Pathogen-targeted biocatalysts

#Pathogenic organisms, including viruses, may be divided into different classes on the basis of their fate after being phagocytized. For instance, organisms that are promptly destroyed when phagocytized (i.e., S. pneumoniae, S. pyogenes) behave as extracellular parasites, damaging tissues only so long as they remain outside phagocytic cells.

#Pathogens that function as extracellular parasites owe their virulence to antiphagocytic surface components. Most pathogenic bacteria for example, maintain capsules comprising high-molecular-weight polysaccharides. The relation between capsules, phagocytosis and virulence is clearly exemplified by S. pneumoniae. A fully encapsulated S (smooth) strain is found to resist phagocytosis (in the absence of antibodies) and is highly virulent for mice, whereas its nonencapsulated R (rough) mutant is readily phagocytized and is essentially avirulent. However, enzymatic removal of the capsular polysaccharide, or combination with antibody, renders the S organisms both nonpathogenic and susceptible to phagocytosis.

#In contrast to extracellular parasites, there are two classes of intracellular parasites, both of which can multiply within phagocytic cells. One class of these intracellular pathogens comprise the obligate intracellular parasites (i.e., rickettsiae, chlamydiae). For these organisms, which include viruses, subtle differences in the cell surface receptors essential for their uptake may be important in resistance.

#FIG. 1 shows the double stranded nucleic acid sequence (SEQ ID No. 1) for a CD4 gene fragment.

#FIG. 2 shows 5' (SEQ ID No.3) and 3' (SEQ ID No.4) amplimers for PCR amplifying a catalytic domain fragment of tissue plasminogen activator (tPA).

#FIG. 3 shows various plasmids used to create a CD4/tPA fusion protein.

#Surface components of pathogens can play a major role in the survival of the pathogen, and therefor in its pathogenicity to a host. For instance, a given surface component can be crucial to the interaction of the pathogen with a host cell or tissue and is thus a critical component of the tissue-tropism of the pathogen. For intracellular pathogens, the surface component may be required for entry into the cell, while in the case of extracellular pathogens, particular surface components may be vital to the evasion of phagocytosis. Other surface components, such as the proteins, lipids and polysaccharides making up cellular walls, nutrient receptors including endocytic and pinocytic components, and chemotaxic receptors, can also be critical to the survival of the pathogen. The present invention provides a biocatalyst, directed against a surface component of a pathogen necessary for pathogenicity, such that action upon this component by the catalytic moiety destroys the functional integrity of this molecule and thereby disrupts pathogenicity.

#A distinct advantage of the biocatalyst of the present invention comes from the catalytic nature of the molecule's action on a pathogen. As stated above, interaction between surface components of pathogens and host cells have been implicated in pathogenicity. Disruption of these interactions has generally been accomplished by way of molecules which bind and mask essential determinants of at least one of the involved surface components. The typical masking molecule, such as a soluble receptor, requires a large number of molecules in circulation in order to effectively neutralize the pathogen, as the masking molecule acts stoichiometrically rather than catalytically. The pathogen-targeted biocatalysts of this invention direct a catalytic moiety to a surface component of the pathogen. The surface component is degraded and normal pathogen/cellular interactions are disrupted, thereby abrogating pathogenicity. The effectiveness of the biocatalyst is enhanced by the fact that the molecule, upon degradation of the surface component, is not "consumed" by the reaction.

#Most current regimens of therapy directed to disrupting pathogenicity by affecting the function of a surface component of a pathogen, act by masking the bound component.