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PEOPLE@HES-SO - Verzeichnis der Mitarbeitenden und Kompetenzen
PEOPLE@HES-SO - Verzeichnis der Mitarbeitenden und Kompetenzen

PEOPLE@HES-SO
Verzeichnis der Mitarbeitenden und Kompetenzen

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Jossen Valentin

Jossen Valentin

Professeur-e HES Associé-e

Hauptkompetenzen

Mammalian Cell Culture

Cell Culture Development

Bioprocess Engineering

Reusable and Single-Use Bioreactors

Data analysis and modeling

Stem cells

  • Kontakt

  • Lehre

  • Publikationen

  • Konferenzen

Hauptvertrag

Professeur-e HES Associé-e

Büro: ENP.19.515

HES-SO Valais-Wallis - Haute Ecole d'Ingénierie
Rue de l'Industrie 23, 1950 Sion, CH
HEI - VS
Bereich
Chimie et sciences de la vie
Hauptstudiengang
Ingénierie des Sciences du vivant
BSc HES-SO en Ingénierie des sciences du vivant - HES-SO Valais-Wallis - Haute Ecole d'Ingénierie
  • Cell Biology
  • Bioscience and Biochemistry: Part Cell Metabolism
  • Biopharmaceutical Development and Manufacturing with Mammalian Cells
  • Lab Biopharmaceutical Development and Manufacturing with Mammalian Cells

2024

Single-Use Systems in Biopharmaceutical Manufacture: State of the Art and Recent Trends
Buchkapitel

Jossen Valentin, Regine Eibl, Gilles Broccard, Dieter Eibl

Dans Pörtner Ralf,  Biopharmaceutical Manufacturing. 2024,  Cham : Springer

Link zur Publikation

Zusammenfassung:

Single-use systems have become increasingly important in recent years. This applies in particular to their use in the manufacture of biopharmaceutical products based on mammalian cell cultures, such as antibodies for the therapy of cancer and autoimmune diseases. Today, developers and manufacturers of such biotherapeutics can choose from a large product portfolio of single-use systems from different suppliers. For example, there are single-use versions of storage bags, filters, mixers, bioreactors, connectors, harvesting and transfer systems, membrane adsorbers, freeze- and -thaw systems, to name just a few examples. Thus, the production of clinical samples in complete single-use production facilities (upstream processing to filling) is already a reality. Based on the definition of the term and the presentation of milestones in the development of single-use technology, the book chapter discusses the most important advantages and existing limitations of the single-use systems currently available on the market. Subsequently, the current state of development of single-use technology is presented on the basis of selected product examples. The focus is on the upstream area and single-use bioreactors. Finally, approaches for process intensification up to complete continuous production resulting from the increasing implementation of single-use systems are deduced.

2021

Development of a Biodegradable Microcarrier for the Cultivation of Human Adipose Stem Cells (hASCs) with a Defined Xeno- and Serum-Free Medium
Wissenschaftlicher Artikel

Francesco Muoio, Stefano Panella, Matias Lindner, Jossen Valentin, Yves Harder, Tiziano Moccetti, Regine Eibl, Michele Mueller, Tiziano Tallone

Applied Sciences, 2021 , vol.  11, no  925, pp.  925-952

Link zur Publikation

Zusammenfassung:

Stirred single-use bioreactors in combination with microcarriers (MCs) have established themselves as a technology that has the potential to meet the demands of current and future cell therapeutic markets. However, most of the published processes have been performed using fetal bovine serum (FBS) containing cell culture medium and non-biocompatible MCs. This approach has two significant drawbacks: firstly, the inevitable potential risks associated with the use of FBS for clinical applications; secondly, non-biocompatible MCs have to be removed from the cell suspension before implantation, requiring a step that causes loss of viable cells and adds further costs and complications. This study aimed to develop a new platform based on a chemically defined xeno- and serum-free cell culture medium and biodegradable MC that can support the growth of human adipose stem cells (hASCs) while still preserving their undifferentiated status. A specific combination of components and manufacturing parameters resulted in a MC prototype, called “BR44”, which delivered the desired functionality. MC BR44 allows the hASCs to stick to its surface and grow in a chemically defined xeno- and serum-free medium (UrSuppe). Although the cells’ expansion rate was not as high as with a commercial non-biodegradable standard MC, those cultured on BR44 maintained a better undifferentiated status in both static and dynamic conditions than those cultured on traditional 2D surfaces.

Chemically Defined Xeno- and Serum-Free Cell Culture Medium to Grow Human Adipose Stem Cells
Wissenschaftlicher Artikel

Stefano Panella, Francesco Muoio, Jossen Valentin, Yves Harder, Regine Eibl, Tiziano Tallone

Cells, 2021 , vol.  10, no  25, pp.  466-502

Link zur Publikation

Zusammenfassung:

Adipose tissue is an abundant source of stem cells. However, liposuction cannot yield cell quantities sufficient for direct applications in regenerative medicine. Therefore, the development of GMP-compliant ex vivo expansion protocols is required to ensure the production of a “cell drug” that is safe, reproducible, and cost-effective. Thus, we developed our own basal defined xeno- and serum-free cell culture medium (UrSuppe), specifically formulated to grow human adipose stem cells (hASCs). With this medium, we can directly culture the stromal vascular fraction (SVF) cells in defined cell culture conditions to obtain hASCs. Cells proliferate while remaining undifferentiated, as shown by Flow Cytometry (FACS), Quantitative Reverse Transcription PCR (RT-qPCR) assays, and their secretion products. Using the UrSuppe cell culture medium, maximum cell densities between 0.51 and 0.80 × 105 cells/cm2 (=2.55–4.00 × 105 cells/mL) were obtained. As the expansion of hASCs represents only the first step in a cell therapeutic protocol or further basic research studies, we formulated two chemically defined media to differentiate the expanded hASCs in white or beige/brown adipocytes. These new media could help translate research projects into the clinical application of hASCs and study ex vivo the biology in healthy and dysfunctional states of adipocytes and their precursors. Following the cell culture system developers’ practice and obvious reasons related to the formulas’ patentability, the defined media’s composition will not be disclosed in this study.

Cellular Agriculture: Opportunities and Challenges
Wissenschaftlicher Artikel

Jossen Valentin, Regine Eibl, Yannick Senn, Géraldine Gubser, Christian van den Bos, Dieter Eibl

Annual Review of Food Science and Technology, 2021 , vol.  12, no  1, pp.  51-73

Link zur Publikation

Zusammenfassung:

Cellular agriculture is the controlled and sustainable manufacture of agricultural products with cells and tissues without plant or animal involvement. Today, microorganisms cultivated in bioreactors already produce egg and milk proteins, sweeteners, and flavors for human nutrition as well as leather and fibers for shoes, bags, and textiles. Furthermore, plant cell and tissue cultures provide ingredients that stimulate the immune system and improve skin texture, with another precommercial cellular agriculture product, in vitro meat, currently receiving a great deal of attention. All these approaches could assist traditional agriculture in continuing to provide for the dietary requirements of a growing world population while freeing up important resources such as arable land. Despite early successes, challenges remain and are discussed in this review, with a focus on production processes involving plant and animal cell and tissue cultures.

Human Adipose Stem Cells (hASCs) Grown on Biodegradable Microcarriers in Serum- and Xeno-Free Medium Preserve Their Undifferentiated Status
Wissenschaftlicher Artikel

Francesco Muoio, Stefano Panella, Jossen Valentin, Matias Lindner, Yves Harder, Michele Mueller, Regine Eibl, Tiziano Tallone

Journal of Functional Biomaterials, 2021 , vol.  12, no  25, pp.  1-28

Link zur Publikation

Zusammenfassung:

Human adipose stem cells (hASCs) are promising candidates for cell-based therapies, butthey need to be efficiently expandedin vitroas they cannot be harvested in sufficient quantities.Recently, dynamic bioreactor systems operated with microcarriers achieved considerable high celldensities. Thus, they are a viable alternative to static planar cultivation systems to obtain highnumbers of clinical-grade hASCs. Nevertheless, the production of considerable biomass in a shorttime must not be achieved to the detriment of the cells’ quality. To facilitate the scalable expansionof hASC, we have developed a new serum- and xeno-free medium (UrSuppe) and a biodegradablemicrocarrier (BR44). In this study, we investigated whether the culture of hASCs in defined serum-freeconditions on microcarriers (3D) or on planar (2D) cell culture vessels may influence the expressionof some marker genes linked with the immature degree or the differentiated status of the cells.Furthermore, we investigated whether the biomaterials, which form our biodegradable MCs, mayaffect cell behavior and differentiation. The results confirmed that the quality and the undifferentiatedstatus of the hASCs are very well preserved when they grow onBR44MCs in defined serum-free conditions. Indeed, the ASCs showed a gene expression profile more compatible with anundifferentiated status than the same cells grown under standard planar conditions

Chemically Defined, Xeno-Free Expansion of Human Mesenchymal Stem Cells (hMSCs) on Benchtop-Scale Using a Stirred Single-Use Bioreactor
Buchkapitel

Misha Teal, Jossen Valentin, Dieter Eibl, Regine Eibl

Dans Turksen Kursad,  Bioreactors in Stem Cell Biology. 2021,  New York, NY : Humana

Link zur Publikation

Zusammenfassung:

In recent years, the use of hMSCs, which may be isolated from adipose tissue among others, for the treatment of diseases has increased significantly. The cell quantities required for such therapeutic approaches, between 1012 and 1013, have thus far been predominantly produced using commercially available multi-tray systems, such as the Cell Factory (Thermo Fisher Scientific) or HYPERStack (Corning), which can be purchased with up to 40 layers. However, the handling of these planar multilayer systems is difficult, and process monitoring opportunities remain limited. Here, automated stirred single-use bioreactors provide a viable alternative to the time-consuming multiplication of cells using such planar systems, while still managing to achieve the desired clinically relevant quantities. In these stirred single-use systems, adherent cells are predominantly cultivated in suspension up to pilot scale using carrier materials, also referred to as microcarriers (MCs).This chapter describes the steps which need to be realized to guarantee successful hMSC expansion within a stirred single-use bioreactor (Eppendorf's BioBLU® 0.3c) operated using MCs under serum- and xeno-free conditions at benchtop scale. The cultivations were performed using an immortalized human adipose-derived mesenchymal stem cell (hASC) line, hence referred to as hASC52telo, and a new chemically defined, xeno-free medium, hence referred to as the UrSuppe formulation. Spinner flask cultivations were performed under comparable process conditions.

2020

An Approach towards a GMP Compliant In-Vitro Expansion of Human Adipose Stem Cells for Autologous Therapies
Wissenschaftlicher Artikel

Jossen Valentin, Francesco Muoio, Stefano Panella, Yves Harder, Tiziano Tallone, Regine Eibl

Bioengineering, 2020 , vol.  7, no  3, pp.  77-100

Link zur Publikation

Zusammenfassung:

Human Adipose Tissue Stem Cells (hASCs) are a valuable source of cells for clinical applications (e.g., treatment of acute myocardial infarction and inflammatory diseases), especially in the field of regenerative medicine. However, for autologous (patient-specific) and allogeneic (off-the-shelf) hASC-based therapies, in-vitro expansion is necessary prior to the clinical application in order to achieve the required cell numbers. Safe, reproducible and economic in-vitro expansion of hASCs for autologous therapies is more problematic because the cell material changes for each treatment. Moreover, cell material is normally isolated from non-healthy or older patients, which further complicates successful in-vitro expansion. Hence, the goal of this study was to perform cell expansion studies with hASCs isolated from two different patients/donors (i.e., different ages and health statuses) under xeno-and serum-free conditions in static, planar (2D) and dynamically mixed (3D) cultivation systems. Our primary aim was I) to compare donor variability under in-vitro conditions and II) to develop and establish an unstructured, segregated growth model as a proof-of-concept study. Maximum cell densities of between 0.49 and 0.65 × 10 5 hASCs/cm 2 were achieved for both donors in 2D and 3D cultivation systems. Cell growth under static and dynamically mixed conditions was comparable, which demonstrated that hydrodynamic stresses (P/V = 0.63 W/m 3 , τ_nt = 4.96 × 10 −3 Pa) acting at N s1u (49 rpm for 10 g/L) did not negatively affect cell growth, even under serum-free conditions. However, donor-dependent differences in the cell size were found, which resulted in significantly different maximum cell densities for each of the two donors. In both cases, stemness was well maintained under static 2D and dynamic 3D conditions, as long as the cells were not hyperconfluent. The optimal point for cell harvesting was identified as between cell densities of 0.41 and 0.56 × 10 5 hASCs/cm 2 (end of exponential growth phase). The growth model delivered reliable predictions for cell growth, substrate consumption and metabolite production in both types of cultivation systems. Therefore, the model can be used as a basis for future investigations in order to develop a robust MC-based hASC production process for autologous therapies.

Numerical Methods for the Design and Description of In Vitro Expansion Processes of Human Mesenchymal Stem Cells
Buchkapitel

Jossen Valentin, Dieter Eibl, Regine Eibl

Dans Herwig Christoph, Pörtner Ralf, Möller Johannes,  Digital Twins. 2020,  Cham : Springer

Link zur Publikation

Zusammenfassung:

Human mesenchymal stem cells (hMSCs) are a valuable source of cells for clinical applications (e.g., treatment of acute myocardial infarction or inflammatory diseases), especially in the field of regenerative medicine. However, for autologous (patient-specific) and allogeneic (off-the-shelf) hMSC-based therapies, in vitro expansion is necessary prior to the clinical application in order to achieve the required cell numbers. Safe, reproducible, and economic in vitro expansion of hMSCs for autologous and allogeneic therapies can be problematic because the cell material is restricted and the cells are sensitive to environmental changes. It is beneficial to collect detailed information on the hydrodynamic conditions and cell growth behavior in a bioreactor system, in order to develop a so called “Digital Twin” of the cultivation system and expansion process. Numerical methods, such as Computational Fluid Dynamics (CFD) which has become widely used in the biotech industry for studying local characteristics within bioreactors or kinetic growth modelling, provide possible solutions for such tasks.

In this review, we will present the current state-of-the-art for the in vitro expansion of hMSCs. Different numerical tools, including numerical fluid flow simulations and cell growth modelling approaches for hMSCs, will be presented. In addition, a case study demonstrating the applicability of CFD and kinetic growth modelling for the development of an microcarrier-based hMSC process will be shown.

Oxygen Mass Transfer in Biopharmaceutical Processes: Numerical and Experimental Approaches
Wissenschaftlicher Artikel

Sören Werner, Jossen Valentin, Dieter Eibl

Chemie Ingenieur Technik, 2020 , vol.  93, no  1-2, pp.  1-21

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Zusammenfassung:

Oxygen supply in aerobic bioprocesses is of crucial importance. For this reason, this paper presents the oxygen demand of different cells and summarizes experimental and numerical possibilities for the determination of oxygen transfer in bioreactors. The focus lies on the volumetric oxygen mass transfer coefficient (kLa) calculation using computational fluid dynamics and state?of?the?art models for surface?aerated and forced?aerated bioreactors. In addition, experimental methods for the determination of the kLa value and the gas bubble size distribution are presented.

2019

Advanced Therapy Medicinal Products (ATMPs) & GMP : 10 years after
Wissenschaftlicher Artikel

Christian van den Bos, Jossen Valentin, Dieter Eibl, Regine Eibl

pharmind, 2019 , vol.  4, pp.  508-513

Single‐Use Bioreactors – An Overview
Buchkapitel

Jossen Valentin, Regine Eibl, Dieter Eibl

Dans Eibl Regine, Eibl Dieter,  Single‐Use Technology in Biopharmaceutical Manufacture, Second Edition. 2019,  New York : John Wiley & Sons, Inc.

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Zusammenfassung:

This chapter aims to give an overview of instrumented single-use bioreactor (SUB) types that are currently available commercially. Based on the milestones in the development of SUBs, it focuses on the three most often applied types: wave-mixed, stirred, and orbitally shaken versions. Their typical working principles are explained, possible fields of application are deduced, and trends in their selection are summarized. The development of SUBs can be described in three phases: the early beginnings; the establishment of disposable membrane bioreactors, multitray cell culture systems, and the first bag bioreactors; and the expansion of wave-mixed, stirred, orbitally shaken, and further SUBs. The most suitable bioreactor for a desired application depends on several factors. The selection of the bioreactor is significantly influenced by the cultivation task, the engineering parameters of the bioreactor, the scale, the production organism, the legal requirements, the infrastructure, the know-how of staff, and the costs for investment and operation.

2018

Einweg-Bioreaktoren: Therapeutische Antikörper
Professioneller Artikel

Regine Eibl, Jossen Valentin, Dieter Eibl

Chemie in unserer Zeit, 2018 , vol.  52, no  4, pp.  230-237

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Zusammenfassung:

Einweg-Bioreaktoren sind in der Entwicklung und Produktion von monoklonalen Antikörpern angekommen. Studien belegen, dass sie richtig ausgewählt und eingesetzt sicherere, schnellere, grünere und kleinere Produktionsverfahren als ihre wiederverwendbaren Gegenspieler erlauben und zu Kosteneinsparungen beitragen. In der Inokulumproduktion haben sich wellendurchmischte Einweg-Bioreaktoren durchgesetzt, während die Anwender in der Fermentation mehrheitlich mit gerührten Einweg-Bioreaktoren arbeiten. Es wird prognostiziert, dass die Anzahl der Einweg-Bioreaktoren weiter zunehmen wird.

Manufacturing human mesenchymal stem cells at clinical scale: process and regulatory challenges
Wissenschaftlicher Artikel

Jossen Valentin, Christian van den Bos, Regine Eibl, Dieter Eibl

Applied Microbiology and Biotechnology, 2018 , vol.  102, no  4, pp.  1-14

Link zur Publikation

Zusammenfassung:

Human mesenchymal stem cell (hMSC)-based therapies are of increasing interest in the field of regenerative medicine. As economic considerations have shown, allogeneic therapy seems to be the most cost-effective method. Standardized procedures based on instrumented single-use bioreactors have been shown to provide billion of cells with consistent product quality and to be superior to traditional expansions in planar cultivation systems. Furthermore, under consideration of the complex nature and requirements of allogeneic hMSC-therapeutics, a new equipment for downstream processing (DSP) was successfully evaluated. This mini-review summarizes both the current state of the hMSC production process and the challenges which have to be taken into account when efficiently producing hMSCs for the clinical scale. Special emphasis is placed on the upstream processing (USP) and DSP operations which cover expansion, harvesting, detachment, separation, washing and concentration steps, and the regulatory demands.

Power Input Measurements in Stirred Bioreactors at Laboratory Scale
Wissenschaftlicher Artikel

Jossen Valentin

Journal of Visualized Experiments (jove), 2018

Link zur Publikation

Zusammenfassung:

The power input in stirred bioreactors is an important scaling-up parameter and can be measured through the torque that acts on the impeller shaft during rotation. However, the experimental determination of the power input in small-scale vessels is still challenging due to relatively high friction losses inside typically used bushings, bearings and/or shaft seals and the accuracy of commercially available torque meters. Thus, only limited data for small-scale bioreactors, in particular single-use systems, is available in the literature, making comparisons among different single-use systems and their conventional counterparts difficult. This manuscript provides a protocol on how to measure power inputs in benchtop scale bioreactors over a wide range of turbulence conditions, which can be described by the dimensionless Reynolds number (Re). The aforementioned friction losses are effectively reduced by the use of an air bearing. The procedure on how to set up, conduct and evaluate a torque-based power input measurement, with special focus on cell culture typical agitation conditions with low to moderate turbulence (100 < Re < 2·10?), is described in detail. The power input of several multi-use and single-use bioreactors is provided by the dimensionless power number (also called Newton number, P0), which is determined to be in the range of P0 ≈ 0.3 and P0 ≈ 4.5 for the maximum Reynolds numbers in the different bioreactors.

Growth Behavior of Human Adipose Tissue-Derived Stromal/Stem Cells at Small Scale: Numerical and Experimental Investigations
Wissenschaftlicher Artikel

Jossen Valentin, Regine Eibl, Matthias Kraume, Dieter Eibl

Bioengineering, 2018 , vol.  5, no  4, pp.  106-136

Link zur Publikation

Zusammenfassung:

Human adipose tissue-derived stromal/stem cells (hASCs) are a valuable source of cells for clinical applications, especially in the field of regenerative medicine. Therefore, it comes as no surprise that the interest in hASCs has greatly increased over the last decade. However, in order to use hASCs in clinically relevant numbers, in vitro expansion is required. Single-use stirred bioreactors in combination with microcarriers (MCs) have shown themselves to be suitable systems for this task. However, hASCs tend to be less robust, and thus, more shear sensitive than conventional production cell lines for therapeutic antibodies and vaccines (e.g., Chinese Hamster Ovary cells CHO, Baby Hamster Kidney cells BHK), for which these bioreactors were originally designed. Hence, the goal of this study was to investigate the influence of different shear stress levels on the growth of humane telomerase reversed transcriptase immortalized hASCs (hTERT-ASC) and aggregate formation in stirred single-use systems at the mL scale: the 125 mL (= SP100) and the 500 mL (= SP300) disposable Corning® spinner flask. Computational fluid dynamics (CFD) simulations based on an Euler–Euler and Euler–Lagrange approach were performed to predict the hydrodynamic stresses (0.06–0.87 Pa), the residence times (0.4–7.3 s), and the circulation times (1.6–16.6 s) of the MCs in different shear zones for different impeller speeds and the suspension criteria (Ns1u, Ns1). The numerical findings were linked to experimental data from cultivations studies to develop, for the first time, an unstructured, segregated mathematical growth model for hTERT-ASCs. While the 125 mL spinner flask with 100 mL working volume (SP100) provided up to 1.68.105 hTERT-ASC/cm2 (= 0.63 × 106 living hTERT-ASCs/mL, EF 56) within eight days, the peak living cell density of the 500 mL spinner flask with 300 mL working volume (SP300) was 2.46 × 105 hTERT-ASC/cm2 (= 0.88 × 106 hTERT-ASCs/mL, EF 81) and was achieved on day eight. Optimal cultivation conditions were found for Ns1u < N < Ns1, which corresponded to specific power inputs of 0.3–1.1 W/m3. The established growth model delivered reliable predictions for cell growth on the MCs with an accuracy of 76–96% for both investigated spinner flask types.

2017

Stirred Bioreactors: Current State and Developments, With Special Emphasis on Biopharmaceutical Production Processes
Buchkapitel

Jossen Valentin, Regine Eibl, Ralf Pörtner, Matthias Kraume, Dieter Eibl

Dans Larroche Christian, Sanroman Maria Angeles, Du Guocheng, Pandey Ashok,  Current Developments in Biotechnology and Bioengineering: Bioprocesses, Bioreactors and Controls. 2017,  online : Elsevier B.V.

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Zusammenfassung:

It is undisputed that stirred bioreactors are the most common type of bioreactors for submerged bioprocesses. Stirred bioreactors for biopharmaceuticals, which represent the strongest-growing branch of modern biotechnological products, are the focus of our chapter. They are available in working volumes from milliliters up to 70 m3 and are used for a wide range of expression systems that require aseptic conditions. Based on a brief summary of some basic rules for the construction and operation of stirred bioreactors, this chapter will review key engineering issues that affect the design of aseptic stirred bioreactors. We will also provide readers with some general information on transport phenomena in stirred bioreactors before discussing special design aspects and performance parameters for expression systems (mammalian and microbial cells) that prevail in commercial biopharmaceutical production processes. Special attention will be paid to single-use stirred bioreactors, which may have advantages over their reusable counterparts and have become widely accepted over the past years.

2016

Theoretical and Practical Issues That Are Relevant When Scaling Up hMSC Microcarrier Production Processes
Wissenschaftlicher Artikel

Jossen Valentin, Cedric Schirmer, Dolman Mostafa Sindi, Regine Eibl, Matthias Kraume, Ralf Pörtner, Dieter Eibl

Stem Cells International, 2016 , vol.  2016, no  4760414

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Zusammenfassung:

The potential of human mesenchymal stem cells (hMSCs) for allogeneic cell therapies has created a large amount of interest. However, this presupposes the availability of efficient scale-up procedures. Promising results have been reported for stirred bioreactors that operate with microcarriers. Recent publications focusing on microcarrier-based stirred bioreactors have demonstrated the successful use of Computational Fluid Dynamics (CFD) and suspension criteria (NS1u, NS1) for rapidly scaling up hMSC expansions from mL- to pilot scale. Nevertheless, one obstacle may be the formation of large microcarrier-cell-aggregates, which may result in mass transfer limitations and inhomogeneous distributions of stem cells in the culture broth. The dependence of microcarrier-cell-aggregate formation on impeller speed and shear stress levels was investigated for human adipose derived stromal/stem cells (hASCs) at the spinner scale by recording the Sauter mean diameter (d32) versus time. Cultivation at the suspension criteria provided d32 values between 0.2 and 0.7 mm, the highest cell densities (1.25 × 106 cells mL−1 hASCs), and the highest expansion factors (117.0 ± 4.7 on day 7), while maintaining the expression of specific surface markers. Furthermore, suitability of the suspension criterion NS1u was investigated for scaling up microcarrier-based processes in wave-mixed bioreactors for the first time.

Novel probes for pH and dissolved oxygen measurements in cultivations from millilitre to benchtop scale
Wissenschaftlicher Artikel

Caspar Demuth, Joel Varonier, Jossen Valentin, Regine Eibl, Dieter Eibl

Applied Microbiology and Biotechnology, 2016 , vol.  100, no  9, pp.  3853-3863

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Zusammenfassung:

pH value and the concentration of dissolved oxygen (DO) are key parameters to monitor and control cell growth in cultivation studies. Reliable, robust and accurate methods to measure these parameters in cultivation systems in real time guarantee high product yield and quality. This mini-review summarises the current state of the art of pH and DO sensors that are applied to bioprocesses from millilitre to benchtop scale by means of a short introduction on measuring principles and selected applications. Special emphasis is placed on single-use bioreactors, which have been increasingly employed in bioprocess development and production in recent years. Working principles, applications and the particular requirements of sensors in these cultivation systems are given. In such processes, optical sensors for pH and DO are often preferred to electrochemical probes, as they allow semi-invasive measurements and can be miniaturised to micrometre scale or lower. In addition, selected measuring principles of novel sensing technologies for pH and DO are discussed. These include solid-state sensors and miniaturised devices that are not yet commercially available, but show promising characteristics for possible use in bioprocesses in the near future.

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Development of a method for reliable power input measurements in conventional and single-use stirred bioreactors at laboratory scale
Wissenschaftlicher Artikel

Stephan Kaiser, Sören Werner, Jossen Valentin, Matthias Kraume, Dieter Eibl

Engineering in Life Sciences, 2016 , vol.  17, no  5, pp.  500-511

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Zusammenfassung:

Power input is an important engineering and scale-up/down criterion in stirred bioreactors. However, reliably measuring power input in laboratory scale systems is still challenging. Even though torque measurements have proven to be suitable in pilot scale systems, sensor accuracy, resolution and errors from relatively high levels of friction inside bearings can become limiting factors at smaller scales. An experimental setup for power input measurements was developed in this study by focusing on stainless steel and single-use bioreactors in the single-digit volume range. The friction losses inside the air bearings were effectively reduced to less than 0.5% of the measurement range of the torque meter. A comparison of dimensionless power numbers determined for a reference Rushton turbine stirrer (NP = 4.17±0.14 for fully turbulent conditions) revealed good agreement with literature data. Hence, the power numbers of several reusable and single-use bioreactors could be determined over a wide range of Reynolds numbers between 100 and >104. Power numbers of between 0.3 and 4.5 (for Re = 104) were determined for the different systems. The rigid plastic vessels showed similar power characteristics to their reusable counterparts. Thus, it was demonstrated that the torque-based technique can be used to reliably measure power input in stirred reusable and single-use bioreactors at the laboratory scale. This article is protected by copyright. All rights reserved

2015

On-line-pH- und DO-Messungen in Mikrocarrier-basierter hMSC-Kultur
Wissenschaftlicher Artikel

Jossen Valentin, Carmen Schirmaier, Gernot John, Dieter Eibl, Regine Eibl

BIOspektrum, 2015 , vol.  24, pp.  422-424

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Zusammenfassung:

Spinner flasks are often used for microcarrie-based cultivations of human mesenchymal stem cells (hMSCs). Normally, they are not equipped with pH and dissolved oxygen (DO) probes. This application note describes the cultivation of hMSCs in single-use spinner flasks equipped with optical pH (SP-HP8) and DO (SP-PSt3) sensors for the first time. While reaching peak cell numbers between 4.1 × 107 cells and 5.9 × 107 cells in two cultivation runs, reliable DO and pH data were delivered.

2014

Scale-up of adipose tissue-derived mesenchymal stem cell production in stirred single-use bioreactors under low-serum conditions
Wissenschaftlicher Artikel

Carmen Schirmaier, Jossen Valentin, Stephan Kaiser, Frank Jüngerkes, Silke Brill, Agnieszka Safavi-Nab, Ann Siehoff, Christian van den Bos, Dieter Eibl, Regine Eibl

Engineering in Life Sciences, 2014 , vol.  14, no  3, pp.  292-303

Link zur Publikation

Zusammenfassung:

Suspension cultures, in which human mesenchymal stem cells (hMSCs) are cultivated on microcarriers in scalable single-use stirred bioreactor types, have been shown to be a promising alternative to planar flask cultures. However, stirred single-use bioreactors were originally developed for production processes with robust, permanent cell lines. hMSCs are adherent primary cells and thus expanding them in such bioreactor systems imposes more stringent requirements on bioreactor systems. For low-serum conditions (5%) and different types of stirred single-use bioreactors, a suspension criteria-based approach for expanding human adipose tissue-derived mesenchymal stem cells (hASCs) from mL- to pilot scale was successfully developed. For process scale-up, experimental and numerical investigations were performed to (1) predict optimum impeller speeds, (2) determine the main engineering parameters (local shear stress, turbulent dissipation rate, Kolmogorov microscale), and (3) verify suspension criteria NS1 and NS1u for rapid process transfer from 100 mL to 2 L and 35 L cultures. Using optimized medium-microcarrier combinations as well as NS1 and NS1u as scale-up factors, total hASC quantities between 3 · 107 (100 mL scale) and 1 · 1010 (35 L scale) were obtained. The cell quantities obtained are the highest reported to date for scalable single-use bioreactors under low-serum conditions.

Modification and qualification of a stirred single-use bioreactor for the improved expansion of human mesenchymal stem cells at benchtop scale
Wissenschaftlicher Artikel

Jossen Valentin, Stephan Kaiser, Carmen Schirmaier, Jaqueline Herrmann, Alexander Tappe, Dieter Eibl, Ann Siehoff, Christian van den Bos, Regine Eibl

Pharmaceutical Bioprocessing, 2014 , vol.  2, no  4, pp.  311-322

Link zur Publikation

Zusammenfassung:

Background: To improve cultivation conditions for human bone-marrow-derived mesenchymal stem cells, we redesigned the commercially available UniVessel® SU bioreactor using results obtained from computational fluid dynamics. The goal was to produce ≥1 × 109 cells and to achieve expansion factors ≥30. Screening studies suggested that microcarrier solid fractions of at least 0.3% are required to reach the appropriate cell densities. Results: The fluid flow pattern found in the most promising modification (#2) was altered by increasing the impeller blade angle and lowering the off-bottom clearance. As a result, the maximum required specific power input was reduced by a factor of 2.2–4.6, depending on the microcarrier concentration, and peak cell densities were 3.4-times higher than in the standard version. Conclusion: The peak cell number of nearly 1.1 × 109 cells (expansion factor = 35), which was achieved in our low-serum cultivations, indicates an improvement in the redesigned UniVessel® SU configuration for bone marrow-derived mesenchymal stem cell expansions.

Mass Production of Mesenchymal Stem Cells — Impact of Bioreactor Design and Flow Conditions on Proliferation and Differentiation
Buchkapitel

Jossen Valentin, Ralf Pörtner, Stephan Kaiser, Matthias Kraume, Dieter Eibl, Regine Eibl

Dans Eberli Daniel,  Cells and Biomaterials in Regenerative Medicine. 2014,  10.5772/59385 : IntechOpen

Link zur Publikation

2013

Expansion of mesenchymal adipose-tissue derived stem cells in a stirred single-use bioreactor under low-serum conditions
Professioneller Artikel

Carmen Schirmaier, Stephan Kaiser, Jossen Valentin, Silke Brill, Frank Jüngerkes, Christian van den Bos, Dieter Eibl, Regine Eibl

23rd European Society for Animal Cell Technology (ESACT) Meeting: Better Cells for Better Health BMCProceedings, 2013 , vol.  7, no  6

Link zur Publikation

2012

Fluid Flow and Cell Proliferation of Mesenchymal Adipose-Derived Stem Cells in Small-Scale, Stirred, Single-Use Bioreactors
Wissenschaftlicher Artikel

Stephan Kaiser, Jossen Valentin, Carmen Schirmaier, Dieter Eibl, Silke Brill, Christian van den Bos, Regine Eibl

Chemie Ingenieur Technik, 2012 , vol.  85, no  1-2, pp.  95-102

Link zur Publikation

Zusammenfassung:

The fluid flow and suspension characteristics inside small-scale, stirred, single-use bioreactors were investigated experimentally and by means of computational fluid dynamics. The required impeller speeds for homogenous suspension were determined for two microcarrier types. The shear stress level and turbulence distribution were predicted using a numerical model, which was verified by particle image velocimetry measurements. In subsequent cultivations of primary mesencymal adipose-derived stem cells, up to 31.4-fold expansion in cell number was achieved for serum concentrations as low as 5 %.

2018

Multiphase CFD simulations for supporting MC-based hMSC process development
Konferenz

Jossen Valentin

5thBioProScale Symposium, 20.03.2018 - 22.03.2018, Berling, Germany

2017

Standardized expansion of human adipose tissue-derived stromal/stem cells (hASCs) in wave-mixed single-use bioreactors with one-dimensional motion
Konferenz

Jossen Valentin, Cedric Schrimer, Regine Eibl, Dieter Eibl, Matthias Kraume, Ralf Pörtner

ECI: Single Use Technologies II Bridging Polymer Science to Bioprocess Applications, 07.05.2017 - 10.05.2017, Tomar, Protugal

Link zur Konferenz

Zusammenfassung:

The large number of realized and ongoing clinical trials with human mesenchymal stem cells (200 by end of June 2016; www.clinicaltrial.gov) demonstrate their great potential in the field of regenerative medicine. However, new and standardized production technology is necessary to generate the number of cells required at the cell quality desired. The previously used parallelized plastic shells that consist of multiple layers (e.g. stacked plate systems) need to be replaced by instrumented, dynamically-mixed and scalable single-use bioreactors. In these systems the growth surface for the adherent growing cells is provided by microcarriers. While the first users have already worked with stirred single-use bioreactors (e.g. Lonza in Walkersville), there have only been two approaches described in the specialist literature for one-dimensional wave-mixed bioreactors, both of which were not completely successful. This is surprising, since these single-use bioreactors have dominated seed inoculum production for some time due to their uncomplicated operation. Furthermore, wave-mixed single-use bioreactors have proven to be successful in microcarrier-based vaccine production. The investigations we present consist of Computational Fluid Dynamics (CFD) simulations and data verification by Particle Image Velocimetry (PIV) measurements, suspension studies in a serum-reduced culture medium with a suitable polystyrene microcarrier, and expansion studies with human adipose tissue-derived stromal/stem cells (hASCs) based on biochemical engineering investigations. They provide an important contribution to the development and production of stem cell-based cell therapeutics in single-use bioreactors with one-dimensional motion, and for the first time demonstrate the suitability of this type of reactor for the expansion of hASCs when working with culture specific suspension criteria (NS1UW and NS1W). The first proof-of-concept expansions of hASCs at a rocking angle/rocking rate combination of 4° and 31 rpm allowed 2.85 x 108 hASCs to be harvested after 9 days of cultivation without a change in the stem cell characteristics. Moreover, the results generated contribute to a better understanding of the parameter-dependent, hydrodynamic cell stress caused by mechanical stress in hASC expansions with microcarriers in wave-mixed bioreactors with one-dimensional motion, and allow for direct comparison with stirred single-use bioreactors, which was previously not possible. Finally, the results obtained are also beneficial for other microcarrier-based production processes, such as those for vaccines (e.g. shortening of process development time).

How to use Computational Fluid Dynamics in the development of cell therapeutics
Konferenz

Jossen Valentin, Regine Eibl, Dieter Eibl

ECI: Scale-up and Manufacturing of Cell-based Therapies V, 15.01.2017 - 19.01.2017, San Diego, USA

Link zur Konferenz

Zusammenfassung:

Computational Fluid Dynamics (CFD) is an established method in fluid mechanics that allows fluidic problems to be solved through numerical methods. In recent years, CFD has established itself as a useful tool in biochemical engineering, where it is mainly used to characterise and optimize devices (e.g. bioreactors, pumps, etc.). By using CFD, fundamental bioengineering parameters (e.g. turbulent dissipation rates, shear gradients) can be predicted independently of time and location. This allows process related parameters to be defined in silico, which reduces the number of experiments and costs. This is particularly important for the development of cell therapeutics, where the starting cell material is restricted and the batch costs are high. Recent economic reports have predicted a significant increase in cell therapeutics over the next few years, especially for human mesenchymal stem cells (hMSCs). This situation can also be seen in the high number of clinical trials (269 trails, August 2016; clinicaltrails.gov) that are currently focusing on using hMSCs for the treatment of illnesses such as myocardial infarction, Crohn’s disease and graft versus host disease. However, large amounts of hMSCs are required for one single therapeutic dose (35-350 million cells per dose), which explains the demand for efficient and scalable in vitro expansion procedures.

Following a brief introduction to CFD, we aim to highlight the capabilities of CFD for the development of bioprocesses and scale-up procedures. For this purpose, we will show how CFD data can be used to support the scale-up of a microcarrier-based hMSC expansion process in stirred and wave-mixed single-use bioreactors. Our presented investigations involve Computational Fluid Dynamics (CFD) simulations and data verification using Particle Image Velocimetry (PIV) measurements, suspension studies in a serum-reduced culture medium with a suitable polystyrene microcarrier, and expansion studies with human adipose tissue-derived stromal/stem cells (hASCs). This combination of biochemical engineering and biological expertise enabled the establishment of a MC-based hMSC expansion process that resulted in up to 1.25 x 106 hMSCs/mL in stirred single-use bioreactors. Initial proof-of-concept expansions of hASCs in wave-mixed single-use bioreactors at a rocking angle/ -rate combination of 4° and 31 rpm resulted in the harvest of 2.85 x 108 hASCs after 9 days of cultivation without changes to the stem cell characteristics. All the investigations performed showed that the suspension criteria NS1U for stirred and NS1UW for wave-mixed bioreactors are beneficial for the cultivation of hMSCs.

2016

Scale-up considerations for cell therapies
Konferenz

Jossen Valentin

Informa conference: Cell therapy manufacturing & gene therapy, 02.12.2016 - 03.12.2016, Brussels, Belgium

Fluid flow simulations in a porous scaffold to improve co-cultivation of human mesenchymal stem cells and human microvascular endothelial cells for bone tissue engineering
Konferenz

Jossen Valentin

BMT Dreiländertagung, 04.10.2016 - 06.10.2016, Basel, Switzerland

Link zur Konferenz

How to use Computational Fluid Dynamics in the development of cell therapeutics
Konferenz

Jossen Valentin

BioTech 2016 Biopharmaceutical Manufacturing and Single-Use Technologies, 05.09.2016 - 06.09.2016, Wädenswil, Switzerland

Overview: Single-use technologies in biopharmaceutical manufacturing
Konferenz

Jossen Valentin

AchemAsia 2016, 09.05.2016 - 12.05.2016, Beijing, China

2015

Successfully employing single-use bioreactors for different expression systems
Konferenz

Jossen Valentin

ECI: Single Use Technologies I Bridging Polymer Science to Bioprocess Applications, 18.10.2015 - 21.10.2015, Leesburg, USA

Link zur Konferenz

Zusammenfassung:

The single-use bioreactor market was estimated to have grown at a CAGR of 20% in 2014. Wave-mixed bioreactors and stirred bioreactors represent the largest segment of today`s single-use bioreactor market, and are preferred by process developers and manufacturers of preclinical and clinical samples when high-value products up to medium scale and products requiring high safety demands are in focus. However, wave-mixed and stirred single-use bioreactors are also becoming increasingly important for commercial biopharmaceutical productions. This is ascribed to the availability of high-productivity cell lines, which require smaller bioreactors. In addition, the availability of bioengineering data and cultivation bags with improved films enables optimized operations with wave-mixed and stirred single-use bioreactors, whereby the risks of leachable migration and adsorption of hydrophobic components are reduced.

Based on a brief description of the current single-use bioreactor market, we aim to highlight the predominance of wave-mixed and stirred versions, and their main applications. In addition, their advantages and limitations are summarized. An insight into our long-term work with wave-mixed and stirred single-use bioreactors is given in the main part of our presentation, and results from cultivations with different expression systems covering plant cells, insect cells and mesenchymal stem cells (1-4) are provided. In this context, the advantageous combination of bioengineering and cell biological expertise is demonstrated for both process development and scale-up.

How can Computational Fluid Dynamics be used in order to rapidly expand bone marrow derived hMSCs in clinically relevant numbers
Konferenz

Jossen Valentin

DECHEMA Himmelfahrtstagung: Scale-up And Scale-down Of Bioprocesses, 11.05.2015 - 13.05.2015, Hamburg, Germany

2014

A successful approach in order to rapidly scaling-up microcarrier-based expansion of human mesenchymal stem cells for allogeneic therapies
Konferenz

Jossen Valentin

3rd International Conference on Tissue Science & Regenerative Medicine, 24.09.2014 - 26.09.2014, Valencia, Spain

Optimization of a benchtop single-use bioreactor for the expansion of adipose tissue and bone marrow derived human mesenchymal stem cells
Konferenz

Jossen Valentin

41st Annual European Society for Arti cial Organs Congress, 17.09.2014 - 20.09.2014, Rome, Italy

Numerische Strömungsuntersuchungen für die Massstabsübertragung der Expansion von humanen mesenchymalen Stammzellen in gerührten Single-Use Bioreaktoren
Konferenz

Jossen Valentin, Stephan Kaiser, Regine Eibl, Dieter Eibl

17. Köthener Rührer-Kolloquium, 26.06.2014 - 26.06.2014, Köthen, Germany

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