TETRACYCLINE COMPLEXES WITH SUSTAINED ACTIVITY

FIELD OF THE INVENTION

The present invention generally relates to controlled release of tetracycline antibiotics. Specifically, it relates to a complex comprising a tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof and a divalent metal carboxylate; a pharmaceutical preparation comprising the complex, methods for manufacturing the complex and the pharmaceutical preparation, and a complex or a pharmaceutical preparation for use in a method for treatment of the human or animal body, in particular for therapy and/or prophylaxis of a bacterial infection; and/or wherein antibiotic activity is maintained over a prolonged period of time; and/or for the therapy and/or prophylaxis of an acute, chronic or recurrent periodontal disease.

BACKGROUND ART

The tetracycline compounds are a class of antibiotics that inhibit protein synthesis by preventing the attachment of aminoacyl-tRNA to the ribosomal acceptor (A) site. The tetracyclines are broad-spectrum agents, exhibiting activity against a wide range of gram-positive and gram-negative bacteria, atypical organisms such as chlamydiae, mycoplasmas, and rickettsiae, and protozoan parasites. The favorable antimicrobial properties of these agents and the absence of major adverse side effects has led to their extensive use in the therapy of human and animal infections, for instance periodontal disease (Chopra et al., Microbiol Mol Biol Rev. 2001 , June 65(2):232-60). Tetracycline molecules comprise a linear fused tetracyclic nucleus (rings designated A, B, C, and D) to which a variety of functional groups are attached. The simplest tetracycline to display detectable antibacterial activity is 6-deoxy-6-demethyltetracycline and so this structure may be regarded as the minimum pharmacophore (Mitscher, L. A. 1978. The chemistry of the tetracycline antibiotics. Marcel Dekker, Inc., New York, N.Y):

Tetracycline antibiotics that are widely clinically used include doxycycline, minocycline and tetracycline. An overview of the most commonly used tetracycline antibiotics is shown below:

Among these, especially doxycycline and minocycline have been used in the treatment of periodontal diseases. However, there are problems with the therapeutic applications of these active substances due to their relatively short retention time and their instability in aqueous environment.

Periodontal diseases are highly prevalent. About 30% of the human population is affected worldwide. They have considerable impact on individuals and society, and are costly to treat. The cost of dental care is the fourth highest of all diseases and consuming between 5 and 10% of all healthcare resources (Batchelor, P. British Dental Journal 2014, 217, 405—409). Representative population studies show that periodontal diseases are widespread and their prevalence has been increasing since 1997 (Micheelis, W. et al. Vierte Deutsche Mundgesundheitsstudie (DMS IV), Deutscher Arzte-Verlag, Koln, 2006). Amongst the adult population in Germany, 52.7% were found to be affected by moderately severe and 20.5% by severe forms of periodontitis. The health insurance expenditure in Germany for the direct treatment of periodontitis amounted to about EUR 1.1 billion ( Statistisches Bundesamt, 2008), not including the costs incurred by secondary diseases.

Periodontitis is a general term describing inflammation condition of the periodontal apparatus which is caused by multi-bacterial induction and has strong relations to various systemic diseases, such as cardiovascular diseases, rheumatoid arthritis, chronic obstructive pulmonary disease and Alzheimer’s disease.

The currently established therapy of periodontitis, according to the recommendations of the German Society of Dental, Oral and Craniomandibular Sciences, is generally performed by manual supra and subgingival debridement (removal of the bacterial plaques) along with the application of antiseptic substances (daily disinfection by mouth washes), which disintegrates the entire oral biofilm and provides an opportunity for recolonization by potential pathogens (Sanz, I. et al. J Evid Based Dent Practice 2012, 12(3), 76-86). Furthermore, adjuvant systemic broad-spectrum antibiotic therapy is applied in advanced disease forms, such as persistent or recurrent localized deep sites (Jepsen, K. et al. Periodontol 2000 2016, 71 (1 ), 812-112). The latter also leads to a non-selective destruction of the biofilm and has to be administered in high doses and over a prolonged period of time in order to reach sufficient therapeutic levels at the particular site of action, i.e. the gingival pocket. Standard adjuvant therapy of periodontitis involves, for instance, systemic administration of doxyciclin (per os) 1 x 200 mg/die for 1 day and 2 x 100 mg/die for further 18 days (Wissenschaftliche Stellungnahme: Adjuvante Antibiotika in der Parodontitistherapie, Deutsche Gesellschaft fOr Zahn- Mund- und Kieferheilkunde, DZZ 2003). As a result, resistance development in oral pathogens is observed. Further, the microbiome in the patient’s intestine is destroyed, which leads to a loss of metabolic support, immune modulation, and enables recolonization by potential pathogens. A focused and targeted therapy would represent a significant improvement in the treatment of periodontitis and conditions associated therewith. A systematic review showed a significant effect of tetracycline formulations on reduction of periodontal probing depth (Matesanz-Perez, P. et al. J Clin Periodontol 2013, 40(3), 227-241).

The presence of periodontopathogenic bacteria varies among periodontitis patients. Nevertheless, the occurrence of certain bacterial species in the subgingival plaques has been found to be closely associated with the etiology of periodontal diseases (Socransky et al., Journal of Clinical Periodontology, 1998, 25, 134-144). The first step towards the development of periodontitis is the colonization of bacteria“yellow complex, "green complex" and "purple complex” in healthy periodontal sites. The Actinomyces species is closely related to members of the“yellow complex” ( Streptococcus sanguis, Streptococcus mitis, Streptococcus gordonii and Streptococcus intermedius), "green complex" ( Eikenella corrodens, Capnocytophaga gingivalis, Capnocytophaga sputigena), and "purple complex" ( Veillonella parvula, Actinomyces odontolyticus). These complexes are followed by colonization by the

members of the bacteria of the so-called “orange complex” including members of the Fusobacterium nucleatum/periodonticum subspecies, Prevotella intermedia, Prevotella nigrescens and Parvimonas micra (formerly known as Peptostreptococcus micros or Micromonas micros). Colonization of healthy periodontal sites by members of the“orange complex” has been found to correlate with the occurrence of gingivitis. The bacteria of the“orange complex” furthermore promote the colonization by bacteria of the so-called “red complex”, which in turn are associated with deep pockets and chronic periodontitis. The“red complex” consisting of the tightly related group Tannerella forsythia, Porphyromonas gingivalis and Treponema denticola strongly relates to clinical measures of periodontal disease and in particular to pocket depth and bleeding on probing. Additionally, Lamont et al. (Microbiology 2002, 148, 1627-1636.) and Daep et al. ( Infect . Immun. 2006, 74, 5756-5762) demonstrated that S. gordonii plays a role in colonization with Porphyromonas gingivalis.

The application of antibiotics or antiseptics within an adhesive slow-release form of administration for topical application is technologically challenging. The oral mucosa is covered by a liquid film and is partially colonized by bacteria. Additionally, it is subject to mechanical stress due to speaking, swallowing and chewing. Although these stress sources are not present in the periodontal pocket per se, the presence of sulcus liquid (the flow rate of which is strongly increased in the case of periodontitis) leads to increased washing out of active substances out of the pocket.

In view of this difficult starting situation, none of the existing products or local antibiotic or antiseptic therapy has been able to solve the existing problems in a satisfactory manner. For instance, Actisite® is a tetracycline containing fiber for periodontal pocket placement requiring a complicated application by a specialized dentist into the periodontal pocket and a further intervention for removal after 10 days. This product is not on the German market anymore. Elyzol®, a metronidazol containing gel, fails to meet the expectations for active ingredient release beyond two days due to lack of sufficient gel adhesion. This product is also not on the German market anymore. Atridox®, a doxycycline containing control led-release product composed of a two syringe mixing system, has also been taken off the German market.

Ligosan®, which is a doxycycline containing application system, and Periochip®, which is a chlorhexidine containing bio degradable chip, are the only products for local application still available on the market. Ligosan® is an application system comprising 14% of doxycycline in a biodegradable gel matrix for application in the periodontal pocket by means of a special location system, which provides a continuous release over 12 days and leads to an improvement of the periodontal pockets. However, the handling and manageability of the gel is complicated and application of the proper amount in all parts of the pocket is difficult, especially when not directly visible interdental spaces in the area of the side teeth are concerned. The gel needs to be inserted by means of a special application system. Furthermore, although it is proven to be effective against bacteria in the deep periodontal pockets, it is not suitable for a complete therapy and does not substitute classical treatment measures (Eickholz, P et al.; J Clin Periodontol 2002; 108-117; Ratka-Kmger, P. et al., J Periodontol. 2005/1 ,76: 66-74). Additionally, due to the high costs and low incentive for the dentists, a general application of this therapy appears to be difficult (see https://www.parodontitis.com/ligosan-slow-releaseR-zur-behandlung-von-zahnfleischtaschen.html). Periochip® contains 2.5mg of chlorhexidine in a biodegradable gelatin chip that can be inserted into the periodontal pockets with tweezers. The chip remains in the pocket for 7-10 days and over time, the drug is released continuously. However, it was found to be not as effective against the bacterial composition as local antibiotics. Further, the costs are not paid by the health insurance (see https://www.parodontitis.com/lokale-behandlung-der-zahnfleischtaschen-mit-periochip-ec40-chxhtml.html). In summary, although some improvement of periodontitis parameters (depth of the pocket and

inflammation parameters) has been achieved with these systems, they require complicated application and furthermore are not reimbursed by the public health insurance companies in Germany such that they cause unbearable costs which have to be paid by the patients.

The systemic application of low-dose doxycycline (Periostat®) as a long-term therapy has also been applied. However, the expected pharmacologic effect does not always correlate to a successful clinical result. For instance, therapy seems to have diminished efficacy in smokers. Furthermore, a high level of compliance on the side of the patient is required; and the results of scientific studies as to whether a clinical improvement is present are controversial. Thus, no general recommendation can be given concerning this therapy (G. Greenstein, Efficacy of submicrobial-dose doxycycline in the treatment of periodontal diseases: a critical evaluation, The international journal of Periodontics & Restorative Dentistry, 2004, 24(6):528-543; I. Needleman et al., A randomized-controlled trial of low-dose doxycycline for periodontitis in smokers, Journal of clinical Periodontoiogy, 2007, 34(4):325-333).

Finally, Arestin® is a subgingival sustained-release product composed of microspheres containing the antibiotic minocycline hydrochloride incorporated into a bioresorbable polymer, poly(glycolide-co-DL-lactide) or PGLA, for professional subgingival administration into periodontal pockets. Each unit-dose cartridge delivers minocycline hydrochloride equivalent to 1 mg of minocycline free base. The continuous drug release is described to be 14 days. Nevertheless, the product is no longer available on the German market (see https://www.parodontitis.com/vom-markt-genommen-atridoxR-actisiteR-arestinR-und-elyzolR.html). The consistency of the material (microspheres) requires application by using special cartridges and renders its handling and placement into the periodontal pocket difficult and not very reliable.

As here presented, there are a few effective therapeutic options, but they all offer opportunities for improvement. Thus, there is a high demand for the development of novel, improved forms for administration of tetracycline antibiotics with sustained activity, in particular for the treatment of periodontitis and related conditions. An ideal drug delivery system for the treatment of periodontitis should combine a simple production, an easy application, the absence of solvents and a controlled release over a prolonged period with a high bioactivity.

PROBLEMS TO BE SOLVED BY THE INVENTION

In view of the above, the present invention aims at providing a form for administration of tetracycline compounds having increased stability; and/or prolonged activity; and/or simpler, easier and/or more reliable applicability and handling. In particular, the present invention aims at providing topical application forms allowing a dentist to apply an antibiotic in a simple, reliable and as painless as possible manner. Further, significantly prolonged and uniform release is aimed at, such that the required activity level is maintained over a prolonged period of time (i.e. beyond 14 days). Finally, it would be desirable that the new forms for administration are preferably safe, reliable and sufficiently cheap to be reimbursed by the public health insurance.

Further objects of the present invention are the provision of a method for manufacturing such administration forms; the provision of a method for treatment of the human or animal body, and/or a compound or a pharmaceutical preparation for use in such method; the provision of a method for therapy or prophylaxis of a bacterial infection, and/or a compound or a pharmaceutical preparation for use in such method; the provision of a method for therapy or prophylaxis of an acute, chronic or recurrent periodontal disease and/or a compound or a pharmaceutical preparation compound for use in in such method.

SUMMARY OF THE INVENTION

The present invention provides a complex comprising a tetracycline compound (TC) or a pharmaceutically acceptable salt thereof and a divalent metal carboxylate (MA2), wherein: the molar ratio TC : MA2 is in the range of 1 : 0.8-3.0; the divalent metal cation M is an earth alkaline metal cation; and A is a carboxylate anion derived from a ¾ ¾4 carboxylic acid.

The present invention further provides a pharmaceutical preparation comprising the complex according to any one of aspects 1—4 and optionally one or more pharmaceutically acceptable excipient(s), preferably in the form of a strand-shaped extrudate

The present invention further provides a method for manufacturing said complex, the method comprising the following steps: (a) mixing a tetracycline compound (TC) or a pharmaceutically acceptable salt thereof and a divalent metal carboxylate (MA2) in a molar ratio of 1 : 0.8-3.0 with an organic solvent to obtain a dispersion, wherein the divalent metal cation M is an earth alkaline metal cation and A is a carboxylate anion derived from a C3-C24 carboxylic acid; (b) heating said dispersion to form the complex; (c) removing the solvent to obtain the complex. The present invention further provides a method for manufacturing the pharmaceutical preparation in the form of a strand shaped extrudate, the method comprising the following steps: (d) comminuting said complex and, if present, one or more pharmaceutically acceptable excipients to obtain an extrusion precursor; (e) extruding said extrusion precursor at a temperature above room temperature; (f) cooling the product of step (e) to obtain the pharmaceutical preparation in the form of a strand-shaped extrudate.

The present invention further provides a complex as defined above and/or a pharmaceutical preparation as defined above for use in a method for treatment of the human or animal body; and/or for use in a method for therapy and/or prophylaxis of a bacterial infection, preferably caused by one or more bacteria selected from the group consisting of Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, Streptococcus gordonii, Fusobacterium nucleatum, Actinomyces naeslundii and Parvimonas micra; and/or wherein antibiotic activity is maintained over a period of at least 21 days; and/or for use in a method for therapy and/or prophylaxis of an acute, chronic or recurrent periodontal disease.

EFFECTS ACHIEVED BY THE INVENTION

The present inventors found that the complexes of tetracycline compounds according to the present invention degrade more slowly in aqueous environments than the corresponding tetracycline compounds. Furthermore, these complexes were found to have a lower solubility in water and to provide a more delayed and sustained release as compared to the corresponding tetracycline compounds or even to sustained release systems, such as Arestin®. Although chelate complexes of tetracycline compounds with magnesium or calcium ions in a ratio of 1 :0.5 have been previously described, the present complexes are different in term of their stoichiometry. The complexation according to the present invention results in modified chemical properties of the compounds. In particular, the solubility, the photometric behavior and the reactivity are influenced in a favorable manner. The active substance remains biologically active over a prolonged period of time and is released in a retarded manner due to the lower solubility in water, increased stability towards degradation in aqueous solution and at the lipophilic nature of the carboxylate anion. Furthermore, the complex formation seems to prevent a conversion of the tetracycline compounds into less active epimers and thus contributes to maintaining the biological activity of the active substances over a longer period of time. These effects can be maintained when the complexes according to the present invention are

administered either alone or in combination with one or more pharmaceutical excipients in a pharmaceutical preparation, e.g., a preparation for topical administration.

Moreover, when the complex is provided in a pharmaceutical preparation such as an extrudate (either as a pure substance or together with an excipient, e.g., a biodegradable polymer), an even longer release and activity can be achieved. Such extrudates can be easily formed and handled (e.g., in order to be brought into a periodontal pocket) and additionally result in a yet further delay of drug release. Finally, they exhibit good adhesion to the oral mucosa and thus excellent application reliability.

The manufacture method described herein leads to the formation of novel tetracycline compound complexes with surprisingly pronounced sustained drug release that is therapeutically desirable. Furthermore, the complexes obtained according to the manufacture method of the present invention have an increased solubility in organic solvents and decreased water solubility. The methods for manufacturing the complexes and preparations according to present invention are simple, versatile, do not require the use of toxic organic solvents, and can be based exclusively on using well established, reliable and safe pharmaceutical ingredients. Furthermore, the complexes according to the present invention are easily formable and can be processed by extrusion, either alone or together with one or more excipient in order to be converted into extrudate having a suitable desired geometry, e.g., suitable for easier placement into a periodontal pocket.

The complexes and preparations were shown to be superior in terms of release behavior and sustained antibacterial activity, especially against oral pathogens associated with the occurrence and progress of periodontitis. This can contribute to an increased efficacy of non-surgical periodontitis therapy and consequently to a decreased amount of surgical treatment necessary, along with a significant cost reduction. Additionally, the burden for the patient caused by treatment with systemically administered antibiotics can be reduced by the substitution for an effective locally applied therapeutic form instead.

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 shows glass vials with dried minocycline/magnesium stearate complexes in different molar ratios. From left to right: minocycline hydrochloride/magnesium stearate in molar ratios of 1 :1 , 1 :2; minocycline free base/magnesium stearate in ratios of 1 :1 and 1 :2 (corresponding to Examples 4, 3, 2 and 1 , respectively).

Fig. 2 shows microphotographs of dried minocycline/magnesium stearate having a molar ratio of minocycline/magnesium stearate of 1 :1 (left, Example 2) and 1 :2 (right, Example 1 ).

Fig. 3 shows microphotographs of dried doxycycline-magnesium stearate having a molar ratio of doxycycline/magnesium stearate of 1 :1 (left, Example 6) and 1 :2 (right, Example 5).

Fig. 4 shows chromatograms of minocycline (left) and the complex according to Example 1 (right) during incubation in a buffer at pH 7.1 (top) and 2.3 (bottom) at 37°C over time (day zero always being the top curve).

Fig. 5 shows UV/Vis absorption spectra of minocycline and the minocycline complex according to Example 1 in ethanol before (left) and after (right) addition of hydrochloric acid.

Fig. 6 shows FTIR-ATR spectra of minocycline, magnesium stearate and the complex according to Example 1 in the area 680-4000 cm_l

Fig. 7 shows release curves for the extrudate of Example 9 consisting of the pure minocycline/magnesium stearate complex 1 :2, as well as together with various PLGA polymers upon extrusion (Examples 10-13).

Fig. 8 shows force-displacement diagrams upon deformation of extrudates on a texture analyzer fitted with a knife edge blade.

Fig. 9 shows a photograph of extrudates obtained according to the present invention comprising Resomer 502 RG (top) and Resomer 503 RG (bottom) containing 11.5% minocycline. Both extrudates were extruded with the 600 pm extrusion device. The larger diameter of the 503 RG extrudate indicates the viscoelastic properties of the polymer.

Fig. 10 shows an experimental assembly for assessment of the adhesive force towards mucous membranes (A) and the geometry of the flow channel specifically used for the measurements (B).

Fig. 11 shows the minimal inhibitory concentrations of eluates (dilutions) vs. Streptococcus gordonii ATCC 10558 and Porphyromonas gingivalis ATCC 33277 up to 42 days.

Fig. 12 shows the inhibitory effect of eluates obtained after 24 hours, 2 days, 7 days, 14 days and 28 days vs. biofilm formation.

Fig. 13 shows UV/VIS spectra (230—450 nm) of minocycline and doxycycline paired with magnesium- and calcium stearate and their complexes after addition of HCI. The ratios represent the molar ratios.

Fig. 14 shows FTIR-ATR-Spectra of minocycline, magnesium and calcium stearate and their complexes; 680-4000 cm ’' . Minocycline: a - u O-H; b - u N-H; c - u C-H; d - u CONh^; e - u C=C (aromatic); f - u C-N Magnesium stearate / Calcium stearate: g - u C-H; h - uas COO ; i - d C-H; j - C¾ rocking-vibration.

Fig. 15 shows FTIR-ATR-Spectra of doxycycline, magnesium and calcium stearate and their complexes; 680-4000 cm ’' . Doxycycline: a - u O-H / u N-H; b - u C-H; c - u CONH ; d - u C=C (aromatic); f - u C-N Magnesium stearate / Calcium stearate: e - u C-H; f - uas COO ; g - d C-H; h - CH rocking-vibration.

Fig. 16 shows agar plates incubated with S. aureus ATCC 29213. Left plate: Discs loaded with ethanol; Middle plate: Discs loaded with ethanolic minocycline complex solution; Right plate: Discs loaded with ethanolic minocycline solution.

Fig. 17 shows release profile of several minocycline complex PLGA compositions in PBS pH 7.0 at 37 °C.

Fig. 18 shows chromatograms of minocycline and the minocycline complex at pH 7.0 and pH 2.3 at 37 °C.

Fig. 19 shows 5 wt% suspension of minocycline HCI (left) and 5 wt% solution of disodium phosphate (right)

Fig. 20 shows Mixed minocycline HCI and disodium phosphate solution (2.5 wt% t Minocycline HCI, 2.5 wt% disodiumphosphate

Fig. 21 shows 1 wt% solution of minocycline HCI (left) and 1 wt% solution of disodium phosphate (right).

Fig. 22 shows mixed minocycline HCI and disodium phosphate solution (0.5 wt% t Minocycline HCI, 0.5 wt% disodiumphosphate.

Fig. 23 shows mixed minocycline HCI and disodium phosphate solution (0.5 wt% t Minocycline HCI, 0.5 wt% disodiumphosphate, but with addition of 400 pi 0.1 M HCI

DETAILED DESCRIPTION OF THE INVENTION

Complexes

The present invention provides a complex comprising a tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof and a divalent metal carboxylate (MA ), wherein: the molar ratio TC : MA is in the range of 1 : 0.8-3.0; the divalent metal cation M is an earth alkaline metal cation; and A is a carboxylate anion derived from a C3-C24 carboxylic acid.

A major problem with the controlled release of tetracyclines represents their chemical instability in water. For example, minocycline shows epimerization reactions with loss of activity. Also doxycycline degrades in an aqueous environment. Therefore, drug degradation prior release should be avoided. We established the hypothesis that the formation of a lipophilic complex might increase the stability of the drug and lead to prolonged release times.

Tetracycline compounds

As used herein, the term“tetracycline compound” (TC) refers to any compound belonging to the family of tetracycline antibiotics, i.e. based on the 6-deoxy-6-demethyltetracycline minimum pharmacophore structure:

Preferably, the tetracycline compound (TC) is selected from minocycline, doxycycline, tetracycline, chlortetracycline, oxytetracycline, rolitetracycline, tigecycline, demeclocycline, lymecycline, meclocycline, methacycline, omadacycline, sarecycline and eravacycline. Preferred tetracycline compounds are also selected from the group consisting of 7-chlortetracycline, 5-hydroxytetracycline, tetracycline, 6-demethyl-7-chlortetracycline, 2-N-pyrrolidinomethyltetracycline, 2-N-lysinomethyltetracycline, N-methylol-7-chlortetracycline, 6-methylene-5-hydroxytetracycline (methacycline), 6-deoxy- 5-hydroxytetracycline (doxycycline), 7-dimethylamino-6-demethyl-6-deoxytetracycline (minocycline), 9-(N,N-dimethylglycylamido)-6-demethyl-6-deoxytetracycline, 9-(N,N-dimethylglycylamido)-minocycline, 9-(t-butylglycylamido)-minocycline (tigecycline), as further described in Table 2 of Chopra et al., Microbiol Mol Biol Rev. 2001, June 65(2):232-60). Further preferred tetracycline compounds include clomocycline (N-methylol-7-chlortetracycline), 9-(N,N-dimethylglycylamido)-6-demethyl-6-deoxytetracycline and 9-(N,N-dimethylglycylamido)-minocycline.

It should be noted that the term“tetracycline compound” or“TC” as used herein denotes the general class of compounds, namely any compound belonging to the family of tetracycline antibiotics, whereas the term“tetracycline” only denotes the specific, preferred compound (4S,4aS,5aS,12aS)-4-(dimethylamino)-1 ,4,4a,5,5a,6, 11 ,12a-octahydro-3,6,10,12,12a-pentahydroxy-6-methyl-1 ,1 1-dioxo-2-naphthacenecarboxamide:

tetracycline

Preferably, the tetracycline compound (TC) is selected from minocycline, doxycycline, tetracycline, chlortetracycline, oxytetracycline, rolitetracycline, tigecycline, demeclocycline, lymecycline, meclocycline, methacycline, omadacycline, sarecycline, eravacycline, clomocycline, 9-(N,N-dimethylglycylamido)-6-demethyl-6-deoxytetracycline and 9-(N,N-dimethylglycylamido)-minocycline. These antibiotics have the advantage of being safe, clinically established and thus reliable and likely to be suitable for various therapeutic applications as described herein.

Most preferably, the tetracycline compound is minocycline (7-dimethylamino-6-demethyl-6-deoxytetracycline or (4S, 4aS,5aR,12aR)-4,7-bis(dimethylamino)-1 , 10,11 ,12a-tetrahydroxy-3,12-dioxo-4a, 5,5a, 6-tetrahydro-4H-tetracene-2-carboxamide) or doxycycline (6-deoxy-5-hydroxytetracycline or (4S,4aR,5S,5aR,6R,12aR)-4-(dimethylamino)-1 ,5,10,11 ,12a-pentahydroxy-6-methyl-3,12-dioxo-4a, 5,5a, 6-tetrahydro-4H-tetracene-2-carboxamide), and yet further preferably minocycline.

minocycline doxycycline

These antibiotics have the additional advantage that their applicability in the treatment of periodontal diseases is clinically established.

Molar Ratio

The molar ratio TC : MA2 is preferably in the range of 1 : 0.8-3.0. The present inventors found that the predominate species obtained when preparing the complexes according to the present invention are those wherein TC : MA2 is 1 :1 or 1 :2. Thus, it is intended that also mixtures wherein the molar ratio TC : MA2 is between 1 :1 and 1 :2 are also within the scope of the present invention. A minor excess of TC, e.g., wherein TC : MA2 = 1 : 0.8, is also within the scope of the present invention. Similarly, an excess of MA2, e.g., beyond a ratio of TC : MA2 of 1 :2 or even 1 :3, is also considered to be within the scope of the present invention, regardless of whether the excess metal cations are coordinated to the TC. Thus, also such mixtures containing complexes with a defined ratio of TC : MA2 together with a certain excess of one of the components are also intended to be covered by the definition of the complex according to the present invention. Preferably, TC: MA2 is 1 :1 or 1 :2; more preferably TC: MA2 is 1 :2. Further preferable ranges for the molar ratio TC : MA2 are, for instance, 1 :0.7-1 :12, 1 :0.7-1 :11 , 10.8-111 , 1 :0.9-1 :1.2, 10.9-111 , 111-12.9, 114-2.5, and 1 :1.8-1 :2.2; more preferably 10.9-111 and/or 119-12.1

Carboxylate Anion

A is a carboxylate anion derived from a C3-C24 carboxylic acid. The lipophilic nature of the carboxylate anion contributes to improving the sustained release of the tetracycline antibiotic from the complexes and preparations according to the present invention.

The carboxylic acid can be saturated or unsaturated. Preferably, A is a carboxylate anion derived from a carboxylic acid selected from arachidic acid (C20), stearic acid (C-| 3), palmitic acid (C-|g), myristic acid (C | 4), lauric acid (C-|2)> capric acid (C-| Q), caprylic acid (C3), hydroxystearic acid, oleic acid, linoleic acid, linolenic acid and ricinoleic acid. The hydroxystearic acid can be preferably 10-hydroxystearic acid or 12- hydroxystearic acid. Most preferably, the carboxylate anion is stearate (C-13).

Pharmaceutically Acceptable Salts, Hydrates and Solvates

As used herein, the term "pharmaceutically acceptable" embraces both human and veterinary use. For example, the term "pharmaceutically acceptable" embraces a veterinarily acceptable compound or a compound acceptable in human medicine and health care. Salts, hydrates and solvates of the tetracycline compounds are those wherein the counter-ion or associated solvent is pharmaceutically acceptable. However, salts, hydrates and solvates having non-pharmaceutically acceptable counter-ions or associated solvents are within the scope of the present invention, for example, for use as intermediates in the preparation of other compounds and their pharmaceutically acceptable salts, hydrates and solvates. Suitable salts include those formed with either organic and inorganic acids or bases. Pharmaceutically acceptable acid addition salts include those formed from hydrochloric, hydrobromic, sulfuric, nitric, citric, tartaric, phosphoric, lactic, pyruvic, acetic, trifluoroacetic, triphenylacetic, sulfamic, sulfanilic, succinic, oxalic, fumaric, maleic, malic, mandelic, glutamic, aspartic, oxaloacetic, methanesulfonic, ethanesulfonic, arylsulfonic (for example p-toluenesulfonic, benzenesulfonic, naphthalenesulfonic or naphthalenedisulfonic), salicylic, glutaric, gluconic, tricarballylic, cinnamic, substituted cinnamic (for example, phenyl, methyl, methoxy or halo substituted cinnamic, including 4-methyl and 4-methoxycinnamic acid), ascorbic, oleic, naphthoic, hydroxynaphthoic (for example 1- or 3-hydroxy-2-naphthoic), naphthaleneacrylic, benzoic, 4-methoxybenzoic, 2- or 4-hydroxybenzoic, 4-chlorobenzoic, 4-phenylbenzoic, benzeneacrylic (for example 1 ,4-benzenediacrylic), isethionic acids, perchloric, propionic, glycolic, hydroxyethanesulfonic, pamoic, cyclohexanesulfamic, salicylic, saccharinic and trifluoroacetic acid. Particularly preferred acid addition salts are hydrochloride, hyclate (hydrochloride hemiethanolate hemihydrate,-HCI ½EtOH ½H20) and acid addition salts formed from maleic or oxalic acid. Pharmaceutically acceptable base salts include ammonium salts, alkali metal salts such as those of sodium and potassium, alkaline earth metal salts such as those of calcium and magnesium and salts with organic bases such as dicyclohexylamine and /V-methyl-D-glucamine. All pharmaceutically acceptable acid addition salt forms of the compounds of the present invention are intended to be embraced by the scope of the present invention.

The complex is formed of a tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof, and a divalent metal carboxylate (MA ), In a preferred embodiment, the ratio TC: MA is 1 :2; and/or M is selected from Mg^+ and Ca^+; and/or A is a carboxylate anion derived from a carboxylic acid selected from arachidic acid (C ), stearic acid (C-| ), palmitic acid (C-|g), myristic acid (C |4), lauric acid (C^), capric acid (C-| Q), caprylic acid (C ), hydroxystearic acid, oleic acid, linoleic acid, linolenic acid and ricinoleic acid. Preferably, the complex consists of said tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof, and the divalent metal carboxylate (MA ),

In a further preferred embodiment, the complex has the formula [(TC)- (MgA )], wherein the tetracycline compound (TC) is minocycline or a pharmaceutically acceptable salt thereof. Preferably, TC is minocycline free base or minocycline hydrochloride, and A is stearate.

Pharmaceutical Preparations

The present invention provides a pharmaceutical preparation comprising the complex as described above and optionally one or more pharmaceutically acceptable excipient(s).

As used herein, the term "pharmaceutical preparation" is intended to encompass a product comprising the claimed compounds in the therapeutically effective amounts, as well as any product that results, directly or indirectly, from combinations of the claimed compounds.

In the pharmaceutical preparation according to the present invention, the total content of the tetracycline compound (TC) is preferably in the range of 5-30 wt.%, 30-50 wt.%, more preferably 8-28 wt.%, 32—48 wt.%, further preferably 10-20 wt.%, yet further preferably 11-15 wt.%, still further preferably 11.5 ± 5 wt.%, 27.9 wt.% ± 5 wt.%, 11.5 ± 2 wt.%, 27.9 wt.% ± 2 wt.%, 11.7-14.75 wt.%, 15.25-19.75 wt.%, and most preferably 11.5 wt.% or 27.9 wt.%, Furthermore, the overall content of the tetracycline complex (preferably, the complex having the formula [(TC) 2(MgA2)]) in the pharmaceutical preparation is in the range of 5-95 wt.%, more preferably 10-80 wt.%, further preferably 15-75 wt.%, yet further preferably 20-60 wt.%, still further preferably 30-50 wt.%, and most preferably 41.2 wt.%.

Excipients

As used herein, the term“excipient” refers to a carrier, a binder, a disintegrator and/or a further suitable additive for a galenic formulation, such as extrudate, cream, gel, emulsion, suspension, liniment, ointment, powder, paste, balm, lotion, eye drops, spray, topical aerosol, topical solution, topical suspension, skin patch and nonwoven. Carriers, which can be added to the mixture, include necessary and inert pharmaceutical excipients, including, but not limited to, suitable release retardants, suspending agents, lubricants, flavorants, sweeteners, preservatives, coatings, granulating agents, dyes, and coloring agents.

A particularly preferable class of excipients are biodegradable polymers, which will be discussed in greater detail below.

Further preferable types of excipients are emulsifiers, such as glycerol monostearate, which may be present preferably in an amount of 5 to 15 wt.%, more preferably 8 to 12 wt.%, most preferably 10 wt.%. By adding such excipients, the release behavior of the preparations, in particular extrudates, can be modulated (e.g., accelerated).

Further preferable types of excipients are plasticizers, such as polyethylene glycol (PEG), which may be present preferably in an amount of 5 to 15 wt.%, more preferably 8 to 12 wt.%, most preferably 10 wt.%. A preferable type of plasticizer is PEG 1500. By adding such excipients, the mechanical behavior of the preparations, in particular of extrudates, can be modulated (e.g., brittleness and/or a tendency towards hardening over time can be reduced).

Routes of Administration and Types of Preparations

Preferably, the pharmaceutical preparation according to the present invention is arranged for topical administration. Topical administration means application to body surfaces such as the skin or mucous membranes to treat ailments via a large range of classes including creams, foams, gels, lotions, and ointments. Many topical medications are epicutaneous, meaning that they are applied directly to the skin. Topical medications may also be inhalational, such as asthma medications, or applied to the surface of tissues other than the skin, such as eye drops applied to the conjunctiva, or ear drops placed in the ear, or medications applied to the surface of a tooth, gum or periodontal pocket.

Further preferably, the pharmaceutical preparation is selected from the group consisting of extrudate, particle, granule, powder, film, strip, compact, chip, paste, cream, gel, emulsion, suspension, liniment, ointment, balm, lotion, eye drops, spray, topical aerosol, topical solution, topical suspension, skin patch and nonwoven. Yet further preferably, the pharmaceutical preparation is arranged for application into a periodontal pocket.

Extrudates

We aimed to develop hot-melt extrudates that emulate the ideals that an ideal drug delivery system for the treatment of periodontitis should combine a simple production, an easy application, the absence of solvents and a controlled release over a prolonged period with a high bioactivity. Hot melt extrusion is a favorable technique because it is a continuous and well established process in the pharmaceutical industry. The dimensions and shapes can be adapted by the extrusion parameters. In addition, the flexibility and swelling behavior can be modified by appropriate selections of the excipients. The extrudates can be cut to the appropriate length. The insertion of the extrudates is an easy and fast process. It requires - in contrast to the application of microparticles and highly viscous gels - no special device.

As used herein, the term “extrudate” refers to any material that has been extruded through a die. Specifically, extrudates refer to dosage forms obtained by shaping a precursor by extrusion into rods which can be either cut into pieces or used as such, e.g. as a sustained release dosage form for topical or a different route of administration. Preferably, the pharmaceutical preparation is in the form of a strand-shaped extrudate, e.g., as shown in Fig. 9. The strand-shaped extrudate preferably has an essentially circular or an essentially elliptical cross section, and/or a maximum cross-sectional diameter in the range of 0.1-10 mm, 0.2-5 mm, 0.3-1 mm, 0.4-0.8 mm, 0.3-0.6 mm, more preferably 0.2-5 mm, further preferably 0.3-1 mm, yet further preferably 0.4-0.8 mm, most preferably 0.3-0.6 mm, 0.3, 0.4, 0.5 or 0.6 mm. A suitable diameter allows for easy, pain-free and reliable placement into a periodontal pocket of a given size.

Biodegradable Polymers

The pharmaceutical preparation according to the present invention preferably comprises, as a possible excipient, one or more biodegradable polymer(s). These may be selected from biodegradable polyesters, such as poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co-glycolic acid) (PLGA), polyhydroxybutyric acid (PHB) or polycaprolactone (PCL); mixed biodegradable polyesters, such as PLA-PCL and PLGA-PCL; biodegradable PEGylated diblock (AB) or triblock (ABA or BAB) copolymers, such as PEG-PLA, PEG-PLGA, PEG-PCL and PEG-PCL-PLGA; and pectins,

Poly(glycolic acid) (PGA), Poly(lactic acid) (PLA) and their copolymers are biodegradable polyesters suitable for use the present invention. They degrade in the body by simple hydrolysis of the ester backbone to non-harmful and non-toxic compounds. The degradation products are either excreted by the kidneys or eliminated as carbon dioxide and water through well-known biochemical pathways. Since PLA/PGA polymers are considered safe, non toxic and biocompatible by regulatory agencies in virtually all developed countries, additional applications of these materials can be brought to market sooner and are more cost effective than those utilizing novel polymers with unproven biocompatibility. Examples of PLA/PGA polymers suitable for use in the present invention are for instance the RESOMER polymers manufactured by Evonik Rohm Pharma GmbH shown in Table 1 below (RESOMER® Biodegradable Polymers for Medical Device Applications Research. RESOMER® Materials by Evonik Rohm GmbH, https://www.sigmaaldrich.com/technical-documents/articles/materials-science/polymer-science/resomer.html, retri eved 25.10.2018).

CLAIMS

1. A complex formed of a tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof and a divalent metal carboxylate (MA2), wherein:

the molar ratio TC : MA2 is in the range of 1 : 0.8-3.0;

the divalent metal cation M is an earth alkaline metal cation; and

A is a carboxylate anion derived from a C3-C24 carboxylic acid,

wherein the tetracycline compound (TC) is selected from minocycline, doxycycline, chlortetracycline, rolitetracycline, tigecycline, demeclocycline, lymecycline, meclocycline, methacycline, omadacycline, sarecycline, eravacycline, clomocycline, 9-(N,N-dimethylglycylamido)-6-demethyl-6-deoxytetracycline and 9- (N,N-dimethylglycylamido)-minocycline.

2. The complex according to any one of the preceding claims, wherein:

the ratio TC: MA2 is 1 :2; or

M is selected from Mg^+ and Ca^+; or

A is a carboxylate anion derived from a carboxylic acid selected from arachidic acid (C20), stearic acid (C-1 ), palmitic acid (C-|g), myristic acid (C | 4), lauric acid (C^), capric acid (C-| Q), caprylic acid (C3), hydroxystearic acid, oleic acid, linoleic acid, linolenic acid and ricinoleic acid.

3. The complex according to any one of the preceding claims, wherein:

the ratio TC: MA2 is 1 :2; and

M is selected from Mg^+ and Ca^+; and

A is a carboxylate anion derived from a carboxylic acid selected from arachidic acid (C20), stearic acid (C-13), palmitic acid (C-|g), myristic acid (C | 4), lauric acid (C^), capric acid (C4 Q), caprylic acid (C3), hydroxystearic acid, oleic acid, linoleic acid, linolenic acid and ricinoleic acid.

4. The complex according to any one of the preceding claims having the formula [(TC)-2(MgA2)], wherein the tetracycline compound (TC) is minocycline or a pharmaceutically acceptable salt, hydrate or solvate thereof.

5. A pharmaceutical preparation comprising the complex according to any one of claims 1-4 and optionally one or more pharmaceutically acceptable excipient(s).

6. The pharmaceutical preparation according to claim 5, which is arranged for topical administration and/or is selected from the group consisting of extrudate, particle, granule, powder, film, strip, compact, chip, paste, cream, gel, emulsion, suspension, liniment, ointment, balm, lotion, eye drops, spray, topical aerosol, topical solution, topical suspension, skin patch and nonwoven.

7. The pharmaceutical preparation according to claim 5 or 6 further comprising, as an excipient, one or more biodegradable polymer(s) selected from biodegradable polyesters, such as poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(lactic-co-glycolic acid) (PLGA), polyhydroxybutyric acid (PHB) or polycaprolactone (PCL); mixed biodegradable polyesters, such as PLA-PCL and PLGA-PCL; biodegradable PEGylated diblock (AB) or triblock (ABA or BAB) copolymers, such as PEG-PLA, PEG-PLGA, PEG-PCL and PEG-PCL-PLGA; and pectins, and/or wherein the total content of the tetracycline compound (TC) is in the range of 5-20 wt.%.

8. The pharmaceutical preparation according to claim 7, wherein the biodegradable polymer(s) have a weight average molecular weight range of 4-250 kDa; have a glass transition temperature of 42—48 °C; have an inherent viscosity [dl/g] (0.1 % in CHCI3 at 25 °C) of 0.03-1.70; and/or are PLGA copolymers (poly(D,L- lactide-co-glycolides)) having a L/G ratio of 5/95-95/5.

9. The pharmaceutical preparation according to claim 7 or 8, wherein the biodegradable polymer(s) are PLGA copolymers having a L/G ratio of 25/75-75/25; having a weight average molecular weight range (kDa) of 5- 90; having an inherent viscosity [dl/g] (0.1 % in CHCI3 at 25 °C) of 0.14-0.82; and/or being terminated with ester, alkyl ester or PEG groups.

10. The pharmaceutical preparation according to any one of claims 7 to 9, wherein the biodegradable polymer(s) are poly(D,L-lactide-co-glycolides) with a PLA/PGA ratio of 50:50, having a weight average molecular weight range of 7,000-38,000 and/or an inherent viscosity [dl/g] (0.1% in CHCI3 at 25 °C) of 0.16-0.44; or poly(D,L- lactide-co-glycolides) with a PLA/PGA ratio of 50:50 and a PEG end group (PLGA-PEG), having a weight average molecular weight range of 30-85 kDa and/or an inherent viscosity [dl/g] (0.1 % in CHCI3 at 25 °C) of 0.45-0.80.

11. The pharmaceutical preparation according to any one of claims 5-10, which is in the form of a strand shaped extrudate.

12. The pharmaceutical preparation according to claim 11 having an essentially circular or an essentially elliptical cross section and/or a maximum cross-sectional diameter of 0.1-10 mm, 0.2-5 mm, 0.3-1 mm, 0.4-0.8 mm, 0.3-0.6 mm, 0.2-5 mm, 0.3-1 mm, 0.6-0.9 mm, 0.4-0.8 mm, 0.3-0.6 mm, 0.3 mm, 0.4 mm, 0.5 mm or 0.6 mm.

13. A method for manufacturing the complex according to any one of claims 1—4, the method comprising the following steps:

(a) mixing a tetracycline compound (TC) or a pharmaceutically acceptable salt, hydrate or solvate thereof and a divalent metal carboxylate (MA2) in a molar ratio of 1 : 0.8-3.0 with an organic

solvent to obtain a dispersion, wherein the divalent metal cation M is an earth alkaline metal cation and A is a carboxylate anion derived from a C3-C24 carboxylic acid;

(b) heating said dispersion to form the complex;

(c) removing the solvent to obtain the complex.

14. A method for manufacturing the pharmaceutical preparation according to claim 11 , the method comprising the following steps:

(d) comminuting the complex according to any one of claims 1—4 and, if present, one or more pharmaceutically acceptable excipients to obtain an extrusion precursor;

(e) extruding said extrusion precursor at a temperature above room temperature;

(f) cooling the product of step (e) to obtain the pharmaceutical preparation in the form of a strand shaped extrudate.

15. The complex according to any one of claims 1—4 or the pharmaceutical preparation according to any one of claims 5-12 for use in a method for treatment of the human or animal body.

16. The complex according to any one of claims 1—4 or the pharmaceutical preparation according to any one of claims 5-12 for use in a method for therapy and/or prophylaxis of (i) a bacterial infection; or (ii) a bacterial infection caused by one or more bacteria selected from the group consisting of Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, Streptococcus gordonii, Fusobacterium nucleatum, Actinomyces naeslundii and Parvimonas micra.

17. The complex according to any one of claims 1—4 or the pharmaceutical preparation according to any one of claims 5-12 for the use according to claim 16, wherein antibiotic activity is maintained over a period of at least 21 days.

18. The complex according to any one of claims 1—4 or the pharmaceutical preparation according to any one of claims 5-12 for use in a method for therapy and/or prophylaxis of an acute, chronic or recurrent periodontal disease.

19. The complex according to any one of claims 1—4 or the pharmaceutical preparation according to any one of claims 5-12 for use in the method according to claim 18, wherein the periodontal disease is selected from dental plaque-induced gingival diseases, periodontitis, chronic periodontitis, aggressive periodontitis, periodontitis as a manifestation of systemic diseases, necrotizing periodontal diseases, abscesses of the periodontium, periodontitis associated with endodontic lesions, peri-implant mucositis, peri-implantitis and endodontic infections.

20. A method for treatment of the human or animal body comprising administering a therapeutically effective amount of the complex according to claim 1 or the pharmaceutical preparation according to claim 5 to a subject in need thereof.

21. The method according to claim 20, wherein the method is for therapy and/or prophylaxis of (i) a bacterial infection; or (ii) a bacterial infection caused by one or more bacteria selected from the group consisting of Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia, Streptococcus gordonii, Fusobacterium nucleatum, Actinomyces naeslundii and Parvimonas micra.

22. The method according to claim 21 , wherein antibiotic activity is maintained over a period of at least 21 days.

23. The method according to claim 20, wherein the method is for therapy and/or prophylaxis of an acute, chronic or recurrent periodontal disease.

24. The method according to claim 23, wherein the periodontal disease is selected from dental plaque-induced gingival diseases, periodontitis, chronic periodontitis, aggressive periodontitis, periodontitis as a manifestation of systemic diseases, necrotizing periodontal diseases, abscesses of the periodontium, periodontitis associated with endodontic lesions, peri-implant mucositis, peri-implantitis and endodontic infections.