Viruses lack the cellular machinery for self-reproduction. The viral genome codes for the proteins that constitute the protective outer shell (capsid) as well as for those proteins required for viral reproduction that are not provided by the host cell. The capsid consists of monomeric subunits of protein and serves to protect the virus's genetic material, detect cells suitable for infection, and initiate the infection by “opening” the target cell to inject DNA into the cytoplasm. After entering the cell, the virus's genetic material begins the destructive process of causing the cell to produce new viruses.
Viruses are classified as either DNA or RNA virus according to the nucleic acid type of their genetic material. The RNA viruses are divided into three groups: Group III—viruses possessing double-stranded RNA genomes, e.g. rotavirus; Group IV—viruses possessing positive-sense single-stranded RNA genomes including for example Hepatitis A virus, enteroviruses, rhinoviruses, poliovirus, foot-and-mouth virus, SARS virus, hepatitis C virus, yellow fever virus, and rubella virus; and Group V—viruses possessing negative-sense single-stranded RNA genomes inclusing, for example, the deadly Ebola and Marburg viruses, and the influenza, measles, mumps and rabies virus. Some negative sense RNA viruses contain also positive sense RNA and are referred to as ambisense viruses (e.g. some bunyaviruses).
The influenza virus is an RNA virus of the family Orthomyxoviridae, which comprises the influenzaviruses, isavirus, and thogotovirus. There are three types of influenza virus: Influenzavirus A, Influenzavirus B, or Influenzavirus C. Influenza A and C viruses infect multiple species, while influenza B virus infects almost exclusively humans.
Influenzavirus A has only one soecies, called the Influenza A virus. It is hosted by birds, but may also infect several species of mammals. Unusually for a virus, the influenza A virus genome is not a single piece of nucleic acid; instead, it contains eight pieces of segmented negative-sense RNA (13.5 kilobases total), which encode 11 proteins.
The influenza virus binds specifically to sialic acid sugars present on the surface of certain cells through the specific receptor hemagglutinin and is taken up through endocytosis into the cell, where the viral RNA (vRNA) is transported into the nucleus. The vRNA is then either exported back into the cytoplasm and translated, or remains in the nucleus, where it is transcribed into new vRNA molecules by RNA-dependent RNA polymerase (RDRP). Newly-synthesised viral proteins are either secreted through the Golgi apparatus onto the cell surface (in the case of neuraminidase and hemagglutinin) or transported back into the nucleus where some form the capsid shell and others bind vRNA that is packaged within the capsid to form new viral genome particles comprising the negative-sense vRNAs, RDRP and other viral proteins.
The newly formed viral particles leave the host cells by entering into plasma membrane protusion with clustered hemagglutinin and neuraminidase molecules. The mature virus then buds off from the cell in a sphere of host phospholipid membrane, acquiring hemagglutinin and neuraminidase with this membrane coat. Similarly to the stage of entry into the cell, the viruses adhere to the cell through hemagglutinin; the mature viruses then detach once the neuraminidase has cleaved sialic acid residues from the host cell. After the release of new influenza virus, the host cell dies.
The RNA-dependent RNA transcriptase lacks RNA proofreading activity and therefore mistakes introduced into the copied polynucleotide are not corrected. The frequency of errors is roughly a single nucleotide insertion error for every 10 thousand nucleotides, which is the approximate length of the influenza vRNA. Hence, nearly every newly produced influenza virus particle has a vRNA sequence that is different from other influenza virus particles. The separation of the genome into eight separate segments of vRNA allows mixing or reassortment of vRNAs, if more than one viral line has infected a single cell. The resulting rapid change in viral genetics produces antigenic shifts and allows the virus to infect new host species and quickly overcome protective immunity. For a compound intended to inhibit a process essential for viral propagation to be effective over time, it should therefore target constant regions of the target proteins and not—as is the case with current vaccines—target viral cell surface proteins that rapidly change their antigenicity.
There are two different replication processes for viruses containing RNA. In the first process, the viral RNA is directly copied using RDRP. In influenza virus, this complex consists of three polypeptides—PB1, PB2, and PA—collectively referred to as P proteins, while in other RNA viruses RDRP consists of a single polypeptide. The P protein complexes are normally associated with viral nucleocapsids, consisting of genomic RNA (vRNA) molecules covered with viral nucleoprotein. PB1 is the best characterized of the three P proteins; it contains five sequence blocks common to all RNA-dependent RNA polymerases and RNA-dependent DNA polymerases. PB2 has cap-binding and endonucleolytic activities which are necessary for viral mRNA synthesis. PA is indispensable for proper plus-strand copy RNA and vRNA synthesis, but no specific function in these processes has been assigned to it. Bipartite nuclear localization signals have been found in each of the three P proteins. Inside the nucleus, the RDRP enzyme uses the vRNA copy as a template to make hundreds of duplicates of the original RNA.
One representative of the RNA positive sense viruses is human hepatitis C virus. The positive sense RNA genome is directly translated into viral proteins without intermediate steps.
The most effective medical approaches to viral diseases thus far are vaccination to provide resistance to infection, and drugs that inhibit the viral proteins such as the cocktail of inhibitors used to treat human immunodeficiency virus (HIV)-AIDS. These drugs act on three critical step during the HIV cycle, i.e. replication, production of infectious viral particles; and fusion with the cellular membrane, thereby blocking entry into the host cell. A fourth step that may be interfered with is the budding, or release of the mature viral particles from the host cell. The three stages: replication, packaging and fusion with the cell membrane (for entry or release of viral particles), are the main essential processes in the viral propagation cycle amenable to manipulation with specific compounds for all enveloped viruses—whether positive or negative sense viruses.
Passive immunization with specific antiviral monoclonal or polyclonal antibodies has also proven effective both as prophylactic and therapeutic antiviral agents, e.g. in the case of human polyclonal antibodies against West Nile Virus or the monoclonal antibody Palivizumab approved for prevention and treatment of infection caused by respiratory syncitial virus.
Antibodies are made up of two identical heavy and two identical light chains. Each antibody has a constant region, which is the same for all immunoglobulins of the same class, and a variable region, which differs between immunoglobulins of different B cells, but is the same for all immunoglobulins produced by the same B cell. The variable regions of the heavy and light chains can be fused together to form a single chain variable fragment (scFv), which retains the original specificity of the parent immunoglobulin. “Designed” monoclonal antibody therapy is already being employed in a number of diseases and in some forms of cancer. Trastuzumab (Herceptin®, Genethech), a humanized monoclonal antibody that acts on the HER2/neu (erbB2) receptor, isl used in breast cancer therapy in patients with tumors overexpressing the HER2/neu receptor.
Virotherapy has been designed as a promising strategy for treatment of various diseases, especially cancer. It consists in the use of viruses by reprogramming viruses to only attack cancerous cells while healthy cells remained undamaged. The viruses are used most commonly as a vector directed to specifically target cells and DNA in particular.
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USPTO Backgrounds