A tetrandrine-porphyrin derivative and its anti-tumor use

Abstract

The application synthesizes a series of novel ligustrazine-porphyrin derivatives with ligustrazine and porphyrin as lead compounds, combines porphyrin PDT and ligustrazine chemotherapy, realizes synergistic anticancer of photodynamic therapy (PDT) and chemotherapy, and obtains a series of antitumor drugs with better activity and smaller side effects, and has potential application value.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry, specifically to a tetramethylpyrazine-porphyrin derivative and its antitumor uses. Background Technology

[0002] Photodynamic therapy (PDT), as a novel non-invasive therapy mediated by reactive oxygen species (ROS), has received increasing attention in recent years and is now used to treat cancers such as pharyngeal cancer, skin cancer, and breast cancer, achieving certain therapeutic effects. Because PDT can selectively target tumor tissue with minimal impact on the function of normal cells, it boasts the advantage of minimal toxic side effects, making it particularly suitable for patients intolerant to traditional cancer treatments such as chemotherapy and radiotherapy. The reactive oxygen species generated during PDT (including type I hydroxyl radicals and type II singlet oxygen) can oxidize biomolecules (proteins, nucleic acids, and lipids) within the system. The oxidation of these essential biomolecules interferes with normal cellular activities, leading to tumor cell apoptosis and necrosis. To fully realize the therapeutic efficacy of PDT, the selection of a suitable photosensitizer is crucial.

[0003] With the development of photodynamic therapy (PDT), the preparation of photosensitizers, especially porphyrin-based photosensitizers, has made remarkable progress. Most photosensitizers used in cancer treatment have a macrocyclic skeleton based on porphyrins. The main advantages of porphyrin compounds in photodynamic research include: 1) the stability of aromatic compounds; 2) efficient absorption of visible light; 3) high reactive oxygen species yield; 4) ease of functionalization and structural diversity; and 5) long triplet lifetime and low dark toxicity. Photofri, the first photosensitizing drug approved by the US FDA, is a classic porphyrin-structured drug.

[0004] Li et al. (Chinese Chemical Letters, 2007, 18(11): 1331-1334) designed and synthesized a novel potential targeted anticancer drug based on porphyrin and 5-fluorouracil, and evaluated its in vitro anticancer activity against human liver cancer cells SMCC-7721 using the MTT assay. Preliminary results showed that the anticancer activity of the coupled compounds was more than twice that of 5-fluorouracil. Porphyrin had no killing effect on tumor cells under light-free conditions, indicating that the coupled compounds actually enhanced the targeting of 5-fluorouracil to tumor tissues, thereby enhancing the anticancer activity. S. Weimin et al. (Bioorg Med Chem, 2008, 16(10): 5665-5671) synthesized a series of porphyrin compounds containing 5-fluorouracil / L-phenylalanine by coupling 5-fluorouric acid / Boc-L-phenylalanine with aminoporphyrin. In vitro anticancer activity studies showed that the introduction of 5-fluorouracil and L-phenylalanine significantly enhanced the phototoxicity of porphyrins. The high selectivity of porphyrin compounds for tumor tissues broadens the prospects for research on porphyrin-linked anticancer drugs.

[0005] Tetramethylpyrazine (TMP) is the main active ingredient extracted from a plant called Ligusticum chuanxiong, and it has a long history and wide application in traditional Chinese medicine. TMP has many pharmacological effects, including vasodilation, antiplatelet aggregation, and regulation of blood circulation. It is often used to treat headaches, migraines, angina pectoris, and stroke. With in-depth research on tetramethylpyrazine, it may have the potential to inhibit the proliferation and metastasis of cancer cells. A series of in vitro studies have shown that tetramethylpyrazine can effectively inhibit the proliferation of various cancer cell lines, including but not limited to breast cancer, liver cancer, and lung cancer. Further research on the mechanism of action of tetramethylpyrazine has revealed that it may exert its anti-cancer effects through multiple pathways, such as regulating the cell cycle, promoting apoptosis, and inhibiting angiogenesis. Specifically, tetramethylpyrazine may affect the growth and metastasis of cancer cells, thereby inhibiting tumor development. These research results provide an important theoretical basis for further exploring the potential application of tetramethylpyrazine in cancer treatment, and also provide new ideas and directions for its clinical application and drug development. However, although in vitro experimental results demonstrate the anticancer potential of tetramethylpyrazine, its efficacy and safety in clinical applications still require further in-depth research and validation. Therefore, further clinical trials and pharmacological studies will help to comprehensively evaluate the potential value of tetramethylpyrazine as an anticancer drug and its clinical application prospects.

[0006] The tetramethylpyrazine-porphyrin derivative designed in this invention exhibits excellent anticancer efficacy under the synergistic effect of PDT and chemotherapy drugs, providing important scientific basis and technical support for expanding the application of porphyrin and its derivatives in the biomedical field. Summary of the Invention

[0007] The technical problem to be solved by the present invention is to provide a tetramethylpyrazine-porphyrin derivative, which exhibits excellent anti-cancer efficacy under the synergistic effect of PDT and chemotherapy drugs, while causing less damage to normal tissues and achieving selective inhibition of tumor cells.

[0008] The first aspect of the present invention is to provide a compound of formula I or formula II and a pharmaceutically acceptable salt thereof, having the following structure:

[0009]

[0010] Where n is an integer selected from 1 to 8;

[0011] R is independently selected from H, halogen, hydroxyl, nitro, CN, C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group, C 6-10 Aryl, C2- 10 Mixed aromatics;

[0012] R' is selected from H, halogen, hydroxyl, nitro, CN, C. 1-6 Alkyl, C 2-6 alkenyl, C 2-6 alkynyl group;

[0013] M is selected from Zn, Ni, and Mn.

[0014] Preferably, n is selected from 1, 2, 3, 4 or 5.

[0015] Preferably, each of R is independently selected from H, halogen, hydroxyl, nitro, CN, C. 1-6 Alkyl group; more preferably, each of R is independently selected from H or Cl;

[0016] Preferably, R' is selected from H, halogen, hydroxyl, nitro, CN, C. 1-6 Alkyl; more preferably, R is selected from H;

[0017] Preferably, M is selected from Zn.

[0018] Another aspect of the present invention provides a method for preparing compound of formula I, the synthetic route of which is as follows:

[0019]

[0020] The definitions of n, R, and R' are as described above.

[0021] The specific reaction steps are as follows:

[0022] In an organic solvent, an alkali, a catalyst, and porphyrin 1 are added, and the mixture is heated and stirred. Then, tetramethylpyrazine derivative 2 is added to the reaction system until the reaction is complete. After post-treatment, the porphyrin-tetramethylpyrazine derivative of formula I is obtained.

[0023] Preferably, the molar ratio of porphyrin derivative 1 to tetramethylpyrazine derivative 2 is 1:(1-1.5), more preferably 1:1-1.2, and even more preferably 1:1.2.

[0024] The alkali is selected from potassium hydroxide, triethylamine, or potassium carbonate, and more preferably potassium carbonate.

[0025] The catalyst is selected from one or two of potassium iodide and sodium iodide.

[0026] Another aspect of the present invention provides a method for preparing compounds of formula II, the synthetic route of which is as follows:

[0027]

[0028] The definitions of n, R, R' and M are as described above.

[0029] The specific reaction steps are as follows:

[0030] The compound of formula I was dissolved in an organic solvent, and the metal salt M was added. The mixture was refluxed and the reaction was monitored by TLC until the reaction was complete. After post-treatment, the tetramethylpyrazine-porphyrin derivative of formula II was obtained.

[0031] Preferably, the molar ratio of compound I to metal salt M is 1:3-7, more preferably 1:5.

[0032] Another aspect of the present invention provides a pharmaceutical composition comprising a compound of formula I or formula II or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier and excipient.

[0033] Another aspect of the present invention relates to the use of a compound of formula I or II and a pharmaceutically acceptable salt thereof or a pharmaceutical composition comprising the thereof in the preparation of an anticancer drug;

[0034] Preferably, the cancer is caused by gastric cancer, non-small lung cancer cells, prostate cancer, bladder cancer, colon cancer cells, or breast cancer; particularly, the cancer is caused by human gastric cancer cells HGC-27, human non-small lung cancer cells A549, human prostate cancer cells DU145, human bladder cancer cells TCCSUP, human colon cancer cells HCT-116, or human breast cancer cells MDA-MB-231.

[0035] Compared with the prior art, the beneficial effects of the present invention are:

[0036] (1) This invention provides a new class of tetramethylpyrazine-porphyrin derivatives with anticancer activity, which broadens the scope of existing anticancer compounds and can be used as lead compounds for further optimization;

[0037] (2) The compound of the present invention uses porphyrin molecules as carriers and utilizes their tumor tissue aggregation effect to target tumor cells, thereby reducing the side effect of killing normal cells.

[0038] (3) The tetramethylpyrazine-porphyrin derivative of the present invention can achieve the synergistic effect of phototherapy and chemotherapy. In addition, the insertion of metal Zn into porphyrin can also enhance the antitumor activity of the compound. Attached Figure Description

[0039] Figure 1 Effects of compound 8d on DU145 cell migration. Detailed Implementation

[0040] The present invention will be specifically illustrated below through examples. In this invention, the following examples are provided to better illustrate the invention and are not intended to limit the scope of the invention. Unless otherwise specified, the materials, reagents, etc., used in the following examples are commercially available.

[0041] Example 1: Synthesis of Porphyrins 4 and 5

[0042]

[0043] Pyrrole is prone to deterioration during storage, requiring a redistillation apparatus to be set up during use. The temperature should be maintained at 140℃, and the mixture should be refluxed in the dark. The initial turbid portion should be discarded, and the clear, transparent freshly distilled pyrrole should be collected. In a 250 mL three-necked flask, 4.8 g of p-hydroxybenzaldehyde and 6.1 mL of benzaldehyde were added, along with 120 mL of propionic acid as a solvent to completely dissolve the raw materials. After refluxing at 135℃ for 0.5 h, 11 mL of pyrrole was measured and diluted with propionic acid to 22 mL. This solution was then slowly added dropwise along the tube wall using a constant-pressure dropping funnel, with the dropping rate strictly controlled at approximately 0.5 h. After the addition was complete, reflux was continued for 1 h. After the reaction was finished, the mixture was cooled to room temperature. 60 mL of anhydrous ethanol was added to the reaction solution, and the mixture was left to stand overnight at 4℃. The mixture was then filtered and dried to obtain a dark purple solid crude product. Dichloromethane:n-hexane in a 3:1 ratio was used as the developing solvent, and the product was purified by silica gel chromatography. The product band was collected, and the solvent was removed under reduced pressure to obtain a blue-purple solid porphyrin 4.

[0044] Porphyrin 4: Yield: 7%, 1H NMR (500 MHz, Chloroform- d ) 8.89 – 8.83 (m, 8H),8.22 (d, J= 6.7 Hz, 6H), 8.11 – 8.06 (m, 2H), 7.75-7.70 (m, 9H), 7.26 (d, J = 7.8 Hz, 2H), -2.76 (s, 2H) ppm.

[0045]

[0046] 4.8 g of p-hydroxybenzaldehyde and 16.8 g of p-chlorobenzaldehyde were added to a 250 mL three-necked flask, and 120 mL of propionic acid was used as a solvent to completely dissolve the raw materials. After refluxing at 135 °C for 0.5 h, 11 mL of pyrrole was measured and diluted with propionic acid to 22 mL. The solution was then slowly added dropwise along the tube wall through a constant pressure dropping funnel, with the dropping rate strictly controlled at approximately 0.5 h. After the addition was complete, reflux was continued for 1 h. After the reaction was completed, the mixture was cooled to room temperature. 60 mL of anhydrous ethanol was added to the reaction solution, and the mixture was left to stand overnight at 4 °C. The mixture was then filtered and dried to obtain a dark purple solid crude product. Dichloromethane was used as the developing solvent, and the product was purified by silica gel column chromatography. The product band was collected, and the solvent was removed under reduced pressure to obtain a blue-purple solid porphyrin 5.

[0047] Porphyrin 5: Yield: 8% 1H NMR (500 MHz, Chloroform- d ) δ 9.05 (d, J = 4.7 Hz,2H), 8.95 – 8.92 (m, 6H), 8.15 (d, J = 7.8 Hz, 6H), 8.01 (d, J = 8.1 Hz, 2H), 7.74 (d, J = 8.0 Hz, 6H), 7.26 (d, J = 6.0 Hz, 2H), -2.82 (s, 2H) ppm.

[0048] Example 2 Synthesis of tetramethylpyrazine derivatives 3a-3e

[0049]

[0050] Prepare a 50 mL round-bottom flask. Accurately weigh 0.1 g (0.6 mmol) of tetramethylpyrazine formic acid, 0.414 g (3 mmol) of anhydrous potassium carbonate, and 48.5 μL (1.5 mmol) of triethylamine. Dissolve them thoroughly in DMF (8-10 mL). After stirring at room temperature / 60℃ for 0.5 h, accurately pipette 87 μL (0.72 mmol) of 1,6-dibromohexane, 96 μL (0.72 mmol) of 1,5-dibromopentane, 79.4 μL (0.72 mmol) of 1,4-dibromobutane, 126 μL (0.72 mmol) of 1,3-dibromopropane, and 63 μL (0.72 mmol) of 1,2-dibromoethane into the reaction system. Monitor the reaction progress using a TCL. After the reaction is complete, extract the reaction solution three times with dichloromethane as the extractant. Collect the dichloromethane and wash with pure water for 3-5 minutes. Next, remove as much DMF as possible. Remove water from dichloromethane using anhydrous Na₂SO₄, then rotary evaporate under reduced pressure to obtain a pale yellow viscous liquid, 3a-3e, as the crude product. Use methanol:dichloromethane (20:1) as the developing solvent, purify by silica gel column chromatography, collect the organic dissolved product, and rotary evaporate under reduced pressure to obtain a pale yellow viscous liquid compound, 3a-3e.

[0051] 3a: 1H NMR (500 MHz, DMSO- d 6) δ 4.29 (t, J = 6.6 Hz, 2H), 3.53 (t, J =6.7 Hz, 2H), 2.59 (s, 3H), 2.51 (s, 3H), 2.47 (s, 3H), 1.83-1.78 (m, 2H), 1.73-1.67 (m, 2H), 1.45-1.39 (m, 4H) ppm.

[0052] 3b: 1H NMR (500 MHz, DMSO- d 6) δ 4.3 (t, J = 6.8 Hz, 2H), 3.52 (t, J =6.2 Hz, 2H), 2.59 (s, 3H), 2.51 (s, 3H), 2.48 (s, 3H), 1.91 – 1.85 (m, 4H), 1.68 – 1.61 (m, 2H) ppm.

[0053] 3c: 1H NMR (500 MHz, DMSO- d 6) δ 4.31 (t, J= 6.4 Hz, 2H), 3.51 (t, J =5.9 Hz, 2H), 2.57 (s, 3H), 2.52 (s, 3H), 2.49 (s, 3H), 1.84 – 1.79 (m, 4H)ppm.

[0054] 3d: 1H NMR (500 MHz, DMSO-d6) δ 4.29 (t, J = 6.5 Hz, 2H), 3.52 (t, J =6.1 Hz, 2H), 2.57 (s, 3H), 2.51 (s, 3H), 2.47 (s, 3H), 1.96-1.91(m, 2H) ppm.

[0055] 3e: 1H NMR (500 MHz, DMSO-d6) δ 4.29 (t, J = 6.6 Hz, 2H), 3.52 (t, J =6.7 Hz, 2H), 2.58 (s, 3H), 2.52 (s, 3H), 2.47 (s, 3H) ppm.

[0056] Example 3 Synthesis of tetramethylpyrazine-porphyrin of Formula I

[0057]

[0058] 0.54 mmol of 3a / 3b / 3c / 3d / 3e (the yield is difficult to calculate due to the intermediate being liquid; a conversion rate of 90% is used) in a 50 mL round-bottom flask was diluted with 15 mL of DMF. 0.55 mmol of porphyrin 4 / porphyrin 5, 0.414 g of anhydrous potassium carbonate, and 0.1 g of potassium iodide were added. The mixture was reacted at 60 °C for 12 h, with TLC monitoring and time adjustments made. After the reaction, the reaction mixture was extracted three times with dichloromethane as the extractant. The dichloromethane was collected and washed 3-5 times with pure water to remove as much DMF as possible. Anhydrous Na₂SO₄ was used to remove water from the dichloromethane, and the mixture was rotary evaporated under reduced pressure to obtain a crude purple solid. The crude solid was purified using a silica gel column chromatography with dichloromethane:acetone = 50:1 as the developing solvent. The third purple band was collected, yielding derivatives 6a-6e and 7a-7e. The synthesized 6a-6e and 7a-7e were reacted with dichloromethane as solvent, and 0.11 g of zinc acetate was added to introduce metallic zinc. The mixture was kept under gentle reflux for 2 hours until the reaction was complete. The unreacted zinc acetate was washed with an appropriate amount of pure water, and the organic phase was dried with anhydrous Na2SO4. The mixture was then dried under reduced pressure to obtain bright purple solids 8a-8e and 9a-9e.

[0059] 6a: Yield: 78%.1 1H NMR (500 MHz, Chloroform- d ) δ 8.89 – 8.84 (m, 8H), 8.21 (d, J J = 6.9 Hz, 6H), 8.12 – 8.08 (m, 2H), 7.75 (m, 9H), 7.28 (d, J J = 8.0 Hz, 2H), 4.48 (t, J J = 6.8 Hz, 2H), 4.24 (t, J J = 6.3 Hz, 2H), 2.77 (s, 3H), 2.57 (s, 3H), 2.55 (s, 3H), 2.05 – 1.99 (m, 2H), 1.97 – 1.93 (m, 2H), 1.76 – 1.70 (m, 2H), 1.68 – 1.62 (m, 2H), -2.77 (s, 2H) ppm. FTMS: C58H50N6O3: 878.3944; [M+H] + : 879.4008.

[0060] 6b: Yield: 82%. 1 1H NMR (500 MHz, Chloroform- d ) δ 8.89 – 8.84 (m, 8H), 8.24 – 8.20 (m, 6H), 8.11 (d, J J = 8.0 Hz, 2H), 7.79 – 7.73 (m, 9H), 7.28 (d, J J = 8.5 Hz,, 2H), 4.53 (t, J J = 6.8 Hz, 2H), 4.28 (t, J J = 6.2 Hz, 2H), 2.80 (s, 3H), 2.60 (s, 3H), 2.57 (s, 3H), 2.10 – 2.00 (m, 4H), 1.84 – 1.78 (m, 2H), -2.77 (s, 2H) ppm. FTMS: C57H48N6O3: 864.3788; [M+H] + : 865.3850.

[0061] 6c: Yield: 85%. 1 1H NMR (500 MHz, Chloroform- d ) δ 9.00 (d,J = 4.8 Hz, 2H), 8.94 – 8.91 (m, 6H), 8.14 (d, J = 7.9 Hz, 6H), 8.10 (d, J = 8.0 Hz, 2H), 7.74 (d, J = 7.8 Hz, 6H), 7.28 (d, J = 8.2 Hz, 2H), 4.61 (t, J = 6.3 Hz, 2H), 4.35 (t, J = 5.8 Hz, 2H), 2.80 (s, 3H), 2.60 (s, 3H), 2.58 (s, 3H), 2.22 – 2.13 (m, 4H) ppm. FTMS: C56H46N6O3: 850.3631; [M+H] + : 851.3704.

[0062] 6d: Yield: 81%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.87 – 8.84 (m, 8H), 8.22 (d, J = 6.4 Hz, 5H), 8.12 (d, J = 8.1 Hz, 2H), 7.80 – 7.71 (m, 9H), 7.29(d, J = 8.1 Hz, 2H), 4.78 (t, J = 6.4 Hz, 2H), 4.43 (t, J = 6.0 Hz, 2H), 2.82(s, 3H), 2.61 (s, 3H), 2.58 (s, 3H), 2.61 – 2.48 (m, 2H), -2.77 (s, 2H) ppm. FTMS: C55H44N6O3: 836.3475; [M+H] + : 837.3538.

[0063] 6e: Yield: 82%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.85 (d, J = 10.8 Hz, 8H), 8.22 (d, J = 7.6 Hz, 6H), 8.13 (d,J = 7.9 Hz, 1H), 7.78 – 7.73 (m, 9H),7.33 (d, J = 8.4 Hz, 2H), 5.03 – 4.90 (m, 2H), 4.71 – 4.58 (m, 2H), 2.89 –2.85 (m, 3H), 2.64 – 2.58 (m, 6H), -2.77 (s, 2H) ppm. FTMS: C54H42N6O3: 822.3318; [M+H] + : 823.3391.

[0064] 7a: Yield: 84%, 1 1H NMR (500 MHz, Chloroform-d) δ 8.91 (d, J = 4.7 Hz,2H), 8.82 (d, J = 8.7 Hz, 6H), 8.14 (d, J = 8.2 Hz, 6H), 8.10 (d, J = 8.2 Hz,2H), 7.74 (d, J = 8.0 Hz, 6H), 7.28 (d, J = 8.2 Hz, 2H), 4.50 (t, J = 6.9 Hz,2H), 4.25 (t, J = 6.3 Hz, 2H), 2.79 (s, 3H), 2.59 (s, 3H), 2.56 (s, 3H), 2.03– 1.98 (m, 2H), 1.97– 1.91 (m, 2H), 1.74 – 1.68 (m, 2H), 1.65 – 1.61 (m, 2H),-2.82 (s, 2H) ppm. FTMS: C58H47Cl3N6O3: 982.2746; [M+H] + : 983.2822.

[0065] 7b: Yield: 80%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.91 (d, J = 4.8 Hz,2H), 8.83(d, J = 4.8 Hz, 6H), 8.16 – 8.12 (m, 6H), 8.10 (d, J = 8.2 Hz, 2H),7.74 (d, J = 8.0 Hz, 6H), 7.27 (d, J= 8.2 Hz, 2H), 4.54 (t, J = 6.8 Hz, 2H),4.28 (t, J = 6.2 Hz, 2H), 2.80 (s, 3H), 2.60 (s, 3H), 2.58 (s, 3H), 2.10 –2.00 (m, 4H), 1.84 – 1.78 (m, 2H), -2.83 (s, 2H) ppm. FTMS: C57H45Cl3N6O3: 966.2619; [M+H] + : 967.2691.

[0066] 7c: Yield: 85%, 1 H NMR (500 MHz, Chloroform- d ) δ 8.90 (d, J = 4.8 Hz,2H), 8.83 – 8.81 (m, 6H), 8.14 (d, J = 8.3 Hz, 6H), 8.10 (d, J = 8.2 Hz, 2H),7.75 (d, J = 8.2 Hz, 6H), 7.28 (d, J = 8.3 Hz, 2H), 4.62 (t, J = 6.3 Hz, 2H),4.34 (t, J = 5.8 Hz, 2H), 2.81 (s, 3H), 2.60 (s, 3H), 2.57 (s, 3H), 2.23 –2.13 (m, 4H), -2.83 (s, 2H) ppm. FTMS: C56H43Cl3N6O3: 952.2642; [M+H] + : 953.2535.

[0067] 7d: Yield: 85%, 1 H NMR (500 MHz, Chloroform- d ) δ 8.89 (d, J = 4.7 Hz,2H), 8.83 – 8.81 (m, 6H), 8.14 (d, J = 7.9 Hz, 6H), 8.10 (d, J = 8.1 Hz, 2H),7.75 (d, J= 8.0 Hz, 6H), 7.30 (d, J = 8.2 Hz, 2H), 4.78 (t, J = 6.4 Hz, 2H),4.44 (t, J = 6.0 Hz, 2H), 2.82 (s, 3H), 2.61 (s, 3H), 2.59 (s, 3H), 2.53 –2.48 (m, 2H), -2.84 (s, 2H) ppm. FTMS: C55H41Cl3N6O3: 938.2306; [M+H] + : 939.2378.

[0068] 7e: Yield: 84%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.89 – 8.80 (m, 8H),8.16 – 8.11 (q, J = 9.6, 9.0 Hz, 8H), 7.74 (d, J = 7.8 Hz, 6H), 7.33 (d, J =8.4 Hz, 2H), 4.96 (s, 2H), 4.65 (s, 2H), 2.88 (d, J = 10.2 Hz, 3H), 2.61 (d, J = 11.1 Hz, 6H), -2.84 (s, 2H) ppm. FTMS: C54H39Cl3N6O3: 954.2249; [M+H] + : 925.2210.

[0069] 8a: Yield: 90%, 1 1H NMR (500 MHz, Chloroform- d ) δ 9.01 – 8.90 (m, 8H),8.26 – 8.20 (m, 6H), 8.11 (d, J = 8.1 Hz, 2H), 7.81 – 7.71 (m, 9H), 7.25 (d,, J = 5.0 Hz, 2H), 4.47 (t, J = 6.8 Hz, 2H), 4.25 (t, J= 6.3 Hz, 2H), 2.74(s, 3H), 2.58 (s, 3H), 2.55 (s, 3H), 2.04 – 1.98 (m, 2H), 1.97 – 1.94 (m,2H), 1.75 – 1.71 (m, 2H), 1.67 – 1.62 (m, 2H) ppm. FTMS:C58H48N6O3Zn:940.31048;[M+H] + :941.3115.

[0070] 8b: Yield: 91%, 1 H NMR (500 MHz, Chloroform- d ) δ 8.99 (d, J = 4.6 Hz,2H), 8.94 (d, J = 3.0 Hz, 6H), 8.24 – 8.21 (m, 6H), 8.11 (d, J = 8.3 Hz, 2H),7.79 – 7.73 (m, 9H), 7.27 d, J = 5.0 Hz, 2H), 4.52 (t, J = 6.8 Hz, 2H), 4.29(t, J = 6.2 Hz, 2H), 2.78 (s, 3H), 2.60 (s, 3H), 2.57 (s, 3H), 2.10 – 2.00(m, 4H), 1.84 – 1.78 (m, 2H) ppm. FTMS:C57H46N6O3Zn:926.2953;[M+H] + :927.2963.

[0071] 8c: Yield: 92%, 11H NMR (500 MHz, Chloroform-d) δ 9.00 (d, J = 4.8 Hz, 2H), 8.94 - 8.91 (m, 6H), 8.14 (d, J = 7.9 Hz, 6H), 8.10 (d, J = 8.0 Hz, 2H), 7.74 (d, J = 7.8 Hz, 6H), 7.28 (d, J = 8.2 Hz, 2H), 4.61 (t, J = 6.3 Hz, 2H), 4.35 (t, J = 5.8 Hz, 2H), 2.80 (s, 3H), 2.60 (s, 3H), 2.58 (s, 3H), 2.22 – 2.13 (m, 4H) ppm. FTMS: C56H44N6O3Zn: 912.2838; [M+H] + : 913.2818.

[0072] 8d: Yield: 91%, 1 1H NMR (500 MHz, Chloroform-d) δ 8.98 – 8.94 (m, 8H), 8.22 (d, J = 6.9 Hz, 6H), 8.12 (d, J = 7.9 Hz, 2H), 7.79 – 7.73 (m, 9H), 7.28 (d, J = 8.6 Hz, 2H), 4.76 (t, J = 6.4 Hz, 2H), 4.43 (t, J = 5.9 Hz, 2H), 2.80 (s, 3H), 2.61 (s, 3H), 2.58 (s, 3H), 2.52 – 2.48 (m, 2H) ppm. FTMS: C55H42N6O3Zn: 898.2610; [M+H] + : 899.2662.

[0073] 8e: Yield: 90%, 11H NMR (500 MHz, Chloroform-d) δ 8.96 – 8.93 (m, 8H), 8.23 – 8.20 (m, 6H), 8.13 (d, J = 8.5 Hz, 2H), 7.78 – 7.78 (m, 9H), 7.32 (d, J = 8.4 Hz, 2H), 4.95 (t, J = 4.8 Hz, 2H), 4.65 (t, J = 4.8 Hz, 2H), 2.87 (s, 3H), 2.62 (s, 3H), 2.60 (s, 3H) ppm. FTMS: C54H40N6O3Zn: 884.2504; [M+H] + : 885.2511.

[0074] 9a: Yield: 92%, 1 1H NMR (500 MHz, Chloroform- d ) δ 9.01 (d, J = 4.6 Hz, 2H), 8.94– 8.90 (m, 6H), 8.14 (d, J = 7.9 Hz, 6H), 8.09 (d, J = 8.1 Hz, 2H), 7.74 (d, J = 8.0 Hz, 6H), 7.28 (d, J = 5.0 Hz, 2H), 4.48 (t, J = 6.8 Hz, 2H), 4.27 (t, J = 6.4 Hz, 2H), 2.76 (s, 3H), 2.59 (s, 3H), 2.56 (s, 3H), 2.02 (p, J = 6.7 Hz, 2H), 1.95 (p, J = 7.1 Hz, 2H), 1.76 – 1.70 (m, 2H), 1.67 – 1.28 (m, 2H) ppm. FTMS: C58H45Cl3N6O3: 1042.1910; [M+H] + : 1043.1958.

[0075] 9b: Yield: 92%, 1 1H NMR (500 MHz, Chloroform- d ) δ 9.01 (d, J= 4.7 Hz, 2H), 8.95 – 8.90 (m, 6H), 8.14 (d, J = 8.0 Hz, 6H), 8.10 (d, J = 8.4 Hz, 2H), 7.74 (d, J = 8.0 Hz, 6H), 7.27 (d, J = 4.1 Hz, 2H), 4.51 (t, J = 6.8 Hz, 2H), 4.28 (t, J = 6.2 Hz, 2H), 2.77 (s, 3H), 2.60 (s, 3H), 2.57 (s, 3H), 2.10 – 1.99 (m, 4H), 1.84 – 1.79 (m, 2H) ppm. FTMS: C57H43Cl3N6O3: 1028.1801; [M+H] + : 1029.1805.

[0076] 9c: Yield: 90%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.98 (d, J = 4.6 Hz, 2H), 8.94 (d, J = 3.0 Hz, 6H), 8.22 (d, J = 6.9 Hz, 6H), 8.11 (d, J = 8.1 Hz, 2H), 7.79 – 7.73 (m, 9H), 7.28 (d, J = 8.3 Hz, 2H), 4.61 (t, J = 6.3 Hz, 2H), 4.35 (t, J = 5.7 Hz, 2H), 2.81 (s, 3H), 2.60 (s, 3H), 2.57 (s, 3H), 2.23 – 2.13 (m, 4H) ppm. FTMS: C56H41Cl3N6O3: 1014.1617; [M+H] + : 1015.1650.

[0077] 9d: Yield: 91%, 1 1H NMR (500 MHz, Chloroform- d ) δ 8.99 (d, J= 4.7 Hz,2H), 8.93 – 8.91 (m, 6H), 8.14 (d, J = 7.9 Hz, 6H), 8.10 (d, J = 8.4 Hz, 2H), 7.74 (d, J = 8.1 Hz, 2H), 7.28 (d, J = 9.1 Hz, 2H), 4.74 (t, J = 6.3 Hz, 2H), 4.42 (t, J = 5.7 Hz, 2H), 2.78 (s, 3H), 2.60 (s, 3H), 2.58 (s, 3H), 2.51 –2.46 (m, 2H) ppm. FTMS: C55H39Cl3N6O3: 1000.1441; [M+H] + : 1001.1497.

[0078] 9e: Yield: 91%, 1 H NMR (500 MHz, Chloroform- d ) δ 8.98 (d, J = 4.7 Hz,2H), 8.94-8.91 (m, 6H), 8.13 (dd, J = 6.8, 4.8 Hz, 8H), 7.74 (d, J = 8.1 Hz, 6H), 7.33 (d, J = 8.4 Hz, 2H), 4.95 (t, J = 4.8 Hz, 2H), 4.65 (t, J = 4.8 Hz,2H), 2.86 (s, 3H), 2.62 (s, 3H), 2.60 (s, 3H) ppm. FTMS: C54H37Cl3N6O3: 986.1301; [M+H] + : 987.1342.

[0079] Example 4: Scratch resistance migration study

[0080] DU145 cells were seeded into 6-well plates and cultured to a cell density of 90%. Using a 20µL pipette tip, a straight scratch was gently made on the surface of the 6-well plate. The width and depth of the scratch were ensured to be consistent by adjusting the pipette tip angle and pressure. PBS buffer was used to wash away free cells and cell debris from the scratch area, making the scratch area clearly visible. Serum-free medium containing 8 days of iodine was prepared to a concentration lower than the IC50 of DU145 cells. 50 Three concentrations of the solution, 4µM, 8µM, and 16µM, were added to 6-well plates, with a negative control group (serum-free culture medium to exclude interference from proliferation under serum conditions). The initial state of the scratches was observed and photographed using an inverted microscope, with the scratch mark marked as time 0 h. The 6-well plates were returned to the incubator for further culture. At 12 h, one plate was removed and exposed to light for 10 min. After 24 hours, photographs of the scratched area were taken again to record the degree of cell migration. The area of ​​the scratched region was calculated using image processing software: migration rate = (initial scratch area - final migration area) / initial scratch area × 100%.

[0081] The test results are attached. Figure 1 , by appendix Figure 1 It was found that the cell migration rate differed significantly from the control group after 24 h, and the effect of 8 days on DU145 cell migration was concentration-dependent. At 12 h, the anti-migration ability of the 16 µM concentration after light treatment was greater than that of the untreated group; while the anti-migration ability of the 4 µM and 8 µM concentrations after light treatment was slightly less than that of the untreated group. The difference may be due to the influence of ROS levels. Low levels of ROS promote cell migration, while high levels inhibit it. Tetramethylpyrazine itself has anti-migration activity. At low concentrations of the 8-day compound, the less ROS generated by light exposure antagonizes the effect of tetramethylpyrazine, resulting in weakened anti-migration ability. At high concentrations, the high ROS levels synergistically enhance the anti-migration effect of tetramethylpyrazine. In conclusion, the 8-day compound has a better anti-migration effect, with an anti-migration rate of 41% against DU145 cells under light exposure, compared to 32% in the untreated group.

[0082] Example 5: MTT colorimetric assay for detecting in vitro antitumor cell activity

[0083] The inhibitory activity of the target compound and control compound on different cell types was determined using the MTT assay. A blank control group, negative control group, positive control group, and experimental group were set up. DMSO was used as the negative control group, 5-FU and Ce6 as the positive control group, and tetramethylpyrazine (TMP), porphyrin compounds (4, 5), and target compounds 6a-9e were used as the experimental group. The blank control group and negative control group each had 6 replicates, while the positive control group and experimental group each had 3 replicates.

[0084] Table 1. In vitro anti-proliferative activity of tetramethylpyrazine-porphyrin derivatives against HGC-27 and A549.

[0085]

[0086] Table 2. In vitro anti-DU145 and TCCSUP proliferation activity of tetramethylpyrazine-porphyrin derivatives

[0087]

[0088]

[0089]

[0090] Table 3. In vitro anti-proliferative activity of tetramethylpyrazine-porphyrin derivatives against HCT-116 and MDA-MB-231.

[0091]

[0092]

[0093]

[0094] Table 4. In vitro anti-H9C2 proliferation activity of tetramethylpyrazine-porphyrin derivatives

[0095]

[0096] As shown in Tables 1, 2, 3, and 4, the tetramethylpyrazine-porphyrin derivatives of this invention exhibited varying degrees of inhibitory effects on different cell lines under both light and dark conditions. After light treatment, the tetramethylpyrazine-porphyrin derivative series compounds showed inhibitory effects on all cell lines. After cells took up the compounds, laser irradiation resulted in a PDT effect, which effectively generated ROS to damage the cells; even without light treatment, the antitumor activity of tetramethylpyrazine also resulted in antiproliferative activity.

Claims

1. A tetramethylpyrazine-porphyrin derivative and its pharmaceutically acceptable salt, having the structure shown in Formula I or Formula II: in: n is selected from 2, 3, 4, 5, or 6; R is selected from H or Cl; R' is selected from H; M is selected from Zn.

2. A method for preparing the compound of formula I as described in claim 1, wherein the reaction route is as follows: in, The definitions of n, R, and R' are as described in claim 1; The specific reaction steps are as follows: In an organic solvent, add an alkali, a catalyst and porphyrin derivative 1, heat and stir, add tetramethylpyrazine derivative 2 to the reaction system until the reaction is complete, and after post-treatment, obtain tetramethylpyrazine-porphyrin derivative of formula I. The catalyst is selected from potassium iodide and sodium iodide.

3. The method according to claim 2, characterized in that: The molar ratio of porphyrin derivative 1 to tetramethylpyrazine derivative 2 is 1:(1-1.5); the base is selected from potassium hydroxide, triethylamine or potassium carbonate.

4. A method for preparing the compound of formula II as described in claim 1, wherein the reaction route is as follows: in, The definitions of n, R, R' and M are as described in claim 1; The specific reaction steps are as follows: Dissolve the compound of formula I in an organic solvent, add the metal salt of M, reflux the reaction, monitor the reaction by TLC until the reaction is complete, and obtain the tetramethylpyrazine-porphyrin derivative of formula II after post-treatment.

5. The method according to claim 4, characterized in that: The molar ratio of compound I to metal salt M is 1:3-7.

6. A pharmaceutical composition comprising the tetramethylpyrazine-porphyrin derivative of claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

7. Use of the tetramethylpyrazine-porphyrin derivative of claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 6 in the preparation of a medicament for treating cancer, wherein the cancer is selected from gastric cancer, non-small cell lung cancer, prostate cancer, bladder cancer, colon cancer cells, and breast cancer.

8. The use as described in claim 7, wherein the cancer is caused by human gastric cancer cells HGC-27, human non-small lung cancer cells A549, human prostate cancer cells DU145, human bladder cancer cells TCCSUP, human colon cancer cells HCT-116, or human breast cancer cells MDA-MB-231.

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