CN119874643B

CN119874643B - Paclitaxel prodrug, and preparation method and application thereof

Info

Publication number: CN119874643B
Application number: CN202510362932.9A
Authority: CN
Inventors: 余文颖; 耿永安
Original assignee: China Pharmaceutical University
Current assignee: China Pharmaceutical University
Priority date: 2025-03-26
Filing date: 2025-03-26
Publication date: 2025-06-27
Anticipated expiration: 2045-03-26
Also published as: CN119874643A

Abstract

The present invention discloses a paclitaxel prodrug and a preparation method and application thereof. The compound of the present invention and its pharmaceutically acceptable salts and esters can block toxic sites through ROS-responsive linking arms, highly respond to the tumor microenvironment, play a selective killing and release effect, and have low toxicity to normal cells. The photosensitive fragment can be used to promote the release of the prodrug, amplify the chemotherapy effect, and can also be used as a self-imaging fragment to play a living imaging function, which can be used for the precise treatment of breast cancer. Therefore, the paclitaxel prodrug of the present invention can be used to prepare drugs for preventing and/or treating diseases related to breast cancer.

Description

Paclitaxel prodrug, and preparation method and application thereof

Technical Field

The invention relates to the field of pharmaceutical chemistry, in particular to a taxol prodrug, a preparation method and application thereof.

Background

Paclitaxel (PTX) is an antitumor tricyclic diterpenoid compound with a complex structure, which is originally separated from the stem and leaf of Taxus chinensis. PTX is a white crystalline powder, odorless, tasteless, poorly water-soluble, but readily soluble in organic solvents, and has a molecular formula of C ₄₇H₅₁NO₁₄ and a molecular weight of 853.92. From its discovery, various studies and developments have been carried over. With the intensive progress of the research, the action mechanism of paclitaxel is elaborated that PTX can bind and stabilize beta-tubulin in microtubules, lead to the formation of parallel microtubules and inhibit cell division in the G2-M phase of the cell cycle, and at the same time, paclitaxel can also cause apoptosis in various different ways, and involves the activation and regulation of various molecular signaling pathways, including unbalance of Bcl-2 family proteins, activation of Caspase cascade reaction, reduction of mitochondrial membrane potential, and cleavage of DNA.

Paclitaxel is one of the most popular natural antitumor drugs. Because of its broad-spectrum antitumor activity, paclitaxel has been widely used clinically for the treatment of breast cancer, ovarian cancer and part of head and neck squamous cell carcinoma and lung cancer. However, because of the poor water solubility of paclitaxel, it has high cardiac and neurotoxicity, and is prone to bone marrow suppression and drug resistance, and its clinical application is limited. Therefore, research to increase the effectiveness and the pharmaceutical property of paclitaxel has been a hot spot and a difficult point of paclitaxel development.

Prodrugs, broadly refers to substances which, by different modification means, convert a pharmaceutically active compound, i.e. the parent drug, to a substance which is inactive in vitro but which, in vivo, is capable of re-releasing the active drug molecule via a specific conversion pathway.

Changes in redox status within tumor cells during tumorigenesis and progression can regulate signaling and death regulation. ROS is a generic term for a class of free radicals, a class of byproducts that are produced during cellular metabolism due to changes in redox state. Hydrogen peroxide, singlet oxygen, superoxide anions, hydroxyl radicals, and the like all belong to ROS. In normal cells, it can be cleared rapidly, while the level of ROS is much higher than in normal cells due to the higher level of redox in tumor cells. Many studies have shown that tumor cells produce ROS levels (> 10 mM) at concentrations of tens or even thousands of times higher than normal cells (< 1 mM).

Photodynamic therapy is a new method for treating tumors by using photosensitizers and laser activated photosensitizers. The irradiation of the focus part with specific wavelength can activate the photosensitive medicine selectively gathered in the focus tissue to trigger photochemical reaction to destroy focus. Photosensitizing drugs in photodynamic therapy (PDT) transfer energy to surrounding oxygen, generating singlet oxygen that is very active. The singlet oxygen can perform oxidation reaction with the nearby biomacromolecule to generate cytotoxicity so as to kill the lesion cells. PDT has the advantage over conventional therapies that it enables accurate and effective treatment with minimal side effects.

Thus, there is an urgent need for prodrugs that combine photodynamic therapy with chemotherapy.

Disclosure of Invention

The invention aims to provide a paclitaxel prodrug for combination of photodynamic therapy and chemotherapy, and also aims to provide a preparation method of the paclitaxel prodrug and application of the paclitaxel prodrug.

The technical proposal is that a compound shown as a general formula I or a general formula II or pharmaceutically acceptable salt thereof,Wherein R ₁ is selected from hydrogen, C ₁-C₆ alkyl;

Wherein R ₂ is selected from C ₁-C₆ alkyl.

Further, the R ₁ is selected from hydrogen, C ₁-C₆ n-alkyl.

Further, the R ₂ is selected from C ₁-C₆ n-alkyl.

Further, the R ₁ is selected from the group consisting of hydrogen, methyl, ethyl, n-propyl, n-butyl, n-pentyl, and n-hexyl.

Further, the R ₂ is selected from the group consisting of methyl, ethyl, n-propyl, n-butyl, n-pentyl, and n-hexyl.

The examples of the present invention provide the following preferred compounds: 。

In some preferred embodiments, the pharmaceutically acceptable salts include, but are not limited to, acid addition salts of the compounds of formula I or formula II with hydrochloric acid, hydrobromic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, citric acid, tartaric acid, lactic acid, pyruvic acid, acetic acid, maleic acid or succinic acid, fumaric acid, salicylic acid, phenylacetic acid, mandelic acid, and also acid salts of the compounds of formula I or formula II with inorganic bases.

The compounds of the general formula I or the general formula II according to the invention can also exist in the form of salts or esters thereof, which are converted in vivo into compounds of the general formula I or the general formula II. For example, within the scope of the present invention, the compounds of the present invention are converted to pharmaceutically acceptable salt forms and used in salt form according to procedures known in the art.

In another aspect, the present invention provides a process for the preparation of the above-described compound or a pharmaceutically acceptable salt thereof,

Wherein R ₁ is selected from C ₁-C₆ alkyl.

Wherein R ₂ is selected from C ₁-C₆ alkyl.

In another aspect, the invention provides a pharmaceutical composition comprising a compound as described above, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers.

The pharmaceutical compositions of the present invention may be administered in a variety of known ways, such as orally, parenterally, by inhalation spray, or via an implanted reservoir. The pharmaceutical composition of the invention can be administered alone or in combination with other drugs. The oral composition may be any orally acceptable dosage form including, but not limited to, tablets, capsules, emulsions, suspensions, dispersions, and solutions. Common pharmaceutically acceptable carriers or excipients include stabilizers, diluents, surfactants, lubricants, antioxidants, binders, colorants, fillers, emulsifiers, and the like.

Sterile injectable compositions can be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. Pharmaceutically acceptable carriers and solvents that can be used include water, mannitol, sodium chloride solution, and the like.

The actual dosage level of the active ingredient in the pharmaceutical compositions of the present invention may be varied to obtain an amount of active ingredient that is effective to achieve the desired therapeutic response for the particular patient, composition and mode of administration, and that is non-toxic to the patient. The dosage level selected will depend on a variety of factors including the activity of the particular compound of the invention or salt thereof employed, the route of administration, the time of administration, the rate of excretion of the particular composition employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular composition employed, the age, sex, weight, general health and past medical history of the patient being treated, and like factors well known in the medical arts.

In another aspect, the invention provides an application of the compound or pharmaceutically acceptable salt thereof in preparing a medicament for preventing and/or treating tumor diseases.

Preferably, the neoplastic disease is breast cancer.

Compared with the prior art, the invention has the advantages that toxic sites can be blocked by the ROS responsive connecting arm, the effect of selectively killing and releasing is achieved in high response to tumor microenvironment, the normal cytotoxicity is low, the photosensitive fragment can be used for promoting the release of the prodrug, amplifying the chemotherapy curative effect, can be used as an autoradiography fragment, plays a living body imaging function, and can be used for the accurate treatment of breast cancer.

Drawings

FIG. 1 is a graph of compound 2 in vitro modeling of release of different ROS levels over time;

FIG. 2 is an in vivo image of a12 h compound 2 mouse, with the left side being the oral group and the right side being the tail vein group;

FIG. 3 is an in vivo image of 24h Compound 2 mice, with the left side being the oral group and the right side being the tail vein group;

FIG. 4 is a plot of in vivo images of 36h Compound 2 mice, with the left side being the oral group and the right side being the tail vein group;

FIG. 5 is a photograph of a 48h compound 2 in vivo image of mice, with the left side being the oral group and the right side being the tail vein group;

fig. 6 is an in vivo image of 72h compound 2 mice, with the left side being the oral group and the right side being the tail vein group.

Detailed Description

The process for the preparation of the compounds of the general formula I or of the general formula II according to the invention is described below in connection with specific examples, which, however, do not constitute any limitation of the invention. The compounds of the present invention may also be conveniently prepared by optionally combining the various synthetic methods described in this specification or known in the art, such combinations being readily apparent to those skilled in the art to which the present invention pertains.

The starting materials, reagents, etc. used in the examples of the present invention are all commercially available. The invention can be prepared into salt form by adopting a salt forming method commonly used in the field, for example, the compound is dissolved in ethanol hydrochloride at room temperature to react to form hydrochloride, or benzenesulfonic acid is added into the compound to react to form benzenesulfonate.

The design concept of the compounds of the general formula I and the general formula II is as follows, paclitaxel is coupled with photosensitizer 5-aminolevulinic acid and derivatives thereof through different redox-responsive connecting arms, and a prodrug combined with photodynamic therapy and chemotherapy is generated. In the compound of the invention, the connection site of the taxol is taxol C-2', the site is connected with one end of a redox-responsive connecting arm, and the other end of the redox-responsive connecting arm is connected with the amino group of 5-aminolevulinic acid and derivatives thereof. Wherein the redox responsive linkage arm is a disulfide or thioether bond. The photosensitizer 5-aminolevulinic acid and its derivatives include 5-aminolevulinic acid, methyl 5-aminolevulinate, ethyl 5-aminolevulinate, propyl 5-aminolevulinate, butyl 5-aminolevulinate, pentyl 5-aminolevulinate and hexyl 5-aminolevulinate.

The thioether prodrugs of the invention are shown in the general formula I:

wherein R ₁ is selected from hydrogen and C ₁-C₆ is n-alkyl.

The disulfide bond prodrug of the invention is shown in a general formula II:

Wherein R ₂ is selected from C ₁-C₆ n-alkyl.

The synthetic route of the compound shown in the general formula I is as follows:

Dropwise adding thionyl chloride into an alcohol (R ₁ -OH) solution in an ice bath, then adding 5-aminolevulinic acid hydrochloride (5-ALA HCl) for reaction, and obtaining a 5-aminolevulinate derivative (5-ALA-R ₁) after the reaction is finished;

Paclitaxel (PTX), thiohydroxy acetic anhydride, triethylamine (TEA) and 4-Dimethylaminopyridine (DMAP) are added into Dichloromethane (DCM) to react, and the reaction is finished to obtain an intermediate PTX-S-COOH;

Adding an intermediate PTX-S-COOH, 2- (7-aza-benzotriazol) -N, N, N ', N' -tetramethyl urea Hexafluorophosphate (HATU), DIPEA and a 5-aminolevulinate derivative (5-ALA-R ₁) into dichloromethane, stirring and reacting to obtain a target product after the reaction is finished;

wherein R ₁ is selected from C ₁-C₆ n-alkyl.

When R ₁ is hydrogen, the synthetic route of the compound shown in the general formula I is as follows:

Adding an intermediate PTX-S-COOH, N-hydroxysuccinimide (NHS) and 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI) into dichloromethane for reaction, and obtaining an active ester intermediate after the reaction is finished;

Adding the active ester intermediate, 5-aminolevulinic acid hydrochloride and N, N-Diisopropylethylamine (DIPEA) into a dichloromethane solution for reaction, and obtaining a target product after the reaction is finished.

The synthetic route of the compound shown in the general formula II is as follows:

2,2' -dithio diacyl diacetic acid, acetyl chloride and nitrogen are added for protection reaction, and the target product 1,4, 5-oxo dithiophene-2, 7-diketone (DTDPA) is obtained after the reaction is finished;

Paclitaxel (PTX), DTDPA, triethylamine and DMAP are added into DCM to react overnight, and an intermediate PTX-S-S-COOH is obtained after the reaction is finished;

The intermediate PTX-S-S-COOH, 2- (7-aza-benzotriazol) -N, N, N ', N' -tetramethyl urea Hexafluorophosphate (HATU), DIPEA and 5-amino levulinate derivative (5-ALA-R ₂) are added into dichloromethane, and stirred for reaction, and the target product is obtained after the reaction is finished, wherein R ₂ is selected from C ₁-C₆ N-alkyl.

Synthesis of intermediates

The synthesis of the intermediate related to the embodiment of the invention is as follows:

1. Synthesis of 2- (((1S, 2R) -1-benzoylamino-3- (((2 aR,4S,4aS,6R,9S,11S,12 aR,12 bS) -6,12 b-diacetoxy-12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-2 a,3, 4a,5,6,9,11,12 a,12 b-dodecahydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxo-9-yl) oxy) -3-oxo-1-phenylpropane-2-oxoethoxy) -2-oxoethyl) thioacetic acid (PTX-S-COOH)

50 Mg of paclitaxel, 15 mg of thiohydroxy acetic anhydride, 10. Mu.l of triethylamine, 1.5 mg of 4-dimethylaminopyridine were weighed out and 2 ml of methylene chloride was added. The reaction was at room temperature overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give intermediate PTX-S-COOH,51 mg, as a yellow solid. 88% yield.

¹H NMR (400 MHz, Chloroform-d)δ8.03 – 7.97 (m, 2H), 7.85 – 7.79 (m, 2H), 7.63 (dd,J= 8.2, 0.8 Hz, 1H), 7.58 – 7.49 (m, 2H), 7.49 – 7.40 (m, 6H), 7.40 – 7.33 (m, 2H), 7.27 (ddt,J= 8.2, 6.9, 2.1 Hz, 1H), 6.64 (p,J= 1.0 Hz, 1H), 6.16 (dddt,J= 7.3, 6.2, 2.6, 1.5 Hz, 1H), 5.69 (d,J= 8.8 Hz, 1H), 5.53 (dd,J= 7.6, 0.8 Hz, 1H), 5.53 – 5.46 (m, 1H), 4.82 (ddt,J= 6.4, 3.8, 2.5 Hz, 1H), 4.42 – 4.32 (m, 2H), 4.30 (d,J= 12.2 Hz, 1H), 3.84 (d,J= 11.5 Hz, 1H), 3.77 (dt,J= 8.8, 2.6 Hz, 1H), 3.54 (d,J= 8.4 Hz, 2H), 3.43 (d,J= 1.4 Hz, 2H), 2.45 (s, 1H), 2.40 – 2.31 (m, 2H), 2.25 – 2.16 (m, 6H), 2.02 (ddd,J= 12.3, 7.7, 6.5 Hz, 1H), 1.82 (t,J= 1.2 Hz, 3H), 1.19 (d,J= 1.0 Hz, 5H).

Electrospray mass spectrometry (ESI-MS) calculated C ₅₁H₅₅NO₁₇S (M-H)^- m/z 984.3, found 984.3.

2. Synthesis of ethyl 5-aminolevulinate

600 Μl of thionyl chloride was added dropwise to 8 ml of ethanol in an ice bath at 0deg.C. Stirring for 10min. Deicing 1g of 5-aminolevulinic acid hydrochloride are added. The reaction was carried out at room temperature for 1 hour. Heated to 78 ℃ and refluxed for 2 hours. Cooling, adding diethyl ether, precipitating solid, grinding, and pulping. Filtered with diethyl ether and acetonitrile and washed. 0.85 g of ethyl 5-aminolevulinate is obtained as the target product. White solid. 89% yield .¹H NMR (400 MHz, Chloroform-d)δ4.13 (q,J= 6.6 Hz, 2H), 3.73 (t,J= 6.5 Hz, 2H), 2.90 (q,J= 6.6 Hz, 1H), 2.80 (q,J= 6.5 Hz, 1H), 2.75 – 2.68 (m, 2H), 2.63 – 2.56 (m, 2H), 1.24 (t,J= 6.6 Hz, 3H).ESI-MS calculated C ₇H₁₃NO₃(M+H)⁺ m/z 160.2, found 160.2.

3. Synthesis of propyl 5-aminolevulinate

600 Μl of thionyl chloride was added dropwise to 8 ml of n-propanol in an ice bath at 0deg.C. Stirring for 10min. Deicing 1 g of 5-aminolevulinic acid hydrochloride are added. The reaction was carried out at room temperature for 1 hour. Heated to 98 ℃ and refluxed for 2 hours. Cooling, adding diethyl ether, precipitating solid, grinding, and pulping. Filtered with diethyl ether and acetonitrile and washed. 0.97 g of intermediate propyl 5-aminolevulinate was obtained. White solid. 93.85% yield .¹H NMR (400 MHz, Chloroform-d)δ4.07 (t,J= 5.7 Hz, 2H), 3.73 (t,J= 6.5 Hz, 2H), 2.90 (q,J= 6.6 Hz, 1H), 2.80 (q,J= 6.6 Hz, 1H), 2.75 – 2.68 (m, 2H), 2.63 – 2.56 (m, 2H), 1.77 – 1.67 (m, 2H), 0.97 (t,J= 8.0 Hz, 3H).ESI-MS calculated C ₈H₁₅NO₃(M+H)⁺ m/z 174.2, found 174.2.

4. Synthesis of butyl 5-aminolevulinate

600 Μl of thionyl chloride was added dropwise to 8 ml of n-butanol in an ice bath at 0deg.C. Stirring for 10min. Deicing 1g of 5-aminolevulinic acid hydrochloride are added. The reaction was carried out at room temperature for 1 hour. Heated to 118 ℃ and refluxed for 2 hours. Cooling, adding diethyl ether, precipitating solid, grinding, and pulping. Filtered with diethyl ether and acetonitrile and washed. 1.01 g of intermediate butyl 5-aminolevulinate was obtained. White solid. 90% yield .¹H NMR (400 MHz, Chloroform-d)δ4.07 (t,J= 6.5 Hz, 2H), 3.73 (t,J= 6.5 Hz, 2H), 2.90 (q,J= 6.6 Hz, 1H), 2.80 (q,J= 6.6 Hz, 1H), 2.75 – 2.68 (m, 2H), 2.63 – 2.56 (m, 2H), 1.61 (p,J= 6.8 Hz, 2H), 1.39 (dt,J= 14.0, 7.0 Hz, 2H), 0.97 (t,J= 7.0 Hz, 3H).ESI-MS calculated C ₉H₁₇NO₃(M+H)⁺ m/z 188.2, found 188.2.

5. Synthesis of 5-aminolevulinic acid pentyl ester

600 Μl of thionyl chloride was added dropwise to 8 ml of n-pentanol in an ice bath at 0deg.C. Stirring for 10min. Deicing 1g of 5-aminolevulinic acid hydrochloride are added. The reaction was carried out at room temperature for 1 hour. Heated to 139 ℃ and refluxed for 2 hours. Cooling, adding diethyl ether, precipitating solid, grinding, and pulping. Filtered with diethyl ether and acetonitrile and washed. 1.13 g of intermediate amyl 5-aminolevulinate was obtained. White solid. Yield .¹H NMR (400 MHz, Chloroform-d)δ4.06 (t,J= 5.9 Hz, 2H), 3.73 (t,J= 6.5 Hz, 2H), 2.90 (q,J= 6.6 Hz, 1H), 2.80 (q,J= 6.6 Hz, 1H), 2.75 – 2.68 (m, 2H), 2.63 – 2.56 (m, 2H), 1.71 – 1.62 (m, 2H), 1.42 – 1.31 (m, 4H), 0.95 – 0.85 (m, 3H).ESI-MS calculated 94% C ₁₀H₁₉NO₃(M+H)⁺ m/z 202.3, found 202.3.

6. Synthesis of hexyl 5-aminolevulinate

600 Μl of thionyl chloride was added dropwise to 8 ml of n-hexanol in an ice bath at 0deg.C. Stirring for 10min. Deicing 1g of 5-aminolevulinic acid hydrochloride are added. The reaction was carried out at room temperature for 1 hour. Heated to 159 ℃ and refluxed for 2 hours. Cooling, adding diethyl ether, precipitating solid, grinding, and pulping. Filtered with diethyl ether and acetonitrile and washed. 1.19 g of intermediate hexyl 5-aminolevulinate was obtained. White solid. 93% yield .¹H NMR (400 MHz, Chloroform-d)δ4.06 (t,J= 6.2 Hz, 2H), 3.73 (t,J= 6.5 Hz, 2H), 2.90 (q,J= 6.6 Hz, 1H), 2.80 (q,J= 6.6 Hz, 1H), 2.75 – 2.68 (m, 2H), 2.63 – 2.56 (m, 2H), 1.68 – 1.59 (m, 2H), 1.46 – 1.25 (m, 6H), 0.92 – 0.86 (m, 3H).ESI-MS calculated C ₁₁H21NO₃(M+H)⁺ m/z 216.3, found 216.3.

7. Synthesis of 1,4, 5-oxo-dithiophene-2, 7-dione (DTDPA)

0.5 G of 2,2' -dithiodiacetic acid, 2ml of acetyl chloride are added. The reaction was carried out at room temperature for 2 hours under nitrogen protection. Add 20 ml toluene three times, spin dry, remove solvent. Adding diethyl ether, pulping, and separating out solid. Suction filtration yields 270 mg of the target product 1,4, 5-oxo-dithiophene-2, 7-dione (DTDPA). White solid, yield 47%.

8. Synthesis of 2- (((1S, 2R) -1-benzoylamino-3- (((2 aR,4S,4aS,6R,9S,11S,12 aR,12 bS) -6,12 b-diacetoxy-12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-2 a,3, 4a,5,6,9,11,12 a,12 b-dodecahydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxo-9-yl) oxy) -3-oxo-1-phenylpropane-2-oxoethoxy) -dithio) acetate (PTX-S-S-COOH)

50 Mg of paclitaxel, 15 mg DTDPA,10 μl triethylamine, 1.5 mg DMAP were added to 2ml DCM. The reaction was carried out overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to obtain intermediate PTX-S-S-COOH,40 mg. Yellow solid, 67% yield.

¹H NMR (400 MHz, Chloroform-d)δ8.03 – 7.97 (m, 2H), 7.85 – 7.79 (m, 2H), 7.63 (dd,J= 8.3, 0.9 Hz, 1H), 7.58 – 7.49 (m, 2H), 7.49 – 7.40 (m, 2H), 7.44 (s, 2H), 7.40 – 7.33 (m, 2H), 7.27 (ddt,J= 8.2, 6.9, 2.1 Hz, 1H), 6.64 (p,J= 1.0 Hz, 1H), 6.16 (dddt,J= 7.3, 6.2, 2.6, 1.5 Hz, 1H), 5.69 (d,J= 8.8 Hz, 1H), 5.53 (dd,J= 7.6, 0.7 Hz, 1H), 5.53 – 5.46 (m, 1H), 4.82 (ddt,J= 6.4, 3.8, 2.5 Hz, 1H), 4.42 – 4.32 (m, 2H), 4.30 (d,J= 12.2 Hz, 1H), 3.84 (d,J= 11.5 Hz, 1H), 3.80 – 3.68 (m, 2H), 3.70 – 3.61 (m, 3H), 2.45 (s, 1H), 2.40 – 2.31 (m, 2H), 2.25 – 2.16 (m, 6H), 2.02 (ddd,J= 12.3, 7.7, 6.5 Hz, 1H), 1.82 (t,J= 1.2 Hz, 3H), 1.19 (d,J= 1.0 Hz, 5H).

ESI-MS calculated C ₅₁H₅₅NO₁₇S₂(M-H)^- m/z 1016.3, found 1016.3.

EXAMPLE 1 Synthesis of Compound 1

(3S, 4R) -4- (((2 aR,4S,4aS,6R,9S,11S,12 aR,12 bS) -6,12 b-diacetoxy-12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5-oxo-2 a,3, 4a,5,6,9,10,11,12 a,12 b-dodecahydro-1H-7, 11-methoxycyclodeca [3,4] benzo [1,2-b ] oxa-9-yl) oxycarbonyl) -1,6,10,13-tetraoxo-1, 3-diphenyl-5-oxa-8-thia-2, 11-diazahexadecane-16-oic acid

60 Mg of the intermediate PTX-S-COOH, 8mg of N-hydroxysuccinimide (NHS), 14 mg of 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI) were added to 2 ml of dichloromethane in an ice bath, and after half an hour of reaction, the reaction was resumed at room temperature for 24 hours. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, and evaporated to dryness under reduced pressure to give an active ester intermediate.

Then, 12 mg of 5-aminolevulinic acid hydrochloride, 26. Mu.l of N, N-Diisopropylethylamine (DIPEA) and the active ester were added to the mixture and reacted overnight in 2ml of methylene chloride. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective product 1,21 mg. White solid. 31% yield.

¹H NMR (400 MHz, Chloroform-d)δ8.18 – 8.14 (m, 2H), 7.87 – 7.71 (m, 2H), 7.64 – 7.56 (m, 1H), 7.55 – 7.28 (m, 11H), 7.15 (d,J= 6.5 Hz, 1H), 6.33 – 6.21 (m, 2H), 6.07 (dd,J= 9.4, 3.3 Hz, 1H), 5.69 (d,J= 7.1 Hz, 1H), 5.44 (d,J= 3.3 Hz, 1H), 4.99 (dd,J= 9.8, 2.3 Hz, 1H), 4.44 (dd,J= 10.9, 6.6 Hz, 1H), 4.35 – 4.16 (m, 2H), 4.07 (d,J= 5.1 Hz, 2H), 3.81 (d,J= 7.1 Hz, 1H), 3.75 – 3.35 (m, 3H), 3.20 (t,J= 14.7 Hz, 2H), 2.57 (s, 4H), 2.51 (s, 3H), 2.44 – 2.14 (m, 7H), 1.94 (d,J= 1.4 Hz, 3H), 1.89 – 1.78 (m, 1H), 1.68 (s, 3H), 1.44 – 1.32 (m, 3H), 1.14 (s, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ203.93, 203.47, 173.43, 171.63, 170.35, 168.23, 168.00, 167.09, 162.57, 142.85, 136.93, 133.82, 133.69, 132.86, 131.99, 130.40, 129.40, 129.11, 128.89, 128.55, 127.81, 127.03, 84.56, 81.28, 79.23, 75.83, 75.68, 75.21, 72.18, 58.56, 52.88, 49.35, 45.79, 43.32, 35.95, 34.43, 32.64, 31.27, 29.84, 26.91, 25.91, 22.91, 22.33, 21.02, 14.93, 9.80.

High resolution mass spectrum HRMS (ESI) calculated C ₅₆H₆₂N₂O₁₉S (M+Na)⁺ m/z 1121.3559, found 1121.3558.

EXAMPLE 2 Synthesis of Compound 2

(2 AR,4S,4aS,6R,9S,11S,12S,12aR,12bS) -9- ((R) -15- (((S) -benzoylamino (phenyl) methyl) -3,6,9,13-tetraoxo-2, 14-dioxa-11-thio-8-azahexadecan-16-yl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5-oxo-3, 4a,5,6,9,10, 12-dehydro-1H-7, 11-methoxycyclodeca [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of methyl 5-aminolevulinate. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective product 2,30 mg as a yellow solid. The yield thereof was found to be 53%. Purity of 97%.¹H NMR (400 MHz, Chloroform-d)δ8.26 (d,J= 9.4 Hz, 1H), 8.22 – 8.10 (m, 2H), 7.90 – 7.82 (m, 2H), 7.65 – 7.56 (m, 1H), 7.52 (dd,J= 8.4, 6.9 Hz, 2H), 7.50 – 7.27 (m, 8H), 7.11 (t,J= 5.3 Hz, 1H), 6.30 (s, 1H), 6.11 (dd,J= 9.5, 3.2 Hz, 1H), 5.70 (d,J= 7.1 Hz, 1H), 5.41 (d,J= 3.2 Hz, 1H), 4.99 (dd,J= 9.7, 2.2 Hz, 1H), 4.33 (d,J= 8.4 Hz, 1H), 4.26 – 4.12 (m, 3H), 3.83 (d,J= 7.0 Hz, 1H), 3.78 (d,J= 15.6 Hz, 1H), 3.73 – 3.67 (m, 2H), 3.65 (s, 4H), 3.51 (d,J= 14.0 Hz, 1H), 3.24 – 3.10 (m, 4H), 2.93 (s, 1H), 2.80 – 2.54 (m, 6H), 2.53 (s, 4H), 2.42 (dd,J= 15.7, 9.5 Hz, 2H), 2.22 (s, 3H), 2.17 (s, 1H), 1.96 (d, J = 1.4 Hz, 3H), 1.69 (s, 3H), 1.24 (s, 3H), 1.14 (s, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ204.04, 203.87, 172.61, 171.40, 170.06, 169.13, 168.71, 168.18, 167.86, 167.15, 143.00, 137.10, 133.77, 132.86, 131.83, 130.43, 129.41, 129.04, 128.88, 128.45, 128.40, 127.88, 127.00, 84.60, 81.21, 79.28, 76.60, 75.87, 75.76, 75.27, 72.27, 72.18, 66.73, 58.64, 52.69, 49.34, 45.70, 43.33, 38.76, 35.75, 35.67, 34.85, 34.71, 32.05, 27.92, 26.92, 22.94, 22.35, 22.04, 20.96, 14.98, 10.46, 9.76.

HRMS (ESI) calculated C ₅₇H₆₄N₂O₁₉S (M+H)⁺ m/z 1113.3897, found 1113.3906.

EXAMPLE 3 Synthesis of Compound 3

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzoylamino (phenyl) methyl) -4,8,11,14-tetraoxo-3, 15-dioxa-6-thio-9-azaheptadecanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,9,11, 12-dehydro-1H-7, 11-methoxycyclodecanoic acid [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diyldiacetic acid ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,16 mg of ethyl 5-aminolevulinate. To 2 ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 3,29 mg as a yellow solid in 50% yield. The purity is 98%.

¹H NMR (400 MHz, Chloroform-d)δ8.27 (d,J= 9.5 Hz, 1H), 8.21 – 8.14 (m, 2H), 7.90 – 7.83 (m, 2H), 7.62 – 7.57 (m, 1H), 7.51 (dd,J= 8.2, 6.8 Hz, 2H), 7.45 – 7.31 (m, 8H), 7.08 (t,J= 5.1 Hz, 1H), 6.30 (s, 1H), 6.13 (dd,J= 9.5, 3.0 Hz, 1H), 5.70 (d,J= 7.1 Hz, 1H), 5.42 (d,J= 3.0 Hz, 1H), 4.99 (dd,J= 9.7, 2.3 Hz, 1H), 4.46 (dd,J= 10.9, 6.6 Hz, 1H), 4.33 (d,J= 8.4 Hz, 1H), 4.23 (dd,J= 8.4, 1.1 Hz, 1H), 4.11 (q,J= 7.1 Hz, 3H), 3.86 – 3.76 (m, 2H), 3.51 (d,J= 13.9 Hz, 1H), 3.15 (dd,J= 14.7, 4.2 Hz, 2H), 2.83 (s, 2H), 2.81 – 2.69 (m, 1H), 2.71 – 2.66 (m, 1H), 2.64 (dq,J= 3.1, 2.0, 1.5 Hz, 2H), 2.54 (s, 3H), 2.44 (dd,J= 15.5, 9.4 Hz, 1H), 2.23 (s, 5H), 1.97 (d,J= 1.5 Hz, 3H), 1.69 (s, 3H), 1.27 – 1.21 (m, 9H), 1.14 (s, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ203.72, 203.56, 172.20, 171.09, 169.74, 168.79, 168.40, 167.86, 167.53, 166.84, 142.68, 136.78, 133.46, 132.55, 131.52, 130.11, 129.09, 128.72, 128.57, 128.14, 128.09, 127.56, 126.68, 84.28, 80.89, 78.98, 75.56, 75.44, 74.96, 71.95, 71.87, 60.80, 58.33, 52.37, 49.02, 45.39, 43.02, 38.51, 35.44, 35.36, 34.55, 34.38, 31.76, 29.52, 27.65, 26.60, 22.64, 22.05, 20.65, 14.67, 13.96, 9.45.

HRMS (ESI) calculated C ₅₈H₆₆N₂O₁₉S (M+H)⁺ m/z 1127.4053, found 1127.4050.

EXAMPLE 4 Synthesis of Compound 4

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzoylamino (phenyl) methyl) -4,8,11,14-tetraoxo-3, 15-dioxa-6-thio-9-azaoctadecanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,9,11, 12-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,18 mg of propyl 5-aminolevulinate. To 2 ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 4,30 mg as a yellow solid in 50% yield. The purity is 95%.

¹H NMR (400 MHz, Chloroform-d)δ8.28 (d,J= 9.5 Hz, 1H), 8.21 – 8.15 (m, 2H), 7.89 – 7.83 (m, 2H), 7.63 – 7.56 (m, 1H), 7.51 (dd,J= 8.3, 6.8 Hz, 2H), 7.45 – 7.31 (m, 8H), 7.08 (t,J= 5.2 Hz, 1H), 6.30 (s, 1H), 6.13 (dd,J= 9.5, 3.0 Hz, 1H), 5.70 (d,J= 7.1 Hz, 1H), 5.42 (d,J= 3.0 Hz, 1H), 4.99 (dd,J= 9.7, 2.3 Hz, 1H), 4.46 (dd,J= 10.9, 6.6 Hz, 1H), 4.33 (d,J= 8.4 Hz, 1H), 4.25 – 4.12 (m, 3H), 4.01 (t,J= 6.7 Hz, 2H), 3.87 – 3.74 (m, 2H), 3.51 (d,J= 13.9 Hz, 1H), 3.15 (dd,J= 14.7, 4.8 Hz, 2H), 2.81 (s, 7H), 2.54 (s, 4H), 2.44 (dd,J= 15.5, 9.4 Hz, 1H), 2.22 (s, 4H), 1.96 (d,J= 1.4 Hz, 3H), 1.69 (s, 3H), 1.63 (q,J= 7.1 Hz, 2H), 1.42 (s, 1H), 1.24 (s, 3H), 1.14 (s, 3H), 0.92 (t,J= 7.4 Hz, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ 204.04, 203.89, 172.64, 171.40, 170.09, 169.15, 168.77, 168.24, 167.91, 167.12, 142.97, 137.06, 133.77, 132.85, 131.84, 130.40, 129.41, 129.04, 128.87, 128.45, 128.42, 127.85, 127.03, 84.58, 81.18, 79.23, 76.59, 75.87, 75.76, 75.26, 72.25, 72.18, 65.31, 58.62, 55.86, 52.78, 49.33, 45.70, 43.80, 43.33, 38.75, 35.71, 34.85, 34.68, 32.08, 28.35, 28.11, 27.92, 26.90, 22.92, 22.41, 22.34, 20.96, 18.72, 17.33, 14.96, 14.07, 12.65, 9.76.

HRMS (ESI) calculated C ₅₉H₆₈N₂O₁₉S (M+H)⁺ m/z 1141.4210, found 1141.4211.

EXAMPLE 5 Synthesis of Compound 5

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzoylamino (phenyl) methyl) -4,8,11,14-tetraoxo-3, 15-dioxa-6-thio-9-azanonadecanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,10,11, 12-dehydro-1H-7, 11-methoxycyclodecanoic acid [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diyldiacetic acid ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,19 mg of butyl 5-aminolevulinate were admixed. To 2 ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give 5,32 mg of the objective compound as a yellow solid in 54% yield. The purity was 96%.

¹H NMR (400 MHz, Chloroform-d)δ8.27 (d,J= 9.5 Hz, 1H), 8.18 (d,J= 7.6 Hz, 2H), 7.86 (d,J= 7.6 Hz, 2H), 7.59 (d,J= 7.3 Hz, 1H), 7.51 (t,J= 7.6 Hz, 2H), 7.46 – 7.31 (m, 8H), 7.07 (t,J= 5.1 Hz, 1H), 6.30 (s, 1H), 6.13 (dd,J= 9.1, 2.8 Hz, 1H), 5.70 (d,J= 7.1 Hz, 1H), 5.42 (d,J= 2.8 Hz, 1H), 5.03 – 4.95 (m, 1H), 4.46 (dd,J= 11.0, 6.6 Hz, 1H), 4.33 (d,J= 8.5 Hz, 1H), 4.27 – 4.13 (m, 3H), 4.06 (t,J= 6.7 Hz, 2H), 3.87 – 3.75 (m, 2H), 3.51 (d,J= 13.8 Hz, 1H), 3.15 (dd,J= 14.7, 4.9 Hz, 2H), 2.72 – 2.61 (m, 4H), 2.54 (s, 4H), 2.45 (dd,J= 15.5, 9.4 Hz, 1H), 2.23 (s, 4H), 1.97 (s, 3H), 1.93 – 1.85 (m, 1H), 1.69 (s, 3H), 1.60 (q,J= 7.2 Hz, 3H), 1.36 (h,J= 7.5 Hz, 3H), 1.24 (s, 4H), 1.14 (s, 3H), 0.92 (t,J= 7.4 Hz, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ204.04, 203.89, 172.64, 171.40, 170.09, 169.15, 168.77, 168.24, 167.91, 167.12, 142.97, 137.06, 133.77, 132.85, 131.84, 130.40, 129.41, 129.04, 128.87, 128.45, 128.42, 127.85, 127.03, 84.58, 81.18, 79.23, 76.59, 75.87, 75.76, 75.26, 72.25, 72.18, 65.31, 58.62, 55.86, 52.78, 49.33, 45.70, 43.80, 43.33, 38.75, 35.71, 34.85, 34.68, 32.08, 28.35, 28.11, 27.92, 26.90, 22.92, 22.41, 22.34, 20.96, 18.72, 17.33, 14.96, 14.07, 12.65, 9.76.

HRMS (ESI) calculated C ₆₀H₇₀N₂O₁₉S (M+H)⁺ m/z 1155.4366, found 1155.4368.

EXAMPLE 6 Synthesis of Compound 6

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzoylamino (phenyl) methyl) -4,8,11,14-tetraoxo-3, 15-dioxa-6-thio-9-azaeicosyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,10,11, 12-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,21 mg of amyl 5-aminolevulinate. To 2 ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 6,34 mg as a yellow solid in 56% yield. The purity was 96%.

¹H NMR (400 MHz, Chloroform-d)δ8.27 (d,J= 9.4 Hz, 1H), 8.20 – 8.16 (m, 2H), 7.89 – 7.84 (m, 2H), 7.60 (t,J= 7.3 Hz, 1H), 7.51 (dd,J= 8.4, 6.8 Hz, 2H), 7.45 – 7.31 (m, 8H), 7.09 (t,J= 5.2 Hz, 1H), 6.30 (s, 1H), 6.12 (dd,J= 9.5, 3.1 Hz, 1H), 5.70 (d,J= 7.1 Hz, 1H), 5.41 (d,J= 3.1 Hz, 1H), 4.99 (dd,J= 9.7, 2.3 Hz, 1H), 4.46 (dd,J= 11.0, 6.6 Hz, 1H), 4.33 (d,J= 8.5 Hz, 1H), 4.25 – 4.13 (m, 3H), 4.04 (t,J= 6.8 Hz, 2H), 3.87 – 3.76 (m, 2H), 3.51 (d,J= 13.9 Hz, 1H), 3.17 (t,J= 5.1 Hz, 2H), 2.84 (s, 4H), 2.64 (p,J= 2.4 Hz, 2H), 2.54 (s, 3H), 2.43 (dd,J= 15.5, 9.3 Hz, 1H), 2.23 (s, 4H), 1.96 (d,J= 1.4 Hz, 3H), 1.69 (s, 3H), 1.60 (q,J= 7.0 Hz, 3H), 1.45 (dd,J= 17.5, 6.6 Hz, 6H), 1.24 (s, 3H), 1.14 (s, 3H), 0.90 (d,J= 6.7 Hz, 3H).

HRMS (ESI) calculated C ₆₁H₇₂N₂O₁₉S (M+H)⁺ m/z 1169.4523, found 1169.4519.

EXAMPLE 7 Synthesis of Compound 7

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzoylamino (phenyl) methyl) -4,8,11,14-tetraoxo-3, 15-dioxa-6-thio-9-azaeicosanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,10,11, 12-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxo-6, 12b (2 aH) -diyldiacetic acid ester

51 Mg of intermediate PTX-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,23 mg of hexyl 5-aminolevulinate. To 2 ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give 7,31 mg of the objective compound as a yellow solid in 51% yield. The purity was 97%.

¹H NMR (400 MHz, Chloroform-d)δ8.34 (d,J= 9.5 Hz, 1H), 8.23 – 8.18 (m, 2H), 7.91 – 7.86 (m, 2H), 7.64 – 7.58 (m, 1H), 7.54 (dd,J= 8.3, 6.8 Hz, 2H), 7.47 – 7.34 (m, 8H), 7.14 – 7.10 (m, 1H), 6.32 (d,J= 4.7 Hz, 2H), 6.15 (dd,J= 9.5, 2.9 Hz, 1H), 5.72 (d,J= 7.1 Hz, 1H), 5.43 (d,J= 3.0 Hz, 1H), 5.02 (dd,J= 9.8, 2.3 Hz, 1H), 4.48 (dd,J= 10.9, 6.6 Hz, 1H), 4.34 (s, 1H), 4.26 (s, 3H), 4.07 (t,J= 6.8 Hz, 2H), 3.86 (d,J= 7.2 Hz, 1H), 3.81 (s, 1H), 3.54 (d,J= 13.9 Hz, 1H), 3.18 (s, 2H), 2.66 (d,J= 2.3 Hz, 2H), 2.57 (s, 3H), 2.52 – 2.39 (m, 2H), 2.25 (s, 4H), 1.99 (d,J= 1.5 Hz, 3H), 1.92 (ddd,J= 13.9, 11.0, 2.4 Hz, 2H), 1.72 (s, 3H), 1.66 – 1.56 (m, 3H), 1.45 (d,J= 1.6 Hz, 1H), 1.39 (s, 2H), 1.28 (s, 5H), 1.27 (s, 3H), 1.16 (s, 3H), 0.91 – 0.89 (m, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ204.05, 203.87, 172.63, 171.42, 170.05, 169.12, 168.70, 168.17, 167.83, 167.16, 143.03, 137.10, 133.76, 132.83, 131.83, 130.43, 129.38, 129.03, 128.88, 128.44, 128.39, 127.89, 127.00, 84.60, 81.19, 79.30, 76.60, 75.90, 75.76, 75.26, 72.28, 72.17, 65.35, 58.64, 52.66, 49.34, 45.69, 43.32, 35.74, 35.66, 34.82, 34.71, 32.06, 32.02, 31.64, 31.57, 31.52, 30.44, 30.32, 30.26, 29.83, 28.63, 27.93, 26.91, 25.65, 22.96, 22.83, 22.66, 22.36, 20.97, 15.00, 14.14, 9.76.

HRMS (ESI) calculated C ₆₁H₇₂N₂O₁₉S (M+H)⁺ m/z 1183.4679, found 1183.4677.

EXAMPLE 8 Synthesis of Compound 8

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -16- ((S) -benzoylamino (phenyl) methyl) -3,6,9,14-tetraoxo-2, 15-dioxa-11, 12-dithio-8-azaheptadec-17-yl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5, 4a,5,6,9,11,12 a-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of methyl 5-aminolevulinate were admixed. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 8,24 mg as a yellow solid in 43% yield. The purity was 96%.

¹H NMR (400 MHz, Chloroform-d)δ8.16 – 8.09 (m, 2H), 7.87 – 7.81 (m, 2H), 7.78 (d,J= 8.6 Hz, 1H), 7.67 – 7.59 (m, 1H), 7.58 – 7.35 (m, 9H), 7.28 (d,J= 3.3 Hz, 1H), 6.99 (t,J= 4.7 Hz, 1H), 6.30 (s, 1H), 6.21 – 6.13 (m, 1H), 5.91 (dd,J= 8.5, 5.6 Hz, 1H), 5.65 (d,J= 7.1 Hz, 1H), 5.48 (d,J= 5.6 Hz, 1H), 4.96 (dd,J= 9.7, 2.3 Hz, 1H), 4.44 (dd,J= 10.9, 6.6 Hz, 1H), 4.33 – 3.99 (m, 4H), 3.76 (d,J= 7.1 Hz, 1H), 3.61 – 3.50 (m, 2H), 3.42 – 3.20 (m, 2H), 2.85 (s, 3H), 2.75 – 2.56 (m, 4H), 2.51 (ddd,J= 14.6, 9.7, 6.5 Hz, 1H), 2.44 (s, 3H), 2.22 (s, 4H), 2.14 (dd,J= 15.6, 9.3 Hz, 2H), 2.09 – 1.99 (m, 1H), 1.95 – 1.84 (m, 5H), 1.68 (s, 3H), 1.46 (dd,J= 22.6, 6.5 Hz, 2H), 1.13 (s, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ203.86, 203.51, 173.11, 171.18, 170.11, 169.12, 168.89, 168.40, 167.61, 167.05, 142.60, 136.93, 134.24, 133.86, 132.98, 131.98, 130.35, 129.42, 129.15, 128.79, 128.66, 127.60, 127.25, 84.57, 81.17, 79.13, 76.54, 75.74, 75.68, 75.11, 72.14, 71.96, 58.48, 53.73, 52.19, 49.65, 45.88, 43.27, 42.36, 40.82, 38.84, 35.80, 35.34, 34.72, 27.58, 26.86, 22.99, 22.14, 20.98, 18.78, 17.44, 15.01, 9.75.

HRMS (ESI) calculated C ₅₇H₆₄N₂O₁₉S₂(M+H)⁺ m/z 1145.3617, found 1145.3615.

EXAMPLE 9 Synthesis of Compound 9

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzamide (phenyl) methyl) -4,9,12,15-tetraoxo-3, 16-dioxa-6, 7-dithio-10-octadecanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5, 4a,5,6,9,10,11, 12-dehydro-1H-7, 11-methoxycyclohexanecarboxylic acid [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diyldiacetic acid ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of ethyl 5-aminolevulinate. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 9,24 mg as a yellow solid in 42% yield. The purity was 97%.

¹H NMR (400 MHz, Chloroform-d)δ8.17 – 8.09 (m, 2H), 7.87 – 7.80 (m, 2H), 7.78 (d,J= 8.5 Hz, 1H), 7.68 – 7.58 (m, 1H), 7.57 – 7.33 (m, 10H), 6.97 (t,J= 5.7 Hz, 1H), 6.30 (s, 1H), 6.22 – 6.08 (m, 1H), 5.92 (dd,J= 8.5, 5.5 Hz, 1H), 5.66 (d,J= 7.1 Hz, 1H), 5.48 (d,J= 5.5 Hz, 1H), 4.97 (dd,J= 9.7, 2.3 Hz, 1H), 4.44 (dd,J= 10.9, 6.6 Hz, 1H), 4.32 – 4.16 (m, 3H), 4.12 – 4.00 (m, 3H), 3.77 (d,J= 7.0 Hz, 1H), 3.64 – 3.46 (m, 2H), 3.42 – 3.20 (m, 2H), 2.73 – 2.66 (m, 2H), 2.60 (ddd,J= 7.4, 5.8, 3.4 Hz, 2H), 2.44 (s, 3H), 2.34 – 2.10 (m, 5H), 1.97 – 1.86 (m, 6H), 1.68 (s, 4H), 1.42 (d,J= 5.5 Hz, 2H), 1.37 (s, 1H), 1.33 (s, 3H), 1.13 (s, 3H).

¹³C NMR (101 MHz, Chloroform-d)δ203.88, 203.56, 172.67, 171.19, 170.10, 169.10, 168.83, 168.38, 167.63, 167.08, 142.63, 136.95, 134.27, 133.86, 133.00, 132.00, 130.37, 129.43, 129.16, 128.81, 128.68, 127.61, 127.24, 84.60, 81.20, 79.17, 75.74, 75.13, 72.16, 71.99, 61.18, 58.51, 53.70, 49.70, 45.89, 43.29, 42.40, 40.80, 35.81, 35.37, 34.74, 32.07, 31.58, 30.46, 30.34, 29.84, 29.51, 27.88, 26.88, 23.01, 22.84, 22.16, 20.99, 15.03, 14.27, 14.24, 9.76.

HRMS (ESI) calculated C ₅₈H₆₆N₂O₁₉S₂(M+H)⁺ m/z 1159.3774, found 1159.3773.

EXAMPLE 10 Synthesis of Compound 10

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzamide (phenyl) methyl) -4,9,12,15-tetraoxo-3, 16-dioxa-6, 7-dithio-10-azanonanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5-oxo-3, 4a,5,6,9,10, 12-12 a-decahydro-1H-7, 11-methoxycyclodeca [3,4] benzo [1,2b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of propyl 5-aminolevulinate. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 10,27 mg as a yellow solid in 47% yield. The purity is 95%.

¹H NMR (300 MHz, Chloroform-d)δ8.17 – 8.07 (m, 2H), 7.86 – 7.82 (m, 2H), 7.74 – 7.30 (m, 11H), 7.28 (t,J= 1.7 Hz, 1H), 6.99 (t,J= 4.6 Hz, 1H), 6.30 (s, 1H), 6.16 (t,J= 8.8 Hz, 1H), 5.91 (dd,J= 8.5, 5.6 Hz, 1H), 5.65 (d,J= 7.0 Hz, 1H), 5.47 (d,J= 5.6 Hz, 1H), 5.02 – 4.90 (m, 1H), 4.44 (dd,J= 10.9, 6.6 Hz, 1H), 4.33 – 4.06 (m, 4H), 3.98 (t,J= 6.7 Hz, 2H), 3.76 (d,J= 7.1 Hz, 1H), 3.66 – 3.47 (m, 2H), 3.43 – 3.17 (m, 2H), 2.74 – 2.48 (m, 5H), 2.44 (s, 3H), 2.21 (s, 3H), 1.91 (d,J= 9.9 Hz, 4H), 1.67 (s, 3H), 1.59 (p,J= 7.1 Hz, 2H), 1.25 (s, 3H), 1.21 (s, 3H), 1.12 (s, 3H), 0.90 (t,J= 7.4 Hz, 3H).

¹³C NMR (75 MHz, Chloroform-d)δ203.88, 203.57, 172.77, 171.17, 170.09, 169.11, 168.76, 168.40, 167.64, 167.05, 142.62, 136.93, 134.26, 133.87, 132.96, 131.98, 130.36, 129.39, 129.14, 128.80, 128.65, 127.61, 127.25, 84.57, 81.16, 79.12, 75.68, 75.09, 72.14, 66.76, 58.46, 53.72, 52.73, 49.68, 45.88, 43.25, 42.35, 40.74, 36.94, 35.33, 34.73, 30.51, 29.85, 29.82, 29.77, 27.81, 26.86, 22.99, 21.99, 20.98, 17.83, 15.02, 10.46, 9.74.

HRMS (ESI) calculated C ₅₉H₆₈N₂O₁₉S₂(M+H)⁺ m/z 1173.3931, found 1173.3934.

EXAMPLE 11 Synthesis of Compound 11

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzamide (phenyl) methyl) -4,9,12,15-tetraoxo-3, 16-dioxa-6, 7-dithio-10-azacycloalkyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8,13, 5-tetramethyl-5-oxo-3, 4a,5,6,10,11, 12-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diyldiacetic acid ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of butyl 5-aminolevulinate. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 11,26 mg as a yellow solid in 45% yield. The purity was 96%.

¹H NMR (300 MHz, Chloroform-d)δ8.18 – 8.06 (m, 2H), 7.87 – 7.28 (m, 14H), 6.98 (t,J= 4.6 Hz, 1H), 6.30 (s, 1H), 6.22 – 6.08 (m, 1H), 5.91 (dd,J= 8.5, 5.5 Hz, 1H), 5.65 (d,J= 7.1 Hz, 1H), 5.47 (d,J= 5.6 Hz, 1H), 4.96 (dd,J= 9.6, 2.2 Hz, 1H), 4.44 (t,J= 8.8 Hz, 1H), 4.33 – 4.06 (m, 4H), 4.03 (t,J= 6.7 Hz, 2H), 3.76 (d,J= 7.1 Hz, 1H), 3.64 – 3.49 (m, 2H), 3.45 – 3.20 (m, 2H), 2.75 – 2.48 (m, 6H), 2.44 (s, 3H), 2.22 (s, 3H), 2.14 (dd,J= 15.6, 9.3 Hz, 1H), 1.90 (d,J= 11.1 Hz, 4H), 1.67 (s, 4H), 1.61 – 1.50 (m, 2H), 1.23 (d,J= 12.5 Hz, 5H), 1.13 (s, 3H), 0.91 (t,J= 7.3 Hz, 4H).

¹³C NMR (75 MHz, Chloroform-d)δ203.88, 203.58, 172.77, 171.18, 170.09, 169.11, 168.85, 168.40, 167.66, 167.06, 142.63, 136.93, 134.26, 133.57, 132.97, 132.07, 130.36, 129.40, 129.14, 128.79, 128.66, 127.61, 127.24, 84.33, 81.17, 79.13, 75.73, 74.77, 71.80, 71.42, 65.08, 58.47, 54.98, 49.69, 45.88, 43.26, 42.29, 40.98, 35.91, 34.74, 30.65, 29.96, 29.75, 29.38, 27.84, 26.87, 22.99, 21.93, 20.99, 19.20, 15.03, 13.83, 9.76.

HRMS (ESI) calculated C ₆₀H₇₀N₂O₁₉S₂(M+H)⁺ m/z 1187.4087, found 1187.4087.

EXAMPLE 12 Synthesis of Compound 12

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzamide (phenyl) methyl) -4,9,12,15-tetraoxo-3, 16-dioxa-6, 7-dithio-10-azaeicosanoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5-oxo-3, 4a,5,6,9,10,12 a-decahydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of amyl 5-aminolevulinate were admixed. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 12,28 mg as a yellow solid in 48% yield. The purity was 96%.

¹H NMR (300 MHz, Chloroform-d)δ8.17 – 8.10 (m, 2H), 7.75 – 7.68 (m, 2H), 7.65 – 7.30 (m, 12H), 7.10 (d,J= 9.4 Hz, 1H), 6.27 (s, 1H), 6.01 (dd,J= 9.2, 3.4 Hz, 1H), 5.67 (d,J= 7.1 Hz, 1H), 5.55 (d,J= 3.5 Hz, 1H), 5.01 – 4.93 (m, 1H), 4.43 (dd,J= 10.8, 6.6 Hz, 1H), 4.34 – 4.15 (m, 2H), 3.80 (d,J= 7.0 Hz, 1H), 3.50 (s, 2H), 2.55 (ddd,J= 15.4, 9.6, 6.4 Hz, 1H), 2.44 (s, 3H), 2.34 (dd,J= 15.6, 9.0 Hz, 2H), 2.22 (s, 3H), 2.18 – 1.99 (m, 3H), 1.89 (d,J= 1.3 Hz, 4H), 1.68 (s, 3H), 1.42 (d,J= 4.4 Hz, 2H), 1.33 (s, 3H), 1.27 (d,J= 4.8 Hz, 6H), 1.21 (s, 4H), 1.13 (s, 3H), 0.86 (dt,J= 8.6, 6.2 Hz, 4H).

¹³C NMR (75 MHz, Chloroform-d)δ203.94, 203.82, 171.19, 169.89, 169.28, 168.60, 168.50, 168.33, 167.75, 167.21, 142.61, 137.70, 136.71, 134.09, 133.61, 133.06, 132.26, 130.37, 129.30, 128.85, 128.35, 127.30, 126.86, 83.76, 81.21, 79.25, 75.68, 74.98, 72.03, 69.94, 68.01, 58.63, 52.85, 51.22, 43.31, 40.43, 39.25, 39.17, 35.78, 34.98, 31.94, 31.55, 30.80, 29.84, 26.94, 23.30, 22.44, 20.47, 20.06, 14.95, 13.88, 9.80.

HRMS (ESI) calculated C ₆₁H₇₂N₂O₁₉S₂(M+Na)⁺ m/z 1223.4063, found 1223.4056.

EXAMPLE 13 Synthesis of Compound 13

(2 AR,4S,4aS,6R,9S,11S,12 aR,12 bS) -9- (((R) -2- ((S) -benzamide (phenyl) methyl) -4,9,12,15-tetraoxo-3, 16-dioxa-6, 7-dithio-10-azabehenoyl) oxy) -12- (benzoyloxy) -4, 11-dihydroxy-4 a,8, 13-tetramethyl-5-oxo-3, 4a,5,6,10,11, 12-dehydro-1H-7, 11-methoxycyclodecyl [3,4] benzo [1,2-b ] oxa-6, 12b (2 aH) -diacetic acid diethyl ester

51 Mg of intermediate PTX-S-S-COOH,24 mg of 2- (7-azabenzotriazol) -N, N, N ', N' -tetramethylurea Hexafluorophosphate (HATU), 20. Mu.l of DIPEA,15 mg of hexyl 5-aminolevulinate. To 2ml of dichloromethane was added. Stir overnight. The reaction was monitored by thin layer chromatography. After the reaction was completed, the organic solvent was removed under reduced pressure, and the residue was extracted (ethyl acetate was dissolved, and washed with water and saturated sodium chloride in this order). The organic phase was dried over anhydrous sodium sulfate, evaporated to dryness under reduced pressure, and purified by silica gel column chromatography to give the objective compound 13,28 mg as a yellow solid in 47% yield. The purity is 98%.

¹H NMR (400 MHz, Chloroform-d)δ8.13 (dt,J= 7.0, 1.3 Hz, 2H), 7.88 – 7.82 (m, 2H), 7.79 (d,J= 8.5 Hz, 1H), 7.68 – 7.60 (m, 1H), 7.59 – 7.29 (m, 10H), 6.96 (t,J= 4.5 Hz, 1H), 6.30 (s, 1H), 6.17 (t,J= 9.0 Hz, 1H), 5.92 (dd,J= 8.5, 5.6 Hz, 1H), 5.65 (d,J= 7.1 Hz, 1H), 5.48 (d,J= 5.6 Hz, 1H), 4.96 (dd,J= 9.7, 2.3 Hz, 1H), 4.44 (dd,J= 10.9, 6.6 Hz, 1H), 4.32 – 4.15 (m, 4H), 4.11 – 3.98 (m, 4H), 3.77 (d,J= 7.0 Hz, 1H), 3.64 – 3.51 (m, 3H), 3.42 3.21 (m, 2H), 2.77 (dd,J= 7.3, 5.5 Hz, 1H), 2.72 – 2.58 (m, 5H), 2.56 – 2.48 (m, 1H), 2.44 (s, 3H), 2.22 (s, 3H), 2.14 (dd,J= 15.5, 9.0 Hz, 1H), 1.94 – 1.89 (m, 4H), 1.68 (s, 3H), 1.62 – 1.55 (m, 3H), 1.21 (s, 3H), 1.13 (s, 3H), 0.93 – 0.85 (m, 6H).

¹³C NMR (101 MHz, Chloroform-d)δ204.19, 203.88, 173.07, 171.48, 170.41, 169.41, 169.15, 168.70, 167.94, 167.39, 142.96, 137.27, 134.59, 134.18, 133.30, 132.29, 130.69, 129.74, 129.47, 129.12, 128.98, 127.93, 127.57, 84.91, 81.50, 79.48, 76.06, 76.01, 75.44, 72.47, 72.28, 65.71, 65.55, 58.81, 54.04, 50.02, 46.21, 43.59, 43.14, 42.70, 41.09, 36.14, 35.66, 35.07, 31.86, 31.85, 30.15, 28.97, 28.92, 28.24, 28.17, 27.20, 26.00, 25.96, 23.31, 22.97, 22.46, 21.29, 15.34, 14.58, 14.45, 10.07.

HRMS (ESI) calculated C ₆₂H₇₄N₂O₁₉S₂(M+H)⁺ m/z 1215.4400, found 1215.4404.

EXAMPLE 14 Compound prodrug Release assay

Compound 2 was formulated as a stock solution of 10 millimoles per liter (mM) using dimethyl sulfoxide (DMSO) as vehicle and stored in a-20 ℃ refrigerator. An appropriate amount of the compound stock solution was diluted with Phosphate Buffer (PBS) to a final concentration of 100 μmol per liter, and then H ₂O₂ (0, 1,2,10 mm) was added and incubation was performed at 37 ℃ and repeated 3 times. Samples were collected at appropriate time intervals (0, 1,2,4,6,8,10,12,24 h) and analyzed directly by High Performance Liquid Chromatography (HPLC) and Mass Spectrometry (MS). Agilent 1100 HPLC and UV detector were used with Agilent C18 column (4.6X105 mm,3.5 μm), mobile phase 85% methanol 0.1% trifluoroacetic acid, flow rate 1.0 ml/min.

As shown in FIG. 1, the prodrug did not substantially decompose in the case of 0mmol/LH ₂O₂ after 24 hours incubation, showing good stability. As ROS concentration increases, the rate and extent of prodrug release increases significantly, exhibiting a positive correlation. Low concentration (1 mmol/L) of monosulfur prodrug released 50.07% + -0.02 and disulfide prodrug released 53.06% + -0.06. High concentration (10 mmol/L) of the monosulfur prodrug released 68.83% + -0.02 and disulfide prodrug released 69.23% + -0.02.

The high-low concentration gap mono-sulfur prodrug releases 18.76% ± 0.04 more and disulfide prodrug releases 16.17% ± 0.03 more. These data indicate that the release of such prodrugs is ROS-responsive.

Example 15 anti-cell proliferation assay (MTT assay)

MDA MB-231 cells in the logarithmic growth phase (the cells were purchased from the cell bank of the institute of Kagaku-Han virus, academy of China) were taken, digested with 0.25% pancreatin in an ultra clean bench, and the digestion was stopped. The culture medium is sucked by a pipetting gun, the adherent cells are blown down and then transferred into a 10ml centrifuge tube, the supernatant is discarded after centrifugation for five minutes at 1000 revolutions per minute (rpm), then a proper amount of culture medium is added for blowing and beating uniformly to prepare single cell suspension, 20 microlitres of the single cell suspension is sucked and added into a cell counting plate, and after counting, the cell concentration is diluted by the culture medium and adjusted to reach 3000/hole. 200 microliters of each well is inoculated in a 96-well plate, after the inoculation is completed, the conventional culture is carried out for 12 h hours, after the cells are attached to the wall, the culture medium is sucked out, 200 microliters of culture medium containing 1 mu M of medicine is added again, 3 parallel wells are arranged for each compound concentration, and the same amount of blank culture medium is added into a control group. After 24h of administration, 20. Mu.l of MTT was added to each well, incubation and culture were continued for 3h, and then the mixture was removed, and the mixture was shaken on a shaker for 10 min, and the absorbance of each well was measured by using a microplate reader at a wavelength of 490/570 nm, and the cell viability (%) = (1-A _{experimental group}/A_{Control group}). Times.100% after the effect of the test compound at different concentrations was calculated according to the corresponding formula, wherein A _{experimental group} represents the absorbance of the experimental group and A _{Control group} represents the absorbance of the control group. The experimental results are shown in Table 1. The data indicate that the prodrugs exhibit similar or even stronger killing effects on cancer cells. Preferably compound 13 exhibits 9% greater inhibition than the positive drug.

Table 1 antiproliferative activity of prodrugs on breast cancer cells

Inhibition (1 μm) data results are the average of three replicates, ±represents Standard Deviation (SD);

example 16 acute toxicity detection of Compounds in mice

ICR mice were kept normally for approximately five weeks, and then the mice were randomly assigned to 6 dose groups of compound 2 and paclitaxel, each group corresponding to a different dose (10 mg/kg, 20 mg/kg, 50 mg/kg, 100 mg/kg, 150 mg/kg, 200 mg/kg), each group having 6 mice (male and female halves). The compound was injected via the tail vein. Abnormal behavior and death were recorded within two weeks after dosing. The maximum tolerance (MTD value) was used as an indicator of acute toxicity determination, i.e., the maximum dose that allowed mice of the entire dose group to remain viable after a single administration.

The MTD value of the positive drug paclitaxel group was 20 mg/kg. It was observed that 20 mg/kg mice had a significant weight loss for two weeks after dosing, whereas the paclitaxel prodrug group did not die even at the highest 200 mg/kg dose, and the low dose group had no significant change in body weight. The result shows that the MTD value of the taxol prodrug in ICR mice is at least 10 times higher than that of taxol which is a positive medicine, and the in-vivo safety is greatly improved.

EXAMPLE 17 in vivo imaging experiments of Compound 2 in mice

The purpose of this example was to explore the fluorescence imaging ability of compounds on tumor-bearing mice. Following tumor formation in mice subcutaneously implanted with 4T1 tumor cells, compound 2 was administered intravenously as well as orally and then scanned with Tanon ABL imaging systems at various time points (0, 1,2,3,4,6,8,12,24,36,48,72 h). After 12h, 24h, 36h, 48h and 72h of administration, a significant near infrared fluorescence signal was observed at the tumor site, as shown in fig. 2-6. The quantitative values of fluorescence signals of in vivo imaging of mice are shown in Table 2. Notably, 12h after injection, near infrared fluorescence signals in tumors of the tail vein injection group have been clearly recognized, in contrast to certain fluorescence signals that can be recognized by tumor-bearing mice 24h of the oral administration group. This suggests that compound 2 has an accumulating effect on tumors in mice and has a certain oral activity.

Table 2 quantitative response values for in vivo imaging of compound 2 mice

As described above, although the present invention has been shown and described with reference to certain preferred embodiments, it is not to be construed as limiting the invention itself. Various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.

Claims

1. A compound as shown in general formula I or general formula II or a pharmaceutically acceptable salt thereof, characterized in that:

Wherein, R ₁ is selected from hydrogen, C ₁ -C ₆ alkyl;

Wherein, R ₂ is selected from C ₁ -C ₆ alkyl.

2. The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein: said _R1 is selected from hydrogen, _C1 - _C6 n-alkyl.

3. The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein: said _R2 is selected from _C1 - _C6 n-alkyl.

4. The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein _R1 is selected from hydrogen, methyl, ethyl, n-propyl, n-butyl, n-pentyl and n-hexyl.

5. The compound or pharmaceutically acceptable salt thereof according to claim 1, wherein _R2 is selected from methyl, ethyl, n-propyl, n-butyl, n-pentyl and n-hexyl.

6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: it is selected from any one of the following compounds: .

7. A method for preparing the compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Wherein, R ₁ is selected from C ₁ -C ₆ alkyl.

8. A method for preparing the compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: Wherein, R ₂ is selected from C ₁ -C ₆ alkyl.

9. A pharmaceutical composition comprising the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers.

10. Use of the compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and/or treating tumor diseases.