Download or read book Assessment and Improvement of Gene Transfer Into Human Hematopoietic Stem Cells written by Dimitri Anthonius Breems and published by . This book was released on 1997 with total page 164 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Genetic Modification of Hematopoietic Stem Cells written by Christopher Baum and published by Humana Press. This book was released on 2011-01-20 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Gene Transfer into Hematopoietic Cells: From Basic Science to Clinical Application Christopher Baum 1. The Potential of Gene Transfer into Hematopoietic Vectors with the potential for stable transgene integration are Cells widely used in basic hematology and clinical trials of gene me- cine. In basic research, both gain-of-function and loss-of-fu- tion situations of individual genes can be created by gene transfer, leading to a wide range of applications in developmental biology, stem cell biology, immunology, leukemia research, and human genetics. With the first evidence of successful modification of murine hematopoietic cells using retroviral gene vectors (1, 2), researchers have also explored the therapeutic potential of this approach. To date, the emerging discipline of gene therapy is a highly diversified field that offers entirely novel approaches to treat a great variety of human diseases (3). All hematopoietic cell types are of major interest in this context, since the modification of the hematopoietic stem cell population may potentially give rise to a completely transgenic hematopoiesis with the potential to cure genetic disorders or fight severe chronic infections, and the targeting of mature cells such as lymphocytes or antigen-p- senting dendritic cells offers all types of transient and semiper- nent modifications of the immune system. The unifying principle of gene medicine is the need to transfer complex nucleic acids cells that do not contribute to the germline (somatic cells).

Download or read book Gene Transfer Into Hemopoietic Stem Cells written by Markus Peter Wilhelmus Einerhand and published by . This book was released on 1992 with total page 108 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Stem Cells from Cord Blood in Utero Stem Cell Development and Transplantation Inclusive Gene Therapy written by W. Holzgreve and published by Springer Science & Business Media. This book was released on 2013-03-09 with total page 265 pages. Available in PDF, EPUB and Kindle. Book excerpt: The title "Stem Cells from Cord Blood, In Utero Stem Cell Develop ment, and Transplantation-Inclusive Gene Therapy" suggests that more than one topic is combined in one workshop. Indeed, at first glance the recovery of stem cells from cord blood has to be seen as separate from the attempts to achieve effective in utero therapy by stem cell trans plantation, because the first issue deals with an innovative stem cell source as an alternative to bone marrow, which is already spreading rapidly in medical practice, whereas the second topic is still strictly ex perimental and only investigated in medical centers with the appropri ate background. It is, however, not only justified, but helpful to com bine the two topics in one workshop and consequently to cover them in the same volume of the Ernst Schering Research Foundation Work shop series, because they are intimately related and both based on the new insights into the biology of stem cells. Professor Werner Arber, the Nobel Laureate from the University of Basel, pointed out in his In- Professor Dr. W. Holzgreve VI Preface The participants of the workshop troductory Lecture that our understanding of hematopoietic stem cells as descendents of totipotent cells and our current approaches to using them in post-and prenatal therapy have been furthered significantly by genetic engineering technologies which are "artificial contributions to the process of biologic evolution".

Download or read book Oversight and Review of Clinical Gene Transfer Protocols written by Institute of Medicine and published by National Academies Press. This book was released on 2014-03-27 with total page 78 pages. Available in PDF, EPUB and Kindle. Book excerpt: Gene transfer research is a rapidly advancing field that involves the introduction of a genetic sequence into a human subject for research or diagnostic purposes. Clinical gene transfer trials are subject to regulation by the U.S. Food and Drug Administration (FDA) at the federal level and to oversight by institutional review boards (IRBs) and institutional biosafety committees (IBCs) at the local level before human subjects can be enrolled. In addition, at present all researchers and institutions funded by the National Institutes of Health (NIH) are required by NIH guidelines to submit human gene transfer protocols for advisory review by the NIH Recombinant DNA Advisory Committee (RAC). Some protocols are then selected for individual review and public discussion. Oversight and Review of Clinical Gene Transfer Protocols provides an assessment of the state of existing gene transfer science and the current regulatory and policy context under which research is investigated. This report assesses whether the current oversight of individual gene transfer protocols by the RAC continues to be necessary and offers recommendations concerning the criteria the NIH should employ to determine whether individual protocols should receive public review. The focus of this report is on the standards the RAC and NIH should use in exercising its oversight function. Oversight and Review of Clinical Gene Transfer Protocols will assist not only the RAC, but also research institutions and the general public with respect to utilizing and improving existing oversight processes.

Download or read book Improving Nuclease Mediated Gene Editing Outcomes in Human Hematopoietic Stem Cells written by Anastasia Lomova and published by . This book was released on 2019 with total page 183 pages. Available in PDF, EPUB and Kindle. Book excerpt: Autologous hematopoietic stem cell (HSC) transplantation, combined with gene editing, could provide an ideal therapeutic option for the treatment of congenital blood diseases, such as hemoglobinopathies, primary immune deficiencies, and storage disorders. Gene editing relies on site-specific induction of a double stranded break (DSB) by targeted nucleases (such as Zinc Finger Nucleases (ZFNs) or CRISPR/Cas9 system), and subsequent gene correction using endogenous cellular repair mechanisms. The two main competing pathways to repair the break are non-homologous end joining (NHEJ), an often-imprecise pathway which can result in insertions and deletions (indels), or accurate homology-directed repair (HDR) pathway which uses a homologous donor template to seamlessly repair the break and incorporate the desired changes. For certain diseases, where a knockout of a gene can result in therapeutic benefit, repair by NHEJ pathway may be favorable. However, for conditions where disruption of a gene can result in an even more severe phenotype than the original disease (such as sickle cell anemia), repair via HDR pathway is critical. Despite advances in nuclease technologies and the ability to efficiently achieve high frequency of site-specific gene disruption, the current progress to reach clinically relevant levels of precise HDR-mediated repair still remains elusive. Therefore, our translational goal is to improve the gene editing outcomes in HSCs, specifically, increase HDR and decrease NHEJ levels, which will be beneficial for treating many diseases of the blood. This dissertation aims to identify the hindrances that limit efficient HDR-mediated editing in HSCs, and investigates several approaches to address these impediments. Our results indicate that one major reason for low gene correction in HSCs is their heightened susceptibility to cell toxicity resulting from the electroporation of the nuclease and homologous donor template. We demonstrate that co-electroporation of mRNA encoding the anti-apoptotic protein BCL2 with gene editing reagents significantly ameliorates the cytotoxicity and increases the yield of gene-corrected HSCs. Next, we show that cell cycle-dependent control of nuclease activity and DNA repair pathways can influence gene editing outcomes to favor the precise DNA modification (HDR) over faulty repair events (NHEJ) in human HSCs. By using a modified version of Cas9 protein with reduced nuclease activity in G1 phase of cell cycle, when HDR cannot occur, and transiently increasing the proportion of cells in HDR-preferred phases (S/G2), we achieve a 4-fold improvement in HDR/NHEJ ratio over the control condition in vitro, and a significant improvement in long-term gene-modified engrafted cells after xenotransplantation of edited human HSCs into immune-deficient mice. Finally, we investigate what cellular elements govern the DNA repair pathway choice and how they can be exploited to shift the balance toward HDR from NHEJ. We test the effects of manipulating the expression levels of several DNA repair factors, that are presumed to be important for pathway choice and progression, on HDR and NHEJ levels in K562 cell line and primary human hematopoietic stem and progenitor cells (HSPCs). Interestingly, we observe differential effects of DNA repair factor manipulation on gene editing outcomes dependent upon the delivery method employed and the types of cells used. These strategies for improving gene editing outcomes in human HSCs have important implications for the field of gene therapy as a whole, and can be applicable to diseases where increased HDR/NHEJ ratio is critical for therapeutic success.

Download or read book Identification and Characterization of Human Hematopoietic Stem Cells Using Gene Transfer and the Novel SCID Transplantation Assay written by André Larochelle and published by . This book was released on 1997 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: The absence of preclinical repopulation assays for primitive human hematopoietic cells has hindered the development of stem cell gene therapy. Current human gene therapy trials employ gene transfer methods optimized using surrogate in vitro colony-forming cell (CFC) and longterm culture-initiating cell (LTC-IC) assays and by inference from studies on other mammalian species. In this thesis, we report the identification and characterization of a novel human hematopoietic cell, capable of repopulating the bone marrow (BM) of nonobese diabetic mice with severe combined immunodeficiency (NOD/SCID), namely the SCID-repopulating cell (SRC). Human BM or cord blood (CB) cells were infected with retroviral vectors and subsequently transplanted into NOD/SCID mice. Whereas CFC and LTC-IC were efficiently transduced with retroviruses, SRCs were transduced rarely, establishing that SRC are biologically distinct from most CFC and LTC-IC. Moreover, characterization of the cell surface phenotype indicated that SRC are exclusively CD3$\sp+$CD38$\sp-$ in contrast to CFC and LTC-IC that are mostly found in the CD34$\sp+$CD38$\sp+$ fraction after multiparameter flow sorting, confirming the distinct character of SRC. In combination with various data obtained in the past decade, these analyses provide the strongest evidence that SRC defines a novel human hematopoietic cell that is more immature than any other cell type detected to date. The inefficient infection of SRC was consistent with the low level of gene marking reported in primates and human gene therapy trials. This inability to infect the most primitive human repopulating cells, in contrast to most CFC and LTC-IC, highlights the need for appropriate pre-clinical models that will predict the outcomes of human clinical trials. As described in chapter 3 of this thesis, primitive BM cells from $\beta$-thalassemia major and sickle cell anemia (SCA) patients can engraft and proliferate in the BM of immune-deficient mice. The BM of transplanted mice contained the entire erythroid lineage from BFU-E to mature erythrocytes expressing human $\gamma$-, $\beta$- or $\rm\beta\sp{s}$-globin as in the original donor. Moreover, human erythroid cells from mice transplanted with SCA bone marrow showed characteristic sickling under reducing conditions in an in vitro assay. This system can thus be used to evaluate gene transfer efficiency into primitive human cells, longevity of expression, expression in the appropriate lineage, and correction of the disease phenotype.

Download or read book Immunopharmacology written by Manzoor M. Khan and published by Springer Science & Business Media. This book was released on 2008-12-19 with total page 275 pages. Available in PDF, EPUB and Kindle. Book excerpt: During the past decades, with the introduction of the recombinant DNA, hybridoma and transgenic technologies there has been an exponential evolution in understanding the pathogenesis, diagnosis and treatment of a large number of human diseases. The technologies are evident with the development of cytokines and monoclonal antibodies as therapeutic agents and the techniques used in gene therapy. Immunopharmacology is that area of biomedical sciences where immunology, pharmacology and pathology overlap. It concerns the pharmacological approach to the immune response in physiological as well as pathological events. This goals and objectives of this textbook are to emphasize the developments in immunology and pharmacology as they relate to the modulation of immune response. The information includes the pharmacology of cytokines, monoclonal antibodies, mechanism of action of immune-suppressive agents and their relevance in tissue transplantation, therapeutic strategies for the treatment of AIDS and the techniques employed in gene therapy. The book is intended for health care professional students and graduate students in pharmacology and immunology.

Download or read book Gene Technology written by Axel R. Zander and published by Springer. This book was released on 2012-01-21 with total page 547 pages. Available in PDF, EPUB and Kindle. Book excerpt: The 11 th meeting in "Modern Trends in Human Leukemia" took place from June 19 to 21, 1994 in Wilsede in the middle of the Liineburger Heide, South of Hamburg. Interwoven with the Leukemia program was the Nato-sponsored Symposium of the ASI-Series "Gene Technology in Analysis of Malignant and Inherited Human Diseases Related to Development" . The Wilsede meeting was continued on a ship of the Neva leading through lake Ladoga and lake Onega. The topics of both meetings included discussion on recent progress isolation and development of hematopoietic stem cells, genes crucial for development and diseases, methods of gene transfer, application of gene transfer; oncogenes and anti-oncogenes as targets for gene therapy; receptors and their ligands in normal development and diseases, immunology and immunotherapy, radiation biology, clinical leukemias and bone marrow transplantation. The Nato workshop concentrated not only on analysis of cell systems useful for somatic gene therapy, but also on actual themes directly related to correction of human diseases. The latter aspects emphasized themes related to biotechnology, the first part was by nature more general. We also included a few contributions that discussed perspectives for the future of gene therapy and possible relationships to evolution.

Download or read book Studies of in Utero Transplantation and Gene Transfer in Murine Fetal Liver Hematopoietic Stem Cells written by Margret L. Casal and published by . This book was released on 1999 with total page 147 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Human Hematopoietic Stem Cells Ex Vivo Expansion Gene Transfer and Molecular Characterization written by Yuk Yin Ng and published by . This book was released on 2003 with total page 132 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book New Methods for Lentiviral Based Hematopoietic Stem Cell Gene Therapy written by Katelyn Masiuk and published by . This book was released on 2019 with total page 101 pages. Available in PDF, EPUB and Kindle. Book excerpt: Hematopoietic stem cell (HSC) transplant with gene therapy has recently emerged as a successful clinical treatment of a number of previously incurable genetic blood diseases. This approach aims to permanently fix genetic defects in HSCs, a rare and specialized type of cell with the unique ability to regenerate the entire blood system throughout a patient's lifetime. In this approach, bone marrow (BM) or mobilized peripheral blood (mPB) is collected from a patient, enriched for HSCs, transduced with an engineered lentiviral vector (LV) encoding the correct genetic sequence, and transplanted back into the patient. After transplant, modified HSCs engraft in the BM and produce healthy blood cells throughout the patient's lifetime. While the last decade of research has yielded major advances including successful Phase I/II gene therapy clinical trials, clinical and commercial scaling of this technology to a broader range of patients and diseases has revealed a number of hurdles. One major limitation is the great expense and difficulty of producing clinical-grade LV, which I address in Chapters 2 and 3 by presenting two methods that improve the efficiency of LV transduction of HSC. In Chapter 4, I demonstrate the successful application of a new LV gene therapy for an autoimmune blood disease. Chapter 2 presents a method to enhance the enrichment of HSCs from the heterogeneous cell population obtained from the collection of bone marrow cells, addressing a critical limitation in creating cost-effective, clinical-grade LV vector. This method utilizes immunomagnetic beads to purify CD34+CD38- cells, a population highly enriched for HSCs beyond standard CD34+ selection. Using immune-deficient xenograft models, we demonstrate that enrichment of CD34+CD38- cells reduces gene therapy culture scale and lentiviral vector requirements by ~10-fold while still maintaining the long-term gene-marked engraftment required for clinical benefit. Therefore, this strategy represents an easily translatable method which can conserve valuable clinical grade LV preparations and could lower the cost per patient, or allow for the treatment of a greater number of patients. Chapter 3 presents a method to further improve HSC transduction efficiency with the use of two compounds: Prostaglandin E2 (PGE2) and poloxamer synperonic F108 (PS-F108). While transduction enhancement with each individual compound has previously been reported, the combination of these compounds leads to a synergistic and marked improvement in LV transduction of HSCs using a globin LV. Remarkably, this synergistic combination achieved a 6-fold improvement in gene transfer to long-term engrafting HSCs while using a LV dose 10-fold lower than the dose in our current clinical protocol. Thus this strategy has two major advantages: it reduces the amount of viral particles required to transduce HSCs, and also allows for better gene transfer and ultimate globin transgene expression, which is critical to improving clinical efficacy. Finally, chapter 4 demonstrates the effectiveness of a newly engineered LV for the treatment of a severe form of genetic autoimmunity called IPEX syndrome. IPEX is caused by mutations in FoxP3, the key lineage-determining transcription factor required for the development and function of regulatory T cells (Treg cells). We developed a new LV using endogenous human FOXP3 regulatory elements to restore FoxP3 expression in a developmentally appropriate manner. We use this LV to transduce HSCs and restore functional Treg development in a mouse model of FoxP3 deficiency and successfully rescue autoimmune defects associated with this phenotype. These findings demonstrate preclinical efficacy for the treatment of IPEX patients by autologous HSC transplant and may provide further insight into new cell therapies for autoimmunity. Collectively, the work described here advances the field of gene therapy by improving the efficiency of the manufacturing process and expanding the range of diseases which can be treated by this method.

Download or read book Transduction and Selection of Hematopoietic Stem Cells for Gene Therapy of Hemoglobin Disorders written by Kanit Bhukhai and published by . This book was released on 2015 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Recent clinical trials conducted in patients with hematopoietic congenital diseases have demonstrated the potential benefit of autologous hematopoietic stem cell (HSC) transplantation combined with gene transfer using integrative lentiviral vectors. However, the level of transduced HSCs was occasionally non optimal, resulting in partial correction of the diseases. In order to achieve high level HSC modification without increasing the concurrent risk of insertional mutagenesis and oncogene activation, we decided to develop methods aimed at selecting genetically modified stem cells rather than increasing their initial transduction rate. In order to demonstrate the feasibility of our approach, drug resistance genes encoding an antibiotic resistant protein or a dealkylating agent were introduced, together with a suicide gene, in a clinical 3-globin lentiviral vector specifically designed for patients with hemoglobin disorders. In vitro evaluation made with a vector encoding the dealkylating protein suggested that its expression was too low to provide full protection to the cells. lnterestingly, we demonstrated that the puromycin resistant gene allowed optimal ex vivo selection of genetically modified puromycin treated human HSC, provided that P-gp transporter inhibitors were added to the cells. Once selected, transduced HSC survived and were able to reconstitute human hematopoiesis in immunodeficient animal. Furthermore, the vector was able to express the therapeutic [3- globin gene for correction of hemoglobin disorders and to produce the suicide protein in vivo, for elimination of transduced stem cells if necessary.

Download or read book In Vivo Gene Transfer Into Mobilized Hematopoietic Stem Cells written by Maximilian Richter and published by . This book was released on 2017 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Pharmaceutical Biotechnology written by Daan J. A. Crommelin and published by CRC Press. This book was released on 2002-11-14 with total page 456 pages. Available in PDF, EPUB and Kindle. Book excerpt: The field of pharmaceutical biotechnology is evolving rapidly. A whole new arsenal of protein pharmaceuticals is being produced by recombinant techniques for cancer, viral infections, cardiovascular and hereditary disorders, and other diseases. In addition, scientists are confronted with new technologies such as polymerase chain reactions, combinatorial chemistry and gene therapy. This introductory textbook provides extensive coverage of both the basic science and the applications of biotechnology-produced pharmaceuticals, with special emphasis on their clinical use. Pharmaceutical Biotechnology serves as a complete one-stop source for undergraduate pharmacists, and it is valuable for researchers and professionals in the pharmaceutical industry as well.

Download or read book Heritable Human Genome Editing written by The Royal Society and published by National Academies Press. This book was released on 2021-01-16 with total page 239 pages. Available in PDF, EPUB and Kindle. Book excerpt: Heritable human genome editing - making changes to the genetic material of eggs, sperm, or any cells that lead to their development, including the cells of early embryos, and establishing a pregnancy - raises not only scientific and medical considerations but also a host of ethical, moral, and societal issues. Human embryos whose genomes have been edited should not be used to create a pregnancy until it is established that precise genomic changes can be made reliably and without introducing undesired changes - criteria that have not yet been met, says Heritable Human Genome Editing. From an international commission of the U.S. National Academy of Medicine, U.S. National Academy of Sciences, and the U.K.'s Royal Society, the report considers potential benefits, harms, and uncertainties associated with genome editing technologies and defines a translational pathway from rigorous preclinical research to initial clinical uses, should a country decide to permit such uses. The report specifies stringent preclinical and clinical requirements for establishing safety and efficacy, and for undertaking long-term monitoring of outcomes. Extensive national and international dialogue is needed before any country decides whether to permit clinical use of this technology, according to the report, which identifies essential elements of national and international scientific governance and oversight.

Download or read book The European Blood and Marrow Transplantation Textbook for Nurses written by Michelle Kenyon and published by Springer. This book was released on 2018-03-14 with total page 318 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book is open access under a CC BY 4.0 license. This textbook, endorsed by the European Society for Blood and Marrow Transplantation (EBMT), provides adult and paediatric nurses with a full and informative guide covering all aspects of transplant nursing, from basic principles to advanced concepts. It takes the reader on a journey through the history of transplant nursing, including essential and progressive elements to help nurses improve their knowledge and benefit the patient experience, as well as a comprehensive introduction to research and auditing methods. This new volume specifically intended for nurses, complements the ESH-EBMT reference title, a popular educational resource originally developed in 2003 for physicians to accompany an annual training course also serving as an educational tool in its own right. This title is designed to develop the knowledge of nurses in transplantation. It is the first book of its kind specifically targeted at nurses in this specialist field and acknowledges the valuable contribution that nursing makes in this area. This volume presents information that is essential for the education of nurses new to transplantation, while also offering a valuable resource for more experienced nurses who wish to update their knowledge.