Technical Field

The invention relates to a stable pharmaceutical composition containing, as the active substance, tenofovir disoproxil fumarate, or a combination of tenofovir disoproxil fumarate with another active substance.

Tenofovir disoproxil fumarate (TDF, Form I) is an extensively used pharmaceutical salt. It is present in the monotherapeutical product Viread, in a combination with emtricitabine in the product Truvada, and in a triple combination with emtricitabine and efavirenz in the product Atripla. It is used in curing AIDS and hepatitis of type B. State of the art

Although API is registered as tenofovir disoproxil fumarate (salt) and in this form it enters the process of preparing the pharmaceutical formulation, it has been found that various solid forms of API are present in respective products. In particular hemifumarate (TDHF), corresponding to a co-crystal of tenofovir disoproxil and fumaric acid, as described in several publications, e.g. in patent publication WO2008143500A1 including final solution of its structure. Tenofovir disoproxil fumarate (1:1) and tenofovir disoproxil hemifumarate (2:1) differ in stoichiometry, and these forms cannot be considered mutual polymorphs.

The phase analysis of the original products has shown that API is present in various crystalline modifications (see Table 1).

Table 1. Comparison of composition of original products with tenofovir

TDF can be converted to the TDHF form by simple stirring in isopropyl alcohol or in water, as described, e.g. in patent publication WO2008143500A1. The observed phase changes in analyzed original products suggest possible connection with the applied technological procedure in formulation of the medicinal form.

This connection has already been discussed in the registration documentation of drugs containing TDF; e.g. the EMEA Scientific Discussion on drug Truvada (emtricitabine- TDF) states that emtricitabine as well as tenofovir disoproxil fumarate is prone to decomposition at elevated humidities and temperatures. The control of the amount of water used during the production process and of the content of unbound water in the final product minimizes the risk of degradation. Drug Viread (TDF) has the following composition (excipients):

Core of tablet

Sodium salt of croscarmellose

Lactose monohydrate

Magnesium stearate (E572)

Microcrystalline cellulose (E460)

Pre-swollen starch

Coat layer of tablet

Triacetin (E1518)

Hyprome!lose (E464)

Lactose monohydrate

Titanium oxide (E171)

Drug Truvada (emtricitabine-TDF) has the following composition (excipients)-.

Core of tablet:

Sodium salt of croscarmellose

Lactose monohydrate

Magnesium stearate (E572)

Microcrystalline cellulose (E460)

Pre-swollen starch (gluten-free)

Coat:

Glycerol triacetate (E1518)

Hypromellose (E464)

Aluminium enamel of indigo carmine (E132) Lactose monohydrate

Titanium oxide (E171)

Whereas the pharmaceutical formulations Viread and Truvada are prepared by the technology of wet granulation, the pharmaceutical formulation Atripla involves both the technology of wet granulation (efavirenz) and dry granulation (emtricitabine and TDF). The above mentioned indicates that, during the formulation process, the drugs Viread and Truvada contend with the conversion of the input API, tenofovir disoproxil fumarate TDF, to TDHF.

Disclosure of Invention

The invention provides a stable pharmaceutical composition for preparation of tablets by wet granulation, which comprises tenofovir disoproxil fumarate, or a combination of tenofovir disoproxil fumarate and another active substance, and a disintegrant selected from the group including crospovidone, maize starch, and hydroxypropylcellulose. Using of a disintegrant from the group including crospovidone, maize starch, and hydroxypropylcellulose instead of sodium salt of croscarmellose prevents the conversion of TDF to TDHF.

In addition, the invention includes a pharmaceutical composition for preparation of tablets by wet granulation, which comprises tenofovir disoproxil fumarate, or a combination of tenofovir disoproxil fumarate and another active substance, the sodium salt of croscarmellose and an acid from the group including fumaric, citric, malic, maleic, L-tartaric, and malonic acids in a 25 to 100 % wt. excess over the amount of the sodium salt of croscarmellose. Using of the combination of the sodium salt of croscarmellose and an acid from the above mentioned group prevents the conversion of TDF to TDHF. Detailed description of invention

Conditions have been found under which the conversion of TDF to TDHF does not take place in the formulation even if the technology of wet granulation is applied.

It has been shown that stability TDF is, in addition to humidity, connected with pH of the environment Stability of the TDF and TDHF forms in environment of various pHs has been studied directly with the API. The conversion of tenofovir TDF in presence of a buffer has been monitored by Raman spectroscopy. In an environment of pH = 2, the sample was completely dissolved and, after evaporation of the solvent, the solid portion did not correspond to any of the above described forms; tenofovir was probably chemically decomposed. In an environment of pH = 3.2, tenofovir remained in the form of TDF. In an environment of pH = 3.8, TDF was converted to TDHF within 6 h and, after several days, TDHF was dissociated to the free base. In an environment of pH = 4.8, TDF was converted to TDHF within 15 min and, after one day, TDHF was dissociated to the free base. In an environment of pH = 6.8, TDF was converted to TDHF within 10 min and, after several hours, TDHF was dissociated to the free base. The conversion of TDF to TDHF seems to run quicker at higher pH. After remaining in an environment of higher pH for a longer period of time, TDHF is dissociated to the free base. Individual excipients can contribute to increased pH and thus to the conversion of TDF to TDHF. From the excipients used, the most basic excipient is sodium croscarmellose that plays a role of the disintegrant in the dosage form and belongs among neutral excipients. The effect of sodium croscarmellose on pH was studied in mixtures of excipients: Avicel pH 101 + polyvinylpyrrolidone (K-25) + lactose monohydrate (hereinafter placebo only), and placebo with sodium croscarmellose (see the table). It is apparent that sodium croscarmellose increases pH of the mixture.

Possible solution is replacement of sodium croscarmellose with another disintegrant, such as carboxymethyl starch, calcium croscarmellose, maize starch, crospovidone or hydroxypropylcellulose (e.g. type LH21). By measuring pH of these mixtures, it has been found that the disintegrants calcium croscarmellose and carboxymethyl starch (containing sodium ions) increase pH as well (see Table 2).

After the granulation of all these mixtures with various disintegrants together with TDF, it has been shown that the disintegrants containing sodium or calcium ions (sodium croscarmellose, calcium croscarmellose, carboxymethyl starch) contribute to the conversion to TDHF (see Figure 1). Table 2. Final pH values of mixture of excipients with various clisinteg rants.

Addition of an acid from the group of fumaric, citric, malic, maleic, l-tartaric or maionic acids, which interact with sodium cations of croscarmellose instead of fumaric acid bound in TDF, has proved another possible solution of preventing undesirable conversion of TDF to TDHF, while keeping sodium croscarmellose as the disintegrant.

10 - 27 mg of citric, fumaric, malic, maleic, l-tartaric or maionic acid in the solid state was admixed to the tableting matter with sodium croscarmellose; consequently, the samples contained the added acid in 25, 50, and 100% excess over sodium croscarmellose. In all cases (with all acids) the conversion to TDHF was successfully prevented (see Figure 2).

Chemical purity of the mixtures with added acid without treatment was comparable with purity of the mixtures without acid. After treatment (80°C, 7 hours), no effect of fumaric acid on the degradation of tenofovir was observed. With a higher than 55% excess of citric acid, the degradation runs already after three hours at 80°C (sum of impurities 1%).

In the case of samples with maionic, maleic, and L-tartaric acids, on one hand, no conversion to TDHF was detected but, on the other hand, the diffraction patterns showed peaks corresponding to an unknown new solid phase of TDF. This new phase has a similar diffraction pattern as TDF (see Fig. 3). However, the positions and intensities of the diffraction peaks differ slightly from those of TDF and several completely new reflexions appear. The ssNMR spectrum of this new phase is also quite similar to that of TDF (Fig. 4). The melting temperature of this new phase is by about 12°C lower than that of TDF (Fig. 5). This new phase can be a TDF hydrate; water content is approximately 1.2%, as determined by the method of Carl Fischer. Due to the fact that TDF crystallizes in crystals which are too small for the X-ray structural analysis from a monocrystal and that the structure of TDF is too complex with large number of torsional angles for evaluation from the powder data, the character of the new phase could only be studied indirectly.

Brief Description of Drawings

Figure 1. Diffraction patterns of granulated mixtures (from the top): with sodium croscarmellose, calcium croscarmellose, with carboxymethyl starch (diffraction peaks in the position 8° 2Θ are characteristic of TDHF), and with other disintegrants - crospovidone, maize starch, hydroxypropylcellulose (overlapping charts at bottom, characteristic peaks of TDHF not detected).

Figure 2. Diffraction patterns of mixtures (TDF + excipients) with added acids (citric, fumaric, malic) and a diffraction pattern of the reference mixture of TDF with excipients without added acid (top), in which peaks corresponding to TDHF are apparent.

Figure 3. Comparison of diffraction patterns of the TDF form and the new solid form of TDF (hydrate).

Figure 4. ssNMR spectra of TDF (top) and of the new solid form of TDF (bottom). Figure 5. DSC curve of the melting of the new solid form of TDF (hydrate).

Analytical methods

X-ray powder diffraction

The diffraction patterns were obtained using a powder diffractometer X'PERT PRO MPD PANalytical; X-ray beam Cu a (λ = 1.542 A), excitation voltage: 45 kV, anodic current: 40 mA, measured range: 2 - 40° 2Θ, step size: 0.01° 2Θ, remaining at reflexion 0.05 s. The measurement was performed on a flat sample of area/thickness 10/0.5 mm. 0.02 rad Soller diaphragms, 10 mm mask, and 1/4° fixed anti-scattering diaphragm were used to correct the primary beam. The irradiated area of tine sample was 10 mm; programmable divergent diaphragms were used. 0.02 rad Soller diaphragms and 5.0 mm anti-scattering diaphragm were used to correct the secondary beam. ss NMR

Instrumentation for the polymorphic studies: Bruker Avance 400 MHz WB (wide bore); method: 13C CP/MAS, 4 mm probe, 13 kHz spinning.

Raman spectroscopy

The samples were measured in glass HPLC vials in a spectrometer FT-Raman RFS100/S, with germanium detector (Bruker Optics, Germany), at wavelength of Nd: YAG laser 1064 nm, in the measuring range from 4000 to -2000 cm ^"1 , with spectral resolution 4.0 cm ^"1 . Data were obtained at 64 accumulations of spectra. The software OMNIC was applied in processing the spectra.

Differential scanning calorimetry (DSC) The thermograms were measured using an apparatus DSC Pyris 1 of the firm Perkin Elmer. The sample weight into the standard Al pan was 2.7 - 3.9 mg and heating rate was 10°C/min. The temperature program used included stabilization of the sample for 1 min at temperature of 20°C and heating up to 150°C with heating rate of 10°C/min. Nitrogen 4.0 of flow rate of 20 ml/min was used as the purge gas. Preparation of samples

Measurement of pH of mixtures with disintegrants. The mixtures were weighed according to Table 3 and finely pulverized in an agate mortar. Individual mixtures were overlayered with 4 ml of purified water and, after filtering off the solids, pH of the filtrate was measured using a digital pH-meter. Granulation of mixtures with disintegrants. 300 mg of TDF were weighed to the mixtures according to Table 3. The weighed mixtures were homogenized by pulverizing in an agate mortar. Subsequently, 150 μΙ of purified water were added. The mixtures were kneaded in vials by means of a spatula for 5 minutes and left on Petri dishes. The air-dried granulate was pulverized in an agate mortar and prepared for the powder X-ray analysis.

Additional addition of an acid. 300 mg of TDF and the respective acid (citric, malonic, malic, tartaric, maleic, fumaric, l-tartaric) in 25, 50, and 100% excess over sodium croscarmellose were weighed to the mixture with sodium croscarmellose (see Table 3). The weighed mixtures were homogenized by pulverization in an agate mortar. Subsequently, 150 μΙ of purified water were added. The mixtures were kneaded in vials by means of a spatula for 5 minutes and left on Petri dishes. The air- dried granulate was pulverized in an agate mortar and prepared for the powder X-ray analysis.

Table 3. Composition of mixtures for measuring pH. The contents of API and

excipients in the mixtures correspond to the weight in one tablet.

Examples

Preparation of the therapeutic composition Example 1 - Replacement of sodium croscarmellose by another disintegrant

The raw materials tenofovir disoproxil fumarate (300 g), lactose monohydrate (202 g), microcrystalline cellulose Avicel pH 101 (50 g), crospovidone (21 g), and polyvinylpyrrolidone K-25 (20 g), which were each independently previously sieved through a screen of mesh size 0.5 mm, were charged into a granulator. The mixture was homogenized for 10 min, then sprinkling with purified water (150 g) was started, and the mixture was granulated. The formed granulate was dried in a drying oven at 60°C. The dried granulate was ground in a vibrating mill and sieved through a screen of mesh size 0.63 mm. The other auxiliary substances crospovidone (21 g), microcrystalline cellulose Avicel pH 112 (50 g) were gradually sieved to the granulate (593 g) and the mixture was homogenized at 19 r.p.m. for 0 min. Finally, magnesium stearate was added (10 g) and the mixture was homogenized at 19 r.p.m. for additional 3 minutes. The tableting matter obtained in the above-described manner was compressed in a rotary pill press and used for production of cores. These cores were then coated with a pre-made coating suspension Opadry II Light Blue Y-30-10671-A. The Form I of TDF was detected in thus prepared tablets.

Crospovidone can be replaced with maize starch (89 g) or hydroxypropylcellulose LH21 (59 g). Example 2 - Addition of fumaric (citric, malic) acid to the formulation with sodium croscarmellose

The raw materials tenofovir disoproxil fumarate (300 g), lactose monohydrate (206 g), microcrystalline cellulose Avicel pH 101 (50 g), sodium croscarmellose (17 g), fumaric acid (10 g), and polyvinylpyrrolidone K-25 (20 g), which were each independently previously sieved through a screen of mesh size 0.5 mm, were charged into a granulator. The mixture was homogenized for 10 min, then sprinkling with purified water (150 g) was started, and the mixture was granulated. The formed granulate was dried in a drying oven at 60°C. The dried granulate was ground in a vibrating mill and sieved through a screen of mesh size 0.63 mm. The other auxiliary substances sodium croscarmellose (21 g), microcrystalline cellulose Avicel pH 112 (50 g) were gradually sieved to the granulate (603 g) and the mixture was homogenized at 19 r.p.m. for 10 min. Finally, magnesium stearate was added (10 g) and the mixture was homogenized at 19 r.p.m. for additional 3 minutes. The tableting matter obtained in the above- described manner was compressed in a rotary pill press and used for production of drug cores. These cores were then coated with a pre-made coating suspension Opadry II Light Blue Y-30-10671-A. The Form I of TDF was detected in thus prepared tablets.

Fumaric acid can be replaced with citric acid (17 g) or malic (12 g) acid.

Example 3 - Addition of malonic acid to the formulation with sodium croscarmellose The raw materials tenofovir disoproxil fumarate (300 g), lactose monohydrate (206 g), microcrystalline cellulose Avicel pH 101 (50 g), sodium croscarmellose (17 g), malonic acid (14 g), and polyvinylpyrrolidone K-25 (20 g), which were each independently previously sieved through a screen of mesh size 0.5 mm, were charged into a granulator. The mixture was homogenized for 10 min, then sprinkling with purified water (150 g) was started, and the mixture was granulated. The formed granulate was dried in a drying oven at 60°C. The dried granulate was ground in a vibrating mill and sieved through a screen of mesh size 0.63 mm. The other auxiliary substances sodium croscarmellose (21 g), microcrystalline cellulose Avicel pH 112 (50 g) were gradually sieved to the granulate (607 g) and the mixture was homogenized at 19 r.p.m. for 10 min. Finally, magnesium stearate was added (10 g) and the mixture was homogenized at 19 r.p.m. for additional 3 minutes. The tableting matter obtained in the above- described manner was compressed in a rotary pill press and used for production of drug cores. These cores were then coated with a pre-made coating suspension Opadry II Light Blue Y-30- 0671-A. The Form I of TDF with admixture of the new solid form of TDF (hydrate) was detected in thus prepared tablets.

Malonic acid can be replaced with maleic acid (16 g) or L-tartaric acid (20 g).

Example 4 - Preparation of new phase of tenofovir disoproxil fumarate

Tenofovir disoproxil fumarate Form I (3 g) was mixed with malonic acid (0.2 g) and purified water (1.5 g). The mixture was dried at room temperature. The X-ray pattern corresponded to the new solid form of TDF (hydrate).

Malonic acid can be replaced with maleic acid (0.22 g) or L-tartaric acid (0.29 g).