Recombinant expression of a biomaterial-targeting BMP-2 containing a peptide tag

含有肽标签的生物材料靶向 BMP-2 的重组表达

• 批准号: 7990589
• 负责人: Bruce Lamb
• 金额: $ 31.22万
• 依托单位: AFFINERGY ,INC
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7990589
• 关键词: Address; Adoption; Adverse event; Affinity; Animal Model; Area; Autologous Transplantation; Binding; Binding Sites; Biochemical; Biocompatible Materials; Biological; Biological Assay; Bone Substitutes; Bone Transplantation; Cell Density; Cells; Cervical; Cervical spine; Chimeric Proteins; Chinese Hamster Ovary Cell; Clinical; Code; Collagen; Complementary DNA; Culture Media; Defect; Development; Diffuse; Dose; Drug Formulations; Ensure; Esophagus; Exhibits; Filler; Goals; Growth Factor; Implant; Incidence; Intellectual Property; Label; Link; Marketing; Mediating; Medical; Orthopedics; Osteogenesis; Patients; Peptides; Pharmaceutical Preparations; Phase; Physicians; Physiological; Population; Porifera; Preclinical Testing; Preparation; Procedures; Production; Proteins; Recombinant Growth Factor; Recombinant Proteins; Recombinant Transgenes; Recombinants; Research; Safety; Sales; Science; Signal Transduction; Site; Spinal Fusion; System; Technology; Testing; Therapeutic; Time; Trachea; Transplanted tissue; Vertebral column; Work; base; bone; bone morphogenetic protein 2; design; dosage; expiration; expression cloning; expression vector; implantation; improved; in vitro Assay; in vitro activity; in vitro testing; milligram; novel; novel therapeutic intervention; osteogenic; osteogenic protein; programs; public health relevance; response; small molecule; tricalcium phosphate; vector

DESCRIPTION (provided by applicant): The emergence of biological therapeutics, including cells, peptides, and proteins, has allowed medical science to address a number of unmet clinical needs. Often these agents are extremely potent, specifically activating endogenous signaling cascades to promote well-characterized biochemical and physiological responses. For example, bone morphogenetic protein-2 (BMP-2) was identified almost 20 years ago as a powerful initiator of osteogenic activity, and has since become part of a leading orthopedic product used for spinal fusion procedures (InFuse from Medtronic). Despite annual sales of nearly $1B, BMP-2 has been shown to trigger bone formation at ectopic sites, such as the trachea or esophagus, when not used precisely as directed by its labeling. So-called off-label use of BMP-2 in cervical spinal fusion procedures has led to numerous patient complications, culminating in an FDA-issued warning to physicians advising against off-label use. Two major issues have led to these complications: 1) large doses of BMP-2 are used in these procedures and 2) the drug is added to a collagen sponge for which BMP-2 has only limited passive affinity. These factors are inextricably linked, in that large doses are applied to ensure that a therapeutic dose is retained over time as the protein diffuses away from the delivery site. Due to these safety concerns, BMP is used in approximately 7-10% of the 237,000 cervical fusion procedures performed annually, as opposed to over 60% of lumbar fusion procedures. Enabling the use of BMP-2 in cervical procedures would result in the near doubling of the $1B market for orthobiologics. Using our core technology, Affinergy has developed a number of peptide-based linker motifs, capable of binding to a range of biomaterials with high affinity. Targets for available peptide linkers include bone substitute materials such as collagen and tricalcium phosphate. Here, we propose the development of a novel transgene for the recombinant production of a BMP-2 protein containing a peptide linker. The resulting molecule will both retain BMP-2 activity and exhibit improved affinity for bone grafting materials over unmodified BMP-2. We believe this product will address both currently unmet needs for BMP-2 delivery by ensuring a therapeutic concentration of protein remains bound to the delivery matrix over time, thereby reducing the initial dosage required. This program is also well-timed, with the impending expiration of considerable intellectual property surrounding BMP-2 within the next 2-3 years. The proposed research program will require: 1) the construction of a BMP-2 expression vector containing the peptide-linker domains within the coding sequence; 2) the establishment of an expression clone to ensure the reproducible production of the protein and 3) in vitro testing to determine the activity of peptide-tagged BMP-2 as well as its affinity for bone substitute materials. When successful, Phase II work would include large-scale culture optimization, testing the modified BMP-2 in a critical-sized defect animal model, and a range of preclinical testing. Affinergy believes that partners, such as large orthopedics companies, would seize the opportunity to market this product. PUBLIC HEALTH RELEVANCE: The use of osteogenic proteins in spinal fusion has given rise to a $1B market in spine orthobiologics. While the use of these products continues to grow, certain indications are not able to employ these products due to a range of safety concerns. Here, we propose the incorporation of a peptide tag into the widely used orthobiologic, BMP-2. This peptide will contain a high affinity binding site for bone void filler materials currently used as carriers for BMP-2 in spinal fusion. We believe this product will (1) be restricted to the implantation site, and (2) allow for a smaller dose of BMP-2; these improvements mitigate BMP-2 safety issues such as ectopic bone formation. In addition, this product would enable a more wide-spread and safer use of BMP-2 in cervical spinal fusions, an indication in which orthobiologics are used in only 7-10% of cases because of the known increased incidence of adverse events when used in the cervical spine fusions. The use of Affinergy's proprietary peptides therefore provides effective leverage toward the near doubling of the orthobiologics market.

描述（由申请人提供）：生物疗法（包括细胞、肽和蛋白质）的出现使医学能够解决许多未满足的临床需求。通常这些药物非常有效，可以特异性激活内源性信号级联，以促进明确的生化和生理反应。例如，骨形态发生蛋白 2 (BMP-2) 在大约 20 年前被认为是成骨活性的强大引发剂，此后已成为用于脊柱融合手术的领先骨科产品的一部分（美敦力的 InFuse）。尽管年销售额接近 10 亿美元，但 BMP-2 已被证明在不严格按照标签指示使用时会触发异位部位（例如气管或食道）的骨形成。在颈椎融合手术中所谓的超说明书使用 BMP-2 导致了许多患者并发症，最终 FDA 向医生发出警告，建议不要超说明书使用。两个主要问题导致了这些并发症：1) 在这些手术中使用了大剂量的 BMP-2，2) 将药物添加到胶原蛋白海绵中，而 BMP-2 对胶原蛋白海绵仅具有有限的被动亲和力。这些因素是密不可分的，因为应用大剂量以确保随着时间的推移，随着蛋白质从递送部位扩散走，保留治疗剂量。由于这些安全问题，每年进行的 237,000 例颈椎融合手术中大约有 7-10% 使用了 BMP，而腰椎融合手术中超过 60% 使用了 BMP。在宫颈手术中使用 BMP-2 将使 10 亿美元的骨科生物制剂市场几乎翻一番。利用我们的核心技术，Affinergy 开发了许多基于肽的连接基序，能够以高亲和力结合一系列生物材料。可用肽接头的靶标包括骨替代材料，例如胶原蛋白和磷酸三钙。在这里，我们建议开发一种新的转基因，用于重组生产含有肽接头的 BMP-2 蛋白。所得分子将保留 BMP-2 活性，并且与未修饰的 BMP-2 相比，对骨移植材料具有更高的亲和力。我们相信，该产品将通过确保治疗浓度的蛋白质随着时间的推移保持与递送基质结合，从而减少所需的初始剂量，从而解决目前未满足的 BMP-2 递送需求。该计划也恰逢其时，因为围绕 BMP-2 的大量知识产权即将在未来 2-3 年内到期。拟议的研究计划将需要：1）构建包含编码序列内肽接头结构域的 BMP-2 表达载体； 2) 建立表达克隆以确保蛋白质的可重复生产，3) 体外测试以确定肽标记的 BMP-2 的活性及其对骨替代材料的亲和力。如果成功，第二阶段工作将包括大规模培养优化、在临界尺寸缺陷动物模型中测试改良的 BMP-2，以及一系列临床前测试。 Affinergy 相信大型骨科公司等合作伙伴将抓住机会推销该产品。 公共健康相关性：在脊柱融合中使用成骨蛋白已经催生了价值 10 亿美元的脊柱骨科生物制品市场。虽然这些产品的使用不断增长，但由于一系列安全问题，某些适应症无法使用这些产品。在这里，我们建议将肽标签纳入广泛使用的骨制品 BMP-2 中。该肽将包含对目前用作脊柱融合中 BMP-2 载体的骨空隙填充材料的高亲和力结合位点。我们相信该产品将 (1) 限制在植入部位，并且 (2) 允许使用较小剂量的 BMP-2；这些改进减轻了 BMP-2 的安全问题，例如异位骨形成。此外，该产品将使 BMP-2 在颈椎融合术中得到更广泛、更安全的使用，在这种情况下，只有 7-10% 的病例使用骨科生物制剂，因为已知使用时不良事件的发生率会增加在颈椎融合中。因此，Affinergy 专有肽的使用为骨科生物制剂市场近乎翻倍提供了有效的杠杆。