Flucuridine phosphamide ester prodrug and application thereof in preparation of medicine for treating liver cancer

Abstract

The invention discloses a floxuridine phosphamide ester prodrug and application thereof in preparation of a medicine for treating liver cancer, and belongs to the technical field of medicine. The invention designs and synthesizes a series of 5-FdU phosphamide ester derivatives which are subjected to specific esterification modification at 3 '-site. Different aromatic acyl, fatty acyl or amino acid residues are introduced to the 3 '-site, so that the lipophilicity of molecules is further adjusted to enhance liver tissue distribution, and the enrichment and activation process of the medicine in the liver is optimized by utilizing the steric hindrance or metabolic lysis characteristic of the 3'-site. Experiments prove that the series of compounds show excellent anti-hepatoma cell proliferation activity and good pharmacokinetic characteristics, and a new choice is provided for clinical treatment of liver cancer.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to a fluorouridine phosphoramide ester prodrug and its use in the preparation of antitumor drugs, particularly drugs for treating liver cancer. Background Technology

[0002] Primary liver cancer is one of the most common malignant tumors worldwide, and its incidence and mortality rates rank among the highest in my country. Because liver cancer has an insidious onset, most patients are diagnosed at an advanced stage, losing the opportunity for surgical resection. Currently, chemotherapy remains one of the important means of comprehensive treatment for liver cancer.

[0003] 5-Fluorouracil (5-FU) and its deoxynucleoside form, 5-fluoro-2'-deoxyuridine (5-FdU), are classic antimetabolite antitumor drugs. In vivo, they induce tumor cell apoptosis by inhibiting thymidylate synthase (TS), thus blocking DNA biosynthesis. However, the clinical application of 5-FdU in liver cancer faces the following major challenges: 1. Metabolic limitations: After entering cells, 5-FdU must be phosphorylated by thymidine kinase (TK) to be converted into a nucleoside monophosphate (5-FdUMP) to exert its activity. In liver cancer cells, kinase activity is often downregulated, leading to low activation efficiency and drug resistance.

[0004] 2. First-pass effect and half-life: 5-FdU is readily degraded and metabolized by dihydropyrimidine dehydrogenase (DPD) in vivo, resulting in low bioavailability and strong systemic toxicity.

[0005] To overcome the aforementioned shortcomings, phosphoramide ester prodrug (ProTide) technology has been introduced into the design of nucleoside analogs. ProTide technology significantly enhances the lipophilicity of the drug by masking neutral ligands (such as amino acid esters and phenoxy groups) at the 5'-phosphate group of the nucleoside, enabling it to enter cells via passive diffusion. More importantly, this technology allows for the direct release of monophosphate nucleosides after entering cells, bypassing the rate-limiting step—kinase phosphorylation—thus effectively overcoming drug resistance caused by kinase deficiency. However, existing ProTide derivatives still have room for improvement in terms of targeting specific liver cancers, metabolic stability, and systemic toxicity control. For example, balancing the stability of the drug in the bloodstream with the rate of release of the active drug within hepatocytes remains a research challenge.

[0006] This invention designs and synthesizes a series of 5-FdU phosphoramide ester derivatives with specific esterification modifications at the 3'-position. By introducing different aromatic acyl, fatty acyl, or amino acid residues at the 3'-position, not only is the lipophilicity of the molecule further modulated to enhance its distribution in liver tissue, but the steric hindrance or metabolic cleavage properties of the 3'-position are also utilized to optimize the enrichment and activation process of the drug in the liver. Experiments have demonstrated that this series of compounds exhibits excellent anti-hepatocellular carcinoma cell proliferation activity and favorable pharmacokinetic characteristics, providing a new option for the clinical treatment of liver cancer. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of existing technologies, such as low bioavailability of 5-fluorouracil (5-FU), strong kinase dependence, easy development of drug resistance in liver cancer treatment, and significant systemic toxicity, and to provide a novel 5-fluorouracil deoxynucleoside phosphoramide ester derivative. This series of compounds significantly enhances lipophilicity through a dual strategy of 5'-phosphoramide esterification and 3'-specific ester modification, enabling intracellular drug delivery bypassing kinase activation and improving selective killing activity against liver cancer cells.

[0008] In a first aspect, the present invention provides a compound as represented by general formula I, or a pharmaceutically acceptable salt, hydrate, solvate, polymorph, or stereoisomer thereof: , in: R1 is selected from the following groups: (1) C1-C 12 Chain-like fatty acyl groups, C3-C 12 Cycloalkyl acyl groups: such as isobutyryl, n-hexanoyl, adamantyl, etc.; (2) Substituted or unsubstituted aryl group: The substituent is selected from halogen, C1-C4 alkyl, C1-C4 alkoxy, acyloxy or cyano; for example p-methoxybenzoyl, 3,5-dimethoxybenzoyl, 3,5-difluorobenzoyl, p-acetoxybenzoyl, p-isopropoxybenzoyl, etc. (3) Amino acyl group: such as L-alanyl, L-valine, etc. The amino group can be protected by protecting groups such as tert-butoxycarbonyl (Boc).

[0009] In some specific embodiments, the compound is selected from one of the following fifteen compounds: .

[0010] In a second aspect, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of the above-mentioned compound or a pharmaceutically acceptable salt, hydrate, solvate, polymorph or stereoisomer thereof, and a pharmaceutically acceptable excipient.

[0011] Furthermore, the dosage form of the pharmaceutical composition is an injection, an oral preparation, or a topical dosage form.

[0012] Thirdly, the present invention provides the use of the above-mentioned compound or its pharmaceutically acceptable salt, hydrate, solvate, polymorph or stereoisomer, or the above-mentioned pharmaceutical composition in the preparation of a medicament for treating liver cancer.

[0013] Furthermore, the liver cancer mentioned is primary liver cancer.

[0014] Furthermore, the drug may be used in combination with one or more other antitumor drugs.

[0015] Compared with the prior art, the present invention has the following significant advantages: Efficiently bypassing kinase dependence: As a prodrug of ProTide, the compounds of this invention can be directly metabolized in cells to release 5-FdUMP monophosphate, independent of the rate-limiting step of thymidine kinase (TK), thus exhibiting stronger sensitivity to drug-resistant liver cancer cell lines lacking or with low TK expression. Synergistic regulation of lipophilicity and metabolic stability: By introducing specific ester groups (such as adamantyl or substituted benzoyl groups) at the 3'-position, the molecule's lipid-water partition coefficient (LogP) is finely regulated. This not only enhances cell membrane permeability but also protects the glycoring structure through steric hindrance, prolonging the drug's half-life in plasma. Excellent antitumor activity: In vitro cytotoxicity experiments show that the compounds of this invention exhibit significantly superior inhibitory activity against various liver cancer cell lines (such as HepG2 and Huh7) compared to the parent drug 5-FU, with some compounds showing significantly higher IC50 values. 50 The value reached the nanomolar level. Reduced systemic toxicity: Because the prodrug remains in an inactive form before entering tumor cells and exhibits a good tendency to accumulate in liver tissue, damage to the hematopoietic system and gastrointestinal mucosa is reduced, improving the safety of drug administration. Attached Figure Description

[0016] Figure 1 This is a record of the metabolic hydrolysis process of compound DL-44 synthesized in this invention under the action of carboxypeptidase Y. 31 PNMR data.

[0017] Figure 2 This is a graph showing the metabolic stability of the compound DL-44 synthesized in this invention in rat plasma.

[0018] Figure 3 This invention compares the tumor volume of liver carcinoma in situ in nude mice with that of the synthesized compounds DL-21 and DL-44, the control group, and the sorafenib treatment group. Detailed Implementation

[0019] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only a part of the embodiments of the present invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the protection scope of the present invention. Example 1

[0020] Synthesis of compounds.

[0021] The synthetic route of compound DL-21 is shown below. All compounds represented by general formula I can be prepared by referring to this synthetic route. The molar ratios of various reactants are based on stoichiometry, and the reaction temperature is room temperature.

[0022] (1) Using 5-fluoro-2'-deoxyuridine as raw material and N,N-dimethylformamide as solvent, under an anhydrous and oxygen-free argon atmosphere, it reacts with tert-butyldimethylsilane chloride to obtain 5-fluoro-2'-deoxyuridine with tert-butyldimethylsilyl group protection at the 5' position.

[0023] (2) Using 5'-O-tert-butyldimethylsilyl-5-fluoro-2'-deoxyuridine as the raw material and 1,4-dioxane as the solvent, 3'-O-tert-butyloxycarbonyl-5'-O-tert-butyldimethylsilyl-5-fluoro-2'-deoxyuridine was reacted with ditert-butyl dicarbonate in the presence of DMAP to obtain 3'-O-tert-butyloxycarbonyl-5'-O-tert-butyldimethylsilyl-5-fluoro-2'-deoxyuridine.

[0024] (3) Using 3'-O-tert-butyloxycarbonyl-5'-O-tert-butyldimethylsilyl-5-fluoro-2'-deoxyuridine as the raw material and tetrahydrofuran as the solvent, it reacts with 1 M tetra-n-butylammonium fluoride to obtain 3'-O-tert-butyloxycarbonyl-5-fluoro-2'-deoxyuridine.

[0025] (4) Using 3'-O-tert-butyloxycarbonyl-5-fluoro-2'-deoxyuridine as a raw material, under anhydrous and oxygen-free conditions and in an argon atmosphere, with 1 M tert-butylmagnesium chloride solution, it reacts with N-[(S)-(2,3,4,5,6-pentafluorophenoxy)phenoxyphosphoryl]-L-alanine isopropyl ester to obtain 3'-O-tert-butyloxycarbonyl-5-fluoro-2'-deoxyuridine-5'-O-[phenyl(isopropoxy-L-alanyl)] phosphate.

[0026] (5) 3'-O-tert-butoxycarbonyl-5-fluoro-2'-deoxyuridine-5'-O-[phenyl(isopropoxy-L-alanyl)] phosphate reacts with trifluoroacetic acid to give 5-fluoro-2'-deoxyuridine-5'-O-[phenyl(isopropoxy-L-alanyl)] phosphate.

[0027] (6) 5-Fluoro-2'-deoxyuridine-5'-O-[phenyl(isopropoxy-L-alanyl)] phosphate, in anhydrous dichloromethane as solvent, reacted with Boc-L-2-aminobutyric acid in the presence of EDCI and DMAP to give 3'-O-(tert-butoxycarbonyl-L-2-aminobutyryl)-5-fluoro-2'-deoxyuridine-5'-O-[phenyl(isopropoxy-L-alanyl)] phosphate, namely compound DL-21. Example 2

[0028] Preparation and characterization of specific compounds.

[0029] The following fluorodeoxyuridine monophosphate prodrugs were prepared according to the synthetic route in Example 1, and were named and characterized. All compounds were characterized by 1H and 1C NMR spectra on a JNM-EPC 600 MHz NMR spectrometer.

[0030]

[0031] DL-19 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 2,4,6-trifluorobenzoate 1 H NMR (400 MHz, Chloroform- d ) δ 7.84 (d, J = 6.1 Hz, 1H), 7.31 (t, J= 7.9 Hz, 2H), 7.23 (d, J = 8.1 Hz, 2H), 7.15 (t, J = 7.3 Hz, 1H), 6.79 –6.70 (m, 2H), 6.25 (ddd, J = 9.2, 5.3, 1.7 Hz, 1H), 5.52 (d, J = 6.2 Hz, 1H), 5.01 (p, J = 6.2 Hz, 1H), 4.43 (dd, J = 6.4, 2.5 Hz, 2H), 4.32 (p, J = 2.4Hz, 1H), 4.02 – 3.90 (m, 2H), 2.51 (dd, J = 14.2, 5.3 Hz, 1H), 2.01 – 1.94 (m, 1H), 1.37 (d, J = 6.4 Hz, 3H), 1.22 (dd, J = 6.3, 1.6 Hz, 7H). DL-21 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl (S)-2-((tert-butoxycarbonyl)amino)butanoate

[0032] 1H NMR (400 MHz, Chloroform- d ) δ 7.83 (d, J = 6.2 Hz, 1H), 7.35 –7.27 (m, 2H), 7.23 – 7.19 (m, 2H), 7.15 (td, J = 7.5, 1.1 Hz, 1H), 6.19 (ddd, J = 9.2, 5.2, 1.7 Hz, 1H), 5.34 – 5.28 (m, 1H), 5.01 (p, J = 6.3 Hz, 1H),4.93 (d, J = 7.9 Hz, 1H), 4.37 (dt, J = 6.1, 2.9 Hz, 2H), 4.23 – 4.11 (m,2H), 4.01 – 3.90 (m, 1H), 3.76 (t, J = 10.4 Hz, 1H), 2.43 (ddd, J = 14.2,5.3, 1.3 Hz, 1H), 1.94 – 1.79 (m, 2H), 1.67 (dd, J = 14.6, 6.9 Hz, 1H), 1.44(s, 9H), 1.36 (d, J = 7.0 Hz, 3H), 1.22 (dd, J = 6.3, 1.3 Hz, 6H), 0.96 (t, J = 7.4 Hz, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.92 (d, J = 7.2 Hz),172.56, 157.08, 156.82, 155.56, 150.38 (d, J = 6.8 Hz), 148.87, 141.92,139.56, 129.91, 125.47, 124.15, 123.81, 120.39 (d, J = 4.7 Hz), 85.21, 83.40(d, J= 7.7 Hz), 80.29, 75.04, 69.61, 66.24, 54.90, 50.51, 37.36, 28.35,25.47, 21.71 (d, J = 5.4 Hz), 21.08 (d, J = 4.9 Hz), 9.98, 1.09. DL-22 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 3,5-dimethoxybenzoate

[0033] 1 H NMR (400 MHz, Chloroform- d ) δ 7.87 (d, J = 6.2 Hz, 1H), 7.35 –7.28 (m, 2H), 7.24 (d, J = 1.1 Hz, 1H), 7.23 (q, J = 1.2 Hz, 1H), 7.18 – 7.11(m, 3H), 6.67 (t, J = 2.4 Hz, 1H), 6.30 (ddd, J = 9.0, 5.3, 1.7 Hz, 1H), 5.48(dt, J = 6.8, 1.7 Hz, 1H), 5.01 (p, J = 6.2 Hz, 1H), 4.51 – 4.38 (m, 2H),4.32 (p, J = 2.5 Hz, 1H), 4.05 – 3.91 (m, 1H), 3.83 (s, 6H), 2.57 (dd, J =14.9, 6.0 Hz, 1H), 2.04 – 1.96 (m, 1H), 1.38 (d, J = 7.0 Hz, 3H), 1.23 (d, J = 7.4 Hz, 6H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.95 (d, J = 7.2 Hz),165.93, 160.87, 157.02, 156.76, 150.43 (d, J= 6.9 Hz), 148.88, 141.94,139.57, 130.87, 129.92, 125.46, 124.18, 123.84, 120.40 (d, J = 4.7 Hz),107.51, 106.21, 85.30, 83.47 (d, J = 7.2 Hz), 75.06, 69.62, 66.30 (d, J = 5.3Hz), 55.74, 50.55, 37.71, 29.77, 29.43, 21.72 (d, J = 4.9 Hz), 21.13 (d, J =4.8 Hz). DL-24 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 4-methoxybenzoate

[0034] 1 H NMR (400 MHz, Chloroform- d ) δ 7.97 (d, J = 8.9 Hz, 2H), 7.88 (d, J = 6.1 Hz, 1H), 7.31 (t, J = 7.9 Hz, 2H), 7.22 (s, 1H), 7.18 – 7.12 (m, 1H),6.93 (d, J = 8.9 Hz, 2H), 6.31 (ddd, J = 9.1, 5.2, 1.7 Hz, 1H), 5.46 (d, J =6.6 Hz, 1H), 5.01 (p, J = 6.3 Hz, 1H), 4.47 – 4.43 (m, 2H), 4.31 (t, J = 2.5Hz, 1H), 4.04 – 3.94 (m, 1H), 3.87 (s, 3H), 3.84 – 3.79 (m, 1H), 2.58 – 2.51(m, 1H), 1.99 – 1.90 (m, 1H), 1.38 (d, J = 7.0 Hz, 3H), 1.22 (d, J = 5.8 Hz,6H). 13C NMR (100 MHz, Chloroform-d) δ 173.00, 172.92, 165.81, 164.04, 157.07,156.80, 150.48, 150.41, 148.98, 141.95, 139.59, 131.98, 129.90, 125.44,124.20, 123.86, 121.37, 120.44, 120.39, 113.93, 85.31, 83.64, 83.56, 74.54,69.60, 66.32, 55.60, 50.54, 37.75, 21.74, 21.69, 21.16, 21.11. DL-25 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl hexanoate

[0035] 1 H NMR (400 MHz, Chloroform- d ) δ 7.83 (d, J = 6.2 Hz, 1H), 7.31 (t, J = 7.9 Hz, 2H), 7.21 (d, J = 8.8 Hz, 2H), 7.19 – 7.11 (m, 1H), 6.21 (ddd, J =9.2, 5.2, 1.7 Hz, 1H), 5.24 (d, J = 6.8 Hz, 1H), 5.01 (p, J = 6.3 Hz, 1H),4.37 (dq, J = 5.9, 2.6 Hz, 2H), 4.15 (p, J = 2.4 Hz, 1H), 4.01 – 3.91 (m,1H), 3.84 – 3.77 (m, 1H), 2.38 (dd, J = 14.1, 5.2 Hz, 1H), 2.32 (t, J = 7.5Hz, 2H), 1.89 – 1.80 (m, 1H), 1.60 (q, J = 7.3 Hz, 2H), 1.36 (d, J = 7.0 Hz,3H), 1.29 (td, J= 8.5, 7.5, 4.3 Hz, 4H), 1.22 (d, J = 7.4 Hz, 7H), 0.89 (t, J = 6.8 Hz, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ 173.43, 173.04, 172.97,157.36, 157.10, 150.44, 150.37, 149.19, 141.94, 139.58, 129.86, 125.38,124.10, 123.76, 120.38, 120.33, 85.15, 83.44, 83.37, 74.10, 69.53, 66.30,66.25, 53.55, 50.48, 37.53, 34.06, 31.26, 24.47, 22.31, 21.70, 21.65, 20.98,20.93, 13.93. DL-26 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 4-isopropoxybenzoate

[0036] 1 H NMR (400 MHz, Chloroform- d ) δ 7.95 (d, J = 8.9 Hz, 2H), 7.88 (d, J = 6.2 Hz, 1H), 7.31 (t, J = 7.9 Hz, 2H), 7.25 (s, 1H), 7.24 – 7.21 (m, 1H),7.18 – 7.12 (m, 1H), 6.89 (d, J = 8.9 Hz, 2H), 6.30 (ddd, J = 9.0, 5.1, 1.7Hz, 1H), 5.01 (p, J = 6.2 Hz, 1H), 4.64 (p, J = 6.0 Hz, 1H), 4.47 – 4.42 (m,2H), 4.31 (p, J = 2.3 Hz, 1H), 4.04 – 3.95 (m, 1H), 3.88 – 3.80 (m, 1H), 2.54(dd, J= 13.8, 5.5 Hz, 1H), 1.94 (ddd, J = 14.2, 9.1, 6.5 Hz, 1H), 1.39 –1.34 (m, 9H), 1.23 (s, 3H), 1.21 (s, 3H). 13 C NMR (100 MHz, Chloroform- d ) δ172.95 (d, J = 7.6 Hz), 165.84, 162.56, 157.05, 156.78, 150.45 (d, J = 6.7Hz), 148.95, 139.58, 132.00, 129.90, 125.44, 124.21, 123.87, 120.84, 120.42(d, J = 4.7 Hz), 115.29, 85.32, 83.62 (d, J = 7.4 Hz), 74.46, 70.26, 69.60,66.37, 50.54, 37.76, 21.94, 21.72 (d, J = 4.7 Hz), 21.14 (d, J = 4.8 Hz). DL-27 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 4-(trifluoromethyl)benzoate

[0037] 1 H NMR (400 MHz, Methanol- d 4) δ 8.22 (d, J = 8.1 Hz, 2H), 7.86 (d, J =6.5 Hz, 1H), 7.81 (d, J = 8.2 Hz, 2H), 7.33 (t, J = 7.8 Hz, 2H), 7.26 – 7.22(m, 2H), 7.16 (t, J = 7.3 Hz, 1H), 6.27 (ddd, J = 7.6, 5.6, 1.6 Hz, 1H), 5.56(d, J = 6.5 Hz, 1H), 4.96 (p, J = 6.3 Hz, 1H), 4.43 (h, J = 4.7 Hz, 3H), 3.92(dq, J = 10.1, 7.1 Hz, 1H), 2.59 (ddd, J = 14.5, 5.7, 1.8 Hz, 1H), 2.25 (ddd,J = 14.8, 8.7, 6.7 Hz, 1H), 1.33 (d, J = 7.1 Hz, 3H), 1.20 (dd, J = 6.3, 1.6Hz, 6H). DL-28 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl isobutyrate

[0038] 1 H NMR (400 MHz, Methanol- d 4) δ 7.81 (d, J = 6.5 Hz, 1H), 7.33 (t, J =7.9 Hz, 2H), 7.26 – 7.21 (m, 2H), 7.19 – 7.14 (m, 1H), 6.15 (d, J = 7.2 Hz,1H), 5.28 – 5.24 (m, 1H), 4.96 (p, J = 6.3 Hz, 1H), 4.34 (dd, J = 6.0, 3.1Hz, 2H), 4.20 (q, J = 2.8 Hz, 1H), 3.95 – 3.85 (m, 1H), 2.59 (p, J = 7.0 Hz,1H), 2.37 (ddd, J = 14.3, 5.6, 1.7 Hz, 1H), 2.10 (ddd, J= 14.7, 8.7, 6.6 Hz,1H), 1.32 (d, J = 7.1 Hz, 3H), 1.20 (dd, J = 6.3, 1.5 Hz, 6H), 1.16 (dd, J =7.0, 1.2 Hz, 6H). 13 C NMR (101 MHz, Chloroform- d ) δ 176.75, 172.95 (d, J = 7.2Hz), 157.21, 156.94, 150.41 (d, J = 6.5 Hz), 149.06, 141.94, 139.57, 129.88,125.42, 124.13, 123.79, 120.38 (d, J = 4.7 Hz), 85.18, 83.52 (d, J = 7.4 Hz),74.11, 69.58, 66.30 (d, J = 5.2 Hz), 50.50, 37.57, 33.77, 21.70 (d, J = 4.8Hz), 21.06 (d, J = 4.8 Hz), 18.85. DL-30 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl (3S,5S,7S)-adamantane-1-carboxylate

[0039] 1 H NMR (400 MHz, Methanol- d 4) δ 7.81 (d, J = 6.5 Hz, 1H), 7.33 (t, J =7.9 Hz, 2H), 7.25 – 7.20 (m, 2H), 7.17 (t, J = 7.3 Hz, 1H), 6.14 (ddd, J =8.8, 5.5, 1.6 Hz, 1H), 5.26 – 5.19 (m, 1H), 4.96 (p, J = 6.3 Hz, 1H), 4.33(dd, J= 6.0, 3.1 Hz, 2H), 4.18 – 4.14 (m, 1H), 3.95 – 3.85 (m, 1H), 2.34(ddd, J = 14.3, 5.6, 1.7 Hz, 1H), 2.15 – 2.05 (m, 1H), 2.00 (s, 3H), 1.90 (d, J = 2.9 Hz, 6H), 1.81 – 1.68 (m, 6H), 1.34 – 1.31 (m, 3H), 1.20 (dd, J = 6.3,1.6 Hz, 6H). 13 C NMR (101 MHz, Chloroform- d ) δ 177.28, 172.96 (d, J = 7.2 Hz),157.22, 156.96, 150.42 (d, J = 6.7 Hz), 149.03, 141.92, 139.56, 129.88,125.41, 124.17, 123.83, 120.39 (d, J = 4.7 Hz), 85.22, 83.66 (d, J = 7.6 Hz),73.95, 69.57, 66.36 (d, J = 5.3 Hz), 50.50, 40.63, 39.28, 38.69, 37.60,36.40, 27.83, 21.71 (d, J = 5.2 Hz), 21.09 (d, J = 4.7 Hz). DL-42 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 4-acetoxybenzoate

[0040] 1 H NMR (400 MHz, Chloroform- d ) δ 7.82 (d, J = 6.2 Hz, 1H), 7.76 (dd, J = 7.9, 1.7 Hz, 1H), 7.68 – 7.62 (m, 1H), 7.37 – 7.30 (m, 3H), 7.24 – 7.16(m, 4H), 6.16 (td,J = 6.1, 1.6 Hz, 1H), 5.00 (p, J = 6.3 Hz, 1H), 4.47 –4.41 (m, 1H), 4.34 (dd, J = 7.3, 3.0 Hz, 2H), 4.02 (dd, J = 4.8, 2.3 Hz, 1H),3.94 (dd, J = 9.7, 5.6 Hz, 2H), 2.42 – 2.34 (m, 1H), 2.31 (s, 3H), 2.12 –2.03 (m, 1H), 1.36 – 1.33 (m, 3H), 1.22 (dd, J = 6.2, 2.9 Hz, 6H). 13 C NMR(100 MHz, Chloroform- d ) δ 173.07 (d, J = 6.9 Hz), 169.05, 164.54, 155.96,155.69, 151.39, 150.42 (d, J = 6.6 Hz), 147.58, 141.28, 138.90, 136.15,132.37, 129.98, 126.69, 125.47, 124.86, 120.11 (d, J = 4.8 Hz), 85.56, 85.02(d, J = 7.0 Hz), 69.97, 69.77, 65.77 (d, J = 5.2 Hz), 50.50, 40.25, 21.71 (d, J = 6.4 Hz), 21.07 – 20.69 (m). DL-44 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)- yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl 3,5-difluorobenzoate

[0041] 1 H NMR (400 MHz, Chloroform- d ) δ 7.87 (d, J = 6.2 Hz, 1H), 7.55 –7.51 (m, 2H), 7.35 – 7.29 (m, 2H), 7.23 (dd, J= 7.6, 1.2 Hz, 2H), 7.16 (t, J = 6.8 Hz, 1H), 7.06 (tt, J = 8.5, 2.4 Hz, 1H), 6.29 (ddd, J = 9.1, 5.3, 1.7Hz, 1H), 5.54 – 5.49 (m, 1H), 5.03 (h, J = 6.3 Hz, 1H), 4.44 (dd, J = 6.5,2.4 Hz, 2H), 4.32 (s, 1H), 4.03 – 3.94 (m, 1H), 3.81 – 3.71 (m, 1H), 2.56(ddd, J = 14.2, 5.3, 1.3 Hz, 1H), 2.00 (d, J = 9.0 Hz, 1H), 1.38 (d, J = 7.1Hz, 3H), 1.23 (dd, J = 6.3, 0.9 Hz, 6H). 13 C NMR (100 MHz, Chloroform- d ) δ172.99 (d, J = 6.8 Hz), 164.36 – 163.68 (m), 161.66 (d, J = 11.7 Hz), 156.98(d, J = 27.5 Hz), 150.40 (d, J = 6.7 Hz), 149.00, 141.99, 139.63, 132.22,129.92, 125.48, 124.08, 123.74, 120.35 (d, J = 4.7 Hz), 112.94 (d, J = 26.9Hz), 109.52, 109.27, 109.02, 85.25, 83.29 (d, J = 7.4 Hz), 75.78, 69.64,66.23, 50.55, 37.62, 21.71 (d, J = 5.2 Hz), 21.23 – 20.85 (m). DL-45 (2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl) oxy)methyl)tetrahydrofuran-3-yl cyclopropanecarboxylate

[0042] 1 H NMR (400 MHz, Chloroform- d ) δ 7.83 (d, J = 6.2 Hz, 1H), 7.30 (dd,J = 8.6, 7.2 Hz, 2H), 7.23 – 7.20 (m, 2H), 7.15 (ddd, J = 8.2, 7.1, 1.1 Hz,1H), 6.23 (ddd, J = 9.1, 5.2, 1.7 Hz, 1H), 5.24 (dt, J = 6.5, 1.6 Hz, 1H),5.01 (h, J = 6.3 Hz, 1H), 4.42 – 4.30 (m, 2H), 4.18 (p, J = 2.4 Hz, 1H), 3.96(ddt, J = 16.1, 9.0, 7.0 Hz, 1H), 3.74 (dd, J = 11.4, 9.4 Hz, 1H), 2.44 –2.37 (m, 1H), 1.84 (ddd, J = 14.1, 9.1, 6.5 Hz, 1H), 1.64 (dd, J = 7.9, 4.6Hz, 1H), 1.36 (d, J = 7.0 Hz, 3H), 1.22 (d, J = 6.2 Hz, 6H), 1.01 (tt, J =4.3, 2.3 Hz, 2H), 0.96 – 0.91 (m, 2H). DL-47 isopropyl ((S)-(((2R,3S,5R)-3-acetoxy-5-(5-fluoro-2,4-dioxo-3, 4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-2-yl)methoxy)(phenoxy) phosphoryl)-L-alaninate

[0043] 1 H NMR (400 MHz, Chloroform- d) δ 7.83 (d, J = 6.2 Hz, 1H), 7.34 –7.28 (m, 2H), 7.22 (dt, J = 7.5, 1.3 Hz, 2H), 7.15 (ddt, J = 8.2, 7.3, 1.1Hz, 1H), 6.21 (ddd, J = 9.1, 5.2, 1.7 Hz, 1H), 5.24 (dt, J = 6.6, 1.7 Hz,1H), 5.01 (p, J = 6.3 Hz, 1H), 4.42 – 4.30 (m, 2H), 4.17 (p, J = 2.5 Hz, 1H),4.00 – 3.90 (m, 1H), 3.76 (dd, J = 11.4, 9.4 Hz, 1H), 2.39 (ddd, J = 14.1,5.3, 1.4 Hz, 1H), 2.09 (s, 3H), 1.86 (ddd, J = 14.0, 9.1, 6.5 Hz, 1H), 1.36(d, J = 7.0 Hz, 3H), 1.22 (dd, J = 6.3, 1.2 Hz, 6H). DL-48(2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl)- 2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl)amino)(phenoxy)phosphoryl)oxy) methyl)tetrahydrofuran-3-yl 4-fluorobenzoate 1 H NMR (400 MHz, Chloroform- d ) δ 8.07 – 8.01 (m, 2H), 7.88 (d, J =6.2 Hz, 1H), 7.32 (dd, J = 8.6, 7.2 Hz, 2H), 7.25 – 7.22 (m, 2H), 7.19 – 7.11(m, 3H), 6.30 (ddd, J = 9.1, 5.2, 1.7 Hz, 1H), 5.49 (dt, J = 6.6, 1.6 Hz,1H), 5.02 (p, J = 6.3 Hz, 1H), 4.49 – 4.42 (m, 2H), 4.32 (q, J = 2.4 Hz, 1H),3.99 (tq, J = 9.0, 7.0 Hz, 1H), 3.80 (dd, J= 11.4, 9.4 Hz, 1H), 2.56 (ddd, J = 14.2, 5.3, 1.3 Hz, 1H), 1.99 (ddd, J = 14.2, 9.1, 6.5 Hz, 1H), 1.38 (d, J =7.1 Hz, 3H), 1.23 (d, J = 6.3 Hz, 6H). DL-51 5-(tert-butyl) 1-((2R,3S,5R)-5-(5-fluoro-2,4-dioxo-3,4- dihydropyrimidin-1(2H)-yl)-2-((((S)-(((S)-1-isopropoxy-1-oxopropan-2-yl) amino)(phenoxy)phosphoryl)oxy)methyl)tetrahydrofuran-3-yl) (tert- butoxycarbonyl)-L-glutamate

[0044] 1 H NMR (400 MHz, Chloroform- d ) δ 7.82 (d, J = 6.2 Hz, 1H), 7.34 –7.27 (m, 2H), 7.21 (d, J = 8.8 Hz, 2H), 7.17 – 7.13 (m, 1H), 6.20 (ddd, J =9.1, 5.2, 1.7 Hz, 1H), 5.01 (p, J = 6.3 Hz, 1H), 4.38 – 4.35 (m, 2H), 4.24 –4.16 (m, 2H), 4.00 – 3.92 (m, 1H), 3.79 (t, J = 10.4 Hz, 1H), 2.51 – 2.41 (m,2H), 2.40 – 2.33 (m, 1H), 2.15 (dd, J = 13.7, 6.6 Hz, 1H), 1.92 – 1.81 (m,2H), 1.46 (s, 9H), 1.43 (s, 9H), 1.36 (d, J = 7.0 Hz, 3H), 1.22 (d, J = 6.2Hz, 6H). 13 C NMR (100 MHz, Chloroform- d ) δ 172.91 (d, J = 7.3 Hz), 172.50,171.33, 157.02, 155.55, 150.40 (d, J= 6.5 Hz), 148.82, 139.53, 129.90,125.45, 123.85, 120.39 (d, J = 4.6 Hz), 85.22, 83.35 (d, J = 7.3 Hz), 82.50,80.06, 74.48, 69.61, 66.25, 53.23, 50.52, 37.48, 30.14, 28.39, 28.09, 21.71(d, J = 5.0 Hz), 21.11 (d, J = 4.8 Hz). Example 3

[0045] In vitro anti-proliferative activity of the compound against liver cancer cells and comparison with normal cells (IC50 value determined by CCK-8 assay).

[0046] (1) Experimental methods HepG2 and Huh7 human liver cancer cells were collected and placed in 96-well plates. Compounds of various concentration gradients were added, resulting in a final volume of 200 mL per well. Cells were incubated at 37°C for 48 hours, followed by the addition of 20 mL of CCK-8 to each well and incubation at 37°C for another 4 hours. The supernatant was removed, and the residue was dissolved in DMSO. Absorbance was recorded at 490 nm using SpectraMax to calculate the cell proliferation inhibition rate and IC50 of the compounds. 50 value.

[0047] IR (%) = [1 - (OD) Drug -OD Blank ) / (OD Control -OD Blank )]×100.

[0048] In this experiment, compound IC 50 The values ​​were calculated and analyzed using the software GraphPad Prism 7.0.

[0049] Table 1. Half-maximal inhibitory concentration (IC50) of the compounds against HepG2 and Huh7 liver cancer cells. 50

[0050] The experimental results are shown in Table 1, where "-" indicates that the assay was not performed. The results show that the inhibitory activity of the tested compounds against HepG2 cells was superior to that of the 5-FU group. Furthermore, the inhibitory activity of compounds DL-21, DL-25, and DL-44 against liver cancer cells was superior to that of compound 11b disclosed in the prior art (CN 118772216 A). In the HepG2 cell line, compound DL-44 showed an activity more than 100-fold higher than that of the 5-FU group, indicating that compounds DL-21, DL-25, DL-44, and this series of prodrugs have potential applications in the treatment of liver cancer-related diseases. Example 4

[0051] Carboxypeptidase Y (EC 3.4.16.1) assay.

[0052] First, dissolve compound DL-44 (5 mg) in acetone-d6 (0.15 mL), then add 0.30 mL of Trizma buffer (pH 7.6). Use at 25°C. 31 P NMR data were used as a control. Then, carboxypeptidase Y (0.1 mg dissolved in 0.15 mL Trizma) was added to the sample solution, and the NMR data were analyzed at specific time points. 31 The P NMR (at 25°C) was measured and the spectrum was recorded by 64 scans. 31 Data recorded by pNMR were analyzed using the MNova program. Carboxypeptidase Y and Trizma buffer were purchased from Beijing Jinming Biotechnology Co., Ltd.

[0053] The study used carboxypeptidase Y. Compound DL-44 and carboxypeptidase Y were dissolved in acetone-d6 and Trizma buffer (pH 7.6) and used at 25°C. 31 P NMR data was recorded continuously for 80 minutes, with scans acquired every 10 minutes. The recorded spectrum is as follows: Figure 1 As shown, compound DL-44 was rapidly hydrolyzed into intermediate metabolites a (δ 5.26 ppm) and b (7.44 ppm) lacking ester motifs. After 80 minutes, compound DL-44 was mainly metabolized into intermediate b. This indicates that compound DL-44 can undergo initial activation under the action of carboxypeptidase Y, releasing intermediate metabolite b to participate in subsequent biological metabolic activation, thus enabling the final delivery of the effective form of 5-fluorodeoxyuridine monophosphate in tumor cells. Example 5

[0054] In vitro metabolic stability assay.

[0055] Compound DL-44 (1 mg / mL in acetonitrile) was incubated in rat plasma at 37°C for 0 min, 0.5 h, 1 h, 2 h, 4 h, 8 h, 24 h, and 48 h. At each time point, each sample solution was diluted and quenched with 600 μL of acetonitrile, shaken for 30 s, and centrifuged at 12000 rpm for 10 min. The solution was then incubated in a container equipped with C... 18 Samples were analyzed by HPLC on an Agilent 1260 HPLC instrument with a column (2.1 mm × 100 mm, 3.5 μm). Compounds were eluted with MeOH / phosphate buffered saline over 20 minutes. Absorbance was measured at 268 nm at a flow rate of 1 mL / min.

[0056] The in vitro metabolic stability of compound DL-44 was studied, and the results are as follows: Figure 2 As shown, compound DL-44 is stable in rat plasma, and more than 50% of the original drug residue was observed after 48 hours of culture. Example 6

[0057] Inhibitory effects of compounds DL-21 and DL-44 on the proliferation of human hepatocellular carcinoma cells in situ in nude mice. Twenty-five Balb / c nude mice were surgically opened, and HepG2 cells (2 × 10⁻⁶ cells / year) were introduced into the mice. 6 50 μL of DL-21 (cells / mL) was seeded into the right lobe of the liver. Three days later, mice were randomly divided into four groups and injected with the drug via the tail vein. DL-21 and DL-44 were administered at a dose of 45 mg / kg / 2 days, every two days, with sorafenib as a control. The vector consisted of 8% Tween 80, 2% dimethyl sulfoxide, and 90% saline. On day 21, mice were euthanized, the liver was dissected, and the length, width, and height of the hepatocellular carcinoma tumor were measured. The tumor data were analyzed using the DRAP package software in R to calculate the inhibition rate. (See...) Figure 3 The results showed that the inhibition rates of DL-21 and DL-44 were 55.84% and 53.33%, respectively, which were significantly better than those of the control group sorafenib (38.98%).

[0058] Although embodiments of the present invention have been shown and described above, it is understood that these embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions, and alterations to the above embodiments within the scope of the present invention without departing from its principles and spirit. The scope of protection of the present invention is defined by the claims and their equivalents.

Claims

1. A compound as shown in general formula I, or a pharmaceutically acceptable salt, hydrate, solvate, polymorph, or stereoisomer thereof: ， in, R1 is selected from: C1-C 12 Chain-like fatty acyl groups, C3-C 12 The alkyl acyl group, substituted or unsubstituted arcarboxyl group, or amino acyl group, wherein the substituent of the arcarboxyl group is selected from halogen, C1-C4 alkyl, C1-C4 alkoxy, acyloxy, or cyano.

2. The compound according to claim 1, characterized in that, R1 is selected from: isobutyryl, n-hexanoyl, adamantyl, p-methoxybenzoyl, 3,5-dimethoxybenzoyl, 3,5-difluorobenzoyl, p-acetoxybenzoyl, p-isopropoxybenzoyl, L-alanyl, L-valine, Boc-protected L-meththioyl, or Boc-protected L-glutamyl.

3. The compound according to claim 1, characterized in that, The compound is selected from one of the following fifteen compounds: 。 4. A pharmaceutical composition, characterized in that, The compound comprises a therapeutically effective amount of any one of claims 1-3, or a pharmaceutically acceptable salt, hydrate, solvate, polymorph or stereoisomer thereof, and a pharmaceutically acceptable excipient.

5. The pharmaceutical composition according to claim 4, characterized in that, The dosage form of the pharmaceutical composition is an injection, an oral preparation, or a topical dosage form.

6. Use of the compound of any one of claims 1-3, or a pharmaceutically acceptable salt, hydrate, solvate, polymorph or stereoisomer thereof, or the pharmaceutical composition of claim 4 or 5, in the preparation of a medicament for treating liver cancer.

7. The use according to claim 6, characterized in that, The liver cancer mentioned is primary liver cancer.

8. The use according to claim 6 or 7, characterized in that, The drug is used in combination with one or more other antitumor drugs.

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