KR101610203B1 - Medical Device Set for Optical Fiber Laser Ablation and Photodynamic Therapy, and Cancer Treatment Therapy Using the Same - Google Patents

Abstract

The present invention relates to a combination of laser ablation and photodynamic therapy using methylene blue nanoparticles to provide a combination of optical fiber laser ablation and optical therapy that can replace conventional radiotherapy with sophisticated equipment, A set of medical devices for epidural therapy, and a method of treating cancer using the same.

Description

[0001] The present invention relates to a medical device set for optical fiber laser ablation and photodynamic therapy, and a cancer treatment using the same. [0001] The present invention relates to an optical fiber laser ablation and photodynamic therapy,

The present invention relates to a medical device set for optical fiber laser ablation and photodynamic therapy and a method for treating cancer using the same.

Lasers have been attracting much attention and attention in the medical field because of their ability to accurately target small and dense areas as concentrated beams with very high luminance and superior directionality compared to other light sources. The medical approach of lasers can be roughly divided into two methods, one is by using thermal energy by laser irradiation and the other is by using photochemical effect.

The photochemical method is to apply a photo-activating action that changes a biomolecule or biological tissue by injecting a light-sensitive agent capable of absorbing the laser light into a living body.

A method using thermal energy is a method of irradiating a laser beam to a living body and absorbing a part thereof to be converted into heat and utilizing a thermal destruction effect generated at this time. Such a treatment is called laser ablation or laser ablation. This method is applied to the incision and removal of the affected part by destroying the living tissue by irradiating the laser with a very small area, and is applied to the coagulation of tissues and blood, the removal of abnormal blood vessels and the treatment of skin diseases.

When the laser ablation treatment is applied to surgery, the blood is instantaneously coagulated or vaporized, resulting in a hemostatic effect and a non-contact type so that there is no infection problem. Particularly, in areas where it is difficult to approach such as lungs, bladder, blood vessels, and vertebrae, laser light is transmitted through an optical fiber to destroy tumors and abnormal tissues. When the functions such as oral cavity and larynx are to be conserved, Has been performed. This laser surgery is incised only in a very small area, so it is lighter in inflammation, less pain and scarring than the operation using a conventional surgical knife, and is actively used for shortening the recovery and hospitalization period.

Generally, laser ablation is used alone to remove cancer or abnormal cells in clinical practice, but it is also applied in combination with other therapies such as radiation therapy. Radiation therapy after laser resection is intended to suppress or completely eliminate the proliferative response of residual tumor cells.

However, in the case of radiation therapy, it is more likely that the patient will suffer from chronic fatigue, anorexia, nausea, vomiting, abdominal cramps, diarrhea, hair loss, anemia, hematopoietic dysfunction, dyspnea, cough, Pain, and other side effects such as dryness, redness, swelling, itching, tissue fibrosis, dental caries, and radiation osteonecrosis of the irradiated area. In addition, since radiation therapy does not have cancer cell targeting ability, there is a problem that it involves side effects such as radiation necrosis of normal cells, exfoliation and dryness of peripheral normal tissues, and skin atrophy.

Numerous references are referenced throughout the specification and are cited therein. The disclosure of the cited document is incorporated herein by reference in its entirety to more clearly describe the state of the art to which the present invention pertains and the content of the present invention.

Korean Patent No. 10-140634 Korea Patent Publication No. 10-2011-0138003

The present inventors noticed that the conventional laser resection alone can not completely remove the tumor tissue and, in order to remove residual tumor tissue, expensive equipment is required when accompanied with radiation therapy, and serious systemic and local side effects are necessarily accompanied Therefore, we have tried to provide a new cancer treatment which can replace the existing treatment. As a result, the combination of treatment with laser ablation and photodynamic therapy with methylene blue nanoparticles proved synergistic effects in cancer treatment and could effectively replace conventional surgery and radiation therapy , Thereby completing the present invention.

Accordingly, an object of the present invention is to provide a medical device set for cancer treatment for optical fiber laser ablation and photodynamic therapy, and a method for treating cancer using the same.

It is another object of the present invention to provide a pharmaceutical composition and formulation for removing residual tumor cells after laser ablation.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

One aspect of the present invention is a set of medical devices for treating cancer comprising (i) an optical fiber probe for laser ablation and (ii) methylene blue nanoparticles, wherein the methylene blue nanoparticles comprise a methylene blue- And a polymer. The present invention also provides a medical device set comprising the same.

Optical fiber for laser ablation Probe

Laser optical fiber probes can be categorized into optical fiber probes for optical activation and fiber optic probes for laser ablation, depending on the purpose of use. Optical fiber probes for optical activation do not damage surrounding tissues, but merely transmit a small amount of light Refers to an optical fiber probe for use in activating a photosensitizer, and the optical fiber probe for laser ablation means an optical fiber probe for use in destroying tumors and abnormal tissues by transmitting laser light through an optical fiber .

The optical fiber probe included in the medical device set of the present invention is a laser ablation probe for destroying and removing a target tissue. The probe is a laser ablation technique that is deeply placed in the human body and transmits a laser beam to a region difficult to approach, It can be used without limitation as long as it is used.

According to an embodiment of the present invention, the optical fiber probe for laser ablation includes broadly all of UV, visible light and IR, and has a wavelength of 500 to 1,500 nm (a wavelength frequently used in an actual laser therapy apparatus) And emits a laser.

According to another embodiment of the present invention, the laser ablation optical fiber probe emits a laser having an output of 1 W to 200 W, and may include a CW and a pulsed laser.

In one embodiment, the medical device set of the present invention may further comprise a guide needle for inserting the optical fiber probe into the tissue, wherein the optical fiber probe is inserted into the tissue through the needle, To the inside.

In the case of an optical fiber probe for optical activation aiming at only activating a photosensitizer, it is used for non-invasively insertion into a space (esophagus, urethra, etc.) in the human body to transmit light to the inner surface of the space, A resection fiber optic probe may be used in a minimally invasive manner by inserting an optical fiber into the tissue (inserting an optical fiber into the needle after stabbing the needle) and delivering light into the tissue.

In this case, the optical fiber probe for laser ablation according to the present invention is inserted into a tissue and then used for laser ablation by transmitting light of 700 to 1500 nm, and then transmits light of 500 to 700 nm for use in photodynamic therapy The function of ablation and light activation may be performed at the same time.

In a preferred embodiment of the present invention, the bundle-type optical fiber probe disclosed in Korean Patent Publication No. 10-1406434 can be used as the laser cut-off optical fiber probe, A front irradiating unit having a planar end portion of the front illuminating optical fiber; And a side irradiating unit that is disposed on the outer periphery of the front irradiating unit and has a sloped surface at an end of the side irradiating optical fiber to reflect the laser beam to the side surface, And the bundle-type optical fiber probe is a bundle-type optical fiber probe.

The bundle-type optical fiber probe includes a front irradiation optical fiber and a side irradiation optical fiber bundled to provide a bundle-type optical fiber probe which can be irradiated in various directions using one optical fiber probe. In the bundled optical fiber probe, FIG. 1A is a perspective view of the optical fiber probe, and FIG. 1B is a view of the laser beam passing through a bundle type optical fiber probe according to an embodiment.

Here, the optical fiber is a fiber made by elongating a transparent dielectric such as quartz glass or plastic elongatedly and having a diameter of 0.1 to 1 mm. The optical fiber is covered with a medium having a high refractive index center and a periphery covered with a medium having a low refractive index. Specifically, the central core portion and the surrounding cladding portion have a double cylindrical shape, May be coated with one or two layers of synthetic resin coating to protect it from impact.

In another preferred embodiment, an optical fiber probe having a ring lens structure disclosed in Korean Patent Application No. 10-2013-0145377 can be used as the optical fiber probe for laser ablation.

The optical fiber probe of the ring lens structure comprises: a main body; A center portion formed at an end of the main body; And a tip of a ring lens structure including a lens portion including a curved surface surrounding the center portion.

The optical fiber probe of the ring lens structure has a ring-shaped lens having a center portion hollowed out by simple processing at the end of the optical fiber without a separate mechanism, and a ring-type lens tip capable of focusing at a desired position or freely adjusting a beam profile FIGS. 2A and 2C are perspective views of an optical fiber probe having a ring lens structure according to an embodiment. FIGS. 2B and 2C are diagrams showing how a laser passes through an optical fiber probe of a ring lens structure according to an embodiment. 2d.

The lens portion of the optical fiber probe of the ring lens structure may have a curved surface or a concave surface. In the case of a convex surface, the curvature R may be at least 1/2 of a width W of the lens portion, , The curvature R may be a relationship of R <-W / 2 with the width W of the lens portion.

The shape of the center of the optical fiber probe of the ring lens structure may be a cone, a cylinder, a cylinder with a convex end, or a cylinder with a concave end, and preferably a cone shape having a central angle of 40 degrees or more and 100 degrees or less .

Methylene blue  Nanoparticle

The methylene blue nanoparticles contained in the medical device set of the present invention are composed only of materials whose composition is in clinical use or derived from human body. Methylene blue is a substance used when performing a dye endoscopy to apply or inject a dye so as to facilitate observation of lesions that are difficult to observe or invisible. For example, its location can easily be confirmed for a surveillance lymph node biopsy And is a substance currently in clinical use as part of a test method using color micro-fluid during gastrointestinal endoscopy.

Methylene blue is a positively charged dye which is very difficult to be introduced into cells by itself. However, among the methylene blue nanoparticles of the present invention, a stable methylene blue / fatty acid complex is formed with fatty acids by electrostatic force and hydrophobic interaction on the molecular structure , Thus indicating a high solubility in solvents.

Fatty acid (fatty acid) contained in the methylene blue nanoparticles of the present invention means a carboxylic acid or a salt thereof. In one embodiment, the fatty acid may be a long chain saturated or unsaturated monocarboxylic acid, preferably oleic acid Or a salt thereof.

The amphiphilic polymer contained in the methylene blue nanoparticle of the present invention is a compound having a hydrophilic moiety and a hydrophobic moiety at the same time, and any polymer capable of forming a micelle structure by gathering molecules can be used without limitation. When a polymeric micelle is formed in a hydrophilic solvent, the hydrophobic portion of the molecule is gathered at the center to form nuclei, and the hydrophilic portion forms an outer portion in contact with water. In water-soluble solvents, hydrophobic substances, such as oil, are located in the inner part of the micelle and stabilize and dissolve in the solvent.

In one embodiment of the present invention, the amphiphilic polymer includes an anionic amphiphilic polymer having a hydrophilic group such as a carboxylate, a sulfonate or phosphate structure, a cationic amphiphilic polymer such as a quaternary ammonium salt containing a hydrophilic group, An amphiphilic polymer having both an anionic site and a cationic site in the molecule, and an amphiphilic polymer having a hydrophilic moiety that is non-ionic, that is, a hydrophilic moiety that does not ionize into a hydrophilic moiety, and preferably, a polyoxyethylene-polyoxypropylene block copolymer Can be used.

In a preferred embodiment, the methylene blue-fatty acid complex is encapsulated in an amphiphilic polymeric micelle in an aqueous environment, thereby allowing the nanoparticles of the present invention to self-assemble.

In one embodiment, the methylene blue nanoparticles of the present invention are prepared by mixing a methylene blue and a fatty acid to form a methylene blue-fatty acid mixture, lyophilizing the mixture to form a complex, and mixing the complex thus obtained with the amphiphilic polymer But is not necessarily limited thereto.

3 is a schematic diagram illustrating the formation of water-dispersed self-assembled nanoparticles containing methylene blue as a photosensitizer for use in the present invention, according to an embodiment.

Another aspect of the present invention is a pharmaceutical composition for removal of residual tumor cells after laser ablation comprising methylene blue nanoparticles, wherein the methylene blue nanoparticles comprise a methylene blue-fatty acid complex and an amphiphilic polymer And to provide a pharmaceutical composition comprising the same.

Yet another aspect of the present invention is a topical dosage form for removal of residual tumor cells after laser ablation, comprising methylene blue nanoparticles, wherein said methylene blue nanoparticles comprise a methylene blue-fatty acid complex and an amphipathic polymer And the like.

Another aspect of the present invention is to provide a medical device kit for treating cancer comprising (i) an optical fiber probe for laser ablation and (ii) the above-described pharmaceutical composition or preparation.

The pharmaceutical composition or topical administration of the present invention may contain a pharmaceutically acceptable carrier. The pharmaceutically acceptable carriers to be contained in the pharmaceutical composition of the present invention are those conventionally used in the present invention and include lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia rubber, calcium phosphate, alginate, gelatin, But are not limited to, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrups, methylcellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. It is not. The pharmaceutical composition of the present invention may further contain a lubricant, a wetting agent, a sweetening agent, a flavoring agent, an emulsifying agent, a suspending agent, a preservative, etc. in addition to the above components. Suitable pharmaceutically acceptable carriers and formulations include, but are not limited to, Remington's Pharmaceutical Sciences (19th ed., 1995).

The medical treatment kit and the pharmaceutical composition or preparation of the present invention can be used in combination with laser resection and photodynamic therapy using methylene blue nanoparticles in order to prevent the radiation therapy that requires conventional surgery and expensive equipment, In addition to being able to replace therapy, it also produces synergistic effects in cancer treatment in a non-surgical way.

FIG. 1A is a perspective view of a bundle type optical fiber probe, which is an embodiment of an optical fiber probe that can be included in the medical treatment apparatus for treating cancer according to the present invention, FIG. 1B is a cross section of a bundle type optical fiber probe, .
FIGS. 2A and 2C are perspective views of an optical fiber probe of a ring lens structure according to an embodiment of an optical fiber probe that can be included in the medical treatment apparatus for treating cancer of the present invention, FIGS. 2B and 2D are perspective views of a ring lens structure And the laser passes through the optical fiber probe.
FIG. 3 is a schematic view showing the formation of aqueous dispersion self-assembled nanoparticles containing methylene blue as a photosensitizer to be included in the medical treatment apparatus for cancer treatment of the present invention, according to an embodiment of the present invention.
Figure 4 is a graph showing the absorbance (a) and the fluorescence spectrophotometer (Hitachi F-7000, wavelength calibrated for excitation and emission) measured with a UV-visible spectrometer (Agilent 8453) And a fluorescence wavelength spectrum (b).
5 is a photograph (b) of the nanoparticles prepared in Example 1, which was observed by Dynamic Light Scattering (a) and a transmission electron microscope (TEM, CM30, FEI / Philips, 200 kV) .
6 is a photograph showing the tissue penetration ability of MB NPs and MB Sol. According to (1) of Example 2. Fig.
FIG. 7 is a photograph showing the imaging diagnosis behavior of MB NPs in an early cancer model by the local cancer target mapping according to (2) of Example 2. FIG.
FIG. 8 is a schematic diagram showing a process of laser ablation using a laser ablation optical fiber and photodynamic treatment using MB NPs in Example 3; FIG.
Fig. 9 shows the results obtained by carrying out the experiment of Example 3. Fig.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention more specifically and that the scope of the present invention is not limited by these embodiments.

Example

Example  1: In the water environment As photosensitizers  Methylene Blue  Self contained Assembly Formation of furnace particle

(1) Hydrogenation of methylene blue / sodium oleate by electrostatic force

20 mg of methylene blue (MB, purchased from Aldrich) and 30 mg of sodium oleate (purchased from Aldrich) were heated in 100 mL of tetrahydrofuran (THF, Daejeon Chemical) at 60 to 90 ° C for 1 to 5 minutes, .

Methylene blue formed a sodium methyl oleate and a stable methylene blue / sodium oleate complex (MBO) by electrostatic force and hydrophobic interaction on the molecular structure, showing high solubility in solvents.

The mixture obtained by dissolving methylene blue in this way was freeze-dried after removing excess sodium oleate and other impurities using a sieve filter (5 μm) to obtain MBO.

(2) Preparation and evaluation of nanoparticles containing MBO and amphiphilic polymer

0.2 mg of the MBO obtained in the above (1) and 20 mg of the ampholytic polymer Pluronic ^® F-68 (purchased from Aldrich) were thoroughly mixed using a solvent THF and the solvent was completely removed. The solvent-removed mixture was evenly dispersed in 2 mL of water to prepare methylene blue nanoparticles (MB NPs).

The absorbance and fluorescence of methylene blue (MB Sol.) Dissolved in water and methylene blue nanoparticles (MB NPs) dispersed in water were measured. The results are shown in FIGS. 4A and 4B, respectively. 4A and 4B, it was confirmed that the absorption wavelength of the MB dissolved in water and the fluorescence wavelength were shifted to a short wavelength region due to the formation of nanoparticles.

The size of the nanoparticles was measured using a Zetasisernano ZS (Malvern Instruments, UK), and the shape of the nanoparticles was observed with a transmission electron microscope (TEM, CM30, PEI / Philips, 200 kV) Respectively. As a result, it was confirmed from FIGS. 5A and 5B that the nanoparticles were spherical particles having a diameter of 80 to 100 nm.

Example  2: MB NPs Accumulation and characterization of cancer target after biopsy injection

(1) Evaluation of tissue permeability of MB NPs and MB Sol.

(MBC) and MB Sol prepared in Example 1 were each applied in 0.5 mL increments, and then, in a water bath, 37 &lt; RTI ID = 0.0 &gt; &Lt; / RTI &gt; Then, the sample was sufficiently washed with DPBS, and the fluorescence image was obtained with a 12-bit CCD camera (Kodak Image Station 4000 MM, ex: 625 nm / em: 700 nm).

It was confirmed that the methylene blue formed nanoparticles embedded in the amphiphilic polymer and the tissue permeability was remarkably improved as compared with that of the single methylene blue.

(2) Evaluation of Local Cancer Markability of MB NPs Using Cancer Model

Sixty μL of MDA-MB-231 cells (1 × 10 ⁷⁾ were dispersed in the medium and injected into the left hip muscle area of a female rat (Balb / c-nu, 5.5 weeks old, Orient Bio). After 4 to 5 weeks, the tumor growth was visually confirmed, and 60 μL of the nanoparticles (MB NPs) containing methylene blue were locally injected into the cancer tissue site and the fluorescence imaging apparatus (IVIS-Spectrum, Perkin-Elmer, USA) To confirm the accumulation of cancer tissue target. For the control experiment of the experiment, methylene blue aqueous solution (MB Sol.) Was prepared in the same manner as in the above experiment and shown in FIGS. 7a and 7b.

In the above experiments, the ability of MB NPs to accumulate tumor tissue was also confirmed in animal models, which confirmed the cancer targeting effect in vivo.

(3) Accumulation of MB NPs and evaluation of photocatalytic properties using early cancer model

In order to carry out the accumulation experiment of nanoparticles in the early cancer model, 1 × 10 ⁶ SCC7 (1 × 10 ⁶ ) dispersed in 60 μL of cell culture was injected into the muscles of the left and right buttocks of male rats (Balb / c-nu, 5.5 weeks old, Orient Bio) (Squamous cell carcinoma) cells were subcutaneously injected. After 2 hours, 60 μL of MB NPs were subcutaneously injected to the site where the left cancer cells were injected. After 2 hours, the material was sufficiently absorbed into the cells, and the laser was irradiated to the left hind for 20 minutes. After the injection of 100 μL of Annexin V / FITC (60 μL) and FITC (60 μL), a fluorescence imaging system (IVIS-Spectrum, Perkin-Elmer, USA) We evaluated the cytotoxicity of MB NPs by phototoxicity in early cancer models.

As a result, it was confirmed that methylene blue as a photosensitizer was hydrophobicized to induce apoptosis using MB NPs introduced into the amphipathic polymer. As a result, it was confirmed that the effect of photodynamic therapy in vivo was demonstrated.

Example  3: With laser ablation MB NPs Confirmation of synergy effect by combination of

(1) Removal of tumor tissue using fiber-optic laser ablation

In the same animal disease model used in Example 2 (2), a guide needle was placed at the center of the cancer tissue and a guiding beam was irradiated to visually confirm that the multifaceted optical fiber was positioned at the center of the tumor. (NT-3, output wavelength: 980 nm ± 20%, Daino Co., Ltd., Korea) for 20 seconds to 20 seconds. The pattern of the process is shown in Fig. Thereafter, the thermal denaturation of the cancer tissue was confirmed by visual observation, and the change in the tumor size (= short axis ² X long axis) was measured and observed for 6 times, once every 2 to 3 days, and is shown in FIGS. 9A and 9B.

(2) Removal of tumor tissue using MB NPs photodynamic therapy

After one day of laser ablation as described above, 60 μL of MB NPs were subcutaneously injected into the cancer tissue site 1 hour later, photodynamic therapy (irradiation of 655 nm laser for 10 minutes) Photodynamic therapy was performed in the same manner over a total of 6 times, and tumor size changes were observed and measured for 15 days.

For comparative experiments, photodynamic therapy was also performed in the same manner on the cancer tissue sites of the same animal disease model made in Examples 2 (2) and 3 (1).

The results of observation of the tumor for 15 days and the change in tumor size are shown in FIGS. 9A and 9B.

As a result, the tumor size was increased with time in the control group, and when the laser resection was performed alone, the residual tumor cells grew again with time. In addition, when the photomynamic therapy using MB NPs alone was performed, the increase in tumor size was prevented, but no significant decrease in the initial tumor size was observed until 15 days. On the other hand, the combination of laser resection and photodynamic therapy using MB NPs showed that synergistic effects were obtained by effectively removing tumors as compared with the case of performing alone.

This is because the photodynamic treatment using methylene blue nanoparticles can be effectively used instead of the conventional radiotherapy after the laser ablation, and as a result, the present invention requires conventional surgery and expensive equipment, accompanies serious systemic and local side effects And to provide a new cancer treatment that can replace radiation therapy.

Claims (15)

(i) an optical fiber probe for laser ablation; (ii) an injection needle for inserting the optical fiber probe into the tissue; And (iii) a methylene blue nanoparticle,
Wherein the optical fiber probe is inserted into the tissue through the inside of the needle to deliver light to the inside of the tissue, and the laser ablation optical fiber probe emits a laser having an output of 1 to 5 W for 10 to 20 seconds, Wherein the particles comprise a methylene blue-oleic acid or salt complex thereof and a polyoxyethylene-polyoxypropylene block copolymer.
delete delete The medical instrument set according to claim 1, wherein the laser ablation optical fiber probe emits a laser having a wavelength of 500 to 1500 nm.
delete The laser cut-off optical fiber probe according to claim 1,
A front irradiating unit having a front irradiating optical fiber positioned at the center, the end of the front irradiating optical fiber being formed in a plane; And
A side irradiating optical fiber is disposed on the outer periphery of the front irradiating unit and an end face of the side irradiating optical fiber is formed with a sloped surface to reflect the laser beam to the side,
Wherein the front irradiation unit and the side irradiation unit are bundled optical fiber probes formed in a bundle.
The laser cut-off optical fiber probe according to claim 1,
main body;
A center portion formed at an end of the main body; And
A lens portion including a curved surface surrounding the center portion,
Wherein the optical fiber probe is a fiber optic probe having a tip of a ring lens structure including a tip of a ring lens structure.
The medical instrument set according to claim 7, wherein the curved surface of the lens portion is a convex surface or a concave surface.
The medical device set according to claim 1, wherein the methylene blue-fatty acid complex is encapsulated in an amphiphilic polymer micelle.
The set of medical devices according to claim 1, wherein the methylene blue nanoparticles are self-assembled in an aqueous environment.
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