Recent advances in biotechnology have resulted in the development of two new categories of products—cell therapy products and gene therapy products. Cell therapy products contain living mammalian cells as one of their active ingredients, while gene therapy products contain pieces of nucleic acid, usually deoxyribonucleic acid (DNA), as one of their active ingredients. Some products combine both categories, resulting in a therapy that uses cells that express a new gene product. Both cell and gene therapy products may be combined with synthetic or natural biomaterials to form tissue constructs.

For the purpose of this chapter, cell and gene therapy products include any product that has live cells or pieces of nucleic acid, however formulated. The following are excluded at this time: (1) tissue-based products in which the cells are removed or killed, (2) enhanced reproductive methods such as in vitro fertilization, (3) non-mammalian cell-based products, (4) traditional vaccines such as live attenuated virus, and (5) noncell, nongene products manufactured by using cells or recombinant-DNA (rDNA) technology, which are discussed under Biotechnology-Derived Articles 1045.1

Cell therapy products are products with live cells that replace, augment, or modify the function of a patient's cells that are diseased, dysfunctional, or missing. Transplantation of bone marrow to replace marrow that has been destroyed by chemotherapy and radiation is an example of a cell-therapy product. These products are also referred to as somatic cell therapy products because nongerm-line cells are used in the product. In addition, cells may be combined with biomaterials. For example, dermal or epidermal cells can be grown on a collagen substrate to produce a sheet of cells for wound and burn therapy. Examples of cell therapy products are shown in Table 1.

Table 1. Examples of Cell Therapy Products

There are three sources of donor cells for cell therapy products: (1) the patient's own cells (autologous cell products), (2) the cells from another human being (allogeneic cell products), and (3) the cells derived from animals such as pigs, primates, or cows (xenogeneic cell products). Autologous cells are not rejected by the patient, but they are not available for many treatments because they are missing, dysfunctional, or diseased. In those situations allogeneic and xenogeneic cells can be used. The advantage of allogeneic cells is that they do not trigger a rejection reaction as strong as that caused by xenogeneic cells. Xenogeneic cells are used when human cells with the desired characteristics are not available or the supply of human donors is too limited. Cell therapy products are sometimes encapsulated in a device that prevents the patient's cells and antibodies from killing the xenogeneic cells. However, the use of xenogeneic cells has the potential to cause zoonoses in humans. Much research is focused on identifying and propagating stem cells, regardless of the source, because stem cells can be manipulated to differentiate either during manufacturing or after administration. Cell lines are preferable to freshly isolated cells because they can be tested extensively for viruses, tumorigenicity, and other features. They also ensure a constant and reproducible product by minimizing donor-to-donor variability.

Cell therapy products can be modified by treatment with DNA or another nucleic acid so that the pattern of gene expression is changed. This new product, a combination of gene therapy and cell therapy, is referred to as an ex vivo gene therapy product. Typically, cells are taken from the patient and modified outside of the body before they are returned to the patient.

Cell therapy products face several unique manufacturing challenges that are addressed in other sections of this chapter. First, cells cannot be terminally sterilized or filtered so removal or inactivation of microorganisms or viruses without killing the cells is problematic. Second, every raw material used in manufacturing has the potential of remaining associated with the cells. Qualification and sourcing of all raw materials is critical to producing a safe and effective product. Third, storage of the cell therapy products may present a challenge. Freezing is the main mode of long-term storage, and some cell therapy products cannot be frozen without changing their basic characteristics, especially those for differentiated functions. These types of products may have to be administered to patients within hours, or days at the most, after completion of the manufacturing process. Fourth, there is often an urgent clinical need to administer a product as soon as possible. Fifth, some products consist of a batch size equivalent to one dose, very often in a small volume. For these last three challenges, traditional analytical methods, especially those for sterility, mycoplasma, and potency, are not always applicable because these methods are not rapid or they are not amenable to small volumes. Even when these traditional methods are performed, the results are not available in time for products requiring rapid release. These products are often released on the basis of the results of new, very rapid or small-volume methods. Currently there are no compendial standards for such methods, although, as stated in the General Notices and 21 CFR 610.9, alternative methods to compendial tests are permissible, provided they are shown to be equivalent. As such new methodologies become properly validated, they will be included in the compendia.

Gene therapy products are products in which nucleic acids are used to modify the genetic material of cells. A retroviral vector used to deliver the gene for factor IX to cells of patients with hemophilia B is an example of a gene therapy product. Gene therapy products can be broadly classified on the basis of their delivery system. Means for delivering gene therapy products include viral vectors (viruses with the genes of interest but usually without the mechanism to self-replicate in vivo), nucleic acids in a simple formulation (naked DNA), or nucleic acids formulated with agents, such as liposomes, that enhance their ability to penetrate the cell. Some types of gene therapy block the expression of a gene by the administration of antisense oligonucleotides, which are complementary to a naturally occurring ribonucleic acid (RNA) and block its expression. Most of the initial clinical work has been done using viral vectors. The choice of a gene vector is complex (see Design Considerations for Gene Vectors under Manufacturing of Gene Therapy Products). The most common viruses used to date include murine retroviruses, human adenovirus, and human adeno-associated viruses (AAVs). Antisense-oligonucleotide products are in clinical development and on the market. Examples of gene therapy products are shown in Table 2.

Table 2. Examples of Gene Therapy Products

Although manufacturing of vectors or nucleic acids can be analogous to that used for rDNA products or vaccines, there are some unique challenges. Analytical methodologies for vectors (see Analytical Methodologies) are still being developed. Methods for quantitating viral vector particles and determining the number of particles that are active (potent) are important areas that are rapidly evolving. Traditional assays for viral dose, such as the plaque assay or the tissue culture infectious dose assay, detect a fraction of the active vector particles. The precision of such assays is about a factor of three (half a log). Further, manufacture of large batches of viral vectors with no, or minimal amounts, of replication-competent viruses (RCVs) is challenging. Detecting a small number of RCV particles in the presence of large amounts of replication-defective vector is difficult. As in cell therapy products, sourcing of raw materials is critical. Removal of adventitious agents or other process contaminants from viral vectors can be impossible. Even defining purity is an issue for enveloped viral vectors, such as retroviruses or herpes viruses, as they incorporate cellular proteins in their envelope when they bud from the cell. This makes it difficult to determine if contaminating extraviral cellular proteins have been adequately removed.

For gene therapy vectors administered directly to patients, there are safety concerns related to the fate of their nucleic acids. For example, alteration of germ-line DNA is undesirable. Integration of gene therapy products into somatic cell DNA carries a theoretical risk of insertional mutation, which could result in modified gene expression and deregulation of the cell. For viral gene therapies, patients may need to be monitored for the presence of RCV. To address the risks associated with specific products, preclinical studies, quality control (QC) testing, and patient monitoring strategies need to be developed in accordance with the applicable regulations and guidance documents. The methodologies used to support these activities, including polymerase chain reaction (PCR) methods, are amenable to compendial standardization.

Clinical uses for cell and gene therapy products, their manufacturing processes, and analytical schemes for determining identity, dose, potency, purity, and safety are rapidly evolving and are as diverse as the products themselves. This informational chapter summarizes the issues and best current practices in the manufacturing, testing, and administration of cell and gene therapy products. Usually informational chapters focus on materials that are already commercially available. This chapter, however, not only discusses products for commercial applications, but it also addresses the production of clinical trial materials and the other unregulated uses, such as bone marrow transplantation. When different approaches can be used for clinical trial material as compared to those used for commercial product, it is so stated.

The traditional compendial perspective is to develop public standards that can be applied to a particular final product without expressly defining production details. Efforts have been made in this chapter to specify when traditional methodologies or standards can be adapted. Novel methodologies applicable to cell and gene therapies are also highlighted. As these new methodologies become properly validated, they will be included in subsequent publications.

This chapter is extensive because of the diverse nature of the products and the special considerations that they require. Manufacturing has been divided into three sections. The first section, Manufacturing Overview, discusses general aspects of manufacturing and process development. The other two manufacturing sections are Manufacturing of Cell Therapy Products and Manufacturing of Gene Therapy Products. The latter section includes a subsection on designing gene vectors. On-Site Preparation and Administration follows the manufacturing sections because the handling of these products at the clinic often requires facilities and expertise not found in a typical hospital. Storage, shipping, and labeling issues are addressed under Storage and Shipping and under Labeling. Regulations, Standards, and New Methodologies summarizes existing guidelines and highlights the need for the development and validation of new methodologies to assess product quality. The final sections of this chapter, Definition of Terms and Abbreviations, list and define the terms and abbreviations referred to in this chapter and those commonly employed in this field.