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Wongsodirdjo P, Caruso AC, Yong AK, Lester MA, Vella LJ, Hung YH, Nisbet RM. Messenger RNA-encoded antibody approach for targeting extracellular and intracellular tau. Brain Commun 2024; 6:fcae100. [PMID: 38585667 PMCID: PMC10996922 DOI: 10.1093/braincomms/fcae100] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2023] [Revised: 02/19/2024] [Accepted: 03/21/2024] [Indexed: 04/09/2024] Open
Abstract
Monoclonal antibodies have emerged as a leading therapeutic agent for the treatment of disease, including Alzheimer's disease. In the last year, two anti-amyloid monoclonal antibodies, lecanemab and aducanumab, have been approved in the USA for the treatment of Alzheimer's disease, whilst several tau-targeting monoclonal antibodies are currently in clinical trials. Such antibodies, however, are expensive and timely to produce and require frequent dosing regimens to ensure disease-modifying effects. Synthetic in vitro-transcribed messenger RNA encoding antibodies for endogenous protein expression holds the potential to overcome many of the limitations associated with protein antibody production. Here, we have generated synthetic in vitro-transcribed messenger RNA encoding a tau-specific antibody as a full-sized immunoglobulin and as a single-chain variable fragment. In vitro transfection of human neuroblastoma SH-SY5Y cells demonstrated the ability of the synthetic messenger RNA to be translated into a functional tau-specific antibody. Furthermore, we show that the translation of the tau-specific single-chain variable fragment as an intrabody results in the specific engagement of intracellular tau. This work highlights the utility of messenger RNA for the delivery of antibody therapeutics, including intrabodies, for the targeting of tau in Alzheimer's disease and other tauopathies.
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Affiliation(s)
- Patricia Wongsodirdjo
- The Florey Institute, Parkville, Victoria 3052, Australia
- Florey Department of Neuroscience and Mental Health, The University of Melbourne, Parkville, Victoria 3052, Australia
| | - Alayna C Caruso
- The Florey Institute, Parkville, Victoria 3052, Australia
- Florey Department of Neuroscience and Mental Health, The University of Melbourne, Parkville, Victoria 3052, Australia
| | - Alicia K Yong
- The Florey Institute, Parkville, Victoria 3052, Australia
| | - Madeleine A Lester
- The Florey Institute, Parkville, Victoria 3052, Australia
- Florey Department of Neuroscience and Mental Health, The University of Melbourne, Parkville, Victoria 3052, Australia
| | - Laura J Vella
- The Florey Institute, Parkville, Victoria 3052, Australia
- Department of Surgery, The Royal Melbourne Hospital, The University of Melbourne, Parkville, Victoria 3052, Australia
| | - Ya Hui Hung
- The Florey Institute, Parkville, Victoria 3052, Australia
- Florey Department of Neuroscience and Mental Health, The University of Melbourne, Parkville, Victoria 3052, Australia
| | - Rebecca M Nisbet
- The Florey Institute, Parkville, Victoria 3052, Australia
- Florey Department of Neuroscience and Mental Health, The University of Melbourne, Parkville, Victoria 3052, Australia
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Mir S, Mir M. The mRNA vaccine, a swift warhead against a moving infectious disease target. Expert Rev Vaccines 2024; 23:336-348. [PMID: 38369742 DOI: 10.1080/14760584.2024.2320327] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2023] [Accepted: 02/14/2024] [Indexed: 02/20/2024]
Abstract
INTRODUCTION The rapid development of mRNA vaccines against SARS-CoV-2 has revolutionized vaccinology, offering hope for swift responses to emerging infectious diseases. Initially met with skepticism, mRNA vaccines have proven effective and safe, reducing vaccine hesitancy amid the evolving COVID-19 pandemic. The COVID-19 pandemic has demonstrated that the time required to modify mRNA vaccines to counter new mutant strains is significantly shorter than the time it takes for pathogens to mutate and generate new variants that can thrive in vaccinated populations. This highlights the notion that mRNA vaccine technology appears to be outpacing viruses in the ongoing evolutionary race. AREAS COVERED This review article offers valuable insights into several crucial aspects of mRNA vaccine development and deployment, including the fundamentals of mRNA vaccine design and synthesis, the utilization of delivery systems, considerations regarding vaccine safety, the longevity of the immune response, strategies for modifying the original mRNA vaccine to address emerging mutant strains, as well as addressing vaccine hesitancy and potential approaches to mitigate reluctance. EXPERT OPINION Challenges such as stability, storage, manufacturing complexities, production capacity, allergic reactions, long-term effects, accessibility, and misinformation must be addressed. Despite these hurdles, mRNA vaccine technology holds promise for revolutionizing future vaccination strategies.
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Affiliation(s)
- Sheema Mir
- College of Veterinary Sciences, Western University of Health Sciences, Pomona, CA, USA
| | - Mohammad Mir
- College of Veterinary Sciences, Western University of Health Sciences, Pomona, CA, USA
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Navarro-Corcuera A, Sehrawat TS, Jalan-Sakrikar N, Gibbons HR, Pirius NE, Khanal S, Hamdan FH, Aseem SO, Cao S, Banales JM, Kang N, Faubion WA, LaRusso NF, Shah VH, Huebert RC. Long non-coding RNA ACTA2-AS1 promotes ductular reaction by interacting with the p300/ELK1 complex. J Hepatol 2022; 76:921-933. [PMID: 34953958 PMCID: PMC8934273 DOI: 10.1016/j.jhep.2021.12.014] [Citation(s) in RCA: 26] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/04/2021] [Revised: 11/01/2021] [Accepted: 12/03/2021] [Indexed: 12/11/2022]
Abstract
BACKGROUND & AIMS Biliary disease is associated with a proliferative/fibrogenic ductular reaction (DR). p300 is an epigenetic regulator that acetylates lysine 27 on histone 3 (H3K27ac) and is activated during fibrosis. Long non-coding RNAs (lncRNAs) are aberrantly expressed in cholangiopathies, but little is known about how they recruit epigenetic complexes and regulate DR. We investigated epigenetic complexes, including transcription factors (TFs) and lncRNAs, contributing to p300-mediated transcription during fibrosis. METHODS We evaluated p300 in vivo using tamoxifen-inducible, cholangiocyte-selective, p300 knockout (KO) coupled with bile duct ligation (BDL) and Mdr KO mice treated with SGC-CBP30. Primary cholangiocytes and liver tissue were analyzed for expression of Acta2-as1 lncRNA by qPCR and RNA in situ hybridization. In vitro, we performed RNA-sequencing in human cholangiocytes with a p300 inhibitor. Cholangiocytes were exposed to lipopolysaccharide (LPS) as an injury model. We confirmed formation of a p300/ELK1 complex by immunoprecipitation (IP). RNA IP was used to examine interactions between ACTA2-AS1 and p300. Chromatin IP assays were used to evaluate p300/ELK1 occupancy and p300-mediated H3K27ac. Organoids were generated from ACTA2-AS1-depleted cholangiocytes. RESULTS BDL-induced DR and fibrosis were reduced in Krt19-CreERT/p300fl/fl mice. Similarly, Mdr KO mice were protected from DR and fibrosis after SGC-CBP30 treatment. In vitro, depletion of ACTA2-AS1 reduced expression of proliferative/fibrogenic markers, reduced LPS-induced cholangiocyte proliferation, and impaired organoid formation. ACTA2-AS1 regulated transcription by facilitating p300/ELK1 binding to the PDGFB promoter after LPS exposure. Correspondingly, LPS-induced H3K27ac was mediated by p300/ELK1 and was reduced in ACTA2-AS1-depleted cholangiocytes. CONCLUSION Cholangiocyte-selective p300 KO or p300 inhibition attenuate DR/fibrosis in mice. ACTA2-AS1 influences recruitment of p300/ELK1 to specific promoters to drive H3K27ac and epigenetic activation of proliferative/fibrogenic genes. This suggests that cooperation between epigenetic co-activators and lncRNAs facilitates DR/fibrosis in biliary diseases. LAY SUMMARY We identified a three-part complex containing an RNA molecule, a transcription factor, and an epigenetic enzyme. The complex is active in injured bile duct cells and contributes to activation of genes involved in proliferation and fibrosis.
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Affiliation(s)
- Amaia Navarro-Corcuera
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Tejasav S Sehrawat
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Nidhi Jalan-Sakrikar
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Hunter R Gibbons
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Nicholas E Pirius
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States
| | - Shalil Khanal
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Feda H Hamdan
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Sayed Obaidullah Aseem
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Sheng Cao
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Jesus M Banales
- Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute, Donostia University Hospital, CIBERehd, Ikerbasque, San Sebastian, Spain
| | - Ningling Kang
- The Hormel Institute, University of Minnesota, Austin, MN, United States
| | - William A Faubion
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Nicholas F LaRusso
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States
| | - Vijay H Shah
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States
| | - Robert C Huebert
- Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, United States; Gastroenterology Research Unit; Mayo Clinic and Foundation, Rochester, MN, United States.
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Recent advancements in lipid–mRNA nanoparticles as a treatment option for cancer immunotherapy. JOURNAL OF PHARMACEUTICAL INVESTIGATION 2022; 52:415-426. [PMID: 35369363 PMCID: PMC8960215 DOI: 10.1007/s40005-022-00569-9] [Citation(s) in RCA: 24] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2021] [Accepted: 03/13/2022] [Indexed: 12/14/2022]
Abstract
Background Cancer remains a serious health concern worldwide, and different approaches are being developed for its treatment. The strategy to use the immune system as an approach for treating cancer has recently gained momentum. Messenger RNA (mRNA) has been assessed as an up-and-coming resource for the evolution of advanced cancer immunotherapies over the past decades. However, degradation in extracellular compartments and during endosomal escape remain obstacles for efficient mRNA delivery and limit the therapeutic applications of this approach. Area covered Lipid-based nanocarriers are gaining significant attention as non-viral mRNA vectors. Various lipid-based nanocarrier types have been developed to enhance the stability of mRNA molecules, facilitate their transfection, and ensure delivery to an intracellular compartment suitable for further processing. This review discusses the development of novel mRNA delivery systems using lipids for effective cancer immunotherapy. Expert opinion mRNAs are superior to other biomolecules for developing therapeutic drugs and vaccines with multiple medical applications that are currently being explored by researchers in various biomedical fields. Lipid-based mRNA nanoparticles can improve the potency of the mRNA by enhancing its stability, enabling its cellular uptake, and facilitating its endosomal escape. Targetability of these therapeutics can be increased by conjugating their surface with the desired ligands or targeting agents. Lipid–mRNA nanoparticles are increasingly being incorporated in cancer immunotherapy applications, including vaccines, monoclonal antibodies, and chimeric antigen receptor T-cell treatment, and several such nanoparticles are being assessed in clinical trials. Further research that assesses key variables for transfection efficiency of lipid–mRNA nanoparticles will expedite the development of improved therapeutics.
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Aune TM, Tossberg JT, Heinrich RM, Porter KP, Crooke PS. Alu RNA Structural Features Modulate Immune Cell Activation and A-to-I Editing of Alu RNAs Is Diminished in Human Inflammatory Bowel Disease. Front Immunol 2022; 13:818023. [PMID: 35126398 PMCID: PMC8813004 DOI: 10.3389/fimmu.2022.818023] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2021] [Accepted: 01/04/2022] [Indexed: 12/12/2022] Open
Abstract
Alu retrotransposons belong to the class of short interspersed nuclear elements (SINEs). Alu RNA is abundant in cells and its repetitive structure forms double-stranded RNAs (dsRNA) that activate dsRNA sensors and trigger innate immune responses with significant pathological consequences. Mechanisms to prevent innate immune activation include deamination of adenosines to inosines in dsRNAs, referred to as A-to-I editing, degradation of Alu RNAs by endoribonucleases, and sequestration of Alu RNAs by RNA binding proteins. We have previously demonstrated that widespread loss of Alu RNA A-to-I editing is associated with diverse human diseases including viral (COVID-19, influenza) and autoimmune diseases (multiple sclerosis). Here we demonstrate loss of A-to-I editing in leukocytes is also associated with inflammatory bowel diseases. Our structure-function analysis demonstrates that ability to activate innate immune responses resides in the left arm of Alu RNA, requires a 5’-PPP, RIG-I is the major Alu dsRNA sensor, and A-to-I editing disrupts both structure and function. Further, edited Alu RNAs inhibit activity of unedited Alu RNAs. Altering Alu RNA nucleotide sequence increases biological activity. Two classes of Alu RNAs exist, one class stimulates both IRF and NF-kB transcriptional activity and a second class only stimulates IRF transcriptional activity. Thus, Alu RNAs play important roles in human disease but may also have therapeutic potential.
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Affiliation(s)
- Thomas M. Aune
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, United States
- Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, TN, United States
- *Correspondence: Thomas M. Aune,
| | - John T. Tossberg
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Rachel M. Heinrich
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Krislyn P. Porter
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, United States
| | - Philip S. Crooke
- Department of Mathematics, Vanderbilt University, Nashville, TN, United States
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Crooke PS, Tossberg JT, Porter KP, Aune TM. Reduced A-to-I editing of endogenous Alu RNAs in lung after SARS-CoV-2 infection. CURRENT RESEARCH IN IMMUNOLOGY 2021; 2:52-59. [PMID: 33969287 PMCID: PMC8084883 DOI: 10.1016/j.crimmu.2021.04.001] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2021] [Revised: 04/19/2021] [Accepted: 04/22/2021] [Indexed: 12/16/2022] Open
Abstract
Due to potential severity of disease caused by SARS-CoV-2 infection, it is critical to understand both mechanisms of viral pathogenesis as well as diversity of host responses to infection. Reduced A-to-I editing of endogenous double-stranded RNAs (dsRNAs), as a result of inactivating mutations in ADAR, produces one form of Aicardi-Goutières Syndrome, with an immune response similar to an anti-viral response. By analyzing whole genome RNA sequencing data, we find reduced levels of A-to-I editing of endogenous Alu RNAs in normal human lung cells after infection by SARS-CoV-2 as well as in lung biopsies from patients with SARS-CoV-2 infections. Unedited Alu RNAs, as seen after infection, induce IRF and NF-kB transcriptional responses and downstream target genes, while edited Alu RNAs as seen in the absence of infection, fail to activate these transcriptional responses. Thus, decreased A-to-I editing may represent an important host response to SARS-CoV-2 infection.
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Affiliation(s)
- Philip S Crooke
- Department of Mathematics, Vanderbilt University, Nashville, TN, 37212, USA
| | - John T Tossberg
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, 37212, USA
| | - Krislyn P Porter
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, 37212, USA
| | - Thomas M Aune
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN, 37212, USA.,Department of Pathology, Microbiology and Immunology, Vanderbilt University Medical Center, Nashville, TN, 37212, USA
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Crooke PS, Tossberg JT, Porter KP, Aune TM. Cutting Edge: Reduced Adenosine-to-Inosine Editing of Endogenous Alu RNAs in Severe COVID-19 Disease. THE JOURNAL OF IMMUNOLOGY 2021; 206:1691-1696. [PMID: 33782089 DOI: 10.4049/jimmunol.2001428] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/29/2020] [Accepted: 02/16/2021] [Indexed: 01/10/2023]
Abstract
Severe COVID-19 disease is associated with elevated inflammatory responses. One form of Aicardi-Goutières syndrome caused by inactivating mutations in ADAR results in reduced adenosine-to-inosine (A-to-I) editing of endogenous dsRNAs, induction of IFNs, IFN-stimulated genes, other inflammatory mediators, morbidity, and mortality. Alu elements, ∼10% of the human genome, are the most common A-to-I-editing sites. Using leukocyte whole-genome RNA-sequencing data, we found reduced A-to-I editing of Alu dsRNAs in patients with severe COVID-19 disease. Dendritic cells infected with COVID-19 also exhibit reduced A-to-I editing of Alu dsRNAs. Unedited Alu dsRNAs, but not edited Alu dsRNAs, are potent inducers of IRF and NF-κB transcriptional responses, IL6, IL8, and IFN-stimulated genes. Thus, decreased A-to-I editing that may lead to accumulation of unedited Alu dsRNAs and increased inflammatory responses is associated with severe COVID-19 disease.
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Affiliation(s)
- Philip S Crooke
- Department of Mathematics, Vanderbilt University, Nashville, TN 37212
| | - John T Tossberg
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37212; and
| | - Krislyn P Porter
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37212; and
| | - Thomas M Aune
- Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37212; and .,Department of Pathology, Microbiology and Immunology, Vanderbilt University, Nashville, TN 37212
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