Intricate Supply Chain Complicates Gene Therapy Manufacturing

By Cathy Yarbrough, Contributing Editor
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This article originally appeared in Life Science Leader magazine.

After three decades of disappointments and clinical setbacks, gene therapy is finally delivering on its promise to alleviate – and possibly even cure – a long list of acquired and hereditary diseases. In 2017, the FDA approved three therapies that use vectors derived from harmless viruses to deliver functional copies of genes to patients’ cells. FDA Commissioner Scott Gottlieb, M.D., hailed gene therapy as “a whole new scientific paradigm for the treatment of serious diseases.”

The newly-approved gene therapies include two CAR T cell treatments. The third gene therapy to gain the FDA’s approval last year treats previously irreversible blindness in children and adults with inherited retinal disease (IRD). (Editor’s note: for more information about the three groundbreaking therapies, please see sidebar “FDA-Approved Gene Therapies.”)

Media coverage has understandably focused on the effectiveness and cost of gene therapies. Less coverage has been dedicated to the challenges of manufacturing these transformative treatments, particularly CAR T cell therapies.

“Manufacturing CAR T cell therapies requires a complex, multi-step process, and all along the way there are ancillary reagents, technology, logistics, and other pieces that further complicate the process,” said Bruce Levine, Ph.D., founding director of the Clinical Cell and Vaccine Production Facility (CVPF) at the University of Pennsylvania (UPenn).

“The complexity of manufacturing gene therapies is reflected in the prices that have been set for them,” added Levine. Under Levine’s direction, CVPF created the CAR T cellular product design that was licensed by Novartis to produce Kymriah, the first gene therapy approved by the FDA.

Unlike traditional small molecule and protein drugs, Kymriah and other gene therapies that are manufactured with patients’ cells are not mass-produced. They are instead individually manufactured on demand. “Each dose of CAR-T is tailored individually to, and manufactured for, each patient using the patient’s own blood cells via pioneering technology and a sophisticated process,” said David Lebwohl, M.D., senior VP and franchise global program head, CAR-T oncology global development at Novartis.

Because the manufacturing process for autologous gene therapies is so complicated, Kite Pharma staff members who manufacture Yescarta, the company’s FDA-approved CAR T cell therapy, must spend three months in hands-on training before they are allowed to “touch” even a bag of patient cells, said Timothy Moore, EVP, technical operations at Kite Pharma, a Gilead company.

Producing autologous gene therapies requires an entirely new approach to supply chain logistics as well as manufacturing. The cGMP-compliant supply chain for CAR-T and other gene therapies manufactured from patients’ cells is circular. “The patients are the beginning and end of the supply chain. Logistically, this is a very big challenge,” said Derek Adams, Ph.D., chief technology and manufacturing officer at bluebird bio, whose investigational CAR-T therapy for relapsed/refractory multiple myeloma is being developed in partnership with Celgene.

The CAR T cell therapy supply chain begins at the medical treatment center where the patient’s peripheral blood mononuclear cells (PBMCs), which include T cells, are extracted through the specialized blood filtration process leukapheresis. The patient’s PBMCs are cryopreserved for transport to the manufacturing plant that will use the cells to produce the patient’s CAR T cell therapy. Novartis has a 180,000-sq.-ft. manufacturing facility that specializes in CAR T cells in Morris Plains, N.J. Kite/Gilead, headquartered in California, manufactures its CAR T cell therapy at the company’s new 108,500-sq.-ft. manufacturing plant in El Segundo, CA, while bluebird bio, headquartered in Massachusetts, is building a 125,000-sq.-ft. gene therapy manufacturing plant in Durham, NC.

Prior to shipment to the manufacturing facility, the patient’s cells are electronically assigned a unique batch number, which enables the cells to be traced throughout the manufacturing process and supply chain.

MANUFACTURING BATCH FOR EACH PATIENT
After the patient’s PBMCs are delivered to the manufacturing plant, they are assigned to the staff responsible for transforming the cells into a CAR T cell therapy. After thawing and washing the PBMCs, staff members isolate and activate the T cells. The T cells are then incubated with the viral vector carrying genetic material encoding the CAR. The transduced T cells are reprogrammed to target cancer cells whose surfaces display a specific antigen. The cells are then multiplied, or expanded, in a culture system to increase to the quantity required for a single therapeutic dose.

Because CAR T cells must be manufactured independently from other patients’ cells, a separate manufacturing batch must be produced for each patient.

After quality and release testing are performed to ensure their safety and potency, the CAR T cells are cryopreserved for transport to the treatment center for administration by infusion to the patient. Prior to the infusion, the patient undergoes lymphodepleting chemotherapy to improve the newly infused CAR T cells’ survival and proliferative capacity.

The average turnaround time from the arrival of the patient’s PBMCs at the manufacturing plant to the delivery of the CAR-T therapy to the treatment center is 17 days for Kite/Gilead and 22 days for Novartis.

In 2018, Novartis plans to train 35 treatment facilities worldwide on collecting patients’ PBMCs and administering Kymriah; Kite/Gilead will train at least 60 treatment centers in 2018.

A major challenge in manufacturing CAR T cells is the high degree of variability in the starting material, said Levine. While well-characterized clonal cell lines are used to produce biologic drugs, the T cells used in the manufacture of CAR-T therapies often vary greatly from patient to patient as a result of disease severity and previous treatments. “For example, some types of chemotherapy can adversely affect the ability of T lymphocytes to divide in culture,” said Levine.

“We may get a low yield in the number or percentage of T cells in the shipment from the patient’s treatment facility,” Levine said. “Or we may get an adequate yield, but the T cells may not grow or transduce well in culture.” CVPF has established criteria for the minimum absolute T cell count that must be met for the center to manufacture CAR T cells for a patient.

Levine also has developed conditional manufacturing pathways to guide CVPF staff in manufacturing a CAR-T therapy from patient T cells with certain characteristics. Thus, the manufacturing approach is adjusted according to the state of each patient’s cells. “If the T cells have condition A, we will implement pathway 1. If it is condition B, pathway 2 will be followed, and so on,” he said.

As a result of these efforts, CVPF has “a good success rate in producing CAR T cells that surpass the criteria.”

Kite/Gilead also has developed a process to manage for the variations in the starting material. As a result, the company succeeded in developing CAR-T therapies from the T cells of 99 percent of the patients in the Yescarta clinical trials, said Moore. The high success rate is due to the manufacturing staff’s extensive training and the “well-defined steps in our robust manufacturing process.”

Unlike CAR T cell therapies, Spark Therapeutics’ Luxturna is not produced with patient cells.

An in vivo gene therapy, Luxturna consists of harmless recombinant adeno-associated virus (AAV) particles that serve as transduction vectors for the RPE65 gene, which encodes a protein necessary for vision. At selected treatment centers worldwide, retinal surgeons trained by Spark Therapeutics administer Luxturna by subretinal injection.

Spark Therapeutics manufactures each dose of Luxturna in a purpose–built, multisuite cGMP facility at the company’s 48,000-sq.-ft. headquarters in Philadelphia. To create the viral vector, scientists insert the RPE65 gene into a plasmid, a small circular DNA molecule that naturally occurs in bacterial cells. In the plasmid, the gene is flanked by AAV inverted terminal repeat sequences that are crucial for virus replication and packaging.

“Successfully integrating a functional gene into a viral genome is a combination of art and science,” said John Furey, COO at Sparks Therapeutics. “A lot of expertise and experience with viral vectors are required to produce a safe, effective gene therapy with the optimal yield and quality required for clinical and commercial manufacturing.”

Like Yescarta and Kymriah, Luxturna is cryopreserved prior to shipment to the patient’s treatment center. Spark Therapeutics has trained retinal surgeons at five treatment centers in the U.S. to administer Luxturna. During the next 12 months, U.S. treatment centers for Luxturna are expected to increase to between eight and 10, said Furey.

Novartis will be responsible for training retinal surgeons outside the U.S., as a result of Spark Therapeutics’ licensing agreement with the global pharmaceutical company. The agreement covers development, registration, and commercialization rights to Luxturna in markets outside the U.S. Spark Therapeutics retains exclusive rights for Luxturna in the U.S. and is responsible for obtaining EMA approval of the gene therapy. Under a separate agreement with Novartis, Spark Therapeutics will manufacture Luxturna for patients outside the U.S.

Unlike most companies with gene therapy products, Spark Therapeutics manufactures its clinical and commercial- grade viral vectors in-house. The consistent cGMP production of large quantities of pure, safe, and potent viral vectors is very time-consuming. Thus, Novartis and Kite/Gilead have turned to CDMOs to produce their lentiviral and retroviral vectors. Bluebird bio depends on four external partners for the manufacture of lentiviral vectors for the company’s four investigational gene therapies.

RELIABLE VIRAL VECTORS: SERIOUS CONCERN
However, securing dependable external sources of high-quality clinical-grade vectors can be challenging, particularly for small biotech companies with new R&D programs in gene therapy. The competition for reliable viral vector sources has become a serious concern in the risk-adverse biopharmaceutical industry.

Thus, bluebird bio plans to continue its CDMO partnerships even after it has obtained the capability of manufacturing viral vectors in-house at its manufacturing facility now under construction in North Carolina. Bluebird bio wants to ensure that a lentiviral vector shortage will never be a problem for the company, said Adams. Kite/Gilead similarly plans to continue using CDMOs after it begins to develop viral vector constructs in-house.

Spark Therapeutics’ in-house capability in manufacturing viral vectors is based on almost two decades of research to improve the safety and efficiency of gene transfer in specific target tissues. The research began at Children’s Hospital of Philadelphia (CHOP)’s Center for Cellular and Molecular Therapeutics (CCMT) under the leadership of Katherine High, M.D., today the president and head of research and development at Spark Therapeutics.

In addition to helping renew the life sciences industry’s interest in gene therapy, CCMT’s achievements enabled the 2013 founding of Spark Therapeutics as a spin-off of CHOP. The new company licensed several of CCMT’s assets, including its proprietary design and production of AAV-based vector product candidates. Furey said that Spark Therapeutics has the most comprehensive manufacturing process for producing AAV-based vector product candidates.

One of the challenges addressed by Spark Therapeutics is the lack of assays to determine whether a finished gene therapy product will perform as designed. To address this problem, the company identified and instituted multiple steps to control the manufacturing process so that Luxturna is made strictly and consistently in compliance. The company also created 40 release tests to evaluate each batch of the gene therapy according to predefined criteria for safety, purity, and potency.

Because they are living drugs, gene therapies are temperature sensitive and have limited lifespans. Therefore, Novartis, Kite/Gilead, Spark Therapeutics, and bluebird bio have enlisted dedicated courier services to deliver their newly produced living drugs to treatment centers. (Courier services also are used to transport patients’ cells from treatment centers to manufacturing facilities for CAR-T therapies and other gene therapies that are developed with patients’ cells.) During transit, courier services electronically monitor in real time the temperature and chain of identity and custody of the cells.

The chances that a patient’s cells are damaged or lost during the manufacturing process or the supply chain are minimal. However, Kite/Gilead has created a process to ensure that patients do not have to undergo second leukapheresis procedures in order to be treated with a CAR T cell therapy. After patients’ PBMCs arrive at the company’s CAR T cell manufacturing plant, staff set aside some of the cells, which are used to create backup bags of cells that are cryopreserved and stored at the facility in case they are needed.

The other challenges in gene therapy manufacturing include the scale-out that would be required to consistently manufacture therapies for diseases such as hemophilia that affect relatively large patient populations. (BioMarin Pharmaceuticals, headquartered in California, and Spark Therapeutics have investigational gene therapy products for hemophilia A and B, respectively.) In addition, there is a lack of standardized assays for evaluating the potency and safety of vectors. However, this challenge is being addressed by academic and industry experts in collaboration with the federal government’s National Institute of Standards and Technology.

One challenge that gene therapy developers have not had to address is the morale and dedication of manufacturing staff. “Our manufacturing facility is operated 24/7, even on holidays,” said Moore. Because a patient’s PBMCs were delivered to Kite/Gilead’s CAR-T manufacturing facility prior to Christmas week, a lot of staff had to work during the holiday. The staff members were willing to do so because they realized that the CAR T cells that they were helping to manufacture were destined for a critically ill patient whose survival could depend on the gene therapy. “Developing and manufacturing gene therapies takes personalized medicine to a new level,” said Moore.

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FDA-APPROVED GENE THERAPIES

In 2017, the FDA approved three autologous gene therapies, all of which are designed to be one-time treatments. Each therapy is under regulatory review in Europe.

• Kymriah*, the first gene therapy product to earn FDA approval was developed by Novartis for patients up to 25 years of age with refractory or relapsed B-cell precursor acute lymphoblastic leukemia (ALL). An overall remission rate of 83 percent characterized ALL patients who were treated with Kymriah, a CAR T cell therapy, in the late-stage clinical trial.
• Yescarta, the second gene therapy approved by the FDA, was developed by Kite Pharma, a Gilead Company, for patients with relapsed or refractory large B-cell lymphoma. In the company’s late-stage clinical trial of the therapy, the overall response rate was 72 percent. In addition, cancer could not be detected in 51 percent of the clinical trial participants treated with the CAR T cell therapy.
• Luxturna was developed by Spark Therapeutics for patients who are 12 months and older who have inherited a mutated RPE65 gene from each parent and as a result have a congenital retinal disease that leads to irreversible blindness. Luxturna, the first FDA-approved gene therapy for a genetic disease, packages a functional copy of the RPE65 gene in a benign genetically engineered viral vector. In the Phase 3 clinical study, statistically significant improvements in visual function occurred in treated patients.

*Kymriah is not the first gene therapy to be approved by regulators. In 2012, the EMA granted marketing authorization to uniQure, a biotech company headquartered in The Netherlands, for Glybera, a gene therapy for adult patients with familial lipoprotein lipase deficiency. UniQure did not seek the FDA’s authorization to market Glybera in the U.S. Because Glybera’s usage has been limited, uniQure announced in 2017 that it would not renew its EMA application to market the drug in Europe.