USE OF HISTONE DEACETYLASE INHIBITORS FOR THE CARE OF PHILADELPHIA-NEGATIVE MYELOPROLIFERATIVE SYNDROMES

Abstract

Method of treating Philadelphia-negative myeloproliferative syndromes in a patient suffering therefrom by administering to the patient diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride, or other pharmaceutically acceptable salts and/or solvates, in a daily dosage amount of from 50 to 100 mg per patient.

Claims

1. Method of treating Philadelphia-negative myeloproliferative syndromes in a patient suffering from said syndromes, said method comprising administering to said patient diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride, or other pharmaceutically acceptable salts and/or solvates, in a daily dosage amount of from 100 to 150 mg per patient.

2. Method according to claim 1, wherein said diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride is in monohydrate form.

3. Method according to claim 1, wherein said diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride is in monohydrate crystal form.

4. Method according to claim 1, wherein said daily dosage is 100 mg per patient.

5. Method according to claim 1, wherein said daily dosage is 150 mg per patient.

6. Method according to claim 1, wherein said Philadelphia-negative myeloproliferative syndrome is polycythemia vera.

7. Method according to claim 1, wherein said Philadelphia-negative myeloproliferative syndrome is essential thrombocythemia.

8. Method according to claim 1, wherein said Philadelphia-negative myeloproliferative syndrome is primary myelofibrosis.

9. Method according to claim 1, wherein said Philadelphia-negative myeloproliferative syndrome is secondary myelofibrosis.

10. Method according to claim 1, wherein said diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride, or other pharmaceutically acceptable salts and/or solvates, is not administered. in combination with other active principles.

11. Method according to claim 1, wherein said diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl-methyl]-ammonium chloride, or other pharmaceutically acceptable salts and/or solvates, is administered in combination with at least one other cytostatic active principle.

Description

DESCRIPTION OF THE INVENTION

[0019] We have now found, and this constitutes one aspect of the present invention, that the administration of diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl methyl]-ammonium chloride, preferably in monohydrate form, more preferably in monohydrate crystal form, to patients suffering from polycythemia vera, essential thrombocythemia or myelofibrosis causes a complete or partial response; in addition, in patients who initially demonstrated a state of splenomegaly, a significant reduction in the volume of the spleen is observed.

[0020] We have also found, and this constitutes a second aspect of the invention, that the therapeutic dose of such an active principle, for the treatment of Philadelphia-negative myeloproliferative syndromes in humans, is significantly lower than that normally used for the care of tumour syndromes and may be from 10 to 150 mg/day/patient, preferably from 30 to 120 mg/day/patient, and even more preferably from 50 to 100 mg/day/patient.

[0021] Purely by way of example, the dose of Zolinza™ (vorinostat) indicated for the treatment of cutaneous T-cell lymphoma is 400 mg/day.

[0022] The present invention relates to the use of diethyl-[6-(4-hydroxycarbamoyl-phenylcarbamoyloxymethyl)-naphthalen-2-yl methyl]-ammonium chloride, preferably in monohydrate form, more preferably in monohydrate crystal form for the therapeutic treatment of Philadelphia-negative myeloproliferative syndromes (polycythemia vera, essential thrombocythemia or idiopathic myelofibrosis). Such an active principle may be used alone, i.e. not in combination with other active principles, or in combination with other cytostatic active principles such as, purely by way of example, hydroxyurea or pipobroman.

[0023] The present invention relates also to the therapeutic use of such an active principle, for the treatment of Philadelphia-negative myeloproliferative syndromes, at daily doses lower than those used for the treatment of tumour pathologies (for example: cutaneous T-cell lymphoma), these doses being from 10 to 150 mg/day/patient, preferably from 50 to 120 mg/day/patient, and even more preferably from 50 to 100 mg/day/patient.

[0024] The term “treatment”, in relation to the present invention, is to be understood as meaning the action of caring for, relieving, mitigating, minimizing, eliminating or blocking the harmful effects resulting from the pathological state or the progression of the disease.

[0025] The inhibitory activity of a particular compound, with respect to histone deacetylases, may be measured in vitro using, for example, an enzyme test which demonstrates the inhibition of at least one of these enzymes. Tests of this type are known in the literature: see, for example, P. A. Marks et al. in J. Natl. Cancer Inst. 92 1210-1215 (2000); L. M. Butler et al. in Cancer Res. 60 5165-5170 (2000); V. M. Richon et al. in Proc. Natl. Acad. Sci. USA 95 3003-3007 (1998); M. Yoshida et al. in J. Biol. Chem. 265 17174-17179 (1990); kits tor measuring the HDAC inhibition are also available commercially (e.g. Biomol International LP (USA); HDAC1 Fluorimetric Drug Discovery Kit, product number BML-AK511-0001).

[0026] The following examples are intended to be illustrative of the invention rather than limiting the scope thereof.

EXAMPLE 1

[0027] Clinical study of ITF2357 in patients suffering from Philadelphia-negative myeloproliferative syndromes. Design: phase II, open, non-randomized study.

[0028] Population in the study: 27 patients with the JAK2.sup.V617F mutation suffering from myeloproliferative syndromes not adequately controlled by the standard pharmacological treatment (hydroxyurea).

[0029] Drug under study: ITF2357 at a dose of 50 mg twice per day.

[0030] Duration of the treatment: up to a maximum of 24 weeks of continual administration.

[0031] Primary objective: to evaluate the efficiency and tolerability of ITF2357 in the treatment of patients having JAK2.sup.V617F-positive myeloproliferative diseases (PV, ET, IM).

[0032] Secondary objective: to evaluate the effect of the treatment on the mutated JAK2 allele burden by quantitative PCR.

[0033] Examination of the preliminary data (duration of the treatment from 4 to 12 weeks) shows that the administration of ITF2357 to patients suffering from polycythemia vera, essential thrombocythemia or myelofibrosis induces a complete or partial response in approximately 60% of cases; in addition, a significant reduction in the volume of the spleen is observed in patients who initially exhibited a state of splenomegaly.

EXAMPLE 2

[0034] Comparative Effect on the Cloning Efficiency of the Human Cell Line SET-2

[0035] The JAK2.sup.V617F mutated, human cell line SET-2 (a megakaryoblastic cell line established from the peripheral blood of a patient with leukemic transformation of essential thrombocythemia) was obtained by the German cell bank DSMZ (catalogue number ACC 608) and grown in 24 well plates in culture medium (RPMI 1640+Hepes buffer 1M+Penicillin 10.000 IU/ml+Streptomycin 10.000 μg/ml+20% foetal calf serum). Previous experiments were carried out to determine the optimal number of cells giving a sufficient number of clones in each Petri dish to make a reproducible counting. In the case of the SET-2 line the number corresponded to 3×10.sup.3 cells/Petri dish. When a sufficient number of cells were obtained, the cells were harvested by centrifugation (10 min at 10.sup.3 rpm) and then suspended in culture medium at the concentration of 90×10.sup.3 cells/ml. 100 μl of the suspension were added to 3.0 ml of methyl-cellulose (Methocult™, catalogue number H4230, Stemcell Technologies) prepared as described by manufacturer and, then, 50 μl of 63 fold concentrated solution of the HDAC inhibitor in 0.1% DMSO was added. For each compound 5 serial dilutions (1000-12 nM range) were tested. The methyl-cellulose solution containing the HDAC inhibitor was then carefully mixed using a sterile plastic Pasteur pipette avoiding the formation of bubbles. At the end 1 ml of the solution was homogeneously dispensed in a Petri dish (35 mm diameter with grid, catalogue number 174926 Nunc) using a syringe with a 18 G needle. For each experimental point 2 Petri dishes were done. The 2 experimental dishes were put in an larger Petri dish along with a reservoir of sterile water to ensure a constant humidity. All the dishes were then maintained at 37° C. in a humidity and CO.sub.2-controlled sterile incubator. After 14 days of growing the number of clones in each dish was determined by using a binocular microscope. The average number of clones for each coupe of dishes was calculated and the percentage inhibition of clones formation was determined in respect to the number of clones obtained in the absence of any compounds (control dishes). The EC.sub.50 value (concentration required to reduce of 50% the number of clones) was calculated using GraphPad Prism 5.0 software and reported in the table below. Values for rhHDAC1 inhibition are also reported: the enzyme was obtained from BPS Biosciences (cat n. 50001) and the test was carried out using a BIOMOL kit, according to supplier instructions.

TABLE-US-00001 TABLE 1 IC.sub.50 nM Code Originator INN Chemical Class EC.sub.50 μM (rhHDAC1) ITF2357 Italfarmaco Givinostat Hydroxamic acid 0.028-0.054* 121 MGCD- MethylGene Mocetinostat Benzamide 1.196** 17 0103 KD- Kalypsys — S,Acetyl-α-mercaptoketone <20% inhib. at 24 5170 1 μM SNDX- Bayer Entinostat Benzamide 1.820** 293 275 Schering *values obtained in two different experiment **p < 0.05 towards ITF2357 in the same experiment

[0036] Although Mocetinostat and Entinostat are considered as class I specific HDAC inhibitors [Zhou, N. et al. J Med Chem 51, 4072 (2008) and Jones, P. et al. J Med Chem 51, 2350 (2008)], KD-5170 has been reported to be a broad spectrum Class I and II-HDAC inhibitor [WO 2007/067795; Payne, J. E. et al. Bioorg Med Chem Lett 18, 6093 (2008)], the same as ITF2357.

[0037] Although, the inhibition of cloning efficiency of SET-2 cells is not proven to be predictive for the cure of the Philadelphia-negative, JAK2.sup.V617F-positive, myeloproliferative syndromes, the potent inhibitory effect of ITF2357, if compared to the other HDAC inhibitors, seems to indicate that the presence of histone deacetylase inhibition is not essential for obtaining the clinical efficacy shown by ITF2357 with respect to Philadelphia-negative myeloproliferative syndromes.

[0038] The tested compounds correspond to the following formulae:

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