Pathophysiology of Biphosphonate-Induced ONJ in a Novel Animal Model

描述（由申请方提供）：近年来，颌骨骨坏死（ONJ）已被确定为双膦酸盐（BP）治疗的潜在并发症。这种骨疾病的特征是坏死骨的积累，暴露在口腔中持续6至8周以上。ONJ最常见于使用强效氮（N-）BP唑来膦酸盐（ZOL）或帕米膦酸盐治疗的癌症患者中，用于抑制骨转移，但在阿仑膦酸盐（ALN）治疗的骨质疏松患者中也报告了较低程度的ONJ。虽然多个病例报告研究表明ONJ的发病率和严重程度与N-BP抑制骨吸收的效力和这些药物的累积剂量相关，但尚未建立基于机制的因果关系。因此，开发动物模型来研究ONJ的病理生理学将是增强我们对这种骨疾病理解的基本工具。我们的长期目标是阐明ONJ的病因和病理生理学，以便为新的有效预防和治疗策略奠定基础。本提案的目的是开发ONJ的动物模型，并确定不同效力的N-BP对坏死骨去除和骨愈合的影响。我们的中心假设是ONJ是一个两阶段过程：1）风险因素启动口腔中导致硬组织坏死的过程; 2）N-BP，已被充分证明可抑制骨吸收，阻止坏死骨的清除，导致坏死骨在颌骨中积聚，其本身延迟骨愈合的启动和后续进展。我们计划利用稻鼠来验证我们的中心假设（Oryzomys palustris）与已建立的牙周炎，并确定ZOL和ALN的影响：1）通过进行牙周病的离体评估，包括牙槽骨的放射学评估、软组织受累和牙齿移动性的评估，来评估牙周损伤的进展，2）通过使用组织化学/组织学技术评估坏死骨积聚来去除坏死骨，并且还通过使用定量组织形态计量学评估骨吸收指数; 3）通过使用定量组织形态计量学和免疫组织化学技术在牙周组织中作为骨愈合的指标的新骨形成和血管生成;和4）使用真实的时间PCR技术与骨吸收、骨形成和血管生成相关的基因的表达。我们推测，众所周知的和理想的抗吸收作用的N-BP在ONJ的发展中具有核心作用。因此，在本项目完成时，我们期望提供直接证据，证明与效力较低的BP ALN相比，强效BP ZOL将在颌骨中诱导更高的坏死骨蓄积，随后会损害骨愈合。我们也期望开发一种ONJ的动物模型，以促进我们对这种骨疾病的认识和理解的进一步发展。项目叙述：颌骨骨坏死（ONJ）是双膦酸盐（BP）治疗的潜在不良副作用。多项病例报告研究表明，ONJ的发生率和严重程度与BP的效力和药物的累积剂量有关。然而，这种骨疾病是知之甚少，部分原因是缺乏适当的动物模型ONJ。拟议的研究将提供深入了解组织水平的机制，为ONJ样病变的发展，沼泽稻大鼠，牙周炎的动物模型，并将阐明的基础上，在细胞和分子水平上，更高的发病率的BP唑来膦酸盐诱导ONJ样病变相比，BP阿仑膦酸盐。

DESCRIPTION (provided by applicant): In recent years, osteonecrosis of the jaw (ONJ) has been identified as a potential complication of bisphosphonate (BP) treatment. This bone disorder is characterized by the accumulation of necrotic bone with exposure in the oral cavity that persists for more than 6 to 8 weeks. ONJ is observed most frequently in cancer patients treated with the potent nitrogen (N-)BPs zoledronate (ZOL) or pamidronate for inhibition of bone metastases, but it has also been reported to a much lesser extent in alendronate (ALN)-treated osteoporotic patients. While multiple case report studies suggest that the incidence and severity of ONJ are associated with the potency of N-BPs to inhibit osteoclastic bone resorption and cumulative dose of these drugs, no mechanism-based cause and effect relationship has yet been established. Therefore, the development of animal models to study the pathophysiology of ONJ will be a fundamental tool to enhance our understanding of this bone disorder. Our long-term goal is to elucidate the causes and pathophysiology of ONJ in order to create the basis for new effective strategies for prevention and treatment. The objectives of this proposal are to develop an animal model for ONJ, and to establish the effects of N-BPs of different potencies on the removal of necrotic bone and bone healing. Our central hypothesis is that ONJ is a two-stage process: 1) risk factors initiate processes in the oral cavity that cause necrosis of hard tissues; and 2) N-BPs, well-proven to inhibit osteoclastic resorption, block the removal of necrotic bone, causing an accumulation of necrotic bone in the jaw that in and of itself delays the initiation and subsequent progress of bone healing. We plan to test our central hypothesis by utilizing rice rats (Oryzomys palustris) with established periodontitis and determine the effects of ZOL and ALN on: 1) the progression of the periodontal damage by performing an ex-vivo evaluation of the periodontal disease, including a radiographic assessment of the alveolar bone, the evaluation of soft tissue involvement and tooth mobility, and by conducting a histometric analysis of the periodontal tissue; 2) the removal of necrotic bone by the assessment of necrotic bone accumulation using histochemical/histological techniques, and also on indices of bone resorption by using quantitative histomorphometry; 3) new bone formation and angiogenesis, as indices of bone healing, in the periodontium by using quantitative histomorphometry and immunohistochemical techniques; and 4) the expression of genes associated with bone resorption, bone formation, and angiogenesis using real time PCR techniques. We postulate that the well- known and desirable anti-resorptive effect of N-BPs has a central role in the development of ONJ. Consequently, at the completion of this project, it is our expectation to provide direct evidence that the potent BP ZOL will induce a higher accumulation of necrotic bone in the jaw, with subsequent impairment of bone healing as compared to that induced by the less potent BP ALN. It is also our expectation to develop an animal model for ONJ that will facilitate further advances in our knowledge and understanding of this bone disorder. Project Narrative: Osteonecrosis of the jaw (ONJ) is a potential adverse side effect of bisphosphonate (BP) treatment. Multiple case report studies suggest that the incidence and severity of ONJ are associated with the potency of BP and cumulative dose of the drug. However, this bone disorder is poorly understood, due in part to lack of appropriate animal models for ONJ. The proposed research will provide insight into a tissue-level mechanism for the development of ONJ-like lesions in marsh rice rats, an animal model for periodontitis, and will elucidate the basis, at the cellular and molecular levels, for the higher incidence of the BP zoledronate to induce ONJ- like lesions as compared with the BP alendronate.