Discovery of a Platelet Derived Growth Factor Peptide-based Mimetic

项目概述/摘要：糖尿病足溃疡是一个严重的健康问题，给患者和社会带来了高昂的成本（Boulton, Vileikyte et al. 2005）。很大一部分足部溃疡对现有的常规治疗仍然没有反应，其相关的并发症和费用促使了广泛的研究，并带来了有希望的突破。重组生长因子、可生物降解基质和生物工程皮肤等效物已被开发出来，以更好地促进愈合（Edmonds, Bates et al. 2000）。这些疗法已经显示出希望，但仍有局限性，包括高成本、免疫原性和需要高剂量、重复的生长因子。血小板衍生生长因子（PDGF）是一种有效的趋化、有丝分裂原和分化因子，可促进软组织形成，rhPDGF-BB目前已被FDA批准用于治疗糖尿病溃疡。重复应用高剂量PDGF增加了治疗的成本和复杂性。一般来说，生物疗法的开发和制造比合成疗法更昂贵。因此，一种具有更长的停留时间的合成替代品可以显著降低当前治疗的成本和复杂性。本提案的目标是产生一种PDGF-BB模拟物，可以固定在胶原蛋白支架上，用于促进软组织和伤口愈合。我们计划使用噬菌体展示技术来鉴定模拟PDGF- b受体的PDGF样信号传导的肽。这些肽将连接到先前确定的高亲和力胶原结合肽。由此产生的混合分子有望允许从胶原基质表面的合成分子的控制，局部递送。我们预计胶原结合肽将显著改善生物活性PDGF模拟物在伤口中的保留。首先，使用噬菌体展示技术，我们将识别与PDGF b受体结合的肽。其次，为了提高肽对b受体的亲和力，将基于初始b受体结合肽的序列构建重点文库，并在b受体上再次筛选。接下来，含有2个b受体结合序列的合成肽将用柔性连接物化学合成，所得二聚体将通过成纤维细胞增殖试验测试pdgf - bb样活性。最后，我们将合成Affinergy现有的胶原蛋白结合序列和新鉴定的PDGF-BB模拟肽的组合，并测试杂交分子与胶原蛋白的结合以及在与胶原基质结合时诱导成纤维细胞增殖的能力。项目简介：糖尿病足溃疡是一个严重的健康问题，给患者和社会带来了高昂的成本。很大一部分足部溃疡对传统治疗没有反应，这促使研究取得了有希望的突破，包括重组生长因子、可生物降解基质和生物工程皮肤等效物。这些疗法已显示出前景，但仍有局限性，包括高成本、免疫原性和需要高剂量的生长因子。在这个项目中，我们将开发一种基于血小板衍生生长因子（PDGF）肽的模拟物，它可以固定在胶原蛋白支架上，用于促进伤口愈合。我们计划使用噬菌体展示技术来鉴定模拟PDGF生物活性的肽。一般来说，生物疗法的开发和制造比合成疗法更昂贵。因此，一种具有更长的停留时间的合成替代品可以显著降低当前治疗的成本和复杂性。

DESCRIPTION (provided by applicant): Project Summary/Abstract: Diabetic foot ulcers are a significant health problem that imposes high costs on both patients and society (Boulton, Vileikyte et al. 2005). A large proportion of foot ulcers remain unresponsive to available conventional treatment and their associated complications and costs have prompted extensive research that has led to promising breakthroughs. Recombinant growth factors, biodegradable matrices, and bioengineered skin equivalents have been developed to better promote healing (Edmonds, Bates et al. 2000). These therapies have shown promise, but still have limitations including high cost, immunogenicity, and the need for high, repeated doses of growth factor. Platelet Derived Growth Factor (PDGF) is a potent chemotactic, mitogen, and differentiation factor that promotes soft tissue formation and rhPDGF-BB is currently FDA approved to treat diabetic ulcers. Repeated application of high dose PDGF increases the cost and complexity of treatment. In general, biologic therapeutics are more expensive to develop and manufacture than synthetic therapeutics. Therefore, a synthetic alternative that also has a longer resident time could significantly reduce the cost and complexity of current therapies. The goal of this proposal is to generate a PDGF-BB mimetic that can be immobilized on a collagen scaffold used to promote soft tissue and wound healing. We plan to use phage display technology to identify peptides that mimic PDGF-like signaling of the PDGF B-receptor. These peptides will be linked to a previously identified high-affinity, collagen-binding peptide. The resulting hybrid molecule is expected to allow controlled, local delivery of a synthetic molecule from the surface of a collagen matrix. We expect that collagen binding peptides will significantly improve the retention of a bioactive PDGF mimetic in a wound. First, using phage display techniques, we will identify peptides that bind to the PDGF B-receptor. Secondly, to increase the affinity of the peptides for the B-receptors, focused libraries will be constructed based on the sequences of the initial B-receptor-binding peptides and screened again on the B-receptors. Next, synthetic peptides containing 2 copies of the B-receptor binding sequence will be chemically synthesized with a flexible linker and the resulting dimers will be tested for PDGF-BB-like activity with a fibroblast proliferation assay. Finally, we will synthesize combinations of Affinergy's existing collagen binding sequences and the newly identified PDGF-BB mimetic peptide and the hybrid molecule will be tested for binding to collagen and for its ability to induce fibroblast proliferation while bound to a collagen matrix. Project Narrative: Diabetic foot ulcers are a significant health problem that imposes high costs on both patients and society. A large proportion of foot ulcers remain unresponsive to conventional treatment and have prompted research that has led to promising breakthroughs, including recombinant growth factors, biodegradable matrices, and bioengineered skin equivalents. These therapies have shown promise, but still have limitations including high cost, immunogenicity, and the need for high doses of growth factor. In this project, we will develop a Platelet Derived Growth Factor (PDGF) peptide-based mimetic that can be immobilized on a collagen scaffold used to promote wound healing. We plan to use phage display technology to identify peptides that mimic PDGF's biologic activity. In general, biologic therapeutics are more expensive to develop and manufacture than synthetic therapeutics. Therefore, a synthetic alternative that also has a longer resident time could significantly reduce the cost and complexity of current therapies.