Blood, Vol. 94 No. 2 (July 15), 1999:
pp. 465-474
Correction of Uroporphyrinogen Decarboxylase Deficiency
(Hepatoerythropoietic Porphyria) in Epstein-Barr Virus-Transformed
B-Cell Lines by Retrovirus-Mediated Gene Transfer: Fluorescence-Based
Selection of Transduced Cells
Antonio Fontanellas,
Frédéric Mazurier,
François Moreau-Gaudry,
Francis Belloc,
Cécile Ged, and
Hubert de
Verneuil
From the Laboratoire de Pathologie Moléculaire et
Thérapie Génique, FR 60 Biologie des Greffes,
Université Victor Segalen Bordeaux 2, Bordeaux, France; and the
Laboratoire d'Hémobiologie, Hôpital du
Haut-Lévêque, CHU de Bordeaux, Bordeaux, France.
Hepatoerythropoietic porphyria (HEP) is an inherited metabolic
disorder characterized by the accumulation of porphyrins resulting from
a deficiency in uroporphyrinogen decarboxylase (UROD). This autosomal
recessive disorder is severe, starting early in infancy with no
specific treatment. Gene therapy would represent a great therapeutic
improvement. Because hematopoietic cells are the target for somatic
gene therapy in this porphyria, Epstein-Barr virus-transformed B-cell
lines from patients with HEP provide a model system for the disease.
Thus, retrovirus-mediated expression of UROD was used to restore
enzymatic activity in B-cell lines from 3 HEP patients. The potential
of gene therapy for the metabolic correction of the disease was
demonstrated by a reduction of porphyrin accumulation to the normal
level in deficient transduced cells. Mixed culture experiments
demonstrated that there is no metabolic cross-correction of deficient
cells by normal cells. However, the observation of cellular expansion
in vitro and in vivo in immunodeficient mice suggested that genetically
corrected cells have a competitive advantage. Finally, to facilitate
future human gene therapy trials, we have developed a selection system
based on the expression of the therapeutic gene. Genetically corrected
cells are easily separated from deficient ones by the absence of
fluorescence when illuminated under UV light.
