Compound for inducing apoptosis and immunogenic death of cancer cells as well as preparation method and application of compound

Abstract

The invention discloses a compound for inducing apoptosis and immunogenic death of cancer cells based on mitochondrial targeting of fatty acid as well as a preparation method and application of the compound, and belongs to the field of medicinal chemistry and preparations. The structural formula of the prodrug is shown as a formula (I) and / or a formula (II). The series of compounds can form a relatively stable self-assembly in an aqueous solution, show high cytotoxicity which is not possessed by pure triphenylphosphine and fatty acid, and induce tumor cells to generate an obvious ICD phenomenon. The medicine can be obtained through simple synthesis, is high in yield, low in preparation cost, high in stability and good in safety, meets the requirements of clinical medication and large-scale industrial production, and has good market prospects and clinical application value.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical chemistry and preparation, and specifically relates to a series of fatty acid-based mitochondrial targeted compounds that induce cancer cell apoptosis and immunogenic death, as well as preparation methods and anti-tumor applications. Background Art

[0002] Immunogenic cell death (ICD) refers to the release of tumor-associated antigens (TAAs) and damage-associated pattern molecules (DAMPs) by tumor cells when they die after being treated with chemotherapy drugs or photodynamic therapy, which further activates tumor-specific T lymphocytes by recruiting and activating antigen-presenting cells (APCs), thereby promoting anti-tumor immunity (1, Krysko, DV; Garg, AD; Kaczmarek, A.; Krysko, O.; Agostinis, P.; Vandenabeele, P., Immunogenic cell death and DAMPs in cancer therapy. Nat Rev Cancer 2012, 12(12), 860-875. 2, Kroemer, G.; Galassi, C.; Zitvogel, L.; Galluzzi, L., Immunogenic cell stress and death. Nat Immunol 2022, 23(4), 487-500.). However, despite the great progress made in the field of anti-tumor, the ICD inducers that can be used in clinical treatment are limited, and only a few drugs have been proven to cause ICD, such as anthracycline drugs doxorubicin and oxaliplatin. In addition, high doses of chemotherapy drugs are required to fully induce ICD in tumor cells, which is difficult to achieve clinically due to severe toxic side effects (Zhou, JY; Wang, GY; Chen, YZ; Wang, HX; Hua, YQ; Cai, ZD, Immunogenic cell death in cancer therapy: Present and emerging inducers. J Cell Mol Med 2019, 23(8), 4854-4865.).Therefore, it is urgent to explore more efficient and accurate new ICD-inducing drugs (1. Chen, C.; Ni, X.; Jia, SR; Liang, Y.; Wu, XL; Kong, DL; Ding, D., Massively Evoking Immunogenic Cell Death by Focused Mitochondrial Oxidative Stress using an AIE Luminogen with a Twisted Molecular Structure. Adv Mater 2019, 31(52), e1904914. 2. Zheng, P.; Ding, BB; Jiang, ZY; Xu, WG; Li, G.; Ding, JX; Chen, XS, Ultrasound-Augmented Mitochondrial Calcium Ion Overload by Calcium Nanomodulator to Induce Immunogenic Cell Death. Nano Lett 2021, 21(5), 2088-2093.).

[0003] Studies have shown that the generation of reactive oxygen species (ROS) and endoplasmic reticulum stress are closely related to cell immunogenic death. Drugs based on direct induction of endoplasmic reticulum stress by ROS can more effectively exert the ICD effect than drugs that indirectly induce endoplasmic reticulum stress, such as doxorubicin (Krysko, DV; Garg, AD; Kaczmarek, A.; Krysko, O.; Agostinis, P.; Vandenabeele, P., Immunogenic cell death and DAMPs in cancer therapy. Nat Rev Cancer 2012, 12 (12), 860-875.). Mitochondria are the main source of ROS in tumor cells, which can interact with the ER through mitochondrial-associated membranes (MAMs) and induce ER stress (1. Zorov, DB; Juhaszova, M.; Sollott, SJ, Mitochondrial Reactive Oxygen Species (Ros) and Ros Induced Ros Release. Physiological Reviews 2014, 94(3), 909-950. 2. Wu, HX; Carvalho, P.; Voeltz, GK, Here, there, and everywhere: The importance of ER membrane contactsites. Science 2018, 361(6401), eaan5835. 3. Csordas, G.; Weaver, D.; Hajnoczky, G., Endoplasmic Reticulum-Mitochondrial Contactology: Structure and Signaling Functions. Trends Cell Biol 2018, 28(7), 523-540.4, Senft, D.; Ronai, ZA, UPR, autophagy, and mitochondria crosstalk underlies the ER stress response. Trends Biochem Sci 2015, 40(3), 141-148.). Therefore, targeting mitochondria, damaging mitochondria and inducing the production of a large amount of ROS, thereby initiating mitochondrial and endoplasmic reticulum stress, is expected to provide ideas for the development of more efficient ICD inducers. Summary of the invention

[0004] Fatty acids are essential nutrients for human life. They have low toxicity in vivo and can only produce significant cytotoxicity to tumor cells at high concentrations (such as >100 μM). The applicant used amino or hydroxylated triphenylphosphine to chemically couple and modify fatty acids to synthesize mitochondrial-targeted fatty acid compounds. It was unexpectedly discovered that it can form relatively stable self-assemblies in aqueous solution and specifically damage mitochondria, generating a large amount of ROS, thereby showing high cytotoxicity that simple triphenylphosphine and fatty acids do not have, and inducing obvious ICD phenomena in tumor cells.

[0005] The purpose of the present invention is to provide a novel fatty acid-based mitochondrial targeted ICD inducer, which can activate specific anti-tumor immunity by inducing tumor cell ICD. To achieve the above purpose, the present invention adopts the following technical solutions:

[0006] The present invention provides a fatty acid-based mitochondrial-targeted cancer cell apoptosis and immunogenic death inducer, the structural formula of which is shown in formula (I) or (II):

[0007]

[0008] Wherein, m=1-8; R comes from different fatty acids.

[0009] Further, the R is:

[0010] wherein n is an integer of 3 to 30; one or more CH2-CH2 in R may be further replaced by CH=CH. Furthermore, R contains 0 to 8 CH=CH.

[0011] Furthermore, m is an integer of 1 to 8; further, m is an integer of 2 to 8; specifically m is 2, 3, 4, 5, 6, 7, 8.

[0012] Further, the R structure is as follows:

[0013]

[0014] More specifically, the compound is characterized in that it is one of the following compounds:

[0015]

[0016] In the present invention, various fatty acids are modified by using the mitochondrial targeting group triphenylphosphonium (TPP) to synthesize a series of mitochondrial targeting fatty acid compounds. Specifically, the triphenylphosphine group provides the compound with the property of targeting mitochondria, and the compound can self-assemble in an aqueous solution to form a nanoparticle structure.

[0017] The research results of the present invention show that the anti-tumor activity of the nanoparticles formed by assembling the above compounds is significantly better than that of fatty acids and triphenylphosphine, and can induce immunogenic cell death of tumor cells.

[0018] The present invention also provides a method for preparing the compound described in any of the above technical solutions, comprising: (1) reacting R precursor fatty acid ROH with N-hydroxysuccinimide under the action of a base or a base / catalyst to obtain an intermediate product; (2) reacting the intermediate product with an amino or hydroxylated derivative of triphenylphosphine under the action of a base or a base / catalyst, and performing post-treatment after the reaction to obtain a drug with a structural formula as shown in formula (I) or (II).

[0019] Further, a method for synthesizing the fatty acid-based mitochondrial-targeted immunogenic death inducer comprises the following steps:

[0020] (1) In the presence of a base or a base / catalyst, a fatty acid reacts with N-hydroxysuccinimide to obtain an intermediate product; in the reaction, the molar ratio of the fatty acid to the N-hydroxysuccinimide is 1:1 to 1.5.

[0021] (2) In the presence of a base or a base / catalyst, the intermediate product reacts with an amino or hydroxylated derivative of triphenylphosphine. After the reaction, the crude product is separated and purified to obtain a drug having a structural formula as shown in formula (I) and (II).

[0022] Preferably, in step (1) and step (2), the base is selected from but not limited to N,N-diisopropylethylamine (DIEA) or 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC); the catalyst is selected from but not limited to 4-dimethylaminopyridine (DMAP).

[0023] Preferably, in step (1) and step (2), the reaction solvent is dichloromethane, dimethylformamide (DMF), dimethyl sulfoxide (DMSO) or a mixture of two or three of the above solvents.

[0024] Preferably, in step (1), the molar ratio of fatty acid to N-hydroxysuccinimide is 1:1 to 1.5.

[0025] Preferably, in step (1), the reaction temperature is 20-50°C and the reaction time is 1-4 hours. More preferably, the reaction temperature is 45°C.

[0026] Preferably, in step (2), the molar ratio of the intermediate product to the amino or hydroxylated derivative of triphenylphosphine is 1:1 to 1.5. The structural formula of the amino or hydroxylated derivative of triphenylphosphine is as follows:

[0027]

[0028] Among them, m=1~8.

[0029] Preferably, the triphenylphosphine amino derivative is 3-aminopropyl (triphenyl) phosphine bromide, 5-aminopentyl (triphenyl) phosphine bromide, 9-aminononyl (triphenyl) phosphine bromide. The triphenylphosphine hydroxylated derivative is 3-hydroxypropyl (triphenyl) phosphine bromide, 5-hydroxypentyl (triphenyl) phosphine bromide, 9-hydroxynonyl (triphenyl) phosphine bromide.

[0030] Preferably, in step (2), the reaction temperature is 20-50°C and the reaction time is 8-24 hours. During the reaction, thin layer chromatography is used to monitor the reaction progress. More preferably, the reaction temperature is 45°C and the reaction time is 8 hours.

[0031] Another object of the present invention is to provide a mitochondrial targeted fatty acid compound self-assembly preparation. Specifically, the mitochondrial targeted fatty acid compound is dissolved in an organic solvent such as dimethyl sulfoxide (DMSO), and slowly injected into deionized water under an ultrasonic water bath to prepare self-assembled nanoparticles (mtDSN). The volume ratio of DMSO to deionized water is fixed at 3-20%.

[0032] Preferably, the volume ratio of DMSO to deionized water is fixed at 5%.

[0033] During the respiratory oxidation process, mitochondria store the generated energy in the mitochondrial inner membrane as electrochemical potential energy, resulting in a negative potential difference (-180mV to -200mV) on both sides of the mitochondrial inner membrane. Driven by this mitochondrial membrane potential, lipophilic cations can be preferentially accumulated in mitochondria (Murphy, MP, Selective targeting of bioactive compounds to mitochondria. Trends in biotechnology 1997, 15 (8), 326-30; Murphy, MP, Targeting lipophilic cations to mitochondria. Biochim. Biophys. Acta. 2008, 1777 (7-8), 1028-31). Triphenylphosphine (TPP) is the most typical and widely used lipophilic cation mitochondrial targeting group. The delocalization of the three benzene ring π electron clouds in its structure makes the entire molecule positively charged, which can be targeted to mitochondria under the drive of mitochondrial membrane potential. Therefore, triphenylphosphine salts can achieve mitochondrial targeting by virtue of the lipophilic cationic properties of triphenylphosphine (Zhang Haifeng, An Lu. Application of triphenylphosphine salts in tumor diagnosis and treatment [J]. Journal of Shanghai Normal University (Natural Science Edition), 2024, 53(01): 137-145.). Salts exist in the form of free ions in aqueous solution, and the drug precursors in the present invention also exist in the form of free ions in the solution environment, and do not require the participation of anions when exercising the targeting effect.

[0034] During preparation, the fatty acid-based mitochondrial targeted cancer cell apoptosis and immunogenic death inducer mentioned in the present invention can form a compound structure that is externally neutral with the corresponding negative ions. The amino or hydroxylated derivative of triphenylphosphine used in the preparation process also exists in the form of cations in the reaction system, and forms an externally neutral intermediate structure with the above negative ions. The above negative ions include but are not limited to fluoride ions, bromide ions, iodide ions or astatine ions.

[0035] The present invention shows that, compared with less toxic fatty acids and triphenylphosphine amino or hydroxylated derivatives, the fatty acid-based mitochondrial-targeted immunogenic death inducer provided by the present invention exhibits a strong cytotoxic effect in various tumor cells and induces immunogenic cell death of tumor cells.

[0036] The present invention also provides the use of the fatty acid-based mitochondrial-targeted immunogenic death inducer in the preparation of anti-tumor drugs.

[0037] Specifically, the tumor is pancreatic cancer, lung cancer, cervical cancer, liver cancer, breast cancer, gastric cancer, ovarian cancer, colorectal cancer, bone cancer, melanoma, oral cancer, brain glioma, prostate cancer, bladder cancer, and kidney cancer.

[0038] Compared with the prior art, the present invention has the following beneficial effects:

[0039] (1) The present invention synthesizes a fatty acid-based mitochondrial-targeted cancer cell apoptosis and immunogenic death inducer, which can self-assemble in aqueous solution to form a nanoformulation without the need for the addition of an amphiphilic polymer excipient. It exhibits a powerful anti-tumor activity that simple fatty acids and triphenylphosphine amino derivatives do not have, and can induce the immunogenic death of tumor cells, providing ideas for the development of efficient ICD inducers.

[0040] (2) The present invention can obtain the drug precursor through simple synthesis, with high yield, low preparation cost, high stability, good safety, and meets the requirements of clinical drug use and large-scale industrial production, and has good market prospects and clinical application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 This is the synthetic route of fatty acid compound 1 in Example 1.

[0042] Figure 2 This is the synthetic route of fatty acid compound 2 in Example 2.

[0043] Figure 3 This is the synthetic route of fatty acid compound 3 in Example 3.

[0044] Figure 4 This is the synthetic route of fatty acid compound 4 in Example 4.

[0045] Figure 5 This is the synthetic route of fatty acid compound 5 in Example 5.

[0046] Figure 6 This is the synthetic route of fatty acid compound 6 in Example 6.

[0047] Figure 7 This is the synthetic route of fatty acid compound 7 in Example 7.

[0048] Figure 8 This is the synthetic route of fatty acid compound 8 in Example 8.

[0049] Fig. 9 This is the synthetic route of fatty acid compound 9 in Example 9.

[0050] Fig.10 This is the synthetic route of fatty acid compound 10 in Example 10.

[0051] Fig.11 This is the synthetic route of fatty acid compound 11 in Example 11.

[0052] Fig.12 This is the synthetic route of fatty acid compound 12 in Example 12.

[0053] Fig.13 This is the synthetic route of fatty acid compound 13 in Example 13.

[0054] Fig.14 This is the synthetic route of fatty acid compound 14 in Example 14.

[0055] Fig.15 This is the synthetic route of fatty acid compound 15 in Example 15.

[0056] Fig.16 This is the synthetic route of fatty acid compound 16 in Example 16.

[0057] Fig.17 This is the synthetic route of fatty acid compound 17 in Example 17.

[0058] Fig.18 This is the synthetic route of fatty acid compound 18 in Example 18.

[0059] Fig.19 This is the synthetic route of fatty acid compound 19 in Example 19.

[0060] Fig. 20 This is the synthetic route of fatty acid compound 20 in Example 20. DETAILED DESCRIPTION

[0061] The present invention will be further described below in conjunction with specific embodiments. The following embodiments are only used to illustrate the present invention and are not intended to limit the scope of application of the present invention. Without departing from the spirit and essence of the present invention, modifications or replacements made to the inventive method, steps or conditions all belong to the scope of the present invention.

[0062] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials and reagents used are reagents and materials that can be obtained from commercial channels unless otherwise specified.

[0063] N-hydroxysuccinimide, CAS No. 6066-82-6; triphenylphosphine, CAS No. 603-35-0; 3-bromopropylamine, CAS No. 5003-71-4; 5-bromopentylamine, CAS No. 51874-27-2; heptanoic acid, CAS No. 142-62-1; lauric acid, CAS No. 143-07-7; oleic acid, CAS No. 112-80-1; linoleic acid, CAS No. 60-33-3; DHA, CAS No. 6217-54-5; 9-aminononyl (triphenyl) phosphonium bromide, CAS No. 2248017-20-9, Merck 906107; 3-hydroxypropyl (triphenyl) phosphonium bromide, CAS No. 51860-45-8, Merck S860166.

[0064] Example 1 Synthesis of fatty acid compound 1

[0065] 1. Synthesis of 3-aminopropyl (triphenyl)phosphonium bromide

[0066] 3-Bromopropylamine (0.6 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. Silica gel column chromatography was used to separate and purify to obtain 0.9 g of a white solid product with a yield of 80.4%.

[0067] 2. Synthetic fatty acid compound 1

[0068] Weigh heptanoic acid (Hep, 58.6 mg, 0.45 mmol), N-hydroxysuccinimide (NHS, 62.1 mg, 0.54 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC, 77.6 mg, 0.50 mmol) and 4-dimethylaminopyridine (DMAP, 61.1 mg, 0.50 mmol) and dissolve in 2 mL of anhydrous dichloromethane (DCM). Stir and heat the reaction solution to 45 ° C for 2 h to obtain Hep-NHS. DCM was dried, 4 mL of anhydrous N, N-dimethylformamide (DMF) was added, and then 3-aminopropyl (triphenyl) phosphine bromide (200.2 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and stirred and heated at 45 ° C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction, the DMF was dried by oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 110.5 mg and the yield was 47.9%. For the synthetic route, see Figure 1 . 1 H NMR (400MHz, CDCl3) δ9.04-9.01 (t, J = 6Hz, 1H), 7.82-7.67 (m, 15H), 3.87-3.80 (m, 2H), 3.50-3.46 (m, 2H), 2 .40-2.36(t,J=8Hz,2H),1.89-1.83(m,2H),1.66-1.59(m,2H),1.36-1.24(m,6H),0.87-0.84(t,J=6Hz,3H). 13 C NMR (100MHz, CDCl3) δ174.8,135.1,135.1,135.1,133.5,133.5,133.5,133.4,133.4,133.4,130.6,130.6,130 .6,130.5,130.5,130.5,118.6,117.8,117.8,38.7,36.4,31.7,29.1,25.9,22.5,21.1,20.6,14.1.HRMS:calcd for[C 28 H 35 NOP] + =432.2451; obsd:432.2442.

[0069] Example 2 Synthesis of fatty acid compound 2

[0070] 1. Synthesis of 3-aminopropyl (triphenyl)phosphonium bromide

[0071] 3-Bromopropylamine (0.6 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. Silica gel column chromatography was used to separate and purify to obtain 0.9 g of a white solid product with a yield of 80.4%.

[0072] 2. Synthetic fatty acid compounds 2

[0073] Weigh lauric acid (Lau, 90.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain Lau-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 3-aminopropyl (triphenyl) phosphine bromide (200.2 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 105.6 mg, with a yield of 40.3%. For the synthetic route, see Figure 2 . 1 H NMR (400MHz, CDCl3) δ9.01-8.98 (t, J = 6Hz, 1H), 7.82-7.67 (m, 15H), 3.88-3.81 (m, 2H), 3.50-3.47 (m, 2H), 2. 40-2.36(t,J=8Hz,2H),1.87-1.82(m,2H),1.65-1.58(m,2H),1.31-1.19(m,16H),0.89-0.86(t,J=6Hz,3H). 13 C NMR (100MHz, CDCl3) δ174.8,135.1,135.1,135.1,133.5,133.5,133.5,133.4,133.4,133.4,130.6,130.6,130.6,130.5,130. 5,130.5,118.7,117.8,117.8,38.7,36.4,31.9,29.6,29.6,29.6,29.4,29.3,26.0,22.7,22.6,21.0,20.5,14.1.HRMS:calcd for[C 33 H 45 NOP] + =502.3233; obsd:502.3227.

[0074] Example 3 Synthesis of fatty acid compound 3

[0075] 1. Synthesis of 3-aminopropyl (triphenyl)phosphonium bromide

[0076] 3-Bromopropylamine (0.6 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. Silica gel column chromatography was used to separate and purify to obtain 0.9 g of a white solid product with a yield of 80.4%.

[0077] 2. Synthetic fatty acid compounds 3

[0078] Weigh oleic acid (OA, 127.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain OA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 3-aminopropyl (triphenyl) phosphine bromide (200.2 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 101.4 mg, with a yield of 33.9%. For the synthetic route, see Figure 3 . 1 H NMR (400MHz, CDCl3) δ8.99-8.96 (t, J = 6Hz, 1H), 7.81-7.67 (m, 15H), 5.37-5.29 (m, 2H), 3.86-3.79 (m, 2H), 3.48 (s, 2H), 2. 39-2.35(t,J=8Hz,2H),2.01-1.98(t,J=6Hz,4H),1.85(s,2H),1.62-1.60(m,2H),1.27(s,20H),0.89-0.86(t,J=6Hz,3H). 13C NMR (100MHz, CDCl3) δ174.8,135.1,135.1135.1,133.5,133.5,133.5,133.4,133.4,133.4,130.6,130.6,130.6,130.5,130.5,130.5,129.9,12 9.8,118.7,117.8,117.8,38.8,36.4,31.9,29.8,29.8,29.5,29.5,29.5 ,29.4,29.3,27.2,25.9,22.7,22.7,22.6,20.9,20.4,14.1.HRMS:calcd for[C 39 H 55 NOP] + =584.4016; obsd:584.4006.

[0079] Example 4 Synthesis of fatty acid compound 4

[0080] 1. Synthesis of 3-aminopropyl (triphenyl)phosphonium bromide

[0081] 3-Bromopropylamine (0.6 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. Silica gel column chromatography was used to separate and purify to obtain 0.9 g of a white solid product with a yield of 80.4%.

[0082] 2. Synthetic fatty acid compounds 4

[0083] Weigh linoleic acid (LA, 126.2 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain LA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 3-aminopropyl (triphenyl) phosphine bromide (200.2 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, and washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 145.8 mg and the yield was 48.9%. For the synthetic route, see Figure 4 . 1 H NMR (400MHz, CDCl3) δ9.11-9.08(t,J=6Hz,1H),7.82-7.61(m,15H),5.41-5.28(m,4H),3.88-3.81(dd,J=16,12Hz,2H),3.49-3.47(m,2H),2.78-2. 75(t,J=6Hz,2H),2.41-2.37(t,J=8Hz,2H),2.07-2.00(m,4H),1.88-1.79 (m,2H),1.64-1.59(m,2H),1.39-1.22(m,14H),0.90-0.87(t,J=6Hz,3H). 13 C NMR (100MHz, CDCl3) δ174.8,135.1,135.1,135.1,133.5,133.5,133.5,1 33.4,133.4,133.4,130.6,130.6,130.6,130.5,130.5,130.5,130.2,130 .2,127.9,127.9,118.7,117.8,117.8,38.7,36.4,31.5,29.7,29.5,29. 4,29.3,27.3,27.2,25.9,25.6,22.7,22.6,21.1,20.6,14.1.HRMS:calcd for[C 39 H 53 NOP] + =582.3859; obsd:582.3860.

[0084] Example 5 Synthesis of fatty acid compound 5

[0085] 1. Synthesis of 3-aminopropyl (triphenyl)phosphonium bromide

[0086] 3-Bromopropylamine (0.6 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. Silica gel column chromatography was used to separate and purify to obtain 0.9 g of a white solid product with a yield of 80.4%.

[0087] 2. Synthetic fatty acid compounds 5

[0088] Weigh DHA (147.8 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain DHA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 3-aminopropyl (triphenyl) phosphine bromide (200.2 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the reaction was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 135.9 mg, with a yield of 42.5%. For the synthetic route, see Figure 5 . 1 H NMR (400MHz, CDCl3) δ9.07-9.05(t,J=4Hz,1H),7.81-7.67(m,15H),5.47-5.25(m,12H),3.87-3.79(dd,J=16,12Hz,2H),3 .49-3.48(d,J=4Hz,2H),2.84-2.82(m,10H),2.49-2.37(m,4H),2.11-2.04(m,2H),1.86(s,2H),0.99-0.95(t,J=8Hz,3H). 13 C NMR (100MHz, CDCl3) δ173.9,135.1,135.1,135.1,133.6,133.6,133.6,133.5,13 3.5,133.5,132.0,130.6,130.6,130.6,130.5,130.5,130.5,129.1,128.6,128.5 ,128.3,128.2,128.2,128.1,127.9,127.9,127.0,127.0,118.7,117.8,117.8,3 8.8,35.8,25.6,25.6,25.6,25.5,23.5,22.6,21.0,20.5,20.5,14.3.HRMS:calcd for[C 43 H 53 NOP] + =630.3859; obsd:630.3858.

[0089] Example 6 Synthesis of fatty acid compound 6

[0090] 1. Synthesis of 5-aminopentyl (triphenyl) phosphonium bromide

[0091] 5-Bromopentylamine (0.7 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. 1 g of white solid product was obtained by separation and purification by silica gel column chromatography with a yield of 83.3%.

[0092] 2. Synthetic fatty acid compound 6

[0093] Weigh heptanoic acid (Hep, 58.6 mg, 0.45 mmol), N-hydroxysuccinimide (NHS, 62.1 mg, 0.54 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC, 77.6 mg, 0.50 mmol) and 4-dimethylaminopyridine (DMAP, 61.1 mg, 0.50 mmol) and dissolve in 2 mL of anhydrous dichloromethane (DCM). Stir and heat the reaction solution to 45 °C for 2 h to obtain Hep-NHS. DCM was dried, 4 mL of anhydrous N,N-dimethylformamide (DMF) was added, and then 5-aminopentyl (triphenyl) phosphine bromide (214.1 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction, the DMF was dried by oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 143.8 mg and the yield was 59.1%. For the synthetic route, see Figure 6 .

[0094] Example 7 Synthesis of fatty acid compound 7

[0095] 1. Synthesis of 5-aminopentyl (triphenyl) phosphonium bromide

[0096] 5-Bromopentylamine (0.7 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. 1 g of white solid product was obtained by separation and purification by silica gel column chromatography with a yield of 83.3%.

[0097] 2. Synthetic fatty acid compound 7

[0098] Weigh lauric acid (Lau, 90.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain Lau-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 5-aminopentyl (triphenyl) phosphine bromide (214.1 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 132.6 mg and the yield was 48.3%. For the synthetic route, see Figure 7 .

[0099] Example 8 Synthesis of fatty acid compound 8

[0100] 1. Synthesis of 5-aminopentyl (triphenyl) phosphonium bromide

[0101] 5-Bromopentylamine (0.7 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. 1 g of white solid product was obtained by separation and purification by silica gel column chromatography with a yield of 83.3%.

[0102] 2. Synthetic fatty acid compound 8

[0103] Weigh oleic acid (OA, 127.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain OA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 5-aminopentyl (triphenyl) phosphine bromide (214.1 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 122.5 mg, with a yield of 39.3%. For the synthetic route, see Figure 8 .

[0104] Example 9 Synthesis of fatty acid compound 9

[0105] 1. Synthesis of 5-aminopentyl (triphenyl) phosphonium bromide

[0106] 5-Bromopentylamine (0.7 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. 1 g of white solid product was obtained by separation and purification by silica gel column chromatography with a yield of 83.3%.

[0107] 2. Synthetic fatty acid compounds 9

[0108] Weigh linoleic acid (LA, 126.2 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain LA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 5-aminopentyl (triphenyl) phosphine bromide (214.1 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the reaction was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 147.4 mg, with a yield of 47.4%. For the synthetic route, see Fig. 9 .

[0109] Example 10 Synthesis of fatty acid compound 10

[0110] 1. Synthesis of 5-aminopentyl (triphenyl) phosphonium bromide

[0111] 5-Bromopentylamine (0.7 g, 2.8 mmol) and triphenylphosphine (1.4 g, 5.3 mmol) were refluxed in butanol (6 mL) at 120 ° C for 6 h and then cooled to room temperature. Benzene and ether were added to the reaction mixture, and the precipitate was washed with ether until it was no longer sticky. The solid was then redissolved in ethanol and ether was added. 1 g of white solid product was obtained by separation and purification by silica gel column chromatography with a yield of 83.3%.

[0112] 2. Synthetic fatty acid compounds 10

[0113] Weigh DHA (147.8 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain DHA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 5-aminopentyl (triphenyl) phosphine bromide (214.1 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, and washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 149.7 mg, with a yield of 45.0%. For the synthetic route, see Fig.10 .

[0114] Example 11 Synthesis of fatty acid compound 11

[0115] 1. Synthetic fatty acid compound 11

[0116] Weigh heptanoic acid (Hep, 58.6 mg, 0.45 mmol), N-hydroxysuccinimide (NHS, 62.1 mg, 0.54 mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC, 77.6 mg, 0.50 mmol) and 4-dimethylaminopyridine (DMAP, 61.1 mg, 0.50 mmol) and dissolve in 2 mL of anhydrous dichloromethane (DCM). Stir and heat the reaction solution to 45 °C for 2 h to obtain Hep-NHS. DCM was dried, 4 mL of anhydrous N,N-dimethylformamide (DMF) was added, and then 9-aminononyl (triphenyl) phosphine bromide (242.3 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction, the DMF was dried by oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 153.2 mg. For the synthetic route, see Fig.11 .

[0117] Example 12 Synthesis of fatty acid compound 12

[0118] 1. Synthetic fatty acid compound 12

[0119] Weigh lauric acid (Lau, 90.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain Lau-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 9-aminononyl (triphenyl) phosphine bromide (242.3 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 150.5 mg. For the synthetic route, see Fig.12 .

[0120] Example 13 Synthesis of fatty acid compound 13

[0121] 1. Synthetic fatty acid compound 13

[0122] Weigh oleic acid (OA, 127.1 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain OA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 9-aminononyl (triphenyl) phosphine bromide (242.3 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 159.3 mg. For the synthetic route, see Fig.13 .

[0123] Example 14 Synthesis of fatty acid compound 14

[0124] 1. Synthetic fatty acid compound 14

[0125] Weigh linoleic acid (LA, 126.2 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain LA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 9-aminononyl (triphenyl) phosphine bromide (242.3 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 156.1 mg. For the synthetic route, see Fig.14 .

[0126] Example 15 Synthesis of fatty acid compound 15

[0127] 1. Synthetic fatty acid compound 15

[0128] Weigh DHA (147.8 mg, 0.45 mmol), NHS (62.1 mg, 0.54 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) and dissolve them in 2 mL of anhydrous DCM. Stir and heat the reaction solution to 45 °C for 2 h to obtain DHA-NHS. DCM was dried, 4 mL of anhydrous DMF was added, and then 9-aminononyl (triphenyl) phosphine bromide (242.3 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol) were added, and the mixture was stirred and heated at 45 °C for 8 h. The reaction process was monitored by thin layer chromatography. After the reaction was completed, the DMF was dried with an oil pump, and washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product was separated and purified by silica gel column chromatography and vacuum dried to obtain a white solid. The yield was 159.2 mg. For the synthetic route, see Fig.15 .

[0129] Example 16 Synthesis of fatty acid compound 16

[0130] 1. Synthetic fatty acid compound 16

[0131] Weigh heptanoic acid (Hep, 58.6 mg, 0.45 mmol), add 4 mL of anhydrous N, N-dimethylformamide (DMF), then add 3-hydroxypropyl (triphenyl) phosphine bromide (200.6 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol), and stir and heat at 45 ° C for 8 h. The reaction process is monitored by thin layer chromatography. After the reaction is completed, the DMF is dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product is separated and purified by silica gel column chromatography, and a white solid is obtained after vacuum drying. The yield is 143.9 mg. For the synthetic route, see Fig.16 .

[0132] Example 17 Synthesis of fatty acid compound 17

[0133] 1. Synthetic fatty acid compound 17

[0134] Weigh lauric acid (Lau, 90.1 mg, 0.45 mmol), add 4 mL of anhydrous N, N-dimethylformamide (DMF), then add 3-hydroxypropyl (triphenyl) phosphine bromide (200.6 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol), and stir and heat at 45 ° C for 8 h. The reaction process is monitored by thin layer chromatography. After the reaction is completed, the DMF is dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product is separated and purified by silica gel column chromatography, and a white solid is obtained after vacuum drying. The yield is 147.2 mg. For the synthetic route, see Fig.17 .

[0135] Example 18 Synthesis of fatty acid compound 18

[0136] 1. Synthetic fatty acid compound 18

[0137] Weigh oleic acid (OA, 127.1 mg, 0.45 mmol), add 4 mL of anhydrous N, N-dimethylformamide (DMF), then add 3-hydroxypropyl (triphenyl) phosphine bromide (200.6 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol), and stir and heat the reaction at 45 ° C for 8 h. The reaction process is monitored by thin layer chromatography. After the reaction is completed, the DMF is dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product is separated and purified by silica gel column chromatography, and a white solid is obtained after vacuum drying. The yield is 158.3 mg. For the synthetic route, see Fig.18 .

[0138] Example 19 Synthesis of fatty acid compound 19

[0139] 1. Synthetic fatty acid compounds 19

[0140] Weigh linoleic acid (LA, 126.2 mg, 0.45 mmol), add 4 mL of anhydrous N, N-dimethylformamide (DMF), then add 3-hydroxypropyl (triphenyl) phosphine bromide (200.6 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol), and stir and heat at 45 ° C for 8 h. The reaction process is monitored by thin layer chromatography. After the reaction is completed, the DMF is dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product is separated and purified by silica gel column chromatography, and a white solid is obtained after vacuum drying. The yield is 157.6 mg. For the synthetic route, see Fig.19 .

[0141] Example 20 Synthesis of Fatty Acid Compound 20

[0142] 1. Synthetic fatty acid compounds 20

[0143] Weigh DHA (147.8 mg, 0.45 mmol), add 4 mL of anhydrous N, N-dimethylformamide (DMF), then add 3-hydroxypropyl (triphenyl) phosphine bromide (200.6 mg, 0.50 mmol), EDC (77.6 mg, 0.50 mmol) and DMAP (61.1 mg, 0.50 mmol), stir and heat at 45 ° C for 8 h. The reaction process is monitored by thin layer chromatography. After the reaction is completed, the DMF is dried with an oil pump, washed with 5% citric acid, saturated NaHCO3 and saturated NaCl in turn, and then dehydrated with anhydrous Na2SO4. The initial product is separated and purified by silica gel column chromatography, and a white solid is obtained after vacuum drying. The yield is 163.4 mg. For the synthetic route, see Fig. 20 .

[0144] Example 21 Preparation and testing of mtDSN nanoformulation

[0145] The fatty acid compounds prepared in Examples 1 to 10 were dissolved in DMSO and slowly injected into deionized water under an ultrasonic water bath to prepare self-assembled nanoparticles (mtDSN). The volume ratio of DMSO to deionized water was fixed at 5%. Uniformly dispersed nanoparticles were obtained. The nanoparticles prepared from fatty acid compound 1 were named mtDSN-1. Similarly, the corresponding nanoparticles were prepared from fatty acid compounds 2 to 5, respectively, and were named mtDSN-2 to 5.

[0146] Example 22 Toxicity test of mtDSN on tumor cells

[0147] The toxicity of the nanoparticles mtDSN-1 to 5 prepared from the fatty acid compounds 1 to 5 in the examples to various tumor cells was investigated, and the specific method was as follows:

[0148] Take cells in the logarithmic growth phase and inoculate them in a 96-well culture plate (2000-5000 cells / well). After culturing in a 37°C cell culture incubator for 24 hours, add mtDSN-1~5, 3-aminopropyl (triphenyl) phosphine bromide (TPP-NH2) and fatty acids (dissolved in DMSO) as control drugs, and 4 replicates for each drug and each concentration. After adding the drugs, place the 96-well cell plate in a cell culture incubator for culture. After incubation for 72 hours, add 30μL MTT solution (5mg / mL) to each well and continue incubation for 4 hours. Discard the culture medium and add 100μL DMSO to each well. After sufficient shaking, measure the absorbance at 490nm with an enzyme reader. Calculate the relative activity of the cells according to the following formula and draw a survival curve to obtain the IC value of the drug on cell growth. 50 (half inhibitory concentration).

[0149] Cell viability (%)=(At-Ab) / (Acon-Ab)×100%.

[0150] Among them, At, Acon and Ab are the absorbance of the experimental group (Treatment), the control group (Control, no fatty acid compound and comparison drug, only cells are added) and the blank well (Blank, no drug is added, no cells are added). The in vitro cytotoxicity results of mtDSN-1~5 on tumor cells are shown in Table 1.

[0151] Table 1. Cytotoxicity test of mtDSN-1-5 in A549 lung cancer, HeLa cervical cancer, Panc2 pancreatic cancer, LM3 liver cancer, 4T1 breast cancer, 7901 gastric cancer, A2780 ovarian cancer, MC38 colorectal cancer, U2OS osteosarcoma, B16F10 melanoma, DU145 prostate cancer, HSC-3 oral cancer, U251 brain glioma, MBT-2 bladder cancer, TK10 kidney cancer tumor cells (IC 50 ±SD, μM)

[0152]

[0153] Table 2. Cytotoxicity of fatty acids and TPP-NH2 in A549 lung cancer, HeLa cervical cancer, Panc2 pancreatic cancer, LM3 liver cancer, 4T1 breast cancer, 7901 gastric cancer, A2780 ovarian cancer, MC38 colorectal cancer, U2OS osteosarcoma, B16F10 melanoma, DU145 prostate cancer, HSC-3 oral cancer, U251 brain glioma, MBT-2 bladder cancer, and TK10 kidney cancer tumor cells (IC 50 ±SD,

[0154] μM)

[0155]

[0156] Note: TPP+Hep, TPP+Lau, TPP+OA, TPP+LA, TPP+DHA are the fatty acids and TPP-NH2 corresponding to the nanoparticles mtDSN-1, mtDSN-2, mtDSN-3, mtDSN-4, and mtDSN-5, respectively.

[0157] The results in Tables 1 and 2 show that after 72 h of co-culture with various tumor cells, the anti-tumor activity of nanoparticles mtDSN-1~5 prepared from fatty acid compounds 1~5 was significantly enhanced compared with fatty acids and TPP-NH2, realizing the transformation of fatty acids from "low tumor toxicity" to "high toxicity anti-tumor".

[0158] Example 23 Detection of mtDSN-induced cancer cell apoptosis

[0159] The ability of the nanoparticles mtDSN-1 to 5 prepared from the fatty acid compounds 1 to 5 in inducing apoptosis of triple-negative breast cancer cells 4T1 was investigated, and the specific method was as follows:

[0160] 4T1 cells were cultured at 1×10 5 Each well was inoculated with 10 μM nanoparticles mtDSN-1-5 (10 μM) prepared from fatty acid compounds 1-5 and incubated for 48 hours. When the experiment was over, the cells were digested with trypsin and washed with PBS and centrifuged to obtain a cell pellet. Annexin V-FITC and PI were then added according to the kit instructions for staining at room temperature in the dark for 15 minutes. Each group of samples was screened and tested on a flow cytometer.

[0161] Table 3. Proportion of apoptosis induced by mtDSN-1-5 in 4T1 cells

[0162]

[0163] As shown in Table 3, after incubation at a concentration of 10 μM for 48 hours, the apoptotic rate of nanoparticles mtDSN-2~5 increased significantly.

[0164] Example 24 Evaluation of mtDSN-1-5 inducing tumor cell ICD and promoting anti-tumor immune activation

[0165] Mouse primary bone marrow cells were extracted from the femur and tibia of healthy C57BL / 6 mice, and were co-stimulated with colony stimulating factor and IL-4 for 5 days to differentiate into BMDCs. 4T1 cells were inoculated in a six-well plate and co-cultured with equal concentrations of free drugs and nanoparticles for 12 hours, then the drugs were withdrawn and replaced with fresh culture medium for another 12 hours. The supernatant was co-cultured with the induced BMDCs for 24 hours. BMDCs incubated with fresh culture medium were used as blank controls. Flow cytometry was used to analyze the CD80 of BMDCs under different treatments. + / CD86 + and the expression of MHCII, to explore whether the small molecule self-assembly mtDSN-1~5 can induce the maturation of antigen-presenting cells by promoting ICD of breast cancer cells.

[0166] Table 4. mtDSN-1~5 induce BMDC maturation

[0167]

[0168] As shown in Table 4, the small molecule self-assembly mtDSN-2~5 can significantly increase the CD80 + / CD86 +The expression of MHCII was verified to achieve the induction of immunogenic death of tumor cells. The above embodiments are only preferred embodiments of the present invention, not all. Based on the embodiments in the embodiments, other embodiments obtained by those skilled in the art without creative work are all within the protection scope of the present invention.

Claims

1. A compound, characterized in that The structure is shown in formula (I) or (II): Wherein, m=1-8; R comes from different fatty acids.

2. The compound according to claim 1, characterized in that The R is: wherein n is an integer of 3 to 30; and one or more CH2-CH2 in R may be further replaced by CH=CH.

3. The compound according to claim 1, characterized in that The R structure is as follows:

4. The compound according to claim 1, characterized in that One of the following compounds:

5. A method for preparing the compound according to any one of claims 1 to 4, characterized in that: include: (1) In the presence of a base or a base / catalyst, the R precursor fatty acid ROH reacts with N-hydroxysuccinimide to obtain an intermediate product; (2) In the presence of a base or a base / catalyst, the intermediate product reacts with an amino or hydroxylated derivative of triphenylphosphine, and after the reaction, post-processing is performed to obtain a drug with a structural formula as shown in formula (I) or (II); the structures of the amino or hydroxylated derivative of triphenylphosphine are as follows:

6. A nanoparticle, characterized in that: It is formed by self-assembly of the compound according to any one of claims 1 to 4.

7. An anti-tumor drug, characterized in that: Containing a therapeutically effective amount of the compound according to any one of claims 1 to 4 or the nanoparticles according to claim 6.

8. The anti-tumor drug according to claim 7, characterized in that: The tumor includes one or more of the following: pancreatic cancer, lung cancer, cervical cancer, liver cancer, breast cancer, gastric cancer, ovarian cancer, colorectal cancer, osteosarcoma, melanoma, oral cancer, brain glioma, prostate cancer, bladder cancer, and kidney cancer.

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