CN106267188A - The novel antibodies of small molecule immune agonist coupling PD 1 antibody and the application in antitumor thereof - Google Patents
The novel antibodies of small molecule immune agonist coupling PD 1 antibody and the application in antitumor thereof Download PDFInfo
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- CN106267188A CN106267188A CN201610671971.8A CN201610671971A CN106267188A CN 106267188 A CN106267188 A CN 106267188A CN 201610671971 A CN201610671971 A CN 201610671971A CN 106267188 A CN106267188 A CN 106267188A
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Abstract
The invention provides novel antibodies and the application thereof of a kind of small molecule immune agonist coupling PD 1 antibody, described novel antibodies is reacted by small molecule immune agonist and PD 1 antibody coupling and obtains, and described novel antibodies includes the compound of below general formula [I]: the novel antibodies in the present invention can be used for immunization therapy and the immunomodulating of malignant tumor.
Description
Technical field
The present invention relates to immunochemistry field, more particularly, it relates to a kind of PD-inhibited to immune system
1 antibody and small molecule immunity agonist chemical combination coupling form novel antibodies, and the preparation of this novel antibodies and anti-swollen
Application in tumor.
Background technology
In recent years, an impressive progress in immunity antitumor field is the immune antitumor action of application PD-1 antibody,
As a breakthrough antibody drug of antineoplastic, its target spot is the expression PD-1 albumen (such as T cell) on immunocyte.
PD-1 is an immune suppressive protein, and suppression and the effect of immunologic balance are excessively played in the immunity to normal human.But swollen
In tumor human body, PD-1 also produces inhibitory action to normal anti tumor immune response;Therefore PD-1 antibody can release this
Immunosuppressive action (Products in China magazine volume 27 6 phase 856 860 in June, 2014) to tumor, this acts on and faces
Bed treatment tumor creates the effect of many long-term treatments.
At present PD-1 antibody the U.S., Japan and other countries listing clinical practice (specifically can be found in following network address:http://www.fiercebiotech.com/biotech/anti-pd-1-cancer-star-nivolumab-wins-
world-s-first-regulatory-approval;http://www.pharmatimes.com/news/japan_
Approves_worlds_first_pd-1_drug%2C_nivolumab_1002153).
But, the effect of PD-1 antibody simply releases the immune suppression effect of " braking ", although therefore its curative effect shows
Write, but only in about 20% to 30% patient effectively.In order to improve its curative effect ratio, a kind of product of needs can be simultaneously
Improve tumour patient autoimmune ability, to produce synergism and higher therapeutic effect.
Summary of the invention
The technical problem to be solved in the present invention is, for the problem that above-mentioned PD-1 antibody curative effect ratio is relatively low, it is provided that one
Plant the novel antibodies of small molecule immune agonist coupling PD-1 antibody and the application in antitumor thereof.
The present invention solves the technical scheme of above-mentioned technical problem and is to provide a kind of small molecule immune agonist coupling PD-1 and resists
The novel antibodies of body, described novel antibodies is reacted by small molecule immune agonist and PD-1 antibody coupling and obtains, described novel anti-
Body includes the compound of below general formula [I]:
In formula [I]: X1Represent OH or SH;R1Represent alkoxyl or alkyl amine group, X2Represent link group;Wherein antibody portion
Dividing and linked with small molecule immune agonist ingredient formed by small molecule immune agonist and described antibody covalent combination, m is
The number (m is defined as coupling degree) of small molecule agonist, is the numeral of 1~10;Antibody moiety refers to the monoclonal antibody for anti-PD-1
(IgG1, IgG2 or IgG4).
When the compound that described small molecule immune agonist is formula 1:
Described X2Expression thiocarbonyl group:
When the compound that described small molecule immune agonist is formula 2-1,2-2,2-3:
Described X2It is expressed as follows group:
When the compound that described small molecule immune agonist is formula 3:
In formula 3, u is the integer of 0~12;
X2It is expressed as follows group:
Wherein u is the integer of 0~12;
When the compound that described small molecule immune agonist is formula 4:
X2It is expressed as follows group:
Wherein PEG is polyethylene groupAs two polyethylene groups areTrimerization second
Glycol group isFour polyethylene groups are
The invention provides a kind of novel bifunctional antibody 10,12,14,16,18, the existing immunity of these novel antibodies presses down
Removing function processed (with the antitumous effect that significantly improves as representative), has activation immunologic function the most simultaneously.
It is in place of the creation of the present invention, it is provided that existing immunosuppressant removing function is (with the antitumous effect significantly improved
For representing), there is novel antibodies activating immunologic function and preparation method thereof the most simultaneously.
It is in place of the creation of the present invention, it is provided that have above-mentioned difunctional, can promote that again active t cell is value-added novel anti-
Body and preparation method thereof.
It is an object of the invention to, it is provided that for preparing coupling precursor micromolecular compound and the synthesis of above-mentioned novel antibodies
The compound of these little molecules of coupling precursor or their salt.
Another object of the present invention is to, it is provided that prepared novel antibodies immunosuppressant release and excite, disease-resistant
Application in the regulation of poison, tumour immunity and knubble biological immunization therapy.
New coupled antibody in the present invention can be used for immunization therapy and the immunomodulating of malignant tumor, and its administrated method can be
Lumbar injection, subcutaneous injection, intramuscular injection and intravenous injection;Or can use the novel antibodies in the present invention and immunocyte
After (such as dendritic cell, natural killer cell NK, lymphocyte, Monocytes/Macrophages, granulocyte etc.) co-culture, separation is exempted from
The method fed back in epidemic disease cyton.
Above-mentioned novel antibodies can be made into the corresponding small molecule immune agonist compound or their salt of preparing them
It is applicable to above-mentioned various medicine, compound medicine, cooperative drug can be made with other drug or make pharmaceutically acceptable
The complex of carrier or conjugate.
The New-type bifunctional antibody of the present invention or their salt can be used for preparing in the medicine of various ratio.
Accompanying drawing explanation
Fig. 1 is interleukin-6 (IL-6) stimulating activity of novel antibodies 10,12,14,16,18.
Fig. 2 is IL-12 (IL-12) stimulating activity of novel antibodies 10,12,14,16,18.
Fig. 3 is the IFN-γ stimulating activity of novel antibodies 10,12,14,16,18.
Fig. 4 is the anti-tumor activity of novel antibodies 10,12,14,16,18.
Fig. 5 is the promotion T cell increment activity of novel antibodies 10,12,14,16,18.
Fig. 6 be small molecule immune agonist 19,20,21,22, the IFN-γ stimulating activity of 20-2.
Fig. 7 is interleukin-6 (IL-6) stimulating activity of small molecule immune agonist 19,20,21,22.
Fig. 8 is the functional character of PD-1 antibody.
Fig. 9,10,11 are the PD-1 antibody function features of novel antibodies of the present invention.
Detailed description of the invention
In order to make the purpose of the present invention, technical scheme and advantage clearer, below in conjunction with drawings and Examples, right
The present invention is further elaborated.Should be appreciated that specific embodiment described herein only in order to explain the present invention, and
It is not used in the restriction present invention.
The invention provides the novel antibodies of a kind of small molecule immune agonist coupling PD-1 antibody, there is following structure
Formula [I] represents:
In formula [I]: X1Represent OH or SH;R1Represent alkoxyl or alkyl amine group, X2Represent link group;Wherein antibody with
Small molecule immune agonist ingredient is linked by small molecule immune agonist and described antibody covalent combination and is formed, and m is little point
The number (m is defined as coupling degree) of sub-agonist, is the numeral of 1~10;Antibody moiety refer to monoclonal antibody for anti-PD-1 (IgG1,
IgG2 or IgG4).
When the compound that described small molecule immune agonist is formula 1:
Described X2Expression thiocarbonyl group:
When the compound that described small molecule immune agonist is formula 2-1,2-2,2-3:
Described X2It is expressed as follows group:
When the compound that described small molecule immune agonist is formula 3:
In formula 3, u is the integer of 0~12;
X2It is expressed as follows group:
Wherein u is the integer of 0~12;
When the compound that described small molecule immune agonist is formula 4:
X2It is expressed as follows group:
Wherein PEG is polyethylene groupAs two polyethylene groups areThree
Polyethylene group isFour polyethylene groups are
Deng.
When small molecule immune agonist be formula 1,2-1,2-2,2-3,4 represent compound time, with PD-1 antibody and other
The Typical Representative structural formula of the novel antibodies formed as antibody moiety as a example by antibody is as follows:
Wherein m is the number between 1 to 10.
When the compound that the little molecule of the precursor of coupling novel antibodies is formula 1,2-1,2-2,2-3,4 representatives, resist with PD-1
The Typical Representative synthesis of the novel antibodies formed as representative antibodies as a example by body and structural formula thereof are as follows:
In the novel antibodies that above-mentioned coupling is formed, antibody is selected from PD-1 monoclonal antibody;Also apply be applicable to other similar solution
Except immunosuppressant antibody, such as PD-L1 antibody, CTLA-4 antibody, TIM-3 antibody, LAG3 antibody, TIGIT antibody etc.;PD-1 is mono-
The anti-monoclonal antibody (IgG1, IgG2 and IgG4) including various anti-human PD-1 albumen, the sequence of described PD-1 albumen is as follows:
1MQIPQAPWPWWAVLQLGWRPGWFLDSPDRPWNPPTFSPALLVVTEGDNATFTCSFSNTS61ESFVLNWYR70MSPSNQ
TDKLAAFPEDR86SQPGQDCRFRVTQLPNGRDFHMSWRARRND118SGTYLCGAISLAPKAQIKE136SLRAELRVTERR
AEVPTAHPSPSPRPAGQFQTLVVGVVGGLLGS181LVLLVWVLAVICSRAARGTIGARRTGQPLKEDP5AVPVFSVDY
GELDFQWREKTPEPPVP241CVPEQTEYATIVFP5GMGTSSPARRGSADGPRSAQPLRPEDGHCSWPL
Described PD-1 antibody is the epitope sequences of antigen for this sequence.
Coupling precursor compound 11 synthesis citing:
20-1 is dissolved in anhydrous DMSO, adds the succinic anhydride of equivalent, mixture natural room temperature in 10 DEG C of coolings
Stir 24 hours.Mixed reactant falls with 20 times of volume of water, separates out a large amount of compound as white solid SZU-101;Yield 88%;
MS(ESI)445.27(M+1);
SZU-101 (1eq), NHS (1.2eq) and EDC (1.3eq) are dissolved in dry DMF, under room temperature, stir 4h, terminate anti-
Should, reactant liquor is poured in dichloromethane, sucking filtration, be dried, obtain active ester 23 white solid, be directly used in next step reaction.Will
Active ester and NH2-PEG2-COOH (compound 24) are dissolved in DMF in equivalent ratio, be stirred at room temperature to reaction terminate (TLC supervise
Survey).Adding water, separate out a large amount of solid, sucking filtration, be dried, column chromatographic isolation and purification (dichloromethane: methanol=10:1) obtains white solid
Compound 19, yield 86%;Fusing point 221 DEG C;High resolution mass spectrum molecular weight HRMS (ESI) theoretical value m/z 603.2746, found
604.2757(M+H)。
19 (1eq), NHS (1.2eq) and EDC (1.3eq) are dissolved in dry DMF, stir 12h under room temperature, terminate reaction, will
Reactant liquor is poured in dichloromethane, sucking filtration, is dried, obtains active ester compound 11 white solid, yield 82%;Fusing point 187 DEG C;High
Resolution Mass Spectrometry molecular weight HRMS (ESI) theoretical value m/z 700.2839, found 701.2842 (M+1).
Coupling precursor compound 13 synthesis citing:
Compound 20-1 (with reference to Chinese patent CN201210382202.8) (1eq) and TGA (1eq) are dissolved in nothing
Water DMF, adds 1.2eq HBTU, 3eq triethylamine and 0.1eq DMAP, is stirred at room temperature, and after reaction terminates, adds water, and sucking filtration is dry
Dry 20-2 faint yellow solid.It is stirred at room temperature in DMF with 4-(N-maleimidomehyl) hexamethylene-1-carboxylic acid (1eq) again,
TLC monitoring raw material disappears, and reactant mixture pours into cold water, and product solid filters, and is dried to obtain compound 20, MS (ESI)
656.27 (M+1). compound 20 (1eq) is added NHS (1.2eq) and the DMF solution stirred overnight at room temperature of EDC (1.3eq), instead
After should completing, obtaining active ester compound 13, fusing point 201 DEG C through preparation liquid phase separation, yield 35% (is calculated from initiation material 20-1
Rise);High resolution mass spectrum molecular weight HRMS (ESI) theoretical value m/z 752.2630, found 753.2632 (M+1).
Coupling precursor compound 15,17 synthesis preparation method, reaction condition are identical with preparing compound 13 with ingredient proportion,
Simply willWithReplace;
Coupling precursor compound 15 synthetic route:
Obtain compound 21, white solid, Mass Spectrometric Identification molecular weight: HRMS (ESI) 630.2356 (M+1).Obtain coupling precursor
Compound 15, white solid, Mass Spectrometric Identification molecular weight: HRMS (ESI) 727.2431 (M+1).
The synthetic route of coupling precursor compound 17:
Obtain compound 22, white solid, Mass Spectrometric Identification molecular weight: HRMS (ESI) 588.1819 (M+1).Obtain coupling precursor
Compound 17, white solid, Mass Spectrometric Identification molecular weight: HRMS (ESI) 685.2022 (M+1).
With New-type bifunctional antibody preparation synthesis implementation example in the present invention that PD-1 antibody is formed:
Identification mass spectrograph model: LDI-1700 laser desorption ionisation flight time mass spectrum (MALDI-TOF-MS) is raw
Business men: Linear Scientific company of the U.S..
Mass spectrum determines that the method for the ratio (coupling degree) of small molecule immune agonist is as follows: Mass Spectrometric Identification gained coupling is novel
The molecular weight of antibody products deducts former PD-1 antibody molecule and measures value added, calculates: molecular weight value added/(little point of coupling precursor
Sub-molecular weight-18)=coupling degree;(1 molecular weight deducting 1 hydrone of the every coupling of micromolecular compound).
PD-1 antibody first-selection is Nivolumab (BMS-936558), Pembrolizumab (MK-3475), AMP-514,
AMP-224, Pidilizumab;But it is also applied for the PD-1 antibody in other source.
The preparation of Nivolumab according in patent WO2006121168 prepare 5C4,17D8,2D3,4H1,4A11,7D3 and
The method of 5F4;And the method announced in CN201380062005.0.
The preparation of Pembrolizumab according to preparation h409A11 in patent WO2008156712 (A1) and
The method announced in US8354509 and US8900587 patent.
The preparation of AMP-514 and AMP-224 is according to the method announced in patent WO201214549 patent.
The preparation of Pidilizumab is according to the method announced in patent WO2009014708 and WO2009114335 patent.
The preparation method of PD-1 antibody may be used without other open method well known in the art, as at Chinese patent
CN201410838610.9;CN201310199947.5;CN201380079581.6;CN201480011008.6;
Method disclosed in CN201310258289.2.
PD-1 antibody can also be bought as source, such as purchased from BioXCell company of U.S. PD-1 antibody, title in market:
Anti h PD-1 (h CD279) IgG1, content 95%;Ke Xin bio tech ltd, Beijing anti-PD-1 human monoclonal resists
Body.PD-1 antibody sources in novel antibodies synthetic method of the present invention is not limited to this.
The reaction density of following PD-1 antibody is 1 equivalent concentration (1eq, relative to other reactant).
The preparation of novel antibodies 10:
The synthesis preparation method of coupling precursor micromolecular compound 1 refer to Chinese patent (CN201210382202.8).
Compound 1 (10eq equivalent concentration) and PD-1 antibody (Nivolumab) are mixed in the DMSO solvent of 1mL, mixed
Compound added triethylamine 0.1mL, with 5 DEG C to 20 DEG C of stirring reactions 12 hours.Reactant mixture mixes with 0 DEG C of pure water of 10mL,
Shake up, be filtered to remove little molecule with the biological filter membrane of 10kD, use pure water eluted product, eluent lyophilization to obtain novel antibodies 10
(yield 60%).Mass Spectrometric Identification molecular weight increases to 1167;Calculating coupling degree is 3.(1167/386=3.02).
The preparation of novel antibodies 12:
Compound 11 (30eq equivalent concentration) and PD-1 antibody (Nivolumab) are mixed in appropriate DMSO solvent, mixed
Compound addition triethylamine regulation pH value was to 8, with 5 DEG C to 20 DEG C of stirring reactions 12 hours.Reactant mixture and 0 DEG C of 10 times of volumes
Pure water mixes, and shakes up, is filtered to remove little molecule with the biological filter membrane of 10kD, is temporarily released from one's regular work thing by pure water rinsing, eluent lyophilization
Obtain novel antibodies 12 (yield 72%).Mass Spectrometric Identification molecular weight increases to 2931;Calculating coupling degree is 5.(2931/585.6=
5)。
(in the preparation method of novel antibodies 12, Nivolumab is replaced with BioXCell company PD-1 antibody: anti
H PD-1 (h CD279) IgG1, obtains novel antibodies 12-2, and the result of its preparation process and gained novel antibodies 12-2 is with novel
Antibody 12 is identical).
The preparation of novel antibodies 14:
Compound 13 (40eq equivalent concentration) and PD-1 antibody (AMP-514) are mixed in appropriate DMSO solvent, mixing
Thing addition triethylamine regulation pH value was to 8, with 5 DEG C to 20 DEG C of stirring reactions 12 hours.0 DEG C of reactant mixture and 10 times of volumes pure
Water mixes, and shakes up, is filtered to remove little molecule with the biological filter membrane of 10kD, is temporarily released from one's regular work thing by pure water rinsing, and eluent lyophilization obtains
Novel antibodies 14 (yield 66%).Mass Spectrometric Identification molecular weight increases to 1917;Calculating coupling degree is 3.
The preparation of novel antibodies 16:
Compound 15 (40eq equivalent concentration) and PD-1 antibody (Pembrolizumab) are mixed in appropriate DMSO solvent
In, mixture addition triethylamine regulation pH value was to 8, with 5 DEG C to 20 DEG C of stirring reactions 12 hours.Reactant mixture and 10 times of volumes
The mixing of 0 DEG C of pure water, shake up, be filtered to remove little molecule with the biological filter membrane of 10kD, be temporarily released from one's regular work thing by pure water rinsing, eluent is freezing
It is dried to obtain novel antibodies 16 (yield 73%).Mass Spectrometric Identification molecular weight increases to 2377;Calculate coupling degree and be about 4.
The preparation of novel antibodies 18:
Compound 17 (40eq equivalent concentration) and PD-1 antibody (AMP-224) are mixed in appropriate DMSO solvent, mixing
Thing addition triethylamine regulation pH value was to 8, with 5 DEG C to 20 DEG C of stirring reactions 12 hours.0 DEG C of reactant mixture and 10 times of volumes pure
Water mixes, and shakes up, is filtered to remove little molecule with the biological filter membrane of 10kD, is temporarily released from one's regular work thing by pure water rinsing, and eluent lyophilization obtains
Novel antibodies 18 (yield 57%).Mass Spectrometric Identification molecular weight increases to 1143;Calculate coupling degree and be about 2.
The immune activation experimental technique (mice) of novel antibodies of the present invention:
1. take Balb/C mouse spleen lymphocyte, by 1x106/ ml/ hole bed board
2. preferred antibody concentration: stimulate 24h by 0.1 μM of dosing.
3. collect supernatant and detect IL-6/IL-12 (effect is as shown in Figure 1, 2) by ELISA method.
The immune activation experimental technique (mice) of small molecular immunity agonist of the present invention:
1. take Balb/C mouse spleen lymphocyte, by 1x106/ ml/ hole bed board
2. choose the concentration of small molecule immune agonist: stimulate 24h by 0.1,10,20 μMs of dosings.
3. collect supernatant and detect IFN-γ/IL-6 (effect is as shown in Figure 6,7) by ELISA method.
The immune activation experimental technique (people) of novel antibodies of the present invention:
1. take volunteer peripheral mononuclear cells, isolate T cell (suspension cell) in 37 DEG C, 5%CO2Containing blood serum medium
Cultivate 7 days, by 1x106/ ml/ hole bed board
2. preferred antibody concentration: stimulate 24h by 0.1 μM of dosing.
3. collect supernatant and detect IFN-γ (as shown in Figure 3) by ELISA method.
The antitumous effect experimental technique of novel antibodies of the present invention:
The Balb/C mice of 68 week old is randomly divided into 7 groups.At mouse back subcutaneous implantation 2.5x105Individual 4T1 tumor
Cell.It is administered with the PD-1 antibody control of PBS blank, 10mg/kg respectively;Comparison medicine and novel antibodies 10,12,14,16,
The dosage of 18 is 10mg/kg, volume 100 μ L;Each group administering mode is lumbar injection.After implantation tumor the 1st day, the
7,15,22,29 days each group be administered respectively.When tumor reaches 1500mm3Or more than body weight 15% time by mice euthanasia.Swollen
Tumor suppression result is as shown in Figure 4.In the visible present invention, novel antibodies process group has the Graft Versus Tumor significantly improved.
The promotion T cell increment experimental technique of novel antibodies of the present invention:
Take volunteer peripheral mononuclear cells, isolate T cell (suspension cell) in 37 DEG C, 5%CO2Train containing blood serum medium
After supporting 7 days, change serum-free medium and be simultaneously introduced novel antibodies and the PD-1 (comparison) (concentration is 1 μM) of the present invention, after 3 days
CCK-8 cell dyeing method detects.Visible novel antibodies of the present invention can promote by strengthening blocking-up PD-1 and the activation natural immunity
Enter T cell increment.Result is as shown in Figure 5.
The PD-1 antibody function characterization experiments method of novel antibodies of the present invention (resists with the specificity two anti-detection PD-1 of PD-1
Body):
1. extraction healthy volunteer's peripheral blood
2. acquirement peripheral blood is in centrifuge tube, 800g, 30min.Separate serum and hemocyte.
3. it is resuspended in normal saline after obtaining hemocyte, adds to peripheral blood lymphocyte separation liquid along wall gently
Layer.
4.800g is centrifuged 30min. and draws tunica albuginea layer (periphery lymphocyte layer).
5. the peripheral blood lymphocyte of isolated is resuspended in 900g in normal saline, 10min.Wash 2 times.
6. counting separates the peripheral blood lymphocyte obtained.
7. taking 1x106 cell, 100ul normal saline is resuspended, uses PD-1 antibody respectively, novel antibodies of the present invention 10,12,
16, and IgG1 (comparison), hatch 30min for 37 degree.
8. brine 3 times, resuspended with 100ul normal saline, add the two of Anti-PD-1-FITC and resist, room temperature is incubated
Educate 30min.
9. brine, machine testing in streaming.
Result is as shown in Fig. 8,9,10,11: compare with the group of matched group and non-coupling it is found that
The PD-1 antibody function feature of the novel antibodies of the present invention remains intact.
The above, the only present invention preferably detailed description of the invention, but protection scope of the present invention is not limited thereto,
Any those familiar with the art in the technical scope that the invention discloses, the change that can readily occur in or replacement,
All should contain within protection scope of the present invention.Therefore, protection scope of the present invention should be with scope of the claims
It is as the criterion.
Claims (10)
1. the novel antibodies of a small molecule immune agonist coupling PD-1 antibody, it is characterised in that described novel antibodies is by little
Molecular immune agonist and the reaction of PD-1 antibody coupling obtain, and described novel antibodies includes the compound of below general formula [I]:
In formula [I]: X1Represent OH or SH;R1Represent alkoxyl or alkyl amine group, X2Represent link group;Wherein antibody moiety with
Small molecule immune agonist ingredient is to be linked shape by small molecule immune agonist coupling precursor and described antibody covalent combination
Becoming, m is the number (m is defined as coupling degree) of small molecule agonist, is the numeral of 1~10;Antibody moiety refers to the list for anti-PD-1
Anti-(IgG1, IgG2 or IgG4);Described small molecule immune agonist coupling precursor is defined as when carrying out covalent combination with antibody
The activated form of small molecule immune agonist.
When the compound that described small molecule immune agonist coupling precursor is formula 1:
Described X2Expression thiocarbonyl group:
When the compound that described small molecule immune agonist coupling precursor is formula 2-1,2-2,2-3:
Described X2It is expressed as follows group:
When the compound that described small molecule immune agonist coupling precursor is formula 3:
In formula 3, u is the integer of 0~12;
X2It is expressed as follows group:
Wherein u is the integer of 0~12;
When the compound that described small molecule immune agonist coupling precursor is formula 4:
X2It is expressed as follows group:
Wherein PEG is polyethylene groupAs two polyethylene groups areTrimerization second two
Alcohol groups isFour polyethylene groups are
The novel antibodies of small molecule immune agonist coupling PD-1 antibody the most according to claim 1, it is characterised in that: institute
Stating PD-1 antibody and include the monoclonal antibody (IgG1, IgG2 or IgG4) of various anti-human PD-1 albumen, the sequence of described PD-1 albumen is preferential
Select as follows:
1MQIPQAPWPWWAVLQLGWRPGWFLDSPDRPWNPPTFSPALLWTEGDNATFTCSFSNTS61ESFVLNWYR70MSP
SNQTDKLAAFPEDR86SQPGQDCRFRVTQLPNGRDFHMSWRARRND118SGTYLCGAISLAPKAQIKE136SLRAELRVT
ERRAEVPTAHPSPSPRPAGQFQTLWGVVGGLLGS181LVLLVWVLAVICSRAARGTIGARRTGQPLKEDPSAVPVFSV
DYGELDFQWREKTPEPPVP241CVPEQTEYATIVFPSGMGTSSPARRGSADGPRSAQPLRPEDGHCSWPL;
Described PD-1 antibody is the epitope sequences of antigen for this sequence.
The novel antibodies of small molecule immune agonist coupling PD-1 antibody the most according to claim 1, it is characterised in that: institute
The PD-1 antibody stated is Nivolumab, Pembrolizumab, AMP-514, AMP-224, Pidilizumab or other anti h
PD-1(h CD279)IgG1。
The novel antibodies of small molecule immune agonist coupling PD-1 antibody the most according to claim 1, it is characterised in that: when
When small molecule immune agonist is the compound of formula 1,2-1,2-2,2-3,4 representatives, the Typical Representative structure of described novel antibodies
Formula is as follows:
Wherein m is the number between 1 to 10.
The novel antibodies of small molecule immune agonist coupling PD-1 antibody the most according to claim 4, it is characterised in that: when
Small molecule immune agonist be formula 1,2-1,2-2,2-3,4 represent compound time, the Typical Representative 10 of described novel antibodies,
12, the structural formula of 14,16,18 is as follows:
6. a small molecule immune agonist coupling precursor compound, it is characterised in that: for new as in claim 5 of preparation
The Typical Representative 12,14,16,18 of type antibody, correspondence includes the compound of following structural formula 11,13,15,17:
7. exciting for synthesizing the small molecule immune of small molecule immune agonist coupling precursor compound as claimed in claim 6
Immunomodulator compounds, it is characterised in that: described small molecule immune agonist compound for include structure as shown below formula 19,20-2,20,
21,22 compound or their salt:
8. according to fitting that the novel antibodies according to any one of claim 1---7 and micromolecular compound or their salt are made
Make compound medicine, cooperative drug for various medicines or with other drug, or make pharmaceutically acceptable carrier
Complex or conjugate.
9. use medicine as described in claim 8, immunosuppressant release and excite, prepared by antibody, antiviral, glycosuria
Application in disease and immunotherapy of tumors.
10. the application of as described in claim 9 medicine, administrated method is lumbar injection, subcutaneous injection, intramuscular injection and quiet
Arteries and veins injection or oral;Or can be used for immunocyte (as dendritic cell, natural killer cell NK, lymphocyte, monokaryon/
Macrophage, granulocyte etc.) co-culture after, the method fed back in isolating immune cells body.
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WO2018032793A1 (en) * | 2016-08-15 | 2018-02-22 | 深圳大学 | Novel antibody obtained by coupling small molecule immuno-agonist and pd-1 antibody, and application thereof in tumor resistance |
CN108379591A (en) * | 2018-04-03 | 2018-08-10 | 深圳大学 | The synthesis and its application of immune agonist target compound |
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WO2023174312A1 (en) * | 2022-03-15 | 2023-09-21 | 中国科学院上海药物研究所 | Anti-human pd-l1 and tlr7 double-targeting nanobody conjugate drug and use thereof in resisting tumor |
Families Citing this family (4)
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---|---|---|---|---|
WO2018127918A1 (en) | 2017-01-05 | 2018-07-12 | Kahr Medical Ltd. | A sirp alpha-cd70 fusion protein and methods of use thereof |
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Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101239980A (en) * | 2008-02-18 | 2008-08-13 | 靳广毅 | Immune receptor regulator couplet precursor, couplet and application thereof |
WO2011134669A1 (en) * | 2010-04-30 | 2011-11-03 | Telormedix Sa | Methods for inducing an immune response |
CN102993265A (en) * | 2012-10-10 | 2013-03-27 | 靳广毅 | Immune receptor regulator couplet as well as preparation method and application thereof, coupling precursor for preparing immune receptor regulator couplet and compound for compounding coupling precursor |
CN103254304A (en) * | 2012-05-30 | 2013-08-21 | 深圳大学 | Immune stimulant couplet, preparation method thereof and antineoplastic application of the immune stimulant couplet |
CN104220441A (en) * | 2012-02-13 | 2014-12-17 | 百时美施贵宝公司 | Enediyne compounds, conjugates thereof, and uses and methods therefor |
CN104974252A (en) * | 2014-04-01 | 2015-10-14 | 上海中信国健药业股份有限公司 | Antibody-small molecule drug conjugate capable of inhibiting tumor growth, and preparation method and application thereof |
Family Cites Families (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101230064A (en) * | 2007-01-22 | 2008-07-30 | 靳广毅 | 9 methylene aryl substituted purine derivative |
CN102439011B (en) * | 2009-02-11 | 2016-05-04 | 加利福尼亚大学校务委员会 | The treatment of TOLL sample receptor modulators and disease |
CN103467590B (en) * | 2013-09-02 | 2016-04-13 | 深圳大学 | Bioconjugation body, the purine compound preparing it, synthetic method, medicinal preparations and its application in immunomodulatory |
CN105311032B (en) * | 2015-11-09 | 2018-09-25 | 深圳大学 | TLR7 agonists are combined application in preparation of anti-tumor drugs with tyrosine kinase inhibitor |
CN105315281B (en) * | 2015-11-16 | 2017-04-19 | 深圳大学 | Compound I and compound II as well as preparation methods and application thereof |
CN105327345A (en) * | 2015-11-19 | 2016-02-17 | 深圳大学 | Vaccine adjuvant and vaccine comprising same |
CN106267188A (en) * | 2016-08-15 | 2017-01-04 | 深圳大学 | The novel antibodies of small molecule immune agonist coupling PD 1 antibody and the application in antitumor thereof |
-
2016
- 2016-08-15 CN CN201610671971.8A patent/CN106267188A/en active Pending
-
2017
- 2017-04-19 WO PCT/CN2017/081007 patent/WO2018032793A1/en active Application Filing
Patent Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101239980A (en) * | 2008-02-18 | 2008-08-13 | 靳广毅 | Immune receptor regulator couplet precursor, couplet and application thereof |
WO2011134669A1 (en) * | 2010-04-30 | 2011-11-03 | Telormedix Sa | Methods for inducing an immune response |
CN104220441A (en) * | 2012-02-13 | 2014-12-17 | 百时美施贵宝公司 | Enediyne compounds, conjugates thereof, and uses and methods therefor |
CN103254304A (en) * | 2012-05-30 | 2013-08-21 | 深圳大学 | Immune stimulant couplet, preparation method thereof and antineoplastic application of the immune stimulant couplet |
CN102993265A (en) * | 2012-10-10 | 2013-03-27 | 靳广毅 | Immune receptor regulator couplet as well as preparation method and application thereof, coupling precursor for preparing immune receptor regulator couplet and compound for compounding coupling precursor |
CN104974252A (en) * | 2014-04-01 | 2015-10-14 | 上海中信国健药业股份有限公司 | Antibody-small molecule drug conjugate capable of inhibiting tumor growth, and preparation method and application thereof |
Cited By (12)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2018032793A1 (en) * | 2016-08-15 | 2018-02-22 | 深圳大学 | Novel antibody obtained by coupling small molecule immuno-agonist and pd-1 antibody, and application thereof in tumor resistance |
CN107281483A (en) * | 2017-04-25 | 2017-10-24 | 深圳大学 | The preparation method of immune activator and composition and its application and composition |
CN107281484A (en) * | 2017-05-31 | 2017-10-24 | 深圳大学 | Composition for preparing hepatitis B vaccine and its preparation method and application |
WO2018219266A1 (en) * | 2017-05-31 | 2018-12-06 | 深圳大学 | Composition for preparing hepatitis b vaccine, preparation method therefor, and application thereof |
CN108379591A (en) * | 2018-04-03 | 2018-08-10 | 深圳大学 | The synthesis and its application of immune agonist target compound |
WO2019192454A1 (en) * | 2018-04-03 | 2019-10-10 | 深圳大学 | Novel small molecule immune agonists and immune targeting compounds and application thereof |
CN108379591B (en) * | 2018-04-03 | 2022-03-29 | 深圳大学 | Synthesis of immune agonist targeting compound and application thereof |
CN115192726A (en) * | 2018-04-03 | 2022-10-18 | 深圳大学 | Synthesis of immune agonist targeting compound and application thereof |
CN111704614A (en) * | 2020-04-15 | 2020-09-25 | 深圳大学 | Novel series immune agonists |
WO2021208459A1 (en) * | 2020-04-15 | 2021-10-21 | 深圳大学 | Immune agonists |
WO2022194153A1 (en) * | 2021-03-16 | 2022-09-22 | 中国科学院上海药物研究所 | Pd-l1 and tlr7 double-targeting nanobody coupling drug and use thereof in anti-tumor |
WO2023174312A1 (en) * | 2022-03-15 | 2023-09-21 | 中国科学院上海药物研究所 | Anti-human pd-l1 and tlr7 double-targeting nanobody conjugate drug and use thereof in resisting tumor |
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