3D Printed Engineered Living Materials for Drug Delivery

项目摘要 传统的药物递送平台受到可以装载到递送系统中的药物量的限制。 虽然对于一些聚合物体系，药物负载能力可以达到50重量%，但是在此期间， 可持续的治疗剂的递送通常是有限的。此外，局部药物的治疗效果 药物递送系统与活性剂的局部生物利用度有关。生物催化装置 因此，原位产生治疗剂可以提供活性药物的更有效的局部递送， 疗效该计划的长期愿景是创造下一代3D打印原位药物 生产和输送装置，包括药物洗脱支架、微针和贴片， 用于局部和持续治疗递送的生物材料（ELM）。ELMs是微生物的复合体 在一个实施方案中，所述方法包括将细胞掺入聚合物基质内，其中所述细胞保持其活力并且可以代谢地 制造出一种治疗化合物尽管ELMs在药物输送方面具有巨大的潜力， 应用中，ELM作为药物洗脱支架或贴片部署的主要挑战是治疗 肠道疾病的特征在于ELM必须（i）可加工成精确的形状因子（例如，患者专用支架） 具有必要的机械性能，（ii）以预定速率生物降解成非细胞毒性组分， 和（iii）在装置的使用寿命内原位生物催化产生和包埋治疗剂。客观 该提案的一个主要目的是开发3D打印树脂，提供可生物降解的水凝胶结构， 刚度，并证明制造一个原型ELM装置持续交付的模型 化合物.在目标1中，我们将用非致病性E. coli Nissle 1917（EcN）， 使用市售3D打印机将其3D打印成水凝胶构建体。我们将配制水性树脂 由可溶性球状蛋白衍生物组成，所述球状蛋白衍生物可与水溶性丙烯酸酯共聚， 光引发的聚合反应。ELM水凝胶的机械性能（刚度， 强度和韧性）和酶降解速率。在Aim中 2、我们将对非致病性EcN进行代谢工程改造，以产生作为模型治疗化合物的小檗碱。 我们将进一步评估Caco-2细胞的细胞毒性和上皮完整性， 榆树。作为这些ELM的验证，我们将使用体外模型来确认小檗碱的产生和洗脱 通过评估Caco-2对促炎刺激的反应，从3D打印的ELM中获得。我们设想这些 3D打印ELM可用作治疗恶性肿瘤或炎症的局部药物输送装置 肠道疾病虽然我们的ELM平台与广泛的微生物兼容， 包括酵母菌和细菌，我们选择专注于E. coli Nissle 1917，a 肠道微生物组中常见的细菌菌株。利用人类的原生微生物 微生物组可以促进我们的平台翻译来治疗肠道疾病。

PROJECT SUMMARY Traditional drug delivery platforms are limited by the amount of drug that can be loaded into the delivery system. While drug loading capacity can reach 50% by weight for some polymeric systems, the time period over which the delivery of the therapeutic can be sustained is often limited. In addition, the therapeutic efficacy of a local drug delivery system is related to the local bioavailability of the active agent. Devices that bio-catalytically produce the therapeutic in situ could thus provide more effective local delivery of the active drug and improve therapeutic efficacy. The long-term vision for this program is to create the next-generation 3D printed in situ drug production and delivery devices, including drug-eluting stents, microneedles, and patches, based on engineered living materials (ELMs) for localized and sustained therapeutic delivery. ELMs are composites of microorganisms incorporated within a polymeric matrix, wherein the cells maintain their viability and can be metabolically engineered to produce a therapeutic compound. Despite the immense potential of ELMs for drug delivery applications, the primary challenges to the deployment of ELMs as drug-eluting stents or patches to treat intestinal diseases are that ELMs must (i) be processable into precise form factors (e.g., patient-specific stents) with the requisite mechanical properties, (ii) biodegrade into non-cytotoxic components at predetermined rates, and (iii) bio-catalytically produce and elute the therapeutic agent in situ for the lifetime of the device. The objective of this proposal is to develop 3D printable resins that afford biodegradable hydrogel constructs with a tunable stiffness, and to demonstrate the fabrication of a prototype ELM device for sustained delivery of a model compound. In Aim 1, we will create aqueous resins with non-pathogenic E. coli Nissle 1917 (EcN) that can be 3D printed into hydrogel constructs using a commercially available 3D printer. We will formulate aqueous resins comprised of soluble globular protein derivatives that can be co-polymerized with water-soluble acrylate monomers upon photo-initiated polymerization. The mechanical properties of the ELM hydrogels (stiffness, strength, and toughness) and rates of enzymatic degradation will be quantified for each resin formulation. In Aim 2, we will metabolically engineer non-pathogenic EcN to produce berberine as a model therapeutic compound. We will further evaluate the cytotoxicity and epithelial integrity of Caco-2 cells in the presence of 3D printed ELMs. As validation of these ELMs we will use an in vitro model to confirm the production and elution of berberine from the 3D printed ELM by evaluating the Caco-2 response to proinflammatory stimulation. We envision these 3D printed ELMs to be used as devices for local drug delivery in the treatment of malignancies or inflammatory diseases affecting the intestines. While our ELM platform is compatible with a broad array of microorganisms that include yeast and bacteria, we have chosen to focus on engineered variants of E. coli Nissle 1917, a commensal strain of bacteria common within the gut microbiome. Using a microorganism native to the human microbiome may facilitate translation of our platform to treat intestinal diseases.