Viral vectors are essential therapeutic tools for gene therapy, genetically modified cell therapies and next-generation vaccines. They are used to deliver therapeutic genes or genetic material coding for antigens into host cells.
Gene therapy, an innovative class of biological products, has opened up new horizons making it possible to solve previously unmet medical needs, particularly in oncology and rare diseases. With more and more cell and gene therapy products coming to market, the FDA has forecast the approval of 10 to 20 new products a year between 2019 and 2025, based on an assessment of the pipeline and the clinical success rates of these products.
Against this backdrop, the bioprocessing of viral vectors, which is the most widespread modality, has become a strategic sector stimulating innovation and development to meet the technical, economic and logistical constraints of the field.
Here we provide an overview of the current challenges in viral vector biomanufacturing, as well as the emerging innovative technologies to meet the growing demand. We highlight the key figures for the viral vector biomanufacturing market and its major players.
The challenges of viral vectors biomanufacturing
Viral vectors are used for a variety of applications, including:
- in vivo gene therapy targeting rare acquired or inherited disorders
- engineering of autologous/allogeneic cells for ex vivo cell therapies
- recombinant vector vaccines
- engineering of plants and the animal gut microbiome.
In the field of gene therapy, adeno-associated viruses (AAVs) are the vector of choice due to their tissue tropism, low immunogenicity and genotoxicity, and efficient transduction with sustained gene expression. Lentiviruses provide highly stable transgene expression, which is why they are used for ex vivo gene therapy, mainly to produce CAR-T cells or modify pluripotent and haematopoietic stem cells. Adenoviruses have become a vector of oncolytic agents and vaccines based on their innate immunostimulatory role and replication activity.
Figure 1: Heterogeneity of viral vectors: size, genomic capacity and main applications. Source: R. Kilgore, 2023.
The development and production of viral vectors are more recent than those of other classes of biologics, such as monoclonal antibodies. There are currently no universal, standardised manufacturing processes as compared to the many gold-standards of monoclonal antibody bioprocessing. In addition, there are major differences between these two types of molecule that prevent methods from fluidly being transposed: viral vectors are larger and more complex molecules, requiring a higher level of control and biosafety (BSL-2 for most viral vectors compared with a BSL-1 classification for mAbs), which adds to production and installation costs.
From small-scale to industrial production
The players involved use different upstream production systems and downstream processes, and face various challenges. In particular, scaling up is a difficulty encountered by many academic laboratories, which are developing production processes on a small (laboratory scale), with cell cultures adhering in multilayer flasks. These processes, which require manual handling, make scale-up difficult if not impractical. For time and cost issues, companies still often produce the first early batches using these ‘laboratory’ processes. They are then faced with major challenges in transferring production processes and scaling-up to manufacture larger batches while meeting regulatory requirements.
The production of viral vectors is preceded by the design and manufacture of plasmids. Plasmid requirements are high and can be a critical step. Unlike CHO, the gold-standard cell line used for monoclonal antibody production, HEK293 cells, used for viral vector production, have fewer stable clones. Most of the time, viral vectors are produced using transient transfection. In other words, each time a recombinant virus is produced, several plasmids are needed to transfect the cells. This puts pressure on the manufacture of plasmids, as demand increases and influences plasmid supply costs.
Meeting the need for strict regulatory compliance
The challenges of meeting regulatory requirements and quality control in the production of viral vectors are crucial, as these products are intended for sensitive medical applications such as gene therapies and vaccines. Regulatory authorities, such as the FDA, Food and Drug Administration (United States) and the EMA, European Medicines Agency (Europe), impose strict standards to guarantee the safety, efficacy and traceability of vectors used in clinical treatments.
Analytical methods for controlling product quality throughout the process are therefore developed right from the research & development of the therapeutic product at laboratory level. Several quality control attributes (QCAs) are commonly analysed, including safety, identity, functionality, purity and viability. Controlling these QCAs enables us to demonstrate that the final product does not vary significantly from one batch to the next.
Determining these critical attributes is part of the Quality by Design (QbD) approach, which aims to integrate product quality right from the development stages. The aim is to ensure upstream that the sources of variability in the process are identified and dealt with, so as to guarantee that the finished product downstream conforms to the predefined characteristics. The QbD approach is not a regulatory requirement but a quality assurance approach, strongly recommended by the health authorities: all the new ICH (International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use), FDA and EMA guidelines use QbD concepts and terminology
Figure 2: Overview of the methodology using process and product knowledge to develop the QbD approach to serve the safety, purity and efficacy of AAV pharmaceutical products. QTPP: Quality Target Product Profile. CQA: Critical Quality Attributes, CPP: Critical Process Parameters. Source: Cytiva.
Innovations in the bioproduction of viral vectors
Evolution of culture methods
The HEK293 cell lines usually used for viral vector production are adherent cells. To overcome the limitations of traditional adherent cell culture methods, new technologies have been developed that offer considerable advantages for industrial production. These include suspension cell culture methods, which offer titers similar to those of adherent culture and support higher cell densities. Innovations in bioreactor suspension culture processes have led to production capacities of 1,000 to 2,000 litres.
Figure 3: Evolution of viral vector production processes using bioreactors for suspension production. Source: A Lamproye, 2023.
Purification technologies
Downstreaming viral vectors involves a number of constraints, particularly in relation to the size of the molecules, the fragility of the viruses and the presence of a large number of impurities (plasmid DNA, cellular proteins, etc).
Immunoaffinity chromatography (IAC) is currently the most widely used method for purifying viral vectors. Dynamic research by academics and industry has identified antibodies and antibody fragments used as affinity ligands specifically for viral vectors. A major breakthrough in IAC technology has been achieved with Cytiva’s AVB ligand, based on a single-domain camelid antibody (VHHs or Nanobodies®). This technology has stimulated the development of ligands designed for the purification of VVs (ThermoFisher’s POROS Capture Select or Repligen’s CaptureSelect)
Figure 4: Parallel between standard downstream process techniques (top) requiring several chromatography, filtration and capture steps; and innovative techniques (bottom) with a single affinity chromatography step. Source: O. Terova, 2016.
Ion exchange chromatography (IEX) is the main alternative to affinity chromatography for the primary capture step and is the predominant technology used for viral vector polishing. IEX has a powerful ability to separate empty capsids from full capsids, which is a significant advantage over affinity chromatography. This separation is important because full capsids are the carriers of therapeutic genetic material in gene therapy applications and higher full-to-empty ratios can increase treatment efficacy.
Recent successes with new methods such as steric exclusion chromatography and supports such as monoliths and membranes can reduce processing times by taking advantage of the size properties of viral vectors. Further developments should be done on continuous chromatography, which could enable the process to be intensified and costs reduced, which is important to facilitate access to these therapies.
The viral vector biomanufacturing market
A growing market
In recent years, the number of viral-vector-based gene therapies approved and in development has increased, with 10 products having received marketing authorisation and 1,229 products in development in the active pipeline. This increased number of approvals and ongoing clinical trials related to viral vector-based gene therapy has led to a clear increase in demand for large-scale viral vector manufacturing.
This dynamic has been reflected in the number of investments and deals by CDMOs and biotech and pharmaceutical companies, either to obtain new production capacity or to acquire existing capacity. In addition to CDMOs and in-house production sites, hospitals and university centres are also building capacity to meet Phase I/II production needs.
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Figure 5: In vivo gene therapies based on approved viral vectors. Source: GlobalData 2024, MabDesign.
in vivo gene therapy pipeline
Figure 6: In vivo gene therapy products based on viral vectors in development. Breakdown by development phase. Source: GlobalData 2024, MabDesign.
The landscape of CDMO players has thus expanded, and by 2022 there will be more than 50 players in Europe and the USA. The market can be defined as concentrated, with 5 players holding 40% of the total available capacity: Catalent, FujiFilm Dyosinth, Lonza, Thermo Fisher Scientific and WuXi Advanced Therapies. These CDMOs have existed for many years and have often developed in the field of gene therapy following acquisitions, such as that of US company Brammer Bio by Thermo Fisher in 2019. With an estimated CAGR of between 10 and 20% between 2024 and 2030 in the gene therapy field, further mergers or acquisitions of companies in the near/medium term are to be expected.
Figure 7: CDMOs active in viral vector production (non-exhaustive list). Sources: adapted from A Lamproye 2023, MabDesign.
Changing business models in biomanufacturing
The biomanufacturing offer available for viral vectors remains a challenge. In vivo gene therapy, where the final product is the viral vector injected directly into the patient, mainly uses AAV, which makes this vector particularly sought-after. On the other hand, volumetric capacities do not always meet the need: the batch sizes required for in vivo gene therapies are increasingly large as their application extends beyond the therapeutic area of rare diseases.
Strategic divergences are emerging, leaving the way open for larger companies with sufficient financial resources to acquire the capacity to internalize production (e.g. Cellectis). This option also enables them to limit the risk of supply shortages and retain their intellectual property. Smaller companies that want to bring their product to market but do not have enough resources to invest in production capacity will turn to outsourcing, sometimes facing long lead times. Nevertheless, speed to market is essential for gene therapy products, which can often benefit from fast-track designations (or orphan drug status, for example) that reduce drug development time, putting even more pressure on CDMOs.
Alternative structures offer a response to the growing challenges of industrialising gene therapies, such as Shared Manufacturing Organizations (SMOs) or modular ‘plug-and-play’ facilities. The SMO concept is based on the idea of pooling production capacity between several players. Rather than building dedicated plants, biopharmaceutical companies, including start-ups and biotech companies, can access shared facilities where production is tailor-made for different customers. Modular plug-and-play facilities represent another key innovation in the production of biomedicines. These facilities are designed to be rapidly deployed, flexible and modular, enabling companies to adapt to production needs in the shortest possible time.
In September of this year, Sanofi inaugurated Modulus, a plant capable of adapting to manufacture up to 4 vaccines or biopharmaceuticals simultaneously, and of reconfiguring itself in a matter of days or weeks to change technological platforms (live attenuated viral vaccines, recombinant protein vaccines or messenger RNA vaccines, as well as biotechnology-derived treatments such as enzymes or monoclonal antibodies).
INITS has invested near Montpellier to create the INITS-SMO (Shared Manufacturing Organization) plant, which is designed to provide biotechnology companies with premises that meet GMP pharmaceutical standards, enabling them to produce their own batches of innovative drug candidates for preclinical and clinical trials.
Perspectives & Conclusion
Approaches such as Artificial Intelligence (AI) and Industry 4.0 may offer an interesting potential for transformation to meet the challenges of viral vector production mentioned above.
In particular, AI-based algorithms could provide more predictive quality control, using machine learning models to reduce production failures. Industry 4.0, which integrates AI directly into its overall system, makes it possible to create digital twins providing simulations of the process with the aim of optimising it. In 2023, Généthon and Thalès have announced a collaboration to develop a digital model that will use Artificial Intelligence to model bioprocessing steps and optimise yields.
75% of in vivo gene therapies currently being developed use viral vectors, which include retroviruses, lentiviruses, adenoviruses and adeno-associated viruses. Thanks to their high infectivity, viral vectors are often very effective for gene transfection. However, there are concerns about their clinical safety due to their tendency to induce immune responses and mutations caused by transgene insertion. As an alternative, non-viral vectors are increasingly being developed in the wake of the COVID19 crisis and with the emergence of mRNA vaccines. In particular, lipid nanoparticles (LNPs) or cationic polymers, which have demonstrated robust gene loading capacity, high safety and practicality. They thus offer interesting potential for gene delivery.
With technological advances and a growing understanding of virus biology, the prospects for development and innovation in the production of viral vectors promise to improve their efficacy, safety and personalisation, paving the way for increasingly targeted and accessible gene therapies and vaccines for a wide range of diseases.
This article is an extract from our Bioprocess watch “USP and DSP for viral vectors” published in December 2024. You can download it in full by clicking on the link below:
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