Biopharmaceutical Manufacturing in Single-Use Bioreactors Current Status and Challenges from a CDMO Perspective Dr

Wissenschaft und Technik Biotechnologie

Biopharmaceutical Manufacturing in Single-Use Bioreactors Current Status and Challenges from a CDMO Perspective Dr. Tina Lütke-Eversloh and Peter Rogge Rentschler Biopharma SE, Laupheim n ABSTRACT n ZUSAMMENFASSUNG Given today's fast growing biopharmaceutical market, a broad Herstellung von Biopharmazeutika in Single-Use-Bio- portfolio of Good Manufacturing Practice (GMP)-compliant reaktoren – Aktueller Status und Herausforderungen aus manufacturing options is available from traditional stainless der Sicht eines Dienstleistungsunternehmens steel equipment to various disposable devices. This article Im Hinblick auf den schnell wachsenden Markt bieten focuses on benefits and drawbacks of implementing single-use leistungsfähige biopharmazeutische Dienstleistungsunterneh- equipment for manufacturing biopharmaceuticals to meet men ein breites Spektrum von Good-Manufacturing-Practice both process efficiency and regulatory guidelines. Depending (GMP)-konformen Herstellungsprozessen an. Dabei kommen on the type of product and the client's requirements, e. g. neben klassischen Edelstahlproduktionsanlagen auch diverse provision of material for clinical trials, single-use facilities Single-Use-Systeme zum Einsatz. Dieser Beitrag fasst die allow fast changeover times and a high degree of flexibility. On wichtigsten Vor- und Nachteile der Implementierung von the other hand, Contract Development and Manufacturing Single-Use-Systemen in der biopharmazeutischen Produktion Organizations (CDMOs) face considerable challenges asso- zusammen. Dabei spielen sowohl die Verbesserung der ciated with single-use plastics such as material integrity and Effizienz als auch die Gewährleistung der behördlichen solid (particulates) or chemical (leachables, extractables) Konformität eine wichtige Rolle. Zum einen erlauben Single- contaminations. In addition, an effective supply chain Use-basierte Prozesse einen schnellen Produktwechsel in der management is required to ensure continuous availability of GMP-Produktionsanlage und damit eine extrem hohe Flex- consumables and consistent operational quality. ibilität. Zum anderen gibt es beachtliche Herausforderungen, die z. B. die Materialbeschaffenheit oder partikuläre sowie chemische Verunreinigungen der Kunststoffmaterialien be- treffen. Weitere Herausforderungen sind die kontinuierliche Bereitstellung der erforderlichen Single-Use-Komponenten sowie die Qualitätssicherung durch ein effektives Manage- ment der Zulieferkette.

Introduction cally, modern biopharmaceuticals Upstream Processing (USP) exhibit improved potency and phar- The manufacturing of biopharma- macokinetic properties, and thus, In general, the mode of operation of ceuticals constitutes a fast growing can be applied in significantly lower stirred bioreactors for mammalian market segment, and many thera- doses. Since many companies out- cell cultures is comparable for both peutic proteins are currently in clini- source the process development and stainless steel and single-use vessels cal trials and more sophisticated material supply for clinical studies, [3, 4]. Chinese hamster ovary (CHO) molecules for specialized therapies the field of Contract Development cells are favored hosts for recombi- are developed. These include mono- and Manufacturing Organizations nant production of therapeutic pro- clonal antibodies (mAb), enzymes, (CDMOs) is growing as well [1, 2]. teins due to the glycosylation pat- hormones, fusion proteins and other In this article, single-use and terns and excellent cultivation tech- recombinant proteins, which are multi-use equipment for mamma- niques. Beside fast and easy fed- used for the treatment of severe lian cell cultures and related USP batch processes yielding high prod- diseases, particularly cancer, auto- techniques for the manufacturing of uct titers, CHO suspension cultures immune and genetic disorders. Typi- therapeutic proteins are compared. can also be subjected to continuous

Pharm. Ind. 80, Nr. 2, 281–284 (2018) © ECV • Editio Cantor Verlag, Aulendorf (Germany) Lütke-Eversloh und Rogge • Pharmaceutical Manufacturing 281 Wissenschaft und Technik Biotechnologie

bioprocessing which allows relative- cause suitable depth filters with sig- tors can be customized, extended ly high volumetric productivities as nificantly improved capacities are and are quickly assembled, which is well as protein uniformity and con- commercially available by now. of special interest if multiple prod- sistency [5–7]. At the authors’ com- ucts are manufactured in the same pany, various USP projects em- GMP suite. The flexible design of ployed continuous cell cultivation single-use devices also allows modu- Advantages of with or without cell retention in lar concepts, which can include ei- Single-Use Equipment both stainless steel and single-use ther fully disposable equipment or bioreactors. However, most of the The main reasons to prefer single- stainless steel/single-use hybrid so- cell culture processes are conducted use equipment are the lower risks of lutions [14]. With respect to opera- in the straightforward fed-batch cross-contaminations, lower clean- tion, utilities and maintenance, full mode with monoclonal antibodies ing and sterilization efforts, higher single-use facilities can reduce the being currently the most frequent flexibility, faster changeover and operating costs for an average mAb type of biopharmaceutical. In the lower investment costs. Reduced process by > 20 % and labor costs last 12 months, more than 50 % of complexity, lower maintenance, by > 10 % [15]. the Good Manufacturing Practice time-savings and regulatory con- (GMP) drug substance batches were formity clearly increased the portion operated in single-use bioreactors at of single-use equipment in biophar- Disadvantages of the authors’ company. maceutical manufacturing during Single-Use Equipment The USP for biopharmaceutical the past years [10–12]. In addition, manufacturing does not only in- more new devices are developed and Besides the benefits mentioned clude the production bioreactor, but existing technologies are contin- above, single-use devices do also also the seed train from cell thawing uously improved by the vendors face some drawbacks; an overview is to inoculation and the harvest pro- such as a single-use bioreactor with provided in Table 1. First, single-use cedure after cultivation. The sub- a nominal volume of 4 900 L [13]. bioreactors are scale-limited due to cultivation steps are typically con- Moreover, installation of single- pressure, gas transfer and mixing is- ducted in single-use shake flasks or use equipment is faster and results sues. Only recently, the commonly single-use wave bioreactors as seed in a smaller footprint due to the lack used maximum volume of 2 000 L train. Several stirred tank bioreac- of CIP and SIP (cleaning/steriliza- was extended to 4 900 L with a work- tors of different scales are imple- tion in place) requirements. Bioreac- ing volume of up to 4 000 L [13]. mented in the upscale cultivation line to generate sufficient cell masses for inoculating the produc- n Table 1 tion bioreactor. Different harvest procedures can Major advantages and drawbacks of single-use equipment for be used to remove cells and cell de- biomanufacturing. bris and finally to provide clarified liquid for the first downstream pro- Item Single-use Stainless steel cess (DSP) step. An initial centrifu- Investment costs low high gation step is included in most pro- Technical complexity low high cesses, and single-use centrifugation Maintenance costs low high devices are available for volumes of Production scale limited not limited up to 6 000 L. Since cell debris and particles cannot be completely re- Consumables demand high medium moved, subsequent filtration with Material supply high medium porous depth filters is necessary to Cleaning efforts none high clarify the harvest broth, followed Sterilization time none high by a final 0.2-µm filtration. Depend- Waste production high medium ing on the process and the facility’s equipment, direct filtration is an al- Energy consumption medium high ternative option [8, 9]. The authors’ Inventory and storage space medium high company’s standard for processes Biological contamination risk low medium conducted in single-use USP units Chemical contamination risk medium low comprises two depth and one Product changeover quick slow membrane filtration steps without cell removal by centrifugation, be- Overall processing time quick medium

Pharm. Ind. 80, Nr. 2, 281–284 (2018) 282 Lütke-Eversloh und Rogge • Pharmaceutical Manufacturing © ECV • Editio Cantor Verlag, Aulendorf (Germany) However, scales larger than 2 000 L are not always required for a CDMO, because modern biopharmaceuti- cals often have a significantly im- proved potency and half-life in the patient’s body. Hence, clearly lower amounts of product are required for the therapy and many new biopro- cesses enable good titers and yields, minimizing the need for production scales of > 2 000 L. The materials have a consider- able impact on employing a single- use-based bioprocess: the demand of consumables is high and suitable strategies for logistics (e. g. long de- livery times) and storage (e. g. shelf life) must be carefully evaluated. Since storage of consumables is lim- ited, material integrity plays a piv- otal role with respect to availability in the case of breaks or leakages or other malfunctions. In general, the continuous supply of material of consistent quality must be guaran- teed, which constitutes the core of the authors’ company supply chain management conducted in close co- operation with the supplier. Never- theless, having a second material supply source (if available) is highly recommendable for a CDMO, which can either be the same supplier with alternative raw material sources or a different supplier. In addition to particulates origi- nating from the plastic material it- self, seals, gaskets or the like, chemi- cal contaminations might cause se- vere problems for the cell culture and/or the final active pharmaceuti- cal ingredient (API) [16]. The main contaminants are differentiated ac- cording to the conditions, which led to their release into the product- containing liquid: whereas leacha- bles migrate under actual process conditions, extractables migrate in- to the test solution under harsh ex- cessive conditions. A multi-stage evaluation concept for leachables Figure 1: Comparison of costs for biopharmaceutical manufacturing using stainless and extractables is therefore per- steel and single-use equipment. (a) Distribution of COGs, (b) relation of costs and formed: after an overall risk assess- annually produced amounts of drug substance, (c) relation of costs and total number of batches per year (source: Rentschler Biopharma SE). ment to evaluate relevant materials, a component-specific risk assess- ment is conducted. Considering

Pharm. Ind. 80, Nr. 2, 281–284 (2018) © ECV • Editio Cantor Verlag, Aulendorf (Germany) Lütke-Eversloh und Rogge • Pharmaceutical Manufacturing 283 Wissenschaft und Technik Biotechnologie

product safety as most important yields a price of approximately 250 [6] Li F, Vijayasankaran N, Shen Y, Kiss R, parameter, the supplier’s data are euros per gram drug substance, Amanullah A. Cell culture process for monoclonal antibody production. mAbs, analyzed and compared to the theo- whereas 280 euros g-1 are estimated 2010, 2: 466–479. retical worst-case biomanufacturing for 36 batches performed in the [7] Schmidt SR. Drivers, opportunities, and conditions. If the theoretical ex- 2 000-L single-use facility. This price limits of continuous processing. BioPro- cess Int, 2017, 15(3): 30–37. tractable concentration might attain advantage of stainless steel over sin- [8] Felo M, Christensen B, Higgins J. Process a critical dose, remaining safety con- gle-use further improves with in- cost and facility considerations in the cerns lead to further investigations. creasing quantities of drug manu- selection of primary cell culture clarifi- cation technology. Biotechnol Prog, For this, standardized tests for facturing, which is typically the case 2013, 29: 1239–1245. single-use equipment were develop- for commercial supply of biophar- [9] Schmidt SR, Wieschalka S., Wagner R. ed in the past few years [17]. maceuticals (fig. 1b). In contrast, the Single-use depth filters. BioProcess Int, 2017, 14(1)i: 6–11. economic benefits reverse in favor [10] Shukla AA, Gottschalk U. Single-use dis- of single-use bioreactors if the total posable technologies for biopharmaceu- number of batches per year is sur- tical manufacturing. Trends Biotechnol, Economic Considerations 2013, 31: 147–154. veyed, and these price trends further [11] Langer ES. Advances in bioprocessing. In addition to balancing the above- bifurcate with lower numbers of BioProcess Int, 2015, 13(11)s: 2–5. mentioned pros and cons, economic batches (fig. 1c). [12] Calcott PH. Disposable components in biomanufacturing: A regulatory per- aspects are taken into account to spective. BioProcess Int, 2015, 13(11)s: decide whether a process is operat- 6–9. ed in a multi- or single-use bioreac- [13] Stanton D. ABEC breaks the 2,000L Conclusions “plastic ceiling” with 4,900L single-use tor. To illustrate this, a typical fed- bioreactor. https://www.biopharma-re- batch process of mAb manufactur- Since recently, the popularity of sin- porter.com/Article/2017/07/12/ABEC- ing is given to compare the costs of gle-use equipment for biomanufactu- breaks-the-2-000L-ceiling-with-4-900L- single-use-bioreactor using the 3 000 L stainless steel and ring highly increased, particularly at [14] Minow B, Rogge P, Thompson K. Imple- 2 000 L single-use facilities. The cal- small- and mid-scale CDMOs. The menting a fully disposable mAb manu- culations include the entire pharma- authors’ company decided to operate facturing facility. BioProcess Int, 2012, 10(6): 48–57. ceutical production process, i. e. both single-use and stainless steel [15] Levine HL, Stock R, Lilja JE, Gaasvik Å, both USP and DSP, to manufacture facilities, enabling a high degree of Hummel H, Ransohoff TC, Jones SD. Sin- 100 kg of drug substance with a pro- flexibility. However, there is no gold- gle-use technology and modular const- ruction. BioProcess Int, 2013, 11(4)s: 40– ductivity of 2 g L-1 and a purifica- en rule for deciding which the most 45. tion yield of 70 %. Capital invest- suitable facility is for all purposes, be- [16] Johnson MW. Understanding particu- ment is not considered, which is es- cause it strongly depends on the bio- lates in single-use bags. BioProcess Int, 2014, 12(4)s: 22–28. timated to be 30–35 million euros process and the customer [18, 19]. [17] Ding W, Madsen G, Mahajan E, O’Con- for a new stainless steel unit and Both technologies have advantages nor S, Wong K. Standardized ex- 20–25 million euros for a single-use and disadvantages with respect to tractables testing protocol for single-use systems in biomanufacturing. Pharm construction. The costs of goods costs, time and safety. Eng, 2014, 34: 1–11. (COGs) are split into direct and indi- [18] Rogge P, Müller D, Schmidt SR. The sin- rect labor costs, suite utilization, gle-use or stainless steel decision pro- cess. BioProcess Int, 2015, 13(11)s: 10– materials (raw materials, water, buf- n LITERATURE 15. fers, etc.), consumables (single-use [19] Schmidt SR. The benefits and limits of [1] Schmidt SR, Schwab B. Seize the mar- materials such as bags, tubing, fil- disposable technologies in manufactur- ket. Eur. Biopharm. Rev, Apr. 2017, 74– ing protein therapeutics. Am Pharm ters), and external costs (e. g. waste 77. Rev, July 2016. management, maintenance). How- [2] Liu, JKH. The history of monoclonal an- tibody development – progress, remain- ever, the choice of materials and ing challenges and future innovations. consumables used for the process Annals Med Surg, 2014, 3: 113–116. might significantly impact the costs. [3] Eible R, Eible D. Disposable bioreactors Correspondence: in cell culture-based upstream process- As shown in fig. 1a, the cost distribu- ing. BioProcess Int, February 2009, Dr. Tina Lütke-Eversloh tion differs only to a minor degree Suppl. 18–23. Senior Process Manager, USP Process between stainless steel and single- [4] Nienow, AW, Isailovic B, Barrett TA. De- Design and Validation sign and performance of single-use, stir- use equipment, whereas materials red-tank bioreactors. BioProcess Int, Rentschler Biopharma SE and labor constitute the main costs Nov. 2016, Suppl. 12–21. Erwin-Rentschler-Str. 21 for both types of facilities. [5] Gronemeyer P, Ditz R, Strube J. Recent 88471 Laupheim (Germany) advances in large-scale production of Conducting 24 batches per year monoclonal antibodies and related pro- e-mail: tina.luetke-eversloh@rentschler- in the 3 000-L stainless steel facility teins. Bioeng, 2014, 1: 188–212. biopharma.com

Pharm. Ind. 80, Nr. 2, 281–284 (2018) 284 Lütke-Eversloh und Rogge • Pharmaceutical Manufacturing © ECV • Editio Cantor Verlag, Aulendorf (Germany)