Pub Date : 2024-08-07DOI: 10.1126/scitranslmed.abo4736
Emma van Groesen, Elma Mons, Ioli Kotsogianni, Melina Arts, Kamaleddin H. M. E. Tehrani, Nicola Wade, Vladyslav Lysenko, Floor M. Stel, Jordy T. Zwerus, Stefania De Benedetti, Alexander Bakker, Parichita Chakraborty, Mario van der Stelt, Dirk-Jan Scheffers, Jairo Gooskens, Wiep Klaas Smits, Kirsty Holden, Peter S. Gilmour, Joost Willemse, Christopher A. Hitchcock, J. G. Coen van Hasselt, Tanja Schneider, Nathaniel I. Martin
Gram-positive bacterial infections present a major clinical challenge, with methicillin- and vancomycin-resistant strains continuing to be a cause for concern. In recent years, semisynthetic vancomycin derivatives have been developed to overcome this problem as exemplified by the clinically used telavancin, which exhibits increased antibacterial potency but has also raised toxicity concerns. Thus, glycopeptide antibiotics with enhanced antibacterial activities and improved safety profiles are still necessary. We describe the development of a class of highly potent semisynthetic glycopeptide antibiotics, the guanidino lipoglycopeptides, which contain a positively charged guanidino moiety bearing a variable lipid group. These glycopeptides exhibited enhanced in vitro activity against a panel of Gram-positive bacteria including clinically relevant methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant strains, showed minimal toxicity toward eukaryotic cells, and had a low propensity for resistance selection. Mechanistically, guanidino lipoglycopeptides engaged with bacterial cell wall precursor lipid II with a higher binding affinity than vancomycin. Binding to both wild-type d-Ala-d-Ala lipid II and the vancomycin-resistant d-Ala-d-Lac variant was confirmed, providing insight into the enhanced activity of guanidino lipoglycopeptides against vancomycin-resistant isolates. The in vivo efficacy of guanidino lipoglycopeptide EVG7 was evaluated in a S. aureus murine thigh infection model and a 7-day sepsis survival study, both of which demonstrated superiority to vancomycin. Moreover, the minimal to mild kidney effects at supratherapeutic doses of EVG7 indicate an improved therapeutic safety profile compared with vancomycin. These findings position guanidino lipoglycopeptides as candidates for further development as antibacterial agents for the treatment of clinically relevant multidrug-resistant Gram-positive infections.
革兰氏阳性细菌感染是一项重大的临床挑战，耐甲氧西林和耐万古霉素菌株一直是令人担忧的问题。近年来，人们开发了半合成万古霉素衍生物来解决这一问题，临床上使用的泰拉万星就是一例，它具有更强的抗菌效力，但也引发了毒性问题。因此，仍然需要抗菌活性更强、安全性更好的糖肽类抗生素。我们介绍了一类高效半合成糖肽抗生素--胍基脂甘肽的开发情况，这种抗生素含有一个带正电荷的胍基，并带有一个可变的脂基。这些糖肽对一系列革兰氏阳性细菌（包括临床相关的耐甲氧西林金黄色葡萄球菌（MRSA）和耐万古霉素菌株）具有更强的体外活性，对真核细胞的毒性极小，而且耐药性选择倾向低。从机理上讲，胍基脂甘肽与细菌细胞壁前体脂质 II 的结合亲和力高于万古霉素。与野生型 d-Ala-d-Ala 脂质 II 和对万古霉素耐药的 d-Ala-d-Lac 变体的结合均得到了证实，这为胍基聚糖肽增强对万古霉素耐药分离菌的活性提供了启示。在金黄色葡萄球菌小鼠大腿感染模型和 7 天败血症存活研究中评估了胍基脂多糖肽 EVG7 的体内疗效，结果均显示其优于万古霉素。此外，与万古霉素相比，EVG7 在超治疗剂量时对肾脏的影响极小至轻微，这表明其治疗安全性有所提高。这些研究结果表明，胍基脂甘肽可作为抗菌剂进一步开发，用于治疗临床相关的耐多药革兰氏阳性感染。
{"title":"Semisynthetic guanidino lipoglycopeptides with potent in vitro and in vivo antibacterial activity","authors":"Emma van Groesen, Elma Mons, Ioli Kotsogianni, Melina Arts, Kamaleddin H. M. E. Tehrani, Nicola Wade, Vladyslav Lysenko, Floor M. Stel, Jordy T. Zwerus, Stefania De Benedetti, Alexander Bakker, Parichita Chakraborty, Mario van der Stelt, Dirk-Jan Scheffers, Jairo Gooskens, Wiep Klaas Smits, Kirsty Holden, Peter S. Gilmour, Joost Willemse, Christopher A. Hitchcock, J. G. Coen van Hasselt, Tanja Schneider, Nathaniel I. Martin","doi":"10.1126/scitranslmed.abo4736","DOIUrl":"10.1126/scitranslmed.abo4736","url":null,"abstract":"<div >Gram-positive bacterial infections present a major clinical challenge, with methicillin- and vancomycin-resistant strains continuing to be a cause for concern. In recent years, semisynthetic vancomycin derivatives have been developed to overcome this problem as exemplified by the clinically used telavancin, which exhibits increased antibacterial potency but has also raised toxicity concerns. Thus, glycopeptide antibiotics with enhanced antibacterial activities and improved safety profiles are still necessary. We describe the development of a class of highly potent semisynthetic glycopeptide antibiotics, the guanidino lipoglycopeptides, which contain a positively charged guanidino moiety bearing a variable lipid group. These glycopeptides exhibited enhanced in vitro activity against a panel of Gram-positive bacteria including clinically relevant methicillin-resistant <i>Staphylococcus aureus</i> (MRSA) and vancomycin-resistant strains, showed minimal toxicity toward eukaryotic cells, and had a low propensity for resistance selection. Mechanistically, guanidino lipoglycopeptides engaged with bacterial cell wall precursor lipid II with a higher binding affinity than vancomycin. Binding to both wild-type <span>d</span>-Ala-<span>d</span>-Ala lipid II and the vancomycin-resistant <span>d</span>-Ala-<span>d</span>-Lac variant was confirmed, providing insight into the enhanced activity of guanidino lipoglycopeptides against vancomycin-resistant isolates. The in vivo efficacy of guanidino lipoglycopeptide EVG7 was evaluated in a <i>S. aureus</i> murine thigh infection model and a 7-day sepsis survival study, both of which demonstrated superiority to vancomycin. Moreover, the minimal to mild kidney effects at supratherapeutic doses of EVG7 indicate an improved therapeutic safety profile compared with vancomycin. These findings position guanidino lipoglycopeptides as candidates for further development as antibacterial agents for the treatment of clinically relevant multidrug-resistant Gram-positive infections.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 759","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141902814","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-07DOI: 10.1126/scitranslmed.adi1625
Nikole L. Warner, Jacob Archer, Stephanie Park, Garima Singh, Kathryn M. McFadden, Taishi Kimura, Katrina Nicholes, Adrian Simpson, Jason T. Kaelber, David W. Hawman, Heinz Feldmann, Amit P. Khandhar, Peter Berglund, Matthew R. Vogt, Jesse H. Erasmus
The recent emergence and rapid response to severe acute respiratory syndrome coronavirus 2 was enabled by prototype pathogen and vaccine platform approaches, driven by the preemptive application of RNA vaccine technology to the related Middle East respiratory syndrome coronavirus. Recently, the National Institutes of Allergy and Infectious Diseases identified nine virus families of concern, eight enveloped virus families and one nonenveloped virus family, for which vaccine generation is a priority. Although RNA vaccines have been described for a variety of enveloped viruses, a roadmap for their use against nonenveloped viruses is lacking. Enterovirus D68 was recently designated a prototype pathogen within the family Picornaviridae of nonenveloped viruses because of its rapid evolution and respiratory route of transmission, coupled with a lack of diverse anti-enterovirus vaccine approaches in development. Here, we describe a proof-of-concept approach using a clinical stage RNA vaccine platform that induced robust enterovirus D68–neutralizing antibody responses in mice and nonhuman primates and prevented upper and lower respiratory tract infections and neurological disease in mice. In addition, we used our platform to rapidly characterize the antigenic diversity within the six genotypes of enterovirus D68, providing the necessary data to inform multivalent vaccine compositions that can elicit optimal breadth of neutralizing responses. These results demonstrate that RNA vaccines can be used as tools in our pandemic-preparedness toolbox for nonenveloped viruses.
{"title":"A self-amplifying RNA vaccine prevents enterovirus D68 infection and disease in preclinical models","authors":"Nikole L. Warner, Jacob Archer, Stephanie Park, Garima Singh, Kathryn M. McFadden, Taishi Kimura, Katrina Nicholes, Adrian Simpson, Jason T. Kaelber, David W. Hawman, Heinz Feldmann, Amit P. Khandhar, Peter Berglund, Matthew R. Vogt, Jesse H. Erasmus","doi":"10.1126/scitranslmed.adi1625","DOIUrl":"10.1126/scitranslmed.adi1625","url":null,"abstract":"<div >The recent emergence and rapid response to severe acute respiratory syndrome coronavirus 2 was enabled by prototype pathogen and vaccine platform approaches, driven by the preemptive application of RNA vaccine technology to the related Middle East respiratory syndrome coronavirus. Recently, the National Institutes of Allergy and Infectious Diseases identified nine virus families of concern, eight enveloped virus families and one nonenveloped virus family, for which vaccine generation is a priority. Although RNA vaccines have been described for a variety of enveloped viruses, a roadmap for their use against nonenveloped viruses is lacking. Enterovirus D68 was recently designated a prototype pathogen within the family Picornaviridae of nonenveloped viruses because of its rapid evolution and respiratory route of transmission, coupled with a lack of diverse anti-enterovirus vaccine approaches in development. Here, we describe a proof-of-concept approach using a clinical stage RNA vaccine platform that induced robust enterovirus D68–neutralizing antibody responses in mice and nonhuman primates and prevented upper and lower respiratory tract infections and neurological disease in mice. In addition, we used our platform to rapidly characterize the antigenic diversity within the six genotypes of enterovirus D68, providing the necessary data to inform multivalent vaccine compositions that can elicit optimal breadth of neutralizing responses. These results demonstrate that RNA vaccines can be used as tools in our pandemic-preparedness toolbox for nonenveloped viruses.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 759","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141902810","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-07DOI: 10.1126/scitranslmed.adn2140
Marisa Zallocchi, Sarath Vijayakumar, Jonathan Fleegel, Lyudmila Batalkina, Katyarina E. Brunette, Dhaval Shukal, Zhiyong Chen, Olivier Devuyst, Huizhan Liu, David Z. Z. He, Ali Sajid Imami, Abdul-Rizaq Ali Hamoud, Robert McCullumsmith, Martin Conda-Sheridan, Luana Janaína De Campos, Jian Zuo
Hearing loss is a major health concern in our society, affecting more than 400 million people worldwide. Among the causes, aminoglycoside therapy can result in permanent hearing loss in 40% to 60% of patients receiving treatment, and despite these high numbers, no drug for preventing or treating this type of hearing loss has yet been approved by the US Food and Drug Administration. We have previously conducted high-throughput screenings of bioactive compounds, using zebrafish as our discovery platform, and identified piplartine as a potential therapeutic molecule. In the present study, we expanded this work and characterized piplartine’s physicochemical and therapeutic properties. We showed that piplartine had a wide therapeutic window and neither induced nephrotoxicity in vivo in zebrafish nor interfered with aminoglycoside antibacterial activity. In addition, a fluorescence-based assay demonstrated that piplartine did not inhibit cytochrome C activity in microsomes. Coadministration of piplartine protected from kanamycin-induced hair cell loss in zebrafish and protected hearing function, outer hair cells, and presynaptic ribbons in a mouse model of kanamycin ototoxicity. Last, we investigated piplartine’s mechanism of action by phospho-omics, immunoblotting, immunohistochemistry, and molecular dynamics experiments. We found an up-regulation of AKT1 signaling in the cochleas of mice cotreated with piplartine. Piplartine treatment normalized kanamycin-induced up-regulation of TRPV1 expression and modulated the gating properties of this receptor. Because aminoglycoside entrance to the inner ear is, in part, mediated by TRPV1, these results suggested that by regulating TRPV1 expression, piplartine blocked aminoglycoside’s entrance, thereby preventing the long-term deleterious effects of aminoglycoside accumulation in the inner ear compartment.
{"title":"Piplartine attenuates aminoglycoside-induced TRPV1 activity and protects from hearing loss in mice","authors":"Marisa Zallocchi, Sarath Vijayakumar, Jonathan Fleegel, Lyudmila Batalkina, Katyarina E. Brunette, Dhaval Shukal, Zhiyong Chen, Olivier Devuyst, Huizhan Liu, David Z. Z. He, Ali Sajid Imami, Abdul-Rizaq Ali Hamoud, Robert McCullumsmith, Martin Conda-Sheridan, Luana Janaína De Campos, Jian Zuo","doi":"10.1126/scitranslmed.adn2140","DOIUrl":"10.1126/scitranslmed.adn2140","url":null,"abstract":"<div >Hearing loss is a major health concern in our society, affecting more than 400 million people worldwide. Among the causes, aminoglycoside therapy can result in permanent hearing loss in 40% to 60% of patients receiving treatment, and despite these high numbers, no drug for preventing or treating this type of hearing loss has yet been approved by the US Food and Drug Administration. We have previously conducted high-throughput screenings of bioactive compounds, using zebrafish as our discovery platform, and identified piplartine as a potential therapeutic molecule. In the present study, we expanded this work and characterized piplartine’s physicochemical and therapeutic properties. We showed that piplartine had a wide therapeutic window and neither induced nephrotoxicity in vivo in zebrafish nor interfered with aminoglycoside antibacterial activity. In addition, a fluorescence-based assay demonstrated that piplartine did not inhibit cytochrome C activity in microsomes. Coadministration of piplartine protected from kanamycin-induced hair cell loss in zebrafish and protected hearing function, outer hair cells, and presynaptic ribbons in a mouse model of kanamycin ototoxicity. Last, we investigated piplartine’s mechanism of action by phospho-omics, immunoblotting, immunohistochemistry, and molecular dynamics experiments. We found an up-regulation of AKT1 signaling in the cochleas of mice cotreated with piplartine. Piplartine treatment normalized kanamycin-induced up-regulation of TRPV1 expression and modulated the gating properties of this receptor. Because aminoglycoside entrance to the inner ear is, in part, mediated by TRPV1, these results suggested that by regulating TRPV1 expression, piplartine blocked aminoglycoside’s entrance, thereby preventing the long-term deleterious effects of aminoglycoside accumulation in the inner ear compartment.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 759","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141902813","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-07DOI: 10.1126/scitranslmed.adg1915
Prashanth Thevkar Nagesh, Yeonhee Cho, Yuan Zhuang, Mrigya Babuta, Marti Ortega-Ribera, Radhika Joshi, Veronika Brezani, Arman Patel, Aditi Ashish Datta, Viliam Brezani, Yun-Cheng Hsieh, Adriana Ramos, Jeeval Mehta, Christopher Copeland, Eleni Kanata, Zhenghui Gordon Jiang, Ioannis Vlachos, John Asara, AlcHepNet Consortium, Gyongyi Szabo
Severe alcohol-associated hepatitis (AH) is a life-threatening form of alcohol-associated liver disease. Liver neutrophil infiltration is a hallmark of AH, yet the effects of alcohol on neutrophil functions remain elusive. Identifying therapeutic targets to reduce neutrophil-mediated liver damage is essential. Bruton’s tyrosine kinase (BTK) plays an important role in neutrophil development and function; however, the role of BTK in AH is unknown. Using RNA sequencing of circulating neutrophils, we found an increase in Btk expression (P = 0.05) and phosphorylated BTK (pBTK) in patients with AH compared with healthy controls. In vitro, physiologically relevant doses of alcohol resulted in a rapid, TLR4-mediated induction of pBTK in neutrophils. In a preclinical model of AH, administration of a small-molecule BTK inhibitor (evobrutinib) or myeloid-specific Btk knockout decreased proinflammatory cytokines and attenuated neutrophil-mediated liver damage. We found that pBTK was essential for alcohol-induced bone marrow granulopoiesis and liver neutrophil infiltration. In vivo, BTK inhibition or myeloid-specific Btk knockout reduced granulopoiesis, circulating neutrophils, liver neutrophil infiltration, and liver damage in a mouse model of AH. Mechanistically, using liquid chromatography–tandem mass spectrometry, we identified CD84 as a kinase target of BTK, which is involved in granulopoiesis. In vitro, CD84 promoted alcohol-induced interleukin-1β and tumor necrosis factor–α in primary human neutrophils, which was inhibited by CD84-blocking antibody treatment. Our findings define the role of BTK and CD84 in regulating neutrophil inflammation and granulopoiesis, with potential therapeutic implications in AH.
{"title":"In vivo Bruton’s tyrosine kinase inhibition attenuates alcohol-associated liver disease by regulating CD84-mediated granulopoiesis","authors":"Prashanth Thevkar Nagesh, Yeonhee Cho, Yuan Zhuang, Mrigya Babuta, Marti Ortega-Ribera, Radhika Joshi, Veronika Brezani, Arman Patel, Aditi Ashish Datta, Viliam Brezani, Yun-Cheng Hsieh, Adriana Ramos, Jeeval Mehta, Christopher Copeland, Eleni Kanata, Zhenghui Gordon Jiang, Ioannis Vlachos, John Asara, AlcHepNet Consortium, Gyongyi Szabo","doi":"10.1126/scitranslmed.adg1915","DOIUrl":"10.1126/scitranslmed.adg1915","url":null,"abstract":"<div >Severe alcohol-associated hepatitis (AH) is a life-threatening form of alcohol-associated liver disease. Liver neutrophil infiltration is a hallmark of AH, yet the effects of alcohol on neutrophil functions remain elusive. Identifying therapeutic targets to reduce neutrophil-mediated liver damage is essential. Bruton’s tyrosine kinase (BTK) plays an important role in neutrophil development and function; however, the role of BTK in AH is unknown. Using RNA sequencing of circulating neutrophils, we found an increase in <i><i>Btk</i></i> expression (<i>P</i> = 0.05) and phosphorylated BTK (pBTK) in patients with AH compared with healthy controls. In vitro, physiologically relevant doses of alcohol resulted in a rapid, TLR4-mediated induction of pBTK in neutrophils. In a preclinical model of AH, administration of a small-molecule BTK inhibitor (evobrutinib) or myeloid-specific <i>Btk</i> knockout decreased proinflammatory cytokines and attenuated neutrophil-mediated liver damage. We found that pBTK was essential for alcohol-induced bone marrow granulopoiesis and liver neutrophil infiltration. In vivo, BTK inhibition or myeloid-specific <i>Btk</i> knockout reduced granulopoiesis, circulating neutrophils, liver neutrophil infiltration, and liver damage in a mouse model of AH. Mechanistically, using liquid chromatography–tandem mass spectrometry, we identified CD84 as a kinase target of BTK, which is involved in granulopoiesis. In vitro, CD84 promoted alcohol-induced interleukin-1β and tumor necrosis factor–α in primary human neutrophils, which was inhibited by CD84-blocking antibody treatment. Our findings define the role of BTK and CD84 in regulating neutrophil inflammation and granulopoiesis, with potential therapeutic implications in AH.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 759","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.science.org/doi/reader/10.1126/scitranslmed.adg1915","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141902812","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-07DOI: 10.1126/scitranslmed.adi4830
Jihoon Han, Jae Hoon Sul, Jeongmi Lee, Eunae Kim, Hark Kyun Kim, Minshik Chae, Jeein Lim, Jongho Kim, Chanhee Kim, Jun-Sik Kim, Yoonsuk Cho, Jae Hyung Park, Yong Woo Cho, Dong-Gyu Jo
Effective intracellular delivery of therapeutic proteins can potentially treat a wide array of diseases. However, efficient delivery of functional proteins across the cell membrane remains challenging. Exosomes are nanosized vesicles naturally secreted by various types of cells and may serve as promising nanocarriers for therapeutic biomolecules. Here, we engineered exosomes equipped with a photoinducible cargo protein release system, termed mMaple3-mediated protein loading into and release from exosome (MAPLEX), in which cargo proteins can be loaded into the exosomes by fusing them with photocleavable protein (mMaple3)–conjugated exosomal membrane markers and subsequently released from the exosomal membrane by inducing photocleavage with blue light illumination. Using this system, we first induced transcriptional regulation by delivering octamer-binding transcription factor 4 and SRY-box transcription factor 2 to fibroblasts in vitro. Second, we induced in vivo gene recombination in Cre reporter mice by delivering Cre recombinase. Last, we achieved targeted epigenome editing in the brains of 5xFAD and 3xTg-AD mice, two models of Alzheimer’s disease. Administration of MAPLEXs loaded with β-site amyloid precursor protein cleaving enzyme 1 (Bace1)–targeting single guide RNA–incorporated dCas9 ribonucleoprotein complexes, coupled with the catalytic domain of DNA methyltransferase 3A, resulted in successful methylation of the targeted CpG sites within the Bace1 promoter. This approach led to a significant reduction in Bace1 expression, improved recognition memory impairment, and reduced amyloid pathology in 5xFAD and 3xTg-AD mice. These results suggest that MAPLEX is an efficient intracellular protein delivery system that can deliver diverse therapeutic proteins for multiple diseases.
{"title":"Engineered exosomes with a photoinducible protein delivery system enable CRISPR-Cas–based epigenome editing in Alzheimer’s disease","authors":"Jihoon Han, Jae Hoon Sul, Jeongmi Lee, Eunae Kim, Hark Kyun Kim, Minshik Chae, Jeein Lim, Jongho Kim, Chanhee Kim, Jun-Sik Kim, Yoonsuk Cho, Jae Hyung Park, Yong Woo Cho, Dong-Gyu Jo","doi":"10.1126/scitranslmed.adi4830","DOIUrl":"10.1126/scitranslmed.adi4830","url":null,"abstract":"<div >Effective intracellular delivery of therapeutic proteins can potentially treat a wide array of diseases. However, efficient delivery of functional proteins across the cell membrane remains challenging. Exosomes are nanosized vesicles naturally secreted by various types of cells and may serve as promising nanocarriers for therapeutic biomolecules. Here, we engineered exosomes equipped with a photoinducible cargo protein release system, termed mMaple3-mediated protein loading into and release from exosome (MAPLEX), in which cargo proteins can be loaded into the exosomes by fusing them with photocleavable protein (mMaple3)–conjugated exosomal membrane markers and subsequently released from the exosomal membrane by inducing photocleavage with blue light illumination. Using this system, we first induced transcriptional regulation by delivering octamer-binding transcription factor 4 and SRY-box transcription factor 2 to fibroblasts in vitro. Second, we induced in vivo gene recombination in Cre reporter mice by delivering Cre recombinase. Last, we achieved targeted epigenome editing in the brains of 5xFAD and 3xTg-AD mice, two models of Alzheimer’s disease. Administration of MAPLEXs loaded with β-site amyloid precursor protein cleaving enzyme 1 (<i>Bace1</i>)–targeting single guide RNA–incorporated dCas9 ribonucleoprotein complexes, coupled with the catalytic domain of DNA methyltransferase 3A, resulted in successful methylation of the targeted CpG sites within the <i>Bace1</i> promoter. This approach led to a significant reduction in <i>Bace1</i> expression, improved recognition memory impairment, and reduced amyloid pathology in 5xFAD and 3xTg-AD mice. These results suggest that MAPLEX is an efficient intracellular protein delivery system that can deliver diverse therapeutic proteins for multiple diseases.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 759","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141902811","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-31DOI: 10.1126/scitranslmed.adk4769
Allyson N. X. Choi, Tanamas Siriphanitchakorn, Milly M. Choy, Justin S. G. Ooi, Menchie Manuel, Hwee Cheng Tan, Lowell Z. Lin, Xin Yap, Duane J. Gubler, Eng Eong Ooi
Dengue viruses (DENVs), like all viruses, evolve to perpetuate transmission of their species in their hosts. However, how DENV genetics influences dengue disease outbreaks remains poorly understood. Here, we examined isolates of the South Pacific dengue virus type 2 (DENV-2) that emerged in the 1970s and caused major dengue outbreaks in islands in this region until it reached Tonga, where only a few mild cases were reported. Phylogenetically, the DENV-2 strain isolated in Tonga segregated into a clade different from those clades infecting populations in other South Pacific islands. We found that this epidemiological observation could be explained by a single histidine-to-arginine substitution in position 86 of the premembrane (prM) protein of the Tonga DENV-2 strain. This mutation attenuated viral protein translation in mammalian cells but not in midgut cells of the mosquito vector Aedes aegypti. In mammalian cells, the prM mutation resulted in reduced translation of the viral genome and subsequent reduced virus replication. In contrast, in mosquito midgut cells, the prM mutation conferred a selective infection advantage, possibly because of the positively charged arginine residue introduced by the mutation. These findings provide molecular insights into the year-long silent transmission of attenuated DENV-2 in Tonga during the 1970s dengue outbreak in the South Pacific.
{"title":"A prM mutation that attenuates dengue virus replication in human cells enhances midgut infection in mosquitoes","authors":"Allyson N. X. Choi, Tanamas Siriphanitchakorn, Milly M. Choy, Justin S. G. Ooi, Menchie Manuel, Hwee Cheng Tan, Lowell Z. Lin, Xin Yap, Duane J. Gubler, Eng Eong Ooi","doi":"10.1126/scitranslmed.adk4769","DOIUrl":"10.1126/scitranslmed.adk4769","url":null,"abstract":"<div >Dengue viruses (DENVs), like all viruses, evolve to perpetuate transmission of their species in their hosts. However, how DENV genetics influences dengue disease outbreaks remains poorly understood. Here, we examined isolates of the South Pacific dengue virus type 2 (DENV-2) that emerged in the 1970s and caused major dengue outbreaks in islands in this region until it reached Tonga, where only a few mild cases were reported. Phylogenetically, the DENV-2 strain isolated in Tonga segregated into a clade different from those clades infecting populations in other South Pacific islands. We found that this epidemiological observation could be explained by a single histidine-to-arginine substitution in position 86 of the premembrane (prM) protein of the Tonga DENV-2 strain. This mutation attenuated viral protein translation in mammalian cells but not in midgut cells of the mosquito vector <i>Aedes aegypti</i>. In mammalian cells, the prM mutation resulted in reduced translation of the viral genome and subsequent reduced virus replication. In contrast, in mosquito midgut cells, the prM mutation conferred a selective infection advantage, possibly because of the positively charged arginine residue introduced by the mutation. These findings provide molecular insights into the year-long silent transmission of attenuated DENV-2 in Tonga during the 1970s dengue outbreak in the South Pacific.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 758","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860759","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-31DOI: 10.1126/scitranslmed.abq5585
Shajedul Islam, Frederico O. Gleber-Netto, Collin F. Mulcahy, Mica D. E. Glaun, Snigdha Srivastava, Patrick J. Hunt, Michelle D. Williams, Carly E. Barbon, Michael Spiotto, Weilu Zhao, Adewale Adebayo, Shamima Akhter, Tongxin Xie, Kala Chand Debnath, Hinduja Naidu Sathishkumar, Blake Myers, Sahana Lothumalla, Ismail Yaman, Jared K. Burks, Javier Gomez, Xiayu Rao, Jing Wang, Karin Woodman, Jobran Mansour, Benjamin Arenkiel, Kate L. Osman, Chandler Haxton, Teresa E. Lever, Katherine A. Hutcheson, Moran Amit
The incidence of human papilloma virus–mediated oropharyngeal squamous cell carcinoma (OPSCC) has increased over the past 40 years, particularly among young individuals with a favorable prognosis; however, current therapy often leads to unfortunate side effects, such as dysphagia. Despite the emphasis on dysphagia in previous studies, there is an important research gap in understanding the correlation between neuronal changes and patient-reported and functional outcomes in patients with OPSCC. To address this issue, we examined pathologic tissue samples from patients with OPSCC using multiplex immunofluorescence staining and machine learning to correlate tumor-associated neuronal changes with prospectively collected patient-reported and functional outcomes. We found that tumor enrichment of adrenergic (TH+) and CGRP+ sensory–afferent nerves correlated with poorer swallowing outcomes. Functional electromyography recordings showed correlations between growing (GAP43+) and immature cholinergic (ChAT+DCX+) nerves and denervation patterns in survivors of OPSCC. A murine model of radiation-induced dysphagia further confirmed that immature cholinergic and CGRP+ nerves were correlated with impaired swallowing. Preclinical interventional studies also supported the independent contributions of CGRP+ and cholinergic (ChAT+) nerves to dysphagia in treated mouse models of OPSCC. Our results suggest that CGRP+ and ChAT+ neuronal signaling play distinct roles in tumor- and radiation-induced dysphagia in OPSCC and offer a comprehensive dataset on the neural landscape of OPSCC. These insights may guide early interventions for swallow preservation and the repurposing of neurology-related drugs, such as CGRP blockers, in clinical oncology and survivorship.
{"title":"Neural landscape is associated with functional outcomes in irradiated patients with oropharyngeal squamous cell carcinoma","authors":"Shajedul Islam, Frederico O. Gleber-Netto, Collin F. Mulcahy, Mica D. E. Glaun, Snigdha Srivastava, Patrick J. Hunt, Michelle D. Williams, Carly E. Barbon, Michael Spiotto, Weilu Zhao, Adewale Adebayo, Shamima Akhter, Tongxin Xie, Kala Chand Debnath, Hinduja Naidu Sathishkumar, Blake Myers, Sahana Lothumalla, Ismail Yaman, Jared K. Burks, Javier Gomez, Xiayu Rao, Jing Wang, Karin Woodman, Jobran Mansour, Benjamin Arenkiel, Kate L. Osman, Chandler Haxton, Teresa E. Lever, Katherine A. Hutcheson, Moran Amit","doi":"10.1126/scitranslmed.abq5585","DOIUrl":"10.1126/scitranslmed.abq5585","url":null,"abstract":"<div >The incidence of human papilloma virus–mediated oropharyngeal squamous cell carcinoma (OPSCC) has increased over the past 40 years, particularly among young individuals with a favorable prognosis; however, current therapy often leads to unfortunate side effects, such as dysphagia. Despite the emphasis on dysphagia in previous studies, there is an important research gap in understanding the correlation between neuronal changes and patient-reported and functional outcomes in patients with OPSCC. To address this issue, we examined pathologic tissue samples from patients with OPSCC using multiplex immunofluorescence staining and machine learning to correlate tumor-associated neuronal changes with prospectively collected patient-reported and functional outcomes. We found that tumor enrichment of adrenergic (TH<sup>+</sup>) and CGRP<sup>+</sup> sensory–afferent nerves correlated with poorer swallowing outcomes. Functional electromyography recordings showed correlations between growing (GAP43<sup>+</sup>) and immature cholinergic (ChAT<sup>+</sup>DCX<sup>+</sup>) nerves and denervation patterns in survivors of OPSCC. A murine model of radiation-induced dysphagia further confirmed that immature cholinergic and CGRP<sup>+</sup> nerves were correlated with impaired swallowing. Preclinical interventional studies also supported the independent contributions of CGRP<sup>+</sup> and cholinergic (ChAT<sup>+</sup>) nerves to dysphagia in treated mouse models of OPSCC. Our results suggest that CGRP<sup>+</sup> and ChAT<sup>+</sup> neuronal signaling play distinct roles in tumor- and radiation-induced dysphagia in OPSCC and offer a comprehensive dataset on the neural landscape of OPSCC. These insights may guide early interventions for swallow preservation and the repurposing of neurology-related drugs, such as CGRP blockers, in clinical oncology and survivorship.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 758","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860820","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-31DOI: 10.1126/scitranslmed.adg3894
Marloes van den Berg, Zhonghua Shi, Wout J. Claassen, Pleuni Hooijman, Christopher T. A. Lewis, Jesper L. Andersen, Robbert J. van der Pijl, Sylvia J. P. Bogaards, Stefan Conijn, Eva L. Peters, Leon P. L. Begthel, Bas Uijterwijk, Johan Lindqvist, Paul R. Langlais, Armand R. J. Girbes, Sandra Stapel, Henk Granzier, Kenneth S. Campbell, Weikang Ma, Thomas Irving, Darren T. Hwee, James J. Hartman, Fady I. Malik, Marinus Paul, Albertus Beishuizen, Julien Ochala, Leo Heunks, Coen A. C. Ottenheijm
Patients receiving mechanical ventilation in the intensive care unit (ICU) frequently develop contractile weakness of the diaphragm. Consequently, they may experience difficulty weaning from mechanical ventilation, which increases mortality and poses a high economic burden. Because of a lack of knowledge regarding the molecular changes in the diaphragm, no treatment is currently available to improve diaphragm contractility. We compared diaphragm biopsies from ventilated ICU patients (N = 54) to those of non-ICU patients undergoing thoracic surgery (N = 27). By integrating data from myofiber force measurements, x-ray diffraction experiments, and biochemical assays with clinical data, we found that in myofibers isolated from the diaphragm of ventilated ICU patients, myosin is trapped in an energy-sparing, super-relaxed state, which impairs the binding of myosin to actin during diaphragm contraction. Studies on quadriceps biopsies of ICU patients and on the diaphragm of previously healthy mechanically ventilated rats suggested that the super-relaxed myosins are specific to the diaphragm and not a result of critical illness. Exposing slow- and fast-twitch myofibers isolated from the diaphragm biopsies to small-molecule compounds activating troponin restored contractile force in vitro. These findings support the continued development of drugs that target sarcomere proteins to increase the calcium sensitivity of myofibers for the treatment of ICU-acquired diaphragm weakness.
{"title":"Super-relaxed myosins contribute to respiratory muscle hibernation in mechanically ventilated patients","authors":"Marloes van den Berg, Zhonghua Shi, Wout J. Claassen, Pleuni Hooijman, Christopher T. A. Lewis, Jesper L. Andersen, Robbert J. van der Pijl, Sylvia J. P. Bogaards, Stefan Conijn, Eva L. Peters, Leon P. L. Begthel, Bas Uijterwijk, Johan Lindqvist, Paul R. Langlais, Armand R. J. Girbes, Sandra Stapel, Henk Granzier, Kenneth S. Campbell, Weikang Ma, Thomas Irving, Darren T. Hwee, James J. Hartman, Fady I. Malik, Marinus Paul, Albertus Beishuizen, Julien Ochala, Leo Heunks, Coen A. C. Ottenheijm","doi":"10.1126/scitranslmed.adg3894","DOIUrl":"10.1126/scitranslmed.adg3894","url":null,"abstract":"<div >Patients receiving mechanical ventilation in the intensive care unit (ICU) frequently develop contractile weakness of the diaphragm. Consequently, they may experience difficulty weaning from mechanical ventilation, which increases mortality and poses a high economic burden. Because of a lack of knowledge regarding the molecular changes in the diaphragm, no treatment is currently available to improve diaphragm contractility. We compared diaphragm biopsies from ventilated ICU patients (<i>N</i> = 54) to those of non-ICU patients undergoing thoracic surgery (<i>N</i> = 27). By integrating data from myofiber force measurements, x-ray diffraction experiments, and biochemical assays with clinical data, we found that in myofibers isolated from the diaphragm of ventilated ICU patients, myosin is trapped in an energy-sparing, super-relaxed state, which impairs the binding of myosin to actin during diaphragm contraction. Studies on quadriceps biopsies of ICU patients and on the diaphragm of previously healthy mechanically ventilated rats suggested that the super-relaxed myosins are specific to the diaphragm and not a result of critical illness. Exposing slow- and fast-twitch myofibers isolated from the diaphragm biopsies to small-molecule compounds activating troponin restored contractile force in vitro. These findings support the continued development of drugs that target sarcomere proteins to increase the calcium sensitivity of myofibers for the treatment of ICU-acquired diaphragm weakness.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 758","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860821","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-31DOI: 10.1126/scitranslmed.adn6605
Prabhu S. Arunachalam, NaYoung Ha, S. Moses Dennison, Rachel L. Spreng, Kelly E. Seaton, Peng Xiao, Yupeng Feng, Veronika I. Zarnitsyna, Dmitri Kazmin, Mengyun Hu, Jordan M. Santagata, Xia Xie, Kenneth Rogers, Lisa M. Shirreff, Claire Chottin, Alexandra J. Spencer, Sheetij Dutta, Katherine Prieto, Jean-Philippe Julien, Mark Tomai, Christopher B. Fox, Francois Villinger, Adrian V. S. Hill, Georgia D. Tomaras, Bali Pulendran
Authorization of the Matrix-M (MM)–adjuvanted R21 vaccine by three countries and its subsequent endorsement by the World Health Organization for malaria prevention in children are a milestone in the fight against malaria. Yet, our understanding of the innate and adaptive immune responses elicited by this vaccine remains limited. Here, we compared three clinically relevant adjuvants [3M-052 + aluminum hydroxide (Alum) (3M), a TLR7/8 agonist formulated in Alum; GLA-LSQ, a TLR4 agonist formulated in liposomes with QS-21; and MM, the now-approved adjuvant for R21] for their capacity to induce durable immune responses to R21 in macaques. R21 adjuvanted with 3M on a 0, 8, and 23–week schedule elicited anti-circumsporozoite antibody responses comparable in magnitude to the R21/MM vaccine administered using a 0-4-8–week regimen and persisted up to 72 weeks with a half-life of 337 days. A booster dose at 72 weeks induced a recall response similar to the R21/MM vaccination. In contrast, R21/GLA-LSQ immunization induced a lower, short-lived response at the dose used. Consistent with the durable serum antibody responses, MM and 3M induced long-lived plasma cells in the bone marrow and other tissues, including the spleen. Furthermore, whereas 3M stimulated potent and persistent antiviral transcriptional and cytokine signatures after primary and booster immunizations, MM induced enhanced expression of interferon- and TH2-related signatures more highly after the booster vaccination. Collectively, these findings provide a resource on the immune responses of three clinically relevant adjuvants with R21 and highlight the promise of 3M as another adjuvant for malarial vaccines.
{"title":"A comparative immunological assessment of multiple clinical-stage adjuvants for the R21 malaria vaccine in nonhuman primates","authors":"Prabhu S. Arunachalam, NaYoung Ha, S. Moses Dennison, Rachel L. Spreng, Kelly E. Seaton, Peng Xiao, Yupeng Feng, Veronika I. Zarnitsyna, Dmitri Kazmin, Mengyun Hu, Jordan M. Santagata, Xia Xie, Kenneth Rogers, Lisa M. Shirreff, Claire Chottin, Alexandra J. Spencer, Sheetij Dutta, Katherine Prieto, Jean-Philippe Julien, Mark Tomai, Christopher B. Fox, Francois Villinger, Adrian V. S. Hill, Georgia D. Tomaras, Bali Pulendran","doi":"10.1126/scitranslmed.adn6605","DOIUrl":"10.1126/scitranslmed.adn6605","url":null,"abstract":"<div >Authorization of the Matrix-M (MM)–adjuvanted R21 vaccine by three countries and its subsequent endorsement by the World Health Organization for malaria prevention in children are a milestone in the fight against malaria. Yet, our understanding of the innate and adaptive immune responses elicited by this vaccine remains limited. Here, we compared three clinically relevant adjuvants [3M-052 + aluminum hydroxide (Alum) (3M), a TLR7/8 agonist formulated in Alum; GLA-LSQ, a TLR4 agonist formulated in liposomes with QS-21; and MM, the now-approved adjuvant for R21] for their capacity to induce durable immune responses to R21 in macaques. R21 adjuvanted with 3M on a 0, 8, and 23–week schedule elicited anti-circumsporozoite antibody responses comparable in magnitude to the R21/MM vaccine administered using a 0-4-8–week regimen and persisted up to 72 weeks with a half-life of 337 days. A booster dose at 72 weeks induced a recall response similar to the R21/MM vaccination. In contrast, R21/GLA-LSQ immunization induced a lower, short-lived response at the dose used. Consistent with the durable serum antibody responses, MM and 3M induced long-lived plasma cells in the bone marrow and other tissues, including the spleen. Furthermore, whereas 3M stimulated potent and persistent antiviral transcriptional and cytokine signatures after primary and booster immunizations, MM induced enhanced expression of interferon- and T<sub>H</sub>2-related signatures more highly after the booster vaccination. Collectively, these findings provide a resource on the immune responses of three clinically relevant adjuvants with R21 and highlight the promise of 3M as another adjuvant for malarial vaccines.</div>","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 758","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.science.org/doi/reader/10.1126/scitranslmed.adn6605","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860758","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-31DOI: 10.1126/scitranslmed.adr6879
{"title":"Erratum for the Research Article “Gene-edited and -engineered stem cell platform drives immunotherapy for brain metastatic melanomas” by N. Kanaya et al.","authors":"","doi":"10.1126/scitranslmed.adr6879","DOIUrl":"10.1126/scitranslmed.adr6879","url":null,"abstract":"","PeriodicalId":21580,"journal":{"name":"Science Translational Medicine","volume":"16 758","pages":""},"PeriodicalIF":15.8,"publicationDate":"2024-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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