Programmable nucleases have enabled rapid and accessible genome engineering in eukaryotic cells and living organisms. However, their delivery into human blood cells can be challenging. Here, we have utilized "nanoblades," a new technology that delivers a genomic cleaving agent into cells. These are modified murine leukemia virus (MLV) or HIV-derived virus-like particle (VLP), in which the viral structural protein Gag has been fused to Cas9. These VLPs are thus loaded with Cas9 protein complexed with the guide RNAs. Highly efficient gene editing was obtained in cell lines, IPS and primary mouse and human cells. Here, we showed that nanoblades were remarkably efficient for entry into human T, B, and hematopoietic stem and progenitor cells (HSPCs) thanks to their surface co-pseudotyping with baboon retroviral and VSV-G envelope glycoproteins. A brief incubation of human T and B cells with nanoblades incorporating two gRNAs resulted in 40 and 15% edited deletion in the Wiskott-Aldrich syndrome (WAS) gene locus, respectively. CD34 cells (HSPCs) treated with the same nanoblades allowed 30-40% exon 1 drop-out in the WAS gene locus. Importantly, no toxicity was detected upon nanoblade-mediated gene editing of these blood cells. Finally, we also treated HSPCs with nanoblades in combination with a donor-encoding rAAV6 vector resulting in up to 40% of stable expression cassette knock-in into the WAS gene locus. Summarizing, this new technology is simple to implement, shows high flexibility for different targets including primary immune cells of human and murine origin, is relatively inexpensive and therefore gives important prospects for basic and clinical translation in the area of gene therapy.
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http://dx.doi.org/10.3389/fgeed.2021.604371 | DOI Listing |
Adv Sci (Weinh)
December 2024
Department of Cardiology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, 310003, China.
Aortic aneurysm is a life-threatening disease caused by progressive dilation of the aorta and weakened aortic walls. Its pathogenesis involves an imbalance between connective tissue repair and degradation. CD34 cells comprise a heterogeneous population that exhibits stem cell and progenitor cell properties.
View Article and Find Full Text PDFInt J Gynecol Pathol
December 2024
Department of Pathology, The First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong, P.R. China.
The incidence of neurotrophic tyrosine kinase receptor (NTRK) fusion uterine sarcoma is extremely low, and reports have been mostly focused on cases localized to the cervix. So far, only 4 cases have been reported of the uterine corpus. In this study, we reported a case of NTRK fusion corpus sarcoma.
View Article and Find Full Text PDFVestn Otorinolaringol
December 2024
Dagestan State Medical University, Makhachkala, Russia.
Unlabelled: This paper presents the results of the evaluation of cellular and humoral immunity in chronic purulent otitis media (CPOM), as well as cytokine status, and studies the effect of azoximers bromide on the immunity system in CPOM.
Objective: To study the clinical and immunologic effectiveness of azoximer bromide in the postoperative period during tympanoplasty in CPOM patients.
Material And Methods: Forty-nine patients with mesotympanitis and epitympanitis were examined.
J Pers Med
November 2024
Department of Internal Medicine, Rehabilitation and Physical Medicine, Medical University of Lodz, 90-419 Lodz, Poland.
: The etiology and causes of osteoarthritis are still being studied at the cellular and molecular level by many scientists around the world. With advances in knowledge, technology, and the need for complexity, new therapeutic approaches-such as restorative medicine-are being developed to protect the patient from endoprosthesis implantation, aiming to simultaneously restore and maintain physical and psychosocial function. The purpose of this study was to evaluate the effectiveness of physiotherapy after the implantation of CD34+ stem cells into the hip joints of patients with osteoarthrosis.
View Article and Find Full Text PDFCancer Sci
December 2024
Department of Medicine and Biosystemic Science, Kyushu University Graduate School of Medicine, Fukuoka, Japan.
In this study, we investigated the measurable residual leukemic stem cell (MR-LSC) population after allogeneic stem cell transplantation (allo-SCT) for high-risk acute myeloid leukemia (AML), utilizing T-cell immunoglobulin mucin-3 (TIM-3) expression as a functional marker of AML leukemic stem cells (LSCs). Analysis of the CD34CD38 fraction of bone marrow cells immediately after achievement of engraftment revealed the presence of both TIM-3LSCs and TIM-3 donor hematopoietic stem cells (HSCs) at varying ratios. Genetic analysis confirmed that TIM-3 cells harbored patient-specific mutations identical to those found in AML clones, whereas TIM-3 cells did not, indicating that TIM-3CD34CD38 cells represent residual AML LSCs.
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