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1.
J Am Chem Soc ; 146(28): 19218-19228, 2024 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-38955767

RESUMO

The messenger RNA (mRNA) vaccines hold great significance in contagion prevention and cancer immunotherapy. However, safely and effectively harnessing innate immunity to stimulate robust and durable adaptive immune protection is crucial, yet challenging. In this study, we synthesized a library of stimuli-responsive bivalent ionizable lipids (srBiv iLPs) with smart molecular blocks responsive to esterase, H2O2, cytochrome P450, alkaline phosphatase, nitroreductase, or glutathione (GSH), aiming to leverage physiological cues to trigger fast lipid degradation, promote mRNA translation, and induce robust antitumor immunity via reactive oxygen species (ROS)-mediated boosting. After subcutaneous immunization, esterase-responsive vaccine (eBiv-mVac) was rapidly internalized and transported into the draining lymph nodes. It then underwent fast decaging and self-immolative degradation in esterase-rich antigen-presenting cells, releasing sufficient mRNA for antigen translation and massive reactive quinone methides to elevate ROS levels. This resulted in broad activation of innate immunity to boost T cell response, prompting a large number of primed antigen-specific CD8+ T cells to circulate and infiltrate into tumors (>1000-fold versus unvaccinated control), thereby orchestrating innate and adaptive immunity to control tumor growth. Moreover, by further combining our vaccination strategy with immune checkpoint blockade, we demonstrated a synergism that significantly amplified the magnitude and function of antigen-specific CD8+ T cells. This, in turn, caused potent systemic antitumor efficacy and prolonged survival with high complete response rate in xenograft and metastasis models. Overall, our generalized stimuli-responsive mRNA delivery platform promises a paradigm shift in the design of potent vaccines for cancer immunotherapy, as well as effective and precise carriers for gene editing, protein replacement, and cell engineering.


Assuntos
Linfócitos T CD8-Positivos , Imunidade Inata , Espécies Reativas de Oxigênio , Espécies Reativas de Oxigênio/metabolismo , Imunidade Inata/efeitos dos fármacos , Animais , Linfócitos T CD8-Positivos/imunologia , Camundongos , Vacinas de mRNA/química , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/química , RNA Mensageiro/imunologia , RNA Mensageiro/genética , Humanos , Camundongos Endogâmicos C57BL
2.
Front Immunol ; 15: 1381508, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38690272

RESUMO

Seasonal influenza remains a serious global health problem, leading to high mortality rates among the elderly and individuals with comorbidities. Vaccination is generally accepted as the most effective strategy for influenza prevention. While current influenza vaccines are effective, they still have limitations, including narrow specificity for certain serological variants, which may result in a mismatch between vaccine antigens and circulating strains. Additionally, the rapid variability of the virus poses challenges in providing extended protection beyond a single season. Therefore, mRNA technology is particularly promising for influenza prevention, as it enables the rapid development of multivalent vaccines and allows for quick updates of their antigenic composition. mRNA vaccines have already proven successful in preventing COVID-19 by eliciting rapid cellular and humoral immune responses. In this study, we present the development of a trivalent mRNA vaccine candidate, evaluate its immunogenicity using the hemagglutination inhibition assay, ELISA, and assess its efficacy in animals. We demonstrate the higher immunogenicity of the mRNA vaccine candidate compared to the inactivated split influenza vaccine and its enhanced ability to generate a cross-specific humoral immune response. These findings highlight the potential mRNA technology in overcoming current limitations of influenza vaccines and hold promise for ensuring greater efficacy in preventing seasonal influenza outbreaks.


Assuntos
Imunidade Humoral , Vírus da Influenza A Subtipo H1N1 , Vacinas contra Influenza , Infecções por Orthomyxoviridae , Vacinas de mRNA , Animais , Feminino , Humanos , Camundongos , Reações Cruzadas/imunologia , Ensaio de Imunoadsorção Enzimática , Células HEK293 , Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Imunidade Humoral/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/química , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Camundongos Endogâmicos BALB C , Vacinas de mRNA/administração & dosagem , Vacinas de mRNA/química , Vacinas de mRNA/genética , Vacinas de mRNA/imunologia , Estações do Ano , Fatores de Tempo , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , Infecções por Orthomyxoviridae/virologia
3.
Adv Mater ; 36(23): e2311574, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38433564

RESUMO

Dendritic cell (DC) maturation is a crucial process for antigen presentation and the initiation of T cell-mediated immune responses. Toll-like receptors play pivotal roles in stimulating DC maturation and promoting antigen presentation. Here, a novel message RNA (mRNA) cancer vaccine is reported that boosts antitumor efficacy by codelivering an mRNA encoding tumor antigen and a TLR7/8 agonist (R848) to DC using supramolecular lipid nanoparticles (SMLNP) as a delivery platform, in which a new ionizable lipid (N2-3L) remarkably enhances the translation efficiency of mRNA and a ß-cyclodextrin (ß-CD)-modified ionizable lipid (Lip-CD) encapsulates R848. The incorporation of R848 adjuvant into the mRNA vaccine through noncovalent host-guest complexation significantly promotes DC maturation and antigen presentation after vaccination, thus resulting in superior antitumor efficacy in vivo. Moreover, the antitumor efficacy is further boosted synergized with immune checkpoint blockade by potentiating the anticancer capability of cytotoxic T lymphocytes infiltrated in tumor sites. This work indicates that SMLNP shows brilliant potential as next-generation delivery system in the development of mRNA vaccines with high efficacy.


Assuntos
Vacinas Anticâncer , Células Dendríticas , Imidazóis , Imunoterapia , Lipídeos , Nanopartículas , Receptor 7 Toll-Like , Receptor 8 Toll-Like , Animais , Nanopartículas/química , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Camundongos , Lipídeos/química , Imidazóis/química , Vacinas de mRNA/química , beta-Ciclodextrinas/química , RNA Mensageiro/genética , RNA Mensageiro/química , Neoplasias/terapia , Linhagem Celular Tumoral , Antígenos de Neoplasias/imunologia , Humanos , Camundongos Endogâmicos C57BL , Lipossomos
4.
Nature ; 621(7978): 396-403, 2023 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-37130545

RESUMO

Messenger RNA (mRNA) vaccines are being used to combat the spread of COVID-19 (refs. 1-3), but they still exhibit critical limitations caused by mRNA instability and degradation, which are major obstacles for the storage, distribution and efficacy of the vaccine products4. Increasing secondary structure lengthens mRNA half-life, which, together with optimal codons, improves protein expression5. Therefore, a principled mRNA design algorithm must optimize both structural stability and codon usage. However, owing to synonymous codons, the mRNA design space is prohibitively large-for example, there are around 2.4 × 10632 candidate mRNA sequences for the SARS-CoV-2 spike protein. This poses insurmountable computational challenges. Here we provide a simple and unexpected solution using the classical concept of lattice parsing in computational linguistics, where finding the optimal mRNA sequence is analogous to identifying the most likely sentence among similar-sounding alternatives6. Our algorithm LinearDesign finds an optimal mRNA design for the spike protein in just 11 minutes, and can concurrently optimize stability and codon usage. LinearDesign substantially improves mRNA half-life and protein expression, and profoundly increases antibody titre by up to 128 times in mice compared to the codon-optimization benchmark on mRNA vaccines for COVID-19 and varicella-zoster virus. This result reveals the great potential of principled mRNA design and enables the exploration of previously unreachable but highly stable and efficient designs. Our work is a timely tool for vaccines and other mRNA-based medicines encoding therapeutic proteins such as monoclonal antibodies and anti-cancer drugs7,8.


Assuntos
Algoritmos , Vacinas contra COVID-19 , COVID-19 , Estabilidade de RNA , RNA Mensageiro , SARS-CoV-2 , Vacinas de mRNA , Animais , Humanos , Camundongos , Códon/genética , COVID-19/genética , COVID-19/imunologia , COVID-19/prevenção & controle , Vacinas contra COVID-19/química , Vacinas contra COVID-19/genética , Vacinas contra COVID-19/imunologia , Meia-Vida , Herpesvirus Humano 3/genética , Herpesvirus Humano 3/imunologia , Vacinas de mRNA/química , Vacinas de mRNA/genética , Vacinas de mRNA/imunologia , Estabilidade de RNA/genética , Estabilidade de RNA/imunologia , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/imunologia , RNA Mensageiro/metabolismo , SARS-CoV-2/genética , SARS-CoV-2/imunologia
6.
Science ; 376(6594): 680-681, 2022 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-35549401
8.
Chem Pharm Bull (Tokyo) ; 69(12): 1141-1159, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34853281

RESUMO

Considerable efforts have been made on the development of lipid nanoparticles (LNPs) for delivering of nucleic acids in LNP-based medicines, including a first-ever short interfering RNA (siRNA) medicine, Onpattro, and the mRNA vaccines against the coronavirus disease 2019 (COVID-19), which have been approved and are currently in use worldwide. The successful rational design of ionizable cationic lipids was a major breakthrough that dramatically increased delivery efficiency in this field. The LNPs would be expected to be useful as a platform technology for the delivery of various therapeutic modalities for genome editing and even for undiscovered therapeutic mechanisms. In this review, the current progress of my research, including the molecular design of pH-sensitive cationic lipids, their applications for various tissues and cell types, and for delivering various macromolecules, including siRNA, antisense oligonucleotide, mRNA, and the clustered regularly interspaced short palindromic repeats (CRISPR)-associated (Cas) system will be described. Mechanistic studies regarding relationships between the physicochemical properties of LNPs, drug delivery, and biosafety are also summarized. Furthermore, current issues that need to be addressed for next generation drug delivery systems are discussed.


Assuntos
Portadores de Fármacos/química , Lipídeos/química , Lipossomos/química , Nanopartículas/química , COVID-19/imunologia , COVID-19/prevenção & controle , COVID-19/virologia , Cátions/química , Concentração de Íons de Hidrogênio , RNA Guia de Cinetoplastídeos/química , RNA Guia de Cinetoplastídeos/metabolismo , RNA Interferente Pequeno/química , RNA Interferente Pequeno/metabolismo , SARS-CoV-2/isolamento & purificação , Vacinas de mRNA/química , Vacinas de mRNA/metabolismo
9.
Acc Chem Res ; 54(23): 4262-4271, 2021 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-34756014

RESUMO

Over the past two decades, research on mRNA-based therapies has exploded, mainly because of the inherent advantages of mRNA, including a low integration probability, transient expression, and simple and rapid in vitro transcription production approaches. In addition, thanks to improved stability and reduced immunogenicity by advanced strategies, the application of mRNA has expanded from protein replacement therapy to vaccination, gene editing and other fields, showing great promise for clinical application. Recently, with the successive launch of two mRNA-based COVID-19 vaccines, mRNA technology has attracted an enormous amount of attention from scientific researchers as well as pharmaceutical companies. Because of the large molecular weight, hydrophilicity, and highly negative charge densities of mRNA, it is difficult to overcome the intracellular delivery barriers. Therefore, various delivery vehicles have been developed to achieve more effective mRNA delivery. In general, conventional mRNA administration methods are based on injection strategies, including intravenous, intramuscular, intradermal, and subcutaneous injections. Although these routes circumvent the absorption barriers to some extent, they bring about injection-related concerns such as safety issues, pain, low compliance, and difficulty in repeated dosing, increasing the need to explore alternative strategies for noninvasive delivery. The ideal noninvasive delivery systems are featured with easy to use, low risks of infection, and good patient compliance. At the same time, they allow patients to self-administer, reducing reliance on professional healthcare workers and interference with bodily functions and daily life. In particular, the noninvasive mucosal delivery of mRNA vaccines can induce mucosal immune responses, which are important for resisting pathogens infected through mucosal routes.Because of the potential clinical benefits mentioned above, we detailed the existing strategies for the noninvasive delivery of mRNA in this review, including delivery via the nasal, pulmonary, vaginal, and transdermal routes. First, we discussed the unique strengths and biological hindrances of each route on the basis of physiology. Next, we comprehensively summarized the research progress reported so far and analyzed the technologies and delivery vehicles used, hoping to provide some references for further explorations. Among these noninvasive routes, nasal and pulmonary delivery are the earliest and most intensively studied areas, mostly owing to their favorable physiological structures: the nasal or pulmonary mucosa is easily accessible, highly permeable and highly vascularized. In contrast, the development of vaginal mRNA delivery is relatively less reported, and the current research mainly focused on some local applications. In addition, microneedles have also been investigated to overcome skin barriers for mRNA delivery in recent years, making microneedle-based delivery an emerging alternative pathway. In summary, a variety of mRNA formulations and delivery strategies have been developed for noninvasive mRNA delivery, skillfully combining appropriate vehicles or physical technologies to enhance effectiveness. We surmise that continuous advances and technological innovations in the development of mRNA noninvasive delivery will accelerate the translation from experimental research to clinical application.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Vacinas de mRNA/química , Administração Cutânea , Administração por Inalação , Animais , COVID-19/prevenção & controle , COVID-19/virologia , Humanos , SARS-CoV-2/isolamento & purificação , Vacinas de mRNA/administração & dosagem , Vacinas de mRNA/imunologia
10.
Microbiol Spectr ; 9(3): e0096521, 2021 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-34756082

RESUMO

The prompt rollout of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mRNA vaccine is facilitating population immunity, which is becoming more dominant than natural infection-mediated immunity. In the midst of coronavirus disease 2019 (COVID-19) vaccine deployment, understanding the epitope profiles of vaccine-elicited antibodies will be the first step in assessing the functionality of vaccine-induced immunity. In this study, the high-resolution linear epitope profiles of Pfizer-BioNTech COVID-19 mRNA vaccine recipients and COVID-19 patients were delineated by using microarrays mapped with overlapping peptides of the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. The vaccine-induced antibodies targeting the RBD had a broader distribution across the RBD than that induced by the natural infection. Half-maximal neutralization titers were measured in vitro by live virus neutralization assays. As a result, relatively lower neutralizability was observed in vaccine recipient sera, when normalized to a total anti-RBD IgG titer. However, mutation panel assays targeting the SARS-CoV-2 variants of concern have shown that the vaccine-induced epitope variety, rich in breadth, may grant resistance against future viral evolutionary escapes, serving as an advantage of vaccine-induced immunity. IMPORTANCE Establishing vaccine-based population immunity has been the key factor in attaining herd protection. Thanks to expedited worldwide research efforts, the potency of mRNA vaccines against the coronavirus disease 2019 (COVID-19) is now incontestable. The next debate is regarding the coverage of SARS-CoV-2 variants. In the midst of vaccine deployment, it is of importance to describe the similarities and differences between the immune responses of COVID-19 vaccine recipients and naturally infected individuals. In this study, we demonstrated that the antibody profiles of vaccine recipients are richer in variety, targeting a key protein of the invading virus, than those of naturally infected individuals. Vaccine-elicited antibodies included more nonneutralizing antibodies than infection-elicited antibodies, and their breadth in antibody variations suggested possible resilience against future SARS-CoV-2 variants. The antibody profile achieved by vaccinations in naive individuals provides important insight into the first step toward vaccine-based population immunity.


Assuntos
Vacinas contra COVID-19/imunologia , COVID-19/imunologia , Mapeamento de Epitopos , Ligação Proteica , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/química , Vacinas de mRNA/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/química , COVID-19/prevenção & controle , Vacinas contra COVID-19/química , Humanos , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinação , Vacinas Sintéticas/imunologia , Vacinas de mRNA/química
11.
J Am Chem Soc ; 143(49): 20529-20545, 2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34841867

RESUMO

Unquestionably, polymers have influenced the world over the past 100 years. They are now more crucial than ever since the COVID-19 pandemic outbreak. The pandemic paved the way for certain polymers to be in the spotlight, namely sequence-defined polymers such as messenger ribonucleic acid (mRNA), which was the first type of vaccine to be authorized in the U.S. and Europe to protect against the SARS-CoV-2 virus. This rise of mRNA will probably influence scientific research concerning nucleic acids in general and RNA therapeutics in specific. In this Perspective, we highlight the recent trends in sequence-controlled and sequence-defined polymers. Then we discuss mRNA vaccines as an example to illustrate the need of ultimate sequence control to achieve complex functions such as specific activation of the immune system. We briefly present how mRNA vaccines are produced, the importance of modified nucleotides, the characteristic features, and the advantages and challenges associated with this class of vaccines. Finally, we discuss the chances and opportunities for polymer chemistry to provide solutions and contribute to the future progress of RNA-based therapeutics. We highlight two particular roles of polymers in this context. One represents conjugation of polymers to nucleic acids to form biohybrids. The other is concerned with advanced polymer-based carrier systems for nucleic acids. We believe that polymers can help to address present problems of RNA-based therapeutic technologies and impact the field beyond the COVID-19 pandemic.


Assuntos
Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , Vacinas contra COVID-19/química , Polímeros/farmacologia , SARS-CoV-2/efeitos dos fármacos , Vacinas de mRNA/química , Animais , Portadores de Fármacos , Humanos
12.
Nat Commun ; 12(1): 6777, 2021 11 22.
Artigo em Inglês | MEDLINE | ID: mdl-34811367

RESUMO

Lipid nanoparticle (LNP)-formulated mRNA vaccines were rapidly developed and deployed in response to the SARS-CoV-2 pandemic. Due to the labile nature of mRNA, identifying impurities that could affect product stability and efficacy is crucial to the long-term use of nucleic-acid based medicines. Herein, reversed-phase ion pair high performance liquid chromatography (RP-IP HPLC) was used to identify a class of impurity formed through lipid:mRNA reactions; such reactions are typically undetectable by traditional mRNA purity analytical techniques. The identified modifications render the mRNA untranslatable, leading to loss of protein expression. Specifically, electrophilic impurities derived from the ionizable cationic lipid component are shown to be responsible. Mechanisms implicated in the formation of reactive species include oxidation and subsequent hydrolysis of the tertiary amine. It thus remains critical to ensure robust analytical methods and stringent manufacturing control to ensure mRNA stability and high activity in LNP delivery systems.


Assuntos
Sistemas de Liberação de Medicamentos , Lipossomos/química , Nanopartículas/química , RNA Mensageiro/química , Potência de Vacina , Aldeídos/química , Cromatografia Líquida , Humanos , Íons/química , Lipídeos/química , Nucleosídeos/química , Oxirredução , Biossíntese de Proteínas , Estabilidade de RNA , Vacinas de mRNA/química
13.
Bioconjug Chem ; 32(12): 2497-2506, 2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34775749

RESUMO

Understanding immune responses toward viral infection will be useful for potential therapeutic intervention and offer insights into the design of prophylactic vaccines. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the COVID-19 pandemic. To understand the complex immune responses toward SARS-CoV-2 infection, here we developed a method to express and purify the recombinant and engineered viral receptor-binding domain (RBD) to more than 95% purity. We could encapsulate RNA molecules into the interior of a virion-sized liposome. We conjugated the purified RBD proteins onto the surface of the liposome in an orientation-specific manner with defined spatial densities. Both the encapsulation of RNAs and the chemical conjugation of the RBD protein on liposome surfaces were stable under physiologically relevant conditions. In contrast to soluble RBD proteins, a single injection of RBD-conjugated liposomes alone, in the absence of any other adjuvants, elicited RBD-specific B cell responses in BALB/c mice, and the resulting animal sera could potently neutralize HIV-1 pseudovirions that displayed the SARS-CoV-2 spike proteins. These results validate these supramolecular structures as a novel and effective tool to mimic the structure of enveloped viruses, the use of which will allow systematic dissection of the complex B cell responses to SARS-CoV-2 infection.


Assuntos
Anticorpos Neutralizantes/imunologia , Vacinas contra COVID-19/uso terapêutico , COVID-19/prevenção & controle , Lipossomos/uso terapêutico , SARS-CoV-2/imunologia , Glicoproteína da Espícula de Coronavírus/uso terapêutico , Adjuvantes Imunológicos/química , Adjuvantes Imunológicos/uso terapêutico , Animais , COVID-19/imunologia , Vacinas contra COVID-19/química , Feminino , Humanos , Imunização , Lipossomos/química , Camundongos Endogâmicos BALB C , Modelos Moleculares , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/química , Vacinas Sintéticas/química , Vacinas Sintéticas/uso terapêutico , Vacinas de mRNA/química , Vacinas de mRNA/uso terapêutico
14.
Viruses ; 13(8)2021 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-34452327

RESUMO

Human T-cell lymphotropic virus type 1 (HTLV-1) infection affects millions of individuals worldwide and can lead to severe leukemia, myelopathy/tropical spastic paraparesis, and numerous other disorders. Pursuing a safe and effective immunotherapeutic approach, we compared the viral polyprotein and the human proteome with a sliding window approach in order to identify oligopeptide sequences unique to the virus. The immunological relevance of the viral unique oligopeptides was assessed by searching them in the immune epitope database (IEDB). We found that HTLV-1 has 15 peptide stretches each consisting of uniquely viral non-human pentapeptides which are ideal candidate for a safe and effective anti-HTLV-1 vaccine. Indeed, experimentally validated HTLV-1 epitopes, as retrieved from the IEDB, contain peptide sequences also present in a vast number of human proteins, thus potentially instituting the basis for cross-reactions. We found a potential for cross-reactivity between the virus and the human proteome and described an epitope platform to be used in order to avoid it, thus obtaining effective, specific, and safe immunization. Potential advantages for mRNA and peptide-based vaccine formulations are discussed.


Assuntos
Epitopos/química , Infecções por HTLV-I/prevenção & controle , Vírus Linfotrópico T Tipo 1 Humano/imunologia , RNA Mensageiro/química , Vacinas de Subunidades Antigênicas/imunologia , Vacinas Sintéticas/imunologia , Vacinas Virais/imunologia , Vacinas de mRNA/imunologia , Sequência de Aminoácidos , Bases de Dados Genéticas , Mapeamento de Epitopos , Epitopos/genética , Epitopos/imunologia , Infecções por HTLV-I/imunologia , Infecções por HTLV-I/virologia , Vírus Linfotrópico T Tipo 1 Humano/química , Vírus Linfotrópico T Tipo 1 Humano/genética , Humanos , Imunização , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/genética , Vacinas Sintéticas/química , Vacinas Sintéticas/genética , Vacinas Virais/química , Vacinas Virais/genética , Vacinas de mRNA/química , Vacinas de mRNA/genética
15.
Mol Ther ; 29(9): 2769-2781, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-33992803

RESUMO

It is well established that memory CD8 T cells protect susceptible strains of mice from mousepox, a lethal viral disease caused by ectromelia virus (ECTV), the murine counterpart to human variola virus. While mRNA vaccines induce protective antibody (Ab) responses, it is unknown whether they also induce protective memory CD8 T cells. We now show that immunization with different doses of unmodified or N(1)-methylpseudouridine-modified mRNA (modified mRNA) in lipid nanoparticles (LNP) encoding the ECTV gene EVM158 induced similarly strong CD8 T cell responses to the epitope TSYKFESV, albeit unmodified mRNA-LNP had adverse effects at the inoculation site. A single immunization with 10 µg modified mRNA-LNP protected most susceptible mice from mousepox, and booster vaccination increased the memory CD8 T cell pool, providing full protection. Moreover, modified mRNA-LNP encoding TSYKFESV appended to green fluorescent protein (GFP) protected against wild-type ECTV infection while lymphocytic choriomeningitis virus glycoprotein (GP) modified mRNA-LNP protected against ECTV expressing GP epitopes. Thus, modified mRNA-LNP can be used to create protective CD8 T cell-based vaccines against viral infections.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Vírus da Ectromelia/imunologia , Ectromelia Infecciosa/prevenção & controle , Proteínas Virais/genética , Vacinas de mRNA/administração & dosagem , Animais , Composição de Medicamentos , Ectromelia Infecciosa/imunologia , Imunização Secundária , Memória Imunológica , Lipossomos , Masculino , Camundongos , Nanopartículas , Peptídeos/química , Peptídeos/genética , Peptídeos/imunologia , Pseudouridina/análogos & derivados , Pseudouridina/química , Proteínas Virais/química , Proteínas Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/química , Vacinas Virais/farmacologia , Vacinas de mRNA/química , Vacinas de mRNA/farmacologia
16.
Mol Ther ; 29(9): 2782-2793, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34058388

RESUMO

We present a live-attenuated RNA hybrid vaccine technology that uses an RNA vaccine delivery vehicle to deliver in vitro-transcribed, full-length, live-attenuated viral genomes to the site of vaccination. This technology allows ready manufacturing in a cell-free environment, regardless of viral attenuation level, and it promises to avoid many safety and manufacturing challenges of traditional live-attenuated vaccines. We demonstrate this technology through development and testing of a live-attenuated RNA hybrid vaccine against Chikungunya virus (CHIKV), comprised of an in vitro-transcribed, highly attenuated CHIKV genome delivered by a highly stable nanostructured lipid carrier (NLC) formulation as an intramuscular injection. We demonstrate that single-dose immunization of immunocompetent C57BL/6 mice results in induction of high CHIKV-neutralizing antibody titers and protection against mortality and footpad swelling after lethal CHIKV challenge.


Assuntos
Anticorpos Neutralizantes/sangue , Febre de Chikungunya/prevenção & controle , Vírus Chikungunya/genética , Lipídeos/química , Vacinas de mRNA/administração & dosagem , Animais , Anticorpos Antivirais/sangue , Febre de Chikungunya/imunologia , Vírus Chikungunya/imunologia , Chlorocebus aethiops , Modelos Animais de Doenças , Composição de Medicamentos , Feminino , Genoma Viral , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Nanoestruturas , Células Vero , Vacinas Virais/administração & dosagem , Vacinas Virais/química , Vacinas Virais/imunologia , Vacinas de mRNA/química , Vacinas de mRNA/imunologia
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