The serum levels of anti-tat antibodies, tat protein, and p24 protein are predictive of disease progression in HIV infected individuals and serve as prognostic markers. The present invention relates to a method for determining the prognosis of an HIV infected individual by measuring serum levels of one or more of these prognostic markers. In addition, a method for monitoring whether an HIV infected individual is in need of immunization with a tat vaccine and a method for monitoring the efficacy of immunization with a tat vaccine are also disclosed.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention generally relates to diagnostic and prognostic tests for disease progression. More particularly, it relates to a prognostic test for the stage of progression of HIV infected patients and a method for treating HIV infected patients based on the prognosis.

2. Description of the Related Art

Following immune activation, HIV-1-infected CD4+ cells synthesize viral proteins and release virions (Zagury et al, 1986). In infected cells, the regulatory transactivating-transcription protein tat, which is not found in the viral particle, but is encoded by the HIV-1 genome, enhances the transcription of viral mRNA through interaction with the cis-acting RNA sequence termed TAR (Cullen, 1991). In contrast to its enhancing effect on viral replication, tat impairs the normal physiologic machinery of infected T cells, thus contributing to their premature death (Zagury et al, 1986). The tat-induced cellular alterations may also be exercised in uninfected T cells, since this protein, released from acutely infected cells into the extracellular compartment (Ensoli et al, 1993), can target uninfected cells by a mechanism involving interaction with integrins (Hynes, 1992) or other cell membrane receptors (Weeks et al, 1993). Immune cells, which carry a large density of integrin receptors, particularly after activation (Hynes, 1987), may be dysregulated by extracellular tat, as shown in vitro (Zagury et al, 1996). Pretreatment of activated PBMCs with tat induces both T cell immune suppression (Viscidi et al, 1995; Chirmule et al, 1995) and apoptosis (Li et al, 1995; Westendorp et al, 1995; Katsikis et al, 1997). Extracellular tat acts not only on T cells but also on antigen processing cells (APCs), e.g., monocytes, macrophages and dendritic cells (Zagury et al, 1998). The tat-induced immunosuppressive effect is further promulgated by the enhanced secretion of the immunosuppressive cytokine, IFN.alpha., by APCs (Zagury et al, 1998). The present inventors showed that PHA-activated PBMC cultures treated with HIV-1, but not untreated cultures, generate CD8.sup.+ suppressive T cells. In this in vitro experimental model, the potent role of tat and IFN.alpha. in HIV-1-induced immune-suppression was confirmed since specific anti-tat and anti-IFN.alpha., but not control antibodies prevented the generation of CD8.sup.+ suppressive T cells (Zagury et al, 1998).

Currently, the prognostic indicator or marker is viral load. However, the work of the present inventors shows that it is not a good prognostic indicator of future disease progression at an early stage of disease when the viral-load remains low, but is merely an indicator of the stage of disease (asymptomatic versus sick). While a test for the viral p24 protein is commercially available, this test is only used to determine whether an individual is infected, and not as a prognostic indicator/marker of disease progression.

Citation of any document herein is not intended as an admission that such document is pertinent prior art, or considered material to the patentability of any claim in the present application. Any statement as to content or a date of any document is based on the information available to applicants at the time of filing and does not constitute an admission as to the correctness of such a statement.

SUMMARY OF THE INVENTION

The present invention is based on the discovery that the immune parameters, tat protein, anti-tat antibody, and p24 protein levels, in HIV infected individuals are the best predictors of disease progression, at least at a non-advanced stage of disease. Using tat protein, anti-tat antibody, and p24 protein as prognostic markers, the present invention provides a method for determining the prognosis of an HIV infected individual by measuring the serum level of the prognostic marker and comparing it with either serum levels of the prognostic marker that are indicative of disease progression or non-progression or comparing it to a past measurement made earlier on the same individual.

The present invention also provides a method for treating an HIV infected individual by immunization with a tat vaccine when the monitoring of an individual's anti-tat antibody or tat protein level determines that a decrease in the level of anti-tat antibody or an increase in tat protein over time warrants the administration of tat vaccine.

Further provided is a method for evaluating the immune response of a non-infected individual as a result of immunization with a tat vaccine by measuring the level of anti-tat antibodies after immunization.

DETAILED DESCRIPTION OF THE INVENTION

To investigate whether particular serum markers in HIV infected individuals, such as antibodies against HIV-1 specificities or the p24 viral marker, are predictive of disease progression, serum was collected from 104 HIV-1 infected non-disease progressing individuals (chosen as individuals who have been infected more than eight years with CD4 cell counts always about 500 per mm.sup.3 and with no clinical sign of disease) at the time of enrollment in the study and at follow-up one to two years later. At the follow-up, 26 of the HIV-1 infected individuals showed signs of disease progression (CD4 cell decrease over 30% or presence of clinical symptoms). In this study, the present inventors discovered that, of the serum markers tested (antibodies to HIV antigens including env, gag, nef and tat proteins, as well as p24 antigenemia, viremia, CD4 cell count and IFN.alpha. titer), anti-tat antibodies and p24 antigenemia were the best predictive markers in the serum, with the anti-tat antibodies being inversely correlated with p24 antigenemia, which is already known as a marker of viral replication. These were the only two serum markers that significantly correlated the two subgroups, infected individuals with serum marker level above the median and subjects with serum marker level below the median, with progression and non-progression of disease.

The serum levels of anti-tat antibodies and p24 protein are used as prognostic markers in prognostic methods and treatments in accordance with the present invention. By "prognosis", it is intended that the progressive state of the HIV infected individual in early infection can be predicted. Thus, as the serum levels of anti-tat antibodies correlate inversely with disease progression, at high serum levels of anti-tat antibodies, the HIV infected individual is in a state of non-progression. The serum level of anti-tat antibodies determines the progression state of an HIV infected individual. As long as the anti-tat antibody levels are still high, the individual is non-progressing. Once the level of anti-tat antibodies begins to drop, then the prognosis is that the infected individual is entering a progressive state.

While the serum level of anti-tat antibodies is the preferred prognostic marker, the serum level of tat protein can also be used as well (complexed or uncompleted). Like the correlation between anti-tat antibodies and p24 protein, the serum level of tat protein correlates inversely with anti-tat antibodies. The higher the serum level of anti-tat antibodies, the lower the serum level of tat protein, and, conversely, the lower the serum level of anti-tat antibodies, the higher the serum level of tat protein.

In addition, besides measuring the serum level of anti-tat antibodies or tat protein as a prognostic marker/indicator, the serum level of p24, which is already a known marker for viral replication, can also be used as a prognostic marker in the present invention because it has been discovered by the present inventors to be highly correlative with disease progression. Tests for p24 to diagnose HIV-1 infection (but not the progressive state of the individual) are commercially available.

The present invention is directed to a method for determining the prognosis of an HIV infected individual by measuring the serum level of one or more of anti-tat antibodies, tat protein, or p24 protein as prognostic markers, and comparing the measured level to levels of the prognostic marker indicative of disease progression or non-progression. It is further contemplated that the measurement of the level of one or more of the anti-tat antibody, tat protein or p24 protein prognostic markers can be combined with the measurement of other markers, such as viral load, CD4 cell count, or IFN.alpha., to provide a more complete determination of the overall status of the HIV-1 infected individual. The level of any of the serum markers discussed above can be measured by any of a variety of assay techniques and is not limited to ELISA used in the Example herein, as would be appreciated by those of skill in the art. For instance these techniques could include the use of blots, beads, plates, immunoprecipitations, monoclonal or polyclonal antibodies against tat protein or fragments, the tat protein or tat peptides, competitive measurements or direct assays, enzymatic or radioactive markers, etc.

The correlation of high serum levels of anti-tat antibodies with non-disease progression among HIV infected individuals and low levels with disease progression strongly suggest that raising the serum level of anti-tat antibodies through active immunization may control viral replication. In this context, the present invention is also directed to a method for treating an HIV infected individual. The serum level of anti-tat antibodies is monitored over time in this method according to the present invention to determine whether the level of anti-tat antibodies has dropped sufficiently or the level of tat protein has increased sufficiently to warrant administration of a tat vaccine. By active immunization with a tat vaccine, the level of anti-tat antibodies is brought back up to a level indicative of non-disease progression. This same individual is monitored over time to confirm that the serum level of anti-tat antibodies is at a level predictive of non-disease progression and that this level is maintained over time. If a drop in serum level of anti-tat antibodies or an increase of tat protein in the serum is observed, further intervention, such as additional immunization with a tat vaccine, can be contemplated.

While the preferred tat vaccine is the tat toxoid vaccine disclosed in WO 96/27389, which contents are herein incorporated entirely by reference, the tat vaccine used in accordance with the present invention is not limited to such a tat toxoid vaccine. As will be appreciated by those skilled in the art, any tat vaccine which achieves the intended purpose of stimulating an immune response and raise the titer of anti-tat antibodies and which is within the skill of the art can be used in accordance with the present invention.

The present invention is further directed to a method for evaluating the immune response of a non-infected individual as the result of immunization with a tat vaccine. This method compares the level of anti-tat antibodies before and after immunization to determine the individual's humoral immune response to tat protein.
 

Claim 1 of 2 Claims

1. A method for determining the prognosis of an HIV infected individual, comprising: measuring the levels of anti-tat antibodies and p24 protein in the serum of an HIV infected individual; comparing the measured levels to a mean anti-tat antibody value and a mean p24 protein value; and determining the prognosis of the HIV infected individual, wherein anti-tat antibody levels above the mean value in combination with p24 protein levels below the mean value is indicative of disease non-progression towards AIDS, and anti-tat antibody levels below the mean value in combination with p24 protein levels above the mean level is indicative of disease progression towards AIDS.

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