JP6545926B2 - Composition for treating hand-foot syndrome - Google Patents

Description

  The present invention relates to a composition for treating or preventing hand-foot syndrome.

  Hand-foot syndrome is a skin disease that develops due to the administration of a chemotherapeutic drug such as an anticancer drug. Common symptoms appear in a limited area of the palms and soles of the foot and are accompanied by pain, rash, erythema, often from tingling sensory abnormalities. If you do not cope with the problem, it will become severe, and it will be difficult to continue the treatment with the causative agent as the patient's QOL declines, and it will often cause problems in the original treatment.

  As treatment for hand-foot syndrome, rest of the hand; urea ointment for moisturizing purpose; topical treatment with external preparations such as heparin analogue preparations or vitamin-containing ointment; prodnisolone or dexamethasone as systemic therapy, or pyridoxine hydrochloride (vitamin B6 And the like (non-patent document 1) and the like have been proposed, but these are all coping treatments and do not aim to cure hand-foot syndrome.

  Until now, as the onset mechanism of hand-foot syndrome, one of the causes is inhibition of proliferation ability of skin basal cells, leakage of anticancer drug from blood vessel (patent document 1), secretion of anticancer drug from eccrine sweat gland (non-patent document 2) It is also suggested that the causal relationship between the anticancer drug and its onset is unknown.

  Many anticancer agents cause DNA damage and induce apoptosis of cancer cells. It has been reported that cancer apoptosis is induced by reactive oxygen species (hereinafter also referred to as "ROS") generated directly or indirectly by anticancer drugs (Non-patent Documents 3 to 8). Furthermore, oxidative damage due to ROS was amplified in the presence of copper (II) ions in experiments using human whole myelocytic leukemia cell line HL-60 cells and HL-60 derived catalase overexpressing HP 100 cells It reports (nonpatent literature 6 and 9).

U.S. Patent No. 6060083

Investigational New Drugs, Vol. 8, p 57-63, 1990 Annals of Oncology 16, 1210-1211, 2005 Hiraoka W et al. , (1998) J. Clin. Invest. 102, 1961-1968 Tada-Oikawa S et al. , (1999) FEBS Lett. , 442, 65-69 Varbiro G et al. , (2001) Free Radic. Biol. Med. , 31, 548-558. Murata M et al. , (2004) Free Radic Biol. Med. , 37, 793-802. Sadzuka Y et al. , (2005) YAKUGAKU ZASSHI, 125, 149-157 Gewirtz. D. A. , (1999) Biochemi. Pharmacol. , 57, 727-741. Murata M et al. , (2004) Free Radic Biol. Med. , 37, 793-802.

  As described above, despite the need for treatment or prevention of hand-foot syndrome that is serious in terms of continuation of the causative drug and improvement of the patient's QOL, the onset mechanism is unknown and there is no effective treatment available It is in. Then, an object of the present invention is to provide a novel composition effective for treatment or prevention of hand-foot syndrome.

  As a result of intensive studies to solve the above-mentioned problems, the present inventors are closely involved in the development of inflammatory symptoms of hand-foot syndrome, as ROS generated upon administration of a chemotherapeutic agent such as an anticancer drug. The present invention has been completed.

  Accordingly, the present invention includes the following features.

  (1) A composition for treating or preventing hand-foot syndrome, which comprises a substance that captures or removes active oxygen as an active ingredient.

  (2) Substances that capture or remove active oxygen include superoxide dismutase, bilirubin, glutathione peroxidase, peroxidase, catalase, ascorbic acid or derivatives thereof, cysteine, glutathione, linoleic acid, tocopherol or derivatives thereof, α-carotene, β- The composition according to the above (1), which is at least one selected from carotenes, flavonoids, uric acid, thioredoxin, or phytochemicals.

  (3) The composition according to the above (1), wherein the substance that captures or removes active oxygen is one that targets superoxide anion.

  (4) The composition according to the above (1), which comprises at least superoxide dismutase as a substance which scavenges or removes active oxygen.

  (5) The composition according to any one of (1) to (4), wherein the hand-foot syndrome is caused by administration of a liposomal formulation of doxorubicin.

  (6) The composition in any one of said (1)-(5) which is a skin external preparation.

  (7) The composition according to (6) above, which is a skin external preparation.

  (6) The composition in any one of said (1)-(5) which is a skin external preparation.

  (7) The composition according to (6) above, which is a skin external preparation.

  (8) Use of a substance that scavenges or removes active oxygen for producing a therapeutic or prophylactic agent for hand-foot syndrome.

  (9) Substances that capture or remove active oxygen are superoxide dismutase, bilirubin, glutathione peroxidase, peroxidase, catalase, ascorbic acid or derivatives thereof, cysteine, glutathione, linoleic acid, tocopherol or derivatives thereof, α-carotene, β- The use according to the above (8), which is at least one selected from carotenes, flavonoids, uric acid, thioredoxin, or phytochemicals.

  (10) The use according to the above (8), wherein the substance that captures or removes active oxygen is one that targets superoxide anion.

  (11) The use according to the above (8), wherein the substance which scavenges or removes active oxygen is superoxide dismutase.

  (12) The use according to any one of the above (8) to (11), wherein the hand-foot syndrome is caused due to the administration of a liposome-ized preparation.

  According to the present invention, a novel composition effective for the treatment or prevention of hand-foot syndrome is provided.

The photograph figure of a rat hind paw sole is shown after the high dose (10 mg / kg) or the low dose (5 mg / kg) PEGL-DOX solution is repeated (it performs 3 times once every three days). It is a photograph figure which shows the result of H & E staining (a), picrosirius red staining (b) and TUNEL staining (c) in the skin tissue collected from the lesion site of the hand-foot syndrome onset rat. The measurement result of the cytokine and the chemokine which were implemented in the skin tissue extract | collected from the lesion site of the hand-foot syndrome onset rat is shown. It is the result of the analysis using each cultured cell of HaCaT and NHDF. (A) and (b) are the results of in vitro tests on chemokines and inflammatory cytokines for which an increase in production was confirmed in tissues collected from lesion sites of hand-foot syndrome onset rats, respectively. It is a result of the in vitro toxicity test of DOX which used each culture cell of HaCaT and NHDF. FIG. 5 is a diagram showing rescue from DOX cytotoxicity by superoxide dismutase (SOD) using HaCaT cultured cells. The H & E stained image of the forelimb shell skin in the hand-foot syndrome onset rat evoked by intradermal injection of doxorubicin (DOX) is shown. ac, saline administration group; df, 1% SOD administration group. It is a histogram which shows surface thickening of the forelimb shell skin of the physiological saline treatment group and the 1% SOD treatment group in the hand-foot syndrome onset rat evoked by intradermal injection of doxorubicin (DOX).

  The present invention relates to a composition for treating or preventing hand-foot syndrome (hereinafter also referred to as “the composition of the present invention”). In the present invention, the hand-foot syndrome to be treated or prevented is a skin condition (numbness, skin hypersensitivity) that develops at the extremity end such as the fingertips, palms and soles due to the administration of a chemotherapeutic drug (especially an anticancer drug) , Pigmentation, redness, heat, erythema, blisters, swelling, keratinization, cracking, nail deformation, pigmentation, etc., but not limited thereto). Hand-foot syndrome is also known as palmar and plantar redness perceptual syndrome, edge erythema, chemotherapeutic drug-induced erythema with chemotherapeutic agent, palm and sole erythema, and hand-foot skin reaction. The term includes all of these.

  In addition, fluorinated pyrimidine antitumor agents (eg, capecitabine) used for the treatment of colorectal cancer and breast cancer as chemotherapeutic agents that may cause onset of hand-foot syndrome; anthracyclines used for malignant solid tumors; recurrence -Modified liposomal doxorubicin preparation (PEGL-DOX) used for advanced ovarian cancer; docetaxel used for a wide range of cancers; molecular targeted drugs used for renal cancer such as sorafenib and sunitinib Although a large number of drugs are known, the hand-foot syndrome in the present invention is not limited by the drug causing the onset, but is directed to any drug as a cause. In one embodiment, the hand-foot syndrome is a hand-foot syndrome resulting from the administration of a liposomalized formulation of doxorubicin.

  The present invention is based on the finding that the symptoms of hand-foot syndrome resulting from the administration of a chemotherapeutic drug such as an anticancer drug involve the ROS generated by the interaction with the chemotherapeutic drug. Therefore, the composition of the present invention is characterized by containing a substance that captures or removes active oxygen as an active ingredient.

The ROS, generally, superoxide anion generated by the one-electron reduction of oxygen molecules ^{(O 2-),} hydrogen peroxide _{(H 2 O} 2), hydroxyl radical (· OH), and singlet oxygen ⁽¹ _{O 2} In the present invention, the active ingredients can be selected or combined so as to target at least one, two, three, and all four in the present invention. Thus, "capturing substance or eliminate active oxygen (ROS)" in the present invention, superoxide anion ^{(O 2-),} hydrogen peroxide _{(H 2 O} 2), hydroxyl radical (· OH), and singlet A substance (eg, an antioxidant) that targets at least one of oxygen ( ¹ O ₂ ) for capture or removal.

  Substances that scavenge or remove active oxygen can capture or remove any one of superoxide anion, hydrogen peroxide, hydroxy radical and singlet oxygen, and so long as they do not adversely affect the human body. It may be derived or synthetic. In one embodiment of the present invention, the substance that captures or removes active oxygen is a substance that targets superoxide anion for capture or removal.

  Substances capable of capturing or removing the superoxide anion include, but are not limited to, superoxide dismutase, bilirubin, ascorbic acid or derivatives thereof (6-ascorbyl stearate, ascorbyl 6-palmitate, 2, 6-dipalmitate ascorbyl, 2,3,5,6-tetrahexyldecanoate ascorbyl, (ascorbyl / tocopheryl) potassium phosphate, ascorbic acid-2-sodium sulfate, sodium ascorbate, ascorbic acid-2-phosphate magnesium Ascorbic acid-2-sodium phosphate, ascorbyl-2-glucoside and the like are known, and the composition of the present invention can contain at least one of them as an active ingredient.

  Substances capable of capturing or removing hydrogen peroxide include, but are not limited to, glutathione peroxidase, peroxidase, catalase, ascorbic acid or derivatives thereof (6-ascorbyl stearate, ascorbyl 6-palmitate, 2 , 6-dipalmitate ascorbyl, 2,3,5,6-tetrahexyl decanoic acid ascorbyl, (ascorbyl / tocopheryl) potassium phosphate, ascorbic acid-2-sodium sulfate, ascorbic acid sodium, ascorbic acid-2-phosphate Magnesium, sodium ascorbate-2-phosphate, ascorbyl-2-glucoside, etc.), vitamin E succinate etc. are known, and the composition of the present invention can contain at least one of them as an active ingredient.

  Substances capable of capturing or removing hydroxy radicals include, but are not limited to, cysteine, glutathione, linoleic acid, tocopherol or derivatives thereof (α-tocopherol, tocopherol acetate, vitamin E succinate etc.), Alpha-carotene, beta-carotene, flavonoids, uric acid, thioredoxin and the like are known, and the composition of the present invention can contain at least one of these as an active ingredient.

  Substances capable of capturing or removing singlet oxygen include, but are not limited to, ascorbic acid and its derivatives (6-ascorbyl 6-stearate, ascorbyl 6-palmitate, ascorbyl 2,6-dipalmitate) Ascorbyl 2,3,5,6-tetrahexyldecanoate, (ascorbyl / tocopheryl) potassium phosphate, ascorbic acid-2-sodium sulfate, sodium ascorbate, sodium ascorbate-2-magnesium ascorbate, ascorbic acid-2- Sodium phosphate, ascorbyl-2-glucoside, etc.), tocopherol and derivatives thereof (α-tocopherol, tocopherol acetate, etc.), β-carotene, riboflavin, uric acid, thioredoxin, etc. are known, and the composition of the present invention At least Can also contain 1 type as an active ingredient.

  In addition to the above, the composition of the present invention is a substance which is known to have high antioxidant activity as an active ingredient, such as α-lipoic acid, coenzyme Q10, xanthophylls (astaxanthin, lutein, zeaxanthin, canthaxanthin, fucoxanthin, Anteraxanthin, violaxanthin, etc., carotenoids (α-carotene, β-carotene, lycopene, etc.), polyphenols (chlorogenic acid, ellagic acid, lignans, sesamin, curcumin, coumarin, oleocantal, oleuropein, resveratrol etc.), butylhydroxyanisole Butylhydroxytoluene, sodium erythorbate, propyl gallate, ferulic acid, caffeic acid, 5- (diethylphosphono) -5-methyl-1-pyrroline N-oxide, flavonoids (flavanones, flavo , Chalcone, catechin, epigallocatechin gallate, flavanonol, aurone, flavan-34-diol (leucoanthocyan), isoflavone, anthocyanin, tannin, rutin etc., plant extract (liquor leaf extract, mulberry tea extract, agetsu extract, Rosemary extract, etc.), manganese N, N′-bis (salicylidiene) ethylenediamine chloride, (acetato-κO) [[2,2- [1,2′-ethanediylbis [(nitrilo-κN) methylidyne]] bis [3, 5-dimethoxyphenolato-κO]] (2-)]-manganese, 2,6-bis (1,1-dimethylethyl) -4-[[(1-ethyl) amino] methyl] phenol hydrochloride, 3-methyl- 1-phenyl- -Pyrazolone, SOD mimetic compound (manganese (III) tetrakis (4-benzoic acid) porphyrin chloride, manganese (III) tetrakis (1-methyl-2-pyridyl) porphyrin, manganese (III) meso-tetrakis (N-methyl-2) -Pyridyl) porphyrin pentachloride, etc.), 7-hydroxyflavone-manganese complex, nordihydroguaiaretic acid, 21- (4- (2, 6 Di-1- pyrrolidinyl) -4-pyrimidinyl) -1-piperazinyl)- pregna-1,4,9 (11) -triene-3,20-dione, (Z) -2-Butenedionate, (−)-2-((4- (2,6-Di-1-pyrrolidinyl-4-) pyrimidinyl) -1-pip Razinyl) methyl) -3,4-dihydro-2,3,7,8-tetramethyl-2H-1-benzopyran-6-ol, 2 HCl, phytochemical (anthocyanin, isoflavone, sesaminol, curcumin, sulforahan, methyl cysteine sulfoxide) Allicin, lutein, lycopene, limonene, phytosterol, β-glucan, saponin, capsaicin, gingerol and the like).

  In the present invention, the composition of the present invention can be prepared by selecting one or more types of substances capable of capturing or removing the above-mentioned ROS regardless of the type of ROS to be targeted.

  In the present invention, the administration route and dosage form of the composition are not particularly limited as long as the therapeutic or preventive effect of hand-foot syndrome is exerted, but administration of a chemotherapeutic agent already known to cause onset of hand-foot syndrome It is preferable not to impair the effect of the chemotherapeutic agent in the case of application to a subject who is scheduled for or a subject who continues to administer the chemotherapeutic agent that caused the onset of the hand-foot syndrome. That is, as described above, cancer apoptosis is reported to be induced by active oxygen (hereinafter also referred to as "ROS") generated directly or indirectly by an anticancer agent (Hiraoka W et al., ( 1998) supra; Tada-Oikawa S et al., (1999) supra; Varbiro G et al., (2001) supra; Murata M et al., (2004) supra; Sadzuka Y et al., (2005) supra; Gewirtz.D.A., (1999) supra), in view of such an intrinsic role by anti-cancer agents, because systemic removal of ROS can significantly reduce the efficacy of chemotherapeutic agents.

  Therefore, the composition of the present invention can, in one embodiment, be in the form of a dosage form that locally delivers a substance that captures or removes ROS, for example, a skin external preparation, to a lesion site of hand-foot syndrome. Such a skin external preparation can be provided, for example, as a skin external medicine.

  In the skin external preparation of the present invention, the compounding amount of the substance that captures or removes ROS is particularly limited as long as it can effectively improve the symptoms of hand-foot syndrome and does not cause other harmful side effects. For example, it can be 0.001 to 30 (w / w)%, preferably 0.01 to 10 (w / w)%, for example 5 (w / w)%.

  In the skin external preparation of the present invention, in addition to the active ingredient, components generally used for skin external preparations, such as, but not limited to, water, surfactants (anionic surfactants, cationic interface Activator, amphoteric surfactant or nonionic surfactant), liquid or solid oil, wax, hydrocarbon oil, higher fatty acid, higher alcohol, ester oil, silicone oil, silicone oil, thickener, film agent, UV absorber, metal Ion blocking agents, lower alcohols, sugars, amino acids, organic amines, polymer emulsions, pH adjusters, skin nutrients, antioxidants, perfumes, etc. are appropriately blended and manufactured according to the desired product form according to the conventional method. be able to.

  The form of the external preparation for skin of the present invention is not particularly limited, and can be provided in the form of an ointment, a gel, a cream, a solution, a lotion, an aerosol, a patch, a cosmetic sheet and the like.

  The external preparation for skin of the present invention may be used alone or in combination with other agents used for treatment of hand-foot syndrome. Such agents include, for example, moisturizers, antibiotics, external steroids, oral steroids, non-steroidal anti-inflammatory analgesics and the like.

  The skin external preparation of the present invention can be used prophylactically for patients who develop hand-foot syndrome due to the administration of a chemotherapeutic agent, as well as for cancer patients who are scheduled to receive a chemotherapeutic agent. Good.

  Hereinafter, the present invention will be described in more detail using examples, but the present invention is not limited to these examples.

1. Preparation of a hand-foot syndrome onset model The frequency of hand-foot syndrome onset depending on the type of drug is as follows: PEGL-DOX (maximum 78%); capecitabine (51-78%); sorafenib (approximately 55) %) (Ministry of Health, Labor and Welfare, 2010). Therefore, in order to create a hand-foot syndrome onset model, it was decided to adopt PEGL-DOX, which has the highest incidence frequency.

(1) Preparation of PEGL-DOX (Doxorubicin PEGylated Liposome Preparation) Previously reported (Sadzuka Y., 2005, except that separation by a filter for unifying particle size and confirmation of average particle size after preparation were not performed. The PEGylated liposome preparation was prepared according to the above.

  That is, L-α-distearoylphosphatidyl-DL-glycerol (PEG-modified lecithin; DSPG) and lecithin (each manufactured by NOF Corporation) are dissolved in a solvent in which methanol and chloroform are mixed 1: 4, and the mixture is heated in a 35 ° C. water bath. The solvent was removed by a rotary evaporator in the above to form a thin film on the surface of the eggplant flask. An aqueous solution of DOX (20% aqueous solution; manufactured by Nippon Kayaku Co., Ltd.) and a sorbitol / lactic acid solution were added to the flask, and the mixture was stirred for 15 minutes in a 60 ° C water bath. Thereafter, ultrasonic irradiation was performed for 3 minutes to obtain a 0.2 w / v% PEGL-DOX solution.

(2) Preparation of hand-foot syndrome model (2-1) Intravenous tail model of PEGL-DOX A high dose (10 mg / kg) or low dose (5 mg / kg) PEGL-DOX solution was purchased from Japan SLC Ltd. 7-week-old female SD rats were given single or multiple doses (three doses once a day) (intravenous tail).

  In the single dose group, flares appeared in the forearm palm, sole of the foot, auricle, and nose immediately after PEGL-DOX administration, but the flares occurred transiently and disappeared in 5 to 10 minutes. On the other hand, in the repeated administration group of PEGL-DOX, inflammation at the periphery of the extremity was observed at a low dose of 5 mg / kg, and the tendency was remarkable at a high dose (10 mg / kg) (FIG. 1).

  Moreover, in the repeated administration group, although the hand and foot were observed to develop, the effect on the whole body skin was not remarkable and only a slight dryness was observed. This closely resembles the symptoms observed when administering Doxil® to humans (Gewirtz. D.A., 1999, supra). Therefore, it was determined that administration of PEGL-DOX developed hand-foot syndrome in the forearm palm and hind foot sole.

(2-2) Local injection model of doxorubicin PEGL-DOX tail intravenous injection requires relatively high concentration of PEGL-DOX to cause severe hand-foot syndrome, so symptoms of hand-foot syndrome There was a problem that the animal died before the onset of Therefore, we attempted to cause hand-foot syndrome by intradermal administration of doxorubicin to the sole of the rat.
When 40 μl of doxorubicin saline solution prepared to 0.08% was intradermally administered to the forelimbs and hindlimbs of 8-week-old female SD rats purchased from Japan SLC Co., Ltd. In addition, inflammatory redness, swelling, exfoliation, etc. were observed on the instep, which closely resembled the symptoms of hand-foot syndrome.
Therefore, this model was used as a second model of hand-foot syndrome onset.

2. Pathological analysis of hand-foot syndrome model rat In order to perform pathological analysis of hand-foot syndrome onset site, skin tissue collected from a high dose (10 mg / kg) PEGL-DOX repetitively-administered rat where hand-foot syndrome developed according to the above (2-1) It was evaluated histologically. Histological evaluation was performed using H & E staining, picrosirius red staining and TUNEL staining, respectively, by comparing with the non-administration control group.

(1) Hematoxylin-Eosin (H & E) Staining Formalin-fixed and paraffin-embedded skin tissue was sectioned to a thickness of 4 μm. After deparaffinization, rehydration and H & E staining were performed, and the histological images of the forelimb palm and hind foot sole were observed with a light microscope.

(2) Picrosirius Red Stain In order to observe the state of collagenous fibers in the dermis in detail, picrosirius red was stained and observed with a polarization microscope.

(3) TUNEL staining TUNEL staining was performed to detect apoptosis. And TUNEL staining using the DeadEnd ^TM Fluorometric TUNEL system kit (product of Promega Corporation), was observed with a fluorescence microscope.

  The results are shown in FIG. H & E staining of the sole skin of high dose PEGL-DOX repeated administration rats reduced or lost the granular layer and decreased the number of cells from the basal layer to the spinous layer, compared with the non-administered control group Showed a rough arrangement, and thinning of the epidermal layer was remarkable (Fig. 2a). On the other hand, fibroblasts in the dermal layer were almost equivalent to the normal state.

  FIG. 2 b shows the results of picrosirius red staining (staining of collagen fibers). There are many collagen fiber bundles that are thick enough to be observed in red and high brightness, and as the brightness is low from yellow to green, disordered or broken state of the array is exhibited. As is clear from FIG. 2 b, in the PEGL-DOX repeated administration rats, the arrangement of collagen fibers in the dermal layer was disturbed, and fiber breakage was remarkable.

  FIG. 2 c shows the results of TUNEL staining. Green color indicates cells in which apoptosis is induced. In the PEGL-DOX administration model, apoptosis of epidermal basal cells was induced.

  The above results indicate that at the site of hand-foot syndrome onset, thinning along with cell loss in the epidermal layer, collagen destruction in the dermal layer, and apoptosis of epidermal basal cells are observed, resulting in inflammatory symptoms. It is thought that

3. Analysis of hand-foot syndrome onset cause (1) in vivo cytokine analysis In order to elucidate the cause of onset of hand-foot syndrome, skin tissue of hind limb after PEGL-DOX administration according to the above (2-1) is collected and expressed Cytokines and chemokines were measured. The collected skin tissue was homogenized and centrifuged to collect the supernatant, and detection was performed using Rat Cytokine Antibody Array (manufactured by RayBiotech).

  The results are shown in FIG. The expression of the chemokines CINC3 and Fractalkine and the production of IL-10 which works to suppress the production of the IL family are remarkable, and at the same time, the expression upregulation of inflammatory cytokines IL-1β and IL-6 was also confirmed.

(2) In Vitro Cytokine Analysis In order to confirm the effect of DOX on cells in skin tissue, further in vitro experiments were performed. In order to compare the effects on the epidermal layer and the dermal layer, HaCaT cells (a human keratinocyte-derived cell line; received from Kobe University School of Medicine) and NHDF (a Kurashiki Spinning Co., Ltd.) that is normal human dermal fibroblasts are respectively compared. Using. Specifically, each culture cell is added with DOX (1.5 μM) and cultured for 24 hours, and then cytokines in the medium are added to Luminex 200 system (Millipore) (FIG. 4a) or Human Cytokine Antibody Array (RayBiotech) Detection (FIG. 4 b). The cytokines to be detected were the chemokines IL-8, GRO (all correspond to rat CINC3) and Fractalkine, which were enhanced in vivo production, and the inflammatory cytokines IL-1 and IL-6.

  In addition, in order to evaluate the contribution of reactive oxygen species (ROS) to the amount of produced cytokines, the same experiment is performed also in a system in which copper (II) chloride (manufactured by Wako Pure Chemical Industries, Ltd.) is additionally added as a copper (II) ion to the medium. Did.

  The results are shown in FIG. In HaCaT, the presence of 1.5 μM DOX alone did not increase the production of the three chemokines (Figure 4a). On the other hand, in the presence of DOX and copper (II) ion (50 and 375 μM), the production amount of each chemokine increased, and GRO and IL-8 increased as the Cu (II) ion concentration increased. On the other hand, in NHDF, there was almost no change due to DOX addition or coexistence of Cu (II) ion, and rather, it was observed that the production amount of various chemokines was suppressed in the presence of DOX.

  Also for inflammatory cytokines, the effect of DOX and / or copper (II) ions was altered by cell type (Figure 4b). HaCaT cells increased the production of IL-1α and IL-6 in the presence of DOX, which production was further enhanced by the addition of Cu (II) ions in the presence of DOX. The production of IL-1β was elevated under coexistence of high concentration of copper (II) ion and DOX. In contrast, NHDF cells did not show a worthy response in the case of DOX alone but stimulated increased production of IL-1β in the presence of copper (II) ions.

  As described above, DOX enhances the production of chemokines IL-8, GRO and Fractalkine, inflammatory cytokines IL-1β, IL-6 and IL-1α in epidermal cells, and enhances the production of IL-1β in dermal cells , And this production enhancement was shown to be recognized in a copper (II) ion dependent manner. These results suggest that ROS whose production is enhanced in the coexistence of DOX and copper (II) ion may be involved in enhanced production of these cytokines.

(3) DOX toxicity evaluation test For the toxicity evaluation of DOX, a toxicity evaluation test using cultured cells HaCaT and NHDF (described above) was performed. That is, 1.5 μM DOX was added to the medium, and the proportion of viable cells after 24 hours was measured using Cell Counting kit-8 (CCK8) (manufactured by DOJINDO LABORATORIES). In the toxicity test in the presence of copper (II) ions, 50 μM or 375 μM copper chloride (manufactured by Wako Pure Chemical Industries, Ltd.) was added to a medium containing 1.5 μM DOX.

  In addition, 1.5 μM DOX, 50 μM copper (II) chloride and 100 μg / ml superoxide dismutase (SOD) (Sigma Aldrich) were added to the HaCaT culture medium to investigate the involvement of DOX in skin tissue and ROS. The survival rate after 12 hours was measured.

  When DOX was not added, the viability of HaCaT cells was reduced by about 20% when 50 μM copper (II) chloride was added, but was similar when high concentrations of 375 μM were added. In the presence of DOX, cell viability decreased rapidly with increasing copper (II) ion concentration (FIG. 5). This suggests that the promotion of ROS production due to the coexistence of DOX and copper (II) ion reduced the cell survival rate. On the other hand, in NHDF cells, no remarkable influence on the survival rate was observed by the combination of DOX and copper (II) ion. The difference in cell damage caused by ROS between HaCaT and NHDF was well correlated with the skin histology of a hand-foot syndrome model animal.

  Also, addition of SOD, which is known to capture ROS, improved the viability of HaCaT cells (Figure 6).

  From the above results, in the pathophysiology of hand-foot syndrome, ROS generated by the interaction between the anticancer drug and copper (II) ion present in the skin promotes the production of chemokines and inflammatory cytokines in epidermal cells, and as a result , Vasodilation, redness, heat sensation, vascular permeability has been enhanced, and it has been shown that the inflammatory symptoms of hand-foot syndrome develop.

  On the other hand, in the cytotoxicity test using HaCaT, addition of SOD showed improvement in cell viability, indicating that the component that captures or removes ROS is effective for treatment or prevention of hand-foot syndrome It was done.

4. Suppression test of onset of hand-foot syndrome In this test, a hand-foot syndrome onset animal model by local injection of (2-2) doxorubicin was used.
40 μl of 1% superoxide dismutase saline solution (SOD) was intradermally administered to 8-week-old female SD rat forelimbs and hind-paws of hindlimbs. A saline only administration group without SOD was also prepared as a negative control. Twenty-four hours after SOD administration, 0.08% doxorubicin was intradermally administered to the same site. Administration of SOD or saline and doxorubicin was only once each, and the onset of hand-foot syndrome and its course were observed for 17 days. On the final day of observation, skin of palm and shell of forelimb and hind limb was collected, formalin fixed, paraffin embedded, and tissue section was prepared. H & E staining was performed to observe the condition of limb skin. The thickness of the epidermis was measured by image processing software Axio Vision 4.8 (manufactured by Carl Zeiss), and comparisons between groups were statistically processed by Welch's t test.

  By administering 0.08% doxorubicin to the soles of rats, symptoms of hand-foot syndrome such as inflammatory redness, swelling, and exfoliation on the soles are observed to a different degree in the SOD administration group and the saline administration group. The 1% SOD administration group (n = 2, (forelimb n = 4, hind limb n = 4) as compared to the physiological saline administration group (n = 2, (forelimb n = 4, hind limb n = 4)) of the negative control 2.) it was observed that the degree of redness and swelling was rather weak and recovery to normal was quick. In the saline administration group, redness and swelling were also observed in the forelimb back. On the other hand, in the 1% SOD administration group, various symptoms in the forearm were significantly reduced.

  When observing the skin histology of the forelimb shell by H & E staining, inflammatory surface hyperplasia was observed in the negative control saline administration group (Fig. 7a to c). The 1% SOD administration group significantly suppressed inflammatory epidermal hyperplasia, and the degree was slight (Fig. 7d-f). When the thickness of the epidermis was measured at 60 to 80 stained tissue images, a significant inhibitory effect was observed in the 1% SOD-administered group (p <0.001) (FIG. 8).

  Thus, the occurrence of hand-foot syndrome or its symptoms could be effectively reduced by pre-administering an antioxidant capable of capturing or removing ROS. Thus, substances that capture or remove ROS have been shown to be useful in the treatment or prevention of hand-foot syndrome.

  According to the present invention, a composition useful for the treatment or prevention of hand-foot syndrome is provided. The composition of the present invention has industrial applicability in that it is provided as a skin external preparation for patients with hand-foot syndrome, such as a medicine for treatment of hand-foot syndrome.

Claims (6)

A composition for treating or preventing hand-foot syndrome, comprising a substance capable of capturing or removing reactive oxygen as an active ingredient,
Here , the composition, wherein the substance capable of capturing or removing active oxygen is at least one selected from glutathione peroxidase, peroxidase, catalase, uric acid, and thioredoxin . The composition according to claim 1 , wherein the hand-foot syndrome is caused by administration of a liposomal formulation of doxorubicin. The composition according to claim 1, which is a skin external preparation. The composition according to claim 3 , which is a skin external preparation. Use of a substance capable of capturing or removing active oxygen for producing a therapeutic or prophylactic agent for hand-foot syndrome,
Here, the use, wherein the substance capable of capturing or removing active oxygen is at least one selected from glutathione peroxidase, peroxidase, catalase, uric acid, and thioredoxin . The use according to claim 5 , characterized in that the hand-foot syndrome develops due to the administration of a liposomal formulation.