PT109480B - POLYMENZIMIDAZOLE POLYMER WITH FUNCTIONED SPACER CHAIN AND ITS METHOD OF OBTAINING TO REMOVE GENOToxic IMPURITIES - Google Patents

Abstract

THE INVENTION REFERS TO A POLYMENZIMIDAZOLE POLYMER WITH A FUNCTIONED SPACING CHAIN, ACCORDING TO GENERAL FORMULA (I) AND ITS RESPECTIVE METHOD OF ACHIEVEMENT, WITH DEOXYRIBONUCLEIC ACID BASES OR COMBICATED WATER, WATER, WATER, WATER, WATER, WATER, WASH GENOToxic IMPURITIES. IN A PARTICULAR WAY THE INVENTION FOCUSES ON OBTAINING A MATERIAL, THAT ESTABLISHES SPECIFIC INTERACTIONS WITH GENOTIC IMPURITIES OF VARIOUS CHEMICAL FAMILIES, TO BE EXPLORED AS A SELECTIVE ADSORBENT.

Description

DESCRIPTION

POLYBENZIMIDAZOLE POLYMER WITH FUNCTIONED SPACER CHAIN AND ITS METHOD OF OBTAINING TO REMOVE GENOToxic IMPURITIES

TECHNICAL FIELD OF THE INVENTION

The present invention relates to a polybenzimidazole polymer (PBI) with a functionalized spacer chain and respective structural functionalization method with deoxyribonucleic acid (DNA) bases, namely adenine, thymine, cytosine or guanine, or with carboxylic acid groups (-COOH) .

PREVIOUS TECHNIQUE

The industrial production of active pharmaceutical ingredients (IFA) often takes place in an organic phase using highly reactive species, which sometimes also originate in the reaction medium itself, and which present a great diversity of functional chemical groups [Genotoxic Impurities: Strategies for Identification and Control, Andrew Teasdale, 2010, John Wiley & Sons, Chapter 9, 221-247; Guidance for Industry Genotoxic and Carcinogenic Impurities in Drug Substances and Products: Recommended Approaches, U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), December 2008]. These compounds, globally referred to as genotoxic impurities (IGT), can persist in the formulation of the final compound. A recent and comprehensive survey of IGT associated with various manufacturing processes of different APIs can be found in the literature [G. Székely et al., Chem. Rev. 115 (2015) 8182-8229].

IGTs are typically electrophilic species that, either formed during the manufacturing process of the IFA, or generated in vivo, can chemically attack the nucleophilic centers present in the DNA molecule, which can induce single or double strand breaks, and damage whose repair mechanisms DNA may not be able to reverse. All of these anomalies compromise the DNA replication process and can lead to genetic mutations, implying an increased risk to the health of the patient, who is administered the IFA [A. Teasdale et al. Org. Process Res. Dev. 17 (2013) 221-230].

Sometimes, reactive species can interact with solvents, or other reagents, present in the production of the IFA and also generate genotoxic compounds. Sulphonic acids are an example of this type of compound [A. Teasdale et al. , Org. Process Res. Dev. 14 (2010) 9991007]. When in the presence of alcohols (used in the manufacturing process in recrystallization steps, or even when washing equipment) they lead to the formation of sulfonate esters, which are substances known to be genotoxic. An example of this kind occurred in 2007 with the production of an antiviral drug Viracept, whose detailed description of the incident is found in the literature [C. Gerber et al. , Toxicol. Lett. 190 (2009) 248-253].

On the other hand, aromatic amine-type compounds constitute another group of IGT, which, although not inherently genotoxic, during their metabolism in vivo, originate electrophilic species. Generally, when these undergo an oxidation, N-hydroxyl compounds are formed which are conjugated as acetates, sulfates or glucuronides. The further decoupling of these compounds results in nitrogen ions (ArN ⁺ H), which are considered to be active genotoxins that bind to DNA [DJ Snodin, Org. Process Res. Dev. 14 (2010) 960-976].

Due to its high toxicity and persistence, the maximum permissible levels for IGT in IFA formulations are strictly regulated by the competent authorities, with a reference value of 1.5 pg / day for the toxicological risk limit [European Medicines Agency Evaluation of Medicines for Human Use, London, 28 June 2006, CPMP / SWP / 5199/02, EMEA / CHMP / QWP / 251344/2006, Committee for Medicinal Products for Human Use (CHMP), Guideline on the Limits of Genotoxic Impurities].

To eliminate these impurities, it is necessary to resort to intermediate and final purification steps in the IFA manufacturing process. The use of conventional purification techniques, such as recrystallization or distillation, for example, involves high energy costs. In addition, as these processes are not selective for the removal of IGT, significant losses of IFA also occur, with a huge economic impact on their production [N. V. V. S. S. Raman et al. , J. Pharmaceut. Biomed. Anal. 55 (2011) 662-667].

Therefore, it is necessary to develop versatile platforms for the different purification steps aimed at the selective removal of IGT, thus promoting maximum recovery of IFA with high purity. For this reason, obtaining a robust material in an organic medium capable of selectively removing an IGT, or a chemically related IGT family, presents itself as a great technological development with high potential for application in an industrial environment.

The polymeric material used in this invention is a linear polybenzimidazole (PBI) polymer that belongs to a class of heterocyclic polymers, the structure of which is shown in Figure 1. PBI is a polymer compatible with most organic solvents, with accessible functionalizable groups. For these reasons it was chosen as a platform that, after a structural modification, allows it to be explored in a purification stage in an industrial production process of an IFA, as an adsorbent material for selective removal of IGT, which has not been described yet. and whose capacity found in terms of IGT removal and selection efficiency in relation to IFA was surprisingly high for a stoichiometry of 0.25 moles of deoxyribonucleic acid base per PBI monomer.

The modification of the PBI with a DNA base is intended to mimic the interaction that occurs in vivo between an IGT, for example of the sulfonate type (electrophilic species), and DNA (nucleophilic centers).

To date, there are no similar approaches described in the literature or in industrial application. For this reason, the invention described here contains a strong component of innovation that opens up numerous possibilities with regard to selective methodologies for purifying IFAs.

The final material obtained, due to its robustness and versatility, allows it to be exploited as a highly specific adsorbent material for the IGT class, in particular the sulfonate family.

On the other hand, the polymer modified with -COOH groups allows the selective elimination of IGT from the family of aromatic amines. This guarantees the removal of electrophilic species, which would result from the in vivo metabolization of these aromatic compounds, if they were present in the IFA when administered to the patient.

The chemistry in functionalization of the PBI polymer it has been exploited for The obtaining of PBI membranes highly reticulated, per reaction of polymer with compounds dihalogenates in reflux (80 ° C) in acetonitrile (MeCN),

to give them greater physical strength, in the formulation of nanofiltration membranes resistant to organic solvents, for example [I. B. Valtcheva et al., J. Membrane Sci. 457 (2014) 62-72; I. B. Valtcheva et al., J. Membrane Sei. 493 (2015) 568-579].

US 7,259,230 and US 8,129,498 describe processes in which inorganic organic hybrid units or amide functions are inserted in the PBI structure, respectively. In both cases, the nitrogen atoms of the PBI imidazole rings are firstly ionized with an alkaline hydride (NaH) and subsequently reacted with said compounds containing a halogenated group of type RC1. In these cases, it is intended to favor the solubility of PBI in organic media in order to allow easier handling in the processing of the resulting polymers. In these two examples, the chemical functionalities introduced in the PBI molecule are directly linked to the polymers.

nitrogen atoms in the imidazole rings of the

US 4,814,400 describes the synthesis of modified PBI with carboxylic acid functions. This is achieved by first reacting the PBI with a halogenated ester, type XR-COOR ¹ . It is only after basic hydrolysis of the ester functions that the polymer functionalized with -COOH groups is obtained.

The present invention describes a direct method for obtaining modified PBI with -COOH functions in a single reaction step, in the presence of a basic compound, for example potassium carbonate (K2CO3), this approach being described for the first time, having surprisingly positive results using this weaker base.

US 2012035333A1 describes a method for obtaining PBI with carboxylic groups by reacting the polymer with cyclic acid anhydrides. In this way, a polymer with amide groups is directly linked to the PBI rings and to the free carboxylic acid groups. In the present invention, using a halogenated carboxylic acid compound with a spacer chain, there is no formation of the amide bond and the carboxylic acid function is separated from the structural structure of the PBI by two or more carbon atoms. This aspect is important to avoid stereochemical impediments that could make it difficult to connect to the IGTs that are intended to be removed.

The relevance of the present invention is related to the fact that it is possible to modify the structure of the polymer

PBI, relatively inert, in order to contain a DNA base or a carboxylic acid type functionality, through a spacer chain between the nitrogen atoms of the PBI imidazole rings and the new chemical functionality inserted.

The presence of this spacer chain presents itself as an enormous advantage, since it facilitates the interaction between the functional groups present in the adsorbent material and the IGT, in which the latter can present several spatial geometries, more or less bulky. In this way, the adsorbent is highly versatile compared to the different geometries of the different IGT molecules.

SUMMARY OF THE INVENTION

The present invention relates to a polybenzimidazole polymer with functionalized spacer chain and respective structural functionalization method with bases of DNA or carboxylic acids, chemically compatible with organic solvents, to be applied in the removal of genotoxic impurities. The invention focuses on obtaining a material, which establishes specific interactions with genotoxic impurities from various chemical families, to be exploited as a selective adsorbent of these.

In one embodiment, the present invention relates to a polybenzimidazole polymer with a functionalized spacer chain and respective structural functionalization method with deoxyribonucleic acid (DNA) bases, namely adenine, thymine, cytosine or guanine, or with carboxylic acid groups.

DESCRIPTION OF THE FIGURES

Figure 1 - Chemical structure of the polybenzimidazole polymer (PBI).

Figure 2 - Polybenzimidazole polymer with functionalized spacer chain: general formula of functionalized polybenzimidazole polymer with a spacer chain R and functional group Z. R represents a spacer chain composed of aromatic or aliphatic hydrocarbons, containing between 2 to 11 carbon atoms and Z represents a DNA base, or a carboxylic acid group.

Figure 3 - Reaction of formation of 9- (3-bromopropyl) adenine: DMF, at room temperature, K2CO3 (2 eq), Br- (CH2) _m ^_ Br (5 eq).

Figure 4 - PBI formation reaction with adenine groups: i) DMSO, 100 ° C-180 ° C, ii) K ₂ CO ₃ (1 eq), Br- (CH ₂ ) _m -Adenine (0.25 eq) , 50 ° C, iii) 100 ° C-150 ° C.

Figure 5 - PBI formation reaction with carboxylic acid groups: i) DMSO, 100 ° C-180 ° C, ii) K ₂ CO ₃ (1 eq), Br (CH ₂ ) _m -COOH (1 eq), 50 ° C, iii) 100 ° C-150 ° C.

Figure 6 - PBI ¹ H NMR spectrum in DMSO-de ·

Figure 7 - PBI ¹ H NMR spectrum with adenine groups in DMSO-de ·

Figure 8 - ¹ H NMR spectra of PBI with carboxylic acid groups, 3-bromopropionic acid, in DMSO-de ·

Figure 9 - PBI ¹ H NMR spectrum with 5-bromovaleric acid carboxylic acid groups in DMSO-de ·

Figure 10 - ¹ H NMR spectrum of PBI with carboxylic acid groups, 11-bromoundecanoic acid, in DMSO-de ·

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a polybenzimidazole polymer with functionalized spacer chain and respective structural functionalization method with bases of DNA or carboxylic acids, chemically compatible with organic solvents, to be applied in the removal of genotoxic impurities. The invention focuses on obtaining a material, which establishes specific interactions with genotoxic impurities from various chemical families, to be exploited as a selective adsorbent of these.

In one embodiment, the present invention relates to a polybenzimidazole polymer with a functionalized spacer chain and respective structural functionalization method with deoxyribonucleic acid (DNA) bases, namely adenine, thymine, cytosine or guanine, or with carboxylic acid groups.

The spacer chain is composed of aromatic or aliphatic hydrocarbons, containing between 2 and 11 carbon atoms.

The most nucleophilic positions of the DNA bases are known to be the endocyclic nitrogen at the N3 and N7 positions of guanine and adenine. These sites are preferentially reactive to the presence of electrophilic species, such as IGTs of the sulfonate type.

Surprisingly it was found that the functionalization of the PBI polymer, of the present invention, results in obtaining a specific material for its binding to certain genotoxic impurities, allowing their elimination and consequently the production of active pharmaceutical ingredients with high purity.

In the present invention, genotoxic impurities refer to families of compounds of the sulfonate or aromatic amine type.

Furthermore, the method of obtaining the polybenzimidazole polymer with functionalized spacer chain is a versatile method, in that it can be included in an industrial process, namely in the pharmaceutical industry, to eliminate or reduce the levels of genotoxic impurities. It can also be included in any synthesis industry, which exploits compounds structurally similar to IGT, for their elimination of the respective final products.

It should be noted once again that the industrial synthesis processes for obtaining active compounds in the pharmaceutical industry take place in reaction media essentially consisting of organic solvents. Currently, adsorbent materials such as resins are designed to perform well in aqueous media, being inefficient in organic media. The fact that the functionalized PBI polymers described in this invention are adsorbent materials resistant to organic solvents gives them an exceptional property to be applied in the context of treatment of post-synthetic solutions of these active pharmaceutical compounds for their purification.

A method of obtaining the polybenzimidazole polymer with a functionalized spacer chain to remove genotoxic impurities has the following steps:

a) Obtaining an aromatic or aliphatic hydrocarbon compound containing at its ends, a halogen atom and a DNA base, or a carboxylic acid group;

b) Dissolution of the polymer in a solvent, at atmospheric pressure;

c) Addition, to the solution obtained in step b), of a base, at atmospheric pressure;

d) Addition, to the solution obtained in step c), of the halogenated compound obtained in step a), at atmospheric pressure;

e) Reaction of the solution obtained in step d), up to a period of 24 h, obtaining the final solution;

f) Precipitation of the final solution with a co-solvent, followed by successive washing / filtration steps.

Preparation of a halogenated DNA base with a spacer chain composed of an aliphatic hydrocarbon.

Based on what is described in the literature, the introduction of molecules into the structure of the PBI polymer occurs by reacting the heterocyclic nitrogen atoms of the polymer with organic compounds containing a terminal halogenated functional group, of the type RX in which R can correspond to a diversity of organic compounds and possible combinations of these with inorganic compounds and X to a halogen.

DNA bases, as an example, adenine do not have a halogenated group in their molecular structure. Therefore, it is first necessary to modify these molecules in order to contain this terminal functional group -X, which will make it possible to link to the PBI, keeping intact the positions that will be used for IGT recognition. This is achieved by reacting the DNA base, for example adenine, with a dibromoalkane, of the XRX type, under controlled conditions so as to have at the end the DNA base, eg adenine, with a spacer chain containing a terminal halogenated group, as shown in Figure 3. X will have to be a good leaving group, selected from: F, Cl, Br, I; and R can be a chain with aromatic groups of the type (R-Ph-R) or a linear chain containing between 2 and 11 carbon atoms.

This same methodology can be applied to all pyrimidine and purine bases by reaction with their most nucleophilic nitrogen atoms.

In the DNA molecule, the bases form hydrogen bonds with their complementary base. In the example of adenine and its complementary base, thymine, these hydrogen bonds are established between the nitrogen atom of position 7 and by one of the two hydrogen atoms of the -NH ₂ group of position 6 of adenine (Figure 3). On the other hand, the binding of DNA bases to ribose is done by groups not involved in interactions with complementary bases, in the case of adenine by the nitrogen atom of position 9. For this reason, this was the position in which the bromoalkane-type spacer chain, RX, after reaction of adenine with a halogenated compound.

The DNA molecule with a spacer chain, which in the present invention, by way of example, refers to the compound 9- (3-bromopropyl) adenine, is described in the literature and is obtained with yields between 8% and 38% [N.

J. Leonard et al. , J. Org. Chem. 34 (1969) 3240-3248; WO 2013151663]. These low yield values contribute to the formation of by-products, such as the adenine-R-adenine dimer and a tricyclic derivative.

The present invention also describes the synthesis of this same compound, whose optimization has enabled it to be obtained with a yield greater than 60%. The development of this synthesis was achieved iteratively through the introduction of a set of innovations that are described below and that allowed to improve the performance of a synthesis that is not trivial.

To obtain 9- (3-bromopropyl) adenine, 2 equivalents (eq.) Of 1,3dibromopropane compared to adenine (in a 1 molar solution of adenine in DMF) were initially used at room temperature in the presence of 2 eq. of potassium carbonate (K ₂ CO ₃ ) for 16 hours. The insolubility of K ₂ CO ₃ in DMF did not allow verifying the complete dissolution of adenine in the 3.7 mL of solution used. However, after the reaction, the reaction mixture was filtered and both the filtrate and the solid residue were analyzed by thin layer chromatography (TLC), verifying that both phases had the same chromatographic profile in the TLC.

In order to avoid loss of the final product, it was decided that at the end of the reaction, the filtration step should not be carried out, but that the organic solvent should be completely evaporated. To remove K ₂ CO ₃ , the solid residue was then washed with acetone, filtered and evaporated. The resulting solid was washed again with methanol (MeOH, 10 ml) and a final product was obtained in 16% yield.

In an attempt to improve the reaction yield, the product was extracted with a solution of brine and dichloromethane (DCM). However, there was a formation of a precipitate in the organic phase which made it difficult to separate the phases. The organic phases were collected and, to avoid possible product losses, acetone was added to dissolve the precipitate from the organic phase. This mixture was filtered and subsequently evaporated and the residue was purified by column chromatography (CC) to obtain the final compound in 27% yield.

Then, the reaction temperature was increased to 50 ° C, but the degradation of the product after its formation and the consequent formation of more secondary products was verified by TLC, compared to the reaction at room temperature. Thus, it was decided to keep the reaction at room temperature to favor the formation of a more stable final product and avoid the formation of secondary products. At this point, the solubility of 9 (3-bromopropyl) adenine in various solvents was studied and found to be soluble in DMF, DMSO and in DCM / MeOH mixtures. Thus, a reaction was carried out in which 3 eq. 1,3-dibromopropane for adenine (in a 1 molar solution of adenine in DMF) at room temperature in the presence of 2 eq. of potassium carbonate (K ₂ CO ₃ ) for 16 hours. After this reaction the solvent was evaporated, the solid residue was washed with a solution of DCM / MeOH (40 ml) in a ratio of 10: 1 to 4: 1 (v / v), filtered, concentrated filtered and the residue was purified by CC, obtaining the final product in 35% yield. Following this same methodology, in the presence of 5 eq. 1,3-dibromopropane in relation to adenine, in DMF, the final product was obtained with a yield of 50% for the same 16 hours of reaction.

In order to increase the yield of this synthesis, 5 eq. 1,3-dibromopropane and the concentration of the initial adenine solution was increased from 1 M to 1.2 M in DMF. In this case, 3 mL of this solution was used for a reaction, monitored by TLC, which ran for only 3 hours, instead of the 16 hours previously used. The resulting product was purified by CC, using a solvent gradient starting with 100% DCM, followed by a mixture of DCM / MeOH in a 10: 1 (v / v) ratio. With this procedure the final product was obtained with a yield of 61%.

According to the present invention, if the DNA base is adenine, the step of obtaining an aromatic or aliphatic hydrocarbon compound containing at its ends, a halogen atom and a DNA base, or a carboxylic acid group comprises the following steps:

a) Dissolution of adenine in DMF, at room temperature and subsequent addition of K ₂ CO ₃ ;

b) Slow addition of an aromatic or aliphatic dihalogenated hydrocarbon;

c) Reaction of the solution obtained in step b) at room temperature, up to a period of 16 hours, obtaining the final solution;

d) Evaporation of the DMF from the final solution, obtaining a solid;

e) Adding a 10: 1 to 4: 1 (v / v) solution of DCM / MeOH to the solid obtained in step d) and filtration;

f) Evaporation of the solvent from the solution obtained in step e), obtaining a solid;

g) Purification by column chromatography of the solid obtained in step f) with a mixture of DCM / MeOH.

Functionalization of PBI based on halogenated DNA with a spacer chain composed of aliphatic hydrocarbon.

After obtaining the modified adenine base with a bromine atom at the end, this unit was incorporated into the PBI polymer. Briefly, the polymer is dissolved in dimethylsulfoxide (DMSO) between 100 ° C and 180 ° C and, subsequently, a basic compound is added, in this case, the base (K2CO3), to deprotonate the nitrogen atoms of the polymer's imidazole rings , and finally the DNA base (ie modified adenine, 9— (3 - bromopropyl) adenine) is added. In this nucleophilic substitution, compounds of the HX type are formed (eg HBr, which are released into the reaction medium, giving the connection between the DNA base (ie adenine) and the PBI through the spacer chain present in the first, as shown in the diagram. Figure 4.

After this reaction, water (40 mL) was added to the reaction medium, to promote precipitation of the modified polymer and also to remove the base in solution, and later washes were performed with MeOH (40 mL) and DCM (40 mL), to remove secondary products that have been formed as well as derivatized adenine.

In the first attempt of this synthesis, a 1% (w / v) PBI solution in DMSO was used in the presence of 2.2 eq. of K2CO3 and 1 eq. of 9- (3-bromopropyl) adenine at room temperature for 24 hours. At the end of that time, water (40 ml) was added to the reaction mixture, but no polymer precipitation occurred. Acetone (80 ml) was then added to this mixture of DMSO and water and left in the freezer (-20 ° C) for 16 hours to obtain a fine powder at the end which was filtered and dried.

To favor product formation, the concentration of the initial PBI solution was increased to 15%. This solution was placed at 170 ° C for 3 hours for complete solubilization of the polymer. After this time, the mixture was allowed to cool to room temperature and 2 eq. of K2CO3 and 1 eq. 9- (3-bromopropyl) adenine. At this stage the mixture had the properties of a viscous gel, not allowing an efficient stirring of the reaction mixture, and therefore it was necessary to increase the temperature to 50 ° C for 24 hours.

At the end of the reaction, water (40 ml) was added, obtaining a solid in the form of small particles, which was washed multiple times with water and then dried under vacuum.

This polymer (50 mg) was placed in contact with 1 ml of IGT solution in a concentration of 100 ppm in DCM, at room temperature, for 24 hours and the percentage of IGT bound to the polymer after this time was quantified. With this polymer, a preliminary value for binding to the IGT of 14% was obtained. In view of this result, he tried to optimize the reaction using 3 eq. sodium hydroxide (NaOH) replacing K ₂ CO ₃ . In this case, a final product of high hardness was obtained, impossible to be processed for use in the tests to remove IGT.

In another approach, the 15% PBI solution was added at 50 ° C 2 eq. of K2CO3 and 2 eq. of 9— (3 - bromopropyl) adenine, the reaction proceeding for another 24 hours at 80 ° C. The final product showed an IGT binding of about 24%. At this point, we tried to add the reagents and the reaction itself at a temperature of 100 ° C. A high hardness solid was obtained again, impossible to be processed for use in IGT removal tests.

It was hypothesized that the addition temperature of the reagents would be very high and it was decided to do this step at 50 ° C and increase the temperature to 100 ° C for the course of the reaction. In these latter conditions it was found that in the presence of 1 or 2 eq. of K2CO3 the polymer obtained had an IGT removal value of 59%, similar for both equivalents used. Thus, it was decided to use only 1 eq. of K2CO3 in the reaction to obtain the PBI polymer modified with adenine.

In the IGT tests, impurities from the reaction were observed, which persisted if the polymer was washed only with water or with DCM. However, after an additional wash with MeOH it was found that these impurities were polymer and did not interfere with the efficient elimination of their binding to IGT.

In the binding reaction of halogenated adenine (9- (3bromopropyl) adenine) to PBI different equivalents of halogenated adenine compared to PBI were explored to select the proportion that gives rise to the polymer modified with adenine with the best adsorption capacity for IGT sulfonate in medium organic. Values for IGT removal between 16% -97% were obtained, higher than those of the starting PBI, which showed only about 10% of IGT binding.

Functionalization of PBI with carboxylic acid with a spacer chain composed of aliphatic hydrocarbon.

The optimized reaction conditions for obtaining the PBI polymer with halogenated adenine (9- (3bromopropyl) adenine), were applied in the synthesis of the modified polymer with carboxylic acids (Figure 5). In this case, the reaction was carried out between the PBI and the halogenated compound which consisted of a carboxylic acid with a halogenated function for binding to the polymer.

The influence of several equivalents of halogenated carboxylic acid against the PBI was also determined to select the proportion that results in a removal of IGT from the aromatic amine by the PBI modified with carboxylic acid. IGT binding values were obtained between 59% -99%, higher than the starting PBI which showed only about 10% IGT binding.

For a better understanding of the invention, some examples of application of the present invention are described below for illustrative and non-limiting purposes.

Examples

As an example, the PBI polymer can be solubilized in a suitable solvent including DMSO, dimethylacetamide (DMAc), dimethylformamide (DMF) or N-methyl-2-pyrrolidone (NMP). A 15% PBI solution in DMSO proved to be adequate to perform the functionalization described in this invention.

The ionization of the nitrogen atoms of the imidazole rings, by deprotonation with a base was then carried out. K2CO3 can be used for this purpose.

The DNA base, or the carboxylic group, can contain a spacer chain composed of an aromatic and / or aliphatic hydrocarbon containing between 2 to 11 carbon atoms, with a halogenated function in a terminal position.

As an example, we present the DNA molecule with a spacer chain, [adenine- (R) -X], where R is independently selected from aromatic and / or aliphatic hydrocarbon chains containing up to 11 carbon atoms, and X is selected from the following atoms: F, Cl, Br or I.

For the reaction to occur, a minimum of 0.1 eq. modified adenine or carboxylic acid derivative, in relation to the imidazole ring nitrogen.

Preferably for removal of IGT the reaction requires about 0.25 eq. of the modified DNA base or 1.00 eq. of halogenated carboxylic acid.

Both the ionization of the imidazole groups and the alkylation can occur at temperatures between 50 ° C and 100 ° C at atmospheric pressure.

Example 1

1.1 Synthesis of 9- (3-bromopropyl) adenine

To a solution of 0.50 g of adenine (3.7 iranol) in 3 ml of DMF, in a 10 ml flask, was added 2 eq. K2CO3 (1 g, 7.2 iranol). The mixture was stirred for about 15 minutes at room temperature. After this time, 5 eq. 1,3-dibromopropane (1.9 mL, mmol). The mixture was stirred for about 3 hours at room temperature. The solvent was removed under reduced pressure and to the residue obtained about 40 ml of a 4: 1 (v / v) solution of DCM / MeOH was added. This mixture was stirred for about 10 minutes, filtered with a 3G porous plate filter and the solvent was removed under reduced pressure. The obtained residue was purified by column chromatography with the eluent 10: 1 (v / v) of DCM / MeOH to obtain 0.55 g of a white solid corresponding to the final product in a yield of 61%. The identification of the final product was performed by H and C nuclear magnetic resonance (NMR) whose spectra were obtained in a 300 MHz Bruker spectrometer for

1.2 Reaction between PBI polymer and 0.25 eq. of 9- (3bromopropyl) adenine

A solution of PBI was prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 (m-phenylene) -5,5 bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser, a magnetic stirrer was added. The system was heated and stirred magnetically, at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture was cooled to about 50 ° C, 1 eq. K2CO3 (1.7 g, 12.5 mmol). In this step, at 30 ° C the solution becomes highly viscous, not allowing the procedure to continue, and at 100 ° C the final polymer obtained has a hardness that does not allow it to be processed for IGT removal tests. During this process the mixture maintained the initial dark brown color. 0.25 eq. 9- (3-bromopropyl) adenine (0.32 mmol, 0.82 g), prepared in example 1.1, and the mixture was stirred for 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction was completed, about 80 ml of distilled water was added as a co-solvent and the polymer immediately precipitated as a yellowish brown solid which was broken manually with the aid of a spatula. This mixture was allowed to stir for a few minutes and was filtered through a 3G porous plate filter, the solid was transferred to a glass beaker (100 ml), about 80 ml of MeOH was added and allowed to stir. a few minutes being filtered then. To the resulting solid, about 80 ml of DCM was added, allowing the solution to stir for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer was separated from the mother liquor by filtration with a porous plate funnel 3G. The collected solid was allowed to air dry or was dried under reduced pressure. The final product was characterized by ¹ H NMR in DMSO-de, whose spectrum is shown in Figure 7. ¹ H NMR (300 MHz, DMSO-de): δ 2.03 (sl), 3.40 (sl), 4.01 (sl) , 7,967.64 (m, 7H), 8.35 (d, J = 6 Hz, 2H), 9.16 and 8.68 (ls, 1H). This spectrum can be compared with that obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6: NMR ^Τ Η (300 MHz, DMSO-d ₆ ): δ 7.84-7.62 (m, 6H), 8.04 (s , 1H), 8.33 (d, J = 7 Hz, 2H), 9.16 (br s, 1H), 13.28 (br s, 2NH).

1.3 Reaction between PBI polymer and 0.25 eq. of l- (3bromopropyl) thymine

A solution of PBI is prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 '- (m-phenylene) -5,5'bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser is added a magnetic stirrer. The system is heated and magnetically stirred at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture is cooled to about 50 ° C, 1 eq. K2CO3 (1.7 g, 12.5 mmol) and 0.25 eq. of 1 - (3 - bromopropyl) thymine (3.2 mmol, 0.79 g) and the mixture is stirred for up to 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction is complete, about 80 ml of distilled water is added as a co-solvent and the polymer immediately precipitates as a yellowish brown solid which is broken manually with the help of a spatula. This mixture is allowed to stir for a few minutes and is filtered through a 3G porous plate filter, the solid is transferred to a glass beaker (100 ml), about 80 ml of MeOH is added and left to stir. a few minutes and then filter. To the resulting solid, about 80 ml of DCM are added and allowed to stir for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer is separated from the mother liquor by filtration with a porous plate funnel 3G. The solid obtained is allowed to dry in air or under reduced pressure. The final product is characterized by ¹ H NMR in DMSO-de, compared to the spectrum obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6.

1.4 Reaction between PBI polymer and 0.25 eq. of l- (3bromopropyl) cytosine

A solution of PBI is prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 (m-phenylene) -5,5 bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser is added a magnetic stirrer. The system is heated and magnetically stirred at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture is cooled to about 50 ° C, 1 eq. K2CO3 (1.7 g, 12.5 mmol) and 0.25 eq. of 1 - (3 - bromopropyl) cytosine (3.2 mmol, 0.74 g) and the mixture is stirred for up to 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction is complete, about 50 ml of distilled water is added as a co-solvent and the polymer immediately precipitates as a yellowish brown solid which is broken manually with the aid of a spatula. This mixture is allowed to stir for a few minutes and is filtered through a 3G porous plate filter, the solid is transferred to a glass beaker (100 ml), about 80 ml of MeOH is added and left to stir. a few minutes and then filter. To the resulting solid, about 80 ml of DCM are added and allowed to stir for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer is separated from the mother liquor by filtration with a porous plate funnel 3G. The solid obtained is allowed to dry in air or under reduced pressure. The final product is characterized by ¹ H NMR in DMSO-de, compared to the spectrum obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6.

Example 2 Reaction between PBI polymer and 1.00 eq. of 3-bromo propionic acid

A solution of PBI was prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 '- (m-phenylene) -5,5'bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser, a magnetic stirrer was added. The system was heated and stirred magnetically, at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture was cooled to about 50 ° C, 1 eq. K ₂ CO ₃ (1.7 g, 12.5 mmol). In this step, at 30 ° C the solution becomes highly viscous, not allowing the procedure to continue, and at 100 ° C the final polymer obtained has a hardness that does not allow it to be processed for IGT removal tests. During this process the mixture maintained the initial dark brown color. 1 eq. of 3-bromopropionic acid (1 g, 13 mmol) and the mixture was stirred for 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction was completed, about 80 ml of distilled water was added as a co-solvent and the polymer immediately precipitated as a yellowish brown solid which was broken manually with the aid of a spatula. This mixture was allowed to stir for a few minutes and is filtered through a 3G porous plate filter, the solid was transferred to a glass beaker (100 ml), about 80 ml of MeOH was added and allowed to stir. a few minutes being filtered then. To the resulting solid, about 80 ml of DCM was added, allowing stirring for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer was separated from the mother liquor by filtration with a porous plate funnel 3G. The collected solid was dried under vacuum. The final product was characterized by ¹ H NMR in DMSO-de, whose spectrum is shown in Figure 8. ¹ H NMR (300 MHz, DMSO-de): δ 2.03 (sl), 8.01-7.59 (m, 9H), 9.21 and 8.81 (bs, 1H). This spectrum can be compared with that obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6.

Example 3

Reaction between PBI polymer and 1.00 eq. of 5bromovaleric acid

A solution of PBI was prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 '- (m-phenylene) -5,5'bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser, a magnetic stirrer was added. The system was heated and magnetically stirred at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture was cooled to about 50 ° C, 1 eq. K2CO3 (1.7 g, 12.5 mmol). In this step, at 30 ° C the solution becomes highly viscous, not allowing the procedure to continue, and at 100 ° C the final polymer obtained has a hardness that does not allow it to be processed for IGT removal tests. During this process the mixture maintained the initial dark brown color. 1 eq. of 5-bromovaleric acid (2.5 g, 13 mmol) and the mixture was stirred for 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction was completed, about 80 ml of distilled water was added as a co-solvent and the polymer immediately precipitated as a yellowish brown solid which was broken manually with the aid of a spatula. This mixture was allowed to stir for a few minutes and is filtered through a 3G porous plate filter, the solid was transferred to a glass beaker (100 ml), about 80 ml of MeOH was added and allowed to stir. a few minutes being filtered then. To the resulting solid, about 80 ml of DCM was added, allowing stirring for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer was separated from the mother liquor by filtration with a porous plate funnel 3G. The collected solid was dried under vacuum. The final product was characterized by ¹ H NMR in DMSO-de, whose spectrum is shown in Figure 9. ¹ H NMR (300 MHz, DMSO-de): δ 1.56 (m), 1.73 (m), 2.04 (m) , 2.30 (m), 7.95-7.64 (m, 7H), 8.35 (s, 2H), 9.18 (si, 1H). This spectrum can be compared with that obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6.

Example 4

Reaction between PBI polymer and 1.00 eq. 11bromoundecanoic acid co

A solution of PBI was prepared, in a 25 ml flask, by solubilizing 2 g of poly-2,2 '- (m-phenylene) -5,5'bisbenzimidazole (commercially available) in 13 ml of DMSO. To the flask with a reflux condenser, a magnetic stirrer was added. The system was heated and magnetically stirred at atmospheric pressure, up to 3 hours, at a temperature between 100 ° C and 180 ° C. After this time, the mixture was cooled to about 50 ° C, 1 eq. K ₂ CO ₃ (1.7 g, 12.5 mmol). In this step, at 30 ° C the solution becomes highly viscous, not allowing the procedure to continue, and at 100 ° C the final polymer obtained has a hardness that does not allow it to be processed for IGT removal tests. During this process the mixture maintained the initial dark brown color. 1 eq. 11-bromoundecanoic acid (3.4 g, 13 mmol) and the mixture was stirred for 24 hours at a temperature between 100 ° C and 150 ° C. After the reaction was completed, about 80 ml of distilled water was added as a co-solvent and the polymer immediately precipitated as a yellowish brown solid which was broken manually with the aid of a spatula. This mixture was allowed to stir for a few minutes and is filtered through a 3G porous plate filter, the solid was transferred to a glass beaker (100 ml), about 80 ml of MeOH was added and allowed to stir. a few minutes being filtered then. To the resulting solid, about 80 ml of DCM was added, stirring for a few minutes and then filtered to obtain the final polymer. Between each wash the polymer was separated from the mother liquor by filtration with a porous plate funnel 3G. The collected solid was dried under vacuum. The final product was characterized by ² Η NMR in DMSO-de, whose spectrum is shown in Figure 10. ² Η NMR (300 MHz, DMSO-de): δ 2.26-1.03 (m), 3.81 (si), 7.93- 7.63 (m, 7H), 8.35 (si, 2H), 9.19 (si, 1H). This spectrum can be compared with that obtained for the PBI starting product in the same deuterated solvent, shown in Figure 6.

Example 5

Determination of removal efficiency, percentage of binding to IGT and adsorption capacity of modified PBI polymers

In these tests, the polymers obtained in examples 1.2 and 2 were used. For the polymer with adenine groups, example 1.2, the IGT used as an example was methyl p-toluene sulfonate (MPTS), a compound representing the sulfonate-type IGTs. For the polymer with carboxylic acid groups, example 2, 4-dimethyl aminopyridine (DMAP), a representative compound of the aromatic amine type IGTs, was used.

To 50 mg of each polymer, 1 ml of a solution of the respective IGT, with a concentration of 100 ppm, prepared in DCM was added. This suspension was stirred for 24 hours at room temperature and 200 rpm. After this time, the mixtures were centrifuged (13,900 rpm, 35 minutes) and the supernatant was filtered, with syringe filters and analyzed by HPLC (High Pressure Liquid Chromatography). These analyzes were performed on a Merck Hitachi equipment coupled to a UV L-2400 detector with a Macherey-Nagel C18 reverse phase analytical column (Nucleosil 100-10, 250 x 4.6 mm).

The percentage of IGT binding was determined by the difference between the initial concentration and the equilibrium concentration, present in the solution resulting from the test, according to the equation:

% Removal = (C ₀ -C _e ) / C ₀ * 100 where Co (mg / L) is the initial concentration of IGT and C _and (mg / L) is the equilibrium concentration of IGT in solution.

The adsorption capacity of each polymer was determined according to the following equation:

q _e = V [C ₀ -C _e ] / M where q _e (mg / g) is the amount of IGT bound to the polymer, Co (mg / L) is the initial concentration of IGT, C _e (mg / L ) is the equilibrium concentration of IGT in solution, V (L) is the volume of solution used and M (g) is the mass of polymer used in the test.

From the results present in Table 1, it was found that the polymer obtained from the reaction with 0.25 eq of 9— (3 - bromopropyl) adenine shows the best percentage of MPTS removal, corresponding to about 96%. For the remaining polymers, the percentage of IGT removal was between 16% -78%. In either case, the removal of IGT was always superior to that observed for the starting PBI polymer, used as a control, with only 10% removal of

IGT. Controls were also made with IGT solutions processed in the same way, but in the absence of polymer, to detect IGT losses by adsorption on the walls of vials, filters or evaporation, in which case the removal is negligible.

Table 1 - Percentage of removal and adsorption capacity of PBI polymers with adenine for IGT methyl ptoluenesulfonate from a solution with a concentration of 100 ppm in DCM.

Equivalents of 9- (3-bromopropyl) adenine added in reaction with PBI Removal (%) q _e (mg MPTS / g polymer) PBI 10 ± 2.8 0.22 ± 0.06 0.10 16 ± 0.4 0.36 ± 0.02 0.25 97 ± 0.2 2.18 ± 0.02 0.50 78 ± 0.1 1.74 ± 0.01 0.75 53 ± 1.3 1.10 ± 0.04 1, 00 59 ± 2.4 1.38 ± 0.07

According to the data it appears that, for DMAP derived from carboxylic acid in the adsorbent with the best IGT, presenting the IGT values at a value of about 99%.

Table 2 shows the use of 1 eq. the reaction with PBI results in characteristics for removal; higher removal of

For the remaining polymers, formed in the presence of different equivalents of carboxylic acid, the binding to IGT varied between 59% -87%.

The results show that polymers containing carboxylic acid groups have a greater affinity for IGT when compared to the starting PBI, used as a control, which shows an IGT removal of only 10%.

Table 2 - Percentage of removal and adsorption capacity of PBI polymers with carboxylic acid for IGT 4dimethyl aminopyridine from a solution with a concentration of 100 ppm in DCM.

Equivalents of 3-bromopropionic acid added in the reaction with PBI Removal (%) q _e (mg DMAP / g polymer) PBI 10 ± 6, 1 0.23 ± 0.14 0.25 64 ± 1.8 0.90 ± 0.02 0.50 89 ± 0.5 1.31 ± 1.57 0.75 87 ± 0.9 1.23 ± 0.02 1, 00 99 ± 0.1 2.37 ± 0.03

Example 6

Removal efficiency, percentage of binding of PBI polymer functionalized with adenine to IGT methyl methanesulfonate in batch and in column chromatography.

In this example, the polymer obtained in example 1.2 was used. The IGT used was methyl methanesulfonate (MSM), a compound belonging to the sulfonate type IGTs.

For the batch procedure, 50 mg of the polymer was added 1 ml of a solution of IGT, with a concentration of 100 ppm, prepared in DCM. The suspension was stirred for 24 hours at room temperature and 200 rpm. After that time the mixture was centrifuged (13,900 rpm, 35 minutes) and the supernatant was filtered, with syringe filters and analyzed by HPLC, as described in the example

5.

For the chromatographic column procedure, 100 mg of polymer was conditioned in a glass column, through which 2 ml of a solution of IGT, with a concentration of 100 ppm, prepared in DCM was passed. The resulting column eluate was analyzed by HPLC, as described in example 5.

A percentage of IGT removal by binding to the polymer of about 96% was determined in both the batch and chromatographic column assays.

Claims (14)

1. Polybenzimidazole polymer with functionalized spacer chain characterized by having the following general formula (I): On what: a) R represents a spacer chain composed of aromatic or aliphatic hydrocarbons, containing between 2 to 11 carbon atoms; b) Z represents a DNA base, or a carboxylic acid group. Polybenzimidazole polymer according to claim 1, characterized in that the DNA base is selected from the following group: adenine, thymine, cytosine or guanine. 3. Method of obtaining the polybenzimidazole polymer with functionalized spacer chain for removal of genotoxic impurities, as defined in claims 1 and 2, characterized by the following steps: a) Obtaining an aromatic or aliphatic hydrocarbon compound containing at its ends, a halogen atom and a DNA base, or a carboxylic acid group; b) Dissolution of the polymer in a solvent, at atmospheric pressure; c) Addition, to the solution obtained in step b), of a base, at atmospheric pressure; d) Addition, to the solution obtained in step c), of the halogenated compound obtained in step a), at atmospheric pressure; e) Reaction of the solution obtained in step d), up to a period of 24 h, obtaining the final solution; f) Precipitation of the final solution with a co-solvent, followed by successive washing / filtration steps. Method according to claim 3, characterized in that, in case the DNA base is adenine, step a), comprises the following steps: a) Dissolution of adenine in DMF, at room temperature and subsequent addition of K ₂ CO ₃ ; b) Slow addition of an aromatic or aliphatic dihalogenated hydrocarbon; c) Reaction of the solution obtained in step b) at room temperature, up to a period of 16 hours, obtaining the final solution; d) Evaporation of the DMF from the final solution, obtaining a solid; e) Adding a 10: 1 to 4: 1 (v / v) solution of DCM / MeOH to the solid obtained in step d) and filtration; f) Evaporation of the solvent from the solution obtained in step e), obtaining a solid; g) Purification by column chromatography of the solid obtained in step f) with a mixture of DCM / MeOH. 5. Method according to claim 3, characterized in that the solvent used for the dissolution of PBI, in step b) is selected from the group of: dimethylsulfoxide (DMSO), N-methyl-2-pyrrolidone (NMP), dimethylacetamide (DMAc). 6. Method according to claim 3, characterized in that step b) is carried out at a temperature between 100 and 180 ° C. Method according to claim 3, characterized in that step b) runs for up to 3 hours. 8. Method according to claim 3, characterized in that step c) is carried out at a temperature between 30 and 100 ° C. Method according to claim 3, characterized in that the base used in step c) is a carbonate. 10. Method, according to step d) 100 ° C. 11. Method, according to step e) 150 ° C. 12. Method, according to whether the co-solvent used in claim 3, characterized at a temperature between 30 and claim 3, characterized at a temperature between 100 and claim 3, characterized in step f) is water. Method according to claim 3, characterized in that the washing solvents used in the successive washing / filtration steps are water, methanol and dichloromethane, respectively. 14. Use of the polybenzimidazole polymer, as defined in claims 1 and 2, characterized in that it is used to remove genotoxic impurities of the sulfonate or aromatic amine type.