Crystalline form of triethylenetetramine tetrahydrochloride and its pharmaceutical use

Abstract

To provide a novel crystalline form of triethylenetetramine tetrachloride which has improved room temperature stability over known forms and over the dichloride.SOLUTION: The novel crystalline form has peaks at 22.9, 25.4, 25.8, 26.6, 34.6, and 35.3±0.1°2θ in an XRPD spectrum, and Raman shifts at 943, 1173, 1527, and 1612±5 cm-1. The crystalline form of triethylenetetramine tetrachloride is useful in the treatment of Wilson's disease. There is also provided a method for producing a crystalline form of triethylenetetramine tetrachloride, the method comprising adding an antisolvent to an aqueous solution of the compound.SELECTED DRAWING: None

Description

[Technical field]

[0001] FIELD OF THEINVENTION The present invention relates to crystalline forms of triethylenetetramine tetrachloride (TETA.4HCl) and methods of making said crystalline forms. The present invention further relates to the treatment of Wilson's disease using crystalline triethylenetetramine tetrachloride. [Background technology]

[0002] 2. Background of the Invention Triethylenetetramine, or 1,2-ethanediamine, N,N'-bis(2-aminoethyl) (TETA) is:

[0003] [ka]

[0004] It has the structure: The dichloride salt (TETA.2HCl) is a polyamine chelator of copper(II). Its copper chelating properties make it useful for the treatment of various conditions, especially Wilson's disease. Wilson's disease is a genetic disorder caused by a mutation in the Wilson's disease protein (ATP7B gene). The condition leads to copper accumulation in the body. The copper chelating ability of TETA.2HCl also leads to its consideration for the treatment of many conditions such as organ damage in diabetic patients, Alzheimer's disease, and cancer (Henriet et al, International Journal of Pharmaceutics 511 (2016) 312-321).

[0005] However, even though TETA.2HCl has been known for many years to be useful in treating Wilson's disease, it has not been a successful treatment. This is, at least in part, because it has proven difficult to provide a suitable form of TETA.2HCl that has sufficient stability at room temperature. Thus, patients must store tablets under cold conditions, a onerous requirement for a treatment that must be taken with every meal for the rest of their lives.

[0006] Studies have also shown that changes in humidity can affect the stability of the salts. Salts are very sensitive to water and exist in different polymorphs depending on the humidity level. High humidity results in instability of the compound. These stability effects lead to challenges in formulating suitable drugs for the treatment of patients, and require storage of materials under special conditions, such as low temperatures. Therefore, there is a need for improved treatments for Wilson's disease that can be delivered orally and are stable under ambient conditions for extended periods of time.

[0007] EP 1778618 describes synthetic techniques for making TETA and its salts, including the diHCl and tetraHCl salts, of which only the diHCl salt is said to be useful in treating Wilson's disease.

[0008] WO 2006 / 027705 describes the synthesis of triethylenetetramine, including triethylenetetramine dihydrochloride form I and form II. The document does not mention the crystalline form of triethylenetetramine tetrahydrochloride. Summary of the Invention

[0009] Summary of the Invention The inventors have surprisingly found that the new crystalline form of TETA.4HCl has improved handling and room temperature stability. It is therefore more useful for formulation into drugs than the known forms of the dichloride or tetrachloride salts. Previously known techniques for producing TETA.4HCl, such as an anti-solvent crystallization process carried out at room temperature and a process involving a high temperature drying step, lead to the crystalline form described herein as Form A. However, the inventors have found that by carefully controlling the manufacturing conditions, particularly the temperature and rate of crystallization, a new crystalline form, identified herein as Form B, can be produced. This new form has good handling and good stability and shelf life properties, making it beneficial for the manufacture of new formulations, such as tablets, for treating Wilson's disease.

[0010] The present invention therefore provides a method for producing a computer-implemented computer-readable recording medium comprising the steps of: (i) an XRPD pattern having at least two peaks selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm -1 A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of The present invention provides a crystalline form of triethylenetetramine tetrachloride having at least one of the following:

[0011] Also provided are pharmaceutical compositions comprising the crystalline forms described herein, together with one or more pharma- ceutically acceptable carriers or diluents.

[0012] Also provided is a method for preparing a crystalline form of triethylenetetramine tetrachloride, comprising adding an anti-solvent to an aqueous solution of triethylenetetramine tetrachloride and recovering the resulting crystals, wherein the addition of the anti-solvent is carried out at a temperature of about 20° C. or less.

[0013] Also provided are crystalline forms of triethylenetetramine tetrachloride, or pharmaceutical compositions containing triethylenetetramine tetrachloride obtained or obtained by the methods described herein.

[0014] There is also provided a crystalline form or pharmaceutical composition as described herein for use in the treatment of the human or animal body by therapy, preferably for use in the prevention or treatment of Wilson's disease.

[0015] Also provided is a method for the prevention or treatment of Wilson's disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a crystalline form or pharmaceutical composition described herein.

[0016] There is also provided the use of a crystalline form or pharmaceutical composition described herein in the manufacture of a medicament for the prevention or treatment of Wilson's disease.

[0017] Specific aspects of the present invention are set out below: 1. Features include: (i) an XRPD pattern having at least two peaks selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm -1 A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of A crystalline form of triethylenetetramine tetrachloride having at least one of the following:

[0018] 2. The crystalline form of embodiment 1 having an XRPD pattern having at least two peaks selected from 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ.

[0019] 3. The crystalline form of embodiment 1 or embodiment 2 having an XRPD pattern with at least three peaks selected from 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ.

[0020] 4. The crystalline form of any one of embodiments 1-3 having an XRPD pattern with peaks at 25.4, 34.6, and 35.3±0.1 degrees 2θ.

[0021] 5. A crystalline form according to any one of embodiments 1-4 containing 10% by weight or less of triethylenetetramine tetrachloride form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

[0022] 6. The crystalline form of embodiment 5 containing 5% or less by weight of triethylenetetramine tetrachloride form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

[0023] 7. In essence (i) an XRPD pattern as defined in any one of claims 1 to 4; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm -1 A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of The crystalline form according to any one of the preceding embodiments comprises form B of triethylenetetramine tetrachloride having the formula:

[0024] 8. A pharmaceutical composition comprising the crystalline form of any one of embodiments 1-7 and a pharma- ceutically acceptable carrier or diluent.

[0025] 9. The pharmaceutical composition of embodiment 8, which is a solid oral dosage form comprising the crystalline form of any one of embodiments 1-7 and a pharma- ceutically acceptable carrier.

[0026] 10. The pharmaceutical composition of embodiment 8 or embodiment 9, comprising 10% by weight or less, preferably 5% by weight or less, and more preferably 2% by weight or less of triethylenetetramine tetrachloride Form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

[0027] 11. The pharmaceutical composition of embodiment 10, wherein the composition is substantially free of triethylenetetramine tetrachloride Form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

[0028] 12. A method for preparing a crystalline form of triethylenetetramine tetrachloride comprising adding an anti-solvent to an aqueous solution of triethylenetetramine tetrachloride and recovering the resulting crystals, wherein the addition of the anti-solvent is conducted at a temperature of about 20° C. or less.

[0029] 13. The method of claim 12, wherein the rate of addition of the anti-solvent to the solution is 0.5 ml / min or less per gram of TETA.4HCl dissolved in the aqueous solution.

[0030] 14. (i) adding an anti-solvent to an aqueous solution of TETA.4HCl at a temperature T1, for a period of time t1, and / or at an addition rate R1; (ii) optionally, adding seed crystals of TETA.4HCl; (iii) optionally, stirring the resulting mixture at T1 for a further period of time t1a; (iv) optionally, reducing the temperature to a temperature T2 and agitating the mixture for a further period of time t2; and (v) recovering the resulting crystals. wherein T1 is about 20° C. or less; T2 is at least 5° C. lower than T1; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 2 hours, and t2 is at least 30 minutes.

[0031] 15. (i) adding an antisolvent to an aqueous solution of TETA.4HCl at a temperature T1, for a period of time t1, and / or at a rate R1; (ii) optionally, adding seed crystals of TETA.4HCl; (iii) optionally, stirring the resulting mixture at T1 for a further period of time t1a; (iv) reducing the temperature to a temperature T2 and stirring the mixture for a further period of time t2; and (v) recovering the resulting crystals. 15. The method of embodiment 14, wherein T1 is about 20° C. or less; T2 is about 10° C. or less; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 3 hours, and t2 is at least 30 minutes.

[0032] 16. (i) adding an antisolvent to an aqueous solution of TETA.4HCl at a temperature T1, for a period of time t1, and / or at a rate R1; (ii) adding seed crystals of TETA.4HCl; (iii) stirring the resulting mixture at T1 for a further period t1a; (iv) reducing the temperature to a temperature T2 and stirring the mixture for a further period of time t2; and (v) recovering the resulting crystals. 16. The method of embodiment 15, wherein T1 is about 15° C. or less; T2 is about 5° C. or less; t1 is at least 1 hour, R1 is 0.2 ml / min / g or less, t1a is at least 4 hours, and t2 is at least 30 minutes.

[0033] 17. The method of any one of aspects 12-16, further comprising drying the recovered crystals at a temperature of less than about 40° C., preferably less than about 30° C.

[0034] 18. The method of any one of aspects 12 to 17, wherein the recovered crystals are combined with a pharma- ceutically acceptable carrier to produce a pharmaceutical composition.

[0035] 19. The method of embodiment 18, further comprising compressing the mixture of the crystals and a pharma- ceutically acceptable carrier to form a tablet, and optionally sugar-coating or film-coating the tablet.

[0036] 20. A crystalline form or pharmaceutical composition obtained by the method of any one of aspects 12 to 19.

[0037] 21. A crystalline form or pharmaceutical composition according to any one of the preceding embodiments for use in a method of treatment of the human or animal body by therapy.

[0038] 22. The crystalline form or pharmaceutical composition for use according to embodiment 21 for use in the prevention or treatment of Wilson's disease. [Brief description of the drawings]

[0039] [Figure 1] 1a and 1b are flow charts illustrating a method for producing TETA.4HCl in crude form. [Diagram 2] FIG. 2 is a flow chart illustrating a method for preparing Form B of TETA.4HCl in substantially pure form. [Diagram 3] 3 depicts the X-ray powder diffraction patterns of TETA.4HCl form A ("Profile 1") and a mixture of TETA.4HCl forms A and B ("Profile 1+2"). Arrows highlight the characteristic peaks of TETA.4HCl form B. [Figure 4] FIG. 4 depicts the X-ray diffraction pattern of form B of TETA.4HCl. [Diagram 5] Figure 5a shows the Raman spectra of Form A (bottom) and Form B (top) of TETA.4HCl. Figure 5b shows the spectra overlaid with Form A on the top line and Form B on the bottom line. The peaks of Form B are highlighted. [Figure 6] FIG. 6 depicts a Heckel plot of TETA.4HCl form A and TETA.4HCl form B. [Figure 7] Figure 7a shows a tablet of TETA.4HCl formed from TETA.4HCl form A. Figure 7b shows an image of the same tablet after aging. Figure 7c shows a Raman analysis of the aged tablet, with the dark areas corresponding to the location of TETA.4HCl form A within the tablet. [Figure 8]Figures 8a and 8b show the FTIR-ATR spectra of TETA.4HCl form B. Figure 8a shows the full spectrum from 4000 to 525 cm-1. Figure 8b shows the fingerprint region from 1800 to 525 cm-1. [Figure 9] Figure 9a shows the X-ray powder diffraction analysis of a sample of Example 7 after manual addition of ethanol at different rates and comparison with reference patterns of Forms A and B. Figure 9b shows a zoomed-in view of Figure 9a at 2θ=20-29°. Figure 9c shows a zoomed-in view of Figure 9a at 2θ=34-37°. [Figure 10] Figure 10a shows an X-ray powder diffraction analysis of a sample of Example 7 after programmed addition of ethanol and a comparison with reference patterns for Forms A and B. Figure 10b shows a zoomed-in view of Figure 10a from 2θ=20-29°. Figure 10c shows a zoomed-in view of Figure 10a from 2θ=34-37°. [Figure 11] 11a and 11b show the mass change versus relative humidity in the DVS analysis of Example 2 and Reference Example 3, respectively. [Figure 12] 12a-12c show XRPD spectral analyses of products made according to the prior art process. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0040] Detailed Description of the Invention Crystalline TETA.4HCl The crystalline form of triethylenetetramine tetrachloride (TETA.4HCl) described herein is identified as Form B. This crystalline form can be characterized by one or more of its XRPD spectrum, its Raman spectrum, its melting point, its FTIR spectrum and its DVS behavior. Details of each of these characteristics of the crystalline form are described below. Typically, the crystalline form of the present invention is characterized by its XRPD spectrum and / or its Raman spectrum, most preferably its XRPD spectrum. Thus, the crystalline form of the present invention typically has the following characteristics: (i) an X-ray powder diffraction (XRPD) pattern having at least two peaks selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6, and 35.3±0.1 degrees 2θ; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm -1 A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of The present invention has at least one of the following:

[0041] Typically, the crystalline form of TETA.4HCl of the present invention has an XRPD pattern with at least two peaks selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ. The XRPD pattern preferably has at least three peaks, more preferably at least four peaks, selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ. More preferably, at least five or all of these peaks are observed in the XRPD pattern. More preferably, the crystalline form of TETA.4HCl of the present invention has an XRPD pattern with at least two peaks, preferably at least three, four, five or all of the peaks, selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.05 degrees 2θ. It is particularly preferred that said crystalline form of TETA.4HCl has an XRPD pattern with peaks at 25.4, 34.6 and 35.3±0.1 degrees 2θ, more preferably 25.4, 34.6 and 35.3±0.05 degrees 2θ.

[0042] Typically, the peaks at 25.4 and 35.3±0.1°2θ are the most intense, especially the peak at 25.4±0.1°2θ. The peak at 25.4±0.1°2θ is preferably at least twice, more preferably at least three times as intense as the next most intense peak. Typically, the peak at 35.3±0.1°2θ is at least twice as intense as the next most intense peak.

[0043] Typically, the XRPD pattern of form B of TETA.4HCl is substantially the same as that depicted in FIG.

[0044] XRPD data can be obtained using a PANALYTICAL X'PERT PRO MPD diffractometer. Diffraction data is typically obtained by diffracting powder samples with a Cu-K α XRPD data is obtained by exposure to X-ray radiation. The X-rays were generated from a Cu anode supplied with 40 kV and 40 mA current. Details of the operating conditions for obtaining the XRPD data are given in the Examples section herein.

[0045] Typically, the crystalline form of TETA.4HCl of the present invention has peaks at 943, 1173, 1527 and 1612±5 cm -1 The Raman spectrum has shifts at two or more of 943, 1173, 1527 and 1612±5 cm -1 Preferably, the crystalline form of TETA.4HCl exhibits at least two, preferably three, and more preferably all four peaks at 943, 1173, 1527 and 1612±2 cm. -1 It is particularly preferred that the crystalline form of TETA.4HCI has a Raman spectrum with shifts at two or more, preferably three, and more preferably all four of the following: 943 and 1173±5 cm -1 , most preferably 943 and 1173±2 cm -1 It is particularly preferred to have a Raman spectrum with a shift in the Raman spectrum from 0 to 100. Typically, the Raman spectrum is similar to that shown in Figure 5a (top spectrum).

[0046] Raman spectra can be obtained, for example, using a Renishaw RA802 Pharmaceutical Analyser, which operates at a laser wavelength of 785 nm. Further operating conditions are described in the Examples section herein.

[0047] The crystalline form of Form B of TETA.4HCl is storage stable. Thus, the XRPD pattern and / or Raman spectrum of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months, is typically identical or substantially identical to that of the crystalline form of the present invention described above. Preferably, at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months, retains the crystalline form, Form B, described herein.

[0048] The crystalline form of Form B of TETA.4HCl is stable in a humid environment. Thus, the XRPD pattern and / or Raman spectrum of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month, preferably 4 months, more preferably 6 months, is typically the same or substantially the same as that of the crystalline form of the present invention described above. Preferably, at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month is the crystalline form, Form B, described herein. Preferably, at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 4 months, preferably 6 months, retains the crystalline form, Form B, described herein.

[0049] Preferably, the storage stability of the crystalline form of the present invention is determined by the XRPD pattern. Thus, the XRPD pattern of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months is preferably identical or substantially identical to that of the crystalline form of the present invention described above. Preferably, at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months retains the same or substantially identical XRPD pattern of the crystalline form, Form B, described herein. Furthermore, the XRPD pattern of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month, preferably 4 months, more preferably 6 months is preferably identical or substantially identical to that of the crystalline form of the present invention described herein. Preferably, at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month, preferably 4 months, more preferably 6 months retains the same or substantially identical XRPD pattern of the crystalline form, Form B, described herein.

[0050] Instead, the storage stability of the crystalline form of the present invention is determined by Raman spectrum. Thus, the Raman spectrum of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months is preferably identical or substantially identical to the Raman spectrum of the crystalline form of the present invention described above. Preferably at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 20° C. for 6 months, preferably 10 months, more preferably 12 months retains the same or substantially identical XRPD pattern as that of the crystalline form, Form B, described herein. Furthermore, the Raman spectrum of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month, preferably 4 months, more preferably 6 months is preferably identical or substantially identical to that of the crystalline form of the present invention described herein. Preferably at least 90% by weight, more preferably at least 95% by weight, more preferably at least 98% by weight of a sample of the crystalline form of the present invention stored at 40° C. and 75% humidity for 1 month, preferably 4 months, more preferably 6 months retains the same or substantially identical Raman spectrum as that of the crystalline form, Form B, described herein.

[0051] A particular advantage of the crystalline forms of the present invention relates to their storage stability. Upon storage of tablets derived from form A of TETA.4HCl, discolored patches are observed after 6 months of storage at 40° C. and 75% humidity. A tablet derived from aged form A of TETA.4HCl is shown in FIG. 7b, which shows the discoloration of the tablet over time. The present invention, and the provision of form B of TETA.4HCl, and in particular substantially pure form B of TETA.4HCl, aims to address this issue. Tablets derived from form B of TETA.4HCl are believed to have a reduced tendency to discolor over time.

[0052] The crystalline forms of the present invention typically have two or more, preferably four or more, more preferably five or six or more, and most preferably all of the 1475, 1525, 16010, 2380, 2435, 2580, 2830 and 2880±5 cm-1 The crystalline form of the present invention preferably has an FTIR spectrum with peaks at 1525, 2435 and 2675±5 cm -1 , and most preferably 1526, 2436 and 2674±2 cm -1 The crystalline form of the present invention has an FTIR spectrum with a peak at 943±2 cm -1 Preferably, the crystalline form having a peak at 943±2 cm is present in an amount of 50% by weight or less, for example 40% by weight or less, preferably 20% by weight or less, more preferably 10% by weight or less. Most preferably, the crystalline form has a peak at 943±2 cm -1 The crystal form is substantially free of a crystal form having a peak at

[0053] FTIR spectra are typically FTIR-ATR spectra and can be obtained using a Nicolet iS5 FT-IR spectrometer in ATR diamond mode. Specific conditions suitable for obtaining FTIR spectra are set forth in further detail in Example 4.

[0054] The crystalline form of the present invention typically has a melting temperature, as measured by DSC, of ​​about 260° C., typically about 259° C. DSC analysis can be performed as described in Example 4. For example, the analysis can be performed using a Toledo DSC3+ device under a 50 mL / min nitrogen flush, providing samples in 40 μL sealed aluminum pans with holes in the lids prior to analysis.

[0055] Analysis of the crystalline forms of the present invention by DVS can also be used to distinguish crystalline form B of the present invention from form A. The crystalline forms of the present invention typically exhibit a weight gain of 50-59% above 90% RH, typically 54-57%. After completion of the sorption and desorption cycle (0%-95% RH), the weight gain of the sample is typically 10% or less, preferably 5% or less. This is in contrast to form A of TETA.4HCl, which exhibits a weight gain of 14-15% after sorption / desorption (0-95% RH).

[0056] Crystalline forms of TETA.4HCl of the present invention typically contain 50% or less by weight, such as 40% or less by weight, preferably 20% or less by weight, and more preferably 10% or less by weight of TETA.4HCl Form A. Preferred crystal forms of TETA.4HCl of the present invention are substantially free of TETA.4HCl Form A. As used herein, free of Form A means that the crystal form contains 5% or less by weight of Form A, preferably 2% or less by weight, more preferably 1% or less by weight, and most preferably 0.5% or less by weight, or 0.1% or less by weight of Form A.

[0057] Form A of TETA.4HCl is a crystalline form obtained under standard crystallization conditions, such as those described in Reference Example 3 herein. Form A is characterized by an XRPD pattern with peaks at 25.2 and 35.7±0.1°2θ, typically at 25.2 and 35.7±0.05°2θ. The XRPD spectrum of Form A preferably also has peaks at 21.8, 26.9 and 28.2±0.1°2θ, typically at 21.8, 26.9 and 28.2±0.05°2θ. In particular, Form A is characterized by the XRPD pattern shown in FIG. 3 ("Profile 1"). Form A also has peaks at 933 and / or 1513±5 cm -1 , typically 933 and / or 1513 ± 2 cm -1 Form A may also be characterized by a Raman spectrum having peaks at 933, 1167, 1513 and 1604±5 cm. -1 Typically 933, 1167, 1513 and 1604±2cm -1 Form A is characterized by a Raman spectrum having a peak at 100° C. Typically, Form A is characterized by a Raman spectrum as shown in FIG. 5a herein (lower spectrum).

[0058] The crystalline form of the present invention preferably contains 50% or less by weight, such as 40% or less, preferably 20% or less, more preferably 10% or less, more preferably 5% or less, 2% or less, 1% or less, and most preferably 0.5% or less or 0.1% or less, of a crystalline form of TETA.4HCl having an XRPD pattern having peaks at 25.2 and 35.7±0.1 degrees 2θ, or having peaks at 21.8, 25.2, 26.9, 28.2 and 35.7±0.1 degrees 2θ.

[0059] The crystalline form of the present invention has a molecular weight of 933 and / or 1513±5 cm -1 Typically 933 and / or 1513 ± 2 cm -1 or peaks at 933, 1167, 1513 and 1604 cm -1 ±5cm -1 Typically 933, 1167, 1513 and 1604±2cm -1 Preferably, the crystalline form of TETA.4HCl having a Raman spectrum with a peak at 0.05% or less, such as 0.05% or less, for example 0.05% or less, preferably 0.05% or less, more preferably 0.05% or less, more preferably 0.05% or less, more preferably 0.05% or less, and most preferably 0.05% or less.

[0060] Preferably, the crystalline form of TETA.4HCl comprises at least 90% by weight of form B. More preferably, the crystalline form consists essentially of form B, i.e., it is substantially pure form B of TETA.4HCl. When the crystalline form consists essentially of form B, it typically comprises at least 95% by weight of form B of TETA.4HCl, more preferably at least 98% by weight, more preferably at least 99% by weight, and most preferably at least 99.5% or 99.9% by weight of form B of TETA.4HCl, where form B of TETA.3HCl is characterized by an XRPD spectrum and / or Raman spectrum described herein, and preferably form B of TETA.4HCl is characterized by an XRPD spectrum described herein.

[0061] The crystals of TETA.4HCl described herein are typically provided in dry form. Thus, they typically contain less than 1% by weight of water, preferably less than 0.5% by weight of water, more preferably less than 0.1% by weight or less than 0.05% by weight of water. Any residual solvent is preferably less than 0.1% by weight, more preferably less than 0.5% by weight.

[0062] Method for producing crystalline TETA.4HCl TETA.4HCl can be prepared by techniques known in the art. For example, TETA free base is commercially available and can be converted to crystalline TETA hydrate and isolated by conventional methods. TETA hydrate can be treated with aqueous HCl to obtain TETA.4HCl salt. Typically, TETA.4HCl salt is isolated in crude form before recrystallization as the Form B polymorph.

[0063] Crystalline forms of TETA.4HCl can be obtained by anti-solvent crystallization processes, typically from aqueous solutions. Such processes involve adding an anti-solvent to an aqueous solution of TETA.4HCl and recovering the resulting crystals. When carried out under standard crystallization conditions, for example, by crystallization at or above room temperature and / or by methods involving drying at elevated temperatures, such methods have been found to lead to a single crystalline form of TETA.4HCl, identified herein as Form A. Crystals of Form A have also been obtained with varying solvent systems.

[0064] For example, the inventors have prepared TETA.4HCl using the methods described in WO 2006 / 027705 and found that these methods lead to the production of crystals of Form A. The inventors have reproduced Example 17 of WO 2006 / 027705, starting from a mixture of triethylenetetramine isomers and using the crystallization conditions described in Example 17 of WO 2006 / 027705. The product was analyzed by XRPD, the results of which are shown in Figures 12a-12c. The resulting product contained peaks characteristic of Form A of TETA.4HCl. However, certain peaks identified as characteristic of Form B of TETA.4HCl, particularly those around 35° 2θ and 25.4° 2θ, were absent, suggesting that the product produced is Form A of TETA.4HCl, which form is identified as produced by standard room temperature crystallization.

[0065] The inventors have found that by using the same solvent system but varying the crystallization conditions, particularly the time and temperature of the process, crystals of Form B can be obtained.

[0066] At temperatures below about 20° C., especially below about 15° C., TETA.4HCl may be produced as Form B. At about 20° C.-30° C., the crystalline form produced may depend on conditions other than just the temperature of crystallization. Thus, above about 20° C., further conditions must typically be controlled to ensure production of Form B. In particular, the crystalline form produced may depend on the rate of crystallization. Thus, slow crystallization favors the formation of Form B, while faster crystallization favors Form A. Even at temperatures between 15-20° C., some crystals of Form A may be produced, unless crystallization is performed slowly. For example, if the addition of an antisolvent is used to form crystals, the antisolvent should preferably be added slowly to the solution to ensure that Form B is produced, and not Form A.

[0067] The crystalline forms of the present invention are typically prepared by crystallization at temperatures below about 20° C., preferably below about 15° C., and more preferably below about 10° C. In one aspect, the preferred temperature for crystallization is below 13° C., more preferably between 7 and 13° C. At temperatures below about 15° C., and particularly below 13° C., Form B is the thermodynamically favored form, and crystallization will generally result in substantially pure Form B.

[0068] All steps of the crystallization process are preferably carried out below 30°C, preferably below about 20°C, preferably below about 15°C, more preferably below about 10°C. If the temperature is above about 15°C, a mixture of Form A and Form B may be produced. If the temperature is above about 30°C, only Form A will result. The temperature is preferably maintained below about 15°C at all times during crystallization to ensure that the product produced is substantially pure Form B. At temperatures between about 15°C and 20°C, crystals of Form B can be produced by conducting the crystallization at a slow rate of anti-solvent addition. In particular, the addition of Form B seeds in combination with slow solvent addition promotes the formation of substantially pure Form B. The addition of anti-solvent in a slow and controlled manner ensures that the crystallization develops from the seeds and that separate nucleation of Form A does not occur.

[0069] Typically, crystallization is carried out by adding antisolvent at a rate of 0.5 ml / min or less of antisolvent added to an aqueous solution of TETA.4HCl per gram of TETA.4HCl dissolved in the aqueous solution. Thus, the preferred rate of addition is 0.5 ml of antisolvent or less per gram of TETA.4HCl per minute, i.e., 0.5 ml / min / g or less. The preferred rate of antisolvent addition is 0.2 ml / min / g or less, more preferably about 0.1 ml / min / g or less. The preferred rate of addition is 0.01-0.2 ml / min / g, most preferably 0.01-0.1 ml / min / g.

[0070] Thus, to provide substantially pure Form B crystals, the crystallization is preferably carried out at about 15° C. or below, and at an addition rate of preferably 5 ml / min / g or below, more preferably 0.2 ml / min / g or below, e.g., about 0.1 ml / min / g. Most preferably, the crystallization is carried out at 13° C. or below, e.g., 7-13° C., and at an addition rate of preferably less than 0.2 ml / min / g, e.g., about 0.1 ml / min / g or below.

[0071] Seed crystals of Form B of TETA.4HCl are preferably added. Seed crystals may be added before, during, or after the antisolvent addition, typically before or during the antisolvent addition, most preferably before the antisolvent addition. If seeds are added during or after the antisolvent addition, they are preferably added before crystal formation is observed.

[0072] A preferred method of crystallization uses TETA.4HCl, preferably purified TETA.4HCl, as the starting material. The presence of impurities in the starting material can affect the ability to crystallize the desired polymorph. Thus, the TETA.4HCl is preferably in isolated form, i.e., it is isolated (and typically purified) from any reaction mixture in which it was produced before crystallization to produce Form B is initiated. Additionally, crude TETA.4HCl is preferably recrystallized before the process to produce Form B is initiated. This also provides a higher purity starting material, allowing for the reliable production of crystals of Form B by following the methods described herein.

[0073] TETA.4HCl is typically dissolved in an aqueous solution prior to crystallization. The solution typically contains 0.01-10 g TETA.4HCl per ml of water. The solution preferably contains 0.1-5 g TETA.4HCl per ml of water, most preferably 0.6-1.2 g TETA.4HCl per ml of water. The volume of anti-solvent used for recrystallization is typically 0.5 ml or less per gram of TETA.4HCl dissolved in the aqueous solution. The preferred amount of anti-solvent is 0.2 ml / g TETA.4HCl or less, more preferably about 0.1 ml / g TETA.4HCl or less. The preferred amount of anti-solvent is 0.01-0.2 ml / g TETA.4HCl, most preferably 0.01-0.1 ml / g TETA.4HCl.

[0074] A preferred method for crystallization of Form B is: (i) adding an anti-solvent to an aqueous solution, preferably a stirred aqueous solution, of TETA.4HCl at a temperature T1, for a period of time t1, and / or at an addition rate R1; (ii) optionally, adding seed crystals of TETA.4HCl; (iii) optionally, stirring the resulting mixture at T1 for a further period of time t1a; (iv) optionally, reducing the temperature to a temperature T2 and agitating the mixture for a further period of time t2; and (v) recovering the resulting crystals. Includes.

[0075] T1 is about 20° C. or less, preferably about 15° C. or less, more preferably about 10° C. or less. To produce substantially pure Form B, T1 is preferably about 15° C. or less, more preferably about 10° C. or less.

[0076] The anti-solvent can be any solvent in which TETA.4HCl is substantially insoluble. Suitable anti-solvents include ethanol, methanol, acetonitrile, propan-2-ol, acetone, and 1,4-dioxane, and mixtures thereof. Methanol and ethanol, and mixtures thereof, are preferred, especially ethanol.

[0077] The solution is typically agitated or mixed, typically by stirring, during the addition. Further agitation of the mixture, such as stirring, may be applied between steps (iii) and (iv).

[0078] The crystallization process is typically carried out for a long period of time. Thus, t1 is typically at least 1 hour, preferably at least 1.5 hours. The addition rate R1 of the antisolvent is typically 0.5 ml / min / g or less. A preferred rate R1 is 0.2 ml / min / g or less, more preferably about 0.1 ml / min / g or less, for example 0.01-0.2 ml / min / g, most preferably 0.01-0.1 ml / min / g.

[0079] If seeds are added, they are typically added before, during or after carrying out step (i). Preferably, seeds are added before step (i) or during the addition of the antisolvent in step (i). If seeds are added during or after the addition of the antisolvent, the mixture is typically stirred at t1a for an additional period of time, preferably at least 2 hours, such as at least 3 or 4 hours, for example about 5 hours. Preferably, an additional stirring period at a lower temperature is also included before recovering the crystals. This additional stirring step is carried out for a period of time t2, preferably at least 30 minutes. The additional stirring step is carried out at a low temperature, T2. T2 is typically lower than T1, and preferably at least 5°C, more preferably at least 10°C, lower than T1. T2 is typically about 10°C or less, preferably about 5°C or less, more preferably about 0°C. It has been found that increasing the time or rate of crystallization and decreasing the temperature of crystallization provide higher purity of Form B. The introduction of seeds also serves to improve the purity of the crystalline form of Form B.

[0080] In the above method, typically T1 is about 20°C or less; T2 is at least 5°C lower than T1; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 2 hours, and t2 is at least 30 minutes. Preferably, T1 is about 20°C or less; T2 is about 10°C or less; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 3 hours, and t2 is at least 30 minutes. More preferably, T1 is about 15°C or less; T2 is about 5°C or less; t1 is at least 1 hour, R1 is 0.2 ml / min / g or less, t1a is at least 4 hours, and t2 is at least 30 minutes. The crystallization is preferably carried out under an inert atmosphere, for example under nitrogen.

[0081] A preferred crystallization method includes at least steps (i), (iv) and (v) above. A more preferred method includes steps (i), (iii), (iv) and (v). A most preferred method includes all of steps (i) to (v). Thus, a preferred method for producing TETA.4HCl is: (i) adding an anti-solvent to an aqueous solution of TETA.4HCl, typically a stirred aqueous solution, at a temperature T1, for a period of time t1, and / or at a rate R1; (ii) optionally, adding seed crystals of TETA.4HCl; (iii) optionally, stirring the resulting mixture at T1 for a further period of time t1a; (iv) reducing the temperature to a temperature T2 and stirring the mixture for a further period of time t2; and (v) recovering the resulting crystals. wherein T1 is about 20° C. or less; T2 is about 10° C. or less; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 3 hours, and t2 is at least 30 minutes.

[0082] A further preferred process suitable for preparing substantially pure Form B of TETA.4HCl is: (i) adding an anti-solvent to an aqueous solution of TETA.4HCl, typically a stirred aqueous solution, at a temperature T1, for a period of time t1, and / or at a rate R1; (ii) adding seed crystals of TETA.4HCl; (iii) stirring the resulting mixture at T1 for a further period t1a; (iv) reducing the temperature to a temperature T2 and stirring the mixture for a further period of time t2; and (v) recovering the resulting crystals. wherein T1 is about 15° C. or less; T2 is about 5° C. or less; t1 is at least 1 hour, R1 is 0.2 ml / min / g or less, t1a is at least 4 hours, and t2 is at least 30 minutes.

[0083] The crystals may be recovered by any suitable means, so long as the temperature of the recovery step is maintained below about 40° C., preferably below about 30° C. Higher temperature steps performed before the crystals are completely dried have been found to lead only to crystals of Form A. Most preferably, recovery of the crystals is carried out at below about 25° C., for example, at or below about 20° C.

[0084] Suitable methods for recovering the crystals include filtration and centrifugation. The resulting crystals are then typically dried at a temperature below about 40° C., preferably below about 30° C. The crystals may be washed, for example, with an anti-solvent before drying. Suitable anti-solvents for washing are those mentioned above, in particular methanol or ethanol, most preferably ethanol. As heating leads to the production of crystals of Form A, drying is typically vacuum drying. Vacuum drying at below about 40° C. is preferred. It is.

[0085] The resulting dry product may be further processed, if desired, for example by grinding or granulation. Crystalline form B is substantially stable to grinding.

[0086] Where relevant, recovery and further processing steps such as washing, drying, grinding, etc. are typically carried out under an inert atmosphere such as nitrogen.

[0087] Pharmaceutical Compositions and Dosage Forms The pharmaceutical composition of the present invention comprises crystalline Form B of TETA.4HCl as described herein, together with one or more pharma- ceutically acceptable carriers or diluents. The pharmaceutical composition can take any suitable form, but is preferably an oral dosage form. For example, the composition can take the form of a tablet, capsule, powder, semi-solid, sustained release formulation, solution, suspension, or any other suitable composition. Tablets, capsules, and powders, especially tablets, are preferred.

[0088] In an alternative embodiment, the compositions are administered parenterally, for example subcutaneously or intravenously.

[0089] Pharmaceutical dosage forms can be produced by subjecting the crystals produced as described herein to further processing steps. Thus, a composition, typically an oral dosage form, can be produced by (a) obtaining form B of TETA.4HCl, for example using the methods described above, (b) optionally milling and / or granulating the obtained crystals, (c) combining form B of TETA.4HCl with a pharma- ceutically acceptable carrier, and (d) optionally mixing form B of TETA.4HCl and the carrier. Suitable carriers are further described below. If the oral dosage form is a tablet, the process can further include (e) compressing the mixture to form a tablet, and optionally sugar-coating or film-coating the tablet. Alternatively, the solid oral dosage form can be a capsule or powder. In this case, the method of the present invention can further include (e) packaging the obtained mixture, for example, in a capsule. Further standard steps, such as milling, granulation, sugar-coating, or film-coating, can be included in the process.

[0090] Pharmaceutical compositions typically contain up to 85% by weight TETA.4HCl, for example up to 50% by weight TETA.4HCl. Preferred compositions are sterile and pyrogen-free.

[0091] Pharmaceutically acceptable carriers suitable for preparing oral dosage forms include, for example, solubilizers such as cyclodextrins or modified cyclodextrins; diluents such as lactose, dextrose, sucrose, cellulose, corn starch or potato starch; lubricants such as silica, talc, stearic acid, magnesium or calcium stearate, and / or polyethylene glycol; binders such as starch, tragacanth gum, gelatin, syrup, acacia, sorbitol, methylcellulose, carboxymethylcellulose or polyvinylpyrrolidone; disaggregating agents such as starch, alginic acid, alginate or sodium starch glycolate; foaming mixtures; dyes; sweeteners; wetting agents such as lecithin, polysorbate, lauryl sulfate; and non-toxic and pharmacologically inactive substances commonly used in pharmaceutical preparations. Such pharmaceutical preparations can be prepared in a known manner, for example by mixing means.

[0092] Compositions of the invention typically contain a high proportion of Form B of TETA.4HCl and a small amount of Form A. Preferably, pharmaceutical compositions of the invention contain 10% or less by weight of Form A of TETA.4HCl. Preferably, compositions of the invention are substantially free of Form A of triethylenetetramine tetrachloride. As used herein, substantially free of Form A means that the composition contains 5% or less by weight, preferably 2% or less by weight, more preferably 1% or less by weight, or 0.5% or less by weight, and most preferably 0.1% or less by weight of Form A of TETA.4HCl.

[0093] Medical Use The compound of the present invention is administered to the subject in a therapeutically effective amount.It will be understood that the specific dose level of a specific subject depends on various factors, including the activity of the specific compound used, age, body weight, general health, sex, diet, administration time, administration route, excretion rate, drug combination, and the severity of the specific disease being treated.The optimal dose level and administration frequency are usually determined by clinical trial.

[0094] A typical daily dose is up to 50 mg per kg of body weight, for example 0.001 to 50 mg per kg of body weight, depending on the age, weight and condition of the subject being treated, the type and severity of the disease, and the frequency and route of administration. Preferably, the daily dose level is 0.05 mg to 2 g, preferably 0.1 mg to 10 mg. The compounds of the present invention are typically administered to patients in a non-toxic amount.

[0095] The present invention also provides the crystalline form defined herein or the composition defined herein for use in a method of treating the human or animal body by therapy.In particular, the crystalline form and composition of the present invention are useful in reducing copper levels in a subject and / or reducing the toxic effects of copper retention in a subject.Therefore, they are useful for treating disorders and diseases associated with elevated copper levels.In particular, they are useful for preventing and treating Wilson's disease.

[0096] TETA itself is a known treatment for Wilson's disease and diseases and disorders associated with elevated copper levels. After administration of the crystalline form of the present invention, the compound dissolves in an in vivo system, and the therapeutic effect of the crystalline form can be expected to be the same as known forms of TETA.

[0097] The subject treated by the present invention may be a human or animal subject, in particular a human or mammal, typically a human. EXAMPLES

[0098] Working Example Reference Example 1: Synthesis of crude TETA.4HCl Crude TETA.4HCl was prepared as shown in Figures 1a and 1b.

[0099] Example 2: Synthesis of Form B of TETA.4HCl Form B of TETA.4HCl in substantially pure form was prepared according to the steps shown in FIG.

[0100] Reference Example 3: Synthesis of Form A of TETA.4HCl The crude TETA 4HCl was dissolved in 2 volumes of purified water under nitrogen, and then the resulting solution was clarified by filtration. The reactor was heated to 70°C (jacket reactor temperature) and when the temperature of the reaction mixture was between 55 and 60°C, 7 volumes of methanol were added at such a rate that the temperature in the mass remained between 55 and 65°C to recrystallize the product.

[0101] After stirring for at least 30 minutes at 65° C. (jacket reactor temperature), the reaction mixture was slowly cooled over a period of at least 2 hours and 30 minutes, observing a cooling rate of about 5° C. per 30 minutes, to a temperature in the mass between 30° C. and 35° C. The suspension was then stirred for at least 1 hour with the temperature in the mass maintained between 30° C. and 35° C.

[0102] The product was filtered through an enamel Nutsch filter with a membrane porosity of 10 μm and washed twice with 1.5 volumes of methanol. Forced filtration was performed after the last wash and the product was dried. The product was dried in a vacuum oven at 60° C. for at least 14 hours.

[0103] Example 4: Analysis of Crystalline Forms of TETA.4HCl X-ray powder diffraction A few milligrams of the samples obtained in Example 2 and Reference Example 3 above were placed between three polymer foils (Kapton® and polypropylene). Kapton® shows a broad peak in the diffractogram with weak intensity around 2θ=5.5°.

[0104] The samples were placed in a PANALYTICAL X'PERT PRO MPD diffractometer configured in transmission mode and analyzed using the conditions shown in Table 1 below. Diffraction data was obtained by dividing the powder sample by Cu-K with a characteristic wavelength (λ) of 1.5418 Å. α The diffraction patterns were obtained by exposure to X-ray radiation. The X-rays were generated from a Cu anode supplied with 40 kV and 40 mA current. Analyses were performed between 2° and 50° (unless otherwise stated). The calibration of the diffractometer was checked before each analysis.

[0105] Figure 3 shows the XRPD pattern of a mixture of Form A and Form B ("Profile 1+2"), as well as the XRPD pattern of crystals prepared according to Reference Example 3 (substantially pure Form A: "Profile 1"). Arrows indicate peaks characteristic of Form B. Figure 4 shows the XRPD pattern of crystals prepared according to Example 2 (substantially pure Form B).

[0106] [Table 1]

[0107] Raman spectroscopy The samples of Example 2 and Reference Example 3 were analyzed by Raman spectroscopy using a Renishaw RA802 Pharmaceutical Analyser with the following conditions:

[0108] [Table 2]

[0109] The spectra are provided in Figures 5a and 5b. In Figure 5b, the lower trace represents Example 2 (Form B), while the upper trace represents Reference Example 3 (Form A). The arrows indicate the shifts specific to Form B. The Raman spectrum of Example 2 shows peaks at 943, 1173, 1527 and 1612 cm -1 The Raman spectrum of Reference Example 3 (Form A) shows bands at 933, 1167, 1513 and 1604 cm -1 The peak is shown in.

[0110] FTIR-ATR analysis Infrared spectra are measured on a Nicolet iS5 FT-IR spectrometer equipped with an iS7 ATR module with the following parameters:

[0111] [Table 3]

[0112] An infrared spectrum was obtained for Example 2 (Form B) and is shown in Figures 8a and b.

[0113] DSC analysis DSC analyses were performed using a Mettler Toledo DSC3+ (serial number B531255222) in 40 μl sealed aluminum pans with lids pierced prior to analysis under a nitrogen flush of 50 mL / min.

[0114] [Table 4]

[0115] An endothermic event corresponding to melting of the sample is observed. The onset and peak temperatures are given in the table below.

[0116] [Table 5]

[0117] Thermogravimetric analysis (TGA) Thermogravimetric analysis was performed using a Pyris 1 TGA analyzer (serial number 537N7052501) under a nitrogen flush of 20 mL / min in sealed aluminum pans that were pierced prior to analysis.

[0118] [Table 6]

[0119] Thermogravimetric analysis showed a weight loss starting at 225°C, which increased after 289.5°C, likely due to decomposition. The weight loss was 10.24%: 2.64% between 225.0 and 289.5°C, and 7.60% between 289.5 and 299.0°C.

[0120] Dynamic Vapor Sorption (DVS) Analysis DVS analysis was performed using an SMS DVS Intrinsic analyser (serial number PF 140088) in open aluminium pans at 25°C with a nitrogen purge gas of 100ml / min. Stability criteria was less than 0.002% weight change over a 5 minute time frame (minimum 10 minutes, maximum 100 minutes).

[0121] [Table 7]

[0122] DVS isotherm plots are provided in Figures 11a (Example 2) and 11b (Reference Example 3). DVS analysis performed on these two samples shows a significant weight gain at high relative humidity values ​​(above 90% RH). - in Example 2 at least +55.8% - At least +60.8% in reference example 3

[0123] The desorption steps behave differently for the two solids: Example 2 reaches almost the reference weight at the second desorption step; in Reference Example 3, its weight is +14.4% greater than the reference weight at the end of the second desorption step (and +7.9% greater relative to the minimum value reached). It is noteworthy that for the latter, a time limit criterion is reached at these steps (and therefore the weight of the sample is not stable).

[0124] Example 5: Haeckel test The purpose of the Heckel test is to compress a test material under controlled conditions to derive the yield pressure of the bulk material. A known weight of material is compressed with a flat surfaced punch moving at a set speed in a 10 mm diameter die. The force on the punch is precisely measured at frequent intervals while the displacement of the punch is used to calculate the volume of the powder. The yield pressure is calculated at slow and fast punch speeds to evaluate the time dependent factor for the deformation of the material. The samples produced in Example 2 and Reference Example 3 were subjected to the Heckel test.

[0125] manner Measurement of true density by helium pychnometry Equipment used: Micromeritics AccuPyc II 1340 Test parameters: Cup size 3.5cm3 Purge Number 5 Purge pressure 19.5psig Run count: 10 Effective filling pressure 19.5psig Equilibration rate 0.02psig Execution accuracy Yes Percentage of full scale 0.05% Tests were performed in duplicate (assuming a target variation of <2% was achieved).

[0126] compression A known weight of pure drug is compressed to theoretical zero porosity using a 10 mm diameter flat faced punch. A compression simulator was used with the following conditions: Tooling: 10mm round flat surface Profile: V-profile Punch speed - slow 0.1mm / sec Punch speed - high speed 300mm / sec Die Lubrication: Yes, with magnesium stearate in acetone Number of repetitions: 3 Elasticity correction: Yes

[0127] During compression, the position of the punch tip is precisely measured, the force is measured by a load cell, and a record of the main compression parameters is made. Temperature and humidity are monitored at intervals during the test.

[0128] The data was analyzed by the Compaction Analysis software program and the Haeckel equation:

[0129]

number

[0130] (Wherein, D=relative density of the molded body) P = applied pressure K=slope of the line in the linear region) The yield pressure (Py) value was generated using [Reference: RW Heckel. Trans. Metall. Soc. AIME 221 (1961)1001-1008].

[0131] Strain Rate Sensitivity (SRS) For some materials, the deformation characteristics change with the rate of applied force. This can be inferred by calculating the strain rate sensitivity. The yield pressure in high speed compression is given by the following equation:

[0132]

number

[0133] is compared with that of low-speed compression using [Reference: RJ Roberts and RC Roe, Chem. Eng. Sci. 42(1987) p903]

[0134] result true density

[0135] [Table 8]

[0136] [Table 9]

[0137] [Table 10]

[0138] [Table 11]

[0139] [Table 12]

[0140] [Table 13]

[0141] [Table 14]

[0142] The Heckel test is a measure of the deformation of the formed tablet. The strength of the compact is an indication of how the residence time affects the adhesion of the compact. The compacts of Reference Example 2 produced at low speed were found to have a moderate tensile strength (6-11 kPb). At high speed, the strength decreased to about 4-6 kPb. On the other hand, Example 3 showed a strength of 7-12 kPb at low speed but 9-11 kPb at high speed, indicating that Example 3 has a greater tensile strength of the compacted product at high compaction fates. Figure 6 shows the strain rate sensitivity of Example 2 and Reference Example 3 at different production speeds.

[0143] Example 6: Aging of tablets The TETA.4HCl obtained according to Reference Example 3 was compressed to form tablets. An image of the tablet is provided in Figure 7a. The tablet was aged for 6 months at 40°C and 75% humidity. After aging, the tablet was observed to have many discoloured patches. An image of the aged tablet is provided in Figure 7b.

[0144] The tablets were analyzed by Raman spectroscopy under the same conditions as set forth in Example 4 above, and the results were compared to the Raman spectra of TETA.4HCl forms A and B. Figure 7c shows areas of the tablet that exhibit the presence of TETA.4HCl form A in the dark areas. As is evident from a comparison of Figures 7b and c, the areas of TETA.4HCl form A correlate with the location of the discolored areas on the tablet surface.

[0145] Trace amounts of TETA.4HCl form B were detected in the tablets that may have formed under compression of the TETA.4HCl to form the tablets. The areas of TETA.4HCl form B do not correlate with the areas of discoloration observed in the aged tablets.

[0146] Example 7: Crystallization Process Crystallization trials for the preparation of Form B were carried out at 20° C. The starting solution was prepared in a mixture of ethanol / water (25:75) and the addition of ethanol was carried out until the (75:25) ratio was reached. The first trials were carried out by adding ethanol manually. Additional trials were then carried out using a syringe pump for slow and controlled rate addition.

[0147] Manual addition (dropping) In these tests, addition was done manually using a micropipette. The solution of TETA.4HCl in a (25:75) ethanol / water mixture was kept under stirring and thermostated at 20° C. The amount of TETA.4HCl starting material is shown in Table 15.

[0148] The anti-solvent (ethanol) was added dropwise at regular intervals. Two addition "rates" were tested. After the addition of the anti-solvent, the solid phase was sampled and analyzed by X-ray diffraction to determine the nature of the solid phase. The conditions for X-ray diffraction were as indicated in Example 4. The last test was performed at a slower addition rate with a seed of Form B present at the beginning of the addition (PE1716E007-L-5). This test leads to a solid phase with no Form A signal observed in the diffractogram.

[0149] The results are shown in Table 15 and the diffractograms obtained after analysis of the solid phase in the suspension at the end of the ethanol addition are shown in Figures 9a, b and c. Figures 9a-c start with the lowest line: the reference for Form A (bottom line in Figures 9a-c). Reference for Form B PE1716E007-L-5 PE1716E007-L-4 PE1716E007-L-3 PE1716E007-L-2 PE1716E007-L-1 PE1716E007-R-1 (top line in Fig. 9a-c) 4 shows the XRPD diffractogram of

[0150] [Table 15]

[0151] Program Addition (Continuous) In this series of tests, the anti-solvent addition was driven by a syringe pump, which allowed for continuous addition at a very slow rate. The same protocol as above was used: a near-saturated solution of the starting material was prepared in a 25:75 ethanol / water mixture, as shown in Table 16. The solution was then saturated with Form B until a solid phase remained in suspension: this ensures that Form B species are present at the start of the anti-solvent addition.

[0152] Two addition rates were tested (for 750 mg starting material in solution): 0.05 mL / min and 0.1 mL / min. Ethanol was added at these selected rates until an ethanol / water ratio of 75:25 was obtained. Two additional tests were performed adding up to a ratio of 82:18, corresponding to a concentration of approximately 12.5%, and up to a ratio of 87.5 / 12.5, corresponding to a concentration of approximately 9% (starting material / total weight).

[0153] X-ray diffraction analysis was performed using the conditions given in Example 4. The results are given in Table 16 below and the diffractograms obtained after analysis of the solid phase in the suspension at the end of the ethanol addition are shown in Figures 10a-10c. These show diffraction profiles with no signal for Form A observed in two runs with a final ethanol / water ratio of 75:25 and in the run with 87.5:12.5. A small shoulder to the left of the peak at 2θ=25.4° (corresponding to Form A) in the run with 82:18.

[0154] Figure 10a-c starts from the lowest line: Reference for Form A (bottom line in Figure 10a-c) Reference for Form B PE1716E007-L-6 PE1716E007-L-7 PE1716E007-L-8 PE1716E007-L-9 (top line in Figure 10a-c) 4 shows the XRPD diffractogram of

[0155] [Table 16]

[0156] Example 8: Crystallization Process The synthesis of triethylenetetramine (trientine) tetrahydrochloride (TETA 4HCl) is outlined in the scheme below.

[0157] [ka]

[0158] In step I, triethylenetetramine (TETA) is converted to the corresponding triethylenetetramine hydrate (TETA hydrate) by stirring in the presence of water and TBME. Isopropanol is added as an antisolvent and optionally seeded with TETA hydrate. The TETA hydrate is crystallized, filtered and isolated. Crude triethylenetetramine tetrahydrochloride (crude TETA 4HCl) is obtained by reacting triethylenetetramine hydrate (TETA hydrate) with aqueous hydrochloric acid in ethanol (step II). Crude triethylenetetramine tetrahydrochloride (crude TETA 4HCl) is recrystallized from a mixture of purified water and ethanol. Crude triethylenetetramine tetrahydrochloride (crude TETA 4HCl) is further purified by recrystallization from a mixture of purified water and ethanol in the presence of Form B seeds to obtain triethylenetetramine tetrahydrochloride (TETA 4HCl) (step III).

[0159] The process can be used to produce TETA in batches of 125 kg to 110-130 kg. The overall yield of the synthesis is about 50%, including two recrystallizations of crude TETA 4HCl.

[0160] Recrystallization to produce Form B crystals is carried out as summarized in the flow charts of Figures 1a, 1b and 2, but with the addition of an additional recrystallization of crude TETA.4HCl at the end of step II (Figure 1b) and before step III (Figure 2). The process can be described as follows:

[0161] Once the equipment has been inerted, all operations are carried out under nitrogen flow.

[0162] Step I: Preparation of triethylenetetramine hydrate (TETA hydrate) Charge triethylenetetramine (nominal amount 125 kg) to the reactor followed by TBME (185±5 kg). Add water (approximately 28 kg) with stirring over ≥15 min while maintaining the temperature at ≤30°C. Optionally, seed the solution with triethylenetetramine hydrate (approximately 0.1 kg) while stirring at 25-35 °C to promote crystallization. Add isopropanol (64±1 kg) at 25-35°C. Heat the suspension to 30-40°C for ≥15 min, followed by slow cooling to 15-25°C for ≥90 min. Cool the suspension to -5 to 5°C and stir for ≥ 30 minutes. The product is filtered and centrifuged, then a sample is taken for analysis (GC assay) and determination of impurities. If the sample is sufficiently pure, the wet TETA hydrate is loaded into a dryer and dried at ≦25°C until it meets the requirements set for the next step. The dried product is analyzed for purity by GC, appearance, residual water by KF, identity by FTIR, sulfated ash and residual solvents by GC.

[0163] Step II: Preparation of crude triethylenetetramine tetrahydrochloride (crude TETA.4HCl) Dissolve triethylenetetramine hydrate (TETA hydrate) in water (85±1 kg) and acidify by dosing concentrated hydrochloric acid (200±5 kg) for ≥1 hour at ≤40°C. Check the pH value (target pH=1.0) and add concentrated hydrochloric acid until the pH is ≤1.0. Cool the reaction mixture to 15-25°C and stir for ≥10 min. Treat the solution with ethanol (672±5 kg) dosed over ≧1.5 hours while maintaining the temperature at ≦30° C. Cool the suspension to -5 to 5°C and stir for ≥ 30 minutes. · The product is filtered and the solid is washed successively with ethanol (1 × 20 kg, then 3 × 25 kg). Dissolve crude triethylenetetramine tetrahydrochloride (crude TETA.4HCl) in water (340±10 kg). Treat the solution with ethanol (909±10 kg) dosed over ≧1.5 hours while maintaining the temperature at ≦30° C. Cool the suspension to -5 to 5°C and stir for ≥ 30 minutes. Filter the product and wash the solid successively with ethanol (1×14 kg, then 3×15 kg).

[0164] Step III: Preparation of triethylenetetramine tetrahydrochloride (TETA.4HCl) Dissolve crude triethylenetetramine tetrahydrochloride (crude TETA.4HCl) in water (340±10 kg). Treat the solution with ethanol (909±15 kg) dosed over ≥1.5 hours while maintaining the temperature at 7-13°C. During the ethanol addition, the solution is seeded with TETA.4HCl (2 wt%). The suspension is stirred for ≥5 hours, then cooled to -5°C and stirred for ≥30 minutes. Filter the product and wash the solid successively with ethanol (1×14 kg, then 3×15 kg). -The purity of the product samples is analyzed by GC. Dry the product at ≦40°C and, if the control parameters for loss on drying are met, grind the product. The ground bulk material is transferred under nitrogen into double layered food grade polyethylene bags, then sealed in aluminium bags. The aluminium bags are inserted into HDPE drums.

[0165] Reprocessing The TETA4HCl obtained after recrystallization is tested for impurities by GC. If the impurity levels are too high, step III can be repeated.

Claims

1. Features: (i) an XRPD pattern having at least two peaks selected from the peaks at 22.9, 25.4, 25.8, 26.6, 34.6 and 35.3±0.1 degrees 2θ; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm －１ A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of A crystalline form of triethylenetetramine tetrachloride having at least one of the following:

2. 2. The crystalline form of claim 1 having an XRPD pattern with peaks at 25.4, 34.6 and 35.3±0.1 degrees 2θ.

3. 3. The crystalline form of claim 1 or claim 2 containing up to 10% by weight of triethylenetetramine tetrachloride form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

4. In essence (i) an XRPD pattern as defined in claim 1 or claim 2; and / or (ii) 943, 1173, 1527 and 1612 ± 5 cm －１ A Raman spectrum having at least two peaks selected from the peaks of the Raman shift of 2. The crystalline form of any one of the preceding claims, consisting of form B of triethylenetetramine tetrachloride having the formula:

5. A pharmaceutical composition comprising the crystalline form of any one of claims 1 to 4 and a pharma- ceutically acceptable carrier or diluent.

6. 6. The pharmaceutical composition of claim 5, which is a solid oral dosage form comprising the crystalline form of any one of claims 1 to 4 and a pharma- ceutically acceptable carrier.

7. 7. The pharmaceutical composition of claim 5 or claim 6, comprising less than 10% by weight, preferably less than 5% by weight, more preferably less than 2% by weight of triethylenetetramine tetrachloride form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1°2θ.

8. 8. The pharmaceutical composition of claim 7, substantially free of triethylenetetramine tetrachloride Form A having an XRPD pattern with peaks at 25.2 and 35.7±0.1 degrees 2θ.

9. 1. A method for preparing a crystalline form of triethylenetetramine tetrachloride comprising adding an anti-solvent to an aqueous solution of triethylenetetramine tetrachloride and recovering the resulting crystals, wherein the addition of the anti-solvent is carried out at a temperature of about 20° C. or less.

10. 10. The method of claim 9, wherein the rate of addition of the anti-solvent to the solution is 0.5 ml / min or less per gram of TETA.4HCl dissolved in the aqueous solution.

11. (i) adding an anti-solvent to an aqueous solution of TETA.4HCl at a temperature T1, for a period of time t1, and / or at an addition rate R1; (ii) optionally, adding seed crystals of TETA.4HCl; (iii) optionally agitating the resulting mixture at T1 for a further period of time t1a; (iv) optionally, reducing the temperature to a temperature T2 and agitating the mixture for a further period of time t2; and (v) recovering the resulting crystals.

11. The method of claim 9 or claim 10, wherein T1 is about 20° C. or less; T2 is at least 5° C. lower than T1; t1 is at least 1 hour, R1 is 0.5 ml / min / g or less, t1a is at least 2 hours, and t2 is at least 30 minutes.

12. (i) adding an anti-solvent to an aqueous solution of TETA.4HCl at a temperature T1, for a period of time t1, and / or at a rate R1; (ii) Seeding with TETA.4HCl; (iii) agitating the resulting mixture at T1 for a further period of time t1a; (iv) reducing the temperature to temperature T2 and agitating the mixture for a further period of time t2; and (v) recovering the resulting crystals.

12. The method of claim 11, wherein T1 is about 15° C. or less; T2 is about 5° C. or less; t1 is at least 1 hour, R1 is 0.2 ml / min / g or less, t1a is at least 4 hours, and t2 is at least 30 minutes.

13. 13. The method of any one of claims 9 to 12, further comprising drying the recovered crystals at a temperature below about 40°C, preferably below about 30°C.

14. The method of any one of claims 9 to 13, wherein the recovered crystals are combined with a pharma- ceutically acceptable carrier to produce a pharmaceutical composition.

15. 15. The method of claim 14, further comprising compressing the mixture of the crystals and a pharma- ceutically acceptable carrier to form a tablet, and optionally sugar-coating or film-coating the tablet.

16. A crystalline form or pharmaceutical composition obtainable by the method according to any one of claims 9 to 15.

17. 13. A crystalline form or a pharmaceutical composition according to any one of the preceding claims for use in the prevention or treatment of Wilson's disease.

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