Automated Production of RALA/Nucleic Acid Nanoparticles from Bench to Patient Dose

RNAi疗法有可能改变医疗保健干预措施，在过去两年中，有两种产品被批准用于治疗危及生命的疾病。RNAi疗法旨在短暂地减少有缺陷的基因以达到治疗目的。这是一个快速增长的市场，有109种基于RNAi的治疗方法正在临床试验中（2018年7月）\[Wu X. and Turnball A.P. 2018\]。然而，围绕RNAi疗法仍然存在一些问题，包括到达适当的组织，然后确保细胞内递送到目的地。最近的研究还表明，那些有潜在健康状况的人，如糖尿病、高血压或吸烟者，肺上皮中ACE-2受体的数量增加\[Leung J.M, 2020\]。研究表明，COVID-19利用ACE-2受体进入细胞，以产生更多的病毒颗粒，从而感染更多的ACE-2受体阳性细胞\[Kuba K. 2005\]。ACE-2受体在许多生物学功能中发挥着作用，但如果能在短时间内降低表达，就能降低病毒的传染性，并有助于扭转平衡，使其朝着健康恢复的方向发展。RNAi可以用来短暂地降低这种ACE-2受体的表达，但前提是有合适的递送系统。pHion （Belfast SME）已经开发出一种安全的RNAi递送解决方案，不会进一步加剧免疫系统，优先将治疗药物递送到肺部，并且具有成本效益，最终使RNAi治疗得到广泛采用。这项创新围绕着一种被称为RALA的肽的使用，这种肽被设计成将RNAi浓缩成纳米颗粒（NPs），这些纳米颗粒具有穿过细胞膜、逃离核内体、将货物高效率地运送到细胞质的必要特性。在旨在减少ACE-2表达的RNAi和RALA肽NPs之间形成的NPs不需要冷链储存，可以保存数月而不失去功能。然而，我们还没有一种适当的方法来支持大规模生产这些NPs。事实上，对于核酸行业来说，最大的障碍之一将是新疗法的制造。因此，与COVID-19带来的具体挑战主题明确一致，该项目旨在加速和优化RALA/RNAi治疗对患者剂量的扩大，以便“为未来做好准备”。拟议的9个月项目旨在开发使用微流体自动化生产功能性NPs的最佳条件，可以很容易地转移到临床剂量。我们还将开发最佳的冻干工艺，以确保高度稳定的功能性产品。最后，有了适当的监管框架和证据，我们可以将我们的过程转移到外部以扩大临床剂量，我们将很好地将这种疗法应用于临床，并将RALA定位为RNAi治疗到肺部的首选递送系统。

RNAi therapeutics have the potential to transform healthcare interventions as evidenced by the approval of 2 products in the last 2 years for life threatening diseases. RNAi therapy is designed to transiently reduce a defective gene for therapeutic purposes. It is a rapidly growing market with 109 RNAi based therapeutics in clinical trials (July 2018) \[Wu X. and Turnball A.P. 2018\]. However, there are still issues that surround the RNAi therapeutics which include getting to the appropriate tissue and then ensuring intracellular delivery to the destination site. Recent studies have also indicated that those with underlying health conditions such as diabetes, high-blood pressure or smokers have an increased number of ACE-2 receptors in the lung epithelium \[Leung J.M, 2020\]. Studies have revealed that COVID-19 uses the ACE-2 receptor to enter cells in order to produce more viral particles that can infect more ACE-2 receptor positive cells \[Kuba K. 2005\]. The ACE-2 receptor plays a role for many biological functions but if expression could be lowered for a short period of time it could reduce the infectivity of the virus and help tip the balance towards healthy recovery. RNAi could be used to transiently reduce expression of this ACE-2 receptor but only if there is an appropriate delivery system. pHion (Belfast SME) have developed a solution for RNAi delivery that is safe, does not further exacerbate the immune system, preferentially delivers the therapeutic to the lung and is cost-effective, ultimately enabling widespread adoption of the RNAi therapy. The innovation centres around the use of a peptide termed RALA that is designed to condense RNAi into nanoparticles (NPs) that have the properties necessary to cross cell membranes, escape endosomes delivering the cargo to the cytoplasm with high efficiency. The NPs formed between the RNAi which is designed to reduce ACE-2 expression and the RALA peptide NPs do not require cold chain storage and can be stored for many months without losing functionality. However, we do not as yet have a methodology in place to support the large-scale production of these NPs. Indeed, for the nucleic acid industry, one of the greatest hurdles will be the manufacture of novel therapeutics. Therefore with clear alignment to the specific theme of challenges as a result of COVID-19, this project is designed to accelerate and optimise the scale-up of the RALA/RNAi therapeutic to patient doses in order to be 'future ready'. The proposed 9 month project is designed to develop the optimal conditions for the automated production of functional NPs using microfluidics that can be readily transferred to clinical doses. We will also develop the optimal lyophilisation process to ensure a highly stable functional product. Finally, with regulatory framework in place and proof that we can transfer our process externally to scale up to clinical doses, we will be well positioned to take this therapy to the clinic and to position RALA as the go-to delivery system for RNAi therapeutics to the lung.