CN104987504B - Pegylation Lapatinib and its injection and preparation method - Google Patents

Abstract

The present invention provides Pegylation Lapatinib and its injection and preparation method.Pegylation Lapatinib, its general formula are：A is selected from single armed or the polyethylene glycol of multi-arm or the derivative of polyethylene glycol；X is selected from Y and M is separately selected from double carboxylic acids with amino or corresponding acyl group substituent；N is selected from amino acid or peptide；LPT is Lapatinib；A=0 or 1；B=0 or 1；C=1 or 2；D=1 or 2；E is equal to the arm number of X.The present invention has synthesized the pegylated medicament of Lapatinib, reduces the toxicity of Lapatinib, improves its water-soluble and biological stability, avoids the drug resistance of cancer cell and cancer stem cell, enhance targeting, significantly improve anticancer therapeutic.

Description

Pegylation Lapatinib and its injection and preparation method

Technical field

The present invention relates to the conjugate field of Lapatinib, in particular to Pegylation Lapatinib and its note Penetrate agent and preparation method.

Background technology

Lapatinib (Lapatinib) is a kind of oral small molecule epidermal growth factor receptor tyrosine kinase inhibition Agent, is mainly used for the treatment ErbB-2 overexpressions of joint capecitabine, and previously receives anthracycline, taxol or bent appropriate The late period of pearl monoclonal antibody treatment or metastatic breast cancer, were approved by the FDA in the United States into market in 2007.Lapatinib can suppress Cancer stem cell, such as U87-MG cancer of the brains stem cell and SUM225 breast carcinoma stem cells.

The molecular weight of Lapatinib is 581.06, belongs to small molecule anti-cancer drug, and there are water-soluble is poor, biological stability Difference, the shortcomings that toxic side effect is big, and the Lapatinib of small molecule can be by p- by diffusing into cancer cell or cancer stem cell Glycoprotein membrane pump goes out, and then causes the resistance to the action of a drug.And the macromolecule coupling anticancer stem cell small-molecule drug energy pole of long-acting slow-release The earth reduces the toxicity of small-molecule drug, greatly lifts its water-soluble and biological stability, strengthens its passive targeting, relatively The price for antibody coupling anticarcinogen (ADC), antibody anticarcinogen, antibody anticancer stem cell drugs or the antibody of immunization therapy It is cheap, and cancer cell or the extremely strong drug resistance of cancer stem cell can be avoided, this is because macromolecule coupling anticarcinogen or height Molecule coupling labeled anticancer stem cell drugs have larger molecular weight, and cancer cell or cancer stem cell can only pass through endocytosis and pinocytosis Effect absorbs it.But it is both anticarcinogen to be directed to and the macromolecule of the Lapatinib of anticancer stem cell drugs is coupled skill Art is reported and product is not yet found.

In view of this, it is special to propose the present invention.

The content of the invention

The first object of the present invention is to provide Pegylation Lapatinib, and the Pegylation Lapatinib is Lapatinib is coupled the macromolecule anticancer drug to be formed with polyethylene glycol and its derivative, and compared with Lapatinib, its toxicity is big Amplitude reduction, water-soluble and biological stability improve, have significant passive targeting, much lower drug resistance, higher it is anti- Cancer curative effect.

The second object of the present invention is to provide a kind of preparation method of the Pegylation Lapatinib, this method With the advantages that easy to operate, reaction condition is gentle, and reaction rate is fast and yield is high.

The third object of the present invention is to provide a kind of injection of the Pegylation Lapatinib, the injection Have the advantages that additive is few, dissolubility is high, drug effect is high, easy preparation.

In order to realize the above-mentioned purpose of the present invention, spy uses following technical scheme：

Pegylation Lapatinib, its general formula are：

A is selected from single armed or the polyethylene glycol of multi-arm or the derivative of polyethylene glycol；X is selected from Y and M separately substitutes selected from double carboxylic acids with amino or corresponding acyl group Thing；N is selected from amino acid or peptide；LPT is Lapatinib；

A=0 or 1；B=0 or 1；C=1 or 2；D=1 or 2；E is equal to the arm number of X.

From above-mentioned general formula, Pegylation Lapatinib is using amino acid as connection chain, by polyethylene glycol with drawing pa The macromolecular substances to be formed are coupled together for Buddhist nun, it not only remains Lapatinib anti-cancer properties, but also with single La Pa Compared for Buddhist nun and showed following advantage：Toxicity is greatly lowered, and water-soluble and biological stability improves, and has significantly by moving-target Tropism and much lower drug resistance.

In above-mentioned general formula, as c=2, represent to be connected to two N-LPT on M.

Preferably, when a, b are 0, c, d are 1；Work as a=0, during b=1, d=1.Idol with above design feature Join thing to be easier to prepare, medicine cost is low.The molecular structural formula of moiety thing is listed below：

Preferably, double carboxylic acids with amino are glutamic acid or aspartic acid.

Preferably, A is selected from the derivative of single armed, the polyethylene glycol of four arms or eight arms or polyethylene glycol.

Preferably, the molecular weight of A is 12000,20000 or 40000 dalton.

Preferably, Y and M are separately selected from

These materials are all the substituents of the amino position of glutamic acid, it is possible to achieve following two purposes：First, utilize paddy The more amidated LPT of the high activity acyl group of propylhomoserin or aspartic acid coupling, second, increase side chain quantity, to improve poly- second two Refine the drug release rate of Lapatinib.In addition, the dimethylaminoethoxyethanol on side chain can reduce and polyethylene glycol Combination difficulty.

Preferably, N is glycine, it is more easy to combine with Lapatinib, therefore product is more easy to obtain.

Preferably, molecular formula isIt is demonstrated experimentally that the medicine of the compound Comprehensive performance more preferably, is clinically more easy to promote.

The preparation method of above-mentioned Pegylation Lapatinib, comprises the following steps：

Step A：Make Lapatinib that amidation process occur with Amino Acid/Peptide, obtain the first intermediate product；

Step B：Make first intermediate product that amidation process occur with M, obtain the second intermediate product；

Step C：Make second intermediate product that amidation process occur with Y, obtain the 3rd intermediate product；

Step D：The 3rd intermediate product is coupled with the derivative of polyethylene glycol or polyethylene glycol by amido link, obtain To product.

Above-mentioned preparation method reduces the difficulty that polyethylene glycol and LPT are directly coupled using amino acid as connecting bridge, and Involved three classes reaction in building-up process (reaction, polyethylene glycol and Y between LPT and amino acid reaction, amino acid it is anti- Should) reaction condition it is all very gentle, therefore the requirement to equipment and production environment is low, and manufacturing cost is low.Further, since synthesized The mechanism reacted in journey is amidation process, and the high selectivity of amidation process, reaction rate is fast, the accessory substance of isomers class It is few, therefore the yield of the synthetic method is high, speed is fast.

Preferably, in the step A, the condition that amidation process occurs with peptide or arbitrary amino acid for Lapatinib is：With HBTU, HOBT, DMF and DIEA are auxiliary agent, are reacted at -2~4 DEG C, and Lapatinib, peptide or arbitrary amino acid, HBTU, Molar ratio between HOBT and DIEA is：1:0.8~1.2:1.3~1.7:1.3~1.7:4.2~4.8.

Above-mentioned reaction is using HBTU (O- benzotriazole-tetramethylurea hexafluorophosphoric acid ester) as condensing agent, HOBT (1- hydroxyls Base benzotriazole) compensation is used as, DIEA (n,N-diisopropylethylamine) is used as alkaloid, DMF (n,N-Dimethylformamide) As solvent, and with the reaction of specific ratio promotion Lapatinib and peptide/arbitrary amino acid, make the combination of acyl group and amino difficult Degree reduces, and reaction yield improves.

Preferably, Lapatinib is made to occur to further include before amidation process with peptide or amino acid：Use tertbutyloxycarbonyl (Boc) amino of the peptide or the amino acid is protected.

After above-mentioned safeguard measure can be substituted to avoid the amino of peptide/arbitrary amino acid, it can not be combined with other amino acid. Specific guard method is referred in the following manner：Amino acid is dissolved in Isosorbide-5-Nitrae-dioxane, it is molten to add sodium hydroxide Liquid, then di-tert-butyl dicarbonate (Boc is added at room temperature₂O), react at room temperature, be finally separating extraction product；Amino acid, The molar ratio of sodium hydroxide is 1:1.2-1.3.

Preferably, in the step A, the method that amidation process occurs with peptide for Lapatinib is：Lapatinib with it is described The reaction of free amino acid that the termination of peptide is related to, obtained product according still further to amino acid in the peptide sequence successively with the peptide The other free amino acids reaction being related to, and the free amino acid in above-mentioned reaction is before participating in reacting：With N- hydroxyl ambers Amber acid imide activates the carboxyl of the free amino acid.Reaction rate is improved by activating, improves yield, reduces coupling difficulty, So that post processing is more prone to.

Preferably, in the step B, the condition of first intermediate product and M amidation process is：Among described first Molar ratio >=2 of product and M.The first intermediate product for controlling suitable molar ratio to allow the M of a molecule to connect two molecules, makes The connection quantity of LPT is double.

Preferably, first intermediate product occurs to further include before amidation process with M：Use n-hydroxysuccinimide Activate the carboxyl of glutamic acid；Similarly, reaction rate is improved by activating, reduces coupling difficulty.It is similar, in described second Between product and Y amidation process occurs before can also be first with the carboxyl of n-hydroxysuccinimide activation Y.

Preferably, in the step C, the condition of second intermediate product and Y amidation process is：Among described second Molar ratio >=2 of product and glutamic acid.Equally it is that the ratio for controlling reactant increases the quantity of the LPT of combination.

Preferably, in the step D, the 3rd intermediate product is made to occur also to wrap before amidation process with polyethylene glycol Include：N, N'- succinimidyl carbonate, N, N'- succinimides yl acetate or N, N'- succinimidyl glutarat The derivative of activated polyethylene glycol or polyethylene glycol.To reduce the coupling difficulty of polyethylene glycol and LPT.

Preferably, the molar ratio of the 3rd intermediate product and polyethylene glycol is more than 1.5；Ensure the 3rd intermediate product with gathering Ethylene glycol is coupled completely.

Preferably, with N, N'- succinimidyl carbonates, N, N'- succinimides yl acetate or N, N'- succinyl The derivative of imido grpup glutarate activated polyethylene glycol or polyethylene glycol, obtains the 4th intermediate product, is produced among the described 3rd With the 4th intermediate product, using dichloromethane as solvent, at 20-30 DEG C amidation process occurs for thing：Select gentle reaction Condition reduces production difficulty.

The injection of Pegylation Lapatinib, including：The Pegylation Lapatinib and solvent, the solvent For physiological saline and/or 95% ethanol.The injection has the advantages that additive is few, dissolubility is high, drug effect is high, easy preparation.

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

(1) the macromolecule coupling drug of Lapatinib has been synthesized, the toxicity of Lapatinib active compound has been reduced, improves water-soluble Property and biological stability, avoid the very strong drug resistance of cancer cell and cancer stem cell, enhance targeting, significantly improve anti- Cancer curative effect.

(2) preparation method of Pegylation Lapatinib is easy to operate, reaction condition is gentle, reaction rate is fast and yield Height, suitable for industrialized production.

(3) different types of Pegylation Lapatinib injection is provided, for facing for Pegylation Lapatinib Bed application provides foundation.

Brief description of the drawings

In order to illustrate more clearly about the embodiment of the present invention or technical scheme of the prior art, below will be to embodiment or existing There is attached drawing needed in technology description to be briefly described.

Fig. 1 to Fig. 5 is respectively the curve of 6-56,4-208,6-162,7-129, LPT to U87 inhibition of cancer cell rates；

Fig. 6 to Figure 10 is the curve of 6-56,4-208,6-162,7-129, LPT to N87 inhibition of cancer cell rates；

Figure 11 to Figure 13 is the curve of 6-56,6-162, LPT to CALU-3 inhibition of cancer cell rates；

Figure 14 to Figure 18 is the curve of 6-56,4-208,6-162,7-129, LPT to BT474 inhibition of cancer cell rates；

Figure 19 to Figure 23 is the curve of 6-56,4-208,6-162,7-129, LPT to SK-BR-3 inhibition of cancer cell rates；

The growth curve of tumour when Figure 24 is physiological saline, 7-129, LPT act on mouse；

Figure 25 to Figure 42 is respectively the Pegylation Lapatinib that embodiment 1-18 is provided¹H-NMR spectrum；

Figure 43 to Figure 55 be respectively embodiment 1,3-5,9-17 provide Pegylation Lapatinib mass spectrogram.

Embodiment

Embodiment of the present invention is described in detail below in conjunction with embodiment, but those skilled in the art will Understand, the following example is merely to illustrate the present invention, and is not construed as limiting the scope of the invention.It is not specified in embodiment specific Condition person, the condition suggested according to normal condition or manufacturer carry out.Reagents or instruments used without specified manufacturer, is The conventional products that can be obtained by commercially available purchase.

To meet simplicity, clearly requiring, the part of compounds digital number below related to, such as 3-59,7-129, P1~P43 etc..

Embodiment 1

SynthesisMethod：

First, prepare

Glycine (19.1274g, 254.7943mmol) is dissolved in Isosorbide-5-Nitrae-dioxane (200mL), adds 2N NaOH solution (80uL, 160mmol), treats that solution temperature is cooled to room temperature, and adds Boc₂O (77.8524g, 356.7120mmol), When reaction 12 is small at room temperature, it is concentrated by evaporation, is finally washed with ethyl acetate (150mL × 2), pH value is adjusted to 4 with hydrochloric acid, then Extracted with ethyl acetate (150mL × 5), merge organic phase, dried with anhydrous sodium sulfate, concentrating filter liquor is dried to obtain product 44.6 grams, yield 100%.

2nd, prepare

In the flask of 50mL add P1 (90.4mg, 0.5163mmol), Lapatinib (300g, 0.5163mmol), HBTU (290.6mg, 0.7745mmol) and HOBT (104.6mg, 107745mmol), is dissolved with DMF (15mL), by solution 0 Cooled down 20 minutes in DEG C cryogenic thermostat reactive bath technique, DIEA (0.41mL, 2.3234mmol) is then slowly added dropwise.2 it is small when after move to Room temperature, is stirred overnight.Reaction solution is poured into saturation NaHCO₃In solution (200mL), it is extracted with ethyl acetate (200mL × 2).So Cleaned with saturated sodium bicarbonate solution (150mL × 2), finally cleaned with saturation NaCl solution (100mL) afterwards.With anhydrous MgSO₄ Dry, filtering, is concentrated under reduced pressure and is evaporated.Silica gel column chromatography, with 5%MeOH/ dichloromethane eluents.Product is collected, is evaporated 380.0mg yield 99.70%.

3rd, prepare

P2 (380.0mg, 0.5147mmol) is added in flask, is dissolved with dichloromethane (10mL), finally instills TFA (0.20mL, 2.5737mmol).It is stirred overnight at room temperature.Stop reaction, evaporated under reduced pressure, is dissolved with MeOH, adds sodium acid carbonate (211.2mg, 2.5137mmol), is stirred 10 minutes.Filter, the addition silica white (3 grams) into filtrate, is spin-dried for making solid molten Liquid, dry method loading.Column chromatography, first with 2%MeOH/ dichloromethane eluents, then with 1%TEA, 5%MeOH/ dichloromethane eluents. Product is collected, is concentrated under reduced pressure and is evaporated to obtain 209.9mg, yield 63.9%.

4th, 3-59 is prepared：

The polyethylene glycol M- of P3 (19.95mg, 0.025mmol) and molecular weight for 12000 single armed is added in cylinder bottle SC-12K (200mg, 0.01667mmol), is dissolved with dichloromethane (5mL), is stirred at room temperature one week, stops reaction, by mixture It is spin-dried for, the dissolving of product water is put into bag filter dialysis.Then product water solution decompression is concentrated, dichloromethane is used after vacuum drying Alkane dissolves, and adds anhydrous ether while stirring and is precipitated out, filtered, vacuum drying obtains product 136.3mg, yield 65.3%.

Embodiment 2

SynthesisMethod：

The polyethylene glycol M-SC- of P3 (9.6mg, 0.015mmol) and molecular weight for 20000 single armed is added in cylinder bottle 20K (200mg, 0.01mmol), is dissolved with dichloromethane (5mL), is stirred at room temperature one week, stops reaction, mixture is spin-dried for, will The dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, it is molten with dichloromethane after vacuum drying Solution, adds anhydrous ether and is precipitated out, filtered, vacuum drying obtains product 182.6mg, yield 89.0% while stirring.

Embodiment 3

SynthesisMethod：

P3 (9.6mg, 0.015mmol) is added in cylinder bottle, and the polyethylene glycol M- for the single armed that molecular weight is 40000 SC-40K (200mg, 0.01mmol), is dissolved with dichloromethane (8mL), is stirred at room temperature one week, stops reaction, mixture is revolved It is dry, the dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, dichloromethane is used after vacuum drying Alkane dissolves, and adds anhydrous ether while stirring and is precipitated out, filtered, vacuum drying obtains product 168.4mg, yield 83.1%.

Embodiment 4

Synthesis

The polyethylene glycol of P3 (19.1433mg, 0.03mmol) and molecular weight for 40,000 four arms is added in cylinder bottle 4ARM-SC-40K (200mg, 0.01mmol), is dissolved with dichloromethane (8mL), is stirred at room temperature two weeks, mixture is spin-dried for, will The dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, it is molten with dichloromethane after vacuum drying Solution, adds anhydrous ether and is precipitated out, filter, obtain product, vacuum drying obtains product 120mg, yield while stirring 55.8%.

Embodiment 5

Synthesis

P3 (101.9mg, 0.16mmol) is added in cylinder bottle, and the polyethylene glycol for eight arms that molecular weight is 40000 8ARM-SG-40K (400mg, 0.01mmol), is dissolved with dichloromethane (8mL), is stirred at room temperature 2 weeks, mixture is spin-dried for, will produce The dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, is dissolved after vacuum drying with dichloromethane, Anhydrous ether is added while stirring to be precipitated out, and is filtered, vacuum drying obtains product 236.9mg, yield 53.6%.

Embodiment 6

Synthesis

First, prepare

Pidolidone (20.0620g, 13.36mmol) is added in flask, is dissolved with water/DMF (100mL/200mL).Room Triethylamine (38.00mL, 272.71mmol) is added under temperature while stirring, then adds Boc₂O (30.36g, 139.09mmol), DMF (40mL) is eventually adding, is stirred overnight.Concentration, is dissolved with ethyl acetate (500mL), is neutralized with 1M HCl (100mL).Point Water phase is separated out, NaCl is added in water phase, untill it is no longer dissolved, water phase is extracted with ethyl acetate, isolates organic phase.Will Above-mentioned organic phase merges, and washes with water, is cleaned with saturated salt solution, is dried with anhydrous magnesium sulfate, filters, and evaporated under reduced pressure obtains product 19.75g yield 58.58%.

2nd, prepare

In the round-bottomed flask of 1L add P4 (20g, 80.8898mmol) and n-hydroxysuccinimide (37.2384g, 323.5592mmol), dissolved with dichloromethane (400mL), then add DMAP (1.9765g, 16.1780mmol), be placed in -5 DEG C cryogenic thermostat reactive bath technique in, be added portionwise DCC (66.7599g, 323.5592mmol) after twenty minutes, 1 it is small when after move to room Temperature, is stirred overnight.Filtering, filtrate is concentrated.Solvent homogenate wet method dress post 16cm × 8.5cm is with petroleum ether.Wet method loading.Column Chromatography, with 30% ethyl acetate/petroleum ether to 100% ethyl acetate gradient, TLC monitorings, potassium permanganate colour developing, collects production Product, are evaporated, vacuum drying oven it is dry final products 41.6462g, yield 100%.

3rd, prepare

P5 (223.2mg, 0.5056mmol) and P3 (709.7mg, 1.1223mmol) is added in barrel reactor, with two Chloromethanes dissolves, and is stirred overnight at room temperature.Direct wet method loading, silica gel column chromatography, with 2% NH₃·H₂O, 4%MeOH/ dichloromethane The NH of alkane to 3%₃·H₂O, 6%MeOH/ dichloromethane gradient, collect product point, are concentrated under reduced pressure and are evaporated to obtain product 620mg, Yield 47.7%.

4th,

P6 (620mg, 0.4168mmol) is added in flask, is dissolved with dichloromethane, addition TFA (0.1597mL, 2.084mmol), it is stirred overnight at room temperature.Evaporated under reduced pressure, is dissolved with MeOH, adds sodium acid carbonate (175.1mg, 2.084mmol) simultaneously Stirring 10 minutes.Filtering, silica white solid solution, dry method loading are made by filtrate.Column chromatography, with 2% NH₃·H₂O, 4% MeOH/ dichloromethane eluents.Product is collected, is concentrated under reduced pressure and is evaporated to obtain 413.3mg, yield 100%.

5th, prepare

In flask add P1 (5.0905g, 29.0587mmol) and n-hydroxysuccinimide (3.6788g, 31.9646mmol), dissolved with dichloromethane, add DMAP (355.0mg, 2.9059mmol), be placed in 0 DEG C of cryogenic thermostat In reactive bath technique, after twenty minutes add DCC (6.5953g, 31.9646mmol), 1 it is small when after move to room temperature, be stirred overnight.Point TLC Plate, is unfolded in 20% ethyl acetate/petroleum ether, and the reaction was complete for phosphomolybdic acid colour developing display.Filtering, it is heavy to be washed with dichloromethane Form sediment.The silica white of 20g is added in filtrate, is spin-dried for, dry method upper prop.With 30% ethyl acetate/petroleum ether to 50% ethyl acetate/ The ethyl acetate elution of petroleum ether to 100%, collects product point, is concentrated under reduced pressure and is evaporated to obtain product 4.8945g, yield 65.98%.

6th, prepare

P7 (220.2mg, 0.1587mmol) and P8 (43.213mg, 0.1587mmol) is added in cylinder bottle, uses dichloro Methane dissolves, and is stirred overnight at room temperature.Wet method loading column chromatography, uses 1.5%NH₃·H₂O, 3%MeOH/ dichloromethane are to 3% NH₃·H₂O, 6%MeOH/ dichloromethane gradient, collect product point, are concentrated under reduced pressure, and vacuum drying obtains product 213.9mg, production Rate 89.8%.

7th, prepare

In flask add P9 (213mg, 0.1385mmol), dissolved with dichloromethane, add TFA (53uL, 0.6925mmol), it is stirred overnight at room temperature.Evaporated under reduced pressure, is dissolved with MeOH, adds sodium acid carbonate (58.2mg, 0.6925mmol) Stirring 10 minutes.Filtering, silica white solid solution, dry method loading are made by filtrate.Column chromatography, uses 3%NH₃·H₂O, 6% MeOH/ dichloromethane eluents.Product is collected, is concentrated under reduced pressure and is evaporated to obtain 166.8mg, yield 100%.

8th, 3-137 is prepared

The polyethylene glycol of P10 (65.0mg, 0.045mmol) and molecular weight for 40,000 four arms is added in cylinder bottle 4ARM-SCM-40K (300mg, 0.015mmol), is dissolved with dichloromethane (8mL), is stirred at room temperature two weeks.Stop reaction, will be mixed Compound is spin-dried for, and the dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, is used after vacuum drying Dichloromethane dissolves, and adds anhydrous ether while stirring and is precipitated out, filtered, vacuum drying obtains product 289.6mg, yield 88.3%.

Embodiment 7

Synthesis

First, prepare

P7 (4.8g, 3.3052mmol) and P5 (694.7187mg, 1.5739mmol.) is added in flask, uses dichloromethane Alkane dissolves, and is stirred overnight at room temperature.Homogenate method dress silicagel column 25cm × 4.5cm.Post separation in wet method, uses 2%NH₃·H₂O/4% MeOH/ dichloromethane is to 2.5%NH₃·H₂O/5%MeOH/ dichloromethane eluents, collect product point, are concentrated under reduced pressure and are evaporated and must produce Product 3.3147g, yield 75.9%.

2nd, prepare

P11 (2.8356g, 0.9497mmol) is added in flask, is dissolved with dichloromethane (50mL), under stirring at room temperature TFA (0.728mLL, 9.4967mmol) is slowly added to, is stirred overnight.Evaporated under reduced pressure, crude product is dissolved with absolute methanol, is added Sodium acid carbonate (797.8177mg, 9.4967mmol) stirring neutralizes.Filtering, silica white solid solution is made by filtrate.Homogenate method Fill silicagel column 20cm × 4.5cm.Dry method upper prop, uses 2.5%NH₃.H₂O/5%MeOH/ dichloromethane is to 5%NH₃.H₂O/10% MeOH/ dichloromethane gradients, collect product, are concentrated under reduced pressure and are evaporated to obtain product 2.5983g, yield 94.8%.

3rd, prepare

P12 (95mg, 0.0328mmol) and P8 (9.37mg, 0.0344mmol) is added in cylinder bottle, uses dichloromethane (10mL) dissolves, and is stirred overnight at room temperature.Dichloromethane wet method homogenate dress column 20cm × 2cm.Wet method upper prop, column chromatography, with 2% NH₃·H₂O, 3%MeOH/ dichloromethane are to 3%NH₃·H₂O, 6%MeOH/ dichloromethane gradient, collect product point, subtract Pressure concentration vacuum drying obtains product 99.8mg, yield 100%.

4th, prepare

P13 (99.8mg, 0.0328mmol) is added in flask, is dissolved with dichloromethane, addition TFA (12.6uL, 0.1640mmol), it is stirred overnight at room temperature.Evaporated under reduced pressure, is dissolved with MeOH, adds sodium acid carbonate (0.3g), is stirred 10 minutes.Cross Filter, silica white solid solution is made by filtrate.Dichloromethane wet method homogenate dress column 25cm × 2.5cm.Column chromatography, uses 4%NH₃· H₂O, 8%MeOH/ dichloromethane are to 5%NH₃·H₂O, 10%MeOH/ dichloromethane gradient, collect product point, and decompression is dense Contracting vacuum drying obtains product 96.5mg, yield 100%.

5th, 3-153 is prepared

The polyethylene glycol of P14 (110.5mg, 0.03759mmol) and molecular weight for 40,000 four arms is added in cylinder bottle 4ARM-SCM-40K (250.6mg, 0.006266mmol), is dissolved with dichloromethane (8mL), is stirred at room temperature two weeks, by mixture It is spin-dried for, the dissolving of product water is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, dichloro is used after vacuum drying Methane dissolves, and adds anhydrous ether while stirring and is precipitated out, filtered, vacuum drying obtains product 252.36mg, yield 77.2%.

Embodiment 8

Synthesis

First, prepare

H is added into the flask of 500mL₂NCH₂CH₂OCH₂CH₂OH (20g, 190.223mmol), uses CH₂Cl₂(300mL) Dissolving, adds triethylamine (53mL, 380.446mmol), Boc is added portionwise under magnetic agitation₂O (49.8192 grams, 228.2670mmol), it is stirred overnight at room temperature.TLC monitorings show that raw material exhausts.Directly it is evaporated, is dissolved with methanol, adds carbonic acid Hydrogen sodium (31.9575g, 380mmol) neutralizes triethylamine, and filtering, adds 60g silica whites and be spin-dried in filtrate.It is wet with petroleum ether Method homogenate dress column 10cm × 8.5cm.Dry method loading, with 20% ethyl acetate/petroleum ether to 50% ethyl acetate/petroleum ether and 2% methanol/ethyl acetate gradient elution.Product is collected, is concentrated under reduced pressure and is evaporated to obtain 35.4485g, yield 90.79%.

2nd, prepare

Under nitrogen protection, P16 (35.4685g, 172.803mmol) is added into the there-necked flask of 1L, with THF (200mL) Dissolving, mechanical agitation, is placed in 0 DEG C of cryogenic thermostat reactive bath technique.After twenty minutes, be added portionwise 60% NaH (27.6485g, 690.12mmol), a large amount of bubbles are released.After twenty minutes, bromoacetic acid (36.0165g, 259.2045mmol) is added, is released a large amount of Simultaneously there is white slurry precipitation in bubble.After adding, 60 DEG C of oil bath overnights are moved to.The reaction was complete for TLC detections display.Water is added dropwise (30mL) neutralizes remaining NaH, adds water (300mL), water phase and organic phase is separated, with petroleum ether/anhydrous ether (100mL × 2) wash water phase.Water phase pH to 4 is adjusted with 1M HCl.Then NaCl is added to solution saturation.Water phase is extracted with ethyl acetate (100mL×12).Dried, filtered with anhydrous sodium sulfate, be concentrated under reduced pressure dry product 20.4653g, yield 44.99%.

3rd, prepare

P16 (20.4653g, 77.729mmol), n-hydroxysuccinimide are added in the flask of 500mL (10.7311g, 93.275mmol) and DMAP (0.9488g, 7.7729mmol), is dissolved with dichloromethane (200mL), is placed in 0 DEG C Cryogenic thermostat reactive bath technique in stir, after 30 minutes add DCC (64.1513g, 310.916mmol).3 it is small when after move to room temperature, It is stirred overnight.TLC shows that raw material exhausts.Filtering, addition 60g silica whites are spin-dried in filtrate.Dress column is homogenized with petroleum ether wet method 20cm×8.8cm.Dry method loading.With the ethyl acetate gradient of 20% ethyl acetate/petroleum ether to 100%, product is collected Point, is concentrated and dried to obtain product 20.6374g, yield 73.88%.

4th, prepare

P10 (644mg, 0.4458mmol) and P17 (1.6050g, 4.4583mmol) is added in 50mL flasks, uses dichloro Methane dissolves, and is stirred overnight at room temperature.5g silica whites are added after the completion of reaction to be spin-dried for.With dichloromethane wet method be homogenized dress column 20cm × 3.5cm.Dry method loading, column chromatography.Use 1%NH₃·H₂O, 2%MeOH/ dichloromethane are to 5%NH₃·H₂O, 10%MeOH/ bis- Chloromethanes gradient elution, collects product point, and be concentrated under reduced pressure dry product 169.4mg, yield 22.27%.

5th, prepare

P18 (320mg, 0.1894mmol) and TFA (0.145mL, 1.8939mmol) is added in 50mL flasks, uses dichloro Methane dissolves, and is stirred overnight at room temperature.TLC display reactions are completed, and are directly evaporated, are dissolved with MeOH, add sodium acid carbonate (2.3863g, 2.841mmol) neutralizes unnecessary TFA, and 5g silica whites are added after the completion of reaction and are spin-dried for.It is even with dichloromethane wet method Slurry dress column 15cm × 2.5cm.Dry method loading, column chromatography.Use 2%NH₃·H₂O, 4%MeOH/ dichloromethane are to 5%NH₃·H₂O, 10%MeOH/ dichloromethane gradients, collect product point, and be concentrated under reduced pressure dry product 228.7mg, yield 76.02%.

6th, 6-135-2 is prepared

P19 (160mg, 0.1006mmol, 10.0) and molecular weight are added in 30mL cylinder bottles as the poly- of 40,000 four arms Ethylene glycol 4ARM-SCM-40K (400mg, 0.01mmol), is dissolved with dichloromethane (3mL), is stirred at room temperature two weeks.By mixture It is spin-dried for, product is dissolved with water, is put into bag filter dialysis.Then the aqueous solution of product is concentrated under reduced pressure, with two after vacuum drying Chloromethanes dissolves, and adds anhydrous ether while stirring and is precipitated out, filtered, vacuum drying obtains product 432.1mg, yield 100%.

Embodiment 9

Synthesis

First, prepare

6-aminocaprolc acid (50g, 381.1847mmol) is dissolved in Isosorbide-5-Nitrae-dioxane in the round-bottomed flask of 1L (300mL), adds 2N NaOH aqueous solutions (18.2967g/230mL, 457.4217mmol) in the case where 0 DEG C of condition stirs, will react Room temperature is moved to, under agitation, adds Boc₂O, 12 it is small when after, reaction solution is concentrated into 30%, Ran Houjia of original system Enter water (300mL), (500mL × 2) are cleaned with ethyl acetate, water phase PH is adjusted to 4 or so with 1M HCl, then uses ethyl acetate Extract (500mL × 5), merge organic phase, use anhydrous Na₂SO₄When drying 2 is small or so, filtering, filtrate is spin-dried for, with toluene azeotropic Water removal three times, is drained under vacuum pump, and 103.79 grams of product, yield 100% are obtained when then vacuum drying oven drying 5 is small.

2nd, prepare

By P20 (20g, 86.4230mmol), n-hydroxysuccinimide (39.7857g, 345.6918mmol) and DMAP (2.1166g, 17.2846mmol) is added in the round-bottomed flask of 2L, is added dichloromethane (500mL), is stirred under the conditions of -5 DEG C Mix 10 minutes, then rapidly join DCC (71g, 345.6918mmol) in batches, when stirring 1 is small under the conditions of -5 DEG C, reaction is moved To being stirred overnight at room temperature.Filtering, filtrate is evaporated, is dissolved with ethyl acetate, add about 60g silica whites be spin-dried for making solid it is molten Liquid, dry method upper prop.Column chromatography, with the ethyl acetate/petroleum ether gradient elution of 10% ethyl acetate/petroleum ether to 50%.Collect Product, is concentrated under reduced pressure and is evaporated to obtain 17.8416 grams of product, yield 64%.

3rd, prepare

P7 (2.5g, 1.7215mmol) and P21 (1.1292g, 3.4428mmol) are placed in 200mL round-bottomed flasks, added Enter dichloromethane (50mL), be stirred overnight at room temperature.In reaction according to TLC monitoring add P21 (649.599mg, 2.0658mmol).After reaction, concentrate.Solvent homogenate wet method dress post is done with dichloromethane.Wet method loading.Column chromatography, is used 1%NH₃·H₂O/2% ethanol/methylenes are to 2%NH₃·H₂O/4% ethanol/methylene gradient elutions, collect product, subtract Pressure concentration is evaporated to obtain 2.3933 grams of product, yield 83.4%.

4th, prepare

P22 (2.1g, 1.2611mmol) is placed in 250mL round-bottomed flasks, dichloromethane (20mL) is added, is stirred in room temperature Mix down and be slowly added to TFA (12.6115mmol, 0.9664mL).After reaction, reaction solution is spin-dried for, is dissolved with absolute methanol, Adding sodium bicarbonate solid (12.6115mmol, 1.0595g) and neutralize unnecessary TFA, filtering, adds 6g silica whites in filtrate, Do solid solution.Solvent homogenate dress column 10cm × 3.5cm is with dichloromethane.Dry method loading.Column chromatography, is first washed with dichloromethane It is de-, then use 3%NH₃·H₂O/6% ethanol/methylenes are eluted to 5%NH₃·H₂O, 10% ethanol/methylene gradient are washed It is de-, product is collected, is concentrated under reduced pressure and is evaporated to obtain 1.5238 grams of product, yield 100%.

5th, 4-181 is prepared

The polyethylene glycol 4ARM-SCM-40K (2.5g, 0.0625mmol) of four arms is placed in 40mL straight tube bottles, Ran Houjia Enter about dichloromethane (30mL), add P23 (782.57mg, 0.5mmol) under stirring at room temperature, since solubility is bad, add DMF (2mL), at room temperature shading are slowly stirred 2 weeks.Mixture is spin-dried for, the dissolving of product water is put into bag filter dialysis.Then The aqueous solution of product is concentrated under reduced pressure, is dissolved after vacuum drying with dichloromethane, anhydrous ether is added while stirring and is precipitated Out, filter, vacuum drying obtains product 2.4836g, yield 96.7%.

Embodiment 10

Synthesis

Single armed polyethylene glycol M-SC-12K (1.0g, 0.08333mmol) is placed in 50mL straight tube bottles, adds dichloromethane (30mL), adds P23 (260.9mg, 0.1667mmol) under stirring at room temperature, since solubility is bad, adds DMF (10mL), Shading is slowly stirred 2 weeks at room temperature.After reaction, mixture is spin-dried for, the dissolving of product water is put into bag filter dialysis.So The aqueous solution of product is concentrated under reduced pressure afterwards, is dissolved after vacuum drying with dichloromethane, anhydrous ether is added while stirring and is sunk Shallow lake comes out, and filtering, vacuum drying obtains product 933.3mg, yield 93%.

Embodiment 11

Synthesis

First, prepare

P3 (2g, 3.1343mmol), P1 (0.5491g, 3.1343mmol), HBTU are added in the flask of 500mL (1.7644mg, 4.7015mmol) and HOBT (635.3mg, 4.7015mmol), is dissolved with DMF (20mL), and solution is low at 0 DEG C Cooled down after twenty minutes in warm isothermal reaction bath, DIEA (2.46mL14.1044mmol) is added dropwise.3 it is small when after move to room temperature, it is stirred Night.Reaction solution is poured into saturation NaHCO₃Solution (200mL), is extracted with ethyl acetate (200mL × 2).Organic phase saturated carbon Sour hydrogen sodium solution (150mL) cleaning, is cleaned with saturation NaCl solution (100mL).With anhydrous MgSO₄Dry, filtering, is concentrated under reduced pressure Solution is evaporated, is then dissolved with dichloromethane.Dress column 10cm × 3.5cm is homogenized with dichloromethane wet method.Wet method loading.Silica gel Column chromatography.With 2%MeOH/ dichloromethane to 4%MeOH/ dichloromethane gradients.Collect product and be evaporated to obtain 2.2062g, produce Rate 88.5%.

2nd, prepare

P24 (2.2062g, 2.7778mmol) is added in 50mL flasks, is dissolved with dichloromethane (5mL), adds TFA (0.212mL, 27.778mmol), is stirred overnight at room temperature.TLC display reactions do not complete, and add 1.5mL.Next day reaction is completed, rotation It is dry, dissolved with MeOH, add sodium acid carbonate (3g) and neutralize, 15g silica whites are added after the completion of reaction and are spin-dried for.With dichloromethane wet method Homogenate dress column 30cm × 3.5cm.Dry method loading.Column chromatography, first with 2%MeOH/ dichloromethane eluents, then uses 1%NH₃.H₂O, 2%MeOH/ dichloromethane is to 4%NH₃.H₂O, 8%MeOH/ dichloromethane gradient, collect product point, are concentrated under reduced pressure, vacuum Dry to obtain product 1.639g, yield 84.88%.

3rd, 6-162 is prepared

The polyethylene glycol 4ARM- of P25 (1.3903g, 2.00mmol) and molecular weight for 40,000 four arms is added in flask SCM-40K (10g, 0.25mmol), is dissolved with dichloromethane (10mL), stirred at room temperature.Some insoluble matters, add DMF It is completely dissolved after (80mL).Reaction is moved in the flask of 2L after two weeks, magnetic agitation adds ether 1.5L, and product is analysed completely Go out, continue magnetic agitation 10min, filter to obtain product, dissolved with dichloromethane, add anhydrous ether while stirring and be settled out Come, filtering, vacuum drying obtains product 8.9895g, yield 88%.

Embodiment 12

Synthesis

First, prepare

P25 (5.0161g, 7.2157mmol) and P5 (1.5167g, 3.4361mmol) are placed in 500mL round-bottomed flasks, Dichloromethane (200mL) is added, reaction is stirred at room temperature.After reaction, add 10g silica whites be spin-dried for making solid it is molten Liquid.Dry method loading, column chromatography.First use 0.5%NH₃·H₂O/1% ethanol/methylenes are to 4%NH₃·H₂O/8% methanol/bis- Chloromethanes gradient elution, collects product, is concentrated under reduced pressure and is evaporated to obtain product 4.006g, yield 34.8%.

2nd, prepare

P26 (5.4g, 3.3718mmol) is added in 500mL round-bottomed flasks, is dissolved with dichloromethane (50mL), room temperature is stirred TFA (2.58mL, 33.7176mmol) is slowly added dropwise under the conditions of mixing, is stirred overnight.Reaction terminates, and evaporated under reduced pressure, crude product is used Absolute methanol dissolves, and adds sodium bicarbonate solid (2.8g, 33.7176mmol) and neutralizes, filtering, and 15g silica whites are added in filtrate It is spin-dried for.Solvent homogenate wet method dress post 15cm × 4.5cm is with dichloromethane.Dry method loading.Column chromatography, uses 1%NH₃.H₂O/2% MeOH/ dichloromethane is to 5%NH₃.H₂O/10%MeOH/ dichloromethane gradients, collect product, are concentrated under reduced pressure, vacuum drying Obtain net product 4.6g, yield 92%.

3rd, prepare

P27 (3.4114g, 2.2719mmol) and P21 (2.2356g, 6.8158mmol) are placed in 500mL round-bottomed flasks In, dichloromethane (200mL) is then added, at room temperature stirring reaction.In reaction according to TLC monitoring add P21 (863.4mmg, 2.6323mmol).Concentrate after reaction.Solvent homogenate wet method dress post is done with dichloromethane.Wet method loading.Column chromatography, with 2% NH₃·H₂O/4% ethanol/methylenes are to 6%NH₃·H₂O/12% methanol/CH₂Cl₂Gradient elution, collects product, and decompression is dense Contracting is evaporated to obtain 2.3933 grams of product, yield 83.4%.

4th, prepare

P28 (2.5g, 1.4581mmol) is added in 500mL round-bottomed flasks, is dissolved with dichloromethane (50mL), room temperature is stirred Mix down and TFA (1.117mL, 14.5812mmol) is slowly added dropwise, be stirred overnight at room temperature.It is spin-dried for after reaction, by crude product nothing Water methanol dissolves, and adds sodium acid carbonate (1.2249g, 14.58124mmol) and neutralizes remaining TFA, filtering, adds in filtrate 8g silica whites make solid solution.Solvent homogenate wet method dress post is done with dichloromethane.Dry method loading.Column chromatography, uses 2%NH₃· H₂O/4% ethanol/methylenes are to 5%NH₃·H₂O/10% ethanol/methylene gradient elutions, collect product, are concentrated under reduced pressure It is evaporated to obtain 3.1 grams of product, yield 100%.

5th, 4-199 is prepared

The polyethylene glycol 4ARM-SCM-40K (1.0g, 0.02409mmol) for four arms that molecular weight is 40000 is placed in 30mL In straight tube bottle, dichloromethane (20mL) is then added, adds P29 (311.2mg, 0.19275mmol) under stirring at room temperature, due to Solubility is bad, adds DMF (2mL).Shading is slowly stirred reaction 2 weeks at room temperature.After reaction, add diethyl ether sedimentation, filtering, Crude product is dissolved with dichloromethane, then the sedimentation that adds diethyl ether, filtering, is collected product drying and is obtained 1.0322g, yield 100%.

Embodiment 13

Synthesis

The polyethylene glycol M-SC-12K (1.0g, 0.08333mmol) of single armed is placed in 50mL straight tube bottles, then adds two Chloromethanes (30mL), is stirred at room temperature lower addition P29 (269.1mg, 0.1667mmol), since solubility is bad, adds DMF (10mL), at room temperature shading are slowly stirred reaction 2 weeks.After reaction, add diethyl ether sedimentation, filtering, crude product dichloromethane Dissolving, then settled with ether, collect product drying and obtain 847.6mg, yield 76%.

Embodiment 14

Synthesis

First, prepare

P27 (4.4g, 2.9303mmol) and P5 (616.0mg, 51.3954mmol) are placed in 500mL round-bottomed flasks, so Dichloromethane (100mL) is added afterwards, and reaction is stirred at room temperature.To after reaction, stop reaction, add 20g silica whites and do Solid solution.Solvent homogenate wet method dress post 12cm × 8.5cm is with dichloromethane.Dry method loading.Column chromatography, uses 3%NH₃· H₂O/6%MeOH/ dichloromethane is to 5%NH₃·H₂O/10%MeOH/ dichloromethane gradients, collect product, are concentrated under reduced pressure It is evaporated to obtain product 1.5g, yield 17%.

2nd, prepare

P31 (1.5g, 0.4725mmol) is added in 500mL round-bottomed flasks, is dissolved with dichloromethane (50mL), in room temperature TFA (0.3621mL, 4.72526mmol) is slowly added dropwise under stirring condition, is placed in and is stirred overnight at room temperature.Reaction terminates, and stops anti- Should, it is spin-dried for, crude product is dissolved with absolute methanol, add sodium acid carbonate and neutralize, filter, proper silica gel powder is added in filtrate and is made Solid solution.Solvent homogenate wet method dress post 3.5cm × 7cm is with dichloromethane.Dry method loading.Column chromatography, uses 3%NH₃·H₂O/ 6%MeOH/ dichloromethane is to 4%NH₃·H₂O/8%MeOH/ dichloromethane gradients, collect product, and the vacuum that is concentrated under reduced pressure is dried 2.2 grams are done to obtain, yield 100%.

3rd, prepare

P32 (2.5g, 0.8663mmol) and P21 (852.4mg, 2.5989mmol) is added in 250mL round-bottomed flasks, is used Dichloromethane (50mL) dissolves, and is stirred overnight at room temperature.Wet method dress silicagel column 12cm × 4.5cm.Wet method loading after the reaction was complete. Use 2%NH₃·H₂O/4%MeOH/ dichloromethane is to 3%NH₃·H₂O/6%MeOH/ dichloromethane gradients, collect product Point, is concentrated under reduced pressure and is evaporated to obtain product 26212g, yield 976%.

4th, prepare

P32 (200mg, 0.06454mmol) is added in 50mL round-bottomed flasks, is dissolved with dichloromethane (5mL), room temperature is stirred Mix down and be slowly added to TFA (0.04945mL, 0.6454mmol), be stirred overnight.Decompression is spin-dried for, and crude product is dissolved with methanol, is added Enter sodium acid carbonate stirring and neutralize TFA.Filtering, silica white solid solution is made by filtrate.Homogenate method dress silicagel column 4.5cm × 7.4cm.Dry method loading.Column chromatography, first with dichloromethane eluent, then uses 2%MeOH/CH₂Cl₂Washed to 5%MeOH/ dichloromethane It is de-, then use 2%NH₃·H₂O/5%MeOH/ dichloromethane is to 4%NH₃·H₂O/8%MeOH/ dichloromethane gradients.Receive Collect product, be concentrated under reduced pressure and be evaporated to obtain product 137.4mg, yield 70.9%.

5th, 4-226 is prepared

Take the polyethylene glycol M-SCM-40K (352.5mg, 0.008813mmol) of single armed to be placed in straight tube bottle, add P33 (72.6mg, 0.02421mmol), is dissolved with dichloromethane (5mL), is stirred 2 weeks under room temperature lucifuge.Concentration, adds ether sedimentation, Filter to obtain crude product；Crude product is dissolved with dichloromethane, adds ether sedimentation, vacuum drying obtains 150mg, yield 43%.

Embodiment 15

Synthesis

First, prepare

P3 (7.1g, 11.1267mmol) is taken to be placed in 500mL flasks, addition Boc-Leu (2.5735g, 11.1267mmol), HBTU (6.2636g, 16.6900mmol), HOBT (2.2552g, 16.6900mmol) and DMF (300mL), Stirred 30 minutes under the conditions of 0 DEG C, be added dropwise DIEA (8.7218mL, 50.0702mmol), 1 it is small when after move to it is stirred at room temperature Night.Reaction solution is poured into saturated sodium bicarbonate solution, is extracted with ethyl acetate (400mL), water mutually uses ethyl acetate again (200mL × 3) extract.Merge organic phase, cleaned (200mL × 2) with saturated sodium bicarbonate solution, then it is clear with saturation NaCl solution Wash (200mL × 2).The anhydrous MgSO of organic phase₄Dry, filtering, is evaporated.Crude product wet method loading crosses silica gel column purification, with 3% Ethanol/methylene collects product, is concentrated and dried to obtain 9.96g, yield 100% to 5% ethanol/methylene gradient elution.

2nd, prepare

Take P34 (9.8394g, 11.5568mmol) to be placed in 500mL flasks, add dichloromethane (70mL) dissolving, add TFA (10mL), is stirred overnight at room temperature.It is evaporated, crude product is dissolved with methanol, adds TFA in sodium acid carbonate and unnecessary.Cross Filter, adds silica white (40g) into filtrate, solid solution is made in revolving, and dry method loading crosses silica gel column purification.First with 3% methanol/ Dichloromethane to 5% ethanol/methylene elutes, then uses 2%NH₃·H₂O/5% ethanol/methylenes are to 3%NH₃·H₂O/ 7% ethanol/methylene gradient elution, collects product, is concentrated and dried to obtain 6.75 grams, and 77.74%.

3rd, prepare

P35 (6.657g, 8.8610mmol) is taken to be placed in 500mL flasks, addition Boc-Phe (2.3509g, 8.8610mmol), HBTU (4.9882g, 13.2915mmol), HOBT (1.7960g, 13.2915mmol) and DMF (400mL), Magnetic agitation 30 minutes at 0 DEG C, be added dropwise DIEA (6.9485mL, 39.8745mmol), 1 it is small when after move to room temperature, it is stirred Night.Reaction solution is poured into saturated sodium bicarbonate solution, is extracted with ethyl acetate (400mL), water mutually uses ethyl acetate again (200mL × 3) extract.Merge organic phase, cleaned (200mL × 3) with saturated sodium bicarbonate solution, then it is clear with saturation NaCl solution Wash (200mL × 3).Organic phase is with anhydrous MgSO₄Dry, filtering, is evaporated.Crude product is dissolved with dichloromethane, wet method loading mistake Silica gel column purification.First use dichloromethane eluent, then with 2% ethanol/methylene to 7% ethanol/methylene gradient elution, receive Collect product, be concentrated and dried to obtain 8.9151g, yield 100%.

4th, prepare

Take P36 (8.8209g, 8.8335mmol) to be placed in 250mL flasks, add dichloromethane (70mL), add TFA (6.7688mL, 88.3350mmol), is stirred overnight at room temperature.It is evaporated, crude product is dissolved with methanol, is added sodium acid carbonate and is neutralized, Filtering, adds silica white (60g) into filtrate, solid solution is made in revolving, and dry method loading crosses silica gel column purification.First use dichloromethane Alkane elutes, and is then eluted with 5% ethanol/methylene, then use 2%NH₃·H₂O/5% ethanol/methylenes are to 4%NH₃· H₂O/8% ethanol/methylene gradient elutions, collect product, are concentrated and dried to obtain 8.8842g, yield 100%.

5th, prepare

Take P37 (8.7882g, 9.7815mmol) to be placed in 1L flasks, add P1 (1.7135g, 9.7815mmol), HBTU (5.5063g, 14.6723mmol), HOBT (1.9825g, 14.6723mmol) and DMF (400mL), 30 are stirred under the conditions of 0 DEG C Minute, be added dropwise DIEA (7.6673mL, 44.0168mmol), 2 it is small when after move at room temperature, be stirred overnight.Reaction solution is poured into In saturated sodium bicarbonate solution, extracted with ethyl acetate (500mL), water is mutually extracted with ethyl acetate (200mL × 3) again.Merge Organic phase is cleaned (200mL × 3) with saturated sodium bicarbonate solution, then is cleaned (200mL × 3) with saturation NaCl solution.Organic phase With anhydrous MgSO₄Dry, filtering, is evaporated, crude product is dissolved with dichloromethane, and wet method loading crosses silica gel column purification.First use dichloro Methane elutes, and then with 2% ethanol/methylene to 7% ethanol/methylene gradient elution, collects product, is concentrated and dried Product 8.2468g, yield 79.87%.

6th, prepare

Take P38 (8.1332g, 10.1936mmol) to be placed in 250mL flasks, add dichloromethane (70mL) dissolving, slowly TFA (10mL) is added, is stirred overnight at room temperature.Next day adds TFA (2mL), stops reaction after the completion of reaction, is evaporated, crude product Dissolved with methanol, add sodium acid carbonate and neutralize, filtering, adds silica white (60g) into filtrate, be spin-dried for that solid solution is made, do Method loading crosses silica gel column purification.Dichloromethane eluent is first used, is then eluted with 2% ethanol/methylene, finally uses 1%NH₃· H₂O/3% ethanol/methylenes are to 4%NH₃·H₂O/8% ethanol/methylene gradient elutions, collect product, are concentrated and dried Product 6.4309g, yield 90.42%.

7th, 7-129 is prepared

Take the polyethylene glycol 4ARM-SCM-40K (3.9847g, 0.09962mmol) of four arms to be placed in 500mL flasks, add P39 (0.7616g, 0.7970mmol), is dissolved with dichloromethane (80mL), is stirred 2 weeks under room temperature lucifuge.Concentration, adds ether Sedimentation, filters to obtain crude product, crude product is dissolved with dichloromethane, then is settled with ether, and filtration drying obtains product 4.3973g, Yield 92.93%.

Embodiment 16

Synthesis

First, prepare

Take P39 (3.6671g, 3.8379mmol) to be placed in 250mL flasks, add P5 (806.7mg, 1.8275mmol), Then dichloromethane (50mL) is added, is stirred 5 days at room temperature.Add methanol to dissolve product, then add silica white (30g) system Into solid solution, dry method loading crosses silica gel column purification.Use 0.5%NH₃·H₂O/1% ethanol/methylenes are to 4%NH₃·H₂O/ 7% ethanol/methylene gradient elution, collects product, is concentrated and dried to obtain product 3.55g, yield 83.62%.

2nd, synthesize

Take P40 (3.5g, 1.6492mmol) to be placed in 500mL flasks, add dichloromethane (50mL) dissolving, then slowly add Enter TFA (2mL), be stirred overnight at room temperature, it is rear to add TFA (4mL) again, overnight.Stop reaction, be spin-dried for, crude product methanol Dissolving, adds sodium acid carbonate and neutralizes, filtering, adds silica white (40g) into filtrate, is spin-dried for that solid solution, dry method loading is made Cross silica gel column purification.0.5L dichloromethane eluents are first used, then use 2%NH₃·H₂O/5% ethanol/methylenes are to 4%NH₃· H₂O/8% ethanol/methylene gradient elutions, collect product, are concentrated and dried to obtain product 3.1333g, yield 93.96%.

3rd, prepare

Take P41 (3.0423g, 1.5045mmol) to be placed in 100mL flasks, add P21 (0.9870g, 3.0091mmol) With dichloromethane (20mL), stir 4 days at room temperature.Concentration, then adds n-hexane sedimentation, and filtration drying obtains product 2.9822g, Yield 8869%.

4th, prepare

Take P42 (2.8756g, 1.2866mmol) to be placed in 500mL flasks, add dichloromethane (20mL), then slowly drop Add TFA (2mL), be stirred overnight at room temperature.It is spin-dried for, crude product is dissolved with methanol, adds TFA in sodium acid carbonate and unnecessary.Cross Solid impurity is filtered out, silica white (30g) is added into filtrate, is spin-dried for that solid solution is made, dry method loading crosses silica gel column purification. Dichloromethane eluent is first used, is then eluted with 2% ethanol/methylene to 5% ethanol/methylene, then use 2%NH₃·H₂O/ 5% ethanol/methylene is to 5%NH₃·H₂O/10% ethanol/methylene gradient elutions, collect product, being concentrated and dried to produce Product 1.9896g, yield 72.43%.

5th, 7-172 is prepared

Take the polyethylene glycol 4ARM-SCM-40K (2.5g, 0.0625mmol) of four arms to be placed in straight tube bottle, add dichloromethane Alkane (20mL) and P43 (1.0724g, 0.5mmol), stir 2 weeks under room temperature lucifuge, are transferred in 200mL flasks with DMF and reacted again 7 days.Concentrated by rotary evaporation, then adds anhydrous ether sedimentation, filters to obtain crude product；Then it is crude product dichloromethane and methanol is molten Solution, adds ether sedimentation, and filtration drying obtains product 1.7192g.

Embodiment 17

Synthesis

Take the polyethylene glycol M-SC-12K (1g, 0.0833mmol) of single armed to be placed in straight tube bottle, add dichloromethane (5mL) With P43 (357.4796mg, 0.1667mmol), stir 2 weeks under room temperature lucifuge, be transferred in 100mL flasks that the reaction was continued with DMF 9 days.Concentrated by rotary evaporation, adds anhydrous ether sedimentation, filters to obtain crude product；Then crude product dichloromethane and methanol are dissolved, Ether sedimentation is added, filtration drying obtains product 0.9927g, yield 84.93%.

Embodiment 18

Synthesis

First, prepare

L-Aspartic acid (15g, 112.70mmol) H2O/DMF (150mL/120mL) is added in flask to dissolve.Room temperature Under the conditions of add triethylamine (31.41mL, 225.39mmol) while stirring, then add Boc2O (25.09g, 114.95mmol), DMF (60mL) stirrings are eventually adding, overnight.Directly it is evaporated with Rotary Evaporators, with ethyl acetate (500mL) Dissolving, is neutralized with 1M HCl (90mL).Water outlet phase is separated, NaCl is added in water phase, untill it is no longer dissolved, with acetic acid second Ester (300mL × 3) extracts.Merge organic phase to be cleaned with water (200ml) and saturated salt solution (200ml).Done with anhydrous sodium sulfate Dry, filtering, is concentrated under reduced pressure, is dried in vacuo to obtain product 302g, yield 100%.

2nd, prepare

Enter in the three-neck flask of 500mL 5-70 (1.8462g, 7.9180mmol), 5-64 (10.6125g, 16.6278mmol), HBTU (8.9146g, 23.7539mmol) and HOBT (3.2096g, 23.7539mmol), uses DMF (300mL) dissolves, and solution cool down 20 minutes in -5 DEG C of cryogenic thermostat reactive bath techniques, then dropwise addition DIEA (12.63mL, 71.2617mmol).Move to and be stirred at room temperature after 30 minutes, overnight.Reaction stops, and reaction solution is poured into saturation NaHCO₃Solution In (200mL), (400mL) is extracted with ethyl acetate, then with saturation NaHCO₃Solution (200mL × 3) cleans organic phase, then uses Saturation NaCl solution (300mL × 2) is cleaned.The anhydrous MgSO of final organic phase₄Dry, filtering, being then concentrated under reduced pressure will be molten Liquid is evaporated.Crude product is dissolved with dichloromethane, 30g silica whites is added and makes solid solution, column chromatography, solvent is made with dichloromethane Wet method dress post is homogenized, uses 1%NH₃.H₂O/2%MeOH/CH₂Cl₂To 2%NH₃.H₂O/4%MeOH/CH₂Cl₂Gradient elution.Collect Product, is evaporated to obtain 6.2g, yield 53.2%.

3rd, prepare

5-75 (6g, 4.0715mmol) is added in flask, uses CH₂Cl₂(50ml) dissolves, and slowly drips under stirring at room temperature Add TFA (3.1199mL, 40.7152mmol), be stirred overnight at room temperature.Reaction terminates, evaporated under reduced pressure, by crude product absolute methanol Dissolving, adds NaHCO₃Solid neutralizes.Filtering, silica white solid solution is made by filtrate.It is wet that solvent homogenate is done with dichloromethane Method fills column.Dry method loading.Column chromatography, uses 2%NH₃.H₂O/2%MeOH/CH₂Cl₂To 2.5%NH₃.H₂O/5%MeOH/CH₂Cl₂ Gradient elution.Product is collected, is concentrated under reduced pressure and is evaporated to obtain 3.3g yields 60%.

4th, prepare

5-78 (1.0g, 0.7280mmol) and P21 (478.1mg, 1.4561mmol) are placed in 200mL round-bottomed flasks, Dichloromethane (50mL) is added, is stirred overnight at room temperature.After reaction, concentrate.Solvent homogenate wet method is done with dichloromethane to fill Column.Wet method loading.Column chromatography, uses 1%NH₃·H₂O/2% ethanol/methylenes are to 2%NH₃·H₂O/4% methanol/dichloromethane Alkane gradient elution, collects product, is concentrated under reduced pressure and is evaporated to obtain product 5-85:808.7mg yield 70%.

5th, prepare

5-85 (2.0g, 1.2602mmol) is placed in 250mL round-bottomed flasks, adds dichloromethane (20mL), room temperature is stirred Mix down and be slowly added to TFA (12.6022mmol, 0.9657mL).After reaction, reaction solution is spin-dried for, is dissolved with methanol, added With unnecessary TFA in sodium bicarbonate solid, filtering, addition silica white makes solid solution, dry method loading in filtrate.Column chromatography 10cm × 3.5cm, solvent homogenate dress column is done with dichloromethane.Dichloromethane eluent is first used, then uses 3%NH₃·H₂O/6% methanol/ Dichloromethane eluent is to 5%NH₃·H₂O/10% ethanol/methylene gradient gradient elutions, collect product, are concentrated under reduced pressure and are evaporated Obtain product 5-99:1.78 grams, yield 95.0%.

6th, 5-123 is prepared

The polyethylene glycol 4ARM-SCM-40K (1.0g, 0.02409mmol) of four arms is placed in 50mL straight tube bottles, adds two Chloromethanes (20mL), is stirred at room temperature lower addition 5-99 (286.6mg, 0.01928mmol), since solubility is bad, adds DMF (15mL), is slowly stirred 2 weeks under room temperature shading.After reaction, mixture is spin-dried for, the dissolving of product water is put into bag filter Dialysis.Then the aqueous solution of product is concentrated under reduced pressure, is dissolved after vacuum drying with dichloromethane, add anhydrous ether while stirring It is precipitated out, is filtered, vacuum drying obtains product 5-123:1.132g yield 100%.

Test above-described embodiment¹H-NMR spectrum and mass spectrogram, as shown in Figure 25-55.

The Performance experiment of Pegylation Lapatinib

Experiment 1, cytotoxicity test

Using MTS detection methods, measure compound is to people's Malignant glioma cells strain (U87), human stomach cancer cell line (N87), human lung carcinoma cell line (CALU-3), Breast cancer lines (BT474) and Breast cancer lines (SK-BR-3) increase The inhibitory action grown.

Totally 13 groups of testing compound.First and second group be respectively Lapatinib and Sorafenib DMSO solution, concentration is 0.1M.Three to eight group is respectively the normal saline solution of 6-56,6-162,7-172,4-181,4-208,7-129.9th to 13 groups be respectively 6-56,6-162,7-172,4-181,7-129 95% ethanol solution.

First, for tetra- plants of cells of U87, CALU-3, N87 and BT474, (compound to be determined is molten using sterile saline Solution)

U87 and CALU-3 are cultivated with EMDM+10%FBS+1%P/S, N87 and BT474 1640+10%FBS+1% P/S is cultivated, and is cultivated in 37 DEG C, the incubator of 5%CO2.Passage in 2~3 days is once.Treat cell length to blake bottle bottom about When 80%, trypsin digestion cell, accurate counting, is accessed in 96 orifice plates, blank group is directly added into the amount in 3000/100 μ L/ holes Not celliferous nutrient solution, 24 it is small when after used after cell attachment.The cell culture fluid in orifice plate is sucked, corresponding addition is to be measured Compound gradient dilution application liquid.Cell is placed in 37 DEG C, 5%CO₂Incubator in cultivate, 72 it is small when after suck nutrient solution, Testing compound gradient dilution application liquid is continuously added, when culture 48 is small.After reaching action time, 10 μ L MTS are added per hole, When 37 DEG C of incubations 3 are small, in detection OD 492nM in Sunrise microplate reader.

2nd, for the measure of SK-BR-3 cells (compound to be determined is dissolved using absolute ethyl alcohol)

SK-BR-3 is cultivated using 1640+10%FBS+1%P/S.Other conditions of cell culture, step with it is above-mentioned Four plants of cells are identical.The mode of action of testing compound is also identical with above-mentioned four plants of cells.

Experimental result

Testing compound as shown in Fig. 1-2 3, can draw to draw a conclusion the curve of inhibition of cancer cell rate from figure：

1st, compared with other testing compounds, solubility of the compound 4-208 in physiological saline is preferable.All to be measuredization Dissolubility of the compound in 95% ethanol is more than the dissolubility in physiological saline.

2nd, all testing compounds all show a degree of inhibitory activity in physiological saline, and solubility compared with The activity of good compound 4-208 is most strong.Pharmaceutical activity in all 95% ethanol of testing compound is more than in physiological saline Pharmaceutical activity.The IC of two compounds of 6-56 and 6-162 can be deduced₅₀Value is about near 20 μM.

3rd, the active compound Lapatinib (LPT) in this experiment shows preferable activity, the positive drug of this experiment The IC of Sorafenib₅₀For a μM rank, this is consistent with literature value, and it is reliable to illustrate the system.

Experiment 2, the animal tolerance test of polyethylene glycol conjugation Lapatinib

The tolerance of animal when the first, investigating the administration of 6-56 and 6-162 singles tail vein

On Balb/c female mices, give the tested material 6-56 and 6-162 single tail vein administration of various dose, specifically For：Mouse is randomly divided into 7 groups, every group 3, is administered one day, the administration same day is D1, to observation post administration mouse weight and clinical condition Shape one week, D7 are put to death.

The results are shown in Table 1.After 6-56 is administered 3-4 days, weight loss 2.8%-10.1%, weight base after being administered 7 days This recovery；And weight is in rising trend always after Ctrl groups and tested material 2 are administered；And all groups are not seen during administration Observe animal dead.Illustrate that the tolerance of 6-56 is relatively low, and 6-162 has no toxic side effect substantially.

The changes of weight of animal when 1 6-56 and 6-162 singles tail vein of table is administered

The tolerance of animal when the 2nd, investigating more tail vein administrations of 6-56 and 6-162

On Balb/c female mices, give the more tail vein administrations of tested material 6-56 and 6-162 of various dose, specifically For：Mouse is randomly divided into 3 groups, every group 3, was administered respectively at 1,6,11,16 day, administration gives (interval time 62 times when natural gift Hour), it is administered 8 times altogether.The administration same day is D1, to observation post administration mouse weight and clinical symptoms, in D16 last doses latter week Put to death.

The results are shown in Table 2, and 6-56 (20mg/kg) administration group mouse rise in administration 6 days or so, Normal-weight；To Medicine during 6-16 days changes of weight it is smaller, remain unchanged substantially.And solvent control group and 6-162 (60mg/kg) administration group mouse Weight is in rising trend always after administration.All groups do not observe animal dead under selected dosage.

The changes of weight of animal during more tail vein administrations of table 2 6-56 and 6-162

3rd, 7-129 drug resistances are tested

BALB/c mouse, is dissolved using absolute ethyl alcohol, ultrasonic wave added, after adding physiological saline again after complete drug dissolution Reach medicine solute effect.Medicine is divided into saline control group and 7-129 drug model groups.Medicine dissolution mechanism is 7.75mg Medicine ultrasound suspending is after 15ul absolute ethyl alcohols plus water 45ul dissolves, 37 DEG C of lasting water-baths.Experiment packet：Control group single, mould Repeatedly (morning and each tail vein injection in afternoon once, every time type group single (co-injection 20mg/kg), control group is multiple, and model group Injection volume is 10mg/kg).

Experimental result

1st, drug resistance experiment single group：By the observation of 1 week, compared with physiological saline, hair indifference；Changes of weight No significant difference；Without nausea,vomiting,diarrhea, appetite, indigestion, dry skin, fash, red swelling of the skin, itching, pain, the back of the body Bitterly, expiratory dyspnea, tired and situations such as have a sleepless night.Intracorporeal organ naked eyes viewing indifference.Mice sleep and enliven situation without Difference；Mouse anus and defecation situation indifference, do not there is diarrhea situation.

2nd, drug resistance tests multiple group：By the observation of 1 week, compared with physiological saline, hair has some setbacks, brightless suitable It is sliding.Changes of weight no significant difference；Without nausea,vomiting,diarrhea, appetite, indigestion, dry skin, fash, red swelling of the skin, It is itching, pain, backache, expiratory dyspnea, tired and situations such as have a sleepless night.Intracorporeal organ naked eyes viewing indifference.Mice sleep With enliven situation indifference；Mouse anus and defecation situation indifference, do not there is diarrhea situation.

Experiment 3, the mouse live body curative effect of polyethylene glycol conjugation Lapatinib

Mouse is into knurl：To after several bottles, pancreatin digestion, PBS washings use serum-free 1640 after counting for SK-BR-3 cell culture Culture medium is resuspended, and is that solution density reaches 1*10E7/mL, is placed on trash ice stand-by.NOD mouse inoculations position shaving, uses one Secondary property asepsis injector is inoculated with SK-BR-3 cells, and 0.2mL/ branch, is wiped to kill the thin of spilling after inoculation using cotton ball soaked in alcohol Born of the same parents, put back in cage and normally raise.

Live body is treated：

Active compound LPT：Physiological saline disperses, and 1g is scattered in 200mL physiological saline, dispersed.Oral administration gavage at noon 20mg/kg.7-129：It is divided into the morning and respectively injects a 10mg/kg (conversion dosage) in the afternoon, when interval time 6-7 is small, amounts to 20mg/kg。

Experiment packet：Active compound multiple dosing group；7-129 multiple dosing groups；Physiological saline group.

Experimental result

Tumor growth curve is as shown in figure 24, and compared with active compound, 7-129 shows tumour the inhibition of higher, says After Pegylation, drug effect significantly improves bright Lapatinib.

Although being illustrated and the invention has been described with specific embodiment, but will be appreciated that without departing substantially from the present invention's Many other change and modification can be made in the case of spirit and scope.It is, therefore, intended that in the following claims Including belonging to all such changes and modifications in the scope of the invention.

Claims (20)

1. Pegylation Lapatinib, its general formula are：

A is selected from single armed or the polyethylene glycol of multi-arm or the derivative substituent of polyethylene glycol；X is selected from Y and M is separately selected from double carboxylic acid substituents with amino or its is corresponding Acyl group substituent substituent；N is selected from 49-Phe ,82-Ser,115-Arg,144-Met,145-Asn ,161-Arg,169-Met Human Connective tissue growth factor base or peptide substituent；LPT is Lapatinib substituent；

A=0 or 1；B=0 or 1；C=1 or 2；D=1 or 2；E is equal to the arm number of A.

2. Pegylation Lapatinib according to claim 1, it is characterised in that when a, b are 0, c, d are 1； Work as a=0, during b=1, d=1.

3. Pegylation Lapatinib according to claim 1, it is characterised in that double carboxylic acids with amino are paddy Propylhomoserin or aspartic acid.

4. Pegylation Lapatinib according to claim 1, it is characterised in that A is selected from single armed, four arms or eight arms The derivative substituent of polyethylene glycol or polyethylene glycol.

5. Pegylation Lapatinib according to claim 1, it is characterised in that the molecular weight of A is 12000,20000 Or 40000 dalton.

6. Pegylation Lapatinib according to claim 1, it is characterised in that Y and M are independently Ground is selected from the substituent of following compound：

7. Pegylation Lapatinib according to claim 1, it is characterised in that N is glycine substituent.

8. Pegylation Lapatinib according to claim 1, it is characterised in that molecular formula is

9. the preparation method of the Pegylation Lapatinib described in claim 1, it is characterised in that comprise the following steps：

Step A：Make Lapatinib that amidation process occur with Amino Acid/Peptide, obtain the first intermediate product；

Step B：Make first intermediate product and double carboxylic acids with amino or its corresponding acyl group substituent generation amidatioon anti- Should, obtain the second intermediate product；

Step C：Make second intermediate product and double carboxylic acids with amino or its corresponding acyl group substituent generation amidatioon anti- Should, obtain the 3rd intermediate product；

Step D：The 3rd intermediate product is coupled with the derivative of polyethylene glycol or polyethylene glycol by amido link, produced Product.

10. the preparation method of Pegylation Lapatinib according to claim 9, it is characterised in that the step A In, the condition that amidation process occurs with peptide/amino acid for Lapatinib is：Using HBTU, HOBT, DMF and DIEA as auxiliary agent, -2 Reacted at~4 DEG C, and Lapatinib, peptide/arbitrary amino acid, the molar ratio between HBTU, HOBT and DIEA are：1:0.8~ 1.2:1.3~1.7:1.3~1.7:4.2~4.8.

11. the preparation method of Pegylation Lapatinib according to claim 10, it is characterised in that make Lapatinib Occur to further include before amidation process with peptide or amino acid：The amino of the peptide or amino acid is protected with tertbutyloxycarbonyl.

12. the preparation method of Pegylation Lapatinib according to claim 9, it is characterised in that the step A In, the method that amidation process occurs with peptide for Lapatinib is：The free amino acid that the termination of Lapatinib and the peptide is related to Reaction, other free amino acids that obtained product is related to the peptide successively according still further to the sequence of amino acid in the peptide are anti- Should, and the free amino acid in above-mentioned reaction is before participating in reacting：The free ammonia is activated with n-hydroxysuccinimide The carboxyl of base acid.

13. the preparation method of Pegylation Lapatinib according to claim 9, it is characterised in that the step B In, first intermediate product is with the condition of double carboxylic acids with amino or its corresponding acyl group substituent amidation process：Institute State molar ratio >=2 of first intermediate product with double carboxylic acids with amino or its corresponding acyl group substituent.

14. the preparation method of Pegylation Lapatinib according to claim 13, it is characterised in that step B：Make institute State the first intermediate product and glutamic acid and occur amidation process, first intermediate product and double carboxylic acids with amino or its is corresponding Acyl group substituent occur amidation process before further include：With the carboxyl of n-hydroxysuccinimide activation glutamic acid.

15. the preparation method of Pegylation Lapatinib according to claim 9, it is characterised in that the step C In, second intermediate product is with the condition of double carboxylic acids with amino or its corresponding acyl group substituent amidation process：Institute State molar ratio >=2 of second intermediate product with double carboxylic acids with amino or its corresponding acyl group substituent.

16. the preparation method of Pegylation Lapatinib according to claim 15, it is characterised in that if in step B, First intermediate product and glutamic acid is occurred amidation process, second intermediate product and double carboxylic acids with amino or its Corresponding acyl group substituent occurs to further include before amidation process：With double carboxylic acids of n-hydroxysuccinimide activation zone amino Or the carboxyl of its corresponding acyl group substituent.

17. the preparation method of Pegylation Lapatinib according to claim 9, it is characterised in that the step D In, the 3rd intermediate product is further included before with polyethylene glycol or derivatives thereof generation amidation process：With N, N'- ambers Imide carbonic ester, N, N'- succinimides yl acetate or N, N'- succinimidyl glutarat activated polyethylene glycol Or the derivative of polyethylene glycol.

18. the preparation method of Pegylation Lapatinib according to claim 17, it is characterised in that in the described 3rd Between the molar ratio of product and polyethylene glycol be more than 1.5.

19. the preparation method of Pegylation Lapatinib according to claim 17, it is characterised in that with N, N'- ambers Amber imide carbonic ester, N, N'- succinimides yl acetate or N, N'- succinimidyl glutarat activate poly- second two The derivative of alcohol or polyethylene glycol, obtains the 4th intermediate product, the 3rd intermediate product is with the 4th intermediate product with two Chloromethanes is solvent, and amidation process occurs at 20-30 DEG C.

20. the injection of the Pegylation Lapatinib described in claim 1, it is characterised in that including：The polyethylene glycol It is physiological saline and/or 95% ethanol to change Lapatinib and solvent, the solvent.