# Patent

## Title
3,5,3′ -triiodothronine sulfate as thyromimetic agent and pharmaceutical formulations thereof

## Abstract

The invention regards the use of triiodothyronine sulfate, commonly named T3S, as a medicament having thyromimetic activity for the treatment of pathologies due to organic deficiency of triiodothyronine (T3), as such or in association with thyroxine (T4), and pharmaceutical formulations thereof.

## Background
This application is the national stage application of corresponding international application number PCT/EP2003/012584 filed Nov. 11, 2003, which claims priority to and the benefit of the Italian application no. MI2002A002394, filed Nov. 13, 2002, all of which are hereby incorporated by reference.
FIELD OF THE INVENTION
The present invention regards the use of 3,5,3′-triiodothyronine sulfate, usually named triiodothyronine sulfate or T3 sulfate or even better T3S, as an active principle, alone or in combination with thyroxine, in the treatment of pathologies due to organic deficiency of 3,5,3′-triiodothyronine. Accordingly, the same is usable for the preparation of thyromimetic pharmaceutical compositions.
BACKGROUND OF THE INVENTION
A number of iodothyronines are present in blood, which are directly produced by thyroid gland or are the result of peripheral metabolism of other iodothyronines. Among them, 3,5,3′-triiodothyronine (acronym T3) is deemed to be the biological active form of thyroid hormone (TH), because it has shown high affinity for the specific receptors of the same and is normally present in serum at a concentration sufficient for the activation of said receptors.
The main secretion product of thyroid gland in the healthy adult is thyroxine, commonly designated with the acronym T4. It is peripherically converted to its biologically active form, T3 (Ref. 1), through enzymatic removal of an iodine atom from the external aromatic ring of the molecule by both type I and type II 5′-iodothyronine monodeiodinases (type I MD and type II MD, respectively). This metabolic pathway is the main mechanism of endogenous production of T3; on consequence, T4 can properly be considered a pro-hormone. On the other hand, a minor part of T3 is also directly secreted by thyroid. On average, the amount of T4 produced in an adult being of 70 Kg weight every day amounts to 100 μg, while the total production of T3 amounts to around 25 μg. 4-8 μg of T3 out of said 25 μg are directly secreted by thyroid and the remaining ones derive from the peripheral conversion of T4.
T3 undergoes two different metabolic pathways. The main metabolic pathway consists in the partial deiodination of the inner aromatic ring by type III 5-iodothyronine monodeiodinase (type III MD) to give 3,3′-diiodothyronine, which is biologically non-active and is further metabolized through deiodination or sulfoconjugation. The other metabolic pathway regards around 20% of the total amount of T3 produced by the body and brings on sulfoconjugation of T3 to give T3S, which is not able to bond to the thyroid hormones (Ref. 2), thus resulting biologically non-active (Ref. 3).
Contrary to what happens with T3, T3S is not deiodinated by type III MD. Rather, it resulted to be an excellent substrate for type I MD (Ref. 4), which converts it very quickly into 3,3′-diiodothyronine sulphate. On consequence it has been widespread common knowledge that, in the healthy adult being, sulfoconjugation of T3 to give T3S represents a way for speeding up the catabolism of T3, so facilitating its biliary and urinary excretion. Actually, it was found that serum levels of T3S, physiologically low in the health adult, are higher when type I MD activity is reduced.
Yet, it has also unexpectedly been found that, just in some body districts and organs, sulfatases exist which, under particular physiological conditions and situations, are able to convert again T3S into its active form T3 (Ref's. 7-9).
Such enzymes have been described in the intestinal microflora as well as in body tissues like liver, kidneys and nervous central system (Ref. 10).
Recently, it has been found that endogenous T3S levels in serum are quite high during intrauterine life and as such are kept by the body, i.e. higher than the ones normally found in the adult being, at least until the forth month of postnatal life (Ref. 11). Considering the essential role played by thyroid hormones during growth, in particular as far as nervous central system functions are involved, suppositions have been made about the possibility that, in this tissue, T3S may also possibly be used by the body as an occasional source of T3, if and when needed, during the first period of life. Studies performed on autoptic specimens of human nervous cerebral tissue post-mortem showed that the amount of T3 in the same results limited by type III MD (Ref. 12). While this enzyme does not attack T3S, it has been surmised that T3S may exceptionally represent an alternative endogenous source of T3 hormone in those tissues which contain sulfatases able to reconvert T3S into its active form, just in case a particular need of the hormone arises in said tissues (Ref's. 8, 13).
Further studies have been performed to ascertain the effective role played by T3S during production and metabolism of thyroid hormones. Said studies have recently demonstrated that it shows thyromimetic effects in hypothyroid rats (Ref. 10) as well as in euthyroid rats (Ref. 14). In both cases T3S has shown a potency of around one fifth that of T3. Moreover both treatments with T3S and with T3 produced a significant reduction of serum levels of thyreotropic hormone (TSH) in euthyroid rats, thus showing to possess similar capability in inhibiting its secretion. On the contrary, in the case of hypothyroid rats, T3S showed a poor capability of inhibiting TSH secretion when compared to T3. It is well known that TSB is a highly responsive indicator to the functional status of thyroid gland and consents to detect the smallest alterations of its hormonal secretion. Actually, its levels are higher under conditions of reduced thyroid functionality, even in those conditions that are defined as sub-clinical, while they are reduced when an excess of thyroid hormones are present. Accordingly, T3S seems unexpectedly non-comparable to T3 as far as its capability of inhibition on formation of TSH is involved.
In conclusion, particularly in view of the latest studies, a clear and complete knowledge of the biological role played by T3S has not yet been reached.
In fact its main, well-grounded and universally accepted, feature is its non-biological activity, i.e. it is a biologically inert metabolite of T3 (Ref's. 2 and 3), and the sulfation pathway is regarded as a metabolic activator of T3 catabolism (Ref. 5).
On the other hand, only in particular tissues and under exceptional critical conditions due to shortage of thyroid hormone in those tissues, it has been shown its potential as an endogenous local source of T3.
As a result, today the skilled technician is still facing a complex, somewhat conflicting, situation, which highlights only some of the biological characteristics of the product and needs more exhaustive in depth studies.
In any case, none of the several documents forming the state-of-the-art discloses, shows or suggests the possibility of using this anomalous metabolite of T3 in therapy. No close prior-art document, either of experimental nature or substantially speculative, either taken alone or in combination with other related documents, suggests the use, or even the potential use of T3S as a medicament, taken as such or preferably in combination with other thyroid hormones or pro-hormones, like, for example T4. The fact that, only in some specific tissues of the body and under particular, peculiar circumstances, part of T3S can be reconverted into T3 does not mean, nor implies, nor suggests that it is possible to generalize this feature to the whole organism through exogenous administration of the product. In particular, there is no suggestion that oral administration of the product, even in protected form according to known methods of the pharmaceutical technique, may render it bioavailable also because it is well known that in those districts where suitable sulfatases are not present the same is rapidly metabolized and excreted through the bile and urines.
SUMMARY OF THE INVENTION
It has now unexpectedly been found, and this is one of the aspects of the present invention, that T3S, as such or in association with other thyroid hormones or pro-hormones, preferably T4, and properly formulated according to the desired application, is particularly useful as a medicament to be used in all those pathologies caused by insufficient production by the body of the needed quantities of active thyroid hormones, in particular T3.


DETAILED DESCRIPTION OF THE INVENTION
In fact, it has unexpectedly been found that the administration of T3S, contrary to what known about its normal metabolism, allows to maintain steady levels of T3 in the body for long times (from 12 to 18 hrs) and that results particularly useful in those cases in which it is needed to supplement thyroid hormone in its most active form.
Particularly preferred in the therapy of hypothyroidism, and this is a main aspects of the present invention, is resulted the association of T3S with T4. The hormonal association which, in theory, should more accurately mime the normal thyroid secretion is represented by a combination of T4 with T3. Actually, pharmaceutical compositions comprising both of said iodothyronines, formulated in proportions similar to the ones of the normal physiologic secretion, have already been tried and marketed. Unfortunately, the oral simultaneous administration of T4 with T3 was not able to reproduce the normal thyroid hormones serum levels, because of pharmacokinetics of T3. In fact, T3 undergoes a very quick absorption and an equally quick elimination after oral administration; its elimination rate is about 20 times higher than the one of T4. For this reason administration of T3 gives raise to a dangerous peak excess in hormone concentration, if compared to the normal physiologic levels, followed by a too much fast drop to sub-physiologic levels. On consequence, today most of the specialised physicians prefer using T4 alone, even if in this way production of T3 only depends on the periferic deiodination of T4, because direct secretion of T3 by thyroid does not exists or is seriously insufficient.
On the contrary, the association of the invention avoids the above problems, because it has unexpectedly been found that, for example, after oral administration, T3S provides T3 serum levels that increase in a gradual way and keep steady for long periods of time, thus preventing the formation of too much high peaks.
Another unespected advantage deriving from the use of T3S in the treatment of pathologies due to organic deficiency of T3 consists in its recently found systemic thyromimetic activity linked to a poor inhibition of TSH secretion. This effect is particularly useful in the case of thyroidectomized patients suffering from thyroid carcinoma, when administration of T4 must be suspended in view of carrying out total body scintigraphy. In such a case administration of T3S instead of T4 may solve patient's necessity, without interfering with the diagnostic examination.
Another further advantage of T3S in the therapy of hypothyroidism regards its autolimitation capability. In fact, it is actively deiodinated by type I MD, which, on its part, is stimulated by thyroid hormones. In hypothyroid subjects type I MD activity is reduced; on consequence also T3S elimination is slowed. As a matter of fact, its effect on the body results greater. On the contrary, in case of over administration, type I MD activity is increased, thus giving more T3S elimination, i.e. limiting possible undesired collateral effects.
Last but not least, a further advantage of T3S is represented by the fact that it is a metabolite normally present in the body, usually non-active, i.e. non-toxic. On consequence problems of hypersensitivity or intolerance following its administration are not reasonably predictable.
Accordingly, another main aspect of the present invention regards pharmaceutical formulations comprising T3S as an active principle, as such or in combination with other thyroid hormones or pro-hormones. Particularly preferred are formulations comprising T3S in association with T4.
Said formulations differ in the dosage of the active principle or principles, or in the type of pharmaceutical form provided, depending on the desired administration kind. Moreover they can also contain useful additives like excipients, diluents, dissolvents, solvents, carriers, dyestuffs, flavourings, sweeteners commonly used in the pharmaceutical technology. The preparation of specific pharmaceutical formulations in response to particular needs of administration is plainly comprised in the general technical field of the present invention.
EXPERIMENTAL SECTION
As an example, absolutely non-limiting for the skilled technician, T3S may be administered for oral use at doses ranging from 5 to 1000 μg, preferably from 10 to 500 μg, more preferably from 25 to 250 μg.
Analogously, when in association with T4, preferred doses range from 10 to 500 μg for T3S and from 10 to 250 μg for T4, more preferably from 25 to 250 μg for T3S and from 25 to 200 μg for T4.
Two representative formulations for oral administration, selected among the preferred ones, are hereinafter enclosed by way of an example. Obviously, said formulations have no limiting effect on the other possible variations, which may also comprise different types of administration, different doses or different components depending on the specific pharmacological application or the particular pathology.
Example A
Oral Formulation Containing T3S

















T3S
50
μg;



Calcium phosphate dibasic anhydrous
103.5
mg;



Mais starch
17.65
mg;



Microcrystalline cellulose
5
mg;



Sodium carboxymethylamide
5
mg;



Talc
5
mg;



Citric acid
2.8
mg;



Magnesium stearate
1
mg










Example B
Oral Formulation Containing T3S and T4 

















T3S
50
μg;



T4 sodium salt
125
μg;



Calcium phosphate dibasic anhydrous
103.5
mg;



Mais starch
17.525
mg;



Microcrystalline cellulose
5
mg;



Sodium carboxymethylamide
5
mg;



Talc
5
mg;



Citric acid
2.8
mg;



Magnesium stearate
1
mg










In particular, when the association is taken into account, the formulations of the present invention will also possibly comprise individually formulated doses of T3S and T4, so that sequential administration is possible. In this case, one suitable kit is provided, which consents distinct administration of said active principles in ways that can differ from patient to patient, depending on the needed therapeutic application. In such a way, the specialized physician will have a wide choice of changing the prescription according to the actual need of the patient.
Just by way of an absolutely non-limitative example, in the case of oral administration, one package containing two individual blisters, which have different shape and/or color and/or different contents and/or doses, may suit the desired scope. Other possibilities exist and are easily available to the expert of the field.
The pharmaceutical compositions of the present invention are usable in the treatment of pathologies due to organic deficiency of triiodothyronine (T3), like, for example, original hypothyroidism from autoimmune thyroid affections, hormonal production defects, thyroidectomy, congenital hypothyroidism, as well as some disorders due to reduced activity of type I 5′-iodothyronine monodeiodinase (type I MD) which is induced, for example, by hypothyroidism, non thyroidal systemic illnesses, fast, selenium shortage and so on.
REFERENCES


    1. Chopra I J. Nature, source and relative biological significance of circulating thyroid hormones. In: Braverman L E., Utiger R D. (eds) The Thyroid, Lippincott, Philadelphia 1991, pp. 126-143.
    2. Spaulding S W., Smith T J., Hinkle P M., Davis F B., Kung M P., Roth J A. Studies on the biological activity of triiodothyronine sulfate. J. Clin. Endocrinol. Metab. 1992, 74, 1062-1067.
    3. Lo Presti J S., Mizuno L., Nimalysuria A., Anderson K P., Spencer C A., Nicoloff J T. Characteristics of 3,5,3′-triiodothyronine sulfate metabolism in euthyroid man. J. Clin. Endocrinol. Metab. 1991, 73, 703-709.
    4. Santini F., Hurd R E., Chopra I J. A study of metabolism of deaminated and sulfoconjugated iodothyronines by rat placental iodothyronine 5-monodeiodinase. Endocrinology 1992, 131, No. 4, 1689-1694.
    5. Otten M H., Mol J A., Visser T J. Sulfation proceding deiodination of iodothyronines in rat hepatocytes. Science 1983, 221, 81-83.
    6. Mol J A., Visser T J. Rapid and selective inner ring deiodination of T4 sulfate by rat liver deiodinase. Endocrinology 1986, 117, 8-12.
    7. Kung M P., Spaulding S W., Roth J A. Desulfation of 3,5,3′-triiodothyronine sulfate by microsomes from human and rat tissues. Endocrinology 1988, 122, 1195-1200.
    8. Santini F., Chopra I J., Wu S Y., Solomon D H., Chua Teco G N. Metabolism of 3,5,3′-triiodothyronine sulfate by tissues of the fetal rat: a consideration of the role of desulfation of 3,5,3′-triiodothyronine sulfate as a source of T3. Pediatr. Res. 1992, 31, 541-544.
    9. De Herder W W., Hazenberg M P., Pennock-Schroeder A M., Hennemann G., Visser T J. Rapid bacteria-dependent in vitro hydrolysis of iodothyronine conjugates by intestinal contents of humans and rats. Med. Biol. 1986, 64, 31-35.
    10. Santini F., Hurd R E., Lee B., Chopra I J. Thyromimetic effects of 3,5,3′-triiodothyronine sulfate in hypothyroid rats. Endocrinology 1993, 133, No. 1, 105-110.
    11. Santini F., Chiovato L., Ghiri P., Lapi P., Mammoli C., Montanelli L., Scartabelli G., Ceccarini G., Coccoli L., Chopra I J., Boldrini A., Pinchera A. Serum iodothyronines in human fetus and the newborn: evidence for an important role of placenta in fetal thyroid hormone homeostasis. J. Cl. Endocrinol. Metab. 1999, 84, No. 2, 493-498.
    12. Santini F., Pinchera A., Ceccarini G., Castagna M., Rosellini V., Mammoli C., Montanelli L., Zucchi V., Chopra I J., Chiovato L. Evidence for the role of the type III-iodothyronine deiodinase in the regulation of 3,5,3′-triiodothyronine content in the human central nervous system. Eur. J. Endocrinol. 2001, 144, 577-583.
    13. Santini F., Cortellazzi D., Baggiani A M., Marconi A M., Beck-Peccoz P., Chopra I J. A study of the serum 3,5,3′-triiodothyronine sulfate concentration in normal and hypothyroid fetuses at various gestational stages. J. Cl. Endocrinol. Metab. 1993, 76, No. 6, 1583-1587.
    14. Chopra I J., Nguyen D. Demonstration of thyromimetic effects of 3,5,3′-triiodothyronine sulfate (T3S) in Euthyroid rats. Thyroid 1996, 6, No. 3, 229-232.

## Claims

1. A solid composition comprising triiodothyronine sulfate_at a dose ranging from 5 to 1000 μg together with pharmaceutically acceptable additives for oral administration to a human as a thyroid hormone substitute therapy.

2. The composition according to claim 1, comprising triiodothyronine sulfate at a dose ranging from 10 to 500 μg.

3. The composition according to claim 2, further comprising from 10 to 250 μg of thyroxine.

4. The composition according to claim 2, comprising triiodothyronine sulfate at a dose ranging from 25 to 250 μg.

5. The composition according to claim 3, further comprising from 25 to 200 μg of thyroxine.

6. A kit comprising (i) a solid composition according to claim 2 and ((ii) a solid composition comprising an effective amount of thyroxine for oral administration to a human.

7. The kit according to claim 6, comprising from 10 to 500 μg of triiodothyronine sulfate and from 10 to 250 μg of thyroxine, in compositions (i) and (ii), respectively.

8. The kit according to claim 7, comprising from 25 to 250 μg of triiodothyronine sulfate and from 25 to 200 μg of thyroxine, in compositions (i) and (ii), respectively.

9. A method of treating a subject with a pathology due to organic deficiency of triiodothyronine comprising oral administration of a solid composition according to claim 1.

10. The method according to claim 9, wherein the triiodothyronine sulfate is administered at a dose ranging from 10 to 500 μg.

11. The method according to claim 10, wherein the triiodothyronine sulfate is administered at a dose ranging from 25 to 250 μg.

12. A method of treating a subject with a pathology due to organic deficiency of triiodothyronine comprising oral administration of a solid composition according to claim 3.

13. The method according to claim 12, wherein the triiodothyronine sulfate is administered at a dose ranging from 25 to 250 μg and the thyroxine is administered at a dose ranging from 25 to 200 μg.

14. The method according to any one of claim 9 or 12, wherein said pathology is selecetd from the group consisting of original hypothyroidism from autoimmune thyroid affections, hormonal production defects, thyroidectomy, and congenital hypothyroidism.

15. The method according to any one of claim 9 or 12, wherein said pathology is due to reduced activity of type I 5′-iodothyronine monodeiodinase.

16. The method according to claim 15, wherein said reduced activity of type I 5′-iodothyronine monodeiodinase is due to hypothyroidism, non thyroidal systemic illness, fast, or selenium shortage.