AU668088B2 - Pharmaceutical dipeptide compositions and methods of use thereof - Google Patents

Description

OPI DATE 02/11/92 AOJP DATE 10/12/92 APPLN. ID 17503 92 PCT NUMBER PCT/US92/02440 INTERN I J N TREATY (PCT) (51) International Patent Classification 5 International Publication Number: WO 92/17191 A61K 37/00, COK 5/00 Al A6K 37/00, C7K 500 Al (43) International Publication Date: 15 October 1992 (15.10.92) (21) International Application Number: PCT/US92/02440 (72) Inventors; and Inventors/Applicants (for US only) KHAVINSON, Vladi- (22) International Filing Date: 1 April 1992 (01.04.92) mir Khatskelevich [RU/RU]; Zoologichesky Pereulok, House Apt. #83, St. Petersburg, 197198 (RU).

MOROZOV, Vyacheslav Grigorievich [RU/RU]; Zo- Priority data: ologichesky Pereulok, House Apt. #83, St. Peters- 678,129 1 Ap.'il 1991 (01.04.91) US bourg, 197198 YAKOVLEV, German Mikhailovich [RU/RU]; ul. Komissara Smirnova, 8-50, St. Petersburg, 194175 SERY, Sergey Vladimirovich [RU/ Parent Application or Grant RU]; 3rd Rabfak per., 12-2-11, St. Petersburg, 139012 (63) Related by Continuation

(RU).

US 678,129 (CIP) Filed on 1 April 1991 (01.04.91) (74) Agents: KENNEY, Ernest et al.; Bacon Thomas, 625 Slaters Lane, Fourth Floor, Alexandria, VA 22314 (US).

(71) Applicant (for all designated State' except US): CYTOVEN [US/US]; 1309, 114th Avenue, Suite 104, Bellevue, (81) Designated States: AU, CA, US.

WA 98004 (US).

Published With international search report.

(54)Title: PHARMACEUTICAL DIPEPTIDE COMPOSITIONS AND METHODS OF USE THEREOF (57) Abstract Methods are provided for the therapy of immunodeficient, immunodepressed or hyperactive immune states and for the prevention and treatment of opportunistic infections in such states comprising administering to a subject a pharmaceutically acceptable composition comprising as an active ingredient the dipeptide L-Glu-L-Trp and/or its pharmaceutically acceptable salts.

U I I a~alnu WO 92/17191 PCT/US92/02440 PHARMACEUTICAL DIPEPTIDE COMPOSITIONS AND METHODS OF USE THEREOF This is a continuation-in-part of copending Serial No. 07/678,129, filed April 1, 1991.

The present invention is directed to dipeptide pharmaceutical compositions and uses thereof, in particular, uses thereof for treatment of immunodepressed states and of opportunistic infections in immunodepressed states.

BACKGROUND OF THE INVENTION Several polypeptides found in the thymus gland have been implicated as playing roles in the development and maintenance of immunological competence in animals, including human beings. Some of these polypeptides have been shown to stimulate the maturation, differentiation and function of T-cells.

For example, a heat-stable fraction isolated from calf thymus extracts, designated as Thymosin fraction 5, has been shown to reconstitute immune functions in thymic-deprived or immunodepressed individuals. Several peptides have been isolated from Thymosin fraction 5, such as Thymosin alphaI (28 amino acids, U.S. Patent No. 4,079,127), Thymosin beta 4 (44 amino acids, Low et al., PNAS, 78,1162-1166 i.L.Y Z~ Z~ IU~=nc~ ;i -E1U-~ t WO 92/17191 PCT/US92/02440 -2- (1981)), Thymosin beta 8 (39 amino acids, U.S. Patent No. 4,389,343) and Thymosin betag (41 amino acids, U.S. Patent No. 4,389,343). However, practical administration of such polypeptides is expensive due to the relatively low yield and complexity of isolation and/or manufacture of such long chain polypeptides. Most importantly, in some cases, these polypeptides produce side reactions in patients.

The present invention is based in part on the discovery that a dipeptide, hereinafter referred to as Thymogen, exhibits a broad range of efficacy for prevention and treatment of opportunistic infections in immunodepressed states, and for therapeutically effective treatment of immunodeficient states. This is believed to be highly unexpected for such a relatively small compound to exhibit such a broad range of activity. Furthermore, we have not found any significant side effects from the use of the dipeptide according to the present invention. Due to its simple nature, the dipeptide is rather inexpensive to manufacture.

As used herein, the terms "immunomodulator" and "immunomodulating" encompass the activity of enhancing or restoring the subject's immune system, as evidenced by measurable blood parameters and/or the patient's improved ability to combat infection or disease, and the ability to heal tissue. Hence, immunomodulation encompasses improvement of the immune system due to an immunodeficient state (for example, caused by removal of the thymus), and/or an immunodepressed state (for example, caused by exposure to radiation). Furthermore, the present invention provides for modulation of the immune system by lowering blood parameters and other indicia of the immune state if these indicia are abnormally -3- WO 92/17191 PCT/US92/02440 elevated. The present invention encompasses the therapeutic method of treating the immunodeficient, immunodepressed or elevated immune state per se, thus providing prophylaxis against infection and disease, as well as a treatment of infection, disease or wound indirectly by enhancing the immune system.

It is therefore an object of the present invention to provide pharmaceutical compositions of the dipeptide Thymogen which have broad immunomodulating activity, as well as activity for other uses such as treatment of infections, disease and wounds (burns, frost bites, and the like), enhancement of metabolic processes, and many other uses.

It is an object of the present invention to provide therapeutic methods for treatment of immunodepressed and immunodeficient states.

It is yet another object of the present invention to provide methods for preventing and treating opportunistic infections in immunodeficient and immunodepressed states.

These and other objects will be apparent from the following description and appended claims.

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention provides pharmaceutical preparations comprising the dipeptide L-Glu-L-Trp, using the normal convention wherein the first named amino acid is the amino terminus and the last named amino acid is the carboxyl terminus. The compositions according to the present invention may be formulated into any convenient formulation which allows for the active ingredient to be absorbed into 1 WO 92/17191 PCT/US92/02440 -4the blood stream. Intramuscular and intranasal forms of application are preferred. The preferred dosage rate of the active ingredient for intramuscular administration is about 50 to 100gg per dose for adults (for a 300 to 1000g total treatment therapy); for infants up to 1 year old about 10/g per dose, for infants 1 to 3 years old about 10 to 20/g per aose; for infants 4 to 6 years old about 20 to 30Ag per dose, for children 7 to 14 years old about 50/g per dose. All of the foregoing dosages are useful for a treatment of 3 to 10 days, depending upon the immunodeficiency level. The treatment may be repeated as needed, usually within 1 to 6 months.

For prophylactic uses against opportunistic infections in immunodeficient or immunodepressed patients, the intramuscular and/or intranasal single daily dose for adults may be from about 50 to and for children about 10 to 50 Ag per dose for treatment over 3 to 5 days.

For treatment of burns, frost bite, or other wounds, including chronic apical periodontitis, the dipeptide may be applied in about 100g doses as a paste or other suitable medium.

For ophthalmology, such as for treatment of infectious eye diseases, the dipeptide may be applied in single daily dosages of about 10g (over 4 to days) or as installations into the conjunctival cavity at about 5/g twice daily over about 4 to days.

The dipeptide may be utilized intramuscularly as an injection solution with the active ingredient in a therapeutically effective immunopotentiating amount of about .001 to .01% by weight. If presented in the Illl~- ~-ceri- ~-mraP~ma~-a;rP1 aaP-na~ -i 1 ir imiiiilii~~y WO 92/17191 PCT/US92/02440 form of a tablet, capsule or suppository it is preferred that the active ingredient be present in an amount of about 0.1mg per tablet, suppository or capsule. If presented in such form, the capsule, suppository or tablet may also contain other conventional excipients and vehicles such as fillers, starch, glucose, etc.

The dipeptide may be obtained by conventional peptide synthesis, including the Merrifield solid state neptide synthesis technique. Typically an amino and !de chain protected derivative of an activated ester of glutamic acid is reacted with protected Ltryptophan. After elimination of the protecting groups and conventional purification, such as by thin layer or GL chromatography, the peptide may be purified such as by, lyophilization, gel purification, and the like.

The purified dipeptide L-Glu-L-Trp, comprises a white powder (if lyophilized; otherwise, it is crystalline), soluble in water, DMF; insoluble in chloroform and ether. [alpha 22 D +12.6; C

H

2 0. Rf 0.65 (butanol: acetic acid: water UV (275 5nm, max). NMR (500MHz): 0.001mol/l of the peptide solution, Trp (3.17; 3.37; 4.57; 7.16; 7.24; 7.71; 7.49); Glu (1.90; 1.96; 2.21; 3.72).

The active dipeptide ingredient of the pharmaceutical preparations according to the present invention may be used as a free peptide or in the form of a water soluble pharmaceutically acceptable salt, such as a sodium, potassium, ammonium or zinc salt. It will be understood that the dipeptide may be administered with other active ingredients which independently WO 92/17191 PCr/US92/02440 -6impart an activity to the composition, such as, antibiotics, interferon, anesthetics, and the like.

The most preferred formulation according to the present invention is a solution for intramuscular injection containing about .001 to .01% by weight (.0001-.001mg/kg body weight, or 10-100ig active ingredient per Iml solvent). The pharmaceutically acceptable vehicle for this injection form may be any pharmaceutically ar' -ptable solvent such as 0.9% aqueous sodium chloride, distilled water, Novocaine solution, Ringer's solution, glucose solution, and the like. The dipeptide containing compositions according to the present invention may be administered in a compatible pharmaceutical suitable for parenteral administration intravenous, subcutaneous, intramuscular). The preparations may be subjected to conventional pharmaceutical operations, such as sterilization, and may contain adjuvants, such as preservatives, stabilizers, wetting agents and the like.

The pharmaceutical preparations according to the present invention demonstrate a high effectiveness in the treatment of immunodepressed and immunodeficient states for the preventing and treatment of opportunistic infections in those states.

Also included within the scope of the present invention are the pharmaceutically acceptable salts of the dipeptide, such as sodium or potassium or strong organic bases, such as guanidine.

The dipeptide containing compositions according to the present invention have activity in the restoration and stimulation of the immune functions.

Thus they are useful in the treatment of t H I .nl-lll~ ^-fiTT U- LaoUIIC WO 92/17191 PCT/US92/02440 -7opportunistic infections of an immunodepressed subject in an immunopotentiating effective amount as described above.

The dipeptide compositions according to the present invention may also be used in veterinary practice as an immunomodulatory agent for prophylaxsis and treatment of hypotrophy in farming animals, fur bearing animals and poultry.

Among the opportunistic infections which may be treated utilizing the compositions according to the present invention are: respiratory diseases, influenza, AIDS, burns, wounds, other open sores, rashes (due to allergic reactions), sun exposure, local trauma (with an ointment), eczemas, psoriasis, and the like. Furthermore, the compositions according to the present invention may be utilized to assist healing in immunodepressed or immunodeficient states, such as for the healing of bone fractures, lesions, gingival diseases, gynecological infections, infralymphatic infections, and the like. The compositions may also be used to enhance the immunodeficient state to increase susceptibility to microbial antibiotics and to enhance the patient's responsive reaction to other types of therapies.

The compositions according to the present invention also may be utilized to enhance metabolic processes; to enhance production of blood insulin; for treatment of irradiated cancer patients, as well as for veterinary uses.

Veterinary uses include treatment of infections and inflammatory diseases, nitrate toxicoses, dyspepsia.

Prophylactic uses in animals include administration to young animals to sustain weight gain and growth i WO 92/17191 PCT/US92/02440 -8and to increase immunity to acute and chronic viral infections.

A particularly important use is for the treatment of Marek's disease in poultry. Administration intramuscularly or preferably by aerosol will be effective as evidenced by continued and sustained growth rates in chicks which receive treatment prior to inoculation with active virus associated with Marek's disease. This treatment may be performed in conjunction with administration of other vaccines known to be effective for this or other diseases.

The following examples are provided to further elucidate the invention, but are not intended to restrict the invention in scope or spirit in any way.

EXAMPLE 1 About 262 adult patients were treated over a period of 2 months on a daily basis with intramuscular injections of solutions containing 100 gsg of Thymogen. These patients were treated for 2 months immediately succeeding exposure to radiption caused from the Chernobyl nuclear accident. s a control, about 18 people exposed to radiation were tested for various blood parameters to establish a baseline.

The results are shown below.

i WO 92/17191 WO 9217191PCI'!US92/02440 -9- THE EFFICIENCY OF RADIATION IMMUNODEFICIENCY CORRECTION TWO MONTHS AFTER IRRADIATION (X±m) Indices Examinated groups Healthy Irradiated (control) Prior to After THYMOGEN therapy therapy Leukocytes, abs 5.6±0.8 3.5±0.4 5.0±1.2 Lymphocytes, 1.98±0.16 0.80±0.24 1.9±0.4 2 abs CD2-DR+, 35.8±0.9 21 ±4 30.0±1.2 CD2-DR+, abs 0.59±0.04 0.16±0.04 0.55±0.06 CD21 49.3±1.5 32 7 48.7±1.8 CD2, abs 0.98±0.09 0.55±0.08 1.13±0.08 E-RFC, 30.2±1.6 22.9±1.9* 27.4±2.4 LMI with ConA, 65.0±t2.1 120 ±17 90 CD19, 22.0±1.7 32 ±3 27 4 LCD19, abs 0.46±0.03 0.26±0.06* 0.51±0.10* 1gm, g/1 1.1 0.4 0.87±0.07 1.00±0.10 IgG, g/1 11.1±0.9 10.2±2.0 10.0±1.0 IgA, g/l 1.70±0.10 1.5±0.4 1.49±0.19 statistically significant (P<0.05) vs. the indices in healthy people; statistically significant (P<0.05) vs. the data obtained prior to immunocorrection with

THYMOGEN;

abs cell concentration presented as 109/l LMI leucocyte migration inhibition; RFC rosette-forming cells.

SUBSTITUTE SHEET WO 92/1 7191 PCT/ US 9 2/02440 EXAMPLE 2 The group of patients described in Example 1 were further treated for a period of 36 months and tested again subsequent to the first stage of therapy (after 4 months) and afte.- the second stage of therapy (6 months). The blood parameters are shown below. As canz seen most of the blood parameters were elevated after both the first and second stages of therapy.

.0 THE PROLONGED THYMOGEN THERAPY TRIALS RESULTS IN IRRADIATED PATIENTS (Xim) Inidices Initerims of examnination prior to after the 1st after the therapy stage of 2nd state THYMOGEN use of THYMOGEN use Leucocytes, abs 3.5±0.5 4.7±0.2 5.5±0.3 Lyriphocytes, abs 1.0±0.5 1.5±0.4 1.9±0.5' CD2-DR+, 12.8±2.6 22.3±0.5 29 3 CD2-DR+, abs 0.13±0.04 0.34±0.05* 0.56±0.08 CD3, 14 3 35 ±4 46± 3* 0 CD3, abs 0.26±0.05 0.49±0.06 0.89:t0.11' CD4, 1.1 19.5±1.7 24.1±1.5 CD4, abs 0.07±0.01 0.28±0.03 0.45±0.04' CD8, 17 ±3 15.4±2.3 22.3±2.2 CD8, abs 0.16±0.04 0.23±0.03 0.40±0.05' CD19, 12.2±1.9 15.0±2.8 21.1±2.1' C1,aba 0.14±0.04 0 W1±.06 0.9006] statistically significant (P<0.05) in comparison with the indices prior to therapy; abs -cell concentration presented as 109 /1.

SUBSTITUTE

SHEET

WO 92/17191 PCT/US92/02440 -11- EXAMPLE 3 The patients described in Example 1 were tested for blood parameters the first few days after exposure to the radiation of the Chernobyl accident. It could be seen from the table below that response to the treatment was observed even after a few weeks? of treatment.

SUBSTITUTE

SHEET

WO 92/17191 WO 9217191PCF/US92/02440 -12- THYMOGEN INFLUENCE ON IMMUNE STATUS IN EARLY TERMS AFTER IRRADIATION AFFECTION (X~tm) Indices Examinated groups Healthy Irradiated (control) Prior to After therapyTHYMOGEN Leukcyte, ab 7-;0.3 heray *therapy* Lekoyes as 5.103 3.810.3 6.4±0.8 Lymphocytes, abs 1.91*0.12 1.15±0.14 2.27±0.16" CD2-DR+, 30.8*1.1 17.6±2.0 31 3 CD2-DR+, abs 0.59±0.04 0 .20±0.03* 0.69±0.08" CD2, 50.6±1.6 47 4 50.9 2.4 CD2, abs 0.9a±0.09 0.55*0.08 1.13±0.07 E-RFC, 29.7*2.5 29.8*2.6 23.4±2.6 LMI with ConA, 66*±4 98*±9* 60*±7' CD19, 22.8*2.2 27.0±2.8 30.5±1.9* CD19, abs 0.47±0.03 0.30±0.05* 0.68±0.04" 1gm, g/1 1.1±0.4 0.51*0.08* 0.58*0.10' IgG, g/l 10.1±0.9 8.6±1.3 9.2 *0.7 IgA, g/l 1.71*0.16 2.07±0.20 1.11±0.09,* C3, g/l 0.57*0.03 0.74*0.07 0.68±0.04 statistically significant (P<0.05) in comparison with the indices in healthy people statistically significant (P<0.05) in comparison with the data obtained prior to THYMOGEN use; LMI leukocyte migration inhibition; abs cells concentration presented as 109 /1.

WO 92/17191 PCT/US92/02440 -13- EXAMPLE 4 A number of (36) breast cancer patients were treated with the Thymogen by injection of daily dosages of 100 ig The patients had been previously treated with radiation therapy (single doses 2 grad; total dose 45-50 grad). It can be seen from the table below the treatments restore their blood parameter levels.

SUBSTITUTE

SHEET

WO 92/17191 WO 9217191PCT/US92/02440 -14- IMMUNITY AND NON-SPECIFIC RESISTANCE INDICES IN BREAST CANCER PATIENTS TREATED WITH THYMOGEN AFTER RADIOTHERAPY (X±m) Indices Prior to After After radiotherapy radiotherapy THYMOGEN use Lymphocytes 1.61±0.18 0.79±0.09 1.72±0.21 T-Wrphocytes 0.83±0.07+ 0.32±0.03 0.92±0.12 (xlO "Active" T- 0.49±0.06 0.19±0.03 0.52±0.07 lymphocytes (XlO 1) T-helpers (OKT4 0.30±0.03 0.12±0.01 0.39±0.04 T-BUP reSsors 0.28±0.04 0.16±0.02 0.21±0.03 (OKT8_)__(xlO_9/1) OKT4 +/OKT8 +1.07±0.09 0.75±0.06 1.86±0.17 DSH a to tuberculin 7.3±0.4 2.6±0.2* 8.7±0.6* 1241b with ConA 68 4 96 7 71 SI c to THYMOGEN 1.23±0.15 1.19±0.13 1.27±0.14* B-lymphocyte (Ig 0.15±0.02 0.11±0.01l 0.17±0.02 Phagocytic index 4.3±0.3 2.06±0.18 3.7±0.2 Cation proteins 1.58±0.09 1.36±0.08 1.49±0.12 C 3 -cmlm 0.75±0.05 0.66±0.04 0.68±0.04 statistically significant (P<0.05) vs. the analogous index before radiotherapy; statistically significant (P<0.05) vs. the analogous index after radiotherapy; a Delayed-Skin Hypersensitivity; b Leukocyte Migration Inhibition; c Sensitivity Index.

SUBSTITUTE SHEET I i WO 92/17191 PCT/US92/02440 EXAMPLE On peripheral blood of human volunteers in vitro.

Cell cultures were incubated and treated. As can be seen from the table below, after 24 hrs. incubation at concentrations of 1 g/ml and 100 Ag/ml, there was statistically no mutagenic effect in these cultures.

SUBSTITUTE

SHEET

cl i i- II~ WO 92/17191 PCT/US92/02440 -16- THE CALCULATION OF CHROMOSOME STRUCTURAL DAMAGES IN HUMAN PERIPHERAL BLOOD LYMPHOCYTES Dose of Number of Metaphases Index of Level of the analysed with reliability mutagenic medicine metaphases chromosome effect structural (numbers) aberrations Control 1000 15 1.5 THYMOGEN 1000 15 1.5 >0.05 0 1 pg/ml THYMOGEN 1000 16 1.6 >0.05 0 100 Pg/ml EXAMPLE 6 About 263 patients were treated with doses of 100 p.g of Thymogen introduced intramuscularly on a daily basis over a period of 3 years after exposure to the radiation at the Chernobyl accident. Blood parameters after 3 years of such treatment were restored to the statistical norm prior to their exposure to the radiation.

SUBSTITUTE SHEET WO 92/17191 WO 9217191PCT/US92/02440 -17- Indices Stati.sti.cal Results of victims examination norma Prior to After therapy theapy THYMOGEN Conven- Leucocytes, 5.210.2 5.8±0.3 5.6±0.4 5.5*1.0 abs Lymnphocytes, 1.96±0.06 2.0±0.3 2.1±0.3 1.8±0.23 abs CD2-DR+, 30.8±1.1 15 3 32 3 18.4±2.5 CD2-DR+, abs 0.59±0.04 0.30±0.0 0.66±0.1 0.34±0.11 6 0 CD3, 55.6±1.9 67.7±2.7 59.2±2.1 61 3 CD3, abs 1.09±0.08 1.33±0.0 1.21±0.1 1.12±0.18 CD4, 35.3±2.7 36.7±2.6 36.2±1.7 38 3 CD4, abs 0.69±0.05 0.72±0.0 0.74±0.0 0.70±0.05 8 CD8, 21.3~±0.9 29.710.9 23.2±2.1 25.0±2.7* CD8, abs 0.41±0.03 0.56±0.0 0.48±0.0 0.46±0.06 2 7 T4/T8 1.64±0.12 1.24±0.1 1.58±0.0 1.52±0.13 0 4 LMI, 59.7±1.7 140 75 6* 107 B-Ig+, 13.8±1.2 10.7±0.3 11.0±0.3 11.2±0.7 B-Ig+, abs 0.29±0.02 0.21±0.0 0.23±0.0 0.20±0.05 1 4 B-IgM+, 6.4±0.7 3.0±0.3* 4.1±0.6* 4.4±0.3 B-IgM+, abs 0.12±0.01 0.062±0. 0.12±0.0 0.00±0.004 002 03 B-IgG-, 4.1±0.5 4.7±0.9 4.8±0.5 4.6±0.5 B-IgG+, abs 0.078±0.008 0.059±0. 0.09±0.0 0.08±0.006 003 07 B-IgA+, 2.2±0.2 2.3±0.3 1.9±0.3 11.98±0.09 U11TIJESHEET WO 92/17191 WO 9217191PCT/US92/02440 -18- I I Indices norma Statitical Reaults of victims examination Prior to therapy After therapy

THYMOGEN

tional B-IgA+, abs 0.041±0.004 0.048±0. 0.04±0.0 0.04±0.002 006 02 1gM, g/l 1.15±0.06 0.53i0.0 1.06:t0.0 1.03±0.13 9 6 IgG, g/1 11.5±0.5 13.2±1.1 10.9±1.3 11.3±1.2 IgA, g/l 1.90±0.08 0.82±0.2 1.2±0.4* 1.1±0.3 -statistically significant (P<0.05) vs. the indices in healthy persons; statistically significant (P<0.05) vs. the data obtained prior to therapy; abs cell concentration presented as 10 9l LMI leucocyte migration inhibition.

SUBSTITUTE SHEET WO 92/17191 PCT/US92/02440 -19- EXAMPLE 7 The patients described above in Example 6 were tested for blood parameters after 6 months of treatment immediately following exposure to the radiation caused by the Chernobyl accident. The results below show that after 6 months those treated with Thymogen showed improvement over those patients who were not treated.

SLISSTITUTE SHEET WO 92/17191 PCr/US92/02440 THYMOGEN USE EFFICIENCY IN ACUTE RADIATION SICKNESS (6 MONTHS AFTER ACCIDENCE) Indices Healthy Pati~ents (suffered in people accidence) (ortatiti Prior to After therapy norma) therapy Without With THYHOGEN THYMO-

GEN

Lymphocytes, 33.9±1.2 32.9±2.4 29.2i2.0 3 0.0±1 .8 Lymphocytes, 1.96±0.06 1.49±0.14 1.39±0.13 1.52±0 abs .12* CD2, 53.6±1.9 38.7±2.7 32 3 49± 3 CD2, abs 1.05±0.05 0.56±0.04 0.44±0.04 0.75±0 CD2-DR-, 30.8±1.1 18.9±1.6 19.7±1.2 20.8±1 .6, CD2-DR+, abs 0.59t0.04 0.30±0.25 0.28±0.02 0.31±0 .02 CD3, 55.6±1.9 39.0±2.4* 37 5 53.4±1 .8 CD3, abs 1.09±0.08 0.58±0.04 0.51±0.03 0.82±0 .04 CD4, 35.3±2.7 20.3±1.3* 18.9±-1.3* 32.6±1 CD4, abs 0.69±0.05 0.30±0.03 0.26±0.03 0.50±0 .04 CDB, 21.3±0.9 19.5±1.5 17.5±1.6 21.2±1 .8 CD8, abs 0.41±0.03 0.29±0.03 0.24±0.03 0.32±0 T4/T8 1.64±0.12 1.04±0.04* 1.08±0.10 1.54±0 LMI 59.7±1.7 106 6* 107 72.7±4 SUBSTITUTE SHEET WO 92/17191 WO 9217191PCTr/US92/02440 -21indices Healthy Patients (suffered in people accidence) (statistic norma) Prior to After therapy therapy Without With THYMOGEN THYMO-

GEN

CD19, %25.00±0.12 18.2±2.1 23 ±3 26.7±2 CD19, abs 0.49±0.04 0.27±0.03 0.31±0.05 0.41±0 B-Ig+, %13.8±1.2 15.8±1.3 16.2±1.7 19.0±1 B-Ig+, abs 0.29±0.02 0.23±0.03 0.23±0.04 0.29±0 B-IgM+, 6.4±0.7 6.3±0.8 5.4±0.5 8.8±0.

7 B-IgM+, abs 0.12±0.01 0.09±0.01 0.08±0.01 0.13±0 .02 B-IgG+, 4.1±0.5 7.8±0.9 7.1±0.8* 6.4±0.

B-IgG+, abs 0.082±0.00 0.098±0.00 0.104±0.0 0.100± 8 7 08 0.007 B-IgA+, 2.20±0.20 1.80±0.15 1.70±0.20 1.8±0.

3 B-IgA+, abs 0.038±0.00 0.033±0.00 0.024±0.0 0.030± 4 4 03 0.002 1gM, g/l 1.15±0.06 1.14±0.08 1.20±0.07 1.07±0 .09 IgG, g/l 11.5±0.5 11.9±1.0 11.7±0.9 10.9±1 .1 IgA, g/l 1.9±1.0 1. 6±0. 8 1608 180 statistically significant (P<0.05) vs.

indices in healthy people; statistically significant (P<0.05) vs.

data obtained before immunocorrection; abs cell concentration presented as LMI leucocyte migration inhibition.

the the SUBSTITUTE

SHEET

n ft I WO 92/17191 PCP'IUS92/02440 -22- EXAMPLE 8 A group of 452 persons were treated with daily dosages of 100 Ag of Thymogen administered intramuscularly over a period of 5-10 days and compared with a random group (250 persons) (not similarly treated) as a control. The cases of respiratory diseases and influenza were recorded for both groups. As can be seen from the table below, the untreated group had a greater occurrence of the diseases and sicknesses, hospitalization or disablement than the group treated with Thymogen.

r"CI' SUBSTITUTE

SHEET

i;

I",

WO 92/17191 PCr/US92/02440 -23- THYMOGEN CLINICO-EPIDEMIOLOGICAL PREVENTIVE EFFICIENCY IN ARD AND FLU Indi.ces Group of Index of observation efficiency THYMOGEN Control sickness rate per 100 9.8 30.4 3.1 persons/month_____ Pneumonia rate/100 0.20 0,50 persons/month___________ Need in hospitalization, 30.6 44.9 1.7 Average term of 6.2 8.8 1.4 hospitalization, days Rate of lingo-ring and 3.9 13.8 complicated cases, The same index in in- 9.8 26.2 2.7 patients, Number of temporary 4.1 7.0 1.7 disablement cases/100 persons/month___________ Number of temporary 26.5 57.6 2.2 disablement days/100 persons./monthII THE DYNAMICS OF ARD AND INFLUENZA RATE IN GROUPS OF OBSERVATION BY MONTHS FROM THE BEGINNING OF INVESTIGATION Ind ices Groups lot 2nd 3rd 4th month month month month sickness rate/100 Tq1YMOGEN 9.6 11.3 9.4 11.0 persons/month Coatrol 28.6 33.4 28.7 30.6 Need in THYMOGEN 27.0 27.1 28.3 28.3 hospitalization, Control 41.2 50.8 48.8 39.0 Avera~ge term of THYMOGEN 6.2 6.2 6;2 hospitalization, Control 9.3 8.4 10.2 11.0 day~s Number of temporary THYMOGEN 3.8 7.3 3.4 3.6 disablement Control 6.9 10.2 7.5 5.6 cases/ 100 persons/month SUBSTITUI 4E SHEET WO 92/17191 PCT/US92/02440 -24- Indices Groups lat 2nd 3rd 4th month month month month Number of temporary THYMOGEN .24.0 48,6 14..3 26..3 disablement days/100 Control 47.9 82.0 51.4 42.8 persons/month EXAMPLE 9 In separate studies, a total of 21 AIDS infected individuals have been studied, including full-blown syndrome, prodromal, and pre-AIDS afflicted individuals who were treated with Thympentin.

Thympentin and TPI are thymic gland peptide extracts previously well characterized. Comparative studies between TPI and 7hymogen reveal that Thymogen is a far more effective cell mediator, restoring normal immunologic indices, including T-cell functional activity and T4/T8 ratios. Method of Administration: SteUile saline containing the sodium salt of the medication is administered either IM, infralymphatically, or intranasally each day for 5 days consecutively every 30 days.

Immunosupressed individuals who have su'stair-,,d radiation injuries were treated with Thymogen with excellent restoration of immunological indices and models for acquired immune deficiency syndrome.

Thymogen may thus benefit AIDS infected individuals by eduing the need to use other medications with toxic side effects, and sustain and or support the individuals by reducing the needs to use other medications with toxic side effects, and sustain and or support the individuals immune indices resulting in a reduction of opportunistic infections.

SUBSTITUTE

SHEET

l l WO 92/17191 PCT/US92/02440 EXAMPLE A total of 159 patients were treated with pyoderma, including furunculitis, cellulitis, and folliculitis, with a control group consisting of 25 patients who were not treated with thymogen. Medications were administered either IM or intranasally for consecutive days. Immunological indices were normalized with disappearance of skin manifestations and relapses were prevented after treatment with Thymogen. Clinical improvement correlated with immunological indices correction. Administration IM, intranasally, or topically as a sterile saline solution of medication for a period of 5 to 10 days at a concentration of 1 Ag/kg body weight.

EXAMPLE 11 A nuntber of patients within the group of 159 patients afflicted with furunculitis, pyoderma, cellulitis, and folliculitis were afflicted with acne vulgaris and acne. The immunological indices corrected and normalized rapidly within the group therapy. The clinical outcome correlated with the correction of immunological indices, and relapses were controlled.

EXAMPLE 12 of 30 patients were treated with psoriasis U patients were used as controls and were untreated with thymogen. All patients had at a least year history of no unsuccessful treatment. The administration of 100 gg IM or intranasally for a period of 10 days resulted in the improvement in 7% of the patients, significant improvement in 60% of patients, and total recovery in 33% of the patients.

SUBSTITUTE

SHEET

I_ ^I_ WO 92/17191 PCT/US92/02440 -26- EXAMPLE 13 A total of 46 female patients with the various disorders (pelvic inflammatory diseases, cervicitis, vaginitis and various tubo-ovarian and adnexal abscesses) were treated and 50 patients were used as controls. Thymogen was applied IM, intranasally at 100 Mg 5 consecutive days or 50 gg intralymphatically for 5 consecutive days in conjunction with conventional therapy. The clinical effect of Thymogen expressed the arresting of pain syndrome, the control of body temperature, e.g. reduction of fever, the decrease of duration of conventional treatment. The normalization of immune status correlated with clinical improvements.

EXAMPLE 14 Patients treated with Thymogen either topically, IM, or intranasally experienced marked reduction of recurrence of herpetic lesions, with substantial reduction in the period between outbreaks. In one study, individuals who experienced 7-10 outbreaks per year experienced less than one outbreak per year after treatment with Thymogen in combination with interferon.

EXAMPLE A total of 37 patients with Herpes Zoster were treated with Thymogen in combination with conventional interferon treatment and 25 control patients with interferon alone. Administration single daily IM or intranasal 100 jg during a period of 10 days resulted in accelerated regression of foci of herpetic infection. There was noted prevention of relapses, and healing occurred on the average SUBSTITUTE

SHEET

WO 92/17191 PCr/US92/02440 -27earlier than control groups. Immunological indices correlated with clinical outcome.

EXAMPLE 16 Patients were treated for gingival disease by subcutaneous administration of thymogen in the area of the gingiva. The treatment resulted in the arresting of gingival disease. Approximately patients were studied with disease and treated and an equal number were treated conventionally without Thymogen for control purposes. Administration of 100 ug Im, Subcutaneously, or by electrophoresis (whereby a small voltage charge to the gums results in a rapid transfer of medication through the gum epithelium) resulted in the arresting of bleeding, more rapid restoration of inflammatory processes, and the decrease of purulent discharge. The treatment resulted in fewer recurrences and prolongation of normal gums. It was also noted that normalization of immunologic indices was achieved with normal coagulation.

EXAMPLE 17 The treatment with toothpaste containing Thymogen will result in a reduction of dental caries.

EXAMPLE 18 25 A total of 46 patients with periapical granulomas and 28 patients with the same disease not treated with Thymogen were used for controls. Instillation of 100 Ag of Thymogen into the foramen at the base of the tooth, or in the composition of the filling paste during 3 days resulted in the accelerated arrestation of the inflammatory process, reduction in pain, and SUBSTITUTE

SHEET

WO 92/17191 PCT/US92/02440 -28increased stability of the underlying dental structures as evidenced by x-ray studies.

EXAMPLE 19 The use of dental toothpaste containing Thymogen will result in the reduction of gingival disease and reduction in dental caries.

EXAMPLE The use of Thymogen 100 pg IM, intranasally, or intralymphatically controls the advance of lymphangitis.

EXAMPLE 21 A total of 186 patients with acute respiratory disease, including upper airway diseases, such as colds, were treated with Thymogen and 87 patients who were not treated with Thymogen were used as controls.

Administration IM or intranasally 100 Ag 3 7 days resulted in a milder course of the viral infection.

There was noted a decrease in the specific signs of upper respiratory infections such rhinorrhea, sore throat, fever, muscle aches, headaches, and ear pain.

Secondary infectious complications were diminished, and the duration of the treatment was also diminished.

EXAMPLE 22 A total of 51 patients were treated with Thymogen with 24 patient controls, administration IM, intranasally, and installation into sinuses with 1 pg/kg dose during a period of 3 10 days resulted in normalization of nasal breathing, the disappearance SUBSTITUTE

SHEET

I

1~1 WO 92/17191 PCT/US92/2440 -29of nasal mucous swelling, the arresting of exudates from affect sinuses, and improved general condition and immune status. The decrease of treatment duration up to 1.7 times compared to controls.

EXAMPLE 23 Thymogen IM or intranasal accompanying conventional therapy (antibiotics) results in accelerated healing of chronic and acute ear infections.

EXAMPLE 24 A total of 41 patients with various eye problems as described and 36 patients in control studies were treated by conventional methods with the first group receiving Thymogen in addition to the conventional treatment. Administration of Thymogen intra ocularly at 18 Mg for 5 consecutive days, or as installation into conjunctival cavity as drops bid for 5 days resulted in more rapid arresting of the inflammatory process and the increase in visual acuity, and the decrease of duration of treatment.

EXAMPLE A total of 156 patients treated with Thymogen and 82 patients in the control study, were administered, medication IM or intranasally 100 pg 5 10 days resulting in accelerated reduction in symptom complexes including joint pain, muscle aches, fevers, chills, and upper respiratory symptoms.

SUBSTITUTE SHEET WO 92/17191 PCT/US92/02440 EXAMPLE 26 A total of 263 patients and 18 control patients sustained exposure to radiation injury. Thymogen was administered IM and/or intranasally 100 Ag for days. A repeated course may be prescribed on the basis of immunological indices, and averages every 4 to 6 months. The results of the treatment are restoration of normal or near normal immune indices with functional activity in the majority of all cases studied. There was an arresting of esthenic syndrome, and an arresting of the somatic pathological exacerbations and reduction of opportunistic infections.

EXAMPLE 27 Thymogen administration IM or intranasally results in the improved immune parameters, functionals activity of lymphocytes and neutrophils, and reduction of post-operative complications and infections associated with bone-marrow compromise, such as, that caused from transplant or radiation exposure.

EXAMPLE 28 A total of 29 patients afflicted with various allergies as described and 17 patients in the control group were treated with Thymogen in dose 1 pg/kg IM or intranasally for 5 7 days resulted in disappearance of allergic reactions.

EXAMPLE 29 A total of 76 patients with 72 patients in control exposed to massive hemotransfusions during postoperative period were treated with Thymogen.

SUBSTITUTE SHEET WO 92/17191 PCT/US92/02440 -31- Thymogen was administered starting from 4-6 day of post-operative period single daily IM or intranasally in does 100 Ag for 5 days. None of studies patients showed clinical manifestation of alloblood rejection while in 17. of control patients the adverse hemotranfusional reactions were observed.

EXAMPLE Thymogen was applied in 76 patients treated with antibiotics for various indications who had unfavorable allergological history. Control group comprised 43 patients. Thymogen was administered IM or intranasally single daily at 100 Mg for 5-10 days, In the majority of case the use of Thymogen prevented the arising of allergic reactions or promoted the less severe course of them while in the control group in 70% of patients the pronounced signs of drug intolerance was marked.

EXAMPLE 31 Thymogen was administered to 17 patients subjected to skin grafting. The control group comprised 27 patients. Thymogen was administered IM or intranasally single daily at 50-100 Mg for 5 days.

In all the patients the use of Thymogen prevented the arising of infections complications and graft rejection. In control group the manifestations of rejection were determined in 8 patients.

EXAMPLE 32 Thymogen was administered to 52 patients suffered from chronic skin diseases caused by antibioticresistant staphylococci. 42 patients with the same pathology bir not ated with the immunomodulator SUBSTITUTE SHEET 1_11~_ WO 92/17191 PC/US92/02440 -32were the control group. Thymogen was administered IM to 27 patients single daily at 100 gg for 5 days and intranasally to 25 patients in the same daily and total dose. The differences between these two methods of application were not noticed. In all the patients with signs of secondary T-immunodeficiency the staphylocci antibiotic-sensitivity to one, few or all antibiotics has been increased sharply (more than 100-fold) what permitted further to choose for each patient the antibiotic with exclusively high activity against pathogen. As a whole, within the examined group of patients the reliable decease of MIC of all studied antibiotics has been marked. The proposed treatment regiment permitted to obtain the complete recovery in 27 patients, significant improvement in 8 patients and moderate improvement in 1 patient.

EXAMPLE 33 Thymogen was used in 37 patients with wounds of various origin, type and localization, the control group comprised 24 patients. Thymogen was administered IM or topically single daily at 100 ug for 10 days. The use of Thymogen speeded up (when compared to the control group) significantly wound healing, reduced therapy duration and prevented the development of infectious complications.

EXAMPLE 34 Administration of Thymogen either intranasally or IM accelerates wound healing, resulting in statistically fewer infections and reduced escar.

SUBSTITUTE

SHEET

i WO 92/17191 PCT/US92/02440 -33- EXAMPLE Thymogen was applied to 44 patients with bone fractures various origin, type and localization.

The control group comprised 28 patients. Thymogen was administered intramuscularly or intranasally single daily at 100 up for 10 days. The use of Thymogen accelerated essentially (in comparison with the control group) the consolidation of fractures, prevented the development of infectious complications, reduced pain syndrome and treatment duration.

EXAMPLE 36 Thymogen was prescribed to 176 patients with chronic osteomyelitis of various ethiology and localization.

The control group comprised 88 patients. Thymogen was administered IM or intranasally single daily at 100 ug for 10 days. The use of thymogen rendered the pronounced positive influence on clinical course what expressed in significant decrease of intoxication syndrome and pain syndrome, disappearance of purulent inflammatory manifestations, speeding up of wound healing, reduction of destruction areas, prevention of relapses.

EXAMPLE 37 A total of 23 patients with cutaneous burns were treated with Thymogen either IM or intranasally with 14 patients for control treated conventionally.

Accelerated wound healing, diminished frequency of infections, and less escar was noted in those individuals treated with Thymogen.

ci SUBSTITUTE SHEET 1 t WO 92/17191 PCT/US92/0244C -34- EXAMPLE 38 A total of 17 patients with frostbite to the extremities where treated with Thymogen either IM or intranasally with 11 patient controls. The rapid healing and restoration of tissue integrity was observed.

EXAMPLE 39 Thymogen administration either IM or intranasally results in less deformity and scarring evidenced by experience in healing fractures, burns, military accidents, and other injuries to the extremities.

EXAMPLE Experimental data supports the finding that Thymogen administered IM, intranasally, or ocular installation results in restoration and regeneration of corneal epithelium with fewer infections and complications related to escar.

EXAMPLE 41 A total of 246 patients with various forms of cancer, and 158 controls after radiation and chemo-therapy, where Thymogen was administered in single 100 ug daily dose for 10 days experienced normalization of immunological indices, the prevention of postoperative infections, the prevention of upper r 25 respiratory infections, and prevention of exacerbations of various secondary complications such as gastritis, cholecystitis, etc. If it was determined necessary based on immunological indices, the treatment regimen was repeated in 4-6 months.

SUBSTITUTE

SHEET

WO 92/17191 PCr/US92/02440 EXAMPLE 42 Patients treated with Thymogen simultaneously during the administration of chemotherapy experienced fewer complications and side effects related to chemotherapy including diminished frequency and intensity of ulcerative lesions, nausea, and other related problems of chemotherapy administration.

EXAMPLE 43 Experimental models support the fact that administration of Thymogen prophylactically results in diminished frequency of spontaneous tumorogenesis.

EXAMPLE 44 Thymogen was applied to 268 persons in combination with the anti-flu vaccination. The control group comprised 197 persons. the vaccination was delivered by air pressure. ,The Thymogen dose was 50 ug delivered in a single dose for 3 consecutive days.

After Thymogen use, it was observed the significant decrease of sickness rate for a period o 12 months compared to controls who received flu-vaccination without Thymogen. In the event of flu, the course of the infection was noted to be less severe and the recovery more rapid when compared to controls.

EXAMPLE Thymogen was applied in 97 pregnant women with Toxenia of first and second half of pregnancy. The control group comprised 54 patients. Thymogen was administered iM and intranasally at 100 ug daily for 10 days. Under the influence of Thymogen, it was observed that the BP normalized, and peripheral edema SUBSTITUTE SHEET WO 92/17191 'PCT/US92/02440 -36was reduced with normalization of the blood chemistry profile, and the restoration of initially altered immunologic indices.

EXAMPLE 46 Thymogen was administered to 34 pregnant women with 27 pregnant women for control. The route of administration is IM or intranasally 100 ug daily for iC days. Signs of clinical improvement were resolution of weakness, dizziness, and increased appetite, and the normalization of the immunological and hematological indices. It was also noted that there was a decrease in fetal hypoxia.

EXAMPLE 47 A total of 19 post-term women and 48 women post-term in the control study were treated. The administration of 100 ug of Thymogen IM )r intranasally over 3 5 days resulted in the effacement of the cervix with thinning at the cervix and the decent of the fetus, with subsequent spontaneous normal delivery.

EXAMPLE 48 A total of 27 patients with pyelonephritis and 19 control patients with pyelonephritis were treated with the administration of Thymogen single daily dose of 100 ug for 5 10 consecutive days in combination with conventional therapy resulted in reduction of fever, the normalization of urina analysis, and the improvement and resolution of the infection. The normal course of delivery in those women treated with Thymogen was without complications.

SUBSTITUTE

SHEET

i WO 92/17191 PCT/US92/02440 -37- EXAMPLE 49 A total of 45 patients with leprosy (Hansen's disease) and 27 infected individuals were tr.. .t i.

Thymogen was administered IM or intranasally i:i single daily doses of 100 ug for 5 days consecutively in additional to conventional therapy. The patients studied had previous documented resistance to treatment by conventional methods. Thymogen administration resulted in resolution of the lesions and prevented relapses, and promoted more rapid healing of specific ulcers. The immunologic indices were normalized.

EXAMPLE Thymogen was administered to 84 young sportsmen. The control group consisted of 44 persons. The Thymogen was administered intranasally single dose 1 ug/kg during 3 days. The use of Thymogen resulted in the reduction of upper respiratory infections and rates of illness 4 fold. In the event of infection, it was noted that the infections was less severe without complications, and the clinical improvement was accompanied by the normalization of immunological indices.

EXAMPLE 51 A total of 33 patients were studied with patients who had relapsing forms of tropical malaria, moderate to severe, and severe cases with 21 patients in the control group. Thynogen was administered at 100 ug single daily doses IM or intranasally for 5 days. the results of such treatment was reduction of hepatolineal syndrome, the normalization of SUBSTITUTE

SHEET

WO 92/17191 PCT/US92/02440 -38hematological and immunological indices, reduction of fever, and prevention of relapses.

EXAMPLE 52 Thymogen was applied in 27 persons with the goal to increase the resistance to excessive solar radiation, in the conditions of hot rarine climates. The control group comprised 24 persons. The administration was intranasally 100 ug for 3 days.

The use of Thymogen prevented the occurrence of upper respiratcry infections in the treated group relativ, to the control group. There was also noted suppression of their immunologic indices.

EXAMPLE 53 Thymogen was applied in 21 patients with hemorrhagic Dengue Fever, and 28 patienits served as controls.

Thyrmogen was administered IM single daily doses of 100 ug for 5 conacutive days in conjunction with conventional therapy. The results of treatment were reduction in fever, reduction of toxic symptoms, significant decrease in hepato-lineal syndrome. It was also noted that the muscular and bone pain eYperienced typically was reduced, and the immunological indices were normalized.

EXAMPLE 54 A total of 48 patients infected and 34 infected controls were examined and treated with administration of Thymogen 100 ug IM or intranasally for 5 10 days resulting in normalization of fever, the reduction of toxic symptoms, and the resolution of icterus (jaundice). The hematological and immunological indices were normalized.

SUBSTITUTE SHEET I i _I WO 92/17191 PCY/US92/02440 -39- EXAMPLE A total of 36 patients infected and 24 patients infected were controls. Administration of Thymogen in 100 ug IM or intranasally for 5 10 days resulted in the reduction of fever, more rapid reduction of toxic symptoms, and the restoration of immu.)logir indices.

EXAMPLE 56 A total of 37 patients infected with pulmonary TB and 26 patients infected as controls were studied and treated. Thymogen was administered at 50 to 100 ug every other day during 5 doses total in combination with convention therapy. The results of the treatment 2 months after the course of Thymogen revealed the disappearance of toxic symptoms, the reabsorption of infiltrates, and resolution of pulmonary cavities. The disappearance of TB bacilli was noted in the sputum. The restoration of initially decreased immune indices was also noted.

EXAMPLE 57 A total of J7 patients, children and adults, with bronchial asthma and 28 similar patients as controls were studied. Thymogen was administered IM single daily doses 1 ug/kg for 5 10 days resulting in less severe clinical symptoms. The significant reduction in bronchial obstruction and laryngotracheitis was noted. The normalization of fever, and the reduction in duration of treatment was noted. In some of the patients it was possible to avoid steroids in the conventional commitment treatment course. In the following year observation there was noted a decrease in the incidence of bronchial asthma 4.2 fold. In SUBSTITUTE SHEET WO 92/17191 PCT/US92/02440 more than half of the patients the disappearance of drug and food allergy manifestations was noted.

EXAMPLE 58 A total 125 patients with 53 patients for control infected with Shigella dysentery were examined.

Thymogen was administered IM single doses of 100 ug for 10 consecutive days with resultant normalization of fever, the reduction of toxemia, and the normalization gastrointestinal disorders and symptoms. Bacterial shedding in the GI track was observed to cease, and the immunclogical indices were normalized.

EXAMPLE 59 A total of 12 patients who had been thymectomized were treated with Thymogen. Prior to therapy these individuals had experienced frequent serious infections including upper respiratory infections.

Thymogen was administered in a sir-?le dose 100 ug daily for 10 days and repeated ev y 4 6 months.

The normalization of immunologic id es was observed, and there was reduction fectious disorders including cutaneous infe 'ons and other chronic exacerbations.

EXAMPLE A total of 39 patients were studied with 27 patient controls. Thymogen was administered IM or intranasally at 100 up for 5 10 days to the study group of patients with the resulting reduction of fever, decrease in toxi< symptoms, the reduction of musculoskeletal pain, and tie reduction or SUBSTITUTE

SHEET

;1_1 WO 92/17191 PCT/US92/02440 -41disappearance of jaundice. Immunological indices were normalized.

EXAMPLE 61 The use of Thymogen as an ingredient or applicant with cosmetics provides for a less allergenic cosmetic with fewer allergic reactions.

EXAMPLE 62 To prevent and treat bronchopneumonia, hypotrophy or nitrate intoxication in cattle or swine, an intramuscular dose of 1 microgram/kg body weight of the dipeptide is administered: cycle of 4-6 months.

To prevent and treat viral diseases, Marek's disease or hypotrophy in poultry, a mist is delivered to the incubator habitat of the poultry in a concentration of dipeptide of about 200/microgram/cu. meter over 1- 3 day cycles.

SUBSTITUTE SHEET

Claims (8)

1. A method for treating psoriasis in a subject including administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to alleviate said psoriasis.

2. A method for assisting in the healing of local skin trauma of skin grafts in a subject including the step of administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to assist said local skin trauma of skin grafts.

3. A method for assisting healing of bone fractures in a subject including the step of administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to assist in healing of said bone fractures.

4. A method for improving the course of treatment for osteomyelitis in a subject including the step of administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to improve the course of treatment for said osteomyelitis. A method for assisting in the healing of burns in a subject including the step of administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to 25 assist in the healing of said burns.

S

6. A method for assisting in the healing of frost bite in a subject including the step of administering to the subject an amount of a dipeptide selected from the group consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to assist in the healing of said frost bite.

7. A method for sustaining weight gain and growth in an animal including the step of administering to the animal an amount of a dipeptide selected from the group T CAWINWORDWENDYNODELEMTBI53CLDOC NVT0 -43- consisting of L-Glu-L-Trp and a pharmaceutically acceptable salt thereof effective to sustain weight gain and growth. DATED: 29 February, 1996 PHILLIPS ORMONDE FITZPATRICK Attorneys for: CYTOVEN I ol* K- Il INTERNATIONAL SEARCH REPORT International Application No. PCT/US92/02440 I. CLASSIFICATION OF SUBJECT MATTER (if several classification symbols aoolv. indicate all) 3 I According to International Patent Classification (IPC) or to both National Classification end IPC IPC A61K 37/00; C07K 5/00 US CL 514/19; 530/331 II. FIELDS SEARCHED Minimum Documentation Searched 4 Clasification Systerr Classification Symbols U.S. 514/19; 530/331 Documentation Searched other than Minimum Documentation to the extent that such Documents are included in the Fields Searched 6 APS, CAS, Biosis, Medline III. DOCUMENTS CONSIDERED TO BE RELEVANT 14 Category" Citation of Document, 1 with indication, where appropriate, of the relevant passagesl7 Relevant to Claim No. 1

8 X Medline Abstracts, Issued 1990, Rodionov et al., 1,4,5,30,61-66 "The immunocorrective Therapy Of Pyoderma Caused By Staphylococci Multiply Resistant To Antibiotics", Abstract No. 90224329, Vestn. Dermatol. Venerol., Volume 1, pages 42-45. See Abstract. X/Y,P Medline Abstract, Issued September 1991, Iakovlev et 1 2 27 6 1 al., "The Biochemical And Immunological Indices In The 66/40,60 Rehabilitation Period Of The Victims Of The Accident On The Komsomolets Atomic Submarine", Abstract No. 92101433, Voen. Med. ZH, Volume 9, pages 28-33. See Abstract. X/Y Bulletin: "Thymogen", published 1989 by 1,2,18,19,34- Cytomed(Lennirgrad), pages 3-10. 35,40,42,43,48 ,55,57,60,62,6 3/3 ,9-19,24- 29,31,41,44- 46,64-66 SSpecial categones of cited documents: 1 later document published aftar the international filing document definng the genral state of the art which is date or priority date and not in conflict with the not cornideed to be of partiuar relevance application but cited to understand the principle or earlier document but published on or after the theory underlying the invention interntionl filing date "X document of particular relevance; the claimed document which may throw doubts on priority claim(s) invention cannot be considered novel or cannot be or which is cited to establih the publication date of considered to involve an inventive step another citation or other special reason (a specified) document of particular relevance; the claimed document referring to an oral dilosur, use, exhibition invention cannot be considered to involve an or other maan inventive step when the document is combined with or other means one or more other such documents, such combination P" document published prior to the international filing date being obvious to a person skilled in the art but later then the priority date claied document member of the same patent family IV. CERTIFICATION Date of the Actual Completion of the International Search 2 Date of Mailing of this International Search Report 2 16 JUNE 1992 24 JUN1992 International Searching Authority' Signature of Authorized Officer 2 0 ISA/US A c C E ISA/US BENNETT CELSA Form PCT/ISA/210 (second sheet)(May 1986) B International ApplicL n No. PCT/tJS92/02440 FURTHER INFORMATION CONTINUED FROM THE SECOND SHEET x, P x/Y Medline Abstract, Issued 1991, Khmellnitskii et I67 "Morphofunctional Characteristics Of Thel Immuunocompetent System In Hypotrophy and Its Correction By Thymogen, Abstract No. 92171772, Arkh. Patol. Volume 53(10), pages 24-27. See Abstract. I Yakovlev et "Resistance Stress Regulation", published 1990 by Nauka Publishers (Leningrad) See pages 90-93. 1,2, 27, 20, 28/: 1,18,1,43 ,48 V. OBSERVATIONiS WHERE CERTAIN CLAIMS WERE FOUND UNSEARCHABLE 1. Claim numbrs_, because they relate to subject matter not required to be searched by this Authortty, namnely: 2. CClaim rasnters because tde relate to puts of the International application that do not comply with the preecribed req~airentrs to stich an extent that no msuugfti Irtunationi search can be curle okx I1). apecWWficy:. 3. Claim mnbrers becauis they am dependent claims not drafted in accordance with the second aid third sentences of PCT LA* 0.4(a). VI. E] OBSERVATMON WHER uNrTy OF WIIvENTioN is LACKiNG 2 This kntematlione Searching Athority founod multiple lInventions lIn this Irrtsrnatiioria application as follows: 1. ElAs al eqired additiona search fes wea tknsly pod by the applicant, this imrwatona search report covers all eearrhabio claims of frthem on kpplcwn 2. As Aoriysomes of the reqired edltona search fees were tlirily prid by the applicant, this Wtsmationa mearch report covers only those cdamns of the International application for which ees were paid, specifically claims: 3.C No reqLars additional search fees were timly paid by the applicat, Coneequertly, this Internatonal search report is reetrtcted to the swertor firs mentioned in the claims; It is covered by claim numbters: 4, As sil searchek"l claim couLd be searched wfth*LX effort justIfying an additional fee, the Intarnatlona Search Authortty did noot invite payment of anry additonal fee. Bemnark on protest C]The additiona search fees were accompwanied by applicant's protest. O No protest eccomnied the payment of additional search foes, Form PCTo!SAI210 Isupplernental shoot12))(Rev. 4-90) B III~SL-P~ International Application No. PCT/US92/02440 III. DOCUMENTS CONSIDERED TO BE RELEVANT (CONTINUED FROM THE SECOND SHEET) Category* Citation of Document, 1 with indication, where appropriate, of the relevant peasges 17 Ralevant to Claim No. 1 a X,P X/Y, P X/Y X,P Medline Abstract, issued September-October 1991, Grigoriants et al., "Immunocorrection In The Combined Treatment Of Patients With Osteomyelitis Developing Following Combined Injuries To The Maxillofacial And Craniocerebral Areas", Abstract No. 92188368, Stomatologiia(Mosk), Volume 5, pages 53-54. See Abstract. Biological Abstract, Volume 92, Issued 1991, Rodionov et al., "Natural Killer Activity In Patients With Chronic Dermatoses", Abstract No. 100385, Vestn. Dermatol. Venerol. Volume 5, pages 4-6. See Abstract. Chemical Abstracts, Volume 116, No.17, Issued 1990, Aliev et al. "Simulation Of Thymus Dysfunction In Guinea Pigs By Using Immunomodulators", Abstract No. 171986U, Izv. Akad. Nauk. Az. SSR, Ser. Biol. Nauk., Volume 1, pages 73-80. See Abstract. Chemical Abstract, Volume 116, No. 17, Issued 1991, Demidov et al., "Effects Of Thymus Preparations And Antituberculous Drugs On Immunological Reactivity And The Course Of Tuberculous Process In Experimental Animals", Abstract No. 165824Y, Probl. Tuberk. Volume 12, pages 52-54. See Abstract. 2,31,35,61-66 1,4,5,39,61- 63/6,7,8,62,64 -66 1,2,61/3,60,63 -66 1,54,61,63-66 Form PCT/SA/210 (extra sheet)(May 1986) B 1- i.