Table 2.

mRNA vaccine delivery systems

Delivery types	Delivery subtypes	Advanntages	Challenges
Carrier based delivery system	Liposomes and their derivatives, mainly lipid nanoparticles(LNPs) [65]	Lipid nanoparticles (LNPs) demonstrate a remarkable mRNA encapsulation efficiency, which is pivotal for the protection of mRNA from nuclease degradation and subsequent stable delivery to the target cells Moreover, LNPs possess a distinct advantage in terms of tissue penetration, which facilitates deeper penetration into tissues and organs, thereby enabling more widespread and efficient cellular uptake. The nanoscale dimensions of LNPs contribute to their enhanced intracellular delivery, as they can easily traverse cellular barriers and accumulate within the target cells In addition to their delivery efficiency, LNPs exhibit low cytotoxicity and immunogenicity, which are critical attributes for their application in therapeutic settings Another notable feature of LNPs is their potent adjuvant properties, which are essential for enhancing the immune response when delivering vaccines or immunotherapies [66–68].	Lipid nanoparticles (LNPs) are susceptible to degradation, showcasing suboptimal stability under storage conditions, with a propensity for aggregation and fusion phenomena, which can compromise their structural integrity and therapeutic efficacy [69].
	Polymers [70]	Certain polymers have demonstrated the ability to significantly enhance the process of endosomal escape, thereby improving the delivery efficiency of therapeutic agents. Additionally, these polymers provide protection for messenger RNAs (mRNAs) against enzymatic degradation, ensuring their stability, and facilitate a safe and effective release of mRNAs into the cytoplasm for subsequent translation [71].	The low purity and high molecular weight of polymer-based delivery vectors, coupled with their high charge density, can result in significant cytotoxicity [72]
	Virus-like replicon particles [73]	Viral replicon particles (VRPs) have the unique capacity to encapsulate self-amplifying RNA (saRNA)-encoded antigens, effectively facilitating their transport to the cytosol. Through in vitro synthesis, viral structural proteins can be produced and utilized for encapsulating saRNAs that encode specific antigens. Extensive researches [74] has illuminated the therapeutic potential of mRNA vaccines administered via VRPs across a diverse array of viral, bacterial diseases, and cancer. This method enhances RNA replication, elicits potent innate immune responses, and promotes the maturation of dendritic cells, contributing to the vaccines' efficacy and immunogenicity.	Viral replicon particles (VRPs) possess a notable disadvantage, as they have been observed to elicit neutralizing antibody responses specifically targeted against the viral surface proteins, as evidenced by studies [75, 76].
	Cationic nanoemulsion (CNE) [77]	CNE can enhance the efficacy of mRNA vaccines by binding to saRNA in a pH-dependent manner, comprising nanoemulsions and cationic lipids. Nanoemulsions can be generated via techniques such as ultrasound, microfluidics, and vigorous stirring [78]. Among the CNE components, the cationic lipid 1.2-diol sn glycerol-3-phosphate choline (DOTAP) stands out for its positive charge, being emulsified with MF59, the identical adjuvant component of the lotion [79]. Additionally, CNE has shown promising therapeutic effects in its ability to deliver saRNA, indicating that lower doses of adjuvant subunits in CNE complexes can elicit substantial immune responses [80]. Numerous studies have been conducted to investigate the stability, toxicity, and biodistribution of CNE, with findings confirming its stability [81].	However, the conclusions regarding the toxicity of CNE vary across different models. One study demonstrated that the toxicity of nanoemulsions on human foetal lung cells (MRC-5) is dose-dependent [82]. In contrast, another investigation revealed that the rabies animal model exhibited suitable tolerance to CNE-delivered self-amplifying mRNA (SAM) vaccines [83]
	Cationic cell-penetrating peptides (CPP) [84, 85]	Cationic peptides, including protamine, a well-established cationic peptide utilized for mRNA transport [86], facilitate the formation of nanosized complexes with mRNAs. These complexes effectively shield the mRNA from enzymatic degradation, maintain immunogenicity across varying temperatures, and preserve the potency of antigen-encoded mRNA vaccines [87]. Protamine's ability to spontaneously condense mRNA via electrostatic interactions serves to protect the enclosed mRNA from degradation by extracellular RNases [88, 89]. Furthermore, the protamine-mRNA complexes demonstrate adjuvant properties, stimulating TLR7/8 to trigger robust innate immune responses [90].	The specific combination ratio and binding strength between protamine and mRNA are crucial factors that can significantly influence the translation process. These parameters may impose limitations on the efficiency of vaccine protein expression, ultimately affecting the overall effectiveness of the vaccine in eliciting an immune response and providing protection [91].
Naked mRNA	-	First, the mRNA cannot be integrated into the genome, reducing the risk of genetic mutations. Second, ribosomes can bind directly to the mRNA in the cytoplasm, causing the mRNA to be translated immediately and rapidly initiating an immune response after vaccination. Third, the final position of the mRNA determines the site of protein expression, allowing for precise control of protein expression [92, 93].	The lack of a carrier during the delivery process can lead to unstable protein translation and expression. However, this can be mitigated by altering the administration method and proper chemical modifications. However, research in this area is relatively limited at present [94].
Dendritic Cell-mRNA Delivery System (DCs-mRNA)	-	Dendritic cells (DCs) serve as the orchestrators of the immune response, exhibiting unparalleled efficiency in their ability to capture and present antigens. This is achieved through a meticulously regulated process involving internalization and proteolytic degradation. Following this intricate mechanism, DCs proceed to present antigens to CD8 + T or CD4 + T cells via major histocompatibility complexes (MHCs), specifically MHC class I (MHCI) or MHC class II (MHCII). By doing so, they initiate an adaptive immune response [95]. The pivotal role of DCs in this context underscores their significance as prime targets for vaccination strategies.	Challenges primarily include the two aspects: Firstly, serum protein aggregation and mRNA degradation upon systemic administration [96], compromising vaccine integrity. Additionally, the second challenge involves the efficient systematic dissemination of mRNA vaccines, ensuring uniform distribution [97] for optimal immune response.