Review of animal use requirements in WHO biologics guidelines – database of suggested guideline revisions

Original text

The endotoxin content of each lot of purified Vi polysaccharide should be determined and shown to be within limits agreed with the NRA. Suitable in vitro methods include the Limulus amoebocyte lysate (LAL) test.

New text

The need for pyrogenicity testing should be assessed during the manufacturing development process and be re-evaluated following any significant changes in the production process or relevant reported production inconsistencies that may influence pyrogenicity. A risk-based approach should be implemented which is suitable to the manufacturing process and the product depending on the potential presence of endotoxins and non-endotoxin pyrogens. The endotoxin content of the final product should be determined using a suitable in vitro assay, such as the recombinant factor C (rFC) or limulus/tachypleus amoebocyte lysate (LAL/TAL) tests. The rFC method is strongly recommended due to concerns over the impact on the sustainability of limulus stocks. The endotoxin content should be consistent with levels found to be acceptable in final product lots used in clinical trials and within the limits agreed upon with the NRA. A monocyte activation test (MAT) may be used for pyrogen testing after a product-specific validation. The use of the rabbit pyrogen test should be avoided due to its inherent variability, high retesting rates, and interspecies differences in pyrogenic responses as compared to humans.

Original text

The purified bulk conjugate should be tested to confirm the absence of toxicity specific to the carrier protein where appropriate (for example, when DT or TT is used as the carrier protein). Alternatively, the absence of specific toxicity of the carrier protein may be demonstrated at the purified carrier protein stage if agreed with the NRA.

New text

The carrier protein should be tested for the absence of specific toxicity of the carrier protein where appropriate (e.g. when tetanus or diphtheria toxoids have been used). Absence of specific toxicity of the carrier protein may also be assessed through validation of the production process. Where available and appropriately validated and with approval from the NRA, an in vitro test should be used. If in vitro assays are scientifically justified as unavailable and inappropriate, a suitable in vivo test may be used.

Original text

The endotoxin content of each lot of purified Vi polysaccharide should be determined and shown to be within limits agreed with the NRA. Suitable in vitro methods include the Limulus amoebocyte lysate (LAL) test.

New text

The need for pyrogenicity testing should be assessed during the manufacturing development process and be re-evaluated following any significant changes in the production process or relevant reported production inconsistencies that may influence pyrogenicity. A risk-based approach should be implemented which is suitable to the manufacturing process and the product depending on the potential presence of endotoxins and non-endotoxin pyrogens. The endotoxin content of the final product should be determined using a suitable in vitro assay, such as the recombinant factor C (rFC) or limulus/tachypleus amoebocyte lysate (LAL/TAL) tests. The rFC method is strongly recommended due to concerns over the impact on the sustainability of limulus stocks. The endotoxin content should be consistent with levels found to be acceptable in final product lots used in clinical trials and within the limits agreed upon with the NRA. A monocyte activation test (MAT) may be used for pyrogen testing after a product-specific validation. The use of the rabbit pyrogen test should be avoided due to its inherent variability, high retesting rates, and interspecies differences in pyrogenic responses as compared to humans.

Original text

The endotoxin content of the final product should be determined using a suitable in vitro assay such as a LAL test. The endotoxin content should be consistent with levels found to be acceptable in vaccine lots used in clinical trials and within the limits agreed with the NRA.
The need for pyrogenicity testing should be determined during the manufacturing development process. It should also be evaluated following any changes in the production process or relevant reported production inconsistencies that could influence the quality of the product with regard to its pyrogenicity. When required, the monocyte activation test (MAT) or rabbit pyrogenicity test may be used for monitoring potential pyrogenic activity subject to the agreement of the NRA.

New text

The need for pyrogenicity testing should be assessed during the manufacturing development process and be re-evaluated following any significant changes in the production process or relevant reported production inconsistencies that may influence pyrogenicity. A risk-based approach should be implemented which is suitable to the manufacturing process and the product depending on the potential presence of endotoxins and non-endotoxin pyrogens. The endotoxin content of the final product should be determined using a suitable in vitro assay, such as the recombinant factor C (rFC) or limulus/tachypleus amoebocyte lysate (LAL/TAL) tests. The rFC method is strongly recommended due to concerns over the impact on the sustainability of limulus stocks. The endotoxin content should be consistent with levels found to be acceptable in final product lots used in clinical trials and within the limits agreed upon with the NRA. A monocyte activation test (MAT) may be used for pyrogen testing after a product-specific validation. The use of the rabbit pyrogen test should be avoided due to its inherent variability, high retesting rates, and interspecies differences in pyrogenic responses as compared to humans.

Original text

Method used: ______________________________________________________________
Strain and type of animals: ___________________________________________________
Number of animals:_________________________________________________________
Route of injection:__________________________________________________________
Volume of injection: ________________________________________________________
Quantity of protein injected: __________________________________________________
Date of start of test: _________________________________________________________
Date of end of test: _________________________________________________________
Specification: _____________________________________________________________
Result: ___________________________________________________________________

New text

Original text

Although there have been advances in the use of animal models, no ideal animal model exists that establishes direct serological or immunological correlates of clinical protection. In the absence of such a model, it is important to ensure consistency of production using modern physical, chemical and immuno-based quality control methods as described in Part A of these WHO Recommendations. Additionally, any changes in critical quality attributes should be assessed for their impact on immunogenicity. Once the physicochemical tests are validated, these non-animal methods are considered more appropriate for use in lot release processes than animal models.

New text

Original text

Manufacturers and regulators should also take note of the decision of the Committee in 2018 to discontinue the inclusion of the general safety (innocuity) test in routine lot release testing requirements for all vaccines in WHO Recommendations, Guidelines and other guidance documents for biological products (3). This test is therefore not included in the manufacturing recommendations provided in Part A of the current document.

New text

Original text

The mouse weight-gain test currently in use to monitor the toxicity of vaccine lots is considered to be insufficiently sensitive and of questionable relevance. A more relevant and validated test should be sought.

New text

The mouse weight-gain test currently in use to monitor the toxicity of vaccine lots is considered to be insufficiently sensitive and of questionable relevance. A more relevant and validated test should be sought. The potential use of the Y-1 adrenal cell assay for cholera toxin as a more specific test for residual toxicity should be investigated. Such a specific test could be used on a-lot-to-lot basis or to validate the production process.

Original text

NATs may be used as an alternative to microbiological methods for culturing mycobacteria or to the in vivo guinea-pig test for the detection of mycobacteria after they have been validated and approved by the NRA

New text

A test for the absence of virulent mycobacteria should be performed. Where available and appropriately validated, an in vitro test should be used (for example a validated nucleic acid amplification test or culture method). If in vitro assays are not available or appropriate, a suitable compendial in vivo test may be used.

Original text

Each virus master seed and working seed lot should be tested in cell cultures and in animals for adventitious viruses relevant to the passage history of the seed virus. For virus grown in hamster or its cultured cells, the neutralized virus should be tested for adventitious agents by inoculating it on to cultures of human cells, mouse cells, simian cells, mosquito cells (e.g. C6/36), baby hamster kidney (BHK)-21 cells and PHK cells.

New text

A strategy for testing adventitious viruses in vaccines must be developed based on a risk assessment. Relevant culture methods and/or specific molecular biology or broad molecular methods should be part of the overall testing package with the agreement of the NRA. In vivo tests may only be used if the risk assessment indicates that this test provides an additional risk mitigation taking into account the overall testing package.