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

Original text

An identity test should be performed that demonstrates that all of the intended pneumococcal polysaccharide serotypes and carrier protein(s) are present in the final product, unless this test has been performed on the final bulk. A serological test, using antibodies specific for the purified polysaccharide may be used.

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Original text

The requirement to test lots of pneumococcal conjugate vaccine for unexpected toxicity (abnormal toxicity) should be agreed with the NRA. Such a test may be omitted for routine lot release once consistency of production has been well established to the satisfaction of the NRA and when good manufacturing practice is in place.

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Original text

Methods that have been used to separate unbound polysaccharide before assay, and that are potentially applicable to pneumococcal conjugates, WHO Technical Report Series No. 977, 2013 WHO Expert Committee on Biological Standardization Sixtieth report include hydrophobic chromatography, acid precipitation, precipitation with carrier protein-specific antibodies, gel filtration and ultrafiltration. The amount of unbound polysaccharide can be determined by specific chemical or immunological tests, or by HPAEC after hydrolysis.

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Original text

The composition of the purified outer-membrane complex should be determine by SDS_PAGE or a similar method and shown to be consistent from lot to lot, have not more than 8% LPS by weight, and pass the rabbit pyrogenicity test when injected into rabbits in amounts of 0.25 ug/kg of body mass

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 bulk conjugate should be tested for the absence of specific toxicity of the carrier protein where appropriate. Anbsence of specific toxicity of the carreir protein may also be assessed through validation of the production process

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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.

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An immunological test, using antibodies specific for the purified polysaccahride, may be used

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The vaccine in the final container should be tested for pyrogenic activity by intravenous injection into rabbits or by a LAL test.

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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

Each final lot should be tested for unexpected toxicity (abnormal toxicity) using a test approved by the national control authority.
This test may be omitted for routine lot release once consistency of production has been well established to the satisfaction of the national control authority and when good manufacturing practices are in place.

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Original text

The endotoxin content of the purified polysaccharide should be determined and shown to be within limits agreed by the national control authority in order to ensure that any pyrogenic activity of the final product is acceptable.
Less that 10IU of endotoxin per ug of polysaccharide when measured by a LAL test can be achieved. Alternatively, polysaccharide preparations should pass the rabbit pyrogenicity test when injected into rabbits in amounts of 1.0ug or purified polysaccharide per kg.

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

Manufacturers are encouraged to avoid the use of materials of animal origin
wherever possible.

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