PCR-based analytics of gene therapies using adeno-associated virus vectors: Considerations for cGMP method...

Area of Science:

• Molecular Biology
• Biotechnology
• Pharmaceutical Sciences

Background:

• Gene therapy has advanced significantly, enabling treatments for numerous genetic diseases.
• Analytical methods are crucial for ensuring the safety, efficacy, and stability of gene therapy products during manufacturing.
• Nucleic acid-based testing is vital for enumeration and identification assays of active pharmaceutical ingredients.

Purpose of the Study:

• To review method development and validation for nucleic acid-based testing of gene therapies.
• To focus on adeno-associated virus (AAV) vectors and their co-transfection factors.
• To discuss improvements in analytical testing, including next-generation sequencing and artificial intelligence.

Main Methods:

• Survey of nucleic acid-based testing methodologies for gene therapies.
• Comparison of quantitative PCR and droplet digital technologies for gene therapy assessment.
• Description of validation parameters for adeno-associated virus (AAV) genome titers.

Main Results:

• Nucleic acid-based methods are reliable and rapid but require consideration of unique biological, technological, and regulatory aspects of gene therapies.
• Quantitative PCR and droplet digital technologies offer different advantages for gene therapy analysis.
• Specific validation parameters for AAV genome titers illustrate the scope of analytical development.

Conclusions:

• Method development and validation are critical for the successful manufacturing of gene therapies.
• Continuous innovation in analytical testing, such as next-generation sequencing and AI, is needed.
• This review serves as an introduction for new researchers and a clarification for industry professionals.