KR101847486B1 - Porphyrin derivative having hydrophilic group and hydrophobic group, and process for preparing the same - Google Patents

Abstract

The present invention provides a porphyrin derivative having a hydrophilic polyethylene glycol group and a hydrophobic alkyl group, and a process for producing the porphyrin derivative.
The porphyrin derivative of the present invention and its production method are excellent in an oxygen production efficiency and anticancer activity and can be usefully used as an anticancer agent or a photosensitizer for photodynamic diagnosis / treatment which requires a photosensitive composition.

Description

TECHNICAL FIELD The present invention relates to a porphyrin derivative having a hydrophilic group and a hydrophobic group and a process for preparing the porphyrin derivative,

More particularly, the present invention relates to a porphyrin derivative having a hydrophilic polyethylene glycol group and a hydrophobic alkane group, and a process for producing the porphyrin derivative. BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a porphyrin derivative having a hydrophilic group and a hydrophobic group,

Photodynamic therapy (PDT) is a therapy using light, photosensitizer, active oxygen in the body, a method of injecting a photosensitizer into the body and irradiating a laser light having a specific wavelength to a disease site to treat the disease . In 1904, von Tappeiner and Jadlbauer demonstrated the need for oxygen in the photoreceptor response, explaining it as oxygen-dependent light sensitization in 1907, and the terminology of photodynamic therapy was established and academic research began. And has been used for the treatment of various diseases such as cancer diagnosis and treatment, autologous bone marrow transplantation, antibiotic treatment, AIDS treatment, skin graft surgery and arthritis, and its application range is gradually increasing.

More specifically, the photodynamic therapy used in cancer treatment is a method in which light is irradiated to a photosensitizer material that absorbs light of a specific wavelength, thereby causing singlet oxygen (singlet oxygen) Or free radicals are produced, and such unicellular oxygen or free radicals are induced by inducing apoptosis of various lesion sites or cancer cells while preserving normal cells as they are.

Photodynamic therapy is selectively accumulated in cancer cells, and it can complement the existing cancer treatment methods such as surgical operation, radiation therapy, side effects of drug therapy, and aftereffects after cancer treatment, and can be performed by local anesthesia alone. Even an institution can preserve its function and form, and it does not leave tissue damage or scarring.

Examples of the photosensitizer include porphyrins, chlorins, bacteriochlorins, phthalocyanine, 5-aminolevulinic acid, porphycenes, etc. , There is a problem that it is difficult to form a formulation capable of parenteral administration due to the hydrophobic photosensitizer in the clinical application of the photodynamic therapy, so it is necessary to increase the water solubility. Therefore, most of the photosensitizers currently used in photodynamic cancer treatments have low solubility when administered in vivo, inadequate interaction with biomolecules, mutual entanglement between photosensitizers, And low photoreactivity. Thus, the effect of treatment is low.

Accordingly, development of a photosensitizer which is more rapidly transmitted to a cancerous lesion in the living body and has excellent photosensitization activity is recognized as an important problem.

Korean Patent Publication No. 10-2016-0053084 (Feb. Korean Registered Patent No. 10-1116570 (Feb. International Patent WO 2010/033678 A2 (Mar. 25, 2010)

The inventors of the present invention have conducted studies to find a photosensitizer which is faster in the in vivo transfer to a cancer lesion site and superior in photosensitization activity, and found that when a hydrophilic polyethylene glycol group and a porphyrin derivative having a hydrophobic alkyl group are hydrophobic / Or hydrophilic / hydrophilic functional group-containing compound, and exhibits excellent anticancer activity upon irradiation with laser light.

Accordingly, it is an object of the present invention to provide a porphyrin derivative having a hydrophilic polyethylene glycol group and a hydrophobic alkyl group, and a process for producing the same.

In the present invention, a compound represented by the following formula (1) is provided.

[Chemical Formula 1]

Wherein m is an integer of 0 to 6 and n is an integer of 0 to 18.

In one embodiment, m may be 3, or n may be 9.

In one embodiment, the compound represented by Formula 1 may be a compound represented by Formula 1a below.

[Formula 1a]

According to one aspect of the present invention, a photosensitive composition comprising the compound is provided.

According to an aspect of the present invention, there is also provided an anticancer agent comprising the compound.

Further, according to one aspect of the present invention, there is provided a photosensitizer for photodynamic diagnosis or treatment comprising said compound.

According to an aspect of the present invention, there is also provided a process for preparing a compound represented by the following general formula (1), which comprises reacting a compound represented by the following general formula (2), a compound represented by the following general formula (3)

[Chemical Formula 1]

(2)

(3)

[Chemical Formula 4]

Wherein m is an integer of 0 to 6 and n is an integer of 0 to 18.

In one embodiment, the compound of formula 2 may be prepared by reacting 4-hydroxybenzaldehyde with triethyleneglycol methyl ether tosylate. The compound of formula 3 may be prepared by reacting 4-hydroxybenzaldehyde with an alkyl- .

In one embodiment, the compound represented by Formula 1 is a compound represented by Formula 1a, the compound represented by Formula 2 is represented by Formula 2a, and the compound represented by Formula 3 is represented by Formula 3a . ≪ / RTI >

[Formula 1a]

(2a)

[Chemical Formula 3]

According to the present invention, since the porphyrin derivative having a polyethylene glycol group and an alkyl group simultaneously contains a hydrophilic functional group and a hydrophobic functional group and has a structure similar to that of a cell membrane, it has an excellent penetration ability into tissue cells in vivo and also has a hydrophobic / hydrophobic functional group- It was found that the production efficiency of singlet oxygen was superior to that of hydrophilic / hydrophilic functional group-containing compound and it was found that it exhibits excellent anticancer activity upon laser light irradiation.

Accordingly, the porphyrin derivatives having hydrophilic polyethylene glycol groups and hydrophobic alkyl groups of the present invention and their production methods can be usefully used as anticancer agents or photo-sensitizers for photodynamic diagnosis / treatment which require a photosensitive composition.

Fig. 1 shows the structure analysis ( ¹ H-NMR) of the hydrophobic substituent.
2 is a structural analysis ( ¹ H-NMR) of the hydrophilic substituent.
3 shows MALDI-TOF-MS measurement results of porphyrin derivative 7 (compound 7) (A), porphyrin derivative 8 (compound 8) (B), and porphyrin derivative 9 (compound 9) (C).
4 shows the results of ¹ H-NMR measurement of porphyrin derivative 7 (compound 7).
5 shows the results of ¹ H-NMR measurement of porphyrin derivative 8 (compound 8).
6 shows the results of ¹ H-NMR measurement of porphyrin derivative 9 (compound 9).
7 is a UV-vis absorption spectrum of porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9).
8 is a fluorescence spectrum of porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9).
Fig. 9 shows the result of measurement of fluorescence change of DPBF molecules of porphyrin derivative 7 (compound 7).
Fig. 10 shows the result of measurement of fluorescence change of DPBF molecules of porphyrin derivative 8 (compound 8).
Fig. 11 shows the result of measuring fluorescence change of DPBF molecules of porphyrin derivative 9 (Compound 9).
12 is a graph showing cell viability at various concentrations of porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9) in the MTT evaluation.
13 is a graph showing cell viability of the porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9) under laser irradiation.

The present invention provides a compound represented by the following formula (1).

[Chemical Formula 1]

Wherein m is an integer of 0 to 6 and n is an integer of 0 to 18.

M is an integer of 0 to 6, preferably 1 to 5, more preferably 2 to 4, and most preferably 3 as a repeating unit of an ethylene glycol group which is a hydrophilic structure. N is an integer of 0 to 18, preferably 3 to 15, more preferably 6 to 12, most preferably 9, as a repeating unit of an alkyl group which is a hydrophobic structure.

When m is 3 and n is 9, it is a compound represented by the following formula (1a).

[Formula 1a]

The compound of Formula 1 may be prepared by reacting a compound of Formula 2, a compound of Formula 3 and a compound of Formula 4 below.

[Chemical Formula 1]

(2)

(3)

[Chemical Formula 4]

Wherein m is an integer of 0 to 6 and n is an integer of 0 to 18.

The compound of Formula 2 may be prepared by reacting 4-hydroxybenzaldehyde with triethyleneglycol methyl ether tosylate. The compound of Formula 3 may be prepared by reacting 4-hydroxybenzaldehyde with an alkyl-Br .

In one embodiment, the compound represented by Formula 1 is a compound represented by Formula 1a, the compound represented by Formula 2 is represented by Formula 2a, and the compound represented by Formula 3 is represented by Formula 3a . ≪ / RTI >

[Formula 1a]

(2a)

[Chemical Formula 3]

The porphyrin derivative having a polyethylene glycol group and an alkyl group of the present invention has hydrophilic functional groups and hydrophobic functional groups at the same time and has a structure similar to that of the cell membrane so that it has excellent penetration into tissue cells in vivo and also has hydrophobic / It has been found that the production efficiency of singlet oxygen is higher than that of a compound containing a hydrophilic functional group and it shows excellent anticancer activity upon laser light irradiation.

Accordingly, the porphyrin derivative having the hydrophilic polyethylene glycol group and the hydrophobic alkyl group of the present invention can be usefully used as a photosensitizer composition, an anticancer agent, or a photosensitizer for photodynamic diagnosis or treatment.

When the compound of the present invention is used as an anticancer agent or a photodynamic agent for photodynamic diagnosis or treatment, the dosage thereof varies depending on the condition and the weight of the patient, the degree of the disease, the drug form, the administration route and the period. May be appropriately selected by a person skilled in the art. For example, the above-mentioned anticancer agent or photodynamic agent for photodynamic diagnosis or treatment may be administered at a dose of 0.0001 to 1000 mg / kg, preferably 0.01 to 1000 mg / kg per day, Or may be administered in divided doses. In addition, the anticancer agent or photodynamic diagnostic or therapeutic agent of the present invention may contain 0.001 to 50% by weight of the compound, based on the total weight of the composition.

The anticancer agent or photodynamic agent for photodynamic diagnosis or treatment of the present invention can be administered to mammals such as rats, mice, livestock and human in various routes such as oral, peritoneal, rectal or intravenous, muscular, subcutaneous, intrauterine Or by intracerebroventricular injection.

Hereinafter, the present invention will be described in more detail through examples and test examples. However, the following examples and test examples are provided for illustrating the present invention, and the scope of the present invention is not limited thereto.

<Examples>

1. hydrophobic substituents ( 1 alkylaldehyde ) Synthesis of

(1) Synthesis method

In a 1000 ml two-necked flask, 150 ml of DMF, 2.7 g of 4-hydroxybenzaldehyde and 8.3 g of K ₂ CO ₃ were added, and the mixture was stirred at 85 ° C for 40 minutes. When the color of the solution became clear in the turbid state, 9.9 g of alkyl-Br (in DMF) was injected to proceed the reaction. The reaction was monitored every hour and the reaction was terminated and cooled to room temperature. The DMF was removed by heating to 70-80 ° C and evaporation. And washed once with methylene chloride (MC) / water. The residue was purified by silica column (400 ml) with hexane: methylene chloride (MC) = 4: 1 solvent. The reaction procedure is shown in Scheme 1 below.

[Reaction Scheme 1]

(2) Structural analysis ^One H-NMR)

The characteristic peaks of the compound were confirmed at 9.88 ppm of CHO-1H (A), 7 to 8 ppm of aromatic-4H (B) and 4.2 ppm of -OCH ₂ -2H (C) .

2. Hydrophilic substituent ( 1 Tegaldehyde ) Synthesis of

(1) Synthesis method

In a 1000 ml two-necked flask, 9 g of triethylene glycol methyl ether tosylate (TEG-OTs), 2.7 g of 4-hydroxybenzaldehyde, 8.3 g of K ₂ CO ₃ and 5.95 g of 18-crown- Were added and kept under vacuum / nitrogen for 40 minutes. After the addition of THF, the reaction was allowed to proceed at reflux at 90 ° C for 12 hours. When the 4-hydroxybenzaldehyde disappeared, the reaction was terminated. The remaining 18-crown-6 was removed by washing once with methylene chloride / water. The silica column (400 ml) was refined with an ethyl acetate solvent for 24 hours. The reaction procedure is shown in Scheme 2 below.

[Reaction Scheme 2]

(2) Structural analysis ^One H-NMR)

The characteristic peaks of the compound were confirmed at 9.88 ppm of CHO-1H (A), 7 to 8 ppm of aromatic-4H (B) and 3.2 to 4.5 ppm of -OCH ₂ CH ₂ -11H (C) Respectively.

3. Synthesis of porphyrin derivatives

Hydrophilic / hydrophilic porphyrin [compound 7], hydrophilic / hydrophobic porphyrin [compound 8] and hydrophobic / hydrophobic porphyrin [compound 9] 3 porphyrin derivatives were synthesized by cross-condensation reaction as shown in Reaction Scheme 3 below.

[Reaction Scheme 3]

(1) Synthesis method

In a 500 ml flask, 1.23 g of 2,2'-Dipyrromethane (DPM), 1.21 g of the hydrophobic substituent (1 alkylaldehyde) synthesized above, 1.21 g of the synthesized hydrophilic substituent (1-Tegaldehyde) Was added to 500 ml of a chloroform solvent and bubbled with nitrogen for 30 minutes. BF ₃ OEt ₂ 0.853 ml were added thereto and stirred in a dark place. After 1 hour, a small amount of P-chloranil (a necessary amount for the test reaction) was added and the porphyrin reaction was confirmed by TLC (pink fluorescence). When the reaction was confirmed, 1.290 g of P-chloriranyl was added and stirred for 1 hour. After completion of the reaction, 5 ml of triethylamine was added to remove the oxidation peak of porphyrin from UV. The polymer was then passed through a silica column (200 ml) with ethyl acetate to remove the polymer, and then the column step was carried out. Silica column (500 ml) hexane / MC (methylene chloride) / EA (ethyl acetate) (gradation) conditions.

(2) Structural analysis

(2-1) Measurement of molecular weight

The MALDI TOF-TOF 5800 System model was used to measure molecular weight under DHB matrix conditions.

The results of MALDI-TOF-MS measurement of porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9) are shown in FIG.

(2-2) ^One 1 H-NMR

Results of ¹ H-NMR measurement of the results of porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9) are shown in FIGS. 4, 5 and 6, respectively.

<Test Example>

1. Evaluation of Optical Properties of Synthetic Derivatives: Absorption and Fluorescence Spectrum Analysis

Absorption and fluorescence spectrum analysis were performed as follows, and the results of the analysis are shown in FIGS. 7 and 8. FIG.

Absorption spectra were measured under a THF solvent condition using a JASCO V-670 spectrophotometer. Fluorescence spectra were measured in a THF solvent using a Luminescence Spectrometer (LS 55 Luminescence Spectrometer, PerkinElmer).

As shown in Fig. 7, the long wavelength shift of the absorption band of porphyrin was observed with the increase of the hydrophilic substituent in the UV-vis absorption spectrum. Further, as shown in FIG. 8, the long wavelength shift of the maximum fluorescence peak was observed in accordance with the increase of the hydrophilic substituent in the fluorescence spectrum.

2. Singlet oxygen  Evaluation of generation efficiency

The porphyrin forms an excited state by the absorption of visible light, and this excited state can react with oxygen molecules to produce an atomic oxygen, one of the active oxygen atoms. Fluorescence changes of 1,3-diphenylisobenzofuran (DPBF) molecules were measured in order to determine the amount of anoxic oxygen produced by porphyrin. The DPBF molecule is a quencher of unilamellar oxygen and is capable of determining the relative amount of oxygen produced from fluorescence reduction. DMSO was used as a solvent and the concentration of each porphyrin derivative was adjusted to 1 μM. First, the same amount of DPBF was introduced into each porphyrin solution, and the fluorescence reduction of DPBF was measured by irradiating 635 nm laser. The results are shown in FIGS. 9 to 11.

As shown in Figs. 9 to 11, in the case of porphyrin derivative 7, the reduction of fluorescence of DPBF was found to be 7.12% lower than that of the initial amount. In the case of porphyrin derivative 8, the decrease of fluorescence of DPBF was 22.4% In the case of porphyrin derivative 9, the fluorescence reduction of DPBF was found to be 17.15% less than the initial amount. It was found that the production efficiency of the oxygen was higher in the order of the hydrophilic / hydrophobic compound (compound 8), the hydrophilic / hydrophilic compound (compound 9) and the hydrophobic / hydrophobic compound (compound 7), and the production efficiency of hydrophilic / hydrophobic porphyrin derivative 8 Respectively.

3. Assessment of cytotoxicity: MTT  evaluation( MTT  assay)

The light yellow substance 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide ( MTT) is a substance that is reduced by the mitochondrial activity of cells. By measuring the degree of reduction, the survival rate (cytotoxicity) of the cells can be grasped relatively.

The cell viability of the control group (the untreated group of the laser [laser]) and the group treated with the laser irradiation were measured, and general cytotoxicity and anticancer activity at the time of laser irradiation were evaluated.

(1) Evaluation method

The cell line MCF-7 (P.3), which is a breast cancer cell, was used as follows. Cells were seeded at a density of 5000 cells / well in a 96-well plate and cultured for 24 hours. The concentration of porphyrin was diluted with water (final DMSO concentration: 1%) at 20, 10, 5, and 2.5 uM and cultured for 12 hours. The porphyrin was dissolved in DMSO and prepared.

After that, the 635 nm laser was treated for 3 minutes only in the laser irradiation group (Light sample), and cultured for 12 hours after the generation of active oxygen. MTT solution (0.5% in PBS) was added to each well and reacted at 37 ° C for 3 hours. After completion of the reaction, the supernatant was removed, and formazan was dissolved in 200 μl of DMSO for 10 minutes. The dissolved formazan is purple and indicates a gradual increase in proportion to the amount. Absorbance was measured at 570 nm using a micro-reader (Microreader).

(2) MTT  Evaluation results

The cell viability at the respective concentrations of the porphyrin derivative 7 (compound 7), porphyrin derivative 8 (compound 8), and porphyrin derivative 9 (compound 9) in the case where the laser was not irradiated is shown in Fig. As shown in Fig. 12, all three compounds did not show cytotoxicity with a cell survival rate of 90% or more. In general, when the cell viability is 75% or more, cytotoxicity can be assumed to be insufficient.

(3) Evaluation of antitumor activity: Phototoxicity

A solution of each compound was irradiated with a laser of 635 nm to generate active oxygen, and cytotoxicity was evaluated. The results are shown in FIG. As shown in FIG. 13, the cell viability of the porphyrin derivative 8 having a hydrophilic / hydrophobic substituent was generally low, and cell viability was remarkably reduced at a solution concentration of 20 μM. As a result, it can be seen that the anticancer activity of the porphyrin derivative 8 upon light irradiation is superior to that of the porphyrin derivative 7 and the porphyrin derivative 9.

Claims (11)

A compound represented by the following formula (1a):
[Formula 1a]

.
delete delete delete A light-sensitive composition comprising a compound of claim 1.
An anticancer agent comprising the compound of claim 1.
A photosensitizer for photodynamic diagnosis or treatment comprising the compound of claim 1.
Reacting a compound of formula (2a), a compound of formula (3a), and a compound of formula (4): &lt; EMI ID =
[Formula 1a]

(2a)

[Chemical Formula 3]

[Chemical Formula 4]

.
The process according to claim 8, wherein the compound of formula (2a) is prepared by reacting 4-hydroxybenzaldehyde with triethylene glycol methyl ether tosylate.
The process according to claim 8, wherein the compound of formula (3a) is prepared by reacting 4-hydroxybenzaldehyde with an alkyl-Br.
delete

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