(29d) Combining Tunable Biomaterials and Flow-Based Membrane Technologies for Improved Biomanufacturing of T Cell Therapies

Introduction: The manufacturing steps used for adoptive cell therapies, such as chimeric antigen receptor T-cell (CAR-T) therapy, include isolation, activation, genetic modification, and expansion of a patient’s T-cells. These processing steps remain mainly based on lab-scale procedures, which are time and labor intensive and afford limited opportunities for scale up increasing the cost of treatment.^3,4 There is an unmet need to develop modular cell therapy production platforms that can recreate elements of the in vivo microenvironment to effectively engineer patients’ cells with increased efficacy and reduced cost.

Approach and experimental methods: In this work, we present a modular manufacturing platform that combines soft hydrogels with existing commercial flow-based membrane devices for improved activation, expansion, and transduction of primary human T-cells. We created thin poly(ethylene glycol) diacrylate (PEGDA) hydrogel films on microfiltration membranes to prepare hydrogel coated membranes (HCM) with bioinspired mechanical properties and cell-activating anti-CD3 and anti-CD28 through biotin-avidin chemistry to regulate T-cell activation and proliferation. T cell activation and proliferation in response to HCM in comparison to state-of-the-art controls were extensively characterized with flow cytometry by analyzing the changes in marker expression of following T-cell markers (CD69, CD25, CCR7, CD45RA, CD45RO, CXCR3, CCR7, and PD-1). We further introduced HCMs into a tangential flow filtration (TFF) device to efficiently transduce the T cells with a model lentiviral system (Lentibrite GFP) under flow. Lastly, we investigated the applicability of the device for selection of specific cell populations (e.g., CD4) by functionalizing HCM with anti-CD8, which allowed immobilization of CD8 T-cells on the membrane and yielded an enriched CD4 T-cell population in the effluent flowed out the device.

Results and conclusions: Culture on antibody-functionalized HCM yielded highly activated and proliferative primary human T-cells that presented a memory phenotype ideal for T-cell therapy applications, especially in comparison to rigid controls. In parallel, T-cell transduction with the HCM under flow in the TFF flow device resulted in a multi-fold increase in transduction efficiency compared to static controls. Finally, the HCM successfully enriched the CD8 target cell population in the TFF device following selection experiments. Together, these results demonstrate that HCM and TFF technologies combine as a modular platform to improve T-cell activation, increase proliferation, enhance transduction, and enrich desired cell populations compared to current industrial biomanufacturing processes. Integrating tunable biomaterials with flow technology presents the potential for improved biomanufacturing of T-cell therapies with relevance to current industrial processes.

References:

1. Guedan, S. Annual review of immunology, 2019; 37:145-171

2. Mohanty, R. Oncology reports, 2019; 42(6):2183-2195

3. Iyer, R. Frontiers in medicine, 2018; 5:150

4. Dai, X. Biotechnology Journal, 2019; 14(4):1800239