CN114181223B - Cephalotaxine derivative and preparation method and application thereof - Google Patents

Abstract

The invention provides a cephalotaxine derivative, which has a structure shown in a formula I or pharmaceutically acceptable salt or stereoisomer thereof. The cephalotaxine derivatives provided by the invention can effectively inhibit the growth of various tumor cells, especially inhibit Ba/F3-BCR/ABL-T315I mutant cells, and are novel and can overcome BCR-ABL ^T315I The protein synthesis inhibitor of drug-resistant mutation can be used for preparing anti-tumor drugs and has wide market application prospect.

Description

Cephalotaxine derivative and preparation method and application thereof

Technical Field

The invention relates to the technical field of medicines, in particular to a cephalotaxine derivative and a preparation method and application thereof.

Background

Chronic myelogenous leukemia (Chronic Myeloid Leukemia, CML) is a malignant myeloproliferative disease, which is manifested clinically by malignant proliferation of granulocytes (including neutrophils, eosinophils, basophils) and the like, and accumulation and concomitant myeloproliferation in blood, leading to anemia, thrombocytopenia and other symptoms in patients. CML accounts for about 20% of adult leukemias, and is one of the major diseases that threatens human life health. According to Globocan 2012 study data, there were 351965 newly increased leukemia cases and 265461 deaths worldwide in 2012, whereas in china, the number of deaths from leukemia in 2010 was estimated to be up to 58000. The occurrence of Philadelphia (Ph) is an important marker for clinical diagnosis of CML, and is derived from the mutual shift between the long arms of human chromosome 22 and chromosome 9, and this shift results in the ABL gene originally located on chromosome 9 being heterozygous downstream of the BCR gene on chromosome 22, thereby producing the fusion gene BCR-ABL, which encodes the occurrence of the BCR-ABL protein kinase produced as the main pathogenic mechanism of CML, and currently, BCR-ABL inhibitors have become one of the main drugs for clinical use in the treatment of CML. The first generation Bcr-Abl inhibitor imatinib is taken as a first small molecule targeted inhibitor designed and synthesized for Bcr-Abl kinase, is approved by FDA to be marketed in 2001, and has strong inhibition activity and good selectivity, thus obtaining great success in clinical treatmentAnd becomes a first-line drug for treating CML. However, with the large-scale use of imatinib, the problem of acquired drug resistance is gradually revealed, and especially for patients in the acceleration phase and the rapid change phase, the drug resistance incidence is as high as 40% -50% and 80%, respectively. Acquired point mutations of Bcr-Abl kinase are the main cause of drug resistance, and currently more than 100 types of point mutations have been discovered successively. To overcome imatinib resistance, second generation Bcr-Abl inhibitors including dasatinib, nilotinib, bosutinib, radotinib, and the like were successively approved for marketing. Although second generation inhibitors are effective against a variety of point mutations, few mutations remain to be crafted. In particular the Bcr-AblT315I mutation, the "goated" amino acid threonine, which is the Bcr-Abl protein, is mutated to isoleucine, approximately 15% -20% of all clinically available mutations, at present Bcr-Abl ^T315I The problem of drug resistance caused by mutations remains a significant challenge in CML treatment. The third generation Bcr-Abl inhibitor, pluratinib, was approved by the FDA for marketing, and although it has a good inhibitory effect on Bcr-AblWT kinase and various mutations including T315I mutation, it does not perform well on mutations such as Y253F/H, E255K/V, while it has a risk of inducing fatal blood coagulation and serious stenosis of blood vessels in clinical use, and thus was suspended for sale by the FDA in 10 months of 2013. Although the drug is then marketed again (12 months 20) after the black frame warning is added, the above problems remain to be solved by the development of novel Bcr-Abl inhibitors or other mechanism of action drugs.

10 months 2012, homoharringtonine (trade name: omacetaxine mepsuccinate) is approved by the U.S. FDA for use in treating chronic myelogenous leukemia patients resistant to two or more Bcr-Abl inhibitors. Homoharringtonine exhibits good inhibitory activity against a variety of Bcr-Abl mutations, including T315I mutations. Homoharringtonine is the first marketed protein inhibitor for the treatment of CML, which binds to the A site of the ribosomal 60S large subunit, competitively inhibits the binding of acetylated transfer RNA to the A site, thereby inhibiting the expression of a downstream series of oncogenic proteins, which are IC inhibitory activity against mouse lymphocytic leukemia cell P388 ₅₀ The value reaches 17 nM. At the same time, the method comprises the steps of,homoharringtonine is also the first natural product for the treatment of CML and is derived from plants of the genus Cephalotaxus (Cephalotaxus) of the family Cephalotaxus, from which about 50 more alkaloid compounds have been isolated and identified, many of which have good anti-CML activity as cephalotaxine-type alkaloids having the same parent nucleus as homoharringtonine, e.g., IC inhibitory activity of harringtonine, isoharringtonine, deoxyharringtonine on mouse lymphocytic leukemia cell P388 ₅₀ The values were 32 nM, 18 nM, 7.5 nM, respectively. Although homoharringtonine pair CML and Bcr-Abl ^T315I Mutant CMLs all exhibit good therapeutic effects, but there are still a number of problems. Homoharringtonine has serious toxic and side effects of myelosuppression, so that the administration mode of the homoharringtonine is changed from intravenous injection to subcutaneous injection in clinical secondary experiments. Subcutaneous injections also present a number of inconveniences to the patient relative to orally administered Bcr-Abl inhibitors. In addition, commercial homoharringtonine is a semisynthetic, the parent nucleus cephalotaxine fragment is extracted and separated from plants, and a side chain part is obtained by chemical synthesis, so that the synthesis is complicated due to the fact that the side chain fragment is complex, and the total yield of the side chain synthesis is only 24.5% at present. Therefore, there is an urgent need to find novel cephalotaxine compounds with high efficiency, low toxicity and stable in vivo metabolism, for treating CML, in particular with Bcr-Abl ^T315I Isomutated CML provides more candidate compounds.

Disclosure of Invention

In view of the above, the technical problem to be solved by the present invention is to provide a cephalotaxine derivative, its preparation method and application, and has high anti-tumor activity.

The invention provides a cephalotaxine derivative, which has a structure shown in a formula I or pharmaceutically acceptable salt or stereoisomer thereof:

a formula I;

wherein R is ₁ Is O;

R ₂ 、R ₃ 、R ₄ independently selected from one or more of H, C C3 alkoxy, halogen, trifluoromethyl, trifluoromethoxy, benzyl, benzyloxy, C1C 3 acyloxy, hydroxyl and amino.

Metabolites of the compounds and pharmaceutically acceptable salts thereof, as well as prodrugs that can be converted in vivo to the structures of the compounds and pharmaceutically acceptable salts thereof, are also encompassed by the claims of the present application.

Preferably, the R ₂ 、R ₃ 、R ₄ And independently selected from one or more of H, methoxy, F, cl, br, benzyloxy and hydroxyl.

Preferably, the cephalotaxine derivative is 3- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine, 3- (2 ' -o-methoxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine, 3- (2 ' -m-fluorobenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine, 3- (2 ' -m-benzyloxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine, 3- (2 ' -p-benzyloxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine, 3- (2 ' -m-hydroxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine or 3- (2 ' -p-hydroxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine.

Preferably, the cephalotaxine derivative has any one of the following structures:

the method for preparing the cephalotaxine derivatives is not particularly limited and can be prepared according to methods well known to those skilled in the art. For example, the person skilled in the art can synthesize the compound by several steps of reactions using phenylpyruvic acid or diethyl succinate as starting material according to the prior art and the common general knowledge.

Preferably, the invention provides a preparation method of the cephalotaxine derivative, which comprises the following steps:

s1) reacting phenylpyruvic acid or phenyl-substituted phenylpyruvic acid derivative with benzyl alcohol to prepare 2-oxo-3-phenylpropionic acid benzyl ester or derivative thereof;

s2) reacting benzyl 2-oxo-3-phenylpropionate or its derivative with methyl (triphenylphosphine alkenyl) acetate to obtain 1-benzyl 4-methyl 2-benzyl maleate andE) -2-benzylidenesuccinic acid 1-benzyl 4-methyl ester or a derivative thereof;

s3) mixing 1-benzyl 4-methyl 2-benzyl maleate with [ ]E) -reducing the 1-benzyl 4-methylester of 2-benzylidenesuccinic acid with a reducing agent to obtain 2-benzyl-4-methoxy-4-oxobutanoic acid or a derivative thereof;

s4) reacting 2-benzyl-4-methoxy-4-oxo butyric acid or a derivative thereof with cephalotaxine to obtain the cephalotaxine derivative shown in the formula I.

Taking phenylpyruvic acid as an example, the reaction route of the preparation method is as follows:

preferably, the reducing agent is selected from palladium carbon and Ru [ (], the reducing agent isS)-DTBM-SegPhos](OAc) ₂ Or Ru [ (]R)-DTBM-SegPhos](OAc) ₂ 。

Preferably, the present invention provides another preparation method of the cephalotaxine derivative, comprising the following steps:

a) Reacting dimethyl succinate with benzaldehyde or benzaldehyde derivative with substituent on phenyl, and acidifying and esterifying the product to obtain 1-methyl 2-benzylidene succinate or its derivative;

b) Reducing the 2-benzylidene succinic acid 1-methyl ester or a derivative thereof to obtain 2-benzyl-4-methoxy-4-oxo butyric acid or a derivative thereof;

c) 2-benzyl-4-methoxy-4-oxo-butyric acid or a derivative thereof reacts with cephalotaxine to obtain the cephalotaxine derivative shown in the formula I.

Taking benzaldehyde as an example, the reaction route of the preparation method is as follows:

the invention provides a pharmaceutical composition comprising the cephalotaxine derivative and an auxiliary agent.

The cephalotaxine derivative or the pharmaceutical composition provided by the invention can be used for treating transitional proliferative diseases or symptoms such as human or other mammal tumors.

The invention provides application of the cephalotaxine derivative or the pharmaceutical composition in preparing antitumor drugs.

Preferably, the tumor is any one or more of transitional proliferative diseases such as leukemia, non-small cell lung cancer, pancreatic cancer, breast cancer, prostatic cancer, liver cancer, skin cancer, epithelial cell cancer, gastrointestinal stromal tumor, histiocytic lymphoma, nasopharyngeal carcinoma, head and neck tumor, colon cancer, rectal cancer, glioma and the like.

Preferably, the non-small cell lung cancer is one or more of lung adenocarcinoma and lung squamous carcinoma.

The compounds of formula I provided above in the present invention may be used in combination with other drugs known to treat or ameliorate similar conditions. When administered in combination, the mode of administration and dosage of the original drug remains unchanged, while the compound of formula I is administered simultaneously or subsequently. When the compound of formula I is administered simultaneously with one or more other drugs, it is preferred to use a pharmaceutical composition containing one or more known drugs together with the compound of formula I. Drug combinations also include administration of the compound of formula I with one or more other known drugs over overlapping time periods. When a compound of formula I is administered in combination with one or more other drugs, the dosage of the compound of formula I or the known drug may be lower than when they are administered alone.

Drugs or active ingredients that may be used in combination with the compounds of formula I include, but are not limited to:

estrogen receptor modulators, androgen receptor modulators, retinoid receptor modulators, cytotoxin/cytostatics, antiproliferative agents, protein transferase inhibitors, HMG-CoA reductase inhibitors, HIV protein kinase inhibitors, reverse transcriptase inhibitors, angiogenesis inhibitors, cell proliferation and survival signaling inhibitors, agents that interfere with cell cycle checkpoints and apoptosis inducers, cytotoxic drugs, tyrosine protein inhibitors, EGFR inhibitors, VEGFR inhibitors, serine/threonine protein inhibitors, bcr-Abl inhibitors, c-Kit inhibitors, met inhibitors, raf inhibitors, MEK inhibitors, MMP inhibitors, topoisomerase inhibitors, histidine deacetylase inhibitors, proteasome inhibitors, CDK inhibitors, bcl-2 family protein inhibitors, MDM2 family protein inhibitors, IAP family protein inhibitors, STAT family protein inhibitors, PI3K inhibitors, AKT inhibitors, integrin blockers, interferon- α, interleukin-12, COX-2 inhibitors, p53 activators, VEGF antibodies, EGF antibodies, and the like.

In one embodiment, drugs or active ingredients that may be used in combination with the compounds of formula I include, but are not limited to: albumin, alendronic acid, interferon, al Qu Nuoying, allopurinol sodium, palonosetron hydrochloride, altretamine, aminoglutethimide, amifostine, amrubicin, an Ya pyridine, anastrozole, dolasetron, aranesp, arglabin, arsenic trioxide, minoxin, 5-azacytidine, azathioprine, BCG or tice BCG, betadine, betamethasone acetate, betamethasone sodium phosphate formulation, bexarotene, bleomycin sulfate, british, bortezomib, busulfan, calcitonin, alezomib injection, capecitabine, carboplatin, kang Shide, cefesone, cet Mo Baijie, daunorubicin, chlorambucil, cisplatin, cladribine, clofaxine, cyclophosphamide, cytarabine, dacarbazine, dactinomycin, daunorubicin, dexamethasone dexamethasone phosphate, estradiol valerate, deniinterleukin 2, dibaume, dulorelin, delazocine, diethylstilbestrol, dafukang, docetaxel, deoxyfluorouridine, doxorubicin, dronabinol, jejunum-166-chitosan complex, eligard, labyrinase, epirubicin hydrochloride, aprepitant, epirubicin, alfuzoxetine, erythropoietin, eplatin, levamisole, estradiol formulations, 17-beta-estradiol, estramustine sodium phosphate, ethinyl estradiol, amifostine, hydroxy phosphate, petrolatum, etoposide, fadrozole, tamoxifen formulations, febuxostat, finasteride, feveride, fluorouridine, fluconazole, fludarabine, 5-fluorodeoxyuridine monophosphate, 5-fluorouracil, fluoxytestosterone, flusteramine, fotemustine, fludarabine, 1-beta-D-arabinofuranosyl cytothiadine-5' -stearoyl phosphate, fotemustine, fulvestrant, gamma globulin, gemcitabine, gemtuzumab, imatinib mesylate, carmustine wafer capsule, goserelin, glatiramer hydrochloride, histrelin, and mefenadine, hydrocortisone, erythro-hydroxynonyladenine, hydroxyurea, chlorpyrifos Titania Bei Moshan antibody, idarubicin, ifosfamide, interferon alpha, interferon-alpha 2, interferon alpha-2A, interferon alpha-2B, interferon alpha-nl, interferon alpha-n 3, interferon beta, interferon gamma-la, interleukin-2, intron A, iressa, irinotecan, ketery, lentinan sulfate, letrozole, leucovorin, leuprorelin, pharmaceutical composition, and pharmaceutical composition Leuprolide acetate, levamisole, levoleucovorin calcium salt, levothyroxine sodium preparation, lomustine, lonidamine, dronabinol, nitrogen mustard, mecobalamin, medroxyprogesterone acetate, megestrol acetate, melphalan, esterified estrogens, 6-azurin, mesna, methotrexate, methyl aminolevulinate, miltefosine, melamycin, mitomycin C, mitotane, mitozepine, trovatam, doxorubicin citrate liposomes, nedaplatin, pegylated feaglutinin, olpreninterleukin, neutrogen, nilutamide, tamoxifen, NSC-631570, recombinant human interleukin 1-beta, octreotide, ondansetron hydrochloride, dehydrohydrocortisone oral solution, oxaliplatin, mitoxan, paclitaxel, prednisone sodium phosphate formulation, peganase, perroxen, penoxsulam, penstatin, streptozotocin, pilocarpine hydrochloride, bicin, plicamycin, porphin sodium, prednimustine, setprednisolone, prednisone, betamethadone, procarbamate, recombinant human erythropoietin, raltitrexed, liratio, rhenium-186 etidronate, meloxicam, dynamics stretch-A, romidep, pilocarpine hydrochloride tablet, octreotide, sarustine, semustine, sirolimus, sibutran, soxazocine, sodium methylprednisolone, pafoglic acid, stem cell therapy, streptozocin, strontium chloride-89, levothyroxine sodium, tamoxifen, tamsulosin, tamonamine, testosterone, temustine, temozolomide, teniposide, propione, methyltestosterone, thioguanine thiotepa, thyroid stimulating hormone, tiludronate, topotecan, toremifene, tolsimizumab, trastuzumab, troxipran, tretinoin, methotrexate tablet, trimethoprim, triptorelin acetate, triptorelin pamoate, ulipran, uridine, valrubicin, visrinone, vinblastine, vincristine, vinorelbine, vitamin Lu Liqin, dexpropimide, clean setami Ding Sizhi, pivannin, a stable preparation of paclitaxel protein, acolbifene, interferon r-lb, afinitak, aminopterin, alzoxifene, asprisnil, altamide, atrasentan, BAY 43-9006, avastin, CCI-779, CDC-501, celecoxib, cetuximab, clenbuterol, ciprofloxacin acetate, dyclonidine, DN-101, doxorubicin-MTC, dSLIM, dutasteride, edotecarin, epothilone, fenretinide, histamine dihydrochloride, histrelin hydrogel implant, holmium-166 DOTMP, ibandronic acid, interferon gamma, intron-PEG, ixabepilone, keyhole-shaped hemocyanin, L-651582, lanle peptide, lasofoxifene, libra, lonafamib, midependrofen, mi Nuoqu acid ester, MS-209, liposomal MTP-PE, MX-6, nafarelin, nemorubicin, neovalvulat, nolatatrex, olimarson, onco-TCS, osiam, paclitaxel polyglutamate, sodium, PN-401, QS-21, quasipuncum, R-1549, raloxifene, frog enzyme, 13-cis-valproic acid, satraplatin, T-138067, tarca, docosahexaenoic acid, thymol, galangustazole, altern, titania, tenuina, TLvalproate, TLvalproine, or combinations thereof.

Compared with the prior art, the invention provides a cephalotaxine derivative which has a structure shown in a formula I or pharmaceutically acceptable salt or stereoisomer thereof. The cephalotaxine derivatives provided by the invention can effectively inhibit the growth of various tumor cells, especially inhibit Ba/F3-BCR/ABL-T315I mutant cells, and are novel and can overcome BCR-ABL ^T315I The protein synthesis inhibitor of drug-resistant mutation can be used for preparing anti-tumor drugs and has wide market application prospect.

Detailed Description

In order to further illustrate the present invention, the cephalotaxine derivatives, and the preparation methods and applications thereof, provided by the present invention are described in detail below with reference to examples.

Example 13 preparation of- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (CY 0076)

Step a: preparation of benzyl 2-oxo-3-phenylpropionate (Compound 1)

1) Phenylpyruvic acid (1.64 g, 10 mmol) was dissolved in 20 mL dry dichloromethane, thionyl chloride (1.2 mL, 14 mmol) and 1 drop of N, N-dimethylformamide were added with stirring, and reacted at low temperature for 30 min. Benzyl alcohol (973 mg, 9 mmol) was slowly added dropwise, and after 1h of reaction, a large amount of ice water was added to precipitate a pale yellow solid, which was filtered under reduced pressure, and the filter cake was sufficiently washed with water and dried under vacuum 2 h. The product is directly put into the next reaction.

2) To the reaction was added 20. 20 mL dry dichloromethane to dissolve, benzyl alcohol (1.08 g, 10 mmol) and pyridine (791 mg, 10 mmol) were slowly added and stirred at room temperature for 2 h. After the reaction, adding a large amount of ice-water mixture for quenching, extracting with dichloromethane for multiple times, combining organic phases, washing with saturated saline solution once, and anhydrous Na ₂ SO ₄ Drying, filtration and spin drying gave a solid 1.8. 1.8 g (Yield: 74%). ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 7.47-7.25 (m, 10 H), 5.31 (s, 2H), 5.29 (s, 2H). MS (ESI), m/z: 255[M+H] ⁺ 。

Step b: 1-benzyl 4-methyl 2-benzyl maleate (Compound 2) and [ ]E) Preparation of 1-benzyl 4-methyl-2-benzylidenesuccinate (Compound 3)

Benzyl 2-oxo-3-phenylpropionate (compound 1) (1 g, 4 mmol) was dissolved in 8 mL toluene, methyl (triphenylphosphine) acetate (1.3 g, 4 mmol) was added and heated to 90 ℃ for reaction overnight. After the reaction was completed, the solvent was removed under reduced pressure, and white solids 2 (464 mg, yield: 38%) and 3 (371 mg, yield: 30%) were separated by column chromatography. Compound 2: ¹ H NMR (400 MHz, DMSO-d ₆ ) δ7.40-7.26 (m, 10H), 6.27 (s, 1H), 5.25 (s, 2H), 5.19 (s, 2H), 3.73 (s, 3H). MS (ESI), m/z: 311[M+H] ⁺ . Compound 3: ¹ H NMR (400 MHz, CDCl ₃ ) δ7.35-7.21 (m, 10H), 6.93 (s, 1H), 5.18 (s, 2H), 4.26 (s, 2H), 3.81 (s, 3H). MS (ESI), m/z: 311[M+H] ⁺ 。

step c: preparation of 2-benzyl-4-methoxy-4-oxobutanoic acid (Compound 4)

1-benzyl 4-methyl 2-benzyl maleate (Compound 2) (463 mg, 1.5 mmol) andE) 1-benzyl 4-methylester of 2-benzylidenesuccinic acid (Compound 3) (371 mg, 1.2 mmol) was dissolved in 6 mL ethyl acetate, the reaction system was replaced with argon, pd/C was added, the reaction system was replaced with argon, and the reaction was stirred at room temperature for 2 h. After the reaction was completed, the cake was washed with celite, and the filtrates were combined, and the solvent was removed under reduced pressure to give a pale yellow oil (553 mg, yield: 92%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.33-7.19 (m, 5H), 3.66 (s, 3H), 3.21-3.13 (m, 2H), 2.82-2.76 (m, 1H), 2.70-2.63 (m, 1H), 2.45-2.40 (m, 1H). MS (ESI), m/z: 223[M+H] ⁺ 。

Step d: preparation of 3- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine

2-benzyl-4-methoxy-4-oxobutanoic acid (compound 4) (89 mg, 0.4 mmol),N, N' -dicyclohexylcarbodiimide (124 mg, 0.4 mmol), 4-dimethylaminopyridine (98 mg, 0.8 mmol) were dissolved in 1mL anhydrous tetrahydrofuran, cephalotaxine (126 mg, 0.4 mmol) was added to the reaction system, and stirred overnight at room temperature. After the reaction, the filtrate was collected by suction filtration under reduced pressure, and separated by column chromatography to give 70 mg (Yield: 34%) as a pale yellow oil. MS (ESI),m/z: 520[M+H] ⁺ 。

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.32-7.17 (m, 5H),δ 6.58 (s, 1H), 6.56(s, 1H),5.88 (s, 2H), 5.83 (s, 1H), 5.02(s, 1H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.88 (m, 2H),2.63-2.69(m, 5H),2.35 (m, 2H),1.75(m, 2H),1.65 (m, 2H).

example 2 3 preparation of- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (CY 0076)

Steps e-f: preparation of 1-methyl 2-benzylidene succinate (Compound 7)

Step e A solution of dimethyl succinate (2.57 g,17.6 mmol) in MeOH (15 mL) was carefully added to a reflux mixture of sodium methoxide (0.84 g,15.5 mmol) and methanol (15 mL). The mixture was refluxed for 1 hour, then benzaldehyde (14.1 mmol) was added over 30 minutes and stirred at reflux temperature for a further 5 hours. After the reaction was completed, methanol was removed in vacuo, and the product was used for the next reaction.

Step f the product obtained in step e was dissolved in 15% NaOH solution (10 mL) and refluxed for 4 hours. After cooling, the reaction mixture was washed with EtOAc (2×10 mL). Next, the aqueous phase was acidified to pH with 2M HCl<2 extracted with EtOAc (3X 10 mL) and the organic layer was dried over anhydrous Na ₂ SO ₄ Drying and concentrating.

Step g the product from step f was dissolved in MeOH (15 mL), amberlyst-15 H+ (0.5 g) was added and the reaction mixture was refluxed for 16 hours. The mixture was filtered through celite and concentrated in vacuo. The residue was subjected to column chromatography on silica gel (PE: etoac=9:1 to 7:3) to give compound 7 (yield 36%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 8.03 (s, 1H), 7.46 – 7.34 (m, 5H), 3.76 (s, 3H), 3.57 (s, 2H).

Step h: preparation of 2-benzyl-4-methoxy-4-oxobutanoic acid (Compound 4)

2-Benzylglycoctane-1-methyl ester (Compound 7) (220 mg, 1 mmol) was dissolved in 5mL anhydrous degassed methanol, the reaction system was replaced with argon, pd/C was added, the reaction system was replaced with argon, and then the reaction system was stirred at room temperature for 2 h. After the reaction was completed, the cake was washed with celite, and the filtrates were combined, and the solvent was removed under reduced pressure to give compound 4 (221 mg, yield: 99%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.05 (s, 1H), 7.22 – 7.18 (m, 2H), 7.15 – 7.08 (m, 3H), 3.54 (s, 3H), 3.10 – 3.02 (m, 2H), 2.72 – 2.65 (m, 1H), 2.56 (dd, J = 17.2, 9.2 Hz, 1H), 2.32 (dd, J = 17.2, 4.8 Hz, 1H). ¹³ C NMR (101 MHz, CDCl ₃ ) δ 179.96, 172.44, 137.95, 129.09, 128.65, 126.83, 51.90, 42.92, 37.39, 34.49.

Step i: preparation of 3- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (CY 0076)

2-benzyl-4-methoxy-4-oxobutanoic acid (Compound 4) (0.36 mmol) and triethylamine (1.35 mmol) were dissolved in 2mL CH ₂ Cl ₂ To the above solution was added 2,4, 6-trichlorobenzoyl chloride (80. Mu.L, 0.51 mmol) by syringe. Stirred at room temperature for 1 hour. 1mL of cephalotaxine (95 mg,0.3 mmol) and N, N-dimethylaminopyridine (DMAP, 7.3mg,0.06 mmol) CH were added to the reaction system ₂ Cl ₂ A solution. The reaction mixture was then stirred at room temperature for 4 hours. After completion of the reaction (monitored by TLC), the mixture was taken up in CH ₂ Cl ₂ Diluted organic layer was sequentially washed with saturated NaHCO ₃ Aqueous solution and brine wash. The combined organic layers were treated with anhydrous Na ₂ SO ₄ Dried, filtered, concentrated in vacuo, and loaded onto a silica gel column packed with 5% triethylamine/PE. The column was eluted with 3:2 PE:EtOAc to afford the product 3- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (CY 0076) 109 mg (70% yield).

The product was characterized by nuclear magnetic resonance.

Example 3 3- (2'RPreparation of-benzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (CY 0076-2)

Synthesized by the method of example 2, in step h, ru [ (. About. ] is usedS)-DTBM-SegPhos](OAc) ₂ (1.7. 1.7 mg, 0.002 mmol) instead of Pd/C. (yield: 60%).

¹ H NMR (400 MHz, CDCl ₃ ) δ7.32-7.17 (m, 5H),δ 6.58 (s, 1H), 6.56(s, 1H),5.88 (s, 2H), 5.83 (s, 1H),5.02(s, 1H),3.62 (d,1H),3.60 (s,3H),3.50 (s,3H),3.36 (m, 1H),3.18-3.12 (m, 1H), 2.88 (m, 2H),2.63-2.69(m, 5H),2.35 (m, 2H),1.75(m, 2H),1.65 (m, 2H).

Example 43- (2'SPreparation of-benzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (CY 0076-1)

Synthesized by the method of example 2, in step h, ru [ (. About. ] is usedR)-DTBM-SegPhos](OAc) ₂ (1.7. 1.7 mg, 0.002 mmol) instead of Pd/C. (yield: 82%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.21 (t, 2H), 7.15 (m, 1H), 6.96 (d, 2H), 6.59 (s, 1H), 6.57 (s, 1H), 5.89 (d, 1H), 5.81 (d, 1H), 5.75 (d, 1H), 5.04 (s, 1H), 3.78 (d, 1H), 3.70 (s, 3H), 3.53 (s, 3H), 3.25 (m, 1H), 3.07 (m, 1H), 2.93(m, 1H), 2.79 (m, 1H), 2.59 (m, 2H), 2.37(m, 2H), 2.24 (dd, 1H), 2.03(m, 3H), 1.89(m,1H), 1.74 (m, 2H).

Example 53 preparation of (2 ' -O-methoxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (HLA 268)

The synthesis was carried out in accordance with example 2 (yield: 68%). ¹ H NMR (400 MHz, CDCl ₃ ) δ7.21-7.17 (m, 1H), 6.98 (m, 4H), 6.58 (s, 1H), 6.56(s, 1H), 5.87 (s, 3H), 5.02(s, 1H), 3.70 (s, 3H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.87 (m, 2H), 2.64-2.70 (m, 5H), 2.37 (m, 2H), 1.78(m, 2H), 1.66 (m, 2H).

Example 6 3- (2'RPreparation of-O-methoxybenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 270)

The synthesis was carried out in accordance with example 3 (yield: 64%). ¹ H NMR (400 MHz, CDCl ₃ ) δ7.21-7.17 (m, 1H), 6.98 (m, 4H), 6.59 (s, 1H), 6.55(s, 1H),5.88 (s, 3H), 5.05(s, 1H),3.72 (s, 3H),3.62 (d, 1H), 3.60 (s, 3H), 3.52 (s, 3H), 3.36 (m, 1H), 3.18-3.12 (m, 1H), 2.88 (m, 2H), 2.64-2.70 (m, 5H), 2.35 (m, 2H), 1.76 (m, 2H), 1.64(m, 2H).

Example 7 3- (2'SPreparation of-O-methoxybenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 293)

The synthesis was carried out in accordance with example 4 (yield: 83%). ¹ H NMR (400 MHz, CDCl ₃ )

δ7.21-7.17 (m, 1H), 6.98 (m, 4H), 6.59 (s, 1H), 6.55(s, 1H), 5.88 (s, 3H), 5.05(s, 1H), 3.72 (s, 3H), 3.62 (d,1H), 3.60 (s, 3H), 3.52 (s, 3H), 3.36 (m, 1H), 3.18-3.12 (m, 1H), 2.88 (m, 2H), 2.64-2.70 (m, 5H), 2.35 (m, 2H), 1.76 (m, 2H), 1.64 (m, 2H).

Example 83 preparation of (2 ' -m-fluorobenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (HLA 309-3)

The synthesis was carried out in accordance with example 2 (yield: 68%). ¹ H NMR (400 MHz, CDCl ₃ )δ 7.34-7.30 (m, 1H),δ 7.02 (m, 3H), 6.59 (s, 1H), 6.55 (s, 1H), 5.88 (s,3H), 5.05(s,1H), 3.62(d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.87-2.93(m, 2H), 2.62-2.68(m, 5H), 2.36 (m, 2H), 1.76(m, 2H), 1.62(m, 2H).

Example 9 3- (2'RPreparation of-m-fluorobenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 309-2)

The synthesis was carried out in accordance with example 3 (yield: 62%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.32 (m, 1H), 7.04 (m, 3H), 6.59 (s, 1H), 6.56 (s, 1H), 5.87 (s, 3H), 5.02 (s, 1H), 3.64 (d, 1H), 3.61 (s, 3H), 3.52 (s,, 3H), 3.35 (m, 1H), 3.18-3.12 (m, 1H), 2.89 (m, 2H), 2.62-2.68 (m, 5H), 2.38 (m, 2H), 1.72(m, 2H), 1.64 (m, 2H).

Example 10 3- (2'SPreparation of-m-fluorobenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 309-1)

The synthesis was carried out in accordance with example 4 (yield: 60%). ¹ H NMR (400 MHz, CDCl ₃ ) δ7.32 (m, 1H), 7.04 (m, 3H), 6.59 (s, 1H), 6.56 (s, 1H), 5.87 (s, 3H), 5.02(s, 1H), 3.64 (d, 1H), 3.61 (s, 3H), 3.52 (s, 3H), 3.35 (m, 1H), 3.18-3.12 (m, 1H), 2.89 (m, 2H), 2.62-2.68 (m, 5H), 2.38 (m, 2H), 1.72 (m, 2H), 1.64 (m, 2H).

Example 11 3- (2'SPreparation of-m-benzyloxybenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 767)

The synthesis was carried out in accordance with example 4 (yield: 76%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.40-7.48 (m, 4H), 7.32-7.22 (m, 2H), 7.04 (m, 3H), 6.59 (s, 1H), 6.56 (s, 1H), 5.87 (s, 3H), 5.14 (s, 2H), 5.02 (s, 1H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.87 (m, 2H), 2.64-2.70 (m, 5H), 2.37 (m, 2H), 1.75(m, 2H), 1.65(m, 2H).

Example 12 preparation of 3- (2 ' -p-benzyloxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (HLA 758)

The synthesis was carried out in accordance with example 4 (yield: 99%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.40-7.48 (m, 4H), δ 7.32 (m, 1H), 7.13(m, 1H), 7.01 (m, 3H), 6.58(s, 1H), 6.52 (s, 1H), 5.88 (s, 3H), 5.16 (s, 2H), 5.03 (s, 1H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.87-2.93 (m, 2H), 2.64-2.70(m, 5H), 2.36 (m, 2H),1.75 (m, 2H), 1.65(m, 2H).

Example 13- (2)SPreparation of' -m-hydroxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (HLA 768)

Using the product of example 11, 3- (2 ' S-m-benzyloxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (50 mg) as a reaction starting material, dissolved in 1ml of methanol, pd/C was added, the reaction system was stirred overnight at room temperature in 1 atm of hydrogen, pd/C was removed by filtration and the filtrate was concentrated. Silica gel column chromatography (PE: etoac=4:1) gave the corresponding debenzylated product 3- (2S ' -m-hydroxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine in 76% yield. ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.20(s, 1H), 7.10-7.00 (m, 2H), 6.84-6.80 (m, 2H), 6.58(s, 1H), 6.52(s,1H), 5.88(s, 3H), 5.03(s, 1H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.34 (m, 1H), 3.18-3.12 (m, 1H), 2.87-2.93 (m, 2H), 2.64-2.70(m, 5H), 2.36 (m, 2H), 1.75(m, 2H), 1.65(m, 2H).

Example 14 3- (2'SPreparation of p-hydroxybenzyl-4 '-methoxy-4' -oxobutanoyl) cephalotaxine (HLA 760)

Using the product of example 12, 3- (2 ' S-p-benzyloxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (50 mg) as a reaction starting material, dissolved in 1ml of methanol, pd/C was added, the reaction system was stirred overnight at room temperature in 1 atm of hydrogen, pd/C was removed by filtration and the filtrate was concentrated. Silica gel column chromatography (PE: etoac=4:1) gave the corresponding debenzylated product 3- (2S ' -p-hydroxybenzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine in 99% yield. ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.02(s, 1H), 7.02-7.00 (m, 2H), 6.84-6.80 (m, 2H), 6.58(s, 1H), 6.52(s,1H), 5.88(s, 3H), 5.03(s, 1H), 3.62 (d,1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.33 (m, 1H), 3.16-3.10 (m, 1H), 2.88-2.94 (m, 2H), 2.66-2.72 (m, 5H), 2.34 (m, 2H), 1.75(m, 2H), 1.65(m, 2H).

EXAMPLE 15 preparation of 3-N- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (designated CY 5120068)

Step e: preparation of triethylethane-1, 2-tricarboxylic acid ester (Compound 5)

Diethyl malonate (728 mg, 4.5 mmol) was dissolved in absolute ethanol, 2.5 ml of a freshly prepared 2M sodium ethoxide solution in ethanol was added dropwise at 0 ℃, and after stirring for 40 min at 0 ℃, ethyl bromoacetate (759 mg, 4.5 mmol) was added dropwise slowly, and the reaction was carried out slowly to room temperature for 3-4 h. The off-white solid is separated out, and the reaction liquid is directly put into the next reaction.

Step f: preparation of 3-phenylpropane-1, 2-tricarboxylic acid triethyl ester (Compound 6)

2.5 mL of ethanol solution of newly prepared 2M sodium ethoxide is slowly added dropwise to the above triethylethane-1, 2-tricarboxylic acid ester (compound 5) solution at 0 ℃, and after stirring for 40 min at 0 ℃, benzyl bromide (770 mg, 4.5 mmol) is slowly added dropwise to the reaction system, and the mixture is slowly warmed to room temperature to react 3-4 h. After the reaction was completed, the mixture was quenched with water, ethanol was removed by spinning under reduced pressure, extracted with ethyl acetate several times, the organic phases were combined, washed with saturated brine once, and dried over Na ₂ SO ₄ Drying, filtration and spin drying gave a white solid 1.2. 1.2 g (Yield: 81%). ¹ H NMR (400 MHz, CDCl ₃ -d ₆ ) δ 7.26-7.20 (m, 3H), 7.08-7.06 (m, 2H), 4.22-4.11 (m, 6H), 3.37 (s, 2H), 2.83 (s, 2H), 1.26-1.22 (m, 9H). MS (ESI), m/z: 337[M+H] ⁺ 。

Step g: preparation of diethyl 2-benzylsuccinate (Compound 7)

3-Phenylpropane-1, 2-tricarboxylic acid triethyl ester (Compound 6) (3.33 g, 10 mmol) was dissolved in 20 mL DMSO, lithium chloride (1.1 g, 25 mmol) was added, water (360 mg, 20 mmol) was added, and the mixture was heated to 160℃to react overnight. Quenching with a large amount of water after the reaction, extracting with ethyl acetate for several times, mixing the organic phases, washing with saturated saline solution once, and anhydrous Na ₂ SO ₄ Drying, filtration and spin drying gave a solid 2.3. 2.3 g (Yield: 87%). ¹ H NMR (400 MHz, CDCl ₃ -d ₆ ) δ7.28-7.14 (m, 5H), 4.12-4.05 (m 4H), 3.13-2.99 (m, 2H), 2.75 (dd, J = 13.2 Hz, 8.0 Hz, 1H), 2.64 (dd, J = 16.8 Hz, 9.2 Hz, 1H), 2.38 (dd, J = 16.8 Hz, 5.2 Hz, 1H), 1.22-1.15 (m, 6H). MS (ESI), m/z: 265[M+H] ⁺ 。

Step h: preparation of 2-Benzylsuccinic acid (Compound 8)

Diethyl 2-benzylsuccinate (Compound 7) (89 mg, 0.34 mmol) was dissolved in 2mL tetrahydrofuran:water (3:1), sodium hydroxide (27 mg, 0.67 mmol) was added, 3 h was stirred at room temperature, tetrahydrofuran was removed by evaporation under reduced pressure, extraction was performed multiple times with ethyl acetate, and the organic phases were combined to give 68 mg (Yield: 96%) as a white solid. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ12.22 (s, 2H), 7.31-7.17 (m, 5H), 3.33-2.86 (m, 2H), 2.77-2.70 (m, 1H), 2.45-2.39 (m, 1H), 2.27-2.22 (m, 1H)。

Step i: preparation of 2-benzyl-4-methoxy-4-oxobutanoic acid (Compound 9)

2-Benzylsuccinic acid (Compound 8) (937 mg, 4.5 mmol) was dissolved in 9 mL methanol and Amberlyst-15H was added ⁺ Reflux reaction 5h was heated. After the completion of the reaction, the mixture was filtered through celite, and the filtrate was collected by column chromatography to give 253. 253 mg (Yield: 25%) as a pale yellow oil. ¹ H NMR (400 MHz, CDCl ₃ -d ₆ ) δ 7.32-7.17 (m, 5H), 3.65 (s, 3H), 3.18-3.12 (m, 2H), 2.82-2.76 (m, 1H), 2.66 (dd, J = 17.2 Hz, 9.2 Hz, 1H), 2.42 (dd, J = 16.8 Hz, 4.4 Hz, 1H)。

Step j: preparation of 3-azido cephalotaxine (Compound 10)

Cephalotaxine (1.5 g, 4.67 mmol) was dissolved in 10mL anhydrous tetrahydrofuran and diphenyl azide phosphonate (3.3 g, 11.9 mmol) was slowly added dropwise under argon at-15℃to 1, 8-diazabicyclo [5.4.0 ]]Undec-7-ene (1.8 g, 11.9 mmol) was allowed to react overnight at room temperature slowly. After the completion of the reaction, the solvent was evaporated under reduced pressure, and a pale yellow solid 863 mg (YIelds: 53%) was isolated by column chromatography. MS (ESI),m/z: 341[M+H] ⁺ 。

step k: preparation of 3-amino cephalotaxine (Compound 11)

3-azido cephalotaxine (compound 10) (540 mg, 1.59 mmol) was dissolved in 4 mL tetrahydrofuran: water (3:1), triphenylphosphine (458 mg, 1.75 mmol) was added and the reaction stirred at room temperature overnight. After the reaction, tetrahydrofuran was removed under reduced pressure, extracted with ethyl acetate several times, the organic phases were combined, washed once with saturated brine, and dried Na ₂ SO ₄ Dried, filtered and spun-dried, and column chromatographed to give solid 250 mg (Yield: 50%). ¹ H NMR (400 MHz, CDCl ₃ -d ₆ ) δ 6.68 (s, 1H), 6.64 (s, 1H), 5.88 (s, 2H), 4.74 (s, 1H), 3.86 (d, J = 4.0 Hz, 1H), 3.69 (s, 3H), 3.06-3.01 (m, 1H), 2.90-2.84 (m, 3H), 2.53-2.45 (m, 3H), 1.92-1.87 (m, 4H), 1.74-1.69 (m, 2H). MS (ESI), m/z: 315[M+H] ⁺ 。

Step l: preparation of 3- (2 ' -benzyl-4 ' -methoxy-4 ' -oxobutanoyl) cephalotaxine (designated CY 5120068)

3-Aminocephalotaxine (Compound 11) (100 mg, 0.32 mmol), 2-benzyl-4-methoxy-4-oxobutanAcid (Compound 9) (106 mg, 0.48 mmol), O- (7-azabenzotriazolyl)N, N, N', N' -tetramethyluronium hexafluorophosphate (181.4 mg, 0.48 mmol), triethylamine (64.4 mg, 0.64 mmol),N, Ndiisopropylethylamine (2 mg, 5%) was dissolved in 2mL dry dichloromethane and stirred overnight at room temperature. After the reaction, quench with water, extract with ethyl acetate multiple times, combine the organic phases, wash with saturated brine once, dry Na ₂ SO ₄ Dried, filtered and spun-dried, and column chromatographed to give solid 65 g (Yield: 39%). MS (ESI),m/z: 519[M+H] ⁺ 。

¹ H NMR (400 MHz, CDCl ₃ ) δ 8.24(brs, 1H), 7.32-7.28 (m, 4H), 7.18-7.14 (m, 1H), 6.58(s, 1H), 6.51 (s, 1H), 5.86 (s, 3H), 5.04 (s, 1H), 3.62 (d, 1H), 3.60 (s, 3H), 3.50 (s, 3H), 3.44 (m, 1H), 3.18-3.12 (m, 1H), 2.80-2.84 (m, 2H), 2.60-2.69(m, 5H), 2.34 (m, 2H), 1.75(m, 2H),1.65 (m, 2H).

EXAMPLE 16 preparation of 3- (4 '-methoxy-2' - (3 '-methoxybenzyl) -4' -oxobutanoyl) cephalotaxine (designated CY 5120024)

The synthesis was as in example 15.

MS (ESI), m/z: 549[M+H] ⁺ 。

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.21-7.17 (m, 1H), 6.98 (m, 4H), 6.59 (s, 1H), 6.55(s, 1H),5.88 (s, 3H), 5.72(brs, 1H), 5.05(s, 1H), 3.70 (s, 3H), 3.62 (d, 1H), 3.63 (s, 3H), 3.52 (s, 3H), 3.36 (m, 1H), 3.18-3.12 (m, 1H), 2.88 (m, 2H), 2.64-2.70(m, 5H), 2.35 (m, 2H), 1.76 (m, 2H), 1.64(m, 2H).

EXAMPLE 17 preparation of 3- (4 '-methoxy-2' - (2 '-chlorobenzyl) -4' -oxobutanoyl) cephalotaxine (designated CY 5120069)

The synthesis was as in example 2.

MS (ESI), m/z: 553[M+H] ⁺ 。

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.66-7.62 (m, 1H), 7.18-7.14 (m, 3H), 6.59 (s, 1H), 6.55(s, 1H), 5.87 (s, 3H), 5.54 (brs, 1H), 5.02(s, 1H), 3.62 (d, 1H), 3.66 (s, 3H), 3.54 (s, 3H), 3.37 (m, 1H), 3.18-3.12 (m, 1H), 2.86 (m, 2H), 2.64-2.70 (m, 5H), 2.36 (m, 2H), 1.74(m, 2H), 1.65 (m, 2H).

Example 18 in vitro cell Activity test of cephalotaxine Compounds

The cytostatic activity of cephalotaxine compounds was evaluated using the method described in the CCK-8 kit. After cells (3000-10000 cells/well) were inoculated on a 96-well cell culture plate for 24 hours, 100. Mu.L of compound solutions of different concentrations were added to each culture well, incubated for 72 hours, 10. Mu.L of CCK-8 solution was added to each culture well, further incubated for 2-3 hours, and absorbance values at 450nm and 650nm were measured with an enzyme-labeled instrument. Processing the raw data in the EXCELL table to obtain cell viability of each processing well. The viability data was then used to calculate IC using a nonlinear regression model on GraphPad Prism software ₅₀ Values. As a result, it was found that a part of cephalotaxine compounds can significantly inhibit the proliferation of Ba/F3-Bcr/Abl-T315I, K562, A549, U937, the half inhibition concentration (IC ₅₀ Table 1) correlated positively with drug concentration. The compounds used were the compounds prepared in examples 1-7, respectively.

IC of Table 1 Compounds against different tumor cells ₅₀ （μM）

The data in Table 1 show that the cephalotaxine compounds prepared by the invention have higher anti-tumor activity than the compounds CY5120068, CY5120024 and CY 5120069.

The above description of the embodiments is only for aiding in the understanding of the method of the present invention and its core ideas. It should be noted that it will be apparent to those skilled in the art that various modifications and adaptations of the invention can be made without departing from the principles of the invention and these modifications and adaptations are intended to be within the scope of the invention as defined in the following claims.

Claims (2)

1. A process for the preparation of cephalotaxine derivatives comprising the steps of:

s1) reacting phenylpyruvate or a phenylpyruvate derivative with benzyl alcohol to prepare benzyl 2-oxo-3-phenylpropionate or a derivative thereof;

s2) reacting benzyl 2-oxo-3-phenylpropionate or a derivative thereof with methyl (triphenylphosphine alkenyl) acetate to obtain 1-benzyl 4-methyl 2-benzyl maleate and (E) -1-benzyl 4-methyl 2-benzylidenesuccinate or a derivative thereof;

s3) reducing 1-benzyl 4-methyl 2-benzyl maleate and (E) -2-benzylidene succinic acid 1-benzyl 4-methyl ester with a reducing agent to obtain 2-benzyl-4-methoxy-4-oxo-butyric acid or a derivative thereof;

s4) reacting 2-benzyl-4-methoxy-4-oxo butyric acid or a derivative thereof with cephalotaxine to obtain a cephalotaxine derivative shown in formula I;

wherein R is ₁ Is O;

R ₂ 、R ₃ 、R ₄ independently selected from one or more of H, methoxy, F, cl, br, benzyloxy and hydroxyl;

the reducing agent is selected from palladium carbon and Ru [ (S) -DTBM-SegPhos](OAc) ₂ Or Ru [ (R) -DTBM-SegPhos](OAc) ₂ 。

2. A process for the preparation of cephalotaxine derivatives comprising the steps of:

a) Reacting dimethyl succinate with benzaldehyde or benzaldehyde derivatives, and acidifying and esterifying the product to obtain 1-methyl 2-benzylidene succinate or derivatives thereof;

b) Reducing the 2-benzylidene succinic acid 1-methyl ester or a derivative thereof to obtain 2-benzyl-4-methoxy-4-oxo butyric acid or a derivative thereof;

c) Reacting 2-benzyl-4-methoxy-4-oxobutyric acid or a derivative thereof with cephalotaxine to obtain a cephalotaxine derivative shown in formula I;

wherein R is ₁ Is O;

R ₂ 、R ₃ 、R ₄ and independently selected from one or more of H, methoxy, F, cl, br, benzyloxy and hydroxyl.