Understanding TFF operations in cell and gene therapy bioprocessing

背景：切向流过滤(TFF)通常用于生物过程序列中的几个步骤，以产生细胞和基因疗法(CGT)。它们通常用于超滤(浓缩)、透析过滤或缓冲液交换，有时用于澄清和/或微粒去除。这一单元操作正在成为回收CGT产品，包括它们的材料(例如，用于成分DNA或RNA提纯，或脂类纳米颗粒生产)的宝贵主力。目前生产中常用的格式是中空纤维，它们有不同的长度、内径、纤维厚度和膜类型。用于TFF的另一种格式是平板格式，但在CGT应用程序中并不常见。目前缺乏关于某些CGT应用的更好格式的信息，也没有系统的研究来比较这两种格式以及影响其在应用于CGT时性能的不同设计参数。这两种格式的放大也略有不同。例如，通过保持流道长度，同时增加膜的宽度以增加表面积，使平板膜的放大变得牢固。湍流促进剂的使用很常见，但这些促进剂对细胞或病毒载体的影响尚不清楚。中空纤维膜的放大是不同的，如果在工艺开发的不同阶段使用不同长度的膜，可能会被证明具有轻微的挑战性。鉴于对CGT产品的需求不断增加，也需要更多地了解所涉及的装置操作，以实现稳健的工艺设计。这项研究旨在研究用于细胞和基因治疗应用的TFF操作。项目描述：该项目将建立在我们对平板膜上单抗的TFF规模表征工作的基础上，并将其扩展到使用中空纤维膜的TFF。Rayat集团目前正在进行一些慢病毒载体生产的工作，重点是在离子交换层析之前的TFF。然而，TFF被应用于过程的不同点，并且用于不同的形式(例如，病毒载体、RNA)或过程组件(例如，DNA、LNP)，因此需要在流程图的不同位置研究TFF。有机会获得各种模式的各种数据，同时，使用计算模型来查看TFF设计的方面，这可能会影响细胞和基因治疗应用的性能。该项目可以产生的过程知识和新方法将造福于生物处理社区。目的：本研究的主要目的是描述和优化细胞和基因治疗生物处理中的TFF操作。具体地说，该项目的目标是：从生产率、产品稳定性和回收率以及杂质去除的角度评估不同形式的TFF的性能。为TFF的生物过程开发应用新的建模方法确定可能影响每种形式的TFF性能的关键参数(例如，中空纤维长度、膜类型、孔径大小、平板湍流促进器等)确定使不同CGT产品和组件(例如，RNA、DNA、LNPs、病毒载体)能够最佳回收的关键TFF条件

Background:Tangential flow filtration (TFF) is often used in several steps within a bioprocess sequence to produce cell and gene therapies (CGT). These are typically for ultrafiltration (concentration), diafiltration or buffer-exchange, and sometimes for clarification and/or particulates removal. This unit operation is becoming an invaluable workhorse for the recovery of CGT products, including their materials (e.g., for component DNA or RNA purification, or lipid nanoparticle production). The current format commonly used in production are hollow fibres which come in different lengths, internal diameters, fibre thickness and membrane type. Another format used for TFF, but not often seen in CGT application, is the flat-sheet format. There is currently a lack of information as to the better format for certain CGT applications, nor are there systematic studies that compare the two formats and the different design parameters that affect their performance when applied to CGT. Scale-up of these two formats are also slightly different. For example, scale-up of flat sheet membranes have been made robust by maintaining the flow-path length while increasing the membrane width for increased surface area. The use of turbulence promoters is common but the effect of these on cells or viral vectors are unknown. Scale-up of hollow fibre membranes is different and may prove slightly challenging if different lengths are used in the different stages of process development. Given the increasing demand for CGT products, the need also grows for increased understanding of the unit operations involved to achieve a robust process design. This study aims to investigate the TFF operation for cell and gene therapy applications.Project Description:The project will build on our work on TFF scale-down characterisation for Mabs on flat-sheet membranes and extend this to TFF using hollow-fibre membranes. There is some work currently underway in the Rayat Group on lentiviral vector production focusing on TFF prior to ion exchange chromatography. However, TFF is applied at different points of the process and for different modalities (e.g., viral vectors, RNA) or process components (e.g., DNA, LNP) and so there is a need to investigate TFF at different places of the flowsheet. There is an opportunity to obtain a wide variety of data for various modalities, and at the same time, to use computation modelling to look at the aspect of TFF design which can affect performance in cell and gene therapy applications. The process knowledge and novel methods this project can generate will benefit the bioprocessing community.Objectives: The main aim of this study is to characterise and optimise TFF operations in cell and gene therapy bioprocessing. Specifically, the project aims to:Evaluate the performance of the different formats of TFF in terms of productivity, product stability and recovery and impurity removal.Apply novel modelling methodologies for the bioprocess development of TFFDetermine the key parameters which may affect the TFF performance for each format (e.g., hollow fibre length, membrane type, pore size, flat-sheet turbulence promoters etc)Identify the key TFF conditions that enable the optimal recovery of different CGT products and components (e.g., RNA, DNA, LNPs, viral vectors)