Biomanufacturing for clinically advanced cell therapies.
ABSTRACT: The achievements of cell-based therapeutics have galvanized efforts to bring cell therapies to the market. To address the demands of the clinical and eventual commercial-scale production of cells, and with the increasing generation of large clinical datasets from chimeric antigen receptor T-cell immunotherapy, from transplants of engineered haematopoietic stem cells and from other promising cell therapies, an emphasis on biomanufacturing requirements becomes necessary. Robust infrastructure should address current limitations in cell harvesting, expansion, manipulation, purification, preservation and formulation, ultimately leading to successful therapy administration to patients at an acceptable cost. In this Review, we highlight case examples of cutting-edge bioprocessing technologies that improve biomanufacturing efficiency for cell therapies approaching clinical use.
Project description:Human induced pluripotent stem cells (iPSCs) are ideal cell sources for personalized cell therapies since they can be expanded to generate large numbers of cells and differentiated into presumably all the cell types of the human body in vitro. In addition, patient specific iPSC-derived cells induce minimal or no immune response in vivo. However, with current cell culture technologies and bioprocessing, the cost for biomanufacturing clinical-grade patient specific iPSCs and their derivatives are very high and not affordable for majority of patients. In this paper, we explored the use of closed and miniature cell culture device for biomanufacturing patient specific neural stem cells (NSCs) from iPSCs. We demonstrated that, with the assist of a thermoreversible hydrogel scaffold, the bioprocessing including iPSC expansion, iPSC differentiation into NSCs, the subsequent depletion of undifferentiated iPSCs from the NSCs, and concentrating and transporting the purified NSCs to the surgery room, could be integrated and completed within two closed 15?ml conical tubes.
Project description:Collagen is a natural polymer found abundantly in the extracellular matrix (ECM). It is easily extracted from a variety of sources and exhibits excellent biological properties such as biocompatibility and weak antigenicity. Additionally, different processes allow control of physical and chemical properties such as mechanical stiffness, viscosity and biodegradability. Moreover, various additive biomanufacturing technology has enabled layer-by-layer construction of complex structures to support biological function. Additive biomanufacturing has expanded the use of collagen biomaterial in various regenerative medicine and disease modelling application (e.g., skin, bone and cornea). Currently, regulatory hurdles in translating collagen biomaterials still remain. Additive biomanufacturing may help to overcome such hurdles commercializing collagen biomaterials and fulfill its potential for biomedicine.
Project description:Colchicine is one of the oldest plant-based medicines used to treat gout and one of the most important alkaloid-based antimitotic drugs with anticancer potential, which is commercially extracted from Gloriosa superba. Clinical trials suggest that colchicine medication could prevent atrial fibrillation recurrence after cardiac surgery. In addition, therapeutic colchicine is undergoing clinical trials to treat non-diabetic metabolic syndrome and diabetic nephropathy. However, the industrial-scale biomanufacturing of colchicine have not yet been established. Clearly, further studies on detailed biorhizome-specific transcriptome analysis, gene expression, and candidate gene validation are required before uncover the mechanism of colchicine biosynthesis and biorhizome-based colchicine biomanufacturing. Annotation of 32312 assembled multiple-tissues transcripts of G. superba represented 15088 unigenes in known plant specific gene ontology. This could help understanding colchicine biosynthesis in G. superba. This review highlights the biorhizomes, rhizome specific genes or gene what expressed with high level in rhizomes, and deep fluid dynamics in a bioreactor specifically for the biomanufacture of colchicine.
Project description:Current limitations to on-demand drug manufacturing can be addressed by technologies that streamline manufacturing processes. Combining the production of two or more drugs into a single batch could not only be useful for research, clinical studies, and urgent therapies but also effective when combination therapies are needed or where resources are scarce. Here we propose strategies to concurrently produce multiple biologics from yeast in single batches by multiplexing strain development, cell culture, separation, and purification. We demonstrate proof-of-concept for three biologics co-production strategies: (i) inducible expression of multiple biologics and control over the ratio between biologic drugs produced together; (ii) consolidated bioprocessing; and (iii) co-expression and co-purification of a mixture of two monoclonal antibodies. We then use these basic strategies to produce drug mixtures as well as to separate drugs. These strategies offer a diverse array of options for on-demand, flexible, low-cost, and decentralized biomanufacturing applications without the need for specialized equipment.
Project description:The modern cell-free protein synthesis (CFPS) system is expanding the opportunity of cell-free biomanufacturing as a versatile platform for synthesizing various therapeutic proteins. However, synthesizing human protein in the bacterial CFPS system remains challenging due to the low expression level, protein misfolding, inactivity, and more. These challenges limit the use of a bacterial CFPS system for human therapeutic protein synthesis. In this study, we demonstrated the improved performance of a customized CFPS platform for human therapeutic protein production by investigating the factors that limit cell-free transcription-translation. The improvement of the CFPS platform has been made in three ways. First, the cell extract was prepared from the rare tRNA expressed host strain, and CFPS was performed with a codon-optimized gene for Escherichia coli codon usage bias. The soluble protein yield was 15.2 times greater with the rare tRNA overexpressing host strain as cell extract and codon-optimized gene in the CFPS system. Next, we identify and prioritize the critical biomanufacturing factors for highly active crude cell lysate for human protein synthesis. Lastly, we engineer the CFPS reaction conditions to enhance protein yield. In this model, the therapeutic protein filaggrin expression was significantly improved by up to 23-fold, presenting 28 ± 5 ?M of soluble protein yield. The customized CFPS system for filaggrin biomanufacturing described here demonstrates the potential of the CFPS system to be adapted for studying therapeutic proteins.
Project description:As breakthrough cellular therapy discoveries are translated into reliable, commercializable applications, effective stem cell biomanufacturing requires systematically developing and optimizing bioprocess design and operation. This article proposes a rigorous computational framework for stem cell biomanufacturing under uncertainty. Our mathematical tool kit incorporates: high-fidelity modeling, single variate and multivariate sensitivity analysis, global topological superstructure optimization, and robust optimization. The advantages of the proposed bioprocess optimization framework using, as a case study, a dual hollow fiber bioreactor producing red blood cells from progenitor cells were quantitatively demonstrated. The optimization phase reduces the cost by a factor of 4, and the price of insuring process performance against uncertainty is approximately 15% over the nominal optimal solution. Mathematical modeling and optimization can guide decision making; the possible commercial impact of this cellular therapy using the disruptive technology paradigm was quantitatively evaluated.
Project description:Recent years have witnessed the expansion of tissue failures and diseases. The uprising of regenerative medicine converges the sight onto stem cell-biomaterial based therapy. Tissue engineering and regenerative medicine proposes the strategy of constructing spatially, mechanically, chemically and biologically designed biomaterials for stem cells to grow and differentiate. Therefore, this paper summarized the basic properties of embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and adult stem cells. The properties of frequently used biomaterials were also described in terms of natural and synthetic origins. Particularly, the combination of stem cells and biomaterials for tissue repair applications was reviewed in terms of nervous, cardiovascular, pancreatic, hematopoietic and musculoskeletal system. Finally, stem-cell-related biomanufacturing was envisioned and the novel biofabrication technologies were discussed, enlightening a promising route for the future advancement of large-scale stem cell-biomaterial based therapeutic manufacturing.
Project description:Engineering organ-specific tissues for therapeutic applications is a grand challenge, requiring the fabrication and maintenance of densely cellular constructs composed of ~108 cells/ml. Organ building blocks (OBBs) composed of patient-specific-induced pluripotent stem cell-derived organoids offer a pathway to achieving tissues with the requisite cellular density, microarchitecture, and function. However, to date, scant attention has been devoted to their assembly into 3D tissue constructs. Here, we report a biomanufacturing method for assembling hundreds of thousands of these OBBs into living matrices with high cellular density into which perfusable vascular channels are introduced via embedded three-dimensional bioprinting. The OBB matrices exhibit the desired self-healing, viscoplastic behavior required for sacrificial writing into functional tissue (SWIFT). As an exemplar, we created a perfusable cardiac tissue that fuses and beats synchronously over a 7-day period. Our SWIFT biomanufacturing method enables the rapid assembly of perfusable patient- and organ-specific tissues at therapeutic scales.
Project description:Adipose tissue engineering offers a promising alternative to current breast reconstruction options. However, the conventional approach of using a scaffold in combination with adipose-derived precursor cells poses several problems in terms of scalability and hence clinical feasibility. Following the body-as-a-bioreactor approach, this study proposes a unique concept of delayed fat injection into an additive biomanufactured and custom-made scaffold. Three study groups were evaluated: Empty scaffold, Scaffold containing 4?cm(3) lipoaspirate and Empty scaffold +2-week prevascularisation period. In group 3, of prevascularisation, 4?cm(3) of lipoaspirate was injected into scaffolds after 2 weeks. Using a well-characterised additive biomanufacturing technology platform, patient-specific scaffolds made of medical-grade-polycaprolactone were designed and fabricated. Scaffolds were implanted in subglandular pockets in immunocompetent minipigs (n?=?4) for 24-weeks. Angiogenesis and adipose tissue regeneration were observed in all constructs. Histological evaluation showed that the prevascularisation?+?lipoaspirate group had the highest relative area of adipose tissue (47.32%?±?4.12) which was significantly higher than both lipoaspirate-only (39.67%?±?2.04) and empty control group (8.31%?±?8.94) and similar to native breast tissue (44.97%?±?14.12). This large preclinical animal study provides proof-of-principle that the clinically applicable prevascularisation and delayed fat-injection techniques can be used for regeneration of large volumes of adipose tissue.
Project description:Survival after acute paediatric (0-14 years), adolescent (15-19 years) and young adult (20-39 years) leukaemia has improved substantially over the last five decades, particularly for acute lymphoblastic leukaemia (ALL) and acute promyelocytic leukaemia. This progress represents one of the most successful achievements in the history of medicine and has been attributed to the development of effective chemotherapy regimens, improvement in supportive care, better risk stratification, use of targeted therapies, and advances in haematopoietic stem cell transplantation. Recent studies have revealed improvement in survival over time for all age groups and subtypes of leukaemia. However, these outcomes varied widely by age and are associated with sociodemographic and clinical factors. The present review concludes that survival and early death after acute leukaemia has greatly improved among young patients. However, inequalities in outcomes remain and are likely a result of multiple factors.