Molecular Therapy. Nucleic Acids最新文献_第2页

Rationalizing the effects of RNA modifications on protein interactions. 使RNA修饰对蛋白质相互作用的影响合理化。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-11-15 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102391

Andrea Vandelli, Laura Broglia, Alexandros Armaos, Riccardo Delli Ponti, Gian Gaetano Tartaglia

{"title":"Rationalizing the effects of RNA modifications on protein interactions.","authors":"Andrea Vandelli, Laura Broglia, Alexandros Armaos, Riccardo Delli Ponti, Gian Gaetano Tartaglia","doi":"10.1016/j.omtn.2024.102391","DOIUrl":"10.1016/j.omtn.2024.102391","url":null,"abstract":"RNA modifications play a crucial role in regulating gene expression by altering RNA structure and modulating interactions with RNA-binding proteins (RBPs). In this study, we explore the impact of specific RNA chemical modifications-N6-methyladenosine (m⁶A), A-to-I editing, and pseudouridine (Ψ)-on RNA secondary structure and protein-RNA interactions. Utilizing genome-wide data, including RNA secondary structure predictions and protein-RNA interaction datasets, we classify proteins into distinct categories based on their binding behaviors: modification specific and structure independent, or modification unspecific and structure dependent. For instance, m⁶A readers such as YTHDF2 exhibit modification-specific and structure-independent binding, consistently recognizing m⁶A regardless of structural changes. Conversely, proteins such as U2AF2 display modification-unspecific and structure-dependent behavior, altering their binding preferences in response to structural changes induced by different modifications. A-to-I editing, which causes significant structural changes, typically reduces protein interactions, while Ψ enhances RNA structural stability, albeit with variable effects on protein binding. To predict these interactions, we developed the catRAPID 2.2 RNA modifications algorithm, which computes the effects of RNA modifications on protein-RNA binding propensities. This algorithm enables the prediction and analysis of RNA modifications' impact on protein interactions, offering new insights into RNA biology and engineering.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102391"},"PeriodicalIF":6.5,"publicationDate":"2024-11-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11664407/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142882580","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineered packaging cell line for the enhanced production of baboon-enveloped retroviral vectors. 用于强化生产狒狒开发的逆转录病毒载体的工程包装细胞系。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-11-13 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102389

Denise Klatt, Lucia Sereni, Boya Liu, Pietro Genovese, Axel Schambach, Els Verhoeyen, David A Williams, Christian Brendel

{"title":"Engineered packaging cell line for the enhanced production of baboon-enveloped retroviral vectors.","authors":"Denise Klatt, Lucia Sereni, Boya Liu, Pietro Genovese, Axel Schambach, Els Verhoeyen, David A Williams, Christian Brendel","doi":"10.1016/j.omtn.2024.102389","DOIUrl":"10.1016/j.omtn.2024.102389","url":null,"abstract":"The baboon endogenous retrovirus (BaEV) glycoprotein is superior to the commonly used vesicular stomatitis virus glycoprotein (VSVg) for retroviral gene transfer into resting hematopoietic stem cells and lymphocyte populations. The derivative BaEVRLess (lacking the R domain) produces higher viral titers compared with wild-type BaEV, but vector production is impaired by syncytia formation and cell death of the HEK293T cells due to the high fusogenic activity of the glycoprotein. This lowers viral titers, leads to increased batch-to-batch variability, and impedes the establishment of stable packaging cell lines essential for the economical production of viral supernatants. Here, we show that knockout of the entry receptor ASCT2 in HEK293T producer cells eliminates syncytia formation, resulting in a 2-fold increase in viral titers, reduced toxicity of viral supernatants, and enables the generation of stable packaging cell lines. In successive steps, we stably integrated BaEVRLess and α-retroviral a.Gag/Pol expression cassettes and isolated clones supporting titers up to 108 to 109 infectious particles/mL, a 10-fold increase in concentrated viral titers. The additional overexpression of CD47 and knockout of β2-microglobulin in the packaging cell line are tailored for future use in in vivo gene therapy applications by reducing non-specific uptake by macrophages and the immunogenicity of viral particles.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102389"},"PeriodicalIF":6.5,"publicationDate":"2024-11-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11638596/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142829445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A novel hyperactive variant of the Sleeping Beauty transposase facilitates non-viral genome engineering. 一种新的睡美人转座酶的过度活跃变体促进了非病毒基因组工程。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-11-04 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102381

Matthias Thomas Ochmann, Csaba Miskey, Lacramioara Botezatu, Nicolás Sandoval-Villegas, Tanja Diem, Zoltán Ivics

{"title":"A novel hyperactive variant of the Sleeping Beauty transposase facilitates non-viral genome engineering.","authors":"Matthias Thomas Ochmann, Csaba Miskey, Lacramioara Botezatu, Nicolás Sandoval-Villegas, Tanja Diem, Zoltán Ivics","doi":"10.1016/j.omtn.2024.102381","DOIUrl":"10.1016/j.omtn.2024.102381","url":null,"abstract":"The Sleeping Beauty (SB) transposon system is a useful tool for genetic applications, including gene therapy. We discovered a hyperactive variant of the SB100X transposase, called SB200X. This mutant, resulting from a specific amino acid replacement (Q124C), showed an ∼2-fold increase in transposition activity in various human and murine cells. Other amino acid replacements in position 124 also led to a hyperactive phenotype. Position 124 is located at the very edge of the linker region that connects the DNA-binding and catalytic domains of the transposase. Consistent with a role of the linker in an autoregulatory mechanism called overproduction inhibition (OPI) in the monophyletic group of mariner transposases, we show that the hyperactivity of Q124C manifests at high concentrations of the transposase, suggesting a partial resistance of SB200X to OPI. We demonstrate that the hyperactive phenotype of Q124C can be combined with features of other useful mutations in the SB transposase. Namely, Q124C improves the transposition efficiency of the previously described K248R variant, while maintaining or even slightly improving its safer genome-wide integration profile. The SB200X transposase could enhance the utility of SB transposon-mediated genome engineering in preclinical and clinical applications.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102381"},"PeriodicalIF":6.5,"publicationDate":"2024-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11626015/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142801584","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

DTMP-prime: A deep transformer-based model for predicting prime editing efficiency and PegRNA activity. DTMP-prime：一个基于深度转换器的预测primer编辑效率和PegRNA活性的模型。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-10-28 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102370

Roghayyeh Alipanahi, Leila Safari, Alireza Khanteymoori

{"title":"DTMP-prime: A deep transformer-based model for predicting prime editing efficiency and PegRNA activity.","authors":"Roghayyeh Alipanahi, Leila Safari, Alireza Khanteymoori","doi":"10.1016/j.omtn.2024.102370","DOIUrl":"10.1016/j.omtn.2024.102370","url":null,"abstract":"Prime editors are CRISPR-based genome engineering tools with significant potential for rectifying patient mutations. However, their usage requires experimental optimization of the prime editing guide RNA (PegRNA) to achieve high editing efficiency. This paper introduces the deep transformer-based model for predicting prime editing efficiency (DTMP-Prime), a tool specifically designed to predict PegRNA activity and prime editing (PE) efficiency. DTMP-Prime facilitates the design of appropriate PegRNA and ngRNA. A transformer-based model was constructed to scrutinize a wide-ranging set of PE data, enabling the extraction of effective features of PegRNAs and target DNA sequences. The integration of these features with the proposed encoding strategy and DNABERT-based embedding has notably improved the predictive capabilities of DTMP-Prime for off-target sites. Moreover, DTMP-Prime is a promising tool for precisely predicting off-target sites in CRISPR experiments. The integration of a multi-head attention framework has additionally improved the precision and generalizability of DTMP-Prime across various PE models and cell lines. Evaluation results based on the Pearson and Spearman correlation coefficient demonstrate that DTMP-Prime outperforms other state-of-the-art models in predicting the efficiency and outcomes of PE experiments.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102370"},"PeriodicalIF":6.5,"publicationDate":"2024-10-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11626815/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142801667","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Local CpG-Stat3 siRNA treatment improves antitumor effects of immune checkpoint inhibitors. 局部CpG-Stat3 siRNA治疗可提高免疫检查点抑制剂的抗肿瘤作用。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-10-09 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102357

Chunyan Zhang, Rui Huang, Lyuzhi Ren, Antons Martincuks, JiEun Song, Marcin Kortylewski, Piotr Swiderski, Stephen J Forman, Hua Yu

{"title":"Local CpG-Stat3 siRNA treatment improves antitumor effects of immune checkpoint inhibitors.","authors":"Chunyan Zhang, Rui Huang, Lyuzhi Ren, Antons Martincuks, JiEun Song, Marcin Kortylewski, Piotr Swiderski, Stephen J Forman, Hua Yu","doi":"10.1016/j.omtn.2024.102357","DOIUrl":"10.1016/j.omtn.2024.102357","url":null,"abstract":"Immune checkpoint blockade (ICB) therapy has significantly benefited patients with several types of solid tumors and some lymphomas. However, many of the treated patients do not have a durable clinical response. It has been demonstrated that rescuing exhausted CD8+ T cells is required for ICB-mediated antitumor effects. We recently developed an immunostimulatory strategy based on silencing STAT3 while stimulating immune responses by CpG, a ligand for Toll-like receptor 9 (TLR9). The CpG-small interfering RNA (siRNA) conjugates efficiently enter immune cells, silencing STAT3 and activating innate immunity to enhance T cell-mediated antitumor immune responses. In the present study, we demonstrate that blocking STAT3 through locally delivered CpG-Stat3 siRNA enhances the efficacies of the systemic PD-1 and CTLA4 blockade against mouse A20 B cell lymphoma. In addition, locally delivered CpG-Stat3 siRNA combined with systemic administration of PD-1 antibody significantly augmented both local and systemic antitumor effects against mouse B16 melanoma tumors, with enhanced tumor-associated T cell activation. Furthermore, locally delivered CpG-Stat3 siRNA enhanced CD8+ T cell tumor infiltration and antitumor activity in a xenograft tumor model. Overall, our studies in both B cell lymphoma and melanoma mouse models demonstrate the potential of combinatory immunotherapy with CpG-Stat3 siRNA and checkpoint inhibitors as a therapeutic strategy for B cell lymphoma and melanoma.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102357"},"PeriodicalIF":6.5,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11605413/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142770477","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Integrating IL-12 mRNA nanotechnology with SBRT eliminates T cell exhaustion in preclinical models of pancreatic cancer. 在胰腺癌临床前模型中，将 IL-12 mRNA 纳米技术与 SBRT 相结合可消除 T 细胞衰竭。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-09-30 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102350

Angela L Hughson, Gary Hannon, Noah A Salama, Tara G Vrooman, Caroline A Stockwell, Bradley N Mills, Jesse Garrett-Larsen, Haoming Qiu, Roula Katerji, Lauren Benoodt, Carl J Johnston, Joseph D Murphy, Emma Kruger, Jian Ye, Nicholas W Gavras, David C Keeley, Shuyang S Qin, Maggie L Lesch, Jason B Muhitch, Tanzy M T Love, Laura M Calvi, Edith M Lord, Nadia Luheshi, Jim Elyes, David C Linehan, Scott A Gerber

{"title":"Integrating IL-12 mRNA nanotechnology with SBRT eliminates T cell exhaustion in preclinical models of pancreatic cancer.","authors":"Angela L Hughson, Gary Hannon, Noah A Salama, Tara G Vrooman, Caroline A Stockwell, Bradley N Mills, Jesse Garrett-Larsen, Haoming Qiu, Roula Katerji, Lauren Benoodt, Carl J Johnston, Joseph D Murphy, Emma Kruger, Jian Ye, Nicholas W Gavras, David C Keeley, Shuyang S Qin, Maggie L Lesch, Jason B Muhitch, Tanzy M T Love, Laura M Calvi, Edith M Lord, Nadia Luheshi, Jim Elyes, David C Linehan, Scott A Gerber","doi":"10.1016/j.omtn.2024.102350","DOIUrl":"10.1016/j.omtn.2024.102350","url":null,"abstract":"Pronounced T cell exhaustion characterizes immunosuppressive tumors, with the tumor microenvironment (TME) employing multiple mechanisms to elicit this suppression. Traditional immunotherapies, such as immune checkpoint blockade, often fail due to their focus primarily on T cells. To overcome this, we utilized a proinflammatory cytokine, interleukin (IL)-12, that re-wires the immunosuppressive TME by inducing T cell effector function while also repolarizing immunosuppressive myeloid cells. Due to toxicities observed with systemic administration of this cytokine, we utilized lipid nanoparticles encapsulating mRNA encoding IL-12 for intratumoral injection. This strategy has been proven safe and tolerable in early clinical trials for solid malignancies. We report an unprecedented loss of exhausted T cells and the emergence of an activated phenotype in murine pancreatic ductal adenocarcinoma (PDAC) treated with stereotactic body radiation therapy (SBRT) and IL-12mRNA. Our mechanistic findings reveal that each treatment modality contributes to the T cell response differently, with SBRT expanding the T cell receptor repertoire and IL-12mRNA promoting robust T cell proliferation and effector status. This distinctive T cell signature mediated marked growth reductions and long-term survival in local and metastatic PDAC models. This is the first study of its kind combining SBRT with IL-12mRNA and provides a promising new approach for treating this aggressive malignancy.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102350"},"PeriodicalIF":6.5,"publicationDate":"2024-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11513558/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142522479","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Optimization of a lentivirus-mediated gene therapy targeting HIV-1 RNA to eliminate HIV-1-infected cells. 优化慢病毒介导的针对 HIV-1 RNA 的基因疗法，以消除受 HIV-1 感染的细胞。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-09-16 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102341

Amanda B Buckingham, Sophia Ho, Finlay Knops-Mckim, Carin K Ingemarsdotter, Andrew M L Lever

{"title":"Optimization of a lentivirus-mediated gene therapy targeting HIV-1 RNA to eliminate HIV-1-infected cells.","authors":"Amanda B Buckingham, Sophia Ho, Finlay Knops-Mckim, Carin K Ingemarsdotter, Andrew M L Lever","doi":"10.1016/j.omtn.2024.102341","DOIUrl":"10.1016/j.omtn.2024.102341","url":null,"abstract":"Persistence of HIV-1 in cellular reservoirs results in lifelong infection, with cure achieved only in rare cases through ablation of marrow-derived cells. We report on optimization of an approach that could potentially be aimed at eliminating these reservoirs, hijacking the HIV-1 alternative splicing process to functionalize the herpes simplex virus thymidine kinase (HSVtk)/ganciclovir (GCV) cell suicide system through targeted RNA trans-splicing at the HIV-1 D4 donor site. AUG1-deficient HSVtk therapeutic pre-mRNA was designed to gain an in-frame start codon from HIV-1 tat1. D4-targeting lentiviral vectors were produced and used to transduce HIV-1-expressing cells, where trans-spliced HIV-1 tat/HSVtk mRNA was successfully detected. However, translation of catalytically active HSVtk polypeptides from internal AUGs in HSVtk ΔAUG1 caused GCV-mediated cytotoxicity in uninfected cells. Modifying these sites in the D4 opt 2 lentiviral vector effectively mitigated this major off-target effect. Promoter choice was optimized for increased transgene expression. Affinity for HIV-1 RNA predicted in silico correlated with the propensity of opt 2 payloads to induce HIV-1 RNA trans-splicing and killing of HIV-1-expressing cells with no significant effect on uninfected cells. Following latency reversing agent (LRA) optimization and treatment, 45% of lymphocytes in an HIV-1-infected latency model could be eliminated with D4 opt 2/GCV. Further development would be warranted to exploit this approach.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102341"},"PeriodicalIF":6.5,"publicationDate":"2024-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11491724/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142470287","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Alternative splicing dysregulation across tissue and therapeutic approaches in a mouse model of myotonic dystrophy type 1. 1 型肌营养不良症小鼠模型中不同组织和治疗方法的替代剪接失调。

IF 6.5 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-09-13 eCollection Date: 2024-12-10 DOI: 10.1016/j.omtn.2024.102338

Sawyer M Hicks, Jesus A Frias, Subodh K Mishra, Marina Scotti, Derek R Muscato, M Carmen Valero, Leanne M Adams, John D Cleary, Masayuki Nakamori, Eric Wang, J Andrew Berglund

{"title":"Alternative splicing dysregulation across tissue and therapeutic approaches in a mouse model of myotonic dystrophy type 1.","authors":"Sawyer M Hicks, Jesus A Frias, Subodh K Mishra, Marina Scotti, Derek R Muscato, M Carmen Valero, Leanne M Adams, John D Cleary, Masayuki Nakamori, Eric Wang, J Andrew Berglund","doi":"10.1016/j.omtn.2024.102338","DOIUrl":"10.1016/j.omtn.2024.102338","url":null,"abstract":"Myotonic dystrophy type 1 (DM1), the leading cause of adult-onset muscular dystrophy, is caused by a CTG repeat expansion. Expression of the repeat causes widespread alternative splicing (AS) defects and downstream pathogenesis, including significant skeletal muscle impacts. The HSA LR mouse model plays a significant role in therapeutic development. This mouse model features a transgene composed of approximately 220 interrupted CTG repeats, which results in skeletal muscle pathology that mirrors DM1. To better understand this model and the growing number of therapeutic approaches developed with it, we performed a meta-analysis of publicly available RNA sequencing data for AS changes across three widely examined skeletal muscles: quadriceps, gastrocnemius, and tibialis anterior. Our analysis demonstrated that transgene expression correlated with the extent of splicing dysregulation across these muscles from gastrocnemius (highest), quadriceps (medium), to tibialis anterior (lowest). We identified 95 splicing events consistently dysregulated across all examined datasets. Comparison of splicing rescue across seven therapeutic approaches showed a range of rescue across the 95 splicing events from the three muscle groups. This analysis contributes to our understanding of the HSA LR model and the growing number of therapeutic approaches currently in preclinical development for DM1.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"35 4","pages":"102338"},"PeriodicalIF":6.5,"publicationDate":"2024-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11465180/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142400781","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Engineered AAV capsid transport mutants overcome transduction deficiencies in the aged CNS 工程化 AAV 胶囊运输突变体克服了老年中枢神经系统的转导缺陷

IF 8.8 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-09-12 DOI: 10.1016/j.omtn.2024.102332

Ivette M. Sandoval, Christy M. Kelley, Luis Daniel Bernal-Conde, Kathy Steece-Collier, David J. Marmion, Marcus Davidsson, Sean M. Crosson, Sanford L. Boye, Shannon E. Boye, Fredric P. Manfredsson

{"title":"Engineered AAV capsid transport mutants overcome transduction deficiencies in the aged CNS","authors":"Ivette M. Sandoval, Christy M. Kelley, Luis Daniel Bernal-Conde, Kathy Steece-Collier, David J. Marmion, Marcus Davidsson, Sean M. Crosson, Sanford L. Boye, Shannon E. Boye, Fredric P. Manfredsson","doi":"10.1016/j.omtn.2024.102332","DOIUrl":"https://doi.org/10.1016/j.omtn.2024.102332","url":null,"abstract":"Adeno-associated virus (AAV)-based gene therapy has enjoyed great successes over the past decade, with Food and Drug Administration-approved therapeutics and a robust clinical pipeline. Nonetheless, barriers to successful translation remain. For example, advanced age is associated with impaired brain transduction, with the diminution of infectivity depending on anatomical region and capsid. Given that CNS gene transfer is often associated with neurodegenerative diseases where age is the chief risk factor, we sought to better understand the causes of this impediment. We assessed two AAV variants hypothesized to overcome factors negatively impacting transduction in the aged brain; specifically, changes in extracellular and cell-surface glycans, and intracellular transport. We evaluated a heparin sulfate proteoglycan null variant with or without mutations enhancing intracellular transport. Vectors were injected into the striatum of young adult or aged rats to address whether improving extracellular diffusion, removing glycan receptor dependence, or improving intracellular transport are important factors in transducing the aged brain. We found that, regardless of the viral capsid, there was a reduction in many of our metrics of transduction in the aged brain. However, the transport mutant was less sensitive to age, suggesting that changes in the cellular transport of AAV capsids are a key factor in age-related transduction deficiency.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"22 1","pages":""},"PeriodicalIF":8.8,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142247942","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Quality by design for mRNA platform purification based on continuous oligo-dT chromatography 基于连续寡聚-dT 色谱的 mRNA 平台纯化质量设计

IF 8.8 2区医学

Molecular Therapy. Nucleic Acids Pub Date : 2024-09-11 DOI: 10.1016/j.omtn.2024.102333

Jixin Qu, Adithya Nair, George W. Muir, Kate A. Loveday, Zidi Yang, Ehsan Nourafkan, Emma N. Welbourne, Mabrouka Maamra, Mark J. Dickman, Zoltán Kis

{"title":"Quality by design for mRNA platform purification based on continuous oligo-dT chromatography","authors":"Jixin Qu, Adithya Nair, George W. Muir, Kate A. Loveday, Zidi Yang, Ehsan Nourafkan, Emma N. Welbourne, Mabrouka Maamra, Mark J. Dickman, Zoltán Kis","doi":"10.1016/j.omtn.2024.102333","DOIUrl":"https://doi.org/10.1016/j.omtn.2024.102333","url":null,"abstract":"Oligo-deoxythymidine (oligo-dT) ligand-based affinity chromatography is a robust method for purifying mRNA drug substances within the manufacturing process of mRNA-based products, including vaccines and therapeutics. However, the conventional batch mode of operation for oligo-dT chromatography has certain drawbacks that reduce this process’s productivity. Here, we report a new continuous oligo-dT chromatography process for the purification of <ce:italic>in vitro</ce:italic> transcribed mRNA, which reduces losses, improves the efficiency of oligo-dT resin use, and intensifies the chromatography process. Furthermore, the quality by design (QbD) framework was used to establish a design space for the newly developed method. The optimization of process parameters (PPs), including salt type, salt concentration, load flow rate and mRNA load concentration both in batch and the continuous mode, achieved a greater than 90% yield (mRNA recovery) along with greater than 95% mRNA integrity and greater than 99% purity. The productivity of continuous chromatography was estimated to be 5.75-fold higher, and the operating cost was estimated 15% lower, when compared with batch chromatography. Furthermore, the QbD framework was further used to map the relationship between critical quality attributes and key performance indicators as a function of critical process parameters and critical material attributes.","PeriodicalId":18821,"journal":{"name":"Molecular Therapy. Nucleic Acids","volume":"2 1","pages":""},"PeriodicalIF":8.8,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142247943","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0