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Next Article 
Blood, Vol. 95 No. 12 (June 15), 2000:
pp. 3645-3651
PLENARY PAPER
Megakaryocyte-targeted synthesis of the integrin
 3-subunit results in the phenotypic correction of
Glanzmann thrombasthenia
David A. Wilcox,
John C. Olsen,
Lori Ishizawa,
Paul F. Bray,
Deborah L. French,
Douglas A. Steeber,
William R. Bell,
Michael Griffith, and
Gilbert C. White II
From the Center for Thrombosis and Hemostasis, Departments of
Medicine and Pharmacology, Cystic Fibrosis/Pulmonary Research and
Treatment Center, University of North Carolina, Chapel Hill, NC; Nexell
Therapeutics Inc., Irvine, CA; the Department of Medicine, Johns
Hopkins University, Baltimore, MD; the Department of Medicine, Mount
Sinai School of Medicine, New York, NY; and the Department of
Immunology, Duke University Medical Center, Durham, NC.
Glanzmann thrombasthenia is an inherited bleeding disorder
characterized by qualitative or quantitative defects of the
platelet-specific integrin,  IIb 3. As a result,
IIb3 cannot be activated and cannot bind to
fibrinogen, leading to a loss of platelet aggregation. Thrombasthenia
is clinically characterized by mucocutaneous hemorrhage with episodes
of intracranial and gastrointestinal bleeding. To develop methods for
gene therapy of Glanzmann thrombasthenia, a murine leukemia virus
(MuLV)-derived vector,  889PlA23, was
transduced into peripheral blood CD34+ cells from 2 patients with thrombasthenia with defects in the 3 gene. The human IIb promoter
was used in this vector to drive megakaryocyte-targeted expression of
the wild-type 3 subunit. Proviral DNA and
IIb3 biosynthesis were detected after in vitro
differentiation of transduced thrombasthenic CD34+ cells
with megakaryocyte growth and development factor. Flow cytometric
analysis of transduced patient samples indicated that 19% of
megakaryocyte progeny expressed IIb3 on the surface
at 34% of normal receptor levels. Treatment of transduced
megakaryocytes with a combination of agonists including epinephrine and
the thrombin receptor-activating peptide induced the
IIb3 complex to form an activated conformation
capable of binding fibrinogen as measured by PAC-1 antibody binding.
Transduced cells retracted a fibrin clot in vitro similar to
megakaryocytes derived from a normal nonthrombasthenic individual.
These results demonstrate ex vivo phenotypic correction of Glanzmann
thrombasthenia and support the potential use of hematopoietic
CD34+ cells as targets for IIb promoter-driven MuLV
vectors for gene therapy of platelet disorders.
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