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Gene therapy has come a long way over a rocky road in the 30 years since the first patients received treatment. Now, the market is rapidly picking up steam, and hundreds of gene therapy clinical trials are in progress. Exciting as that sounds, successful clinical trials represent only half the battle. Without robust production and manufacturing processes, even the most effective therapy will remain out of reach of patients.

As new gene therapies make their way through the approval process, companies are scrambling to develop manufacturing practices capable of producing high volumes of clinical grade product while keeping costs down. To meet demand worldwide, manufacturers will need custom platforms designed with gene therapy in mind. Existing equipment and techniques developed for other biological products, such as mAbs or vaccines, aren’t good enough.

Making the transition from laboratory scale to commercial production won’t be as easy as just scaling up systems already in place. Protocols that work well to produce gene therapy vectors for research just aren’t geared toward producing the consistency and quality needed for clinical manufacturing.

“Now we are in this really exciting phase, where we need to take technology that has been developed in a lab setting and industrialize the process, even though it hasn’t really been thought about for large scale,” said Mark Szczypka, PhD, senior director, viral vector & gene therapy technologies at Pall Biotech. “Most laboratory processes are not very efficient to make large viral vector numbers, so now you have to figure out how to take it to the next level to treat the patient population in a cost-effective manner.”

Gene therapy relies on viral vectors to infect the patient’s cells and deliver a healthy, functional copy of a gene into the body. Different gene therapy applications require different vectors, each with its own growth and purification requirements, meaning a one-size-fits-all manufacturing platform won’t likely suit everyone’s needs.

“Customers do not have a lot of choices if they want to purify their viral vectors,” said Catherine Allioux, PhD, global product manager for chromatography at Pall. “There are not a lot of tools specifically developed for gene therapy,”

The two most commonly used vectors, adeno-associated viruses (AAVs) and lentiviruses (LV), each require very different handling. Lentiviruses are more fragile than AAVs, and conditions ideal for purifying AAVs can destroy the infectivity of LVs, rendering them useless.

Downstream purification is particularly important for clinical production. Contaminants that could affect performance, or stimulate an immune response in the patient, must be removed. For instance, host cell DNA and proteins are released when the cells break open to release the virus, and that cell debris gets filtered out. Empty, broken, or partially filled viral capsids also arise during viral synthesis. These incompletely-assembled virus particles ramp up immunogenicity without conferring any therapeutic benefit, so it’s important to get rid of them during the purification process. This is tricky, because empty capsids and therapy-bearing viruses look a lot alike on the outside. The centrifugation method used to separate them in the laboratory isn’t practical to scale up. However, differences in charge allow the unwanted capsid particles to be removed by ion exchange chromatography.

Many current chromatography techniques are expensive, low yield, and can reduce the infectivity of the virus. Conventional packed bed chromatography methods don’t lend themselves to large scale production. Pall Biotech’s Mustang® Q single-use membrane chromatography capsule, provides a scalable, cost effective option for viral particle purification.

“It is just a plug and play technology, the opposite of the classical chromatography,” said Allioux. “The membrane comes ready to use, reducing the labor involved in setting up and running the system.”  Ion-exchange chromatography is very versatile, and works with multiple types of viral vectors. Mustang Q membrane can be adapted for the very different chemistries of these vectors. Not only can the system efficiently separate empty capsids from full in various different serotypes of AAVs, but it’s also proven effective at purifying the more delicate LVs without diminishing infectivity.

“Mustang Q membrane is already used in a very large scale for recombinant proteins and also mAbs,” said Allioux. She pointed to Pall’s long history of optimizing membrane chromatography in these other arenas. “This is experience we can bring to gene therapy processing,” she said.

“We know that people developing gene therapy processes now face so many hurdles and issues to solve upstream that they would like to have their life simplified in downstream,” Allioux said. “They have a well proven technology with Mustang capsules.”

Indeed, researchers trying to get new gene therapies off the ground face an uphill battle to optimize large scale processes both for upstream viral production and downstream purification. Single-use technologies have begun to catch on as a way to simplify some of these issues. These systems eliminate expensive, time-consuming cleaning steps, and they are often easier to set up than their multi-use counterparts. They also lend themselves well to automation, which improves speed and lowers costs.

“The reason we promote single-use is because when you use traditional methods, such as stainless steel, you have to do cleaning validation every time you change out your line,” Szczypka pointed out. “It’s costly and it takes up time.”

Mustang Q chromatography capsules are designed to fit in well with a completely closed single-use production system. “Pall is developing an end-to-end platform solution that can allow you to optimize your conditions, and have everything in a single-use system that is entirely closed,” said Szczypka. “In a nutshell, what we’re trying to do is streamline the overall process, simplify it, and automate it.”

“It’s a complete integrated solution for customers that want to develop gene therapy manufacturing processes,” he said. “Our goal is to be the leader in helping bring gene therapy to patients and we’re well on our way.”

 

To learn more about Mustang chromatography capsules for gene therapy, download full application note here: www.pall.com/genetherapy

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