Year: 1997
Type: dissertation
Abstract: The application of somatic gene transfer as a potential treatment in human\ndisease has progressed from speculation to reality in a short time\n[4,20,21,84,85,87,105,117,174]. In May 1989 the first clinical marker gene\nprotocol took place [145], followed by the first gene therapy protocol for the\ncorrection of adenosine deaminase deficiency in September 1990 [3,111.\nSubsequently, the number of gene marking and gene therapy trials grew\nexponentially. In 1996 more than 200 clinical protocols were approved\nworldwide [51 and more than 1,500 patients are now enrolled in gene transfer\ntrials (96). However, up to now the gene therapy trials have not brought clear\nclinical benefits for patients [5,31,51,98]. An important explanation for the lack\nof therapeutic effects is the very low gene transfer efficiency in human cells.\nOne of the main targets for gene therapy is the hematopoietic system\nbecause of well-developed procedures for bone marrow transplantation, the\nmany types and wide distribution of hematopoietic cells in the body and the\nexistence of many diseases and treatments that affect hematopoietic cells. The\ntarget cell for gene transfer is the primitive hematopoietic stem cell (HSC), which\nis present at low frequency in the bone marrow, is responsible for long-term\nhematopoiesis and gives rise to all hematological and lymphoid lineages.\nAlthough many research groups have reported transfer and long-term expression\nof genes in a significant percentage of murine hematopoietic cells [77,142,177]'\nthe achievements in larger outbred animals [81,93,150], nonhuman primates\n[14,164,1651 and humans [17,29,36,44,48,72,861 show only a small\npercentage of long-term expression in a limited number of recipients.\nThe aim of this study was to develop protocols which improve gene transfer\nof human HSC. For this purpose suitable assays were developed to determine the\nfrequency and proliferative capacity of different classes of human HSC.\nSubsequently, culture conditions for human HSC were studied. As a result, using\nnewly developed human HSC assays and optimized culture conditions, improved\nhuman HSC gene transfer protocols could be proposed. In the presented studies\nwe have opted for clinically relevant and acceptable protocols. This included the\nuse of target cells which are most frequently used in clinical autologous stem cell\ntransplantation, namely mobilized peripheral blood stem cells (PBSC) [1611 and\nserum-free transfection cultures in which target cells and virus-producing cells\nwere separated.
Author: Dimitri Breems
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1579374098