Microfluidics production of self-amplifying RNA-based therapeutics vaccine

摘要：THERAPEUTIC-SAM@SCALE将支持英国制造战略，以开发用于COVID-19自扩增RNA疫苗的技术为基础。我们将确定用于癌症的自扩增RNA治疗性疫苗必须考虑的关键材料和制造变量。这将转移至GMP生产规模并进行体内验证。通过这一过程，我们将支持国家和国际供应新型热稳定疫苗系统。背景资料：为应对COVID-19，Perrie一直与英国政府疫苗工作组合作，支持COVID-19自扩增RNA疫苗的生产。基于RNA的疫苗联合收割机结合了减毒活疫苗和亚单位疫苗的积极属性。然而，这些RNA疫苗只有在纳米颗粒递送系统中配制时才有效，并且脂质纳米颗粒（LNP）已被用于当前的COVID-19疫苗。脂质纳米颗粒保护RNA免于降解，促进内体逃逸和细胞特异性靶向，并且可以与佐剂共同递送。此外，它们具有以不依赖规模的方式制造的潜力。鉴于这些疫苗的积极反应，该项目现在将把这项技术应用于治疗性疫苗。治疗性疫苗是在感染发生后使用的疫苗，旨在诱导免疫力以改变疾病的进程。然而，在开发治疗性癌症疫苗时，情况更为复杂。与通常被认为是免疫系统外来的细菌和病毒不同，癌细胞更接近于我们正常的健康细胞。此外，肿瘤通常对个体是独特的，并且具有其自身的区别性抗原。因此，为了开发有效的治疗性癌症疫苗，需要疫苗的个性化。使用自扩增RNA疫苗可以通过对肿瘤细胞内的DNA进行快速测序来识别独特的癌症肽来解决这一问题。这些肽又可以在自扩增RNA内编码并掺入RNA疫苗中。挑战：目前生产脂质纳米粒的方法是乙醇注射法;这种批量生产方法涉及多单元操作（多达18个），整个生产在无菌区进行。复杂性是巨大的，因此扩大规模和技术转让是一个重大挑战。我们现在将开发这项技术来开发用于治疗癌症的治疗性疫苗，我们将研究配方，以解决目前与SAM-LNP疫苗相关的热不稳定性。应对挑战：该项目旨在开发热稳定的SAM-LNP，可以使用灵活和自适应的工艺制造，可以在快速响应模式下按比例放大和缩小。为了实现这一目标，目标是：1）研究脂质部分在控制SAM-LNP疫苗的疫苗效力中的作用，2）通过利用冷冻干燥方案形成热稳定的疫苗系统，以及3）开发创新的微流体和在线监测工艺，以创建SAM-LNP疫苗的连续制造选项。

Summary: THERAPEUTIC-SAM@SCALE will support the UK Manufacturing strategy to build on the technology developed for COVID-19 self-amplifying RNA vaccines. We will identify the key material and manufacturing variables that must be considered for self-amplifying RNA therapeutic vaccines for cancer. This will be transferred to GMP manufacturing scale and validated in vivo. Through this process we will support the national and international supply of novel thermostable vaccine systems. Background: In response to COVID-19, Perrie has been working with the UK Government Vaccine Task Force to support the manufacture of a COVID-19 self-amplifying RNA vaccine. RNA-based vaccines combine the positive attributes of both live-attenuated and subunit vaccines. However, these RNA vaccines are only effective when formulated within a nanoparticle delivery system, and lipid nanoparticles (LNPs) has been adopted for the current COVID-19 vaccines. Lipid nanoparticles protect the RNA against degradation, facilitate endosomal escape and cell specific targeting, and can be co-delivered with adjuvants. Furthermore, they have the potential to be manufactured in a scale-independent manner. Given the positive responses being demonstrated with these vaccines, this project will now apply this technology to therapeutic vaccines. A therapeutic vaccine is one in which the vaccine is used after infection occurs, aiming to induce immunity to alter the course of disease. However, the situation is more complicated when developing therapeutic cancer vaccines. Unlike bacteria and viruses that are generally recognised as foreign to our immune system, cancer cells more closely resemble our normal, healthy cells. Furthermore, tumours are generally unique to the individual and have their own distinguishing antigens. As a result, to develop effective therapeutic cancer vaccines, personalisation of the vaccine is needed. The use of self-amplifying RNA vaccines can address this through the rapid sequencing of the DNA within tumour cells to identify unique cancer peptides. These peptides can in turn be coded within self-amplifying RNA and incorporated into RNA vaccines. Challenge: The current method for manufacturing lipid nanoparticles is an ethanol injection method; this batch production method involves multi-unit operations (up to 18) with the entire production being carried out in an aseptic area. The complexity is immense and hence scale up and technology transfer is a major challenge. We will now develop this technology to develop therapeutic vaccines for the treatment of cancer and we will investigate formulations to address thermo-instabilities currently associated with SAM-LNP vaccines.Addressing the Challenge: This project aims to develop thermostable SAM-LNPs which can be manufactured using flexible and adaptive processes, which can be scaled up and down in a rapidly responsive mode. To achieve this, the objectives are: 1) investigate the role of lipid section in controlling vaccine efficacy of SAM-LNP vaccines, 2) format a thermostable vaccine system by exploiting the freeze-drying protocols and 3) develop innovative microfluidic and inline monitoring processes to create continuous manufacturing options for SAM-LNP vaccines.