Drug – bio-affecting and body treating compositions – Whole live micro-organism – cell – or virus containing – Genetically modified micro-organism – cell – or virus
Reexamination Certificate
2000-06-26
2003-02-25
Guzo, David (Department: 1636)
Drug, bio-affecting and body treating compositions
Whole live micro-organism, cell, or virus containing
Genetically modified micro-organism, cell, or virus
C424S093100, C424S093600, C435S069100, C435S069700, C435S235100, C435S320100, C435S455000, C435S456000, C435S325000, C435S366000, C530S350000, C530S387100, C530S387300, C530S388220
Reexamination Certificate
active
06524572
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The current invention relates to the use of a fusion protein ligand to couple with an antibody species to target recombinant viruses to specific cell or tissue of interest. The current invention can be used in viral vector based gene therapy. In the use for gene therapy, current invention broadens the spectrum of diseases amenable to gene therapy using viral vectors, enhances the viral transfection efficiency in cells or tissues that are refractory to the viruses, and finally provides a safer and more flexible system for gene targeting. Current invention can also be used in experimental setting to selectively transfect specific cells or tissues of interest in a mixed cell or tissue environment.
2. Description of Prior Art
Infectious microorganism, especially those by virus, is characteristically tissue and/or species specific. This characteristic is named viral tropism. It has been known that the tropism is mainly associated with the fact that viral entry requires interactions between viral surface proteins and cellular surface receptors and, in some case, also cellular coreceptors and the fact that the viral receptors are expressed in a tissue and/or species specific manner. The tropism, however, presents a limitation in the ability to use viral vectors for gene therapy if intended target cells are not viral native host cells. It is difficult to transfect recombinant viruses to cells lacking these receptors. Therefore, studies of viral receptor expression, receptor-virus interaction and mechanism of viral entry is very important to gene therapy research.
In the case of adenovirus, infection by adenovirus requires binding of viral fiber protein to the extracellular domains of a recently identified 46-kD membrane protein named coxsackievirus/adenovirus receptor (CAR) (Bergelson, et al., 1997, Science 275, 1320-3.). Fiber is a trimer with a structure similar to a knob. The interaction between CAR and viral fiber is highly specific, and of high affinity (Louis, et al., 1994, J Virol 68, 4104-6, Bergelson, et al., 1998, J Virol 72, 415-9.). The structure of the fiber knob in complex with extracellular domain of CAR has been delineated recently (Bewley, et al., 1999, Science 286, 1579-83, Roelvink, et al., 1999, Science 286, 1568-71.). The identified binding site for CARs is on the side of the fiber knob with three CAR monomers bound per fiber knob trimer. The multivalency of CAR binding may contribute to the high efficiency of adenovirus infection. Following initial binding, the viral protein penton base binds via its Arg-Gly-Asp (RGD) motif to &agr;
v
&bgr;
3
or &agr;
v
&bgr;
5
integrins of cell membrane and this binding activates virus internalization via receptor-mediated endocytosis (Wickham, et al., 1993, Cell 73, 309-19.). Inside coated vesicles conformational changes of viral protein trigger the passage of adenovirus core particle through the cell membrane (Wickham, et al., 1994, J Cell Biol 127, 257-64, Wang, et al., 1998, J Virol 72, 3455-8.). Both high affinity binding by adenovirus and endocytosis events by the host cells are necessary for the adenoviral transfection to occur. Because of its ability to transfect a variety of quiescent tissues or cells and to maintain a long-term transgene expression adenovirus has been preferred over other available gene delivery systems for gene therapy. In fact, a majority of clinical trials currently underway use recombinant adenoviral vector based gene delivery systems.
Retroviruses are another example in which presence of viral receptors on host cells are critical for viral entry. Retrovirus infects cells in a two step mechanism. These viruses contain two envelope glycoprotein subunits designated surface (SU) and transmembrane (TM) which form an oligomeric complex on the viral surface and mediate viral entry. The SU protein contains the viral receptor binding determinants whereas the TM protein contains a hydrophobic transmembrane region and a separate hydrophobic segment that mediates virus-cell membrane fusion (Weiss, 1993, The Retroviridae 2, 1-107.). The first step of infection is the attachment of the viral particle via the surface protein of the retrovirus envelope (env) protein and that is followed by viral and cellular membrane fusion for viral uptake. The env protein is largely responsible for the tissue or species specificity of the retroviral infectivity. In the infection by human immunodeficiency virus (HIV), the soluble cell surface receptor is CD4 membrane protein. While G protein-coupled chemokine receptors, such as CXCR4 or CCR5, each acts as coreceptors for the syncytium-inducing T-cell tropic X4 strains (Feng, et al., 1996, Science 272, 872-7.) and primary non-syncytium-inducing macrophage tropic R5 strains, respectively (Deng, et al., 1996, Nature 381, 661-6.). It is also evident that many primary HIV isolates are in fact dual tropic, having the ability to utilize both CXCR4 and CCR5 as coreceptors, and are named as R5X4 isolates.
Another example is adeno-associated viruses (AAV). AAV has a linear single-stranded DNA and only undergo productive infection if the infected cells are co-infected with a helper virus (e.g., adeno- or herpesvirus) otherwise the genome becomes integrated in a latent state at a specific site on a human chromosome (Bems, 1996, Fields Virology.). Recombinant adeno-associated viruses are typically made by replacing viral genes with desired genes of interest or by simply adding the terminal AAV DNA sequences (ITRS) to these genes. In the case of type 2 AAV, membrane associated heparan sulfate proteoglycan is a receptor for viral infection (Summerford and Samulski, 1998, J Virol 72, 1438-45.).
Other examples include negative strand RNA viruses. These viruses infect cells by a variety of different mechanisms. For example, Influenza A viruses which have a segmented RNA genome, contain a surface hemagglutinin protein which binds to cell surface sialic acid receptors and mediates viral entry in a low pH endosome following receptor-mediated endocytosis (Lamb and Krug, 1996, Fields Virology.). The positive strand RNA viruses also infect cells by receptor mediated entry. For example, among the picomaviruses, different members of the immunoglobulin protein superfamily are used as cellular receptors by poliovirus, by the major subgroups of rhinoviruses, and by coxsackie B viruses, whereas an integrin protein is used by some types of ecoviruses and a low density lipoprotein receptor is used by minor subgroups of rhinoviruses (Rueckert, 1996, Fields Virology.). Following receptor binding, it is not yet fully understood what role receptor-mediated endocytosis plays for picomaviral entry, if indeed it is required. Paramyxoviruses containing a non-segmented RNA genome have two surface viral proteins, the hemagglutinin (HN) and fusion protein (F), required for viral entry which occurs at neutral pH. These viruses utilize sialic acid receptors, or protein receptors, such as CD46 used by measles virus, for viral entry (Lamb and Kolakofsky, 1996, Fields Virology.). Rhabdoviruses (e.g., VSV) also have a non-segmented RNA genome, contain a surface protein (G) which also binds to specific cell surface receptors and mediates viral entry in a low pH endosome. In some cases, however, a specific phospholipid, in steady of protein peptide, appears to be one of the receptors for VSV (Wagner and Rose, 1996, Fields Virology.). The herpesviruses which have large double-stranded DNA genomes, contain a number of surface glycoproteins involved in viral entry and utilize various cell surface receptors. For example, herpes simplex virus and cytomegalovirus entry involves binding to a heparin sulfate cell surface receptor and herpes simplex viruses use other proteins (e.g., HVEM) for viral entry (Montgomery, et al., 1996, Cell 87, 427-36.). In contrast, Epstein-Barr virus entry is initiated by binding to a completely distinct cell surface receptor, CR2 (Wolf, et al., 1993, Intervirology 35, 26-39.). Strategies have been described that allow one to engineer herpes simplex v
Guzo David
Rainbow Therapeutic Company
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