Regulatory considerations for messenger RNA vaccines: A comprehensive review on World Health Organization guidelines on development, production, and clinical evaluation

Abstract

The messenger RNA (mRNA) vaccines have revolutionized vaccine development by enabling faster production compared to traditional methods. Comprehensive regulatory requirements for mRNA vaccines are required to ensure uniform quality, safety, and efficacy in this rapidly growing technology. Globally accepted World Health Organization (WHO) guidelines provide broad, adaptable recommendations to all national regulatory authorities, whereas others, such as the European Medicines Agencies (EMA’s) draft guideline, provide European Union specific legally binding, highly detailed technical requirements such as impurity thresholds, validated functional testing, and strict stability protocols. The distinctions reflect WHO’s function as a worldwide standard-setter against EMA’s regional enforcement responsibilities. For India, as one of the world’s major vaccine producers and a significant supplier to numerous countries and UN agencies, relying exclusively on WHO’s generalized framework may be insufficient. India presents unique issues such as varying population needs, disease burdens, and cold chain infrastructure limits. Developing India-specific mRNA vaccination rules that are aligned with WHO and incorporate analytical detail from EMA would boost domestic innovation, facilitate global export acceptance, and assure swift, safe responses to future public health catastrophes.

INTRODUCTION

A vaccine is a biological preparation that offers active acquired immunity to a specific infectious illness, according to the World Health Organization (WHO). Usually, it contains a substance that mimics a pathogen, like a virus or bacteria, that triggers the body’s immune system to identify and combat the pathogen should it be encountered again. The pathogen’s toxins, one of its surface proteins, or weakened or dead versions can all be used to create vaccines.[1]

How does a vaccine work

When a disease-causing organism enters our body, it triggers two responses:

• The symptoms (like fever, nausea, rash, etc.) we first experience

• The immune system’s response, which becomes stronger over time and helps fight the infection.

  • Recognition

  • Producing antibodies

  • Builds adaptive immune system

  • Memory creation

  • Immune system training from vaccines is safe and does not cause illness. By halting the onset of disease, they shield us for years, if not our lives.[2]

Types of vaccine

There are numerous techniques to synthesize vaccines, which determines their classification. Figure 1, depicts the illustration of various types of vaccines.

Figure 1.

Illustration of various types of vaccines.[1] (a) live-attenuated, (b)whole inactivated, (c) split inactivated, (d) recombinant viral vector, (e) recombinant bacterial vectors, (f) virus-like particles, (g) DNA or RNA (h) Recombinant subunits and (i) Synthetic peptides

Live attenuated vaccines

In this type of vaccines, the pathogens are genetically weakened to reduce their growth and hence do not cause disease in the host.

Examples: Vaccines against measles, mumps, rubella, and varicella.

Inactivated or dead vaccines

The bacterium that causes the disease is either destroyed or rendered inactive, typically by a chemical (formalin, etc.) or thermal (high temperature) treatment. When given, these vaccines produce a strong immune response that closely resembles the majority of attacks.

Examples: Vaccines against typhoid, influenza, salk polio, and hepatitis A

Acellular or subunit vaccines

The entire virus or bacteria is not present in acellular vaccines. Rather, they contain proteins or polysaccharides from the bacterial or viral surface, which are the components that our immune system targets since it perceives them as “foreign.” Acellular vaccines come in a variety of forms:

1. Toxoid vaccine

2. Conjugate vaccine

3. Recombinant vaccine

4. DNA/RNA vaccine: messenger RNA (mRNA) vaccine is an example of this category.[1]

While most studies explain WHO’s mRNA vaccine guidelines, this paper goes a step further by assessing their regulatory significance, comparing them to other important frameworks such as the European Medicines Agencies (EMA’s), and identifying gaps pertinent to India. It underlines the importance of a harmonized but India-specific regulatory pathway, balancing global standards with national requirements, to improve compliance, facilitate exports, and handle the unique issues encountered by one of the world’s largest vaccine producers.

MESSENGER RNA VACCINES

The application of mRNA vaccines is among the most recent and fascinating developments in vaccine technology. Using the genetic code of the pathogen, mRNA vaccines can be produced rapidly, in contrast to traditional vaccines, which can take months or even years to produce. “mRNA is probably going to be at least one of the vaccines of the future.”

An mRNA vaccine does not cause illness; instead, the RNA material instructs our body on how to produce a certain protein that is specific to the virus. Antibodies that identify the protein are produced as part of the immunological response that the protein initiates. That way, if a person is ever exposed to that virus in the future, the body would like have the tools (antibodies) to fight against it. Figure 2, portrays the working of mRNA vaccines.

Figure 2.

How messenger RNA vaccines work[3]

Advantages. mRNA vaccines can be prepared quickly and having high degree of adaptability We might be able to modify the formulation’s mRNA to target a different antigen and produce a large amount of superior vaccine material really fast. The COVID-19 vaccine from Pfizer-BioNTech is one example.[3] Figure 3, explains about the pros and cons of mRNA vaccines.[4]

Figure 3.

Pros versus cons of messenger RNA vaccines.[6] mRNA: Messenger RNA

Synthetic messenger RNA types

Three categories of synthetic mRNAs are available for therapeutic use:

• Non-replicating mRNA (nr RNA) is a regular mRNA that has been modified to function better and be safer

• Self-amplifying mRNA (sa RNA) produces copies of itself in the body, requiring only a tiny amount. It’s large and difficult to transport, therefore it can be divided into two sections to aid

• Circular mRNA (circ RNA) forms a loop, making it more stable and long-lasting in the body.[5]

How messenger RNA vaccines differ from traditional vaccines

Both conventional and mRNA vaccines have benefits and drawbacks. Table 1 lists some of the main distinctions between the vaccines.

Table 1.

Messenger RNA versus traditional vaccines; key differences[4,5,6,7,8,9,10]

Specifications	mRNA vaccines	Traditional vaccines
Development timeline	Fast to develop and produce	Slower, takes years
Effectiveness and safety	Generally safe and effective for most people	Highly effective but may not be safe for everyone
Shelf life	Up to 6 months	1–3 years
How long immunity lasts	May need boosters	Some offer long-term immunity, others need boosters
How it works	Gives body instructions to make virus protein	Uses parts of the virus (weakened or inactivated)
Advantages	Fast production, reduces risk of infection and severe illness	Proven, may offer lifelong immunity
Risks	Side effects: pain, fatigue, headache, allergic reactions	Side effects: pain, fever, may not be safe for all
CMC controls	Advanced analytics: purity, sequence, integrity	Standard QC, antigen testing
Nonclinical data	Includes biodistribution, genetic safety	Routine toxicology
Postmarket surveillance	Enhanced real-time monitoring	Routine pharmacovigilance

mRNA=Messenger RNA, CMC=Chemistry, manufacturing, and controls, QC=Quality control

Challenges faced by messenger RNA vaccines

mRNA vaccine regulation aligns with traditional vaccines but faces challenges from novel technology, proprietary methods, and the need for locally adapted WHO guidelines. Figure 4: Details about challenges of mRNA vaccines

Figure 4.

Outlines the challenges of messenger RNA vaccines[11]

NEED OF SEPARATE GUIDELINES FOR MESSENGER RNA VACCINES

Compared to conventional vaccines that directly deliver the antigen (such as an inactivated virus or protein), this is very different. Regulators must evaluate particular hazards such as mRNA stability, cellular absorption, and nanoparticle behavior – areas not addressed by conventional vaccine frameworks – due to this essentially distinct process.

Additional non – Clinical safety and biodistribution studies

Because mRNA enter cells and can spread throughout the body, regulators need non-clinical studies that mimic gene therapy evaluations, like biodistribution, genotoxicity, and prolonged toxicology. In order to handle new safety issues for mRNA products, these are required and not typical for the majority of vaccine types.

Enhanced post-market surveillance and pharmacovigilance

Rare significant side effects, such as anaphylaxis and myocarditis, were discovered during the rapid development of mRNA COVID 19 vaccines. Regulatory agencies now employ rigorous real-time safety monitoring, long-term follow-up procedures, and adaptable risk management updates as a result, which goes beyond the standard post-market monitoring that is customary for conventional vaccines.

Classification challenges and evolving legal frameworks

Initially governed by traditional vaccine regulations rather than gene therapy laws, mRNA vaccines employ synthesized nucleic acids. Because of this discrepancy, authorities (such as the Food and Drug Administration, EMA, and WHO) created new scientific and legal guidelines especially for mRNA prophylactic vaccines, which resulted in developing classification schemes and temporary recommendations.[12]

REGULATORY CONSIDERATIONS OF MESSENGER RNA VACCINES

In 1990, it was shown that transient protein expression may be induced in vivo by direct injection of in vitro generated mRNA. Since then, mRNA has become a key platform for vaccine research due to developments in mRNA stability, decreased immunogenicity, and scalable manufacturing.

Without substantially changing the manufacturing process or quality standards, this platform enables quick changes to the antigen-coding sequence. Nevertheless, additional production, testing, and regulatory monitoring adjustments can be necessary if modifications impact crucial product attributes or cellular reactions.

World Health Organization guidelines for messenger RNA vaccines

This document outlines key regulatory considerations for mRNA vaccine development. As novel platforms, mRNA vaccines require phase-appropriate data and a comprehensive approach to address their unique characteristics.

General considerations of the messenger RNA vaccines

A mRNA vaccine’s intended usage should specify the ailment, population, target pathogen, and antigen (s). Crucial elements of the examination encompass:

• The rationale behind antigen selection and any genetic alterations is quite evident.

• Description in detail of the replicon genes in self-amplifying mRNA, if any

• Details of the complete formulation, including safety data, quality controls, production procedures, and LNPs

• If necessary, details about the dosage, administration method, any equipment or diluent used, and compatibility.

Special considerations of the messenger RNA vaccines

mRNA vaccines are made using linear DNA from polymerase chain reaction or plasmids. The mRNA includes coding regions, UTRs, a 5′ cap, and poly (A) tail for stability and translation. It can be modified to reduce inflammation or boost immune response.

Manufacture and control of messenger RNA vaccines

To guarantee quality, safety, and effectiveness, rigorous monitoring, characterization, and release testing of mRNA vaccines are crucial from a regulatory standpoint. WHO good manufacturing practice standards for biological products suggest evaluating both essential and non-critical quality features using a risk-based approach with in-process controls for every batch. Every material used in the manufacturing needs to have its identification and quality confirmed. Post-approval modifications must adhere to International Council for Harmonization (ICH) lifecycle management criteria, and clinical trial applications should contain adequate risk-related information.

Process development and in-process controls

The manufacturing processes should be validated to show that they meet their important and key parameters and produce a product that consistently satisfies the predetermined quality criteria. Evidence of the consistent and repeatable removal of contaminants associated to the process and product to levels suitable for human consumption should be part of this.

Product characterization

In order to help the manufacturer steer the development and improvement of their process and analytical tests, characterization refers to studies and analyses that are not regularly conducted on every lot but that give them valuable information about the structure, performance, and safety of their product.

It is anticipated that a toxicological risk assessment will be conducted for impurities and residuals that have known or possible harmful effects.

Nonclinical evaluation of messenger RNA vaccines

The intended clinical usage should be taken into consideration when evaluating prospective mRNA vaccines in a nonclinical setting. The following WHO guidelines should serve as the foundation for the planning, execution, and analysis of nonclinical research, including the selection of relevant studies pertaining to the product’s “pharmacology” (in the case of vaccines, immunogenicity, and proof-of-concept) and toxicology.

1. WHO guidelines on nonclinical evaluation of vaccines; and

2. WHO Guidelines on the nonclinical evaluation of vaccine adjuvants and adjuvanted vaccines.

Clinical evaluation of messenger RNA vaccines

Clinical studies should follow the guidelines outlined in the WHO Guidelines on Clinical Evaluation of Vaccines: Regulatory Expectations and the WHO Guidelines for Good Clinical Practice for Pharmaceutical Product Trials.

As with other vaccine types, safety, immunogenicity, and efficacy data should be recorded in clinical trials. However, the potential issues listed below should be given special attention because they may be more pertinent to mRNA vaccines than to other vaccine types that may already have a license and that regulators may be more accustomed to.[13]

The overview of European medicines agency messenger RNA vaccine guidelines

The European Union (EU) adopted its own mRNA vaccine regulations to ensure a robust, harmonized framework tailored to the unique scientific and technical aspects of mRNA technology. Unlike traditional vaccines, mRNA vaccines involve novel manufacturing processes, delivery systems, and quality control challenges that require specific regulatory oversight. By establishing clear, enforceable guidelines, the EU aims to guarantee vaccine safety, efficacy, and quality for its population, support rapid and reliable vaccine development and approval, and maintain public trust. Additionally, these regulations help align manufacturers’ practices across member states, facilitating consistent supply and enabling quick response during public health emergencies. Table 2 enlists some of the main distinctions among WHO[12] and EMA mRNA Vaccine Guidelines.[14]

Table 2.

Regulatory differences: World Health Organization versus European Medicines Agency messenger RNA vaccine guidelines[13,14]

Category	WHO (2021)	EMA draft (2025)
Legal status	Nonbinding, advisory for global NRAs	Legally enforceable in accordance with Directive 2001/83/EC, an EU regulatory framework
Analytical detail	High-level, adaptable guidance	Specific, comprehensive analytical requirements (for example, ELISA for dsRNA and RT-PCR for identification)
Starting materials definition	General GMP control; less prescriptive	Clearly outlines the testing expectations for the initial materials (such as the DNA template, nucleotides, and 5’ cap reagents)
Impurity control	Requires risk-based reasoning and control	Requires dsRNA measurement, established thresholds, and confirmed orthogonal testing
Stability studies	General requirements for cold chain and shelf life	Stability studies with explanations are necessary for all suggested storage conditions
Container closure system	Briefly mentioned	Complete validation and specific needs are listed
Testing for functionality	Adaptable and encouraged depending on the situation	It is required that in vitro expression or translation be proven and managed
Reference standards	Encouraged, not mandatory	Required for key attributes (identity, integrity, dsRNA) aligned with ICH Q2(R2)
Emergency use pathways	Addresses accelerated pathways for public health emergencies	Not addressed in the draft guideline
Impurity thresholds	Based on risk assessment and general standards	Must be defined, tested, and justified with process validation data

EMA=European Medicines Agency, WHO=World Health Organization, NRAs=National Regulatory Authorities, EU=European Union, ELISA=Enzyme-Linked Immunosorbent Assay, RT-PCR=Reverse transcriptase-polymerase chain reaction, GMP=Good manufacturing practices, ICH=International Council for Harmonization

4.3 Need of Indian regulatory framework for mRNA vaccines: Table 3 comprises the aspects explaining the need of Indian Regulatory framework for mRNA

Table 3.

Rationale for developing India’s National messenger RNA vaccine guidelines[13]

Aspect	Rationale
1. WHO guidance is generalized	WHO guidelines are intended for global usage, but India, as a key manufacturer, need specific, enforceable, and context-sensitive norms
2. Local legal and regulatory needs	WHO guidelines are not legally enforceable; India must incorporate them into national legislation consistent with the Drugs and Cosmetics Act of 1940 and Rules of 1945
3. Support for indigenous innovation	Tailored national guidelines can help Indian mRNA developers like Genova Biopharma and Bharat Biotech by aligning R and D capacity and clinical infrastructure
4.Export assurance	A national mRNA guideline enhances credibility with global regulators (e.g., EMA, USFDA, UNICEF) and facilitates export acceptance under SNRAs
5.Customization to national needs	India faces unique challenges: population diversity, logistics (cold chain limitations), and disease burden. A local guideline enables adaptation of WHO principles to Indian realities
6. Pandemic preparedness	enables pre-approved regulatory channels (e.g., EUA, platform switching, variant updates) for speedier outbreak response
7. Harmonization with global standards	Follows WHO/ICH/EMA guidelines while establishing India-specific thresholds, testing standards, and review schedules
8. Reduce regulatory ambiguity	Codified standards eliminate uncertainty, resulting in clarity, consistency, and quick approvals

WHO=World Health Organization, EMA=European Medicines Agency, ICH=International Council for Harmonization, mRNA=Messenger RNA, SNRAs=Strict National Regulatory Regimes, EUA=Emergency use authorization, USFDA=U.S. Food and Drug Administration, UNICEF=United Nations Children’s Fund

CONCLUSION

mRNA vaccines represent a transformative leap in immunization, offering unmatched speed, adaptability, and scalability in responding to infectious diseases. While WHO provides high-level, adaptable guidelines for global use, the EMA enforces more prescriptive, binding standards with detailed technical requirements, such as impurity thresholds, stability protocols, and validated functional testing. For India, one of the world’s largest vaccine producers and a trusted supplier to numerous countries and UN agencies, relying solely on WHO’s broad framework is inadequate. India’s diverse population, high disease burden, logistical constraints, and need for rapid outbreak response demand a harmonized, India-specific regulatory framework. Such guidelines should align with WHO’s global principles while integrating EMA’s analytical rigor to ensure vaccine safety, quality, and efficacy. This approach would strengthen domestic innovation, facilitate global export approvals, enhance pandemic preparedness, and safeguard public trust through transparent, evidence-based communication.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.