Chapter-5 Quality Assurance of Vaccines

“Pharmaceutical Quality Management”
PHR-619
Chapter 5
“Quality Assurance of Vaccines”
(Part-1)
By: Dr. Amjad Khan

Assistant Professor, Department of Pharmacy, KUST
Topic: Quality Assurance of Vaccines
Outlines
➢ Introduction to Vaccines
➢ Composition of Vaccine
➢ Manufacturing Plan for Vaccines
➢ Stability of Vaccines
➢ Factors Affecting Stability of Vaccines

➢ Evaluation of Stability of Vaccines
Course: Pharmaceutical Quality Management, PHR-619-- Instructor: Dr. Amjad Khan, Assistant. Prof. Department of Pharmacy, KUST -- Email: dr.amjad@kust.edu.pk
Vaccine
➢ A vaccine is a biological preparation that improves immunity against a particular disease
➢ A vaccine typically contains an agent that resembles a disease-causing microorganism,

and is often made from weakened or killed forms of the microbe or its toxins.
➢ Vaccine was introduced by Edward Jenner in 1796.
Types of Vaccine
➢ Vaccines can be of various types, depending on their design and processes involved
in their manufacture. Vaccines for human use may contain;
o Whole killed or attenuated organisms (e.g., bacteria or viruses)
o Antigens derived from portions of a pathogen, either by;
▪ Partitioning and purification
▪ Derived using recombinant technology
Classification of Vaccines
Vaccines may be;
➢ Live attenuated whole cell or virus
➢ Inactivated/killed vaccines
❑ Whole cell or virus vaccine
❑ Recombinant proteins
❑ Subunit
▪ Polysaccharides
▪ Proteins
▪ Modified toxins
Regulation of Vaccines
➢ Vaccines are regulated by FDA as biological products.
➢ For the U.S., national requirements are codified in 21 CFR, the 200 and 600
sections, with additional recommendations available in FDA Points to
Consider and Guidance documents (www.fda.gov).
➢ International guidance is available from the International Conference on

Harmonization (ICH) (www.ich.org) and the World Health Organization
Components of Vaccine
Components of Vaccine (Antigens)

➢ Antigen is the main component of vaccine and may be;
➢ Whole Organism/ Virus
➢ Components/ Subunits
➢ Recombinant Proteins
➢ Different vaccine antigens are often combined in one final formulation

in order to elicit immunity against multiple diseases and to reduce the
number of separate administrations needed to achieve immunity to the
various vaccine antigens.
Components of Vaccine (Adjuvants)

➢ Adjuvants are agents incorporated into vaccine formulations to enhance the
immune responses generated by the vaccine antigens. Specifically, they can;
o Increase the amount of antibody produced
o Direct the immune response (Th1 or Th2)
o Increase the duration of antibody presence (persistence)
o Produce a combination of these effects.
➢ Aluminum compounds are the most widely used adjuvants
Components of Vaccine (Anti Microbial Preservative)

➢ Antimicrobial preservatives are added to inhibit the growth of microorganisms that may
be introduced from repeated puncture of multi dose vials.
➢ The microbial preservatives currently used in vaccines includes;
▪ Thimerosal
▪ 2-phenoxyethanol
▪ Benzethonium chloride
▪ Phenol
Components of Vaccine (Stabilizers)
➢ The primary purpose of stabilizers is to protect certain vaccines from adverse conditions
such as heat or to serve as a cryo-preservative during the lyophilization process, usually
the freezing step.
➢ The particular materials chosen for this purpose includes;

➢ Sugars (sucrose or lactose),
➢ Amino acids (glycine or glutamic acid [monosodium salt]),
➢ Glycerol
➢ Proteins (human serum albumin or gelatin)
Components of Vaccine (Manufacturing Residuals)

➢ Manufacturing residuals are the residual amounts of any of the
materials used in the manufacturing process.
➢ Manufacturing residuals are but not limited to;

➢ Pyrogenic substances
➢ Residual Moisture
Components of Vaccine (Cell Derived Residuals)
➢ Killed bacterial component vaccines may contain significant amount of cell-derived

residuals
➢ Common cellular residuals are

❑ Proteins,
❑ Nucleic acids,
❑ Polysaccharides.
➢ Assays for these components are routinely conducted, if appropriate, to ensure purity
Components of Vaccine (Inactivating Chemical Agents)

➢ Several chemical agents have been used to inactivate bacteria and viruses or to detoxify
toxins in vaccine production processes.
➢ Examples of inactivating chemical agents are;

❑ Formaldehyde
❑ b–propiolactone
❑ Glutaraldehyde
❑ Hydrogen peroxide
➢ Inactivating agent should be removed from the final product as thoroughly as possible. The
upper limit for formaldehyde is 0.02%, equivalent to 0.1 mg per 0.5 mL vaccine dose.
Manufacturing Plan for Vaccine

During manufacturing of vaccines, manufacturers need to consider the following factors:
• Physical facilities
• Raw materials and process aids
• Actual manufacturing process, including
a. initial process (production of virus/bacteria and recombinant materials);
b. downstream processes (purification or chemical modification, if applicable);
• Antigen modifications such as conjugation
• Storage of process intermediates and final bulk
• In-process and final product testing regimens and control schemes
• Addition of adjuvants, if applicable
• Formulation and filling
• Container–closure system
• Stability program that supports the dating period of the product
Stability of Vaccine
Vaccines are large and complex molecular assemblies that are highly
susceptible to environmental factors that may significantly affect their activity.
Factors affecting Stability of Vaccine
➢ Temperature is the main factor affecting stability of vaccines
➢ The impact of humidity is not relevant for the vast majority of vaccines due to;
❑ Their liquid formulation
❑ The protective nature of the packaging, with appropriate closure system
➢ Other environmental factors (e.g. light) might be considered
Evaluation of Stability of Vaccine
➢ The objectives of stability studies are to;
❑ Determine shelf-life and storage conditions, and to support licensing
❑ Monitor vaccine stability in the post-licensure period

❑ Support manufacturing changes by demonstrating comparability of product
manufactured by different processes
➢ Vaccine stability testing is based on the determination of the change in a vaccine property
which may be a direct or an indirect indicator of vaccine immunogenicity or efficacy.
➢ Vaccine products should be evaluated to;
❑ Define suitable conditions for storage

❑ To place appropriate time limits for the exposure to these conditions
➢ Evaluation of the stability of vaccine products has three general purposes;

❑First, the products are shown to maintain an acceptable analytical profile throughout
manufacture and use to preserve safety and effectiveness.
❑Second, stability studies across several product batches provide an effective way to
characterize the inherent properties of the product.
❑The third use, demonstrating manufacturing consistency in the product.
➢ The stability of vaccine products depends on the nature of a vaccine antigen, the
product formulation, and the control of vaccine storage prior to use.
➢ Vaccine products are evaluated with programs that include real-time long-term
storage under prescribed conditions
Protocol for Stability Study of Vaccine

➢ Design of vaccine stability studies should clearly indicate;
❑ Purpose of the study
❑ Analysis of the results
❑ Interpretation of the data
➢ The study protocol should include the stability assay format, as well as the number of and
intervals between stability study time points.
➢ The results of a vaccine stability study may either be subject to acceptance criteria, or may
undergo statistical analysis to estimate key vaccine stability characteristics.
Protocols for Stability Study of Vaccine

The overall experimental plan for evaluating the stability profile of a product or intermediate
batches includes;
❑ Specific definition of the storage conditions and relevancy of these conditions
❑ The length of time for which the samples will be stored at each condition
❑ Sample analysis time during time course of storage
❑ The analytical measurements at each time point
❑ Itemization of analytical procedure
Protocols for Stability Study of Vaccine

➢ For stability studies that occur early in product development, the studies may be
conducted to confirm the suitability of the product formulation and/or storage conditions.
➢ Later in development, stability studies are typically conducted to provide data supporting
product dating period or intermediate hold time, to provide more elaborate product
characterization, and to evaluate manufacturing consistency.
➢ Stability studies should be conducted over a duration sufficient to determine the point of
loss of acceptable potency or other relevant parameters.
Stability Testing (Stability Indicating Parameters and Test Frequency)

➢ Stability indicating parameters are selected on case by case basis
➢ For live attenuated vaccines, the titre is main stability-indicating parameter
➢ Other parameters which can affect efficacy and safety include;

❑ Antigen content,
❑ pH,
❑ Specific toxicity,
❑ Antimicrobial agent content,
❑ Sterility,
❑ Adjuvant content and degree of adsorption
❑ Changes in physicochemical properties
Stability Testing (Stability Indicating Parameters and Test Frequency)

➢ For non-live vaccines, potency will have to be studied on formulated vaccines
➢ Appropriate time points for testing should be chosen to take into account
❑ The characteristics of the vaccine in question
❑ The rate of change of the parameter measured
❑ The purpose of testing,
❑ Study design and subsequent data analysis.
Stability Testing of Vaccines (Intermediates)
“The unformulated active (immunogenic) substances that are

processed before final formulation and can be stored for long periods of time
before further processing”.
Stability Testing of Vaccines (Intermediates)

➢ Vaccine production processes involves production of intermediates like;
❑ Bulk purified antigens,
❑ Bulk adsorbed/ adjuvanted antigens
❑ Final bulks
➢ Intermediates are usually not processed immediately and storage periods of up to several
years are possible
➢ Stability testing should be performed at different stages of production, namely single

harvests, monovalent bulks, multivalent bulks and final bulks.
Analytical Measurements (Potency Testing)

➢ The potency assay is generally the key analytical result, predicting whether a
vaccine remains suitable for use and produce the expected clinical response. This
assay can take many forms, depending on individual vaccines.
❑ An infectivity assay for a live virus vaccine
❑ A measure of the proportion of conjugated polysaccharide for a polysaccharide

protein conjugate vaccine

➢ Other analytical measurements that have a link to the potency of the product are;
❑ Degradation profile
❑ Dissociation of a carrier protein from conjugated vaccines
❑ Dissociation of an adjuvant from an antigen complex
➢ Other tests related to physico-chemical characteristics includes;

❑ General safety
❑ Degree of aggregation
❑ pH of the product
❑ Moisture content
❑ Preservative
❑ Evaluation of Container and closure system
➢ Every effort should be made to use quantitative assays that result in a defined value
➢ Descriptive results reported only as pass or fail should not be used if the assay can
provide a defined value.
➢ Validation of the assays is another issue of critical importance for the stability
assessment of a vaccine.
Analytical Measurements (Cumulative age of Antigen in Final Product)

➢ Total age of all components at the end of shelf-life is considered as cumulative age
of the product.
➢ In practice, stability data on the final product should include the data generated on
the intermediates of different ages used in the final formulation.
➢ The stability of the characteristics of a final product should be guaranteed during

the whole shelf-life, irrespective of the age of the intermediates at the time they are
used in the production process.
Stability Testing (Vaccine Container and Closure System)

➢ Important for vaccines in liquid form
➢ The impact of the closure system on vaccine stability and quality in general should
be tested by exposing samples to and maintaining them in different positions during
a certain period of time.
➢ These positions should mimic possible situations that may occur during the
transport and storage of the vaccine and that provide contact between vaccine and
the closure system.
Stability Testing of Freeze Dried Vaccines
➢ Data to support proposed use of vaccine after reconstitution, maximum storage

period and storage conditions should be generated
➢ Residual moisture, reconstitution period and appearance of reconstituted vaccine

should be defined.
➢ The stability of a diluent should be tested
Stability Testing of Combined Vaccine

➢ Each vaccine component (after combination) should be tested to support initial
licensure of combined vaccines.
➢ Determination of the shelf-life of a combined vaccine should be based on the

shortest shelf-life component.
➢ Data generated on monovalent vaccines should support stability of a combined

vaccine but extrapolation of data is not allowed.
Labelling of Vaccines
➢ Label on vaccine container should be of suitable quality for the proposed storage and in
general should meet national requirements for labelling
➢ Label should clearly indicate;

❑ Recommended storage conditions
❑ Expiry date
➢ Instructions for protection from environmental factors (light or freezing) should be stated
➢ If vaccine vial monitors (VVM) are to be used, adequate stability data should be generated
to support selection of appropriate VVM for a specific vaccine.
References
1. United State Pharmacopeia (USP/NF), 2018
2. WHO Technical Report Series No. 962. World Health Organization, Geneva, 2011

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“Pharmaceutical Quality Management”

By: Dr. Amjad Khan

➢ Factors Affecting Stability of Vaccines

➢ A vaccine is a biological preparation that improves immunity against a particular disease

➢ A vaccine typically contains an agent that resembles a disease-causing microorganism,

➢ Vaccine was introduced by Edward Jenner in 1796.

o Antigens derived from portions of a pathogen, either by;

▪ Partitioning and purification

▪ Derived using recombinant technology

➢ Vaccines are regulated by FDA as biological products.

➢ International guidance is available from the International Conference on

Components of Vaccine (Antigens)

➢ Different vaccine antigens are often combined in one final formulation

Components of Vaccine (Adjuvants)

o Direct the immune response (Th1 or Th2)

o Increase the duration of antibody presence (persistence)

o Produce a combination of these effects.

➢ Aluminum compounds are the most widely used adjuvants

Components of Vaccine (Anti Microbial Preservative)

➢ The microbial preservatives currently used in vaccines includes;

Components of Vaccine (Stabilizers)

➢ The particular materials chosen for this purpose includes;

Components of Vaccine (Manufacturing Residuals)

➢ Manufacturing residuals are but not limited to;

Components of Vaccine (Cell Derived Residuals)

➢ Killed bacterial component vaccines may contain significant amount of cell-derived

➢ Common cellular residuals are

Components of Vaccine (Inactivating Chemical Agents)

➢ Examples of inactivating chemical agents are;

Manufacturing Plan for Vaccine

Factors affecting Stability of Vaccine

➢ Temperature is the main factor affecting stability of vaccines

❑ The protective nature of the packaging, with appropriate closure system

➢ Other environmental factors (e.g. light) might be considered

Evaluation of Stability of Vaccine

➢ The objectives of stability studies are to;

❑ Determine shelf-life and storage conditions, and to support licensing

❑ Monitor vaccine stability in the post-licensure period

Evaluation of Stability of Vaccine

➢ Vaccine products should be evaluated to;

❑ Define suitable conditions for storage

Evaluation of Stability of Vaccine

➢ Evaluation of the stability of vaccine products has three general purposes;

Evaluation of Stability of Vaccine

Protocol for Stability Study of Vaccine

❑ Analysis of the results

❑ Interpretation of the data

Protocols for Stability Study of Vaccine

❑ Specific definition of the storage conditions and relevancy of these conditions

❑ Sample analysis time during time course of storage

❑ The analytical measurements at each time point

❑ Itemization of analytical procedure

Protocols for Stability Study of Vaccine

Stability Testing (Stability Indicating Parameters and Test Frequency)

➢ For live attenuated vaccines, the titre is main stability-indicating parameter

➢ Other parameters which can affect efficacy and safety include;

Stability Testing (Stability Indicating Parameters and Test Frequency)

❑ The characteristics of the vaccine in question

❑ The rate of change of the parameter measured

❑ The purpose of testing,

❑ Study design and subsequent data analysis.

Stability Testing of Vaccines (Intermediates)