1. Peroxisomal Biogenesis Disorders and Bone Disease Eric T. Rush, MD Clinical Geneticist Medical Director, Osteogenesis Imperfecta Clinic Assistant Professor of Pediatrics and Internal Medicine University of Nebraska Medical Center

2. Let’s start off with pathways!

3. Background How I got here (the short version) –As a genetics fellow, I started doing research through the metabolic bone/OI clinic and found that I liked it –Had the rare pleasure of meeting many families and patients with PBDs in Omaha in August of 2011 with Drs. Rizzo, Braverman, and Raymond

4. Background A clear need was expressed by several parents for treatment of bone issues Over the past two years, I have interacted in some way with several families who are now being treated In recommending treatment, we borrow heavily from the treatment of other disorders such as osteoporosis and osteogenesis imperfecta

5. What is the problem? 1) A number of children with PBD have decreased bone mineral density and may also have: –Bone pain –Fractures –Impaired mobility –Apparently decreased energy 2) There is precisely zero literature on this exact subject

6. What isn’t Bone?

7. What is Bone? Cells –Osteoprogenitor – immature progenitor cells which differentiate into osteoblasts –Osteoblast – produces osteoid and then mineralize that osteoid into mature bone –Osteocyte – osteoblasts which become trapped in bone matrix and remain in bone lacunae. Probably work to maintain nearby bone –Osteoclast – resorb both the mineral and collagenous matrix of regions of bone that are damaged

8. What is Bone? Extracellular matrix –Organic 85 - 90% Collagen I 1- 2% Osteocalcin 8-14% Other proteins and glycosaminoglycans –Inorganic Hydroxyapatite, a calcium and phosphorous containing mineral

9. Bone Anatomy

10. How is Bone Made?

11. How is Bone Repaired?

12. Disuse Osteoporosis Bone loss due to skeletal unloading –Neurologic or muscular diseases –Immobilization, often described in bedridden patients or with long-term space flight Bone loss is regional and correspond to regions which are not under load –In non ambulatory patients, we see bone loss predominantly in femur with relative spinal sparing

13. Disuse Osteoporosis A negative calcium balance is observed in paraplegics and astronauts prior to loss of BMD Changes to PTH, Vitamin D, GH, and sex hormones have all been implicated.

14. Riddle Me This… Is disuse osteoporosis the only explanation for bone disease in PBD, or is there something more?

15. Bone in Osteogenesis Imperfecta “Brittle Bone Disease” Mutations to one of the two collagen I genes lead to either abnormal or insufficient collagen in the bone. –Collagen I is 85-90% of the organic matrix of bone Classically leaves bones fragile, shortened, and bowed

16. Mild (Mild Nondeforming) Straight long bones i.e. no intrinsic long bone deformity.Straight long bones i.e. no intrinsic long bone deformity. Minimal vertebral crush fractures.Minimal vertebral crush fractures. Absence of chronic bone pain or minimal pain controlled by simple analgesicsAbsence of chronic bone pain or minimal pain controlled by simple analgesics Normal or near normal growth velocity and height.Normal or near normal growth velocity and height. Fully ambulant (other than at times of acute fracture).Fully ambulant (other than at times of acute fracture). Lumbar spine bone mineral density Z-score usually ≥ -2.0Lumbar spine bone mineral density Z-score usually ≥ -2.0 Moderate (Common Variable OI) Variable bowing of long bones related to immobilization for recurrent fractures.Variable bowing of long bones related to immobilization for recurrent fractures. Vertebral crush fractures.Vertebral crush fractures. Anterior bowing of legs and thighs.Anterior bowing of legs and thighs. Lumbar spine bone mineral density z – score ≤ - 2.5.Lumbar spine bone mineral density z – score ≤ - 2.5.

17. Severe OI (Progressively Deforming OI) Progressive deformity of long bones and spine (unrelated to fractures). Multiple vertebral crush fractures Marked impairment of linear growth Cases intermediate between severe and extremely severe have few rib fractures but crumpled long bones. Extremely Severe OI Continuously beaded ribs due to multiple fractures by birth. Crumpled (concertina-like) long bones. All vertebrae hypoplastic/crushed. Respiratory distress leading to perinatal death Perinatally lethal course

18. Treatments – Vitamin D Vitamin D is not a vitamin, it’s a sterol hormone!

19. Effects of Vitamin D Vitamin D wants to keep Calcium in the blood –Increases intestinal absorption of calcium –Decreases renal elimination of calcium –Increases resorption of calcium and phosphorus from bone Parathyroid hormone level is regulated by calcium level in blood, so low vitamin D causes high PTH

20. Effects of Vitamin D The aim is to keep the Vitamin D level within in the normal range (30-50 ng/mL) –Not helpful and likely harmful to have levels much above 50. This will prevent elevated PTH and decrease bone resorption Giving bisphosphonate to people who have very low Vitamin D levels can predispose to hypocalcemia

21. The Bisphosphonate Story In the 1930’s, it was discovered that polyphosphates were able to act as water softeners by inhibiting calcium salts crystals In the 1960’s, it was found that pyrophosphate exists in the body and it was hypothesized that this was the body’s own water softener. Initially, the desire was to inhibit abnormal calcification

22. The Bisphosphonate Story It was hypothesized that a disorder where pyrophosphate metabolism was abnormal would be detrimental to bone calcification. –Indeed this is correct, and the disorder is called hypophosphatasia Early attempts to exploit this discovery were disappointing because pyrophosphate is degraded by stomach acid

23. The Bisphosphonate Story Bisphosphonates were one of the experimental classes devised to be acid stable –Retained bone mineral affinity –Inhibited abnormal calcification Unexpectedly, the bisphosphonates were found to stabilize dissolution of hydroxyapatite crystals –Found that they inhibit bone resorption Experimentally found to improve osteoporosis

24. Pyrophosphate vs. Bisphosphonate

25. How do they work? Bisphosphonates are taken up by osteoclasts They directly inhibit a number of enzymes in the mevalonate pathway, which is in the cholesterol pathway –Inhibit production of farnesyl diphosphate (FPP) and geranylgeranyl diphosphate (GGPP) FPP and GGPP are crucial for the activation of certain proteins that regulate the function of osteoclasts

26. Treatments - Bisphosphonates First case report of bisphosphonate use in a patient with OI published in 1987 –Other single cases published in 1997 First systematic study of thirty children aged three to six with severe OI and use of pamidronate published in obscure journal from Massachusetts in 1998.

27. New England Journal of Medicine 1998 Oct 1;339(14):947-52. “Administration of pamidronate resulted in … a mean annualized increase of 41.9 percent in bone density and the deviation of bone mineral density improved from -5.3 to -3.4. The cortical width of metacarpals increased by 27 percent a year. The mean incidence of radiologically confirmed fracture decreased by 1.7 per year.”

28. Treatments - Bisphosphonates Dozens of similar studies suggest: –Increased density –Fewer fractures –Reduced bone pain –Improved functional status Limitations –Most studies observational, no really large RCTs –Not sure that it is ethical to do an RCT now

29. 35 Years of Medical, Surgical, and Rehabilitative Management Two children with comparable moderate OI, untreated 1975 vs Treated 2010 Treated from 15 months

30. “Good artists copy, great artists steal” Pablo Picasso

31. Caveats with Bisphosphonates Known side effects of bisphosphonates –Fever/flu-like reaction with first infusion –Hypocalcemia, which can be with any infusion Less common side effects –Osteonecrosis of the jaw –Bisphosphonate-associated inflammatory reaction

32. Caveats for Peroxisomal Disorders Side effect profile is much more significant –Bisphosphonate-associated inflammatory reaction seems to be more common High fever, malaise, low blood pressure –Vomiting appears to be more common –Hypocalcemia may be more common –Have not seen osteonecrosis of the jaw (nor do I expect to)

33. Suggestions for Bisphosphonate Therapy in Patients with PBDs Patients should be clinically well for at least one week prior to infusion Labs prior to infusion: Complete metabolic profile, phosphorus, magnesium, vitamin D level, urine N-telopeptides For initial infusions, patients should be admitted overnight between the first and second infusion

34. Suggestions for Bisphosphonate Therapy in Patients with PBDs For patients with adrenal insufficiency, I would consider stress dosing steroids during the first infusion. Acetaminophen and diphenhydramine should be given First dose in the first cycle between 0.125-0.25 mg/kg. Very low. Second dose in the first cycle between 0.25-0.5 mg/kg. Still fairly low. Ionized calcium after infusions

35. Suggestions for Bisphosphonate Therapy in Patients with PBDs Venue for subsequent infusions should depend on the success of the first infusion –Patients who sail initially should be fine in an infusion center If stress dose steroids are not given prior to the infusion, providers should have a low threshold for stress dose later if concerns develop

36. Suggestions for Bisphosphonate Therapy in Patients with PBDs Dosage for subsequent infusions –Less than two years old: 0.5 mg/kg q2m –Two to three years old: 0.75mg/kg q3m –Over three months: 1mg/kg q4m PBD protocol dose 3 mg/kg Omaha protocol dose 4.5 mg/kg/year Montreal protocol dose 9 mg/kg/year

37. Surveillance for Bone Density in PBDs Yearly DEXA while on treatment Same labs yearly as the patient had before infusions, including urine NTX Yearly vitamin D with 25-OHD level around 40 Yearly assessment for well-being and function by a provider familiar with PBDs

38. Thanks! Future scaring major at Monsters University Exhausted from watching big brother

39. References Nuttall JM, Hettema EH, Watts DJ. Farnesyl diphosphate synthase, the target for nitrogen-containing bisphosphonate drugs, is a peroxisomal enzyme in the model system Dictyostelium discoideum. Biochem J. 2012 Nov 1;447(3):353-61. doi: 10.1042/BJ20120750. PubMed PMID: 22849378; PubMed Central PMCID: PMC3465988. Poll-The BT, Engelen M. Peroxisomal leukoencephalopathy. Semin Neurol. 2012 Feb;32(1):42-50. doi: 10.1055/s-0032- 1306385. Epub 2012 Mar 15. Review. PubMed PMID: 22422205. Bone H. Future directions in osteoporosis therapeutics. Endocrinol Metab Clin North Am. 2012 Sep;41(3):655-61. doi: 10.1016/j.ecl.2012.05.003. Epub 2012 Jun 28. Review. PubMed PMID: 22877435. Diab DL, Watts NB. Bisphosphonates in the treatment of osteoporosis. Endocrinol Metab Clin North Am. 2012 Sep;41(3):487-506. doi: 10.1016/j.ecl.2012.04.007. Epub 2012 Jun 9. Review. PubMed PMID: 22877426. Nagotu S, Kalel VC, Erdmann R, Platta HW. Molecular basis of peroxisomal biogenesis disorders caused by defects in peroxisomal matrix protein import. Biochim Biophys Acta. 2012 Sep;1822(9):1326-36. doi: 10.1016/j.bbadis.2012.05.010. Epub 2012 May 19. Review. PubMed PMID: 22617146. Russell RG. Bisphosphonates: mode of action and pharmacology. Pediatrics. 2007 Mar;119 Suppl 2:S150-62. Review. Devogelaer JP, Malghem J, Maldague B, Nagant de Deuxchaisnes C. Radiological manifestations of bisphosphonate treatment with APD in a child suffering from osteogenesis imperfecta. Skeletal Radiol. 1987;16(5):360-3. Shaw NJ. Bisphosphonates in osteogenesis imperfecta. Arch Dis Child. 1997 Jul;77(1):92-3. Allgrove J. Bisphosphonates. Arch Dis Child. 1997 Jan;76(1):73-5. Williams CJ, Smith RA, Ball RJ, Wilkinson H. Hypercalcaemia in osteogenesis imperfecta treated with pamidronate. Arch Dis Child. 1997 Feb;76(2):169-70. Aström E, Söderhäll S. Beneficial effect of bisphosphonate during five years of treatment of severe osteogenesis imperfecta. Acta Paediatr. 1998 Jan;87(1):64-8. PubMed PMID: 9510450. Glorieux FH, Bishop NJ, Plotkin H, Chabot G, Lanoue G, Travers R. Cyclic administration of pamidronate in children with severe osteogenesis imperfecta. N Engl J Med. 1998 Oct 1;339(14):947-52.

40. References Fu L, Tang T, Miao Y, Zhang S, Qu Z, Dai K. Stimulation of osteogenic differentiation and inhibition of adipogenic differentiation in bone marrow stromal cells by alendronate via ERK and JNK activation. Bone. 2008 Jul;43(1):40-7. doi: 10.1016/j.bone.2008.03.008. Epub 2008 Mar 29. PubMed PMID: 18486585. Duque G, Li W, Adams M, Xu S, Phipps R. Effects of risedronate on bone marrow adipocytes in postmenopausal women. Osteoporos Int. 2011 May;22(5):1547-53. doi: 10.1007/s00198-010-1353-8. Epub 2010 Jul 27. PubMed PMID: 20661545. Pitts CJ, Kearns AE. Update on medications with adverse skeletal effects. Mayo Clin Proc. 2011 Apr;86(4):338-43; quiz 343. doi: 10.4065/mcp.2010.0636. Epub 2011 Mar 9. Review. PubMed PMID: 21389249; PubMed Central PMCID: PMC3068894. Di Martino MT, Arbitrio M, Guzzi PH, Leone E, Baudi F, Piro E, Prantera T, Cucinotto I, Calimeri T, Rossi M, Veltri P, Cannataro M, Tagliaferri P, Tassone P. A peroxisome proliferator-activated receptor gamma (PPARG) polymorphism is associated with zoledronic acid-related osteonecrosis of the jaw in multiple myeloma patients: analysis by DMET microarray profiling. Br J Haematol. 2011 Aug;154(4):529-33. doi: 10.1111/j.1365-2141. Id Boufker H, Lagneaux L, Fayyad-Kazan H, Badran B, Najar M, Wiedig M, Ghanem G, Laurent G, Body JJ, Journé F. Role of farnesoid X receptor (FXR) in the process of differentiation of bone marrow stromal cells into osteoblasts. Bone. 2011 Dec;49(6):1219-31. doi: 10.1016/j.bone.2011.08.013. Epub 2011 Aug 26. PubMed PMID: 21893226.2011.08622.x. Epub 2011 Apr 26. PubMed PMID: 21517810.