Current Limitations Of Retroviral Vectors

There are several problems investigators face in developing retroviral vectors that will be clinically effective. First, the vector genomes themselves have limited capacity for insertion of foreign sequences, based on the packaging constraints imposed by the viral core proteins; MuLV-based vectors cannot greatly exceed the 8.3-kb size of the MuLV genome. The stability of the engineered vectors can also be a concern. The presence of 2 copies of the vector genome in a viral particle and the process of reverse transcription both contribute to a relatively high level of rearrangement and instability, which is also influenced by the nature of the inserted sequences. The retroviral life cycle and the process of reverse transcription preclude the use of intron-containing sequences in a vector unless the gene is inserted in reverse orientation in the vector, and even with a cDNA copy of a gene, cryptic splice sites can become apparent when the gene is placed inside the retroviral vector.

Major improvements are also required in the overall efficiency of delivery of retroviral vectors. This will involve both the Env-directed entry process and overcoming any postentry blocks to transduction that might occur in specific cells, notably the inability to transduce nondividing cells. Once delivered to a target cell, improvements will be needed in the ability of the vectors to sustain gene expression long term, and for the therapeutic gene and its controlling sequences to respond to appropriate stimuli—be they natural developmental or physiological signals, or regulatory drugs administered to the patient. Finally, cost-effective ways to manufacture the vector at high enough titers will be required, with appropriate assurances of safety.

Specific issues related to the transduction of nondividing cells are dealt with in Section V.B, and manufacturing and safety concerns are discussed in Section VII. Below, we review current attempts to improve the specificity of gene delivery with retroviral vectors and the strategies designed to improve gene expression from those vectors.

0 0

Post a comment