RU2796427C1 - Method of preparing an antiglaucomatous drainage with antiproliferative properties - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to medicine, in particular to ophthalmology, and is intended to reduce intensity of postoperative scarring in glaucoma surgery. Method of preparing an antiglaucomatous drainage with antiproliferative properties includes electroforming a solution from polyhydroxycarboxylic acids mixture, laying the resulting microfibrous material with fiber diameter of 0.2-20 microns on a drum collector and subsequently cutting the tape into separate drainage in the form of 15-25 mm long, 3-5 mm wide and 150-300 microns thick strings. At the same time, acetonitrile is used as a solvent for preparing the polyhydroxycarboxylic acid mixture, 250-500 mcg of everolimus per drainage is added to the polyhydroxycarboxylic acid solution, after electroforming the resulting drainage is incubated in cyclosporine A solution with concentration of 1.6 mg/ml for 30 minutes, as a result, an additional 3-3.5 mcg of cyclosporine A, which is the everolimus synergist, is adsorbed on the drainage surface and in the drainage structure.

EFFECT: invention provides a long-term improvement in the glaucoma surgical treatment effectiveness and minimizes the likelihood of complications development.

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Description

The invention relates to medicine, namely to ophthalmology, and is intended to reduce the intensity of postoperative scarring in glaucoma surgery.

A microinvasive method of surgical treatment of open-angle glaucoma using drainage based on highly porous xenocollagen, which is proposed to be saturated with the Superlymph cytokine complex in order to prevent postoperative scarring, is known (RF patent No. ).

This drainage has a long period of biodegradation (for several years) [Anisimova S.Yu., Anisimov S.I., Larionov E.V. Glaucoma Surgery XXI. - M.: April, 2012. - 170s], which can lead to excessive proliferation of connective tissue elements, as well as to drainage calcification. The release of the "Superlymph" cytokine complex by drainage occurs within 1-2 days, while the most active phase of inflammation in the surgical wound, including proliferation, occurs within 1 month after the injury. Other disadvantages of the method include the fact that the saturation of the drainage with a complex of cytokines occurs for a long time - for 5 days, and the need to sterilize the drainage by autoclaving can reduce the functional activity of the cytokines used.

The closest adopted for the prototype is a method for the prevention of postoperative scarring in the surgical treatment of glaucoma, including implantation during surgery for glaucoma of a porous bioresorbable drainage based on polyhydroxycarboxylic acids, which is pre-saturated with everolimus, while the drainage is placed in a 2% suspension of crushed everolimus powder in physiological saline and are exposed to ultrasonic waves with a power of 630 W at a frequency of 22 kHz for 6 minutes in an operating room ex tempore or in the process of making a drain before its sterilization (RF patent No. glaucoma V. N. Germanova and others)

The disadvantage of this method is the predominant accumulation of the antiproliferative agent on the surface of the drainage material and inside the pores, which leads to an increase in the rate and time limitation of the release of the antiproliferative agent from the drainage (up to 13 days). At the same time, a "depot" with high concentrations of the drug is created directly at the site of drainage implantation, which leads to apoptosis of cells inside the drainage, an increase in the intensity of the inflammatory reaction in the early stages after surgery, and further formation of a capsule around the drainage. However, the advantage of the method is that, due to the extremely low solubility of everolimus in water, the concentration of the drug that penetrates into the surrounding tissues, not directly in contact with the drainage, does not exceed therapeutic values and effectively prevents scarring in the postoperative period [Germanova V.N., Sarbaeva N.N., Karlova E.V., Volova L.T., Nefedova I.F., Radaikina M.V. Early cellular response of eye tissues to the implantation of bioresorbable drains saturated with immunosuppressants with a selective mechanism of action. Morphological records. - 2020. - No. 3(28). - S. - 15-20].

Thus, there is a need to slow the release of everolimus from the drainage material and avoid direct contact of the immunosuppressant with surrounding tissues.

The objective of the invention is to create a method for manufacturing antiglaucomatous bioresorbable drainage with antiproliferative properties.

The technical result is a long-term increase in the effectiveness of surgical treatment of glaucoma while minimizing the likelihood of complications.

The technical result is achieved by adding everolimus powder in the amount of 250-500 μg per 1 drain to the initial solution of polyhydroxycarboxylic acids, in which acetonitrile is used as a solvent; the preparation of a drainage blank is carried out using electrospinning, during which microfibers with a diameter of 0.2-20 μm are formed from the initial solution of polyhydroxycarboxylic acids with everolimus, and their laying is carried out on a drum collector; to obtain separate drainages, a polymer “tape” is cut; after drying, the drains are placed in a solution of cyclosporine A with an active substance content of 1.6 mg/ml for 30 minutes, as a result of which an additional 3-3.5 μg of cyclosporine A, which is a synergist of everolimus, is adsorbed on the surface and in the structure of the drains. The finished drainage has the form of a strip 15-25 mm long, 3-5 mm wide, 150-300 microns thick.

According to experimental data, drainage ensures the release of cyclosporin A within 3.4±0.5 days and everolimus within 58.3±2.9 days.

As polyhydroxycarboxylic acids, polylactide, polyglycolide, polycaprolactone in various proportions can be used.

The developed method makes it possible to obtain antiglaucomatous bioresorbable drains with antiproliferative properties by including the inhibitor of the mechanistic target rapamycin everolimus in the polymer composition and additional subsequent saturation of the drain with cyclosporine A.

Everolimus has an antiproliferative effect both on immunocompetent cells and directly on human Tenon's capsule fibroblasts. To the greatest extent, the antiproliferative effect of everolimus is expressed within concentrations of 0.5-10.0 μg/ml, at which the rate of proliferation of human Tenon's capsule fibroblasts slows down by 1.7-7.4 times without a cytotoxic effect, which was proved in an experiment on culture cells.

Cyclosporin A is a synergist of everolimus, has an anti-inflammatory effect by inhibiting the synthesis of interleukin-2 by T-lymphocytes, and also has a direct antiproliferative effect on human Tenon's capsule fibroblasts, most pronounced in the concentration range of 0.05-2.0 μg / ml. At these concentrations, cell proliferation slows down by 1.5-5.4 times without a cytotoxic effect, according to in vitro experiment data. Prolonged delivery of CsA to the intervention area at the earliest possible time after surgery contributes to a decrease in the severity of the inflammatory response in response to the intervention and an additional decrease in the intensity of fibroblast proliferation.

The inclusion of everolimus directly into the fiber structure during drainage electrospinning provides an important advantage: under these conditions, everolimus desorption occurs slowly and gradually, in the process of drainage biodegradation, with the flow of intraocular fluid, while the tissues surrounding the drainage do not directly contact everolimus. Due to this release of everolimus, the drainage does not undergo encapsulation, which contributes to the long-term preservation of the aqueous humor outflow pathways. At the same time, the duration of everolimus release (58.3±2.9) days is sufficient for stable functioning of the filtration pad.

The microfiber structure provides an increase in the rigidity and elasticity of the drainage, which facilitates the process of its implantation in comparison with porous models. The distance between the fibers is optimal and sufficient for the outflow of intraocular fluid through the drainage structure.

The size and shape of the drainage allow the drainage to be used for both penetrating and non-penetrating interventions for glaucoma. Drainage implantation is possible with a long axis perpendicular to the limbus between superficial and deep scleral flaps with positioning of the distal end directly under the conjunctiva in the zone of formation of the filtration cushion, which is the optimal technique for performing surgical intervention for glaucoma and contributes to the most effective prevention of sclero-scleral and sclero-conjunctival adhesions with a minimum likelihood of complications associated with drainage, namely, its eruption and displacement.

Thus, the proposed method for manufacturing antiglaucoma drainage gives it the following advantages:

- prolongs the desorption of the selective immunosuppressant (up to 58.3 ± 2.9 days), providing a long-term antiproliferative effect;

- reduces the likelihood of drainage encapsulation;

- does not have a cytotoxic effect;

- has an anti-inflammatory effect due to additional saturation of CsA;

- has an optimal period of biodegradation, excluding drainage calcification;

- has a size and shape that is convenient for implantation in both penetrating and non-penetrating glaucoma surgery;

- easy to manipulate and implant due to increased rigidity and elasticity.

In practice, the method is carried out as follows.

In the initial polymer solution, in which acetonitrile is used as a solvent, everolimus powder (95%), Sigma-Aldrich is added in the amount of 250-500 μg per 1 drain. The production of a drainage blank is carried out using an electrospinning machine. A 6.0% polymer solution with everolimus is placed in the perfusor. With the help of a stepper motor, a dosed supply of the polymer is carried out. A high voltage is generated by a high voltage power supply. Under the action of electrostatic forces, an initial stream of polymer is created, which is then split into separate microfibers under the influence of the hydrodynamic moment and the resistance force of the medium. At the same time, acetonitrile completely evaporates due to its high volatility. The laying (winding) of fibers takes place on a drum collector. To obtain individual drains, a polymer "tape" removed from the collector is cut. As a result, bioresorbable drainages based on polyhydroxycarboxylic acids with a microfibrous structure (fiber diameter 0.2-20 μm, distance between fibers 20-100 μm) are obtained, having the form of strips 15-25 mm long, 3-5 mm wide, 150-300 mm thick. microns containing 250-500 micrograms of everolimus. After drying, the drains are placed in a solution of cyclosporine A, Sandoz with an active substance content of 1.6 mg/ml for 30 minutes, as a result of which an additional 3-3.5 μg of cyclosporine A, which is a synergist of everolimus, is adsorbed on the surface and in the structure of the drains.

Clinical examples:

Example 1

The drainage is made according to the specified method from a mixture of polylactide and polycapronolactone in a ratio of 60:40, has a length of 20 mm, a width of 4 mm, a thickness of 200 microns, contains 300 micrograms of everolimus and 3.5 micrograms of cyclosporine A.

Implanted with a long axis perpendicular to the limbus in the right eye of a Soviet chinchilla rabbit during penetrating hypotensive surgery. The distal end of the drain is located under the superficial scleral flap, the proximal end is between the sclera and Tenon's capsule above the scleral flap.

IOP before surgery 18 mm Hg. Art.

The next day after surgery: mild general hyperemia, well-functioning filtration cushion (AF) with H3E3V2S0 characteristics according to the IBAGS morphological classification of AF. The cornea is transparent, there is no edema. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

IOP 8 mm Hg. Art.

1 week after surgery: no hyperemia. Drainage is visualized. FP: H2E3V2S0. There are no signs of inflammation. IOP 8 mm Hg. The cornea is transparent. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

1 month after surgery: the eye is calm. Drainage is visualized. FP: H2E3V2S0. IOP 9 mm Hg. The cornea is transparent. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

Histological examination (staining with hematoxylin, pikrofuksin): drainage is resorbed, in the area of drainage implantation there is a small amount of mononuclear cells, fibroblasts, single SCIT, single vessels. The capsule is missing. Filter cavities are visualized. Surrounding tissues are intact.

Example 2

The drainage is made according to the specified method from a mixture of polylactide and polycaprolactone in a ratio of 70:30, has a length of 25 mm, a width of 4 mm, a thickness of 150 microns, contains 350 micrograms of everolimus and 3.5 micrograms of cyclosporine A.

Implanted with a long axis perpendicular to the limbus in the right eye of a Soviet chinchilla rabbit during penetrating hypotensive surgery. The distal end of the drain is located under the superficial scleral flap, the proximal end is between the sclera and Tenon's capsule above the scleral flap.

IOP before surgery 21 mm Hg. Art.

The next day after surgery: moderate hyperemia, AF: H2E3V2S0. The cornea is transparent, there is no edema. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

IOP 10 mm Hg. Art.

1 week after surgery: no hyperemia. Drainage is visualized. FP: H2E3V2S0. There are no signs of inflammation. IOP 11 mm Hg. The cornea is transparent. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

1 month after surgery: the eye is calm. Drainage is visualized. FP: H2E3V2S0. IOP 11 mm Hg. The cornea is transparent. The anterior chamber is of medium depth, the moisture is transparent. The lens is transparent. The optic disc is pale pink, the boundaries are clear. On the fundus without focal pathology.

Histological examination (staining: hematoxylin, picrofuchsin): inside the drainage material there is a small amount of mononuclear cells, fibroblasts, the capsule is not expressed. Above the drain is a wide filtering cavity. Surrounding drainage tissues are intact.

Staining: hematoxylin, picrosirius red: thin collagen fibers were revealed inside the drainage.

Claims (1)

A method for manufacturing antiglaucomatous drainage with antiproliferative properties, including electrospinning a solution from a mixture of polyhydroxycarboxylic acids with laying the resulting microfiber material with a fiber diameter of 0.2-20 μm on a drum collector and then cutting the resulting tape into separate drainages in the form of strips 15-25 mm long, wide 3-5 mm, 150-300 µm thick, characterized in that acetonitrile is used as a solvent for preparing a mixture of polyhydroxycarboxylic acids, everolimus is added to the resulting solution of polyhydroxycarboxylic acids in an amount of 250-500 µg per one drain, after electrospinning, the resulting drain is kept in solution cyclosporine A at a concentration of 1.6 mg/ml for 30 minutes, as a result of which an additional 3-3.5 μg of cyclosporine A, which is a synergist of everolimus, is adsorbed on the surface and in the structure of the drains.