Bridging the Gap Between Osteoporosis and Osteonecrosis of the Jaw: A Rationale for an Approach to Preventing and Treating BRONJ with MK4

John Neustadt, ND; and Steve Pieczenik, MD, PhD

Osteonecrosis of the jaw (ONJ) has been defined as “exposed bone in the mandible, maxilla, or both that persists for at least 8 weeks, in the absence of previous radiation and of metastases in the jaws.” ^1,2 ONJ caused by bisphosphonates (eg, Fosamax^®, Zometa^®, Aredia^®, Actonel^®), called bisphosphonate-related ONJ (BRONJ), is an iatrogenic disease and a significant concern to patients and dentists. In 2003, Marx described 36 cases of BRONJ.³ Since then, others have reported additional cases,^4-7 and many dental professionals, particularly oral and maxillofacial surgeons, have identified numerous unpublished cases.

The original publication by Marx reported 36 diagnosed cases under treatment,³ which increased to 76 cases as of 2005.⁸ Among the other publications, Migliorati reported 5 cases,⁴ Ruggerio et al reported 63 cases,⁷ Carter and Gross reported 4 cases,5 and Estilo et al reported 13 cases.⁶ In a previous article Marx also mentioned 43 additional cases reported to him by colleagues nationwide who have sought advice regarding prevention and management of this condition.⁸

Currently, bisphosphonates (BP) are the standard of care for millions of men and women with osteoporosis, and are also widely used for patients with Paget’s disease, bone malignancies, and multiple myeloma in patients with breast and prostate cancer. The rationale for using bisphosphonates in osteoporosis is that these medications increase bone mineral density and decrease fracture risk. Bone mineral density is enhanced by inhibiting resorption of trabecular bone by osteoclasts. Oral bisphosphonates are prescribed and include etidronate (Didronel^®) risedronate (Actonel), tiludronate (Skelid^®), alendronate (Fosamax), and ibandronate (Boniva^®). In 2006 alendronate was the most widely prescribed oral BP, accounting for 37.7% of the osteoporosis drug market and generating $2 billion in sales in the United States alone.⁹

The most potent bisphosphonates are delivered intravenously (IV) and are indicated to stabilize metastatic cancer (primarily breast and prostate) deposits in bone, and to treat the bone resorption defects of multiple myeloma and correct severe hypercalcemia. These are pamidronate (Aredia) and zoledronate (Zometa, Reclast^®). Additionally, intravenous zoledronate was approved by the FDA in 2007 for the treatment of postmenopausal osteoporosis.

In 2007 the risk of ONJ in patients with cancer treated with high doses of intravenously applied BPs was estimated in the range of 1% to 10%.¹ Until recently, BRONJ was claimed to be associated solely with intravenous bisphosphonate use. The relative risk of BRONJ from oral bisphosphonates has historically been calculated at 0.05 to one case each 100,000 persons-years of exposure for oral BPs.^1,2

However, a retrospective article published in 2009 concluded that the actual risk of BRONJ from oral bisphosphonates is 4%.¹⁰ This was the first large institutional study in the United States evaluating the epidemiology of BRONJ. Clinicians may find it instructive to understand the risk factors for BRONJ identified in this study (Table 1) so that they can best manage their patients on bisphosphonates.

Alarmingly, a more recent article published in 2011 concluded that the risk for BRONJ from oral bisphosphonates was actually 8%,¹¹ double what had been previously reported. The increased incidence of BRONJ was based on a retrospective analysis of data collected from 11 different clinical centers across Europe. Importantly, the majority of patients (57%) who experienced BRONJ had no risk factors as defined by the American Association of Oral and Maxillofacial Surgery.

Clinicians will undoubtedly encounter patients during their careers who are taking bisphosphonate medications. Recognizing the risk factors for BRONJ, understanding the pathophysiology of BRONJ, and enacting a rational approach to prevent BRONJ in patients is crucial to providing safe and effective treatments.

What the authors propose in this article is a rational approach for moving BRONJ away from a risk management and quality assurance model that is currently being used by dentists, to a preventive model. This approach as outlined herein admittedly requires additional study, including clinical trials, but the conceptual framework based on a broad review of the ONJ literature provides a way to begin to address this situation in a more proactive, as opposed to reactive, way.

The etiology of BRONJ has been extensively studied.^12-15 Although no consensus has yet been reached as to the underlying pathophysiology, hypotheses include decreased angiogenesis leading to avascular necrosis,^15,16 suppression of bone turnover due to attenuated osteocyte viability and resultant microfractures,^12,17,18 fibroblast suppression,¹⁹ immunosuppression,^20,21 decreased viability of oral keratinocytes,²² and oral microbial biofilms migrating to exposed jawbone from dental procedures (eg, tooth extraction).²³ All of this leads to a decreased wound healing that could contribute to the initiation and propagation of BRONJ.²

While it is the authors’ position that these pathological variables are involved in BRONJ, they firmly believe that decreased collagen production is the initiating factor in BRONJ, from which all other pathophysiological mechanisms follow. Although the possibility of this was hinted at in a 2010 study by German researchers Simon and Niehoff et al,¹⁴ until now it had not been incorporated into a comprehensive model for BRONJ.

The Neustadt-Pieczenik Collagen Deficit and Restoration Hypothesis

What is now just starting to emerge in dental research, but has been well described in the general medical literature on osteoporosis and bone histology, is the preeminent role collagen plays in bone health, fracture healing, and fracture prevention. Bone matrix is a two-phase system in which the mineral phase provides the stiffness and the collagen fibers provide the ductility and ability to absorb energy (ie, the toughness). Alterations of collagen properties can therefore affect the mechanical properties of bone and increase fracture susceptibility.²⁴

Type I collagen is the most abundant type of collagen and is widely distributed in almost all connective tissues with the exception of hyaline cartilage. It is the major protein in bone, skin, tendon, ligament, sclera, cornea, and blood vessels. Type I collagen comprises approximately 95% of the entire collagen content of bone and about 80% of the total proteins present in bone.²⁵

Maciej J.K. Simon, Jorg Wiltfang, and Yahya Acil from the Department of Oral and Maxillofacial Surgery, and Peter Niehoff and Bernard Kimig from the Department of Radiotherapy (Radiooncology), University Medical Center Schleswig-Holstein, Kiel, Germany, investigated the effects of 28 days of zoledronate and pamidronate exposure on collagen production in gingival fibroblasts, osteoblasts, and human osteogenic osteosarcoma cells (SaOS-2) in vitro.¹⁴ To better relate the scientific research to clinical processes the researchers used the most common drugs associated with ONJ: zoledronic acid and pamidronate.

Human osteoblasts and fibroblasts were isolated from iliac crest and gingiva, respectively, of healthy patients who were not taking bisphosphonate medications. Untreated cells were used as the control group and inorganic pyrophosphate (PP) was used as a positive control. Cells were cultured for 28 days in growth medium. Each cell line was cultured in 10 groups of six-well plates. Every third day the growth medium was renewed with zoledronic acid, pamidronate, and PP at four different concentrations each: 1 µm, 5 µm, 10 µm, and 20 µm. Gene expression was evaluated by real-time polymerase chain reaction (PCR) (RT-PCR) after reverse transcription of the corresponding mRNA. Collagen expression was analyzed by enzyme-linked immunosorbent assay (ELISA).

At higher than 1 µm, zoledronic acid and pamidronate were toxic to cells. Thus, higher concentrations yielded insufficient gene expression and collagen production for analyses. At all concentrations used, zolendronate reduced gene expression to < 16%. Thus, gene amplification was only possible for cells cultured in 1 µm pamidronate, which did not suppress gene expression as much as zoledronic acid. Pamidronate significantly reduced gene expression compared to control (P < 0.05). Fibroblasts showed a maximum of 31% gene expression, osteoblasts 56%, and SaOS-2 14%. The mean expression of type I collagen production significantly decreased in all cell lines (P < 0.05) compared to control by treatment with 1 µm each zolendronic acid and pamidronate.

This study demonstrated for the first time that zolendronic acid and pamidronate are directly toxic to osteoblasts, fibroblasts, and SaOS-2 cells and significantly negatively affect collagen production. The reduction in extracellular matrix production of these cell lines by BP exposure could possibly explain why patients experience BRONJ and decreased wound healing. However, it is unclear why osteonecrosis is mainly found in the jaws and not other bones in these patients. Since many factors can influence the clinical outcomes and negative sequelae of BP therapy, further investigation needs to be pursued to verify these findings, test other BPs such as alendronate and ibandronate, and further investigate the clinical implications of these data. Notwithstanding these limitations, in this study type I collagen was reduced in all three cell lines treated with zoledronate and pamidronate, which provides compelling in vitro evidence for the possible in vivo effects of the collagen-destroying potential of BPs.

Decreased bone collagen production as indicated by bone resorption markers leads to increased susceptibility to fractures and osteoporosis. Markers of bone turnover, particularly markers of bone resorption, have been shown in prospective epidemiologic studies to be associated with fracture risk,^26-31 and this association appears to be independent of bone mineral density (BMD).³²

The limitations of BMD in predicting fracture risk are well documented in multiple studies. In 1996 a meta-analysis of prospective cohort studies totaling about 90,000 person years of observation time and more than 2,000 fractures was published in the British Medical Journal.³³ The researchers concluded that BMD predicts less than 50% of patients who progress to fractures. Another study published in 2004 that analyzed data from 7,806 men and women from the Rotterdam Study, a prospective, population-based cohort study of men and women aged 55 years and older, concluded bone density scans only predict 44% of women and 21% of men age 65 and older who progress to a nonvertebral fracture.³⁴

In a larger meta-analysis of 15,259 men and 44,902 women from 13 different cohorts totaling 250,000 person years published in 2004, researchers evaluated the 10-year risk of a fragility fracture in postmenopausal women with osteoporosis (T scores of –2.5 or less).³⁵ They concluded that in men and women the ability of bone density scans to predict hip fractures was only 22%. More recently, in 2006 the North American Menopause Society (NAMS) published a position statement on the predictive value of bone density scans in which they concluded fracture risk “depends largely on factors other than BMD.” ³⁶

Since collagen is not detected by x-rays it is not part of a bone density scan report. Thus, in only reporting the density of bones the scans are evaluating bone quantity and not bone quality. Any discussion of bone quality would be incomplete without considering the pivotal role of bone collagen.

Developed largely from research on osteoporosis, the Neustadt-Pieczenik Collagen Deficit and Restoration Hypothesis posits that collagen destruction represents the keystone for understanding, preventing, and treating osteoporosis and BRONJ. With respect to BRONJ, the authors’ hypothesis is that a “collagen deficit” creates a microenvironment more susceptible to microfractures from mastication. This in turn provides a pocket for bacteria to seed and reproduce. While decreased angiogenesis and immune modulation may contribute to collagen degradation and necrotizing tissue infections with a markedly impaired wound healing capacity in BRONJ, if collagen degradation itself can be prevented and reversed, BRONJ may be effectively prevented and treated. This is the essence of the Neustadt-Pieczenik Collagen Deficit and Restoration Hypothesis and its application to BRONJ.

In osteoporosis research, vitamin K has been extensively studied for its ability to stimulate collagen production, promote bone health, and decrease fracture risk. Vitamin K is a group of structurally similar, lipid-soluble, 2-methyl-1,4-napthoquinones, which include phylloquinone (K1), menaquinones (K2), and menadione (K3).³⁷ Plants synthesize vitamin K1 while bacteria can produce a range of vitamin K2 forms, including the conversion of K1 to K2 by bacteria in the small intestines. Vitamin K3 is synthetic and, because of its toxicity, has been banned by the US Food and Drug Administration for use in dietary supplements, but remains in limited use as an antineoplastic agent. In contrast to vitamin K3, no known toxicity exists for the vitamin K1 and K2 forms.

There are two major forms of vitamin K2. These are menaquinone-4 (menatetrenone, MK4) and menaquionone-7 (MK7). Among the vitamin K analogues, the form most researched for osteoporosis treatment and bone health is MK4. MK4 is produced via conversion of K1 in the body, in the testes, pancreas, and arterial walls.³⁸ While major questions still surround the biochemical pathway for the transformation of K1 to MK4, studies demonstrate that the conversion is not dependent on gut bacteria, occurring in germ-free rats^39,40 and in parenterally administered K1 in rats.^41,42 In fact, tissues that accumulate high amounts of MK4 have a remarkable capacity to convert up to 90% of the available K1 into MK4.^39,40 Recently it has been shown that K1 is converted in vitroto MK4 in the MG-63 human osteoblastic cell line⁴³ and that in humans the UbiA prenyltransferase containing 1 (UBIAD1) enzyme is responsible for conversion of K1 to MK4.⁴⁴

MK7 is instead not produced in humans, but converted from phylloquinone in the intestines by gut microbiota.⁴⁵ However, bacteria-derived menaquinones appear to contribute minimally to overall vitamin K status.^46,47 MK4 and MK7 are both found in the United States in dietary supplements for bone health. The US FDA has not approved any form of vitamin K for the prevention or treatment of osteoporosis; however, MK4 and MK7 each beneficially affects surrogate markers of bone quality and health.

The molecular mechanisms for the beneficial effects of vitamin K on bone health and other conditions has been extensively studied. The family of K vitamins, including phylloquinone, MK4 and MK7, are fat-soluble vitamins that act as coenzymes for a vitamin K-dependent carboxylase enzyme that catalyzes carboxylation of the amino acid glutamic acid, resulting in its conversion to gamma-carboxyglutamic acid (Gla). This carboxylation reaction is essential for formation of bone collagen, which allows bone to deform upon impact, for example, during mastication or a fall, without fracturing. Although vitamin K-dependent gamma-carboxylation occurs only on specific glutamic acid residues in a small number of proteins, it is critical to the calcium-binding function of those proteins.

Three vitamin K-dependent proteins have been isolated in bone: osteocalcin, matrix Gla protein (MGP), and protein S. Osteocalcin is a protein that is synthesized by osteoblasts and is regulated by the active form of vitamin D, 1,25-(OH)2D3, also called calcitriol. The mineral-binding capacity of osteocalcin requires vitamin K-dependent gamma-carboxylation of 3 glutamic acid residues.

Elevated uncerbaxoylated osteocalcin (ucOC) is associated with increased fracture risk,^48,49 and phylloquinoine,⁵⁰ MK4,^51-54 and MK7^55,56 have all been shown in clinical trials to reduce ucOC.

This classical view that the osteoprotective effects of vitamin K are solely due to its effects on osteocalcin and MGP may not be correct, or at least incomplete. In a 1996 letter published in Nature researchers bred osteocalcin-deficient mice to compare the effect on bone histomorphology in animals that are unable to synthesize osteocalcin.⁵⁷ Compared to wild type mice the genetically altered, osteocalcin-deficient mice demonstrated greater bone strength at 6 months of age without any differences in bone mineralization. This suggests that the skeletal benefits of vitamin K may be mediated by another pathway.

While studies showed an inverse correlation with ucOC and fracture risk and all three forms of vitamin K decrease ucOC, ucOC is a surrogate marker. As such, its value is limited and it may actually not explain or predict the fracture-prevention effects of vitamin K.

In addition to stimulating carboxylation of osteocalcin, vitamin K also has signal transduction and transcriptional regulatory functions. MK4 and MK7 both induce osteoblast differentiation by binding to and activating the steroid and xenobiotic receptor (SXR)/pregnane X receptor (PXR) in vitro, which upregulates alkaline phosphatase and MGP.^58-60 And recently MK4 and MK7 were both shown in vitroto inhibit nuclear factor κB (NF-κB), which is involved in osteoclastogenesis, function, and survival.⁶¹

The most clinically relevant end-point is reduction of fractures in prospective, randomized clinical trials. In this regard only MK4 has demonstrated the ability to decrease fractures in clinical trials.^54,62,63 In addition to decreasing fractures by up to 87%, clinical trials in Japan and Indonesia show that MK4 attenuates medication- and disease-induced bone loss as determined by bone density scans.^{51,53,54,62-75} In Japan, MK4 has been approved by the Ministry of Health for the prevention and treatment of osteoporosis since 1995.⁶⁶

It is important to note that vitamin K1 is preferentially used by the liver as a clotting factor. Vitamin K2 on the other hand is used preferentially in other organs, such as the brain, vasculature, breasts, and kidneys. Coagulation studies in humans using 45 mg/day of vitamin K2 (as MK4)53 and even up to 135 mg/day (45 mg tid) of K2 (as MK4),⁷⁶ showed no significant increase in pathologic coagulation risk. Even doses in rats as high as 250 mg/kg body weight did not alter the tendency for blood-clot formation to occur.⁷⁷

The one caveat to this is for people taking the blood-clotting medication Coumadin^® (warfarin). Since warfarin, which was originally used as a rat poison, decreases blood-clot risk by interrupting the vitamin K-dependent clotting factors, taking vitamin K in any amount may interfere with the actions of warfarin and increase blood-clot risk.

MK4 and Fracture Risk and Bone Loss Clinical Trials

Since the fundamental premise of the Neustadt-Pieczenik Collagen Deficit and Restoration hypothesis is that preventing and treating osteoporosis and BRONJ may be successfully done by stimulating bone collage production, all three forms (phylloquinone, MK4, and MK7) may be useful in this regard. However, since microfractures are a component of BRONJ pathogenesis^12,17,18 and only MK4 has been shown to reduce fractures it thus represents the leading therapeutic candidate for preventing and treating osteoporosis and BRONJ.

A clinical trial published in the Journal of Bone and Mineral Research in 2000, evaluated the efficacy of 45 mg daily MK4 for reducing fracture risk and improving bone mineral density.⁵⁴ This randomized, controlled, open-label study lasted 2 years and enrolled 241 osteoporotic women (average age 67 years). The control group (n = 121) received 150 mg elemental calcium per day, while the treatment group (n = 120) consumed 150 mg elemental calcium plus 45 mg MK4 daily. After 24 months of treatment, the control group sustained 30 vertebral fractures for a fracture rate of 30.3%, while the treatment group sustained 13 vertebral fractures for a fracture rate of 10.9%. The decrease in fractures in the MK4 group was highly statistically significant (p = 0.0273). Similarly, the control group exhibited two fractures in the femoral neck, compared to no femur fractures in the treatment group.

A second clinical trial published in 2005 in the journal Bone, evaluated the ability of MK4 to reduce fractures in elderly Alzheimer’s patients in a randomized controlled study design.⁶² This study enrolled 200 ambulatory women with Alzheimer’s disease (average age 78 years) for the treatment group. These women (n = 200) received daily 45 mg MK4, 1000 IU vitamin D2 and 600 mg calcium. The control group (n = 100) was recruited from healthy, active, elderly women in the community without cognitive deficits, and received no intervention.

After 2 years of the study, the control group suffered 22 fractures (15 hip, 2 distal forearm, 2 proximal femur, 1 each at proximal humerus, ribs, and pelvis), while there were only three fractures (2 hip, 1 proximal femur) sustained in the treatment group. Overall there were 86% fewer nonvertebral fractures (p = 0.0003) in the treatment group compared to controls and 87% fewer hip fractures (p = 0.0022) in the treatment group compared to controls.

Importantly, in this study there were no differences in the number of falls sustained in the treatment group compared to the control group. Thus, it is concluded that MK4 reduces fractures independent of the number of falls someone sustains. The importance of this cannot be underestimated, since falling is the number one cause of osteoporotic fractures and death from osteoporosis. This decrease in fractures independent of the number of falls someone sustains is likely due to MK4 stimulating collagen production in bone and not simply adding calcium to bones. Collagen provides flexibility to the bone, while calcium provides rigidity. When someone falls, collagen allows the bone to absorb and disperse the force of the fall without breaking.

A 2006 meta-analysis published in the Archives of Internal Medicine by Cockayne et al at the University of York in England evaluated clinical trials on MK4 and fracture risk.⁶³ They identified 13 randomized, controlled trials of the effect of MK4 on osteoporosis. Of those, seven had fracture risk as an end point and thus were included in their meta-analysis. They concluded that 45 mg of MK4 decreases vertebral fracture by 60%, hip fracture by 73%, and all nonvertebral fractures by 81%.

MK4 has also been shown to stop and reverse bone loss, and reduce fracture risk, from medical conditions and medications. In clinical trials MK4 (45 mg daily) prevented bone loss and/or fractures caused by corticosteroids (eg, prednisone, dexamethasone, prednisolone), anorexia nervosa, cirrhosis of the liver, osteoporosis, menopause (estrogen deficiency), disuse from stroke, immobilization (eg, extended illness, hospitalization), Parkinson disease, phenytoin therapy, testosterone deficiency (eg, aging, prostate cancer treatment), primary biliary cirrhosis, leuprolide treatment (for prostate cancer), and other diseases and medications.^53,65-75

MK4 may also decrease fracture healing time by stimulating bone collagen production. In a letter to Nature in 1960, Bouckaert and Said reported that complete transverse fractures experimentally induced into rabbit and rat femurs completely healed in the presence of vitamin K.⁷⁸ While no clinical trials to date have been conducted to confirm this finding, the aforementioned basic science provides the underlying biochemical understanding for their observation. Hoefert and Schmitz et al¹² provide compelling evidence that jaw microcracks are a fundamental requirement for the etiology of BRONJ. Promoting collagen production using MK4 may decrease the predisposition to microfractures and promote the healing of microfractures when they do occur, thereby playing a fundamental role in preventing and treating BRONJ.

Conclusion

In conclusion, the role of collagen as elaborated in the Neustadt-Pieczenik Collagen Deficit and Restoration Hypothesis postulated in this article cannot be ignored in the pathophysiology and potential prevention and treatment of osteoporosis and BRONJ. Collagen degradation is important in the pathogenesis of osteoporosis and likely plays a pivotal role in the etiology of BRONJ as well. Studies from the medical literature support the safety and efficacy of MK4 as a potential therapeutic agent in preventing and treating osteoporosis and BRONJ and should be the subject of future research.

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About the Authors

John Neustadt, ND
Medical Director
Montana Integrative Medicine, Inc.

President
Nutritional Biochemistry, Inc. (NBI)
Bozeman, Montana

Steve Pieczenik, MD, PhD
Board Certified Psychiatrist
Chairman of the Board
Nutritional Biochemistry, Inc. (NBI)
Bozeman, Montana