Three decades of messenger RNA vaccine development

In the paper

“In a proof-of-concept study, we demonstrated that nucleoside-modified mRNA can be co-delivered with the clinically approved TLR agonist MPLA, achieving functional CTL responses like unmodified mRNA, but with strongly reduced type I IFN responses [157].”

What the sources say

• Not checked yetcites [157] source

In the paper

“By the dual”

What the sources say

• Not checked yetcites [196]

In the paper

“Indeed, as the mRNA molecules only have to reach the cytosol in order to be translated at the ribosomes, they avert the risk of being integrated into the host genome.”

In the paper

“In contrast, no significant changes were detected with unmodified mRNA, which only evoked a substantial immune activation [44].”

What the sources say

• Not checked yetcites [44] source

In the paper

“The fact that preparing a new mRNA construct tailored to a specific disease can be done in a straightforward and fast way, makes mRNA also an ideal candidate to trigger immunity against infectious diseases.”

In the paper

“In 2004, Karikó and colleagues observed, while exposing ex vivo human DCs to mRNA from different sources, that these cells tolerated mammalian mRNA, while strong inflammatory cytokine responses were detected upon the delivery of mRNA derived from bacteria, necrotic mammalian cells and IVT mRNA [39,40].”

What the sources say

• Not checked yetcites [39] source
• Not checked yetcites [40]

In the paper

“For one, mRNA can code full-length proteins, thus avoiding restrictions concerning a patient's MHC haplotype.”

In the paper

“Interestingly, also the MHC class II pathway can be targeted using mRNA as a source of antigen: this either after the secretion and recycling of the mRNA expressed proteins, or via direct shuttling of antigens from the cytosol to the lysosomes, e.g., promoted by the incorporation of a lysosomal targeting sequence in the mRNA construct [9,10].”

What the sources say

• Not checked yetcites [9]
• Not checked yetcites [10]

In the paper

“One of the key advantages of using antigen-encoding mRNA, is that it provides an easy way to evoke MHC-I presentation and elicit CTL responses.”

In the paper

“In turn, type I IFNs bind autocrine or paracrine receptors, activating the Janus kinase-signal transducer activator of transcription (JAK-STAT) pathway, which regulates the gene expression of hundreds of proteins involved in antiviral immunity [35,36].”

What the sources say

• Not checked yetcites [35]
• Not checked yetcites [36]

In the paper

“Antigen signalling by DCs can either originate from the cytoplasm, which involves the presentation of antigenic peptides in major histocompatibility complex (MHC) class I molecules, or alternatively from the lysosomes that harbour fragments of endo-or phagocytized antigens which are loaded into MHC class II.”

What the sources say

• Not checked yetcites [8]

In the paper

“DCs are specialized in the uptake, processing and presentation of protein antigens to lymphocytes, which links innate and adaptive immune responses.”

What the sources say

• Not checked yetcites [5][6][7]

In the paper

“A first-in-human trial demonstrated the clinical feasibility and safety of this approach for advanced melanoma cancers: all vaccinated patients developed CD4 + and CD8 + T cell responses against the selected antigens, with some patients showing objective antitumor responses [15].”

What the sources say

• Not checked yetcites [15] source

In the paper

“Antireverse cap (ARCA) modifications can ensure the correct cap orientation at the 5' end, which yields almost complete fractions of mRNA that can efficiently bind the ribosomes [54].”

What the sources say

• Not checked yetcites [54]

In the paper

“The company CureVac AG found that by stabilizing the mRNA with either a phosphorothioate backbone, or by its precipitation with the cationic protein protamine, antigen expression was diminished, but stronger immune-stimulating capacities could be obtained [62,63].”

What the sources say

• Not checked yetcites [62]
• Not checked yetcites [63]

In the paper

“Correlations were found between the elongation of the poly(A) tail of mRNA and the duration of expression, and specific modifications at the 3' UTR were reported that can slow-down the decay of the poly(A) tail by deadenylation [57,58].”

What the sources say

• Not checked yetcites [57] source
• Not checked yetcites [58]

In the paper

“Taken together, these findings resulted in a paradigm of the design of mRNA formulations for vaccination.One strategy is to use mRNA which is fully optimized to obtain a strong adjuvant effect, another is to work with 'modified' mRNA with high translation capacity, and thus improved antigen bio-availability [43,62].”

What the sources say

• Not checked yetcites [43] source
• Not checked yetcites [62]

In the paper

“As such, the cytosolic presence of a few mRNA molecules can ensure extensive antigen presentation to CTLs, while proteins have to rely on less efficient cross-presentation pathways.”

In the paper

“The binding of mRNA molecules to these danger sensors leads to downstream signalling via specific adaptor molecules (i.e.MyD88 for TLR7/8 and TRIF for TLR3), eventually resulting in the production of type I IFN and other pro-inflammatory cytokines (e.g., IL-6 and TNF-α).”

In the paper

“Moreover, using a high throughput system, Paunovska et al. were able to test hundreds of LNPs for the nucleic acid delivery to macrophages or endothelial cells after i.v.delivery, but found almost no in vitro-in vivo correlations [91].”

What the sources say

• Not checked yetcites [91]

In the paper

“These ionizable lipids (pKa < 7), originally optimized for siRNA delivery, have a neutral to mildly cationic charge under physiological pH conditions.This offers certain benefits over the permanently-charged lipids, the foremost of which is that ionizable lipids have been associated with a reduced toxicity and a prolonged blood circulation lifetime [100,101].”

What the sources say

• Not checked yetcites [100]
• Not checked yetcites [101]

In the paper

“Intradermal injections of granulocyte-macrophage colony-stimulating factor (GM-CSF) contributed to the infiltration of monocytes, and to the migration of mRNA-transfected, matured DCs to the LNs [86,87].”

What the sources say

• Not checked yetcites [86]
• Not checked yetcites [87] source

In the paper

“Only years later, the involvement of immune cells in the uptake of unformulated mRNA was identified: skin-resident DCs were capable of engulfing naked mRNA via macropinocytosis and could trigger T cell immunity in mice [78].”

What the sources say

• Not checked yetcites [78]

In the paper

“The addition of saturated phospholipids, such as DPPC and DSPC, increases the phase transition temperature of the cationic liposomes and supports the formation of such a lamellar lipid bilayer structure [102].”

What the sources say

• Not checked yetcites [102] source

In the paper

“Upon i.d.application, both vaccine components, sequence-engineered mRNA and protamine-complexed mRNA, were taken up by both immune cells and non-immune cells, with the highest frequencies detected within macrophages, DCs and neutrophils [79].”

What the sources say

• Not checked yetcites [79] source

In the paper

“They concluded that APCs could take up antigens that were produced by the transfected muscle cells, and that cross-presentation of the myocyte-expressed antigens was the mechanism leading to the induction of CTL responses [85].”

What the sources say

• Not checked yetcites [85]

In the paper

“The earliest studies have explored the i.m., subcutaneous (s.c.), and intradermal (i.d.) routes, showing detectable protein expression levels at the injection site, which also resulted in adaptive immune responses [4,51,75].”

What the sources say

• Not checked yetcites [4]
• Not checked yetcites [51]
• Not checked yetcites [75]

In the paper

“Over the years, it became clear that the chosen injection route plays a major role regarding the cell types that come in contact with the mRNA and the potency of the evoked immune response.”

In the paper

“Indeed, tumor-infiltrating CD8a + cross-presenting DCs were mainly responsible for mRNA uptake after injection in different subcutaneous murine tumor models, resulting in mRNA expression for >5 days [83,84].”

What the sources say

• Not checked yetcites [83]
• Not checked yetcites [84]

In the paper

“Recently, an i.n.administered TriMix mRNA vaccine was shown to be safe and well tolerated in a doseescalation clinical study against HIV-1 infection, while obtaining moderate HIV-specific T cell responses at high doses (total mRNA dose of 1.2 mg) [89].”

What the sources say

• Not checked yetcites [89]

In the paper

“Notwithstanding, the potency of this self-adjuvant effect of mRNA must be weighed against the risk of any adverse reaction inherent to it, including inflammation reactions and auto-immune events [68].”

What the sources say

• Not checked yetcites [68]

In the paper

“Since mRNAs are large and polyanionic molecules, they cannot passively diffuse across the cell membrane.Therefore, mRNA molecules rely on active transportation via endocytic uptake mechanisms.”

In the paper

“Note that the physicochemical and structural characteristics of mRNA LNPs will depend on the lipid composition, the ratio of mRNA to the total amount of (cationic) lipids, and the LNP synthesis (for a detailed review on these parameters also see [107]).”

What the sources say

• Not checked yetcites [107] source

In the paper

“They proposed that in the rapid-mixing step at acidic pH, (”

What the sources say

• Not checked yetcites [111] source
• Not checked yetcites [112] source

In the paper

“Importantly, depending on the administration route, specific extracellular barriers might be encountered which prompts specific particle optimizations.”

In the paper

“Although all of these cell types were able to internalize mRNA LNPs, only monocytes and myeloid DCs showed high translation of the mRNA.”

What the sources say

• Not checked yetcites [127] source

In the paper

“Although this was not further investigated in this study, prior reports have linked the occurrence of cholesterol nanocrystals to an improved transfection efficiency [114,115].”

What the sources say

• Not checked yetcites [114]
• Not checked yetcites [115]

In the paper

“Notably, bench-top microfluidic mixing devices are available for the production of unloaded liposomes or mRNA LNPs via this ethanol-dilution principle, with the potential to be scaled up for clinical or industrial purposes [116].”

What the sources say

• Not checked yetcites [116] source

In the paper

“In another study, Yanez Arteta et al. indicated that the structural arrangement of mRNA LNPs based on D-Lin-MC3-DMA, DSPC, cholesterol and a PEG-lipid exhibited a disordered inverse hexagonal phase containing D-Lin-MC3-DMA, cholesterol, water and mRNA.”

What the sources say

• Not checked yetcites [113] source

In the paper

“The mechanism of the formation of these ionizable LNP systems (with different nucleic acid payloads) has recently been elucidated by Kulkarni and colleagues [111,112].”

What the sources say

• Not checked yetcites [111] source
• Not checked yetcites [112] source

In the paper

“With respect to the resulting immune response, they showed that i.d.delivery resulted in an earlier activation of skin DCs and a more efficient migration to draining LNs, as well as a prolonged antigen availability at the injection site.”

What the sources say

• Not checked yetcites [127] source

In the paper

“Furthermore, this has also supported the discovery of new lead ionizable lipids holding greatly improved potency and safety for mRNA transfection [117][118][119].”

What the sources say

• Not checked yetcites [117][118][119]

In the paper

“Together, this resulted in a superior immunogenicity compared to i.m. delivery, as i.d.injection resulted in the highest antibody titers and CD4 + T cell activation.”

What the sources say

• Not checked yetcites [127] source

In the paper

“This may reduce the colloidal stability of the mRNA LNPs, followed by the aggregation of the particles and a premature release of the mRNA cargo [139,140].”

What the sources say

• Not checked yetcites [139]
• Not checked yetcites [140]

In the paper

“Recently, Vu et al. elucidated how nanoparticles mediate complement activation, and found this to be quite universal over a range of clinically approved liposomes [148].”

What the sources say

• Not checked yetcites [148]

In the paper

“These PEG-lipids can gradually diffuse out of the LNPs, and thus temporarily confer stealth properties to the LNP system, while achieving higher transfection efficiencies compared to the longer, persistent PEG-lipids [139,144].”

What the sources say

• Not checked yetcites [139]
• Not checked yetcites [144]

In the paper

“These mRNA nanoparticles had a relatively small particle size (<50 nm), and an uncharged, strongly PEGylated outer-surface.”

What the sources say

• Not checked yetcites [135] source

In the paper

“In the specific case of cancer immunotherapy, it was proven essential to evoke a systemic immune response to obtain antitumor immunity, which makes i.v.delivery a highly appealing administration route for mRNA-based cancer vaccines [92,136,137].”

What the sources say

• Not checked yetcites [92] source
• Not checked yetcites [136] source
• Not checked yetcites [137] source

In the paper

“Merely a few surface-bound immunoglobulin molecules are sufficient to trigger complement activation.”

In the paper

“Alternatively, the incorporation of PEGylated lipids is widely used to provide a”

What the sources say

• Not checked yetcites [143]

In the paper

“It is also possible to tweak the serum stability of LNPs by modifying their helper lipid composition.”

In the paper

“However, PEGylation also has its limitations: in patients with pre-existing anti-PEG antibodies, infusion of PEGylated LNPs is contra-indicated, as this could lead to an even stronger complement activation as well as an accelerated blood clearance of PEGylated nanoparticles.”

What the sources say

• Not checked yetcites [143]
• Not checked yetcites [148]
• Not checked yetcites [155]

In the paper

“Reports suggest that nanoparticles up to 200 nm are able to enter the lymphatic vessels, whereas larger particles are withheld at the injection site [128][129][130].”

What the sources say

• Not checked yetcites [128][129][130]

In the paper

“However, recent findings suggest that these nanoparticle-mediated infusion reactions are triggered by an acute intoxication of pulmonary intravascular macrophages [152][153][154].”

What the sources say

• Not checked yetcites [152][153][154]

In the paper

“Perche et al. prepared mannosylated nanoparticles loaded with mRNA, which could be used as an active targeting strategy to enhance the transfection of splenic DCs [126].”

What the sources say

• Not checked yetcites [126]

In the paper

“While it is possible that the observed site-specific targeting could be attributed to the LNP surface charge, the involvement of a different biomolecular corona cannot be excluded.”

In the paper

“Here, it was found that chronic activation of the type I IFN pathway leads to the induction of regulatory DCs, the deletion of virus-specific B cells and strongly impaired T cells responses [180][181][182].”

What the sources say

• Not checked yetcites [180][181][182]

In the paper

“If type I IFNs are produced after recognition of MHC-presented antigens by T cells, they act as a strong 3 rd signal cytokine to stimulate T cell activation and proliferation.”

In the paper

“Indeed, type I IFN signalling induces DC maturation, improved antigen processing and presentation, and enhances DC migration to enriched T cell zones (e.g., via CCR7) [172][173][174][175].”

What the sources say

• Not checked yetcites [172][173][174][175]

In the paper

“By regulating other pro-inflammatory cytokines, type I IFNs are also involved in the regulation of other immune cell types, such as Natural Killer (NK) cell responses [179].”

What the sources say

• Not checked yetcites [179]

In the paper

“For a self-amplifying mRNA LNP vaccine, Pepini et al. reported that the antigen expression was dramatically enhanced in the absence of IFN-α/β, which correlated with improved immunogenicity and IgG antibody responses [167].”

What the sources say

• Not checked yetcites [167]

In the paper

“Comparative studies performed by our research group and others demonstrated that nucleoside-modified mRNA LNPs outperformed unmodified mRNA LNPs in translation capacity, but failed to elicit successful CTL responses due to the drastically reduced production of type I IFNs [119,157].”

What the sources say

• Not checked yetcites [119]
• Not checked yetcites [157] source

In the paper

“In an opinion article, De Beuckelaer et al. suggested that the type I IFN response by mRNA vaccines might have dual effects on naïve T cells, depending on the duration and relative timing between type I IFN signalling and T cell priming [169].”

What the sources say

• Not checked yetcites [169]

In the paper

“Interestingly, by lowering the lipid-to-mRNA ratio to a lower lipid content, Kranz et al. demonstrated that DOTMA-DOPE mRNA LNPs become negatively charged and specifically target DCs in the spleen [92,158].”

What the sources say

• Not checked yetcites [92] source
• Not checked yetcites [158]

In the paper

“This might explain why the early type I IFN response induced by an mRNA vaccine, might not only limit the mRNA-encoded antigen expression, but even exert deleterious effects on T cell responses [169].”

What the sources say

• Not checked yetcites [169]

In the paper

“In humoral immunity, type I IFNs promote the generation of long-lived antibody responses by the indirect enhancement of helper T cell responses and by direct immune-stimulatory effects on B cells [177,178].”

What the sources say

• Not checked yetcites [177]
• Not checked yetcites [178]

In the paper

“As such, this sparks opportunity for combining mRNA vaccines with PD-1 checkpoint inhibitor treatments.”

In the paper

“Breckpot and colleagues demonstrated the reprogramming of DCs after the intratumoral administration of TriMix mRNA (i.e.CD40L, caTLR4 and CD70), or using mRNA encoding a fusokine consisting of IFN-β and the ectodomain of the TGF-β receptor II [84,202].”

What the sources say

• Not checked yetcites [84]
• Not checked yetcites [202]

In the paper

“Indeed, the expression of the MLKL provokes a type of immunogenic cell death, called necroptosis, which enables the release of tumor-specific antigens and natural DAMPs that trigger antitumor responses.”

What the sources say

• Not checked yetcites [204]

In the paper

“In addition, it is of crucial importance to keep a close eye on several immunosuppressive mechanisms that might hamper the evoked immunity.”

In the paper

“This is a condition of temporary facial paralysis, which has been linked to autoimmunity [188].”

What the sources say

• Not checked yetcites [188]

In the paper

“By the localized expression of these immunomodulatory molecules, this strategy was able to break DC tolerance and to activate tumor-specific T cell responses.”

What the sources say

• Not checked yetcites [203] source

In the paper

“To give two examples, Thran et al. evaluated an LNP formulation of sequence-engineered mRNA coding for prophylactic or therapeutic antibodies.This could provide protection against rabies infection, botulinum intoxication, and tumors [201].”

What the sources say

• Not checked yetcites [201] source

In the paper

“There is, however no doubt that distinct mRNA vaccines, varying in mRNA design, formulation and administration route, will exhibit differences in kinetics and cell-specificity for both mRNA expression and innate immune activation.”

In the paper

“This includes the generation of transgenic reporter mouse strains, such as the IFN-β reporter mice and Cre recombinase (mRNA) reporter mice, as well as the development of fluorescent imaging mRNA probes that allow to detect mRNA at high resolution [90,158,[192][193][194][195].”

What the sources say

• Not checked yetcites [90,158,[192][193][194][195]

In the paper

“This mRNA platform represented a valid alternative to classic mRNA cancer vaccines, providing a strong infiltration of T cells able to eliminate tumors in mice.”

What the sources say

• Not checked yetcites [197] source

In the paper

“Firstly, it is essential to ensure adequate intracellular delivery of mRNA molecules, preferably by targeting APCs in vivo.”

In the paper

“Two papers showed that the i.v.administration of unmodified mRNA LNPs triggered a rapid and systemic induction of type I IFNs, which involved the selective targeting of DCs and macrophages and the activation of the TLR7 pathway[92,136].”

What the sources say

• Not checked yetcites [92] source
• Not checked yetcites [136] source

In the paper

“Both parameters are of crucial importance to the vaccine outcome, but are inherently linked to the structural properties of the mRNA molecule itself.”

In the paper

“However, it is still waiting for a more extended clinical experience on how patients respond to mRNA vaccines, including more comparative research to select for the best suited mRNA platform and administration route, as well as to show clear therapeutic benefits over other vaccine strategies.”

In the paper

“We discussed that the design of these LNPs asks for a multi-objective solving approach in which the overall mRNA transfection capacity will depend on trade-offs between different requirements to cross the extra-and intracellular barriers, while precautions are needed to prevent potential nanoparticle inherent (immune) toxicities.”

In the paper

“mRNA represents an attractive source of antigen in vaccination approaches”

In the paper

“Both parameters are of crucial importance to the vaccine outcome, but are inherently linked to the structural properties of the mRNA molecule itself.”

In the paper

“Nowadays, there appears to be a trend towards the use of lipid-based nanoparticles for both localized and systemic delivery of mRNA.”

In the paper

“Nanoformulation widens the administration and delivery options for mRNA vaccines”

In the paper

“Fundamental research should further focus on how to control the type I IFN activity of mRNA vaccines, which might vary depending on the mRNA formulation, administration route, the individual patient and the intended therapeutic application.”

In the paper

“For clinical translation of these mRNA LNP systems, it will also be critical to meet pharmaceutical-and regulatory product requirements, going from scale-up manufacturing under GMP conditions, quality control of these products, stability testing, and good safety and efficacy assessments during the pre-clinical stage in relevant animal models.”

In the paper

“Over the years, a fairly large knowledge gap about the in vivo behaviour of mRNA has been filled, both after the delivery of naked mRNA or when formulated in nanoparticles, and this for various immunization routes.”

In the paper

“For prophylactic mRNA vaccines against infectious diseases, (pre-)clinical studies (in non-human primates) show accumulating evidence that mRNA vaccination is feasible, generally well-tolerated, and potentially beneficial over other traditional vaccine approaches.”

In the paper

“More specifically, although the mRNA-induced type I IFN response might add to the immunogenicity of the mRNA vaccine, it also interferes with the mRNA-encoded antigen expression and possibly even lead to deleterious immune effects.”

In the paper

“The self-adjuvant effect of mRNA is a double-edged sword for vaccine efficacy”

In the paper

“Each of these elements can be optimized and modified in order to modulate the stability, translation capacity, and immune-stimulatory profile of mRNA.”

In the paper

“Both unformulated and nanoparticulate mRNA are used for direct in vivo vaccination”

In the paper

“The choice of the amino lipids (cationic or ionizable lipids) and helper lipids will have an impact on the structural arrangement and functional properties of the formulated mRNA LNPs.”

In the paper

“As such, mRNA approaches which do not necessarily rely on the self-adjuvant effect of mRNA are one the rise, such as the replacement of type I IFN responses by more controllable adjuvant systems, or using mRNA for passive immunization approaches.”

In the paper

“Therefore, the establishment of effective combination therapies that could increase the response rate of checkpoint inhibitors is an area of intensive investigation [220,221].”

What the sources say

• Not checked yetcites [220]
• Not checked yetcites [221]

In the paper

“Contradictory results have been reported about the role of type I IFN responses on the immunogenicity of mRNA vaccines (see also section 2.3).”