WO1997000083A1

WO1997000083A1 - Method of treating cd4+ t cell lymphopenia in immuno-compromised patients

Info

Publication number: WO1997000083A1
Application number: PCT/US1996/010362
Authority: WO
Inventors: Robert G. L. Shorr; Mike A. Clark; David H. Goddard
Original assignee: Enzon Inc
Current assignee: Enzon Pharmaceuticals Inc
Priority date: 1995-06-16
Filing date: 1996-06-14
Publication date: 1997-01-03
Anticipated expiration: 1997-12-16
Also published as: AU6280296A

Abstract

The present invention is directed to methods of treating CD4+ T cell lymphopenia in HIV-infected patients. The methods include administering an effective amount of adenosine deaminase or related enzymatic material to a patient in need thereof. In preferred aspects of the invention, the method is employed in conjunction with HIV-infected patients having CD4+ T cell levels of less than about 200/νl. Effective amounts of the enzyme range from about 5 to about 50 IU/kg/week. In one particularly preferred aspect of the invention, the adenosine deaminase is conjugated to one or more strands of polyethylene glycol to prolonged activity in vivo.

Description

METHOD OF TREATING CD4⁺ T CELL LYMPHOPENIA IN IMMUNO COMPROMISED PATIENTS

BACKGROUND OF THE INVENTION

The present invention relates to methods of increasing CD4+ T cells in patients infected with the human immunodeficiency virus (HIN). In particular, the present invention relates to a novel use of adenosine deaminase for treating CD4+ T cell lymphopenia. Acquired immunodeficiency syndrome (AIDS) is a disease characterized by a progressive loss of function of the immune system. As a result, those afflicted with the syndrome are susceptible to a variety of opportunistic infections. The etiologic agent of AIDS is a cytopathic retrovirus designated the human immunodeficiency virus (HIN). One of the major targets of the HIN in humans is T helper cells (CD4+ cells). The infection of T helper cells by HIN results in a profound dysregulation of the immune system including both depleted numbers and impaired functioning of T lymphocytes. Although the exact mechanism is unknown, the number of T helper cells predictably declines during HIN infection. Clinicians monitor this decline as an indicator of disease progression. Research in the AIDS field has focused on several goals: preventing HIN infections, treating HIN-infected individuals with anti-viral agents, treating opportunistic infections with antimicrobial agents, and restoring the immune system of infected patients.

HIV gains entry into T helper cells by binding to receptors on the cell surface. Initial contact between the virus and the T cell is believed to occur by interaction of the external viral glycoprotein gpl20 and the CD4 protein present on the surface of the T cell. Recently, it has been postulated that a second receptor on the T cells, the CD26 receptor may also be involved with facilitating HIN entry into T cells. CD26 also binds adenosine deaminase (ADA) on the surface of T lymphocytes and may play a role in T cell activation. In HIN infection, the immune system is chronically activated. This chronic activation is believed to play a primary role in the pathology of the disease. The persistence of the virus and viral replication are thought to play a primary role in mamtaining this state of immune activation. Evidence of this phenomenon includes spontaneous lymphocyte proliferation, activation of monocytes, expression of T cell activation antigens on the surface of T helper cells, increased cytokine expression and elevated levels of ADA in the serum and erythrocytes. Thus, the increase in ADA may be a consequence of activation of the immune system in response to infection. In view of this observation, there have been suggestions that plasma ADA and/or the isoenzyme ADA₂ would be useful markers of the disease's progress. See AIDS 1990 4:365-373 and J. of Acquired Immun. Defic. Synd. 4:178-82 (1991). Alternatively, it has also been postulated that increase in ADA in HIN disease is an adaptive response by the immune system to decreasing numbers of T lymphocytes. In contrast, Renouf et al. (Clin. Chem. 35:1478-1481, 1989) report that lymphocyte ADA levels are reduced in patients with HIV infection compared to uninfected controls. Nonetheless, in HTV-infected patients, the loss of CD4+ cells is associated with lymphocyte activation which does not result in cell proliferation as one would normally expect. Instead, this activation results in cell death by apoptosis which is also known as programmed cell death. This physiological suicide mechanism usually functions as a part of homeostasis i.e. normal tissue turnover. Apoptosis is essential in the normal maturation of the immune system. Apoptic cell death is part of the normal development of self-tolerance. Apoptosis, however, can be triggered in immature thymocytes by several events including specific activation of the T cell receptor CD3 complex. Mature T cells are usually resistant to these apoptic stimuli and respond to T cell receptor stimulation by cell proliferation and cytokine secretion. Abnormal induction of apoptosis in mature T cells, however, is also known. Apoptosis in CD4+ T cells has also been observed in HIN-infected individuals, especially upon T cell receptor activation. It has been speculated that the complexing of the external glycoprotein gpl20 of HIN with the CD4+ receptor may cause apoptosis even in noninfected cells. See Science vol. 260 May 28, 1993, 1269-1270.

In view of the foregoing, it would be a significant advance in the field of AIDS-related treatment if CD4+ lymphopenia could be addressed therapeutically. It would also be advantageous to effectively treat HIN-initiated apoptosis of CD4 + T cells. The present invention addresses these needs.

SUMMARY OF THE INVENTION

In one aspect of the present invention there is provided a method of treating CD4+ T cell lymphopenia in humans. The method includes administering an effective amount of an adenosine degrading agent to a human in need of such treatment to provide a significant increase in the CD4+ levels and dierefore reduce or eliminate the lymphopenia.

In another aspect of the invention there is provided a method of reducing CD4+ T cell apoptosis in susceptible mammals. A still further aspect of this invention includes a method of increasing the therapeutic effect of reverse transcriptase inhibitor agents or anti-viral agents used in the treatment of mammals.

These methods also include administering an effective amount of an adenosine degrading agent to the patient in need thereof in order to reduce the T cell apoptosis and/or increase the therapeutic effect of the anti- viral agent in patients receiving the agents. The invention also includes a new use of adenosine deaminase or other adenosine degrading substances for treatment of certain medical conditions in mammals. The new use includes administering an effective amount of the adenosine deaminase in combination with one or more reverse transcriptase inhibitor agents or anti-viral agents a patient in need of such treatment. In this aspect of the invention, the adenosine deaminase is a useful adjunct in the treatment of Acquired Immunodeficiency Disease (AIDS) by assisting in the metabolism of various anti¬ viral drugs into active metabolites. For example, it has been found that adenosine deaminase appears to metabolize agents such as dideoxyinosine, halogenated derivatives of dideoxypurines and dideoxyadenosine and its derivatives. Further, the adenosine deaminase metabolites of many deoxynucleoside analogs have anti- viral activity which, in some cases, is activity greater than that of the parent compound. Thus, the adenosine deaminase agents can be used in treatments as a facilitator which acts to assist in the release of active agents from prodrugs. In addition, the anti-viral activity of these types of agents can be completely abrogated in the presence of adenosine deaminase inhibitors. Consequently, administering me adenosine deaminase agents described herein can block or limit the abrogation of anti-viral activity of the concomitantly administered anti- viral agent.

The adenosine degrading agents useful in carrying out the methods described herein include enzymes obtained from suitable mammalian sources as well as those prepared using recombinant techniques. One particularly preferred adenosine degrading agent is bovine adenosine deaminase (ADA). Such agents are administered in amounts which are described as being "an effective amount" . As will be described in more detail below, amounts which are sufficient to substantially reduce or eliminate the lymphopenia and/or T cell apoptosis generally range from about 5 IU/kg/week to about 50 IU/kg/week and preferably from about 10 IU/kg/week to about 30 IU/kg/week.

In particularly preferred aspects of the invention, the adenosine degrading agent is a bovine adenosine deaminase enzyme administered as a polyethylene glycol conjugate such as that available from Enzon, Inc. under the trademark ADAGEN'

(PEG-ADEMASE, BOVINE).

For purposes of the present invention, "lymphopenia" shall be understood to mean T-cell levels of less than 200 cells/ μl, as measured by fluorescence activated cell sorting (FACS). The term "opportunistic infection" shall be understood to mean an infection which usually occurs only in patients whose resistance is compromised or lowered by an unrelated condition, disease or as a result of drug therapy.

As a result of the present invention, the artisan is provided with a useful alternative or supplement to currently available AIDS therapies. While the treatment methods of the present invention are not anti-viral treatments, they nonetheless cause increases in CD4+ T cell lymphocytes and substantially reduce CD4+ T cell apoptosis, especially when given in combination with an anti- viral agent such as AZT. Patients receiving effective amounts of the agents described herein will therefore be better able to combat opportunistic-infections and have an improved quality of life.

For a better understanding of the present invention, reference is made to me following description and its scope will be pointed out in the appended claims.

DETAILED DESCRIPTION OF THE INVENTION

In one preferred aspect of d e invention, a method of treating CD4+ T cell lymphopenia in HIV-infected patients is provided. The method includes admimstering an effective amount of an adenosine degrading agent such as an adenosine enzyme to a human in need thereof, whereby the rate of decline of die CD4+ T cell counts in the human is substantially reduced or reversed. Since the progressive decline in the T cell count is the hallmark of AIDS caused by infection with HIV, the ability to arrest and/or prevent falls in the CD4+ cell count below about 200/μl; or even reverse such low cell counts, will reduce or eliminate the patient's susceptibility to opportunistic infections. In preferred aspects of this embodiment, die method includes maintaining CD4+ levels of at least about 200 cells/μl or raising CD4+ cell counts above that level by administering a sufficient amount of d e adenosine degrading agents described herein to a patient in need of such treatment. The amounts of adenosine degrading agent used in me methods of die present invention are generally described as amounts which effectively achieve the sought after therapeutic effect, i.e. reversing the lymphopenic condition or increasing the therapeutic effect of anti-viral agents which inhibit reverse transcriptase in mammals. The maximal dosage is me highest dosage which does not cause clinically important side effects. The lowest dosage contemplated is mat which is still effective enough to achieve the results described above. Naturally the doses will vary somewhat depending upon me adenosine degrading enzyme or analog or isoenzyme used but me adenosine degrading agent will usually be administered in amounts ranging from about 5 IU/kg/week to about 50 IU/kg/week and preferably in amounts ranging from about 10 IU/kg/week to about 30 IU/kg/week. It will be understood mat die doses are described as being based on me amount of adenosine deaminase (ADA) administered. Such doses are given for purposes of illustration and d ose of ordinary skill in the art will determine optimal dosing of me adenosine degrading agent (ADA or related compound) based on clinical experience. It is to be understood tiiat the administration of adenosine degrading agent will also be dependent upon die individual patient. For example, while it is contemplated diat the adenosine degrading agent will be administered on a chronic basis in an effort to combat the HIV effects, it will also be understood from the foregoing that episodic administration of the adenosine degrading agent on an as needed basis is also contemplated.

In one particularly preferred aspect of the invention, the adenosine degrading agent administered is bovine adenosine deaminase (ADA) covalently conjugated to polyethylene glycol strands which is available under the tradename ADAGEN^*, (PEG-ADEMASE, bovine) a product of Enzon, Inc., (Piscataway, NJ). This particular agent is administered in accordance with the methods of the present invention parenterally in pharmaceutically acceptable solutions in amounts ranging from 10 IU/kg/week to about 30 IU/kg/week based on the ADA. Intramuscular administration of die ADA conjugate is particularly preferred. The ADA- polyethylene glycol conjugates can also be prepared using the techniques described in U.S. Patent No. 5,324,844, the disclosure of which is hereby incorporated by reference. The '844 patent describes, among other things, forming a substantially hydrolysis-resistant urethane bond between the epsilon amino groups of enzymes and a functionalized terminal group. The linkage by which the enzyme is joined to die polymer strand(s) can be any moiety known in the art which sufficiently unites the enzyme and polymer so that the conjugate may be administered in a pharmaceutically acceptable manner. In addition to me urethane linkage, amide linkages are also preferred. An example of amide-linked polymer enzymes is found in U.S. Patent No. 5,349,001, the disclosure of which is incorporated by reference herein. The '001 patent describes, inter alia cyclic imide-activated polyalkylene oxides and conjugation tiiereof with therapeutic proteins and enzymes induding adenosine deaminase.

ADAGEN is currently used to treat human patients with severe combined immunodeficiency disease (SCID) secondary to an inborn mutation inactivating the gene for adenosine deaminase. In this condition, ADA levels are essentially undetectable (less tiian 1 % of normal levels; see Hirschhorn, Pediatnc Research 33 (suppl. ):535-541, 1993). ADA deficiency results in accumulation of adenosine metabolites, especially deoxyadenosine triphosphate, which inhibit DNA synthesis and thus cause deatii of T cells. ADAGEN treatment reverses SCID by reducing deoxyadenosine triphosphate levels. See Blaese and Culver, Immunodeficiency Reviews 3:329-249 (1992).

It has been surprisingly found tiiat a separate and unexpected benefit is realized when adenosine degrading agents such as adenosine deaminase are administered to HIV-infected patients in order to treat or prevent CD4 + lymphopenia. It is believed that accumulation of adenosine metabolites, especially deoxyadenosine triphosphate, does not play a significant role in the lymphopenic condition and therefore administration of adenosine degrading agents could not have been predicted to successfully treat die lymphopenic condition. The adenosine degrading agents which can be administered in accordance with the methods of the present invention can be prepared or obtained from a variety of sources, including recombinant or mammalian-extracted ADA. Although bovine adenosine deaminase is particularly preferred when it is included as part of a polyethylene glycol conjugate, it is to be understood that the methods described and claimed herein can also be carried out using gene therapy techniques. In this aspect of the invention, ADA or a similar adenosine degrading agent is administered using, for example, the procedures described by Blaese and Culver in Immunodeficiency Reviews 3:329-349 (1992) and die references cited dierein. The disclosure of the aforementioned Immunodeficiency Reviews 3:329-349 (1992) is incoφorated by reference herein.

Cultures are established from CD34+ hematopoietic progenitor cells isolated from the blood of an HIV-infected human patient. See Chelucci et al., Blood 85:1181-1187, 1995, the disclosure of which is incorporated by reference herein. A portion of the culture is treated by retroviral-mediated transduction widi the

LASN vector. The LASN vector contains the human adenosine deaminase gene. Treated CD34+ cells containing the transduced ADA gene are then reinfused into the patient. Alternatively, ADA can be obtained from mammalian sources such as bovine, ovine, human, etc. It is to be understood that other substances including pro-enzymes, fractions of enzymes, and catalytic antibodies can also be included in the present invention.

As used herein, the expression "adenosine degrading agents" means all suitable substances which demonstrate in vivo activity to reduce adenosine levels and treat CD4+ T cell lymphopenia and/or combat the HJN-mediated CD4+ T cell apoptosis. These substances are prepared by using techniques known to those of ordinary skill in the art such as tissue culture, extraction from plant or animal sources or by recombinant DΝA methodologies. Transgenic sources of enzymes, pro-enzymes and fractions thereof are also contemplated. Such materials are obtained from transgenic animals, i.e. mice, pigs, cows, etc. wherein the enzyme is expressed in milk, blood, or tissues. The catalytic antibodies can be prepared using recombinant technologies. The method by which the enzymatic substance is prepared or obtained for the treatment methods and conjugates of the present invention is not limited to those described herein. Substantially non-antigenic polymer substances can be included in the ADA conjugates. Within this group of substances are alpha-substituted polyalkylene oxide derivatives such as metiioxypolyethylene glycols or other suitable alkyl substituted derivatives such as C_rC₄ alkyl groups. It is preferred, however, that the non-antigenic material be a monomethyl-substituted PEG homopolymer. Alternative polymers such as other polyethylene glycol homopolymers, polypropylene glycol homopolymers, other alky 1-poly ethylene oxides, bis-polyethylene oxides and co¬ polymers or block co-polymers of poly (alkylene oxides) are also useful. In those aspects of the invention where PEG-based polymers are used, it is preferred that they have molecular weights of from about 1,000 to about 100,000. Molecular weights of about 2,000 to 40,000 are preferred and molecular weights of about

5,000 are especially prefened.

Alternative non-antigenic polymeric substances include materials such as dextran, polyvinyl pynolidones, polysaccharides, starches, polyvinyl alcohols, polyacryl amides or other similar non-immunogenic polymers. Those of ordinary skill in the art will realize that the foregoing is merely illustrative and not intended to restrict the type of non-antigenic polymeric substances suitable for use herein. As stated above, a covalent modification of the adenosine degrading agent, enzyme or enzyme-like material with PEG or a related substantially related non- antigenic polymer is prefened to provide the hydrolysis-resistant conjugate. The covalent modification reaction of an adenosine degrading agent includes reacting a substance having the desired adenosine degrading activity with a substantially non- antigenic polymeric substance under conditions sufficient to effect conjugations while maintaining at least a portion of the ADA activity. See, for example U.S. Patent Nos. 4,179,337 and 5,122,614, the disclosure of which are incorporated by reference herein. While the references incorporated herein describe epsilon amino group modifications of lysines found on the ADA, other conjugation methods are also contemplated. For example, modification of carboxylic acid groups and odier reactive amino acid groups are also within the scope of the present invention. Covalent linkage by any atom between the adenosine degrading agent and polymer is possible. Moreover, non-covalent associations between polymer and adenosine degrading agent such as lipophilic or hydrophilic interactions are also contemplated. Following the conjugation reaction, the desired product is recovered using known techniques and purified using column chromatography or similar apparatus if necessary. Depending upon the reaction conditions, the conjugates have from about 1 to about 25 polymeric strands attached to each molecule of the enzyme-like substance. By controlling die molar excess of the polymer reacted with the enzyme, for example, the artisan can tailor the number of polymeric strands attached. Conjugates containing from about 5 to about 20 polymeric strands are prefened while conjugates containing from about 10 to 18 polymeric strands are most prefened.

In alternative aspects of the invention, the adenosine degrading agent is administered in a liposome. A non-limiting example of such liposomes can be found in U.S. Patent No.4,534,899, die disclosure of which is incorporated herein by reference. Still further aspects of the invention include administering die adenosine degrading agent in the form of an oral dosage form which allows the agent to be bioavailable. It is to be understood from the foregoing that the delivery system selected or the means of administering the adenosine degrading agent to d e patient is not to be construed nanowly and it is contemplated that any pharmaceutically acceptable delivery system can be used to carry out the inventive treatment methods.

Patients suffering from AIDS due to HIV infection have a severe lowering in the numbers of T lymphocytes circulating in their blood. This results in their resistance to infection being severely impaired. While Applicants are not bound by any particular tiieories, it is believed that the administration of ADA to CD4+ T lymphopenic or apoptoic patient in need thereof provides clinical benefits by reducing the level of circulating adenosine and thus reducing apoptosis of CD4 + T lymphocytes. Alternatively, ADA may act by blocking the binding of HJN to the CD4+ T lymphocytes by competitively blocking the gpl20 binding coreceptor sites which are designated CD26 and it has been postulated tiiat in order for the HIN to gain entry into the T cells, this coreceptor must also be available. In addition, ADA- based therapies may also have a direct effect on CD4+ cells. For example, ADAGEN is an FDA approved drug which has been shown to promote maturation of T lymphocytes. As a result, the benefits from the ADA-based treatment method include an increase in patient's personal sense of well being, decreases in the numbers of infections and a sustained increase in die numbers of circulating CD4+ T cells in their blood.

In yet a still further aspect of die invention, ti ere is provided anotiier method of treating HJN-infected humans. In particular, tiiere is provided a metiiod enhancing the therapeutic effect of antiviral agents in HIN-infected humans. This method, as described above, would appear every weekly This aspect includes administering an effective amount of an adenosine degrading agent as described above in combination with an effective amount of an antiviral agent. Suitable antiviral agents include without limitation, 3' azidothymadine (AZT, zidovudine),

2' ,3'-dideoxyinosine (ddl, didanosine), , 3 '-dideoxycytidine (DDC, zalcitabine) and d e like and mixtures thereof . The exact amounts of such antiviral agents will be apparent to those of ordinary skill in the art based on clinical experience. Generally, however, it is contemplated tiiat these agents be administered in FDA- approved amounts, such as those set forth in the Physician's Desk Reference, cunent addition. For purposes of the present invention, the term "in combination with" shall be understood to mean that the adenosine degrading agent is administered at not only the same time as the antiviral agent but also within a time which allows the combination of antiviral and adenosine degrading agent to have its synergistic effect. Thus, the adenosine degrading agent can be administered at die start of, during or substantially immediately after a course or antiviral agent(s). In this aspect of die invention, synergistic effects are observed.

In yet a still further aspect of the invention, there is provided a method of treating adenosine-induced T-cell toxicity in humans. The metiiod includes administering an effective amount of an adenosine degrading agent to a patient in need thereof . This method of treatment is to be contrasted with the treatments used for SCID (described above) since it is not necessary that patients in need of die treatment described herein, to anest adenosine induced toxicity, have the severely deficient ADA levels observed in patients presenting witii SCID. The amounts of the adenosine degrading agent administered in this aspect of the invention will be in the same range as those described herein for the other treatments described.

EXAMPLES The following examples serve to provide further appreciation of the invention but are not meant in any way to restrict the effective scope of the invention.

EXAMPLE I Modification of bovine adenosine deaminase witii SC-PEG

In this example, bovine adenosine deaminase (ADA) obtained from Sigma Chemical Co. (St. Louis, MO) is conjugated wid the activated poly(etiιylene glycol)-N-succinimide carbonate (SC-PEG) described in U.S. Patent number 5,324,844. The PEG polymer has a molecular weight of about 5,000. 10 mg of ADA in 92 Mm NaOAc pH 5.8/10 % glycerol/18 % EtOH is dialyzed witii 0.1 M sodium phosphate buffer solution, pH 7.0 using a Centricon-10 (a product of die Amicon Corporation of Beverly, MA). The final concentration of the ADA is -0.5 mg/ml. Next, 100 mg of SC-PEG (10-fold excess by weight = 120 molar excess) is added to the enzyme solution and die reaction mixture was stined for 1 hr. at room temperature. The reaction is quenched by adding 0.1 M glycine. The unreacted PEG is removed by dialysis into a buffer solution having a pH of 6.5. The extent of modification is checked by SDS-gel.

EXAMPLE II

In this example, the ability to use adenosine deaminase in the treatment of and/or prevent severe T cell lymphopenia in patients with HIN was demonstrated. ADAGEN ^* (PEG-ADEMASE,bovine) is a product of Enzon, Inc., (Piscataway, NJ). The drug was administered to 17 patients enrolled in this study. Entry criteria into the study included a CD4- T lymphocyte count of about 200 cells/μl or less and no past history of substance abuse. Patients remained under die care of their treating physician, and no alterations were made to odier drug therapy. The patients were evaluated at baseline to determine tiieir history, including documentation ofthe numbers of opportunistic infections, physical examination, and a baseline hematologic profile including T cell subsets.

The patients received parenteral (intramuscular) ADAGEN, 1500 IU intramuscularly once weekly for 12 weeks in combination with their cunent AZT or other anti-viral regimen. This data was compared to historical data used as a control Kahn et al. NEJM 327, 581 (1992) and Eron et al. NEJM 333, 1662 (1995), the contents of each are incorporated by reference herein. In Kahn et al., for example, the decline in efficacy over time for AZT is documented. Anotiier study (Spruance, S.L. et al. Annals of Internal Medicine Vol. 120, No. 5, March 1, 1994 pp360-368) reported tiiat HIV patients receiving zidovudine (AZT) experienced a chronic decrease in CD4⁺ cells over the initial 12 week period of tiie study. The patients receiving ADAGEN were monitored with T cell subset measurements. The results of the T cell subset measurements shown below indicate that die CD4+ T cell subset measurements for die patients receiving ADAGEN in combination with their cunent anti-viral therapy were either higher or did not decrease as much as would have been expected in view of d e previous study. Patient Initial Duration of Final CD4+ Change in CD4+ Treatment Count CD4+ Count (Weeks) Count

1. 22 12 16 - 6

2. 42 12 20 - 22

3. 162 12 177 + 15

4. 28 12 16 - 12

5. 22 12 13 - 9

6. 0 12 5 + 5

7. 22 12 17 - 5

8. 48 12 14 - 34

9. 55 12 143 + 88

10. 70 12 120 + 50

11. 108 12 186 + 78

12. 165 9 140 - 25

13. 170 11 252 + 82

14. 171 12 186 + 15

15. 112 9 98 - 14

16. 210 17 305 + 95

17. 160 19 150 - 10

The Table in particular demonstrates that the chronic decline in CD4⁺ cells known to occur in patients receiving anti- viral therapy widi agents such as AZT can be significantly reduced or even reversed by administering adenosine deaminase in combination with die anti- viral agent. Indeed, die average CD4⁺ cell count improved an average of 17.1 in the patients treated with ADAGEN. The historical control data indicates tiiat an average decrease of about 23 to 25% in CD4⁺ cells would have been expected. An additional result observed was die fact tiiat none of the patients in the smdy had an adverse reaction caused by d e ADAGEN. The patients in die study also did not experience any opportunistic infections during the course of tiie study. The historical data suggested from about 6.7 to about 8.5 % of die patients would have had at least one opportunistic infection during tiie course of the study. Most of die patients reported some improvement in well-being from being an on the treatment regimen. Finally, no deaths were observed in die ADAGEN treated patient group whereas die control data would suggest diat from about 10-20 % would have died during the course of the study.

EXAMPLE III In this example, ADA is admimstered by gene therapy methods following the procedures described in Blaese and Culver, Immunodeficiency Reviews 3:329-349 (1992) and the references cited dierein.

Cultures are established from CD34+ cells isolated from tiie blood of an HIV-infected human patient by the methods of Chelucci et al., (Blood 85:1181- 1187, 1995). The CD34+ cells may optionally be rendered resistant to HIV infection by transformation with an anti-HIV vector such as that of Buonocore et al. , PNAS 90:2695-2699, 1993) or that of Marasco et al., (PNAS 90:7889-7893, 1993), the disclosures of which are incorporated by reference herein. A portion of me culture is then treated by retroviral-mediated transduction witii the LASN vector described by Blaese and Culver, supra. The LASN vector contains the human adenosine deaminase gene. Treated T cells containing the transduced ADA gene are then reinfused into the patient. The patient exhibits increased levels of ADA resulting in increased levels of CD4+ T lymphocytes and decreased occunence of opportunistic infections. While there have been described what are presently believed to be die prefened embodiments of die present invention, those skilled in die art will realize that changes and modifications may be made tiiereto without departing from the spirit of the invention. It is intended to claim all such changes and modifications as fall widiin the true scope of the invention.

Claims

WHAT IS CLAIMED IS;

1. A method of treating CD4+ T cell lymphopenia in a human patient infected with a human immunodeficiency virus, comprising admimstering an effective amount of an adenosine degrading agent.

2. The method of claim 1, wherein said adenosine degrading agent is an adenosine deaminase enzyme.

3. The method of claim 1, wherein said adenosine degrading agent is selected from the group consisting of bovine adenosine deaminase, human adenosine deaminase, adenosine-degrading catalytic antibodies and mixmres thereof.

4. The method of claim 2, wherein said adenosine deaminase enzyme is administered to said mammal in the form of a polyethylene glycol conjugate.

5. The metiiod of claim 4, wherein said adenosine deaminase enzyme is derived from a bovine source.

6. The method of claim 5, wherein said effective amount of said adenosine deaminase is from about 1 to about 100 mg/kg/week.

7. The method of claim 6, wherein said effective amount of said adenosine deaminase is from about 5 to about 50 mg/kg/week.

8. The method of claim 7, wherein said effective amount of said adenosine deaminase is from about 10 to about 30 mg/kg/week.

9. The method of claim 1, wherein said adenosine degrading agent is admimstered to said human via gene therapy .

10. A method of increasing CD4+ T lymphocytes in humans infected with a human immunodeficiency virus, comprising administering an effective amount of an adenosine degrading agent to a human in need thereof .

11. A method of preventing T cell apoptosis in an human immunodeficiency virus-infected human, comprising administering an effective amount of an adenosine degrading agent to said human.

12. A metiiod of enhancing the therapeutic effect of antiviral agents in human immunodeficiency virus-infected humans, comprising administering an effective amount of an adenosine degrading agent in combination with an effective amount of an antiviral agent.

13. The method of claim 12, wherein said anti- viral agent is selected from the group consisting of zidovudine, didanosine, zalcitabine and mixtures thereof.