ES3021807B2 - CARBOSILANE DENDRIMER AND ITS USE FOR THE ELIMINATION OF PATHOGENIC MICROORGANISMS IN SUSPENSION OR IN BIOFILM - Google Patents

Description

[0001] DESCRIPTION

[0004] CARBOSILANE DENDRIMER AND ITS USE FOR THE ELIMINATION OF

[0005] PATHOGENIC MICROORGANISMS IN SUSPENSION OR IN BIOFILM

[0007] TECHNICAL SECTOR

[0008] The present invention relates to the eradication of cells in suspension and attached to a surface, including mixed biofilms (yeasts and bacteria in a common culture), using a cationic carbosilane dendrimer having on its periphery ammonium groups modified with a hydroxyl function (compound BDNG001).

[0010] PRIOR ART STATEMENT

[0011] The elimination of pathogenic microorganisms, especially the most concerning and clinically relevant ones, is an increasingly complex process due to the emergence of antimicrobial resistance. Therefore, current treatments often require the use of highly toxic compounds or extremely high concentrations to achieve complete eradication, particularly if these microorganisms are eukaryotes or present as microbial biofilms. For this reason, it is essential to develop new alternatives that can promote the elimination of potentially problematic microorganisms while minimizing environmental impact and, in particular, side effects in individuals receiving these antimicrobial therapies.

[0013] The use of dendritic systems with antibacterial and antifungal activity is widely recognized in the literature (de la Mata, F.J. WIREs Nanomed Nanobiotechnoly 2022, e1871). Similarly, their antibiofilm activity has been observed with promising results (Heredero-Bermejo, I et al. Pharmaceutics. 2020, 12(10):918; Gómez-Casanova, N. et al. JFungi (Basel). 2021)

, 7(7):574; Gómez-Casanova, N. et al. Pharmaceutics2022

(14, 1604). Generally, these compounds are systems that have positive charges on the periphery along with a hydrophobic backbone, such as carbosilane (CBS), which facilitates interaction with the bacterial membrane. An important advantage of dendritic systems is that they are polyfunctional molecules with a well-defined structure (de la Mata, F.J.WIREs Nanomed Nanobiotechnol.2022, e1871).

[0015] The formation of these molecules is usually carried out by a series of repetitive steps in which the degree of functionalization of the dendritic system is increased each time, giving rise to molecules of a globular nature with branches that arise from a central multivalent group.

[0017] The process of treating and eliminating yeasts is quite complex and complicated, especially since they are eukaryotic cells. Therefore, most antifungals currently used are highly toxic to the cells of the treated individual, which can sometimes also be targets of these treatments (Kagan, S. et al. Antimicrob Agents Chemother. 2012, 56(11):5603-11). In addition, biofilms, especially mixed biofilms formed by yeasts and bacteria, pose an even greater problem because their treatment requires very high concentrations of the compound to be completely destroyed (Lara HH, et al. Pathogens. 2020, 9(10):784).

[0019] The present invention arises from the need to provide an innovative solution for eliminating microorganisms, both in their planktonic form and in their biofilm state. Thus, the invention provides a new compound capable of acting as an active ingredient in antiseptics and/or disinfectants, being effective both for the treatment of a single species and for the treatment of microbial communities composed of different species.

[0021] DESCRIPTION OF THE INVENTION

[0022] The compound to which the invention refers is a CBS dendritic system, similar to those described in WO2014016460, but in this invention it is a cationic dendritic system with hydroxyl groups on the periphery attached to the ammonium group by an alkyl chain.

[0024] In particular, a first object of the invention is a cationic carbosilane dendrimer (hereinafter also referred to as BDNG001) comprising a cyclotetrasiloxane core, wherein each silicon atom of the core is linked by a carbosilane (CH<2>)<2>{Si(Me)<2>}(CH<2>)<2>S(CH<2>)<2> structure to a peripheral structure comprising an ammonium group linked by an alkyl chain (CH<2>)<2> to a hydroxyl group, according to the following formula:

[0025]

[0028] In the present invention, "dendritic compound" refers to a highly branched macromolecule where the growth units, branches or ramifications have a carbosilane backbone.

[0030] In particular, the dendrimer of the present invention is cationic. Therefore, it may comprise counterions to neutralize its charge. Thus, in particular embodiments of the invention, the claimed dendrimer comprises at least one anion selected from a group consisting of at least one halide (preferably chloride, bromide, or iodide), hydroxide, triflate, nitrate, and sulfate, as well as any combinations thereof.

[0032] The invention also relates to the use of the claimed carbosilane dendrimer (or a pharmaceutical composition comprising it) as an antibacterial or antifungal agent, i.e., for the elimination of pathogenic microorganisms (including yeasts and bacteria), either suspended in a liquid medium (such as planktonic cells) or attached to a surface in a biofilm state. A “biofilm” is understood to be a collection of highly organized cells attached to a surface, which may be biotic or abiotic. Biofilms may be composed of a single species of microorganism or several, surrounded by an extracellular matrix. Eradicating a biofilm is extremely difficult, especially once it has become established, and this difficulty is even greater with mixed biofilms, i.e., those composed of more than one species. Their treatment is very limited because either the compounds are toxic or high concentrations of the biocide are required for their elimination. Furthermore, its presence increases the acquisition of resistance, another major problem currently encountered. Thus, one of the main advantages of the invention is that it has proven effective in treating mixed biofilms, that is, those composed of different microorganisms such as yeasts and bacteria. The compound's effectiveness against biofilms has been demonstrated using colorimetric and agar plating methods. The claimed dendrimer is completely water-soluble and exhibits no interaction with the media required for the in vitro study of microorganisms. Additionally, it is a compound with low cytotoxicity.

[0034] In particular, the pharmaceutical composition comprising the dendrimer of the invention may further comprise at least one second active ingredient selected from a group consisting of an antibiotic, anti-inflammatory and antiviral, as well as any combinations thereof.

[0036] The invention also relates to a method for the removal of at least one biofilm made up of a single species of microorganisms or at least two different species of microorganisms, either during its formation or once it has formed, characterized in that it comprises applying to the biofilm a solution (preferably aqueous) of the dendrimer that is the subject of the invention for a time preferably between 1 hour and 72 hours.

[0038] Additionally, the cationic carbosilane dendrimer of the invention is the object of the invention for use as a medicament, as well as for use in the prevention and/or treatment of at least one disease caused by at least one pathogenic microorganism.

[0040] The invention also relates to the use of the claimed dendrimer for the manufacture of a medicament for the treatment of at least one disease caused by at least one pathogenic microorganism.

[0042] Finally, the invention relates to the use of the claimed dendrimer as a carrier for at least one drug, preferably with antibacterial and/or antifungal properties.

[0044] NOMENCLATURE IN BIO-FILMS

[0045] The nomenclature defined below for biofilms has been previously standardized in several publications mentioned above, as well as in a book chapter (Gómez-Casanova, N., Candida and Candidiasis, 2022, chapter in IntechOpen). Dendritic compounds with these characteristics have been evaluated using the resazurin colorimetric method based on cell metabolism to quantify the viability of biofilm cells (quantitative method), and the drop plate method (agar plate inoculation, qualitative method).

[0047] Nomenclature for the treatment of forming biofilms:

[0048] ⋅ CMIB: Minimum Biofilm Inhibitory Concentration, defined as the minimum concentration capable of inhibiting biofilm formation, but which may not kill the cells. It is determined using the resazurin colorimetric method.

[0049] ⋅ CMBB or CMFB: Minimum Bactericidal or Fungicidal Biofilm Concentration, defined as the minimum concentration capable of preventing the formation of a biofilm and killing all cells. It is determined by the drop plate method.

[0051] Nomenclature for the treatment of previously established biofilms:

[0052] ⋅ CMDB: Minimum Concentration that Damages a Biofilm, defined as the minimum concentration capable of seriously damaging an established biofilm, but which might not kill all cells. It is determined by the colorimetric method with resazurin.

[0053] ⋅ CMEB: Minimum Concentration for Biofilm Eradication, defined as the minimum concentration capable of completely eradicating a previously established biofilm. In this case, all cells comprising the biofilm would be dead. This is the most difficult target to achieve in in vitro and in vivo studies of new compounds. It is determined by the drop plate method.

[0055] DESCRIPTION OF THE FIGURES

[0056] Figure 1. Percentage of viability of BDNG001 in previously established biofilms of C. albicans.

[0057] Figure 2. Percentage of viability of BDNG001 in previously established biofilms of S. aureus.

[0058] Figure 3. Percentage of viability of BDNG001 in previously established mixed biofilms of C. albicans and S. aureus.

[0059] Figure 4. Percentage of cell viability in the BDNG001 HeLa cell line at 24 hours.

[0060] Figure 5. Percentage of cell viability in the BDNG001 HeLa cell line at 48 hours.

[0061] EXAMPLES

[0062] The synthesis procedures of the cationic dendrimer BDNG001 are described below, as well as the study of its antifungal, antibacterial and antibiofilm activity against some pathogenic microorganisms.

[0064] Example 1.- Synthesis of the dendrimer BDNG001 (G0Si4O4Si(S-NMe2OHBr)4).

[0065] This compound can be obtained from G0Si4O4Si(S-NMe2)4(0.150 g, 0.13 mmol) (Heredero-Bermejo et al. Pharmaceutics 2020, 12, 918) by reaction with Br(CH<2>)<2>OH (0.077 g, 0.62 mmol), without ruling out other agents that introduce the ethanol group, preferably in THF (tetrahydrofuran) as a solvent, or others such as DMF (dimethylformamide), acetone, DMSO (dimethyl sulfoxide), heating for several days (e.g.

[0066] 3 days at 60ºC). Product BDNG001 is obtained as a pale yellow solid.

[0068] Example 2.- Microbial activity of BDNG001 in planktonic cells

[0069] The use of the compound of the invention, BDNG001, a dendritic system with –OH groups on its periphery, is proposed for the treatment of microorganisms, both prokaryotes (bacteria) and eukaryotes (yeast-like fungus), as an alternative to traditionally used drugs against these microorganisms. The compound is suitable for the evaluation of its antibacterial activity based on ISO 20776-1:2006. For this purpose, bacteria were grown in Mueller-Hinton liquid medium and, after 24 hours of incubation, the amount of microorganism was adjusted to an inoculum of 10<8 CFU/mL of bacteria. Subsequently, 2 mL of the adjusted inoculum was added to 8 mL of sterile distilled water. Finally, 100 µL of the compound, 100 µL of Mueller-Hinton medium (twice concentrated), and 5 µL of the adjusted inoculum were added to a multiwell plate. The plates were incubated for 24 hours at 37°C. After that time, the absorbance was quantified at 630 nm.

[0071] On the other hand, the standardized methods for studying antifungal susceptibility are based on the European Committee on Antimicrobial Susceptibility Testing (EUCAST). For this purpose, yeasts were grown on a Sabouraud agar plate. Then, several colonies were collected and homogenized in sterile distilled water until a density of 0.5 McFarland (10⁶ CFU/mL) was reached (inoculum). Next, a 1:10 dilution was made in RPMI-1640 twice-concentrated medium, supplemented with MOPS and glucose. Finally, 100 µL of the compound and 100 µL of the adjusted inoculum were added to a multiwell plate. The plates were incubated for 24 hours at 37 °C. After this time, they were analyzed at 530 nm.

[0072] Being a water-soluble compound, it does not require any procedure to improve its solubility during the testing process. Concentration gradients up to 512 mg/L were tested in both bacteria and yeast. Each concentration was performed in triplicate, as was the untreated bacterial control. Control wells of water, culture medium, and compound were also included to monitor for possible contamination or interference of the compound on the absorbance reading.

[0074] For these tests, a strain of Candida albicans from the Spanish Collection of Type Strains (CECT1002), a strain of Staphylococcus aureus (CECT240) and a strain of Escherichia coli (CECT515) were used.

[0076] Example 3.- Activity of BDNG001 on forming and established bacterial and fungal biofilms

[0077] The use of the compound of the invention, BDNG001, a dendritic system with –OH groups on its periphery, is proposed for the treatment of bacterial and fungal biofilms as an alternative to traditionally used drugs. The compound is suitable for evaluating its activity against both developing and established biofilms. The procedures and nomenclature commonly used to test antibiofilm activity are described in Heredero-Bermejo, I et al. Pharmaceuticals.

[0078] 2020,12(10):918; Fernandez, J. et al.Polymers (Basel).2021

, 13(13):2127; Gómez-Casanova, N. et al. JFungi (Basel).2021

, 7(7):574; Gómez-Casanova, N., Candida and Candidiasis, 2022 Chapter in IntechOpen). For these tests, the yeast C. albicans (CECT1002) was used as the representative eukaryotic strain and the bacterium S. aureus (CECT240) as the representative prokaryotic strain.

[0080] Biofilms can be treated before they form, either as a preventative measure or once established. For the treatment of forming biofilms in S. aureus bacteria, cells were grown on Mueller-Hinton medium. The following day, the inoculum was adjusted to 0.5 McFarland. A 1:100 dilution was then performed in Tryptic Soy Broth medium supplemented with twice-strength glucose. Finally, 50 µL of compound BDNG001 and 50 µL of the bacterial suspension were added to a Nunc flat-bottom multiwell plate. The plates were incubated for 24 hours at 37°C. After this time, the biofilms were carefully washed with phosphate-buffered saline (PBS). Next, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin (previously filter-sterilized). The microplates were incubated for 20 hours in the dark, and measurements were taken at 570 nm and 600 nm. These data provided the CMIB value. Finally, 5 µL from each well were plated onto plate count agar (PCA) and incubated at 37 °C for 24 hours. These data provided the CMBB value.

[0082] On the other hand, the standardized methods for studying yeast biofilm development present slight differences. Candida was grown overnight in YPD medium (yeast extract, peptone, and dextrose) and adjusted to 0.5 McFarland in RPMI-1640 medium supplemented with MOPS and glucose. Then, 50 µL of compound BDNG001 and 50 µL of the fungal suspension were added to a flat-bottomed Nunc multiwell plate. The plates were incubated for 48 hours at 37°C. After this time, the biofilms were carefully washed with PBS. Then, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin. The multiwell plates were incubated for 20 hours in the dark and measured at 570 nm and 600 nm. These data provided the CMIB value. Finally, 5 µL from each well were plated onto Sabouraud agar plates and incubated at 37 °C for 24 hours. These data provided the CMFB value.

[0084] In both bacteria and yeast, concentration gradients up to 512 mg/L (maximum concentration tested) were tested. Each concentration was performed in triplicate, as well as the untreated bacterial control. Control wells of water, culture medium, and compound were also included to monitor for possible contamination or interference from the compound on the absorbance reading.

[0086] For the treatment of previously established S. aureus biofilms, cells were grown overnight in Mueller-Hinton medium. The following day, the inoculum was adjusted to 0.5 McFarland. A 1:100 dilution was then performed in Tryptic Soy Broth medium supplemented with glucose. Next, 100 µL of the bacterial suspension was added to a flat-bottom Nunc multiwell plate. The plates were incubated for 24 hours at 37°C. After this time, the biofilms were carefully washed with PBS and treated with compound BDNG001. The multiwell plates were again incubated for 24 hours at 37°C. Finally, the biofilms were carefully washed again with PBS. Next, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin. The multiwell plates were incubated for 20 hours in the dark and measured at 570 nm and 600 nm. These data provided the CMDB value. Finally, 5 µL from each well were plated onto PCA agar plates and incubated at 37 °C for 24 hours. These data provided the CMEB value.

[0088] On the other hand, the previously established standardized methods for the study of yeast biofilms again show slight differences. Candida was grown overnight in YPD medium and adjusted to 0.5 McFarland in RPMI-1640 medium supplemented with MOPS and glucose. Finally, 100 µL of the fungal suspension was added to a flat-bottomed Nunc multiwell plate. The plates were incubated for 48 hours at 37 °C. After this time, the biofilms were carefully washed with PBS and treated with the concentration gradient of compound BDNG001. Again, the plates were incubated for 48 hours at 37 °C. After this time, the wells were washed with PBS. Next, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin. The multiwell plates were incubated for 20 hours in the dark, and the absorbance was measured at 570 nm and 600 nm. These data provided the CMDB value. Finally, 5 µL from each well were plated onto Sabouraud agar plates and incubated at 37 °C for 24 hours. These data provided the CMEB value.

[0090] In both bacteria and yeast, concentration gradients up to 1024 mg/L (maximum concentration studied) were tested. Each concentration was performed in triplicate, as well as the untreated bacterial control. Control wells of water, culture medium, and compound were also included to monitor for possible contamination or interference of the compound on the absorbance reading.

[0092] Example 4.- Activity of BDNG001 on mixed biofilms in formation and established

[0093] The use of the compound of the invention, BDNG001, a dendritic system with –OH groups on its periphery, is proposed for the treatment of mixed biofilms as an alternative to traditionally used drugs. The compound BDNG001 was tested on mixed eukaryotic-prokaryotic biofilms, with C. albicans as the eukaryotic representative and S. aureus as the prokaryotic microorganism. The results obtained were based, as previously mentioned, on the nomenclature described in the chapter by Gómez-Casanova, N., Candida and Candidiasis, 2022, published in IntechOpen, and various articles in the literature (Heredero-Bermejo, I et al. Pharmaceuticals. 2020, 12(10):918; Fernandez, J. et al.

[0094] Polymers (Basel).2021

, 13(13):2127; Gómez-Casanova, N. et al. JFungi (Basel).2021

, 7(7):574).

[0096] For the treatment of forming mixed biofilms, the inoculum of each species must initially be adjusted separately. First, the S. aureus (bacteria) species, previously grown in Mueller-Hinton liquid medium, was adjusted to 0.5 McFarland. A 1:100 dilution was then performed. Second, C. albicans (yeast), previously grown in YPD medium, was adjusted to 0.5 McFarland. Finally, both species were mixed in equal parts, resulting in a 1:1 adjusted inoculum. The experiment was performed using RPMI-1640 medium. After homogenizing the mixture, 50 µL of the inoculum suspension and 50 µL of the compound were added to a flat-bottomed Nunc multiwell plate. The plates were incubated for 24 hours at 37 °C. After this time, the biofilms were carefully washed with PBS. Then, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin (previously sterilized by filtration). The multiwell plates were incubated for 20 hours in the dark, and the absorbance was measured at 570 nm and 600 nm. These data provided the CMIB value. Finally, 5 µL from each well were plated onto PCA agar and Sabouraud agar plates and incubated at 37 °C for 24 hours. These data provided the CMBB/CMFB value. Concentration gradients were tested up to 512 mg/L (maximum concentration tested).

[0098] For the treatment of previously established mixed biofilms, the inoculum of each species must initially be adjusted separately. First, the S. aureus (bacteria) species, previously grown in Mueller-Hinton liquid medium, was adjusted to 0.5 McFarland. A 1:100 dilution was then performed. Second, C. albicans (yeast), previously grown in YPD medium, was adjusted to 0.5 McFarland. Finally, both adjusted species were mixed in equal parts (1:1). The experiment was performed using RPMI medium. After homogenizing the mixture, 100 µL of the suspension was inoculated into a flat-bottomed Nunc multiwell plate. The plates were incubated for 24 hours at 37 °C. After this time, the biofilms were carefully washed with PBS and treated with compound BDNG001. The multiwell plates were incubated again for 24 hours at 37°C. After this time, the biofilms were carefully washed again with PBS. Then, 100 µL of PBS was added to each well along with 20 µL of 0.01% resazurin (previously sterilized by filtration). The multiwell plates were incubated for 20 hours in the dark, and the absorbance was measured at 570 nm and 600 nm. These data provided the CMDB value. Finally, 5 µL from each well were plated onto PCA agar and Sabouraud agar plates and incubated at 37°C for 24 hours. These data provided the CMEB value. Concentration gradients were tested up to 1024 mg/L (the maximum concentration studied).

[0100] In both procedures, treatment of forming mixed biofilms and previously established mixed biofilms, each concentration was performed in triplicate, as well as the untreated bacterial control. Control wells of water, culture medium, and compound were also included to monitor for possible contamination or interference of the compound on the absorbance reading.

[0102] Example 5.- Biocompatibility of BDNG001

[0103] The biocompatibility of the main compound of the invention, BDNG001, was studied. For this purpose, HeLa cells (ATCC® CCL-2™) were used. The assays were performed in 24-well plates (NUNC™) in Dulbecco’s Modified Eagle medium supplemented with 10% fetal bovine serum and 1% of an antibiotic mixture composed of: 10,000 U penicillin, 10 mg streptomycin, and 25 μg AmB per mL. The cells were adjusted to a density of 1 × 10⁴ cells/well in 500 μL of fresh medium. The plates were incubated at 37°C and 5% CO₂ for 5 days, until a confluent monolayer was obtained. After the incubation period, the medium was discarded and a concentration gradient of the diluted compound was added to fresh medium. The control contained only fresh medium. The plates were incubated for 24 and 48 hours. Afterward, the medium was discarded and the wells were carefully washed with PBS. Finally, 500 μL of fresh medium and 50 μL of tetrazolium microculture (MTT, 5 mg/mL) were added to each well. The plates were incubated again for 4 hours at 37°C. After incubation, the medium was discarded and 500 μL of DMSO was added to dissolve the crystals. Finally, the absorbance of the wells was measured at 570 nm. These experiments were also performed in triplicate and repeated three times.

[0105] To assess these results, the percentage of viability of HeLa cells was calculated. Percentage of viability is defined as: OD<570>compound/OD<570>control × 100 (the control is considered to be 100% viability). Cells were considered non-cytotoxic when only a viability reduction of <10% was obtained, values between 10–25% were considered low cytotoxicity, and values between 25–40% were considered moderately cytotoxic.

[0106] RESULTS

[0107] The activity assays of the dendrimer of the present invention were preferably performed using water as the solvent, although the possibility of using other solvents is not ruled out. The use of water, however, eliminates the use of solvents that are inherently harmful to microorganisms. This characteristic is extremely relevant, especially in the field of fungal infections, where most antifungals used clinically are poorly soluble in water. This property of the dendrimer of the invention facilitates its easy handling and use. The concentration ranges studied preferably reach maximum concentrations of 512 mg/L for planktonic cells and forming microbial biofilms and up to 1024 mg/L for previously established microbial biofilms.

[0109] In the tests performed, the antimicrobial activity of the main compound of the invention, BDNG001, was tested against bacterial planktonic cells (S. aureus and E. coli) and yeasts (C. albicans). All strains used belong to the stock of the Spanish Type Strain Collection (CECT). Table 1 shows the minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC). The results shown in this table support the good antimicrobial activity, especially against C. albicans.

[0111] Table 1. MIC, MBC and MFC values (mg/L) of different microorganisms (one yeast and two bacteria) using the main compound of the invention, BDNG001. CECT: Spanish Collection of Type Strains.

[0113]

[0116] The main compound of the invention has been tested on forming and previously established biofilms of eukaryotic (C. albicans) and prokaryotic (S. aureus) microorganisms. The compound is readily soluble in water and no aggregates are observed in contact with the culture media used. The results obtained were very satisfactory, as can be seen in Table 2, which shows the values obtained for a forming biofilm: Minimum Biofilm Inhibitory Concentration (MBIC), Minimum Biofilm Fungicidal Concentration (MBFC), and Minimum Biofilm Bactericidal Concentration (MBBC); and values for a previously established biofilm: Minimum Biofilm Damage Concentration (MBDC) and Minimum Biofilm Eradication Concentration (MBEC). Figures 1 and 2 show the percentage of viability for each concentration tested on previously established C. albicans biofilms. S. aureus albicans.

[0118] Table 2. CMIB, CMFB, CMBB, CMDB and CMEB values (mg/L) obtained for compound BDNG001.

[0120]

[0123] The compound of the invention BDNG001 has been tested on forming and previously established mixed biofilms of C. albicans (yeast) and S. aureus (bacteria). The results obtained were extremely satisfactory, as can be seen in Table 3, which shows the values obtained for a forming mixed biofilm: CMIB and CMBB/CMFB; and values for an established biofilm: CMDB and CMEB. Figure 3 shows the percentage of viability in previously established mixed biofilms for each concentration tested with BDNG001.

[0125] Table 3. CMIB, CMFB/CMBB, CMDB and CMEB (mg/L) values obtained for compound BDNG001 in mixed biofilm treatments.

[0128]

[0131] These results obtained are highly relevant. Very few studies have been conducted on mixed biofilms. The dendrimer of the present invention has completely eradicated a previously established biofilm of two completely different species at low concentrations.

[0132] Finally, the cytotoxicity produced after 24 and 48 hours of incubation of the compound with the HeLa cell line was evaluated. This invention guarantees an absence of toxicity at the main concentrations that affect biofilms, including some of the most difficult to eliminate, mixed biofilms. These results are shown in Figures 4 and 5. The viability percentages shown in these figures demonstrate the good biocompatibility of the dendrimer of this invention.

Claims (16)

1. CLAIMS 1. Cationic carbosilane dendrimer comprising a cyclotetrasiloxane core, wherein each silicon atom of the core is linked by a carbosilane (CH<2>)<2>{Si(Me)<2>}(CH<2>)<2>S(CH<2>)<2> structure to a peripheral structure comprising an ammonium group linked by an alkyl chain (CH<2>)<2> to a hydroxyl group, according to the following formula: 2. Carbosilane dendrimer, according to claim 1, wherein the charge of said dendrimer is neutralized by at least one anion selected from a group consisting of at least one halide, hydroxide, triflate, nitrate and sulfate, as well as any combinations thereof. 3. Carbosilane dendrimer, according to claim 2, wherein the halide is selected from a group consisting of chloride, bromide and iodide. 4. Use of a carbosilane dendrimer, according to any one of claims 1 to 3, for the elimination of at least one species of pathogenic microorganism, wherein said microorganism is either in suspension or adhered to a surface, forming a biofilm 5. Use, according to claim 4, wherein said microorganism is selected from a group consisting of yeasts and bacteria. 6. Use according to claim 4 or 5, wherein the biofilm is made up of a single species of microorganisms or at least two different species of microorganisms. 7. Method for eliminating at least one species of pathogenic microorganism, characterized in that it comprises applying to the pathogenic microorganism a solution of the dendrimer defined according to any one of claims 1 to 3. 8. Method for removing at least one biofilm made up of a single species of microorganisms or at least two different species of microorganisms, either during its formation or once formed, characterized in that it comprises applying to the biofilm a solution of the dendrimer defined according to any one of claims 1 to 3. 9. Method according to claim 7 or 8, wherein the application of the dendrimer solution is carried out for a time of between 1 hour and 72 hours. 10. Method according to any one of claims 7 to 9, wherein the solution is an aqueous solution. 11. Cationic carbosilane dendrimer according to any one of claims 1 to 3 for use as a medicament. 12. Cationic carbosilane dendrimer according to any one of claims 1 to 3 for use in the prevention and/or treatment of at least one disease caused by at least one pathogenic microorganism. 13. Use of a pharmaceutical composition characterized in that it comprises a carbosilane dendrimer according to any one of claims 1 to 3 for the elimination of at least one species of pathogenic microorganism, wherein said microorganism is either in suspension or attached to a surface, forming a biofilm. 14. Use of a composition according to claim 13, wherein said composition further comprises at least a second active ingredient selected from a group consisting of an antibiotic, anti-inflammatory and antiviral, as well as any combinations thereof. 15. Use of a carbosilane dendrimer according to any one of claims 1 to 3 as a delivery vehicle for at least one drug. 16. Use, according to claim 15, wherein the drug is a drug with antibacterial and/or antifungal properties.