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Manufacturing and Testing a Clinical-Grade AAV Vector
Developing a cure for a rare disease: Cure AP-4 case study
Chris Edwards’ daughter was born in 2013 and began showing symptoms early on. Searching for a diagnosis, Chris and his wife were unable to find the root cause for the disease affecting their daughter. At age two, she walked with low muscle tone. At age three, she had an absence seizure. Doctors recommended genetic testing to determine a potential cause for her disease. At three and a half years of age, Chris’s daughter was diagnosed with Spastic Paraplegia 47 (SPG47), a subtype of a larger genetic disorder known as Adapter-Protein 4 Hereditary Spastic Paraplegia (AP-4 HSP).
In 2017, gene replacement therapy was suggested as a possible treatment, and strong efforts to push the development of a SPG47 therapy ensued. Mouse studies were initiated, as well as tests in non-human primates (NHPs). In 2023, proof of concept (POC) studies of a gene therapy for SPG47 was successful, but lack of strong financial opportunities was low, resulting in low interest from companies. Despite initial difficulties in funding, efforts to produce a viable AAV product initiated with the University of Sheffield. As of July 2023, clinical plasmid manufacturing with Charles River Laboratories is almost completed, with a human trial set for early 2024 with Boston Children’s hospital.
Ultra-rare diseases or orphan diseases like SPG47 are extremely low prevalence and have unique challenges due to their scarcity. Since they affect such a small population, research and development efforts for treatments and cures are often limited. There are over 7,000 rare diseases, affecting more than 400 million people worldwide. Less than 10% of those affected by rare diseases, including SPG47, have cures. To get a rare disease treatment from concept to clinic takes a dedicated team, including a manufacturing partner that understands time and resources can be limited.
Cell and gene therapy solutions
Gene therapy development follows several stages:
- Discovery: lead constructs are discovered and identified
- Preclinical testing: potency is evaluated in vitro and in vivo to confirm productivity of the potential therapeutic
- Toxicology testing: evaluates safety and biodistribution in vivo
- Clinical testing: safety, dose, efficacy, and benchmarking against current therapy is evaluated
- Commercialization: GMP-grade product is produced for an FDA-approved therapy
Gene therapy development: overcoming plasmid challenges with a platform manufacturing approach
Our gene therapy solutions operate with a phase-appropriate approach to manufacturing based on the stage of therapeutic development. Preclinical stages of development are supported with research grade plasmid DNA and non-GMP master cell banks (MCBs). Clients in Phases 1 or 2 of development have access to High-quality (HQ) plasmid DNA, as well as GMP MCBs for up to 30 patients’ worth of materials. Phase 3 development is supported with GMP grade plasmid DNA as well as GMP grade MCBs for up to 200 patients. For commercial-stage products, support for up to 100,000 patients worth of GMP-grade plasmid DNA and GMP MCBs is available.
Supporting the different gene therapy development stages: Cure AP4’s strategy
Charles River tailored the SPG47 therapeutic development process using a two-plasmid system, the first being an AAV backbone with the gene of interest, and the second being a helper AAV9 packaging plasmid. During the optimization of this process, two challenges were encountered. The first was the large packaging plasmid size, which is not standard. The second was a sequence discrepancy, which was identified prior to production. The plasmid had to be re-synthesized, and HQ plasmid production was rescheduled. Despite these challenges, the phase-appropriate approach allowed our team to appropriately identify and correct the problem, ensuring that the overall program timelines were kept on track. As a result, plasmid manufacturing was able to be completed on time.
Facilitating this timely plasmid manufacturing was our eXpDNA™ platform, a versatile plug and play toolbox for complex plasmids that provides extensive platform manufacturing processes, 100% in-house analytics, and over 30 years of experience in biologics testing.
Also essential to Cure AP4’s success was the consideration taken for vector manufacturing and testing. It’s critical that the starting materials, from plasmids to production cell lines, can support scaling and standardizing the manufacturing process. Charles River takes this into account, and provides a full range of gene therapy solutions, including AAV production through the nAAVigation® platform. This proprietary technology covers the AAV production needs of partners like Cure AP4, from preclinical R&D phase, through early clinical testing, to commercialization.