Gene Delivery Using Viral Vectors - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 253

Reverse genetics techniques have also been developed for the introduction of
specific mutations into the influenza A virus genes. These techniques allowed the
generation of influenza virus vectors expressing foreign antigens. The expression of
tumor-associated antigens (TAAs) by influenza virus vectors might represent a prom-
ising way to induce potent cellular immune responses against cancer cells, leading
to tumor regression. The small interfering RNAs (siRNAs) expressing plasmid con-
structs were made and targeted to sequences encoding the ribonucleoprotein mem-
ber, nucleoprotein (NP) and/or PA, of the influenza virus genome. RNA interference
(RNAi) can also be used to inhibit protein expression and replication of influenza
virus. RNAi treatment holds potential as a new approach to prevent avian influenza
[223-225] .
5.6.4 Human Papillomavirus
Papillomaviruses (PVs) are small, nonenveloped, icosahedral DNA viruses that can
replicate in the nucleus of squamous epithelial cells. Generally they consist of a single
molecule of double-stranded circular DNA around 8000 bp in size within a spherical
protein coat of 72 capsomeres. PVs are normally classified by the species they infect
(e.g., bovine, human, rabbit) and the type within species. The DNA of many PVs,
together with over 50 human viruses, has been cloned and sequenced. Even though,
there is a high degree of sequence divergence between species, all PVs share some
common features of genome organization. The genome is subdivided into an early
region containing proteins E1-E8, a late region containing genes L1 and L2 , and a long
control region (LCR) of transcription that includes the promoter and enhancer for the
viral early genes and the origin of replication. The early region encodes genes neces-
sary for viral DNA replication, cellular proliferation, and cellular transformation in
some viruses. The E1 and E2 genes encode E1 and E2 polypeptides that bind to the
LCR region and induce replication from the origin in the LCR region. The E5, E6, and
E7 proteins of the different PVs have proliferation and sometimes transforming activi-
ties. The late region, about 3 kb, codes for the capsid proteins. L1 is the major capsid
protein and is relatively well conserved among all the PV types. The L1 protein is about
500 amino acids in size and probably induces the major humoral and cell-mediated
responses to viral infection. The L2 proteins are about 500 amino acids in size and
account for only a small proportion of the virion mass. The LCR region contains an
origin of replication with binding sites for E1 and E2 and other cis -acting sequences in
the promoter and enhancer region.
Many viral vectors suffer from the disadvantage of random integration into the
chromosome, making the expression of the cloned genes dependent on the chromo-
somal context of the inserted DNA for the delivery of therapeutic genes into cells.
Papillomaviruses (PVs) are potentially attractive vector systems because of their
extrachromosomal replication in target cells. The PVs are small DNA viruses that
infect humans and a wide range of animals. Generally human papillomaviruses
(HPVs) induce benign proliferative squamous epithelial and fibroepithelial lesions
(warts and papillomas) in their natural hosts. Some HPVs are also involved in the
pathogenesis of anogenital cancer, specifically in cancer of the cervix [226] .