CN106138061A - Prevention or weaken the complex of pulmonary fibrosis and preparation thereof and purposes - Google Patents

Abstract

The present invention relates to complex of a kind of biguanides and EGFR-TKI or ALK-TKI and application thereof, this complex can be used for preventing or slowing down interstitial pneumonia, the interstitial pneumonia that particularly EGFR-TKI or ALK-TKI produces when treating nonsmall-cell lung cancer, it can avoid the complication that EGFR-TKI or ALK-TKI produces when treating nonsmall-cell lung cancer, it is effectively improved the therapeutic effect of EGFR-TKI or ALK-TKI, reduces generation and the development of complication for patients.

Description

Prevention or weaken the complex of pulmonary fibrosis and preparation thereof and purposes

Technical field

The invention belongs to field of medical application, particularly to preventing or weakening the compound of pulmonary fibrosis Thing and preparation thereof and purposes.

Background technology

Pulmonary carcinoma is the malignant tumor that M & M is the highest in global range；Non-small cell lung Cancer (Non Small Cell Lung Cancer, NSCLC) accounts for the 80%～85% of all cases of lung cancer. EGF-R ELISA (Epidermal growth factor receptor, EGFR) and Anaplastic lymphoma kinase ALK Alk receptor tyrosine kinase (anaplasticlymphoma kinase, ALK) is NSCLC's Occur evolution plays an important role.EGFR tyrosine kinase inhibitor (Tyrosine Kinase inhibitor, TKI) and targeted drug that ALK-TKI is representative to specific NSCLC Patient achieves significant curative effect, but occurs serious complication in TKI therapeutic process-- Matter pneumonia；Although its sickness rate is low, but once occur often patient to be produced detrimental effects.

Gefitinib (gefitnib, Iressa) is the EGFR-TKI of representative, is at present by many State ratifies and is widely used in progress or the little molecular targeted agents of intractable NSCLC, has height Selectivity and hypotoxic advantage, in nearly late result and the quality of life of improving advanced NSCLC In effect demonstrate encouraging prospects.III recent phase clinical study results shows, right The NSCLC patient of EGFR sudden change, a line uses targeted drug Erlotinib, routinizes with a line Treatment is compared, and can improve the Progression free survival time of nearly 3 times.

Interstitial pneumonia is gefitinib and the rare but fatal untoward reaction of Erlotinib.Japan Scholar Okamoto et al reports the first and causes serious interstitial pneumonia because taking gefitinib And the case of death.Dyspnea i.e. occurs, although giving heavy dose of class after patient medication 8d Steroid hormone is treated, but still dead after being administered 13d.The research of U.S. FDA finds, in U.S. State uses the incidence rate of patient's interstitial lung disease of gefitinib to be 1%, the most about 1/3 patient Die from interstitial pneumonia.Prevention interstitial pneumonia caused by EGFR-TKIs is to improve such patient The key of prognosis.17-hydroxy-11-dehydrocorticosterone is one for the treatment of most important measure of this kind of interstitial pneumonia, but Curative effect is unsatisfactory and side effect is bigger.Therefore, if a kind of new, effective measures can be found The generation of prevention interstitial pneumonia, then for strengthening the safety of TKI, improving patient's prognosis all To there is important scientific meaning and clinical value.

Showing after deliberation, pulmonary fibrosis is the pathological change that interstitial pneumonia is most basic, its morbidity Mechanism mainly includes fibroblastic gathering and extracellular matrix excess deposition etc..The most clinical Upper majority is the generation by preventing or weaken pulmonary fibrosis, prevents the generation of interstitial pneumonia.

Metformin is a kind of classical medicine treating diabetes, economic security, better tolerance, Owing to its blood sugar lowering mechanism is the picked-up of suppression gastrointestinal tract glucose, increase surrounding tissue to islets of langerhans The sensitivity of element, suppresses the gluconeogenesis that liver, kidney are excessive, therefore to normal person without hypoglycemic activity； Along with going deep into of research, prior art discloses many metformin and become answering of subassembly with other Compound；Such as, CN103271907 discloses a kind for the treatment of being made up of berberine and metformin The compositions of diabetes；CN103860532 discloses and a kind of is made up of memantine and metformin For treating the compositions of dementia of the Alzheimer type disease；Additionally, now technology also disclose by Metformin and meglitinides or the compound formed with DPP-4 inhibitor etc..But It is that prior art yet there are no about the biguanides based on metformin and gefitinib etc. EGFR-TKI or ALK-TKI combination is used for preventing or weakening pulmonary fibrosis thus for treating interstitial The report of property pneumonia.

Summary of the invention

The invention provides a kind of for preventing or weakening by EGFR-TKI or ALK-TKI treatment non- The complex of the pulmonary fibrosis produced during small cell lung cancer and preparation thereof.

The present invention provides a kind of complex preventing or weaken pulmonary fibrosis, wherein, this complex Including the biguanides that parts by weight are 2-4 part and the TKI of 2-5 part；Described TKI is selected from EGFR-TKI or ALK-TKI.

Further improving, biguanides is selected from metformin and salt, phenformin or chlorine Guanidine；Described EGFR-TKI selected from gefitinib, Erlotinib, Conmana, Afatinib, AZD9291, CO-1686 or HM61713；For Buddhist nun or color is auspicious replaces selected from gram azoles for described ALK-TKI Buddhist nun.

When biguanides is metformin, and EGFR-TKI is gefitinib, and, diformazan is double The parts by weight of guanidine are 3 parts, when the parts by weight of gefitinib are 2 parts, to pulmonary fibrosis Weaken effect the most obvious.

Another aspect of the present invention provides the oral formulations being made up of complex and adjuvant, is preferably Tablet.

This tablet is the bilayer tablet being made up of the release layer of label and parcel label, described label It is made up with excipient of metformin；Described release layer is made up of gefitinib and adjuvant.

Preferably, adjuvant is Powderd cellulose, beta-schardinger dextrin-, arabic gum, polyvinylpolypyrrolidone And micropowder silica gel；Excipient be lactose, polyethylene maleic anhydride, ethylene glycol monostearate, Polyvinylpolypyrrolidone and myristyl alcohol；The weight portion of described bilayer tablet is as follows:

Label:

Release layer:

After the present invention is by making bilayer tablet by complex, after making the entrance of this tablet internal, Gefitinib first plays a role, and afterwards, metformin slowly plays a role, and both play and i.e. control Treat the effect of nonsmall-cell lung cancer, the generation of pulmonary fibrosis can also be prevented or weaken simultaneously.Will After gefitinib mixes with Powderd cellulose and beta-schardinger dextrin-, it is possible to decrease gefitinib medicine is at water Precipitation in solution, improves the bioavailability of gefitinib；Simultaneously by metformin and slow release After material polyethylene maleic anhydride and ethylene glycol monostearate make slow release label, label is made to exist 1, the cumulative defaultlogic of 2,6,12 and 24 hours respectively reach 30-32%, 65-67%, 80-82%, 85-88% and more than 90%, release is steady, the effect of slow release to make this double-layer tablet have Really, and 24h release can be reached obtain purpose.

Another aspect of the present invention provides the purposes of this complex, and it is in preparation prevention or weakens lung Application in Fibrotic medicine.The most described pulmonary fibrosis be by EGFR-TKI or ALK-TKI produces when treating nonsmall-cell lung cancer.

The complex that the present invention provides can be additionally used in preparation prevention or slows down the medicine of interstitial pneumonia Thing, described interstitial pneumonia is produced when being and treated nonsmall-cell lung cancer by EGFR-TKI or ALK-TKI Raw complication.

Another aspect of the present invention additionally provides the application of tablet, this tablet can be used for preparation prevention or Weaken the medicine of pulmonary fibrosis.Preferably, pulmonary fibrosis is to be controlled by EGFR-TKI or ALK-TKI Produce when treating nonsmall-cell lung cancer.

The beneficial effects of the present invention is, the present invention provides answering of a kind of biguanides and TKI Compound and application thereof, this complex can be used for preventing or weakening pulmonary fibrosis, particularly EGFR-TKI Or the pulmonary fibrosis that ALK-TKI produces when treating nonsmall-cell lung cancer, thus play prevention or subtract The generation of weak interstitial pneumonia, it can avoid EGFR-TKI or ALK-TKI to treat non-small cell lung The complication produced during cancer, is effectively improved the therapeutic effect of EGFR-TKI or ALK-TKI, subtracts The generation of few complication for patients and development.

Accompanying drawing explanation

Fig. 1 is the result that immunofluorescence technique detection fibers index α-actin expresses；

Fig. 2 is the table of western blot technology for detection Fibrosis Markers α-actin and COL1A1 Reach, the result of the expression of detection TGF-β downstream key signal pathway protein；

Fig. 3 is HE dyeing and the result of Masson dyeing；

Fig. 4 is the result of Elisa method detection hydroxyproline content；

Fig. 5 is the result that SABC detection α-actin expresses；

Fig. 6 is western blot detection COL1A1, α-actin and TGF-β downstream key signal The result of the expression of pathway protein.

Specific embodiment mode

1 one kinds of complex prevented or weaken pulmonary fibrosis of embodiment

This complex includes metformin and the gefitinib of 2 weight portions of 3 weight portions.

2 one kinds of bilayer tablets of embodiment

Label:

Release layer:

3 one kinds of bilayer tablets of embodiment

Label:

Release layer:

Zoopery

Experimental example 1 metformin is obviously reduced gefitinib stimulates the research of the pulmonary fibrosis caused

1.1 experimental technique

Lung fibroblast strain HFL-1 is divided into 6 groups, respectively matched group, P9-CM group, P9-CM+ gefitinib group, P9-CM+ gefitinib+metformin group, R9-CM group, R9-CM+ Metformin group；

Matched group is without special handling；P9-CM group gives at the conditioned medium of PC-9 cell Reason cell 48h；PC-9+ gefitinib group gives the PC-9 processed from gefitinib 30 μMs The conditioned medium of cell processes cell 48h；PC-9+ gefitinib+metformin group give from At the conditioned medium associating metformin 20 μMs of the PC-9 cell that gefitinib 30 μMs processes Reason cell 48h；R9-CM group gives to process cell 48h from the conditioned medium of PC-9GR cell； R9-CM+ metformin group gives to combine metformin from the conditioned medium of PC-9GR cell 20 μMs process cell 48h；

After 48h, use the expression of immunofluorescence technique detection fibers index α-actin；Adopt With western blot technology for detection Fibrosis Markers α-actin, COL1A1 and TGF-β The expression of downstream key signal pathway protein.

2.2 experimental result

From figure 1 it appears that compared with matched group, through P9-CM group, R9-CM group and After the HFL-1 cell 48h that P9-CM+ gefitinib group processes, the expression of α-actin albumen is bright Aobvious increase；Compare with P9-CM group, through the HFL-1 cell that P9-CM+ gefitinib group processes After 48h, the expression of α-actin albumen the most substantially increases, it follows that, gefitinib is not only The content of pulmonary fibrosis mark α-actin albumen can be significantly improved, and can be further Improve the content of the HFL-1 intracellular α-actin albumen after P9-CM group processes；With P9-CM+ gefitinib group compares, after P9-CM+ gefitinib+metformin group processes HFL-1 cell, the expression of α-actin albumen significantly reduces；Compare with R9-CM group, pass through HFL-1 cell after the process of R9-CM+ metformin group, the expression of α-actin albumen is significantly dropped Low；It follows that the condition that metformin not only can be obviously reduced from PC-9 cell strain is trained The pulmonary fibrosis mark that foster base and the conditioned medium from PC-9GR cell strain cause The expression of α-actin albumen；And gefitinib can be obviously reduced and sting the pulmonary fibrosis caused The expression of mark α-actin albumen.

From figure 2 it can be seen that compared with matched group, non-through R9-CM group and P9-CM+ Ji For Buddhist nun's group process HFL-1 cell 48h after, COL1A1, α-actin, pSmad2, pSmad3, The expression of pSTAT3, pAKT and dpERK1/2 albumen substantially increases；Compare with P9-CM group, warp After crossing the HFL-1 cell 48h that P9-CM+ gefitinib group processes, COL1A1, α-actin, The expression of pSmad2, pSmad3, pSTAT3, pAKT and dpERK1/2 albumen substantially increases；With P9-CM+ gefitinib group compares, after P9-CM+ gefitinib+metformin group processes In HFL-1 cell, COL1A1, α-actin, pSmad2, pSmad3, pSTAT3, pAKT Expression with dpERK1/2 albumen substantially reduces；Compare with R9-CM group, through R9-CM+ diformazan Biguanide group process after HFL-1 cell in, COL1A1, α-actin, pSmad2, pSmad3, The expression of pSTAT3, pAKT and dpERK1/2 albumen substantially reduces；It follows that metformin Can be obviously reduced gefitinib and lung that the conditioned medium from PC-9GR cell strain causes is fine Dimensionization mark α-actin and the expression of COL1A1 albumen, significantly inhibit TGF-β signal and lead to The expression of key downstream, road albumen, significantly inhibits the key mechanism of pulmonary fibrosis.

1.3 conclusion

From showing that metformin can be obviously reduced the lung fiber that gefitinib causes above Change.

Experimental example 2 metformin is obviously reduced the pulmonary fibrosis that the bleomycin that gefitinib increases the weight of causes Research

2.1 experiment material

6 week old health male SD rats 75 are (dynamic purchased from the experiment of great Ping hospital of Third Military Medical University Thing center), body weight 200g-210g；

Gefitinib, purchased from Tocris Bioscience company of Britain；

Metformin, purchased from sigma company of the U.S.；

HE and Masson staining kit, is stepped Newbiotics Inc by Foochow and produces；

Enzyme-linked Immunosorbent Assay (enzyme linked immunosorbent assay, ELISA) tries Agent box, is produced by Wuhan You Ersheng company.

The preparation of 2.2 animal models and packet

After 75 SD rat adaptabilities are fed one week, it is divided into by the method for table of random number Matched group, bleomycin group, metformin group, gefitinib group and gefitinib metformin Combination group, often organizes each 15；

By 75 SD rats respectively with after 1% pentobarbital sodium intraperitoneal injection of anesthesia, lie on the back fixing On Mus plate, ethanol disinfection after cervical region unhairing, longitudinal incision throat skin, sudden and violent after chorista Leak pipe；

Wherein, matched group injects physiological saline solution 200 μ l, and remaining each group is injected to tracheal strips Bleomycin (5mg/kg, 5g/L), rapidly that animal is upright after injection, rotate, make medicinal liquid It is evenly distributed in lung；

Each experimental group rat started also to need continuous gavage 21 days in intratracheal injection the same day, wherein, Matched group and bleomycin group give normal saline 2ml gavage, and gefitinib group gives Ji Fei and replaces Buddhist nun's 200mg/kg gavage；Metformin group gives metformin 300mg/kg gavage；Ji Fei replaces Buddhist nun's co-administrated metformin group gives gefitinib 200mg/kg associating metformin 300mg/kg and fills Stomach；After intratracheal injection the 21st day, kill Mus and take lung；

2.3 observation index

The dyeing of left pneumolith wax slice row HE and Masson dyeing；SABC detection α-actin Expression；Take right lung Elisa method detection hydroxyproline content；Western blot detects COL1A1, α-actin and the expression of TGF-β downstream key signal pathway protein.

2.4 experimental result

From figure 3, it can be seen that after HE staining, compared with matched group, through rich next mould After element group and gefitinib group process the lung of mice, lung septal thickening, pulmonary interstitial edema and fiber Change capillary injection amount all to dramatically increase；Compare with bleomycin group, through gefitinib group After processing the lung of mice, lung septal thickening, pulmonary interstitial edema and fibrosis capillary injection amount Also dramatically increase；Compare with bleomycin group, after the lung that metformin group processes mice, Lung septal thickening, pulmonary interstitial edema and fibrosis capillary injection amount all significantly reduce；With Ji Non-compare for Buddhist nun's group, after the lung that gefitinib co-administrated metformin group processes mice, between lung Every thickening, pulmonary interstitial edema and fibrosis capillary injection amount all significantly reduce；It follows that Metformin can significantly reduce lung septal thickening, pulmonary interstitial edema and fibrosis capillary injection, It is obviously reduced lung injury degree；Can draw through masson dyeing, compared with matched group, warp After crossing the lung of bleomycin group and gefitinib group process mice, lung fibrosis degree increases the weight of； Compare with bleomycin group, after the lung that gefitinib group processes mice, lung fibrosis Degree also increases the weight of；Compare with bleomycin group, after the lung that metformin group processes mice, Lung fibrosis degree substantially weakens；Compare with gefitinib group, through gefitinib diformazan After biguanide combination group processes the lung of mice, lung fibrosis degree substantially weakens；It follows that Metformin has been obviously reduced the fibrosis of lung tissue.

Figure 4, it is seen that compare with matched group, through bleomycin group and gefitinib After group processes the lung of mice, the content of hydroxyproline dramatically increases；Compare with bleomycin group, After the lung that gefitinib group processes mice, the content of hydroxyproline also dramatically increases；With rich Bleomycin group compares, and after the lung that metformin group processes mice, the content of hydroxyproline shows Write and reduce；Compare with gefitinib group, process mice through gefitinib co-administrated metformin group Lung after, the content of hydroxyproline significantly reduces；It follows that metformin can significantly reduce The content of hydroxyproline, is obviously reduced tissue collagen deposition.

From figure 5 it can be seen that compared with matched group, process mice through bleomycin group After lung, the expression of α-actin albumen substantially increases；Compared with bleomycin group, non-through Ji Organize the lung of process mice for Buddhist nun after, the expression of α-actin albumen the most substantially increases, it follows that, Gefitinib can increase the weight of the table of the pulmonary fibrosis mark α-actin albumen that bleomycin causes Reach；Compare with bleomycin group, after the lung that metformin group processes mice, α-actin The expression of albumen substantially reduces；Compare with gefitinib group, join through gefitinib metformin Process the lung of mice by group after, the expression of α-actin albumen significantly reduces；It follows that two First biguanide not only can be obviously reduced the pulmonary fibrosis mark α-actin egg that bleomycin causes White expression；And the lung fiber that the bleomycin that gefitinib increases the weight of causes can be obviously reduced Change the expression of mark α-actin albumen.

From fig. 6 it can be seen that compared with matched group, through bleomycin group and gefitinib Group process mice lung after, COL1A1, α-actin, pSmad2, pSTAT3, pAKT and The expression of dpERK1/2 albumen substantially increases；Compare with bleomycin group, through gefitinib group After processing the lung of mice, COL1A1, α-actin, pSmad2, pSmad3, pSTAT3 and pAKT The expression of albumen substantially increases；Compared with bleomycin group, process mice through metformin group Lung after, COL1A1, α-actin, pSmad2, pSTAT3, pAKT and dpERK1/2 albumen Expression substantially reduce；Compared with gefitinib group, through gefitinib co-administrated metformin group After processing the lung of mice, COL1A1, α-actin, pSmad2, pSTAT3, pAKT and dpERK1/2 The expression of albumen substantially reduces；Cause it follows that metformin can be obviously reduced bleomycin Pulmonary fibrosis mark α-actin and the expression of COL1A1 albumen；And can significantly subtract The pulmonary fibrosis mark α-actin that the bleomycin that weak gefitinib increases the weight of causes and The expression of COL1A1 albumen；And significantly inhibit the table of TGF-β signal path key downstream albumen Reach, significantly inhibit the key mechanism of pulmonary fibrosis.

2.5 conclusion

From showing that metformin can be obviously reduced the lung fiber that bleomycin causes above Change；And it is obviously reduced the pulmonary fibrosis that the bleomycin that gefitinib increases the weight of causes.

Claims (9)

1. the complex prevented or weaken pulmonary fibrosis, it is characterised in that described complex includes Parts by weight are biguanides and the TKI of 2-5 part of 2-4 part；Described TKI is selected from EGFR-TKI or ALK-TKI.

2. complex as claimed in claim 1, it is characterised in that described biguanides is selected from two First biguanide and salt, phenformin or proguanil；Described EGFR-TKI is selected from gefitinib, strategic point Buddhist nun, Conmana, Afatinib, AZD9291, CO-1686 or HM61713 are replaced in Lip river；Institute State ALK-TKI selected from gram azoles for Buddhist nun or Ceritinib.

3. complex as claimed in claim 2, it is characterised in that described biguanides is diformazan Biguanide, described EGFR-TKI is gefitinib, and the parts by weight of described metformin are 3 Part, the parts by weight of described gefitinib are 2 parts.

4. contain the complex described in claim 3 and tablet that adjuvant is made, it is characterised in that institute Stating tablet is by label and the bilayer tablet that forms of release layer of parcel label, described label by Metformin is made with excipient；Described release layer is made up of gefitinib and adjuvant.

5. the application of the complex as described in claim 1-3 is arbitrary, it is characterised in that described compound Thing prevents in preparation or weakens the application in the medicine of pulmonary fibrosis.

Apply the most as claimed in claim 5, it is characterised in that described pulmonary fibrosis is by EGFR-TKI Or produce during ALK-TKI treatment nonsmall-cell lung cancer.

7. the application of the complex as described in claim 1-3 is arbitrary, it is characterised in that described compound Thing prevents in preparation or slows down the application in the medicine of interstitial pneumonia, described interstitial pneumonia The complication produced when being and treated nonsmall-cell lung cancer by EGFR-TKI or ALK-TKI.

8. the application of tablet as claimed in claim 4, it is characterised in that described tablet is pre-in preparation Anti-or weaken the application in the medicine of pulmonary fibrosis.

Apply the most as claimed in claim 8, it is characterised in that described pulmonary fibrosis is by EGFR-TKI Or produce during ALK-TKI treatment nonsmall-cell lung cancer.