WO2019008210A2 - Microparticles comprising an endoscopic resection composition - Google Patents

Abstract

The present invention relates to a combined preparation comprising at least one microparticle and a composition, preferably an aqueous pharmaceutical solution, comprising a polymer derived from water soluble cellulose and hyaluronic acid, preferably comprising carboxymethylcellulose and hyaluronic acid. Likewise, it makes reference to the use of said combined preparation in the preparation of a medicament for the treatment of mucosal lesions by means of endoscopic resection, for example, the resection of polyps and / or tumors of the gastrointestinal mucosa.

Description

 MICROPARTICLES THAT INCLUDE A COMPOSITION FOR THE RESECTION

 ENDOSCOPIC

The present invention is within the field of pharmacy and medicine, and more specifically refers to microparticles comprising a composition for endoscopic resection, improving the properties of said composition and its application in the treatment of polyps and / or tumors, particularly of the mucosa of the gastrointestinal tract.

STATE OF THE PREVIOUS TECHNIQUE

The treatment of pre-malignant (or early neoplastic) digestive lesions has undergone a great change, mainly due to the development of new endoscopes and the appearance of new instruments. With the development of endoscopic submucosal resection (commonly called mucosectomy) and endoscopic mucosal dissection, early neoplastic lesions of the gastrointestinal mucosa are resectable endoscopically, practically in their entirety, through these minimally invasive techniques.

Europe, America, China, India and Japan are the areas with the highest prevalence of gastrointestinal tumors, potentially resectable (Pavithran K, et al., "Gastric Cancer in India." Gastric Cancer, 2002; 5: 240-3). Generally, the use of this technique is accepted as a treatment option for cases of early tumors where the probability of lymph node metastases is low. Because mucosectomy is a minimally invasive technique, low cost compared to conventional surgery, and it manages to cure the lesion, endoscopic resection (both mucosectomy and submucosal dissection) is taking a leading role in these countries (Ono H, et al., "Endoscopy mucosal resection for treatment of early gastric cancer" Gut. 2001; 2 (48): 225-9). In the West, the most popular technique is mucosectomy, simpler and safer, and of shorter duration, than submucosal endoscopic dissection (more developed in Asian countries such as Japan).

In order to improve the efficiency of resection, as well as the ease and safety of endoscopic submucosal resection, conventionally a lifting agent has been used, traditionally a physiological saline solution, associated with an organic dye that is commonly injected under the mucosa of the area designated for resection, resulting in an elevation of the lesion that allows to delimit and eliminate damaged tissue.

However, there are limiting factors in endoscopic resection such as the lack of an ideal submucosal lifting agent or injection solution. Occasionally, it is difficult to accurately remove the area of the lesion using conventional agents due, among other factors, to a low degree of protrusion, and especially to the fact that the lifting agent diffuses immediately after injection into the peripheral tissue, causing the disappearance of the protuberance or wheal before the end of the resection process which forces the surgeon to repeat the injection of the lifting agent. One of the few products commercially available for use as a solution for submucosal injection is Glyceol ^® (Uraoka T, et al., "Submucosal injection solution for gastrointestinal tract mucosal endoscopy resection and submucosal dissection endoscope." Drug Des Devel Ther. 2009; 2: 131-8). The use of hypertonic solutions +/- adrenaline and the same mixing sequence but with Glycerol 10% has also been described. The glycerol solution is economical and easy to prepare but its high viscosity and its short-lived effect can limit its use. Also, from a rheological point of view, it is a fluid with Newtonian characteristics. On the other hand, due to its high viscosity, it presents difficulties to inject it submucosally. It can also produce "fumes" that make it difficult to perform the technique. Hyaluronic acid (HA) can also be used, but its high cost and viscosity when injecting it can limit its use (Jung YS, Park DI I. "Submucosal injection solutions for mucosal endoscopy resection and submucosal endoscope dissection of gastrointestinal neoplasms". Gastrointest Interv. 2013; 2: 73-77). Hui P et al. recently reported that HA is more effective than saline for maintenance of mucosal elevation (Hui P, Long ZY, Jun HX, Wei W, Yong HJ, Peng LH. "Endoscopy resection with hyaluronate solution for gastrointestinal lesions: systematic review and meta-analysis "Surg Laparosc Endose Percutan Tech. 2014; 24 (3): 193-8). Also, patent application KR 20110057877 seems to describe a solution of hyaluronic acid between 0.1 and 1% that were tested for use as an injectable in endoscopic submucosal dissection procedures.

Both hyaluronic acid and hydroxypropylmethylcellulose (HPMC) are very viscous and must often be diluted to facilitate their injection. Also, HPMC has been associated with tissue damage and inflammation at the site of injection (Uraoka T, et al., "Submucosal injection solution for gastrointestinal tract mucosal endoscopy resection and submucosal dissection endoscope." Drug Des Devel Ther. 2008; 2: 131-8); "Endoscopy mucosal resection and endoscopy submucosal dissection", Gastrointestinal endoscopy 2008, 68: 1 1 1-18). Patent application WO 03/074108 describes the use of 0.83% HPMC for submucosal injection in mucosal resection or polypectomy interventions. Specifically, an in vivo test is described in which the injection of said solution is effected by means of a 23G needle in two groups of animals in which alternate resection area labeling procedures have been used, resulting in average lifting times 36 minutes and 38 minutes respectively.

Also, a solution of carboxymethylcellulose (CMC) at a concentration greater than 2% has been described as optimal for submucosal injection in endoscopic submucosal dissection (EDS) procedures. In particular, submucosal injection of a 0.5% solution at 1.5% in an in vitro model did not allow the mucosal layer of the muscle layer to be separated. On the other hand, the submucosa injection of a solution of between 2% and 3.5% of carboxymethylcellulose characterized by a viscosity of 200 mPa * S, managed to separate both layers. Based on these results, a concentration of 2.5% was selected for experimentation in an animal model. Because a CMC solution greater than 2% is very viscous, a special 18G needle It was necessary to carry out said submucosal injection (Yamasaki et al. "A novel method of submucosal endoscopy dissection with blunt abrasion by submucosal injection of sodium carboxymethylcellulose: an animal preliminary / study." Gastrointestinal Endoscopy 2006; 64 (6), 958- 965; Uraoka T, et al., "Submucosal injection solution for gastrointestinal tract mucosal endoscopy resection and submucosal dissection endoscope." Drug Des Devel Ther. 2008; 2: 131- 8).

 Hikichi et al. 2012 ("Novel Injection Technique: Submucosal Endoscope Dissection by Submucosal Injection of Sodium Carboxymethylcellulose for Early Gastric Cancer"; Gastrointestinal Endoscopy, 2012, 75 (4S), Sa1765) mention a study in gastric cancer patients at an early stage in which uses a solution for endoscopic resection of sodium carboxymethylcellulose at a concentration of 1.5%. This solution was injected into the submucosa layer with a 25G needle. The average duration of treatment was 31.4 minutes from the injection of the submucosal solution until the end of the procedure. However, the average duration of the elevation is not indicated, and a duration of more than 60 minutes is sometimes necessary.

 Combinations of CMC and hyaluronic acid have been described for other uses, for example in US 2004241155 A1 reference is made to the use of solutions of hyaluronic acid with water-soluble cellulose derivative polymers of low viscosity and a molecular weight less than 100 kDa for its Use in ophthalmic or joint operations, mentioning a combination of hyaluronic acid with 1% CMC. Likewise, EP 1992362 A2 refers to compositions comprising hyaluronic acid and a preservative (eg, benzalkonium chloride) for ophthalmic, otic or nasal use. The use of anionic cellulose derivatives of a molecular weight between 70 and 700 kDa is mentioned.

 Additionally, it is an additional advantage that the composition is stable at room temperature, not needing to maintain the cold chain.

In conclusion, there is currently a need to find a composition for use as an elevating agent in endoscopic treatments comprising a resection of a portion of the gastrointestinal mucosa. Said solution must be of low cost, easily available, and have an optimum viscosity, stable from the physical-chemical and microbiological point of view, and which additionally remains stable at room temperature. An optimum viscosity is that which on the one hand allows it to be easy to inject, allowing the use of standard injection needles (for example, 21 G, 23 G or 25 G), and on the other hand it provides a good elevation of the lesion for a time prolonged (for example, at least 45 minutes, preferably around 60 minutes or more). The duration of the endoscopic treatment will depend on the size and location of the lesion and the ideal composition will avoid the need for reinjection during the endoscopic procedure due to loss of consistency of elevation (also called bump or wheal). Also, said composition must be non-toxic, lacking side effects, such as tissue damage, bleeding and / or inflammation of the injection tissue, thus allowing a safe endoscopic intervention. Finally, the ideal composition is one that allows sterilization without losing its rheological properties.

DESCRIPTION OF THE FIGURES

Figure 1.- Viscosity curve (squares) and flow curve (triangles) of Formulation 1 - Sample M 1502 Glycerol at time 0 (replicate 1).

 Figure 2.- Viscosity curve (squares) and flow curve (triangles) of Formulation 1 - Sample M 1502 Glycerol at time 6 months (replicate 1).

Figure 3.- Viscosity curve (squares) and flow curve (triangles) of Formulation 2 - Sample M 1502 Ac. Hyaluronic at time 0 (replicate 1).

 Figure 4.- Viscosity curve (squares) and flow curve (triangles) of Formulation 2 - Sample M 1502 Ac. Hyaluronic at 6 months time (replicated 1).

 Figure 5.- Viscosity curve (squares) and flow curve (triangles) of Formulation 3 - Sample M1502 Without Adrenaline - refrigerator, at time 0 (replicate 1).

 Figure 6.- Viscosity curve (squares) and flow curve (triangles) of Formulation 3 -

Sample M1502 Without Adrenaline - refrigerator, on time 6 months (replicated 2).

 Figure 7.- Viscosity curve (squares) and flow curve (triangles) of Formulation 4 -

Sample M1502 Without adrenaline - room temperature, at time 0, (replicate 2).

Figure 8.- Viscosity curve (squares) and flow curve (triangles) of Formulation 4 -

Sample M1502 Without adrenaline - room temperature, at time 6 months (replicate 1).

 Figure 9.- Viscosity curve (squares) and flow curve (triangles) of Formulation 5 -

Sample M1502 With Adrenaline - refrigerator, at time 0 (replicated 2).

 Figure 10.- Viscosity curve (squares) and flow curve (triangles) of Formulation 5 - Sample M1502 With Adrenaline - refrigerator, at time 6 months (replicate 1).

 Figure 11.- Viscosity curve (squares) and flow curve (triangles) of Formulation 6 - Sample M1502 With Adrenaline - room temperature, at time 0 (replicate 2).

Figure 12.- Viscosity curve (squares) and flow curve (triangles) of Formulation 6 - Sample M1502 With Adrenaline - room temperature, at time 6 months (replicate 1).

Figure 13.- Transmission and backscattering profiles of Formulation 3 (without adrenaline), at time 0, refrigerator.

 Figure 14.- Transmission and backscattering profiles of Formulation 3 (without adrenaline), in time 30 days, refrigerator.

 Figure 15.- Transmission and backscattering profiles of Formulation 4 (without adrenaline), at time 30 days, room temperature.

Figure 16.- Transmission and backscattering profiles of Formulation 5 (with adrenaline), at time 0, refrigerator. Figure 17.- Transmission and backscattering profiles of Formulation 5 (with adrenaline), in time 30 days, refrigerator.

 Figure 18.- Transmission and backscattering profiles of Formulation 5 (with adrenaline), on time 90 days, refrigerator.

Figure 19.- Transmission and backscattering profiles of Formulation 6 (with adrenaline), at time 30 days, room temperature.

 Figure 20.- Transmission and backscattering profiles of Formulation 6 (with adrenaline), at 90 days, at room temperature.

 Figure 21.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 3, conserved at 8 ° C

 Figure 22.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 3, conserved at 25 ° C

 Figure 23.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 3, at 37 ° C

Figure 24.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 6, conserved at 8 ° C

 Figure 25.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 6, conserved at 25 ° C

 Figure 26.- Viscosity curve (squares) and flow curve (triangles) of the formulation combined with microparticles, MP 1: 6, at 37 ° C

 Figure 27.- Transmission and backscattering profiles of the microparticulated formulations, MPP. 0 days

 Figure 28.- Transmission and backscattering profiles of the microparticulated formulations, MPP. 30 days

Figure 29.- Optical micrographs of the microparticles before being injected

 Figure 30.- Optical micrographs of the MPP 1: 3 and MPP 1: 6 microparticles before being injected.

 Figure 31.- Optical micrographs of the microparticles after crossing the 23 G needle.

Figure 32.- Speak of solution 1 without MPP (AH 0.003% + CMC 0.2%) at time 0.

 Figure 33.- Speak of solution 1 without MPP (AH 0.003% + CMC 0.2%.) After 90 minutes.

 Figure 34.- Talk about solution 2 (0, 1% CMC + 0, 12% AH MPP) at time 0.

Figure 35.- Speak of solution 2 with MPP (0, 1% CMC + 0, 12% AH MPP) after 90 minutes. DESCRIPTION OF THE INVENTION The combination of the same particles with the composition of the invention means that said composition does not need to be kept cold, maintaining its physicochemical properties, and in particular, a suitable viscosity and stability.

 To date, the same particles have been developed as active ingredient vehicles. The use of this type of systems, provides a series of benefits both technological and biopharmaceutical in the pharmaceutical, cosmetic and food sectors, such as:

 Stabilization of unstable active principles, against different external factors Improvement of the dispersion of insoluble or poorly soluble substances in aqueous media. Conversion of liquid active substances into solids in order to improve their handling and conservation.

 - Vehiculation of several incompatible assets.

 Decreased irritant effect caused by some drugs in contact with the gastrointestinal mucosa.

 Improvement in the handling of toxic substances.

 - Masking of organoleptic characteristics.

 Modified or controlled release over time of the vehicles' assets.

The authors of the present invention propose the elaboration of a microparticulate coarse dispersed system formed by the combination of a colloidal solution and microparticles, these being free of active principles. The nature of the microparticles makes them possessing "per se" of different properties, which suppose a novel strategy for mucosectomy. Among them is an improvement in stability.

Thus, the examples of the invention demonstrate that the combined preparations of the invention comprising the microparticles present the following technical advantages over the endoscopic solution without combining with the microparticles: - Mucoadhesiveness, prolongs the contact time and the permanence of the formulation in the place of action. Additionally, the methodology used for the manufacture of the final formulation allows the obtaining of dispersed microspheres in a mucilage, which will improve, even more mucoadhesion. This property can be used if the components of the matrix or those used in the synthesis not only have forming properties but also therapeutic properties such as anti-inflammatory, bacteriostatic, fungicidal, healing, etc. For example, early inhibition of inflammation or antiseptic activity will reduce tissue adhesion or possible infection, respectively. Thus the greater contact with the mucosa will highlight all these innate therapeutic activities to the microparticle itself and not to any vehiculated assets. - Moistometric suitability: its small size and morphology makes it optimal to be injected. Its size, at the same time rude, will be very suitable to increase the height of the habón. - Lynch capacity: in addition to the volume they occupy "per se", the same processed particles are systems capable of absorbing the solution in which they are dispersed, greatly increasing their weight and hence the height of the wheal produced after injection. This characteristic will depend to a large extent on the process of maturation or processing time of the sample, structure of the microspheres and dehydration processes.

-Time of residence and degradation: The use of microparticles increases the dwell time of the wheal and therefore, the maneuvering time for the intervention. Likewise, the lower degradation will have an impact on the anti-adhesion capacity of the fabric.

- Anti-adhesion capacity of tissue: tissue adhesion (binding of damaged tissue to adjacent tissues), generally formed during surgical procedures, can lead to undesirable clinical complications, such as postoperative pain, intestinal obstruction, second intervention and incomplete organ function after Surgery. The microparticles can constitute a physical barrier that keeps damaged tissues separated during the healing process thanks to their mechanical properties and dependent stimulation. -Porosity: these are microspheres with the ability to control the release of assets if necessary. As already explained, the objective of the microspheres is not transport, although this does not exempt them from being able to be used additionally as vehicles for substances that help in the intervention as a hemostatic, anesthetic or analgesic.

It has also been observed that the combined preparation of the invention retains the same effects well in prolonged interventions (> 60 minutes, and has even been found to work well in 90-minute interventions). However, the optimal duration of wheal when using the mucosectomy solution is preferable to be approximately 30 minutes. This fact has allowed the damaged tissue to be removed with precision, avoiding damage to the area of irrigated tissue, and reducing the risk of fragmentation of the polyp and hemorrhages, with a faster recovery of the patient, even in long interventions. The higher efficacy is secondary to a greater permanence of the compound at the intestinal level compared to the standard treatment, a greater expansion of the tissue by injecting a smaller amount of product, an optimum viscosity, the total reabsorption of the product administered, as well as a physical-chemical stability and prolonged microbiological. COMBINED PREPARATION OF THE INVENTION

Therefore, a first aspect of the invention relates to a combined preparation, hereinafter "combined preparation of the invention", comprising: a) a component A, which is a microparticle, and b) a component B, which is a composition, hereinafter the composition of the invention, or composition (B), which in turn comprises:

 I) a polymer derived from water soluble cellulose in a concentration of 0.005% to 2%; Y

 II) hyaluronic acid in a concentration of 0.0001% to 0.5%.

 The present invention relates to a composition wherein the components A and B are juxtaposed, in the form of a combined preparation.

It should be emphasized that the term "combined preparation" or also referred to as "juxtaposition", in this specification, means that the components of the combined preparation do not need to be present as a bond, for example in a true composition, in order to be available for their combined application , separate or sequential. In this way, the expression "juxtaposed" implies that it is not necessarily a true combination, in view of the physical separation of the components.

Unexpectedly, during the synthesis process, the microparticles of the invention incorporated the mucosectomy solution in its external structure, remaining embedded in the external surface of the microparticle. Therefore, we can affirm that they are not a vehicle of drugs but that they form, together with the mucosectomy solution, a treatment in themselves.

In a preferred embodiment of this aspect of the invention the microparticle (A) is dispersed in the composition (B), the microparticle (A) absorbs the composition (B), which is embedded in the microparticle (A), or a combination from both.

In another preferred embodiment the microparticle is made by an internal ionic gelation emulsification technique.

In another preferred embodiment the microparticle is obtained by a process comprising the preparation of an A / O emulsion in which the internal or aqueous phase is formed by a polymer or mixture of polymers, and the external or oil phase is composed of a oil in which a surfactant is dispersed.

When we refer to polymer mixture this can refer to a physical or chemical substance. In addition to water, different solvents can be used in the method of preparation of microparticles that influence the physicochemical properties of the intervening molecules, such as solutions of vasoconstrictor extracts such as fluid extract or tincture (2-4ml_) of ruscus, bilberry , horse chestnut or meliloto, or the colloidal solution of hyaluronic acid and cellulose in which you are dispersed. The polymers were:

 Of natural origin:

• Chitosan • Exopolysaccharides of bacterial origin

 • Cellulose derivatives: Hydroxypropylmethylcellulose, sodium methylcellulose, cellulose acetophthalate, sodium carboxymethylcellulose.

 • Pectin

 In synthesis:

 • Poloxamer 127

 • Eudragit® S 100, L-100, L 12.5, S 12.5, FS30D, RS, FS and NM

 • Shellac

 • PLGA

Other polymers could also be used. Some are "smart" polymers because of their rapid response to changes in environmental physicochemical conditions, which involve pronounced changes in their properties. The stimuli to which the polymers respond can be:

• Physical: such as temperature, ionic strength, solvents, radiation, electric fields, mechanical stress, pressure, sonic radiation and magnetic fields;

 • Chemicals: such as pH, specific ions and chemical agents;

 • Biochemical: as substrates of enzymes, related ligands and other biological agents. In another preferred embodiment the polymer of the microparticle is selected from the list consisting of: alginate, chitosan, exopolysaccharide of bacterial origin, cellulose or cellulose derivatives, pectin, poloxamer 127 or any other type of poloxamer, Eudragit® S 100, L -100, L 12.5, S 12.5, FS30D, RS, FS and NM, Shellac, PLGA, or any of their combinations.

In another preferred embodiment, where the polymer of the microparticle is alginate, more preferably sodium alginate. In another preferred embodiment, the surfactant of the oil phase of the microparticle is polysorbate 80. In another preferred embodiment, the aqueous phase of the microparticle further comprises fluid extract or tincture of ruscus, bilberry, horse chestnut or melilot, or the colloidal solution of hyaluronic and cellulose. Preferably the fluid extract or the dye is in a volume of between 2 ml and 4 ml.

In another preferred embodiment, the oil of the oil phase of the microparticle is selected from olive oil, rosemary oil, argan oil, liquid paraffin, or any of its combinations, contemplating the use of other medicinal oils.

In another preferred embodiment once the aqueous phase of the A / O emulsion is formed from which the microparticle is formed, a calcium source is added, in the form of an insoluble salt, which is dispersed in the aqueous phase. More preferably the source of calcium is calcium carbonate, and even more preferably the calcium carbonate is in a proportion between 0.5-1%.

In another preferred embodiment the oil phase of the A / O emulsion from which the microparticle is formed is composed of a homogeneous solution of polysorbate 80 in oil, preferably in a proportion of between 2-4%.

In another preferred embodiment for forming the oily external phase emulsion of the A / O emulsion from which the microparticle is formed, the two phases are subjected to mechanical stirring, preferably at a rate of between 300 and 2500 rpm, more preferably between 500-2000 rpm, for a time comprised between 5-40 minutes, more preferably for 10-30 minutes.

In another preferred embodiment the emulsion obtained in the previous step is added an acid medium, preferably acetic acid, and more preferably in a proportion of between 4 and 8%, to solubilize the calcium ions and result in the formation of the pre-gelled particles.

In another preferred embodiment the formation of the microparticle further comprises adding a solution of calcium chloride, preferably 5-10% (w / v), to the emulsion resulting from the preceding claim to produce the hardening of the polymer droplets under mechanical agitation , preferably at a speed of between 300 and 2500 rpm, more preferably at a speed of between 500 and 2000 rpm, for a time comprised between 5 and 40 minutes, more preferably for 10-30 minutes In another preferred embodiment the mirroparticles obtained in the previous claim are cleaned with bidistilled water using successive centrifugation cycles

In another preferred embodiment the same particles are matured in calcium solution for a time of between 1 to 15 minutes, preferably between 2 to 10 minutes and subsequently dried at 20-40 ° C, preferably at 25-37 ° C. In another preferred embodiment the combined preparation of the invention further comprises another active ingredient, preferably vitamin K1 or phytomenadione.

In another preferred embodiment the viscosity of said composition (B), hereinafter the composition of the invention, is from 5 to 100 mPa * s, preferably from 5 to 50 mPa * s, more preferably from 10 to 40 mPa * s Examples of viscosities within the most preferred range are from 15 to 30 mPa * s and from 20 to 40 mPa * s. The term "hyaluronic acid" as used in the invention refers to a polysaccharide that includes at least one constituent unit consisting of a glucuronic acid and N-acetylglucosamine. It also includes pharmaceutically acceptable salts thereof, which are not particularly limited, and include, for example, a sodium salt, a potassium salt, a calcium salt, a zinc salt, a magnesium salt, an ammonium salt, ammonium alkyl salt, and the like. The term "hyaluronic acid" also includes derivatives thereof such as those described in EP2537867 A1. Preferably, said hyaluronic acid is the sodium salt of hyaluronic acid.

In the present invention when mention is made of the molecular weight of the polymers used in the invention, one skilled in the art will understand that reference is made to the average molecular weight. Methods for the determination of the average molecular weight are well known to a person skilled in the art and include, for example, tests using the osmotic method (membrane or vapor pressure osmometry), terminal group analysis, light scattering or light scattering laser , sedimentation equilibrium by analytical ultracentrifugation, viscometry, fractionation techniques of the polymer sample with on-line detection: liquid exclusion chromatography (SEC / GPC) and desorption laser mass spectrometry / matrix-assisted ionization: mass spectroscopy (MALDI) , and combinations thereof.

Hyaluronic acid can be high or low molecular weight. Typically high molecular weight hyaluronic acid is considered to be those chains of said polysaccharide of more than 1,000 kDa, preferably more than 1,500 kDa or 1,800 kDa. While low molecular weight hyaluronic acid is usually considered to be one that has a molecular weight of less than 1,000 kDa, preferably less than 800 kDa, more preferably less than 600 kDa, even more preferably less than 300 kDa or less than 250 kDa. In a particular embodiment, the hyaluronic acid of the composition of the present invention is of low molecular weight, preferably has an average molecular weight of 500 to 800 kDa.

Likewise, hyaluronic acid can also be defined by its viscosity. Preferably, the hyaluronic acid of the composition (preferably pharmaceutical composition) of the present invention is of low viscosity for example, said hyaluronic acid has a viscosity in 1% aqueous solution of 100 to 300 mPa * s, preferably 150 to 250 mPa * s. Hyaluronic acid of medium or high viscosity could also be used.

The viscosity of a pharmaceutical solution is typically measured by the use of a viscometer. Today there are different types of viscometer in many applications, which meet different measurement purposes. A person skilled in the art will know how to choose the most suitable viscometer, for example a rotational viscometer (also referred to as a spindle), such as Brookfield LV (for example using spindle 3 at 30 rpm) or the Haake Viscometer VT500. A person skilled in the art will know how to adjust the spindle (also called cylinder or plate) and the rpm depending on the pharmaceutical solution to be analyzed. It should be considered that the final viscosity of the gel will depend on the characteristics of the starting raw material, its concentration and temperature.

In a preferred embodiment, the hyaluronic acid of the composition of the present invention is hyaluronic acid of low molecular weight, preferably of very low molecular weight, and of low viscosity. Preferably, it is sodium hyaluronate (e.g., Uromac®, Nakafarma, ES; Morales et al., The Journal of Urology 1996, 156, 45-48).

In a particular embodiment, optionally in combination with one or more of the characteristics of the other embodiments of the invention, the hyaluronic acid concentration is from 0.0001% to 0.3%, which includes from 0.001% to 0.1% and from 0.0001% to 0.09%, preferably from 0.001% -0.012%, more preferably from 0.002% to 0.008%, and even more preferably from 0.003% to 0.006%.

Water soluble cellulose derivative compounds are well known to a person skilled in the art. These include but are not limited to methylcellulose, ethylcellulose, methyl ethylcellulose, ethylhydroxyethylcellulose, methylhydroxyethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, carboxymethylcellulose, carboxymethylhydroxyethyl cellulose, alkyl cellulose and combinations thereof.

In a particular embodiment of the composition of the invention, optionally in combination with one or more of the characteristics of the other embodiments of the invention, the concentration of the polymer soluble in water-soluble cellulose is 0.01% to 1%, more preferably from 0.1% to 0.5%. In a more preferred embodiment, the concentration of water soluble cellulose-derived polymer is from 0.1% to 0.4%, more preferably from about 0.2% to about 0.3%.

In another preferred embodiment, the present invention relates, optionally in combination with one or more of the features of the other embodiments of the invention, to a (preferably aqueous) composition comprising: a polymer derived from water-soluble cellulose in a concentration from 0.005% to 0.4%; and hyaluronic acid in a concentration of 0.0001% to 0.09%.

Preferably, the viscosity of said composition is from 5 to 100 mPa * s, more preferably from 5 to 50 mPa * s, even more preferably from 10 to 40 mPa * s.

Said polymer soluble in water-soluble cellulose (for example: carboxymethylcellulose) typically has a molecular weight of more than 500 kDa, preferably of more than 800 kDa, more preferably of about 900 kDa. Carboxymethylcellulose or CMC is an organic compound, derived from cellulose, composed of carboxymethyl groups bonded to some hydroxyl groups, present in polymers of glucopyranose, and of general formula RnOCH 2 -COOH. The term as used herein also includes pharmaceutically acceptable salts thereof, such as sodium or potassium salts. It is often used as sodium carboxymethyl cellulose, also called carmellose sodium. Preferably, said carboxylmethyl cellulose is sodium carboxymethyl cellulose.

In a particular embodiment of the composition of the invention, optionally in combination with one or more of the characteristics of the other embodiments of the invention, the concentration of carboxymethylcellulose is from 0.005% to 2% or from 0.005% to 1.9% or from 0.005% to 1, 8% or from 0.005% to 1.7% or from 0.005% to 1.6% or from 0.005% to 1.5%. Preferably, the concentration of carboxymethylcellulose is from 0.025% to 1.5%, more preferably from 0.01% to 1%, even more preferably from 0.1% to 0.5%.

In another particular embodiment of the pharmaceutical composition of the invention, the concentration of carboxymethylcellulose is lower than 2%, 1.9%, 1.8%, 1.7%, 1.6% or 1.5%, preferably lower than 1%, more preferably less than 0.5%.

In a particular preferred embodiment, the concentration of carboxymethylcellulose is from about 0.3% to about 0.2%.

The carboxymethylcellulose may be of high, medium or low viscosity. Thus, for example, the following table defines a CMC solution based on its high, medium or low viscosity, providing viscosity ranges for each of said classifications where said values have been measured in a 2% aqueous solution (for example, example in water or physiological saline) and at 20 ° C. These conditions were those used in the preparation of the example solution.

La viscosidad puede ser expresada en centiPoises(cP). 1 Poise= dynaxseg/cm2= 1 g/cmxseg. En ocasiones la viscosidad es expresada en miliPascales-segundo (mPa.s), medida de Presión en SI. 1 Pascal= 1 Newton x m²; Factor de conversión: (1 cP = 10^~3 Pa s = 1 mPa-s).

Viscosity can be expressed in centiPoises (cP). 1 Poise = dynaxseg / cm2 = 1 g / cmxsec. Sometimes the viscosity is expressed in milliPascals-seconds (mPa.s), Pressure measurement in SI. 1 Pascal = 1 Newton xm ² ; Conversion factor: (1 cP = 10 ^{~ 3} Pa s = 1 mPa-s).

In a particular embodiment, optionally in combination with one or more of the features of other embodiments of the invention, water-soluble cellulose derivative polymer (for example: carboxymethylcellulose) has a viscosity of 500 to 4,500 mPa * s, preferably 1,000 to 3,000 mPa * s, more preferably from 1,500 to 2,500 mPa * s, even more preferably from 2,200 mPa * s to 2,300 mPa * s (for example: 2,237 mPa * s), when the ranges of viscosity of said polymer soluble in water-soluble cellulose (eg, CMC) are measured in a 1% aqueous solution (for example in water or physiological saline) at 25 ° C, for example with a Brookfield LV viscometer using screw 3 to 30 rpm.

 The rheological properties of the CMC mixture with sucrose have been described above (Cancela, M.A et al., 2005 "Effects of temperature and concentration on carboxymethylcellulose with sucrose rheology" Journal of Food Engineering, Vol. 71, pp 419-424). and it has been proven that both with sucrose and without it, it behaves like Pseudo - plastics. The variation of the deformation speed against the tangential stress is exponential; and by increasing the concentrations of CMC and sucrose (glucose + fructose), the viscosity increases. The opposite happens when the temperature increases.

 In a particular preferred embodiment, optionally in combination with one or more of the characteristics of the other embodiments of the invention, said carboxymethylcellulose is sodium carboxymethylcellulose, has a viscosity greater than 1,000 mPa * s in a 1% aqueous solution and is found in said composition at a concentration of 0.2% to 0.3%. The composition (B) (preferably pharmaceutical solution) of the invention may further comprise other polysaccharides, preferably having pseudoplastic rheological properties. Examples of polysaccharides are hydroxypropylmethylcellulose, xanthan gum, carrageenan, gellan gum, guar gum, locust bean gum, and sacchar. In addition, combinations thereof can be used. In a particular embodiment, the composition (preferably pharmaceutical solution) of the invention comprises CMC as the sole polysaccharide.

 Also, the composition (preferably pharmaceutical solution) of the invention may contain other active ingredients and / or pharmaceutical excipients such as an osmotic pressure regulating agent, a pH regulating agent, a preservative, a coloring agent, and one or more active ingredients (for example: a vasoconstrictor or hemostatic agent).

Said composition is preferably an aqueous composition. Typically said aqueous composition comprises water or an aqueous solution. Non-limiting examples of aqueous solutions include:

 • Water,

 • Normal saline solution (saline solution): contains 0.9% sodium chloride or 154 mmol / L.

 • Hypertonic saline solution: contains 3% to 5% sodium chloride or 513-855 mmol / L.

 • Hypotonic saline solution: contains 0.45% sodium chloride or 77 mmol / L.

 • Ringer's solution with lactate: e.g.,: 102 mmol / L of sodium chloride; 28 mmol / L sodium lactate; 4 mmol / L of potassium chloride, and 1.5 mmol / L of calcium chloride.

 • 5% dextrose solution: they provide a concentration of 278 mmol / L of glucose.

• Plasmalyte type solution: mixture similar to the lactated Ringer, with the presence of magnesium, acetate and gluconate ions. • Hypertonic glucosed serum: contains 10%, 20%, 40% glucose, with concentrations of 278x2, 278x4, 278x8 mmol / L.

 • Glucosaline serum: typically contains 0.45% sodium chloride and 5% glucose.

• Albumin solution: contains between 5-25% sterile human albumin in water for injection.

 In a preferred embodiment, said aqueous solution comprises physiological saline.

In a particular embodiment, optionally in combination with one or more of the features of the other embodiments of the invention, the composition (preferably pharmaceutical solution) of the invention further comprises an osmotic pressure regulating agent and / or the pH in a 1 -30%, preferably selected from a sugar, a polyalcohol and a salt and combinations thereof. In a particular embodiment of the composition (preferably pharmaceutical solution) of the invention, said regulatory agent is in a concentration of 10% -20%, preferably 16-18%, more preferably 17.5%. Examples of polyalcohols include mannitol, xylitol, erythritol, treitol, ribitol, myoinisitol, galactitol, sorbitol, glycerol, derivatives and combinations thereof. Sorbitol, glycerol and combinations thereof being especially preferred.

 In a particular embodiment, said osmotic pressure regulating agent and / or pH is a sugar, preferably a monosaccharide and / or a disaccharide. The term disaccharide can include any disaccharide. Examples of disaccharides include lactose, trehalose, sucrose, maltose, isomaltose, cellobiose, isosacarose, isotrehalosa, sorbose, turanosa, melibiose, gentiobiose, and mixtures thereof. Preferably, lactose, trehalose, sucrose, and combinations thereof. The term monosaccharide can include any monosaccharide, such as, for example, mannose, glucose (dextrose), fructose (levulose), galactose, xylose, ribose or any combination thereof. In a particular embodiment, optionally in combination with one or more of the features of the other embodiments of the invention, said sugar is a monosaccharide. In a preferred embodiment, said sugar is fructose. Said sugar can also be a polysaccharide, for example inulin which is constituted by fructose units. Said sugar (eg, fructose) can also act as a preservative and adjuvant agent of carboxymethylcellulose in relation to its pseudoplastic effect.

Said salt is preferably sodium chloride, although other salts such as potassium chloride, sodium citrate, magnesium sulfate, calcium chloride, sodium hypochlorite, sodium nitrate, mercury sulphide, sodium chromate and magnesium dioxide, as well as Phosphate and calcium salts can also be used. In a particular embodiment, optionally in combination with one or more of the characteristics of the other embodiments of the invention, the composition (preferably pharmaceutical solution) of the invention comprises a saline solution, preferably physiological saline (0.9% NaCl). Preferably, said composition (preferably pharmaceutical solution) has an osmolarity of 500-3,000 mOs / L, preferably 1,500-2,500 mOs / L, more preferably 1,700 mOs / L.

 Examples of pH regulating agents include Tris-HCl buffer, acetate buffer, citrate buffer and phosphate buffer or combinations thereof. The term "acetate buffer", "citrate buffer" and "phosphate buffer" as used herein may refer to a buffer system comprising an organic acid and (acetic acid, citric and phosphoric acid, respectively) and a salt of the same. Each one of them can be added in a sufficient amount. The pH of the composition according to the present invention is in the range of 3 to 8, preferably in the range of 4 to 7, more preferably between 5 and 6.

 In a particular embodiment, we verify that the osmolarity and viscosity of the solution combining hyaluronic acid and carboxymethylcellulose is much higher than that of the solutions of each of the products separately.

 The composition (preferably pharmaceutical solution) of the invention may further comprise a dye, preferably water-soluble, such as indigo carmine, methylene blue, tartrazine, erythrosine and quinoline yellow, more preferably indigo carmine or methylene blue. Typically, a diluted dye is usually used. The dye stains the mucosa and facilitates the evaluation of the depth of the lesion, and delimit the edges exactly (Larghi A, Waxman I. "State of the art on mucosal endoscopy resection and submucosal dissection endoscope", Gastrointest Endo Clin North Am. 2007; 17: 441-69).

 The composition (preferably pharmaceutical solution) of the invention may further comprise one or more active ingredients. Said active principle can be, for example, a vasoconstrictor agent. Epinephrine (also called adrenaline) is usually used. Alternative vasoconstrictive agents that can be used alone or in combination with epinephrine include but are not limited to norepinephrine, caffeine, theophylline, and phenylephrine. Each of them can be added in an appropriate amount to control the bleeding during the resection of the lesion. In a preferred embodiment, said vasoconstrictor agent is epinephrine in a concentration of 0.000025-0.5%, preferably 0.00025-0.05%, more preferably 0.001-0.01%, even more preferably 0.005%.

Other active ingredients that may be used in the composition of the invention include but are not limited to inulin (anti-inflammatory / antibacterial) for example in a concentration of 0.1 to 50%; Citric acid (antioxidant / coagulant / correct pH) for example in a concentration of 0.1 to 20%; zinc (healing / antioxidant) for example in a concentration of 0.1 to 20%; amino acids such as glutamine, alanine and / or arginine (immunomodulatory amino acids and which promote the healing process) for example in a concentration of 0.1 to 20%.

In a particular embodiment, the composition (preferably pharmaceutical solution) of the present invention is essentially free of one or more preservatives, such as benzyl alcohol, phenol, m-cresol, chlorobutanol and benzalkonium chloride. In another embodiment, a Preservative can be included in the formulation, particularly when the formulation is a multi-dose formulation. The preservative concentration may be in the range of from about 0.1% to about 2%, more preferably from about 0.5% to about 1%.

In a particular preferred embodiment, the composition (preferably pharmaceutical solution) of the invention, optionally in combination with one or more of the characteristics of the other embodiments of the invention, comprises or consists of:

 - Hyaluronic acid from 0.0001% to 5% (preferably, from 0.0001% to 0.09%)

 - Sodium carboxymethylcellulose from 0.005% to 2% (preferably, from 0.005% to 0.4%)

- Fructose or inulin c.s.p. pH: 5-6

 - Physiological serum c.s.p,

 - optionally, epinephrine 0.0001-0.01%

 optionally, a dye (0.01-0.1 ml, preferably 0.05 ml, 0.1 to 5% solution), wherein preferably said hyaluronic acid is of low molecular weight and low viscosity and / or the carboxymethylcellulose is of high viscosity.

Preferably, the composition (preferably pharmaceutical solution) of the invention, optionally in combination with one or more of the characteristics of the other embodiments of the invention, comprises or consists of:

 - Hyaluronic acid 0.003%

 - Sodium carboxymethylcellulose 0.2% to 0.3%

 - Fructose or inulin c.s.p. pH: 5-6

 - Physiological serum c.s.p.

 - optionally, 0.005% epinephrine

 optionally, a dye (0.01-0.1 ml, preferably 0.05 ml, 0.1 to 5% solution).

In other particular embodiments the composition of the invention further comprises citric acid in a concentration of 0.5% to 3% (eg, 1%), Zinc in a concentration of 0.5% to 3% (eg, 2%), Glutamine and Alanine (eg, Dipeptiven ^® ) in a concentration of 2% to 10% (eg, 5%) and polyethylene glycol (eg, PEG400) in a concentration of 0.5% to 3% (eg, 1%).

In addition to those described in the examples, the following formulations were tested in patients obtaining similar results:

TO)

 Hyaluronic acid 0.003%

Sodium carboxymethylcellulose 0.2% to 0.3% Fructose csp pH: 5-6

 Physiological serum c.s.p.

 Citric acid 1%

 Zinc 2%

 Glutamine and Alanine (Dipeptiven®) 5%

 PEG 400 1%,

 Indigo carmine 1% - 0.05ml_

 (pH: 5-6).

 B)

 Hyaluronic acid 0.003%

 Sodium carboxymethylcellulose 0.2% to 0.3%

 Inulin 10%

 Physiological serum c.s.p.

 Citric acid 1%

 Zinc 2%

 Glutamine and Alanine (Dipeptiven®) 5%

 PEG 400 1%,

 Indigo carmine 1% - 0.05ml_

 (pH: 5-6).

The composition (preferably pharmaceutical solution) of the invention is preferably a sterile composition (preferably solution). Nowadays it is admitted that a product can be considered as sterile when the probability of survival of any microorganism is less than 10 ^".6 There are several methods well known to a person skilled in the art for the sterilization of pharmaceutical compositions, which can be classified Generically in physical and chemical sterilization methods, physical agents include heat sterilization techniques, which can be dry or humid, ultraviolet or ionizing radiation and sterilizing filtration systems. typically to the use of liquid or gaseous antiseptics (eg, ozone) In the present invention the sterilizing agent is preferably a physical agent.

In a particular embodiment, optionally in combination with one or more of the characteristics of the other embodiments of the invention, said composition (preferably solution) is obtained by a method comprising a sterilizing filtration step with a filter between 0.2 μη- 1 μηι, preferably 0.45 μηι. For example, a 0.45 μηι antibacterial air filter can be used. The material of the membrane can be an acrylic copolymer on a nylon support. In a particular embodiment, optionally in combination with one or more of the characteristics of the other embodiments of the invention, said composition / solution is obtained by a method comprising a step of wet heat sterilization. Existing methodologies can be classified according to the temperature used:> 100 ° C (eg steam pressure (autoclave), around 100 ° C (flowing steam) or less than 100 ° C (tindalization) In a preferred embodiment, said wet heat sterilization method uses a temperature equal to or lower than 100 ° C, preferably the sterilization step is by tindalization.The failure to subject the composition of the invention to temperatures higher than 100 ° C, allows to avoid a loss of viscosity of the solution, since heat at high temperatures typically lowers the viscosity of a solution with pseudoplastic characteristics.

PROCEDURE FOR OBTAINING A COMPOSITION (B) OF THE INVENTION In another aspect, the present invention relates to a process for obtaining a composition (B), composition of component B, comprising the following steps: a) mixing a water-soluble cellulose-derived polymer gel of a viscosity of 3,000 to 5,000 mPa * s in 2% aqueous solution with an aqueous solvent;

 b) mixing the solution obtained in step a) with hyaluronic acid; Y

 c) optionally, during step a) or after obtaining said composition in a) a dye and / or one or more active ingredients or excipients are incorporated.

 Preferably, said mixing process is carried out until a composition with a viscosity of 5 to 100 mPa * s, more preferably 5 to 50 mPa * s, even more preferably 10 to 40 mPa * s, is achieved.

 In a preferred embodiment, the concentration of said polymer soluble in water-soluble cellulose in the composition according to the second aspect of the invention is from 0.005% to 2%; and the concentration of hyaluronic acid is from 0.0001% to 0.5%. Preferably, in said composition the concentration of said polymer soluble in water-soluble cellulose is from 0.005% to 0.4%; and the concentration of hyaluronic acid is from 0.0001% to 0.09%.

 Said mixing procedure in a) and b) is generally carried out under constant stirring of 200 rpm at 500 rpm (preferably about 300 rpm) and at constant heat at a temperature of 45 ° C to 55 ° C (preferably about 50 ° C) .

It is also contemplated that in the aqueous solvent used in step a) other excipients and / or active principles have been previously dissolved as described in other aspects of the invention, preferably an osmotic pressure regulating agent has been dissolved and / or the pH, more preferably fructose or inulin.

Said gel of a polymer soluble in water-soluble cellulose of a viscosity of 3,000 to 5,000 mPa * s in 2% solution can be prepared for example by a process comprising: - dissolving said polymer at 2% in an aqueous solvent maintaining constant agitation of 200 rpm at 500 rpm and constant heat at a temperature of 45 ° C to 55 ° C until reaching a desired viscosity between 3000 to 5000 mPa * s.

 Preferably, said stirring process is carried out between 3 and 6 hours, preferably around 5 hours.

 Said polymer soluble in water-soluble cellulose is preferably carboxymethylcellulose. Also, said aqueous solvent is preferably physiological saline. Other characteristics of the composition as well as particular and preferred embodiments thereof have been described hereinabove.

Said process for obtaining a pharmaceutical composition of the invention may further comprise a step of sterilization. Possible methods of sterilization have been described above.

 In a particular embodiment, said step of sterilization is effected by filtration with a filter between 0.2 μπι and 1 μηι, preferably of 0.45 μηι. In another particular embodiment, said sterilization step of sterilization is carried out by heat sterilization, preferably by tindalization or autoclaving.

 Said method may comprise an additional step of aseptic filling of the container (for example: a syringe) containing the composition of the invention.

 In another aspect the present invention relates to a composition obtained or obtainable by a process as described in the second aspect of the invention. Preferably, said composition is a composition (B) of the invention, obtained by a process as just described.

 USES OF COMBINED PREPARATION OF THE PRESENT INVENTION

 The combined preparation of the present invention is preferably a combined pharmaceutical preparation. Said pharmaceutical combination preparation is formulated to be compatible with the selected administration route. Methods for carrying out said administration are known to a person skilled in the art. These include, for example, parenteral injections (preferably excluding the intravascular route) such as subcutaneously, intraarticularly, mucosally, submucosally. The oral, nasal, ophthalmic, rectal or topical route is also contemplated. As well as formulations of controlled, delayed or sustained release. In a particular embodiment, said formulation is for submucosal injection.

 In still another aspect, the invention relates to the combined preparation of the invention for use as a medicament.

In a related aspect, the invention relates to the use of the combined preparation (preferably pharmaceutical combination preparation) of the invention as a vehicle for administration of compounds useful in diagnostic methods, of surgical and / or therapeutic treatment. Said compounds include active substances with pharmacological activity as well as radioisotopes or other compounds commonly used for diagnostic purposes.

 In a related aspect, it relates to the combined preparation of the invention for use, preferably as a lubricant, in the treatment of syndromes or diseases affecting the joints.

 Also, the invention relates to the combined preparation of the invention, for use in a method of treatment that requires tissue separation, wherein said combined preparation is preferably administered by injection, more preferably by endoscopic injection, into one of the tissues to be separated or in a tissue located between them.

In a preferred embodiment, the invention relates to the combined preparation for use in a method of treatment that requires the separation of different tissue layers, wherein said composition is administered by injection, preferably by endoscopic injection, into one of the tissues of the tissue. separate or in a tissue located between both.

Said combined preparation, preferred characteristics and particular embodiments thereof, as well as the method for obtaining same have been described in previous aspects of the invention.

 In a further aspect, the invention is directed to the use of a combined (preferably pharmaceutical) preparation in the manufacture of a medicament for the treatment of mucosal lesions comprising the resection of a portion of the mucosa, preferably of the mucosa of the tract gastrointestinal. However, it is not limited to the gastrointestinal tract, and can be performed at any location, skin and soft tissues, and mucous membranes of the body, such as, for example, the mucosa of the urogenital tract. Therefore, the resection of a portion of the mucosa is preferably performed from the mucosa of the gastrointestinal tract or the urogenital tract (bladder, vagina, ...), more preferably from the urogenital tract.

 In a related aspect, the invention relates to the combined (preferably pharmaceutical) preparation for use in a method for the treatment of mucosal lesions where said method comprises resection of a portion of the mucosa. Also, in another aspect it relates to a method for the treatment of mucosal lesions in a patient, wherein said method comprises the injection, typically endoscopic, of a therapeutically effective amount of said combined preparation for the resection of a portion of the mucosa. .

Also, the invention also relates to the combined preparation of the invention for use in the treatment of lesions in the mucosa of the gastrointestinal tract. In a related aspect it relates to the use of the combined preparation of the invention in the manufacture of a medicament for the treatment of lesions in the mucosa of the gastrointestinal tract. Finally, it also refers to a method for the treatment of lesions in the mucosa of the gastrointestinal tract, wherein said method comprises the injection, typically endoscopic, of a therapeutically effective amount of said combined preparation.

 The invention also relates to the use of the combined (preferably pharmaceutical) preparation of the invention for submucosal injection or as a lifting agent in a method of treatment (eg, surgical treatment method) of endoscopic resection.

 The combined preparation according to the present invention is especially suitable for use in mucosal resection. Said resection typically comprises endoscopic resection of damaged lesions or tissue in the mucosal layer, such as neoplastic lesions (e.g., tumors at an early stage) or pre-neoplastic lesions (e.g., polyps).

Several techniques for mucosal resection have been described. Specific examples of endoscopic mucosal resection techniques include mucosal endoscopic resection (MRS) or mucosectomy, endoscopic submucosal dissection (ESD), laparoscopic mucosal resection, uteroscopic mucosal resection, transurethral bladder tumor resection, and laser mucosectomy. The composition according to the present invention can be used for any of these mucosal resection techniques.

 In a particular embodiment, said endoscopic resection is selected from the group consisting of a mucosectomy and a submucosal endoscopic dissection. Although it is not standardized, mucosectomy is usually performed when the lesion is <20-30 mm, whereas endoscopic submucosal dissection is generally performed for larger lesions (up to 6-7 cm) (B.-H. Min, et. al., "Clinical outcomes of endoscopic submucosal dissection (ESD) for treating early gastric cancer: comparison with mucosal endoscopy resection after circumferential precutting (EMR-P)", Digestive and Liver Disease, 2009; 3 (41): 201-9) . Preferably, said endoscopic resection is an endoscopic mucosal resection (EMR) or mucosectomy.

In general, mucosectomy comprises delimiting the area of the mucosa to be resected (limits of the lesion), injecting a combined preparation, preferably sterile, into the submucosa, typically by using a syringe to which an injection needle is attached. . Said combined preparation elevates the area to be excised and separates it from the rest of the layers of the gastrointestinal tract so as not to cause more damage than is strictly necessary. The volume to be infiltrated varies depending on the size of the lesion. In this intervention, a critical step is the identification of the edges of the lesion before it is removed. Usually, mucosectomy can completely resect> 90% of mucous lesions (in one or several sessions). While endoscopic submucosal dissection consists of a block resection of a large tumor surface, usually colon-rectal. For this a combined preparation is injected, which elevates the lesion, at the level of the sub-mucosa. The mucosa adjacent to the lesion is incised with an adequate margin before the incision of the submucosal layer. For this, a total or partial incision of the circumferential mucosa is initially performed according to the established protocol and characteristics of the lesion (B.-H. Min, et al., "Clinical outcomes of endoscopic submucosal dissection (ESD) for treating early gastric cancer: comparison with mucosal endoscope resection after circumferential precutting (EMR- P) ", Digestive and Liver Disease, 2009; 3 (41): 201-9; Yamamoto K, et al.," Colorectal submucosal endoscope dissection: Recent technical advances for safe and successful procedures. "World J Gastrointest Endose 2015 October 10; 7 (14): 1 114-1 128).

 The combined preparation according to the present invention is preferably applied by injection into the mucosa or surrounding tissue, such as the submucosa, mucosa, or epithelium. Among them, administration by submucosal injection is preferred.

Examples of the organism where the combined preparation can be applied in accordance with the present invention include the digestive mucosa in organs such as the esophagus, stomach, duodenum, bile duct, small intestine, large intestine, colon, rectum and also the mucosa of respiratory organs (eg, the lungs), or the mucosa of genitourinary organs (eg, the urinary bladder, the urethra, the vagina, and the uterus.) Among them, the mucosa of the upper digestive tract (from the esophagus to the stomach or the duodenum) and the mucosa of the lower digestive tract (the small intestine, jejunum, ileum inferior to the duodenum), and large intestine (colon, rectum) is typically preferred, said treatment comprising the injection of the combined (preferably pharmaceutical) preparation of the invention in the submucosal layer.

In a preferred embodiment, the injection site is the submucosa of an organ of the digestive tract, also referred to herein as the gastrointestinal tract. The combined (preferably pharmaceutical) preparation of the invention can be used in combination with a direct inhibitor of smooth muscle peristalsis and locally sprayed into the digestive tract through a sprayer or forceps during an endoscope, eg, digestive tract surgery by laparotomy, endoscopic surgery, endoscopic examination of the digestive tract or other medical practice in which peristalsis of the digestive tract needs to be suppressed.

 The combined preparation of the invention can be used with different devices for endoscopic resection. Normally, polypectomy loops of oval, multifilament morphology of varying sizes (between 30 and 10 mm) are used. Tables 1 and 2 of the article "Endoscopy mucosal endoscopy resection and endoscopy dissection", Gastrointestinal endoscopy 2008, 68: 1 11-18 details mucosectomy devices (EMR) and endoscopic submucosal dissection, respectively, available on the market.

The injection of the combined (preferably pharmaceutical) preparation of the invention is generally effected by an endoscopic injection needle. In a preferred embodiment, optionally in combination with one or more of the features described in other aspects of the invention, the combined preparation of the invention is administered by endoscopic injection into the submucosal layer of an organ, of the gastrointestinal tract.

The diameter of the endoscopic injection needle (G) is standardized in terms of the outer diameter of the needle, and a larger gauge number means an outer diameter of the smaller needle. The size number of an endoscopic injection needle for use with the pharmaceutical solution of the invention is selected depending on the site of surgery, but is generally 21 to 25 G, preferably 23 G. Even when they have the same caliber number, endoscopic injection needles produced by different manufacturers may have different internal diameter. In general it is preferred to use a needle with the smallest possible diameter, it being also possible to use needles with a size less than 25G such as 25sG, 26, 26sG, 27G, 28G, 29G, 30G, 31G, 32G, or 33G. The size of the needle to be used will be chosen according to the viscosity characteristics of the combined preparation and the apparatus used to administer it.

 Endoscopic injection needles designed to minimize injection pressure are commercially available. In a particular embodiment, the combined (preferably pharmaceutical) preparation of the invention can be injected by an operator, preferably without difficulty, even when an endoscopic injection needle having a diameter of 23 G or higher numbering is used. Generally, the effective tube length of an endoscopic injection needle is 1,000 mm or more, preferably 1,500 to 2,500 mm.

The injection pressure of a combined preparation for submucosal injection can be measured, for example, using a 5 or 10 ml plastic luer-lock syringe that is filled with a measuring solution. An endoscopic injection needle with a diameter of 23G and an effective tube length of 1600 mm which is connected to the syringe can be used. The syringe is attached to a texture analyzer (EZ Test 500N manufactured by Shimadzu Corporation), and the piston of the syringe is pushed at a constant speed of 100 mm / min. The force required to discharge the measurement solution into the syringe through the tip of the endoscopic injection needle is measured at 25 ° C and is defined as the injection pressure. If the injection pressure is 14 kgf or more, the measurement solution is discharged through the tip of the endoscopic injection needle but there are leaks around the piston of the syringe, in addition the piston of the syringe does not move even pushing with the hand instead of the texture analyzer. When the injection pressure is about 1 1 kgf, the measurement solution can be discharged through the tip of the needle when the piston of the syringe is pushed by hand rather than with the texture analyzer. In a particular embodiment, optionally in combination with one or more of the features described in other aspects of the invention, the injection pressure of the combined (preferably pharmaceutical) preparation of the invention, measured as described in the preceding paragraph, is preferably from 0.1 to 12 kgf, more preferably from 0.25 to 10 kgf, even more preferably from 0.5 to 10 kgf, with particular preference from 1 , 0 to 7 kgf.

 In another particular embodiment, optionally in combination with one or more of the features described in other aspects of the invention, the retention time of the combined preparation is at least 30 minutes, preferably at least 45 minutes, for example between 30 minutes. minutes and 1 hour. More preferably, the retention time of the combined preparation is 60 or more minutes. Preferably, upon completion of the retention time, the complete reabsorption of the combined preparation progressively occurs. The retention time of the combined preparation is defined as the period of time in which a protuberance or wheal of sufficient height is maintained that allows the endoscopist to distinguish the boundaries of the lesion and remove it. Preferably, the average height of the protrusion in the retention time is at least 3 mm, preferably 4 mm or more, for example between 4 mm and 10 mm, between 5 mm and 8 mm or between 6 and 7 mm. Said protuberance is generally equivalent to 20% to 40%, preferably 30% to 35% of the cavity of the organ in which the mucosal resection is carried out.

 The combined preparation according to the present invention can be loaded and stored in a container. In another aspect of the invention there is further provided a kit comprising a container containing the combined (preferably pharmaceutical) preparation of the invention and, optionally, instructions for the use thereof as a combined preparation for use in such a treatment method. and as described in previous aspects of the invention. Preferably, for use by submucosal injection in a method of endoscopic resection treatment.

 The containers in which the pharmaceutical combination preparation is supplied can be any conventional container that can maintain the pharmaceutical formulation of the invention, such as a syringe, preferably luer-lock syringes, for example between 10 to 50 ml, a vial or a blister. The present invention can provide an assembly comprising one or more containers containing the combined preparation of the invention and an endoscopic injection needle. Alternatively, said kit may contain one or more endoscopic injection needles pre-loaded with the combined preparation of the invention.

It is contemplated that any embodiment discussed in the present specification may be implemented with respect to any, composition, pharmaceutical composition, combined preparation, kit, medical use, treatment procedure, and / or method of manufacturing a medicament of the invention. , and vice versa. It will be understood that the particular embodiments described herein are shown by way of illustration and not as limitations of the invention. The main characteristic features of the present invention can be employed in various embodiments without departing from the scope of the invention. invention. Those skilled in the art will recognize, or be able to determine using no more than routine experimentation, numerous equivalents to the specific procedures described herein. It is considered that these equivalents are within the scope of the present invention and are contemplated by the claims.

All publications and patent applications are incorporated herein by reference to the same extent as if each publication or individual patent application was specifically and individually indicated to be incorporated by reference.

 The use of the word "a" or "one" may mean "one," but it is also consistent with the meaning of "one or more," "at least one," and "one or more than one." The use of the term "other" may also refer to one or more. The use of the term "or" in the claims is used to mean "and / or" unless it is explicitly stated that it refers only to alternatives or the alternatives are mutually exclusive.

 As used in this specification and claims, the words "understand" (and any form of understanding, such as "understand" and "comprise"), "have" (and any form of having, such as "have" and "have"). has ")," include "(and any form of including, such as" includes "and" include ") or" contain "(and any form of containing, such as" contains "and" contain ") are inclusive or open and they do not exclude elements or stages of the procedure not mentioned, additional. As used herein, the term "consisting essentially of" limits the scope of a claim to the materials or steps specified and to those that do not materially affect the basic (s) and novel feature (s) ( s) of the claimed invention. As used herein, the term "consisting of" excludes any element, step or ingredient not specified in the claim except, for example, impurities usually associated with the element or limitation.

 The term "or combinations thereof" as used herein refers to all permutations and combinations of the items listed preceding the term. For example, "A, B, C or combinations thereof" is intended to include at least one of: A, B, C, AB, AC, BC or ABC, and if the order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC or CAB. Continuing with this example, combinations containing repetitions of one or more points or terms, such as BBB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, and so on are expressly included. The person skilled in the art will understand that typically there is no limit on the number of points or terms in any combination, unless otherwise evident from the context.

As used herein, approach words such as, without limitation, "about", "about", "approximately" refer to a condition which, when modified as such, is understood to not necessarily be absolute or perfect but would be considered sufficiently close for those skilled in the art to ensure the designation of the condition as present. The extent to which the description can vary will depend on how large a change can be instituted and still recognize a person skilled in the art that the modified characteristic feature still has the characteristics and capabilities required of the characteristic feature not modified. In general, but subject to the foregoing analysis, a numerical value in the present document that is modified by an approximation word such as "approximately" may vary from the value set to ± 1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%. Preferably said variation is 0%.

 DEFINITIONS

 The term, "aqueous composition" as used herein, refers to a liquid or semi-solid composition (eg, a solution, suspension or gel) containing water, optionally in combination with other mutually miscible solvents (organic solvents per example, soluble in water), and one or more chemical substances dissolved in it.

 The term "combined pharmaceutical preparation" and "pharmaceutical composition" refers to a combined preparation or composition that does not contain agents considered to be toxic or infectious at a concentration harmful to the subject to which it is administered by the appropriate route of administration. Preferably, said combined preparation or pharmaceutical composition is sterile.

 The term "pharmaceutically acceptable salt" as used herein refers to the relatively non-toxic organic and inorganic addition salts: acids of the compounds as described herein. These salts can be prepared in situ during the final isolation and purification of the compounds or by separately reacting the purified compound in its free base form with a suitable organic or inorganic acid and isolating the salt thus formed. Representative salts include the salts hydrobromide, hydrochloride, sulfate, bisulfate, nitrate, acetate, oxalate, valerate, oleate, palmitate, stearate, laurate, borate, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate, naphthylate, mesylate, glucoheptonate, lactobionate and laurylsulfonate, and the like. These may include cations based on the alkali and alkaline earth metals, such as sodium, lithium, potassium, calcium, magnesium and the like, as well as non-toxic ammonium, quaternary ammonium, and amine cations including, but not limited to ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like (see, for example, Berge SM, et al, "Pharmaceutical Salts," J.Pharm.Sci., 1977; 66: 1-19 which is incorporated in the present document by reference).

 The term "therapeutically effective amount" as used herein refers to an amount that is effective, upon administration of an individual dose to a subject of the composition of the invention.

The term "subject" as used herein, refers to a mammalian subject. Preferably, it is selected from the group consisting of a human being, in companion animals, non-domestic farm animals and zoo animals. For example, The subject can be selected from a human being, dog, cat, cow, pig, sheep, horse, bear, and so on. In a preferred embodiment, said mammalian subject is a human subject.

 The term "neoplasia" or "neoplastic lesion" as used herein covers dysplasia, precancerous lesions, cancerous lesions, neoplastic cells, tumors, benign tumors, malignant tumors, solid tumors, carcinomas, etc., preferably located in the skin and Soft tissues of any part of the body.

 As used herein, the term "precancerous lesion" includes syndromes represented by abnormal neoplastic growth, including dysplastic syndromes. Non-limiting examples include, in addition to the dysplastic syndromes, nevi, polyposis syndromes, intestinal polyps, precancerous lesions of the cervix (i.e., cervical dysplasia), prosthetic dysplasia, bronchial dysplasia, breast, and / or bladder, whether The lesions are clinically identifiable as not.

 The term "pseudoplastic" as used herein refers to a fluid in which the apparent viscosity or consistency decreases with an increase in the shear rate. Thus, the decrease in fluid viscosity is not time dependent, and the viscosity of the fluid decreases almost instantaneously by the application of pressure at the moment when the fluid is injected into the tissue with the endoscopic injection needle but quickly regains its viscosity initial just after the release of the pressure. Pseudoplasticity can be evaluated using the Casson performance value. A higher Casson yield value means that the resting viscosity is higher and, therefore, the elevation can be maintained for a longer period of time without diffusion. A too high value of Casson performance causes a high injection pressure, which affects the manageability during injection. The Casson yield value of the pharmaceutical composition of the invention is preferably from 0.1 to 100, more preferably from 0.5 to 75, and even more preferably from 1 to 50.

EXAMPLES OF THE INVENTION

Example 1.- Study efficacy and safety: comparison with glycerol 10%

Material and methods

Retrospective study with 20 patients who attended the Endoscopy Consultation of the Poniente Hospital, in Ejido (Almería) for the performance of a submucosal endoscopic mucosectomy. A solution of 10% glycerol, 5% fructose, 0.005% adrenaline and methylene blue was used in 10 patients. In another 10 the new study solution: extemporaneous solution of Ac. Hyaluronic acid (AH) of low density (UROMAC ^® ) at 0.003%, Carboxymethylcellulose sodium (CMC) 1500-4500 (94224, Guimana) at 0.2% characterized by a viscosity of 1,500-2,500 mPa * S at 1% dilution, Fructose csp pH = 5-6, Adrenaline 0.005%, Methylene Blue 1 drop (0.05 mi) in Physiological Serum csp 50 ml_. The solution was conditioned in syringes of BD Plastipak ™ 50 mL luer-lock syringe and stored at 2-8 ° C until use.

Data collected: number of lesions and size (cm), number of resections for complete removal of the lesion (sessions), number of injections of the solution, volume injected into each lesion (mL), duration of the wheal useful during mucosectomy ( minutes), time (minutes) from the beginning of the endoscopic intervention until the start of reabsorption, vascularization of the tissue (Good / Regular / Bad), generation of fumes that make visibility during the surgical intervention (YES / NO), bleeding during surgical intervention (YES / NO), type of bleeding (LEV / MODERATE / MASSIVE), complications of the surgical intervention, evolution of the tissue (in the revisions) after the administration of the drug (inflammation / tissue damage).

Both solutions were processed under sterile conditions in Horizontal Laminar Flow Cabin and filtered inside the horizontal laminar flow cabin with a mini spike plus v® filter (Braun) sterilizing filter. The following physical-chemical stability conditions were considered: a pH = 5-6, absence of visual precipitation of solutes, as well as visual absence of particles during 30 days in both solutions. To determine the microbiological stability, samples of both solutions were cultured at days 0, 7, 15, 30 days after their elaboration. Likewise, given the momentary impossibility of determining the viscosity, we opted for the determination of the osmolarity of both solutions.

The solution was administered with syringes B / BRAUN Omnifix ^R syringe luer-lock 10 mL, which were attached to a catheter 200 to 240 cm long with a 23-gauge endoscopic injection needle. The minimum working channel required for this material is 2.0-2.8mm (Interject ™ Contrast - Injection Therapy Needle Catheter - Boston Scientific).

Results

 In all patients, the borders of the polyps were perfectly delimited before their removal, using diluted methylene blue. The dye stained the submucosa and allowed the perfect assessment of the depth of the eschar in the patients, as well as delimited the edges exactly. The handles used were oval and multifilament morphology with sizes of 30 and 10 mm, facing the large lesions using "Piecemeal" endoscopic resection. Monopolar current was used with coagulation mixture according to the recommendations of the manufacturer of the electrosurgical unit.

All the patients had between 3-6 lesions, with a surface of between 15-40 mm of surface and a depth of the tissue excised between 0.5-1 mm. We did not observe a difference in the number of sessions for the elimination of the lesions according to the use of one solution or another. With the 10% Glycerol solution, the patients required between 100-200 mL of solution and it was necessary to reinject the solution to 6 patients. While with the study solution an average of 50 mL of solution was needed and it was not necessary to reinject any patient. After the mucosectomies performed, we observed when inserting the solution in the intestinal sub-mucosa a "wheal", which separated during the 30-45 minutes that the interventions lasted the area of the polyps of the area irrigated by blood vessels and musculature, compared to the 15 minutes observed with Glycerol 10% which forced us to reinject patients sometimes. The use of the new mucosectomy formula gave us an optimal elevation of the lesion during a prolonged time, conferring greater safety.

All patients are in clinical follow-up without presenting any complication associated with the intervention. By introducing adrenaline, the potential risk of hemorrhage was reduced and recovery was faster. Subsequently, the solution injected into the sub-mucosa was reabsorbed without problems and we did not show signs of inflammation or tissue damage secondary to the solution used. However, patients are still being followed up and this should be confirmed in future visits. We did not evidence massive bleeding in any of the patients or complications during the intervention.

The vascularization of the tissue submitted to the surgical intervention was good. In 3 patients moderate smoke presence was evidenced with the 10% Glycerol solution, which partially obscured the visibility of the area to be removed, not being observed with the new solution.

Both solutions presented microbiological stability for at least 30 days. The mean osmolarity of the study solution was 1,710 mOs / L, while that of Glycerol 10% was 809 mOs / L. However, from the physico-chemical point of view, and in the absence of studies, we could only recommend a 24-hour stability between 2-8 ° C in both solutions.

Discussion

 After mucosectomies, in our patients we optimized the result of the new formula, whose effect is mechanical-pharmacological. When the solution is introduced into the intestinal sub-mucosa, a "wheal" is produced, which separates the area of the lesion from the area irrigated by blood vessels and musculature for about 45 minutes. Thus, when the polyp is removed, the risk of bleeding, injury and recovery is reduced faster. Subsequently, this solution injected into the sub-mucosa seems to be reabsorbed without causing adverse effects, allowing sufficient time for the mucosectomy to be performed.

 Our formula could have a great potential of use for injection of the sub-mucosa, such as in mucosectomy of early neoplastic lesions of the digestive tract and in endoscopic sub-mucosal dissection. With our preparation the "wheal" that was generated seems more durable than with Glycerol, Physiological Serum, or other products separately.

Conclusions

The advantages of our solution are: high permanence of the compound at intestinal level, high tissue expansion by injecting with a smaller amount of product, optimal viscosity, total reabsorption of the product administered, physical-chemical and microbiological stability, and all at very low cost.

Example 2.- Study efficacy and safety: retrospective study of the composition of the invention in 10 additional patients

Material and methods

 Retrospective study in 10 patients who attended the Endoscopy Consultation of the Poniente Hospital, in Ejido (Almería) for the endoscopic resection of the colonic mucosa of flat lesions≥15mm.

Study solution: Ac. Hyaluronic (AH) 150-250 centipoise (UROMAC ^® ) at 0.003%, Carboxymethylcellulose sodium (CMC) 1500-4500 (94224, Guinama) at 0.2%, characterized by a viscosity of 1,500-2,500 mPa * s, Fructose csp pH = 5-6 and Methylene Blue 0.05 ml_ in Physiological Serum csp 50 ml_. To elaborate the mucosectomy solution, we used a CMC concentration of 0.2% in the final solution, starting from a 2% stock solution - 4,500 mPa.s. In the process of elaboration of the Carboxymethylcellulose (CMC) 2% of high viscosity ((4,474 mPa.s in 2% solution) in saline 0.9% we control at all times the agitation (soft, with magnetic stirrer, and the temperature that will be at all times 50 ° C) until after about 5 hours of agitation we achieve a gel with the desired viscosity.The following protocol was used for the preparation of the study solution:

Weigh 35g of fructose in a beaker and dilute with 100 ml of physiological saline, stir until dissolved. Weigh 20 g of carboxymethylcellulose of high viscosity to 2% and add to the previous solution. We continue in continuous agitation while adding 1 ml_ of 1% adrenaline, 6ml_ of low viscosity hyaluronic acid and 1 drop of methylene blue. We make up to 200ml_ with SF. Shake everything until its complete homogenization.

Sterility is achieved by sterilizing filtration in Horizontal Laminar Flow Cabin (CFLH) class 100, located in a space (room) with partially controlled conditions (Class 100,000). Transfer to the CFLH and pour the content of the beaker with a cut-off transserver and 0.22 micron filter into syringes of BD PlastipakTM luer-lock syringe 50 mL and stored at 2-8 ° C until use.

All patients underwent surgery after deep sedation, by an anesthetist, with midazolam and IV pethidine. The edges of the polyps were perfectly delimited to patients before their removal, using diluted methylene blue. The dye stained the mucosa and allowed the perfect assessment of the depth of the eschar in the patients, as well as delimited the edges exactly. The handles used were oval and multifilament morphology with sizes of 30 and 10 mm, facing the large lesions using "Piecemeal" endoscopic resection. Monopolar current was used with coagulation mixture according to the recommendations of the manufacturer of the electrosurgical unit. Data collected: demographic, number of lesions, number of injections of the solution, volume injected (mL), duration of the wheal useful during the mucosectomy (minutes), time (minutes) from the beginning of the endoscopic intervention until the beginning of reabsorption is observed, bleeding during the surgical intervention, type of bleeding), complications of the surgical intervention, tissue evolution (in the reviews in months 1, 3) after the administration of the drug (inflammation / tissue damage).

The study solution was prepared under sterile conditions in Horizontal Laminar Flow Cabin and was filtered inside the horizontal laminar flow cabin with a mini spike plus v ^® filter (Braun) sterilizing filter.

The solution was administered with syringes B / BRAUN Omnifix® 10 mL luer-lock syringe, which were attached to a 200 to 240 cm long catheter with a 23-gauge endoscopic injection needle. The minimum working channel required for this material is 2.0-2.8mm (InterjectTM Contrast - Injection Therapy Needle Catheter - Boston Scientific).

Results

The 10 patients of the study (90% males of 63.5 years) presented an average of 3 colonic lesions (range: 1-5), with an average size of 27 mm (range: 15-50 mm), requiring a total of 24 mL of mucosectomy solution per patient. It was not necessary to reinject any patient due to loss of consistency of the "habón". The solution remained in the intestinal submucosa for an average of 72 minutes. After the mucosectomies performed, we observed several cases in which the intervention lasted up to 90 minutes (3 patients) and in which the "wheal" generated, to separate the mucosal layer of the "own" muscularis, was effective throughout the intervention, not observing complications during the intervention in any patient. Subsequently, the solution injected into the submucosa was reabsorbed, in all cases, without problems and we did not show signs of inflammation or tissue damage secondary to the solution used, neither during the intervention nor during the follow-up visits in months 1 and 3 after the performance of mucosectomy.

Discussion

 The use of the new formula for mucosectomy provided an optimal elevation of the large colonic lesions for more than 60 minutes, giving greater security to the intervention. All patients are currently in clinical follow-up without presenting any additional complication secondary to the intervention.

In our study we observed that introducing the solution into the intestinal sub-mucosa produced a "wheal", consisting of time, which separated up to 90 minutes, the area of the lesion from the area irrigated by blood vessels and musculature. Thus, when the polyp is removed, the potential risk of complications is reduced. This prolonged permanence of the solution in the intestinal submucosa is mainly due to the mixture of two mucopolysaccharide substances. Viscosity and osmolarity are key to the efficacy and safety of the solution, as the study presented here shows. In our study, the solution was reabsorbed completely without causing security problems. Precisely there lies part of the innovation, in using a CMC of very high viscosity, which surprisingly allowed that in association with AH we need low concentrations of both components to achieve an optimal clinical result in terms of efficacy and safety. In the present study we have used a low viscosity AH but it is also possible to use medium-high viscosity AH.

Conclusions

 In this study, it is demonstrated that the pharmaceutical composition of the invention, used in endoscopic colonic Mucosa Resection in large lesions, remains in the intestinal submucosa for more than 60 minutes, in some cases up to 90 minutes. This fact allowed in all cases to perform an effective and safe intervention in the extirpation of large intestinal polyps due to the greater permanence of the compound at the intestinal level compared to standard treatments, using a smaller amount of product.

Example 3 - Characterization and stability study of the solution of the invention: comparison with 10% glycerol solution and 0.2% hyaluronic acid solution Material and methods

 Study carried out by the UGC of Pharmacy of the Hospital de Poniente and the Department of Galenics and Pharmaceutical Technology of the Faculty of Pharmacy. University of Granada during 2016. The procedure for preparing the formulations indicated below is as described in Example 2.

Composition of the formulations

1. A freshly prepared sample (2/15/16 - Lot: M1502) with glycerol 10% + fructose csp pH = 5-6 + 0.05 mL of methylene blue 1% + physiological saline csp 50 mL. Stored from the start ^at room T (Formulation 1).

2. A freshly prepared sample (2/15/16 - Lot: M1502) with hyaluronic acid 0.4% + fructose csp pH = 5-6 + 0.05 mL of methylene blue 1% + physiological saline csp 50 mL. Stored from the start ^at room T (Formulation 2).

 3. Two freshly prepared samples (2/15/16 - Lot: M1502) of the study solution WITHOUT adrenaline (Carboxymethylcellulose 0.2% + Hyaluronic acid 0.003% + fructose csp pH = 5-6 + 0.05 mL blue of methylene 1% + physiological saline csp 50 mL). Stored from the start in a refrigerator (2 ° -8 ° C) (Formulation 3), and at room temperature (Formulation 4).

 4. Two freshly prepared samples (2/15/16 - Lot: M1502) of the study solution with adrenaline (Carboxymethylcellulose 0.2% + Hyaluronic acid 0.003% + fructose csp pH = 5-6 + 0.05 mL blue of methylene 1% + physiological saline csp 50 mL) Stored from the beginning in refrigerator (2 ° -8 ° C) (Formulation 5), and room temperature (Formulation 6).

PH study

The pH determination of the prepared formulations was carried out with a Crison micropH 200, Model 2000 pH meter. The pH was determined at different times. A significant variation of the pH over a suitable value for the mucosectomy gels could indicate a degradation of the solution or a wrong elaboration.

Theological study

The rheological characterization of the formulations was performed at 25 ° C using a HAAKE Rheostress 1 rotational rheometer (Thermo Fisher Scientific, Karlsruhe, Germany) with a parallel plate geometry configuration, with a fixed bottom plate and a movable top plate (Haake PP60 Ti, 6 cm in diameter). Different spaces between plates were tested until a separation of 0.1 mm was selected.

The device consists of the following elements: Haake VT500 viscometer and thermostatic bath with water recirculation system (Haake C25P). The Rheometer is connected to a computer equipped with the HAAKE software RheoWin ^® Job Manager V. 3.3 to carry out the test and RheoWin ^® Data Manager V 3.3 (Thermo Electron Corporation, Karlsruhe, Germany) to carry out the data analysis obtained.

The viscosity and flow curves were recorded for 3 minutes during the acceleration or ascent period from 0 to 100 s ^_1 , 1 minute at 100 s ^"1 (constant velocity period), and finally 3 minutes during the descent period of 100 to 0 s ^{_ 1.} The viscosity values at 100 s ^"1 were determined at and ten days for the samples stored at 4 and 25 ° C in triplicate. Optical characterization by "multiple light scattering"

In order to predict long-term stability, the formulations were evaluated by means of multiple light scattering, using the Turbiscan® Lab device (Formulaction Co., L'Union, France). The light source is a pulsed near infrared (λ = 880 nm). The undiluted samples are placed and maintained in a cylindrical glass cell that is completely scanned by a reading head. In this way, a light flow pattern is obtained as a function of the height of the sample, which corresponds to the macroscopic fingerprint of the sample at a given moment. The measurements were made in triplicate and at room temperature.

Stability study

 The stability study was carried out in parallel to the characterization, in order to analyze the variations in pH, viscosity, rheological behavior and appearance, since they may be related to structural changes that may occur within the formulations.

 Each master formula was prepared in sufficient quantity and stored in aliquots in topaz vials at different temperatures: 4 ° C and 25 ° C.

The complete characterization of each formula will be carried out 24 hours after its preparation, this being day 0. The study period was 6 months.

The number of determinations made by formula, time and temperature have been 3. The mean values and the standard deviation were calculated for each of the tests. All the results were subjected to a statistical treatment of ANOVA for a confidence level of 95%, in order to check if there were significant differences between the means compared.

Optical microscopy

 The morphology of the same particles was determined by light microscopy in an Olympus BX40 microscope and Sony Cyber-shot DSC-W210 camera (Tokyo, Japan). Clean microparticles resuspended in bidistilled water were used.

 For this, a sample of 0.25 g of clean microparticles was taken and diluted in 5 ml of bidistilled water. Once the sample was placed on a slide, a 0.1% (w / v) methylene blue solution was added in bidistilled water.

RESULTS AND DISCUSSION

PH study

 The data shown below correspond to the formulations 3 to 6. The measurements were made after 6 months from the preparation of the formulations.

 • Formulation 3: NO adrenaline in refrigerator 5.29.

• Formulation 4: NO adrenaline T ^a room 5.99.

 • Formulation 5: WITH adrenaline in refrigerator 5.84.

• Formulation 6: WITH adrenaline T ^a environment 5.02.

The results obtained show a slightly acid pH. The pH values in all formulations are between 5-6. This interval corresponds to the processing pH. No significant variations were observed in the pH value based on the composition or time elapsed in the study. This fact will influence the rheological stability of the formulations since an increase in pH would increase the viscosity of the systems with sodium carboxymethylcellulose (Voigt and Bornschein, 1982).

 After one year from the elaboration, the values of the formulation without adrenaline were: 5.60 (conserved in a refrigerator) and 4.93 (conserved at room temperature). A significant change in value is observed at 25 ° C.

 Regarding the pH values of the formulas without combining both polymers:

 • N2803 Sun Mucosectomy (CMC without AH) in refrigerator: 4.68

 • N2803 Sun Mucosectomy (AH without CMC) in a refrigerator: 5.49 Theological Study

Table 1 presents the mean viscosity values (mPa-s) of the samples at 100 s ^_1 , at time 0 and after 6 months.

 Table 1. Viscosity values as a function of time (mean ± standard deviation), n = 3.

The samples that combine carboxymethylcellulose with hyaluronic acid have a much higher viscosity (about ten times greater) than the rest (samples with glycerol or samples with hiaiuronic acid). This is due to the viscous synergism which has its origin in the interaction between the components of a system, so that the viscosity of the latter turns out to be greater than the sum of the viscosities of the components separately.

The presence of adrenaline does not significantly affect the viscosity value.

The preservation of the samples at low temperatures in the refrigerator shows a remarkable increase in the viscosity value with respect to the samples stored at room temperature. This is due to the fact that, as the temperature increases, the viscous forces are overcome by the kinetic energy, resulting in a decrease in viscosity (Cancela et al., 2005: "Effects of temperature and concentration on carboxymethylcellulose with sucrose rheology "Journal of Food Engineering, Vol. 71, pp 419-424).

 These results should be taken into account for the conservation of the samples only in the case of not achieving the necessary efficiency for the realization of the musectomy with the viscosity of the samples maintained at room temperature.

 Regarding the influence of the conservation time, no significant changes were observed, being able to consider all the samples stable during the 6 months of study.

 In addition to the temperature, the viscosity can be very affected by variables such as the deformation velocity gradient and pressure, among others, these being the most important.

 The variation that the samples undergo with the velocity gradient allows to classify the different types of fluids that can be found from the rheological point of view.

Thus the rheological characterization served not only to evaluate the stability of the formulations but also to know the flow behavior of the final system. The behavior of the formulations is one of the essential criteria in the development of sanitary products, since it intervenes in the functional properties of the final product during the administration (mechanical behavior), the quality control, the design of basic operations such as pumping, mixing , packaging, storage and physical stability.

Figures 1 to 12 show the viscosity (squares) and flow (triangles) curves of the mucosectomy formulations studied. In the Fluidity Curve, the shear stress versus the deformation velocity (τ vs D) is represented, while in the Viscosity Curve the viscosity is plotted as a function of the deformation velocity (μ vs D).

As can be seen, formulation 1 (at 6 months) and formulation 2 (at time 0 and 6 months) present a Newtonian behavior, the flow curve is a straight line that starts at the origin, that is, there is a relationship linear between the shear stress and the deformation speed. Furthermore, it is observed in the viscosity curve that the viscosity is constant for any applied deformation velocity. This behavior continues to be maintained throughout the study time, 6 months, in the formulation made with hyaluronic acid. However, in the case of the glycerol-based formulation, this behavior varies with time, showing a plastic flow at time 0. According to the figure, this type of fluid behaves like a solid until it exceeds a minimum shear stress (effort threshold) and from that value behaves like a liquid. However, it can be seen that at intermediate deformation speeds (50 s ^_1 ), a practically constant value of apparent viscosity is reached, which implies a Newtonian behavior.

As for the formulation of the invention, the viscosity tends to decrease as the shear rate increases. This behavior is typical of pseudoplastic fluids. In this case, formulations made with carboxymethylcellulose in combination with hyaluronic acid have a high viscosity at rest that can reach values close to 35 or 16 mPa.s, at 4 ° C and 25 ° C, respectively. Suitable from the technological point of view if you want to avoid sedimentation as it would make it difficult for particles of other active ingredients and / or pharmaceutical excipients such as an osmotic pressure regulating agent, pH regulating agent, a coloring agent, and a vasoconstrictor agent or hemostatic are added. Ideal also from the physiological point of view, the high viscosity will provide a wheal or protuberance suitable to accurately eliminate the injury.

 This behavior is similar, regardless of the storage temperature or the presence of adrenaline.

Thixotropy is a property associated with the construction and breakage of structures under an effort. Thixotropic fluids are characterized by a change in their internal structure when an effort is applied. The chains that make up its molecules break, the viscosity decreases when a force is applied, and after a time of rest it increases again, when the force ceases due to the reconstruction of its structures; this means that they exhibit a relationship viscosity - time. The area of the hysteresis cycle can be considered as an estimate of the degree of thixotropy; and it is generally accepted that a larger area of the hysteresis cycle will have stronger thixotropic properties and therefore a recovery of its slower structure.

The thixotropy values of the samples tested are shown in Table 2:

 Table 2. Thixotropy values of the formulations tested The samples studied showed hardly appreciable values of thixotropy and therefore, are fluid practically independent of the time of application. This insignificant dependence of the viscosity with time is due to the fact that the formulations with sodium carboxymethylcellulose (CMC-Na) and hyaluronic acid present a structure sufficiently rigid to remain unaltered with shearing with a very slight structural collapse. This behavior is maintained after 6 months from its preparation.

 There are many models that have tried to explain the behavior of non-Newtonian fluids. In this work the data obtained were set to the following: Newton, Bingham, Ostwald-De-Waele, Herschel-Bulkley and Casson. Table 3 shows the best model for each formulation and the values of each parameter, respectively. The criterion for selecting the best model was based on the adjusted one with the highest linear correlation coefficient (r). For formulations 1 and 2 it was also necessary to take into account the Chi-square value.

From the results obtained for the formulations with CMC-Na, it is deduced that the Herschel-Bulkley method is the rheological model that best adjusts the experimental data.

Best rheological model

 CODE DESCRIPTION SAMPLE

 0 days 6 months

 Very close

 1 M1502 10% glycerol in fridge Herschel-Bulkley

Newton M1502 0.4% hyaluronic in

 2 Newton Newton

 fridge

 M1502 WITHOUT adrenaline in

 3 Herschel-Bulkley Herschel-Bulkley

 fridge

M1502 WITHOUT adrenaline t ^a

 4 Herschel-Bulkley Herschel-Bulkley

 ambient

 M1502 WITH adrenaline in

 5 Herschel-Bulkley Herschel-Bulkley

 fridge

M1502 WITH adrenaline t ^a

 6 Herschel-Bulkley Herschel-Bulkley

 ambient

 Table 3. Rheological models that best predict the behavior of the formulations studied.

Optical characterization by means of multiple light scattering. The equipment has an optical head with an infrared light source and two detectors (T and BS) that run the entire height of the sample inside the cell. of Cristal. With the collected data of light intensity in Transmission (T) (backscattering) and Reflection (BS), more commonly called "Backscattering", profiles were obtained that allowed us to characterize the sample and detect processes such as Sedimentation, Flocculation, Coalescence, Separation of phases, Flotation, etc. In short, this technique allows to detect changes in size or location within the samples and allows to evaluate the physical stability, avoiding the dilution of the formulation. Another important advantage is the ability to detect destabilization phenomena long before the human eye and is considered a device that predicts long-term stability, being able to detect the destabilization of the formula before the classical stability methods. The tests were carried out at 25 ° C and the sample took 0, 30 days and 90 days.

 Figures 13 to 20 show the transmission and backscattering profiles of the formulations tested.

 In order to interpret them, it must be taken into account that the left side of the curves corresponds to the lower part of the vial, while the right side corresponds to the upper part. Point out that the region below 5 mm marks the metal base and backscatter above 52 mm the beginning of the free surface of the sample.

When a sedimentation phenomenon occurs, the reflection signal increases over time, in the lower part of the vial. When the sample suffers a creaming phenomenon, an increase occurs in the upper part of the vial. If the destabilization process occurs through aggregation, the backscattering increases over time along the entire road.

If the signal has a deviation less than or equal to ± 2%, it can be considered that there will be no significant differences in drop size. Variations of 10% indicate instability of the formulations.

The superposition of the signal of transmission and / or reflection from hours 0 to 24 shows the stability of the formulations, indicating the absence of destabilization processes. East The pattern is repeated over time and independently of the presence of adrenaline or depending on the storage temperature. Therefore, it could be concluded that the formulations constitute homogeneous dispersions.

Example 4. Preparation of the microparticles of the combined preparation of the invention

For the preparation of the microparticles, the internal ionic gelation emulsification technique has been selected.

This technique consists in the preparation of an A / O emulsion in which the internal or aqueous phase is formed by a polymer (sodium alginate) or a mixture of polymers (of different nature), while the external or oily phase consists of an oil in which a surfactant agent (polysorbate 80) is dispersed.

The steps to follow are those:

 Sodium alginate (2-5%) or the mixture of alginate with various polymers dissolves under mechanical agitation in distilled water. When we talk about polymer mixture, it can refer to physics or chemistry. In addition to water, the use of different solvents in the microcapsule preparation method is estimated to have an influence on the physicochemical properties of the intervening molecules, such as solutions of vasoconstrictor extracts such as the fluid extract or tincture (2-4ml_) of ruscus, bilberry, horse chestnut or meliloto, or the colloidal solution of hyaluronic and cellulose in which you are dispersed. The polymers selected as coavalent agents can be:

Of natural origin:

 • Chitosan

 • Exopolysaccharides of bacterial origin

 · Derivatives of cellulose: Hydroxypropylmethylcellulose, sodium methylcellulose, cellulose acetophthalate

 • Pectin

In synthesis:

 • Poloxamer 127

· Eudragit ^® S 100, L-100, L 12.5, S 12.5, FS30D, RS, FS and NM

 • Shellac

 • PLGA

Some polymers have been called "intelligent" because of their rapid response to changes in environmental physicochemical conditions, which involve pronounced changes in their properties. The stimuli to which the polymers respond can be: • Physical: such as temperature, ionic strength, solvents, radiation, electric fields, mechanical stress, pressure, sonic radiation and magnetic fields;

 • Chemicals: such as pH, specific ions and chemical agents;

 • Biochemical: as substrates of enzymes, related ligands and other biological agents.

Preferably the addition of vitamin K 1 or phytomenadione (5-10 mg) is made as preventive of bleeding during the operation, in the aqueous phase or in the oil phase.

Once the aqueous phase is formed, a calcium source is added, in the form of insoluble salt, calcium carbonate (0.5-1%) that is dispersed in the aqueous phase, progressively, the oil phase is added, composed of a solution homogeneous polysorbate 80 (2-4%) in oil, this may be of a different nature:

• Vegetable with anti-inflammatory properties:

 - Olive oil

 - Rosemary oil

 - Argan oil

 · Mineral: Liquid paraffin

To form the oily external phase emulsion, the two phases are subjected to mechanical agitation 500-2000 r.p.m. for 10-30 minutes.

After this time, acetic acid (4-8%) is added to the emulsion to solubilize the calcium ions and give rise to the formation of the pre-gelled particles. The next step was to add a solution of calcium chloride at 5-10% (w / v) to produce the hardening of the polymer droplets under mechanical agitation at 500-2000 r.p.m. for 10-30 minutes.

The particles are cleaned with bidistilled water using successive centrifugation cycles. To favor its optimum conservation, it is matured in calcium solution for 2-10 minutes and desiccated at 25-37 ° C to favor the elimination of water, thus eliminating the possible contamination or degradation of the same. .

Example 5. Studies of microparticles

PH study

 For the formulations combined with microparticles the values obtained were: Proportion of microparticles is 1: 3 (p: v) (1 gram of microparticles in 3 ml_ of musectomy solution)

CODE DESCRIPTION SAMPLE pH t = 24 hours (04/25/2017)

 1 to N2803 Sun Mucosectomy 1 (CMC + AH) in refrigerator 5.56

2a Microparticles stored in a refrigerator 4.50 Theological study

Table 4 shows the viscosity values of the selected solution in combination with the microparticles. In this case, the proportions studied (microparticles: medium) were two: 1: 3 and 1: 6.

The study was carried out under different conditions of preservation temperature (8 ° C, 25 ° C); In addition, the viscosity at 37 ° C of samples stored under refrigeration was determined, since this parameter could condition not only the conservation of the microparticles but their behavior once administered.

As can be seen as the temperature increases, the formulation undergoes a marked decrease in viscosity. The higher viscosity occurs at 8 ° C which will reduce the sedimentation rate of the particles and / or dispersed microparticles, guaranteeing the stability of the system. However, as expected, when the colloidal reference solution is combined with the alginate microparticles (regardless of the proportion), the viscosity is greatly increased. This increase occurs at all the temperatures studied, so that the new microparticulate formulation could be kept at room temperature and still have a viscosity higher than the reference solution at 8 ° C. Which would represent an advantage in terms of conservation.

It is interesting to note the increase in viscosity suffered by most formulations over time (whatever the temperature). This must be taken into account during the administration and selection of the needle.

Figures 21 to 26 show the viscosity and flow curves of the formulations that combine the colloidal dispersion and the microparticles. The viscosity tends to decrease as the shear rate increases, showing, like the previous profiles, a pseudoplastic behavior. This profile is similar, regardless of the storage temperature or determination (8, 25, 37 ° C) and the proportion of microparticles (1: 3 or 1: 6). All the formulations and different study conditions were adjusted to the Herschel-Bulkley rheological model. These profiles and models are still maintained 30 days after their preparation (graphics not shown). As expected, the microparticles give the formulation an increase in thixotropy and, therefore, a recovery of its slower structure (Table 5).

 Table 5. Thixotropy values of the microparticulated formulations (thixotropy values at 30 days not yet available)

Figures 27 and 28 show the transmission and backscattering profiles of the microparticulate formulations.

The instability of the formulations is obvious as indicated by the distances between spectra, with values equal to or greater than 10%. This instability is due both to changes in the location of microparticles (the transmission spectra clearly shows how the transmission progresses in the upper half of the vial as time progresses) and to the increase in size after the union of several microparticles (the spectra of BS increase over time in the lower half of the vial), this fact indicates the formation of flocs and therefore a rapid destabilization but easy redispersion. As the conservation time passes, there is a decrease in the percentages of transmission and increase in the BS percentages. This is probably due to the increase in the size of the flocs.

Optical microscopy

Finally, the microparticles were observed under optical microscopy before being injected and after passing through a 23G needle, in order to know if they would maintain their morphology and size after being administered. The formulations constituted by the proportions 1: 3 and 1: 6 (microparticles: reference medium) were tested. The microphotographs taken by optical microscopy (Fig. 29 to 31) show that the synthesis methodology gives rise to spherical microparticles with well-defined edges, presenting great sphericity (Figure 29).

They also reveal the differentiated state of the MP (Fig. 30). Regarding the observations made after passing through the needle, no deformations or size changes were observed for the dispersions tested (Figure 31).

Example 6. Test to determine the height and duration of wheal of a mucosectomy solution and analogs in vivo model.

Introduction Endoscopic mucosal resection or mucosectomy is an endoscopic technique that allows removal of lesions of the gastrointestinal mucosa using a polypectomy loop and the use of a submucosal injection. For this, it is essential to elevate the mucosa and that this elevation lasts in time after said injection. These conditions together with the experience of the endoscopist can guarantee optimal conditions of efficacy and safety in the intervention. Therefore, we propose the performance of this trial where several analogues of a mucosectomy solution were compared in an in vivo model.

Methodology

For the preparation of the solutions of AH (Uromac®, Nakafarma - low viscosity - less than 300 kDa), sodium CMC (2237 mPa * S diluted 1% at 25 ° C) and sodium AH + CMC was used a saline solution 0 , 9%, mechanical stirring 500-1000 rpm at 50-75 ° C.

For the preparation of the microparticles, the internal ionic gelation emulsification technique was used. This technique consists in the elaboration of an A / O emulsion in which the internal or aqueous phase is formed by a polymer (sodium alginate) or a mixture of polymers, while the external or oil phase is composed of an oil in which a surfactant is dispersed (eg, polysorbate 80).

Sequence of synthesis of the microparticles

Sodium alginate (2-5%) was dissolved under mechanical stirring in distilled water. Once the aqueous phase was formed, an inorganic calcium salt, calcium carbonate (0.5-1%) was added and dispersed in the aqueous phase. Progressively, the oil phase was added, composed of a homogeneous solution of polysorbate 80 (2-4%) in oil, in this case soybean oil was used.

To form the oily external phase emulsion both phases were subjected to mechanical agitation 500-2000 rpm for 10-30 minutes. After this time, the emulsion was added with acetic acid (4-8%) to form the pregelled particles.

Then, a 5-10% (w / v) calcium chloride solution was added to produce the polymer droplet hardening under mechanical stirring at 500-2000 rpm. for 10-30 minutes.

The particles were cleaned with bidistilled water using successive centrifugation cycles. Then, it was matured in calcium solution for 2-10 minutes and desiccated at 25-37 ° C.

Finally, combinations of each solution with MPP were prepared in a 1: 3 ratio. Test in vivo model

The test to determine the height of the wheal at the gastrointestinal level was performed in pig stomach, with extrapolar results in the gastrointestinal area and internal mucous membranes of the body (bladder, vagina). The stomachs of three pigs were extracted the day before the test and were kept in a refrigerator, in Hanks' solution, until their use. The stomach was divided into fractions and these were placed in a tray, which was placed on a thermostated bath at 37 ° C.

The solutions studied were:

- Solution 1: AH 0.003%

 - Solution 2: CMC 0.2%

 - Solution 3: AH 0.003% + CMC 0.2%

 - Solution 4: AH 0.12% + CMC 0.1%

 In addition to these solutions, the influence of these in combination with powdered microparticles (MPP) was also studied. For this, combinations of each solution with MPP were prepared in a 1: 3 ratio. Once the solutions were prepared, 10 mL was injected into the submucosa of the stomach with a 23G needle. The height of the wheal formed was determined at different times (start, 1, 3, 5, 7, 10, 15, 30, 60 and 90 minutes).

0 1 3 5 7 10 15 30 60 90

Samples / Time

 min min min min min min min min min min

0.003% AH 1 .5 1 .4 1 .2 1 .0 1 .0 0.9 0.9 0.9 0.8 0.8

Height Habón 0.2% CMC 1 .9 1 .7 1 .6 1 .7 1 .6 1 .6 1 .5 1 .6 1 .2 1 .0 in cm 0.1% CMC +

1 .8 1 .8 1 .6 1 .6 1 .6 1 .5 1 .5 1 .4 1 .5 1 .5 0.12% AH Table 1: Height of wheal (cm) in pork stomach (solutions WITHOUT microparticles). The results obtained are expressed as the average of the 3 replicas made for each solution.

 Table 2: Height of wheal (cm) in pork stomach (solutions WITH microparticles). The results obtained are expressed as the average of the 3 replicas made for each solution.

Conclusions

To date, microparticles have been used mainly as vehicles or drug delivery systems, incorporating them inside. However and unexpectedly, during the synthesis process the microparticles of the invention have incorporated the mucosectomy solution into their external structure. Therefore, we can affirm that they are not a vehicle of drugs but that they form, together with the mucosectomy solution, a treatment in themselves.

As shown in tables 1 and 2, the use of a microparticle solution (0.1% CMC + 0.12% AH), contrary to what is expected by a person skilled in the art, does not generate, as shown in the examples, a higher wheal, consistent or with greater mucoadhesion than that obtained by the solution of 0.1% CMC + 0.12% AH. However, having these analogous formulations can have a great potential with respect to the physical-chemical stability profile and the control of the technique. 7.- Test to determine the height and duration of wheal of a solution of mucosectomy and analogs in the ex vivo model.

The same methodology was followed as in example 6.

Essays in ex vivo model

The test to determine the height of the wheal at the gastrointestinal level was performed in pig colon, with extrapolar results in the gastrointestinal area and the rest of the internal mucous membranes of the organism. The colon was extracted from three pigs the day before the test and kept in a refrigerator, in Hanks' solution, until its use. The tests were performed at 37C.

The solutions to study were the following: • Solution 1: AH 0.003% + CMC 0.2%.

 • Solution 2: AH 0, 12% + CMC 0, 1%.

In addition to the solutions alone, the influence of these in combination with the powdered microparticles (MPP) was also studied. For this, combinations of each solution with MPP were prepared in the ratio 1: 3.

Once the solutions were prepared, 10 mL was injected into the submucosa of the colon with a 23G needle. The height of the wheal formed was measured with a ruler at different times (start, 1, 3, 7, 15, 30, 60, 90 and 120 minutes).

Solutions without microparticles In Figures 32 and 33, CMC + AH 1 corresponds to the solution composed of AH 0.003% + CMC 0.2% and CMC + AH 2 corresponds to the solution with 0.12% AH + 0.1% CMC .

In Table 1 and 2 shows the heights (in cm) of the different habons formed.

Solutions with microparticles

Figure 34, 35 shows the wheals formed by the solution with microparticles in powder (MPP) at time 0 and 90 min.

 Table 1: Height of wheal (cm) in pig colon (solutions WITHOUT microparticles). The results obtained are expressed as the average of the 3 replicas made for each solution.

 Table 2: Height of wheal (cm) in pig colon (solutions WITH microparticles). The results obtained are expressed as the average of the 3 replicas made for each solution.

Conclusions

The use of a solution of CON or WITHOUT microparticles (0.1% CMC + 0.12% AH) in colon does not differ from the results obtained in stomach. Therefore, we consider extrapolar our results to the gastrointestinal area and to the rest of the body's internal mucous membranes, skin and soft tissues, such as the urogenital tract (bladder, vagina, ...).

Contrary to what is expected by a person skilled in the art, with the MPP solution, a wheal higher or more consistent than that obtained by the solution of 0.1% CMC + 0.12% AH without MPP is not generated.

However, having these analogous formulations can have a great potential regarding the physico-chemical profile, stability, controlled release of active ingredients, mucoadhesion that may be of interest before or after the intervention, assuming an alternative with technical advantages of interest.

Claims

1. - A combined preparation comprising: a) a component A, which is a microparticle, and b) a component B, which is a composition which in turn comprises:  I) a polymer derived from water soluble cellulose at a concentration of 0.005% to 2%; Y  II) hyaluronic acid in a concentration of 0.0001% to 0.5%;  2. The combined preparation according to the preceding claim, wherein the microparticle (A) is dispersed in the composition (B), the microparticle (A) absorbs the composition (B) that remains embedded in the microparticle (A), or a combination of both. 3. The combined preparation according to any of claims 1-2, wherein the microparticle is made by an internal ionic gelation emulsification technique. 4. - The combined preparation according to any of claims 1-3, wherein the microparticle is obtained by a process comprising the preparation of an A / O emulsion in which the internal or aqueous phase is formed by a polymer or mixture of polymers, and the external or oil phase is composed of an oil in which a surfactant is dispersed. 5. The combined preparation according to claim 4, wherein the polymer of the microparticle is selected from the list consisting of: alginate, chitosan, exopolysaccharide of bacterial origin, cellulose or cellulose derivatives, pectin, poloxamer 127, Eudragit® S 100, L - 100, L 12.5, S 12.5, FS30D, RS, FS and NM, Shellac, PLGA, or any combination thereof. 6. - The combined preparation according to any of claims 4-5, wherein the polymer of the microparticle is alginate, preferably sodium alginate. 7. The combined preparation according to any of claims 4-6, wherein the surfactant of the oil phase of the microparticle is polysorbate 80. 8. The combined preparation according to any of claims 4-7, wherein the aqueous phase of the microparticle also comprises fluid extract or tincture (2-4mL) of ruscus, bilberry, horse chestnut or meliloto, or the hyaluronic colloidal solution and cellulose. 9. The combined preparation according to any of claims 4-8, wherein the oil of the oil phase of the microparticle is selected from olive oil, rosemary oil, argan oil, liquid paraffin, or any combination thereof. 10. - The combined preparation according to any of claims 4-9, wherein once formed the aqueous phase of the emulsion A / O from which the microparticle is formed, a source of calcium is added, in the form of insoluble salt , which is dispersed in the aqueous phase. 1. The combined preparation according to claim 10, wherein the calcium source is calcium carbonate (0.5-1%) 12. - The combined preparation according to any of claims 4-1 1, wherein the oil phase of the emulsion A / O from which the microparticle is formed is composed of a homogeneous solution of polysorbate 80 in oil, preferably in a proportion between 2 and 4%. 13. The combined preparation according to any of claims 4-12, wherein to form the oily external phase emulsion of the A / O emulsion from which the microparticle is formed, the two phases are subjected to mechanical agitation to a speed of 300 - 2500 rpm, more preferably between 500-2000 rpm for a time comprised between 5-40 minutes, more preferably for 10-30 minutes. 14. The combined preparation according to any of claims 4-13, wherein to the emulsion obtained in the preceding claim is added an acid medium, preferably acetic acid and more preferably acetic acid in a proportion of between 4 and 8% , to solubilize calcium ions and lead to the formation of pre-gelled particles. 15. The combined preparation according to any of claims 4-14, wherein the formation of the microparticle further comprises adding a solution of calcium chloride 5-10% (w / v) to the emulsion resulting from the preceding claim to produce the hardening of the polymer droplets under mechanical agitation, preferably at a speed between 300 and 2500 rpm, more preferably at a speed between 500 and 2000 rpm, for a time comprised between 5 and 40 minutes, more preferably for 10-30 minutes . 16. The combined preparation according to any of claims 4-15, wherein the mirroparticles obtained in the preceding claim are cleaned with bidistilled water using successive centrifugation cycles. 17. - The combined preparation according to any of claims 4-16, wherein the microparticles are matured in calcium solution for a time between for a time of between 1 to 15 minutes, preferably 2 to 10 minutes, and then dried to 25-37 ° C. 18. The combined preparation according to any of claims 1-17, further comprising another active ingredient, preferably vitamin K1 or phytomenadione. 19. - The combined preparation according to any of claims 1-18, wherein the viscosity of said composition is from 5 to 100 mPa * s, preferably from 5 to 50 mPa * s, more preferably from 10 to 40 mPa * s 20. The combined preparation according to any of claims 1-19, wherein the concentration of the water soluble cellulose derivative polymer of the composition (B) is 0.01% to 1%, more preferably 0.1% to 0.5%. 21. - The combined preparation according to any of claims 1-20, wherein the hyaluronic acid concentration of the composition (B) is from 0.0001% to 0.3%, preferably from 0.001% to 0.1%. 22. The combined preparation according to any of claims 1-21, wherein component B is a composition which in turn comprises a. a cellulose-derived polymer soluble in water in a concentration of 0.005% to 0.4%; Y  b. hyaluronic acid in a concentration of 0.0001% to 0.09%. 23. The combined preparation according to any of claims 1-22, wherein the polymer derived from the water soluble cellulose of the composition (B) is selected from methylcellulose, hydroxypropylcellulose, carboxymethylcellulose, and hydroxypropylmethylcellulose, and combinations thereof. 24. - The combined preparation according to any of claims 1-23, wherein the hyaluronic acid of the composition (B) has a molecular weight of less than 1,000 kDa, preferably less than 800 kDa, more preferably from 500 to 800 kDa. 25. - The combined preparation according to any of claims 1-24, wherein the hyaluronic acid of the composition (B) has a viscosity in 1% aqueous solution of 100 to 300 mPa * s, preferably 150 to 250 mPa * s . 26. The combined preparation according to any of claims 1-25, wherein the hyaluronic acid of the composition (B) is the sodium salt of the hyaluronic acid. 27. The combined preparation according to any of claims 1-26, wherein the concentration of the hyaluronic acid of the composition (B) is from 0.001% to 0.012%, preferably from 0.002% to 0.008%, more preferably from 0.003% to 0.006% %. 28. The combined preparation according to any of claims 1-27, wherein the concentration of the water-soluble cellulose derivative polymer of the composition (B) is from 0.1% to 0.4%, more preferably from about 0, 2% to approximately 0.3%. 29. The combined preparation according to any of claims 1-28, wherein the water-soluble cellulose derivative polymer of the composition (B) is carboxymethylcellulose, preferably sodium carboxymethylcellulose. 30. The combined preparation according to any of claims 1-29, wherein said carboxymethylcellulose of the composition (B) has a molecular weight of more than 500 kDa, preferably of more than 800 kDa, preferably of about 900 kDa. 31. - The combined preparation according to any of claims 1-30, wherein said carboxymethylcellulose of the composition (B) has a viscosity of 500 to 4500 mPa * s in a 1% aqueous solution, preferably from 1,000 to 3,000 mPa * s , more preferably from 1,500 to 2,500 mPa * s, even more preferably from 2,200 mPa * s to 2,300 mPa * s. 32. The combined preparation according to any of claims 1-31, wherein the sodium carboxymethyl cellulose of the composition (B) has a viscosity greater than 1,000 mPa * s in a 1% aqueous solution, and is in said composition at a 0.2% to 0.3% concentration 33. - The combined preparation according to any of claims 1-32, wherein the composition (B) further comprises an agent regulating the osmotic pressure and / or the pH in a concentration comprised between 1% and 30%, preferably selected from a sugar, a polyalcohol and a salt, or any of their combinations. 34. The combined preparation according to claim 33, wherein the composition regulating agent (B) is in a concentration of 10% to 20%, preferably 16 to 18%, more preferably 17.5%.  35. - The combined preparation according to any of claims 33-34, wherein the regulatory agent of the composition (B) is a sugar, preferably fructose or inulin.  36. - The combined preparation according to any of claims 1-35, wherein the composition (B) is an aqueous composition, preferably comprising physiological serum or glucosaline serum.  37. - The combined preparation according to any of claims 1-36, wherein the composition (B) has a pH of 3 to 8, preferably 4 to 7, and more preferably 5 to 6.  38. - The combined preparation according to any of claims 1-37, wherein the composition (B) has an osmolarity of 500 to 3,000 mOs / L, preferably 1,500 to 2,500 mOs / L, more preferably about 1,700 mOs / L. 39. - The combined preparation according to any of claims 1-38, wherein the composition (B) further comprises a dye, preferably indigo carmine or methylene blue.  40. - The combined preparation according to any of claims 1-39, wherein the composition (B) further comprises one or more active ingredients.  41. - The combined preparation according to claim 40, wherein said active ingredient is a vasoconstrictor agent, preferably epinephrine, more preferably where the epinephrine concentration is 0.000025% to 0.5%.  42. - The combined preparation according to any of claims 1-41, wherein the composition (B) comprises:  - Hyaluronic acid, in a concentration of 0.0001% to 0.09%  - Sodium carboxymethylcellulose, in a concentration of 0.005% to 0.4%  - Fructose or inulin c.s.p. pH from 5 to 6  - Physiological serum c.s.p,  - optionally, epinephrine in a concentration of 0.0001% to 0.01%  optionally, a dye (from 0.01 to 0.1 ml, preferably 0.05 ml, from a solution of 0.1% to 5%).  43. - The combined preparation according to any of claims 1-42, wherein the composition (B) consists of:  - Hyaluronic acid, in a concentration of 0.0001% to 0.09%  - Sodium carboxymethylcellulose, in a concentration of 0.005% to 0.4%  - Fructose or inulin c.s.p. pH from 5 to 6  - Physiological serum c.s.p,  - optionally, epinephrine in a concentration of 0.0001% to 0.01%  optionally, a dye (from 0.01 to 0.1 ml, preferably 0.05 ml, from a solution of 0.1% to 5%).  44. - The combined preparation according to claim 42, wherein the composition (B) comprises:  - Hyaluronic acid, at a concentration of 0.003%  - Sodium carboxymethylcellulose in a concentration of 0.2% to 0.3%  - Fructose or inulin c.s.p. pH from 5 to 6  - Physiological serum c.s.p.  - optionally, epinephrine at a concentration of 0.005%  optionally, a dye (from 0.01 to 0.1 ml, preferably 0.05 ml, from a solution of 0.1% to 5%). 45.- The combined preparation according to claim 43, wherein the composition (B) consists of:  - Hyaluronic acid, at a concentration of 0.003% - Sodium carboxymethylcellulose in a concentration of 0.2% to 0.3% - Fructose or inulin csp pH from 5 to 6  - Physiological serum c.s.p.  - optionally, epinephrine at a concentration of 0.005%  optionally, a dye (from 0.01 to 0.1 ml, preferably 0.05 ml, from a solution of 0.1% to 5%).  46. - The combined preparation according to any of claims 1-45, for use as a medicament. 47. - The combined preparation according to any of claims 1-46, wherein the composition (B) comprises: a. a cellulose-derived polymer soluble in water in a concentration of 0.005% to 0.4%; and b. hyaluronic acid in a concentration of 0.0001% to 0.3%; for use in a method of treatment that requires the separation of different tissue layers, wherein said composition is preferably administered by injection, more preferably by endoscopic injection, in one of the tissues to be separated or in a tissue located therebetween. 48. - The combined preparation according to any of claims 1-47, for use in a treatment method that requires tissue separation, wherein said composition is preferably administered by injection, preferably by endoscopic injection, into one of the tissues at separate or in a tissue located between both. 49. - The combined preparation according to any of claims 1-48, in a method for the treatment of mucosal lesions wherein said method comprises the resection of a portion of the mucosa, preferably of the mucosa of the gastrointestinal tract or of the urogenital tract , more preferably from the mucosa of the gastrointestinal tract. 50. The combined preparation according to claim 49, which is administered by submucosal endoscopic injection. 51. - The combination preparation according to any of claims 47-50, wherein said method of treatment is an endoscopic mucosal resection, preferably selected from the group consisting of i) an endoscopic submucosal resection (EMR) and ii) an endoscopic dissection submucosa (EDS), more preferably it is EMR. 52. - The combined preparation according to any of claims 47-51, wherein the injection pressure, preferably when using an endoscopic injection needle of a diameter 23G is from 0.1 to 12 kgf, more preferably from 0.25 to 10 kgf, even more preferably from 0.5 to 10 kgf, particularly preferably from 1.0 to 7 kgf. 53. The combined preparation according to any of claims 47-52, wherein the retention time of the composition (B) is at least 45 minutes, preferably 60 or more minutes. 54. The combined preparation according to any of claims 47-53, wherein the average height of the protrusion in the retention time is at least 3 mm, preferably 4 mm or more. 55. - The combined preparation according to any of claims 1-54, wherein the ratio of component A (microparticles) to component B (composition comprising a polymer derived from water soluble cellulose and hyaluronic acid) is 0.5: 5 56. - The combined preparation according to any of claims 1-54, wherein the ratio of component A (microparticles) to component B (composition comprising a polymer derived from water soluble cellulose and hyaluronic acid) is about 1: 3 . 57.- A kit comprising a container containing the combined preparation according to any of claims 1-54 and, optionally, instructions for the use thereof in a treatment method according to any of claims 46 or 47-54. . 58.- The kit according to the preceding claim for use in the treatment of lesions in the mucosa of the gastrointestinal tract, preferably in the treatment of tumors and / or polyps.