WO1994008618A1

WO1994008618A1 - Oral tolerance and immune suppression in the treatment of aids

Info

Publication number: WO1994008618A1
Application number: PCT/US1993/009797
Authority: WO
Inventors: Larry I. Benowitz; J. Roberto Trujillo; Carleen A. Irwin
Original assignee: Children's Hospital
Priority date: 1992-10-13
Filing date: 1993-10-13
Publication date: 1994-04-28
Also published as: AU5359094A

Abstract

A method of diagnosis and treatment of AIDS-related disorders has been developed based on the presence of an autoimmune response, evoked by infection with the human immunodeficiency virus (HIV), which then leads to the destruction of cells in the immune and nervous systems.

Description

ORAL TOLERANCE AND IMMUNE SUPPRESSION IN THE TREATMENT OF AIDS

Background of the Invention

The present invention is in the general area of treatment of AIDS, in particular treatment of AIDS Dementia Complex.

Over 100,000 cases of the acquired immunodeficiency syndrome (AIDS) have been reported in the United States alone. It is believed that the main cause of AIDS is infection with a retrovirus that initially infects CD4+ T lymphocytes (helper T cells) , then spreads to other tissues. An additional 1.5 to 2 million Americans are estimated to have been infected with the Human Immunodeficiency Virus type 1 (HIV-1) , but do not yet have AIDS. Most of these people will eventually develop the disease, for which no cure exists. Worldwide, the numbers are much larger.

AIDS is characterized by a steady drop in the number of CD4+ helper T lymphocytes. The loss of the T lymphocytes greatly increases the susceptibility of the patient to infection and to a greater likelihood of cancer. Typically, when HIV infection is first detectable by standard diagnostic tests, the CD4+-cell concentration is often close to the normal level of about 800 cells per cubic millimeter of blood, and the patient feels well. Usually within six months to a year, chronic lymphadenopathy develops. Over time, the number of the patient's CD4+ T cells continues to drop to below 400 and the patient's cell mediated immunity declines in effectiveness, leading to chronic infections of the skin and mucous membranes, then disseminated systemic infections. Throughout the course of HIV infection people may also develop cancers and disorders of the central nervous system.

An aspect of the disease is the so-called AIDS Dementia Complex (ADC) , which occurs in up to two- thirds of adult AIDS patients, as reported by Price et al , Science 239: 586-592 (1988). ADC is characterized by poor memory, inability to concentrate, apathy, and psychomotor retardation. For 80% of affected patients, the symptoms progress rapidly, with the full-blown dementia complex developing within a year. The histopathologic features of ADC consist of gliosis of the cerebral cortex and subcortical nuclei, focal necrosis of gray and white matter, perivascular inflammation, atypical enlargement of oligodendrocyte nuclei, the formation of microglial nodules and ultinucleated giant cells, and demyelination in the white matter.

HIV-1 is a double stranded RNA retrovirus. A DNA polymerase first makes a single-strand DNA copy of the viral RNA, then a ribonuclease destroys the original RNA, the polymerase makes a second DNA copy, using the first one as a template. The polymerase and ribonuclease together are called the reverse transcriptase. The viral double-stranded DNA migrates to the cell nucleus where a viral enzyme splices the HIV genome into the host cell's DNA. The viral genome is then replicated with the cell. Periodically the cells produce new virus particles. The particles or virions are assembled and migrate to the periphery of the cell as they are produced, where they attach to the inside of the cell membrane. As the precursors aggregate, they bind to one another and form a spherical structure that bulges outward under the cell membrane. This buds out from the cell, carrying the envelope protein, which is made and transported to the cell surface independently of the core protein. The envelope protein consists of a glycoprotein, gpl20, which rests outside the cell, and gp41, which is embedded stemlike in the membrane. To infect a cell, gpl20 binds to the receptor molecule CD4, present in large numbers on T4 cells, but also present in at least trace amounts on many other kinds of cells. In the nervous system, only the smallest fraction of neurons or glia show evidence of viral infection, suggesting a mechanism other than direct viral attack (Price et al , Science 239: 586-592). Giulian et al (Science 250: 1593-1596) have reported that infection of macrophages with HIV-1 leads to the production of a factor toxic to neurons, while others have shown that gpl20, the HIV coat protein, is itself highly toxic to neurons (Brenneman et al , Nature 355: 639-642) , perhaps through calcium-dependent mechanisms involving a synergistic action with excitatory amino acids in activating the NMDA class of glutamate receptors (Lipton et al , Neuron 7: 111-118).

Classically, AIDS has been viewed as a direct consequence of the destruction of CD4-positive lymphocytes by the HIV-1 virus. However, this view is difficult to reconcile with the observations that only a minute fraction of lymphocytes become infected by the virus (Harper, et al., Proc. Natl. Acad. Sci., USA, 83:772-776, 1986), and that, at least initially, patients with AIDS show enhanced activity of the immune system (Dalgleish, J. Royal Coll. Physicians, London, 26:152-158, 1992). Kion and Hoffmann, Science 253: 1138-1140 (1991), recently reported that mice exposed to cells from another mouse strain developed antibodies that react with the major glycoproteins of the HIV-1 virus, gpl20 and gp24, along with anti- idiotypic antibodies to major histocompatibility components. See also Dalgleish, J. Royal College of Physicians London 26(2), 152-158 (1992). Accordingly, it is possible that autoimmunity could be a major factor leading to the collapse of the immune system in AIDS. However, as recognized by Hoffmann, et al., the mechanism proposed by Hoffmann, et al., Proc. Natl. Acad. Sci. USA 88: 3060-3064 (1991), is difficult to reconcile with respect to the different data observed in patients divided into groups consisting of homosexuals, hemophiliacs, Africans, and babies. Moreover, it is difficult to understand how one could utilize this theory to assist in the control or treatment of AIDS patients, or even test it.

It is therefore an object of the present invention to provide a method, and compositions used therein, for the diagnosis and treatment of aspects of AIDS involving autoimmune reactions.

It is a further object of the present invention to provide a method, and compositions used therein, for the diagnosis and treatment of AIDS related dementia.

Summary of the Invention

A method for diagnosis and treatment of AIDS- related disorders has been developed based on the presence of an autoimmune response, evoked by infection with the human immunodeficiency virus (HIV) , which then leads to the destruction of cells in the immune and nervous systems. Patients with HIV infection, and particularly those with the AIDS- Dementia Complex (ADC) , were examined and found to carry antibodies in their sera or cerebrospinal fluid (CSF) to brain proteins. Proteins from normal human brain were separated on SDS-polyacrylamide gels, transferred to nitrocellulose membranes, and probed with sera and CSF from patients who were either HIV-1- positive but who did not evidence signs of dementia (HIV+/ADC-) , patients with AIDS who did show the AIDS- dementia complex (ADC) , and normal controls. Results of these studies showed that sera from several HIV- infected patients contained antibodies that reacted with particular human brain proteins. Sera from a normal control group did not contain these autoantibodies. The results supported the proposal that autoimmunity could contribute to the clinical manifestations of AIDS. Such autoimmunity could arise if HIV antigens induced the development of an immune response, cellular or humoral, which then led to an attack of normal cells in the nervous system (or other systems) whose molecular constituents resemble viral antigens.

To determine whether immunological cross- reactivity exists between viral antigens and components of the human body, antibodies to a major determinant of HIV-1 pathogenicity, the V3 loop of the gp 120 protein, were used to identify proteins present in the normal human brain and other organs that might be targets of such an autoimmune response. These studies led to the identification of three proteins present in the brain and in some other tissues, including liver, which were recognized by anti-gpl20 V3 monoclonal antibodies. The cDNA encoding one of these proteins was isolated and expressed.

The presence of the anti-HIV antibodies that cross-react with normal human proteins should be useful as a diagnostic predictor of patients who may develop AIDS related dementia and other aspects of AIDS-related disorders. Moreover, cDNA encoding the cross-reactive portions of the protein, or the protein itself, can be used to make peptides for induction of tolerance in AIDS patients who have, or who may develop, AIDS related dementia based on the reactivity of the peptides with either the patient's T lymphocytes or autoantibodies.

Immunologically cross-reactive proteins are found on other tissues. The same process can be used to identify and treat patients having disorders arising from the presence of an autoimmune reaction involving these other tissues. The methodology can also be used to identify other proteins involved in an autoimmune response in AIDS patients, for development of additional diagnostics and treatments. Brief Description of the Drawings

Figure 1 is a photograph of SDS-PAGE: Proteins from the frontal cortex of a normal subject were separated by SDS-polyacrylamide gel electrophoresis, transferred to nitrocellulose membranes, and probed with sera from either control subjects, subjects carrying the HIV-1 virus but without signs of the AIDS-dementia complex (ADC) , or subjects with ADC. Prominent proteins recognized by antisera from one or more patients are indicated by arrows . Proteins from the frontal cortex of a normal 40-year old male (presumed HIV-negative) were solubilized (approximately 1 mg protein/ml buffer containing 1% SDS [sodium dodecyl sulfate], 5% β-mercaptoethanol, 0.125 M Tris-HCl pH 6.8, 10% glycerol, heated at 95°C 5 min) and separated by SDS-polyacrylamide gel electrophoresis (SDS PAGE) , with a 10 cm resolving gel (10% acrylamide, pH 8.8) and 3 cm stacking gel (4.5% acrylamide, pH 6.8), as described by U.K. Laemmli (Nature 227:680-685, 1970), for approximately 1200 V- h, until the tracking dye (bromphenol blue) was 1 cm from the end; the gel was equilibrated 1 h in 0.125 M Tris-glycine buffer pH 8.8, 20% methanol, and the proteins transferred electrophoretically to nitrocellulose membrane (0.22 μm pore size; Bio-Rad) , using the same buffer at 0.6 A for 4 h, as described by Towbin et al. (Proc. Natl. Acad. Sci., USA 76:4350- 4354, 1979). The membrane containing the electrophoretically transferred proteins was cut into strips about 2 mm wide, which were reacted with sera from the different populations as indicated. Membranes were first blocked in 5% BSA, then reacted with sera diluted 1:1000 (in PBS containing 2% BSA + 0.2% Tween-20) . Following a wash, binding of human antibodies to proteins was visualized using a biotinylated secondary antibody to human IgG (made in sheep) , streptavidin-biotin-[HRP] complex (1/500) , and developed with diaminobenzidine (DAB) .

Figures 2A-2C are a Western blot using a panel of monoclonal antibodies, raised in mice against defined viral proteins, to probe the western blot containing the brain proteins from a normal subject (an automobile accident victim presumed not to have had AIDS) . The antibodies were raised to the so- called V3 loop of gpl20 (lane 1) , undefined sequences in gpl20 (lanes 2 and 3) , gp41 (lane 4) , p24 (lane 5) , pl7 (lane 6), Vpu (lane 7), and Vpr (lane 8; the latter two are regulatory proteins of the virus) . Figure 2A uses a panel of antibodies to various molecular constituents of the HIV-1 virus to probe western blots containing the proteins of the same normal subject shown in Fig. 1. Only the antibodies to the V3 region of gpl20 (lane 1) reacted with specific proteins. Figure 2B and 2C reacts proteins from two other normal human subjects with either the monoclonal antibody generated to amino acids 308-322 of gpl20 (left strip in B and C) or to the entire length of gpl20 (right strips in B and C) . Subsequent work has shown that the second antibody reacts with the same general region of gpl20 as the first. In these studies, the human brain proteins were separated by SDS-PAGE and transferred to nitrocellulose as in Fig. 1. Strips were blocked with PBS buffer containing 5% BSA + 5% sheep serum. Primary antibodies were diluted 1:500; secondary antibodies (biotinylated, made in sheep to rouse IgG) were diluted 1:1000 and the binding visualized with biotin- streptavidin-HRP (1:500) reagent and visualized with DAB. Brain proteins from the other 2 human subjects (also presumed to be HIV-negative) were prepared from frontal cortex and diluted approximately 1 mg/ml in standard SDS buffer (Laem li, 1970). Figure 3 is a western blot of the binding of the anti-V3 antibody to human brain proteins which is displaced by recombinant gpl20. Proteins from the brain of a normal human subject were separated by SDS- PAGE and transferred to nitrocellulose as before. The monoclonal antibody was preincubated with decreasing concentrations of recombinant gpl20 protein, and then used to stain strips of the filter. Lane 7 is the control without competition from recombinant protein. Monoclonal antibody to the V3 region of gpl20 was diluted at 1:500 and mixed with decreasing concentrations of recombinant gpl20 (100 μg/ml, expressed in baculovirus) prior to being used on western blots of human brain proteins. At concentrations from 1:10 down to 1:40 (lanes l to 4) , recombinant gpl20 completely blocked the binding of the antibody to the human brain proteins; at 1:200 (lane 5) it was still somewhat effective, and at 1:1000 (lane 6) there was no noticeable competition, as evidenced by the fact that the major staining bands are as intense as in the control, lane 7. Gels, transfers, and antibody staining were carried out exactly as in Fig. 2. Recombinant gp 120 was prepared in a baculovirus expression system and purchased from American Bio-Technologies, Inc. (Cambridge, MA) .

Figure 4 is an immunoblot using antibodies to the V3 region of gpl20 cross-reacted with proteins from different parts of the nervous system and from some other organs. Lane l contains viral proteins, and the heavily stained band is believed to be gpl20 itself. Lane 2 is from spleen, lane 3 is from lung, lane 4 is from liver, lane 5 is from kidney, lane 6 is from spinal cord, lane 7 is from cerebellum, and lane 8 is from cerebral cortex. In all instances, tissue was solubilized at 10 mg wet weight tissue to 1 ml SDS buffer. Detailed Description of the Invention

The present method and compositions are based upon the discovery that there is an immunological cross-reactivity between components of the HIV-1 virus and proteins in various human tissues, which support the idea that some elements of AIDS result from an autoimmune reaction occurring in patients infected with the virus. The results described herein demonstrate that AIDS patients have antibodies directed against certain self-proteins, including brain proteins which are normally sequestered from the immune system. Moreover, antibodies directed against one critically important component of the virus, the V3 loop of the gpl20 protein, cross-react strongly and specifically with several proteins of the human brain. It is believed that such cross-reactivity occurs in HIV-1-infected patients, contributing to the disease process. From this, it follows that the disease can be treated, at least in part, by suppressing the production of antibodies to viral antigens that are not shared by human proteins.

An autoimmune disease is a malfunction of the immune system of patients, including humans. In a patient afflicted with such a disease, the immune system cannot or does not distinguish between certain exogenous (foreign) substances within the patient and autologous tissues or substances. As a result, the immune system treats these autologous tissues (self antigens) and substances as if they were foreign and evokes the proliferative immune defense that is usually reserved for use against exogenous (foreign) tissues or invading organisms. In general the immune defense consists of either or both the generation of antibodies or activation of cytotoxic T lymphocytes. As used herein, "antibodies" to self-proteins that are cross-reactive with viral components includes either immunoglobulins or molecules located on T lymphocytes such as the T cell receptor, or T cell receptor complex.

As used herein, an "autoantigen" is any substance normally found within a patient which, in an abnormal situation, is no longer recognized as part of the patient itself by the lymphocytes or antibodies of that patient, and is therefore attacked by the immune system as though it were a foreign substance, for example, the brain proteins, or portions of the brain proteins, described herein. The term "autoantigen" is also intended to encompass "analogs", as used herein, to encompass substances that differ from autoantigens or fragments by the deletion, addition or substitution of one or more amino acids, moieties or substituents, but are so structurally related to the autoantigen as to have the same type (but not necessarily the same degree) of suppressive activity when orally or enterally administered. Fragments and analogs can be synthesized using conventional chemical synthetic techniques, e.g., by using the well-known Merrifield peptide synthesis technique described, for example, in Merrifield, R.B. Fed. Proc. Am. Soc. Ex. Biol. 21:412, 1962 and J. Am. Chem. Soc. 85:2149, 1963; and Mitchel, A.R. J. Am. Chem. Soc. 98:7357, 1976. Analogs can be constructed, for example, by identifying an equivalent amino acid sequence (Tarn., J. , et al. J. Am. Chem. Soc, 105:6442, 1983); and using one of the general peptide synthesis methods referred to above or disclosed elsewhere herein. Alternatively, suppressive fragments or analogs can be prepared using recombinant DNA techniques according to methods well- known in the art.

Immunological unresponsiveness to sequestered tissue antigens such as brain antigens may be attributed partially to the location of these self- antigens behind natural anatomical barriers or to the lack of expression of class I or class II MHC molecules in the target organ (see, for example. Miller, et al., FASEB J. 5:2560 (1991)).

As used herein, the term "autoimmune suppressive fragments" includes any peptide or polypeptide containing partial amino acid sequences or moieties of autoantigens and possessing the ability to suppress or prevent an autoimmune response, in particular against the brain proteins described below, regardless of which tissue they are located on. The term includes autoantigens that are active against a specific autoimmune disease, as well as autoimmune-suppressive fragments or analogs thereof as defined above. As used herein the term "autoimmune-disease suppressive agent" or "autoimmune suppressive agent" further includes non-peptide compounds or compositions which can be administered to a patient to suppress, prevent or delay the clinical onset or manifestation of a specific autoimmune disease. In general, a peptide consists of less than one hundred amino acids; more preferably, it consists of as few as six to eight amino acids, more preferably eight to sixteen amino acids. The length of the peptide used to induce tolerance is determined by the number of amino acids required for cross-reactivity.

As employed herein, the term "treatment" refers to prophylactic administration of an agent to prevent an autoimmune disease in susceptible individuals or to administration of an agent for the treatment of an active autoimmune disease in an affected individual, in particular, a patient who is HIV positive, and/or who has AIDS.

Expression of the brain protein and preparation of peptides.

The identification and isolation of brain proteins which are cross-reactive with antibodies against the hypervariable region of HIV gpl20 (V3) is described herein. The isolation and cloning of a cDNA encoding one of these cross-reactive brain proteins is also described. As used herein, "brain protein" refers to any one of the brain proteins, or peptide portions or analogs thereof (consisting of at least six to eight consecutive amino acids) , reactive with monoclonal antibody directed against V3 or reactive with antibodies directed against other HIV antigens. The minimum length of peptide eliciting an immunogenic response is approximately six to eight amino acids. The brain proteins are believed to be autoantigens in AIDS patients. The proteins that are cross-reactive to antibodies against the V3 loop of gp 120 are found in organs other than brain, such as liver and kidney. Diagnostic Agents.

The antigenic peptides or proteins derived from the autoantigens have the potential to identify patients at risk for particular clinical manifestations or patients in particular prognostic groups. The peptides can be used in combination in assays, such as the solid phase assay, to classify patients. Specifically, the peptides that are bound by autoantibodies in patients characterized by specific disorders, such as central nervous system involvement or renal disease, are selected and combined in an assay, such as an ELISA for a test to detect the collection autoantibodies that bind this particular collection of peptides. The peptides, or antibodies to the peptides, can be labelled using radioactive labels, enzyme labels or fluorescent labels. Using a mixture of peptides may increase the efficiency and reliability of such assays, as compared with using a single autoantigen, or a single peptide.

The peptides can be used in solution or immobilized to a solid substrate, such as a gel suitable for affinity chromatography, or a multi-well plate, using standard techniques such as the commercially available cyanogen bromide. Patients having a predisposition for autoimmune manifestations resulting from AIDS or HIV infection can be identified using other anti-HIV antibodies, besides those directed against the V3 loop of gp 120, to screen for proteins other than the brain proteins described below that are cross-reactive with the anti- HIV antibodies. Antibodies against HIV from patients with AIDS or HIV infection can also be used in the same manner, i.e., after isolation of the antibodies (for example, using immobilized HIV components on an ion-exchange or affinity matrix) , one can perform western blots with tissue extracts, both from the patient and from normal controls, then probe with the patients' own antibodies.

One of these proteins has been partially sequenced, and antigenic determinants common to this normal human protein and the V3 loop of gpl20 may serve as one fragment for use in immunosuppression. Other polypeptides that are shared between proteins of the HIV-1 virus and human tissues are presented in Table 1. Whereas it is probably neither feasible nor desirable to utilize all polypeptides shared in common between the HIV-1 virus and human tissues for immunosuppression, optimal choices can be selected more precisely by methods similar to those proposed by PCT/US91/00682 "Assays and Treatment for Autoimmune Diseases" by Harley. Sera from HIV-infected patients can be used in sensitive immunoassays to identify proteins from normal donor human tissues to which there are circulating antibodies; the identity of the proteins with cross-reactive epitopes can be determined, e.g., by mixing sera of AIDS-infected patients with detergent extracts of tissues (in a ratio of 10 mg wet weight tissue: l ml 1% Triton x-100 in phosphate buffered saline following tissue homogenization and precipitation of insoluble material) , allowing antibody-antigen complexes to form, immunoadsorbing these onto Protein G-Sepharose™ beads (Sigma Chemical Co.), precipitating the beads, removing the supernatant, and separating the proteins by SDS-PAGE, transferring the proteins to PVDF membrane (Bio-Rad) , visualizing precipitated bands with Ponceau-S staining, and then having these microsequenced by a university or commercial facility. Sequences or precipitated proteins that resemble peptide sequences in HIV-1 proteins (variants of which are now entered in national data bases) can be identified using widely-available standard computer programs designed for these purposes (e.g., GenBank) .

Through methods similar to these, one of the proteins that is present in normal human tissues and which is cross-reactive with a commercial monoclonal antibody to the V3 region of the gpl20 surface glycoprotein of the HIV-1 virus, has been partially characterized.

The foregoing provides a means of identifying autoantigens in patients infected with the HIV-1 virus. Peptides shared in common between HIV components and patients' own proteins can be used to suppress autoimmune aspects of AIDS by presenting these peptides through means that will induce tolerance (e.g., orally, enterally, or as nasal sprays) . In addition, if it were discovered that particular epitopes of normal human proteins commonly become targets of autoantibodies following HIV-1 infection, this information could be valuable in the prevention of autoimmune aspects of AIDS. First, in designing vaccines against HIV-1 infection, epitopes in viral proteins that resemble epitopes in native proteins should be omitted or mutated through genetic engineering. Second, the polypeptide sequences that are determined as being shared in common between native proteins and HIV-1 proteins might be administered alone (e.g. , orally, enterally, or as nasal sprays) , or in combination with HIV-l proteins, genetically altered as suggested above, administered systemically to suppress autoimmune aspects of AIDS while at the same time fostering immunity to the distinctive components of the virus.

TABLE 1: Sequence similarities between normal human proteins and proteins of the HIV-l virus.

Reference: Altschul, Stephen F., Warren Gish, Webb Miller, Eugene W. Myers, and David J.

Lipman (1990). Basic local alignment search tool. J. Mol. Biol. 215, 403-410.

1 RIQRGPGRA 9 (HIV-l gpl20 protein, V3 loop) RIQRGP R+

60 RIQRGPVRG 68 (Human riboso al protein S17)

Identities = 7/9 (77%) , Positives = 8/9 (88%)

2 IQRGPGRAFV 11 (HIV-l gp 120 protein, V3 loop) I RGP RAF+

291 IVRGPCRAFI 300 (Human α-1-microglobulin bikunin, inter-α-trypsin inhibitor) I

Identities = 7/10 (70%), Positives = 8/10 (80%)

96 GLEGLIHSQRRQDILDLWIY 115 (HIV-l nef protein) +++GL++++RR + LW Y 181 SVDGLLNAHRRNAHVPLWNY 200 (Human gamma-glutamyl carboxylase, hGC)

Identities = 7/20 (35%) , Positives = 15/20 (75%)

65 WTTYWGLHTGEKEWH 80 (HIV-l vif protein) V+ W+LH EKEWH 392 VIINLWALHHNEKEWH 407 (Human cytochrome P450cl7; steroid 17-alpha-hydroxylase)

Identities = 9/16 (56%), Positives = 11/16 (68%)

161 PPLPSVRKLTEDRWNK 176 (HIV-l vif protein) P L SVR++ +++W+K 32 PQLQSVRRIRDQKWHK 47 (Human beta B3 crystallin beta B3 crystallin)

Identities = 7/16 (43%), Positives = 13/16 (81%)

277 RMYNPTNILDIKQGPKESFQSYVDRFYKSLRAEQTDPAVKNWMTQTLLVQNAN 329 (HIV gag protein)

+M ++ +1+ + + +YV FY + + Q + N+-I- L V N

220 KMLDAEDIVGTARPDEKAIMTYVSSFYHAFSGAQKAETAANRICKVLAVNQEN 272 (Non-muscle α-actinin) I

Identities = 11/53 (20%), Positives = 24/53 (45%)

285 LDIKQGPKESFQSYVDRFYKSLRAE 309 (HIV gag protein)

L KQ P + FQ+Y++ +LR++ 136 LGKKQVPPDLFQPYIEEICQNLRGD 160 (Human beta-adrenergic receptor kinase)

Identities = 9/25 (36%) , Positives = 16/25 (64%)

285 LDIKQGPKESFQSYVDRFYKSLRAE 309 (HIV gag protein)

L KQ P + FQ+Y++ +LR++

136 LGKKQVPPDLFQPYIEEICQNLRGD 1 16600 ((HHuummaen β-adrenergic receptor kinase 1)

Identities = 9/25 (36%), Positives = 16/25 (64%)

235 SDIAGTTSTVEEQIQWMF 252 (HIV gag protein)

S 1+ ++S+V+++ QW+F 277 SSITTSSSAVQQEEQWLF 294 (Human transmembrane protein tyrosine kinase, ROSl)

Identities = 7/18 (38%), Positives = 15/18 (83%)

9 RGKKADELEKVRLRPGGKKKYKLKHIVWAANEL 41 (HIV gag protein) RG + D L+ V + + K +YK H +W EL 243 RGTRLDGLDLVDTWKSFKPRYKHSHFIWNRTEL 275 (Human liver/bone/kidney-type alkaline l phosphatase)

Identities = 12/33 (36%) , Positives 18/33 (54%)

9 IAALWAIILAIWWTIVFIEYRRIKKQR 37 (HIV-l vpu protein) I+++ V+++L +V+ + F RR K+QR 538 IGGVAVGWLLLVLAGVGFFIHRRRKNQR 566 (Human protein tyrosine kinase)

Identities = 10/29 (34%), Positives 20/29 (68%)

PLGIIAIAALWAIILAIWWTIVFIEYRRIK 34 (HIV-l Vpu protein)

40 ALVIFAWFLVGVLGNALWWVTAFEAKRTIN 71 (Human C5a anaphylatoxin receptor)

Identities = 12/32 (37%), Positives = 17/32 (53%)

30 YRRIKKQRRIDCLLDRITERAEDSGNESEGDREKLSKLVE 69 (HIV-l vpu protein) YR++ Q LL+ I R + +N+ G R +L++ V+

41 YRKVLGQLSARKLLQDIMSRQQGESNQERGARARLGRQVD 80 (Human growth hormone-releasing factor)

Identities = 12/40 (30%) , Positives = 22/40 (55%)

5 GIIAIAALWAIILA 19 (HIV-l vpu protein)

GIIAI +LWA+++A

63 GIIAIYGLWAVLIA 77 (Human vacuolar H+ ATPase proton channel subunit) '

Identities = 10/15 (66%) , Positives = 14/15 (93%)

35 RQARRNRRRRWRERQR 50 (HIV-l env protein)

RQ RR RRRR R +R 40 RQRRRRRRRRMRRMRR 55 (Human arginine-rich gene)

Identities = 10/16 (62%) , Positives 11/16 (68%)

27 PPPNPEGTRQARRNRRRRWRERQRQ 51 (HIV-l env protein) PPP TR+ R +R+RR + +RQ

55 PPPTRAETREERMERKRREKIERRQ 79 (Human small nuclear ribonucleoprotem, U1-70K)

Identities = 11/25 (44%), Positives 16/25 (64%)

35 RQARRNRRRRWRERQRQIHSISER 58 (HIV-l env protein) R+ R+R R +RER R+ ER 222 RDRDRDRERERRERSRERDKERER 245 (Human small nuclear ribonucleoprotein, U1-70K)

Identities 10/24 (41%), Positives = 14/24 (58%)

23 YQSNPPPNPEGTRQARRNRRRRWRERQRQIHSISER 58 (HIV-l env protein)

Y+ P P+P R+ R+R R +RER R+ ER N

396 YDERPGPSPLPHRDRDRDRERERRERSRERKDERER 431 (Human UI RNA-associated 7OK protein) '

Identities = 14/36 (39%), Positives = 20/36 (55%)

23 YQSNPPPNPEGTRQARRNRRRRWRERQRQIHSISER 58 (HIV-l env protein) Y+ P P+P R+ R+R R +RER R+ ER 219 YDERPGPSPLPHRDRDRDRERERRERSRERKDERER 254 (Human small nuclear ribonucleoprotein, Ul-

70K)

Identities = 14/36 (39%), Positives = 20/36 (55%)

27 PPPNPEGTRQARRNRRRRWRERQRQ 51 (HIV-l env protein)

PPP TR+ R +R+RR + +RQ

55 PPPTRAETREERMERKRREKIERRQ 79 (Human small nuclear ribonucleoprotein, U1-70K) Identities = 11/25 (44%) , Positives = 16/25 (64%)

35 RQARRNRRRRWRERQ 49 (HIV-l env protein) RQ +R+RRRR+R Q

17 RQRQRSRRRRRRSCQ 31 (Human testis specific protamine 1, PI)

Identities = 9/15 (60%), Positives = 12/15 (80%)

38 RRNRRRRWRERQRQIHSISER 58 (HIV-l env protein)

+R R RR+R R Q + + R

18 QRQRSRRRRRRSCQTRRRAMR 38 (Human testis specific protamine 1, PI)

Identities = 8/21 (38%), Positives = 12/21 (57%)

35 RQARRNRRRRWRERQ 49 (HIV-l env protein)

RQ +R+RRRR+R Q 17 RQRQRSRRRRRRSCQ 31 (Human protamine 1)

Identities = 9/15 (60%), Positives = 12/15 (80%)

38 RRNRRRRWRERQRQIHSISER 58 (HIV-l env protein)

+R R RR+R R Q + + R

18 QRQRSRRRRRRSCQTRRRAMR 38 (Human protamine 1)

Identities = 8/21 (38%), Positives = 12/21 (57%)

32 EGTRQARRNRRRRWRERQRQIHSISERILST 62 (HIV-l env protein) E++R++RR R+ + E QRQ + I +L+T

11 EAARRSRRIDRHLRSESQRQRREIKLLLLGT 41 (Human transducin Gx-alpha-subunit, GNAZ)

Identities = 12/31 (39%), Positives = 21/31 (67%)

12 LLKAVRLIKFLYQSNPPPN 30 (HIV env protein) +LK++ LI FL Q+N++PN t

87 VLKTIVLIPFLEQNNSSPN 105 (Human NKG2-C gene Type II integral membrane protein) '

Identities = 10/19 (52%), Positives = 16/19 (94%)

78 RKGRRRRTPKKTKAHSSSASDKSISTRTGNSQPEKKQKK 116 (HIV-l tat protein) R RRRR+ T++ + S K +R+ +++ KK+KK 341 RERRRRRSRSGTRSPKKPRSPKRKLSRSPSPRRHKKEKK 379 (Human arginine rich nuclear protein)

Identities = 13/39 (33%) , Positives = 23/39 (58%)

APEGSLGSYNEPSSCTSEQDAAAQGLVSPGDEI 39 (HIV-l tat protein)

+P G++G+ EP++ + +++AQGL +P I 706 GPKGNVGPQGEPGPPGQQGNPGAQGLPGPQGAI 738 (Human collagen alpha 1(V) chain)

Identities = 11/33 (33%), Positives = 22/33 (66%)

65 MCFLNKGLGIWYERKGRRRRTPKKTKAHSSSASDKS 100 (HIV-l tat protein) +C ++ + IW++ + +++ +KT + ++++ D++ 174 LCLTERQIKIWFQNRRMKWKKENKTAGPGTTGQDRA 209 (Human homeo box cl protein)

Identities = 6/36 (16%) Positives = 24/36 (66%)

65 MCFLNKGLGIWYERKGRRRRTPKKTKAHSSSASDKS 100 (HIV-l tat protein)

+C ++ + IW++ + +++ +KT + ++++ D++ 56 LCLTERQIKIWFQNRRMKWKKENKTAGPGTTGQDRA 91 (Human homeobox protein, HOX2.3)

Identities = 6/36 (16%) , Positives = 24/36 (66%)

7 APEGSLGSYNEPSSCTSEQDAAAQGLVSPGDEI 39 (HIV-l tat protein) +P G++G+ EP++ + +++AQGL +P I 676 GPKGNMGPQGEPGPPGQQGNPGPQGLPGPQGPI 708 (Human alpha-1 type XI collagen, COLllAl)

Identities = 10/33 (30%), Positives = 22/33 (66%)

77 ERKGRRRRTPKKTKAHSSSASDKSISTRTGNSQPEKKQKKT 117 (HIV-l tat protein)

E+K+R+R + +++A+ S+ KS ++T + ++++++

141 EHKSRKRTSESRSRARKRSSKSKSHRSQTRSRSRSRRRRRS 181 (Human son3 protein gene, partial)

Identities = 9/41 (21%) , Positives = 27/41 (65%)Pharmaceutical Compositions for inducing tolerance.

t

At present, therapy for autoimmune diseases is non-specific in that it cannot be directed at the underlying cause. Immunosuppressants which are currently in use include cyclosporin, glucocorticoids, methotrexate, azathioprine, cyclophospha ide, non- steroidal antiinflammatory agents, anti alarials, and other non-specific therapeutics such as sun screens. Usage and dosage of these drugs is dictated by the disease manifestations. Glucocorticoids, for example, are used in high dosages to treat some neurologic complications of systemic lupus erythematosus (SLE) . Both azathioprine and cyclophosphamide are used as an attempt to halt or reverse renal damage. Limiting side effects are common for all of the immunosuppressants. This generally relates to their non-specific effect on not only the portion of the immune system involved in SLE but also on the normally functioning, protective immune system. For example, the dose of cyclophosphamide is generally regulated, and limited, by toxicity to hematopoiesis.

The peptides common to the HIV virus and human tissues can be used therapeutically in combination with a pharmaceutically acceptable carrier to diminish the effects of the response to the autoantigens, alone or in combination with general immunosuppressants, i.e., either to induce tolerance or to competively bind the autoantibodies. The peptides can be administered in a dosage effective to block autoantibodies, or as a vaccine to block the autoantibodies. The peptide acts as a functional antagonist by binding to antibody that does not stimulate or activate the immune cells and thereby block the immune response to the autoantigens.

Peptides used as vaccines can be administered orally, intramuscularly or subcutaneously. Other compounds will be administered according to standard procedures used by those skilled in the art. As defined herein, a pharmaceutical carrier is usually inert by itself but may have biological activity. For example, a vaccine may consist of immunogenic peptides or proteins in combination with an adjuvant (an agent which enhances but does not elicit an antigen-specific immune response) .

Therefore, patients who are seropositive for AIDS could be given orally recombinant portions of gpl20 and p41 that resemble portions of brain proteins or MHC components, synthesized by conventional genetic engineering: cDNAs are available from commercial sources (e.g. , Boehringer-Mannheim) ; PCR can also be used to generate cDNAs using primers based upon the published HIV-l genome. At the same time, immunity to HIV-l can be boosted systemically by presenting recombinant molecules of the AIDS viral coat protein that do not resemble normal human tissues, either as a separate method for vaccination or in combination with induction of tolerance against the cross-reactive proteins or peptides.

A variation on this method would use other means of immunosuppression. For example, portions of gpl20 and p41 that resemble portions of brain proteins, or MHC constituents, could be injected, and when the immune reaction is evident, the production of cells making anti-gpl20 and anti-p41 immunoglobulins suppressed with cyclophosphamide; when the titer of such antibodies has been lowered, the patient is boosted with portions of the HIV-l coat proteins that do not mimic MHC components.

Alternatively, the peptides used for treatment can include peptides homologous to an identified antigenic sequence. These peptides, either free or bound to a carrier, can be delivered to a patient in order to decrease the amount of circulating antibody with a particular specificity. In addition, knowledge of the cross-reacting epitopes between a foreign antigen and an autoantigen allow for re-induction of tolerance. It is well known in experimental models of the immune response that the response can be suppressed and tolerance induced by administering the antigen through particular routes, e.g., oral.

The amino acid sequences can also be used to make agents for neutralizing circulating antibodies or immobilized on substrates in extracorporeal devices for specific removal of autoantibodies, using methodology known to those skilled in the art.

Oral administration or feeding of autoantigenic proteins, e.g., the autoantigen and fragments or analogs thereof induces antigen-specific suppression, presumably through CD8+ T-cells (suppressor cells) that are elicited through such feeding. These cells mediate the suppression (down-regulation) of the immune response. This down-regulation permits suppression of clinical symptoms (such as inflammation) associated with an autoimmune disease.

One method for suppressing the clinical manifestation of an autoimmune disease in a patient in need of such treatment, is to orally or enterally (i.e., by tube feeding) administering to the patient an effective amount of a suppressive agent selected from the group consisting of (i) an autoantigen; (ii) a fragment of the autoantigen which when orally administered to a patient suppresses the symptoms of the autoantigen or a fragment thereof; (iii) an analog of the autoantigen or fragment thereof with immunosuppressive activity, and (iv) combinations of any or all of the foregoing.

The term "suppression" is intended to include prevention of the clinical manifestations or symptoms, as well as complete elimination or at least measurable attenuation of such manifestations and symptoms. To be an "effective amount" the amount of the suppressive agent should be sufficient to cause a measurable and preferably a statistically significant attenuation of at least one clinical symptom associated with the autoimmune disorder. Particular attention would be paid to the CD4 + lymphocyte count.

In general terms, an effective amount of a peptide for suppressing or attenuating clinical symptoms is from about 0.1 to about 15 mg/kg/day, preferably from about 3 to about 10 mg/kg/day (for a 70 kg adult, the effective amount is from about 30 to about 1000 mg/day, preferably from about 300 to about 600 mg/day) . In rodents, the equivalent effective amount of the suppressant agent is from about 100 μg to about 10 mg per rodent. Of course, the broad ranges given above may need to be adjusted according to factors such as the age, sex and physical condition of the subject to be treated, the severity of the disease state, and the specific suppressive activity of the agent or agents to be administered. However, such refinement of suitable dosages can be determined by persons of ordinary skill in the field using no more than routine experimentation. For example, optimum dosage can be established using serially diluted preparations of the active agents of the present invention in connection with a suitable testing procedure. Alternatively, a matrix of dosages and frequency of administration can be established and groups of experimental subjects can be assigned to each point on the matrix in order to determine the optimum conditions.

Such effective amounts can be dispensed to a patient in need of treatment in one daily oral dose or in two or more divided daily oral doses. In man, such divided daily oral doses may be administered, preferably, three times/day over a period of from about 90 to about 120 days, or even longer to insure sufficient uptake and absorption of the autoantigen. fragment or analog. Treatment may continue or be resumed if symptoms persist or recur.

The immunosuppressant or active agent is administered in an effective dose to a patient in a suitable pharmaceutical carrier. Pharmaceutical carriers are known to those skilled in the art. For example, compounds for oral administration can be encapsulated in an enteric coating or provided in combination with a binder such as stearate or lactose, or in solution. Acceptable solutions include sterile water, saline, and buffered solutions at physiological pH.

Administration of autoantigens (or autoimmune suppressive fragments or analogs thereof) in aerosol form is also effective in treating autoimmune disease in patients. Administration of autoantigens in aerosol form is more effective in preventing and treating autoimmune diseases in patients than administration of the same autoantigens in solid form via the oral route and uses a smaller quantity of such autoantigens in an aerosol form than when administered in a solid dosage form. The aerosol administration of autoantigens has been found to be effective in suppressing both cell-mediated and antibody-mediated autoimmune responses.

As used herein, the term "aerosol" refers to finely divided solid or liquid particles that may be created using a pressurized system such as a nebulizer. The liquid or solid source material contains autoantigens and/or autoimmune disease suppressive fragments and analogs thereof as defined herein.

Induction of tolerance is dose-dependent; over a broad dosage range of aerosol material it has been reported that suppression (or attenuation) of clinical manifestations of the disease increases with increasing dosage levels of the aerosolized autoimmune-suppressive agent. A reported advantage of aerosol administration is that less peptide is required for induction of immunity. Another advantage is that it decreases the time of exposure of the autoimmugenic agents to degradative gastric juices, which may act to reduce the efficacy of such agents.

The present invention will be further understood with reference to the following non-limiting examples.

Example 1: Determination that HIV-l infected patients have antibodies that cross-react with human brain proteins.

Proteins from normal human brain were separated on SDS-polyacrylamide gels, transferred to nitrocellulose membranes, and probed with sera and CSF from patients who were either HIV-1-positive but who did not evidence signs of dementia (HIV+/ADC-) , patients with AIDS who did show the AIDS-dementia complex (ADC) , and normal controls.

The results are shown in Figure 1. None of the 14 control patients appeared to have antibodies that reacted with any brain proteins, as evidenced by the absence of distinct staining bands other than the two background bands seen in all cases; among the 10 patients who carried the virus but did not show signs of ADC, numbers 3 and 4 had antibodies against proteins of about 65 kDa, and number 8 had antibodies against a 120 kDa protein. Among the ADC patients, numbers 7, 8, 10 and 12 had antibodies against brain proteins between 90 and 120 kDa in molecular weight.

No statistically significant differences were found between groups. The data do, however, provide support for an autoimmune response in AIDS.

Example 2: Determination of whether or not the antibodies in HIV-infected patients could be reacting with either viral proteins in the brain or with brain proteins that resemble components of the virus.

Whether or not the antibodies in HIV-infected patients could be reacting with either viral proteins in the brain or with brain proteins that resemble components of the virus was then determined, since this could come about if the immune response against the virus leads to the development of antibodies that adventitiously cross-react with cognate regions of normal brain proteins. A panel of monoclonal antibodies, raised in mice against defined viral proteins, were used to probe the western blot containing the brain proteins from a normal subject (an automobile accident victim presumed not to have had AIDS), as described with reference to Figure 2.

The antibodies were raised to the so-called V3 loop of gpl20 (lane 1) , undefined sequences in gpl20 (lanes 2 and 3) , gp41 (lane 4) , p24 (lane 5) , pl7 (lane 6) , Vpu (lane 7) , and Vpr (lane 8) . Vpu and Vpr are viral regulatory proteins. The results shown in Figure 2A indicate that only the anti-V3 antibody (from DuPont/New England Nuclear, raised against a synthetic peptide representing amino acids 308-322 of the viral protein) cross-reacts with brain proteins. The reactive brain proteins have estimated molecular weights of 105, 52, 27 and 17 kDa. Figures 2B and C show that these same proteins are found in two other normal human brains and that they are recognized by another monoclonal antibody raised against gpl20, labeled as mAb2, also from DuPont/NEN, which also reacts with a peptide in the V3 region.

To determine the specificity of this binding, it was determined whether the binding could be competed away using isolated viral gpl20 protein or recombinant gpl20. In Figure 3, the monoclonal antibody to the V3 region of gpl20 was diluted 1:500 and mixed with decreasing concentrations of recombinant gpl20 (100 μg/ml, expressed in baculovirus) prior to being used on western blots of human brain proteins. At concentrations from 1:10 down to 1:40 (lanes l to 4) , recombinant gpl20 completely blocked the binding of the antibody to the human brain proteins; at 1:200

(lane 5) it was still somewhat effective, and at

1:1000 (lane 6) there was no noticeable competition

(lane 7) . Thus, the binding of the antibody is specific to a domain in the endogenous brain proteins that appears to resemble the V3 region of the gpl20 protein of the HIV-l virus.

Example 3: Cloning of a cDNA encoding a brain protein immunoreactive with antibody directed against the V3 region of HIV.

To identify one of the native proteins that immunologically resembles a portion of gpl20, a lambda ZAP human brain cDNA expression library (Stratagene) was screened with the same antibody used on the western blots (Dupont/NEW 9205) . This antibody reacted with proteins being expressed by several lambda clones; one clone expressed a protein that reacted particularly well, so the plaques from this region of the plate were collected and replated for secondary screening at low density. The single plaque was purified, the DNA was excised from the lambda phage as a plasmid, plasmid DNA was transformed into E. coll , from which DNA was prepared and sequenced using the SK and the lambda ZAP reverse primers described by Stratagene. Additional primers were made from these initial sequences.

The cloned DNA has an .EcoRI site at one end and in the middle, which allowed sequencing of an internal portion of the clone. To date, the nucleotide sequence does not match any other in GenBank, nor does the amino acid sequence match anything yet reported.

The nucleotide sequence (Sequence ID No. 1) is as follows:

GGGGCTGACTTTGCTGCTGTGGGGGTGGGGACTGGGGTGCAGGCCCCACTNNTG GGTGTCAATANTGNGGCTACCATTNCTCCTGTGNGGGTGGGGACTGGGGTGCAG CCCCCACTACNNGNTGTCAATACTGGGGCTGCCTTTACTNCTGTGGGGGTGGGG ACTGGGGTGCAGNCCCCACTNCTGGGTGTCAATNCTGGGGCTNCCNTTNCTNCT GTGGGGGTGGGGACTGGGGTGCAGGGAGGGGACGGGTGAATAAACAGACACAGG

TGGNCAGAGGCAGACAANAGCCCATGAATTCAGCGTGGGCCCAGCTCCCATGGT

TGNCTCGCGGCCCAGGGGATNGTGCTTGGTGGNCAAGGAGTGTGAGGCCATCAG

AGCTCCACAANGCGGTGTGGNGTGCTGCACNNCCCAACCCTNCCTCCCAAAACG

TCANCCAGNTNTTCACCGCATGAATACGTGAGTGGTCAGCCAGGTGTTCAACGC

ATGAATAAGTGAGTGAATGAATGAATGAATGAGTAATTTGGTGGCTGACCCAAT

GAGTGAATGAGGGAACAGGNGANTCCGTGAGCANCTCTGAGTGAAGAAGAGTGA

NCCTCGAAGGAGAGTGATGTCAGGAGATGGTANTTTNTGNCAATTCGNCCACTG

NGAGCCCACGTGGTATGTCTCTCATCGGAGCCACAGGGNAGNTATAGGNCGTNC

TCAGGCACACGGGGGTCTCGGGNTGGCNAACAANANNAAAAAATGAATNGANGG

NATGNATGGAAANNAANAAANTTGGGTGGNTTGGANGNAATGAANTGAATTGAA

GGGAAAAAGNCGGAAACACGTTGANNCAACNNCTTGAGTGGAAGAAAGANTTGA

GCCNTCGGAANGAAGGAGTGATGTNAAGGAAGAAGGGCNGGGTTGNTTGACCAA

ANNGGTCCCACTTGGGAAGACAANGTGGGGATGTNTCTCCATCGTANNACAANA

GGGGGAGNAAATAGACCGTGNTCAGGAAANANGGGGGNCTCGACNCCCCCGCGG

GTGTCGACNCCCCTNCCTCCCTCCAGACCTGTNCACACTGCCCCTTTGNAGGGA

GCCCNTCCTGGTCCTCAGTGTTTATANNTGCTGTCCCCNTCTACTGGGTGCTCC

CCTGGCCGTGACATCACCACCAGCCAGANTGGCTTTTGTTCTCCTAAACATCTC

TCCTCCCCTCACCCACCATGAGCTCCACGGGCTGAGGTCTCGGGCCACCCCTCC

TCCCTCCCACCTCTTTCTCTCTTTCTCCAGGTGCCTTGTAGGTGCTTGGTGAGT

TTCCTCATTCNACAAGTCCAGGCATTCATTCATGTGTGTGTTACACACNTAANN

NCAGAGNAGNAN

Example 4: Tissue specificity of the proteins cross- reactive with the anti-V3 antibodies.

The tissue specificity of the proteins that cross-react with the anti-gpl20 V3 antibodies was then determined. Lane 1 in Figure 4 contains the proteins of the HIV-l virus. As expected, the antibody to the

V3 loop of gpl20 recognizes a band at 120,000 Da, the viral gpl20 protein. No cross-reactivity is seen with proteins present in either the spleen or lung (lanes 2 and 3, respectively) , but several cross-reactive proteins are seen in the liver (lane 4) and kidney

(lane 5) . In the nervous system, the bands previously found in the frontal cortex were also present in the spinal cord, cerebellum, and another part of the cerebral cortex (lanes 6, 7 and 8, respectively). Thus, proteins that cross-react with antibodies to a defined region of the gpl20 protein may exist in all parts of the nervous system and in some, but not all, other organs, at least at levels detectable by this technique. The absence of immuno-staining in the spleen would argue against the cross-reactive proteins being MHC components.

Claims

We claim.

1. A method for treating AIDS-related disorders comprising: determining in a patient sample the presence of human proteins immunoreactive with anti-HIV antibodies; isolating peptides from the immunoreactive proteins which bind to the anti-HIV antibodies; administering an effective amount of the immunoreactive peptides to the patient to induce tolerance to the protein.

2. The method of claim 1 wherein the peptide is isolated from a protein present in the brain.

3. The method of claim 2 wherein the patient has AIDS-related dementia.

4. The method of claim 2 wherein the protein is selected from the group of proteins expressed in a brain tissue cDNA library having a molecular weight of 17,000, 27,000, 52,000, 65,000, 90,000, 105,000, and 120,000 D immunoreactive with an antibody to gpl20.

5. The method of claim 4 wherein the protein is encoded by the nucleotide sequence (Sequence I.D. No.

1):

GGGGCTGACTTTGCTGCTGTGGGGGTGGGGACTGGGGTGCAGGCCCCACTNNTG

GGTGTCAATANTGNGGCTACCATTNCTCCTGTGNGGGTGGGGACTGGGGTGCAG

CCCCCACTACNNGNTGTCAATACTGGGGCTGCCTTTACTNCTGTGGGGGTGGGG

ACTGGGGTGCAGNCCCCACTNCTGGGTGTCAATNCTGGGGCTNCCNTTNCTNCT

GTGGGGGTGGGGACTGGGGTGCAGGGAGGGGACGGGTGAATAAACAGACACAGG

TGGNCAGAGGCAGACAANAGCCCATGAATTCAGCGTGGGCCCAGCTCCCATGGT

TGNCTCGCGGCCCAGGGGATNGTGCTTGGTGGNCAAGGAGTGTGAGGCCATCAG

AGCTCCACAANGCGGTGTGGNGTGCTGCACNNCCCAACCCTNCCTCCCAAAACG

TCANCCAGNTNTTCACCGCATGAATACGTGAGTGGTCAGCCAGGTGTTCAACGC

ATGAATAAGTGAGTGAATGAATGAATGAATGAGTAATTTGGTGGCTGACCCAAT

GAGTGAATGAGGGAACAGGNGANTCCGTGAGCANCTCTGAGTGAAGAAGAGTGA

NCCTCGAAGGAGAGTGATGTCAGGAGATGGTANTTTNTGNCAATTCGNCCACTG

NGAGCCCACGTGGTATGTCTCTCATCGGAGCCACAGGGNAGNTATAGGNCGTNC

TCAGGCACACGGGGGTCTCGGGNTGGCNAACAANANNAAAAAATGAATNGANGG NATGNATGGAAANNAANAAANTTGGGTGGNTTGGANGNAATGAANTGAATTGAA GGGAAAAAGNCGGAAACACGTTGANNCAACNNCTTGAGTGGAAGAAAGANTTGA GCCNTCGGAANGAAGGAGTGATGTNAAGGAAGAAGGGCNGGGTTGNTTGACCAA ANNGGTCCCACTTGGGAAGACAANGTGGGGATGTNTCTCCATCGTANNACAANA GGGGGAGNAAATAGACCGTGNTCAGGAAANANGGGGGNCTCGACNCCCCCGCGG GTGTCGACNCCCCTNCCTCCCTCCAGACCTGTNCACACTGCCCCTTTGNAGGGA GCCCNTCCTGGTCCTCAGTGTTTATANNTGCTGTCCCCNTCTACTGGGTGCTCC CCTGGCCGTGACATCACCACCAGCCAGANTGGCTTTTGTTCTCCTAAACATCTC TCCTCCCCTCACCCACCATGAGCTCCACGGGCTGAGGTCTCGGGCCACCCCTCC TCCCTCCCACCTCTTTCTCTCTTTCTCCAGGTGCCTTGTAGGTGCTTGGTGAGT TTCCTCATTCNACAAGTCCAGGCATTCATTCATGTGTGTGTTACACACNTAANN NCAGAGNAGNAN.

6. The method of claim 2 wherein the peptide is administered orally.

7. The method of claim 2 wherein the peptide is administered as an aerosol.

8. The method of claim 1 further comprising boosting immunity to HIV-l by presenting systemically recombinant molecules of the AIDS viral coat that do not resemble portions of gpl20 or p41.

9. A method for vaccinating a patient against AIDS comprising administering to a human patient an effective amount of peptides derived from HIV which are not cross-reactive with antibodies against human proteins to elicit an immune reaction.

10. The method of claim 9 wherein the peptides are derived from HIV gp 120 coat protein.

11. A method for identifying humans susceptible to AIDS-related disorders comprising screening proteins and tissues from a human with antibodies, including T cell receptors, immunoreactive with human immunodeficiency virus proteins.

12. The method of claim 11 wherein the tissue is brain tissue or spinal cord tissue.

13. The method of claim 11 wherein the antibodies are directed against HIV gpl20.

14. The method of claim 13 wherein the antibodies are directed against the V3 region of gpl20.

15. The method of claim 12 wherein the antibodies are directed against HIV gpl20 and are reactive with proteins selected from the group of proteins having molecular weights of 17,000, 27,000, 52,000, 65,000, 90,000, 105,000 and 120,000 D.

16. The method of claim 15 wherein the antibodies are reactive with a protein encoded by the nucleotide sequence (Seq. I.D. No. 1):

GGGGCTGACTTTGCTGCTGTGGGGGTGGGGACTGGGGTGCAGGCCCCACTNNTG GGTGTCAATANTGNGGCTACCATTNCTCCTGTGNGGGTGGGGACTGGGGTGCAG CCCCCACTACNNGNTGTCAATACTGGGGCTGCCTTTACTNCTGTGGGGGTGGGG ACTGGGGTGCAGNCCCCACTNCTGGGTGTCAATNCTGGGGCTNCCNTTNCTNCT GTGGGGGTGGGGACTGGGGTGCAGGGAGGGGACGGGTGAATAAACAGACACAGG TGGNCAGAGGCAGACAANAGCCCATGAATTCAGCGTGGGCCCAGCTCCCATGGT TGNCTCGCGGCCCAGGGGATNGTGCTTGGTGGNCAAGGAGTGTGAGGCCATCAG AGCTCCACAANGCGGTGTGGNGTGCTGCACNNCCCAACCCTNCCTCCCAAAACG TCANCCAGNTNTTCACCGCATGAATACGTGAGTGGTCAGCCAGGTGTTCAACGC ATGAATAAGTGAGTGAATGAATGAATGAATGAGTAATTTGGTGGCTGACCCAAT GAGTGAATGAGGGAACAGGNGANTCCGTGAGCANCTCTGAGTGAAGAAGAGTGA NCCTCGAAGGAGAGTGATGTCAGGAGATGGTANTTTNTGNCAATTCGNCCACTG NGAGCCCACGTGGTATGTCTCTCATCGGAGCCACAGGGNAGNTATAGGNCGTNC TCAGGCACACGGGGGTCTCGGGNTGGCNAACAANANNAAAAAATGAATNGANGG NATGNATGGAAANNAANAAANTTGGGTGGNTTGGAMGNAATGAANTGAATTGAA GGGAAAAAGNCGGAAACACGTTGANNCAACNNCTTGAGTGGAAGAAAGANTTGA GCCNTCGGAANGAAGGAGTGATGTNAAGGAAGAAGGGCNGGGTTGNTTGACCAA ANNGGTCCCACTTGGGAAGACAANGTGGGGATGTNTCTCCATCGTANNACAANA GGGGGAGNAAATAGACCGTGNTCAGGAAANANGGGGGNCTCGACNCCCCCGCGG GTGTCGACNCCCCTNCCTCCCTCCAGACCTGTNCACACTGCCCCTTTGNAGGGA GCCCNTCCTGGTCCTCAGTGTTTATANNTGCTGTCCCCNTCTACTGGGTGCTCC CCTGGCCGTGACATCACCACCAGCCAGANTGGCTTTTGTTCTCCTAAACATCTC TCCTCCCCTCACCCACCATGAGCTCCACGGGCTGAGGTCTCGGGCCACCCCTCC TCCCTCCCACCTCTTTCTCTCTTTCTCCAGGTGCCTTGTAGGTGCTTGGTGAGT TTCCTCATTCNACAAGTCCAGGCATTCATTCATGTGTGTGTTACACACNTAANN NCAGAGNAGNAN.

17. The method of claim 11 wherein the antibodies are reactive with tissue extracted from kidney and liver.

18. The method of claim 11 wherein the antibodies are T cell receptors.

19. The method of claim 11 wherein the antibodies are labeled with a label selected from the group consisting of fluorescent labels, radioactive labels, and enzyme labels.

20. The method of claim 11 wherein the antibodies are reactive with non-MHC proteins.

21. The method of claim 11 wherein the antibodies are reactive with a protein selected from the group consisting of human ribosomal protein S17, human α-1-microglobulin bikunin, human gamma-glutamyl carboxylase, human cytochrome P450cl7, human beta B3 crystallin, non-muscle α-actinin, human beta- adrenergic receptor kinase, human β-adrenergic receptor kinase 1, human transmembrane protein tyrosine kinase, human liver/bone/kidney-type alkaline phosphatase, human protein tyrosine kinase, human C5a anaphylatoxin receptor, human growth hormone-releasing factor, human vacuolar H+ ATPase proton channel subunit, human arginine-rich gene, human small nuclear ribonucleoprotein, human UI RNA-associated 7OK protein, human testis specific protamine, human transducin Gx-alpha-subunit, human NKG2-C gene Type II integral membrane protein, human arginine-rich nuclear protein, human collagen alpha 1(V) chain, human homeo box cl protein, human homeobox protein H0X2.3, human alpha-1 type XI collagen, and human son3 protein gene.

22. The method of claim 11 further comprising identifying peptide fragments from the human proteins which are immunoreactive with the anti-HIV antibodies.

23. The method of claim 22 further comprising administering an effective amount of peptide immunoreactive with the anti-HIV antibodies to a human to decrease the reaction of antibodies in the patient to the proteins immunoreactive to the anti-HIV antibodies.

24. A brain protein isolated by immunoreaction with an antibody immunoreactive with a component of human immunodeficiency virus.

25. The brain protein of claim 24 selected from the group of proteins expressed in a brain tissue cDNA library having a molecular weight of 17,000, 27,000, 52,000, 65,000, 90,000, 105,000 and 120,000 Daltons and immunoreactive with an antibody to the V3 region of HIV gpl20.

26. The brain protein of claim 25 encoded by the nucleotide sequence (Sequence I.D. No. 1) : GGGGCTGACTTTGCTGCTGTGGGGGTGGGGACTGGGGTGCAGGCCCCACTNNTG GGTGTCAATANTGNGGCTACCATTNCTCCTGTGNGGGTGGGGACTGGGGTGCAG CCCCCACTACNNGNTGTCAATACTGGGGCTGCCTTTACTNCTGTGGGGGTGGGG ACTGGGGTGCAGNCCCCACTNCTGGGTGTCAATNCTGGGGCTNCCNTTNCTNCT GTGGGGGTGGGGACTGGGGTGCAGGGAGGGGACGGGTGAATAAACAGACACAGG TGGNCAGAGGCAGACAANAGCCCATGAATTCAGCGTGGGCCCAGCTCCCATGGT TGNCTCGCGGCCCAGGGGATNGTGCTTGGTGGNCAAGGAGTGTGAGGCCATCAG AGCTCCACAANGCGGTGTGGNGTGCTGCACNNCCCAACCCTNCCTCCCAAAACG TCANCCAGNTNTTCACCGCATGAATACGTGAGTGGTCAGCCAGGTGTTCAACGC ATGAATAAGTGAGTGAATGAATGAATGAATGAGTAATTTGGTGGCTGACCCAAT GAGTGAATGAGGGAACAGGNGANTCCGTGAGCANCTCTGAGTGAAGAAGAGTGA NCCTCGAAGGAGAGTGATGTCAGGAGATGGTANTTTNTGNCAATTCGNCCACTG NGAGCCCACGTGGTATGTCTCTCATCGGAGCCACAGGGNAGNTATAGGNCGTNC TCAGGCACACGGGGGTCTCGGGNTGGCNAACAANANNAAAAAATGAATNGANGG NATGNATGGAAANNAANAAANTTGGGTGGNTTGGANGNAATGAANTGAATTGAA GGGAAAAAGNCGGAAACACGTTGANNCAACNNCTTGAGTGGAAGAAAGANTTGA GCCNTCGGAANGAAGGAGTGATGTNAAGGAAGAAGGGCNGGGTTGNTTGACCAA ANNGGTCCCACTTGGGAAGACAANGTGGGGATGTNTCTCCATCGTANNACAANA GGGGGAGNAAATAGACCGTGNTCAGGAAANANGGGGGNCTCGACNCCCCCGCGG GTGTCGACNCCCCTNCCTCCCTCCAGACCTGTNCACACTGCCCCTTTGNAGGGA GCCCNTCCTGGTCCTCAGTGTTTATANNTGCTGTCCCCNTCTACTGGGTGCTCC CCTGGCCGTGACATCACCACCAGCCAGANTGGCTTTTGTTCTCCTAAACATCTC TCCTCCCCTCACCCACCATGAGCTCCACGGGCTGAGGTCTCGGGCCACCCCTCC TCCCTCCCACCTCTTTCTCTCTTTCTCCAGGTGCCTTGTAGGTGCTTGGTGAGT TTCCTCATTCNACAAGTCCAGGCATTCATTCATGTGTGTGTTACACACNTAANN NCAGAGNAGNAN.

27. A method for screening for AIDS-related dementia comprising screening a patient sample for the presence of anti-HIV V3 antibodies, including T cell receptors, and correlating the presence of the antibodies with the presence of proteins in brain or spinal cord tissue or fluid from the patient that is immunoreactive with the anti-HIV V3 antibodies.

28. A peptide isolated from human tissue that is immunoreactive with an antibody directed to or immunoreactive with an HIV component.

29. The peptide of claim 28 consisting of between six amino acids and one hundred amino acids.

30. The peptide of claim 28 immunoreactive with an antibody to HIV gpl20.

31. The peptide of claim 30 immunoreactive with an antibody to HIV gpl20 V3.

32. The peptide of claim 28 derived from a protein is selected from the group of proteins expressed in a brain tissue cDNA library having a molecular weight of 17,000, 27,000, 52,000, 65,000, 90,000, 105,000 and 120,000 D immunoreactive with an antibody to gpl20.

33. The peptide of claim 28 in a pharmaceutically acceptable carrier.

34. The peptide of claim 28 in an oral formulation.

35. The peptide of claim 28 wherein the peptide is a portion of a protein encoded by the nucleotide sequence (Seq. I.D. No. 1) :

GGGGCTGACTTTGCTGCTGTGGGGGTGGGGACTGGGGTGCAGGCCCCACTNNTG GGTGTCAATANTGNGGCTACCATTNCTCCTGTGNGGGTGGGGACTGGGGTGCAG CCCCCACTACNNGNTGTCAATACTGGGGCTGCCTTTACTNCTGTGGGGGTGGGG ACTGGGGTGCAGNCCCCACTNCTGGGTGTCAATNCTGGGGCTNCCNTTNCTNCT GTGGGGGTGGGGACTGGGGTGCAGGGAGGGGACGGGTGAATAAACAGACACAGG TGGNCAGAGGCAGACAANAGCCCATGAATTCAGCGTGGGCCCAGCTCCCATGGT TGNCTCGCGGCCCAGGGGATNGTGCTTGGTGGNCAAGGAGTGTGAGGCCATCAG AGCTCCACAANGCGGTGTGGNGTGCTGCACNNCCCAACCCTNCCTCCCAAAACG TCANCCAGNTNTTCACCGCATGAATACGTGAGTGGTCAGCCAGGTGTTCAACGC ATGAATAAGTGAGTGAATGAATGAATGAATGAGTAATTTGGTGGCTGACCCΛAT GAGTGAATGAGGGAACAGGNGANTCCGTGAGCANCTCTGAGTGAAGAAGAGTGA NCCTCGAAGGAGAGTGATGTCAGGAGATGGTANTTTNTGNCAATTCGNCCACTG NGAGCCCACGTGGTATGTCTCTCATCGGAGCCACAGGGNAGNTATAGGNCGTNC TCAGGCACACGGGGGTCTCGGGNTGGCNAACAANANNAAAAAATGAATNGANGG NATGNATGGAAANNAANAAANTTGGGTGGNTTGGANGNAATGAANTGAATTGAA GGGAAAAAGNCGGAAACACGTTGANNCAACNNCTTGAGTGGAAGAAAGANTTGA GCCNTCGGAANGAAGGAGTGATGTNAAGGAAGAAGGGCNGGGTTGNTTGACCAA ANNGGTCCCACTTGGGAAGACAANGTGGGGATGTNTCTCCATCGTANNACAANA GGGGGAGNAAATAGACCGTGNTCAGGAAANANGGGGGNCTCGACNCCCCCGf-^G GTGTCGACNCCCCTNCCTCCCTCCAGACCTGTNCACACTGCCCCTTTGNA& ,A GCCCNTCCTGGTCCTCAGTGTTTATANNTGCTGTCCCCNTCTACTGGGTGCTCC CCTGGCCGTGACATCACCACCAGCCAGANTGGCTTTTGTTCTCCTAAACATCTC TCCTCCCCTCACCCACCATGAGCTCCACGGGCTGAGGTCTCGGGCCACCCCTCC TCCCTCCCACCTCTTTCTCTCTTTCTCCAGGTGCCTTGTAGGTGCTTGGTGAGT TTCCTCATTCNACAAGTCCAGGCATTCATTCATGTGTGTGTTACACACNTAANN NCAGAGNAGNAN.

36. The peptide of claim 28 wherein the peptide are derived from a protein selected from the group consisting of human ribosomal protein S17, human α-1- microglobulin bikunin, human gamma-glutamyl carboxylase, human cytochrome P450cl7, human beta B3 crystallin, non-muscle α-actinin, human beta- adrenergic receptor kinase, human β-adrenergic receptor kinase 1, human transmembrane protein tyrosine kinase, human liver/bone/kidney-type alkaline phosphatase, human protein tyrosine kinase, human C5a anaphylatoxin receptor, human growth hormone-releasing factor, human vacuolar H+ ATPase proton channel subunit, human arginine-rich gene, human small nuclear ribonucleoprotein, human UI RNA-associated 7OK protein, human testis specific protamine, human transducin Gx-alpha-subunit, human NKG2-C gene Type II integral membrane protein, human arginine-rich nuclear protein, human collagen alpha 1(V) chain, human homeo box cl protein, human homeobox protein HOX2.3, human alpha-1 type XI collagen, and human son3 protein gene.