Please click Additional Files below to see the full abstract

Alves, Paula Marques

Diniz, Cláudia

eliciano, Ana M

Louro, Ana Filipa

Serra, Margarida

Vicente, Pedro

English

Engineering Conferences International

 Poster Number 197 TOWARDS LARGE-SCALE PRODUCTION OF HUMAN-INDUCED PLURIPOTENT STEM CELL-DERIVED EXTRACELLULAR VESICLES IN STIRRED-TANK BIOREACTORS  Ana Filipa Louro, iBET/ITQB-NOVA, Portugal ana.louro@ibet.pt Ana Meliciano, iBET/ITQB-NOVA, Portugal Pedro Vicente, iBET/ITQB-NOVA, Portugal Cláudia Diniz, iBET/ITQB-NOVA, Portugal Paula Marques Alves, iBET/ITQB-NOVA, Portugal Margarida Serra, iBET/ITQB-NOVA, Portugal   Key Words: Acellular therapy, hiPSC, Extracellular Vesicles, Stirred-tank bioreactors, Cardiac regeneration.  A significant bottleneck in the advancement of Extracellular Vesicle (EV)-based therapies is the efficient large-scale manufacture of clinical-grade EV. Upstream challenges include the selection of an appropriate EV parent cell and its large-scale expansion. We previously identified human induced Pluripotent Stem Cells (hiPSC) as a source of native EV for cardiac regeneration and pinpointed the bioactive signatures of hiPSC-EV. Currently, we are developing scalable bioprocesses for hiPSC expansion in Stirred-tank Bioreactors (STB) and modulating critical process parameters to intensify EV production.  Briefly, hiPSC were expanded as 3D aggregates in STB (DasGip Eppendorf AG), operated in perfusion (D=1.3 day-1). Throughout hiPSC expansion, the dissolved O2 concentration was controlled at low levels (4% O2). EV were isolated by density gradient ultracentrifugation, probed for specific markers, and characterized in terms of particle size distribution and morphology. EV bioactivity was assessed in human umbilical vein endothelial cells. A 2.9-fold increase in cell concentration was observed in STB compared to the static 2D monolayer culture, which resulted in a 3.1 increase in total particles isolated per mL of conditioned medium. hiPSC-EV produced in STB presented a cup-shape morphology and were positive for EV markers. Tube formation assays showed increased pro-angiogenic activity for hiPSC-EV produced in STB versus static 2D monolayer culture.  Overall, our study validates hiPSC as cell biofactories for large-scale EV production in STB and provides insights into manufacturing EV-based products. Ongoing work aims at optimizing EV production yields and potency by further manipulating process parameters such as dissolved O2, stirring rate and operation mode.   Funding: FCT PhD fellowships SFRH/BD/145767/2019 and UI/BD/151255/2021; CARDIOPATCH Interreg SUDOE (SOE4/P1/E1063), BRAV3 (H2020-SC1-BHC-874827); iNOVA4Health (UIDB/04462/2020; UIDP/04462/2020); LS4FUTURE (LA/P/0087/2020).   

Towards large-scale production of human-induced pluripotent stem cell-derived extracellular vesicles in stirred-tank bioreactors

https://dc.engconfintl.org/cgi/viewcontent.cgi?filename=0&article=1241&context=ccexviii&type=additional

Towards large-scale production of human-induced pluripotent stem cell-derived extracellular vesicles in stirred-tank bioreactors

Abstract

Similar works

Full text

Available Versions

Engineering Conferences International