Developing and evaluating bone targeting agents to mimic the skeletal effects of mechanical loading

项目概要/摘要 骨质疏松症是一种全身性骨骼疾病，其特征是骨量减少， 骨组织退化，导致骨脆性和骨折敏感性增加。分之一 预计50岁以上的女性中有三分之一和男性中有五分之一会遭受骨折， 生活质量、疼痛、发病率和死亡率增加的限制。然而，大多数情况下 具有非骨选择性药物递送的骨骼系统疾病。内脏器官可能暴露于 大部分药物进入骨腔，而极低的药物浓度到达骨腔。 众所周知，机械负荷是可以增加骨形成和骨量的关键因素。 宇航员在太空飞行中由于重力降低和骨骼卸载而失去骨量。最近我们 证实抑制FAK（粘着斑激酶）模拟骨中机械负荷的作用 细胞（Sato et al. Nat Commun，2020）。然而，FAK及其同源物PYK 2在几乎所有的细胞中广泛表达， 组织和器官。因此，FAK抑制剂治疗可能在骨外引起严重的副作用。到 为了解决这一问题，我们将开发通过双膦酸盐（BP）缀合的骨靶向FAK抑制剂。 这一高风险、高回报的建议为肌肉骨骼研究带来了可观的潜在效益 领域(1)通过BP缀合的骨靶向将消除FAK抑制剂的骨骼外副作用。（二） 二膦酸盐可引起副作用，如颌骨骨坏死和细胞毒性。但我们的 新的药理学上无活性的BP解决了这一潜在问题。(3)BP结合物可能会增加 FAK抑制剂的功效比原来高出约10至1000倍的骨形成， 偶联方法递送到靶位点并选择性地以高浓度释放活性药物（FAK抑制剂）。 骨代谢部位。(4)BP-FAK抑制剂不会引起血清钙升高，尽管目前 骨形成诱导药物，如特立帕鲁肽（PTH 1 -34）和阿巴帕鲁肽（PTH类似物）。(5)FAK抑制剂 正在进行一些癌症治疗的临床试验。因此，双膦酸盐结合的FAK抑制剂 作为骨特异性骨形成诱导剂的临床相关药物。(6)这种BP共轭可以减少 由于双膦酸盐生物材料的性质，与羟基磷灰石结合并停留 在网站上。(7)我们的新配方策略可能会被用于解决口服可用性限制。 本提案的目的1将开发和评价BP偶联的骨靶向FAK抑制剂（VS-6063）， 体外（成骨细胞、破骨细胞和骨细胞）。在目标2中，我们将执行后肢卸载动物模型， 废用骨质疏松症来定义FAK抑制剂在骨中的治疗作用。这将提供数千个新的 候选FAK底物用于最终开发可药用BP-FAK抑制剂缀合物。采取 总之，这些研究将开发新的骨靶向药物，以模拟机械传导 并治疗固定引起的骨质流失。

Project Summary/ Abstract Osteoporosis is a systemic skeletal disease characterized by low bone mass and microarchitectural deterioration of bone tissue with a consequent increase in bone fragility and susceptibility to fracture. One in three women and one in five men over the age of 50 years are predicted to suffer a fracture leading to limitations in the quality of life, pain, morbidity, and increased mortality. However, it is difficult to treat most diseases of the skeletal system with non-bone selective drug delivery. Visceral organs may be exposed to the bulk of the pharmaceutical drug, while very low drug concentrations reach the bone compartment. Mechanical loading is well known as a key factor that can increase bone formation and bone mass. Astronauts lose bone mass during space flight due to reduced gravity and skeletal unloading. Recently, we demonstrated that inhibiting FAK (Focal adhesion kinase) mimics the effects of mechanical loading in bone cells (Sato et al. Nat Commun, 2020). However, FAK and its homolog, PYK2 are widely expressed in almost all tissues and organs. Therefore, FAK inhibitor treatment may cause severe side-effects outside of bone. To address this problem, we will develop bone-targeted FAK inhibitors via bisphosphonate (BP) conjugation. This high-risk, high-reward proposal brings considerable potential benefit to the musculoskeletal research field. (1) Bone targeting via BP-conjugation will eliminate extra-skeletal side effects of FAK inhibitors. (2) Bisphosphonates can cause side effects such as osteonecrosis of the jaw and cytotoxicities. However, our novel pharmacologically-inactive BP addresses this potential concern. (3) The BP-conjugates may increase FAK inhibitors' efficacy to around 10 to 1000 times higher bone formation than the original because this conjugation method delivers to the target sites and releases the active drug (FAK inhibitor) selectively at high bone metabolic sites. (4) There can be no serum calcium elevation by BP-FAK inhibitors, although current bone formation inducible drugs like teriparatide (PTH1-34) and abaloparatide (PTH analog). (5) FAK inhibitors are under several clinical trials for certain cancer therapies. So, bisphosphonate-conjugated FAK inhibitors are clinically relevant drugs as bone-specific bone formation induced agents. (6) This BP-conjugation can reduce administration frequencies due to bisphosphonate biomaterial property that binds to hydroxyapatite and stays on the sites. (7) Our new formulation strategies may be pursued to address oral availability limitations. Aim 1 of this proposal will develop and evaluate BP-conjugated bone-targeting FAK inhibitor (VS-6063) in vitro (osteoblasts, osteoclasts, and osteocytes). In aim 2, we will perform hindlimb unloading animlal model as disuse osteoporosis to define the therapeutic action of FAK inhibitors in bone. This will offer thousands of new candidate FAK substrates for the final development of a druggable BP-FAK inhibitor conjugate. Taken together, these studies will develop novel bone-targeted pharmacologic agents to mimic mechanotransduction and treat immobilization-induced bone loss.