1. Introduction

Vaccines protect against millions of microbes and save thousands of lives from diseases each year ^[1]. Due to the use of extensive vaccines, the smallpox-causing virus (variola) has been eliminated and the cases of measles, polio, and various other childhood ailments have significantly declined throughout the world ^[2]. Traditional vaccine technologies including subunit vaccines and, live attenuated (or weakened) and inactivated (or killed) pathogens, offer long-lasting protection against various lethal ailments ^[3]. Despite this achievement, there exist key obstacles for the development of successful vaccines against diverse infectious disease-causing pathogens, particularly those better efficient to avoid adaptive immunity ^[1][4]. Besides, for the utmost evolving virus vaccines, the major difficulty is not the efficiency of established technologies but the demand for quick and large-scale production. Moreover, traditional vaccine technologies might be invalid for non-infectious disorders like cancer. Therefore, there is an urgent need for the progression of more versatile and effective vaccine platforms.

Nucleic acid-based treatments have developed as promising substitutes for traditional vaccine approaches. In animals, the first data of the use of successful in vitro transcribed (IVT) mRNA was reported in 1990, while reporter mRNAs were administered into in vivo mice model, and subsequently, protein expression was identified ^[5]. In 1992, a later study revealed that the use of messenger RNA encoding vasopressin in the brain hypothalamus can induce biological effects in rats ^[6]. Nonetheless, these primary but significant results could not make considerable investments in evolving mRNA-based vaccines, mainly due to concerns related to higher innate immunogenicity, mRNA instability, as well as inefficient drug delivery in vivo. Alternatively, the field approached DNA- and/or protein-based therapeutics ^[7][8]. Nonetheless, since the discovery of mRNA, it has been known the matter of consistent basic as well as applied research for many ailments ^[9][10]. In the initial decades of mRNA discovery, the main attention was on comprehensive investigations of structural as well as functional characteristics of eukaryotic mRNA and its metabolism. This is in order to make approaches for mRNA-based recombinant technology readily accessible to a wider research arena. In the late 1990s, preclinical investigation of IVT mRNAs was introduced for various applications, such as protein replacement and vaccination tools for cancer as well as infectious diseases ^{[6][11][12][13][14][15][16][17][18]}. Therefore, accumulated information allowed the latest scientific and technological improvements to overcome various difficulties related to mRNA, including its short half-life as well as adverse immunogenicity.

The administration of the mRNA-based vaccine has some benefits over subunit, live attenuated and inactivated virus, and DNA vaccines. Firstly, safety: there is no possible concern of insertional mutagenesis or infection since mRNA is non-integrating and non-infectious. Moreover, mRNA is generally destroyed under normal cellular conditions, and its in vivo half-life could be controlled via the use of diverse modification systems as well as delivery methods ^{[19][20][21][22]}. Besides, the innate immunogenicity of the mRNA could be downregulated to enhance the safety profile ^[19][22][23]. Secondly, efficacy: several modifications provide mRNA not only high stability but also translatability ^[19][22][23]. Efficient in vivo delivery of mRNA can be obtained by formulating them into carrier molecules such as polymers, peptides, lipid nanoparticles, micelles, allowing faster uptake and enhancing protein expression in the cytosol ^[20][21]. mRNA is considered the minimal hereditary material; hence, anti-vector mediated immunity is prevented, and these vaccines could be used recurrently. Finally, production: mostly due to the high yielding capability of IVT mRNA during transcription reactions, these vaccines have the potential for fast, low-priced, as well as scalable manufacturing.

The mRNA vaccine is a rapidly developing field; many preclinical investigations have been reported over the past few years ^{[24][25][26][27][28][29][30]}. Multiple human clinical trials have been initiated IVT mRNA-based therapeutics as protein-replacement therapy in the field of oncology ^{[31][32][33][34]}, cardiology ^[35][36], endocrinology ^[37], hematology ^[38][39], pulmonary medicine ^[38][40], or the treatment of other diseases ^[14][41]. To further advance this revolutionary approach, unresolved issues like targeted mRNA delivery and its intricated pharmacology require to be developed.

2. Challenges and Safety Issues in the Development of mRNA-Based Vaccines against Novel Antigens

The potential benefit of RNA-based therapeutics is their rapid development with lower side effects. The single-stranded IVT mRNA therapeutics are free from the hazard of genomic integration with the host cell and are efficient to generate high-quality viral protein. Besides, mRNAs are rapidly expressed and thus allowing the protein to be manufactured inside the cell. Experimental analyses of mRNA for rapid biological therapy has revealed outstanding tolerability as well as safety profile and reported that mRNA-based vaccines have no potential platform-inherent concerns ^[42]. However, for a majority of other mRNA-based therapeutic applications, such as protein replacement therapy, where preclinical and clinical trials are still very limited, researchers are unclear about the potential challenges and types of safety issues that should be considered ^[43]. Nevertheless, vaccine developers have pointed out several challenges and safety trials for developing novel vaccines against new infections e.g., SARS-CoV2.

Like other viruses, the novel coronavirus is an RNA virus, which exerts its effect via ‘S’ protein consisting of 1273 amino acid residues. This virus showed high mutation efficacy and genetic instability that may hamper immune induction ^[44]. Therefore, it is vital to understand genetic changes in the coding as well as non-coding sequences, genetic variant, pathogenicity, and host–pathogen interrelations. Genetic alteration in the ‘S’ protein likely to stimulate its folding pattern, which may change antigenicity and, consequently, may disturb vaccine design ^[45][46][47]. Copious reports advocated that mutations in the target proteins may be interrelated with drug resistance, leading to vaccine inefficacy. Therefore, immunogen selection for the mRNA-based vaccine development should be carefully considered and designed.

It is also critical to focus that insecurity over long-durable protection still exists against COVID-19. In several reports, there is evidence of reinfections. In such cases, it is required to report how long a protective immune induction will be continued in a patient ^[48][49][50]. Regarding the immunogenicity of the COVID-19 mRNA therapeutic in elderly individuals, it has been demonstrated that after the second inoculation, serum neutralizing titer was noticed in all the populations. Surprisingly, the binding as well as antibody neutralizing efficiency was comparable to those stated among vaccine receivers from 18 to 55 years old and were over the average of a group of control populations ^[51].

In the present pandemic situation, several researchers advised that immune inductions against coronavirus can lead to ADE (antibody-dependent enhancement) ^[52][53]. Although it was reported that immunization of the COVID19 receptor-binding domain did not mediate ADE in rodents ^[54], this principle could contribute to the pathology of several feline coronaviruses and flavivirus ^[54], notably dengue virus ^[55]. Nevertheless, ADE should be taken into consideration while evolving therapeutics against novel viruses ^[56][57]. In addition, considering the emergency need for a COVID-19 vaccine worldwide, being over-precautions, the authority should not stop the release of safe, well-tolerated, and efficient vaccines to the general populations ^[57][58][59]. Some reports demonstrated the safety concern of vaccine-enhanced disorder for inactivated vaccine candidates, remarkably vaccine-associated enhanced respiratory disease (Table 31) ^{[60][61][62][63]}.

Although mRNA vaccines can be manufactured with a minimum time; however, large-scale production of these therapeutics remains a challenging task owing to its huge uncertainty to meet the demand during the pandemic. Additionally, in recent years, nucleic acid-based vaccines could not produce effective platforms for human infections and other diseases using temperature-sensitive lipid nanoparticles, which may hamper for scaling up vaccine manufacture ^[60]. For example, BNT162b2 and the Pfizer-BioNTech COVID-19 vaccines, are lipid nanoparticle-formulated nucleoside-modified mRNA vaccines that encode SARS-CoV-2 spike protein ^[43]. Following the issuance of the emergency use authorization (EUA) for the BNT162b2 vaccine by the U.S. Food and Drug Administration in December 2020, the vaccination scheme of this vaccine was launched later in the U.S and other countries. However, the plan for massive immunization has been delayed by the stringent requirement of storage and transportation of the BNT162b2 vaccine. Besides, cold chain transportation is not available in many COVID-19 epidemic areas, so potent mRNA vaccines with enhanced temperature stability will be highly preferred in the future. Despite huge challenges and risk factors involved in the development of mRNA-based vaccines in clinical trials, several mRNA-based vaccines in human trials have been reported in the literature that has been summarized in Table 31.