although BMP signaling is required for tHO, no single T1-BMPR (ACVR1/ALK2, BMPR1a/ALK3, or BMPR1b/ALK6) alone is necessary for this disease. This suggests that, unlike with genetic forms of HO such as FOP, these receptors have functional redundancy in the setting of tHO. Based on these data we developed an approach based on broad-spectrum T1-BMPR blockade using the ligand trap Alk3-Fc to prevent tHO formation. Our findings suggest that Alk3-Fc treatment during the earliest period after injury is sufficient to block tHO and provides a translational approach to targeting this disease process.
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