CN101128425A

CN101128425A - Dimeric IAP inhibitors

Info

Publication number: CN101128425A
Application number: CNA2006800053464A
Authority: CN
Inventors: S·M·康登; M·G·拉波特; 邓一军; S·R·里平
Original assignee: TetraLogic Pharmaceuticals Corp
Current assignee: Medville Co., Ltd.
Priority date: 2005-02-25
Filing date: 2006-02-27
Publication date: 2008-02-20
Anticipated expiration: 2026-02-27
Also published as: ZA200707023B; CN101128425B

Abstract

Molecular mimics of Smac are capable of modulating apoptosis through their interaction with cellular IAPs (inhibitor of apoptosis proteins). The mimetics are based on a monomer or dimer of the N-terminal tetrapeptide of IAP-binding proteins, such as Smac/DIABLO, Hid, Grim and Reaper, which interact with a specific surface groove of IAP. Also disclosed are methods of using these peptidomimetics for therapeutic purposes. In various embodiments of the invention the Smac mimetics of the invention are combined with chemotherapeutic agents, including, but not limited to topoisomerase inhibitors, kinase inbhibitors, NSAIDs, taxanes and platinum containing compounds use broader language.

Description

IAP dimer inhibitor

Cross reference and relevant application: the application requires the right of priority of following application, incorporate the full content of following application into the application by reference at this: the U.S. Provisional Application the 60/656th that is entitled as " peptide mimics (PEPTIDOMIMETICS) " that on February 25th, 2005 submitted to, No. 201, the U.S. Provisional Application the 60/668th that is entitled as " the immunotherapy purposes of SMAC stand-in (IMMUNOTHERAPEUTICUSES OF SMAC MIMETICS) " that on April 5th, 2005 submitted to, No. 344, the U.S. Provisional Application of submitting on June 20th, 2005 the 60/692nd that is entitled as " separately or with the peptide mimics (PEPTIDOMIMETICS OF SMAC ALONE OR IN COMBINATION WITHTOPOISOMERASE INHIBITORS) of topoisomerase bonded SMAC ", No. 111, the U.S. Provisional Application of submitting on August 9th, 2005 the 60/706th that is entitled as " as the peptide mimics (PEPTIDOMIMETICS OF SMAC AS CIAP INHIBITORS) of the SMAC of CIAP inhibitor ", No. the 60/729th, 853, the U.S. Provisional Application that No. 649 and on October 25th, 2005 submit to is entitled as " separately or with the peptide mimics that contains platinic compound and taxanes bonded SMAC (PEPTIDOMIMETICS OF SMAC ALONE OR INCOMBINATION WITH PLATINUM CONTAINING COMPOUNDS AND TAXANES) ".

Background technology

Apoptosis (apoptosis) has played vital role in the growth of all multi-cell organisms and homeostasis.Apoptosis (Apoptotis) can be in cell be caused by the external factor (outside approach) such as chemokine or via causing such as incident in the cell of dna damage (inner approach).The change of apoptosis path has involved polytype human pathology, comprising: dysplasia, cancer, autoimmune disorder and neurodegenerative disease.A kind of binding mode of chemotherapeutic drug is via apoptotic necrocytosis.

Apoptosis between species be guard and mainly realize that by the activatory caspase described caspase belongs to the L-Cysteine HCL Anhydrous family that its substrate is had aspartic acid specific.These aspartate specific protease (" caspase ") that contain halfcystine produce in cell as the proenzyme of catalytically inactive and be processed into active proteolytic enzyme through proteolysis during apoptosis.Effector caspase one is activated, promptly becomes the reason of the proteolytic cleavage of the wide spectrum cell target spot that finally causes necrocytosis.In the normal survivaling cell of not accepting the apoptosis stimulation, most caspase keeps non-activity.If caspase is activated singularly, the conservative protein families of differentiation that their proteolytic activity can be called IAP (inhibitor of apoptosis protein matter) suppresses.

Proteinic IAP family suppresses apoptosis by activation that prevents caspase proenzyme (procaspase) and the enzymic activity that suppresses ripe caspase.Identified the Mammals IAP that some are special, comprised XIAP, c-IAP1, c-IAP2, ML-IAP, NAIP (neuronal apoptosis inhibitor protein matter), Bruce and Survivin, they all show the activity of anti-apoptotic in cell cultures.Be to substitute the proteic functional capabilities of P35 (a kind of anti-apoptotic gene) by IAP in baculovirus (baculovirus), to have found IAP at first.The IAP in the organism from the fruit bat to the mankind is described, and known IAP crosses expression in multiple human cancer.Generally speaking, IAP comprises that one to three kind baculovirus LAP IAP repeats (BIR) territory, and most IAP also has C-terminal fourth finger motif.The BIR territory self is the zinc-binding domain that contains about 70 residues of 4 α monocycles and 3 β chains, has halfcystine and histidine residues with zinc ion coordination.Think that thereby the BIR territory produces the anti-apoptotic effect by suppressing caspase and suppressing apoptosis thus.XIAP most grow up and fetal tissue in express at large.Proved that XIAP cross expressing in tumour cell given the provide protection that resists multiple short apoptotic stimulation and improved chemotherapeutical tolerance.Consistent with above-mentioned situation, for the patient who suffers from acute myelogenous leukemia, proved between the protein level of XIAP and the survival rate to have very strong dependency.Shown that the downward modulation of XIAP being expressed by antisense oligonucleotide makes tumour cell dead responsive to what bring out in vitro and in vivo via short apoptosis agent widely.Proved that also Smac/DIABLO deutero-peptide makes a large amount of different tumor cell lines to via the drug-induced apoptosis sensitivity of multiple short apoptosis.

Yet, to stand in the apoptotic normal cell in indication, must eliminate the retarding effect that IAP mediates--to small part by be called as Smac ( sEond mItochondrial aCtivator of cAspases, second kind of plastosome activator of caspase) process that mitochondrial protein is implemented.Smac (perhaps DIABLO) is synthesized as the precursor molecule of 239 seed amino acids; 55 residues of N-terminal work as introducing the plastosome targeting sequence of removing the back.The mature form of Smac contain 184 amino acid and in solution as the oligomer functionating.There has been proposal that Smac and various fragment thereof the target as identify therapeutic agents is used.

Use N-terminal synthetic Smac in tenuigenin of plastosome targeting sequence, the N-terminal of described plastosome targeting sequence is removed and space in the mitochondrial film of target subsequently through proteolysis at the ripening period that becomes mature polypeptide.When bringing out apoptosis, Smac and cytochrome c are released into tenuigenin from plastosome together, and Smac combines and make the caspase activation at this with IAP, have wherein eliminated the restraining effect of IAP pair cell apoptosis.Although cytochrome c induces the multimerization of Apaf-1 to activate caspase-9 proenzyme and procaspase-3, Smac has eliminated the restraining effect of most IAP.Smac interacts with all IAP basically, has found out that up to now IAP comprises XIAP, c-IAP1, c-IAP2 and ML-IAP.Therefore, Smac apoptotic main conditioning agent in the Mammals seemingly.

Shown that Smac works as the IAP antagonist, the enzymic activity that it has not only improved the proteolytic activity of caspase proenzyme but also has improved ripe caspase, both all depend on itself and IAP physical interaction ability.X-ray crystallography shows that preceding four amino acid (AVPI) of ripe Smac combine with the part of IAP.This N-terminal sequence is in conjunction with IAP and to block its anti-apoptotic effect necessary.

The fundamental biological knowledge of IAP antagonist shows that IAP can replenish or the collaborative curative effect that strengthens other chemotherapeutic/antineoplastic agents and/or radiation.As the result of dna damage and/or cell metabolism interruption, apoptosis is brought out in expection chemotherapeutic/antineoplastic agent and radiation.

The directional focusing of cancer drug design at present is the selectivity of pair cell apoptotic signal approach activation in tumour when normal cell is escaped injury.The tumour-specific character that report specificity antineoplastic agent (such as TRAIL) has been arranged.Relevant with tumour necrosis factor one of bringing out among some members that apoptotic part (TRAIL) is tumour necrosis factor (TNF) superfamily, described tumour necrosis factor (TNF) superfamily is by bringing out apoptosis with participating in of death receptor.TRAIL and very complicated receptor system interaction, this system comprises two kinds of death receptors and three kinds of bait acceptors (decoy receptor) in the mankind.TRAIL is used as antineoplastic agent separately and in conjunction with other medicament, described other medicament comprises chemotherapeutic agent and ionizing rays.TRAIL can be in crossing the cell express survival factors Bcl-2 and Bcl-XL activated cell apoptosis and can propose therapeutic strategy at chemotherapeutic agent being had the tumour that obtains resistance.TRAIL combines with its homoreceptor and activates the cascade reaction of the caspase that utilizes adaptor molecule (as FADD).At present, five kinds of TRAIL acceptors have been identified.The receptor-mediated apoptosis signals conduction of these two kinds of TRAIL-R1 (DR4) and TRAIL-R2 (DR5), and DcR1, DcR2 and these three kinds of non-functional acceptors of osteoprotegerin (OPG) can be used as the bait acceptor and work.When strengthening medicament that DR4 and DR5 express and TRAIL and be used in combination, can show collaborative anti-tumor activity.

The beneficial effect that TRAIL produces has been shown in the cancer of several types.For example, the instillation of the intravesical of BCG vaccine has been brought out the Th1 immune response and has been a roentgenism x for the treatment of at superficial bladder cancer, and described Th1 immune response causes the generation of the antitumor cell factor (comprising TRAIL) and the infiltration damage of immunocyte.In vitro study shows tests the apoptosis that the interferon-alpha (INF-α) of bladder cancer curative effect has been caused the TRAIL mediation that produces via autocrine in the human bladder cancer cell lines at present in clinical study.The cyclical level of osteoprotegerin in suffering from the patient of bladder cancer (the bait acceptor of TRAIL) is also increasing and is having negative correlation with the degree and the prognosis of tumor stage, tumour.

In addition, shown that the treatment that the TRAIL by NK (natural killer) cell expresses via IL-2 (interleukin II) strengthens, and the cytotoxic effect that is expressed as whole tumour cells of TRAIL is required.At present, approved is used for IL-2 (a kind of cytokine) treatment of melanoma and renal cell carcinoma.

Because cancer cells duplicates and/or the inhibition of dna damage reparation will impel the nuclear dna break, so inducing cell enters the apoptosis path.Topoisomerase is important for the cell processes such as dna replication dna and reparation, and to be a class reduce supercoiled enzyme among the DNA by one of dna molecular or two chains are disconnected and connect once more to topoisomerase.This zymoid restraining effect is weakened the ability of cellular replication and reparation damage dna and activated inner apoptosis path.

The major avenues of approach that causes necrocytosis that the dna damage that is mediated by topoisomerase causes comprises by the short apoptosis molecule that is discharged by plastosome such as the activation of the caspase in the Smac pair cell matter.Closely control participating in of these apoptosis effector paths by the upstream regulation path, described upstream regulation path is to replying via standing the dna damage generation that topoisomerase enzyme inhibitor brings out in the apoptotic cell.By guaranteeing inhibition to the cell response of dna damage to bringing out via topoisomerase enzyme inhibitor with DNA part bonded protein kinase.These kinases (described kinase whose limiting examples comprises Akt, JNK and P38) that are commonly referred to " DNA transmitter " come mediated dna reparation, cell cycle arrest and/or apoptosis by making a large amount of substrate (comprising some downstream kinases) phosphorylation.

The chemotherapy platinum medicine belongs to the dna modification agent of generic categories.The dna modification agent can be the reactive chemical compound of any height, various nucleophilic group bondings in described compound and nucleic acid and the protein and mutagenesis, carcinogenic or cytotoxic effect.The dna modification agent is worked by different mechanism and is obtained identical net result, and described different mechanism is for the formation that destroys among DNA function and necrocytosis, the dna damage/DNA cross-bridges between atom or key and bring out and cause the Nucleotide mispairing that suddenlys change.Three kinds of limiting examples that contain the dna modification agent of platinum are cis-platinum (cisplatin), carbon platinum (carboplatin) and oxaliplatin (oxaliplatin).

It is believed that cis-platinum is by combining and disturb the repair mechanisms of DNA with DNA, finally cause necrocytosis to come kill cancer cell.Carbon platinum and oxaliplatin are the cis-platinum derivatives that adopts same function mechanism.Highly reactive platinum complex forms in cell and by forming in the chain with the dna molecular covalent attachment and the synthesizing of the crosslinked DNA of suppressing of interchain DNA.

Shown that NSAID (non-steroidal anti-inflammatory drug) (NSAID) brought out apoptosis in colorectal cell.NSAID seemingly via Smac from plastosome discharge and bring out apoptotic (PNAS, November 30,2004, vol.101:16897-16902).Therefore, expection NSAID and Smac stand-in are united the activity that the activity that makes every kind of medicine is increased to the medicine that surpasses every kind of independent use.

The people's such as Shi that submit to September 28 calendar year 2001 and authorize on January 31st, 2006 the United States Patent (USP) the 6th that is entitled as " being used to regulate procedural apoptotic composition and method (Compositions and method forRegulating Apoptosis) ", 992, the stand-in of pointing out the N-terminal part of Smac for No. 063 provide feasible drug candidate, incorporate the full content of this patent into this paper by reference at this.

In addition, the U. S. application the 10/777th that being entitled as of the people such as McLendon that submit on February 12nd, 2004 " is applied to diagnose carrying molecule and peptide mimics (IAP-Binding Cargo Moleculesand Peptidomimetics For Use In Diagnostic and Therapeutic Methods) in conjunction with IAP with methods of treatment ", in No. 946, shown that carrying molecule (cargo molecule) can be connected in the N-terminal of Smac tetrapeptide peptide mimics, incorporates the full content of this application into this paper by reference at this.

Summary of the invention

The invention provides simulation and the compound of the tertiary structure of IAP bonded Smac or the active compound of N-terminal part of simulation Smac.The present invention also comprises the steric isomer of simulated compound described herein.The present invention also provides the method for using these stand-in adjusting apoptosis and being further used for therapeutic purpose.The present invention also provides intermediate and uses these intermediate preparation to regulate the method for apoptotic compound, and described compound is to regulate apoptotic by simulation and the tertiary structure of IAP bonded Smac or the N-terminal part activity of simulation Smac.

Compound of the present invention has following general formula (I):

Wherein R1 and R2 be independently H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, the optional alkyl that replaces or

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroaralkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio or R5a and R5b are randomly replaced by following radicals independently: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Or randomly, R5a is connected by following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene bridge (alkynylene bridge) or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms are replaced by N, O or S;

R6a and R6b be independently H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, low alkyl group, the optional alkyl that replaces or

Wherein R7a and R7b are H, alkyl, cycloalkyl, haloalkyl independently; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring; R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroaralkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroaralkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

R3a and R3b are H, halogen, alkyl, aryl, aralkyl, amino, virtue amino, aryl alkyl amino, hydroxyl, alkoxyl group, aryloxy, aralkyl hydroxyl, dialkyl amido, amide group, sulfonamido or amidino groups independently;

M and n are 0,1,2 or 3 independently;

X and Y are O, N, S or C=C independently; With

R9a, R9b, R10a, R10b are H, alkyl, the optional alkyl that replaces, aryl, heteroaryl, the optional aryl that replaces, the optional heteroaryl that replaces independently, and perhaps R9a and R10a can be connected and form fragrance or non-aromatic ring abreast independently or with R9b and R10b via 4-8 the optional atom that replaces such as C, N, O or S; With

When Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms are replaced by N, O or S; And R11a and R11b independently for do not exist, H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces; Or R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms are randomly replaced by N, O or S;

As Wa and Wb not during covalent attachment, Wa and Wb are H, Cl, Br, F, alkyl, CN, CO independently ₂H; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are randomly by N, O or S replacement; Or Wa is H, Cl, Br, F, alkyl, CN, CO ₂H and Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, wherein one or more carbon atoms can be replaced by N, O or S, and R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

Another embodiment of the invention is that The compounds of this invention is with TRAIL or combine and activate other chemistry of TRAIL acceptor or the therapeutic combination of biotechnological formulation.Because following discovery makes TRAIL receive sizable concern recently: promptly, the apoptosis sensitivity that multiple cancer cells type is brought out TRAIL, and as if most normal cell have tolerance to this effect of TRAIL.Cell to TRAIL tolerance can produce by multiple different mechanism, described mechanism comprise acceptor disappearance, bait acceptor existence or during DISC forms, compete caspase-8 proenzyme FLIP cross expression.In the tolerance of TRAIL, Smac stand-in have increased the susceptibility of tumour cell to the TRAIL that impels necrocytosis and increase, expect both clinical correlation for apoptoticly in the tumour of TRAIL tolerance actively increase, clinical response improves, duration of the reaction prolongs and final patient's survival rate improves.In order to support this viewpoint, the reduction of the level of the verified XIAP that causes via external antisense therapy makes the melanoma cell of tolerance and kidney cancer cell to TRAIL sensitivity people such as (, 2004) Chawla-Sarkar.The disclosed Smac stand-in of this specification sheets combine and suppress the interaction of they and caspase with IAP, wherein strengthened the apoptosis that TRAIL brings out.

In another embodiment of the invention, Smac stand-in and BCG vaccine are united the treatment that is used for bladder cancer.The nominal target XIAP of Smac stand-in crosses in a high proportion of bladder cancer patients and expresses.In the research of using antisense XIAP, transitional cell bladder carcinoma cell line is to bringing out the apoptotic chemotherapeutic sensitivity that is acted on by the TRAIL path.The invention provides the Smac stand-in, described Smac stand-in and BCG one are used from the treatment of superficial bladder cancer disease/carcinoma in situ.If vaccine is replied and produced TRAIL, then the disclosed Smac stand-in of this specification sheets will be by strengthening the effectiveness that this is used for improving the BCG vaccine.

Similarly, the Smac stand-in will increase the apoptosis that observed TRAIL brings out among the patient who is just using IL-2 treatment melanoma and renal cell carcinoma.Because IL-2 brings out the activity that the NK cell promotes that TRAIL expresses, the treatment of therefore uniting the activator (as the Smac stand-in) of caspase-9 will produce more effective clinical response.

Another embodiment of the invention provides Smac stand-in, strengthens both and brings out apoptotic effect thereby described Smac stand-in and topoisomerase enzyme inhibitor play synergy.Topoisomerase enzyme inhibitor suppresses duplicating and repairing of DNA, thereby promotes apoptosis, and described topoisomerase enzyme inhibitor uses as thermochemistry therapeutical agent (chemothemotherapeutic agent).Topoisomerase enzyme inhibitor is by being suppressed at the damage that enzyme required in the DNA repair process quickens DNA.Therefore, induce in the tenuigenin from mitochondrial cytochrome c and Smac input cell by the dna damage that causes via topoisomerase enzyme inhibitor.

Topoisomerase enzyme inhibitor (the camptothecine of I class, Hycamtin (tomptecan), SN-38, Rinotecan (irinotecan), Hycamtin, BNP 1350,9-amino-irinotecan (camptothecan), lurtotecan, lattice are auspicious for health (grimatecan), exatecan, amsacrine and fluorine are for health (diflomotecan)) and the topoisomerase enzyme inhibitor (Etoposide of II class, anthracene nucleus element (anthracycyline), anthraquinone and podophyllotoxin) especially in following clone, demonstrate the powerful synergy with Smac stand-in of the present invention: multidrug resistant glioblastoma clone (T98G), breast cancer cell line (MDA-MB-231) and ovarian cancer cell line (OVCAR-3) also have other.For example, other topoisomerase enzyme inhibitor comprises: aclacinomycin A, camptothecine, daunorubicin, Dx, Ellipticine, epirubicin and mitoxantrone (mitaxantrone).

In another embodiment of the invention, chemotherapeutic/antineoplastic agent can be the platiniferous compound.In one embodiment of the invention, the platiniferous compound is a cis-platinum.Cis-platinum can play synergy with the Smac peptide mimics and strengthen restraining effect to IAP, and described IAP for example but is not limited to: XIAP, cIAP-1, c-IAP-2, ML-IAP etc.In another embodiment, the platiniferous compound is a carbon platinum.Carbon platinum can play synergy with the Smac peptide mimics and strengthen restraining effect to IAP, and described IAP includes but not limited to: XIAP, cIAP-1, c-IAP-2, ML-IAP etc.In another embodiment, the platiniferous compound is an oxaliplatin.Oxaliplatin can play synergy with the Smac peptide mimics and strengthen restraining effect to IAP, and described IAP includes but not limited to: XIAP, cIAP-1, c-IAP-2, ML-IAP etc.

In another embodiment of the invention, playing synergistic chemotherapeutic/antineoplastic agent with compound according to the present invention is Taxan.Taxan is antimitotic mitotic inhibitor or microtubule polymerization agent.Taxan includes but not limited to Docetaxel and taxol.

Taxan is characterized as being such compound: it is by suppressing the assembling of tubulin depolymerization promotion microtubule, thereby by the pole corpuscle damage, bring out irregular spindle body and suppress spindle microtubule kinetics and stop cell cycle progression.Compare with other microtubule toxic agents such as vinca alkaloids, colchicine and cryptophycine, unique mechanism of action of Taxan is to suppress the polymerization of tubulin.Microtubule be by the α 'beta '-tubulin with in the mitotic division process by participating in high dynamic (dynamical) cell aggregation thing that related protein that spindle body tissue and function guarantee that institute's separated DNA integrity plays a crucial role constitutes.

In another embodiment, the inner cell apoptosis pathway of any activation and/or cause Smac or cytochrome c has and the synergistic potentiality of Smac stand-in from the medicament that plastosome discharges.

The Smac peptide mimics of any kind of the release of activation inside or external path or Smac and chemotherapeutic/antineoplastic agent and/or radiotherapeutic combination can provide tumoricidal more efficiently means.The apoptotic restraining effect that Smac peptide mimics and IAP interact and blocking-up mediates IAP, and chemotherapeutic/antineoplastic agent and/or radiotherapy are to cause the inside cell apoptosis pathway of apoptosis and necrocytosis to kill somatoblast on one's own initiative by activation.As hereinafter in greater detail, embodiment of the present invention provide the combination of Smac peptide mimics and chemotherapeutic/antineoplastic agent and/or radiation, and it provides the synergy of the cell proliferation of the non-expectation of antagonism.Synergy between Smac peptide mimics and chemotherapeutic/antineoplastic agent and/or the radiotherapy can improve chemotherapeutic/antineoplastic agent and/or radiotherapeutic curative effect.This will allow to make at present the dosage of chemotherapeutic/antineoplastic agent to reduce and chemotherapeutic/antineoplastic agent or radiocurable effect strengthen, the dosage that more efficiently dosage regimen wherein is provided and more can have tolerated for chemotherapeutic/antineoplastic agent and/or radiotherapy.

For easy and purpose example, come principle of the present invention is described by mainly referring to its embodiment.In addition, in order to obtain a large amount of details has been proposed in the following description in thorough understanding of the present invention.Yet, for the ordinary skill in the art, it is evident that and can implement the present invention being not limited under the situation of these details.In other situation,, known method and structure are not described in detail for fear of the present invention being caused unnecessary fuzzy understanding.

Description of drawings

Fig. 1 be to use the fluorescence polarization assay method describe Smac tetrapeptide of the present invention (AVPI) and effectively the Smac stand-in to the graphic representation of the RA of XIAP BIR-3.The result shows the Smac stand-in with respect to the Smac tetrapeptide, and binding affinity has increased by 30,000 times.

Fig. 2 is the transformation period figure that shows three kinds of Smac stand-in of the present invention are used single dose 1mg/kg in to rat vein after.The result shows that the stand-in transformation period of being tried reaches 6h.

Fig. 3 shows that Smac stand-in of the present invention optionally resist the graphic representation of ovarian cancer cell line SK-OV-3 propagation.In this MTT test, the Smac stand-in show anticancer character to the adiaphorous concentration of normal diploid cell line MRC-5 the time.

Fig. 4 shows that use has demonstrated the chemical synergism to the Smac stand-in of the melanoma cell of the apoptosis effect of TRAIL tolerance.The test that is used for cell proliferation shows that this cell is to the antiproliferative effect tolerance of Smac stand-in of the present invention when use the present invention numbers 1 Smac peptide mimics processing breast cancer cell line MDA-MB-231 cell separately.By contrast, as detected by the corresponding minimizing that forms at colony, when numbering 1 and TRAIL unite when using, the cell that causes killing increases by 100 times antiproliferative effect and has increased by 1000 times.

Detailed Description Of The Invention

Also it must be noted that, as employed in this specification and the claims, unless in addition clearly explanation of context, singulative comprises plural implication. Unless otherwise defined, otherwise the implication that all technology used herein and scientific terminology have all with those of ordinary skills usually understand identical. Although can use any similarly or be equal to those method described herein in the practice of embodiment of the present invention or test, description is preferred method at present. All that to herein mention are by reference announced and list of references is incorporated this paper into. Any part of this paper should be interpreted as admitting that the present invention haves no right because invention formerly shifts to an earlier date the date of this published content.

As used herein, term " about " refers to employed numeral is added or deducts the numerical value of this numeral 10%. Therefore, " about 50% " refers to the scope of 45%-55%.

Except as otherwise noted, " alkyl " refers to have side chain or unbranched saturated or undersaturated (being thiazolinyl, alkynyl) aliphatic alkyl group, and it can have nearly 12 carbon atoms. When moieties used as the part of another term, for example " alkylamino ", described moieties can be saturated hydrocarbon chains, but also can comprise undersaturated alkyl carbochain, such as " enamino " and " alkynes is amino ". The example of specific alkyl group comprises methyl, ethyl, n-pro-pyl, isopropyl, normal-butyl, isobutyl group, sec-butyl, the tert-butyl group, n-pentyl, 2-methyl butyl, 2,2-dimethyl propyl, n-hexyl, 2-methyl amyl, 2,2-dimethylbutyl, n-heptyl, 3-heptyl, 2-methyl hexyl etc. Term " low alkyl group ", " C₁-C ₄Alkyl " identical with " alkyl of 1-4 carbon atom " meaning and interchangeablely be used for representing methyl, ethyl, 1-propyl group, isopropyl, cyclopropyl, 1-butyl, sec-butyl or the tert-butyl group. Unless describe, the alkyl group of replacement can contain can be identical or different 1,2,3 or 4 substituting groups. The example of the alkyl group of above-mentioned replacement includes but not limited to: cyanogen methyl, nitro methyl, methylol, triphen methoxy, propionyloxy methyl, aminomethyl, carboxymethyl, carboxyethyl, carboxylic propyl group, alkoxy carbonyl methyl, allyloxy carbonyl ammonia methyl, carbamoyloxy group methyl, methoxy, ethoxyl methyl, tert-butoxy methyl, acetoxy-methyl, chloromethyl, bromomethyl, iodomethyl, trifluoromethyl, 6-hydroxyl hexyl, 2,4-dichloro (normal-butyl), 2-amino (isopropyl), 2-carbamyl oxygen ethyl etc. Alkyl group can also be replaced by carbon ring group. Example comprises cyclopropyl methyl, cyclobutylmethyl, cyclopentyl-methyl and cyclohexyl methyl group, and corresponding ethyl, propyl group, butyl, amyl group, hexyl groups etc. The methyl of concrete substituted alkyl for replacing is such as the methyl group that replaces via the substituting group identical with " the Cn-Cm alkyl of replacement " group. The example of the methyl group that replaces comprises following radicals, for example: methylol, shielded methylol (such as the tetrahydropyran oxygen ylmethyl), acetoxy-methyl, carbamyl oxygen methyl, trifluoromethyl, chloromethyl, carboxymethyl, bromomethyl and iodomethyl.

" amino " refer to uncle ammonia (namely-NH₂), secondary amine (namely-NRH) and tertiary amine (namely-NRR). Concrete secondary amine and tertiary amine are alkylamine, dialkylamine, arylamine, diaryl amine, aromatic yl alkyl amine and alkyl diaryl amine. Concrete secondary amine and tertiary amine are methyl amine, ethylamine, propyl group amine, isopropylamine, aniline, benzyl amine, dimethyl amine, diethylamide, dipropylamine and diisopropylamine (disopropylamine).

When " aryl " uses separately or uses as the part of another term, refer to the carbocyclic ring aromatic group, if no matter whether condense the carbon atom number of appointment or do not specify number, nearly 14 carbon atoms can be arranged. Concrete aromatic yl group comprise phenyl, naphthyl, xenyl, phenanthryl, aphthacene base (naphthacenyl) etc. (referring to: such as Lang ' s Handbook of Chemistry (Dean, J.A., ed) the 13rd edition, the table 7-2[1985]). In one specific embodiment, aryl is phenyl. Except as otherwise noted; the phenyl that replaces or the aryl of replacement represent to use 1,2,3,4 or 5 phenyl or aryl that substituting group replaces, and described substituting group is selected from halogen (F, Cl, Br, I), hydroxyl, shielded hydroxyl, cyano group, nitro, alkyl (such as C₁-C ₆Alkyl), alkoxyl is (such as C₁-C ₆Alkoxyl), the group of benzyloxy, carboxyl, shielded carboxyl, carboxymethyl, shielded carboxymethyl, methylol, shielded methylol, aminomethyl, shielded aminomethyl, trifluoromethyl, alkyl sulfonamide, aryl sulfonic acid amides, heterocyclic radical sulfonamide (heterocyclylsulfonylamino), heterocyclic radical (heterocyclyl), aryl or other appointments. One or more methines (CH) and/or methylene (CH in these substituting groups₂) group can use again and those similar groups as noted above replace. The example of term " phenyl of replacement " includes but not limited to list or two (halogen) phenyl group, as: 2-chlorphenyl, 2-bromophenyl, 4-chlorphenyl, 2,6-dichlorophenyl, 2,5-dichlorophenyl, 3,4-dichlorophenyl, 3-chlorphenyl, 3-bromophenyl, 4-bromophenyl, 3,4-dibromo phenyl, 3-chloro-4-fluorophenyl, 2-fluorophenyl etc.; List or two (hydroxyl) phenyl group, as: 4-hydroxy phenyl, 3-hydroxy phenyl, 2,4-dihydroxy phenyl and shielded hydroxy derivatives thereof etc.; Nitrobenzophenone is such as 3-or 4-nitrobenzophenone; Benzonitrile base, for example 4-benzonitrile base; List or two (low alkyl group) phenyl group, such as 4-tolyl, 2,4-xylyl, 2-tolyl, 4-(isopropyl) phenyl, 4-ethylbenzene, 3-(n-pro-pyl) phenyl etc.; List or two (alkoxyl) phenyl group, for example 3,4-Dimethoxyphenyl, 3-methoxyl group-4-benzyloxy phenyl, 3-methoxyl group-4-(1-chloromethyl) benzyloxy-phenyl, 3-ethoxyl phenenyl, 4-(isopropyl) phenyl, 4-(tert-butoxy) phenyl, 3-ethoxy-4-methoxyphenyl etc.; 3-trifluoromethyl or 4-trifluoromethyl; List or dicarboxyl phenyl or (shielded carboxyl) phenyl group are such as the 4-carboxyl phenyl; List or two (methylol) phenyl or (shielded methylol) phenyl are such as 3-(shielded methylol) phenyl or 3,4-two (methylol) phenyl; List or two (aminomethyl) phenyl or (shielded aminomethyl) phenyl are such as 2-(aminomethyl) phenyl or 2,4-(shielded aminomethyl) phenyl; Or single or two (N-(Methanesulfomide)) phenyl, such as 3-(N-Methanesulfomide)) phenyl. In addition, term " phenyl of replacement " the dibasic phenyl group of expression and trisubstd phenyl group and quaternary phenyl group, substituting group is different in described dibasic phenyl group, for example, 3-methyl-4-hydroxyphenyl, 3-chloro-4-hydroxyphenyl, 2-methoxyl group-4-bromophenyl, 4-ethyl-2-hydroxyphenyl, 3-hydroxyl-4-nitrobenzophenone, 2-hydroxyl-4-chlorphenyl etc.; Substituting group is different in described trisubstd phenyl group, for example, and 3-methoxyl group-4-benzyloxy-6-Methanesulfomide, 3-methoxyl group-4-benzyloxy-6-benzsulfamide; Substituting group is different in described quaternary phenyl group, for example, and 3-methoxyl group-4-benzyloxy-5-methyl-6-benzsulfamide. Concrete substituted-phenyl group is 2-chlorphenyl, 2-aminophenyl, 2-bromophenyl, 3-methoxyphenyl, 3-ethyoxyl-phenyl, 4-benzyloxy phenyl, 4-methoxyphenyl, 3-ethyoxyl-4-benzyloxy phenyl, 3,4-diethoxy phenyl, 3-methoxyl group-4-benzyloxy phenyl, 3-methoxyl group-4-(1-chloromethyl) benzyloxy-phenyl, 3-methoxyl group-4-(1-chloromethyl) benzyloxy-6-Methanesulfomide phenyl group. The aromatic ring that condenses also can use the substituting group of appointment herein to replace, for example, and to use 1,2 or 3 substituting group to replace such as the identical mode of alkyl group that is replaced.

As used herein, term alkylidene atomic group comprises the saturated side chain of the difunctionality that contains 1-30 carbon atom or unbranched hydrocarbyl group, and comprises, for example, and methylene (CH₂), ethylidene (CH₂CH ₂), propylidene (CH₂CH ₂CH ₂), 2-methyl propylidene (CH₂CH(CH ₃)CH ₂), hexylidene ((CH₂) ₆) etc. Low-grade alkylidene comprises 1-10, the more preferably alkylidene group of 1-5 carbon atom.

The alkylidene atomic group that replaces comprises having 1-30 carbon atom and have 1-5 the saturated side chain of substituent difunctionality or unbranched alkylidene atomic group or group. The alkylidene atomic group of rudimentary replacement refers to have 1-10 carbon atom, preferably has a 1-5 carbon atom and have 1-5 substituent substituted alkylene atomic group. Substituting group can include but not limited to those substituting groups for alkyl group.

As used herein, term thiazolinyl atomic group comprises side chain, cyclic hydrocarbon group or the unbranched hydrocarbyl group of 2-30 the carbon atom that contains at least one carbon-carbon double bond, and example has vinyl, positive acrylic, isopropenyl, n-butene base, uncle's cyclobutenyl, octenyl, decene, four decene, own decene, icosa alkene base, two tetradecene bases etc. The term low-grade alkenyl comprises 2-10 the carbon atom that contains at least one carbon-carbon double bond, the alkenyl group of preferred 2-5 carbon atom. One or more carbon-carbon double bonds can have cis or anti-configuration independently. The thiazolinyl atomic group that replaces refers to have 1-5 substituent thiazolinyl atomic group or low-grade alkenyl group, and described substituting group can include but not limited to those substituting groups for alkyl group.

Term alkenylene atomic group comprises difunctional branched or unbranched hydrocarbyl group or the group that contains 2-30 carbon atom and at least one carbon-carbon double bond. " lower alkenylene " comprises 2-10 that contains a carbon-carbon double bond, more preferably the alkenylene group of 2-5 carbon atom. The alkenylene atomic group that replaces refers to have 1-5 substituent alkenylene atomic group or low-grade alkenyl group, and described substituting group can include but not limited to those substituting groups for alkyl group.

Term alkynyl atomic group or group refer to have the straight or branched alkyl of 2-12 carbon atom and at least one triple bond, and some embodiment comprises the alkynyl group of 2-6 carbon atom with a triple bond. The alkynyl that replaces will comprise such as defined 1,2 in alkyl or 3 substituting groups for replacing. Alkynylene comprises difunctional branched or the unbranched hydrocarbon chain that contains 2-12 carbon atom and at least one carbon carbon triple bond; Some embodiment comprises the alkynylene group of 2-6 carbon atom with a triple bond. The alkynylene that replaces will comprise such as defined 1,2 of alkyl groups or 3 substituting groups for replacing.

" heterocyclic group ", " heterocycle ", " heterocycle (heterocycle) ", " heterocyclic radical " or " heterocycle (heterocyclo) " are separately with when rolling into a ball as composite base when using such as the part in the heterocycloalkyl, be used interchangeably, and refer to any atom of specifying number that has, usually from 5 to the single, double of about 14 annular atomses or three ring fillings or undersaturated fragrance (heteroaryl) ring or non-aromatic ring, wherein said annular atoms is carbon and at least one hetero atom (nitrogen, sulphur or oxygen). In one specific embodiment, this group is introduced 1-4 hetero atom. Typically, 5 yuan of rings have 0-2 two keys and 6 or 7 yuan of rings have 0-3 two keys and nitrogen or sulfur heteroatom can be randomly oxidized (such as SO, SO₂), and any nitrogen heteroatom can be randomly quaternized. Concrete nonaromatic heterocycles comprises morpholinyl (morpholino), pyrrolidinyl, Oxyranyle, oxygen cyclobutyl, tetrahydrofuran base, DHF base, 2H-pyranose, THP trtrahydropyranyl, thiiranes group (thiiranyl), sulfuration trimethylene, tetrahydrochysene sulfuration trimethylene, '-aziridino, azetidinyl, 1-methyl-2-pyrrole radicals, piperazinyl and piperidyl. " Heterocyclylalkyl " group is to be covalently attached to the as defined above as defined above heterocyclic group of alkyl group.

5 yuan of heterocycles that contain a sulphur or oxygen atom and 1-3 nitrogen-atoms comprise thiazolyl, thiadiazolyl group,  azoles base and  di azoly, and the example of described thiazolyl has thiazol-2-yl and thiazol-2-yl N-oxide; The example of described thiadiazolyl group has 1,3,4-thiadiazoles-5-base and 1,2,4-thiadiazoles-5-base; The example of described  azoles base has  azoles-2-base; The example of described  di azoly has 1,3,4- diazole-5-base and 1,2,4- diazole-5-base. The concrete 5 yuan of heterocycles that comprise 2-4 nitrogen-atoms comprise imidazole radicals, triazolyl, tetrazole radical, and the example of described imidazole radicals has imidazoles-2-base; The example of described triazolyl has 1,3,4-triazole-5-base, 1,2,3-triazoles-5-base and 1,2,4-triazole-5-base; The example of described tetrazole radical has 1H-TETRAZOLE-5-base. Concrete benzo-fused 5 yuan of heterocycles are benzoxazol-2-base, benzothiazole-2-base and benzimidazolyl-2 radicals-Ji. Contain 1-3 nitrogen-atoms and randomly concrete 6 yuan of heterocycles such as pyridine radicals, pyrimidine radicals, triazine radical, pyridazinyl and the pyrazinyl of sulphur or oxygen atom, the example of described pyridine radicals is pyridine-2-base, pyridin-3-yl and pyridin-4-yl; The example of described pyrimidine radicals is pyrimidine-2-base and pyrimidine-4-yl; The example of described triazine radical is 1,3,4-triazine-2-base and 1,3,5-triazines-4-base; The example of described pyridazinyl is pyridazine-3-base. The other example that is used for 5 and 6 yuan of ring systems of the substituting group of optional substituted heterocycle and above-mentioned discussion is found in the people's such as W.Druckheimer No. the 4th, 278,793, United States Patent (USP).

The aralkyl atomic group refers to aryl substituent and has from about 6 groups to about 20 carbon atoms (and all of various scope and the concrete number of carbon atom wherein make up and subgroup is closed), wherein preferably approximately 6 to about 12 carbon atoms. Aromatic alkyl group can randomly be substituted. Nonrestrictive example comprises, for example benzyl, menaphthyl, benzhydryl, trityl, phenethyl and two phenethyls. The aromatic alkyl group that replaces will comprise the one or more substituting groups on aryl or alkyl that define as for the alkyl group that replaces.

Cycloalkyl aryl atomic group or group refer to the cycloalkyl atomic group that condenses with aromatic yl group, comprise all combinations of independent alkyl-cycloalkyl aryl, cycloalkyl and the aromatic yl group that replaces with two same atoms.

Cycloalkyl atomic group or group more specifically comprise the carbocyclic ring alkyl atomic group that unit price is saturated, the carbocyclic ring alkyl atomic group that described unit price is saturated is comprised of the one or more rings in its structure and has from about 3 to about 14 carbon atoms (and all combinations and the subgroup of various scope and the concrete number of carbon atom are wherein closed), wherein preferably from about 3 to about 7 carbon atoms. Multiring structure can be bridge joint or the ring structure that condenses. This ring can use the one or more substituting groups for alkyl group randomly to replace. The example of group of naphthene base includes but not limited to: cyclopropyl, cyclobutyl, cyclopenta, cyclohexyl, ring octyl group and adamantyl. The group of naphthene base that replaces will contain the one or more substituting groups that define just like for the alkyl group that replaces.

The cycloalkyl-alkyl atomic group more specifically refers to contain naphthenic substituent and has from the about 4 alkyl atomic groups to about 20 carbon atoms (and all combinations and the subgroup of various scope and the concrete number of carbon atom are wherein closed), wherein preferably from about 6 to about 12 carbon atoms, and can include but not limited to: methyl cyclopropyl, methylcyclohexyl, isopropylcyclohexyl-and butyl cyclohexyl groups. Cycloalkyl-alkyl atomic group or group can use the one or more substituting groups for alkyl group randomly to replace, and described substituting group includes but not limited to: hydroxyl, cyano group, alkyl, alkoxyl, alkylthio, halogen, haloalkyl, hydroxyalkyl, nitro, amino, alkylamino and alkylamino.

When " heteroaryl " uses separately with as the part of compound group such as heteroaryl alkyl group, refer to have the aromatic ring of any single, double of specified atom number or three rings, wherein at least one ring is for containing 1-4 heteroatomic 5 yuan of rings, 6 yuan of rings or 7 yuan of rings, and described hetero atom is selected from: nitrogen, oxygen and sulphur (above-mentioned Lang ' s Handbook of Chemistry). Included in this definition is the group of any dicyclo, and wherein any above-mentioned hetero-aromatic ring and phenyl ring condense. Following loop systems is by the example of the heteroaryl groups of term " heteroaryl " expression (replacement or unsubstituted): thienyl, furyl, imidazole radicals, pyrazolyl, thiazolyl, isothiazolyl,  azoles base, different  azoles base, triazolyl, thiadiazolyl group,  di azoly, tetrazole radical, thiatriazole base,  triazolyl, pyridine radicals, pyrimidine radicals, pyrazinyl, pyridazinyl, thiazinyl,  piperazine base, triazine radical, thiadiazine base,  diazine, dithiazine base, two  piperazine bases,  thiazinyl, tetrazine base, thiophene triazine radical,  triazine radical, two thiadiazine bases, imidazolinyl, dihydro-pyrimidin base, tetrahydro-pyrimidine base, tetrazolium [1,5-b] pyridazinyl and purine, and benzo-fused derivative, for example, Benzoxazinyl, benzofuranyl, benzothiazolyl, diazosulfide base, BTA base, benzimidazolyl and indyl. Especially, " heteroaryl " comprising: 1,3-thiazole-2-base, 4-(carboxymethyl)-5-methyl isophthalic acid, the 3-thiazol-2-yl, 4-(carboxymethyl)-5-methyl isophthalic acid, 3-thiazol-2-yl sodium salt, 1,2,4-thiadiazoles-5-base, the 3-methyl isophthalic acid, 2,4-thiadiazoles-5-base, 1,3,4-triazole-5-base, the 2-methyl isophthalic acid, 3,4-triazole-5-base, 2-hydroxyl-1,3,4-triazole-5-base, 2-carboxyl-4-methyl isophthalic acid, 3,4-triazole-5-base sodium salt, 2-carboxyl-4-methyl isophthalic acid, 3,4-triazole-5-base, 1,3- azoles-2-base, 1,3,4- diazole-5-base, the 2-methyl isophthalic acid, 3,4- diazole-5-base, 2-(methylol)-1,3,4- diazole-5-base, 1,2,4- diazole-5-base, 1,3,4-thiadiazoles-5-base, 2-mercaptan-1,3,4-thiadiazoles-5-base, 2-(methyl mercapto)-1,3,4-thiadiazoles-5-base, 2-amino-1,3,4-thiadiazoles-5-base, 1H-TETRAZOLE-5-base, 1-methyl isophthalic acid H-tetrazolium-5-base, 1-(1-(dimethylamino) second-2-yl)-1H-TETRAZOLE-5-base, 1-(carboxymethyl)-1H-TETRAZOLE-5-base, 1-(carboxymethyl)-1H-TETRAZOLE-5-base sodium salt, 1-(methanesulfonic acid)-1H-TETRAZOLE-5-base, 1-(methanesulfonic acid)-1H-TETRAZOLE-5-base sodium salt, 2-methyl isophthalic acid H-tetrazolium-5-base, 1,2,3-triazole-5-base, the 1-methyl isophthalic acid, 2,3-triazole-5-base, the 2-methyl isophthalic acid, 2,3-triazole-5-base, the 4-methyl isophthalic acid, 2,3-triazole-5-base, pyridine-2-base N-oxide, 6-methoxyl group-2-(n-oxide)-pyridazine-3-base, 6-hydroxyl pyridazine-3-base, 1-picoline-2-base, 1-picoline-4-base, 2-hydroxy pyrimidine-4-base, 1,4,5,6-tetrahydrochysene-5,6-dioxo-4-methyl-triazine-3-is basic partially, 1,4,5,6-tetrahydrochysene-4-(formyl methyl)-5, the 6-dioxo-triazine-3-is basic partially, 2,5-dihydro-5-oxo-6-hydroxyl-triazine-3-is basic partially, 2,5-dihydro-5-oxo-6-hydroxyl-inclined to one side triazine-3-base sodium salt, 2,5-dihydro-5-oxo-6-hydroxy-2-methyl-inclined to one side triazine-3-base sodium salt, 2,5-dihydro-5-oxo-6-hydroxy-2-methyl-triazine-3-is basic partially, 2,5-dihydro-5-oxo-6-methoxyl group-2-methyl-triazine-3-is basic partially, 2,5-dihydro-5-oxo-triazine-3-is basic partially, 2,5-dihydro-5-oxo-2-methyl-triazine-3-is basic partially, 2,5-dihydro-5-oxo-2, the 6-dimethyl-triazine-3-is basic partially, tetrazolium [1,5-b] pyridazine-6-base and 8-Aminotetrazole [1,5-b]-pyridazine-6-base. The optional group of " heteroaryl " comprising: 4-(carboxymethyl)-5-methyl isophthalic acid, the 3-thiazol-2-yl, 4-(carboxymethyl)-5-methyl isophthalic acid, 3-thiazol-2-yl sodium salt, 1,3,4-triazole-5-base, the 2-methyl isophthalic acid, 3,4-triazole-5-base, 1H-TETRAZOLE-5-base, 1-methyl isophthalic acid H-tetrazolium-5-base, 1-(1-(dimethylamino) second-2-yl)-1H-TETRAZOLE-5-base, 1-(carboxymethyl)-1H-TETRAZOLE-5-base, 1-(carboxymethyl)-1H-TETRAZOLE-5-base sodium salt, 1-(methanesulfonic acid)-1H-TETRAZOLE-5-base, 1-(methanesulfonic acid)-1H-TETRAZOLE-5-base sodium salt, 1,2,3-triazole-5-base, 1,4,5,6-tetrahydrochysene-5,6-dioxo-4-methyl-triazine-3-is basic partially, 1,4,5,6-tetrahydrochysene-4-(2-formyl methyl)-5, the 6-dioxo-triazine-3-is basic partially, 2,5-dihydro-5-oxo-6-hydroxy-2-methyl-inclined to one side triazine-3-base sodium salt, 2,5-dihydro-5-oxo-6-hydroxy-2-methyl-triazine-3-is basic partially, tetrazolium [1,5-b] pyridazine-6-base and 8-Aminotetrazole [1,5-b] pyridazine-6-base.

" inhibitor " refers to reduce or prevents compound or the minimizing of IAP protein and caspase protein bound or prevent the inhibiting compound of Apoptosis of IAP protein, thereby perhaps be combined the compound of release Smac with inhibition IAP effect with IAP BIR territory in the mode that is similar to Smac amino terminal part.

" pharmaceutically acceptable salt " comprises acid-addition salts and base addition salts. Those salt that " pharmaceutically acceptable acid-addition salts " refers to keep the biological effect of free alkali and character and do not form with biology or other unexpected forms with inorganic acid and organic acid, the example of described inorganic acid has hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid etc., described organic acid can be selected from the organic acid of (araliphatic), heterocycle, carboxyl and the sulfonic group classification of aliphatic, alicyclic, aromatic series, aralkyl, and example has formic acid, acetic acid, propionic acid, glycolic, gluconic acid, lactic acid, pyruvic acid, oxalic acid, malic acid, maleic acid, malonic acid (maloneic acid), butanedioic acid, fumaric acid, tartaric acid, citric acid, L-aminobutanedioic acid, ascorbic acid, glutamic acid, ortho-aminobenzoic acid, benzoic acid, cinnamic acid, mandelic acid, pamoic acid, phenylacetic acid, methanesulfonic acid, ethyl sulfonic acid, p-methyl benzenesulfonic acid, salicylic acid etc.

Term " analogies ", " simulating peptide " and " peptide mimics " herein can Alternates, and are often referred to three grades of integrated structures or active peptide, partial peptide or the non-peptide molecule of the selected native peptides of simulation or protein functional domain (as: binding motif or active site). These simulating peptide comprise peptide and the non-peptide medicament of restructuring or chemical modification, and example is small-molecule drug analogies as described further below.

As used herein, when term " pharmaceutically acceptable ", " can tolerate on the physiology " and grammatical variants thereof refer to composition, carrier, diluent and reagent, be used interchangeably, and expression can be applied to mammal and not produce the material of unexpected physiological effect, the example of described unexpected physiological effect as feel sick, dizzy, rash or epigastric upset.

When " providing " refers to be applied directly to therapeutic agent in the target tissue or on the target tissue or to the agent of patient's administering therapeutic, make thus therapeutic agent that the tissue of its target is had positive impact when being combined with therapeutic agent.

As used herein, " curee " or " patient " refers to animal or mammal, includes but not limited to the mankind, dog, cat, horse, milk cow, pig, sheep, goat, chicken, monkey, rabbit, rat, mouse etc.

As used herein, term " treatment " refers in order to treat, to resist, to alleviate, to prevent or to improve patient's the illness of not expecting or the means of disease. Embodiment of the present invention relate to the promotion Apoptosis, and the cell death that causes thus.

As used herein, term " treatment effective dose " or " effective dose " can Alternates, and refer to that the present invention treats the amount of compound component. For example, the treatment effective dose for the treatment of compound is the predetermined amount that obtains as calculated Expected Results, and described Expected Results namely promotes Apoptosis or effectively preferably by eliminating IAP to apoptotic inhibitory action, more preferably be combined to impel cell to apoptotic sensitivity with caspase by suppressing IAP.

" analogies " or " peptide mimics " are the synthetic compounds with three-dimensional structure (i.e. " core peptide motif "), and described three-dimensional structure is based on the three-dimensional structure of selected peptide. Described peptide motif provides have the expectation biologically active analogies of (namely in conjunction with IAP), wherein simulated compound there is no reduction in conjunction with activity, and usually with identical as the binding affinity of the native peptides of analogies model or greater than the binding affinity as the native peptides of analogies model. For example, in analogies of the present invention, find X₃And X₄Be very similar to non-peptide material. The compound of peptide mimics can have other characteristics that promotes that its treatment is used, such as the permeability that increases cell, improve affinity and/or affinity and prolong biological half-life.

Especially, the layout strategy of stand-in, peptide mimics can obtain easily in the art and can easily be applicable to the present invention (referring to, as Ripka ﹠amp; Rich, Curr.Op.Chem.Biol.2,441-452,1998; People such as Hruby, Curr.Op.Chem.Biol.1,114-119,1997; Hruby ﹠amp; Balse, Curr.Med.Chem.9,945-970,2000).The peptide backbone of a class in the stand-in between the simulation atom, also simulation is the skeleton of non-peptide partly or entirely, and comprises the side group of the functionality of same simulation natural amino acid residue side group.Known in the art in the structure of peptide mimics of tolerance proteolytic enzyme, usually in order to the chemical bond of the several types that replaces peptide bond, as ester, thioesters, thioamides, contrary acid amides, reductive carbonyl, dimethylene and kadsura longepedunculata ketonic bond.Another kind of peptide mimics comprises in conjunction with another peptide or proteinic non-peptide small molecules, but described non-peptide small molecules is not the structural simulation thing of the native peptides of necessity.Occur in the formation of another class peptide mimics from combinatorial chemistry and a large amount of chemicals library.These generally include novel template, though described template is uncorrelated with the native peptides structure, thereby have effect (the above-mentioned Ripka﹠amp that the necessary functional group who is positioned on the non-peptide framework plays original peptide " form " stand-in; Rich, 1998).Tetrapeptide stand-in of the present invention are open and claimed types in people's such as Shi No. the 6th, 992,063, the United States Patent (USP).

Proved the apoptosis that peptide or its stand-in in conjunction with IAP can strengthen cell according to the present invention.

Preferred core is in conjunction with the stand-in of IAP part.Stand-in described herein are suitably little, and since characterized well with IAP in conjunction with the relevant constitutional features of chase, so can synthesize a large amount of simulated compounds.The added benefit of the compound of this size comprises that having improved the solubleness in the aqueous solution more easily is delivered to selected target spot in vivo with making.

In one embodiment, by use other side chains to the amino acid of one or more naturally occurring 20 genes encodings or D amino acid side chain replace modify the present invention in conjunction with the peptide of IAP to generate peptide mimics, for example use following radicals to replace: for example alkyl, low alkyl group, 4 yuan of cycloalkyl, 5 yuan of cycloalkyl, 6 yuan of cycloalkyl to 7 yuan cycloalkyl, acid amides, acid amides low alkyl group, acid amides two (low alkyl group), lower alkoxy, hydroxyl, carboxyl and lower member ester derivatives thereof, and 4 yuan of heterocycles, 5 yuan of heterocycles, 6 yuan of heterocycle to 7 yuan heterocycles.For example, can prepare proline analogs, wherein the ring size of proline residue is by 5 yuan to 4 yuan, 6 yuan or 7 yuan of variations.Cyclic group can be saturated or unsaturated, and if undersaturated words can be fragrance or non-fragrance.Heterocyclic group can contain one or more nitrogen, oxygen and/or sulfur heteroatom.The example of these groups comprises the furazan base, imidazolidyl, imidazolyl, imidazolinyl, isothiazolyl, different  azoles base, morpholinyl (as morpholino),  azoles base, piperazinyl (as the 1-piperazinyl), piperidyl is (as piperidino, piperidino-(1-position only)), pyranyl, pyrazinyl, pyrazolidyl, pyrazolinyl, pyrazolyl, pyridazinyl, pyridyl, pyrimidyl, pyrrolidyl (as the 1-pyrrolidyl), pyrrolinyl, pyrryl, thiadiazolyl group, thiazolyl, thienyl, thio-morpholinyl (as thiomorpholine generation) and triazolyl.These heterocyclic groups can be that replace or unsubstituted.When a group was substituted, substituting group can be alkyl, alkoxyl group, halogen, oxygen or replacement or unsubstituted phenyl.Peptide mimics also can have amino-acid residue, and described amino-acid residue carries out chemically modified by phosphorylation, sulfonation, biotinylation or adding or the part of removing other.

The invention provides tertiary structure or the active compound of simulation Smac N-terminal part of simulation and IAP bonded Smac.The present invention also comprises the steric isomer of simulated compound described herein.The present invention also provides the method for using these stand-in adjusting apoptosis and being further used for therapeutic purpose.The present invention also provides intermediate and uses these intermediate preparation to regulate the method for apoptotic compound, and described compound is to regulate apoptotic by simulation and the tertiary structure of IAP bonded Smac or the N-terminal part activity of simulation Smac.

According to the present invention, provide compound of the present invention with following general formula (I):

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently, or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Or R5a and R5b randomly replace for following radicals independently: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Or randomly, R5a is connected by following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene bridge or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms are replaced by N, O or S;

R6a and R6b be independently H, tert-butoxycarbonyl, benzyloxy carbonyl, ethanoyl, trifluoroacetyl group, alkyl, low alkyl group, the optional alkyl that replaces or

Wherein R7a and R7b are H, alkyl, cycloalkyl, haloalkyl independently; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroaralkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroaralkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

M and n are 0,1,2 or 3 independently;

X and Y are O, N, S or C=C independently;

R9a, R9b, R10a, R10b are H, alkyl, the optional alkyl that replaces, aryl, heteroaryl, optional aryl, the heteroaryl that replaces independently, or R9a and R10a can be connected to form aromatic nucleus or non-aromatic ring via 4-8 optional atom such as C, N, O or the S that replaces independently or with R9b and R10b abreast;

When Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be that N, O or S replace; And R11a and R11b independently for do not exist, H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces; Or R11a and R11b form alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms are randomly replaced by N, O or S;

When Wa and Wb were not covalent attachment, Wa and Wb were H, Cl, Br, F, alkyl, CN, CO independently ₂H; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be by N, O or S replacement; Or Wa can be H, Cl, Br, F, alkyl, CN, CO ₂H, and Wb and R11a be alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be by N, O or S replacement; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

Constitute compound of the present invention and comprise Smac stand-in and intermediate thereof.The present invention includes the steric isomer (stereosiomer) of every kind of disclosed compound.Usually, compound of the present invention comprises the covalently bound homodimer of the tetrapeptide stand-in of the covalently bound dimer of tetrapeptide stand-in of tetrapeptide stand-in, Smac of Smac and Smac.Homodimer is that wherein essentially identical tetrapeptide stand-in are covalently bound those stand-in.

Experiment flow hereinafter is a flow process of announcing the disclosed compound of No.WO2004/007529 first at PCT about being used for preparing, incorporates its full content into this paper by reference herein.Based on the U.S. Provisional Application of submitting on July 15th, 2004 the 60/588th, No. 050, the U. S. application the 11/184th that is entitled as " IAP Binding Compounds " that on July 15th, 2005 submitted to, also describe suitable peptide and peptide mimics for No. 503, incorporated its full content into this specification sheets by reference herein.

As Nikolovska-Coleska, people such as Z. are described, and (Analytical Biochemistry (2004) vol.332:261-273) uses multiple fluorogenic substrate to measure the binding affinity of compound entity of the present invention and XIAP and with K _DThe value report.Briefly, (AbuRPF-K (5-Fam)-NH2) and 40nM XIAP-BIR3 mix 15min in 100L contains the 0.1M potassium phosphate buffer of pH7.5 of 100 μ g/ml bovines with tried peptide and the fluorescently-labeled peptide of 5nM of various concentration in room temperature (RT).After hatching, use 485nm exciter filter and 520nm emission spectral filter in Victor ²The last measurement of V polarization value (polarization values, mP).Use GraphPad Prism to determine IC from curve according to the non-linear least square analytical procedure ₅₀Value.The K that compound described herein obtains _DThe value scope is: K _D＜0.1 μ M (A), K _D=0.1-1 μ M (B), K _D=1-10 μ M (C), and K _D＞10 μ M (D).

Flow process I

2-(2-bromo-6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (2):

(3.2g 17.9mmol) adds to and contains 1 (5.4g, CCl 17.0mmol) with NBS ₄(50mL) in the solution.With heterogenetic reaction mixture reflux (80-85 ℃) 2h, TLC analyzes and shows that 1 consumes fully when 2h.[TLC analyzes, hexane/EtOAc of 4: 1, R _f(8)=0.4; R _f(2)=0.5].Reaction mixture is cooled to room temperature, inclines to silicagel column then.Use the EtOAc/ hexane of 10-15% that product is carried out the white solid 2 that wash-out obtains 4.4g (65%). ¹H NMR(DMSO，300MHz)δ11.74(s，1H)，7.56(m，1H)，7.02(d，J＝9.3Hz，1H)，6.88(m，1H)，3.99(m，1H)，3.22(m，2H)，2.97(m，1H)，2.58(dd，J＝13.5，9.3Hz，1H)，1.9-1.5(4H)，1.40(s，9H)ppm。

1,4-two-[2-(6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester] benzene (4):

(3.3g, the solution of toluene 8.3mmol) (25mL), EtOH (25mL) and water (1mL) outgases to containing 2 under high vacuum.Add K ₂CO ₃(4.5g, 32.5mmol), 3 (0.97g, 5.8mmol) and (Ph ₃P) ₄(0.29g 0.25mmol) and with the mixture of gained stirs 5h at 100 ℃ to Pd.[TLC analyzes, hexane/EtOAc of 4: 1, R _f(2)=0.5; R _f(4)=0.3].Reaction mixture filters and uses 5% EtOAc/ hexane wash by short silicagel pad.Concentrated filtrate also obtains the linen high fluorescent solid 4 of 3.0g (98%) via fast silica gel chromatogram (20% EtOAc/ hexane) purifying crude product. ¹H NMR(CDCl ₃，300MHz)δ8.6-8.4(m，2H)，7.65(m，2H)，7.57(brs，4H)，7.05(m，2H)，7.90(m，2H)，4.22(br s，2H)，3.4-3.1(m，6H)，2.90(m，2H)，1.8-1.3(m，26H)ppm。

Flow process II

1,4-two-2-[1-(2-acetoxyl group-ethyl)-6-fluoro-1H-indol-3-yl methyl]-tetramethyleneimine-1-carboxylic acid uncle Butyl ester } benzene (6):

(3.0g, DMF 4.2mmol) (10mL) solution add to 60%NaH, and (0.67g is in dry DMF 17.0mmol) (10mL) suspension with 4 at 0 ℃.Stirred reaction mixture 1h and then be cooled to 0 ℃ at room temperature.(2.8g, DMF 16.8mmol) (5mL) solution adds to reaction mixture and remove ice bath after adding will to contain 5.At room temperature behind the 2h, LC/MS and TLC analyze and show that 4 consume fully.[TLC analyzes, hexane/EtOAc of 2: 1, R _f(4)=0.4; R _f(6)=0.8].Reaction mixture is cooled to 0 ℃ and add NH ₄The saturated aqueous solution of Cl.Use the diethyl ether extraction product.Ethereal extract makes water, salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain the pale solid 6 of 1.4g by NP-HPLC (silica gel, the EtOAc/ hexane of 10-100% is through 30min) purifying crude product. ¹H NMR(CDCl ₃，300MHz)δ7.68(m，2H)，7.54(s，4H)，7.12(m，2H)，6.94(m，2H)，4.25(m，4H)，4.14(m，6H)，3.4-3.1(6H)，2.60(dd，J＝9.6，13.8Hz，2H)，1.90(s，6H)，1.83(m，2H)，1.7-1.3(m，24H)ppm。

Flow process III

Acetate 2-(2-{4-[1-(2-acetoxyl group-ethyl)-6-fluoro-3-tetramethyleneimine-2-ylmethyl-1H-indoles-2-yl]- Phenyl }-6-fluoro-3-tetramethyleneimine-2-ylmethyl-indol-1-yl)-ethyl ester (7):

(1.4g, DCM 1.58mmol) (20mL) solution is cooled to 0 ℃ will to contain 6.Add TFA (5mL) via transfer pipet, and make reactant be warming up to room temperature and it is monitored analyze up to TLC and to show and 6 consume fully (～2h).TLC analyzes, 10% MeOH/DCM, R _f(6)=0.7; R _f(7)=0.2.Remove in rotatory evaporator and to desolvate and residue is dissolved among the EtOAc.Use saturated NaHCO ₃Twice of solution washing EtOAc solution also uses the salt water washing once.The aqueous cleaning solution that uses EtOAc to be combined strip and with organic extract through anhydrous Na ₂The SO4 drying is filtered and the concentrated yellow solid 7 that obtains 1.2g (amount), and this yellow solid 7 uses under situation about not being further purified. ¹H NMR(CDC13，300MHz)δ8.05(dd，J＝8.4，5.4Hz，2H)，7.56(s，4H)，7.13(dd，J＝9.9，2.4Hz，2H)，6.99(m，2H)，4.60(d，J＝9.9Hz，2H)，4.51(m，2H)，4.26(m，4H)，4.15(m，4H)，3.63(m，2H)，3.54(m，2H)，3.5-3.3(m，4H)，2.41(m，2H)，1.89(s，6H)，1.8-1.5(m，6H)，1.43(s，18H)，1.09(s，18H)ppm。

1,4-two-{ acetate 2-{3-[1-(uncle 2--butoxy carbonyl amino-3,3-dimethyl-butyryl radicals)-tetramethyleneimine-2-base Methyl]-6-fluoro-indoles-1-yl }-ethyl ester } benzene (8):

To contain uncle Boc-L--Leu-OH (0.82g, 3.54mmol) and HATU (1.41g, anhydrous NMP (15mL) solution 3.70mmol) is cooled to 0 ℃.Behind the 15min, via syringe add the N-methylmorpholine (0.46g, 0.5mL, 4.54mmol).Behind the 15min, add and to contain 7 (1.10g, (10mL) solution of DCM 1.61mmol) also makes reaction mixture be warming up to room temperature through 16h, and when 16h, TLC analyzes and shows that 7 consume that [TLC analyzes, hexane/EtOAc of 2: 1, R fully _f(7)=0.01; R _f(8)=0.8].Use the diethyl ether diluted reaction mixture and with rare HCl solution washing once, water washing five times to be to remove excessive N MP, uses saturated NaHCO ₃The aqueous solution and salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain the pale solid 8 of 1.3g (73%) by NP-HPLC (silica gel, the EtOAc/ hexane of 10-100% is through 30min) purifying crude product. ¹H NMR(CDCl ₃，300MHz)δ8.05(dd，J＝5.4，8.4 Hz，2H)，7.56(s，4H)，7.11(dd，J＝2.4，9.9Hz，2H)，6.98(m，2H)，5.43(d，J＝9.9Hz，2H)，4.51(m，2H)，4.26(m，6H)，4.17(m，6H)，3.2-3.7(m，8H)，2.41(dd，J＝12，13Hz，2H)，1.88(s，6H)，1.7-1.5(m，4H)，1.43(s，18H)，1.04(s，18H)ppm。

Flow process IV

Acetate 2-{2-(4-{1-(2-acetoxyl group-ethyl)-3-[1-(2-amino-3,3-dimethyl-butyryl radicals)-pyrroles Alkane-2-ylmethyl]-6-fluoro-1H-indoles-2-yl }-phenyl)-3-[1-(2-amino-3,3-dimethyl-butyryl radicals)- Tetramethyleneimine-2-ylmethyl]-6-fluoro-indoles-1-yl }-ethyl ester (9):

(1.3g, DCM 1.17mmol) (5mL) solution is cooled to 0 ℃ will to contain 8.Add the DCM (25mL) of 20%TFA via transfer pipet, and make reactant be warming up to room temperature and it is monitored analyze up to TLC and to show and 8 consume fully (～2h).TLC analyzes, 10% MeOH/DCM, R _f(8)=0.7; R _f(9)=0.3.Remove in rotatory evaporator and to desolvate and by RP-HPLC (method: solvent orange 2 A: water w/0.1%v/v HOAc, solvent B:ACN w/0.1%v/v HOAc, Dynamax MicrosorbC18 60  8 μ, 41.4mm * 25cm; Flow: 40mL/min; Detector: 254nm) purifying residue.Collection contains the cut of product and uses saturated NaHCO ₃Aqueous solution neutralization.Use the EtOAc extraction product and use salt water washing organic extract, through anhydrous Na ₂SO ₄The pale solid 9 that obtains 0.80g (75%) is filtered and concentrated to drying. ¹H NMR(CDCl ₃，300MHz)δ8.09(dd，J＝5.1，8.7Hz，2H)，7.51(s，4H)，7.13(m，2H)，7.0(m，2H)，4.41(m，2H)，4.25(m，4H)，4.16(m，4H)，3.6-3.0(m，6H)，2.86(m，2H)，2.39(m，2H)，1.91(s，6H)，1.8-1.4(m，12H)，1.04(s，18H)ppm。

1,4-two-acetate 2-[3-(1-{2-[2-(uncle-butoxy carbonyl-methyl-amino)-propionamido]-3, the 3-diformazan Base-butyryl radicals }-tetramethyleneimine-2-ylmethyl)-6-fluoro-indoles-1-yl]-ethyl ester } benzene (10):

To contain Boc-L-N (Me) Ala-OH (0.27g, 1.32mmol) and HATU (0.54g, anhydrous NMP (15mL) solution 1.43mmol) is cooled to 0 ℃.Behind the 15min, via syringe add the N-methylmorpholine (0.17g, 0.2mL, 1.68mmol).Behind the 15min, add and to contain 9 (0.50g, (10mL) solution of DCM 0.55mmol) also makes reaction mixture be warming up to room temperature through 16h, and when 16h, TLC analyzes and shows that 9 consume that [TLC analyzes, hexane/EtOAc of 3: 2, R fully _f(9)=0.01; R _f(10)=0.5].Use the diethyl ether diluted reaction mixture and with rare HCl solution washing once, water washing five times to be to remove excessive N MP, uses saturated NaHCO ₃The aqueous solution and salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain the pale solid 10 of 0.64g (91%) by NP-HPLC (silica gel, the EtOAc/ hexane of 10-100% is through 30min) purifying crude product. ¹H NMR(CDCl ₃，300MHz)δ8.05(m，2H)，7.58(brs，4H)，7.13(m，2H)，6.97(m，2H)，4.75(m，2H)，4.60(d，J＝9.3Hz，2H)，4.50(m，2H)，4.25(m，4H)，4.16(m，4H)，3.70(m，2H)，3.57(m，2H)，3.5-3.2(m，4H)，2.85(brs，6H)，2.42(m，2H)，1.88(s，6H)，1.8-1.4(m，8H)，1.52(s，18H)，1.33(m，6H)，1.04(brs，18H)ppm。

Flow process V

1,4-two-acetate 2-(3-{1-[3,3-dimethyl-2-(2-methylamino--propionamido)-butyryl radicals]-tetramethyleneimine-2- Ylmethyl }-6-fluoro-indoles-1-yl)-ethyl ester } benzene (11):

(0.64g, DCM 0.5mmol) (20mL) solution is cooled to 0 ℃ will to contain 10.Add TFA (5mL) via transfer pipet, and make reactant be warming up to room temperature and it is monitored analyze up to TLC and to show and 10 consume fully (～2h).Remove in rotatory evaporator and to desolvate and by RP-HPLC (method: solvent orange 2 A: water w/0.1%v/v HOAc, solvent B:ACN w/0.1%v/v HOAc, Dynamax Microsorb C18 60  8 μ, 41.4mm * 25cm; Flow: 40mL/min; Detector: 254nm) purifying residue.Collection contains the cut of product and uses saturated NaHCO ₃Aqueous solution neutralization.Product uses EtOAc to extract and use salt water washing organic extract, through anhydrous Na ₂SO ₄The pale solid 11 that obtains 0.50g (93%) is filtered and concentrated to drying. ¹H NMR(CDCl ₃，300MHz)δ8.04(m，2H)，7.83(d，J＝9.3Hz，2H)，7.55(m，4H)，7.12(m，2H)，6.99(m，2H)，4.60(d，J＝9.3Hz，2H)，4.57(m，2H)，4.24(m，4H)，3.73(m，2H)，3.55(m，2H)，3.41(m，2H)，3.30(m，2H)，3.08(m，2H)，2.40(s，6H)，2.38(m，2H)，1.87(s，6H)，1.8-1.3(m，16H)，1.04(br s，18H)ppm。

1,4-two-N-(1-{2-[6-fluoro-1-(2-hydroxyl-ethyl)-1H-indol-3-yl methyl]-tetramethyleneimine-1-carbonyl Base }-2,2-dimethyl-propyl group)-2-methylamino--propionic acid amide } benzene (12):

At 0 ℃ NaOH (1M, 5mL, the excessive) aqueous solution added to and to contain 11 (0.48g is in MeOH 0.44mmol) (5mL) solution.After the adding, remove ice bath and at room temperature reaction mixture is stirred 1h.Make water/EtOAc diluted reaction mixture and layering.Organic phase is used the salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.By RP-HPLC (method: solvent orange 2 A: water w/0.1%v/v HOAc, solvent B:ACN w/0.1%v/v HOAc, Dynamax Microsorb C18 60  8 μ, 41.4mm * 25cm; Flow: 40mL/min; Detector: 254nm) purifying residue.Collection contains the cut of product, and freezing and freeze-drying obtains the cotton-shaped white solid 12 of 0.19g. ¹H NMR(CDCl ₃，300MHz)δppm。 ¹³C NMR(CDCl ₃，75MHz)δ7.8-7.4(m，8H)，7.11(m，2H)，6.95(m，2H)，4.57(d，J＝9.3Hz，2H)，4.4-4.0(m，6H)，3.8-3.4(m，8H)，3.2-3.0(m，3H)，2.6-2.4(m，14H)，2.38(m，6H)，2.2-1.5(m，12H)，1.29(d，J＝6.9Hz，6H)，1.00(s，18H)ppm。

Embodiment:

Embodiment 1

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroaralkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroaralkyl are randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroaralkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps, in some cases, R5a is connected via following radicals with the R5b residue: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene bridge or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

R12a, R12b, R13a, R13b, R14a and R14b are H, Cl, Br, F, alkyl, cycloalkyl, hydroxyl, alkoxyl group, amino, alkylamino, cyano group or CO independently ₂H;

R3a and R3b are H, halogen, alkyl, aryl, aralkyl, amino, virtue amino, aryl alkyl amino, hydroxyl, alkoxyl group, aryloxy, aralkyl hydroxyl, dialkylamino, amide group, sulfonamido or amidino groups independently;

X and Y are O, N, S or C=C independently;

R11a and R11b do not exist or are H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces independently; Or R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms can be replaced by N, O or S;

Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, heteroaryl or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S;

Numbering	R8a	R7a	R5a	^＊) stereochemistry of position	X	Y	R11	Wa-Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,
Numbering	R8a	R7a	R5a	^＊) stereochemistry of position	X	Y	R11	Wa-Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,	1	H	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	H	H	A
2	H	S-Me	S-iPr	S	O	O	na	Instead-(CH=CH)	H	H	S-iPr	S-Me	H	H	H	H	A	1	H	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	H	H	A
2	H	S-Me	S-iPr	S	O	O	na	Instead-(CH=CH)	H	H	S-iPr	S-Me	H	H	H	H	A	3	H	S-Me	S-iPr	S	O	O	na	CH2CH2	H	H	S-iPr	S-Me	H	H	H	H	A
4	H	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	H	H	H	H	H	A	3	H	S-Me	S-iPr	S	O	O	na	CH2CH2	H	H	S-iPr	S-Me	H	H	H	H	A
4	H	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	H	H	H	H	H	A	6	H	S-Me	S-iPr	S	N	N	NH	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	H	H	A
7	H	S-Me	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Me	H	H	H	H	A	6	H	S-Me	S-iPr	S	N	N	NH	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	H	H	A
7	H	S-Me	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Me	H	H	H	H	A	8	Me	S-Me	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	H	H	A
9	H	S-Et	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Et	H	H	H	H	B	8	Me	S-Me	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	H	H	A
9	H	S-Et	S-tBu	S	O	O	na	1, the 4-phenyl	H	H	S-tBu	S-Et	H	H	H	H	B	10	Me	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	H	H	A
11	H	S-Et	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Et	H	H	H	H	A	10	Me	S-Me	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	H	H	A
11	H	S-Et	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	S-Et	H	H	H	H	A	12	H	S-Me	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	H	H	A
13	Me	S-Me	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	H	H	A	12	H	S-Me	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	H	H	A
13	Me	S-Me	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	H	H	A	14	H	S-Et	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Et	H	H	H	H	B
15	Me	S-Me	S-(2R-Et OH)	S	O	O	na	1, the 4-phenyl	H	H	S-(2R- EtOH )	S-Me	Me	H	H	H	A	14	H	S-Et	S-cHex	S	O	O	na	1, the 4-phenyl	H	H	S- cHex	S-Et	H	H	H	H	B
15	Me	S-Me	S-(2R-Et OH)	S	O	O	na	1, the 4-phenyl	H	H	S-(2R- EtOH )	S-Me	Me	H	H	H	A	16	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
17	H	S-Me	S-iPr	S	N	N	H	2, the 5-thiophene	H	H	S-iPr	S-Me	H	H	F	H	A	16	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
17	H	S-Me	S-iPr	S	N	N	H	2, the 5-thiophene	H	H	S-iPr	S-Me	H	H	F	H	A	18	Me	S-Me	S-cHex	S	N	N	H	2, the 5-thiophene	H	H	S- cHex	S-Me	Me	H	F	H	A
19	H	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	F	H	A	18	Me	S-Me	S-cHex	S	N	N	H	2, the 5-thiophene	H	H	S- cHex	S-Me	Me	H	F	H	A
19	H	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	F	H	A	20	Me	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	F	H	A
21	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-iPr	S-Me	Me	H	H	H	B	20	Me	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	F	H	A
21	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-iPr	S-Me	Me	H	H	H	B	22	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-tBu	S- Me	Me	H	H	H	B
23	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A	22	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-tBu	S- Me	Me	H	H	H	B
23	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A	24	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	H	H	A
25	Me	S-Me	S-(2R-Et OBn)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A	24	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	H	H	A
25	Me	S-Me	S-(2R-Et OBn)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A	26	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A
27	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOH	S-Me	Me	H	F	H	A	26	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A

											)
											)							28	Me	S-Me	S-(2R-Et OH)	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-(2R- EtOH )	S-Me	Me	H	F	H	A
29	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-iPr	S-Me	Me	H	F	H	A	28	Me	S-Me	S-(2R-Et OH)	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-(2R- EtOH )	S-Me	Me	H	F	H	A
29	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-iPr	S-Me	Me	H	F	H	A	30	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	F	H	A
31	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOH )	S-Me	Me	H	F	H	A	30	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	F	H	A
31	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOH )	S-Me	Me	H	F	H	A	32	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A
33	Me	S-Me	S-(2R-Et OBn)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A	32	Me	S-Me	S-(2R-Et OH)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A
33	Me	S-Me	S-(2R-Et OBn)	S	N	N	H	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A	34	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	H	H	A
35	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A	34	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	H	H	H	A
35	Me	S-Me	S-iPr	S	N	N	H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A	36	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-tBu	S-Me	Me	H	H	H	B
37	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-iPr	S-Me	Me	H	H	H	B	36	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-tBu	S-Me	Me	H	H	H	B
37	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	R-OH	R-OH	S-iPr	S-Me	Me	H	H	H	B	38	Me	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	F	H	A
39	H	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	F	H	A	38	Me	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	Me	H	F	H	A
39	H	S-Me	S-cHex	S	N	N	H	1, the 4-phenyl	H	H	S- cHex	S-Me	H	H	F	H	A	40	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-iPr	S-Me	Me	F	H	H	B
41	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	F	H	H	A	40	Me	S-Me	S-iPr	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-iPr	S-Me	Me	F	H	H	B
41	Me	S-Me	S-tBu	R	N	N	H	1, the 4-phenyl	S-OH	S-OH	S-tBu	S-Me	Me	F	H	H	A	42	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A
43	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	42	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R- EtOB n)	S-Me	Me	H	F	H	A
43	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	44	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A
45	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OAc	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	44	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A
45	H	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OAc	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	46	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
47	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OAc	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A	46	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A

487	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
487	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A	49	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OAc	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
50	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	49	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OAc	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
50	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	51	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
52	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A	51	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
52	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A	53	H	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	F	H	A
54	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	F	H	H	A	53	H	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OMe	1, the 4-phenyl	H	H	S-iPr	S-Me	H	H	F	H	A
54	Me	S-Me	S-tBu	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	F	H	H	A	55	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A
56	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	F	H	H	A	55	Me	S-Me	S-iPr	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	F	H	H	A
56	Me	S-Me	S-(2R-Et OH)	S	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	F	H	H	A	57	Me	S-Me	S-iPr	S	N	N	Me	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
58	Me	S-Me	S-(2R-Et OH)	S	N	N	Me	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	57	Me	S-Me	S-iPr	S	N	N	Me	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
58	Me	S-Me	S-(2R-Et OH)	S	N	N	Me	1, the 4-phenyl	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A	59	Me	S-Me	S-tBu	S	N	N	Me	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
60	Me	R-Me	R-tBu	R	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	R-tBu	R- Me	Me	H	F	H	C	59	Me	S-Me	S-tBu	S	N	N	Me	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
60	Me	R-Me	R-tBu	R	N	N	CH ₂ CH ₂ OH	1, the 4-phenyl	H	H	R-tBu	R- Me	Me	H	F	H	C	61	H	R-Me	R-iPr	R	O	O	na	1, the 4-phenyl	H	H	R-iPr	R- Me	H	H	H	H	D

Additional embodiments:

Embodiment 2

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl are randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps, in some cases, R5a is connected via following radicals with the R5b residue: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

X and Y are O, N, S or C=C independently;

R11a and R11b independently for do not exist, H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces; Or R11a and R11b form alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms can be replaced by N, O or S;

Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	X	Y	R	Wa-Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,
Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	X	Y	R	Wa-Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,	62	H	H	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	H	H	H	H	H	B
62	H	H	S-tBu	S	N	N	CH ₂CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	H	H	H	F	H	B	62	H	H	S-iPr	S	O	O	na	1, the 4-phenyl	H	H	S-iPr	H	H	H	H	H	B
62	H	H	S-tBu	S	N	N	CH ₂CH ₂ OH	1, the 4-phenyl	H	H	S-tBu	H	H	H	F	H	B	63	H	Me	S-tBu	S	N	N	CH ₂CH ₂ OAc	1, the 4-phenyl	H	H	S-tBu	Me	H	H	F	H	B

Additional embodiments:

Embodiment 3

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein each quilt of alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl is randomly replaced for following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps, in some cases, R5a is connected via following radicals with the R5b residue: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene bridge or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

X is O, N, S or C=C;

Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, heteroaryl or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S.

Numbering

R8a

R7a

R5a

( ^＊) of putting of standing body

X

R11a

Wa-Wb

R3a

R3b

R5b

R7b

R8b

R12

R13

R14

K _DScope

64	Me	S- Me	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
64	Me	S- Me	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A	65	H	H	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	H	H	H	F	H	B
66	Me	S- Me	S- tBu	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A	65	H	H	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	H	H	H	F	H	B
66	Me	S- Me	S- tBu	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A	67	Me	S- Me	S- iPr	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
68	H	H	S- iPr	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-iPr	H	H	H	F	H	C	67	Me	S- Me	S- iPr	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-iPr	S-Me	Me	H	F	H	A
68	H	H	S- iPr	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-iPr	H	H	H	F	H	C	69	Me	S- Me	S- tBu	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
70	Me	R- Me	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	R-Me	Me	H	F	H	A	69	Me	S- Me	S- tBu	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-tBu	S-Me	Me	H	F	H	A
70	Me	R- Me	S- iPr	S	N	CH ₂CH ₂O Ac	1, the 4-phenyl	H	H	S-iPr	R-Me	Me	H	F	H	A	71	Me	R- Me	S- tBu	S	N	CH ₂CH ₂O H	1, the 4-phenyl	H	H	S-tBu	R-Me	Me	H	F	H	B

Flow process VI

Referring to: Macor, J.E.; Blank, D.H.; Post, R.J.; Ryan, K.Tetrahedron Lett.1992,33 (52), 8011-8014.

2-(2-ethoxycarbonyl-vinyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (14):

(130mL g, DCM 0.26mol) (250mL) solution pack into and are provided with in the 2L 3 neck round-bottomed flasks of top stirring and nitrogen inlet with oxalyl chloride.This solution is cooled to-78 ℃.Dropwise add DMSO (20mL, DCM 0.28mol) (30mL) solution.Behind the 30min, dropwise add alcohol 13 (40g, DCM 0.20mol) (200mL) solution.Behind the 30min, (140mL 1.00mol) adds to this solution with TEA.Shifting this solution to ice/water-bath (0 ℃) also continues to stir 30min[NB: reaction mixture is stiff white soup compound].TLC analyzes and shows that starting material does not have residue [hexane/EtOAc of 1: 1, Rf (13)=0.4; R _f(acetaldehyde)=0.6].Reaction mixture uses DCM (200mL) dilution and uses H in succession ₂O, 1M HCl, saturated NaHCO ₃With the salt water washing.The DCM layer is through Na2SO ₄Drying is filtered and the concentrated oily matter crude product (40g) that obtains 2-formyl-tetramethyleneimine-1-carboxylic acid tert-butyl ester, and this crude product uses under situation about not being further purified. ¹H NMR(CDCl ₃，300MHz)δ9.50(d，J＝24Hz，1H)，4.20-4.03(m，1H)，3.60-3.40(m，2H)，2.20-1.87(m，4H)，1.43(s，9H)ppm。

With NaH (60%, 10.0g, 0.25mol) and anhydrous THF (200mL) pack into be provided with that the top is stirred and the 2L 3 neck round-bottomed flasks of nitrogen inlet in.(53.8g 0.24mol) added to stirred mixture lentamente with triethyl phosphine acetate through 20 minutes.Dropwise add 2-formyl-tetramethyleneimine-1-carboxylic acid tert-butyl ester crude product (40g, THF 0.20mol) (75mL) solution.Solution becomes orange, and continues to stir 1h up to analyzing [hexane/EtOAc of 1: 1, R by TLC _f(acetaldehyde)=0.6; R _f(14)=0.8] show no acetaldehyde residue.Use EtOAc and salt solution to dilute this solution and layering.The EtOAc layer is with 1M HCl, salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated and obtains yellow oil 14 (67g), and oily matter 14 uses under situation about not being further purified. ¹H NMR(CDCl ₃，300 MHz)δ6.92-6.76(m，1H)，5.82(d，1H)，4.56-4.32(m，1H)，4.25-4.12(m，2H)，3.48-3.27(m，2H)，2.20-1.98(m，1H)，1.91-1.72(m，2H)，1.43(s，9H)，1.25(t，3H)ppm。

2-(3-hydroxyl-propenyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (15):

With 14 (67g, 0.20mol) and DCM (400mL) pack into and be provided with in the 2L 3 neck round-bottomed flasks that stir the top.This solution is cooled to-78 ℃.(30mL 0.20mol) adds this solution lentamente with boron trifluoride ethyl ether complex.Stirred reaction mixture 30min.With suitable speed add DIBAL (be 1M in DCM, 600mL, 0.6mol).This solution is stirred 2h and with after 30min uses EtOAc (100mL) to handle to remove remaining reagent at-78 ℃.Make reaction mixture be warming up to-5 ℃.Make the reaction mixture cancellation carefully by dropwise adding 1M HCl.Use DCM and H ₂The O diluted reaction mixture also makes it become acid with dissolved aluminum salt.Use rare HCl aqueous solution, water and salt water washing organic phase after the layering in succession.The DCM layer is through Na ₂SO ₄Drying is filtered and is concentrated.By flash chromatography (SiO ₂, the EtOAc/ hexane of 25%-80%) and the purifying residue obtains yellow oil 15 (36g, 79%).[TLC analyzes, hexane/EtOAc of 1: 1, R _f(14)=0.8; R _f(15)=0.2]. ¹H NMR(CDCl ₃，300 MHz)δ5.73-5.52(m，2H)，4.39-4.16(m，1H)，4.15-4.04(m，2H)，3.46-3.25(m，2H)，2.92(brs，1H)，2.08-1.93(m，1H)，1.92-1.79(m，2H)，1.78-1.62(m，1H)，1.42(s，9H)ppm。

Instead-2S-(3-sulfonyloxy methyl oxygen-propenyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (16):

((19g is in DCM 84mmol) (100mL) solution 100mmol) to add to 15 for 10g, 13.9mL with triethylamine.This solution is cooled to 0 ℃ and dropwise add methylsulfonyl chloride (9.6g, 6.5mL is among DCM 84mmol) (20mL).Behind the 1h, TLC analyzes and shows that 15 consume [hexane/EtOAc of 1: 1, R fully _f(15)=0.2; R _f(16)=0.6].Add salt solution and use DCM (3 * 50mL) extraction products.Merge organic extract and use 1N HCl, water, salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated and obtains 16,16 of 21.4g and use under the situation of purifying not having. ¹H NMR(CDCl ₃，300 MHz)δ4.4-4.0(m，2H)，3.42-3.21(m，3H)，3.0(s，3H)，2.00-1.6(m，4H)，1.42(s，9H)ppm。

Flow process VII

2-{3-[ethanoyl-(2-bromo-5-fluoro-phenyl)-amino]-propenyl }-tetramethyleneimine-1-carboxylic acid tert-butyl ester (18):

0 ℃ with 2-bromo-5-fluoroacetanilide (17,53.4g, (9.2g is in dry DMF 0.23mol) (150mL) suspension 0.23mol) to add to 60%NaH with aliquot.Behind the 1h, add DMF (20mL) solution of the crude product (about 0.19mol) of mesylate 16 from feed hopper in mode dropwise.Reaction mixture is spent the night be warming up to room temperature.Reaction mixture is cooled to 0 ℃ once more, and makes its cancellation and neutralize up to pH=7 via adding rare HCl aqueous solution by adding salt solution carefully.Use diethyl ether and water diluted mixture thing and layering.Organic phase washes with water several times to remove DMF, uses the salt water washing subsequently, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain 66g oily matter 18 by fast silica gel chromatogram method (MeOH/DCM of 0.5%-2%) purifying crude product.[TLC analyzes, hexane/EtOAc of 1: 1, R _f(16)=0.5; R _f(17)=0.6; R _f(18)=0.4]. ¹H NMR(CDCl ₃，300 MHz)δ7.64(m，1H)，7.01(m，2H)，5.52(m，1H)，5.39(app dd，J＝6.0，15.3Hz，1H)，4.77(app dd，J＝4.5，13.8Hz，1H)，4.24(m，1H)，3.67(app dd，J＝7.5，13.8Hz，1H)，3.32(m，2H)，1.90(m，1H)，1.81(m，3H)，1.75(m，2H)，1.57(m，1H)，1.43(m，9H)ppm。

2-(1-ethanoyl-6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (19):

Under nitrogen atmosphere, in room temperature with (n-Bu) ₄NCl (41.5g, 0.15mol), K ₂CO ₃(20.6g, 0.15mol), NaHCO ₂(10.2g, 0.15mol) and Pd (OAc) ₂(3.35g, (66g is in dry DMF 0.15mol) (350mL) solution 0.015mol) to add to 18.Uneven mixture is immersed (85 ℃) oil bath of preheating.Behind the 1h, TLC analyze to show residue some 18, therefore add more catalyzer (1g).1.5h after, add other catalyzer (0.6g).Adding post-heating 1.5h, analyzing [TLC analysis, 2% MeOH/DCM, R by TLC _f(18)=0.7; R _f(19)=0.8] prove that 18 have consumed fully.The reaction mixture of heat is transferred to the ice-water bath cooling, filters with the diethyl ether dilution and by Celite (celite) pad then.Solid uses diethyl ether washing and water that filtrate is washed several times to remove DMF, uses the salt water washing then once, through anhydrous Na ₂SO ₄Drying is filtered and the concentrated crude product 19 that obtains 52.5g, and crude product 19 uses under situation about not being further purified. ¹H NMR(CDCl ₃，300MHz)δ8.18(m，1H)，7.60(m，1H)，7.18(m，1H)，7.05(dt，J＝2.4，8.7Hz，1H)，4.13(m，1H)，3.41(m，1H)，3.33(m，2H)，3.17(app dd，J＝14.1，38.1Hz，1H)，2.61(s，3H)，1.83(m，3H)，1.69(m，1H)，1.49(s，9H)ppm。

2-(6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester (20):

SILVER REAGENT MeOH (480mL) solution that will contain crude product 19 (48g) is cooled to 0 ℃.Add a part of NaOH (1M, 144mL) aqueous solution.Behind the 30min, TLC analyzes and shows that [TLC analyzes, hexane/EtOAc of 3: 2, R fully in starting material consumption _f(19)=0.7; R _f(20)=0.8].Use 1NHCl neutralization reaction mixture and use the DCM extraction product.The DCM extract makes water, salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Crude product is adsorbed on the silica gel of 200mL and adopts chromatography (hexane of 80%-65%/EtOAc) obtains the dense thick oily matter 20 of 31.7g. ¹H NMR(CDCl ₃，300 MHz)δ8.11(brs，1H)，7.65-7.57(m，1H)，7.04(m，1H)，6.96(s，1H)，6.87(t，J＝2.8Hz，1H)，4.16-4.09(m，1H)，3.45-3.14(m，3H)，2.76-2.63(m，1H)，1.75(brs，4H)，1.58(s，9H)ppm。

Flow process VIII

2-{1-[2-(tert-butyl-dimethyl-silanyloxy)-ethyl]-6-fluoro-1H-indol-3-yl methyl }-tetramethyleneimine -1-carboxylic acid tert-butyl ester (22):

At 0 ℃, under nitrogen atmosphere, (3.0g, dry DMF 9.42mmol) (40mL) solution add 60%NaH, and (0.45g is in DMF 11.3mmol) (10mL) mixture with 20 via feed hopper.Behind the 1h, via syringe be incorporated in bromide 21 among the DMF (5mL) (2.47g, 2.22mL, 10.3mmol).Behind the 30min, make reaction mixture be warming up to room temperature and the other 30min of stirring.By adding saturated NH ₄The Cl aqueous solution and dilute with water come the cancellation reaction.Product uses diethyl ether extraction and water that the ethereal extract that merges is washed several times to remove DMF, uses the salt water washing, through anhydrous Na ₂SO ₄Drying is filtered and the concentrated yellow oil 22 that obtains 4.49g (amount), and yellow oil 22 is used under situation about not being further purified.TLC analyzes [hexane/EtOAc of 3: 1, R _f(20)=0.4; R _f(22)=0.7]. ¹H NMR(CDCl ₃，300MHz)δ7.68(m，1H)，7.12(d，J＝3.3Hz，1H)，7.03(s，1H)，6.98(t，J＝3.2Hz，1H)，4.26-4.23(m，3H)，4.05-3.99(m，2H)，3.55-3.27(m，3H)，2.75(m，1H)，1.88(brs，4H)，1.67(s，9H)，1.33(m，1H)，1.06-1.00(m，3H)，0.95(s，9H)，0.23-0.14(m，2H)ppm。

2-[6-fluoro-1-(2-hydroxyl-ethyl)-1H-indol-3-yl methyl]-tetramethyleneimine-1-carboxylic acid tert-butyl ester (23):

(4.49g, anhydrous THF (50mL) solution 9.42mmol) is cooled to 0 ℃ will to contain 22.Via syringe add the tetra-n-butyl Neutral ammonium fluoride (1M in THF, 14mL, 14mmol).Behind the 1h, analyze [hexane/EtOAc of 3: 1, R by TLC _f(22)=0.7; R _f(23)=0.1] proved response is complete, therefore uses the EtOAc diluting reaction.Use 1M HCl, water, salt solution with EtOAc solution washing twice, through anhydrous Na ₂SO ₄The tawny oily matter 23 (＞100% that obtains 3.9g is filtered and concentrated to drying; Contained the contaminating impurity of TBS by some), tawny oily matter 23 uses under situation about not being further purified. ¹H NM[R(CDCl ₃，300MHz)δ7.59(brs，1H)，7.01-6.85(m，3H)，4.19-4.10(m，3H)，3.90(brs，2H)，3.38-3.31(m，2H)，3.15(dd，J＝1.4，4.6Hz，1H)，2.68(m，1H)，1.79-1.72(m，4H)，1.47(d，J＝10.9Hz，9H)ppm。

2-{6-fluoro-1-[2-(toluene-4-sulphonyl oxygen)-ethyl]-1H-indol-3-yl methyl }-tetramethyleneimine-1-carboxylic acid uncle Butyl ester (12):

0 ℃ with triethylamine (1.13g, 1.56mL 11.2mmol) add to 23 (3.4g, in anhydrous DCM (50mL) solution 9.38mmol), add subsequently p-TsCl (1.79g, 9.38mmol) and DMAP (0.12g, 0.94mmol).Behind the 30min, make reaction mixture rise to room temperature.In case 23 consumption (at room temperature～30min), are used the DCM diluted reaction mixture and are used 1M HCl, salt solution washed twice, through anhydrous Na fully ₂SO ₄Drying is filtered and is concentrated.(3: 1 hexane/EtOAc) purifying tosylate crude product obtains the white foam 24 of 3.67g (76%), and this white foam 24 is analyzed through TLC and is uniform material [hexane/EtOAc of 3: 1, R by the fast silica gel chromatogram method _f(23)=0.1; R _f(24)=0.3]. ¹H NMR(CDCl ₃，300MHz)δ7.64-7.45(m，3H)，7.11(t，J＝2.5Hz，2H)，6.85(dd，J＝0.8，3.3Hz，1H)，6.79(s，1H)，6.73(t，J＝3.6Hz，1H)，4.25(s，4H)，4.08(brs，1H)，3.34(brd，J＝9.6Hz，2H)，3.20-3.09(m，1H)，2.64-2.57(m，1H)，2.36(s，1H)，1.75(brs，4H)，1.53(s，9H)ppm。

Flow process IX

1,2-two [2-(6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-carboxylic acid tert-butyl ester] ethane (25):

(2.47g, DMF 7.75mmol) (30mL) solution add to 60%NaH, and (0.34g is in dry DMF 8.50mmol) (20mL) suspension with 20 via feed hopper at 0 ℃.Behind the 1h, the shift reaction mixture is to-40 ℃ of baths (ACN/ dry ice).At-40 ℃, (3.65g, DMF 7.06mmol) (20mL) solution adds to cold anion solutions from feed hopper with tosylate 24.Behind the 30min, only observe starting material, therefore be warming up to 0 ℃ lentamente through 2h by the TLC analysis.Behind 0 ℃ of 2-3h, by adding saturated NH ₄Cl aqueous solution cancellation reaction.Use diethyl ether and water diluted mixture thing and layering.Wash ether layer with water several times to remove DMF, use the salt water washing then once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain the white foam 25 of 3.27g (70%) by positive HPLC (the EtOAc/ hexane of 10-100% is through 30 min) purifying crude product, white foam 25 is analyzed [hexane/EtOAc of 3: 1 (two kinds of moving phase), R by TLC _f(20)=0.8; R _f(24)=0.55; R _f(25)=0.5] be homogeneous substance. ¹H NMR(CDCl ₃，300MHz)δ7.61-7.52(m，1H)，6.82(t，J＝9.6Hz，1H)，6.68-6.61(m，1H)，6.48-6.46(m，1H)，4.34(s，2H)，3.93(m，1H)，3.34-3.26(m，2H)，3.17-3.01(m，1H)，2.05(m，1H)，1.70-1.58(m，4H)，1.50(s，9H)ppm。

1,2-two [2-(6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine] ethane (26):

At 0 ℃ trifluoroacetic acid (2mL) added to and to contain 25 (3.27g is in DCM 4.93mmol) (10mL) solution.Behind the 3h, add the TFA (2mL) of other part and in 1h, finish reaction.Remove in rotatory evaporator and to desolvate and residue is dissolved in DCM and uses saturated NaHCO ₃Twice of solution washing, salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated and obtains yellow foam 26, and yellow foam 26 uses under situation about not being further purified. ¹H NMR(CDCl ₃，300MHz)δ7.31(dd，J＝5.1，8.7Hz，1H)，6.92(s，1H)，6.77(ddd，J＝2.4，9.6，11.1Hz，1H)，6.44(dd，J＝2.4，9.9Hz，1H)，4.41(s，2H)，3.65-3.55(m，1H)，3.24-3.16(m，1H)，3.01-2.96(m，1H)，2.92(d，J＝7.8Hz，2H)，2.15-1.99(m，1H)，1.96-1.84(m，2H)，1.76-1.67(m，1H)ppm。

1,2-22,2,2-three fluoro-1-[2-(6-fluoro-1H-indol-3-yl methyl)-tetramethyleneimine-1-yl]-ethyl ketone } ethane (27):

At 0 ℃, (2.17g, 1.44mL 10.3mmol) add to and contain 26 (2.28g, 4.93mmol with TFAA; Theoretical yield based on abovementioned steps) and TEA (2.49g, 3.43mL is in DCM 24.6mmol) (50mL) solution.Behind the 30min, reaction mixture uses the DCM dilution and uses saturated NaHCO ₃Twice of solution washing, salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.(4: 1-1: the purifying crude product of 1 hexane/EtOAc) obtains 27,27 of 2.66g (82%, 2 step) and analyzes [hexane/EtOAc of 2: 1, R through TLC by the fast silica gel chromatogram method _f(26)=0.01; R _f(27)=0.5] be homogeneous substance. ¹H NMR(CDCl ₃，300MHz)δ7.70(dd，J＝5.4，9.0Hz，1H)，6.84(ddd，J＝1.8，9.3，10.5Hz，1H)，6.62(dd，J＝1.8，10.2Hz，1H)，6.44(s，1H)，4.36(s，2H)，4.29-4.28(m，1H)，3.60(app t，J＝7.2Hz，2H)，3.23(dd，J＝2.4，14.1Hz，1H)，2.51(dd，J＝9.9，14.1Hz，1H)，1.92-1.84(m，2H)，1.72-1.66(m，1H)，1.57-1.56(m，1H)ppm。

Flow process X

1-(2-{3,10-two fluoro-14-[1-(2,2,2-three fluoro-ethanoyl)-tetramethyleneimine-2-ylmethyl]-6,7-dihydro-pyrazine And [1,2-a; 4,3-a '] two indoles-13-ylmethyl }-tetramethyleneimine-1-yl)-2,2,2-three fluoro-ethyl ketones (28):

At room temperature (2.66g 4.06mmol) is dissolved among the pure TFA (25mL) with non-annularity dimer 27.Behind the 3h, except that desolvating and making the residue that is generated be dissolved in EtOAc, use saturated NaHCO in rotatory evaporator ₃Twice of solution washing, salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and the concentrated yellow foam that obtains the diastereomer of 2.65g (amount) indyl indoline.[TLC analyzes: hexane/EtOAc of 3: 1, R _f(27)=0.3; R _f(indyl indoline)=0.6-0.7].

(1.10g 4.84mmol) adds as for 1, and (2.65g is 4.05mmol) in the mixture for the indyl indoline crude product of 4-two  alkane (50mL) with the DDQ of a part.Behind the 2-3h, use the EtOAc diluted reaction mixture and filter by the Celite pad.Wash solid and use saturated NaHCO with EtOAc ₃The aqueous solution is used the salt water washing once then with filtrate washing five times.The washing soln that merges with EtOAc again extracting twice and the organic extract that makes merging through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.(4: 1 hexane/EtOAc) the purifying crude product obtains the 1.94g pale solid 28 in (73%, 2 step), and pale solid 28 is analyzed (hexane/EtOAc of 2: 1, R by TLC by the fast silica gel chromatogram method _f(indyl indoline)=0.6-0.7; R _f(28)=0.55] be homogeneous substance.NB: product 2, the fluorescence of 2 '-two indoles (28) are quite strong and be easy to grind purifying by the MeOH with SILVER REAGENT and obtain white solid. ¹H NMR(CDCl ₃，300MHz)δ8.06(dd，J＝5.1，8.1Hz，1H)，7.03-6.93(m，2H)，4.49(d，J＝9.0Hz，1H)，4.40(m，1H)，4.12(d，J＝9.0Hz，1H)，3.75-3.69(m，2H)，3.57-3.51(m，2H)，2.85(dd，J＝10.5，12.9Hz，1H)，1.78-1.74(m，2H)，1.51-1.45(m，1H)ppm。

Flow process XI

3,10-two fluoro-13,14-two-tetramethyleneimine-2-ylmethyl-6,7-dihydro-pyrazine also [1,2-a; 4,3-a '] two indoles (29):

To contain 28 (1.94g, 2.97mmol) and K ₂CO ₃(2.05g, MeOH 14.8mmol) (60mL) mixture is at 60 ℃ of heating 1.5h.Reaction mixture is cooled to room temperature and uses EtOAc and the water dilution.Layering is also used EtOAc aqueous phase extracted three times.The organic extract salt water washing that merges is through anhydrous Na ₂SO ₄Drying is filtered and also to be concentrated the yellow solid 29 that obtains 1.57g (amount), and yellow solid 29 is not advancing-going on foot under the situation of purifying to use.TLC analyzes, hexane/EtOAc of 1: 1, R _f(28)=0.9; R _f(29)=0.01. ¹H NMR(CDCl ₃，300MHz)δ7.65(m，1H)，6.98(app d，J＝8.2Hz，1H)，6.90(app t，J＝8.3Hz，1H)，4.31(s，2H)，3.97(brs，3H)，3.54(m，1H)，3.31(m，1H)，3.14(m，1H)，2.97(m，1H)，1.83(m，1H)，1.68(m，2H)，1.42(m，1H)ppm。 ¹³C NMR(CDCl ₃，75MHz)δ160.6(d，J _C-F＝238.7Hz)，136.2(d，J _C-F＝12.0Hz)，127.1，125.4，120.8(d，J _C-F＝10.2Hz)，109.8，108.9(d，J _C-F＝24.6Hz)，95.3(d，J _C-F＝26.3Hz)，59.6，45.6，41.6，31.0，30.7，24.5ppm。

[1-(2-{14-[1-(uncle 2--butoxy carbonyl amino-3-methyl-butyryl radicals)-tetramethyleneimine-2-ylmethyl]-3,10- Two fluoro-6,7-dihydro-pyrazine also [1,2-a; 4,3-a '] two indoles-13-ylmethyl }-tetramethyleneimine-1-carbonyl)-the 2-first Base-propyl group]-t-butyl carbamate (30):

To contain Boc-L-Val-OH (0.69g, 3.18mmol) and HATU (1.27g, anhydrous NMP (4mL) solution 3.34mmol) is cooled to 0 ℃.Behind the 15min, via syringe add DIPEA (0.45g, 0.61mL, 3.50mmol).Behind the 15min, adding contains 29, and (0.70g, (4mL) solution of NMP 1.52mmol) also makes reaction mixture rise to room temperature through 2h, complete [TLC analysis, hexane/EtOAc of 2: 1, R of TLC analysis demonstration 29 consumption when 2h _f(29)=0.01; R _f(30)=0.5].With the diethyl ether diluted reaction mixture and use rare HCl solution washing once, make to wash five times with water to remove excessive N MP, with saturated NaHCO ₃The aqueous solution and salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.(3: 1 hexane/EtOAc) the purifying crude product obtains the light yellow solid 30 of 1.09g (83%) by the fast silica gel chromatogram method. ¹H NMR(CDCl ₃，300MHz)δ8.04(dd，J＝5.1，8.7Hz，1H)，6.98(m，2H)，5.33(d，J＝9.3Hz，1H)，4.50(m，1H)，4.49(d，J＝8.1Hz，1H)，4.24(dd，J＝7.2，9.3Hz，1H)，4.11(m，2H)，3.67(dd，J＝3.0，13.5Hz，1H)，3.56(m，2H)，2.73(app t，J＝12.9Hz，1H)，1.99(dd，J＝7.2，13.5Hz，1H)，1.70-1.17(m，2H)，1.43(s，9H)，1.01(d，J＝7.2Hz，3H)，0.98(d，J＝7.5Hz，3H)ppm。 ¹³C NMR(CDCl ₃，75MHz)δ171.2，160.4(d，J _C-F＝238Hz)，155.7，136.6(d，J _C-F＝12.0Hz)，127.2，124.8，122.0(d，J _C-F＝9.7Hz)，109.2，108.5(d，J _C-F＝24.0Hz)，95.0(d，J _C-F＝26.3Hz)，79.4，57.7，56.9，47.3，41.7，31.8，29.7，28.4，28.3，23.8，19.7，17.7ppm。

Flow process XII

2-amino-1-(2-{14-[1-(2-amino-3-methyl-butyryl radicals)-tetramethyleneimine-2-ylmethyl]-3, the 10-difluoro -6,7-dihydro-pyrazine also [1,2-a; 4,3-a '] two indoles-13-ylmethyl }-tetramethyleneimine-1-yl)-3-methyl-Ding -1-ketone (31):

(1.09g, DCM 1.27mmol) (20mL) solution is cooled to 0 ℃ will to contain 30.Via transfer pipet add TFA (4mL) and monitoring reaction analyze up to TLC and show and 30 consume fully (～2h).TLC analyzes, 10% MeOH/DCM, R _f(30)=0.5; R _f(31)=0.4.Remove in rotatory evaporator and to desolvate and make residue be dissolved in EtOAc.With saturated NaHCO ₃The aqueous solution is with twice of EtOAc solution washing and use the salt water washing once.The washes aqueous solution that merges is stripped with EtOAc and is made organic extract through anhydrous Na ₂SO ₄Drying is filtered and the concentrated yellow solid 31 that obtains 0.83g (amount), and yellow solid 31 uses under situation about not being further purified. ¹H NMR(CDCl ₃，300MHz)δ8.09(dd，J＝5.1，8.7Hz，1H)，6.97(m，2H)，4.52(m，1H)，4.50(d，J＝8.7Hz，1H)，4.11(m，1H)，3.71(brd，J＝11.1Hz，1H)，3.51-3.32(m，2H)，2.74(app t，J＝12.6Hz，1H)，2.30(brs，4H)，1.92(m，1H)，1.68(m，2H)，1.41(m，1H)，1.03(m，6H)ppm。 ¹³C NMR(CDCl ₃，75MHz)δ174.3，171.4，160.6(d，J _C-F＝232.5Hz)，136.8(d，J _C-F＝7.5Hz)，127.4(d，J _C-F＝3.7Hz)，125.0，122.4(d，J _C-F＝7.5Hz)，109.6，108.7(d，J _C-F＝22.5Hz)，95.2(d，J _C-F＝22.5Hz)，58.0，47.3，41.9，30.0，28.5，28.5，24.1，19.9，17.6ppm。

Penult intermediate (32):

To contain Boc-L-N (Me) Ala-OH (0.49g, 2.45mmol) and HATU (0.98g, anhydrous NMP (4mL) solution 2.56mmol) is cooled to 0 ℃.Behind the 15min, via syringe add DIPEA (0.35g, 0.47mL, 2.69mmol).Behind the 15min, adding contains 31, and (0.77g, (4mL) solution of NMP 1.17mmol) also makes reaction mixture rise to room temperature through 2h, complete [TLC analysis, hexane/EtOAc of 1: 1, R of TLC analysis demonstration 31 consumption when 2h _f(31)=0.01; R _f(32)=0.5].Reaction mixture uses ether to dilute and with rare HCl solution washing once, makes to wash five times with water to remove excessive N MP, uses saturated NaHCO ₃The aqueous solution and salt water washing once, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.(1: 1 hexane/EtOAc) the purifying crude product obtains the light yellow solid 32 of 0.92g (76%) by the fast silica gel chromatogram method. ¹H NMR(CDCl ₃，300MHz)δ8.05(m，1H)，7.65-6.90(m，2H)，4.53(m，3H)，4.13(m，1H)，3.70-3.52(m，4H)，2.82(m，2H)，2.72(app t，J＝11.1Hz，1H)，1.70(m，1H)，1.64(s，3H)，1.53(s，9H)，1.45-1.25(m，2H)，1.34(d，J＝7.0Hz，3H)，1.05-0.88(m，6H)ppm。 ¹³C NMR(CDCl ₃，75MHz)δ171.3，170.4，160.4(d，J _C-F＝232.5Hz)，136.6(d，J _C-F＝7.5Hz)，127.2，124.7，122.1，109.2，108.5(d，J _C-F＝22.5Hz)，95.0(d，J _C-F＝22.5Hz)，57.8，55.5，47.4，41.7，31.6，29.9，29.7，28.4，23.8，19.3，18.0ppm。

Flow process XIII

N-{1-[2-(3,10-two fluoro-14-{1-[3-methyl-2-(2-methylamino--propionic acid amide)-butyryl radicalies]-tetramethyleneimine-2- Ylmethyl }-6,7-dihydro-pyrazine also [1,2-a; 4,3-a '] two indoles-13-ylmethyl)-tetramethyleneimine-1-carbonyl]-2- Methyl-propyl group }-2-methylamino--propionic acid amide (33):

(0.92g, DCM 0.89mmol) (15mL) solution is cooled to 0 ℃ will to contain 32.Via transfer pipet add TFA (3mL) and monitoring reaction analyze up to TLC and show and 32 consume fully (～3h).TLC analyzes, 10% MeOH/DCM, R _f(32)=0.4; R _f(33)=0.3.Remove in rotatory evaporator and to desolvate and make residue be dissolved in EtOAc.Use saturated NaHC0 ₃The aqueous solution is with twice of EtOAc solution washing and use the salt water washing once.The washings that merges strip with EtOAc and with organic extract through anhydrous Na ₂SO ₄The crude product 33 that obtains 0.73g is filtered and concentrated to drying.By RP-HPLC (method: solvent orange 2 A: water w/0.1%v/v HOAc, solvent B:ACN w/0.1%v/v HOAc, Dynamax Microsorb C18 60  8 μ, 41.4mm * 25cm; Flow: 40mL/min; Detector: 272nm) purifying crude product.Use saturated NaHCO ₃Aqueous solution dilution contains the cut of product and extracts with EtOAc.Use salt water washing EtOAc extract, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Residue is dissolved in minimum ACN, is diluted with water to muddiness, freezing and freeze-drying obtains cotton-shaped white solid 33. ¹H NMR(DMSO，300MHz)δ8.04-7.86(m，2H)，7.38(app dd，J＝2.3，10.5Hz，1H)，6.90(app dt，J＝2.3，9.9Hz，1H)，4.68(app d，J＝8.7Hz，1H)，4.34-4.23(m，2H)，3.98(d，J＝8.7Hz，1H)，3.46(m，2H)，2.94(app q，J＝6.4Hz，1H)，2.70(t，J＝12.8Hz，1H)，2.12(s，3H)，1.94(m，1H)，1.58(m，2H)，1.35(m，1H)，1.16-1.07(m，2H)，1.03(d，J＝7.0Hz，3H)，0.85(m，6H)ppm。 ¹³C NMR(CDCl ₃，75 MHz)δ175.0，170.8，160.6(d，J _C-F＝232.5Hz)，136.8(d，J _C-F＝7.5Hz)，127.4(d，J _C-F＝3.7Hz)，125.0，122.3(d，J _C-F＝7.5Hz)，109.5，108.7(d，J _C-F＝22.5Hz)，95.2(d，J _C-F＝22.5Hz)，60.4，57.9，55.3，47.6，42.0，35.2，31.7，30.0，28.6，24.0，19.7，19.6，18.2ppm。Mass spectrum, m/z=[415.6] (M+2)+/ 2.

Embodiment:

Embodiment 4:

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b, as aziridine or azetidine ring;

R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps randomly, R5a is connected via following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

R12a, R12b, R13a, R13b, R14a and R14b are H, Cl, Br, F, alkyl, cycloalkyl, hydroxyl, alkoxyl group, amino, alkylamino, cyano group or CO independently ₂H; With

R3a and R3b are H, halogen, alkyl, aryl, aralkyl, amino, virtue amino, aryl alkyl amino, hydroxyl, alkoxyl group, aryloxy, aralkyl hydroxyl, dialkylamino, amide group, sulfonamido or amidino groups independently.

Numbering	R8a	R7a	R5a	( ^＊) of putting of standing body	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,
Numbering	R8a	R7a	R5a	( ^＊) of putting of standing body	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,	72	Me	S-Me	S-(2R-Et OH)	S	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A
73	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A	72	Me	S-Me	S-(2R-Et OH)	S	H	H	S-(2R-Et OH)	S-Me	Me	H	F	H	A
73	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A	74	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A
75	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	F	H	H	A	74	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A
75	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	F	H	H	A	76	Me	S-Me	H	S	H	H	H	S-Me	Me	H	F	H	B
77	Me	S-Me	S-Me	S	H	H	S-Me	S-Me	Me	H	F	H	B	76	Me	S-Me	H	S	H	H	H	S-Me	Me	H	F	H	B
77	Me	S-Me	S-Me	S	H	H	S-Me	S-Me	Me	H	F	H	B	78	Me	S-Me	S-(2S-Et OH)	S	H	H	S-(2S-EtO H)	S-Me	Me	H	F	H	A
79	Me	S-Me	S-Et	S	H	H	S-Et	S-Me	Me	H	F	H	A	78	Me	S-Me	S-(2S-Et OH)	S	H	H	S-(2S-EtO H)	S-Me	Me	H	F	H	A
79	Me	S-Me	S-Et	S	H	H	S-Et	S-Me	Me	H	F	H	A	80	Me	S-Me	S-(2S-Et OH)	S	H	H	S-(2S-EtO H)	S-Me	Me	F	H	H	A
81	H	H	S-iPr	S	H	H	S-iPr	H	H	H	F	H	B	80	Me	S-Me	S-(2S-Et OH)	S	H	H	S-(2S-EtO H)	S-Me	Me	F	H	H	A
81	H	H	S-iPr	S	H	H	S-iPr	H	H	H	F	H	B	82	Me	S-Me	S-sBu	S	H	H	S-sBu	S-Me	Me	F	H	H	A
83	Me	S-Me	S-cHex	S	H	H	S-cHex	S-Me	Me	F	H	H	A	82	Me	S-Me	S-sBu	S	H	H	S-sBu	S-Me	Me	F	H	H	A
83	Me	S-Me	S-cHex	S	H	H	S-cHex	S-Me	Me	F	H	H	A	84	Me	S-Me	S-tBu	S	H	H	S-tBu	S-Me	Me	H	F	H	A
85	Me	S-Me	S-cHex	S	H	H	S-cHex	S-Me	Me	H	F	H	A	84	Me	S-Me	S-tBu	S	H	H	S-tBu	S-Me	Me	H	F	H	A
85	Me	S-Me	S-cHex	S	H	H	S-cHex	S-Me	Me	H	F	H	A	86	Me	S-Me	S-(2R-Et OH)	R	S-OH	S-OH	S-(2R-Et OH)	S-Me	Me	H	F	H	A
87	Me	S-Me	S-iPr	R	S-OH	S-OH	S-iPr	S-Me	Me	H	F	H	A	86	Me	S-Me	S-(2R-Et OH)	R	S-OH	S-OH	S-(2R-Et OH)	S-Me	Me	H	F	H	A
87	Me	S-Me	S-iPr	R	S-OH	S-OH	S-iPr	S-Me	Me	H	F	H	A	88	Me	S-Me	S-(2R-Et OMe)	R	S-OH	S-OH	S-(2R-Et OMe)	S-Me	Me	H	F	H	A
89	Me	S-Me	S-(2R-Et OtBu)	R	S-OH	S-OH	S-(2R-Et OtBu)	S-Me	Me	H	F	H	B	88	Me	S-Me	S-(2R-Et OMe)	R	S-OH	S-OH	S-(2R-Et OMe)	S-Me	Me	H	F	H	A
89	Me	S-Me	S-(2R-Et OtBu)	R	S-OH	S-OH	S-(2R-Et OtBu)	S-Me	Me	H	F	H	B	90	Me	S-Me	S-(2S-Et OH)	R	S-OH	S-OH	S-(2S-EtO H)	S-Me	Me	H	F	H	A
91	Me	R-Me	S-iPr	S	H	H	S-iPr	R-Me	Me	H	F	H	B	90	Me	S-Me	S-(2S-Et OH)	R	S-OH	S-OH	S-(2S-EtO H)	S-Me	Me	H	F	H	A
91	Me	R-Me	S-iPr	S	H	H	S-iPr	R-Me	Me	H	F	H	B	92	Me	S-Me	R-iPr	S	H	H	R-iPr	S-Me	Me	H	F	H	C
93	Me	R-Me	R-iPr	S	H	H	R-iPr	R-Me	Me	H	F	H	C	92	Me	S-Me	R-iPr	S	H	H	R-iPr	S-Me	Me	H	F	H	C

Additional embodiments:

Embodiment 5

R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroaralkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps randomly, R5a is connected via following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

R12a, R12b, R13a, R13b, R14a and R14b are H, Cl, Br, F, alkyl, cycloalkyl, hydroxyl, alkoxyl group, amino, alkylamino, cyano group or CO independently ₂H and;

Numbering	R8a	R7a	R5a	( ^＊) stereochemistry of position	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope
Numbering	R8a	R7a	R5a	( ^＊) stereochemistry of position	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope	94	Me	S-Me	S-(2R-EtOH)	S	H	H	S-(2R-EtOH)	S-Me	Me	H	H	H	A

Flow process XIV

Contain two-(Boc-allylglycine) kind (34)

To contain Boc-L-allyl group-Gly-OH (0.115g, 0.53mmol) and HATU (0.20g, anhydrous NMP (3mL) solution 0.53mmol) is cooled to 0 ℃.Behind the 10min, via syringe add diisopropylethylamine (0.1mL, 0.58mmol).Behind the 5min, adding contains 29, and (0.11g, (3mL) solution of NMP 0.23mmol) also makes reaction mixture rise to room temperature through 16h, complete [TLC analysis, 5% MeOH/DCM, R of TLC analysis demonstration 29 consumption when 16h _f(29)=0.4].With the diethyl ether diluted reaction mixture and with saturated NaHCO ₃Solution washing once, use rare HCl solution washing once with the salt solution washed twice, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.By RP-HPLC (Dynamax Microsorb C18 60  8 μ, 41.4mm * 250mm; Flow: 40mL/min; Detector: 254nm uses the ACN/ water that contains 0.1%AcOH to carry out the gradient elution of 20-100% through 30min) the purifying crude product.Use the EtOAc dilution to contain the cut of product, use saturated NaHCO ₃The aqueous solution washs, through anhydrous Na ₂SO ₄The faint yellow solid 34 that obtains 0.067g (73%) is filtered and concentrated to drying. ¹H NMR(CDCl ₃，300MHz)δ8.06(dd，J＝5.4，8.7Hz，1H)，7.02-6.92(m，2H)，5.88-5.79(m，1H)，5.42(d，J＝8.4Hz，1H)，5.19-5.11(m，2H)，4.49(dd，J＝6.9，15.9Hz，2H)，4.16-4.08(m，1H)，3.65-3.41(m，3H)，2.75(app t，J＝12.9Hz，1H)，2.54-2.35(m，2H)，1.75-1.68(m，3H)，1.46-1.43(m，9H)，1.21-1.19(m，1H)ppm。Mass spectrum, m/z=877.7 (M+Na)+.

Flow process XV

Closed loop replacement(metathesis)reaction (RCM) product (35):

At room temperature ((0.067g is in anhydrous DCM (30mL) solution 0.08mmol) 12mol%) to add to 34 for 9.2mg, 0.01mmol with the Ge Labu catalyzer of the first-generation (Grubbs ' catalyst).Reaction mixture is heated 6h under refluxing, TLC analyzes and shows most starting material when 6h.(7mg, 0.009mmol 11mol%) add to reaction mixture with other Ge Labu catalyzer then.Behind the 2d, evaporating solvent also passes through NP-HPLC (SiO ₂The EtOAc of hexane/EtOAc to 100% of 20% is through 20min) purifying residue crude product, thus obtain faint yellow solid as the alkene 35 (alkene isomers: 15mg and 24mg) of the expectation of separable isomers (unspecified alkene geometry) mixture.[TLC analyzes, hexane/EtOAc of 1: 1, R _f(34)=0.7; R _f(35)=0.6].35 isomers A: ¹H NMR (CDCl ₃, 300MHz) δ 7.80 (app t, J=9.0Hz, 1H), 7.00 (dd, J=1.5,11.07Hz, 1H), 6.92 (ddd, J=2.1,9.3,11.2Hz, 1H), 5.62 (app d, J=7.8Hz, 1H), 5.56 (brs, 1H), 4.79 (m, 1H), and 4.56-4.47 (m, 2H), 4.12 (app d, J=6.9Hz, 1H), 3.85 (dd, J=3.6,13.5Hz, 1H), and 3.52-3.49 (m, 1H), 3.41 (m, 1H), 2.73 (m, 1H), 2.45-2.40 (m, 2H), 1.70-1.64 (m, 4H), 1.44 (s, 9H), 1.07-1.05 (m, 1H) ppm.Mass spectrum, m/z=849.7 (M+Na)+.35 isomers B: ¹H NMR (CDCl ₃, 300MHz) δ 7.85 (brs, 1H), 7.0 (dd, J=1.5,9.3Hz, 1H), 6.92 (ddd, J=2.4,9.3,11.1Hz, 1H), 5.66 (brs, 1H), 5.56 (app d, J=7.5Hz, 1H), 4.80 (brs, 1H), and 4.62-4.51 (m, 2H), 4.17-4.09 (m, 1H), 3.75-3.52 (m, 3H), 2.69-2.60 (m, 2H), 2.47 (app d, J=15.3Hz, 1H), 1.69 (m, 3H), 1.45 (s, 9H), 1.18 (m, 1H) ppm.Mass spectrum, m/z=849.7 (M+Na)+.

Flow process XVI

The product (36) that connects alkyl:

5%Pd/C (25mg) is added to the isomers A of alkene 35, and (15mg is in EtOAc 0.02mmol) (5mL) solution.At H ₂Use under the atmosphere Pa Er device (Parr apparatus) jolt reaction mixture (～45-50PSI).2.5h after, TLC analyzes and shows unreacted starting materials.Add other 5%Pd/C (20mg) then and use the Pa Er device to make this mixture stand hydrogenation once more.1.5h after, by Celite (Celite ) filtering mixt and use EtOAc rinsing solid.Vacuum concentrated filtrate obtains faint yellow solid 36.

(24mg 0.03mmol) stands as at described same reaction conditions of isomer A and working specification 35 isomers B.With product (36) with merge from isomers A hydrogenant product.Compound 36 is through being separated into flaxen solid (35mg, 85%).[TLC analyzes, hexane/EtOAc of 1: 1, R _f(36)=0.3]. ₁H NMR(CDCl ₃，300MHz)δ7.85(dd，J＝5.4，8.1Hz，1H)，7.00(dd，J＝1.8，9.9Hz，1H)，6.92(ddd，J＝2.4，9.2，11.1，1H)，5.70(d，J＝7.8Hz，1H)，4.79-4.78(m，1H)，4.57-4.51(m，2H)，4.13-4.09(m，1H)，3.75(dd，J＝3.6，12.9Hz，1H)，3.53-3.41(m，2H)，2.49(app t，J＝12.3Hz，1H)，1.68-1.62(m，3H)，1.46(s，9H)，1.08-1.02(m，1H)，0.93-0.89(m，1H)ppm。Mass spectrum, m/z=851.7 (M+Na)+.

Flow process XVII

The free diamines (37) that connects alkyl:

(0.035g, DCM 0.04mmol) (10mL) solution is cooled to 0 ℃ will to contain 36.Add TFA (1mL) via transfer pipet, and make reactant rise to room temperature and it is monitored analyze up to TLC and to show and 36 consume fully (～1h).Remove in rotatory evaporator and to desolvate and residue is dissolved in EtOAc.Use saturated NaHCO ₃Twice of solution washing EtOAc solution is also used the salt water washing once.The washes aqueous solution that merges strip with EtOAc and with organic extract through anhydrous Na ₂SO ₄Drying is filtered and the concentrated yellow solid 37 that obtains 0.025g (amount), and yellow solid 37 uses under situation about not being further purified. ¹H NMR(CDCl ₃，300MHz)δ7.92-7.87(m，1H)，7.00(dd，J＝2.1，9.9Hz，1H)，6.91(ddd，J＝2.4，9.3，11.1Hz，1H)，4.81(m，1H)，4.52(d，J＝8.1Hz，1H)，4.13-4.09(m，1H)，3.79-3.76(m，2H)，3.42-3.39(m，2H)，2.50(app t，J＝12.9Hz，1H)，1.89(m，4H)，1.65-1.61(m，3H)，1.33-1.24(m，4H)，1.09-1.03(m，1H)，0.94-0.82(m，1H)ppm。Mass spectrum, m/z=3 15.3 (M+H)+/ 2; M/z=629.5 (M+H)+.

Flow process XVIII

Contain two-[Boc-N (Me)-L-Ala] big ring (38):

To contain Boc-N-methyl-L-Ala-OH (0.022g, 0.11mmol) and HATU (0.03g, anhydrous NMP (4mL) solution 0.09mmol) is cooled to 0 ℃.Behind the 10min, via syringe add diisopropylethylamine (0.05mL, 0.29mmol).Behind the 5min, adding contains 37, and (0.025g, (3mL) solution of NMP 0.04mmol) also makes reaction mixture rise to room temperature through 24h, complete [TLC analysis, 5% MeOH/DCM, R of TLC analysis demonstration 37 consumption when 24h _f(38)=0.3].Use the diethyl ether diluted reaction mixture also with saturated NaHCO ₃Solution washing once uses rare HCl solution washing once and use the salt solution washed twice, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated and obtains yellow oil 38 (35mg), and yellow oil 38 is used under situation about not being further purified.

Flow process XIX

The Smac stand-in (39) of big ring:

(0.035g, DCM 0.04mmol) (10mL) solution is cooled to 0 ℃ will to contain 38.Via transfer pipet add TFA (1mL) and make reactant rise to room temperature and it is monitored analyze up to TLC and to show and 38 consume fully (～2h).Remove in rotatory evaporator and to desolvate and residue is dissolved in EtOAc.Use saturated NaHCO ₃Twice of solution washing EtOAc solution is also used the salt water washing once.The washes aqueous solution that merges is stripped with EtOAc and is made organic extract through anhydrous Na ₂SO ₄Drying is filtered and the concentrated yellow solid 39 that obtains.By RP-HPLC (Dynamax MicrosorbC18 60  8 μ, 41.4mm * 250mm; Flow 40mL/min; Detector 254nm, the ACN/ water gradient of 20-100% that contains 0.1%AcOH is through 30min) the purifying crude product.Contain the cut of product and use saturated NaHCO with the EtOAc dilution ₃Solution washing is through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.With this substance dissolves in minimum ACN, dilute with water, freezing and freeze-drying obtain white cotton-shaped solid 39 (0.002g). ¹H NMR(CDCl ₃，300MHz)δ8.16(app d，J＝8.1Hz，1H)，7.90-7.86(m，1H)，7.00(app d，J＝8.7Hz，1H)，6.93(ddd，J＝3.0，7.5，9.0Hz，1H)，4.80(m，2H)，4.52(d，J＝8.4Hz，1H)，4.11(d，J＝8.4Hz，1H)，3.79(dd，J＝3.9，12.9Hz，1H)，3.51(m，1H)，3.44(m，1H)，3.11-3.09(m，1H)，2.46(s，3H)，1.35-1.25(m，8H)，1.06(m，1H)，0.89-0.85(m，1H)ppm。Mass spectrum, m/z=400.5 (M)+/ 2; M/z=799.7 (M)+; M/z=821.7 (M+Na+).

Embodiment:

Embodiment 6

Z is a key; The alkylidene group of 1-6 atom, alkenylene, alkynylene; Or alkylidene group, alkenylene or the alkynylene of optional 1-6 the carbon atom that replaces; Sulfide (S-), oxysulfide (SO-), sulfo group (SO ₂-) or disulphide (SS-) group; Aryl, aryl alkylene, heteroaryl, heteroaryl alkylidene group or optional aryl, aryl alkylene, heteroaryl, the heteroaryl alkylidene group that replaces; Amino or replace amino group; Sauerstoffatom;

M and n are 0,1,2 or 3 independently;

Numbering	R8a	R7a	m	n	Z	R3a	R3b	R7b	R8b	R12	R13	R14	K _DRange, μ M
Numbering	R8a	R7a	m	n	Z	R3a	R3b	R7b	R8b	R12	R13	R14	K _DRange, μ M	95	Me	Me	1	1	Z-CH＝CH-	H	H	Me	Me	H	F	H	A
96	Me	Me	1	1	E-CH＝CH-	H	H	Me	Me	H	F	H	A	95	Me	Me	1	1	Z-CH＝CH-	H	H	Me	Me	H	F	H	A
96	Me	Me	1	1	E-CH＝CH-	H	H	Me	Me	H	F	H	A	97	Me	Me	1	1	-CH ₂CH ₂-	H	H	Me	Me	F	H	H	A
98	Me	Me	1	1	-CH ₂CH ₂-	H	H	Me	Me	H	F	H	A	97	Me	Me	1	1	-CH ₂CH ₂-	H	H	Me	Me	F	H	H	A
98	Me	Me	1	1	-CH ₂CH ₂-	H	H	Me	Me	H	F	H	A	99	H	Me	1	1	-SS-	H	H	Me	H	H	F	H	A
100	Me	Me	1	1	-CH(OH)CH(OH)-	H	H	Me	Me	H	H	H	A	99	H	Me	1	1	-SS-	H	H	Me	H	H	F	H	A
100	Me	Me	1	1	-CH(OH)CH(OH)-	H	H	Me	Me	H	H	H	A	101	Me	Me	1	1	-CH ₂CH ₂-	S-OH	S-OH	Me	Me	H	F	H	A

Another embodiment:

Embodiment 7

R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps randomly, R5a is connected by following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

Numbering	R8a	R7a	R5a	(*) stereochemistry of position	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope
Numbering	R8a	R7a	R5a	(*) stereochemistry of position	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope	102	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A
103	Me	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	Me	H	F	H	A	102	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	F	H	A
103	Me	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	Me	H	F	H	A	104	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	H	H	A
105	Me	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	Me	H	H	H	A	104	H	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	H	H	A
105	Me	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	Me	H	H	H	A	106	H	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	H	H	H	H	A
107	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	H	H	A	106	H	S-Me	S-(2R-EtO H)	S	H	H	S-(2R-EtO H)	S-Me	H	H	H	H	A
107	Me	S-Me	S-iPr	S	H	H	S-iPr	S-Me	Me	H	H	H	A	108	H	S-Et	S-iPr	S	H	H	S-iPr	S-Et	H	H	H	H	A
109	Me	S-Et	S-iPr	S	H	H	S-iPr	S-Et	Me	H	H	H	A	108	H	S-Et	S-iPr	S	H	H	S-iPr	S-Et	H	H	H	H	A
109	Me	S-Et	S-iPr	S	H	H	S-iPr	S-Et	Me	H	H	H	A	110	Me	S-Me	S-(2S-EtO H)	S	H	H	S-(2S-EtO H)	S-Me	Me	H	F	H	A
111	Me	S-Me	S-Allyl	S	H	H	S-Allyl	S-Me	Me	H	F	H	A	110	Me	S-Me	S-(2S-EtO H)	S	H	H	S-(2S-EtO H)	S-Me	Me	H	F	H	A
111	Me	S-Me	S-Allyl	S	H	H	S-Allyl	S-Me	Me	H	F	H	A	112	Me	S-Me	S-iPr	R	S- OH	S- OH	S-iPr	S-Me	Me	H	F	H	A
113	Me	S-Me	S-sBu	R	S- OH	S- OH	S-sBu	S-Me	Me	H	F	H	A	112	Me	S-Me	S-iPr	R	S- OH	S- OH	S-iPr	S-Me	Me	H	F	H	A
113	Me	S-Me	S-sBu	R	S- OH	S- OH	S-sBu	S-Me	Me	H	F	H	A	114	Me	S-Me	S-(2S-EtOt Bu)	R	S- OH	S- OH	S-(2S-EtOt Bu)	S-Me	Me	H	F	H	A
115	Me	S-Me	S-(2S-EtO H)	R	S- OH	S- OH	S-(2S-EtO H)	S-Me	Me	H	F	H	A	114	Me	S-Me	S-(2S-EtOt Bu)	R	S- OH	S- OH	S-(2S-EtOt Bu)	S-Me	Me	H	F	H	A
115	Me	S-Me	S-(2S-EtO H)	R	S- OH	S- OH	S-(2S-EtO H)	S-Me	Me	H	F	H	A	116	Me	S-Me	S-(2R-EtO Me)	R	S- OH	S- OH	S-(2R-EtO Me)	S-Me	Me	H	F	H	A
117	Me	S-Me	S-(2S-EtO Me)	R	S- OH	S- OH	S-(2S-EtO Me)	S-Me	Me	H	F	H	A	116	Me	S-Me	S-(2R-EtO Me)	R	S- OH	S- OH	S-(2R-EtO Me)	S-Me	Me	H	F	H	A
117	Me	S-Me	S-(2S-EtO Me)	R	S- OH	S- OH	S-(2S-EtO Me)	S-Me	Me	H	F	H	A	118	Me	S-Me	S-(2R-EtO H)	R	S- OH	S- OH	S-(2R-EtO H)	S-Me	Me	H	F	H	A

Flow process XX

2-amino-N-[1-(2-{1-[6-(3-{1-[2-(2-amino-propionic acid amide)-3-methyl-butyryl radicals]-tetramethyleneimine-2-base Methyl }-indoles-1-yl)-six-2, the 4-diynyl]-1H-indol-3-yl methyl }-tetramethyleneimine-1-carbonyl)-the 2-first Base-propyl group]-preparation of propionic acid amide (43):

A. (1S-{2-methyl isophthalic acid S-[2S-(1-Propargyl-1H-indol-3-yl methyl)-tetramethyleneimine-1-carbonyl]-third The base carbamyl }-ethyl)-t-butyl carbamate (41):

Propargyl bromide [0.06mL, 0.410mmol, (80%wt/ toluene)] is added to 40 (0.150g in THF 0.319mmol) (2mL) solution, adds NaH[0.015g subsequently, 0.410mmol, (60% dispersion in the mineral oil)].Reaction mixture at room temperature stirred spend the night.Water (2mL) is added to reaction mixture and uses ethyl acetate (3 * 30mL) extraction products.Make water, salt water washing acetic acid ethyl ester extract and through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.By HPLC purifying crude product. ¹H NMR(CDCl ₃，300MHz)：δ8.0(s，1H)，7.9(d，J＝9.9Hz，1H)，7.38(d，J＝9.9Hz，1H)，7.3-7.1(m，3H)，6.8(m，1H)，4.8(s，2H)，4.62(m 1H)，4.5-4.4(m 1H)，4.4-4.0(m，2H)，3.7-3.5(m，2H)，3.4(m，1H)，2.5(m，1H)，2.4(s，1H)，2.2-1.8(m，4H)，1.48(s，9H)，1.35(d，J＝9.9Hz，3H)，1.05(d，J＝5.5Hz，3H)，0.95(d，J＝5.5Hz，3H)ppm。

B.{1-[1-(2-{1-[6-(3-{1-[2-(uncle 2--butoxy carbonyl amino-propionamido)-3-methyl-butyryl Base]-tetramethyleneimine-2-ylmethyl }-indoles-1-yl)-six-2, the 4-diynyl]-1H-indol-3-yl methyl }-pyrroles Alkane-1-carbonyl)-2-methyl-propyl group carbamyl]-ethyl }-t-butyl carbamate (42):

(0.070g, (0.040g is in acetonitrile 0.077mmol) (2mL) solution and reaction mixture is immersed oil bath (～100 ℃) the backflow 5min of preheating 0.385mmol) to add to 41 with neutralized verdigris (II).Then water is added to reaction mixture (2mL) and uses EtOAc (3 * 30mL) extraction products.Use NH ₄The OH aqueous solution (5mL), water, salt water washing organic extract and through anhydrous Na ₂SO ₄Drying is filtered and the concentrated crude product that obtains. ¹H NMR(CDCl ₃，300MHz)：δ8.0(s，2H)，7.9(d，J＝9.9Hz，2H)，7.38(d，J＝9.9Hz，2H)，7.3-7.1(m，6H)，6.8(m，2H)，4.8(s，4H)，4.62(m 2H)，4.5-4.4(m 2H)，4.4-4.0(m，4H)，3.7-3.5(m，4H)，3.4(m，2H)，2.5(m，2H)，2.2-1.8(m，8H)，1.48(s，18H)，1.35(d，J＝9.9H，6H)，1.05(d，J＝5.5Hz，6H)，0.95(d，J＝5.5Hz，6H)ppm。

C.2-amino-N-[1-(2-{1-[6-(3-{1-[2-(2-amino-propionamido)-3-methyl-butyryl radicals]-tetramethyleneimine -2-ylmethyl }-indoles-1-yl)-six-2, the 4-diynyl]-1H-indol-3-yl methyl }-tetramethyleneimine-1-carbonyl Base)-2-methyl-propyl group]-propionic acid amide (43):

TFA (1mL) is added to 42, and (0.030g is in DCM 0.029mmol) (5mL) solution and make reaction mixture at room temperature stir 30min.Then with NaHCO ₃The aqueous solution (3mL) adds to reaction mixture.Concentrated reaction mixture, dilute with water also uses DCM (3 * 30mL) extraction products.Water, salt water washing organic extract and through anhydrous Na ₂SO ₄Dry.Remove in rotatory evaporator and to desolvate and by the reversed-phase HPLC purified product. ¹H NMR(DMSO，300MHz)δ8.0(s，2H)，7.9(d，J＝9.9Hz，2H)，7.38(d，J＝9.9Hz，2H)，7.3-7.1(m，6H)，6.8(m，2H)，4.8(s，4H)，4.62(m 2H)，4.5-4.4(m 2H)，4.4-4.0(m，4H)，3.7-3.5(m，4H)，3.4(m，2H)，2.5(m，2H)，2.2-1.8(m，8H)，1.35(d，J＝9.9Hz，6H)，1.05(d，J＝5.5Hz，6H)，0.95(d，J＝5.5Hz，6H)ppm。

Embodiment:

Embodiment 8

R3a and R3b are H, halogen, alkyl, aryl, aralkyl, amino, virtue amino, aryl alkyl amino, hydroxyl, alkoxyl group, aryloxy, aralkyl hydroxyl, dialkylamino, amide group, sulfonamido or amidino groups independently; With

Wa and Wb are H, Cl, Br, F, alkyl, CN or CO independently ₂H.

Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	Wa	Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	The KD scope
Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	Wa	Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	The KD scope	119	Me	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	Me	H	H	H	A

120	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	H	H	A
120	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	H	H	A	121	Me	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	Me	H	H	H	A

Flow process XXI

N-{1-cyclohexyl-2-[2-(1-{2-[2-(3-{1-[2-cyclohexyl-2-(2-methylamino--propionamido)-ethanoyl]- Tetramethyleneimine-2-ylmethyl }-indoles-1-yl)-oxyethyl group]-ethyl }-1H-indol-3-yl methyl)-tetramethyleneimine-1- Base]-2-oxo-ethyl }-preparation of 2-methylamino--propionic acid amide (51):

Compound (46):

At 0 ℃, with NaH (60%, 0.025g, (0.17g is in dry DMF 0.56mmol) (5mL) solution 0.62mmol) to add to indoles 45.Behind the 1h, in extremely rapid succession add the bromine ether (0.16g, 0.68mmol) and n-Bu ₄NCl (0.021g, 0.05mmol).Make reaction mixture slowly rise to room temperature and the lasting 16h of stirring.By adding saturated NH ₄Cl aqueous solution cancellation reaction is also used the diethyl ether extraction product.Water repeatedly washs the ethereal extract of merging to remove excessive DMF, uses the salt water washing then and through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Crude product is merged to the material that forms (0.11g 45,0.36mmol) and by positive HPLC (the EtOAc/ hexane of 10-100%) purifying obtain the colorless oil 46 of 0.18g and the indoles in 0.18g monoalkylation generation under similar reaction conditions. ¹H NMR(CDCl ₃，300 MHz)δ7.75-7.66(m，2H)，7.25-7.07(m，6H)，6.85-6.80(m，2H)，4.13(m，8H)，3.62(brs，4H)，3.40-3.15(m，4H)，2.63(m，2H)，2.04(brs，4H)，1.53(s，18H)ppm。

Compound (47):

(0.18g, DCM 0.27mmol) (8mL) solution is cooled to 0 ℃ will to contain 46.Add trifluoroacetic acid (2mL) and make reaction mixture keep 1h at 0 ℃.By the saturated NaHCO of careful adding ₃Aqueous solution cancellation reaction is also used the EtOAc extraction product.Use NaHCO ₃The organic extract that the aqueous solution, salt water washing merge and through anhydrous Na ₂SO ₄The light yellow oil 47 that obtains 0.075g is filtered and concentrated to drying.Crude product is directly used in next step reaction. ¹H NMR(CDCl ₃，300MHz)δ8.49(br s，2H)，7.57(app.d，J＝8.2，Hz，2H)，7.26-7.052(m，6H)，6.82(s，2H)，4.11(m，8H)，3.57(m，4H)，3.29-2.96(m，6H)，1.95-1.58(m，8H)ppm。 [2-(2-{1-[2-(2-{3-[1-(uncle 2--butoxy carbonyl amino-2-cyclohexyl-ethanoyl)-tetramethyleneimine-2-base Methyl]-indoles-1-yl }-oxyethyl group)-ethyl]-1H-indol-3-yl methyl }-tetramethyleneimine-1-yl)-the 1-hexamethylene Base-2-oxo-ethyl]-t-butyl carbamate (48):

With HATU (0.15g, 0.38mmol) and the N-methylmorpholine (0.042g 0.42mmol) adds to and contains the N-Boc-Cyclohexylglycine (0.09g is in anhydrous NMP (2mL) solution 0.34mmol).(0.075g, anhydrous NMP (2mL) 0.16mmol) also stirs 16h with reaction mixture behind the 15min, to add 47.The dilute with water reaction mixture is also used the diethyl ether extraction product.Make water, salt solution repeatedly wash the ethereal extract of merging to remove excessive N MP, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.Obtain the colorless oil 48 of 0.085mg by positive HPLC (the EtOAc/ hexane of 50-100%) purifying crude product. ¹H NMR(CDCl ₃，300MHz)δ7.87(d，J＝7.62Hz，2H)，7.27-7.10(m，6H)，6.84(s，2H)，5.36(d，J＝9.37Hz，2H)，4.45(m，2H)，4.30(app.t，J＝6.4Hz，2H)，4.18-4.12(m，6H)，3.70-3.58(m，6H)，3.35(dd，J＝14.0，2.9Hz，2H)，2.41(dt，J＝11.1，2.3Hz，2H)，2.04-1.57(m，20H)，1.44(s，18H)，1.37-1.07(m，10H)ppm。

2-amino-1-(2-{1-[2-(2-{3-[1-(2-amino-2-cyclohexyl-ethanoyl)-tetramethyleneimine-2-ylmethyl]-Yin Diindyl-1-yl }-oxyethyl group)-ethyl]-1H-indol-3-yl methyl }-tetramethyleneimine-1-yl)-2-cyclohexyl-ethyl ketone (49):

(0.085g, DCM 0.08mmol) (8mL) solution is cooled to 0 ℃ will to contain 48.Add trifluoroacetic acid (2mL) and reaction mixture is kept 30min at 0 ℃.Add the TFA (1mL) of part in addition and at 0 ℃ of stirred reaction mixture 1h.By the saturated NaHCO of careful adding ₃Aqueous solution cancellation reaction is also used the EtOAc extraction product.Use NaHCO ₃The organic extract that the aqueous solution, salt water washing merge is through anhydrous Na ₂SO ₄The light yellow oil 49 that obtains 0.068g is filtered and concentrated to drying.Crude product is directly used in next step reaction. ¹H NMR(CDCl ₃，300MHz)δ7.90(d，J＝7.6Hz，2H)，7.31-7.12(m，6H)，6.84(app t，J＝14Hz，2H)，4.47(m，minor rotomer)，4.15(m，4H)，3.66-3.36(m，8H)，2.80(m，2H)，2.42(m，2H)，2.04-0.83(m，30H)ppm。

Acid amides (50):

With HATU (0.083g, 0.21mmol) and the N-methylmorpholine (0.024g 0.23mmol) adds to and contains the N-Boc-N-methylalanine (0.041g is in anhydrous NMP (2mL) solution 0.19mmol).Behind the 15min, add 49 (0.068g, 0.09mmol) anhydrous NMP (2mL) and stirred reaction mixture 16h.The dilute with water reaction mixture is also used the diethyl ether extraction product.Make water, salt solution repeatedly wash the ethereal extract of merging to remove excessive N MP, through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.(0.05g 49,0.06mmol) also obtain the colorless oil 50 of 32mg by positive HPLC (the EtOAc/ hexane of 10-100%) purifying to the material merging that forms under similar reaction conditions with crude product. ¹H NMR(CDCl ₃，300MHz)δ7.86(d，J＝7.6Hz，2H)，7.27-7.08(m，6H)，6.84(s，2H)，4.70(m，2H)，4.58(app.t，J＝7.6Hz，2H)，4.19(m，2H)，4.15-4.04(m，6H)，3.76-3.60(m，6H)，3.37(m，2H)，2.83(s，6H)，2.42(m，2H)，1.98-1.55(m，20H)，1.51(s，18H)，1.33(d，J＝7.0Hz，6H)，1.30-0.98(m，10H)ppm。

N-{1-cyclohexyl-2-[2-(1-{2-[2-(3-{1-[2-cyclohexyl-2-(2-methylamino--propionamido)-ethanoyl]- Tetramethyleneimine-2-ylmethyl }-indoles-1-yl)-oxyethyl group]-ethyl }-1H-indol-3-yl methyl)-tetramethyleneimine-1- Base]-2-oxo-ethyl }-2-methylamino--propionic acid amide (51):

(0.0.062g, DCM 0.055mmol) (8mL) solution is cooled to 0 ℃ will to contain 50.Add trifluoroacetic acid (2mL) and reaction mixture is kept 1h at 0 ℃.By the saturated NaHCO of careful adding ₃Aqueous solution cancellation reaction is also used the EtOAc extraction product.Use NaHCO ₃The organic extract that the aqueous solution, salt water washing merge is through anhydrous Na ₂SO ₄Drying is filtered and is concentrated.By reversed-phase HPLC (the ACN/ water w/0.1%HOAc of 10-100%) purifying crude product, obtain the white solid 512HOAc of 0.047g after the freeze-drying. ¹H NMR(DMSO，300MHz)δ7.91(d，J＝8.7Hz，2H)，7.71(d，J＝7.8Hz，2H)，7.32(d，J＝7.5Hz，2H)，7.05(m，2H)，6.99-6.92(m，4H)，4.39(app.t，J＝6.6Hz，2H)，4.15(m，6H)，3.57(m，4H)，3.52-3.23(m，4H)，3.06-2.91(m，4H)，2.46(s，6H)，2.31(m，2H)，2.15(s，12H)，1.91-1.65(m，12H)，1.55-1.49(m，8H)，1.07(d，J＝7.0Hz，6H)，1.16-0.93(m，10H.)ppm。

Embodiment:

Embodiment 9

Wa and Wb are H, Cl, Br, F, alkyl, CN, CO independently ₂H.

Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	Wa	Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,
Numbering	R8a	R7a	R5a	( ^＊) of position body putting is upright	Wa	Wb	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,	122	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	H	H	A
123	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	F	H	A	122	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	H	H	A
123	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	H	F	H	A	124	Me	S-Me	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Me	Me	H	F	H	A
125	Me	S-Et	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Et	Me	F	H	H	B	124	Me	S-Me	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Me	Me	H	F	H	A
125	Me	S-Et	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Et	Me	F	H	H	B	126	Me	S-Me	S-cHex	S	H	H	H	H	S-eHex	S-Me	Me	F	H	H	A
127	Me	S-Me	S-cHex	S	Me	Me	H	H	S-cHex	S-Me	Me	F	H	H	A	126	Me	S-Me	S-cHex	S	H	H	H	H	S-eHex	S-Me	Me	F	H	H	A
127	Me	S-Me	S-cHex	S	Me	Me	H	H	S-cHex	S-Me	Me	F	H	H	A	128	Me	S-Et	S-cHex	S	Me	Me	H	H	S-cHex	S-Et	Me	F	H	H	A
129	H	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	Cl	H	H	A	128	Me	S-Et	S-cHex	S	Me	Me	H	H	S-cHex	S-Et	Me	F	H	H	A
129	H	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	Cl	H	H	A	130	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	Cl	H	H	A
131	H	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	H	Cl	H	H	A	130	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	Cl	H	H	A
131	H	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	H	Cl	H	H	A	132	Me	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	Me	Cl	H	H	A
133	Me	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	Me	F	H	H	A	132	Me	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	Me	Cl	H	H	A
133	Me	S-Me	S-iPr	S	H	H	H	H	S-iPr	S-Me	Me	F	H	H	A	134	H	H	S-cHex	S	H	H	H	H	S-cHex	H	H	H	F	H	C
135	Me	S-Me	H	S	H	H	H	H	H	S-Me	Me	H	F	H	C	134	H	H	S-cHex	S	H	H	H	H	S-cHex	H	H	H	F	H	C
135	Me	S-Me	H	S	H	H	H	H	H	S-Me	Me	H	F	H	C	136	Me	S-Et	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Et	Me	H	F	H	C

137	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	Me	H	H	A
137	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	Me	Me	H	H	A	138	H	H	S-cHex	S	H	H	H	H	S-cHex	H	H	H	H	H	B
139	H	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	H	H	H	A	138	H	H	S-cHex	S	H	H	H	H	S-cHex	H	H	H	H	H	B
139	H	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	H	H	H	A	140	Me	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	Me	H	F	H	A
141	Me	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	Me	H	H	H	A	140	Me	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	Me	H	F	H	A
141	Me	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	Me	H	H	H	A	142	H	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	H	H	H	H	A
143	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	H	H	H	A	142	H	S-Me	S-tBu	S	H	H	H	H	S-tBu	S-Me	H	H	H	H	A
143	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	S-Me	H	H	H	H	A	144	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	H	H	H	H	H	A
145	Me	S-Me	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Me	Me	H	H	H	B	144	Me	S-Me	S-cHex	S	H	H	H	H	S-cHex	H	H	H	H	H	A
145	Me	S-Me	S-(2R-EtOH)	S	H	H	H	H	S-(2R-EtOH)	S-Me	Me	H	H	H	B	146	Me	S-Me	S-(CH ₂) ₄NH ₂	S	H	H	H	H	S-(CH ₂) ₄NH ₂	S-Me	Me	Me	H	H	A

Other embodiment:

Embodiment 10

Wa and Wb are H, Cl, Br, F, alkyl, CN, CO independently ₂H; With

R11a and R11b form alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S.

Numbering	R8a	R7a	R5a	Wa	Wb	( ^＊) positionization put learn three-dimensional	R11a-R11b	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,
Numbering	R8a	R7a	R5a	Wa	Wb	( ^＊) positionization put learn three-dimensional	R11a-R11b	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DScope,	147	H	S-Me	S-iPr	H	H	S	CH ₂CH ₂CH ₂ CH ₂CH ₂CH ₂	H	H	S-iPr	S-Me	H	H	H	H	A
148	Me	S-Me	S-cHe x	H	H	S	CH ₂CH ₂CH ₂ CH ₂CH ₂CH ₂	H	H	S-cHe x	S-Me	Me	H	H	H	A	147	H	S-Me	S-iPr	H	H	S	CH ₂CH ₂CH ₂ CH ₂CH ₂CH ₂	H	H	S-iPr	S-Me	H	H	H	H	A
148	Me	S-Me	S-cHe x	H	H	S	CH ₂CH ₂CH ₂ CH ₂CH ₂CH ₂	H	H	S-cHe x	S-Me	Me	H	H	H	A	149	Me	S-Me	S-tBu	H	H	S	CH ₂CH _2CH ₂ CH ₂CH ₂CH ₂	H	H	S-tBu	S-Me	Me	H	H	H	A
150	H	S-Me	iPr	H	H	S	(R，R)-CH ₂C H(OH)CH(O H)CH ₂	H	H	iPr	S-Me	H	H	F	H	A	149	Me	S-Me	S-tBu	H	H	S	CH ₂CH _2CH ₂ CH ₂CH ₂CH ₂	H	H	S-tBu	S-Me	Me	H	H	H	A
150	H	S-Me	iPr	H	H	S	(R，R)-CH ₂C H(OH)CH(O H)CH ₂	H	H	iPr	S-Me	H	H	F	H	A	151	Me	S-Me	2R-Et OH	H	H	S	(R，R)-CH ₂C H(OH)CH(O H)CH ₂	H	H	2R-Et OH	S-Me	Me	H	F	H	A
152	Me	S-M e	2R-Et OH	H	H	S	(S，S）-CH ₂CH (OH)CH(OH )CH ₂	H	H	2R-Et OH	S-Me	Me	H	F	H	A	151	Me	S-Me	2R-Et OH	H	H	S	(R，R)-CH ₂C H(OH)CH(O H)CH ₂	H	H	2R-Et OH	S-Me	Me	H	F	H	A
152	Me	S-M e	2R-Et OH	H	H	S	(S，S）-CH ₂CH (OH)CH(OH )CH ₂	H	H	2R-Et OH	S-Me	Me	H	F	H	A	153	Me	S-M e	cHex	H	H	S	CH ₂CH ₂CH ₂	H	H	cHex	S-Me	Me	H	H	H	B
154	Me	S-Me	cHex	H	H	S	CH ₂CH ₂OCH ₂	H	H	cHex	S-Me	Me	H	H	H	A	153	Me	S-M e	cHex	H	H	S	CH ₂CH ₂CH ₂	H	H	cHex	S-Me	Me	H	H	H	B

							CH ₂OCH ₂CH ₂
							CH ₂OCH ₂CH ₂										155	Me	S-Et	cHex	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	cHex	S-Et	Me	H	H	H	B
156	Me	S-Me	2R-Et OH	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	2R-Et OH	S-Me	Me	H	H	H	A	155	Me	S-Et	cHex	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	cHex	S-Et	Me	H	H	H	B
156	Me	S-Me	2R-Et OH	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	2R-Et OH	S-Me	Me	H	H	H	A	157	Me	S-Et	2R-Et OH	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	2R-Et OH	S-Et	Me	H	H	H	B
158	Me	S-Me	cHex	H	H	S	C(O)CH ₂CH ₂C H ₂C(O)	H	H	cHex	S-Me	Me	H	H	H	A	157	Me	S-Et	2R-Et OH	H	H	S	CH ₂CH ₂OCH ₂ CH ₂OCH ₂CH ₂	H	H	2R-Et OH	S-Et	Me	H	H	H	B
158	Me	S-Me	cHex	H	H	S	C(O)CH ₂CH ₂C H ₂C(O)	H	H	cHex	S-Me	Me	H	H	H	A	159	Me	S-Me	cHex	H	H	S	C(O)C ₆H ₄C(O)	H	H	cHex	S-Me	Me	H	H	H	A
160	Me	S-Me	2R-Et OH	H	H	S	CH ₂CH ₂	H	H	2R-Et OH	S-Me	Me	H	H	H	A	159	Me	S-Me	cHex	H	H	S	C(O)C ₆H ₄C(O)	H	H	cHex	S-Me	Me	H	H	H	A
160	Me	S-Me	2R-Et OH	H	H	S	CH ₂CH ₂	H	H	2R-Et OH	S-Me	Me	H	H	H	A	161	Me	S-Me	2R-Et OH	H	H	S	CH ₂CH ₂CH ₂ CH ₂	H	H	2R-Et OH	S-Me	Me	H	H	H	B

Another embodiment:

Embodiment 11

R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Perhaps randomly, R5a is connected by following radicals with the R5b residue: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

X is O, N, S or C=C;

Wa is H, Cl, Br, F, alkyl, CN, CO ₂H;

R11b does not exist or is H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces;

Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S.

Numbering	R8a	R7a	R5a	Wa	R11b	( ^＊) upright of position body putting	X	Wb-R11a	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DPαvγε
Numbering	R8a	R7a	R5a	Wa	R11b	( ^＊) upright of position body putting	X	Wb-R11a	R3a	R3b	R5b	R7b	R8b	R12	R13	R14	K _DPαvγε	162	Me	S-Me	S-(2R- EtOH)	H	-	S	O	CH ₂CH ₂CH ₂	H	H	S-(2R- EtOH	S-Me	Me	H	H	H	D
163	Me	S-Me	S-cHex	H	-	S	O	CH ₂CH ₂CH ₂	H	H	S-cHex	S-Me	Me	H	H	H	A	162	Me	S-Me	S-(2R- EtOH)	H	-	S	O	CH ₂CH ₂CH ₂	H	H	S-(2R- EtOH	S-Me	Me	H	H	H	D

In mammalian cell, by at least two kinds of independently machine-processed activation that realize caspase, described mechanism excites via different caspases but causes identical execution thing (effector)--the activation of caspase.Except that cytochrome c activating mechanism (sometimes being called " inner dead path "), the caspase cascade reaction is activated via the activation that is positioned at the death receptor on the cytolemma by " outside dead path " mechanism.The example of death receptor comprises DR4, DR5 and TNF-R1 (and other members of cytokine receptor TNF group).Corresponding part (respectfully) respectively is TRAIL and TNF-α.Caspase-8-proenzyme has brought out auto-activation with combining of death receptor, and wherein the inhibition prodomain of caspase-8-proenzyme is cleaved and remove.Caspase-8 is discharged by acceptor and subsequently can activating effect thing--caspase (caspase-3, caspase-6, caspase-7), and the same with caspase-9 activation pathway, the result passes through effector--caspase and apoptoticly bring out proteolytic cleavage cell target spot.

The present invention relates generally to the Smac peptide mimics, prepares method of Smac peptide mimics and uses thereof, comprises the method for preparing peptide mimics described above.In one embodiment of the invention, Smac peptide mimics (this paper is called the Smac stand-in) plays chemical synergists (chemopotentiatingagent).Term " chemical synergists " refers to play increases organism, tissue or cell to chemical compound or be called the medicament of the treatment or the radiotherapeutic susceptibility effect of " chemotherapeutic " or " chemicals ".One embodiment of the invention are therapeutic compositions of Smac stand-in.Another embodiment of the present invention is the therapeutic composition of Smac stand-in, and it can be used as chemical synergists and biological or chemical therapeutical agent or radiation and works.Another embodiment of the invention is by using the method that the Smac peptide mimics suppresses tumor growth in vivo.Another embodiment of the invention is by using the method that Smac stand-in and biological or chemical therapeutical agent or chemoluminescence suppress tumor growth in vivo.Another embodiment of the invention is to use the method that Smac stand-in treatment of the present invention suffers from the patient of cancer by using Smac stand-in of the present invention or associating biological or chemical therapeutical agent or chemoluminescence separately.

In a preferred embodiment of the invention, can comprise the combination of alkylating agent, vegeto-alkali, antitumor antibiotics, metabolic antagonist, topoisomerase enzyme inhibitor, hormone medicine, NSAIDs, somatomedin, cytokine, mitotic inhibitor and these materials with the suitable biological and chemical therapeutical agent that the Smac stand-in are used simultaneously.

In another embodiment of the invention, cell is in the original place or in individuality, and the using of pharmaceutical composition that contact procedure is contained the Smac stand-in for the treatment of significant quantity influences, and wherein said individuality can be simultaneously or stood to be used for the treatment of the radiation or the chemotherapy of neural proliferative disease in advance.Morbific cell is a tumour, for example still is not limited to: bladder cancer, mammary cancer, prostate cancer, lung cancer, carcinoma of the pancreas, cancer of the stomach, colorectal carcinoma, ovarian cancer, kidney, liver cancer, melanoma, lymphoma, sarcoma and combination thereof.Yet this cell can also be the immortal tumour cell that is used for tumor cell culture.

The Smac stand-in can also be used for the treatment of autoimmune disorder.The apoptosis defective of in tumour, finding, think that the defective owing to the elimination immunity system autoreaction sexual cell ability of apoptosis tolerance plays a key effect in the cause of disease of autoimmune disorder.Autoimmune disorder is characterised in that the create antagonism antibody of himself organ and molecule of immune cell, or the directtissima tissue, causes the latter's destruction.Those id reaction sexual cells are stood the performance that apoptotic obstacle causes disease.The defective that apoptosis is regulated in the clear and definite autoimmune disorder is as systemic lupus erythematous or rheumatoid arthritis.

In one embodiment, the cause of disease also comprises the abnormality proliferation of cell, those in the disease of for example any autoimmune disorder of described cell or the pair cell apoptosis tolerance owing to the crossing expression of IAP or PROTEIN B cl-2 family member.The example of this type of autoimmune disorder includes but not limited to collagen diseases, as rheumatoid arthritis, systemic lupus erythematous, husky general Cotard, CREST syndrome (calcinosis, Raynaud's syndrome, esophageal dysmotility, telangiectasis), dermatomyositis, vasculitis (Mo Busi Wei Genashi disease) and Sj  gren Cotard; Kidney disease is as Goodpasture, rapid progress type glomerulonephritis and II type membranoprolifer ative glomerulonephritis; Endocrinopathy is as type i diabetes, the multiple incretopathy-moniliosis of autoimmunity-ectoderm malnutrition (APECED), autoimmunity parathyropathy, pernicious anemia, sexual gland insufficiency, primary Mo Busiadisenshi disease (Morbus Addison ' s), hyperthyroidism, Hashimoto thyroiditis and primary solid edema (primary myxedema); Dermatosis is as pemphigus vulgaris, bullous pemphigoid, herpes gestationis, epidermolysis bullosa and EMM; Hepatic diseases is as primary biliary cirrhosis, autoimmune cholangitis, autoimmunity 1 type hepatitis, autoimmunity 2 type hepatitis, primary sclerosing cholangitis; Sacred disease is as multiple sclerosis, myasthenia gravis, Lan Bote-Eton myasthenic syndrome, acquired neuromyotonia, Guillain-Barre﹠1﹠ syndrome (wooden Le-Fei Xue syndrome), stiff-man syndrome, cerebellar degeneration, ataxia, opsoklonus, sensorium neuropathy and achalasia; Hematologic disease is as autoimmune hemolytic anemia, idiopathic thrombocytopenic purpura (Mo Busi Wal Hough); With the infectious diseases of autoimmunity reaction, as AIDS, malaria and chagas disease.

The pharmaceutical composition that the present invention comprised comprises the Smac stand-in and the pharmaceutically acceptable carrier of the treatment significant quantity of dosage form, and wherein the Smac stand-in suppress the activity of IAP, has therefore promoted apoptosis.Another embodiment of the invention is to unite the Smac stand-in of the treatment significant quantity that contains dosage form of use and the composition of pharmaceutically acceptable carrier with biological or chemical therapeutical agent and/or radiotherapy, wherein the Smac stand-in suppress the activity of IAP, thereby have promoted apoptosis and improved chemotherapy and/or radiotherapeutic curative effect.

The present invention comprises that also preparation contains the method for pharmaceutical composition of Smac stand-in, and described method includes but not limited to treat the combination of the Smac stand-in and the pharmaceutically acceptable vehicle (exipient) of significant quantity.

In one embodiment of the invention, be used for promoting the Smac peptide mimics of the treatment significant quantity of apoptotic therapeutic composition to combine with at least one IAP.In one embodiment, IAP can be XIAP.In another embodiment, IAP can be ML-IAP.In another embodiment, IAP can be cIAP-1 or cIAP-2.In yet another embodiment, IAP can be polytype IAP.

Embodiment of the present invention comprise that also treatment is in the patient's who needs the treatment situation method, and the using of Smac peptide mimics of wherein treating significant quantity is delivered to the patient and the Smac peptide mimics combines with at least one IAP.In one embodiment, IAP can be XIAP.In another embodiment, IAP can be ML-IAP.In another embodiment, IAP can be cIAP-1 or cIAP-2.In yet another embodiment, IAP can be polytype IAP.This method may further include uses chemotherapeutic simultaneously.Described chemotherapeutic can be, but be not limited to alkylating agent, metabolic antagonist, antitumor antibiotics, Taxan, hormones preparation, monoclonal antibody, glucocorticosteroid, mitotic inhibitor, topoisomerase I inhibitor, topoisomerase II inhibitor, immunomodulator, cell growth factor, cytokine and nonsteroidal anti-inflammatory compound.

The Smac stand-in are preferably used with significant quantity.Significant quantity is to produce the amount of formulation of anticipation reaction separately or with other medications.This can only comprise the process of the disease that temporarily slows down, but preferably includes the process that permanently stops disease or delay the outbreak of disease or ward off disease or the appearance of illness.This can monitor by the method for routine.Usually, the dosage of active compound should from every day about 0.01mg/kg to 1000mg/kg every day.Expect preferred every day via intravenously, intramuscular or intradermal administration and once or several times the dosage range of 50-500mg/kg suit.As long as chemotherapeutic or radiation make system to Smac peptide mimics sensitivity, just can be in chemotherapy or radiation, afterwards or before carry out using of Smac peptide mimics.

Generally speaking, normal experiment in the clinical trial will be determined the concrete scope of optimum treatment effect at every kind of therapeutical agent and every kind of dosage regimen, and will be adjusted in the effective and safe scope using of concrete patient according to patient's state with to the reactivity of initial application.Yet, to adjust final dosage regimen according to the judgement that clinical attending doctor considers following factor, described factor such as time length of the usefulness of age, patient's illness and build size, Smac peptide mimics, treatment and the treatment severity of disease.For example, the dosage regimen of Smac peptide mimics can be every day with 2-4 time (preferred 2 times) the Orally administered 1mg-2000mg/ of gradation days, preferred 1-1000mg/ days, and more preferably 50-600mg/ days, to reduce growth of tumor.Also can use intermittent treatment (as one in three weeks weeks or three weeks all around).

Under initial application dose in hyporeactive situation of experimenter, can adopt the higher dosage that patient's tolerance the allows higher effective dose of different, more concentrated route of delivery (or via).Expection multiple dosing every day can obtain the suitable systemic concentrations of compound.Usually, use maximal dose, i.e. the highest safe dose of the reliable medical judgment of basis.Yet those of ordinary skill in the art should understand for medical reasons, psychological causes or any other reason in fact, and the patient can stake out a claim lower dosage or tolerable dose.

Can use multiple route of administration.Certainly, selected ad hoc fashion will depend on the severity of the specific chemotherapeutic agent of selection, the illness for the treatment of and produce the required dosage of treatment effect.Generally speaking, can use medically acceptable any method of application to implement method of the present invention, described any method of application is meant the active compound that produces effective concentration and any way that does not cause unacceptable side effect clinically.That such method of application includes but not limited to is oral, rectum, part, nose, intracutaneous, suction, intraperitoneal, intravesical or parenteral approach.That term " parenteral " comprises is subcutaneous, intravenously, intramuscular or infusion.Intravenously or intramuscular approach are specially adapted to purpose of the present invention.

In one aspect of the invention, influence normal tissue unfriendly as the other biological or chemotherapeutic or radiotherapeutic Smac peptide mimics of being with or without described herein, but make the chemotherapy/radiation scheme sensitivity of tumour cell other.As if though be not wishing to be bound by theory, because the apoptosis that this tumour-specific brings out, remarkable and disadvantageous side effect minimizing is as unsuitable vasorelaxation or shock.Preferably, composition or method are designed to allow make cell or tumour to chemotherapy or radiotherapy sensitivity by the Smac peptide mimics of using at least a portion before chemotherapy or radiotherapy.The part that the inclusion of radiotherapy and/or chemotherapeutic can be used as treatment plan is included further to strengthen killing and wounding tumour cell by the Smac peptide mimics.

In optional embodiment of the present invention, the Smac stand-in are united the therapy of another kind of form and are used, and the therapy of described another kind of form includes but not limited to be selected from following another kind of therapy: radiotherapy, immunotherapy, photodynamic therapy, chemotherapy and their combination.

The anticancer chemotherapy agent that associating Smac stand-in are used can be specifically with tumorigenesis tissue or cell as any therapeutical agent of target spot, include but not limited to alkylating agent, vegeto-alkali, antitumor antibiotics, metabolic antagonist and the topoisomerase enzyme inhibitor that contains hexamethyl melamine, busulfan, carbon platinum, carmustine, Amboclorin, cis-platinum, endoxan, Dacarbazine, hexamethyl melamine, ifosfamide, lomustine, melphalan, chlormethine, oxaliplatin, Procarbazine, streptozocin, Temozolomide, plug is for group, Uramustine, Docetaxel, Etoposide, irinotecan, taxol, tenisopide, vincristine(VCR), vinealeucoblastine(VLB), vindesine, vinorelbine, bleomycin, gengshengmeisu, daunorubicin, epirubicin, hydroxyurea, idarubicin, mitomycin, mitoxantrone, Plicamycin, azathioprine, capecitabine, CldAdo, cytosine arabinoside, fludarabine, Fluracil, the Ro 2-9757 deoxynucleoside, gemcitabine, mercaptopurine, methotrexate, Nelzarabine, pemetrexed, pentostatin, Tioguanine, irinotecan, Hycamtin, BNP 1350, SN 38,9-amino-irinotecan, lurtotecan, lucky for health (gimatecan), difluoro is for health, Dx, epirubicin, idarubicin, Nemorubicin, mitoxantrone, loxoxantrone, Etoposide and their combination.

Can also use simultaneously with immunosuppressor as Smac stand-in described herein.Immunotherapy comprises using of immune-active agent, and described immune-active agent is selected from bacille Calmette-Guerin vaccine (BCG), Interferon, rabbit and other activate the medicament of infected patient's immune system and their combination specifically.

Pharmaceutical composition in one embodiment of the invention can be before the Smac stand-in be used, add other biology preparation, chemotherapeutic or antineoplastic agent (vide infra) and/or radiation together or afterwards.As used herein, term " pharmaceutically acceptable carrier " refers to solid or liquid filling agent, the thinner that one or more are compatible or is suitable for being applied to human encapsulate (encapsulating) material.Term " carrier " thus organic or inorganic composition, natural or synthetics that expression and activeconstituents combination are convenient to use.The component of pharmaceutical composition can with molecule blend of the present invention, and mutually there not to be the interaction blend that weakens required drug effect in fact each other.

Delivery system of the present invention is designed to include regularly and discharges, postpones to discharge or the slowly-releasing release delivery system, like this Smac peptide mimics send preferential generation, and have the competent time and feasible position sensitivity to be treated arranged.The Smac peptide mimics can unite radiation and/or other anticancer chemical agent uses (vide infra).This type systematic can avoid multiple to use Smac peptide mimics compound, is curee and facilitating property of doctor, and can be applicable to some composition of the present invention especially.

Polytype release delivery system can use, and for those of ordinary skills known and can be used for the context of the invention, it includes but not limited to the polymeric matrix system, for example poly-(rac-Lactide-glycollide), poly-barkite multipolymer, polycaprolactone, polyesteramide, poe, multi-hydroxybutyrate and polyanhydride.The medicine that comprises polymer particulates body is above described in No. the 5th, 075,109, United States Patent (USP) for example to some extent.Delivery system also comprises following non-polymer system: lipid or neutral fat, hydrogel release system, Sylastic system, the system based on peptide, wax coating, the conventional tackiness agent of use and the compressed tablet of vehicle, partially fused implant or the like, described lipid comprises sterol, as cholesterol, cholesterol ester and lipid acid; Described neutral fat is direactive glyceride, two glyceryl ester and Witepsol W-S 55 for example.Concrete example includes but not limited to: (a) corrosion system, wherein contain active compound with form in the matrix, and as at United States Patent (USP) the 4th, 452,775,4,667,014,4,748,034 and 5,239, those that describe in No. 660; (b) diffusion system, wherein active ingredient with the speed of control by seeing through in the polymkeric substance, as at United States Patent (USP) the 3rd, 832,253 and 3,854, described in 480.In addition, can use the hardware delivery system based on pump, wherein some is suitable for implanting.

The use that long-term slowly-releasing discharges implant is desirable.The long-term release of Shi Yonging herein refers to make up row implant side by side with the activeconstituents of delivery treatments level at least 30 days, and preferred 60 days.It is that those of ordinary skills are known and comprise aforesaid some release system that long-term slowly-releasing discharges implant.

Described pharmaceutical composition can exist with dosage device easily and can be by the known any method preparation of pharmaceutical field.All methods include following step: introduce promoting agent to unite with the carrier that constitutes one or more auxiliary agents or vehicle (exipients).Usually, the solid carrier by active compound evenly and densely being introduced liquid vehicle, segmentation or both prepare composition, then if necessary, this product are shaped.

In one embodiment of the invention, with peptide mimics dimer described above and pharmaceutically acceptable vehicle (exipient) combination.

The composition that is suitable for parenteral administration comprises the sterile aqueous preparations of chemical synergists (as the Smac peptide mimics) easily, and described sterile aqueous preparations is preferably oozed with blood of receptor etc.This aqueous compositions can use suitable dispersion agent or wetting agent and suspension agent to prepare according to known method.Injectable sterile preparation can also be injectable sterile solution or the suspension in atoxic parenteral acceptable diluent or solvent, for example 1, and the solution in the 3-butanediol.In acceptable carrier and solvent, what can adopt is water, Ringer's solution and isoosmotic sodium chloride solution.In addition, aseptic expressed oil is conventional solvent or the suspension medium that adopts.For this purpose, the expressed oil of any gentleness that can adopt comprises synthetic direactive glyceride or two glyceryl ester.In addition, lipid acid such as oleic acid can be used for injectable preparation.Be suitable for the carrier formulation that oral, subcutaneous, intravenously, intramuscular etc. use and can see Remington ' s Pharmaceutical Sciences, Mack Publishing Co., Easton, PA incorporates its full content into this paper by reference.

Other chemotherapeutic. the chemotherapeutic that is suitable for associating the present invention use includes but not limited at " Modern Pharmacology with Clinical Applications ", Sixth Edition, CraigStitzel, Chpt.56, the chemotherapeutic of describing among the pg 639-656 (2004) is incorporated its full content into this paper herein by reference.The chemotherapeutic drug that this reference is described comprises alkylating agent, metabolic antagonist, antitumor antibiotics, plant derivation product such as Taxan; Enzyme, hormone preparation such as glucocorticosteroid; Other preparations such as cis-platinum, monoclonal antibody; Immunomodulator such as Interferon, rabbit and cell growth factor.Other classifications that are applicable to chemotherapeutic comprise mitotic inhibitor and the similar thing of non-steroidal estrogen antagonist.Other suitable chemotherapeutics comprise topoisomerase I and topoisomerase II inhibitor, kinase inhibitor and any medicament that can activate outside or inner cell apoptosis pathway or Smac from plastosome release.

The suitable biology preparation and the object lesson of chemotherapeutic include but not limited to cis-platinum, card Jie's nitrogen (BCNU), 5 FU 5 fluorouracil (5-FU), cytosine arabinoside (Ara-C), gemcitabine, methotrexate, daunorubicin, Zorubicin, dexamethasone, Hycamtin, Etoposide, taxol, vincristine(VCR), tamoxifen, TNF-α, TRAIL, Interferon, rabbit (its α and β type), Thalidomide and melphalan.The object lesson of the chemotherapeutic that other are suitable comprises nitrogen mustards (as endoxan), alkylsulfonate, nitrosourea, ethyleneimine, triazene, antifol, purine analogue, pyrimidine analogue, anthracycline antibiotics, bleomycin, mitomycin, gengshengmeisu, Plicamycin, vinca alkaloids, epipodophyllotoxin, Taxan, glucocorticosteroid, the altheine enzyme, oestrogenic hormon, male sex hormone, progestogen, prolan B, Sostatin LAR, hydroxyurea, procarbazine, mitotane, hexamethyl melamine, carbon platinum, mitoxantrone, monoclonal antibody, LEVAMISOLE HCL, Interferon, rabbit, interleukin-, filgrastim and Sargramostim.Chemotherpeutic compositions also comprises other members' (promptly except TRAIL) of TNF superfamily compound or formulation example such as BCG, and they can induce chemokine synthetic after the intravesical treatment.NSAID can also unite Smac stand-in use of the present invention and can comprise selectivity and nonselective cox 2 inhibitor, celecoxib and rofecoxib (rofecoib).

The radiotherapy experimental program. in addition, in the certain methods of embodiment of the present invention, the Smac peptide mimics can be united chemoluminescence or the use of other modality of cancer treatment that is used to suppress growth of tumour cell.For example, but be not limited to, radiotherapy (or radiotherapy) is the medical usage as the ionizing rays of the cancer therapy part of the control malignant cell that is suitable for using in embodiment of the present invention.Though radiotherapy is often used as the part of curative therapy, it also uses as palliative treatment once in a while, and it is impossible wherein curing and its target is the alleviation of symptom.Radiotherapy is generally used for treating tumour.It can be used as original therapy.Usually also radiotherapy can be combined with surgical operation and/or chemotherapy.Using the most common tumor of radiation therapy treatment is mammary cancer, prostate cancer, the rectum cancer, brain neck cancer, gynecological cancer, bladder cancer and lymphoma.Radiotherapy only is applied to relate to the regional area of tumour usually.Irradiation field also often comprises draining lymph node.But it is possible uncommon that radiotherapy is applied to whole body or whole skin surfaces.Usually use every day radiotherapy up to 35-38 part (dosage of every day is a part).These little frequent dosage make healthy cell grow backward, repair the damage that is caused by radiation.Radiotherapeutic three main divisions are external beam radiotherapy or teletherapy, brachytherapy or sealed source radiotherapy and unencapsulated source radiotherapy, and above-mentioned division is the suitable example that the present invention treats experimental program.Difference between them relates to source positions: outside in the body outside, and sealing and unencapsulated source radiotherapy have the radioactive substance that send inside.Brachytherapy sealed source is drawn usually later on, and unencapsulated source injects body.Using of Smac peptide mimics can be before this treats experimental program, carry out simultaneously with this treatment experimental program.

Fig. 1 shows Smac tetrapeptide (AVPI) and the effective binding affinity of Smac stand-in (numbering 17) and XIAPBIR-3, and the Smac stand-in that this figure discloses numbering 17 have significantly increased by 30,000 times with respect to the binding affinity of Smac tetrapeptide.

In rat, investigate numbering 1, numbering 122 and number transformation period of 123 these 3 kinds of Smac stand-in.The IV dosage of every kind of Smac stand-in is 1mg/kg.Fig. 2 shows that the eventually end of Smac stand-in eliminates the transformation period and reach about 6h, wherein numbers 1 and has the longest transformation period.

Biology preparation and chemotherapeutic/antineoplastic agent and radiation are brought out apoptosis by the apoptosis path that activates outside or inside, and because the Smac stand-in reduce the inhibition of apoptosis protein matter (IAP), therefore eliminated the retardance of pair cell apoptosis, the combination of chemotherapeutic/antineoplastic agent and radiation and Smac stand-in will act synergistically and promote apoptosis.In order to show the synergistic effect of Smac stand-in and general chemistry therapeutical agent, select a different set of tumor cell line and tested typical compound and gamma-rays from the chemotherapeutics of different machinery classification.

Use as before by Hansen, M.B., Nielsen, S.E. and Berg, K. ((1989) J.Immunol.Methods 119 203-210) describes and its content is all incorporated this paper into by reference 72h MTT tests and distinguishes specificity and the nonspecific action of Smac stand-in to cell killing.Briefly, with the SK-OV-3 cell inoculation in the McCoyShi substratum that contains 10% fetal bovine serum albumin (every hole 20,000) of 96 orifice plates and 37 ℃ of overnight incubation.Second day, add the test compound of different concns (0.003-10 μ M) and culture plate is cultivated 72h in addition at 37 ℃.In each hole, add the 5mg/mL MTT reagent of 50 microlitres then and culture plate is cultivated 3h at 37 ℃.When culture cycle finished, the DMSO that adds 50 microlitres in each hole was with dissolved cell and use microwell plate microplate reader (Victor ²1420, Wallac is Finland) in the optical density (OD) (OD) of 535nm place metering orifice.Cell survival rate (CS) calculates via following formula:

CS=(handling the average OD of the OD/ control wells in hole) * 100%

Use the Smac stand-in of ovarian cancer cell line test No. 116, SK-OV-3 and MRC-5 cell use as normal cell contrast.Fig. 3 shows for kill tumor cell numbering 116 than negative control more effective 100,000 times, and normal (non-tumour) cell remains unaffected.

By using the point that GraphPad Prism calculating dose response curve and 50%CS point intersect to obtain EC ₅₀, EC ₅₀Be defined as the drug level that produces 50%CS.These results show to can be used as monotherapy with XIAP bonded Smac stand-in or unite with chemotherapy and are used for treatment for cancer.

Annexin V/propidium iodide dyeing-bring out apoptotic ability in order to show the Smac stand-in has carried out the dyeing of annexin V-isothiocyanate fluorescein.Briefly according to working specification (Invitrogen, Carlsbad, CA), with cellular exposure in the Smac of different concns stand-in 18-24h and remove from test board by tryptic digestion subsequently.Cell pellet shape also is suspended in again in the test damping fluid (by manufacturers's supply) then.To cultivate 1h in annexin V and the propidium iodide adding cell product and in the room temperature lucifuge.After cultivating, in every pipe, add other damping fluid (200 μ l), and by flow cytometer sample is analyzed immediately.As estimating and analyze, when existing, the Smac stand-in greatly promoted apoptosis by flow cytometer by annexin/PI dyeing.Compared with the control, the apoptosis cell purpose that causes of IAP antagonist enlarges that (the annexin positive/propidium iodide feminine gender-right lower quadrant (lower right quadrant) is dose-dependent and is attributable to apoptosis decrease and not via the ratio that increases non-viable non-apoptotic cell.

The melanoma cell that has proved the Smac stand-in that use chemical synergism shows tolerance (Chawla-Sarkar.Clin.Cancer Res. (2001)) to the apoptosis effect of chemotherapeutic agent TRAIL.(the MTT test Fig. 4) discloses when use the present invention numbers 1 Smac peptide mimics processing breast cancer cell line MDA-MB-231 cell cell proliferation test, and cell only tolerates the antiproliferative effect of Smac stand-in of the present invention.By contrast, when numbering 1 associating TRAIL uses,, have and make 1000 times the increase of antiproliferative effect of 100 times of cell killing increases as the same by detecting in corresponding forfeiture during colony forms.Control peptide stand-in (numbering 62) fail with TRAIL produce synergy and result's (data not shown) show numbering 62 separately or associating TRAIL all do not have antiproliferative activity.Behind the 4h, TRAIL brings out a spot of apoptosis (if any) of MDA MB-231 cell separately.Use separately the processing of numbering 121 also to fail to bring out significant apoptosis (be about cell total amount 10%).By contrast, behind 4h, TRAIL makes the apoptosis activity increase by 4 times with the combination of numbering 121.

The compound that contains and do not contain the different concns of 0.4ng/ml TRAIL by adding comes analysis of cells to form the ability of great-hearted colony.Briefly, with cell with 100 cell inoculations in every hole in the 2ml growth medium of 12 hole gauge lattice.Remove this substratum behind the 24h and replace with the Smac stand-in of the different concns in the growth medium that contains 1%DMSO.Behind test 72h, remove the Smac stand-in of different concns and replace with the growth medium of 2ml.Culture plate is reentered into incubator 7-10 days, and during this period, colony propagation is 10 cells of each colony extremely at least, and it can be counted by eyes.Use 0.01% crystal violet solution (H ₂Weightmeasurement ratio among the O) culture plate is dyeed 30 minutes.Use tap water washing culture plate to remove any residual Viola crystallina, drying is also counted colony.(CA) S shape dose response (variable slope) equation in is analyzed inhibiting data for GraphPad software, San Diego to use GraphPad Prism.50% inhibition concentration (EC ₅₀) be that enzymic activity has been reduced by 50% drug level with respect to the control sample that does not have medicine.

In the Study of cytotoxicity of T98G cell, observe example--the synergy of Hycamtin and camptothecine of two kinds of topoisomerase enzyme inhibitors.The synergy of maximum is compared with the cytotoxicity of every kind of independent compound is added up, and necrocytosis is than the 50-60% that has more of expection.The result shows that Hycamtin and camptothecine can play synergy as numbering 1 Smac stand-in with the present invention, are used to promote apoptosis.Synergistic total volume is 457, wherein number 1 and topoisomerase enzyme inhibitor such as Hycamtin between maximum synergy compare with the cytotoxicity of one Smac and Hycamtin is added up, necrocytosis is than the 30%-40% that has more of expection.

Potential interacts between the medicine in order further to estimate, use the program (Prichard of MacSynergy II by name, M.N., K.R.Aseltine and C.Shipman, Jr.1993.MacSynergy II.User ' s manual.Univ.of Michigan, Ann Arbor) matrix and every kind of activity that compound is independent of every kind of medicine and the conversion of the dual serial dilution ordering of Smac stand-in have been tested.Test taxol and be numbered 122 the synergy of Smac peptide mimics in the OVCAR3 cell.The synergy of maximum is 10-20% after testing, and its expression necrocytosis is added up desired many than the cytotoxicity that each compound is independent.

Taxan is the compound that suppresses mitotic division (mitotsis) by obstruction based on the unzipping of the spindle body of microtubule.Taxan, taxol and Smac stand-in commonly used by the test different concns draw these data.The dosage range of taxol by about 0.0 to about 500.0nM.For numbering 122, dosage range is about 125.0 to about 8000.0nM.Total collaborative capacity is about 170.

Think that the mechanism of action of the compound that contains platinum is in conjunction with DNA and disturbs its repair mechanism, finally causes necrocytosis.Cis-platinum and the synergy of Smac peptide mimics in the OVCAR-3 cell have been tested.The synergy of maximum is added up with the cytotoxicity that each compound is independent and is compared, and necrocytosis is than the 40-50% that has more of expection.Cis-platinum and Smac stand-in medicine by the test different concns draw these data.For cis-platinum, dosage range is about 0.0 to about 166,500.0nM.For numbering 122, dosage range is about 500.0 to about 32,000.0nM.Total collaborative capacity is about 434.Use the combination of carbon platinum and Smac stand-in to carry out similar test.Synergy between Smac peptide mimics, numbering 122 and the carbon platinum.

This effective synergy makes the use of IAP antagonist Smac peptide mimics become possibility, thereby has improved the curative effect of commercially available antineoplastic compound (such as but not limited to taxol, cis-platinum and carbon platinum).This effect will allow to reduce the tolerance difference antineoplastic compound required dosage and/or improve speed of answer at commercially available dosage.

The invention is not restricted to the embodiment of above description and example, and can in appended claim scope, carry out changes and improvements.

Claims

1. compound, it has following formula (II):

Wherein R1 and R2 are H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, the optional alkyl that replaces independently, or

Or

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Or randomly, R5a is connected by following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene bridge or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

R6a and R6b are H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, low alkyl group, the optional alkyl that replaces independently, or

Or

Wherein R7a and R7b are H, alkyl, cycloalkyl, haloalkyl independently; Or R8a and R7a can independently or form ring with R8b and R7b;

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a are with R8b and R7b independently or form ring;

M and n are 0,1,2 or 3 independently;

X and Y are O, N, S or C=C independently;

R9a, R9b, R10a, R10b are H, alkyl, the optional alkyl that replaces, aryl, heteroaryl, optional aryl, the heteroaryl that replaces independently, and perhaps R9a is independent with R10a or can be connected and form fragrance or non-aromatic ring via 4-8 the optional atom that replaces such as C, N, O or S abreast with R9b and R10b; With

Wherein when Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S; And R11a and R11b independently for do not exist, H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces; Or R11a and R11b form alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be by N, O or S replacement; With

When Wa and Wb were not covalent attachment, Wa and Wb were H, Cl, Br, F, alkyl, CN, CO2H independently; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms can be replaced by N, O or S; Or Wa can be alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces for H, Cl, Br, F, alkyl, CN, CO2H and Wb and R11a together, and wherein one or more carbon atoms are by N, O or S replacement; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

2. compound according to claim 1, it comprises homodimer.

3. compound according to claim 1, wherein the alkyl of R5a and R5b or the optional alkyl that replaces are independently selected from methyl, ethyl, sec.-propyl, isobutyl-, sec-butyl, the tertiary butyl, cycloalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl or heteroarylalkyl.

4. compound according to claim 1, wherein R7a and R7b are independently selected from methyl, methyl fluoride, difluoromethyl, ethyl, fluoro ethyl and cycloalkyl.

5. compound according to claim 1, wherein the alkyl of the described optional replacement of R5a and R5b is independently selected from alkoxylate and hydroxylated alkyl.

6. compound according to claim 1, wherein R3a and R3b are independently selected from H, hydroxyl, alkoxyl group, aryloxy, alkylamino, dialkylamino, amide group, sulfonamido or amidino groups.

7. compound according to claim 1, wherein when Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S; And R11a and R11b does not exist or be H, low alkyl group, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces independently; Or R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyalkyl chain of 2-12 carbon atom together, and wherein one or more carbon atoms are replaced by N, O or S; And X and Y are selected from N, O, S or C=C.

8. compound according to claim 1, wherein when Wa and Wb be non-covalent in conjunction with the time, Wa and Wb are H, Cl, Br, F, alkyl, CN, CO2H independently; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyalkyl chain of 2-12 carbon atom together, and wherein one or more carbon atoms are replaced by N, O or S; And X and Y are selected from N, O, S or C=C.

9. compound according to claim 1, wherein when Wa is H, Cl, Br, F, alkyl, CN, CO2H, Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are replaced by N, O or S; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces, and X and Y are selected from N, O, S or C=C.

10. compound according to claim 1, it has following formula (III):

Wherein R1 and R2 are H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, the optional alkyl that replaces or the compound of following formula independently:

Or

R6a and R6b are H, tert-butoxycarbonyl, benzyloxycarbonyl, ethanoyl, trifluoroacetyl group, alkyl, low alkyl group, the optional alkyl that replaces or the compound of following formula independently

Or

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroaralkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b;

M and n are 0,1,2 or 3 independently;

X and Y are O, N, S or C=C independently; With

R12a, R12b, R13a, R13b, R14a, R14b are H, Cl, Br, F, alkyl, cycloalkyl, hydroxyl, alkoxyl group, amino, alkylamino, cyano group or CO independently ₂H; And

Wherein when Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms can be replaced by N, O or S; And R11a and R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces independently; Or R11a and R11b form alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene (alkynlyene) or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; With

When Wa and Wb were not covalent attachment, Wa and Wb were H, Cl, Br, F, alkyl, CN, CO2H independently; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; Or Wa is H, Cl, Br, F, alkyl, CN, CO ₂H and Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are replaced by N, O or S; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

11. compound according to claim 11, wherein R3a and R3b are independently selected from H, hydroxyl, alkoxyl group, aryloxy, alkylamino, dialkylamino, amide group, sulfonamido or amidino groups.

12. compound according to claim 11, wherein the alkyl of the optional replacement of R5a and R5b is independently selected from alkoxylate and hydroxylated alkyl.

13. compound according to claim 1, it has following formula (IV):

Wherein R5a and R5b are H, alkyl, cycloalkyl, cycloalkylalkyl, Heterocyclylalkyl, aryl, aralkyl, heteroaryl, heteroarylalkyl independently; Or respectively use following radicals randomly to replace: hydroxyl, sulfydryl, halogen, amino, carboxyl, alkyl, haloalkyl, alkoxyl group or alkylthio; Or randomly, R5a is connected by following radicals with R5b: alkylidene group, alkenylene, the alkynylene bridge of the alkylidene group of 2-12 carbon atom, alkenylene, alkynylene or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms can be replaced by N, O or S;

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroarylalkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b;

X and Y are O, N, S or C=C independently;

M and n are 0,1,2 or 3 independently; With

Wherein when Wa and Wb are covalent attachment, Wa and Wb are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces, and wherein one or more carbon atoms are replaced by N, O or S; And R11a and R11b independently for do not exist, H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces; Or R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom together, and wherein one or more carbon atoms are replaced by N, O or S; With

When described Wa and Wb were not covalent attachment, Wa and Wb were H, Cl, Br, F, alkyl, CN, CO2H independently; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; Or Wa is H, Cl, Br, F, alkyl, CN, CO ₂H and Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces of 2-12 carbon atom together, and wherein one or more carbon atoms can be replaced by N, O or S; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

14. compound according to claim 14, wherein the alkyl of the optional replacement of R5a and R5b is independently selected from alkoxylate and hydroxylated alkyl.

15. compound according to claim 14, wherein R3a and R3b are selected from H, hydroxyl, alkoxyl group, aryloxy, alkylamino, dialkylamino, amide group, sulfonamido or amidino groups.

16. compound according to claim 1, it has following formula V:

M and n are 0,1,2 or 3 independently; With

When described Wa and Wb were not covalent attachment, Wa and Wb were H, Cl, Br, F, alkyl, CN, CO2H independently; And R11a and R11b form alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of 2-12 carbon atom or alkylidene group, alkenylene, alkynylene or the alkoxyl group alkylidene chain of optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; Or Wa is that H, Cl, Br, F, alkyl, CN, CO2H and Wb and R11a are alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

17. compound according to claim 16, wherein the alkyl of the optional replacement of R5a and R5b is independently selected from alkoxylate and hydroxylated alkyl.

18. compound according to claim 16, wherein R3a and R3b are selected from H, hydroxyl, alkoxyl group, aryloxy, alkylamino, dialkylamino, amide group, sulfonamido or amidino groups.

19. compound according to claim 1, it has following formula (VI):

R8a and R8b are H, hydroxyl, alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl or heteroarylalkyl independently, and wherein alkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heteroaryl and heteroaralkyl are respectively randomly replaced by following radicals: halogen, hydroxyl, sulfydryl, carboxyl, alkyl, alkoxyl group, amino and nitro; Or R8a and R7a can independently or form ring with R8b and R7b;

X is O, N, S or C=C; With

Wherein Wa and Wb are not covalent attachment, and Wa and Wb are H, Cl, Br, F, alkyl, CN, CO independently ₂H; And Wb and R11a be alkylidene group, alkenylene, the alkynylene chain of key, alkylidene group, alkenylene, alkynylene, aryl, aryl alkylene, aralkyl alkylidene group, heteroaryl, heteroaryl alkylidene group or optional 2-12 the carbon atom that replaces together, and wherein one or more carbon atoms are by N, O or S replacement; And R11b does not exist or be H, alkyl, optional alkyl, hydroxyalkyl, the alkoxyalkyl that replaces.

20. a pharmaceutical composition, it comprises:

Compound, described compound are selected from formula (I), (II), (III), (IV), (V) and compound (VI); With

Pharmaceutically acceptable vehicle.

21. one kind is brought out apoptotic method in cell, it comprises cell is contacted with the compound that is enough to bring out apoptotic amount in cell that described compound is selected from formula (I), (II), (III), (IV), (V) and compound (VI).