WO1995009646A1

WO1995009646A1 - METHOD OF TREATING SEPTIC SHOCK USING THYMOSIN β¿4?

Info

Publication number: WO1995009646A1
Application number: PCT/US1994/010879
Authority: WO
Inventors: Allan L. Goldstein; Mirela O. Fagarasan
Original assignee: The George Washington University Medical Center
Priority date: 1993-10-07
Filing date: 1994-09-27
Publication date: 1995-04-13
Also published as: AU7879294A; EP0725652A1

Abstract

A method of treating septic shock in a mammal including the administration of a septic shock-treating effective amount of Tβ4 to the mammal.

Description

METHOD OF TREATING SEPTIC SHOCK USING THYMOSIN β.

The present invention relates to a method of treating septic shock in mammals.

Description of Background Art

Thymosin β_Λ ("T?₄") is a peptide which has been reported as containing 43 amino acids. Amino acid sequence information on T?₄ is disclosed in U.S. Patent No. 4,297,276, herein incorporated by reference. T_/β₄ has been found to be present in numerous tissue types in mammals and has also been implicated in a wide variety of cellular and physiological processes including inducing terminal deoxynucleotidyl transferase activity of bone marrow cells, stimulating secretion of hypothalamic luteinizing hormone releasing hormone and luteinizing hormone, inhibiting migration and enhancing antigen presentation of macrophages, and inducing phenotypic changes in T-cell lines in vitro. Septic shock is a condition in which infection is widely disseminated in many areas of the body, the infection generally being disseminated through the blood from one tissue to another and causing extensive damage. Septic shock can occur with numerous medical conditions, including (1) peritonitis caused by the spread of infection from the uterus and fallopian tubes; (2) peritonitis resulting from rupture of the gut, sometimes caused by intestinal disease or wounds; (3) generalized infection resulting from spread of a simple infection; (4) generalized gangrenous infection resulting specifically from gas gangrene bacilli; and (5) infection spreading into the blood from the kidney or urinary tract. Septic shock is of critical concern from a clinical viewpoint because, among other reasons, this condition frequently leads to death.

Although septic shock is a somewhat common clinical phenomenon, the mechanisms involved as well as the pathological changes remain poorly understood. For example, despite the treatment of bacterial infection, many patients ^'deteriorate further, which may be due to clinical sequelae of hypotension with low systemic vascular resistance, renal insufficiency, adult respiratory distress syndrome, severe coagulopathy and severe metabolic dysfunctions. Thus, there is an urgent need in the art for effective methods of treating septic shock.

Summary of the Invention In accordance with the present invention, a method of treating septic shock in mammals includes administering a septic shock-treating effective amount of TjS₄ to said mammals.

Description of the Preferred Embodiments The terms "Thymosin ?₄" and "T_/S₄" refer to peptides having the amino acid sequence disclosed in U.S. Patent No. 4,297,276, supra.

Mammalian septic shock occurs in association with a series of events in the mammal's body referred to as the "sepsis cascade" . The sepsis cascade typically begins with bacterial infection of the mammalian host resulting in release of bacterial toxins, introduction of endotoxin, activiation of host defense systems, i.e., plasma protein systems as well as cellular defense systems including endothelial cells, macrophages, monocytes and neutrophils, with release of proinflammatory mediators including cytokines, lipid metobolites, proteases, toxic oxygen products, nitric oxide and adhesion proteins. According to one aspect of the present invention, effective amounts of T/3₄ are administered to a subject to reduce blood free radical levels in the subject, and thereby treat or prevent septic shock in the subject or obstruct progression of sepsis cascade in the subject. T/3₄ has been found to reduce blood free radical levels almost as much as SOD (super oxide dismutase) , an enzyme that eliminates free radicals. When a septic shock-treating or a septic shock-preventing effective amount of T3₄ is administered to a mammal, the blood levels of pathological mediators of bacteria-induced lethality are decreased in the mammal.

T3₄ has been found to obstruct the sepsis cascade in mammals. During sepsis, peroxidation of lipids in blood is increased (mMol of malonyldialdehyde) , but returned to normal or about normal with T?₄ administration. Sepsis also reduces circulating blood leves of glutathione. However, administration of T/3₄ returns circulatory gluthathine leves to normal or about normal . As noted above, administration of T3₄ during sepsis decreases blood hydroperoxide levels and blood glutathione levels. Administration of T3₄ during sepsis also decreases cerebellar cGMP levels, and decreases the blood levels of arachidonic acid metabolites such as Tx/3₂ and 6-keto-PGF^, PAF, and cytokines such as IL- lα and TNF-c .

While not wishing to be bound to any particular theory, it is believed that reducing blood levels of pathological mediators of bacteria-induced lethality in a mammal decreases the amount of infection in the mammal which, in turn, aids in obstructing the spesis cascade, and in preventing and treating septic shock. Thus, according to the present invention, methods of treating and preventing septic shock in mammals are provided. The methods of the present invention include administration of septic shock-treating effective amounts, septic shock-preventing effective amounts and sepsis cascade progression-obstructing effective amounts of T3₄ to mammals.

According to preferred embodiments of the present invention, effective amounts of T3₄ are administered to subjects to treat or prevent septic shock in the subjects, or obstruct progression of sepsis cascade in the subjects. In these embodiments, the subjects preferably are human.

According to preferred embodiments of the present invention, compositions containing T/3₄ may be formulated in a conventional manner for administration by any suitable route. Suitable routes of administration include, but are not limited to, oral, rectal, nasal, topical, vaginal, and parenteral (including subcutaneous, intramuscular, intravenous and intradermal) . Particularly preferred embodiments utilize oral or parenteral administration, with parenteral administration being a more preferred embodiment. It will be appreciated that the preferred route may vary with the condition, age and species of the recipient . While not essential, in preferred embodiments, T?₄ is administered as part of a pharmaceutical formulation. The formulations of the present invention comprise T/3₄ together with one or more pharmaceutically acceptable carriers and optionally with other therapeutic ingredients. The carrier(s) are "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. The formulations include those suitable for oral, rectal, nasal, topical (including buccal and sublingual) , vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. The formulations may conveniently be presented in unit dosage form, e.g., tablets and sustained release capsules, and may be prepared by any suitable pharmaceutical methods .

Such methods include, but are not limited to, the step of bringing into association T/3₄ with the carrier which constitutes one or more accessory ingredients.

In general, the formulations are prepared by uniformly and intimately bringing into association T3₄ with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product. Formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of TS_4; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion, etc.

A tablet may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine T3₄ in a free-flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, preservative, surface-active or dispersing agent. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein. Formulations suitable for topical administration include lozenges comprising T3₄ in a flavored basis, usually sucrose and acacia or tragacanth; pastilles comprising T/3₄ in an inert basis such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising Tβ₄ to be administered in a suitable liquid carrier.

Formulations suitable for topical administration to the skin may be presented as ointments, creams, gels and pastes comprising T/3, and a pharmaceutically acceptable carrier, or may utilize a transdermal patch containing the ingredient to be administered.

Formulations for rectal administration may be presented as a suppository with a suitable base comprising, for example, cocoa butter or a salicylate. Formulations suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size, for example, in the range from about 20 to about 500 microns which is administered in the manner in which snuff is taken, i.e., by rapid inhalation through the nasal passage from a container of the powder held close up to the nose. Suitable formulations wherein the carrier is a liquid, for administration, as for example, a nasal spray or as nasal drops, include aqueous or oily solutions of the active ingredient.

Formulations suitable for vaginal administration may be presented as tampons, creams, gels, pastes, foams or spray formulations containing, in addition to T_/S₄, suitable carriers. Formulations suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may optionally contain anti- oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents . The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

It should be understood that in addition to the ingredients particularly mentioned above the formulations of this invention may include other suitable agents having regard to the type of formulation in question, for example, those suitable for oral administration may include flavoring agents.

A proposed dose for administration of the compositions in the present invention is a septic shock-treating, septic shock-preventing or sepsis cascade progression-obstructing effective amount of T/S₄, which can be in a range of from about 0.4 to about 4 mg of Tjβ₄ per kg of body weight of recipient (mg/kg) , preferably from about 1 to about 4 mg/kg. A dose can be administered to the patient daily, one or more times per day of administration, e.g., two or three times per day, and doses can be administered one or more days per week, e.g., two, three, four, five, six or seven days per week. The invention is applicable to native (i.e., naturally occurring) Tβ_t as well as synthetic Tβ₄ and recombinant Tβ₄ having the amino acid sequence of native Tjβ₄, biologically active amino acid sequences substantially similar thereto, or a biologically active abbreviated sequence form thereof, and their biologically active analogs (including muteins) having substituted, deleted, elongated, replaced, or otherwise modified sequences which possess bioactivity substantially similar to that of native T3₄.

In accordance with one embodiment of the present invention, T3₄ can be administered in combination with a therapeutically effective amount of another substance useful in treating septic shock such as, for example, antibiotics, or antibodies (polyclonal or monoclonal) directed to antigens located on endotoxins. Of course, the acceptable dosage range of the other substance will depend upon its properties (i.e., the acceptable dosage range will depend upon what other substance is being administered) .

T?₄ and another substance useful in treating septic shock can be administered "in combination" which, as defined herein, includes various schemes designed to administer T3₄ and the other substance to a subject, whether or not the other substance and T3₄ are administered separately or together, such that the desired dosages of T3₄ and the other substance are present in the subject at the same time. Any suitable scheme can be used to administer T/3₄ and another substance useful in treating septic shock "in combination" in accordance with the present invention.

Suitable dosages of either T/3₄ alone or T/3₄ in combination with another substance useful in treating septic shock may be administered 1 to 6 times or more per day. The precise dose administered will depend on the age, condition and other factors of the recipient.

The following examples are for illustrative purposes only, and are not to be construed in a limiting sense.

Example 1 Synthetic T/3₄ was provided by Alpha 1 Biomedicals, Inc. (Two Democracy Center, 6903 Rockledge Drive, Ste. 1200, Bethesda, Maryland 20817) . T/3₄ was prepared by solid phase peptide synthesis.

Swiss-Webster mice 4-6 weeks of age (20 - 25g) were housed 6 per cage. The mice were divided into 2 groups: endotoxic mice (endotoxin 60 mg/kg i.p. in acute treatment) and endotoxic mice treated with T ₄, 5 minutes and 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24 hours post administration of the endotoxin. Survival was recorded 2 times a day for 7 days. The results are presented in Table I below. As can be seen, T3₄ protected against lethal endotoxin shock and increased the survival rate to as much as 100%.

Example 2 Swiss-Webster mice 4-6 weeks of age (20-25g) were divided into groups as follows.

1. Mice treated with a lethal dose of endotoxin (60 mg/kg) .

2. Mice treated with 60 mg/kg endotoxin followed by an injection of 100 μg T3₄ (5 minutes post administration of endotoxin) .

3. Mice treated with 60 mg/kg endotoxin followed by 2 injections of 100 μg/T_/8₄ (immediately following and 2 hours post administration of endotoxin) .

4. Mice treated with 60 mg/kg endotoxin followed by 3 injections of 100 μg Tβ_i (immediately following, 2 and 4 hours post administration of endotoxin) . The results presented in Tables I and II below indicate that T0₄, administered 3 times post administration of endotoxin, increased the survival rate of mice treated with endotoxin to 100%.

TABLE I

Experimental Groups Survival of Swiss Webster Mice Following Lethal Endotoxin Dose and T/5₄ (No. mice alive)

0 hrs 24 hrs 48hrs 72hrs

Endotoxin 60 mg/kg 7 1 1 1 /kg 7 4 3 3

Endotoxin 60 mg/kg + T 3₄ 100 μg 2x

Endotoxin 60 mg/kg + T/S₄ 100 μg 3x

TABLE II

Protective Effect of Υβ_t on Survival of Mice Treated with Lethal Doses of Endotoxin

Number of Mice Alive

EXPERIMENTAL GROUPS 0 24 48 72 7 14 hr. hr. hr hr. day day

Endotoxin 60 mg/kg 8 o

Endotoxin 60 mg/kg 8 8 8 8 8 8 +T_/8₄ 100 μg 3x

The results presented in Tables III - V below show effects of T/3₄ respectively on mice cerebellar cGMP levels , blood serum TNF-α_ levels , and blood serum TNF- a and IL- lα levels . TABLE III

Tβ_t effects on cGMP cerebellar levels in mice treated with lethal doses of endotoxin

TABLE IV

Serum TNF-α levels in mice treated with lethal doses of endotoxin

EXPERIMENTAL GROUPS TNF-α^* (pg/ml)

1 hr 3 hr 5 hr

Endotoxin 60 mg/kg 1098 ± 783 ± 38 623 ± 51 106

Endotoxin 60 mg/kg 823 + 79 535 ± 47 480 ± 45 + 1β_t 100 μg

TABLE V

Serum TNF-α^* and IL-lα. levels^* in mice protected against endotoxin-induced lethality by pretreatment with SOD and T/3₄

Pretreatment TNF-α. (pg/ml) IL-1 α (pg/ml)

None 4462 ± 123 1137 ± 123

SOD (3.3 x 10⁴ μ/kg) 63 ± 5.1 53 ± 3.1

1β_t 100 μg/mice 321 ± 26.2 298 + 20

* Mice were pretreated with SOD (super oxide dismatase) , 30 minutes before endotoxin administration XX blood for determined of TNFα. and IL-lα^* levels was collected 1 hr after endotoxin administration. Example 3 Using the same materials and methods as in examples 1 and 2, the time and dose-dependency of the protective effect of T3₄ on endotoxin lethality were studied. The results are presented in Tables II and III below.

As can be seen in Tables VI and VII, T_/δ₄ had a protective effect on endotoxin toxicity when given immediately following, 2 and 4 hours after endotoxin treatment. The most effective protective dose was 100 μg T3₄ administered three times. Also, T/3₄ partially increased the survival of mice treated with endotoxin when it was administered three times in doses of 50 μg and 20 μg T3₄. 10 μg T3₄ has also sometimes shown positive activity.

SUBSTmJTΕSHEET(RULE26) TABLE VI

Experimental Groups Survival of Swiss Webster Mice Following Lethal Endotoxin Dose and T/3₄ (No. mice alive)

0 hrs 24 hrs 48 hrs 72 hrs 96 hrs 120 hrs

Endotoxin 60 mg/kg 8 4 4 4 4 4

Endotoxin 60 mg/kg 8 8 8 8 8 8 10 + T/3₄ 100 μg, 3x; TjS₄ was admini¬ stered immediately following, 2 and 4 hrs. after endotoxin.

15 Endotoxin 60 mg/kg 8 + T(8₄ 100 μg, 3x; TjS₄ was admini¬ stered 2, 4 and 6 hrs. after endotoxin.

20 Endotoxin 60 mg/kg 8 + T/3₄ 10 μg, 3x; T/3₄ was admini¬ stered immediately following, 2 and 4

25 hrs. after endotoxin

TABLE VII Experimental Groups Survival of Swiss Webster Mice

Following Lethal Endotoxin

Dose and Tβ_t

5 (No. mice alive)

0 hrs 24 hrs 48 hrs 72 hrs 96 hrs 5 days 6 days

Endotoxin 60 mg/kg 10 4 3 3 3 3 3

Endotoxin 60 mg/kg 10 9 7 7 7 7 7

hrs. after en otox n.

15 Endotoxin 60 mg/kg 10 5 4 4 3 3 3 + T(8₄ 20 μg, 3x; i.p. Tj8₄ was admini¬ stered immediately following, 2 and 4

20 hrs. after endotoxin. 0 3 3 3 3 3 3

25

Example 4

Using the same methods and materials as in examples 1-3, the effect which Tβ₄ has on blood levels of IL-lα, TNF-α, PAG, Tx3₂ and 6-keto-PGFiQ., which are pathological mediators of endotoxin induced lethality, was studied. The results are presented in Tables IV- IX below.

As can be seen in Tables VIII-XIII, Tβ₄ decreased IL-lα, TNF-α serum levels as well as PAF, Tx/3₂ and 6- keto-PGF^ plasma levels after administration of a lethal dose of endotoxin.

TABLE VIII

EXPERIMENTAL GROUPS IL-lα pg/ml (serum levels]

1 hr 3hr Endotoxin 60 mg/kg 492 + 45, 2 550 ± 37, 1

Endotoxin 60 mg/kg + 147,5 ± 15,1 207, 5 + 19,3 T_/δ₄ 100 μg administered simultaneously

TABLE IX

EXPERIMENTAL GROUPS PAF (pg/ml) (plasma levels)

0.5 hr. 1 hr 2 hr 3 hr

Endotoxin 60 78 + 6 279 + 17 127 + 13 55 + 3 mg/kg

Endotoxin 60 63 ± 7 161 + 17 66 + 5 46 + 5 mg/kg + T|S₄ 100 μg admin¬ istered simultaneously TABLE X

EXPERIMENTAL GROUPS Txβ, (pg/ml) (plasma levels)

0.5 hr. 1 hr 2 hr 3 hr

Endotoxin 60 1442 + 103 2937 ± 258 912 ± 105 695 ± 65 mg/kg

Endotoxin 60 209 ± 9,5 607 + 53 196 + 18 112 + 9 mg/kg + Tβ, 100 μg admin¬ istered simultaneously

TABLE XI

EXPERIMENTAL GROUPS 6-keto-PGFlα (pg/ml) (plasma levels)

0.5 hr. 1 hr 2 hr 3 hr Endotoxin 60 341 ± 31 1141 + 112 897 ± 75 811 ± 7 mg/kg

Endotoxin 60 141 + 9 261 + 23 147 + 19 121 + 13 mg/kg + T3₄ 100 μg admin- istered simultaneously

TABLE XII

EXPERIMENTAL GROUPS PAF (pg/ml) (serum levels] 1 hr 3 hr 5 hr Endotoxin 60 938 662 567 mg/kg

Endotoxin 60 807 591 496 mg/kg + T?₄ 100 μg admin- istered simultaneously TABLE XI I I

EXPERIMENTAL GROUPS TNF-α (pg/ml) (serum level)

1 hr 3 hr 5 hr

Endotoxin 60 938 662 567 mg/kg

Endotoxin 60 807 591 496 mg/kg + T/3₄ 100 μg admin¬ istered immediately after endotoxin

While the invention has been described and illustrated with details and references to certain preferred embodiments, those skilled in the art will appreciate that various modifications, changes, omissions, and substitutes can be made without departing from the spirit of the invention.

Example 5

Using a septic shock model in rats (Sprague- Dawley, male, 200-225 g each) , the effect of thymosin β__ alone and together with antibiotics was studied. Peritonitis was induced in rats in the following way. A 1-cm incision was made into the peritoneal which exposed the cecum. A tight ligature was placed around the cecum with 4-0 suture distal to the insertion of the small bowel, forming an area of devitalized tissue while maintaining bowel continuity. A puncture wound was made with 16-gauge needle into the anti-mesenteric surface of the cecum and a small amount of fecal contents was expressed through the wound. The cecum was replaced into the peritoneal cavity, and the anterior peritoneal wall and skin were closed with surgical staples. Each animal was given a bolus of normal saline (15 ml/kg) for hydration and allowed to recover overnight.

The results presented in Table XIX show T?₄ increased the survival of rats.

TABLE XIX

Survival of Sprague-Dawley rats following septic shock,antibiotics, and thymosin β_t

EXPERIMENTAL GROUPS 0 hr 24 hr 48 hr 72 hr

Rats with Septic Shock + antibiotic* 10 ck + in 10

* gentamicin sulfate, 1 mg/rat

** At 0 hr, 2 hr, and 4 hr post induction of peritonitis

The results presented in Tables XX to XXIII below show effects of Tβ₄ respectively on rat cytokine levels , blood malonyldialdehyde levels , glutathione levels and arachidonic acid metabolic levels .

TABLE xx

The effect of β_t on Cytokine Levels in a septic shock model in rat

EXPERIMENTAL GROUPS TNFα (pg/ml) IL.ot (pg/ml)

Control undetectable undetectable

Rats with Septic 2658 ± 197 1387 + 123 Shock

Rats with Septic 1259 + 138 853 + 62 Shock + T3₄ 1 mg/rat x 3 as above TABLE XXI

The effect of Tβ₄ on lipid peroxidation in a septic shock model in rat

EXPERIMENTAL GROUPS Blood Levels of malonyldialdehyde (nNol MDA)

Control 15.3 ± 1.3 18.6 ± 1.1

Rats with Septic 49.1 ± 3.5 46.3 ± 3.8 Shock

Rats with Septic 19.6 ± 1.7 16.8 ± 1.5 + T3₄ lmg/rat x 3 as above

TABLE XXII

The effect of Tβ_t on glutathione levels in a rat septic shock model

EXPERIMENTAL GROUPS RBC GSH (u oles/ml cells)

Control 1.38 + 0.1 1.46 ± 0.13

Rats with Septic Shock 0.43 ± 0.05 0.29 + 0.04

Rats with Septic Shock 1.45 ± 0.11 1.39 ± 0.12

+ Tβ mg/rat x 3 as above

TABLE XXIII

The effect of T/3₄ on arachidonic acid metabolite levels in a septic shock model in rat

EXPERIMENTAL GROUPS 6-keto-PGF, (pg/ml) T x β. (per/ml)

Control 58 ± 3.5 89 + 7.6

Rats with Septic Shock 1828 ± 145.3 3622 ± 295

Rats with Septic Shock 829 ± 59 1627 + 123

+ TS₄ 1 mg/rat x 3 as above

The. results presented in Tables XXIV and XXV below demonstrate that T/3₄ increases the survival rate of rats with septic shock. TABLE XXIV

Number of Rats Alive

EXPERIMENTAL GROUPS 0 h 24 h 48 h 72 h

Rats with Septic Shock 10 2 2 0

Rats with Septic Shock 10 3 3 3 + T_/S₄ 1 mg/rat x 3 as above

TABLE XXV

Number of Rats Alive

EXPERIMENTAL GROUPS

0 h 24 h 48 h 72 h

Rats with Septic Shock 10 1 0 0

Rats with Septic Shock 10 4 3 3 + Tβ_a 1 mg/rat x 3 as above

SUBSTITUTE SHEET {RULE 26)

Claims

What is claimed is:

1. A method of obstructing progression of a sepsis cascade in a mammal in which a sepsis cascade is occurring, comprising administering to said mammal a sepsis cascade progression-obstructing amount of thymosin β_t (T3₄) .

2. The method of claim 1, wherein said mammal is human.

3. The method of claim 2, wherein the T?₄ is administered at a dosage from about 0.4 to about 4 mg per kg of body weight of said mammal.

4. The method of claim 2, wherein the T3₄ is administered at a dosage from about 1 to about 4 mg per kg of body weight of said mammal.

5. The method of claim 3, wherein the T3₄ is administered parenterally.

6. The method of claim 5, wherein the T/3₄ is administered intravenously.

7. The method of claim 1, wherein the T3₄ is synthetic Tc^.

8. The method of claim 1, wherein said amount of T3₄ is sufficient to reduce in said mammal at least one member selected from the group consisting of cerebellar cGMP levels, blood malonyldialdehyde levels, blood glutathione levels, blood PAF levels, blood Tx/3₂ levels, blood 6-keto-PGFiC levels, blood IL-lα. levels and blood TNFα levels.

9. A method of preventing septic shock from occurring in a mammal following endotoxin introduction in said mammal, comprising administering to said mammal a septic shock-preventing effective amount of Tβ₄, wherein septic shock is prevented in said mammal for at least 72 hours after administration of said T3₄.

10. The method of claim 9 wherein septic shock is prevented in said mammal for at least 120 hours after administration of said T/3₄.

11. The method of claim 9 wherein septic shock is prevented in said mammal for at least 14 days after administration of said T3₄.

12. The method of claim 9 wherein said mammal is human.

13. The method of claim 12, wherein the T/?₄ is administered at a dosage from about 0.4 to about 4 mg per kg of body weight of said mammal.

14. The method of claim 12, wherein the T/3₄ is administered at a dosage from about 1 to about 4 mg per kg of body weight of said mammal .

15. The method of claim 13, wherein the T3₄ is administered parenterally.

16. The method of claim 15, wherein the T/3₄ is administered intravenously.

17. The method of claim 9, wherein the T3₄ is synthetic T/3₄.

18. The method of claim 9, wherein said amount of TjS₄ is sufficient to reduce in said mammal at least one member selected from the group consisting of cerebellar cGMP levels, blood malonyldialdehyde levels, blood glutathione levels, blood PAF levels, blood Tx/3₂ levels, blood 6-keto-PGF^ levels, blood IL-lα levels and blood TNFo. levels.

19. A method of treating septic shock in a mammal which comprises administering to said mammal a septic shock-treating effective amount of T/?₄.

20. The method of claim 19, wherein said mammal is human.

SUBSTITUTE SHEET (RULE 2tt

21. The method of claim 20, wherein the T?₄ is administered at a dosage from about 0.4 to about 4 mg per kg of body weight of said mammal .

22. The method of claim 20, wherein the T?₄ is administered at a dosage from about 1 to about 4 mg per kg of body weight of said mammal .

23. The method of claim 22, wherein the Tβ₄ is administered parenterally.

24. The method of claim 23, wherein the T3₄ is administered intravenously.

25. The method of claim 19, wherein the Tβ₄ is synthetic T/3₄.

26. The method of claim 19, wherein said amount of T_/β₄ is sufficient to reduce in,said mammal at least one member selected from the group consisting of cerebellar cGMP levels, blood malonyldialdehyde levels, blood glutathione levels, blood PAF levels, blood Tx3₂ levels, blood 6-keto-PGF^ levels, blood IL-lα levels and blood TNFα. levels.

27. A pharmaceutical dosage unit containing an amount of T3₄ selected from the group consisting of a mammalian sepsis cascade progression-obstructing effective amount of T3₄, a mammalian septic shock- preventing effective amount of T/?₄ and a mammalian septic shock-treating effective amount of T?₄, wherein said TjS₄ is provided in a carrier which is pharmaceutically acceptable for administration to a mammal.

28. The pharmaceutical dosage unit of claim 27, containing from about 0.4 to about 4 mg T3₄ per kg of mammalian body weight .

29. The pharmaceutical dosage unit of claim 27, containing from about 1 to about 4 mg T/3₄ per kg of mammalian body weight.

30. The pharmaceutical dosage unit of claim 28 which is in a form for parenteral administration, wherein the carrier is a sterile liquid carrier suitable for parenteral administration.

31. The pharmaceutical dosage unit of claim 30 which is in a form for intravenous administration.

32. The pharmaceutical dosage unit of claim 31, wherein the T3₄ is dissolved in a sterile isotonic saline solution.

33. The pharmaceutical dosage unit of claim 27, wherein said T3₄ is present in said pharmaceutical dosage unit in an amount sufficient to reduce in said mammal at least one member selected from the group consisting of cerebellar cGMP levels, blood malonyldialdehyde levels, blood glutathione levels, blood PAF levels, blood TXJ8₂ levels, blood 6-keto-PGF^ levels, blood IL-lα. levels and blood TNFα; levels.

34. The method of claim 1, wherein said administration of T/3₄ reduces blood free radical levels in said mammal.

35. The method of claim 9, wherein said administration of Tβ₄ reduces blood free radical levels in said mammal .

36. The method of claim 19, wherein said administration of Tβ₄ reduces blood free radical levels in said mammal.