Patent Application: US-91606397-A

Abstract:
the subject invention concerns the production and novel use of human anti - hiv iga antibodies to confer passive immunity to hiv when administered in vivo to a subject . the iga antibodies of the subject invention are selectively transported to and secreted in the mucosal tissue of the subject . the iga antibodies can confer protection from hiv infection at locations within the body where hiv penetration usually occurs .

Description:
the subject invention concerns a method for protecting an individual from infection with the human immunodeficiency virus ( hiv ). specifically , iga antibodies to hiv are administered to an individual to provide that person with a passive immunity to hiv infection . in a preferred embodiment , a source of anti - hiv iga antibodies for passive immunity directed at the hiv virus would be from the breast milk of women who are seropositive for hiv antibody and had demonstrated that such antibody protected their newborn from hiv infection . preferably , such iga antibodies would be obtained from a pool of several different women infected with various strains of hiv . thus , the purified antibodies would represent an antibody preparation capable of protecting a person from a wide variety of hiv strains and hiv mutants . the purified immunoglobulin a from such sources is fully assembled including the secretory piece which is added by the mother &# 39 ; s secretory mammary glands . such an antibody can be applied onto the mucosal surface directly but lacks the capability of returning to the mucosa if injected systemically . however , the secretory piece of these molecules can be chemically removed , thereby allowing the iga antibody to be transported to the mucosa . in another embodiment of the invention , iga antibodies are produced through the creation of human hybridomas that produce antibody reactive with hiv . methods for producing such hybridomas are known to those skilled in the art . the source of the initial human lymphocytes is preferably an individual who has been exposed to the hiv virus and was undergoing surgery for hysterectomy , appendectomy of tonsillectomy . all of these organs are a rich source of b lymphocytes secreting iga antibody . the b lymphocytes are isolated using techniques that are specific for the tissue from which they are derived and immortalized using standard techniques for the production of human hybridomas . these hybridomas are then distributed into 96 well tissue culture plates containing the proper media and growth requirements . thereafter , cell supernatants from the wells containing hybridomas are removed and tested not only for reactivity with either hiv virus particles or recombinant hiv proteins , but also for their ability to neutralize the virus and protect human cd4 - bearing cells from infection . the hybridomas from wells containing such activity are removed , expanded and finally processed to obtain a single clone . the supernatants from the clone are retested using the same criteria to insure that the cloning process has been successful . the isotype of the secreted antibody is determined for each clone in order to identify those clones producing hiv - specific iga antibody . these human hybridoma cells are then expanded and the antibody isolated and purified . in a further embodiment , human b lymphocytes secreting iga antibodies can be obtained using severe combined immunodeficiency mice having a human immune system ( scid - hu mice ) reconstituted from human peripheral blood or fetal thymus , bone marrow , and lymph node . these mice can then be immunized with hiv viral particles or hiv encoded proteins including gp160 , gp120 , gp41 and peptide fragments thereof . after successive boosts with hiv antigen the human b lymphocytes can be rescued using epstein bar virus ( ebv ) transformation or phage library repertoires . in an alternative embodiment , anti - hiv monoclonal iga antibodies of the subject invention can be &# 34 ; humanized &# 34 ; through genetic engineering techniques known to those of ordinary skill in the art . specifically , monoclonal antibodies to hiv are prepared by first immunizing an appropriate animal , such as a mouse , with hiv viral particles or hiv proteins . hybridomas are then produced and screened for the production of hiv - reactive iga antibodies using standard techniques . after selecting high - affinity anti - hiv iga antibody - producing clones that are capable of neutralizing hiv , the cells are genetically engineered so that the variable region of the non - human antibody is joined to the constant regions from a human iga antibody to form a chimeric antibody . in a related procedure , the hypervariable regions from a non - human antibody combining site derived from an anti - hiv antibody can be &# 34 ; grafted &# 34 ; onto framework regions of human iga antibody . this latter technique , known as complementarity - determining region ( cdr ) grafting , provides for the production of a humanized iga antibody having a pre - selected non - human antibody binding site specific for a given epitope on an antigen . the iga antibodies prepared according to the subject invention are then purified for use in humans . if the antibody has the secretory piece still attached , then it may optionally be removed prior to administering the antibody to the subject . the administration of the iga antibodies may be according to any suitable method known in the art . for example , the antibodies can be injected intramuscularly or intravenously . the invention contemplates the administration of antibodies reactive to a single epitope or to multiple epitopes present on hiv viral particles or proteins . further , the antibodies used according to the subject invention may consist of monoclonal , polyclonal or a mixture of monoclonal and / or polyclonal antibodies . the antibodies may comprise whole antibody or fragments thereof , such as f ( ab ) 2 fab , etc . the antibodies can be administered in physiologically acceptable formulations that will minimize the immune response of the recipient to the antibodies . it is expected that additional administrations of the anti - hiv iga antibodies will be required over time as mucosal antibody titer decreases in an individual . dosage levels of the anti - hiv iga antibody to be administered , as well as appropriate formulations of the antibody preparation , can be readily determined by a person of ordinary skill in the art . the antibodies of the subject invention can be used to protect uninfected people , particularly people at risk for infection with hiv , such as medical care providers . moreover , the antibodies of the subject invention are also contemplated for use in assays for hiv , or for use as molecular weight standards , or as an inert protein in an assay . it should be understood that the embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and the scope of the appended claims . grimes , r . m ., a . g . randall , and l . n . nickey ( 1990 ) &# 34 ; 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