Automating mosquito microdissection for a malaria PfSPZ vaccine

摘要 尽管每年投资超过30亿美元用于强化控制措施，但2018年，2.28亿例 疟疾病例比2015年增加了约1600万例，死亡人数没有减少。的影响 现有的疟疾控制措施已趋于稳定。此外，世卫组织估计， 由于当前全球气候变化， 2019冠状病毒病疟疾。需要新的工具，特别是疫苗。只有广泛部署一个 有效的疫苗有希望真正消除或根除，特别是在面对突发性 像COVID-19这样的发展。超过98%的疟疾死亡是由恶性疟原虫引起的 （Pf）.因此，预防Pf疟疾的疫苗是优先事项。Sanaria将于2021年进入其第三阶段临床试验 Pf子孢子（SPZ），PfSPZ疫苗，并计划获得FDA的上市许可（许可证）， EMA在2022/2023年。在接下来的5 - 10年里，我们的目标是降低商品成本（COG）和效率， PfSPZ疫苗的生产，使他们能够最有效和经济地使用的个人谁遭受 最多的是疟疾。蚊子的显微切割是PfSPZ疫苗产品提取的关键步骤， 并确保10，000倍纯化远离无关的蚊子部分作为起始材料， 下游纯化程序，然后达到99.9%的最终产品纯度。到目前为止，蚊子 唾液腺PfSPZ已证明体内感染性/效力上级优于从全唾液腺中提取的那些 蚊子，或生长在外面的蚊子。然而，蚊子唾液腺的提取是一个限速， 这是生产PfSPZ疫苗中劳动密集型、昂贵步骤。本提案的总体目标是 为了能够在基于PfSPZ的cGMP生产中实现临时半自动化解剖装置， 开发一个包含多个自动化步骤的综合解剖系统 蚊子定向的下游，用于商业规模的制造。机器人的独特应用 技术，最先进的计算机视觉和机器学习算法，以及软件系统， 在洁净室中对非常小的昆虫进行生产规模的处理不仅提高了制造能力， 也代表了自动化领域的一系列里程碑式创新。在这个项目的成功涉及一个 由调查人员、制造和质量专家组成的高技能多学科团队将决定性地导致 在分离蚊子唾液腺的关键步骤中进一步简化和优化过程， 生产高效PfSPZ疫苗。最初定义疫苗的突破 这是远远优于竞争技术在安全性和保护效力，将继续下去，因为我们 在拟议的研究中取得进展，使疫苗提取因效率提高而更具成本效益， 减少人为错误和操作员疲劳，缩短时间，大大缩短培训周期，以及 提高产品纯度，在抗击疟疾的斗争中部署一种高效工具。

ABSTRACT Despite annual investments of >$3 billion for intensive control measures, in 2018, the 228 million cases of malaria were an increase of ~16 million cases over 2015, and no decrease in number of deaths. The impact of available malaria control measure has plateaued. Moreover, WHO estimates deaths from malaria could double across sub-Saharan Africa this year due to disruptions in access to control measures due to the current global COVID-19 pandemic malaria. New tools, especially a vaccine, are needed. Only broad deployment of an effective vaccine holds the promise of true elimination or eradication, especially in the face of sudden developments like COVID-19. More than 98% of all deaths from malaria are caused by Plasmodium falciparum (Pf). Thus, a vaccine against Pf malaria is the priority. Sanaria is moving in 2021 to Phase 3 clinical trials of its Pf sporozoite (SPZ), PfSPZ Vaccine, and is planning for marketing authorization (licensure) from FDA and EMA in 2022/2023. Over the next 5-10 years we aim to decrease the cost of goods (COGs) and efficiency of production of PfSPZ vaccines so they can be used most effectively and economically by individuals who suffer the most from malaria. Microdissection of mosquitoes is a crucial step in extraction of PfSPZ vaccine products, and ensures a 10,000-fold purification away from irrelevant mosquito parts as the starting material for downstream purification procedures that then achieve a final product purity of 99.9%. To-date, mosquito salivary gland PfSPZ have demonstrated in vivo infectivity/potency superior to those extracted from whole mosquitoes, or grown outside a mosquito. However, extraction of mosquito salivary glands is a rate-limiting, labor-intensive, expensive step in production of PfSPZ-based vaccines. The overarching aim of this proposal is to enable implementation of an interim semi-automated dissection device in cGMP production of PfSPZ-based vaccines against malaria, and develop an integrated dissection system incorporating multiple automation steps downstream of mosquito orientation, for commercial-scale manufacturing. The unique application of robotic technology, state-of-the art computer vision and machine learning algorithms, and software systems to production-scale processing of very small insects in cleanrooms not only advances manufacturing capabilities, but also represents a spectrum of milestone innovations in automation. Success in this project involving a highly-skilled multi-disciplinary team of investigators, manufacturing and quality experts will decidedly lead to further streamlining and process optimization during the key step of isolating mosquito salivary glands for manufacture of our highly effective PfSPZ-based vaccines. The breakthroughs that initially defined a vaccine that is far superior to competing technologies in both safety and protective efficacy, will continue, as we advance in the proposed studies to make vaccine extraction more cost-effective due to greater efficiencies, mitigation of human error and operator fatigue, reduced timeframes, greatly reduced training periods, and increased product purity, towards deployment of a highly-impactful tool in the fight against malaria.