Design and synthesis of bone-selective osteogenic oxysterol-bisphosphonate analogues

 描述(由申请人提供)：骨质疏松症直接影响1000万美国人，另有3400万人患有骨质疏松症并面临发展为骨质疏松症的风险。双膦酸类药物，例如阿仑膦酸(ALN，福善美)，可以通过减缓破骨细胞性骨吸收来提高骨密度和降低骨折风险；然而，许多现有的抗吸收疗法都存在不良副作用和临床疗效有限的问题。对于骨质疏松症的治疗干预需要新的和改进的策略，特别是安全地促进合成代谢骨生长的新疗法。同时解决骨吸收和骨形成的双重治疗方法也可能有所帮助。目前，只有一种FDA批准的骨合成代谢药物Forteo(Teriparatide)在治疗骨质疏松症方面具有显著的临床益处，但出于安全考虑，其使用受到严格限制。多能间充质干细胞(MSCs)是多种细胞类型的前体，包括成骨细胞和脂肪细胞。新骨的形成是由骨髓间充质干细胞的成骨分化推动的，这一过程可能会因年龄、生活方式因素和更年期发生的激素变化而失去平衡。Parhami等人。研究发现，当将特定的氧化甾醇应用于骨髓间充质干细胞时，可以诱导成骨，同时抑制其脂肪生成。到目前为止，最有前途的专利半合成氧固醇OXY133在体外显示出更强的成骨分化能力，包括在原代大鼠、兔和人MSCs中，并在大鼠和兔脊柱融合以及颅骨和股骨缺损模型中刺激体内强劲的局部骨形成。在SBIR第一阶段研究期间，我们已经开始评估骨合成代谢Oxy133和阿仑磷酸钠(ALN)的药物结合物，ALN是一种久负盛名的抗吸收药物，也是一种骨靶向剂。我们已经设计出了化学偶联的方法，并表征了所得偶联物的生物物理和生物学性质。Oxy133-ALN结合物在体外表现出与骨矿物质的强烈结合，并刺激Hedgehog(HH)信号通路和MSCs的成骨。在这一应用中，我们建议进一步开发Oxy133-ALN结合物，作为治疗骨质疏松症的潜在双重治疗药物，促进成骨细胞的骨形成(Oxy133的功能)和抑制破骨细胞的骨吸收(ALN的功能)。在我们成功的第一阶段研究的基础上，我们建议进行第二阶段的研究，作为三个具体目标的一部分：目标1：开发可扩展的合成Oxy133-ALN结合物的方法。目的：评价Oxy133-ALN结合物对破骨细胞性骨吸收的抑制作用及双重治疗的可能性。目的：测定Oxy133-ALN结合物的组织分布特性，并评价部分Oxy133-ALN结合物在OVX小鼠模型中的疗效。

 DESCRIPTION (provided by applicant): Osteoporosis directly affects 10 million Americans and another 34 million are osteopenic and at risk for developing osteoporosis. Bisphosphonate drugs, for example alendronic acid (ALN, Fosamax), can improve bone density and reduce fracture risk by slowing osteoclastic bone resorption; however, many of the existing anti-resorptive therapies are plagued with untoward side effects and limited duration of clinical benefits. New and improved strategies for therapeutic intervention in osteoporosis are needed, particularly with new treatments that safely promote anabolic bone growth. A dual therapy approach, addressing both resorption and formation of bone could also be helpful. Presently, there is only one FDA approved bone anabolic agent, Forteo (teriparatide), that confers significant clinical benefits in osteoporosis, but its use is severely restricted due to safety concerns. Multipotent mesenchymal stem cells (MSCs) are precursors of a variety of cell types, including osteoblasts and adipocytes. Formation of new bone is driven by osteoblastic differentiation of MSCs, a process that can be thrown off balance by age, lifestyle factors and hormonal changes that occur with menopause. Parhami et al. discovered that specific oxysterols induce osteogenesis when applied to MSCs while inhibiting their adipogenesis. The most promising proprietary semi-synthetic oxysterol to date, OXY133, displays increased potency for osteogenic differentiation in vitro, including in primary rat, rabbit, and human MSCs, and it stimulates robust localized bone formation in vivo in rat and rabbit spine fusion and crania and femoral defect models. During SBIR Phase I research, we have begun evaluating drug conjugates of osteo- anabolic Oxy133 and Alendronate (ALN), a well-established anti-resorptive drug that also serves as a bone- targeting agent. We have worked out methods for chemical conjugation and characterized biophysical and biological properties of the resulting conjugates. Oxy133-ALN conjugates display strong in vitro binding to bone mineral and stimulate Hedgehog (Hh) pathway signaling and osteogenesis in MSCs. In this application, we propose to further develop Oxy133-ALN conjugates as potential dual therapy agents for osteoporosis, stimulating bone formation by osteoblasts (function of Oxy133), and inhibiting bone resorption by osteoclasts (function of ALN). Expanding on our successful Phase I studies, we propose to perform Phase II studies as part of 3 Specific Aims: Aim 1: Development of scalable methods for the synthesis of Oxy133-ALN conjugates. Aim 2: Evaluation of the inhibition of osteoclastic bone-resorption by Oxy133-ALN conjugates and the possibility of a dual therapy. Aim3: Determination of Oxy133-ALN conjugate tissue distribution properties and evaluation of select Oxy133- ALN conjugates for efficacy in an OVX mouse model.