Gene transfer by lentiviral vectors is limited by nuclear translocation and rescued by HIV-1 pol sequences

Gene-transfer vectors based on lentiviruses are distinguished by their ability to transduce non-dividing cells(1,2). The HIV-1 proteins Matrix, Vpr and Integrase have been implicated in the nuclear import of the viral genome in non-dividing cells(3-5). Here we show that a sequence within polis also required in cis. It contains structural elements previously associated with the progress of reverse transcription in target cells(6-9). We restored these elements in cis within late-generation lentiviral vectors(10,11). The new vector transduced to a much higher efficiency several types of human primary cells, when both growing and growth-arrested, including haematopoietic stem cells assayed by longterm repopulation of NOD/SCID mice. On in vivo administration into SCID mice, the vector induced higher plasma levels of human clotting factor IX (F.IX) than non-modified vector. Our results indicate that nuclear translocation of the genome is a rate-limiting step in lentiviral infection of both dividing and non-dividing cells, and that it depends on protein and nucleic acid sequence determinants. Full rescue of this step in lentivirus-based vectors improves performance for gene-therapy applications.