Lentiviral expressed growth factors as a novel therapy in wound healing

描述（由申请人提供）：美国每年约有100万人被诊断患有下肢静脉溃疡，全球另有300万人被诊断患有下肢静脉溃疡。目前，有几种伤口愈合治疗的方法。重组PDGF（Regranex，Ortho-McNeil）已被美国食品药品监督管理局（FDA）批准为局部施用的凝胶，其通过促进参与伤口修复的细胞的趋化性募集和增殖而与内源性PDGF-BB在生物学上类似地起作用。伤口愈合护理界的共识是，Regranex在临床应用中表现出有限的潜力。尽管其有限的治疗成功，Regranex是非常昂贵的，因此超出了老年医疗保险患者的手段，他们代表了大多数慢性伤口人群。因此，迫切需要用于伤口愈合的新疗法。 该提案代表了修订后的项目，重点是开发一种新的基因治疗策略，用于使用表达低氧诱导因子1（HIF-11）的自失活（SIN）慢病毒载体（LV）加速伤口愈合，HIF-11是一种已知诱导具有伤口愈合特性的生长因子的转录因子。当直接应用于伤口时，用在LV中表达的HIF-11治疗可能潜在地作为比生长因子更好的伤口愈合疗法。已知LV以高效率稳定表达基因，因此假设在伤口部位稳定表达HIF-11和HIF-11诱导的生长因子将导致加速伤口愈合。 在本提案的目标1中，我们将开发表达HIF-11作为治疗有效负载的SIN慢病毒载体，以及表达绿色荧光蛋白（GFP）的对照载体。这些载体还将表达突变型人tmpk（TMPK）基因，作为优异的安全特征。一旦伤口愈合过程完成，这种自杀基因允许转导的细胞从体内消除，从而限制任何潜在的并发症，因为HIF-1和由HIF-1表达诱导的生长因子的长期表达。慢病毒载体将根据标准程序生产，并在放行检测前进行质量控制。目的2：利用小鼠胚胎成纤维细胞（MEFs）检测SIN.LV.HIF-11和SIN.LV.GFP的体外转导效率。在目的3中，我们将使用糖尿病小鼠模型测试LV延长HIF-11诱导的生长因子的表达和在伤口部位诱导伤口愈合过程的能力。在第一阶段，Lentigen公司提出的研究将最终产生原理性结果的证明，导致开发一种新的，改进的和安全的基因疗法用于治疗慢性伤口。我们的新疗法有望在全球临床环境中用于治疗伤口愈合。公共卫生相关性本提案的重点是开发一种新的基因治疗策略，以加速伤口愈合。仅在美国就有350万慢性皮肤溃疡患者（Wound Healing Society，2003）。全球数字超过1400万人。200万糖尿病患者在其一生中会患上足部溃疡。目前，唯一可用于治疗糖尿病足溃疡的药物是Regranex，其成功率非常有限。我们的新疗法将是负担得起的，预计将在全球临床环境中用于治疗伤口愈合。

DESCRIPTION (provided by applicant): Approximately 1 million people are diagnosed with venous leg ulcers annually in the U.S. and another 3 million worldwide. Currently, there are several approaches to wound healing treatment. Recombinant PDGF (Regranex, Ortho-McNeil) has been approved by the Food and Drug Administration (FDA) as a topically applied gel that acts biologically similarly to endogenous PDGF-BB by promoting chemotactic recruitment and proliferation of cells involved in wound repair. The consensus in the wound healing care community is that Regranex exhibited limited potential in clinical use. Despite its limited therapeutic success, Regranex is very expensive and is therefore beyond the means of elderly Medicare patients who represent the majority of the chronic wound population. Therefore, new therapies for wound healing are urgently needed. This proposal represents the revised project focused upon developing a novel gene therapy strategy for accelerated wound healing using Self Inactivating (SIN) lentiviral vectors (LV) that express hypoxia-inducible factor 1 (HIF-11), a transcription factor known to induce growth factors with wound healing properties. Treatment with HIF-11 expressed in LV might potentially serve as a much better therapy for wound healing than growth factors when applied directly to the wound. LVs are know to stably express genes with high efficiency, so it is hypothesized that stable expression of HIF-11 and HIF-11-induced growth factors at the wound site would lead to accelerated wound healing. In Aim 1 of this proposal we will develop SIN lentiviral vector that express HIF-11 as therapeutic payload, as well as control vector expressing green fluorescence protein (GFP). These vectors will also express the mutant human tmpk (TMPK) gene as an excellent safety feature. This suicide gene allows the transduced cells to be eliminated from the body once the wound healing process has been completed, thus limiting any potential complications because of prolonged expression of HIF-1 and growth factors induced by HIF-1 expression. Lentiviral vectors will be manufactured according to standard procedures and quality control will be performed prior to release for testing. In Aim 2 we will test the transduction efficiency of SIN.LV.HIF-11 and SIN.LV.GFP in vitro using mouse embryonic fibroblasts (MEFs). In Aim 3 we will test the ability of the LVs to prolong the expression of HIF-11-induced growth factors and to induce wound healing process at the wound site using diabetic mouse model. During Phase I Lentigen Corp. proposes studies that will ultimately generate proof of principle results, leading towards development of a novel, improved and safe gene therapy for the treatment of chronic wounds. Our new therapy is expected to be used in worldwide clinical settings for treatment of wound healing. PUBLIC HEALTH RELEVANCE This proposal focuses upon developing a novel gene therapy strategy for accelerated wound healing. There are 3.5 million patients with chronic skin ulcers in the U.S. alone (Wound Healing Society, 2003). The global figure is more than 14 million individuals. Two million people with diabetes will develop foot ulcers during their lifetimes. Currently, the only drug available for the treatment of diabetic foot ulcers is Regranex and it has a very limited success. Our new therapy will be affordable and is expected to be used in worldwide clinical settings for treatment of wound healing.