Molecular Therapy. Methods & Clinical Development最新文献_第6页

High value of ⁶⁴Cu as a tool to evaluate the restoration of physiological copper excretion after gene therapy in Wilson's disease. 高价值的64Cu作为评估Wilson病基因治疗后生理性铜排泄恢复的工具。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-06-09 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.06.001

Oihana Murillo, Maria Collantes, Cristina Gazquez, Daniel Moreno, Ruben Hernandez-Alcoceba, Miren Barberia, Margarita Ecay, Blanche Tamarit, Anne Douar, Veronica Ferrer, Jean Philippe Combal, Ivan Peñuelas, Bernard Bénichou, Gloria Gonzalez-Aseguinolaza

{"title":"High value of 64Cu as a tool to evaluate the restoration of physiological copper excretion after gene therapy in Wilson's disease.","authors":"Oihana Murillo, Maria Collantes, Cristina Gazquez, Daniel Moreno, Ruben Hernandez-Alcoceba, Miren Barberia, Margarita Ecay, Blanche Tamarit, Anne Douar, Veronica Ferrer, Jean Philippe Combal, Ivan Peñuelas, Bernard Bénichou, Gloria Gonzalez-Aseguinolaza","doi":"10.1016/j.omtm.2022.06.001","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.06.001","url":null,"abstract":"Wilson's disease (WD) is an inherited disorder of copper metabolism associated with mutations in ATP7B gene. We have shown that the administration of an adeno-associated vector (AAV) encoding a mini version of human ATP7B (VTX-801) provides long-term correction of copper metabolism in a murine WD model. In preparation of a future clinical trial, we have evaluated by positron emission tomography (PET) the value of 64Cu biodistribution, excretion pattern, and blood kinetics as pharmacodynamic biomarkers of VTX-801 effects. Six-week-old WD mice were injected intravenously with increasing doses of VTX-801 and 3 weeks or 3 months later with [64Cu]CuCl2. Untreated WD and wild-type (WT) mice were included as controls. Control WD mice showed increased hepatic 64Cu retention, reduced fecal excretion of the radiotracer, and altered 64Cu blood kinetics (BK) compared with WT mice. VTX-801 treatment in WD mice resulted in a significant reduction of hepatic 64Cu accumulation, the restoration of fecal 64Cu excretion, and the correction of 64Cu BK. This study showed that VTX-801 restores physiological copper metabolism in WD mice, confirming the mechanism of action of VTX-801, and demonstrated the translational potential of [64Cu]CuCl2-PET to explore VTX-801 pharmacodynamics in a minimally invasive and sensitive manner in WD patients.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"98-106"},"PeriodicalIF":4.7,"publicationDate":"2022-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/52/1d/main.PMC9234538.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40568521","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 5

Biodistribution and environmental safety of a live-attenuated YF17D-vectored SARS-CoV-2 vaccine candidate. 一种以yf17d为载体的SARS-CoV-2减毒候选疫苗的生物分布和环境安全性

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-06-09 Epub Date: 2022-03-16 DOI: 10.1016/j.omtm.2022.03.010

Li-Hsin Li, Laurens Liesenborghs, Lanjiao Wang, Marleen Lox, Michael Bright Yakass, Sander Jansen, Ana Lucia Rosales Rosas, Xin Zhang, Hendrik Jan Thibaut, Dirk Teuwen, Johan Neyts, Leen Delang, Kai Dallmeier

{"title":"Biodistribution and environmental safety of a live-attenuated YF17D-vectored SARS-CoV-2 vaccine candidate.","authors":"Li-Hsin Li, Laurens Liesenborghs, Lanjiao Wang, Marleen Lox, Michael Bright Yakass, Sander Jansen, Ana Lucia Rosales Rosas, Xin Zhang, Hendrik Jan Thibaut, Dirk Teuwen, Johan Neyts, Leen Delang, Kai Dallmeier","doi":"10.1016/j.omtm.2022.03.010","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.03.010","url":null,"abstract":"New platforms are needed for the design of novel prophylactic vaccines and advanced immune therapies. Live-attenuated yellow fever vaccine YF17D serves as a vector for several licensed vaccines and platform for novel candidates. On the basis of YF17D, we developed an exceptionally potent COVID-19 vaccine candidate called YF-S0. However, use of such live RNA viruses raises safety concerns, such as adverse events linked to original YF17D (yellow fever vaccine-associated neurotropic disease [YEL-AND] and yellow fever vaccine-associated viscerotropic disease [YEL-AVD]). In this study, we investigated the biodistribution and shedding of YF-S0 in hamsters. Likewise, we introduced hamsters deficient in signal transducer and activator of transcription 2 (STAT2) signaling as a new preclinical model of YEL-AND/AVD. Compared with YF17D, YF-S0 showed improved safety with limited dissemination to brain and visceral tissues, absent or low viremia, and no shedding of infectious virus. Considering that yellow fever virus is transmitted by Aedes mosquitoes, any inadvertent exposure to the live recombinant vector via mosquito bites is to be excluded. The transmission risk of YF-S0 was hence compared with readily transmitting YF-Asibi strain and non-transmitting YF17D vaccine, with no evidence for productive infection of mosquitoes. The overall favorable safety profile of YF-S0 is expected to translate to other vaccines based on the same YF17D platform.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"215-224"},"PeriodicalIF":4.7,"publicationDate":"2022-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/5a/19/main.PMC8925082.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40310665","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 3

In vivo generation of CAR T cells in the presence of human myeloid cells. 在人骨髓细胞存在的情况下，体内生成CAR - T细胞。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-06-09 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.06.004

Naphang Ho, Shiwani Agarwal, Michela Milani, Alessio Cantore, Christian J Buchholz, Frederic B Thalheimer

{"title":"In vivo generation of CAR T cells in the presence of human myeloid cells.","authors":"Naphang Ho, Shiwani Agarwal, Michela Milani, Alessio Cantore, Christian J Buchholz, Frederic B Thalheimer","doi":"10.1016/j.omtm.2022.06.004","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.06.004","url":null,"abstract":"Pre-clinical humanized mouse models are a powerful tool to evaluate immunotherapies. NSG-SGM3 mice reconstituted with human stem cells (huSGM3) develop pronounced human myeloid cells due to transgenic expression of stem cell factor, granulocyte-macrophage colony-stimulating factor, and interleukin-3 (IL-3) compared with the widely used humanized NSG (huNSG) model. We assessed in vivo generation of CD19-CAR T cells in huSGM3 mice upon single intravenous injection of the T cell-specific lentiviral vectors (LVs) CD4-LV and CD8-LV. While in vivo CAR T cell generation was clearly detectable in individual mice, generation appeared less efficient than previously observed for huNSG mice. Especially for the CD4-LV group, this correlated with increased IL-15 and decreased GM-CSF levels, indicating activation of monocytes and macrophages. Co-culture assays identified macrophages as a potential barrier for gene transfer. Refining CD4-LV and CD8-LV with a less immunogenic surface by using modified packaging cells substantially improved the transduction of lymphocytes in vitro in the presence of macrophages, as well in vivo in huSGM3 mice. Notably, two mice that developed less CAR T cells showed high interferon-α or -β levels before vector injection. Our data emphasize the relevance of innate immune responses for in vivo generation of CAR T cells, which can be overcome by vector surface engineering.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"144-156"},"PeriodicalIF":4.7,"publicationDate":"2022-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/5c/61/main.PMC9249670.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40477166","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 6

Quantification of transgene expression in GSH AAVS1 with a novel CRISPR/Cas9-based approach reveals high transcriptional variation. 用一种新的基于CRISPR/ cas9的方法定量GSH AAVS1的转基因表达，揭示了高转录变异。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-06-09 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.06.003

Anne Inderbitzin, Tom Loosli, Roger D Kouyos, Karin J Metzner

{"title":"Quantification of transgene expression in GSH AAVS1 with a novel CRISPR/Cas9-based approach reveals high transcriptional variation.","authors":"Anne Inderbitzin, Tom Loosli, Roger D Kouyos, Karin J Metzner","doi":"10.1016/j.omtm.2022.06.003","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.06.003","url":null,"abstract":"Genomic safe harbors (GSH) are defined as sites in the host genome that allow stable expression of inserted transgenes while having no adverse effects on the host cell, making them ideal for use in basic research and therapeutic applications. Silencing and fluctuations in transgene expression would be highly undesirable effects. We have previously shown that transgene expression in Jurkat T cells is not silenced for up to 160 days after CRISPR-Cas9-mediated insertion of reporter genes into the adeno-associated virus site 1 (AAVS1), a commonly used GSH. Here, we studied fluctuations in transgene expression upon targeted insertion into the GSH AAVS1. We have developed an efficient method to generate and validate highly complex barcoded plasmid libraries to study transgene expression on the single-cell level. Its applicability is demonstrated by inserting the barcoded transgene Cerulean into the AAVS1 locus in Jurkat T cells via the CRISPR-Cas9 technology followed by next-generation sequencing of the transcribed barcodes. We observed large transcriptional variations over two logs for transgene expression in the GSH AAVS1. This barcoded transgene insertion model is a powerful tool to investigate fluctuations in transgene expression at any GSH site.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"107-118"},"PeriodicalIF":4.7,"publicationDate":"2022-06-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/7d/5a/main.PMC9234542.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40568522","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Targeted biallelic integration of an inducible Caspase 9 suicide gene in iPSCs for safer therapies. 诱导型Caspase 9自杀基因在iPSCs中的靶向双等位基因整合，用于更安全的治疗。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-31 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.011

Stephanie Wunderlich, Alexandra Haase, Sylvia Merkert, Kirsten Jahn, Maximillian Deest, Helge Frieling, Silke Glage, Wilhelm Korte, Andreas Martens, Andreas Kirschning, Andre Zeug, Evgeni Ponimaskin, Gudrun Göhring, Mania Ackermann, Nico Lachmann, Thomas Moritz, Robert Zweigerdt, Ulrich Martin

{"title":"Targeted biallelic integration of an inducible Caspase 9 suicide gene in iPSCs for safer therapies.","authors":"Stephanie Wunderlich, Alexandra Haase, Sylvia Merkert, Kirsten Jahn, Maximillian Deest, Helge Frieling, Silke Glage, Wilhelm Korte, Andreas Martens, Andreas Kirschning, Andre Zeug, Evgeni Ponimaskin, Gudrun Göhring, Mania Ackermann, Nico Lachmann, Thomas Moritz, Robert Zweigerdt, Ulrich Martin","doi":"10.1016/j.omtm.2022.05.011","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.011","url":null,"abstract":"Drug-inducible suicide systems may help to minimize risks of human induced pluripotent stem cell (hiPSC) therapies. Recent research challenged the usefulness of such systems since rare drug-resistant subclones were observed. We have introduced a drug-inducible Caspase 9 suicide system (iCASP9) into the AAVS1 safe-harbor locus of hiPSCs. In these cells, apoptosis could be efficiently induced in vitro. After transplantation into mice, drug treatment generally led to rapid elimination of teratomas, but single animals subsequently formed tumor tissue from monoallelic iCASP9 hiPSCs. Very rare drug-resistant subclones of monoallelic iCASP9 hiPSCs appeared in vitro with frequencies of ∼ 3 × 10-8. Besides transgene elimination, presumably via loss of heterozygosity (LoH), silencing via aberrant promoter methylation was identified as a major underlying mechanism. In contrast to monoallelic iCASP9 hiPSCs, no escapees from biallelic iCASP9 cells were observed after treatment of up to 0.8 billion hiPSCs. The highly increased safety level provided by biallelic integration of the iCASP9 system may substantially contribute to the safety level of iPSC-based therapies.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"84-94"},"PeriodicalIF":4.7,"publicationDate":"2022-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/87/a3/main.PMC9234009.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40477167","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 5

PAM-flexible dual base editor-mediated random mutagenesis and self-activation strategies to improve CRISPRa potency. pam柔性双碱基编辑器介导的随机突变和自激活策略提高CRISPRa效力。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-29 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.005

Cia-Hin Lau, Siping Huang, Raymond H W Lam, Chung Tin

{"title":"PAM-flexible dual base editor-mediated random mutagenesis and self-activation strategies to improve CRISPRa potency.","authors":"Cia-Hin Lau, Siping Huang, Raymond H W Lam, Chung Tin","doi":"10.1016/j.omtm.2022.05.005","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.005","url":null,"abstract":"VP64 is the smallest transactivation domain that can be packaged together with the sgRNA into a single adeno-associated virus (AAV) vector. However, VP64-based CRISPRa often exerts modest activation to the target gene when only one sgRNA is used. Herein, we used PAM-flexible dual base editor-mediated mutagenesis and self-activation strategies to derive VP64 variants with gain-of-function mutations. First, we generated an HEK293FT transgenic clone to stably expressing pTK-CRISPRa-GFP. The sgRNA of CRISPRa was designed to target the TK promoter, thereby allowing self-activation of CRISPRa-GFP. Base editors were then used to randomly mutagenesis VP64 in this transgenic cell. VP64 with enhanced potency would translate into increment of GFP fluorescence intensity, thereby allowing positive selection of the desired VP64 mutants. This strategy has enabled us to identify several VP64 variants that are more potent than the wild-type VP64. ΔCRISPRa derived from these VP64 variants also efficiently activated the endogenous promoter of anti-aging and longevity genes (KLOTHO, SIRT6, and NFE2L2) in human cells. Since the overall size of these ΔCRISPRa transgenes is not increased, it remains feasible for all-in-one AAV applications. The strategies described here can facilitate high-throughput screening of the desired protein variants and adapted to evolve any other effector domains.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"26-37"},"PeriodicalIF":4.7,"publicationDate":"2022-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/e8/50/main.PMC9198377.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40399239","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Generation of hypoimmunogenic induced pluripotent stem cells by CRISPR-Cas9 system and detailed evaluation for clinical application. CRISPR-Cas9系统制备低免疫原性诱导多能干细胞及其临床应用的详细评价。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-29 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.010

Yuko Kitano, Sayaka Nishimura, Tomoaki M Kato, Anna Ueda, Kaho Takigawa, Masafumi Umekage, Masaki Nomura, Ayane Kawakami, Haruna Ogawa, Huaigeng Xu, Akitsu Hotta, Naoko Takasu, Masayoshi Tsukahara

{"title":"Generation of hypoimmunogenic induced pluripotent stem cells by CRISPR-Cas9 system and detailed evaluation for clinical application.","authors":"Yuko Kitano, Sayaka Nishimura, Tomoaki M Kato, Anna Ueda, Kaho Takigawa, Masafumi Umekage, Masaki Nomura, Ayane Kawakami, Haruna Ogawa, Huaigeng Xu, Akitsu Hotta, Naoko Takasu, Masayoshi Tsukahara","doi":"10.1016/j.omtm.2022.05.010","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.010","url":null,"abstract":"In order to expand the promise of regenerative medicine using allogeneic induced pluripotent stem cells (iPSCs), precise and efficient genome editing of human leukocyte antigen (HLA) genes would be advantageous to minimize the immune rejection caused by mismatches of HLA type. However, clinical-grade genome editing of multiple HLA genes in human iPSC lines remains unexplored. Here, we optimized the protocol for good manufacturing practice (GMP)-compatible CRISPR-Cas9 genome editing to deplete the three gene locus (HLA-A, HLA-B, and CIITA genes) simultaneously in HLA homozygous iPSCs. The use of HLA homozygous iPSCs has one main advantage over heterozygous iPSCs for inducing biallelic knockout by a single gRNA. RNA-seq and flow cytometry analyses confirmed the successful depletion of HLAs, and lineage-specific differentiation into cardiomyocytes was verified. We also confirmed that the pluripotency of genome-edited iPSCs was successfully maintained by the three germ layers of differentiation. Moreover, whole-genome sequencing, karyotyping, and optical genome mapping analyses revealed no evident genomic abnormalities detected in some clones, whereas unexpected copy number losses, chromosomal translocations, and complex genomic rearrangements were observed in other clones. Our results indicate the importance of multidimensional analyses to ensure the safety and quality of the genome-edited cells. The manufacturing and assessment pipelines presented here will be the basis for clinical-grade genome editing of iPSCs.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"15-25"},"PeriodicalIF":4.7,"publicationDate":"2022-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9198376/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40399242","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 12

Selective B cell depletion upon intravenous infusion of replication-incompetent anti-CD19 CAR lentivirus. 静脉输注复制无能的抗cd19 CAR慢病毒后选择性B细胞耗竭。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-29 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.006

Craig M Rive, Eric Yung, Lisa Dreolini, Scott D Brown, Christopher G May, Daniel J Woodsworth, Robert A Holt

{"title":"Selective B cell depletion upon intravenous infusion of replication-incompetent anti-CD19 CAR lentivirus.","authors":"Craig M Rive, Eric Yung, Lisa Dreolini, Scott D Brown, Christopher G May, Daniel J Woodsworth, Robert A Holt","doi":"10.1016/j.omtm.2022.05.006","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.006","url":null,"abstract":"Anti-CD19 chimeric antigen receptor (CAR)-T therapy for B cell malignancies has shown clinical success, but a major limitation is the logistical complexity and high cost of manufacturing autologous cell products. If engineered for improved safety, direct infusion of viral gene transfer vectors to initiate in vivo CAR-T transduction, expansion, and anti-tumor activity could provide an alternative, universal approach. To explore this approach we administered approximately 20 million replication-incompetent vesicular stomatitis virus G protein (VSV-G) lentiviral particles carrying an anti-CD19CAR-2A-GFP transgene comprising either an FMC63 (human) or 1D3 (murine) anti-CD19 binding domain, or a GFP-only control transgene, to wild-type C57BL/6 mice by tail vein infusion. The dynamics of immune cell subsets isolated from peripheral blood were monitored at weekly intervals. We saw emergence of a persistent CAR-transduced CD3+ T cell population beginning week 3-4 that reaching a maximum of 13.5% ± 0.58% (mean ± SD) and 7.8% ± 0.76% of the peripheral blood CD3+ T cell population in mice infused with ID3-CAR or FMC63-CAR lentivector, respectively, followed by a rapid decline in each case of the B cell content of peripheral blood. Complete B cell aplasia was apparent by week 5 and was sustained until the end of the protocol (week 8). No significant CAR-positive populations were observed within other immune cell subsets or other tissues. These results indicate that direct intravenous infusion of conventional VSV-G-pseudotyped lentiviral particles carrying a CD19 CAR transgene can transduce T cells that then fully ablate endogenous B cells in wild-type mice.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"4-14"},"PeriodicalIF":4.7,"publicationDate":"2022-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/21/27/main.PMC9198363.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40399240","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 4

Structural basis for the neurotropic AAV9 and the engineered AAVPHP.eB recognition with cellular receptors. 神经嗜性AAV9和工程化AAVPHP的结构基础。细胞受体的eB识别。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-29 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.009

Guangxue Xu, Ran Zhang, Huapeng Li, Kaixin Yin, Xinyi Ma, Zhiyong Lou

{"title":"Structural basis for the neurotropic AAV9 and the engineered AAVPHP.eB recognition with cellular receptors.","authors":"Guangxue Xu, Ran Zhang, Huapeng Li, Kaixin Yin, Xinyi Ma, Zhiyong Lou","doi":"10.1016/j.omtm.2022.05.009","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.009","url":null,"abstract":"Clade F adeno-associated virus (AAV) 9 has been utilized as therapeutic gene delivery vector, and it is capable of crossing blood brain barrier (BBB). Recently, an AAV9-based engineering serotype AAVPHP.eB with enhanced BBB crossing ability further expanded clade F AAVs' usages in the murine central nervous system (CNS) gene delivery. In this study, we determined the cryo-electron microscopy (cryo-EM) structures of the AAVPHP.eB and its parental serotype AAV9 in native form or in complex with their essential receptor AAV receptor (AAVR). These structures reveal the molecular details of their AAVR recognition, where the polycystic kidney disease repeat domain 2 (PKD2) of AAVR interacts with AAV9 and AAVPHP.eB virions at the 3-fold protrusions and the raised capsid regions between the 2- and 5-fold axes, termed the 2/5-fold wall. The interacting patterns of AAVR to AAV9 and AAVPHP.eB are similar to what was observed in AAV1/AAV2-AAVR complexes. Moreover, we found that the AAVPHP.eB variable region VIII (VR-VIII) may independently facilitate the new receptor recognition responsible for enhanced CNS transduction. Our study provides insights into the recognition principles of multiple receptors for engineered AAVPHP.eB and parental serotype AAV9, and further reveal the potential molecular basis underlying their different tropisms.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"52-60"},"PeriodicalIF":4.7,"publicationDate":"2022-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/34/ff/main.PMC9198364.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40399241","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 13

Impact of genetically modified organism requirements on gene therapy development in the EU, Japan, and the US. 转基因生物对欧盟、日本和美国基因治疗发展的影响。

IF 4.7

Molecular Therapy. Methods & Clinical Development Pub Date : 2022-05-28 eCollection Date: 2022-09-08 DOI: 10.1016/j.omtm.2022.05.012

Gentaro Tajima, Seoan Huh, Natalie Anne Schmidt, Judith C Macdonald, Tobias Fleischmann, Keith Merrell Wonnacott

{"title":"Impact of genetically modified organism requirements on gene therapy development in the EU, Japan, and the US.","authors":"Gentaro Tajima, Seoan Huh, Natalie Anne Schmidt, Judith C Macdonald, Tobias Fleischmann, Keith Merrell Wonnacott","doi":"10.1016/j.omtm.2022.05.012","DOIUrl":"https://doi.org/10.1016/j.omtm.2022.05.012","url":null,"abstract":"Advanced therapies are emerging as an important class of medicinal products; among these, gene therapies are advancing at an exceptional rate. However, one of the major challenges for gene therapies relates to the additional regulatory requirements for genetically modified organisms. In this paper, we provide an overview of the regulatory requirements for genetically modified organisms in the European Union, Japan, and the United States. We share our experience in managing these requirements and their impact on the adeno-associated virus gene therapies that are under development at Pfizer. Specifically, we discuss the relative complexity of the approval process and the impact of risk assessment expectations on the clinical development of genetically modified organisms. We also compare the regulatory processes and timelines of various regions based on our experience with adeno-associated viral vectors. Finally, we propose that genetically modified organisms, for which pathogenicity and replication competency are well controlled, should be regulated solely under medicinal product regulations and be exempt from additional requirements for genetically modified organisms. Even if an exemption is not implemented, it should still be possible to significantly reduce the sponsor and agency burden by simplifying and harmonizing documentation and data requirements as well as timelines for applications for genetically modified organisms.","PeriodicalId":517056,"journal":{"name":"Molecular Therapy. Methods & Clinical Development","volume":" ","pages":"74-83"},"PeriodicalIF":4.7,"publicationDate":"2022-05-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/95/98/main.PMC9207611.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40558897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 2