SC IV S. Raw Materials of Biological Origin for the Production of Cell-based and Gene Therapy Medicinal Products

(Ph. Eur. general texts 5.2.12)

This general chapter is published for information.

It contains sections on the quality requirements of raw materials used for the production of cell-based and gene therapy medicinal products for human use. The provisions of the chapter do not exclude the use of different production and control methods. It is the responsibility of the manufacturer of a raw material to qualify (prove to be suitable for the intended use) the raw material in accordance with the requirements given in this general chapter.

However, it is ultimately the responsibility of the user of a raw material to ensure it is of suitable quality for the specific use.

The quality of the raw materials may be considered according to the stage of development of the cell-based or gene therapy medicinal product, thereby acknowledging the inherent evolution of the quality profile of the product during its pharmaceutical and clinical development. Nevertheless, patient safety needs to be ensured in early phase clinical development. The aim is to have an appropriate qualification strategy for the raw materials when used for the production of cell-based/gene therapy medicinal products. It should be noted that changes in raw materials during the lifecycle of the cell-based/gene therapy medicinal product may affect the quality of the medicinal product and thus require additional studies to demonstrate comparability.

The impact of the raw material on the quality, safety and efficacy of the cell-based/gene therapy medicinal product is evaluated using a risk-based approach. Raw materials are used in order to consistently yield an active substance or medicinal product of a specified quality in terms of, for example, biological activity, purity/impurity profile, the risk of adventitious agents (bacteria, viruses, etc.) and stability.

From a risk perspective, the use of raw materials free from human or animal substances is preferred.

The biological nature of a raw material used for the production of cell-based/gene therapy medicinal products places special requirements on its quality. Examples of the critical quality attributes specific to each class of raw material are given in this general chapter.

1 SCOPE

This general chapter applies to raw materials of biological origin used for the production of cell-based/gene therapy medicinal products. The raw materials used in the manufacture of active substances are not intended to form part of the active substance. The raw materials can be extracted from various biological sources or produced by recombinant DNA technology.

This general chapter applies to the following classes of raw materials:

— sera and serum replacements;

— proteins produced by recombinant DNA technology such as growth factors, cytokines, hormones, enzymes and monoclonal antibodies;

— proteins extracted from biological material such as enzymes and polyclonal antibodies;

— vectors.

The principles of this general chapter may also be applied to other classes of biological raw materials where appropriate.

Medical devices, plastics and chemically synthesised raw materials, such as basal media (purely composed of chemicals), synthetic peptides or synthetic polynucleotides, are not within the scope of this general chapter.

2 RISK ASSESSMENT

Evaluation of the impact of the raw material on the quality, safety and efficacy of cell-based/gene therapy medicinal products must be performed by the user of the raw material. No single measure or combination of measures can guarantee the quality, functionality and safety of a raw material for its intended use. Therefore, a risk assessment must consider the biological origin and traceability of the raw material, the production steps applied to it and the ability of the drug product manufacturing process to control or remove the raw material from the final medicinal product.

Any risk factor must be evaluated in relation to the clinical benefit/risk of the cell-based or gene therapy medicinal product. When evaluating the risk posed by the raw material to the final medicinal product, the exposure of a patient to residual amounts of raw material with potential harmful effects (e.g. adverse immune reactions) should be considered in relation to the clinical benefit/risk of the cell-based or gene therapy medicinal product.

3 GENERAL REQUIREMENTS

3-1 ORIGIN

The origin of the raw material and if relevant any biological substances used for the production of the raw material must be known. Special attention must be paid to risks related to the sourcing (including pooling) of the substances used for the production of the raw material. Depending on the source of the raw material and the substances used in its production, raw materials can be divided into 3 categories:

1) raw materials of human or animal origin;

2) raw materials produced using substances of human or animal origin;

3) raw materials free from substances of human or animal origin.

Traceability of all raw materials is required, with particular attention to those materials with an inherent safety concern i.e. those of human or animal origin.

Due to the inherent risk of transmitting adventitious agents, it is recommended to minimise, wherever possible, the use of raw materials of human or animal origin. If such raw materials are required for the production of cell-based/gene therapy medicinal products, appropriate measures are taken to minimise the risks of transmitting adventitious agents such as viruses, prions, bacteria and protozoa.

For human blood and tissue-derived materials, only carefully evaluated donors who have been adequately tested for infectious transmissible agents may be used. These materials comply with appropriate EU and/or national legislation applicable to transplantation and transfusion. Traceability measures enable each donation to be followed from the donation to the raw material and to the final product, and vice-versa.

When raw materials of animal origin are used, these animals fulfil specific health requirements and should be fit for human consumption and reared under controlled conditions, when applicable. If the origin of the animals is not fully traceable (e.g. animals collected from the wild), information on their geographic location at the time of sourcing should be considered.

When vectors or proteins produced by recombinant DNA technology are used as raw materials, traceability to the master cell bank/virus seed lot is required.

For all raw materials of human or animal origin, or raw materials produced using substances of human or animal origin, a viral risk assessment is performed according to the requirements of general chapter 5.1.7. Viral safety. The extent of viral safety testing is dependent on the results of the initial risk assessment. In addition, a risk assessment with respect to transmissible spongiform encephalopathies is carried out and suitable measures are taken to minimise such risks as described in general chapter 5.2.8. Minimising the risk of transmitting animal spongiform encephalopathy agents via human and veterinary medicinal products.

3-2 PRODUCTION

All raw materials are produced within a suitable quality management system and production facilities.

Suitable in-process controls are in place to ensure that the production process is under control and consistently produces raw materials of defined quality.

Quality attributes for raw materials include identity, purity and biological activity where applicable, and they are to be demonstrated using appropriate, qualified control methods. Relevant specifications in terms of identity, purity/impurity profile and assays are to be established.

The production process is optimised to consistently minimise and/or remove adventitious agents and harmful impurities, whilst retaining the quality of the raw material. This can be achieved using one or a combination of the following measures:

— using validated inactivation/removal procedures such as gamma sterilisation or low pH during chromatography, where possible;

— demonstrating the ability of a production process to minimise, remove or inactivate adventitious agents or harmful impurities;

— testing for adventitious agents or harmful impurities.

A raw material is sterile and produced under aseptic conditions and/or subject to terminal sterilisation, unless otherwise justified. If the raw material is not sterile, the level of microbial contamination must be known.

Additives, such as stabilisers, may be added to the raw material. In cases where antibiotics and stabilisers of biological origin are used in the production of the raw material, their presence is justified and careful consideration is given to their selection, use, quality and concentration in the raw material, as well as their impact on the actual raw material itself.

3-3 GENERAL QUALITY REQUIREMENTS

Raw materials must meet pre-defined quality requirements for identity, purity and biological activity. In order to ensure the function of the raw material, it is subject to testing using appropriately qualified methods. The identity test must reflect the uniqueness of the raw material and distinguish it from other related or similar substances. Impurities include both process-related substances (e.g. in the case of recombinant proteins: host-cell-derived proteins (HCP), host-cell-derived DNA and vector-derived DNA (residual DNA), other biological or chemical substances) and product-related substances (e.g. aggregates and degradation products). The content of a raw material may be expressed either in absolute or relative terms. The assay for determination of biological activity may be used to establish the content.

3-3-1 IDENTIFICATION

The identity tests are specific for the particular raw material and address the molecular structure/composition or other relevant physico-chemical, biological or immunochemical properties. Methods used in the determination of biological activity and purity may also serve to identify the raw material. Identification may be carried out by comparison with a defined reference material or a representative batch of the raw material.

3-3-2 TESTS

Tests that may be applicable to raw materials include the following (see also the sections below for specific raw materials):

Appearance

Liquid or reconstituted freeze-dried raw materials comply with the limits defined for the particular raw material with regard to degree of opalescence (2.2.1) and degree of coloration (2.2.2).

Solubility

Freeze-dried raw materials dissolve completely in the prescribed volume of reconstituting liquid within a specified time, at a specified temperature, as defined for the particular raw material.

Osmolality (2.2.35)

Within the limits defined for the particular raw material.

pH (2.2.3)

Within the limits defined for the particular raw material.

Elemental impurities

Within the limits defined for the particular raw material.

Total protein (2.5.33)

Within the limits defined for the particular raw material.

Related substances

The content of product-related substances is within the limits defined for the particular raw material.

Microbiological control

Depending on the raw material concerned, it complies with the test for sterility (2.6.1) or the microbial contamination is determined (2.6.12).

Viral contaminants

Depending on the raw material concerned, relevant virus contamination is determined.

Bacterial endotoxins (2.6.14)

Less than the limit defined for the particular raw material.

Mycoplasmas (2.6.7)

Raw materials are free from mycoplasmas.

Stabiliser

Where applicable, it complies with the limits defined for the particular raw material.

Water (2.5.12)

Freeze-dried raw materials comply with the limits defined for the particular raw material.

3-3-3 ASSAY

Content

The content (e.g. protein content)/composition of the raw material is determined by an appropriate qualified method.

Biological activity

Where relevant, the biological activity is determined by a suitable assay. Where relevant (e.g. for enzymes), the biological activity is expressed per milligram of total protein (specific activity).

3-3-4 REFERENCE MATERIAL OR REFERENCE BATCH

An appropriate reference material or a representative batch of the raw material is used to perform the above-mentioned identification, tests and assay. Where available, the use of established reference standards, such as European Pharmacopoeia reference standards or WHO International Standards, is recommended.

3-4 STORAGE

The shelf life and storage conditions are defined.

3-5 LABELLING

The label states the expiry date, conditions for storage and use and any code that may be required for traceability including the biological origin of the raw material.

4 SERa AND SERUM REPLACEMENTS

4-1 DEFINITION

Sera from human or animal sources and serum replacements (including platelet lysates and other undefined growth additives, conditioned media, blood and other cellular components) are used as growth additives for cell culture. Sera and serum replacements used to promote cellular growth are typically complex biological mixtures, whose exact composition is not always possible to define. Due to this complex nature, special attention is given to verifying the consistency and performance of every batch.

Bovine serum

If bovine serum is used, it complies with the monograph Bovine Serum (2262).

Human serum and platelet lysates

Human serum and platelet lysates used as raw materials for the production of cell-based/gene therapy medicinal products are human blood-derived materials, which can originate from the recipient (autologous) or from another individual (allogeneic).

Conditioned media

Conditioned media, isolated and purified from cultured cell supernatant, may also be used to enhance cell proliferation due to various growth factors and cytokines secreted by the cells into the medium.

Other growth additives with undefined composition

Cell and/or tissue lysates may be used as growth additives.

Composite media

Composite media contain growth additives such as bovine serum, growth factors etc. The principles described in this section of the general chapter apply to individual ingredients of biological origin and/or biologically active ingredients of the composite media.

4-2 PRODUCTION

Due to potential differences in quality between batches of serum, cell or tissue lysate, suitable measures are implemented to verify the consistency of each batch before using them as raw materials for the production of cell-based/gene therapy medicinal products.

Because of the inherent risk of transmitting infectious agents from pooled plasma, pooled sera, or other derivatives from pooled allogeneic human blood or plasma, consideration is given to limit the number of donations which are pooled, unless sufficient methods for inactivation/removal of viruses are applied during production, where applicable.

For conditioned media, a cell bank system is preferred. The removal of the cells from the media must be ensured and potential impurities originating from these cells determined if possible.

4-3 IDENTIFICATION

It is recognised that the exact qualitative composition of sera and serum replacements may be difficult to determine. However, the approximate protein composition in both cases may be determined by, for example, protein electrophoresis. Where relevant, tests for total protein content or any chemical additives are performed. For human serum, the electrophoretic pattern corresponds to that of an appropriate serum reference batch. Alternatively, identity may be determined by comparison of albumin content with an appropriate serum reference batch. For serum replacements, the electrophoretic pattern or the use of markers secreted by cells/platelets may be used. Human origin is determined by a suitable immunochemical method (2.7.1), unless otherwise justified.

4-4 TESTS

See section 3-3-2.

Haemoglobin

Where relevant, within the limits defined for the particular raw material.

Cell-derived impurities

Where relevant, within the limits defined for the particular raw material.

Specific tests for viral contaminants

For bovine serum, the tests for viral contaminants specified in the monograph Bovine serum (2262) apply. For human serum, the tests for viral safety specified in the monograph Human plasma for fractionation (0853) apply.

4-5 ASSAY

The serum or serum replacement must show cell growth promoting properties that are within the limits defined for the particular raw material. More than one type of assay may be necessary to show suitability for the intended use.

5 PROTEINS PRODUCED BY RECOMBINANT DNA TECHNOLOGY

5-1 DEFINITION

Proteins and peptides produced by recombinant DNA technology, which are used as raw materials, include growth factors, cytokines, hormones, enzymes and monoclonal antibodies.

Growth factors, cytokines and hormones They are substances typically used for stimulation or inactivation, growth promotion or differentiation of cells in cell culture systems.

Other proteins Enzymes (e.g. collagenases), as raw materials, may be used for extraction of active substances from tissues and/or fluids. Other proteins (e.g. fibronectin) may be used as culture supports or media components.

Monoclonal antibodies Used as raw materials, they include immunoglobulins and fragments of an immunoglobulin with defined specificity. Antibodies can either be conjugated (chemically modified) or non-conjugated. Typical chemical modifications include fluorescent labelling and conjugation to magnetic beads. Antibodies, as raw materials, may be used for selection, activation/stimulation, isolation or purification of cells in cell culture.

5-2 PRODUCTION

Production of proteins using recombinant DNA technology is based on a well-characterised host-vector system, using a master cell bank and, if applicable, a working cell bank derived from the master cell bank. The expressed protein is extracted and purified using a variety of techniques, such as extraction, precipitation, centrifugation, concentration, filtration and/or chromatography.

During protein production using recombinant DNA technology, process-related impurities including residual host-cell or vector DNA and host-cell proteins must be reduced to acceptable levels. Particular attention must also be given to product-related impurities.

5-3 IDENTIFICATION

Identity is established by appropriate, qualified methods, such as electrophoresis (2.2.31), peptide mapping (2.2.55), isoelectric focusing (2.2.54) or liquid chromatography (2.2.29). For antibodies, identification is based on immunoglobulin class, isotype and/or specificity. In addition to the above-mentioned methods, immunochemical methods (2.7.1) and determination of activity are also considered suitable for identification.

5-4 TESTS

See section 3-3-2.

Host-cell-derived proteins and residual host-cell or vector DNA

Where relevant for the particular raw material, the content of residual host-cell or vector DNA and/or protein is determined using a suitable method unless the production process has been qualified to demonstrate suitable clearance. The content is within the limits defined for the particular raw material.

Related proteins

Related proteins (e.g. polyclonal antibodies with undefined specificities, glycoforms, degradation and oxidation products, oligomers and aggregates) are determined using liquid chromatography, electrophoretic or immunological methods and are within the limits defined for the particular raw material.

5-5 ASSAY

Content

The protein content is determined by an appropriate qualified method, for example by liquid chromatography (2.2.29) or UV spectrophotometry (2.2.25).

Biological activity

The biological activity of a recombinant protein is determined using, for example, cell proliferation, cell differentiation or an enzyme assay. Several acceptable bioassays may exist for a particular protein. For antibodies, cell-based immunoassays and assays based on ligand-binding and affinity may be used.

Where relevant, the biological activity is expressed per milligram of total protein (specific activity).

6 PROTEINS EXTRACTED FROM BIOLOGICAL MATERIAL

6-1 DEFINITION

Proteins extracted from biological material and used as raw materials include enzymes (e.g. porcine - derived trypsin and endonucleases), polyclonal antibodies, other proteins of biological origin (e.g. albumin and transferrin) and peptides of biological origin. They may be of human, animal, plant or microbiological origin.

Proteins extracted from biological material are used in a wide range of applications such as growth promotion, differentiation or purification of cultured cells and extraction of active substances from tissues and/or fluids.

6-2 PRODUCTION

Proteins are extracted from the blood or tissue of animals or humans, or from plant or microbiological sources using mechanical and/or chemical techniques. They are then subjected to further purification processes using a variety of techniques such as centrifugation, filtration, chromatography and concentration.

Polyclonal antibodies are produced by immunisation with a specific antigen, followed by purification. Antibody purification involves selective enrichment or specific isolation of antibodies from serum based on physicochemical fractionation, class-specific affinity and/or antigen-specific affinity.

During production of these proteins, process-related impurities, such as blood components, tissue fragments or contaminating proteins, must be reduced to acceptable levels. Particular attention is given to product-related impurities.

6-3 IDENTIFICATION

Identity is established by appropriate, qualified methods, such as electrophoresis (2.2.31), isoelectric focusing (2.2.54) peptide mapping (2.2.55), liquid chromatography (2.2.29) and immunochemical methods (2.7.1).

6-4 TESTS

See section 3-3-2.

Process-related impurities

Substances derived from the starting material (e.g. blood components, tissue fragments or contaminating proteins) are determined using suitable methods and are within the limits defined for the particular raw material.

Related proteins

Related proteins (e.g. antibodies with undefined specificity, degradation and oxidation products, oligomers and aggregates) are determined using suitable methods and are within the limits defined for the particular raw material.

6-5 ASSAY

Content

The protein content is determined using an appropriate qualified method, for example by liquid chromatography (2.2.29) or UV spectrophotometry (2.2.25).

Biological activity

Where relevant, the biological activity of a protein is determined using, for example, enzyme assays, immunoassays or assays based on cell proliferation/differentiation. For trypsin, the assay may be performed as described in the monograph Trypsin (0694).

Where relevant, the biological activity is expressed per milligram of total protein (specific activity).

7 VECTORS

Vectors that may be used as raw materials in the production of cell-based and gene therapy medicinal products include DNA vectors (e.g. plasmids, transposon vectors) as well as viral vectors and bacteria (e.g. modified Lactococcus species). Vectors are usually considered as starting materials, thus not under the scope of this general chapter. In cases where vectors are not considered as starting materials, such as vectors used as helper plasmids or helper viruses, the principles of this general chapter and the principles of production and quality control as outlined in general chapter 5.14.Gene transfer medicinal products for human use are to be followed.