非标记的天然产物靶点识别和确证方法及应用
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摘要
天然产物是新药创制的重要来源,其靶点的识别及确证对阐明药物作用机制十分重要。常见的靶点识别和确证方法中,用探针标记天然产物的方法费时费力,可能会降低或改变天然产物的活性。近年来开发的无需对天然产物进行化学修饰的方法,逐渐成为天然产物靶点研究的重要手段。这些方法包括直接法和间接法,其中直接法主要基于蛋白质的亲和性、稳定性等原理,间接法则从生理反应或生化特征的改变推断药物靶点。本文对近年来非标记天然产物的靶点识别和确证研究中采用的方法进行综述,以期为天然产物靶点研究提供参考。
Natural products areone source of new drugs,and their target identification is pivotal forelucidating the mechanism of action.Most methods of target discovery and validation utilize labeling natural products with probes This is time-consuming and laborious,and often results in activity decrease or change of the natural products.Recent methods withoutchemicalmodificationhave become the main force in the target identification of natural products including direct or indirect methods.Direct methods are mainly based on the principle of affinity and stability of protein,and indirect methods infer the drug target from the change in physiological responses or biochemical signatures.This review summarizes recent methods for target identification and validation of label-freenatural products,providing new ideas and strategies for future research in natural products.
Natural products areone source of new drugs,and their target identification is pivotal forelucidating the mechanism of action.Most methods of target discovery and validation utilize labeling natural products with probes This is time-consuming and laborious,and often results in activity decrease or change of the natural products.Recent methods withoutchemicalmodificationhave become the main force in the target identification of natural products including direct or indirect methods.Direct methods are mainly based on the principle of affinity and stability of protein,and indirect methods infer the drug target from the change in physiological responses or biochemical signatures.This review summarizes recent methods for target identification and validation of label-freenatural products,providing new ideas and strategies for future research in natural products.
引文
[1]Newman DJ,Cragg GM.Natural products as sources of new drugs from 1981 to 2014[J].J Nat Prod,2016,79:629-661.
[2]Shen B.A new golden age of natural products drug discovery[J].Cell,2015,163:1297-1300.
[3]Paolini GV,Shapland RH,van Hoorn WP,et al.Global mapping of pharmacological space[J].Nat Biotechnol,2006,24:805-815.
[4]Mohamad SZ,Mohd F,Liggi S,et al.Global mapping of traditional Chinese medicine into bioactivity space and pathways annotation improves mechanistic understanding and discovers relationships between therapeutic action(sub)classes[J].Evid Based Complement Alternat Med,2016,2016:2106465.
[5]Yue R,Shan L,Yang X,et al.Approaches to target profiling of natural products[J].Curr Med Chem,2012,19:3841-3855.
[6]Koshelev IuA.Affinity chromatography and proteomic screening as the effective method for S100A4 new protein targets discov-ery[J].Mol Biol(Mosk),2014,48:868-872.
[7]Niphakis MJ,Cravatt BF.Enzyme inhibitor discovery by activity-based protein profiling[J].Annu Rev Biochem,2014,83:341-377.
[8]Zweerink S,Pollmann T,Ninck S,et al.Activity-based protein profiling with natural product-derived chemical probes in human cell lysates[J].Methods Mol Biol,2017,1491:23-46.
[9]Chen X,Wong YK,Wang J,et al.Target identification with quantitative activity based protein profiling(ABPP)[J].Proteomics,2017,17:1600212.
[10]Chang J,Kim Y,Kwon HJ.Advances in identification and validation of protein targets of natural products without chemical modification[J].Nat Prod Rep,2016,33:719-730.
[11]Schenone M,Dancik V,Wagner BK,et al.Target identification and mechanism of action in chemical biology and drug discovery[J].Nat Chem Biol,2013,9:232-240.
[12]Lee H,Lee JW.Target identification for biologically active small molecules using chemical biology approaches[J].Arch Pharm Res,2016,39:1193-1201.
[13]Long MJ,Poganik JR,Ghosh S,et al.Subcellular redox targeting:bridging in vitro and in vivo chemical biology[J].ACS Chem Biol,2017,12:586-600.
[14]Sleno L,Emili A.Proteomic methods for drug target discovery[J].Curr Opin Chem Biol,2008,12:46-54.
[15]Chernobrovkin A,Marin-Vicente C,Visa N,et al.Functional Identification of Target by Expression Proteomics(FITExP)reveals protein targets and highlights mechanisms of action of small molecule drugs[J].Sci Rep,2015,5:11176.
[16]Yang HQ,Li XJ.Chemical proteomics and discovery of drug targets[J].Acta Pharm Sin(药学学报),2011,46:877-882.
[17]Haupt VJ,Schroeder M.Old friends in new guise:repositioning of known drugs with structural bioinformatics[J].Brief Bioin-form,2011,12:312-326.
[18]Lomenick B,Hao R,Jonai N,et al.Target identification using drug affinity responsive target stability(DARTS)[J].Proc Natl Acad Sci U S A,2009,106:21984-21989.
[19]Lomenick B,Jung G,Wohlschlegel JA,et al.Target identifica-tion using drug affinity responsive target stability(DARTS)[J].Curr Protoc Chem Biol,2011,3:163-180.
[20]DeArmond PD,Xu Y,Strickland EC,et al.Thermodynamic analysis of protein-ligand interactions in complex biological mixtures using a shotgun proteomics approach[J].J Proteome Res,2011,10:4948-4958.
[21]Strickland EC,Geer MA,Tran DT,et al.Thermodynamic analysis of protein-ligand binding interactions in complex biological mixtures using the stability of proteins from rates of oxidation[J].Nat Protoc,2013,8:148-161.
[22]Molina MD,Jafari R,Ignatushchenko M,et al.Monitoring drug target engagement in cells and tissues using the cellular thermal shift assay[J].Science,2013,341:84-87.
[23]Jafari R,Almqvist H,Axelsson H,et al.The cellular thermal shift assay for evaluating drug target interactions in cells[J].Nat Protoc,2014,9:2100-2122.
[24]Savitski MM,Reinhard FB,Franken H,et al.Tracking cancer drugs in living cells by thermal profiling of the proteome[J].Science,2014,346:1255784.
[25]Kost GC,Yang MY,Li L,et al.A novel anti-cancer agent,1-(3,5-dimethoxyphenyl)-4-[(6-fluoro-2-methoxyquinoxalin-3-yl)ami-nocarbonyl]piperazine(RX-5902),interferes with beta-catenin function through Y593 phospho-p68 RNA helicase[J].J Cell Biochem,2015,116:1595-1601.
[26]Jung J,Kim Y,Song J,et al.KRIBB53 binds to OCT4 and enhances its degradation through the proteasome,causing apoptotic cell death of OCT4-positive testicular germ cell tumors[J].Carcinogenesis,2018,39:838-849
[27]Tanabe N,Kuboyama T,Tohda C.Matrine directly activates extracellular heat shock protein 90,resulting in axonal growth and functional recovery in spinal cord injured-mice[J].Front Pharmacol,2018,9:446.
[28]Lomenick B,Olsen RW,Huang J.Identification of direct protein targets of small molecules[J].ACS Chem Biol,2011,6:34-46.
[29]West GM,Tucker CL,Xu T,et al.Quantitative proteomics approach for identifying protein-drug interactions in complex mixtures using protein stability measurements[J].Proc Natl Acad Sci U S A,2010,107:9078-9082.
[30]Wallace MA,Kwon DY,Weitzel DH,et al.Discovery of manassantin A protein targets using large-scale protein folding and stability measurements[J].J Proteome Res,2016,15:2688-2696.
[31]Xu Y,Wallace MA,Fitzgerald MC.Thermodynamic analysis of the geldanamycin-Hsp90 interaction in a whole cell lysate using a mass spectrometry-based proteomics approach[J].J Am Soc Mass Spectrom,2016,27:1670-1676.
[32]Wang J,Wu J,Li X,et al.Identification and validation nucleolin as a target of curcumol in nasopharyngeal carcinoma cells[J].JProteomics,2018,182:1-11.
[33]Jin Y,Yoon YJ,Jeon YJ,et al.Geranylnaringenin(CG902)inhibits constitutive and inducible STAT3 activation through the activation of SHP-2 tyrosine phosphatase[J].Biochem Pharmacol,2017,142:46-57.
[34]Ishii T,Okai T,Iwatani-Yoshihara M,et al.CETSA quantitatively verifies in vivo target engagement of novel RIPK1 inhibitors in various biospecimens[J].Sci Rep,2017,7:13000.
[35]Franken H,Mathieson T,Childs D,et al.Thermal proteome profiling for unbiased identification of direct and indirect drug targets using multiplexed quantitative mass spectrometry[J].Nat Protoc,2015,10:1567-1593.
[36]Miettinen TP,Peltier J,Hartlova A,et al.Thermal proteome profiling of breast cancer cells reveals proteasomal activation by CDK4/6 inhibitor palbociclib[J].EMBO J,2018,37:e98359.
[37]Reinhard FB,Eberhard D,Werner T,et al.Thermal proteome profiling monitors ligand interactions with cellular membrane proteins[J].Nat Methods,2015,12:1129-1131.
[38]Nabet B,Roberts JM,Buckley DL,et al.The dTAG system for immediate and target-specific protein degradation[J].Nat Chem Biol,2018,14:431-441.
[39]Ngo VN,Davis RE,Lamy L,et al.A loss-of-function RNAinterference screen for molecular targets in cancer[J].Nature,2006,441:106-110.
[40]Wittrup A,Ai A,Liu X,et al.Visualizing lipid-formulated siRNA release from endosomes and target gene knockdown[J].Nat Biotechnol,2015,33:870-876.
[41]Henderson MC,Azorsa DO.High-throughput RNAi screening for the identification of novel targets[J].Methods Mol Biol,2013,986:89-95.
[42]Ngo VN.Identification of pathogenetically relevant genes in lymphomagenesis by shRNA library screens[J].Methods Mol Biol,2013,971:245-263.
[43]Shalem O,Sanjana NE,Hartenian E,et al.Genome-scale CRISPR-Cas9 knockout screening in human cells[J].Science,2014,343:84-87.
[44]Wang T,Wei JJ,Sabatini DM,et al.Genetic screens in human cells using the CRISPR-Cas9 system[J].Science,2014,343:80-84.
[45]Ran FA,Hsu PD,Wright J,et al.Genome engineering using the CRISPR-Cas9 system[J].Nat Protoc,2013,8:2281-2308.
[46]Kurata M,Yamamoto K,Moriarity BS,et al.CRISPR/Cas9library screening for drug target discovery[J].J Hum Genet,2018,63:179-186.
[47]Lu D,Li L,Deng XM.CRISPR/Cas9 and its application in drugs development[J].Acta Pharm Sin(药学学报),2018,53:11-20.
[48]Lamb J,Crawford ED,Peck D,et al.The connectivity map:using gene-expression signatures to connect small molecules,genes,and disease[J].Science,2006,313:1929-1935.
[49]Winter GE,Buckley DL,Paulk J,et al.Drug development.Phthalimide conjugation as a strategy for in vivo target protein degradation[J].Science,2015,348:1376-1381.
[50]Bondeson DP,Mares A,Smith IE,et al.Catalytic in vivo protein knockdown by small-molecule PROTACs[J].Nat Chem Biol,2015,11:611-617.
[51]You QD,Lu MC,Jiang ZY.Protein degradation as an innovative strategy in drug discovery[J].Acta Pharm Sin(药学学报),2017,52:1777-1782.
[52]Marine S,Bahl A,Ferrer M,et al.Common seed analysis to identify off-target effects in siRNA screens[J].J Biomol Screen,2012,17:370-378.
[53]Cho SW,Kim S,Kim Y,et al.Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases and nickases[J].Genome Res,2014,24:132-141.
[54]Kasap C,Elemento O,Kapoor TM.DrugTargetSeqR:a genomicsand CRISPR-Cas9-based method to analyze drug targets[J].Nat Chem Biol,2014,10:626-628.
[55]Luo L,Carson JD,Molnar KS,et al.Conformation-dependent ligand regulation of ATP hydrolysis by human KSP:activation of basal hydrolysis and inhibition of microtubule-stimulated hydrolysis by a single,small molecule modulator[J].J Am Chem Soc,2008,130:7584-7591.
[56]Wu D,Wang W,Chen W,et al.Pharmacologic inhibition of dihydroorotate dehydrogenase induces apoptosis and differentia-tion in acute myeloid leukemia cells[J].Haematologica,2018:188185.DOI:10.3324/haematol.2018.188185.
[57]Subramanian A,Narayan R,Corsello SM,et al.A next genera-tion connectivity map:L1000 platform and the first 1000000profiles[J].Cell,2017,171:1437-1452.e1417.
[58]Zhang XF,Kang YB,Su JH,et al.Application of connectivity map technology in traditional Chinese medicines[J].J Zhejiang Univ(Agric Life Sci)(浙江大学学报·农业与生命科学版),2016,42:543-550.
[59]Qu XA,Rajpal DK.Applications of connectivity map in drug discovery and development[J].Drug Discov Today,2012,17:1289-1298.
[60]Lv C,Wang YC,Liu RH,et al.Application of connectivity map database to research on Chinese materia medica[J].Chin Herb Med,2016,8:117-120.
[61]Lv C,Wu X,Wang X,et al.The gene expression profiles in response to 102 traditional Chinese medicine(TCM)compo-nents:a general template for research on TCMs[J].Sci Rep,2017,7:352.
[62]Lv C,Zeng HW,Wang JX,et al.The antitumor natural product tanshinone IIA inhibits protein kinase C and acts synergistically with 17-AAG[J].Cell Death Dis,2018,9:165.
[63]Wang L,Xiao ZY,Zhou WX,et al.Exploration of the experi-mental conditions of drug affinity responsive target stability[J].Int J Pharm Res(国际药学研究杂志),2016,43:485-490.
[64]Saxena C.Identification of protein binding partners of small molecules using label-free methods[J].Expert Opin Drug Discov,2016,11:1017-1025.
[2]Shen B.A new golden age of natural products drug discovery[J].Cell,2015,163:1297-1300.
[3]Paolini GV,Shapland RH,van Hoorn WP,et al.Global mapping of pharmacological space[J].Nat Biotechnol,2006,24:805-815.
[4]Mohamad SZ,Mohd F,Liggi S,et al.Global mapping of traditional Chinese medicine into bioactivity space and pathways annotation improves mechanistic understanding and discovers relationships between therapeutic action(sub)classes[J].Evid Based Complement Alternat Med,2016,2016:2106465.
[5]Yue R,Shan L,Yang X,et al.Approaches to target profiling of natural products[J].Curr Med Chem,2012,19:3841-3855.
[6]Koshelev IuA.Affinity chromatography and proteomic screening as the effective method for S100A4 new protein targets discov-ery[J].Mol Biol(Mosk),2014,48:868-872.
[7]Niphakis MJ,Cravatt BF.Enzyme inhibitor discovery by activity-based protein profiling[J].Annu Rev Biochem,2014,83:341-377.
[8]Zweerink S,Pollmann T,Ninck S,et al.Activity-based protein profiling with natural product-derived chemical probes in human cell lysates[J].Methods Mol Biol,2017,1491:23-46.
[9]Chen X,Wong YK,Wang J,et al.Target identification with quantitative activity based protein profiling(ABPP)[J].Proteomics,2017,17:1600212.
[10]Chang J,Kim Y,Kwon HJ.Advances in identification and validation of protein targets of natural products without chemical modification[J].Nat Prod Rep,2016,33:719-730.
[11]Schenone M,Dancik V,Wagner BK,et al.Target identification and mechanism of action in chemical biology and drug discovery[J].Nat Chem Biol,2013,9:232-240.
[12]Lee H,Lee JW.Target identification for biologically active small molecules using chemical biology approaches[J].Arch Pharm Res,2016,39:1193-1201.
[13]Long MJ,Poganik JR,Ghosh S,et al.Subcellular redox targeting:bridging in vitro and in vivo chemical biology[J].ACS Chem Biol,2017,12:586-600.
[14]Sleno L,Emili A.Proteomic methods for drug target discovery[J].Curr Opin Chem Biol,2008,12:46-54.
[15]Chernobrovkin A,Marin-Vicente C,Visa N,et al.Functional Identification of Target by Expression Proteomics(FITExP)reveals protein targets and highlights mechanisms of action of small molecule drugs[J].Sci Rep,2015,5:11176.
[16]Yang HQ,Li XJ.Chemical proteomics and discovery of drug targets[J].Acta Pharm Sin(药学学报),2011,46:877-882.
[17]Haupt VJ,Schroeder M.Old friends in new guise:repositioning of known drugs with structural bioinformatics[J].Brief Bioin-form,2011,12:312-326.
[18]Lomenick B,Hao R,Jonai N,et al.Target identification using drug affinity responsive target stability(DARTS)[J].Proc Natl Acad Sci U S A,2009,106:21984-21989.
[19]Lomenick B,Jung G,Wohlschlegel JA,et al.Target identifica-tion using drug affinity responsive target stability(DARTS)[J].Curr Protoc Chem Biol,2011,3:163-180.
[20]DeArmond PD,Xu Y,Strickland EC,et al.Thermodynamic analysis of protein-ligand interactions in complex biological mixtures using a shotgun proteomics approach[J].J Proteome Res,2011,10:4948-4958.
[21]Strickland EC,Geer MA,Tran DT,et al.Thermodynamic analysis of protein-ligand binding interactions in complex biological mixtures using the stability of proteins from rates of oxidation[J].Nat Protoc,2013,8:148-161.
[22]Molina MD,Jafari R,Ignatushchenko M,et al.Monitoring drug target engagement in cells and tissues using the cellular thermal shift assay[J].Science,2013,341:84-87.
[23]Jafari R,Almqvist H,Axelsson H,et al.The cellular thermal shift assay for evaluating drug target interactions in cells[J].Nat Protoc,2014,9:2100-2122.
[24]Savitski MM,Reinhard FB,Franken H,et al.Tracking cancer drugs in living cells by thermal profiling of the proteome[J].Science,2014,346:1255784.
[25]Kost GC,Yang MY,Li L,et al.A novel anti-cancer agent,1-(3,5-dimethoxyphenyl)-4-[(6-fluoro-2-methoxyquinoxalin-3-yl)ami-nocarbonyl]piperazine(RX-5902),interferes with beta-catenin function through Y593 phospho-p68 RNA helicase[J].J Cell Biochem,2015,116:1595-1601.
[26]Jung J,Kim Y,Song J,et al.KRIBB53 binds to OCT4 and enhances its degradation through the proteasome,causing apoptotic cell death of OCT4-positive testicular germ cell tumors[J].Carcinogenesis,2018,39:838-849
[27]Tanabe N,Kuboyama T,Tohda C.Matrine directly activates extracellular heat shock protein 90,resulting in axonal growth and functional recovery in spinal cord injured-mice[J].Front Pharmacol,2018,9:446.
[28]Lomenick B,Olsen RW,Huang J.Identification of direct protein targets of small molecules[J].ACS Chem Biol,2011,6:34-46.
[29]West GM,Tucker CL,Xu T,et al.Quantitative proteomics approach for identifying protein-drug interactions in complex mixtures using protein stability measurements[J].Proc Natl Acad Sci U S A,2010,107:9078-9082.
[30]Wallace MA,Kwon DY,Weitzel DH,et al.Discovery of manassantin A protein targets using large-scale protein folding and stability measurements[J].J Proteome Res,2016,15:2688-2696.
[31]Xu Y,Wallace MA,Fitzgerald MC.Thermodynamic analysis of the geldanamycin-Hsp90 interaction in a whole cell lysate using a mass spectrometry-based proteomics approach[J].J Am Soc Mass Spectrom,2016,27:1670-1676.
[32]Wang J,Wu J,Li X,et al.Identification and validation nucleolin as a target of curcumol in nasopharyngeal carcinoma cells[J].JProteomics,2018,182:1-11.
[33]Jin Y,Yoon YJ,Jeon YJ,et al.Geranylnaringenin(CG902)inhibits constitutive and inducible STAT3 activation through the activation of SHP-2 tyrosine phosphatase[J].Biochem Pharmacol,2017,142:46-57.
[34]Ishii T,Okai T,Iwatani-Yoshihara M,et al.CETSA quantitatively verifies in vivo target engagement of novel RIPK1 inhibitors in various biospecimens[J].Sci Rep,2017,7:13000.
[35]Franken H,Mathieson T,Childs D,et al.Thermal proteome profiling for unbiased identification of direct and indirect drug targets using multiplexed quantitative mass spectrometry[J].Nat Protoc,2015,10:1567-1593.
[36]Miettinen TP,Peltier J,Hartlova A,et al.Thermal proteome profiling of breast cancer cells reveals proteasomal activation by CDK4/6 inhibitor palbociclib[J].EMBO J,2018,37:e98359.
[37]Reinhard FB,Eberhard D,Werner T,et al.Thermal proteome profiling monitors ligand interactions with cellular membrane proteins[J].Nat Methods,2015,12:1129-1131.
[38]Nabet B,Roberts JM,Buckley DL,et al.The dTAG system for immediate and target-specific protein degradation[J].Nat Chem Biol,2018,14:431-441.
[39]Ngo VN,Davis RE,Lamy L,et al.A loss-of-function RNAinterference screen for molecular targets in cancer[J].Nature,2006,441:106-110.
[40]Wittrup A,Ai A,Liu X,et al.Visualizing lipid-formulated siRNA release from endosomes and target gene knockdown[J].Nat Biotechnol,2015,33:870-876.
[41]Henderson MC,Azorsa DO.High-throughput RNAi screening for the identification of novel targets[J].Methods Mol Biol,2013,986:89-95.
[42]Ngo VN.Identification of pathogenetically relevant genes in lymphomagenesis by shRNA library screens[J].Methods Mol Biol,2013,971:245-263.
[43]Shalem O,Sanjana NE,Hartenian E,et al.Genome-scale CRISPR-Cas9 knockout screening in human cells[J].Science,2014,343:84-87.
[44]Wang T,Wei JJ,Sabatini DM,et al.Genetic screens in human cells using the CRISPR-Cas9 system[J].Science,2014,343:80-84.
[45]Ran FA,Hsu PD,Wright J,et al.Genome engineering using the CRISPR-Cas9 system[J].Nat Protoc,2013,8:2281-2308.
[46]Kurata M,Yamamoto K,Moriarity BS,et al.CRISPR/Cas9library screening for drug target discovery[J].J Hum Genet,2018,63:179-186.
[47]Lu D,Li L,Deng XM.CRISPR/Cas9 and its application in drugs development[J].Acta Pharm Sin(药学学报),2018,53:11-20.
[48]Lamb J,Crawford ED,Peck D,et al.The connectivity map:using gene-expression signatures to connect small molecules,genes,and disease[J].Science,2006,313:1929-1935.
[49]Winter GE,Buckley DL,Paulk J,et al.Drug development.Phthalimide conjugation as a strategy for in vivo target protein degradation[J].Science,2015,348:1376-1381.
[50]Bondeson DP,Mares A,Smith IE,et al.Catalytic in vivo protein knockdown by small-molecule PROTACs[J].Nat Chem Biol,2015,11:611-617.
[51]You QD,Lu MC,Jiang ZY.Protein degradation as an innovative strategy in drug discovery[J].Acta Pharm Sin(药学学报),2017,52:1777-1782.
[52]Marine S,Bahl A,Ferrer M,et al.Common seed analysis to identify off-target effects in siRNA screens[J].J Biomol Screen,2012,17:370-378.
[53]Cho SW,Kim S,Kim Y,et al.Analysis of off-target effects of CRISPR/Cas-derived RNA-guided endonucleases and nickases[J].Genome Res,2014,24:132-141.
[54]Kasap C,Elemento O,Kapoor TM.DrugTargetSeqR:a genomicsand CRISPR-Cas9-based method to analyze drug targets[J].Nat Chem Biol,2014,10:626-628.
[55]Luo L,Carson JD,Molnar KS,et al.Conformation-dependent ligand regulation of ATP hydrolysis by human KSP:activation of basal hydrolysis and inhibition of microtubule-stimulated hydrolysis by a single,small molecule modulator[J].J Am Chem Soc,2008,130:7584-7591.
[56]Wu D,Wang W,Chen W,et al.Pharmacologic inhibition of dihydroorotate dehydrogenase induces apoptosis and differentia-tion in acute myeloid leukemia cells[J].Haematologica,2018:188185.DOI:10.3324/haematol.2018.188185.
[57]Subramanian A,Narayan R,Corsello SM,et al.A next genera-tion connectivity map:L1000 platform and the first 1000000profiles[J].Cell,2017,171:1437-1452.e1417.
[58]Zhang XF,Kang YB,Su JH,et al.Application of connectivity map technology in traditional Chinese medicines[J].J Zhejiang Univ(Agric Life Sci)(浙江大学学报·农业与生命科学版),2016,42:543-550.
[59]Qu XA,Rajpal DK.Applications of connectivity map in drug discovery and development[J].Drug Discov Today,2012,17:1289-1298.
[60]Lv C,Wang YC,Liu RH,et al.Application of connectivity map database to research on Chinese materia medica[J].Chin Herb Med,2016,8:117-120.
[61]Lv C,Wu X,Wang X,et al.The gene expression profiles in response to 102 traditional Chinese medicine(TCM)compo-nents:a general template for research on TCMs[J].Sci Rep,2017,7:352.
[62]Lv C,Zeng HW,Wang JX,et al.The antitumor natural product tanshinone IIA inhibits protein kinase C and acts synergistically with 17-AAG[J].Cell Death Dis,2018,9:165.
[63]Wang L,Xiao ZY,Zhou WX,et al.Exploration of the experi-mental conditions of drug affinity responsive target stability[J].Int J Pharm Res(国际药学研究杂志),2016,43:485-490.
[64]Saxena C.Identification of protein binding partners of small molecules using label-free methods[J].Expert Opin Drug Discov,2016,11:1017-1025.