EXPRESSION OF THE BLOOD-CLOTTING FACTOR-VIII CDNA IS REPRESSED BY A TRANSCRIPTIONAL SILENCER LOCATED IN ITS CODING REGION

Hemophilia A is caused by a deficiency of factor-VIII procoagulant (fV III) activity. The current treatment by frequent infusions of plasma-d erived fVIII concentrates is very effective but has the risk of transm ittance of blood-borne viruses (human immunodeficiency virus [HIV], he patitis viruses). Use of recombinant DNA-derived fVIII as well as gene therapy could make hemophilia treatment independent of blood-derived products. So far, the problematic production of the fVIII protein and the low titers of the fVIII retrovirus stocks have prevented preclinic al trials of gene therapy for hemophilia A in large-animal models. We have initiated a study of the mechanisms that oppose efficient fVIII s ynthesis, We have established that fVIII cDNA contains sequences that dominantly inhibit its own expression from retroviral as well as from plasmid vectors. The inhibition is not caused by instability of the fV III mRNA (t(1/2), greater than or equal to 6 hours) but rather to repr ession at the level of transcription. A 305-bp fragment is identified that is involved in but not sufficient for repression. This fragment d oes not overlap the region recently identified by Lynch et al (Hum Gen e Ther 4:259, 1993) as a dominant inhibitor of RNA accumulation, The r epression is mediated by a cellular factor (or factors) and is indepen dent of the orientation of the element in the transcription unit, givi ng the repressor element the hallmarks of a transcriptional silencer. (C) 1995 by The American Society of Hematology.