.:Research
Viral Vectors for Gene Therapy
Adenovirus
Adenoviruses are viruses of the family Adenoviridae. They were first isolated from human adenoids, from which the name is derived. Adenoviruses have been isolated from a large number of species and tissue types. The human Ad family consists of more than 50 serotypes that can infect and replicate in a wide range of organs, such as the respiratory tract, the eye, urinary bladder, gastrointestinal tract and liver. Adenoviruses are medium-sized (60-90 nm), nonenveloped icosahedral viruses containing double-stranded DNA. The Ad genome consists of a double-stranded linear DNA molecule (size: ~36 Kb) with overlapping transcription units on both strands. Extensive splicing results in the production of over 50 proteins, 11 of which are structural virion proteins.
The viral life-cycle occurs in an early and a late phase, divided by the onset of viral DNA replication. Adenoviral genes fall into three major groups dependent on the time-course of their expression during the viral replicative cycle: early (E1A, E1B, E2, E3, and E4), delayed (IX and IVa2), and the major late transcription unit (see Figure 2). The latter is processed into five mRNAs (L1-L5) that share the same carboxy-terminus. These transcription units are transcribed by the cellular RNA polymerase II, while the viral associated (VA) RNA is transcribed by RNA polymerase III. The viral genome contains two identical origins for DNA replication within each terminal repeat. The E2 region encodes proteins required for replication, including the viral polymerase, and proteins from the E1 and E4 regions also contribute to efficient DNA replication. The genes products of the E3 region are involved in immune surveillance and suppression, but are non-essential for infection in vitro.
The Ad genome is packaged in a protein capsid. The fiber protein projects from the virion and the carboxy-terminal knob domain forms a high-affinity complex with a host cell surface receptor protein. For the majority of Ad serotypes this receptor is the Coxsackie-adenovirus receptor (CAR). In addition to attachment, efficient virus internalization requires an interaction between the viral penton base and the cellular integrin alpha v receptor. After entry the virus rapidly escapes from the endosome and transport to the nucleus is accompanied by gradual disruption of the virus particle. The viral genome is imported through the nuclear pore and associates with the nuclear matrix to facilitate initiation of the primary transcription events. The Ad genome is transcribed and replicated at discrete replication centers in the nucleus of the infected cell and the viral DNA does not normally integrate into the host genome.
Adenoviral infection causes an initial non-specific host response with synthesis of cytokines (tumor necrosis factor, interleukin 1 and 6) followed by a specific response of cytotoxic T lymphocytes (CTLs) directed against virus-infected cells that display viral peptide antigens. In addition, there is activation of B cells and the necessary CD4-positive T cells, leading to a humoral response. Serologic surveys found antibodies against Ad serotypes 1, 2, and 5 in 40-60% of children.
