CN114853848A - Polypeptide compound and application thereof in preparing medicament for treating hepatic fibrosis diseases - Google Patents

Polypeptide compound and application thereof in preparing medicament for treating hepatic fibrosis diseases Download PDF

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CN114853848A
CN114853848A CN202210570877.9A CN202210570877A CN114853848A CN 114853848 A CN114853848 A CN 114853848A CN 202210570877 A CN202210570877 A CN 202210570877A CN 114853848 A CN114853848 A CN 114853848A
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proline
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hepatic fibrosis
asparagine
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CN114853848B (en
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刘建利
王惠嘉
彭欢
李谋
赵金礼
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Shaanxi HuiKang Bio Tech Co Ltd
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Abstract

The invention discloses a polypeptide compound and application thereof in preparing a medicament for treating hepatic fibrosis diseases, wherein the amino acid sequence of the polypeptide compound is X-proline-asparagine-proline-Y, wherein X, Y respectively represents amino acid at the N end and the C end; x represents any one of phenylalanine, histidine, isoleucine, lysine, leucine, proline, glutamine, serine and tryptophan, and Y represents threonine; or the saidX represents asparagine, and Y represents any one of cysteine, aspartic acid, phenylalanine, leucine, serine and tryptophan. IC of the polypeptide compounds of the present invention 50 The values are all lower than 100 mu M, the proliferation inhibition effect on HSC-T6 is obvious, and the obvious dose dependence is shown, so that the compound is expected to be developed into a novel medicament for treating hepatic fibrosis diseases.

Description

Polypeptide compound and application thereof in preparing medicament for treating hepatic fibrosis diseases
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to a group of polypeptide compounds and application of the compounds in preparation of medicines for treating hepatic fibrosis diseases.
Background
Hepatic fibrosis is a chronic disease formed by long-term interaction of various factors, has high morbidity worldwide, and is an important subject to be solved urgently in the field of global public health at present. A large number of previous researches widely prove the reversibility of early hepatic fibrosis, people have accurate grasp on various aspects of the occurrence reason, the formation mechanism, the related cells, the signal path and the like of the hepatic fibrosis, and fully realize the characteristics of the complexity, the multi-target property and the like of the hepatic fibrosis formation process; in recent years, more and more polypeptide compounds are used for the development of the treatment of hepatic fibrosis diseases; a large number of researches and clinical experiences show that the polypeptide compound has the advantages of small side effect, easy passing through a blood brain barrier, high absorption rate and the like when used for treating hepatic fibrosis diseases, so that the development of the polypeptide medicine for treating the hepatic fibrosis diseases is trendy.
Disclosure of Invention
The invention aims to provide a polypeptide compound with remarkable inhibitory activity on HSC-T6.
The amino acid sequence of the polypeptide compound is X-proline-asparagine-proline-Y (abbreviated as XPNPY), wherein X, Y respectively represents amino acid at the N end and the C end; x represents any one of phenylalanine, histidine, isoleucine, lysine, leucine, proline, glutamine, serine and tryptophan, and Y represents threonine; or X represents asparagine, and Y represents any one of cysteine, aspartic acid, phenylalanine, leucine, serine and tryptophan.
In the amino acid sequence of the polypeptide compound, all the amino acid configurations are L-shaped configurations.
The polypeptide compound exists in the form of acetate, is synthesized by a solid phase method, amino acid protected by Fmoc is used as a raw material, 2-CTC Resin (the degree of substitution (SD 1.0mmol/g) is used as a solid phase carrier, DIC/HOBt is used as a condensing agent, pentapeptide is finally synthesized, finally the pentapeptide is cracked from Resin and purified by a high-efficiency liquid phase, and finally the purity of each monomer compound is over 95 percent.
The polypeptide compound has obvious effect of inhibiting the proliferation of HSC-T6 and can be used for preparing medicaments for treating hepatic fibrosis diseases.
The invention has the following beneficial effects:
research shows that HSC cells (hepatic stellate cells) are main cells forming hepatic fibrosis, and can achieve the effect of resisting the hepatic fibrosis by inhibiting the proliferation of the HSC cells and promoting the apoptosis of the HSC cells, and research finds that rat hepatic stellate cells (HSC-T6) almost have all characteristics of active stellate cells, so that the rat hepatic stellate cells (HSC-T6) take HSC-T6 as a model, screen a large amount of tripeptides by a high-throughput screening method, and find that proline-asparagine-proline (PNP) tripeptide fragments can obviously inhibit the proliferation of HSC-T6, so that the rat hepatic stellate cells take PNP fragments as core fragments, and a specific amino acid combination is respectively added to the C end and the N end of the rat hepatic stellate cells for modification, so that the activity of the rat hepatic stellate cells can be increased, and the lipophilicity of the rat hepatic stellate cells can be changed, and the pharmacokinetics and pharmacodynamics can be influenced. The experimental result shows that the IC of the polypeptide compound of the invention 50 The values are all lower than 100 mu M, the inhibition activity to HSC-T6 is obvious, and the obvious dose dependence is shown, so that the compound is expected to be developed into a novel medicament for treating hepatic fibrosis diseases.
Drawings
FIG. 1 is the MS diagram of FPNPT, 575.44 ═ M + H + ，597.44＝M+Na + 。
FIG. 2 is the MS diagram of HPNPT, 565.43 ═ M + H + ，587.34＝M+Na + 。
FIG. 3 is the MS diagram of IPNPT, 541.43 ═ M + H + ，563.53＝M+Na + 。
FIG. 4 is a MS diagram of KPNPT, 556.53 ═ M + H + ，578.54＝M+Na + 。
FIG. 5 is the MS diagram of LPNPT, 541.43 ═ M + H + ，563.43＝M+Na + 。
FIG. 6 is the MS diagram of PPNPT, 525.43 ═ M + H + ，547.43＝M+Na + 。
FIG. 7 is an MS diagram of SPNPT, 515.33 ═ M + H + ，537.43＝M+Na + 。
FIG. 8 is a MS diagram of WPNPT, 614.34 ═ M + H + ，636.35＝M+Na + 。
FIG. 9 is the MS diagram of NPNPNPC, 543.43 ═ M + H + 。
FIG. 10 is the MS diagram of NPNPNPD, 556.33 ═ M + H + ，578＝M+Na + 。
FIG. 11 is the MS diagram of NPNPNPF, 588.34-M + H, 610.44-M + Na + 。
FIG. 12 is the MS diagram of NPNPNPL, 554.43 ═ M + H + ，576.34＝M+Na + 。
FIG. 13 is the MS diagram of NPNPNPS, 528.23 ═ M + H + ，550.33＝M+Na + 。
FIG. 14 is the MS diagram of NPNPNPW, 627.34 ═ M + H + 。
Detailed Description
The invention will be further described in detail with reference to the following figures and examples, but the scope of the invention is not limited to these examples.
Example 1
Synthesis of FPNPT
Figure BDA0003659105190000031
Selecting 2-CTC Resin with the degree of substitution of 1.0mmol/g as a solid phase carrier, weighing 10g of 2-CTC Resin in a reaction column, adding 80mL of DCM swelling Resin for 10min, and filtering. Dissolving 2-CTC Resin with Fmoc-Thr (tBu) -OH of 1.1 time molar equivalent and DIEA of 4 times molar equivalent by 80mL of DCM to prepare a reaction solution, adding the reaction solution into a reaction column, and introducing N 2 Stirring, reacting for 1.5h, adding 10mAnd (3) sealing the L anhydrous methanol for 30min, filtering to remove the reaction liquid, washing with IPA and DMF for 3min twice, and washing the amino acid reaction liquid attached to the resin peptide to obtain the Fmoc-Thr (tBu) -2-CTC resin peptide. Then, a 20% volume fraction PIP/DMF solution was added and N was added 2 Stirring, reacting for 30min, removing the alpha-amino Fmoc protecting group, removing the reaction solution by suction filtration, and washing with IPA and DMF for 3min each time twice. Dipping the resin peptide into a test tube by using a glass rod, adding 2mL of 0.05g/mL ninhydrin ethanol solution (Kaiser color development method), keeping the temperature in a water bath kettle at 95 ℃ for 2min, and detecting by visual observation to show that the resin is positive (black).
After the deprotection reaction is complete, the next amino acid ligation is initiated. Completely dissolving 2-CTC Resin 2 times molar equivalent of Fmoc-Pro-OH, 2.5 times molar equivalent of DIC and 2.5 times molar equivalent of HOBt in 80mL of DMF to prepare reaction liquid, adding the reaction liquid into a reaction column, and introducing N 2 Stirring and reacting for 1.5 h. Similarly, Kaiser color method is used to detect whether the reaction is complete, after the reaction is complete, the reaction solution is removed by suction filtration, and IPA and DMF are washed twice, each time for 3 min. Subsequently, the operation steps of deprotection-condensation-deprotection are repeated, and Fmoc-Asn (trt) -OH, Fmoc-Pro-OH and Fmoc-Phe-OH are connected in sequence according to the amino acid sequence of phenylalanine-proline-asparagine-proline-threonine (FPNPT). After the peptide chain synthesis is finished and the Fmoc protecting group is removed, washing with IPA and DMF twice each for 3min, washing with anhydrous methanol for 3 times each for 3min, and carrying out suction filtration and drying at normal temperature to obtain the resin peptide of FPNPT.
Placing the resin peptide of FPNPT obtained by solid phase synthesis in a 250mL beaker, adding 100mL of cutting fluid under stirring, reacting for 2h, cutting the resin and each side chain protecting group, wherein the volume ratio of the cutting fluid is TFA: TIS: PhSCH 3 :H 2 O88: 4:5: 3. And (3) carrying out suction filtration, washing the resin with DCM for 3 times, carrying out rotary evaporation on the filtrate at 40 ℃ for concentration, separating out a concentrated solution by using methyl tert-butyl ether, fully stirring, centrifuging at 3500r/min for 10min, and volatilizing the methyl tert-butyl ether at normal temperature to obtain crude peptide solid powder, namely the FPNPT crude product.
And (3-5 g of FPNPT crude product is taken in a 100mL beaker, dissolved by ultrasonic with 50mL of pure water, filtered by a 0.45-micron filter membrane and subjected to chromatographic separation and purification. Chromatographic conditions are as follows: LC3000 semi-preparative liquid chromatograph, reversed-phase column chromatography filler chromatographic column (50mm is multiplied by 350mm, 30-50 μm); mobile phase: phase a was 0.1% trifluoroacetic acid in water and phase B was 0.1% trifluoroacetic acid in methanol, flow rate: 30mL/min, sampling 100% of phase A, removing impurities by gradient elution of 10% -40%, and performing equal gradient elution after the main product peaks. Sample introduction amount: 50mL, the detection wavelength is 215nm, an LC3000 workstation monitors the elution process, a sample collection bottle collects the main product peak, the effluent liquid with the purity higher than 95% is merged after analysis, and after rotary evaporation concentration, a freeze dryer freezes for 72 hours, thus obtaining the FPNPT pure freeze-dried powder.
Sampling the purified FPNPT freeze-dried powder with ultrapure water, filtering with a 0.22-micrometer filter membrane with the concentration not more than 50ppm, and performing mass spectrometry by using a Waters Acquity ultra-efficient liquid-mass chromatograph. Chromatographic conditions are as follows: an Acquity UPLC C18 chromatography column (2.1mm × 50mm, 1.7 μm); mobile phase: phase A is 0.1% formic acid water, phase B is 0.1% formic acid acetonitrile, according to the procedure of the mass spectrum detection method, the operation procedure is 1% -31%, the flow rate: 0.3mL/min, time 15min, sample size: 2 μ L. Mass spectrum detection: scan 100-.
The Fmoc-Phe-OH in example 1 was replaced with equimolar amounts of Fmoc-His (trt) -OH, Fmoc-Ile-OH, Fmoc-Lys (boc) -OH, Fmoc-Leu-OH, Fmoc-Pro-OH, Fmoc-Gln (trt) -OH, Fmoc-Ser (otbu) -OH and Fmoc-Trp-OH, respectively, and histidine-proline-asparagine-proline-threonine (HPNPT), isoleucine-proline-asparagine-proline-threonine (IPNPT), lysine-proline-asparagine-proline-threonine (KPNPT), leucine-proline-asparagine-proline-threonine (LPNPT) and the like, in the same manner as in example 1, respectively, The pure products of proline-asparagine-proline-threonine (PPNPT for short), glutamine-proline-asparagine-proline-threonine (QPNPT for short), serine-proline-asparagine-proline-threonine (SPNPT for short) and tryptophan-proline-asparagine-proline-threonine (WPNPT for short) are shown in the figure 2-8.
Example 2
Synthesis of NPNPC
Figure BDA0003659105190000051
Figure BDA0003659105190000061
Selecting 2-CTC Resin with the degree of substitution of 1.0mmol/g as a solid phase carrier, weighing 10g of 2-CTC Resin in a reaction column, adding 80mL of DCM swelling Resin for 10min, and filtering. Dissolving 2-CTC Resin with Fmoc-Cys (trt) -OH of 1.1 times of molar equivalent and DIEA of 4 times of molar equivalent by 80mL of DCM to prepare a reaction solution, adding the reaction solution into a reaction column, and introducing N 2 Blowing and stirring, reacting for 1.5h, adding 10mL of anhydrous methanol, sealing for 30min, filtering to remove reaction liquid, sequentially washing with IPA and DMF twice for 3min each time, and washing amino acid reaction liquid attached to the resin peptide to obtain the Fmoc-Cys (trt) -2-CTC resin peptide. Then, a 20% volume fraction PIP/DMF solution was added and N was added 2 Stirring, reacting for 30min, removing the alpha-amino Fmoc protecting group, removing the reaction solution by suction filtration, and washing with IPA and DMF for 3min each time twice. Dipping the resin peptide into a test tube by using a glass rod, adding 2mL of 0.05g/mL ninhydrin ethanol solution (Kaiser color development method), keeping the temperature in a water bath kettle at 95 ℃ for 2min, and detecting by visual observation to show that the resin is positive (black).
After the deprotection reaction is complete, the next amino acid ligation is initiated. Completely dissolving 2-CTC Resin 2 times molar equivalent of Fmoc-Pro-OH, 2.5 times molar equivalent of DIC and 2.5 times molar equivalent of HOBt in 80mL of DMF to prepare reaction liquid, adding the reaction liquid into a reaction column, and introducing N 2 Stirring and reacting for 1.5 h. Similarly, Kaiser color method is used to detect whether the reaction is complete, after the reaction is complete, the reaction solution is removed by suction filtration, and IPA and DMF are washed twice, each time for 3 min. Subsequently, the operation of deprotection-condensation-deprotection is repeatedSequentially connecting Fmoc-Asn (trt) -OH, Fmoc-Pro-OH and Fmoc-Asn (trt) -OH according to the amino acid sequence of asparagine-proline-cysteine (NPNPC for short). After the peptide chain synthesis is completed and the Fmoc protecting group is removed, washing with IPA and DMF twice each for 3min, washing with anhydrous methanol for 3 times each for 3min, and carrying out suction filtration and drying at normal temperature to obtain the resin peptide of NPNPC.
Placing the resin peptide of NPNPC obtained by the solid phase synthesis in a 250mL beaker, adding 100mL of cutting fluid under stirring, reacting for 2h, cutting the resin and each side chain protecting group, wherein the volume ratio of the cutting fluid is TFA: TIS: PhSCH 3 :H 2 O88: 4:5: 3. And (3) carrying out suction filtration, washing the resin for 3 times by using DCM, carrying out rotary evaporation on the filtrate at 40 ℃ for concentrating, separating out concentrated solution by using methyl tert-butyl ether, centrifuging for 10min at 3500r/min after fully stirring, and volatilizing the methyl tert-butyl ether at normal temperature to obtain crude peptide solid powder, namely the NPNPC crude product.
And (3-5 g of the NPNPNPC crude product is taken in a 100mL beaker, dissolved by ultrasonic with 50mL of pure water, filtered by a 0.45-micron filter membrane and then subjected to chromatographic separation and purification. Chromatographic conditions are as follows: LC3000 semi-preparative liquid chromatograph, reversed-phase column chromatography filler chromatographic column (50mm is multiplied by 350mm, 30-50 μm); mobile phase: phase a was 0.1% trifluoroacetic acid in water and phase B was 0.1% trifluoroacetic acid in methanol, flow rate: 30mL/min, sampling 100% of phase A, removing impurities by gradient elution of 10% -40%, and performing equal gradient elution after the main product peaks. Sample introduction amount: 50mL, the detection wavelength is 215nm, an LC3000 workstation monitors the elution process, a sample collection bottle collects the main product peak, the effluent liquid with the purity higher than 95% is merged after analysis, and the mixture is frozen for 72 hours by a freeze dryer after rotary evaporation concentration, so that the NPNPC pure product freeze-dried powder is obtained.
Preparing a sample of the purified NPNPC freeze-dried powder by ultrapure water, filtering with a 0.22 mu m filter membrane, and performing mass spectrometry identification by adopting a Waters Acquity ultra-efficient liquid-mass chromatograph. Chromatographic conditions are as follows: an Acquity UPLC C18 chromatography column (2.1mm × 50mm, 1.7 μm); mobile phase: phase A is 0.1% formic acid water, phase B is 0.1% formic acid acetonitrile, according to the procedure of the mass spectrum detection method, the operation procedure is 1% -31%, the flow rate: 0.3mL/min, time 15min, sample size: 2 μ L. Mass spectrum detection: scan 100-.
Pure products of asparagine-proline-aspartic acid (NPNPL), asparagine-proline-serine (NPNPNPS) and asparagine-proline-tryptophan (NPW) can be obtained by replacing Fmoc-Cys (trt) -OH in example 2 with equimolar amounts of Fmoc-Asp (otbu) -OH, Fmoc-Phe-OH, Fmoc-Leu-OH, Fmoc-Ser (otbu) -OH and Fmoc-Trp-OH respectively, and by the same procedures as in example 2, the characterization results are shown in FIGS. 10-14.
Example 3
The polypeptide compounds of the invention have inhibition effect on the proliferation of HSC-T6
1. Cell culture
(1) Cell resuscitation
Sterilizing an ultra-clean workbench and experimental articles by ultraviolet rays for 0.5h, re-heating reagents such as a culture medium and the like at room temperature, wiping the surface of the workbench by 75% of alcohol for sterilization, turning on sterile air after turning off an ultraviolet lamp, igniting the alcohol lamp, and operating in a sterile environment around the outer flame of the alcohol lamp. And (3) quickly taking out the cryopreserved HSC-T6 cells from the liquid nitrogen tank, checking the cell types, cell numbers and cryopreserved dates in the records, thawing the cells in a water bath kettle at 37 ℃, continuously and lightly shaking the cell cryopreserving tube to accelerate thawing, and simultaneously taking care to prevent the seal of the cryopreserving tube from contacting liquid in the water bath kettle to cause cell pollution. In a clean bench sterile environment, transferring the unfrozen cells to a 15mL centrifuge tube with 9mL 10% FBS High-DMEM culture medium by using a pipettor, pumping until the cells are uniformly mixed, centrifuging at the normal temperature of 800rpm for 5min, discarding supernatant, adding 1mL culture medium into the centrifuge tube to resuspend the cells, pumping until the cells are uniformly mixed, and transferring to 25cm of 1mL 10% FBS High-DMEM culture medium 2 In a culture flask, at 5% CO 2 Culturing in an incubator with 95% saturated humidity, and changing the culture solution once within 24 h.
(2) Cell passage
The state of the cells was observed by an inverted light microscope, and when the bottom of the flask was filled with 80% or more of the cells, the passaging operation was carried out. The medium in the flask was removed under sterile conditions, the cells were washed 3 times with PBS wash, 3mL each, then 3mL of 0.25% trypsin was added and allowed to spread evenly over the flask, which was then placed in an incubator for digestion for about 2 min. When cells are observed to be round and large pieces are shed under a microscope, the digestion is proved to be complete, 4mL of 10% FBS High-DMEM medium and trypsin are added to stop the reaction, and the reaction solution is slowly blown until adherent cells are completely stripped. The cell suspension was then transferred to a 15mL sterile centrifuge tube, centrifuged at 800rpm for 5min at room temperature, the supernatant was discarded, 2mL of 10% FBS High-DMEM medium was added to resuspend the cells, and the tube was repeatedly blown up to form a single cell suspension for further passaging or inoculation.
(3) Cell cryopreservation
Freezing and storing the cells in the same step as the step of subculturing the cells, digesting, centrifuging and removing supernatant, adding 1mL of cell freezing solution of DMSO: FBS: DMEM-1: 2:7(v/v/v) into a centrifuge tube, repeatedly blowing and beating until the cells are uniformly mixed, then transferring the cells into a cell freezing tube, sealing and storing the cells by a sealing film, marking information such as cell types, freezing date and the like, standing the cells in a refrigerator at the temperature of-20 ℃ for 2 hours, standing the cells in the refrigerator at the temperature of-80 ℃ overnight, and storing the cells in a liquid nitrogen tank.
2. Cell processing and MTT assay
Adherent culture: subculturing HSC-T6 cells, adding 3mL of 0.25% trypsin into carbon dioxide incubator for 2min when the cells are more than 80% of the bottom of the culture flask, adding fresh High-DMEM medium to prevent over digestion, centrifuging to collect cells, blowing the cells into single cell suspension in the fresh medium, inoculating the cells into 96-well culture plate, adding 180 μ L of 10% FBS High-DMEM medium and 20 μ L of cell suspension into each well, culturing at 37 deg.C and 5% CO 2 And culturing for 24h in an incubator with 95% saturated humidity, and observing whether the cells are completely attached to the wall under a microscope. After the cells are attached to the wall, the supernatant is removed, and serum-free High-DMEM medium is added for continuous culture for 12 h.
Grouping and molding: the experiment set up control group, 5ng/mL TGF-beta 1 group, add 5ng/mL TGF-beta 1 different concentrations XPNPY (diluted to 6 concentrations in 10% FBS-containing High-DMEM medium: 100. mu. mol/L, 50. mu. mol/L, 25. mu. mol/L, 12.5. mu. mol/L, 6.25. mu. mol/L, 3.125. mu. mol/L). After 12h of serum-free medium treatment, old medium was removed and each group was incubated with 100. mu.L of 5ng/mL TGF-. beta.1 for 2h, followed by 100. mu.L of 10% FBS High-DMEM fresh medium per well and 100. mu.L of medium-diluted working medium, which were repeated 3 times per well to exclude the effect of irrelevant variables on the experiment.
And (3) MTT method color development: adding drug into 96-well plate, culturing in constant temperature incubator for 48 hr, removing supernatant, adding 100 μ L of 5mg/mL MTT solution into each well, 37 deg.C, and 5% CO 2 And continuously culturing in a 95% saturated humidity incubator for 4h, removing old culture solution, adding 150 μ L of analytical pure DMSO into each well, shaking in a constant temperature horizontal shaker at 37 deg.C for 10min, and measuring absorbance at 490nm with an enzyme linked immunosorbent detector.
3. Data analysis
Data analysis was performed using software SPSS 20.0, absorbance values at different concentrations for each compound
Figure DA00036591051951229519
Representing, applying one-way analysis of variance, P<0.05 showed that the difference was significant, and the cell growth inhibition rate was calculated according to the following formula:
inhibition rate (1-drug OD/control OD) × 100%
Inhibition rate>0 the compound has effect in inhibiting cell proliferation<0 the compound has effect in promoting cell proliferation, and calculating IC with SPSS 20.0 according to inhibition rate and sample concentration 50 The value is obtained. The results are shown in Table 1.
TABLE 1 inhibitory Effect of the polypeptide Compound of the present invention on the proliferation of HSC-T6: (
Figure BDA0003659105190000091
n＝3)
Figure BDA0003659105190000092
Figure BDA0003659105190000101
Figure BDA0003659105190000111
Note: the concentration of TGF-beta 1 is 5ng/mL, a P<0.05 compared to the TGF-. beta.1 group, A P<0.01 compared to the TGF-. beta.1 group, b P<0.05 compared with the control group, B P<0.05 compared to the control group.
TABLE 2 IC of the inhibitory Effect of the polypeptide-based Compound of the present invention on the proliferation of HSC-T6 cells 50 Value of
Figure BDA0003659105190000112
As can be seen from Table 2, IC's of FPNPT, HPNPT, IPNPT, KPNPT, LPNPT, PPNPT, QPNPT, SPNPT, WPNPT, NPNPC, NPNPNPD, NPNPF, NPNPNPL, NPNPNPS, NPNPNPW 50 The values are all lower than 100 mu M, the inhibition activity on HSC-T6 is obvious, and obvious dose dependence is shown. Wherein FPNPT (IC) 50 Value 37.328 μ M), NPNPD (IC) 50 32.328 mu M), the two pentapeptides have the best activity, can effectively inhibit the proliferation of HSC-T6, and is worthy of further evaluation on the pharmacological activity of animals.

Claims (4)

1. A polypeptide compound for treating hepatic fibrosis diseases, which is characterized in that: the amino acid sequence of the polypeptide compound is X-proline-asparagine-proline-Y, wherein X, Y respectively represents amino acids at the N end and the C end;
x represents any one of phenylalanine, histidine, isoleucine, lysine, leucine, proline, glutamine, serine and tryptophan, and Y represents threonine;
or X represents asparagine, and Y represents any one of cysteine, aspartic acid, phenylalanine, leucine, serine and tryptophan.
2. The polypeptide compound of claim 1, wherein: in the amino acid sequence of the polypeptide compound, all the amino acid configurations are L-shaped configurations.
3. The polypeptide compound of claim 1, wherein: the polypeptide compound exists in the form of acetate.
4. Use of the polypeptide compound of claim 1 for the preparation of a medicament for the treatment of liver fibrosis.
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WO2001088538A2 (en) * 2000-05-12 2001-11-22 Dumas David P Compositions and methods for epitope mapping

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Publication number Priority date Publication date Assignee Title
WO2001088538A2 (en) * 2000-05-12 2001-11-22 Dumas David P Compositions and methods for epitope mapping

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XIANGRONG YANG等: "Quantitative structure–chromatographic retention relationship of synthesized peptides (HGRFG, NPNPT) and their derivatives", ANALYTICAL BIOCHEMISTRY, vol. 597, pages 1 - 12 *

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