CN101287500A - Multi-drug ligand conjugates - Google Patents
Multi-drug ligand conjugates Download PDFInfo
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- CN101287500A CN101287500A CNA2006800383623A CN200680038362A CN101287500A CN 101287500 A CN101287500 A CN 101287500A CN A2006800383623 A CNA2006800383623 A CN A2006800383623A CN 200680038362 A CN200680038362 A CN 200680038362A CN 101287500 A CN101287500 A CN 101287500A
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Abstract
Described herein are compounds, pharmaceutical compositions and methods for treating pathogenic cell populations in a patient. The compounds described herein include conjugates of a plurality of cytotoxic drugs and vitamin receptor binding ligands. The plurality of drugs may be the same or different. Similarly, the vitamin receptor binding ligands may be the same or different. The conjugates also include a linker that is formed from one or more spacer linkers, heteroatom linkers, and releasable linkers.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
The application requires the U.S. Provisional Patent Application sequence number 60/709 of submission on August 19th, 2005, the U.S. Provisional Patent Application sequence number 60/787 that on March 30th, 950 and 2006 submitted to, rights and interests under 558 35U.S.C. § 119 (e), described document integral body by reference is attached to herein.
Technical field
The present invention relates to be used for compositions and the method that targeted drug discharges.Especially, the present invention relates to contain the ligand conjugates of two or more medicines and analog and derivant, for example in conjunction with the chemical compound of vitamin receptor and the conjugate of two or more medicines.
Background
Immune system provides identification and eliminates the mode of tumor cell, other pathogenic cell and invasive foreign pathogens.Though immune system provides strong defence line usually, there are a lot of wherein cancerous cell, other pathogenic cell or infectant to escape host immune response and hypertrophy or lasting and the pathogenic situation about accompanying of host.Chemotherapeutics and radiotherapy that tumor is for example duplicated in elimination have been developed.But many existing chemotherapeutics and radiotherapy scheme have adverse side effect, because their effect not only destroys pathogenic cell, and influence for example cell of hemopoietic system of normal host cell.The explanation of the adverse side effect of these cancer therapy drugs is new selective and the little therapy of host toxicity had needs to pathogenic cell colony to developing.
Research worker has been developed by making this type of cell of cytotoxic compound targeting to destroy the therapeutic scheme of pathogenic cell.A plurality of this type of schemes are used the toxin of puting together with antibody, and these antibody combine with antigenic specificity, or pass through the pathogenic cell overexpression, make with expectation that to be released into Normocellular toxin minimum.Develop some immunotoxin of being made up of the antibody of the specific antigen on the guiding pathogenic cell with this method, for example ricin, Pseudomonas exotoxin, diphtheria (Diptheria) toxin are connected with tumor necrosis factor antibody with toxin.These immunotoxin targeting pathogenic cells for example have by the tumor cell of the specific antigen of antibody recognition (Olsnes, S., Immunol.Today, 10, pp.291-295,1989; Melby, E.L., Cancer Res., 53 (8), pp.1755-1760,1993; Better, M.D., PCT international publication number WO announced on May 30th, 91/07418,1991).
The method of population of pathogenic cells style such as cancerous cell or allos pathogen is to strengthen the immunne response of host to pathogenic cell among the another kind of targeting host, avoids also can having the administration needs of the chemical compound of independent host toxicity.It is reported, a strategy of immunotherapy be make antibody for example many subunit antibodies of genetic engineering preparation combine with tumor cell surface, constant region with the lip-deep antibody of exposed cell, thereby induce by the process of various immune-mediated and to kill tumor cell (De Vita, V.T., Biologic Therapy of Cancer, 2d ed.Philadelphia, Lippincott, 1995; Soulillou, J.P., U.S. Patent number 5,672,486).But because be difficult to define tumor specific antigen, so these methods are very complicated.
Summary of the invention
Set forth the ligand conjugates of medicine and analog thereof and derivant herein.These conjugates comprise the part in conjunction with cell receptor, but these parts are covalently bound with two or more medicines of targeted cells.Described herein conjugate also can comprise the multivalence joint that part is connected with medicine.
In one embodiment, set forth the drug release conjugate of bind receptor.This drug release conjugate contains part, two or more medicines or its analog or the derivant and the optional multivalence joint of cell surface receptor, and this conjugate can be represented by following formula usually
(B)-(L)-(D)
n
Wherein (B) represents the part of bind receptor; (D) representative (L) is represented the multivalence joint by medicine or its analog or the derivant of the part targeted cells of bind receptor, and n is the integer greater than 1.Multivalence joint (L) can contain covalently bound a plurality of joints mutually.For example, multivalence joint (L) can contain one or more interval base joint (l
s), and/or separable joint (l
r), interconnection separately and by one or more hetero atom joint (l
H) be connected with medicine with part.Can select these different joints,, make up multivalence joint (L) by any sequence arrangement.For example, can make up multivalence joint (L) by one or more following bivalence joints:
-(L)-
-(l
r)
c-
-(l
s)
a-
-(l
s)
a-(l
r)
c-
-(l
r)
c-(l
s)
a-
-(l
H)
b-(l
r)
c-
-(l
r)
c-(l
H)
b-
-(l
H)
d-(l
r)
c-(l
H)
e-
-(l
s)
a-(l
H)
b-(l
r)
c-
-(l
r)
c-(l
H)
b-(l
s)
a-
-(l
H)
d-(l
s)
a-(l
r)
c-(l
H)
e-
-(l
H)
d-(l
r)
c-(l
s)
a-(l
H)
e-
-(l
H)
d-(l
s)
a-(l
H)
b-(l
r)
c-(l
H)
e-
-(l
H)
d-(l
r)
c-(l
H)
b-(l
s)
a-(l
H)
e-
-(l
s)
a-(l
r)
c-(l
H)
b-
-[(l
s)
a-(l
H)
b]
d-(l
r)
c-(l
H)
e-
Wherein a, b, c, d and e be integer for example at the integer of about 4 scopes of 0-, (l
s), (l
H) and (l
r) be respectively the basic joint in interval, separable joint, hetero atom joint.Other illustrative examples of bivalence joint that can be used for making up described multivalence joint herein is at u.s. patent application serial number 10/765, be described among 336 (also can find at U.S. Patent Application Publication No. US 2005/0002942 A1) and the PCT international publication number WO 2006/012527, described document integral body by reference is attached to herein.
Should be understood that the multivalence joint can be connected with two or more medicines the part of bind receptor by multiple node configuration, include but not limited to following exemplary general formula:
B-L
1-D
1-L
2-D
2 B-L
1-D
1-L
2-D
2-L
3-D
3
D
1-L
1-B-L
2-D
2 D
1-L
1-B-L
2-D
2-L
3-D
3
Wherein B is the part of bind receptor, (L
1), (L
2) and (L
3) each multivalence joint naturally, constitute by one or more intervals base, separable and/or hetero atom joint, and (D
1), D
2And D
3Each is medicine or its analog or derivant naturally.Other modification comprise other medicines or its analog or derivant, other joint and (B), (L) and other configuration of (D) arranging separately be also included within herein.
In a kind of modification, be included in the described herein drug release conjugate in conjunction with the part of an above receptor, the drug release conjugate includes but not limited to following exemplary general formula:
Wherein each B is the part of bind receptor, (L
1), (L
2) and (L
3) each multivalence joint naturally, constitute (D by one or more intervals base, separable and/or hetero atom joint
1), D
2And D
3Each is medicine or its analog or derivant naturally.Other modification comprise other medicines or its analog or derivant, other joint and (B), (L) and other configuration of (D) arranging separately be also included within herein.In a kind of modification, the part of bind receptor is in conjunction with same receptor, and in another kind of modification, the part of bind receptor is in conjunction with different receptors.
In the exemplary of a described herein drug release conjugate, the multivalence joint comprises at least one separable joint (l
r).In the exemplary of another described herein drug release conjugate, the multivalence joint comprises at least two separable joint (l
r)
2In another illustrative aspects, multivalence joint (L) comprises at least one separable joint (l
r), this joint is not the separable joint of disulphide (disulfide).In another illustrative aspects, multivalence joint (L) has at least two separable joint (l
r)
2, one of them separable joint is not the separable joint of disulphide.Can recognize that when more than one separable joint was included in the multivalence joint, those separable joints can be adjacent.Can recognize that also when two separable joints in the multivalence joint were adjacent, these two separable joints can be worked in coordination with and be caused drug release.
In another embodiment, the multivalence joint comprises at least one by the basic joint in amino acids formed peptide interval, and in one aspect, peptide comprises natural amino acid and stereoisomer thereof.In yet another aspect, peptide is only formed by natural amino acid and stereoisomer thereof.
Described herein part generally includes the part of cell surface receptor.Can be used for the exemplary part of described conjugate herein and include but not limited to vitamin and the albumen or its analog or the derivant that exist with the bonded other parts of vitamin receptor, transport protein or other and the bonded surface of vitamin specificity; Peptide part by the storehouse Screening and Identification; Tumor cell-specific peptide; Tumor cell-specificity is fit; Tumor cell-specificity sugar; Tumor cell-specific monoclonal or polyclonal antibody; The Fab of antibody or scFv (being the strand variable domain) fragment is the Fab fragment of the antibody of targeting EphA2 for example, or on the metastatic carcinoma cell specific expressed or unique other albumen that reaches; The little organic molecule that obtains by combinatorial libraries; Somatomedin is EGF, FGF, insulin and insulin like growth factor for example; And homeopeptide; Somatostatin and analog thereof; Transferrins; Protein-lipid complex; Bile salt; Select albumen; Steroid hormone; The peptide that contains Arg-Gly-Asp; Biostearin; Various gala agglutinins; δ-opioid receptor part; The CCK A receptors ligand; The ligands specific of angiotensin AT1 or AT2 receptor; Peroxisome proliferation-activated receptors λ part; Beta-Lactam antibiotic is penicillin for example; The little organic molecule that comprises antimicrobial drug; With with preferential bonded other molecule of receptor-specific of expressing on tumor cell surface or infectious organisms; Antimicrobial drug and according to the other medicines of the binding pocket of the suitable special receptor of receptor or other cell surface protein crystal structure design; Tumor antigen or on tumor cell surface the part of preferential other molecule of expressing, or the fragment of any of these molecule.The extracellular epi-position that the tumor-specific antigen that can play part-drug conjugate binding site effect comprises Ephrin family protein film is EphA2 for example.In normal cell, EphA2 expresses and is limited to cell-cell contact, but in metastatic cancer cell, EphA2 is distributed in whole cell surface.Therefore, the EphA2 on the metastatic cell can combine with the Fab fragment of the antibody of for example puting together with medicine or its analog or derivant, and albumen can not combine with the Fab fragment on the normal cell, obtains the ligands specific-drug conjugate of metastatic carcinoma cell.
Described herein medicine and various analog thereof and derivant generally are to eliminate, kill, disturb and/or reduce the medicine of the pathogenic cell population growth that comprises infectant, cancer, tumor etc.In addition, can be used for the medicine of described conjugate herein and various analog thereof and derivant and can have the multiple mechanism of action, they include but not limited to alkylating agent, microtubule inhibitor comprise stable and/or make microtubule form unsettled those, kinases (CDK) inhibitor that they comprise 'beta '-tubulin agent, cyclin dependent is CDKN1a, CDKN1b etc. for example; Topoisomerase enzyme inhibitor; Protein synthesis inhibitor; Kinases inhibitor comprises inhibitor such as Ras, Raf, PKC, PI3K; Transcription inhibitor; Antifol, heatshock protein blocker etc.
In another embodiment, set forth Pharmaceutical composition.This Pharmaceutical composition contains pharmaceutically acceptable carrier, excipient and/or the diluent of described drug release conjugate and applied in any combination herein.
In another embodiment, be set forth in the method for eliminating pathogenic cell colony in the host animal with pathogenic cell colony.An illustrative aspects, each member of pathogenic cell colony has part or its analog or the bonded available binding site of derivant with bind receptor, and this binding site is by the unique expression of pathogenic cell, overexpression or preferential the expression.This method comprises and gives the described drug release conjugate of host or the step of described its Pharmaceutical composition herein herein.
The accompanying drawing summary
Figure 1A represents with respect to folic acid (, 1.0), the RA of 9 pairs of folacin receptors of embodiment (■, 0.24).
Figure 1B is illustrated in excessive folic acid and exists (ο) and no excessive folic acid () to exist down, in 9 pairs of KB cells of embodiment
3The bonded activity of H-thymidine; The IC of embodiment 9
50Be about 58nM.
Fig. 2 represents that embodiment 11 (■, 0.21) is to the RA of folacin receptor with respect to folic acid (, 1.0).
Fig. 3 is illustrated in excessive folic acid and exists (ο) and no excessive folic acid () to exist down, and embodiment 11 (multiple medicines thing conjugate) is right
3The bonded activity of H-thymidine; The IC of embodiment 11
50=5nM.
Fig. 4 represents that with respect to the excessive folic acid of embodiment 11+ (b), embodiment 11 (a) is to the cell in vitro cytotoxic activity of three kinds of different tumor cell lines (KB, 4T-lc12 and ID8-cl15).
Fig. 5 A represents with respect to untreated control (■), 1 μ mol/kg TIW (6 dosage) () and 2 μ mol/kg TIW (6 dosage)
11 pairs of Balb/c mices of embodiment in the activity of the positive M109 tumor of FR.
Fig. 5 B represents with respect to untreated control (■), 1 μ mol/kg TIW (6 dosage) () and 2 μ mol/kg TIW (6 dosage)
11 pairs of Balb/c mices of embodiment weight do not have influence.
Fig. 6 represents with respect to untreated control (), has () and do not exist under (■) at 40 μ mol/kg EC20 (rhenium complex), and 1 μ mol/kg TIW embodiment 11 continues the activity of 2 weeks (6 dosage) to the positive KB tumor of FR; Independent embodiment 11 shows 5/5 complete reaction; Embodiment 11+EC20 shows 0/5 complete reaction.
Fig. 7 represents with respect to untreated control (), has () and do not exist under (■) at 40 μ mol/kg EC20 (rhenium complex), and 1 μ mol/kg TIW embodiment 11 continues 2 weeks (6 dosage) nu/nu mice weight not to be had influence.
Fig. 8 represents with respect to untreated control (a), has (b) and do not exist under (c) at 40 μ mol/kg EC20 (rhenium complex), and 1 μ mol/kg TIW embodiment 11 continues 2 weeks (6 dosage) to implanting the activity of s.c. people's xenogenesis KB tumor in the nude mouse; Independent embodiment 11 shows 5/5 complete reaction; Embodiment 11+EC20 shows 0/5 complete reaction.
Fig. 9 represents with respect to untreated control (a), has (b) and do not exist under (c) at 40 μ mol/kg EC20 (rhenium complex), and 1 μ mol/kg TIW embodiment 11 continues 2 weeks (6 dosage) nude mouse weight not to be had influence.
Figure 10 represents the mixture of not puting together baseline medicine, ametycin and desacetyl vinblastine list hydrazides (desacetylvinblastine monohydrazide) with respect to 0.5 μ mol/kg TIW (b), 1 μ mol/kg TIW (c) and 2 μ mol/kg TIW (d), and with respect to untreated control (a), 2 μ mol/kg TIW embodiment, 11 (e) are to the activity of folacin receptor positive human tumor in the nude mouse.
Figure 11 represents that with respect to contrast (a) 2 μ mol/kg TIW embodiment 11 continue 2 weeks (e) nude mouse weight not to be had influence.Under all three kinds of dosage (0.5 μ mol/kg TIW (b), 1 μ mol/kgTIW (c), 2 μ mol/kg TIW (d)) of the mixture of not puting together baseline medicine, ametycin and desacetyl vinblastine list hydrazides, weight loss takes place.Before the 20th day, stop high dose (d).
Figure 12 represents that with respect to contrast (a) 2 μ mol/kg TIW embodiment 11 continued for 2 weeks, to three kinds of size 250mm in the nu/nu mice
3(b), 500mm
3(c) and 750mm
3The activity of big KB tumor (d).
Figure 13 represents with respect to contrast (a), with respect to the conjugate of single medicine ametycin (b), desacetyl vinblastine list hydrazides (c) only, or the mixture of these two kinds of single medicine conjugates (d), the activity of embodiment 11 (e).
Figure 14 represents that with respect to contrast (a) 2 μ mol/kg TIW embodiment, 11 (b) continue the negative 4T1 tumor of folacin receptor non-activity in 2 all treatments B ablb/c mices.Data show that because of not having folacin receptor on these tumors, embodiment 11 (b) does not have any effect to tumor with respect to contrast (a) among Figure 14.
Figure 15 represents in the positive KB cell of 12 couples of FR of embodiment
3The bonded activity of H-thymidine.
Describe in detail
Set forth the ligand conjugates of medicine and analog thereof and derivative herein. These conjugates comprise the part in conjunction with cell receptor, and this part comprises the part of cell surface receptor, but they comprise that with targeted cells two or more medicines of pathogenic cell are covalently bound. Described conjugate also can comprise the multivalence joint that part is connected with medicine herein.
The medicine of having set forth bind receptor discharges conjugate, and it comprises part (B), multivalence joint (L) and two or more medicines or drug analogue or the medicaments derivative (D) of bind receptorn, wherein n is more than or equal to 2. Described medicine discharges in the conjugate in this article, the part of bind receptor (B) and two or more medicines (D)nSeparately by the independent hetero atom joint (l that selectsH) and multivalence joint (L) combination. Multivalence joint (L) comprises the combination of one or more intervals base joint, hetero atom joint and separable joint and any order thereof.
In one embodiment, set forth the medicine release conjugate of bind receptor. Medicine discharges conjugate and comprises part for example part, two or more medicines or its analog or derivative and the optional multivalence joint of cell surface receptor, and this medicine discharges conjugate and usually can be represented by following formula
(B)-(L)-(D)
n
Wherein (B) represents the part of bind receptor; (D) representative is by medicine or its analog or the derivative of the part targeted cells of bind receptor; (L) represent the multivalence joint, n is the integer greater than 1. Multivalence joint (L) can comprise a plurality of mutually covalently bound joints. For example, multivalence joint (L) can comprise one or more interval base joint (ls) and/or separable joint (lr), they are separately by one or more hetero atom joint (lH) be connected with other joint, part and medicine. Can select these different joints, and can arrange in any order, consist of multivalence joint (L).
For example, can consist of multivalence joint (L) by one or more following divalence joints:
-(L)-
-(l
r)
c-
-(l
s)
a-
-(l
s)
a-(l
r)
c-
-(l
r)
c-(l
s)
a-
-(l
H)
b-(l
r)
c-
-(l
r)
c-(l
H)
b-
-(l
H)
d-(l
r)
c-(l
H)
e-
-(l
s)
a-(l
H)
b-(l
r)
c-
-(l
r)
c-(l
H)
b-(l
s)
a-
-(l
H)
d-(l
s)
a-(l
r)
c-(l
H)
e-
-(l
H)
d-(l
r)
c-(l
s)
a-(l
H)
e-
-(l
H)
d-(l
s)
a-(l
H)
b-(l
r)
c-(l
H)
e-
-(l
H)
d-(l
r)
c-(l
H)
b-(l
s)
a-(l
H)
e-
-(l
s)
a-(l
r)
c-(l
H)
b-
-[(l
s)
a-(l
H)
b]
d-(l
r)
c-(l
H)
e-
Wherein a, b, c, d and e be integer for example at the integer of about 4 scopes of 0-, (ls)、(l
H) and (lr) be respectively interval base joint, separable joint, hetero atom joint. Can be used for making up other illustrative examples of the divalence joint of described multivalence joint herein at u.s. patent application serial number 10/765, be described among 336 (also can find at U.S. Patent Application Publication No. US 2005/0002942 A1) and the PCT international publication number WO2006/012527, described document by reference integral body is attached to herein.
Be appreciated that the multivalence joint can be connected with two or more medicines by the part of various structures configuration in connection with acceptor, includes but not limited to following exemplary general formula:
B-L
1-D
1-L
2-D
2 B-L
1-D
1-L
2-D
2-L
3-D
3
D
1-L
1-B-L
2-D
2 D
1-L
1-B-L
2-D
2-L
3-D
3
Wherein B is the part of bind receptor, (L1)、(L
2) and (L3) multivalence of respectively doing for oneself joint, basic, separable and/or hetero atom joint consists of (D by one or more intervals1)、D
2And D3Respectively do for oneself medicine or its analog or derivative. Comprise other medicines or its analog or derivative, other joint and (B), other modification of (L) and other configuration of (D) arranging separately is also included within herein.
In a kind of modification, be included in herein in conjunction with the part of more than one acceptors that described medicine discharges in the conjugate, include but not limited to following exemplary general formula:
Wherein each B is the part of bind receptor, (L1)、(L
2) and (L3) multivalence of respectively doing for oneself joint, basic, separable and/or hetero atom joint consists of (D by one or more intervals1)、D
2And D3Respectively do for oneself medicine or its analog or derivative. Comprise other medicines or its analog or derivative, other joint and (B), other modification of (L) and other configuration of (D) arranging separately is also included within herein. In a kind of modification, the ligand binding same receptor of bind receptor, in another kind of modification, the ligand binding of bind receptor is isoacceptor not. Shown in following formula, be appreciated that to be included in herein by more than one multivalence joint described medicine discharges in the conjugate. Be appreciated that in one aspect, select the number of joint according to the configuration of the part of bind receptor and medicine.
For example, assemble in the exemplary of a part of mode that forms multivalence joint or multivalence joint at its center tap covalency, hetero atom joint, interval base joint and separable joint are joined together to form following formula multivalence group:
Wherein this formula also can be expressed as
(l wherein
s)
1Be tripeptides Asp-Asp-Asp, (l
s)
2Be Cys, (l
r)
1Be S-S, (l
s)
3Be CH
2CH
2, (l
H)
1Be O, (l
r)
2Be C (O) NHNH, (l
s)
4Be ω-Lys, (l
s)
5Be C (O) CH
2CH
2, (l
r)
3Be S-S, (l
s)
6Be CH
2CH
2
Can be used for the part of the cell surface receptor of described conjugate herein and include but not limited to vitamin and the albumen or its analog or the derivant that exist with the bonded other parts of vitamin receptor, transport protein or other and the bonded surface of vitamin specificity; Peptide part by the storehouse Screening and Identification; Tumor cell-specific peptide; Tumor cell-specificity is fit; Tumor cell-specificity sugar; Tumor cell-specific monoclonal or polyclonal antibody; For example the lead Fab fragment of antibody of EphA2 of the Fab of antibody or scFv (being the strand variable domain) fragment, or on the metastatic carcinoma cell specific expressed or unique other albumen that reaches; The little organic molecule that obtains by combinatorial libraries; Somatomedin is EGF, FGF, insulin and insulin like growth factor for example; And homeopeptide; Somatostatin and analog thereof; Transferrins; Protein-lipid complex; Bile salt; Select albumen; Steroid hormone; The peptide that contains Arg-Gly-Asp; Biostearin; Various gala agglutinins; δ-opioid receptor part; The CCK A receptors ligand; The ligands specific of angiotensin AT1 or AT2 receptor; Peroxisome proliferation-activated receptors λ part; Beta-Lactam antibiotic is penicillin for example; The little organic molecule that comprises antimicrobial drug; With with preferential bonded other molecule of receptor-specific of expressing on tumor cell surface or infectious organisms; Antimicrobial drug and according to the other medicines of the binding pocket of the suitable special receptor of receptor or other cell surface protein crystal structure design; Tumor antigen or on tumor cell surface the part of preferential other molecule of expressing, or the fragment of any of these molecule.The extracellular epi-position that the example that can play the tumor-specific antigen of part-medicine or its analog or the effect of derivant conjugate binding site comprises Ephrin family protein film is EphA2 for example.In normal cell, EphA2 expresses and is limited to cell-cell contact, but in metastatic cancer cell, EphA2 is distributed in whole cell surface.Therefore, the EphA2 on the metastatic cell can combine with the Fab fragment of the antibody of for example puting together with medicine or its analog or derivant, and albumen can not combine with the Fab fragment on the normal cell, obtains the ligands specific-drug conjugate of metastatic carcinoma cell.
In one embodiment, the part of bind receptor is a vitamin or in conjunction with its analog or the derivant of vitamin receptor, for example can be in conjunction with vitamin and the analog and the derivant of vitamin receptor.
Can comprise carnitine, inositol, thioctic acid, Vitamin B6, ascorbic acid, nicotinic acid, pantothenic acid, folic acid, riboflavin, thiamine, biotin, vitamin B according to the vitamin of described method and chemical compound use herein
12, vitamin A, D, E and K, other relevant vitamin molecules, its analog and derivant and combination thereof.The analog of these vitamin and their bind receptor and derivant constitute exemplary target to entity, and these entities can be by described multivalence joint (L) and medical compounds or their analog or derivant coupling herein, preparation drug release conjugate.
An illustrative aspects, vitamin can be folic acid, folacin or another kind of molecule in conjunction with folacin receptor.Spendable exemplary folacin comprises folinic acid, pteroyl polyglutamic acid, pteroic acid and other amino acid derivativges thereof; With for example tetrahydrochysene pterin, dihydrofoilic acid, tetrahydrofolic acid and their denitrification assorted (deaza) and two remove aza analogues in conjunction with the pteridine of folacin receptor.Term " denitrification is assorted " and " two denitrifications are assorted " analog are meant the art-recognized analog that replaces the carbon atom of one or two nitrogen-atoms in the natural folic acid structure that has.For example, go aza analogues to comprise that the 1-denitrification is assorted, the 3-denitrification is assorted, the 5-denitrification is assorted, the 8-denitrification is assorted and 10-removes aza analogues.Two go aza analogues for example to comprise 1, and 5-two denitrifications are assorted, 5, and 10-two denitrifications are assorted, 8, and 10-two denitrifications are assorted and 5, and 8-two removes aza analogues.Aforementioned folacin is called " folic acid " as usual, reacts they and the bonded ability of folacin receptor.Other analog in conjunction with folacin receptor comprises aminopterin; Methotrexate; N
10-methopterin; 2-deaminizes-hydroxyl folic acid; Remove aza analogues for example assorted methopterin of 1-denitrification or the assorted methopterin of 3-denitrification; With 3 ', 5 '-two chloro-4-amino-4-'-deoxy-ns
10-methyl pteroylglutamic acid (dichioromethotrexate).Can combine the antibody that other the suitable part that causes receptor-mediated drug release conjugate endocytosis transhipment comprises folacin receptor with folacin receptor.Therefore, an illustrative aspects, the available and compound Herba Catharanthi Rosei chemical compound of antibody folacin receptor causes this complex transmembrane transport.
The exemplary of vitamin D 3-analogies and/or derivant also comprises the analog and the derivant of biotin, chemical compound of biological example born of the same parents element, biotinylsulfoxide, oxybiotin and other and biotin receptors bind etc.Can recognize that the analog of described other vitamin and derivant are also included within herein herein.
The Any shape of described conjugate should be included in herein, and these shapes are by the part and the decision of various multivalence joint ways of connecting of its Chinese medicine, bind receptor.In one aspect, the general three shape of described conjugate is linear herein.In yet another aspect, the general three shape of described conjugate is " Y " or " T " shape herein.In yet another aspect, the general three shape of described conjugate is " X " shape or cross herein.At another
In a described herein drug release conjugate exemplary, the multivalence joint comprises at least one separable joint (l
r).In another exemplary of described in this article drug release conjugate, the multivalence joint comprises at least two separable joint (l
r)
2In another illustrative aspects, multivalence joint (L) comprises at least one separable joint (l
r), this joint is not the separable joint of disulphide.In another illustrative aspects, multivalence joint (L) has at least two separable joint (l
r)
2, one of them separable joint is not the separable joint of disulphide.Can recognize that when more than one separable joint was included in the multivalence joint, those separable joints can be adjacent.Be further appreciated that when two separable joints in the multivalence joint were adjacent, these two separable joints can be worked in coordination with and be caused drug release.
Term used herein " separable joint " is called the joint that can rupture again, is meant the joint that comprises at least one key that can rupture (for example pH instability, unstable, the easy oxidation of acid or the unsettled key of enzyme) under physiological condition.It should be understood that this type of physiological condition that causes bond fission comprises the standard chemical hydrolysis that for example takes place under physiological pH, or because compartmentation for example has in the endosome of the pH lower than kytoplasm pH to organelle due to.
Be appreciated that described scissionable bond can be in the one or both ends of appointing of separable joint herein, two adjacent atoms in the separable joint connected and/or with other joint or (B) and/or (D) connect.In the situation that scissionable bond connects two adjacent atoms in the separable joint, after the bond fission, separable joint breaking is two or more fragments.Perhaps, be positioned at separable joint and another part for example hetero atom joint, basic joint, another separable joint, medicine or its analog or derivant at interval at scissionable bond, or the situation between vitamin or its analog or the derivant, after the bond fission, separable joint separates with other parts.
Can by for example on the scissionable bond or near change replace and for example to comprise the α side chain at the disulfide bond ortho position of can rupturing; Increase in the part with hydrolyzable silicon-oxygen key substituent hydrophobicity on the silicon; Make and form hydrolyzable ketal or acetal alkoxyl homologization partly etc., the unstability of regulating scissionable bond.
The exemplary mechanism of described herein bivalence joint breaking comprises following 1,4 and 1,6 splitting mechanism
Wherein X is endogenous or exogenous nucleophile, glutathion or biological reductant etc., any is vitamin or its analog or derivant among Z or the Z ', or medicine or its analog or derivant, or the vitamin or the drug moiety that are connected with the other parts of multivalence joint.Although be appreciated that the above splitting mechanism of describing is a cooperation mechanism, can adopt the discrete step of any number, realize multivalence joint final fracture be shown in end-product.For example can recognize that also can make bond fission by the carbamate moiety acid catalysis is eliminated, it can obtain anchimeric assistance by the stabilisation that β sulfur aryl described in the above example or disulphide provide.In those modification of this embodiment, separable joint is a carbamate moiety.Perhaps, can cause cracking, cause fracture, form mercaptides by the nucleophilic attack disulfide group.This mercaptides can form corresponding thiirane in intermolecular displacement carbonic acid or carbamic acid part.Containing under the multivalence joint situation of benzyl, after the exemplary fracture of disulfide bond, the phenyl mercaptan salt that obtains can further rupture by the intermediate that forms resonance stabilized, discharges carbonic acid or carbamic acid part.In any case therein, can by can with the relevant any mechanism of chemistry, metabolism, physiology or biotic factor that exists, the separable character of the described herein exemplary multivalence joint of realization.
The exemplary mechanism of other of separable joint bond fission comprises the complementary cracking of following oxygen:
Wherein Z is vitamin or its analog or derivant, or medicine or its analog or derivant, or the vitamin or the drug moiety of respectively doing for oneself and being connected with the other parts of multivalence joint, for example comprise the medicine or the vitamin part of one or more intervals base joint, hetero atom joint and/or other separable joint.In this embodiment, the carbamate acid catalysis is eliminated, caused CO
2Discharge the benzyl cation that formation can be caught by water or any other Lewis alkali with the nitrogen moiety that is connected with Z.
After another exemplary mechanism relates to the bond fission of multivalence joint, separable, at interval base and hetero atom joint by the functional group's chemistry that discharges auxiliary other bond fission or cracking is called the anchimeric assistance fracture again or cracked mode is arranged.The exemplary of this multivalence joint or its part comprises the chemical compound with following formula:
Wherein X is for example nitrogen, oxygen or a sulfur of hetero atom, n is selected from 0,1,2 and 3 integer, R is hydrogen or substituent group, this substituent group comprises can inductivity or the alkoxyl etc. for example of the substituent group by aromatic ring resonance stabilized positive charge, any is vitamin or its analog or derivant among Z or the Z ', or medicine or its analog or derivant, or the vitamin or the drug moiety that are connected with the other parts of multivalence joint.Can recognize that other substituent group can be present on aromatic ring, benzyl carbon, carbamate nitrogen, alkanoic acid or the methylene bridge, includes but not limited to hydroxyl, alkyl, alkoxyl, alkylthio group, halogen etc.Complementary cracking can comprise the mechanism that relates to benzyl intermediate, benzyne intermediate, lactone cyclisation, oxygen intermediate, β-elimination etc.Can recognize that also the cracking, anchimeric assistance mechanism also can be impelled separable joint initial cracking behind separable joint breaking.
In this embodiment, but the hydroxyl alkane acid of cyclisation impels the methylene bridge cracking by for example oxonium ion, and impels cracking after the bond fission of bond fission or separable joint.Perhaps, acid catalysis oxonium ion-assistance cracking methylene bridge can be started this exemplary multivalence joint or its segmental cracking cascade.Perhaps, but the acid-catalyzed hydrolysis carbamate can impel the hydroxyl alkane acid β of cyclisation to eliminate, and impels the methylene bridge fracture by for example oxonium ion.Can recognize that the bond fission under described in this article metabolism, physiology or the cell condition or cracked other chemism can cause this cracking cascade.Can recognize that the bond fission under described in this article metabolism, physiology or the cell condition or cracked other chemism can cause this cracking cascade.
In one embodiment, described herein multivalence joint is a following formula: compound
Wherein n is the integer that is selected from 1-about 4; R
aAnd R
bIndependently be selected from hydrogen and alkyl separately, comprise for example C of optional branching of low alkyl group
1-C
4Alkyl; Or R
aAnd R
bBe combined together to form carbocyclic ring with the carbon atom that links to each other; R is the acyl group of the optional alkyl that replaces, optional replacement or the nitrogen-protecting group group of suitably selecting; And (
*) represent the junction point of the other parts of medicine, vitamin, developer, diagnostic reagent, other multivalence joint or conjugate.
In another embodiment, described herein multivalence joint comprises following formula: compound
Wherein m is the integer that is selected from 1-about 4; R is the acyl group of the optional alkyl that replaces, optional replacement or the nitrogen-protecting group group of suitably selecting; And (
*) represent the junction point of the other parts of medicine, vitamin, developer, diagnostic reagent, other multivalence joint or conjugate.
In another embodiment, described herein multivalence joint comprises following formula: compound
Wherein m is the integer that is selected from 1-about 4; R is the acyl group of the optional alkyl that replaces, optional replacement or the nitrogen-protecting group group of suitably selecting; And (
*) represent the junction point of the other parts of medicine, vitamin, developer, diagnostic reagent, other multivalence joint or conjugate.
In another embodiment, separable, at interval base and hetero atom joint can be arranged by auxiliary other bond fission of functional group's chemistry or the cracked mode that discharge after the bond fission of multivalence joint, auxiliary other bond fission of chemistry or cracking are called anchimeric assistance fracture or cracking again.The exemplary of this multivalence joint or its part comprises the chemical compound with following formula:
Wherein X is for example nitrogen, oxygen or a sulfur of hetero atom, and n is selected from 0,1,2 and 3 integer, and R is hydrogen or substituent group, and comprising can inductivity or the alkoxyl etc. for example of the substituent group by aromatic ring resonance stabilized positive charge, symbol (
*) representative forms the junction point of other interval base, hetero atom or the separable joint of multivalence joint, or the junction point of medicine or its analog or derivant or vitamin or its analog or derivant.Can recognize that other substituent group can be present on aromatic ring, benzyl carbon, alkanoic acid or the methylene bridge, includes but not limited to hydroxyl, alkyl, alkoxyl, alkylthio group, halogen etc.Complementary cracking can comprise the mechanism that relates to benzyl intermediate, benzyne intermediate, lactone cyclisation, oxygen intermediate, β-elimination etc.Can recognize that also the cracking, anchimeric assistance mechanism also can be impelled separable joint initial cracking behind separable joint breaking.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and the separable joint that is joined together to form the 3-of multivalence shown in following formula thiosuccimide-1-base alkoxyl methoxy base:
Wherein n is the integer of 1-6, and alkyl is optional to be substituted, and methyl is optional to be replaced by other alkyl or the optional aryl that replaces, their each free independent R group representatives of selecting.Symbol (
*) represent the multivalence linker fragment and the junction point of the other parts of described conjugate herein.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and the separable joint that is joined together to form the 3-of multivalence shown in following formula thiosuccimide-1-base alkyl-carbonyl:
Wherein n is the integer of 1-6, and alkyl is optional to be substituted.Symbol (
*) represent the multivalence linker fragment and the junction point of the other parts of described conjugate herein.In another embodiment; the multivalence joint comprises hetero atom joint, the basic joint in interval and the separable joint that is joined together to form multivalence 3-sulfane base sulfonyl alkyl (dibasic silicyl) oxygen base, and wherein dibasic silicyl is replaced by alkyl and/or the optional aryl that replaces.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and the separable joint that is joined together to form the dithio of multivalence shown in following formula alkyl-carbonyl hydrazide group or multivalence 3-thiosuccimide-1-base alkyl-carbonyl hydrazides:
Wherein n is the integer of 1-6, and alkyl is optional to be substituted, and (B) of hydrazides and multivalence joint (L), (D) or other parts form hydrazone.Symbol (
*) represent the multivalence linker fragment and the junction point of the other parts of described conjugate herein.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and the separable joint that is joined together to form the 3-of multivalence shown in following formula thiosuccimide-1-base alkoxyl alkoxyl alkylidene radical:
Wherein each n is for independently being selected from the integer of 1-6, and each alkyl is independently selected and chosen wantonly by for example alkyl or the optional aryl that replaces and replaces, wherein (B) of alkylidene radical and multivalence joint (L), (D) or other parts formation hydrazone.Symbol (
*) represent the multivalence linker fragment and the junction point of the other parts of described conjugate herein.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint, and they are joined together to form multivalence 3-sulfenyl or 3-dithio aryl-alkoxy carbonyl; 3-sulfenyl or 3-dithio aryl-alkyl amino carbonyl; Multivalence 3-sulfenyl or 3-dithio alkoxy carbonyl, or multivalence 3-sulfenyl or 3-dithio alkyl amino-carbonyl, wherein (B) of alkyl-carbonyl and multivalence joint (L), (D) or other parts form carbonic ester, carbamate or urea.For example, alkyl is an ethyl.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint, they are joined together to form multivalence 3-dithio alkyl amino, and (B), (D) or other parts wherein amino and multivalence joint (L) form vinylogy (vinylogous) amide.For example, alkyl is an ethyl.
In another embodiment, the multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint, and they are joined together to form multivalence 1-alkoxyl cycloalkylidene oxygen base, multivalence alkylidene amino carbonyl (dicarboxyl arlydene) carboxylic acid ester groups, multivalence 3-dithio alkoxy carbonyl, multivalence 3-dithio alkoxy carbonyl hydrazide group, multivalence.
In another embodiment, the multivalence joint comprises at least one by the basic joint in amino acids formed peptide interval.In one aspect, peptide comprises natural amino acid and stereoisomer thereof.In yet another aspect, peptide is only formed by natural amino acid and stereoisomer thereof.
At interval other illustrative examples of base and separable joint is shown in table 1 and 2, wherein (
*) represent the junction point of another joint and vinca alkaloids or its analog or derivant, or with the junction point of the part of bind receptor.
Table 1: interval base and the hetero atom joint and the combination thereof of imagination
Table 2: separable and the hetero atom joint and the combination thereof of imagination
The medicine of any kind of all can be included in the described herein drug release conjugate.In an exemplary, according to one or more pathogenic cell colonies activity are selected medicine.In one aspect, those pathogenic cells are the cancerous cell that comprise solid tumor.
In another exemplary, according to the medicine that the active selection of one or more pathogenic cell colonies is had specific function mechanism.The exemplary mechanism of action comprises alkylating agent; The microtubule inhibitor, they comprise makes microtubule form stable and/or unsettled those inhibitor, comprises the 'beta '-tubulin agent; The kinases of cyclin dependent (CDK) inhibitor; Topoisomerase enzyme inhibitor; Protein synthesis inhibitor; Kinases inhibitor comprises inhibitor such as Ras, Raf, PKC, PI3K; Transcription inhibitor; Antifol; Heatshock protein blocker etc.
Exemplary alkylating agent includes but not limited to ametycin BI etc.The kinases of exemplary cyclin dependent (CDK) inhibitor comprises but is not limited to CYC202, seliciclib, R-roscovitine, AGM-1470 etc.Exemplary topoisomerase enzyme inhibitor includes but not limited to doxorubicin, other anthracycline etc.Exemplary protein synthesis inhibitor includes but not limited to bruceantin etc.The exemplary kinases inhibitor that comprises inhibitor such as Ras, Raf, PKC, PI3K includes but not limited to L-779,450, R115777 etc.Exemplary transcription inhibitor includes but not limited to α-amanitine (amanatin), D actinomycin D etc.Exemplary antifol includes but not limited to methotrexate etc.Exemplary heatshock protein blocker includes but not limited to Ge Erde toxin etc.
Comprise that the exemplary microtubule inhibitor that makes microtubule form stable and/or unsettled those inhibitor comprises 'beta '-tubulin agent, microtubule toxin etc.Include but not limited to and the bonded inhibitor of Herba Catharanthi Rosei binding site with the exemplary microtubule toxin of the receptors bind of selecting, for example the not new A (phomopsin A) in arenastatin, dolastatin, halichondrin B (halichondrin B), maytansine, side, rhizomycin, rice aspergillin (ustiloxin), vinblastine, vincristine etc.; With the bonded stabilizing agent of paclitaxel binding site for example discodermalide, Epothilones (epothilone), taxol, paclitaxel etc.; With the bonded inhibitor of colchicine binding site for example colchicine, combretastatin, curacin A, podophyllotoxin, steganacine etc.; With with bonded other inhibitor in undefined site for example from beads algal rim peptide (cryptophycin), tubulysins etc.
In a described herein drug release conjugate embodiment, at least a in these medicines is microtubule inhibitor or its analog or derivant.In another embodiment, at least a in these medicines is the DNA alkylating agent.In another embodiment, at least a in these medicines is the DNA alkylating agent, at least a other drug in these medicines is the microtubule inhibitor, described herein alkaloid comprises all members in Herba Catharanthi Rosei indole-indoline alkaloid family, such as but not limited to vindesine, vinblastine, vincristine, vinblastine, vindoline, leurosine, vinorelbine, a miaow card, sibutramine, toltrazuril, Changchun flower acid etc. and analog and derivant.
In another described herein drug release conjugate embodiment, at least a in these medicines is P-glycoprotein (PGP) inhibitor.In another embodiment, be included at least a PGP of the being inhibitor in these medicines on the described drug release conjugate herein, at least a other drug that is included in these medicines on the drug release conjugate is the PGP substrate.For example, according to this back embodiment, the PGP substrate is the DNA alkylating agent.About this embodiment, can recognize that with PGP inhibitor and the pairing of PGP substrate, for example the DNA alkylating agent includes but not limited to that it is the overall efficiency of the medicine of PGP substrate in addition that any mitomycin such as ametycin, Mitomycin A etc. can improve.In the described in this article separable conjugate, behind the endocytosis, PGP inhibitor medicaments and PGP substrate medicine all discharge in cell.By this mode, the PGP inhibitor medicaments can improve the overall effectiveness and/or the activity of PGP substrate medicine.In addition, the PGP inhibitor can reduce PGP expresses, and this reduces one or more medicines that are included on the described multiple medicines thing conjugate again herein and flows out from pathogenic cell.Can recognize that mitomycin or its analog or derivant for example the ametycin following adjustment that can be used as PGP inhibitor or PGP are manipulated.Can recognize that also vinca alkaloids or its analog or derivant for example vinblastine analog and derivant can be the PGP substrates, can be by the PGP inhibitor or adjust down, prevent that it from flowing out from pathogenic cell.
In another described herein drug release conjugate embodiment, at least a in these medicines is vinca alkaloids or its analog or derivant.Described herein vinca alkaloids comprises all members in Herba Catharanthi Rosei indole-indoline alkaloid family, such as but not limited to vindesine, vinblastine, vincristine, vinblastine, vindoline, leurosine, vinorelbine, a miaow card, sibutramine, toltrazuril, Changchun flower acid (vinblastinoic acid) etc. and analog and derivant.
As mentioning herein, the spendable Herba Catharanthi Rosei medicine in the described herein conjugate comprises in Herba Catharanthi Rosei indole-indoline alkaloid family all members for example vindesine, vinblastine, vincristine, vinblastine, vindoline, leurosine, vinorelbine, a miaow card, sibutramine, toltrazuril, Changchun flower acid etc. and analog and derivant.For example, this type of analog and derivant comprise U.S. Patent number 4,203, the carbonyl azide of 3-described in 898 thing (carboxazides); U.S. Patent number 4,166, the N of the deacetylate of 4-described in 810 vinblastine-3-carbonyl hydrazides
2-alkyl and other derivant; Leurosine hydrazides described in the Neuss et al.Tetrahedron Lett.783 (1968); Hydrazide derivatives described in the Barnett et al.J.Med.Chem.21:88 (1978); U.S. Patent number 3,392,173 and 3,387, the ester derivant of C-4 described in 001; The dicarboxylic acid derivatives that oxidation obtains described in the Langone et al.Anal.Biochem.95:214 (1979); With Herba Catharanthi Rosei hydrazides described in EP 0 247 792 A2.In each above-mentioned patent and the publication disclosure of the synthetic route of relevant preparation Herba Catharanthi Rosei chemical compound and reaction condition by reference integral body be attached to herein.
In an exemplary, the Herba Catharanthi Rosei medicine is a following formula: compound
Wherein:
R
1And R
2In one be H, another is an ethyl, and R
3Be H, or R
1Be ethyl, R
2And R
3Be combined together to form-O-;
R
4, R
7And R
8Independently be selected from H, alkyl and acyl group separately
R
5And R
6Independently be selected from alkyl separately;
R
9For-the NHNHR group, wherein R is H, alkyl or acyl group;
R
10Be H or acyl group; And
R
11Be ethyl.
In one aspect, the Herba Catharanthi Rosei medicine is following formula chemical compound, wherein R
4And R
8H respectively does for oneself; And R
5, R
6, R
9And R
10The methyl of respectively doing for oneself.
In another embodiment, set forth the drug release conjugate of bind receptor, this conjugate contains part, multivalence joint (L), vinca alkaloids medicine or its analog or derivant and another kind of medicine or its analog or the derivant of bind receptor, and wherein the part of bind receptor, vinca alkaloids and other medicines are separately by hetero atom joint (l
H) combine with multivalence joint (L).Multivalence joint (L) comprises the combination of one or more intervals base joint, hetero atom joint and separable joint and any order thereof.
In another embodiment, be included at least a aclamycin of being or its analog or derivant in these medicines on the described drug release conjugate herein.Aclamycins and analog thereof and derivant may be PGP efflux pump substrates.In one aspect, at least a other drug that is included in these medicines on the described herein drug release conjugate is for example mitomycin or its analog or a derivant of DNA alkylating agent.
In another embodiment, be included at least a DNA of being synthetic inhibitor or its analog or derivant in these medicines on the described drug release conjugate herein.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is that spindle forms inhibitor or its analog or derivant.In one aspect, be included at least a DNA of being synthetic inhibitor or its analog or derivant in these medicines on the described drug release conjugate herein, at least a other drug that is included in these medicines on the described herein drug release conjugate is that spindle forms inhibitor or its analog or derivant.
In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is microtubule stabilizer or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is microtubule synthetic inhibitor or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is to destroy microtubule stabilizer or its analog or derivant.
In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is inducer of apoptosis or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is taxol or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is antifol or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is methotrexate or its analog or derivant.On the one hand, be included at least a in these medicines on the described drug release conjugate herein and be for example methotrexate of antifol or its analog or derivant, at least a other drug that is included in these medicines on the described herein drug release conjugate is taxol or its analog or derivant.
In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is folic acid or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is human epidermal growth factor acceptor-2 (HER-2) inhibitor or its analog or derivant.In another embodiment, being included at least a in these medicines on the described drug release conjugate herein is for example cisplatin etc. of radiolabeled chemotherapeutics.In one aspect, be included at least a in these medicines on the described drug release conjugate herein and be for example methotrexate of antifol or its analog or derivant, at least a other drug that is included in these medicines on the described herein drug release conjugate is folic acid or its analog or derivant.On the other hand, being included at least a in these medicines on the described drug release conjugate herein is taxol or its analog or derivant, and at least a other drug that is included in these medicines on the described herein drug release conjugate is HER-2 inhibitor or its analog or derivant.On the other hand, being included at least a in these medicines on the described drug release conjugate herein is taxol or its analog or derivant; At least a other drug that is included in these medicines on the described herein drug release conjugate is for example cisplatin etc. of radiolabeled chemotherapeutics; And at least a other drug that is included in these medicines on the described herein drug release conjugate is HER-2 inhibitor or its analog or derivant.
Can be by the described herein drug release conjugate of conventional synthetic method preparation.Select synthetic method according to selected hetero atom joint and the functional group and the separable joint that are present on the basic joint in interval.Generally speaking, at Richard C.Larock, " Comprehensive OrganicTransformations, a guide to functional group preparations; " VCHPublishers, Inc.New York (1989) and Theodora E.Greene ﹠amp; Peter G.M.Wuts, " Protective Groups ion Organic Synthesis, " the 2nd edition, John Wiley ﹠amp; Sons has among the Inc.New York (1991) the crucial elaboration that forms reaction is arranged, and described document integral body by reference is attached to herein.Other synthetic route and reaction condition are set forth in U.S. Patent Application Publication No. US 2005/0002942 A1.
For example, the described herein drug release conjugate of useable linear and convergent synthesis path of preparing.The exemplary intermediate that can be used for this type of route comprises the intermediate that contains the multivalence joint, this multivalence joint comprises coupling group at each end, this coupling group is fit to the part of bind receptor or its analog or derivant and vinca alkaloids or its analog or derivant covalently bound.Other the exemplary intermediate that can be used for this type of route comprises the part that contains the bind receptor that is connected with the multivalence joint or the intermediate of its analog or derivant, and the multivalence joint contains coupling group.Other the exemplary intermediate that can be used for this type of route comprises and contains the vinca alkaloids that is connected with the multivalence joint or the intermediate of its analog or derivant that the multivalence joint contains coupling group.In arbitrary situation, coupling group can be nucleopilic reagent, electrophilic reagent or its precursor.
In an exemplary synthetic intermediate embodiment, coupling group is the Michael receptor, the multivalence joint comprise (O) NHN=that has formula-C ,-NHC (O) NHN=or-CH
2The separable joint of C (O) NHN=.An illustrative aspects, coupling group and multivalence joint are combined together to form the derivant of following formula: compound or its protection:
Wherein (D) can form the analog or the derivant of described hydrazone herein for vinca alkaloids or its, and n is an integer for example 1,2,3 or 4.Another illustrative aspects of the drug release conjugate intermediate of described bind receptor in this article, second joint is covalently bound by the alkyl hydrosulfide nucleophile and the following formula chemical compound that are included on second joint.In another illustrative aspects, the part of bind receptor or its analog or derivant are covalently bound by the alkyl hydrosulfide nucleophile and the following formula chemical compound that are included on this part.
In another exemplary, coupling group is for example nitrogen, oxygen or a sulfur of hetero atom, and the multivalence joint comprises one or more hetero atom joints and one or more interval base joint, and they are covalently bound with coupling group with the part of bind receptor.An illustrative aspects, described herein intermediate comprises the derivant of following formula: compound or its protection:
Wherein X is oxygen, nitrogen or sulfur, and m is integer for example 1,2 or 3, wherein (B), l
sAnd l
HDefine same this paper.At an illustrative aspects, l
HFor-NH-, m is 1.At another illustrative aspects, l
HFor-NH-, m is 1, X is-S-.
In another exemplary, described herein intermediate comprises the derivant of following formula: compound or its protection:
Wherein Y is a for example electron-withdrawing substituent of H or substituent group, includes but not limited to nitro, cyano group, halogen, alkyl sulphonyl, carboxylic acid derivates etc., wherein (B) and l
sDefine same this paper.
At another herein in the exemplary of described intermediate, coupling group is the Michael receptor, the multivalence joint comprises one or more hetero atom joints and one or more interval base joint, and they are covalently bound with coupling group with the part of bind receptor.An illustrative aspects, coupling group and multivalence joint are joined together to form the derivant of following formula: compound or its protection:
Wherein X is oxygen, nitrogen or sulfur, and m and n are the independent integer of selecting for example 1,2 or 3, wherein (B), l
sAnd l
HDefine same this paper.In another illustrative aspects, vinca alkaloids or its analog or derivant are covalently bound by the alkyl hydrosulfide nucleophile and the following formula chemical compound that are included on the vinca alkaloids.
In another illustrative aspects, described intermediate comprises the derivant of following formula: compound or its protection:
Wherein AA is one or more aminoacid; for example they are selected from natural amino acid or its stereoisomer; X is nitrogen, oxygen or sulfur; Y is a for example electron-withdrawing substituent of hydrogen or substituent group; include but not limited to nitro, cyano group, halogen, alkyl sulphonyl, carboxylic acid derivates etc.; n and m are the independent integers of selecting for example 1,2 or 3, and p is an integer for example 1,2,3,4 or 5.
AA also can for example have any aminoacid of following general formula for any other aminoacid:
-N(R)-(CR′R″)
t-C(O)-
Wherein R is hydrogen, alkyl, acyl group or suitable nitrogen-protecting group, R ' and R, and " be hydrogen or substituent group, when occurring, they are independently selected separately at every turn, and t is an integer for example 1,2,3,4 or 5.For example, R ' and/or R " independent corresponding to but the hydrogen that is not limited to exist on the natural amino acid or the side chain derivant of methyl, benzyl, hydroxymethyl, sulfidomethyl, carboxyl, carboxyl methyl, guanidine radicals propyl group etc. and derivant and protection for example.Above-mentioned formula comprises all stereoisomer modification.For example, aminoacid can be selected from agedoite, aspartic acid, cysteine, glutamic acid, lysine, glutamine, arginine, serine, ornithine, threonine etc.Another illustrative aspects of the described in this article drug release conjugate intermediate in conjunction with vitamin receptor, medicine or its analog or derivant comprise the alkyl hydrosulfide nucleophile.
Available conventional synthetic route prepares each above intermediate.Other synthetic route and reaction condition have argumentation in U.S. Patent Application Publication No. US 2005/0002942 A1 and PCT international publication number WO2006/012527.
The aforementioned exemplary embodiment is used to illustrate described the present invention herein, should not be construed as or be considered as by any way described the present invention herein to be limited.For example, the chemical compound of being represented usually by following exemplary vitamin-drug conjugate intermediate will be included among described herein the present invention
R wherein
1And R
2Independent separately is for example methyl of hydrogen or alkyl; l
HBe the hetero atom nitrogen etc. of oxygen, sulfur, the optional nitrogen that replaces or optional protection for example.The part of two or more medicines and optional other bind receptor for example folic acid and analog thereof and derivant can be at (l
H) or other functional group of existing for example covalently bound with this exemplary intermediate on amide nitrogen or carbonyl, sour carboxylic acid ester groups or the guanidine amino.
In another embodiment, described folic acid part intermediate has following formula
Wherein m, n and q are the integers that independently is selected from 0-about 8; AA is an aminoacid, R
1Be hydrogen, alkyl or nitrogen-protecting group, medicine optional with (
*) the atom connection.In one aspect, AA is a natural amino acid natural or the non-natural configuration, on the other hand, and fragment (NH-AA-C (O)-)
nIn one or more AA are hydrophilic amino acids.On the other hand, fragment (NH-AA-C (O)-)
nIn one or more AA be Asp and/or Arg.In yet another aspect, integer o is 1 or bigger.On the other hand, integer m is 2 or bigger.The part of medicine or its analog or derivant and optional other joint and other bind receptor can be 2, the segmental free NH side chain of ω-diaminourea alkanoic acid or be connected with the following formula chemical compound on by the terminal carboxylate that wherein free valency is represented.
In another embodiment, described folic acid part intermediate has following formula
Wherein m, n, q and p are for independently being selected from the integer of 0-about 8; AA is an aminoacid, R
1Be hydrogen, alkyl or nitrogen-protecting group, medicine optional with (
*) the atom connection.In one aspect, AA is a natural amino acid natural or the non-natural configuration.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are hydrophilic amino acids.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are Asp and/or Arg.On the other hand, integer o and p are 1 or bigger.On the other hand, integer m is 2 or bigger.The part of medicine or its analog or derivant and optional other joint and other bind receptor can be 2, the segmental free NH side chain of ω-diaminourea alkanoic acid, are connected with the following formula chemical compound on cysteinyl (cyteinyl) sulfydryl of being represented by free valency wherein or terminal carboxylate.
In another embodiment, described folic acid part intermediate has following formula
Wherein m, n, q, p and r are for independently being selected from the integer of 0-about 8; AA is an aminoacid, R
1Be hydrogen, alkyl or nitrogen-protecting group, medicine optional with (
*) the atom connection.In one aspect, AA is a natural amino acid natural or the non-natural configuration.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are hydrophilic amino acids.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are Asp and/or Arg.In yet another aspect, integer o, p and r are 1 or bigger.On the other hand, integer m is 2 or bigger.The part of medicine or its analog or derivant and optional other joint and other bind receptor can be 2, the segmental free NH side chain of ω-diaminourea alkanoic acid, are connected with the following formula chemical compound on by cysteinyl sulfydryl, seryl hydroxyl or terminal carboxylate that wherein free valency is represented.
In another embodiment, described and contained mitomycin as the folic acid part intermediate of one of medicine and have following formula
Wherein m, n and q are for independently being selected from the integer of 0-about 8; AA is an aminoacid.In one aspect, AA is a natural amino acid natural or the non-natural configuration.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are hydrophilic amino acids.On the other hand, (NH-AA-C (O)-)
nOne or more AA in the fragment are Asp and/or Arg.On the other hand, integer o is 1 or bigger.On the other hand, integer m is 2 or bigger.The part of medicine or its analog or derivant and optional other joint and other bind receptor can be 2, segmental other free NH side chains of ω-diaminourea alkanoic acid or be connected with the following formula chemical compound on by the terminal carboxylate that wherein free valency is represented.
In another embodiment, the folic acid part multiple medicines thing conjugate that contains mitomycin and vinca alkaloids and have following formula has been described
In another embodiment, the folic acid part multiple medicines thing conjugate that contains mitomycin, aclamycin and vinca alkaloids and have following formula has been described
In another embodiment, set forth Pharmaceutical composition.This Pharmaceutical composition contains pharmaceutically acceptable carrier, excipient and/or the diluent of described drug release conjugate and applied in any combination herein.
In another embodiment, be set forth in the method for eliminating pathogenic cell colony in the host animal with pathogenic cell colony.An illustrative aspects, the film of pathogenic cell colony has part or its analog or the bonded available binding site of derivant with bind receptor, and this binding site is by the unique expression of pathogenic cell, overexpression or preferential the expression.This method comprises and gives the described drug release conjugate of host or the step of described its Pharmaceutical composition herein herein.
The pathogenic cell colony of available described herein method treatment includes but not limited to for example epithelial cancer of ovary, mammary gland, colon, lung, nose, larynx, brain of cancer; With other kind tumor cell; Infectant, activated huge have a liking for cell, activated mononuclear cell etc.
Described herein drug release conjugate can be used for people's clinical medicine and for animals.Therefore, the host animal with pathogenic cell colony for the treatment of with the drug release conjugate can be the people, or in situation for animals, can be laboratory animal, farming animals, domestic animal or wild animal.Can give host animal described drug release conjugate herein, these animals include but not limited to the people; Laboratory animal such as rodent (for example mice, rat, hamster etc.), rabbit, monkey, chimpanzee; Domestic animal is Canis familiaris L., cat and rabbit for example; Farming animals is cow, horse, pig, sheep, goat for example; With the wild animal of stable breeding for example Bears, panda, lion, tiger, leopard, resemble, zebra, giraffe, gorilla, dolphin and whale.
Described herein drug release conjugate can be used for treating multiple pathogen and pathogenic cell in the host animal." pathogenic cell " used herein expression cancerous cell; Infectant is antibacterial and virus for example; The cell of antibacterial or viral infection; That can cause morbid state activated hugely has a liking for cell; With unique expression, preferentially express or the overexpression ligand receptor for example vitamin receptor or with the pathogenic cell of any other type of vitamin D 3-analogies or the bonded receptor of derivant.Pathogenic cell also can comprise any cell that causes morbid state, and this morbid state causes disease symptoms to alleviate after treating with the drug release conjugate.Pathogenic cell can also be a morbific host cell under some environment, for example host disease is produced graft reaction but under other environment non-pathogenic immune system cell.
Therefore, pathogenic cell colony can be the oncogenicity cancer cell population, and they comprise benign tumor and malignant tumor, or it can be a non-tumorigenic.Cancer cell population can spontaneously produce, or by for example germ line mutation or the somatic mutation generation of host animal of this class process, or it can cause by chemistry, virus or radiation.The present invention can be used for treating this type of cancer for example cancer, sarcoma, lymphoma, Hodgkin, melanoma, mesothelioma, Burkitt lymphoma, nasopharyngeal carcinoma, leukemia and myeloma.Cancer cell population can include but not limited to oral cavity, thyroid, endocrine, skin, stomach, esophagus, larynx, pancreas, colon, bladder, bone, ovary, cervix uteri, uterus, mammary gland, testis, prostate, rectum, kidney, liver and pulmonary carcinoma.
Pathogenic cell colony is in the embodiment of cancer cell population therein, and the effect of administered agents release conjugate is the therapeutic response by tumor epithelial cell reduces or eliminates or the tumor cell proliferation inhibition is measured.In the situation of tumor, elimination can be that the cell in primary tumo(u)r cell or transitional cell or the primary tumo(u)r separation process is eliminated.Also should comprise with drug release conjugate prophylactic treatment, prevent by any Therapeutic Method comprise surgical operation remove tumor, radiotherapy, chemotherapy or biotherapy and remove tumor after tumor recover.Prophylactic treatment can be to carry out initial therapy with the drug release conjugate, for example by multiple dose scheme treatment every day, and/or can be other treatment or the serial therapy of taking after a few days or several months at interval after the initial therapy.Therefore, eliminate above-mentioned any pathogenic cell colony and comprise the number that reduces pathogenic cell, suppress pathogenic cell propagation, prevent the prophylactic treatment that pathogenic cell recovers, or the pathogenic cell treatment that causes disease symptoms to alleviate.
In the situation of eliminating cancerous cell, described herein method can with for example other immunotherapy coupling of the surgical operation of removing tumor, radiotherapy, chemotherapy or biotherapy, immunotherapy includes but not limited to the treatment of monoclonal antibody therapy, immunomodulator, immune effector cell adoptive transfer; Hemopoietic growth factor, cytokine and vaccination treatment.
Described herein method also is applicable to the pathogenic cell colony that causes multiple infectious disease.For example, the present invention is applicable to this type of pathogenic cell colony such as antibacterial; The fungus that comprises yeast; Virus; The cell of viral infection; Mycoplasma and parasite.The infectious biological of available described herein drug release conjugate treatment is any infectious biological that causes the animal morbidity recognized in the art, and they comprise biology for example Gram-negative or gram-positive cocci or bacillus bacterioid.Deformed rod strain for example, the klebsiella spp kind, the Providence strain, the yersinia genus strain, Irving's strain, the enterobacteria kind, sramana's strain, husky thunder strain, the gas bacillus specie, the escherich's bacillus kind, the pseudomonas kind, the shigella dysenteriae kind, the vibrio kind, the Aeromonas kind, crooked strain, the hammer strain, the staphylococcus kind, lactobacillus species, the microsphere strain, the Moraxella kind, bacillus specie, the clostridium kind, the rod-like stem strain, the Erichsen bacillus specie, the microsphere strain, the mycobacterium kind, the Neisser strain, the haemophilus kind, the bacteroid kind, Liszt's strain, the Erysipelothrix kind, the acinetobacter calcoaceticus kind, the Bacillus brucellae kind, the Pasteur strain, the vibrio kind, the Flavobacterium kind, the Fusobacterium kind, the streptobacillus kind, the sheath bacillus specie, the Legionnella kind, the treponema kind, the borrelia vincentii kind, the leptospira kind, the unwrapping wire strain, the Nocardia kind, Li Keshi body kind and any other antibacterial kind that in host animal, causes disease, available described herein drug release conjugate treatment.
Especially pay close attention to be to the drug-fast antibacterial of antibiotic for example antibiotic resistance hammer strain and staphylococcus (Staphlococcus) plant, or to antibiotic sensitive but infect again so that the final antibacterial that produces the drug resistance biology with causing behind the antibiotic therapy.Do not exist under the antibiotic situation, to antibiotic sensitive but with causing the antibacterial that infects consequently final generation drug resistance biology again can use described drug release conjugate treatment herein behind the antibiotic therapy, or available described herein drug release conjugate and the antibiotic therapeutic alliance that is lower than the dosage that normally gives host animal, avoid producing these strains.
Also available described herein drug release conjugate treatment is by the virus disease that causes of DNA and RNA viruses for example.This type of virus includes but not limited to DNA viruses for example papillomavirus, parvovirus, adenovirus, herpesvirus and vaccinia virus; RNA viruses is arenavirus, coronavirus, rhinovirus, breathing syncytial virus, influenza virus, picorna virus, paramyxovirus, reovirus, retrovirus retrovirus, slow virus and baculovirus for example.
The disease that also available described herein drug release conjugate treatment is caused by following reason: comprise any fungus, mycoplasma kind, the parasite of yeast or cause other infectious biologicals of Animal diseases.The example of the fungus of available described herein method and drug release conjugate treatment comprises and grows into mycete or toruloid fungus, comprises and for example causes for example fungus of following disease: tinea, histoplasmosis, blastomycosis, aspergillosis, cryptococcosis, sporotrichosis, coccidioidomycosis, paracoccidioidomycosis, mucormycosis, chromomycosis, dermatomycosis, Protothecosis, fusaridiosis, pityriasis, mycetoma, paracoccidioidomycosis, paeohyphomycosis, pseudallescheriasis, sporotrichosis, trichosporosis, pneumocystis infects and the beads disease.
Also available described herein drug release conjugate treatment parasitic infection, include but not limited to the infection that following parasite causes: cestode belongs to for example cestode, Hymenolepsis; Bothriocephalus and echinococcus kind; Trematodiasis for example Fasciolopsis, Heterophyes(Heterophyes), Metagonimus, Clon, sheet trematodiasis belongs to; Paragonimus and schistosomicide kind; Ascarid is Enterobius, Trichocephalus, Ascaris, Ancylostoma, Necator, Strongyloides, Trichinella, Wuchereria, Bu Shi Filaria, Loa Onchocerca for example; With imperial nematicide kind; Amebicide is Naegleria and Acanthamoeba kind for example; For example Plasmodium, trypanosoma, Leishmania, toxoplasma, Entamoeba, Giardia, Isospora, Cryptosporidium and enterocyte microsporidian belong to (Enterocytozoon) with protozoon.
If these cells are preferentially expressed for example receptor of vitamin or its analog or derivant of ligand receptor, then the pathogenic cell of drug release conjugate guiding can also be the cell with endogenous pathogens, for example the cell of virus, mycoplasma, parasite or bacterial infection.
In one embodiment, when part and receptor, transport protein or other and ligand specificity's knot are incorporated in the protein binding that preferential surface of expressing exists on the pathogenic cell, but drug release conjugate internalization is in the target pathogenic cell.This internalization for example can realize by receptor-mediated endocytosis.If the drug release conjugate contains separable joint, then part and Herba Catharanthi Rosei chemical compound can be at the cell internal disintegrations, and the Herba Catharanthi Rosei chemical compound can work on its cell internal target.
In another exemplary, the part of drug release conjugate can combine with pathogenic cell, is closely associated in Herba Catharanthi Rosei chemical compound and pathogenic cell surface.By separable joint breaking, discharge the Herba Catharanthi Rosei chemical compound then.For example, if separable joint is a disulfide group, then can discharge the Herba Catharanthi Rosei chemical compound by protein disulfide isomerase.The Herba Catharanthi Rosei chemical compound is absorbed by pathogenic cell then, and this pathogenic cell combines with the drug release conjugate of bind receptor, or the Herba Catharanthi Rosei chemical compound can be absorbed by another pathogenic cell adjacent closely with it.Perhaps, can discharge the Herba Catharanthi Rosei chemical compound by the intracellular protein disulfide bond isomerase, wherein separable joint is a disulfide group.Also can for example eliminate the acid-catalyzed hydrolysis that mechanism is carried out, or form mechanism release Herba Catharanthi Rosei chemical compound by oxonium ion generation anchimeric assistance cracking or lactone (lactonium) ion by hydrolysis mechanism by above-mentioned some β.The selection of one or more separable joints will determine to discharge from conjugate the mechanism of Herba Catharanthi Rosei chemical compound.Can recognize that this selection can be determined in advance by drug release conjugate condition to be used.
In another exemplary, when joint does not contain separable joint, the ligand moiety of drug release conjugate can combine with pathogenic cell, makes the lip-deep Herba Catharanthi Rosei targeting compounds of pathogenic cell pathogenic cell, is convenient to attack with bonded other molecule of Herba Catharanthi Rosei chemical compound.Perhaps, in this embodiment, in conjunction with after, can make drug release conjugate internalization in target cell, and ligand moiety and Herba Catharanthi Rosei chemical compound can keep associating in cell, and the Herba Catharanthi Rosei chemical compound is brought into play its effect and is not needed to dissociate with ligand moiety.
In another embodiment again, or the time with the bonded above-mentioned embodiment combination of wherein drug release conjugate and vitamin receptor or another ligand receptor, conjugate can with solubility vitamin receptor in the serum or serum albumin albumin bound for example, cause the length of the circulation time of conjugate, and conjugate is to the height of the unconjugated Herba Catharanthi Rosei chemical compound of the specific activity of pathogenic cell colony than unconjugated Herba Catharanthi Rosei chemical compound.
Part for example the binding site of vitamin can comprise can with the receptor of the bonded part of ligand specificity, wherein by pathogenic cell colony only table reach, overexpression or preferential expressed receptor or other albumen.The lower receptor of concentration that the albumen that is existed by the unique expression of pathogenic cell, overexpression or preferential surface of expressing does not normally exist in non-pathogenic cell or exists provides selectivity to eliminate the method for pathogenic cell.The affinity of the receptors bind on drug release conjugate and cancerous cell or other type pathogenic cell can be very high.Part self can have high binding affinity, maybe can improve binding affinity by the part that uses chemical modification.
In conjoint therapy, described herein drug release conjugate can give with any other known drug, and no matter whether these known drugs are the other drug of targeting.Exemplary other medicines include but not limited to peptide, oligopeptide, retro-inverso oligopeptide, albumen, wherein at least one non-peptide bond replaces albumen analog, apoprotein, glycoprotein, enzyme, coenzyme, enzyme inhibitor, aminoacid and derivant thereof, receptor and other memebrane protein, antigen and the antibody thereof of peptide bond; Hapten and antibody thereof, hormone, lipid, phosphorus matter, liposome, toxin, antibiotic, analgesic, bronchodilator, beta blocker, antimicrobial drug, depressor; Cardiovascular drug comprises anti-arrhythmic, cardiotonic glycoside, anti-anginal drug, vasodilation; Central nervous system's medicated bag is drawn together analeptic, psychotropic, antimanic drugs and antidepressants; Antiviral agents; Antihistaminic; Anticarcinogen comprises chemotherapeutic, tranquilizer, antidepressants, H-2 antagonist, anticonvulsant, Bendectin, prostaglandin and prostaglandin analogue; Muscle relaxant, anti-inflammatory substance, analeptic, decongestant drug, Bendectin, diuretic, spasmolytic, anti-asthmatic, antiparkinsonism drug, expectorant, antitussive, mucolytic; Mineral and nourishing additive agent.
In another illustrative aspects, other drug is optional certainly can the immunoreactive chemical compound of stimulation of endogenous.Suitable compound includes but not limited to cytokine or immune cell growth factor, for example interleukin-11-18, stem cell factor, basic FGF, EGF, G-CSF, GM-CSF, FLK-2 part, HILDA, MTP-1 α, TGF-α, TGF-β, M-CSF, IFN-α, IFN-β, IFN-γ, solubility CD23, LIF and combination thereof.
Can use the treatment of these immunostimulation factors effectively to make up.In one embodiment, for example treat for example about 0.1MIU/m in every day multiple dose scheme of effective dose
2/ dosage/Ri-Yue 15MIU/m
2The IL-2 of the amount of/dosage/day; About 0.1MIU/m in every day multiple dose scheme for example
2/ dosage/Ri-Yue 7.5MIU/m
2The IFN-α of/dosage/day can use with the drug release conjugate, eliminates in having the host animal of pathogenic cell, reduces or inhibition pathogenic cell (MIU=hundred million international units; m
2=ordinary people's approximation surface area).In another embodiment, can use IL-12 and IFN-α, in another embodiment again, can use IL-15 and IFN-α by the treatment effective dose of above-mentioned interleukin and interferon by the treatment effective dose of above-mentioned interleukin and interferon.In an alternate embodiment, can unite by above-mentioned treatment effective dose and use IL-2, IFN-α or IFN-γ and GM-CSF.Also can use any other effective combination of cytokine, comprise the combination of other interleukin and interferon and colony stimulating factor.
In addition, other medicines can be any medicines as known in the art, they are cell toxicant or cytostatic medicament, the infiltrative medicine of promotion tumor, suppress the medicine of tumor cell proliferation, promote apoptosis medicine, reduce the medicine of the anti-apoptosis activity of target cell, the disease that the infectant that can be used for these medicines treating causes, strengthen endogenous immune response, or can be used for treating the morbid state that the pathogenic cell of any kind causes pathogenic cell.Exemplary suitable other medicines comprise corticoid and corticosteroid, alkylating agent, antiandrogen, antiestrogen, androgen, aclamycin and aclamycin derivant, estrogen; Antimetabolite is cytosine arabinoside, purine analogue, pyrimidine analogue and methotrexate, busulfan, carboplatin, chlorambucil, cisplatin and other platinum compounds for example; Tamoxifen, taxol, paclitaxel, paclitaxel derivant, Tai Suo
Cyclophosphamide, daunomycin, rhizomycin, the T2 toxin, plant alkaloid, prednisone, hydroxyurea, teniposide, mitomycin, discodermolides, non-vinca alkaloids microtubule inhibitor, Epothilones, tubulysin, cyclopropyl benzo [e] indolone, open loop (seco-) cyclopropyl benzo [e] indolone, O-Ac-open loop-cyclopropyl benzo [e] indolone, bleomycin and any other antibiotic, chlormethine, nitroso ureas, colchicine, colchicine derivative, different colchicine, muscoril, the trityl cysteine, Halicondrin B; Dolastatin is dolastatin 10 for example; Amanitine is α-amanitine, camptothecine, irinotecan and other camptothecin derivative for example; Ge Erde toxin and Ge Erde toxin derivant, estramustine, nocodazole, MAP4, NSC-3096, vindesine, vinblastine, vincristine, vinblastine, vindoline, leurosine, vinorelbine, a miaow card, sibutramine, toltrazuril, Changchun flower acid, maytansine and analog and derivant; Gemcitabine; Antiinflammatory and short scorching medicine; Peptide and peptide analogue signal transduction inhibitor; With any other art-recognized medicine or toxin.The other medicines that can be used for conjoint therapy comprise penicillin, cephalosporin, vancomycin, erythromycin, clindamycin, rifampicin, chloromycetin; Aminoglycoside antibiotic, gentamycin, amphotericin B, acyclovir, trifluridine, ganciclovir, zidovudine, amantadine, ribavirin and any other Antimicrobe compound recognized in the art.The analog of any above-mentioned other medicines or derivant also can be used for conjoint therapy.
In another exemplary, provide Pharmaceutical composition.When giving one or more dosage, this Pharmaceutical composition contains the drug release conjugate of the amount of pathogenic cell colony in effective elimination host animal.Preferably by parenteral for example Intradermal, subcutaneous, intramuscular, intraperitoneal, intravenous or the intrathoracic host animal drug release conjugate that gives.Perhaps, can use any effective dose and suitable therapeutic dosage forms to comprise slow release formulation by other effective method orally give host animal drug release conjugate for example medically.The Exemplary excipients that can be used for peroral dosage form includes but not limited to corn starch, gelatin, lactose, magnesium stearate, sodium bicarbonate, cellulose derivative and primojel.
The example of parenteral dosage forms comprises and is dissolved in for example aqueous solution of the activating agent of liquid alcohol, glycol, ester and amide of isotonic saline solution, 5% glucose or other pharmaceutically acceptable liquid-carrier of knowing.Parenteral dosage forms of the present invention can be the soluble lyophilized products form that contains the drug release conjugate of doses.Aspect of embodiment of the present invention, can give the slow release formulation as known in the art of any number, for example can use U.S. Patent number 4,713,249; 5,266,333 and 5,417, biodegradable saccharide matrix described in 982, described document is attached to herein by reference, maybe can use slow-releasing pump (for example osmotic pumps).
Can be before giving the drug release conjugate, give other medicines in the host animal conjoint therapy afterwards or simultaneously, and the other medicines that give can be to contain the part that medicine discharges the same compositions of conjugate, or with the part of drug release conjugate different components.The other medicines of effective dose can be used for any this type of conjoint therapy.
In another illustrative aspects, can use more than one drug release conjugate.For example, can be by the administering drug combinations scheme, with the conjugate that contains different ligands for example folic acid-Herba Catharanthi Rosei and vitamin B
12Combined Treatment host animals such as-Herba Catharanthi Rosei conjugate.In another exemplary, the available conjugate that contains more than one parts for example contains a plurality of folic acid or a plurality of vitamin B
12A conjugate of molecule, or the same conjugate that contains ligand combination for example with folic acid and vitamin B
12The Herba Catharanthi Rosei compound treatment host animal that part is puted together.In addition, independently the drug release conjugate uses the drug release conjugate that contains different types of Herba Catharanthi Rosei chemical compound.
The unit daily dose of drug release conjugate can alter a great deal, and depends on molecular weight, its route of administration and the tissue distribution of host's disease, the morbid state of being treated, conjugate; With treat the probability of the other medicines coupling in for example radiotherapy or the conjoint therapy with other.The effective dose that gives host animal depends on body surface area, weight and the doctor evaluation to patient's disease.Effective dosage ranges can be the about 1mg/kg of for example about 1ng/kg-; The about 500 μ g/kg of about 1 μ g/kg-; The about 100 μ g/kg of about 1 μ g/kg-.
Can use any effective scheme that gives the drug release conjugate.For example, can give the drug release conjugate, or graded is by multiple dose scheme administration every day by single dose.In addition, can use staggered dosage regimen for example 1-3 days/replace the treatment every day week, for defining purpose of the present invention, with this intermittence or every day staggered dosage regimen be considered as being equal to treatment every day, and comprise in the present invention.In an exemplary, handle host animal with drug release conjugate multiple injection, eliminate pathogenic cell colony.In one embodiment, for example in 12-72 hour interim or 48-72 hour interim, to the host repeatedly (preferred about 2 to about 50 times) injectable drug discharge conjugate.Can in a few days or interim several months, give host animal other drug release conjugate injection after initial injection, other drug release conjugate injection can prevent the morbid state recurrence due to the pathogenic cell.
An illustrative aspects, can be used for vitamin or its analog or the derivant of drug release conjugate, comprise with activated huge have a liking for receptor specific expressed on the cell for example with the bonded folacin receptor of folic acid bonded those or its analog or derivant.The conjugate that folic acid connects for example can be used for killing or suppressing to cause the activated huge cytophilic activity of host disease state.When suffering from the activated huge host animal of having a liking for cell-mediated morbid state, this type of huge effect of having a liking for the cell-targeting conjugate be the Herba Catharanthi Rosei chemical compound that will put together activated huge have a liking for concentrate in the cell colony and have a liking for cell and associate with huge, kill and activatedly hugely have a liking for cell or suppress the huge cell function of having a liking for.Eliminating, reduce or making the activated huge effect of having a liking for the cell colony inactivation is to stop or reduce the activated huge morbidity characteristic of having a liking for the cell-mediated morbid state of being treated.Knownly hugely have a liking for cell-mediated exemplary disease and comprise rheumatoid arthritis, ulcerative colitis, Crohn disease, psoriasis, osteomyelitis, multiple sclerosis, atherosclerosis, pulmonary fibrosis, sarcosis, systemic scleroderma, organ-graft refection (GVHD) and chronic inflammatory disease by activated.Usually continue to give the drug release conjugate, until the sx or the elimination of morbid state.
Can by parenteral for example Intradermal, subcutaneous, intramuscular, intraperitoneal or intravenous give the pharmaceutically acceptable carrier of host animal drug release conjugate and applied in any combination, activatedly hugely have a liking for cell or suppress activated huge cytophilic function to kill.Perhaps, can give host animal drug release conjugate, can give effective dose by standard or slow release formulation by other effective ways medically.Can use separately or unite and use described Therapeutic Method with activated huge Therapeutic Method of having a liking for cell-mediated morbid state of treatment that other is generally acknowledged.
Following exemplary is not intended to and should not be considered as restrictive scheme.For example, in each chemical compound that provides in this article, the amino acid whose spatial chemistry that is used to form joint can be chosen wantonly and be selected from natural L configuration or non-natural D configuration.HPLC by NMR, MS and/or UV spectrum and/or explanation characterizes each embodiment; The characteristic signal that explanation is in case of necessity selected.
Method embodiment
Suppress tumor growth in the mice
After intravenous (i.v.) has the animal of tumor, estimate the anti-tumor activity of described chemical compound in subcutaneous Balb/c mice with M 109 tumors herein.In the subcutaneous tissue of right axillary fossa, inoculate 1 * 10
6Tumor (the t of M109 cell
0=60mm
3Mean tumour volume) after about 11 days, time (TIW) on every Wendesdays injects the PBS (contrast) that gives mice (5/ group) 1500nmol/kg drug release conjugate or be equal to dose volume, continuous 3 weeks by i.v..Every interval 2 days or 3 days is with the tumor growth of each processed group of kind of calliper.With equation V=a * b
2/ 2 calculate gross tumor volume, and wherein " a " is length of tumor, and " b " is width, and unit is a millimeter.
Suppress tumor growth in the mice
After intravenous (i.v.) has the animal of tumor, estimate the anti-tumor activity of described chemical compound in subcutaneous nu/nu mice with KB tumor herein.In the subcutaneous tissue of right axillary fossa, inoculate 1 * 10
6Tumor (the t of KB cell
0=50-100mm
3Mean tumour volume) after about 8 days, time (TIW) on every Wendesdays injects the PBS (contrast) that gives mice (5/ group) 5 μ mol/kg drug release conjugates or be equal to dose volume, continuous 3 weeks by i.v..Every interval 2 days or 3 days is with the tumor growth of each processed group of kind of calliper.With equation V=a * b
2/ 2 calculate gross tumor volume, and wherein " a " is length of tumor, and " b " is width, and unit is a millimeter.
It is synthetic to suppress cell DNA
With the described herein chemical compound of cell in vitro poison evaluation of measuring, this mensuration is used to predict that medicine suppresses the ability of folacin receptor-positive KB cell growth.Chemical compound is made up of the folic acid that the chemotherapeutics with separately by described scheme preparation herein is connected.Under 37 ℃,, make the KB cellular exposure reach 7h in the folic acid-drug conjugate of prescribed concentration at least 100 times of excessive folic acid existence or not.With new system culture medium flushing cell once, under 37 ℃, incubation is 72 hours in the new system culture medium then.With
3The H-thymidine is estimated cell viability in conjunction with mensuration.
As shown in Ben Wentu, the dose dependent cytotoxicity can be surveyed, in most applications, and IC
50Value (is bonded to new synthetic DNA
3The H-thymidine reduces by 50% required drug conjugate concentration) in low nanomolar concentration scope.In addition, in the presence of excessive free folic acid, the cytotoxicity of these conjugates lowers, and proves that the observed cytosis of killing is by combining mediation with folacin receptor.
Relative affinity is measured
According to preceding method (Westerhof, G.R., J.H.Schornagel, etal. (1995) Mol.Pharm.48:459-471), measure affinity with respect to the folacin receptor (FR) of folic acid through revising a little.In brief, with intensive being seeded in the 24 porocyte culture plates of the positive KB cell of FR, allow itself and plastics adhere to 18h.In designation hole, the trier of progressive concentration or folic acid exists and not in the presence of, with replenishing 100nM
3The RPMI that does not contain folic acid (FFRPMI) of H-folic acid replaces the incubation culture medium that consumes.At 37 ℃ of following incubation 60min, the PBS with pH 7.4 washes 3 times then with cell.The 1%SDS/PBS of 500 μ l pH 7.4 is added each hole.The collecting cell pyrolysis product contains its adding in each bottle of 5mL flicker mixture then, and counting is measured radioactivity then.The negative control pipe only contains
3H-folic acid/FFRPMI (uncontested dose).The positive control pipe contains final concentration 1mM folic acid, and the CPM that will record in these samples (representing the labelling of non-specific binding) deducts from all samples.Need to prove that it is bonded with the FR on the KB cell that relative affinity is defined as displacement 50%
3The inverse of the molar ratio of H-folic acid required compound, folic acid is decided to be 1 to the relative affinity of FR.
The 4T-1 gross tumor volume is measured
6-7 mice in age in week (female Balb/c kind is) derives from Harlan, Inc., Indianapolis, IN.Before carrying out this test, raise mice with the Harlan feedstuff that does not contain folic acid and amounted to for 3 weeks, and raise mice with the Harlan feedstuff that does not contain folic acid at duration of test.In the subcutaneous tissue of right axillary fossa, inoculate folacin receptor-negative 4T-1 tumor cell (1 * 10
6Cell/animal).Behind about 5 days of the inoculated tumour, when the 4T-1 tumor average volume is～100mm
3The time, time (TIW) on every Wendesdays injects the PBS (contrast) that gives mice (5/ group) 3 μ mol/kg drug release conjugates or be equal to dose volume, continuous 3 weeks by i.v..Every interval 2 days or 3 days is with the tumor growth of each processed group of kind of calliper.With equation V=a * b
2/ 2 calculate gross tumor volume, and wherein " a " is length of tumor, and " b " is width, and unit is a millimeter.
Gravimetry
(PTI) given number of days behind tumor inoculation shown in sample drawing described in the related neoplasms stereometry, is measured the weight change percentage rate of mice in mice (5 mice/groups).
The universal method of preparation folic acid-peptide
By the sequential grammar of polymer support, for example based on the Fmoc-strategy of acid-sensitivity Fmoc-AA-Wang resin, preparation contains the described joint of peptide herein with standard method.For example, by method shown in the flow process 1, synthesized by the aminoacid of Wang resin carrying and the aminoacid of Fmoc protection, preparation contains the peptidyl fragment Pte-Glu-(AA) of folic acid
n-NH (CHR
2) CO
2H (3).
(a) 20% piperidines/DMF; (b) Fmoc-AA-OH, PyBop, DIPEA, DMF; (c) Fmoc-Glu-O-t-Bu or Fmoc-Glu (γ-O-t-Bu)-OH, PyBop, DIPEA, DMF; (d) N
10(TFA)-Pte-OH; PyBop, DIPEA, DMSO; (e) TFAA, (CH
2SH)
2, i-Pr
3SiH; (f) NH
4OH, pH 9-10.
In this article in this exemplary of described method, R
1Be Fmoc, R
2Amino acid side chain for the due care of needs.Wang is a 2-chlorine trityl resin, and DIPEA is a diisopropylethylamine.Use standard coupling method is PyBOP and other method described herein or as known in the art for example, and wherein coupling agent exemplarily as activator, is guaranteed effective coupling.Behind each coupling step, under standard conditions, for example use piperidines, tetrabutylammonium fluoride (TBAF) etc. to handle after, the Fmoc protecting group is removed.By described in the flow process 1, the aminoacid construction unit of use due care is Fmoc-Glu-OtBu, N for example
10-TFA-Pte-OH etc. in step (b), are represented the aminoacid construction unit of due care by Fmoc-AA-OH.Therefore, AA is meant the amino acid starting material of any due care.Be appreciated that term amino acid used herein is meant to have the amine separated by one or more carbon and any reagent of carboxylic acid functional, they comprise natural α and beta amino acids; With these amino acid whose amino acid derivativges and analog.Especially, during described in this article folic acid-peptide is synthetic, also can use serine that the protected aminoacid of side chain for example protects, threonine, cysteine, aspartic acid etc.In addition, during described in this article folic acid-peptide was synthetic, raw material also can comprise γ, δ or longer homologous amino acid.In addition, during described in this article folic acid-peptide is synthetic, raw material also can comprise amino acid analogue for example nor-leucine, 2-amino-3-methylpentanoic acid, Beta-methyl threonine, Beta-methyl cysteine, the β with homologue side chain or another kind of branched structure, Beta-Dimethylcysteine etc.
Will be referred to coupling order (step (a) ﹠amp of aminoacid (AA) of the Fmoc protection of formula Fmoc-AA-OH; (b)) carry out " n " inferior, the preparation solid phase is supported peptide (2), wherein n is an integer, can equal 0-about 100.After last coupling step finishes, remaining Fmoc group is removed (step (a)), make peptide be coupled to glutamate derivatives (step (c)) in order, deprotection and the pteroic acid coupling of protecting with TFA (step (d)).Then, after handling with trifluoroacetic acid, dithioglycol and tri isopropyl silane, make peptide and polymer support dissociate (step (e)).These reaction conditions cause removing simultaneously t-Bu, t-Boc and the Trt protecting group of the amino acid side chain part that can form due care.After alkali treatment, the TFA protecting group is removed (step (f)), obtain containing the peptidyl fragment (3) of folic acid.
Chemical compound embodiment
According to the universal method of method embodiment 7 (flow process 1), in the following order, make the Cys-NH of 4-methoxyl group trityl (the MTT)-protection of Wang resin-bonded
2Reaction: 1) a.Fmoc-As ρ (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 2) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 3) a.Fmoc-Arg (Pbf)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 4) a.Fmoc-As ρ (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 5) a.Fmoc-Glu-OtBu, PyBOP, DIPEA; B.20% piperidines/DMF; 6) N
10-TFA-pteroic acid, PyBOP, DIPEA.With MTT, tBu and Pbf protecting group TFA/H
2O/TIPS/EDT (92.5: 2.5: 2.5: 2.5) remove; NH with pH=9.3
4The OH aqueous solution is removed the TFA protecting group.Select
1H NMR(D
2O)δ(ppm)8.68(s，1H，FAH-7)，7.57(d，2H，J＝8.4Hz，FA H-12 & 16)，6.67(d，2H，J＝9Hz，FA H-13 & 15)，4.40-4.75(m，5H)，4.35(m，2H)，4.16(m，1H)，3.02(m，2H)，2.55-2.95(m，8H)，2.42(m，2H)，2.00-2.30(m，2H)，1.55-1.90(m，2H)，1.48(m，2H)；MS(ESI，m+H
+)1046.
According to the universal method of method embodiment 7 (flow process 1), in the following order, make the Cys-NH of 4-methoxyl group trityl (the MTT)-protection of Wang resin-bonded
2Reaction: 1) a.Fmoc-beta-amino alanine (NH-MTT)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 2) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 3) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 4) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 5) a.Fmoc-Glu-OtBu, PyBOP, DIPEA; B.20% piperidines/DMF; 6) N
10-TFA-pteroic acid, PyBOP, DIPEA.With MTT, tBu and a.2% hydrazine/DMF of TFA protecting group; B.TFA/H
2O/TIPS/EDT (92.5: 2.5: 2.5: 2.5) remove.
Reagent for being used to prepare shown in the following table:
| Reagent | (mmol) | Equivalent | Amount |
| H-Cys (4-methoxyl group trityl)-2-chlorine trityl-resin (loading 0.56mmol/g) | 0.56 | 1 | 1.0g |
| Fmoc-beta-amino alanine (NH-MTT)-OH | 1.12 | 2 | 0.653g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Glu-OtBu | 1.12 | 2 | 0.477g |
| N 10TFA-pteroic acid (being dissolved in 10ml DMSO) | 0.70 | 1.25 | 0.286g |
| DIPEA | 2.24 | 4 | 0.390ml |
| PyBOP | 1.12 | 2 | 0.583g |
The following coupling step that carries out: in the peptide synthesising container, add resin, Freamine, DIPEA and PyBOP.Bubbling feeds argon 1h, usefulness DMF and IPA washing 3 *.DMF solution with 20% piperidines makes Fmoc deprotection 3 * (10min), makes each aminoacid coupling then.Continue coupling until finishing all 6 coupling steps.At last, with the DMF solution washing 3 * (5min) of resin, the TFA protecting group on the pteroic acid is dissociated with 2% hydrazine.
With following reagent peptide analogues is dissociated from resin: 92.5% (50ml) TFA, 2.5% (1.34ml) H
2O, 2.5% (1.34ml) tri isopropyl silane, 2.5% (1.34ml) dithioglycol, dissociation steps operation is as follows: add the 25ml reagent that dissociates, bubbling 1.5h inhales and removes solution (drain), with residue reagent wash 3 *.Be evaporated to about 5mL, use ether sedimentation.Centrifugal, drying.Purification by the following method: post-Waters NovaPak C
18300 * 19mm; Buffer A=10mM ammonium acetate, pH 5; B=CAN; 1%B-20%B, 40 minutes, flow velocity 15ml/min obtained 350mg (64%); HPLC-RT 10.307min, 100% is pure,
1H HMR collection of illustrative plates is consistent with the ownership structure,
MS(ES-)：1624.8，1463.2，1462.3，977.1，976.2，975.1，974.1，486.8，477.8.
According to the universal method of method embodiment 7 (flow process 1), in the following order, make the Cys-NH of 4-methoxyl group trityl (the MTT)-protection of Wang resin-bonded
2Reaction: 1) a.Fmoc-beta-amino alanine (NH-IvDde)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 2) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 3) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 4) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 5) a.Fmoc-Glu-OtBu, PyBOP, DIPEA; B.20% piperidines/DMF; 6) N
10-TFA-pteroic acid, PyBOP, DIPEA.With MTT, tBu and a.2% hydrazine/DMF of TFA protecting group; B.TFA/H
2O/TIPS/EDT (92.5: 2.5: 2.5: 2.5) remove.
Reagent for being used to prepare shown in the following table:
| Reagent | (mmol) | Equivalent | Amount |
| H-Cys (4-methoxyl group trityl)-2-chlorine trityl-resin (loading 0.56mmol/g) | 0.56 | 1 | 1.0g |
| Fmoc-beta-amino alanine (NH-IvDde)-OH | 1.12 | 2 | 0.596g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Asp(OtBu)-OH | 1.12 | 2 | 0.461g |
| Fmoc-Glu-OtBu | 1.12 | 2 | 0.477g |
| N 10TFA-pteroic acid (being dissolved in 10ml DMSO) | 0.70 | 1.25 | 0.286g |
| The Fm-propane thioic acid | 0.70 | 1.25 | 199.08 |
| DIPEA | 2.24 | 4 | 0.390ml |
| PyBOP | 1.12 | 2 | 0.583g |
The following coupling step that carries out: in the peptide synthesising container, add resin, amino acid whose DMF solution, DIPEA and PyBOP.Bubbling feeds argon 1h, with DMF and IPA washing 3 * 10ml.(3 * 10ml) (10min) make each aminoacid coupling then to make the Fmoc deprotection with the DMF solution of 20% piperidines.Continue coupling until finishing 6 coupling steps.When coupling finishes,, TFA protecting group on the pteroic acid and the IvDde protecting group on the beta-amino alanine are dissociated with the DMF solution washing 3 * 10ml (5min) of resin with 2% hydrazine.At last, in DMF, use DIPEA and PyBop free amino and the coupling of Fmoc-propane thioic acid with the beta-amino alanine.Bubbling feeds argon 1h, with DMF and IPA washing 3 * 10ml.With resin dry 30min under argon.
With following reagent peptide analogues is dissociated from resin: 92.5% (50ml) TFA, 2.5% (1.34ml) H
2O, 2.5% (1.34ml) tri isopropyl silane, 2.5% (1.34ml) dithioglycol, dissociation steps operation is as follows: add the 25ml reagent that dissociates, bubbling 1.5h inhales and removes solution, with residue reagent wash 3 *.Be evaporated to about 5mL, use ether sedimentation.Centrifugal, drying.Purification by the following method: post-Waters NovaPak C
18300 * 19mm; Buffer A=10mM ammonium acetate, pH 5; B=CAN; 1%B-20%B, 40 minutes, flow velocity 15ml/min obtained 450mg (65%);
1H HMR collection of illustrative plates is consistent with the ownership structure.
According to the universal method of method embodiment 7 (flow process 1), in the following order, make the Cys-NH of the MTT-protection of Wang resin-bonded
2Reaction: 1) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 2) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 3) a.Fmoc-Arg (Pbf)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 4) a.Fmoc-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 5) a.Fmoc-Glu (γ-OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 6) N
10-TFA-pteroic acid, PyBOP, DIPEA.With MTT, tBu and Pbf protecting group TFA/H
2O/TIPS/EDT (92.5: 2.5: 2.5: 2.5) remove; With the NH of TFA protecting group with pH=9.3
4The OH aqueous solution is removed.
1H NMR collection of illustrative plates is consistent with the ownership structure.
According to the universal method of method embodiment 7 (flow process 1), in the following order, make the D-Cys-NH of the MTT-protection of Wang resin-bonded
2Reaction: 1) a.Fmoc-D-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 2) a.Fmoc-D-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 3) a.Fmoc-D-Arg (Pbf)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 4) a.Fmoc-D-Asp (OtBu)-OH, PyBOP, DIPEA; B.20% piperidines/DMF; 5) a.Fmoc-D-Glu-OtBu, PyBOP, DIPEA; B.20% piperidines/DMF; 6) N
10-TFA-pteroic acid, PyBOP, DIPEA.With MTT, tBu and Pbf protecting group TFA/H
2O/TIPS/EDT (92.5: 2.5: 2.5: 2.5) remove; With the NH of TFA protecting group with pH=9.3
4The OH aqueous solution is removed.
1H NMR collection of illustrative plates is consistent with the ownership structure.
Under 0 ℃, with 2-[(benzotriazole-1-base-(oxygen base ketonic oxygen base)-ethyl disulfide group]-pyridine HCl (601mg) and 378 μ L DIPEA add in the 5ml DCM solution of deacetylation vinblastine hydrazides (668mg) successively.Allow reactant be warming up to room temperature, stirred 3 hours.TLC (15%MeOH/DCM) shows conversion fully.Mixture is through silica gel column chromatography (1: 9MeOH/DCM) purification.Flow point evaporation with merging is dissolved in DCM again with concentrate, uses 10%Na
2CO
3, the salt water washing, dry (MgSO
4), be evaporated to 550mg (80%); HPLC-RT 12.651min, 91% is pure,
1H HMR collection of illustrative plates is consistent with the ownership structure, MS (ESI+): 984.3,983.3,982.4,492.4,491.9,141.8.Other details of this method sees that U.S. Patent Application Publication No. US 2005/0002942 A1 is described, and described document integral body by reference is attached to herein.
By following flow preparation ametycin-ethyl curing (disulfide) propanoic acid
(a) diisopropylethylamine (DIPEA), MeOH.
To amino-ethyl curing propanoic acid (81mg, add in 2mL methanol (MeOH) solution 0.372mmol) DIPEA (0.13mL, 0.746mmol).In this solution, slowly add mitomycin-A (100mg, MeOH 0.286mmol) (3.0mL) solution.With the solution stirring 3h that obtains.TLC (20%MeOH/CHCl
3) the analysis showed that and react completely.The solvent decompression is removed, with residue silicagel column purification.Gradient elution (10%-20%MeOH/CHCl
3/ 0.5%TEA), obtain pure product fractions (110mg, 77%).
Select
1H NMR signal (CDCl
3) δ (ppm)
3.50(d，1H)，3.56(dd，1H)，3.90(t，2H)，4.15(d，1H)，4.25(t，1H)，4.68(dd，1H).
Press the preparation of embodiment 7 methods.
In the polypropylene centrifuge bottle, (82mg 0.084mmol) is dissolved in 5mL water, and bubbling feeds argon 10min with embodiment 2.In another flask, at 0.1N NaHCO
3Bubbling feeds argon 10min in the solution, uses 0.1N NaHCO
3Solution transfers to about 6.9 with the pH of joint solution.With the vinblastine hydrazide derivatives (embodiment 6,91mg, 5mL oxolane (THF) solution 0.092mM) slowly adds in the above solution.Under argon, the settled solution that obtains is stirred 15min-1h.By analytical type HPLC (10mM ammonium acetate, pH=7.0 and acetonitrile) monitoring reaction process.With THF evaporation, aqueous solution is filtered, (the XTerra post is on 19 * 300mM) to be expelled to preparation-HPLC post.With 1mM sodium phosphate pH=7.0 and acetonitrile eluting, obtain containing the pure flow point of product, behind the lyophilization 48h, with its separation (78mg, 50%); C
83H
103N
19O
26S
2Actual mass: 1845.68; MW:1846.95; HPLC-RT 15.113min., 100% is pure,
1The HHMR collection of illustrative plates is consistent with the ownership structure,
MS(ES-)：1846.6，1845.5，933.3，924.2，923.3，922.5，615.6，614.7，525.0.
Figure 21 A and 21B represent the RA of folic acid with respect to embodiment 9; With 9 couples of embodiment
3The bonded effect of H-thymidine; The IC of conjugate
50(58nM); Folic acid and conjugate competition prove the specificity combination of conjugate in conjunction with folacin receptor.According to method embodiment 4 and 3, measure respectively.
Figure 1B is illustrated in (ο) and does not have under the existence of () excessive folic acid, in 9 pairs of KB cells of embodiment
3The bonded activity of H-thymidine; The IC of embodiment 9
50Be about 58nM.
In the polypropylene centrifuge bottle, embodiment 3 (56mg) is dissolved in 7.5mL water, bubbling feeds argon 10min.In another flask, at 0.1N NaHCO
3Bubbling feeds argon 10min in the solution, uses 0.1N NaHCO
3Solution transfers to 6.9 with the pH of embodiment 3 solution.7.5mL oxolane (THF) solution of embodiment 6 (44mg) is slowly added in embodiment 3 solution.Under argon, the settled solution that obtains is stirred 15min-1h.By analytical type HPLC (10mM ammonium acetate, pH=7.0 and acetonitrile) monitoring reaction process.With the THF evaporation, aqueous solution is filtered, through preparation-HPLC purification,, obtain pure flow point with 1mM sodium phosphate pH=7.0 and acetonitrile eluting, they are merged, evaporation transfers to pH 4.0 with the aqueous solution that obtains with 0.1N HCl at ambient temperature.Behind the lyophilization 48h, embodiment 10 is separated (61mg, 64%).
1H HMR collection of illustrative plates conforms to the ownership structure with the LCMS data.
Embodiment 11
Method A: prepare embodiment 11 by the following method:
(a) be DCC, DIPEA, THF; (b) be water/THF, pH 8.5
Under argon, with ametycin-ethyl curing propanoic acid (34.4mg 0.069mmol) is dissolved in anhydrous THF (1mL), add successively then N-hydroxyl succinamide (7.9mg, 0.069mmol), dicyclohexylcarbodiimide (14.2mg, 0.069mmol).(0.024mL 0.138mmol), stirs 3h with the mixture that obtains to add diisopropylethylamine.In the polypropylene centrifuge bottle, (embodiment 9, and 26mg 0.014mmol) is dissolved in 3mL water with the folic acid vinblastine.Use 0.1NNaHCO
3The pH of solution is slowly transferred to 8.5.To add in this folic acid vinblastine solution by the 3mL THF solution of the activatory mitomycin c derivative of described preparation herein.The solution that obtains is stirred 15min-1h under argon, wherein by analytical type HPLC (10mM ammonium acetate and acetonitrile pH=7.0) monitoring reaction process.THF decompression is removed, aqueous solution is filtered, (the X-terra post is on 19 * 300mm) to be expelled to preparation-HPLC post.With 1mM sodium phosphate (pH=7.0) and acetonitrile eluting, obtain pure flow point, with its evaporation, lyophilizing 48h obtains 12mg (50%, based on the raw material that reclaims).
1H NMR and mass spectrometric data are supported respectively to belong to structure shown in Fig. 9 and 10.C
103H
127N
23O
32S
4Actual mass 2325.79; MW 2327.51.HPLC-RT20.054min., 99% is pure,
1H HMR collection of illustrative plates is consistent with the ownership structure,
MS(ES+)：
1552.5，116.0，1165.3，1164.3，1148.4，744.9，746.4，745.6.
Method B: at room temperature, under argon, dry DMF (4.5mL) is expelled to embodiment 10 (103mg, 48.7 μ mol) and embodiment 8 (NO
2-PySSCH
2CH
2-MMC, 33.4mg, 1.25 equivalents) mixture in.Inject DIPEA (84.9 μ L, 10 equivalents) and DBU (72.9 μ L, 10 equivalents) successively to the solution that obtains.At room temperature, under argon, reactant mixture was stirred 20 minutes, then under agitation, transfer in the ether (50mL).The suspension that obtains is centrifugal, will precipitate with ether (15mL * 2) washing, be dissolved in phosphate buffer (pH 6.8 for 9mL, 1.25mM) then, through preparation HPLC (post: Waters XTerra RP18,7 μ m, 19 * 300mm; Mobile phase: the A=1.25mM phosphate buffer, pH 6.8, the B=acetonitrile; Method: 10%B-40%B, 25min, flow velocity 25mL/min) purification.11.72-13.88 minute flow point is collected, and lyophilization obtains the 105.8mg material, and this material contains 99.2mg and 6.6mg phosphate.
Method C: prepare embodiment 11 by the following method, yield 34%:
(a) NHS, DCC-resin, DIPEA, THF; (b) embodiment 7, DIPEA, DMSO
Fig. 2 represents the RA of folic acid (, 1.0) with respect to embodiment 11 (■, 0.21).The data representation conjugate has high RA to folacin receptor among Fig. 2.Measure by method embodiment 4.
Figure 1B and 3 represents that respectively embodiment 9 (having a kind of medicine) and 11 (having a pair of medicine) are right
3The bonded effect of H-thymidine, the IC of embodiment 9 conjugates
50(58nM) and the IC of embodiment 11
50(5nM).Data among Figure 1B and 3 show that also folic acid combines folacin receptor with the conjugate competition, proves the binding specificity of conjugate.Measure by method embodiment 3.In addition, to confirm that specific activity to folacin receptor only has an a kind of embodiment 9 of medicine high more than 10 times for the embodiment 11 with two kinds of medicines.
Fig. 4 represents the cell in vitro cytotoxic activity of embodiment 11 (a) to three kinds of different tumor cell lines (KB, 4T-lc12 and ID8-cl15).In addition, the cytotoxic activity that Fig. 4 is illustrated in embodiment 11 under the existence of excessive folic acid (b) descends, and shows that 11 pairs of folacin receptors of embodiment work.
Fig. 5 A and 5B represent two kinds of various dose (1 μ mol/kg ﹠amp; 2 μ mol/kg) M109 pulmonary carcinoma tumor active and to the influence (with Balb/c mouse assay M109 gross tumor volume) of Balb/c mice weight in 11 pairs of Balb/c mices of embodiment.Measure according to method embodiment 1 and 6 respectively.Embodiment 11 suppresses the solid tumor growth, but the weight to mice does not almost have influence under two kinds of dosage.Even after stopping administration on the 20th day, higher dosage (2 μ mol/kg) is the strong inhibition tumor growth also in addition.Vertical line is corresponding to last administration day (the 20th day).Test with 5 animals, under higher dosage 2 μ mol/kg, comprehensive reaction all appears in all 5 animals.
Fig. 6 represents to compare with contrast (a), has (b) and does not exist under (c) situation at 40 μ mol/kg EC20 (rhenium complex), and embodiment 11 is when 1 μ mol/kg TIW, and continuous 2 weeks are to the activity of FR positive KB tumor.Vertical dotted line is represented last administration day.These figure show that embodiment 11 suppresses the solid tumor growth, and its inhibitory action is stoped (competitiveness) by the EC20 rhenium complex.In addition, these figure show, with respect to contrast, handle the not weight of appreciable impact experimental animal with embodiment 11.EC20 (rhenium complex) is the following formula: compound with the rhenium chelating
The preparation method of EC20 has argumentation in U.S. Patent Application Publication No. US 2004/0033195 A1, its synthetic method is described and is attached to herein by reference.Measure according to method embodiment 2.With the combining of folacin receptor in, EC20 is the competitor of embodiment 11, the result shows that embodiment 11 has specific effect.
When Fig. 8 is illustrated in 40 μ mol/kg EC20 (rhenium complex) and adds (b) and do not add (c), embodiment 11 when 1 μ mol/kg TIW to nude mouse in the activity of people's xenogenesis KB tumor of the positive s.c. transplanting of folacin receptor.Data among Fig. 8 show that by (b) and (c) contrast, embodiment 11 suppresses the solid tumor growth, and its inhibitory action is stoped (the competitive prevention) by the EC20 rhenium complex.In addition, data also show among Fig. 8, with respect to contrast (a), and the weight of appreciable impact test nude mouse animal model not after handling with embodiment 11.
Figure 10 represents with respect to untreated control (a), with respect to 0.5 μ mol/kg TIW (b); The mixture of not puting together baseline medicine, ametycin and deacetylate vinblastine list hydrazides of 1 μ mol/kg TIW (c) and 2 μ mol/kg TIW (d), 2 μ mol/kg TIW embodiment, 11 (e) are to the activity of folacin receptor positive human tumor in the nude mouse.Data representation embodiment 11 among Figure 10 suppresses the solid tumor growth, and complete reaction all appears in all 5 experimental animals.On the contrary, the mixture process with 0.5 μ mol/kg TIW (b) or 1 μ mol/kg TIW (c) baseline medicine complete reaction all do not occur 5 arbitrary merely hitting of experimental animal.As represent the shown in Figure 11 of baseline medicine and the effect of 11 pairs of experimental animal weight of embodiment, before the 20th day, because of toxicity occurring, termination gives the mixture (d) of 2 μ mol/kg TIW baseline medicines of high dose.
Figure 11 shows that during handling with respect to contrast (a), embodiment 11 (e) is the weight of appreciable impact experimental animal not.Data among Figure 11 show; compare with embodiment 11; with with the contrast (a) compare; the mixture of not puting together baseline medicine, ametycin and deacetylate vinblastine list hydrazides (0.5 μ mol/kg TIW (b) and 1 μ mol/kg TIW (c)) long time treatment with low dosage causes experimental animal weight significantly to descend.In addition, the mixture that high dose (2 μ mol/kg TIW (d)) is not puted together the baseline medicine causes weight decline maximum, causes this test to be ended.
Described herein chemical compound can be used for treating tumor big or that set up.For example, 11 pairs of big tumors of embodiment are effective.Figure 12 represents that 2 μ mol/kg TIW 11,2 weeks of embodiment are to big (250mm
3, 500mm
3And 750mm
3) activity of s.c.KB tumor.When tumor reaches as corresponding to one of three target volumes shown in the vertical arrows of gross tumor volume the time, begin to handle with embodiment 11.Data among Figure 12 show that embodiment 11 suppresses big tumor growth, and complete reaction appears in experimental animal.
Figure 13 represents with respect to contrast (a); The mixture (d) of each independent single medicine conjugate, mitomycin C conjugate (b) and deacetylate vinblastine list hydrazides conjugate (c) or these single medicine conjugates, continuous two weeks of 1 μ mol/kg TIW embodiment, 11 (e) are handled the activity to the s.c.KB tumor of setting up.Par by 1 μ mol/kg TIW gives each drug conjugate, and continuous two weeks handle.This figure shows that the performance of embodiment 11 is better than the mixture of arbitrary single medicine conjugate or two kinds of single medicine conjugates.Be unexpectedly, the performance of the mixture of single medicine conjugate significantly is not better than the single medicine conjugate of administration respectively, and with respect to contrast, the dosage regimen of any single medicine conjugate does not all have significance,statistical.Only embodiment 11 chemical compounds are better than contrast.In addition, these Notes of Key Datas have the Herba Catharanthi Rosei medicine and the mitomycin medicine has synergism to single conjugate.
Embodiment 12-14
According to described method and condition preparation herein, comprise above method described in the embodiment 11.The thiosulfonates that preparation needs or other details of pyridine radicals dithio-activatory vinblastine and maleimide-activatory vinblastine derivant have argumentation in U.S. Patent Application Publication No. US 2005/0002942 A1.Other details of the E09 that preparation needs has argumentation in U.S. Patent Application Publication No. US 2005/0165227 A1, the disclosure of described document is attached to herein by reference.
Embodiment 12
Figure 15 represents that 12 couples of 100nM embodiment are in the positive KB cell of FR
3The H-thymidine is in conjunction with the activity with respect to the burst length.Measure by method embodiment 3.
Embodiment 14
Claims (52)
1. the drug release conjugate of a bind receptor, described conjugate comprises:
(a) part of bind receptor;
(b) multivalence joint; With
(c) two or more medicines or its analog or derivant;
The part of wherein said bind receptor is covalently bound with described multivalence joint;
Described two or more medicines or its analog or derivant and described multivalence joint are covalently bound; And
Described multivalence joint comprises one or more following elements that are selected from: basic joint, separable joint and hetero atom joint and combination thereof at interval.
2. drug release conjugate in conjunction with vitamin receptor, described drug release conjugate has following formula:
(B)-(L)-(D)
n
Wherein (L) is selected from (l
s)
a, (l
H)
b(l
r)
cAnd combination; (l wherein
r) be separable joint, (l
s) be the basic joint in interval, and (l
H) be the hetero atom joint;
(B) be part in conjunction with vitamin receptor, n is the integer greater than 1, and when occurring, (D) is independent medicine or its analog or the derivant of selecting at every turn; And
A, b and c independently are 0,1,2,3 or 4 separately.
3. drug release conjugate in conjunction with vitamin receptor, described conjugate has following formula:
(B)-(L)-(D)
n
Wherein (L) is selected from (l
s)
a(l
H)
bAnd combination, wherein (l
s) be the basic joint in interval, and (l
H) be the hetero atom joint; And a and b independently are 0,1,2,3 or 4 separately; And
(B) be part in conjunction with vitamin receptor, n is the integer greater than 1, and when occurring, (D) is the independent medicine of selecting at every turn.
4. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises at least one separable joint, described separable joint is not a disulphide.
5. claim 1,2 or 3 drug release conjugate, wherein said hetero atom joint is nitrogen, oxygen or sulphur atom, or is selected from formula-NHR
1NHR
2-,-SO-,-S (O)
2-and-NR
3O-, wherein R
1, R
2And R
3Independently be selected from aryl alkyl, the heteroaryl of aryl, the replacement of hydrogen, alkyl, aryl, aryl alkyl, replacement, the heteroaryl and the alkoxyalkyl of replacement separately.
6. claim 1; 2 or 3 drug release conjugate; wherein said interval base joint is selected from carbonyl; thiocarbonyl; alkylidene; cycloalkylidene; the alkylidene cycloalkyl; the alkylidene carbonyl; the cycloalkylidene carbonyl; the carbonylic alkyl carbonyl; 1-alkylidene butanimide-3-base; 1-(carbonylic alkyl) butanimide-3-base; the alkylidene sulfinyl; the sulfonyl alkyl; alkylidene sulfinyl alkyl; alkylidene sulfonyl alkyl; carbonyl tetrahydrochysene-2H-pyranose; the carbonyl tetrahydrofuran base; 1-(carbonyl tetrahydrochysene-2H-pyranose) butanimide-3-base and 1-(carbonyl tetrahydrofuran base) butanimide-3-base, wherein said interval base joint is optional separately by one or more X
1Substituent group replaces;
Each X wherein
1Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately.
7. the drug release conjugate of claim 6, wherein said hetero atom joint is a nitrogen, and wherein said X
1Substituent group is combined together to form heterocycle with the interval base joint that described hetero atom joint is connected with them.
8. the drug release conjugate of claim 7, wherein said heterocycle is selected from pyrrolidine, piperidines, oxazolidine, isoxazole alkyl, Thiazolidine, isothiazolidine, ketopyrrolidine, piperidones, oxazolidone, isoxazole alkyl ketone, thiazolidone, isothiazolidine ketone and butanimide.
9. claim 1; 2 or 3 drug release conjugate; wherein said separable joint is selected from methylene; 1-alkoxyl alkylidene; 1-alkoxyl cycloalkylidene; 1-alkoxyl alkylidene carbonyl; 1-alkoxyl cycloalkylidene carbonyl; the carbonyl aryl carbonyl; carbonyl (carboxyl aryl) carbonyl; carbonyl (dicarboxyl aryl) carbonyl; halo alkylidene carbonyl; alkylidene (dialkyl group silicyl); alkylidene (alkylaryl silicyl); alkylidene (diaryl silicyl); (dialkyl group silicyl) aryl; (alkylaryl silicyl) aryl; (diaryl silicyl) aryl; oxygen base ketonic oxygen base; oxygen base ketonic oxygen base alkyl; the sulfonyl alkyl; the imino group alkylidene radical; carbonyl alkylidene radical imino group; imino group ring alkylidene radical; carbonyl ring alkylidene radical imino group; the alkylidene sulfonyl; the alkylidene sulfenyl; alkylidene aryl sulfenyl and carbonyl alkylthio group, wherein said separable joint are optional separately by one or more X
2Substituent group replaces;
Each X wherein
2Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately.
10. the drug release conjugate of claim 9, wherein said hetero atom joint is a nitrogen, and wherein said X
2Substituent group is combined together to form heterocycle with the separable joint that described hetero atom joint is connected with them.
11. the drug release conjugate of claim 10, wherein said heterocycle are selected from pyrrolidine, piperidines, oxazolidine, isoxazole alkyl, Thiazolidine, isothiazolidine, ketopyrrolidine, piperidones, oxazolidone, isoxazole alkyl ketone, thiazolidone, isothiazolidine ketone and butanimide.
12. claim 1,2 or 3 drug release conjugate; wherein said hetero atom joint is a nitrogen; and wherein said separable joint and described hetero atom joint are combined together to form divalent group; described divalent group comprises alkylidene aziridine-1-base, alkylidene carbonyl aziridine-1-base, carbonylic alkyl aziridine-1-base, alkylidene sulfinyl aziridine-1-base, sulfinyl alkyl aziridine-1-base, sulfonyl alkyl aziridine-1-base or alkylidene sulfonyl aziridine-1-base, and wherein said separable joint is optional separately by one or more X
2Substituent group replaces;
Each X wherein
2Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately.
13. the drug release conjugate of claim 12; wherein said hetero atom joint is a nitrogen; and described separable joint and described hetero atom joint are combined together to form divalent group, and described divalent group comprises alkylidene aziridine-1-base, carbonylic alkyl aziridine-1-base, sulfinyl alkyl aziridine-1-base or sulfonyl alkyl aziridine-1-base.
14. the drug release conjugate of claim 13, wherein said interval base joint is selected from carbonyl, thiocarbonyl, alkylidene carbonyl, cycloalkylidene carbonyl, carbonylic alkyl carbonyl and 1-(carbonylic alkyl) butanimide-3-base, and wherein said interval base joint is optional separately by one or more X
1Substituent group replaces;
Each X wherein
1Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately;
And wherein said interval base joint and described separable joint are connected to form the aziridine amide.
15. claim 1,2 or 3 drug release conjugate, at least a in wherein said medicine or its analog or the derivant are to be selected from following vinca alkaloids: vinblastine, deacetylation vinblastine, vindesine, sulfo-vindesine and analog and derivant.
16. claim 1,2 or 3 drug release conjugate, wherein said medicine is medicine or its medicaments derivative that contains the nitrogen-atoms of two key bondings, wherein said separable joint is selected from alkylidene carbonylamino and 1-(alkylidene carbonylamino) butanimide-3-base, and wherein said separable joint and described medicine nitrogen are connected to form hydrazone.
17. claim 1,2 or 3 drug release conjugate, wherein said medicine is medicine or its medicaments derivative that contains sulphur atom, described separable joint is selected from alkylidene sulfenyl and carbonyl alkylthio group, and wherein said separable joint and described medicine sulfur are connected to form disulphide.
18. the drug release conjugate of claim 6, the part of wherein said bind receptor is the folic acid that contains nitrogen, and described interval base joint is selected from alkylidene carbonyl, cycloalkylidene carbonyl, carbonylic alkyl carbonyl and 1-(carbonylic alkyl) butanimide-3-base, and wherein said interval base joint and described folic acid nitrogen are connected to form acid imide or alkylamide.
19. the drug release conjugate of claim 18, wherein each X
1Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, carboxyl, carboxyalkyl, guanidine alkylation, the R of alkyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately.
20. the drug release conjugate of claim 6, wherein said hetero atom joint is a nitrogen, and described interval base joint is selected from alkylidene carbonyl, cycloalkylidene carbonyl, carbonylic alkyl carbonyl and 1-(carbonylic alkyl) butanimide-3-base, and wherein said interval base joint is optional separately by one or more X
1Substituent group replaces, and described interval base joint and described nitrogen are connected to form amide.
21. the drug release conjugate of claim 6, wherein said hetero atom joint is a sulfur, and described interval base joint is selected from alkylidene and cycloalkylidene, and wherein said interval base joint is optional separately by carboxyl substituted, and described interval base joint and described sulfur are connected to form mercaptan.
22. the drug release conjugate of claim 6, wherein said hetero atom joint is a sulfur, and described interval base joint is selected from 1-alkylidene butanimide-3-base and 1-(carbonylic alkyl) butanimide-3-base, and described interval base joint and described sulfur are connected to form the basic mercaptan of butanimide-3-.
23. the drug release conjugate of claim 9, wherein said hetero atom joint is an oxygen, and described separable joint is selected from methylene, 1-alkoxyl alkylidene, 1-alkoxyl cycloalkylidene, 1-alkoxyl alkylidene carbonyl and 1-alkoxyl cycloalkylidene carbonyl, and wherein said separable joint is optional separately by one or more X
2Substituent group replaces, and described separable joint and described oxygen are connected to form acetal or ketal.
24. the drug release conjugate of claim 9, wherein said hetero atom joint is an oxygen, and described separable joint is methylene, and the aryl that wherein said methylene is optionally substituted replaces, and described separable joint and described oxygen are connected to form acetal or ketal.
25. the drug release conjugate of claim 9, wherein said medicine is medicine or its medicaments derivative that contains nitrogen-atoms, described hetero atom joint is a nitrogen, and described separable joint is selected from carbonyl aryl carbonyl, carbonyl (carboxyl aryl) carbonyl, carbonyl (dicarboxyl aryl) carbonyl, and described separable joint and described hetero atom joint nitrogen are connected to form amide, and also are connected to form amide with described medicine nitrogen.
26. the drug release conjugate of claim 9, wherein said medicine is medicine or its medicaments derivative that contains oxygen atom, described hetero atom joint is a nitrogen, and described separable joint is selected from carbonyl aryl carbonyl, carbonyl (carboxyl aryl) carbonyl, carbonyl (dicarboxyl aryl) carbonyl, and described separable joint and described hetero atom joint nitrogen are connected to form amide, and also are connected to form ester with described medicine oxygen.
27. the drug release conjugate of claim 9, wherein said hetero atom joint is a nitrogen, and described separable joint is selected from imino group alkylidene radical, carbonyl alkylidene radical imino group, imino group ring alkylidene radical and carbonyl ring alkylidene radical imino group, and wherein said separable joint is optional separately by one or more X
2Substituent group replaces, and described separable joint and described nitrogen are connected to form hydrazone.
28. the drug release conjugate of claim 9, wherein said hetero atom joint is an oxygen, and described separable joint is selected from alkylidene (dialkyl group silicyl), alkylidene (alkylaryl silicyl), alkylidene (diaryl silicyl), (dialkyl group silicyl) aryl, (alkylaryl silicyl) aryl and (diaryl silicyl) aryl, and wherein said separable joint is optional separately by one or more X
2Substituent group replaces, and described separable joint and described oxygen are connected to form silanol.
29. claim 1,2 or 3 drug release conjugate, wherein said medicine is medicine or its medicaments derivative that contains nitrogen-atoms, and described separable joint is halo alkylidene carbonyl, and described halo alkylidene carbonyl is optional by one or more substituent X
2Replace;
Each X wherein
2Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately; And
The nitrogen of described separable joint and described medicine or medicaments derivative is connected to form amide.
30. claim 1,2 or 3 drug release conjugate, wherein said medicine is medicine or its medicaments derivative that contains oxygen atom, and described separable joint is alkylene oxide group carbonyl or halo alkylidene carbonyl, and described alkylene oxide group carbonyl or halo alkylidene carbonyl are optional by one or more X
2Substituent group replaces;
Each X wherein
2Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately;
And the oxygen of described separable joint and described medicine or medicaments derivative is connected to form carbonic ester or ester.
31. claim 1,2 or 3 drug release conjugate, wherein said hetero atom joint is an oxygen, and described interval base joint is optional by one or more X
11-alkylidene butanimide-3-base that substituent group replaces, and described separable joint is selected from methylene, 1-alkoxyl alkylidene, 1-alkoxyl cycloalkylidene, 1-alkoxyl alkylidene carbonyl, 1-alkoxyl cycloalkylidene carbonyl, and wherein said separable joint is optional separately by one or more X
2Substituent group replaces;
Each X wherein
2Substituent group independently is selected from aryl, aryl alkyl, the aryl alkyl of replacement, heteroaryl, the heteroaryl of replacement, carboxyl, carboxyalkyl, alkyl carboxylic acid ester group, alkyl chain alkanoic acid ester group, guanidine alkylation, the R of alkyl, alkoxyl, alkoxyalkyl, hydroxyl, hydroxy alkyl, amino, aminoalkyl, alkyl amino alkyl, dialkyl aminoalkyl, halogen, haloalkyl, mercaptoalkyl, alkylthio alkyl, aryl, replacement
4-carbonyl, R
5-carbonylic alkyl, R
6-acyl amino and R
7-acylaminoalkyl, wherein R
4And R
5Independently be selected from aminoacid, amino acid derivativges and peptide separately, and R wherein
6And R
7Independently be selected from aminoacid, amino acid derivativges and peptide separately;
And wherein said interval base joint and described separable joint are connected to form butanimide-1-base alkyl acetal or ketal with described hetero atom joint separately.
32. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint, they are combined together to form 3-thiosuccimide-1-base alkoxyl methoxy base, and wherein said methyl is optional to be replaced by the aryl of alkyl or replacement.
33. claim 1,2 or 3 drug release conjugate; wherein said multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint; they are combined together to form 3-thiosuccimide-1-base alkyl-carbonyl, and wherein said carbonyl and described medicine or its analog or derivant form the acyl group aziridine together.
34. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprise hetero atom joint, basic joint and separable joint at interval, they are combined together to form 1-alkoxyl cycloalkylidene oxygen base.
35. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprise basic joint, hetero atom joint and separable joint at interval, they are combined together to form alkylidene amino carbonyl (dicarboxyl arlydene) carboxylic acid ester groups.
36. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint, they are combined together to form dithio alkyl-carbonyl hydrazides, and wherein said hydrazides forms hydrazone with described medicine or its analog or derivant.
37. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises hetero atom joint, the basic joint in interval and separable joint, they are combined together to form 3-thiosuccimide-1-base alkyl-carbonyl hydrazides, and wherein said hydrazides forms hydrazone with described medicine or its analog or derivant.
38. claim 1,2 or 3 drug release conjugate; wherein said multivalence joint comprises hetero atom joint, the basic joint in interval, hetero atom joint, the basic joint in interval and separable joint; they are combined together to form 3-sulfane base sulfonyl alkyl (dibasic silicyl) oxygen base, and wherein said dibasic silicyl is replaced by alkyl or the optional aryl that replaces.
39. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprise multiple interval base joint, described interval base joint is selected from natural amino acid and stereoisomer thereof.
40. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint, they are combined together to form 3-dithio alkoxy carbonyl, and wherein said carbonyl forms carbonic ester with described medicine or its analog or derivant.
41. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint, they are combined together to form 3-dithio aryl-alkoxy carbonyl, wherein said carbonyl forms carbonic ester with described medicine or its analog or derivant, and described aryl is optional is substituted.
42. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises hetero atom joint, the basic joint in interval, separable joint, the basic joint in interval and separable joint, they are combined together to form 3-thiosuccimide-1-base alkoxyl alkoxyl alkylidene radical, wherein said alkylidene radical forms hydrazone with described medicine or its analog or derivant, each alkyl is independently selected, and described oxygen base alkoxyl is optional by alkyl or the optional aryl replacement that replaces.
43. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprise separable joint, basic joint and separable joint at interval, they are combined together to form 3-dithio alkoxy carbonyl hydrazides.
44. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint, they are combined together to form 3-dithio alkyl amino, and wherein said amino forms vinylogous amide with described medicine, drug analogue or medicaments derivative.
45. the drug release conjugate of claim 44, wherein said alkyl are ethyl.
46. claim 1,2 or 3 drug release conjugate, wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint, they are combined together to form 3-dithio alkyl amino-carbonyl, and wherein said carbonyl forms carbamate with described medicine, drug analogue or medicaments derivative.
47. the drug release conjugate of claim 46, wherein said alkyl are ethyl.
48. claim 1,2 or 3 drug release conjugate; wherein said multivalence joint comprises separable joint, the basic joint in interval and separable joint; they are combined together to form 3-dithio aryl-alkoxy carbonyl, and wherein said carbonyl forms carbamate or carbamoyl aziridine with described medicine, drug analogue or medicaments derivative.
49. the drug release conjugate of claim 3, wherein at least a medicine is a hapten.
50. the drug release conjugate of claim 53, wherein said hapten are selected from fluorescein and dinitrophenyl hapten.
51. a Pharmaceutical composition, described compositions comprise claim 1,2 or 3 drug release conjugate and pharmaceutically acceptable carrier, diluent or excipient.
52. method of in host animal, eliminating pathogenic cell colony with pathogenic cell colony, the available binding site that each member of wherein said pathogenic cell colony has vitamin or its analog or derivant, and wherein said binding site is by the unique expression of described pathogenic cell, overexpression or preferential the expression, and described method comprises and gives described host's claim 1,2 or 3 drug release conjugate or the step of its Pharmaceutical composition.
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|---|---|---|---|
| US70995005P | 2005-08-19 | 2005-08-19 | |
| US60/709,950 | 2005-08-19 | ||
| US60/787,558 | 2006-03-30 |
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| CN201410100643.3A Division CN103893778A (en) | 2005-08-19 | 2006-08-18 | Multi-drug ligand conjugates |
| CN201410100748.9A Division CN103893779A (en) | 2005-08-19 | 2006-08-18 | Multi-drug ligand conjugates |
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| CN107847605A (en) * | 2015-11-25 | 2018-03-27 | 乐高化学生物科学股份有限公司 | Antibody drug conjugate and its correlation technique comprising branch joint |
| CN107847609A (en) * | 2015-03-13 | 2018-03-27 | 恩多塞特公司 | For treating the conjugate of disease |
| US11413353B2 (en) | 2015-11-25 | 2022-08-16 | Legochem Biosciences, Inc. | Conjugates comprising self-immolative groups and methods related thereto |
| US11654197B2 (en) | 2017-03-29 | 2023-05-23 | Legochem Biosciences, Inc. | Pyrrolobenzodiazepine dimer prodrug and ligand-linker conjugate compound of the same |
| US11707533B2 (en) | 2019-09-04 | 2023-07-25 | Legochem Biosciences, Inc. | Antibody-drug conjugate comprising antibody against human ROR1 and use for the same |
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| CN106687121A (en) * | 2014-05-14 | 2017-05-17 | 亚历克斯利维斯兹奇管理和控股有限公司 | Improved polyethyleneimine polyethyleneglycol vectors |
| CN107847609A (en) * | 2015-03-13 | 2018-03-27 | 恩多塞特公司 | For treating the conjugate of disease |
| CN107847605A (en) * | 2015-11-25 | 2018-03-27 | 乐高化学生物科学股份有限公司 | Antibody drug conjugate and its correlation technique comprising branch joint |
| CN113599533A (en) * | 2015-11-25 | 2021-11-05 | 乐高化学生物科学股份有限公司 | Antibody-drug conjugates comprising branched linkers and methods related thereto |
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| CN107847605B (en) * | 2015-11-25 | 2021-12-24 | 乐高化学生物科学股份有限公司 | Antibody-drug conjugates comprising branched linkers and methods related thereto |
| US11413353B2 (en) | 2015-11-25 | 2022-08-16 | Legochem Biosciences, Inc. | Conjugates comprising self-immolative groups and methods related thereto |
| US11654197B2 (en) | 2017-03-29 | 2023-05-23 | Legochem Biosciences, Inc. | Pyrrolobenzodiazepine dimer prodrug and ligand-linker conjugate compound of the same |
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| US11707533B2 (en) | 2019-09-04 | 2023-07-25 | Legochem Biosciences, Inc. | Antibody-drug conjugate comprising antibody against human ROR1 and use for the same |
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