Automated Expansion of Primary Human T Cells in Scalable and Cell‐Friendly Hydrogel Microtubes for Adoptive Immunotherapy

Abstract

Adoptive immunotherapy is a highly effective strategy for treating many human cancers, such as melanoma, cervical cancer, lymphoma, and leukemia. Here, a novel cell culture technology is reported for expanding primary human T cells for adoptive immunotherapy. T cells are suspended and cultured in microscale alginate hydrogel tubes (AlgTubes) that are suspended in the cell culture medium in a culture vessel. The hydrogel tubes protect cells from hydrodynamic stresses and confine the cell mass less than 400 µm (in radial diameter) to ensure efficient mass transport, creating a cell-friendly microenvironment for growing T cells. This system is simple, scalable, highly efficient, defined, cost-effective, and compatible with current good manufacturing practices. Under optimized culture conditions, the AlgTubes enable culturing T cells with high cell viability, low DNA damage, high growth rate (≈320-fold expansion over 14 days), high purity (≈98% CD3+), and high yield (≈3.2 × 10 cells mL hydrogel). All offer considerable advantages compared to current T cell culturing approaches. This new culture technology can significantly reduce the culture volume, time, and cost, while increasing the production.

Engineered Microenvironment for Manufacturing Human Pluripotent Stem Cell-Derived Vascular Smooth Muscle Cells

“…To address the challenge, we previously developed a scalable and high-yield cell culture method for expanding hPSCs and other human cells (Li et al., 2018a, Li et al., 2018b, Lin et al., 2018a, Lin et al., 2018b). With this method, cells are processed into microscale alginate hydrogel tubes (AlgTubes) that are suspended in the cell culture medium.…”

Engineered human brown adipocyte microtissues improved glucose and insulin homeostasis in high fat diet-induced obese and diabetic mice

“…However, the matrix is expensive. Our group recently developed a novel microbioreactor termed AlgTubes, in which cells are cultured in alginate hydrogel microtubes [36,37,[67][68][69][70][71][72]. AlgTubes are scalable and have low cost.…”

“…Fabricating 3D BA microtissues in shaking plates: Single BAPs were suspended in differentiation medium I in a low adhesion 6-well plate (Corning, #3471) shaking at 75 rpm. Detailed methods of culturing cells in shaking plates can be found in our previous publications [35][36][37][38][39]. Differentiation medium II was used after three days and refreshed once a week.…”

“…However, microwells are unsuitable for producing microtissues at large scales, which are needed for future extensive animal studies, drug screens, and clinical applications. Previously, our lab used 3D suspension culture (e.g., shaking plates, spinning flasks, stirred tank bioreactors) to prepare human pluripotent stem cells at large scales [35][36][37][38][39]. We also pioneered in culturing stem cells at high density in thermoreversible PNIPAAm-PEG hydrogels [35,[39][40][41][42][43]61].…”

Advances in engineering local drug delivery systems for cancer immunotherapy

“…Currently, the commercially available superparamagnetic Dynabeads coated with anti-CD3 antibodies (aCD3) and anti-CD28 antibodies (aCD28) are considered the standard for ex vivo T cell isolation, activation, and expansion (Neurauter et al, 2007). Biomaterials can facilitate the ex vivo expansion and activation of T cells prior to their administration (Lin et al, 2018;Schluck, Hammink, Figdor, Verdoes, & Weiden, 2019). Specifically, hydrogel-based scaffolds have been investigated for use in local cell delivery as well as in the ex vivo expansion and activation of T cells (Monette, Ceccaldi, Assaad, Lerouge, & Lapointe, 2016;Weiden et al, 2018).…”