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Oligodeoxyribonucleotide Uptake in Primary Human Hematopoietic
Cells Is Enhanced by Cationic Lipids and Depends on the
Hematopoietic Cell Subset
Ralf Kronenwett,
Ulrich Steidl,
Michael Kirsch,
Georg Sczakiel, and
Rainer Haas
From Klinische Kooperationseinheit Molekulare
Hämatologie/Onkologie and Forschungsschwerpunkt Angewandte
Tumorvirologie, Deutsches Krebsforschungszentrum, Heidelberg, Germany;
and Medizinische Klinik und Poliklinik V, Universität
Heidelberg, Heidelberg, Germany.
The use of antisense oligodeoxyribonucleotides (ODN) is a potential
method to switch off gene expression. The poor cellular uptake of ODN
in primary cells still is a limiting factor that may contribute to the
lack of functional efficacy. Various forms of cationic lipids have been
developed for efficient delivery of nucleic acids into different cell
types. We examined the two cationic lipids DOTAP and DOSPER to improve
uptake of ODN into primary human hematopoietic cells. Using a
radiolabeled 23-mer, ODN uptake into blood-derived mononuclear cells
could be increased 42- to 93-fold by DOTAP and 440- to 1,025-fold by
DOSPER compared with application of ODN alone. DOTAP was also effective
for delivery of ODN into leukocytes within whole blood, which may
resemble more closely the in vivo conditions. As assessed by
fluorescein isothiocyanate-conjugated ODN both cationic lipids
enhanced cytoplasmic accumulation of ODN in endosome/lysosome-like
structures with a partial shift of fluorescence to the whole cytoplasm
and the nucleus following an incubation of 24 hours. ODN uptake by
cationic lipids into different hematopoietic cell subsets was examined by dual-color immunofluorescence analysis with subset-specific monoclonal antibodies. We found a cell type-dependent delivery of ODN
with greatest uptake in monocytes and smallest uptake in T cells.
CD34+ cells, B cells, and granulocytes took up ODN at an
intermediate level. Uptake of ODN into isolated CD34+
cells could be increased 100- to 240-fold using cationic lipids compared with application of ODN alone. Stimulation of
CD34+ cells by interleukin-3 (IL-3), IL-6, and stem cell
factor did not significantly improve cationic lipid-mediated ODN
delivery. Sequence-specific antisense effects in clonogenic assays
could be shown by transfection of bcr-abl oncogene-directed
antisense ODN into primary cells of patients with chronic myelogenous
leukemia using this established protocol. In conclusion, cationic
lipids may be useful tools for delivery of antisense ODN into primary hematopoietic cells. These studies provide a basis for clinical protocols in the treatment of hematopoietic cells in patients with
hematologic malignancies and viral diseases by antisense ODN.
Blood, Vol. 91 No. 3 (February 1), 1998:
pp. 852-862
© 1998 by The American Society of Hematology.
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