Hepatitis B virus binding proteins and uses thereof

Details Hepatitis B virus binding proteins and uses thereof Hepatitis B virus binding proteins and uses thereof

1999-09-30

2003-07-08

Housel, James (Department: 1648)

Drug, bio-affecting and body treating compositions

Antigen, epitope, or other immunospecific immunoeffector

Virus or component thereof

C435S189000, C435S139000, C435S235100, C514S04400A, C530S350000, C530S390100, C536S023600

Reexamination Certificate

active

06589534

ABSTRACT:

FIELD AND BACKGROUND OF THE INVENTION
The present invention relates to a group of genes, and the proteins encoded thereby, which are capable of interfering with Hepatitis B virus (HBV) infection and to methods for identifying, purifying, isolating and characterizing related genes and gene products. The present invention further relates to isolation of soluble forms of the cellular receptor(s) for HBV on hepatocytes from bodily fluids, including, but act limited to, urine, and to purification of these soluble form binding proteins by means including, but not limited to, affinity columns. The present invention further relates to the use of these genes and their translation products to establish experimental models for HBV infection, whether in cell culture or in animals. The present invention further relates to the use of these genes and their translation products for therapeutic purposes. The present invention further relates to the use of these genes and their translation products to screen for additional binding protein interactions. The present invention further relates to the use of these genes and their translation products to prepare specific detectors of these proteins, including, but not limited to, antibodies, phage-display libraries, specific PCR primers, lectin, DNA probes, RNA probes, and non-antibody proteins for diagnostic and therapeutic purposes.
Hepatitis B virus (HBV) is an enveloped RNA virus that infects human liver and replicates via reverse-transcription of the pregenomic RNA Infected patients develop acute hepatitis, which is often self-limiting, but may develop into chronic hepatitis with high risk of liver cirrhosis and primary liver carcinoma in roughly 10% of all cases. The World Health Organization estimates that there will be 400 million carriers Worldwide in year 2000. Effective vaccines exist, but anti viral drugs with good and long term efficacy are not available. Little is known about how HBV infects liver cells and the HBV cellular receptor(s) remain unknown. Many proteins have been identified which bind to the viral envelope associated proteins, HBsAg, or related proteins, but none are considered genuine HBV receptors (reviewed in De et al., 1991 and in references cited therein). Some of these binding proteins are found in serum and some in hepatocytes. None of these molecules have been convincingly tied to infectivity, disqualifying them as genuine HBV receptors. These molecules are of three types, S binding proteins, preS2 binding proteins, and preS1 binding proteins. A brief summary of the characteristics of the three groups is provided herein.
The S binding proteins: HBsAg containing only the S protein binds to a 34-kDa liver protein, which is identified as the phospholipid-binding protein endonexin II (also known as annexin V). Endonexin II has calcium channel activity and it thought to be located primarily, but not exclusively, intracellularly. The biological significance of this remains unclear, as the observed interaction may simply reflect the known ability of endonexin 11 to bind phospholipids, which are abundant in HBsAg lipoprotein. It was subsequently demonstrated that delipidated HBsAg had a drastically diminished capacity to bind endonexin II, leading to speculation that it might play a role in a postbinding membrane fusion event.
It has also been demonstrated that plasma membranes, derived from human liver, contain apolipoprotein H (Apo H), a 46-kDa protein which binds HBsAg. This protein is a glycoprotein with four N-linked carbohydrate chains, which is present in the serum and is not an integral transmembrane protein of the hepatocyte. Its role in infection is uncertain, Moreover, it has been proven that the interaction between Apo H and HBsAg involves triglycerides and not HBV proteins. However, Apo H might play a role in delivery of the virus from the periphery to the liver.
Since binding of these molecules does not involve the preS determinant, they are unlikely to be the sole component of HBV attachment.
The preS2 binding proteins: Some researchers presumed that HBV binds to liver cells via a polymerized form of human serum albumin (pHSA) because a correlation between high viremia and the presence of a so-called pHSA receptor was observed. The preS2-specific domain does possess a pHSA binding activity, however, only pHSA from human or chimpanzee serum binds to preS2. Moreover, pHSA binds to liver cells, albeit in a non-species specific fashion. Furthermore, membranes from fresh human liver are able to bind natural HBs spheres or recombinant preS2 when they art pretreated with pHSA. These observations would suggest that the preS2 domain acts via pHSA as a species- and organ-specific attachment site of HBV except that identification of the exact binding site for pHSA within the preS2 domain is controversial.
The potential importance of pHSA binding for HBV infection has reduced by the observation that native albumin in physiologic concentrations blocks the binding of pHSA to HBsAg. This finding is especially significant considering that the minute concentration of natural pHSA present is serum is negligible when compared with the serum albumin concentration.
The N-linked glycan at the amino end of the preS2 domain has also been suggested as a potential binding site for human hepatocytes on the preS2 domain. This suggestion stems from an unusual glycan structure composed of one mannose chain and two complex chains which is liver specific and able to bind directly to HepG2 cells. Selective removal of this preS2 glycan reduces the preS2 binding by 70%.
It has also been reported that anti-idiotypic antibodies, raised against an epitope localized in the N-terminal part of preS2 protein, recognized human fibronectin, a component of the extracellular matrix. This binding is thought to be species specific because no binding was found between the preS2-associated epitopes with mouse liver. It is currently hypothesized that fibronectin may contribute to the initial binding of the circulating virus.
The preS1 binding proteins: Many researchers suggest possible roles for preS1 binding molecules in viral entry, although no conclusive evidence that these proteins play a role in permissive infection is available.
A portion of preS1, identified as being involved in attachment to HepG2 cells, is highly homologous to the Fc moiety of the &agr;-chain of immunoglobulin A (IgA). Since IgA binds to liver plasma membranes, a common receptor for the attachment of HBV and IgA to human liver cells has bean proposed. However, known receptors for IgA do not appear to be the receptors for HBV.
Anti-idiotypic antibodies have been used to paratope anti-preS(21-47) antibodies, which may represent a mirror image of the binding site on the receptor and thus be able to react with the receptor. These antibodies reacted with a 35-kDa protein and with three other related components of 40-, 43-, and 50-kDa in HepG2 membrane extracts. The 35-kDa protein, identified as the human liver glyceraldehyde-3-phosphate-dehydrogenase (GAPD) is a key enzyme for glycolysis, and the 50-kDa protein seems to contain intrachain disulfide bonds.
In addition, 31-kDa proteins that gross-linked in vitro to a synthetic preS1 peptide (amino acids 21 to 47) has also been identified
Other researchers also identified a 50-kDa protein in normal human serum which interacts with the epitopes localized within the preS1 and preS2 domains. They characterized this molecule as a glycoprotein with N-linked carbohydrate chains, which requires intact disulfide bonds in order to bind preS proteins. This 50-kDa protein blocks the binding of the preS1- and preS2-specific MAbs to HBV. This protein was detected on the surface of human hepatocytes by specific monoclonal antibodies, but not on hepatocytes from other species or in HepG2 cell membranes.
It has also been argued that the asialoglycoprotein receptor on the surface of hepatocytes a responsible for the binding of HBV, mediated by an epitope located in the preS1 domain,
As the expression of the asialoglycoprotein receptor is exclusive to hepatoc

Affiliated with

Also associated with

Rating

Hepatitis B virus binding proteins and uses thereof does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Hepatitis B virus binding proteins and uses thereof, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Hepatitis B virus binding proteins and uses thereof will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-3064656