Gene Delivery Using Viral Vectors - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 252

in this tissue. The stereotactic injection of only a few microliters of vector prepa-
ration allows the transduction of tens to hundreds of thousands of neurons in the
absence of major inflammatory or vector-specific immune response [218] . Various
studies are under progress in animal models of human neurodegenerative disorders,
for example, in primates, probing the potential value of the lentivector-mediated
transfer of sequences coding for various growth promoting or antiapoptotic factors
in Parkinson's disease. Lentivectors also seem to be the vehicle of choice for the
genetic alteration of retinal cells. HIV vector-mediated gene therapy appears prom-
ising for the treatment of recessive forms of inherited retinal degeneration. Finally,
one potential area of lentivector application is the ex vivo transduction of cells oth-
erwise difficult to transfect. One typical example is the human pancreatic islet cell,
but high-efficiency lentivector-mediated gene transfer has been obtained in a very
large number of either primary cells or well-known lines [219,220] . Lentiviruses
have also been successfully used for transfection of diabetic mice with the gene
encoding platelet-derived growth factor (PDGF).
5.6.3 Influenza Virus
Influenza viruses A, B, and C are very similar in overall structure. The size of virus
particle is 80-120 nm in diameter, and they are usually roughly spherical, although
filamentous forms can occur. These filamentous forms are more common in influenza
C, which can form cordlike structures up to 500 m long on the surfaces of infected
cells. However, despite these varied shapes, the viral particles of all influenza viruses
are similar in composition. These are made of a viral envelope containing two main
types of glycoproteins wrapped around a central core. The central core contains
the viral RNA genome and other viral proteins that package and protect this RNA.
RNA tends to be single stranded, but in some specific cases it is double stranded.
Unusually for a virus, its genome is not a single piece of nucleic acid; instead, it
contains seven or eight pieces of segmented negative-sense RNA, with each piece
containing either one or two genes. For example, the influenza A genome contains
11 genes on 8 pieces of RNA, encoding for 11 proteins: hemagglutinin (HA), neur-
aminidase (NA), nucleoprotein (NP), M1, M2, NS1, NS2(NEP), PA, PB1, PB1-F2,
and PB2. Influenza viruses can only replicate in living cells. Influenza infection and
replication is a multistep process: first, the virus must bind to and enter the cell, then
deliver its genome to a site where it can produce new copies of viral proteins and
RNA, assemble these components into new viral particles, and finally exit the host
cell [221] .
The study of reverse genetics has also allowed us to use influenza viruses for the
improvement of viral expression vectors. Initially in the influenza virus vectors, the
viral HA and the NA genes were manipulated to incorporate foreign genes for expres-
sion. But the expression level was not satisfactory, and the size of foreign genes able
to be incorporated was limited. By focussing on the NS1 gene, which is located on
the shortest of the eight influenza virus genome segments, those expressed with a
comparatively higher efficiency than the other viral genes, a new influenza vector
system has been developed by the manipulation of the NS gene [222] .