Expression of human adenosine deaminase in mice after transplantation of
genetically-modified bone marrow
M Kaleko, JV Garcia, WR Osborne and AD Miller
Program in Molecular Medicine, Fred Hutchinson Cancer Research Center,
Seattle, WA 98104.
A high titer retroviral vector was used to transfer a human adenosine
deaminase (h-ADA) cDNA into murine bone marrow cells in vitro. The h- ADA
cDNA was linked to the retroviral promoter, and the vector also contained a
neomycin phosphotransferase gene as a selectable marker. Infected marrow
was transplanted into syngeneic W/Wv recipients, and h- ADA expression was
monitored for 5.5 months. Several weeks after transplantation, h-ADA was
detected in the erythrocytes of all nine recipients, eight of which
expressed levels equal to the endogenous enzyme. This level of expression
persisted in two of six surviving mice, while expression in three others
stabilized at lower, but readily detectable, levels. Only one mouse had no
detectable h-ADA after 5.5 months. Vector DNA sequences with common
integration sites were found in hematopoietic and lymphoid tissues of the
mice at 5.5 months, providing evidence that hematopoietic stem cells had
been infected. Furthermore, all mice transplanted with marrow that had been
selected in G418 before infusion had multiple vector copies per genome.
While this category included the two highest h-ADA expressors, it also
included the negative mouse. Thus, multiple copies of the vector were not
sufficient to guarantee long-term h-ADA expression. Mice were monitored for
"helper virus" infections with an assay designed to detect a wide range of
replication-competent retroviruses, including those endogenous to the mouse
genome. No helper virus was detected in the two highest h-ADA expressors,
ruling out helper-assisted vector spread as a cause of the high h-ADA
expression. These results help provide a foundation for the development of
somatic gene therapy techniques to be used in the treatment of human
disease.
Volume 75,
Issue 8,
pp. 1733-1741,
04/15/1990
Copyright © 1990 by The American Society of Hematology
