Innovative cell and gene therapies could hold the keys to treating and potentially curing a wide range of genetic and acquired diseases. Among the arsenal of tools available to researchers and biopharmaceutical experts, lentiviral vectors have long been recognized as a versatile platform for delivering genetic material into target cells. However, as the field progresses and therapies move closer to clinical applications, addressing and overcoming common pitfalls associated with lentiviral cell and gene therapies becomes crucial.
Confidence in the therapeutic potential of lentiviruses hinges on accurately quantifying and monitoring the viral load during production and throughout the therapy process. To meet this demand for precision and sensitivity, many experts turn to Droplet Digital™ PCR (ddPCR™) technology. By harnessing the power and absolute quantification of ddPCR technology, cell and gene therapy manufacturers can confidently navigate the intricacies of lentiviral cell and gene therapies, bringing us closer to a future where these groundbreaking treatments become a reality for countless individuals in need.
Understanding Your Technology Options
When it comes to quantifying nucleic acids in lentiviral cell and gene therapies, choosing the right technology is essential. Among the options available, the ddPCR workflow is high-quality and reliable data. Unlike traditional PCR methods that rely on amplification and fluorescence signal detection, ddPCR technology partitions the sample into 20,000 individual reactions within water-in-oil droplets. This partitioning enables absolute quantification of the target, providing accurate measurements of target concentrations without the need for standard curves.
Compared to other digital PCR technologies, ddPCR technology offers several advantages that make it particularly well-suited for lentiviral cell and gene therapies. Firstly, the high number of partitions generated by ddPCR technology ensures robust and statistically significant data, minimizing the impact of low-abundance targets or sample heterogeneity. Secondly, the droplet-based format of ddPCR assays allows for higher tolerance to inhibitors commonly found in biological samples, ensuring accurate quantification even in complex matrices. These features make ddPCR technology a reliable and precise tool for cell and gene therapy manufacturers seeking the sensitivity and precision required to navigate the challenges associated with lentiviral cell and gene therapies.
Do You Really Know Your Lentiviral Titer?
The precise determination of infectious viral titer is a critical factor in lentiviral cell and gene therapies, directly influencing treatment outcomes and patient safety. Inaccurate viral titer determination can lead to detrimental effects, including under-dosing or over-dosing of patients, potentially compromising the efficacy of treatments or causing adverse reactions.
Biotechnology experts are already leveraging ddPCR technology to measure lentiviral vectors infectious titers.. For example, scientists from Biogen published a study in Human Gene Therapy Methods describing their ddPCR method and reporting its superior performance compared to qPCR and fluorescence-activated cell sorting (FACS). Similarly, Lonza scientists have presented their development of high-throughput ddPCR assays for lentivirus infectious titer determination, saying they chose ddPCR technology because it “has emerged as a reliable, cutting-edge technology to quantify the absolute copy number of any gene of interest without using a standard curve.”
Getting Your Lentiviral Vector Copy Number Right
In addition to viral titer, cell and gene therapy developers need to know how efficiently their lentiviral vector integrates the gene of interest into the target cells. Failed transgene integration can offer no therapeutic effect, while vector copy numbers that are too high can result in overly potent and potentially dangerous products. In 2020, NIH researchers demonstrated the utility of ddPCR assays for assessing the average number of lentiviral vectors integrated into multiple types of engineered T cell therapies. In 2022, another group of researchers enabled greater industry standardization by developing lentiviral standards for determining vector copy number assay sensitivity, subsequently using these standards “to show the superior precision of Droplet Digital PCR over quantitative PCR for vector copy number determination.”
Realizing the Promise of Lentiviral Cell and Gene Therapies
In summary, the adoption of ddPCR technology has revolutionized the landscape of cell and gene therapy development and manufacturing. Its versatility as a powerful tool for accurate quantitation of lentiviral titer, vector copy number, and much more has profound implications for improving the safety and efficacy of these advanced therapies. From the early stages of development to the scale-up of commercial manufacturing, ddPCR assays provide scientists and manufacturers with the sensitivity and precision needed to navigate the challenges of cell and gene therapy production confidently. As the field continues to evolve, ddPCR technology remains at the forefront, driving advancements and enabling breakthroughs in the realm of personalized medicine.