Using Protein Structure to Study Network Pharmacology

Hauptman Woodward InstituteNovember 5, 2009Philip E. BourneUniversity of California San Diegopbourne@ucsd.edu

Big Questions in the LabCan we improve how science is disseminated and comprehended?What is the ancestry of the protein structure universe and what can we learn from it?Are there alternative ways to represent proteins from which we can learn something new?What really happens when we take a drug? August 14, 2009

The truth is we know very little about how the major drugs we take workWe know even less about what side effects they might haveDrug discovery seems to be approached in a very consistent and conventional wayThe cost of bringing a drug to market is huge >$800M The cost of failure is even higher e.g. Vioxx - >$4.85Bn

Motivation

The truth is we know very little about how the major drugs we take work receptors are unknownWe know even less about what side effects they might have - receptors are unknownDrug discovery seems to be approached in a very consistent and conventional wayThe cost of bringing a drug to market is huge ~$800M drug reuse is a big businessThe cost of failure is even higher e.g. Vioxx - $4.85Bn - fail early and cheaply

Motivation

A.L. Hopkins Nat. Chem. Biol. 2008 4:682-690Why Dont we Do Better?A Couple of ObservationsGene knockouts only effect phenotype in 10-20% of cases , why? redundant functions alternative network routes robustness of interaction networks

35% of biologically active compounds bind to more than one target

Paolini et al. Nat. Biotechnol. 2006 24:805815

ImplicationsEhrlichs philosophy of magic bullets targeting individual chemoreceptors has not been realized

Stated another way The notion of one drug, one target, one disease is a little nave in a complex system

How Can we Begin to Address the Problem?Systematic screening for multiple targetsIntegration of knowledge from multiple sourcesAnalyze the impact on the complete network

2. What is the ancestry of the protein structure universe?

4. What really happens when we take a drug? Valas, Yang & Bourne 2009 Current Opinions in Structural Biology 19:1-6

*PKAPhosphoinositide-3 Kinase (D) and Actin-Fragmin Kinase (E)ChaK (Channel Kinase)E. Scheeff and P.E. Bourne 2005 PLoS Comp. Biol. 1(5): e49.

ImplicationsThe ATP binding cassette is preserved yet the enzyme has evolved to bind a variety of different substratesThe evolutionary history of the protein kinase-like superfamily can be traced with careful analysisSo taking this a step further The Role of Evolution

What if only the binding pocket was conserved and the global structure of the protein has changed?A drug could potentially bind to distinctly different gene familiesThe Role of Evolution

If this is True What ifWe can characterize a protein-ligand binding site from a 3D structure (primary site) and search for that site on a proteome wide scale?

We could perhaps find alternative binding sites (off-targets) for existing pharmaceuticals and NCEs?

We could use it for lead optimization and possible ADME/Tox prediction

We might be able to construct a site similarity network for a given proteome to define multiple targets for dirty drugsThe Role of Evolution

What Do Off-targets Tell Us?One of four things:NothingA possible explanation for a side-effect of a drugA possible repositioning of a drug to treat a completely different conditionA multi-target strategy to attack a pathogen

Today I will give you examples of 2, 3 and 4 while illustrating the complexity of the problemThe Role of Evolution

AgendaComputational Methodology

Side Effects - The Tamoxifen Story

Repositioning an Existing Drug - The TB Story

Salvaging $800M The Torcetrapib Story

The Future? - The TB Drugome

Swiss-Prot - 20 Year Celebrationwww.pdb.org info@rcsb.org

Need to Start with a 3D Drug-Receptor Complex - The PDB Contains Many ExamplesComputational Methodology

Generic NameOther NameTreatmentPDBidLipitorAtorvastatinHigh cholesterol1HWK, 1HW8TestosteroneTestosteroneOsteoporosis1AFS, 1I9J ..TaxolPaclitaxelCancer1JFF, 2HXF, 2HXHViagraSildenafil citrateED, pulmonary arterial hypertension1TBF, 1UDT, 1XOS..DigoxinLanoxinCongestive heart failure1IGJ

Swiss-Prot - 20 Year Celebrationwww.pdb.org info@rcsb.org

A Reverse Engineering Approach to Drug Discovery Across Gene FamiliesCharacterize ligand binding site of primary target (Geometric Potential)Identify off-targets by ligand binding site similarity(Sequence order independent profile-profile alignment)

Extract known drugs or inhibitors of the primary and/or off-targetsSearch for similar small moleculesDock molecules to both primary and off-targetsStatistics analysis of docking score correlationsComputational Methodology

What we Search AgainstThe Human Target ListComputational Methodology

Initially assign Ca atom with a value that is the distance to the environmental boundary

Update the value with those of surrounding Ca atoms dependent on distances and orientation atoms within a 10A radius define i

Conceptually similar to hydrophobicity or electrostatic potential that is dependant on both global and local environmentsCharacterization of the Ligand Binding Site - The Geometric PotentialXie and Bourne 2007 BMC Bioinformatics, 8(Suppl 4):S9Computational Methodology

Discrimination Power of the Geometric Potential

Geometric potential can distinguish binding and non-binding sites

1000Geometric Potential ScaleComputational MethodologyXie and Bourne 2007 BMC Bioinformatics, 8(Suppl 4):S9

Chart4

0.40951122851.8002407617

0.40951122852.850837194

0.51519154563.1919162441

0.58124174373.346952176

0.58124174373.4272060701

0.59445178343.2794659468

0.713342143.2010360048

0.83223249672.9894575566

0.6076618232.967570131

0.52840158522.876372524

0.713342142.7997665341

0.66050198152.6392587459

0.79260237782.8198300077

0.80581241742.6757377886

0.91149273452.5553569474

1.2153236462.4094407763

0.87186261562.3109473607

0.88507265522.3419545471

0.97754293262.3000036479

1.03038309112.0957210083

0.92470277412.1632072374

0.9643328931.9808120235

0.76618229851.8020647138

1.24174372521.7801772881

0.9643328931.687155729

1.00396301191.6360850691

1.04359313081.6415569255

0.83223249671.5868383614

1.01717305151.5102323715

1.09643328931.4318024295

1.26816380451.2694706891

0.93791281371.2767664976

1.01717305151.2840623062

1.10964332891.307773684

1.16248348751.1527377522

1.42668428011.1454419436

1.51915455751.1527377522

1.75693527081.032356911

1.55878467641.0578922409

1.79656538970.9393353518

1.90224570671.0688359537

1.70409511230.8645533141

1.63804491410.8353700799

2.04755614270.7952431328

1.94187582560.7022215737

2.11360634080.7277569037

2.10039630120.7022215737

2.19286657860.6164958231

2.20607661820.6055521103

2.29854689560.6037281582

2.40422721270.6055521103

2.45706737120.5690730675

2.39101717310.5325940247

2.35138705420.5307700726

2.45706737120.5125305512

2.49669749010.4322766571

2.65521796570.4523401306

2.19286657860.3939736621

1.91545574640.3939736621

2.08718626160.3611425236

1.77014531040.2991281509

1.84940554820.3593185715

1.86261558780.3301353372

1.87582562750.2735928209

1.54557463670.2298179696

1.24174372520.2790646773

1.09643328930.222522161

1.05680317040.2298179696

1.08322324970.1787473097

1.10964332890.1787473097

0.84544253630.1769233575

0.6076618230.1459161712

0.54161162480.1331485062

0.50198150590.1386203626

0.19815059450.0948455113

0.356671070.091197607

0.17173051520.1112610805

0.14531043590.0729580856

0.17173051520.0839017984

0.06605019820.0729580856

0.06605019820.052894612

0.05284015850.0419508992

0.07926023780.0273592821

0.05284015850.0218874257

0.02642007930.012767665

0.02642007930.0164155693

00.0200634735

00.0145916171

00.0091197607

00.0091197607

00.0109437128

00.0091197607

00.0109437128

00.0036479043

00.0072958086

00.0018239521

00

00.0018239521

00.0018239521

00

00

binding site

non-binding site

Geometric Potential

Sheet1

Orient Sphere PotentialWithout Ligand BindingWith Ligand Binding

00.1002190920.0321850333

10.04326181490.0126659732

20.04114171270.0124824084

30.03947639230.0126353791

40.03744591860.0129719146

50.03545527980.0109221073

60.03350447590.011075078

70.03235590280.0105855718

80.03077689130.0116563666

90.02865125650.0117481491

100.02792094980.0114116135

110.02655881730.0126353791

120.02559392740.0122988435

130.02437674970.0123600318

140.02324145490.0119317139

150.02200435970.0123600318

160.02121098120.012849538

170.02026822170.0125435966

180.01882863240.0111668604

190.01828311550.0130331029

200.01721199920.0126965673

210.01631239420.0125741908

220.0155577440.0130331029

230.01478317640.0131860735

240.01392229980.0136449856

250.01343321570.0138897387

260.01273499830.0133696384

270.01218284220.0140733036

280.01134409610.013063697

290.01102209730.0134920149

300.01063260040.0123600318

310.00972193020.0139203329

320.00941320970.0130331029

330.00892523210.0134308267

340.00858110280.0126965673

350.00815841010.0131860735

360.00777001980.0135532032

370.00720458540.013063697

380.00701979570.0126659732

390.00660263570.0125435966

400.00656612040.0123294377

410.00624633460.0134614208

420.00573401350.0127577556

430.00545959520.0130025087

440.00518628350.0119929022

450.0050081330.0127577556

460.00486317820.0130942911

470.00441060940.0115951784

480.00437741360.0121458729

490.00416828030.0134308267

500.00389607520.0125130025

510.00377103780.0117175549

520.00365927880.0119929022

530.00334391910.010860919

540.00319675120.0113198311

550.00308831180.0116563666

560.00299204410.0106161659

570.00275192810.0108915132

580.00265566040.0103714128

590.00250406650.011075078

600.00241443790.010860919

610.00233808770.0111056722

620.00230157240.0100348773

630.00214112620.0087805177

640.00204485850.0089334883

650.00201387580.0096677477

660.00181470130.0085357645

670.00174277720.0079238818

680.00165868120.008627547

690.00157679840.0085663587

700.00145729370.0089946766

710.00139200870.0070366518

720.00129352790.0076179404

730.0011651710.0069142752

740.00116738410.0057822921

750.00109656650.0059964511

760.0010091510.0063023925

770.00102132270.0052621918

780.00102353580.0053539742

790.00090292450.0059046687

800.00081993520.0056599156

810.00076350240.0051398152

820.00074911750.0044667442

830.0007247740.00443615

840.00067719340.0044667442

850.00060194970.0042525852

860.00057207350.0041608028

870.0005289190.0037018907

880.00049461680.0022945604

890.00045035570.0031206021

900.00040056210.0033959493

910.00041605350.0029982255

920.00034191630.0029064431

930.00036625980.0022333721

940.00034744890.0019580248

950.00027995090.0018968366

960.00029433570.0019274307

970.00023679640.0013155479

980.00022573110.0012237655

990.00020802670.0014379245

1000.00193531250.0142262742

Sheet1

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Without Ligand Binding

With Ligand Binding

Sheet2

geom potentialbinding residuenon-binding residuebinding trianglenon-binding trianglebinding tetrahedronnon-binding tetrahedron

61.02974828384.0101344380.50558185754.38698102160.28529366974.768877981310085246934699566281202495

70.99603628643.61140621060.57718160034.00906952350.28529366974.39374053419754725239590989281186566

81.0798054923.30339878170.49827576133.66138389540.31473678874.032403736310574322234183098310171223

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101.05937397842.79835831280.59471623123.09141034530.36651606683.375954417410373661440770162361143349

111.06448185682.55386308570.56695306562.82444453350.36651606683.109997534310423341538864103361132056

121.12884112462.37815363690.59763866972.60801672910.39595918572.874044110711053111640959191390122037

131.0767407652.20145061560.66923841252.42661744760.39088278592.65589493810542880445855074385112774

141.11556064072.0513447620.6721608512.23371823580.42337174482.444316381510922684046050696417103790

151.22384766261.92607821710.58302647732.07338012910.42134118482.26198766561198252013994705741596048

161.16357469761.76534878210.66047109711.95617777570.47718158282.10247947741139230984524439747089275

171.2003514221.68425799250.7203810861.80518060840.49038022231.94982451271175220374934097048382793

181.15131578951.56885074250.72914840141.66709332440.51373166151.8085444861127205274993783650676794

191.20648087611.47507279830.78759717111.56830848360.54317478041.66985502491181193005393559453570905

201.19115724091.38695057360.83435618681.479613620.56652621961.55271435931166181475713358155865931

211.19830827071.28866333950.8416622831.36351279250.59799989851.4534014951173168615763094658961714

221.19626511931.21169969660.84458472151.2927067090.63251941721.34732960971171158545782933962357210

231.19524354361.14864606660.85919691391.20418808950.63251941721.25755473691170150295882733062353398

241.18196305981.08796172450.91326202581.12267517460.65587085641.17331425421157142356252548064649821

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261.18502778690.97370090420.97901689171.00705901840.82745317021.06491085981160127406702285681545218

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281.19217881660.87617795651.02577590740.89968227590.81730037060.94291877141167114647022041980540038

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301.17991990850.77919000921.01262493420.79380360850.99192852430.83055887681155101956931801697735267

311.24938705460.73531996851.0476941960.74357402850.98989796440.7766986633122396217171687697532980

321.17991990850.72286209981.03015956510.71559527121.00614244380.7332478133115594587051624199131135

331.27084014380.67715777161.01408615350.67845181860.9990354840.6944128801124488606941539898429486

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351.23610657080.61418057031.14559588520.62073186261.0548758820.60520322161210803678414088103925698

361.27799117360.59102269171.14267344670.59967068781.07619676130.56832298791251773378213610106024132

371.32702680610.55181479811.14705710440.56684521671.05386060210.54345355831299722078512865103823076

381.2258908140.53125549331.19089368170.52943739791.15132747860.5105769261200695181512016113421680

391.24223602480.51222476141.22596294350.50295671581.13508299910.48142128791216670283911415111820442

401.20954560310.47943687381.18212636620.47365612741.20716787650.45127652471184627380910750118919162

411.2861637790.44940041731.23034660120.45448957711.20716787650.43439074731259588084210315118918445

421.23814972210.44405041231.19673855870.43065255711.23864155540.3991826059121258108199774122016950

431.22384766260.40805252181.30340756330.40624274371.2782374740.3839689207119853398929220125916304

441.22997711670.39704679731.31655853640.38099577061.30971115290.3625143276120451959018647129015393

451.25960281140.37962106681.37062364840.36680811731.2782374740.3392934397123349679388325125914407

461.20341614910.36479391021.30340756330.35332544051.29752779330.32271382117847738928019127813703

471.2861637790.34308817571.33117072890.32732942981.34930707140.3060870991125944899117429132912997

481.24734390320.3359803121.46998655680.31979499281.42849890860.28696401491221439610067258140712185

491.23610657080.31190528961.44368461040.30274337221.40210162950.2726217018121040819886871138111576

501.24019287350.29371527271.49482728390.28022818331.45185034770.25750221911214384310236360143010934

511.24427917620.29547313151.40277047170.27304623461.46200314740.245373662121838669606197144010419

521.18604936250.27835311561.50505581860.26678956761.51784354540.2314081584116136421030605514959826

531.17379045440.25618880931.40861534860.24559620971.5828214630.22175241411493352964557415599416

541.19013566530.24992166061.4714477760.23872268821.53814914460.2093648004116532701007541815158890

551.24836547890.24189665321.41884388330.22290477661.48840042640.198437323712223165971505914668426

561.27186171950.23150235781.46706411830.21092017511.65896746030.1896058502124530291004478716348051

571.15438051650.2192737751.46998655680.20373822631.5828214630.1771240342113028691006462415597521

581.10738803530.21484091381.41153778710.19994897731.62140210160.168692920710842811966453815977163

591.13088427590.20054875771.45245192590.18778813161.66099802020.156093351711072624994426216366628

601.14722948680.19565732451.49628850310.17597977421.7229300980.1507473664112325601024399416976401

611.10738803530.18342874181.48021509150.16386298961.70465505860.1440354465108424001013371916796116

621.11658221640.18350517041.53135776490.16293770781.66505914010.1377003361109324011048369816405847

631.07367603790.17517444841.48459874920.15108528941.78181633590.1272438714105122921016342917555403

641.07469761360.15973586261.37208486760.14394740171.78283161580.122957662910522090939326717565221

651.09104282450.1564494311.41738266410.13376930261.73815929740.11356097510682047970303617124822

660.98888525660.14987656771.43930095270.13147812881.74018985740.10670775159681961985298417144531

671.02872670810.13963512971.3501665790.12751263561.64678410070.102209587610071827924289416224340

681.01544622430.13428512471.37354608690.12204906721.64678410070.09954837029941757940277016224227

690.98786368090.13344440961.44952948740.11372153161.65490634040.09034008719671746992258116303836

700.98684210530.12633654591.47875387220.11270812781.70160921870.085818372696616531012255816763644

710.90715920240.11991653991.47290899530.10446871421.71988425810.082450637488815691008237116943501

720.89387871850.10944581591.36185633290.09737488761.70770089850.08014267898751432932221016823403

730.90000817260.10722938531.3925419370.09874077971.64678410070.07364271448811403953224116223127

740.89694344560.09813437681.42176632180.09142664781.64069242090.0682967298781284973207516162900

750.87242562930.09790509091.39400315620.08843049751.56759226360.06535290458541281954200715442775

760.80908793720.08888651111.34578292130.0790895581.5543936240.06097249367921163921179515312589

770.81215266430.08560007951.30048512480.0778117881.61835626170.05767541017951120890176615942449

780.73859921540.08315436291.32532585190.07547655311.5005837860.05181131177231088907171314782200

790.73246976140.07704007151.25810976680.07499188181.50870602570.04783126097171008861170214862031

800.71918927750.07642864241.23180782040.06512220991.37671963040.04366280547041000843147813561854

810.68139097740.06840363491.19089368170.05979082461.29346667340.039659204667895815135712741684

820.60783752860.06504077471.0900695540.05855711571.31275699270.0365740759595851746132912931553

830.66198103960.06236577221.10468174640.05454756151.15538859840.0322643168648816756123811381370

840.59047074210.06381791641.09153077330.05067019041.04675364230.0295795489578835747115010311256

850.5812765610.0525829060.99362908410.04758591791.06299812170.0260940606569688680108010471108

860.56901765280.05434076470.93079665670.04362042480.91070612720.0206067717557711637990897875

870.56084504740.05021361810.80367058270.03718751360.87923244830.0192172865549657550844866816

880.48933474990.04746218690.75545034780.03251704390.7777044520.0155198429479621517738766659

890.47809741750.04020146590.62832427380.0301818090.68226813540.0140361553468526430685672596

900.43621281460.03806146390.59179379270.02546727830.6396263770.0107626225427498405578630457

910.4096518470.03691503430.44421064940.02066462550.4995177420.0087843724401483304469492373

920.37900457670.03072431420.40183529140.01696349850.4172800650.006358661371402275385411270

930.35755148740.03225288710.29516628670.0137470430.34722574750.0046630181350422202312342198

940.36163779010.03103002880.24694605180.01017809910.24468247120.0029909257354406169231241127

950.31260215760.02606216710.16511777430.0075784980.21117823240.001931148930634111317220882

960.31055900620.02354002190.14466070490.00559575150.17259759380.00113042863043089912717048

970.28604118990.02224073490.07598340050.00326051660.10761967614.47E-04280291527410619

980.26969597910.02285216410.0365304810.00176244140.07208487741.65E-042642992540717

990.26458810070.01926001790.01607341173.97E-040.04061119852.36E-05259252119401

100000000179332518011420

total100100100100100100978881308410684362269579984954246177

Sheet2

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

binding residue

non-binding residue

Geometry Potential

Ratio (%)

Sheet4

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

00

binding tetrahedron

non-binding tetrahedron

Geometry Potential

Ratio (%)

Sheet6

binding residuenon-binding residuebinding trianglenon-binding trianglebinding tetrahedronnon-binding tetrahedronbinding sitenon-binding site

01.0347.5981.50614041864.51998286341.10881617572.8993415450.40951122851.800240761799310746219511310110215068831987

11.1025.6871.00409361244.9170264770.77182302424.07910387630.40951122852.8508371946368043313012303671212004311563

21.19011040785.1871.07360778564.96989901860.63050331564.65051397660.51519154563.19191624415817335613912435958241702391750

31.20444908744.7361.09677917664.83230252530.83704750524.82323767760.58124174373.3469521765886697914212091677250679441835

41.12865892384.381.10450297374.59119893980.97836721384.73596189420.58124174373.42720607015516194114311488390246143441879

51.16348143144.0061.11995056774.31616580960.9674964674.54636410550.59445178343.27946594685685665614510800189236289451798

61.13890083783.7471.12767436474.01651477631.13055766934.3291174410.713342143.201036004855652994146100503104224998541755

71.21469100153.4381.13539816173.74563791541.01097945434.08535710150.83223249672.9894575566593486221479372593212329631639

81.1470943693.1951.26670271113.51704310211.10881617573.8222983470.6076618232.9675701315604518316488005102198657461627

91.17986849382.94401513691.2512551173.24356857281.0870746823.57946156080.52840158522.8763725245764163716281162100186036401577

101.13890083782.75353126971.18174094383.04178968961.0870746823.36042551180.713342142.79976653415563894315376113100174652541535

111.1266105412.55201715630.99636981542.82554379190.88053049243.11556845280.66050198152.6392587459550360931297070281161926501447

121.14914275182.40544215380.98864601842.65393782540.94575497342.94372982370.79260237782.8198300077561340201286640887152995601546

131.11227186142.2483318521.03498880052.47965426230.92401347972.73879720270.80581241742.6757377886543317981346204785142344611467

141.16348143142.10225179651.09677917662.35076994891.05446244162.57205735730.91149273452.5553569474568297321425882297133678691401

151.17986849381.97851506021.11995056772.21776933021.02185020112.4239809841.2153236462.4094407763576279821455549494125982921321

161.18396525941.8728085340.88823665712.05891192070.95662572022.271440770.87186261562.3109473607578264871155151988118054661267

171.14914275181.7815261761.11222677071.95384622760.90227198612.13123384020.88507265522.3419545471561251961444889083110767671284

181.18396525941.68981957771.14312195881.83623179830.76095227742.00399513750.97754293262.3000036479578238991484594770104154741261

191.11432024421.59754732580.88823665711.73660122671.07620393521.88033520371.03038309112.095721008354422594115434549997727781149

201.21878776711.50470942020.93457943931.65491454871.18491140341.77374176450.92470277412.1632072374595212811214141010992187701186

211.30277146191.41109374081.05816019161.56903163720.93488422651.66595540240.9643328931.980812023563619957137392618686585731086

221.19625555621.34646781111.09677917661.46924120910.88053049241.56984814090.76618229851.80206471385841904314236764818159058988

231.19215879061.28721559110.93457943931.39394879161.04359169471.48963369181.24174372521.78017728815821820512134880967742194976

241.12865892381.22223612791.10450297371.31126301031.03272094791.39335326410.9643328931.6871557295511728614332811957241773925

251.15528790021.17839796991.06588398861.25343491681.31536036531.32903162751.00396301191.636085069156416666138313641216907476897

261.1061267131.10493370191.18174094381.19324893981.16316990981.26703811471.04359313081.641556925554015627153298581076585279900

271.08564288491.07346921751.11222677071.13074504321.28274812481.1936925930.83223249671.586838361453015182144282941186204063870

281.15119113461.01202508961.02726500351.07707321911.01097945431.13856800761.01717305151.51023237155621431313326951935917577828

291.17372334540.98119696561.07360778561.03227343141.06533318841.0669733891.09643328931.43180242955731387713925830985545483785

301.22083614990.94598502571.28215030510.98623475571.09794542891.0160817561.26816380451.269470689159613379166246781015280996696

311.22902968110.87994496191.10450297370.94259392771.35884335250.96624836120.93791281371.276766497660012445143235861255021971700

321.28023925110.87138945041.11222677070.89167962841.07620393520.91318252991.01717305151.28406230626251232414422312994746177704

331.28638439950.81312712431.09677917660.85463288151.32623111210.86350306061.10964332891.30777368462811500142213851224487984717

341.25156189190.7883797771.20491233490.82022376711.29361887160.82175075681.16248348751.152737752261111150156205241194270988632

351.27614248550.75946073411.13539816170.78673382771.21752364390.77105153081.42668428011.1454419436623107411471968611240074108628

361.26794895430.71427914871.27442650810.75288421111.28274812480.73003037341.51915455751.1527377522619101021651883911837942115632

371.1983039390.7024004221.2589789140.71311990361.20665289710.69939919011.75693527081.03235691158599341631784411136350133566

381.23722321230.67970356921.39800726040.67783157841.56538754210.66655532721.55878467641.057892240960496131811696114434643118580

391.28228763390.63049170151.11222677070.65237442871.47842156760.63101776721.79656538970.939335351862689171441632413632796136515

401.16143304860.62497657841.29759789910.64170400651.13055766930.5952493191.90224570671.068835953756788391681605710430937144586

411.24131997790.59167371971.13539816170.59770350131.34797260570.57200656181.70409511230.864553314160683681471495612429729129474

421.21469100150.56346174381.34394068120.56093650341.21752364390.54049030671.63804491410.835370079959379691741403611228091124458

431.29457793070.5501688831.37483586930.54075461871.17404065660.51284143092.04755614270.795243132863277811781353110826654155436

441.18396525940.52506800221.09677917660.52129208821.36971409940.48842499151.94187582560.702221573757874261421304412625385147385

451.27819086830.51375492911.2512551170.50342812291.48929231440.4627001852.11360634080.727756903762472661621259713724048160399

461.15323951740.49445199831.28987410210.48276666861.40232633980.4425166982.10039630120.702221573756369931671208012922999159385

471.30686822750.4762096681.2280837260.45355289091.5110338080.41557972792.19286657860.616495823163867351591134913921599166338

481.12865892380.45266433481.2358075230.4439215361.30448961840.39682005222.20607661820.605552110355164021601110812020624167332

491.1778201110.42501801251.51386421560.41294933671.5110338080.38512171092.29854689560.603728158257560111961033313920016174331

501.20649747020.41950288941.22035992890.39700364951.26100663120.36174426892.40422721270.60555211035895933158993411618801182332

511.24336836070.39312928791.34394068120.38641315561.42406783350.34523575432.45706737120.56907306756075560174966913117943186312

521.25156189190.39072525991.27442650810.36675080451.46755082070.32545632192.39101717310.53259402476115526165917713516915181292

531.17577172820.3767253321.38255966630.34732823821.40232633980.30981363852.35138705420.53077007265745328179869112916102178291

541.22288453270.36590720591.35938827530.33453971711.40232633980.29651831962.45706737120.51253055125975175176837112915411186281

551.19011040780.34264469951.21263613190.32406911561.36971409940.28491618182.49669749010.43227665715814846157810912614808189237

561.20444908740.3345841351.30532169610.30880281861.6306120230.26455952862.65521796570.45234013065884732169772715013750201248

571.1163686270.33104879961.28215030510.29157827931.43493858030.25401562882.19286657860.39397366215454682166729613213202166216

581.12046539260.31273576271.17401714680.28506412641.2718773780.24589605641.91545574640.39397366215474423152713311712780145216

591.1573362830.29604897991.28987410210.27591234111.52190455480.2339475862.08718626160.36114252365654187167690414012159158198

601.14299760340.28678640141.2512551170.26456252871.22839439070.22207607851.77014531040.29912815095584056162662011311542134164

611.04057846330.2777359431.09677917660.2512544741.29361887160.21297523071.84940554820.35931857155083928142628711911069140197

621.06925582250.2659279231.03498880050.24310179181.22839439070.20462476991.86261558780.33013533725223761134608311310635141181

631.04877199450.2529178891.00409361240.23271111851.25013588430.1942155551.87582562750.27359282095123577130582311510094142150

641.02828816650.24563509821.27442650810.22579732431.21752364390.1827481021.54557463670.2298179696502347416556501129498117126

651.06515905690.24004926841.15084575580.21356830111.30448961840.171030521.24174372520.279064677352033951495344120888994153

661.05901390850.23177658380.91913184520.20621490151.28274812480.16208359771.09643328930.22252216151732781195160118842483122

671.06925582250.22237259181.18946474090.19506490971.33710185890.15623442711.05680317040.229817969652231451544881123812080126

680.92382064360.20745347681.18946474090.18571330371.16316990980.14567128661.08322324970.17874730974512934154464710775718298

690.94430447160.20031209941.243531320.18055793121.25013588430.14070718781.10964332890.17874730974612833161451811573138498

700.96273991680.19041316051.11995056770.17516277391.29361887160.13314559550.84544253630.17692335754702693145438311969206497

710.88285298760.18574651791.06588398860.16573123961.18491140340.12381385940.6076618230.14591617124312627138414710964354680

720.96683668240.17789807350.98864601840.15849773241.02185020110.12177434590.54161162480.1331485062472251612839669463294173

730.89514328440.17259507050.93457943930.14842677211.02185020110.11221172150.50198150590.1386203626437244112137149458323876

740.9299657920.1628375451.16629334980.1430715791.10881617570.10380353870.19815059450.09484551134542303151358010253951552

750.74561133980.15894867610.91913184520.13567821521.0870746820.10149465550.356671070.0911976073642248119339510052752750

760.7333210430.14721136290.88823665710.1321613720.9674964670.09437559910.17173051520.1112610805358208211533078949051361

770.80911120670.14021139891.07360778560.12300958661.09794542890.08596741630.14531043590.07295808563951983139307810144681140

780.7333210430.13307002160.98864601840.11621568480.89140123930.07948330270.17173051520.0839017984358188212829088241311346

790.70669206660.13038316670.91140804820.11313844690.93488422650.07475009230.06605019820.072958085634518441182831863885540

800.68006309020.11949433390.90368425120.10866246460.80443526470.06763103590.06605019820.05289461233216901172719743515529

810.69030500420.11525193150.88823665710.10046981830.78269377110.0640522670.05284015850.041950899233716301152514723329423

820.61451484050.1042923920.80327488990.093516060.6631155560.05712561750.07926023780.027359282130014751042340612969615

830.66572441060.10103988350.71831312270.08672215820.61963256880.05106479920.05284015850.02188742573251429932170572654412

840.53872467690.08951469040.72603691970.07753040870.53266659420.04515790640.02642007930.012767665263126694194049234727

850.54077305970.08753490260.59473237040.07569205880.47831286010.03936645790.02642007930.0164155693264123877189444204629

860.56535365330.08223189960.49432300920.06626052460.46744211330.033863619700.02006347352761163641658431760011

870.55716012210.07827232410.47887541520.06130497270.31525165780.029380538200.0145916171272110762153429152708

880.43630553680.07268649420.47115161810.05151376120.39134688550.024089347600.0091197607213102861128936125205

890.424015240.06922186560.4016374450.04539924960.20654418960.0199910800.009119760720797952113619103905

900.40557979480.06483804980.4248088360.0386453120.26089792370.015469517200.0109437128198917559672480406

910.35437022470.06087847430.31667567780.03141180480.22828568320.012063914500.0091197607173861417862162705

920.36051537310.05776737920.25488530160.02737542780.17393194910.008850718800.0109437128176817336851646006

930.32774124830.05126236220.27805669270.02018188470.08696597460.006907408800.003647904316072536505835902

940.31954771710.0455351190.23171391060.01550608170.04348298730.003944342100.007295808615664430388420504

950.29087035790.04277755740.11585695530.01063045810.04348298730.002578252900.001823952114260515266413401

960.29291874070.03931292880.08496176720.00763314850.02174149360.0013083671001435561119126800

970.24375755340.03690890080.03861898510.004475982408.08E-0400.0018239521119522511204201

980.2273704910.03507052640.04634278210.00235788360.01087074682.69E-0400.001823952111149665911401

990.23146725660.029272576507.19E-0405.77E-05001134140180300

10000000000203140750100014

total10010010010010010010010048819141429312947250224491995197318757054826

Sheet6

binding site

non-binding site

Geometric Potential

Sheet5

binding residue

non-binding residue

Geometric Potential

Frequency (%)

Sheet3

meanstd

00.90375772951.9026478516

10.69763754562.4892976058

20.50737276043.3930553353

30.79276993824.2968130648

40.85619153324.1699698747

50.88790233074.154114476

61.28428729985.0103060092

70.8720469325.692088156

81.07816711595.2639923894

91.15744410975.8664975424

101.10987791345.2322815919

110.90375772955.089583003

120.77691453945.4066909783

130.8720469325.6603773585

140.98303472335.2639923894

150.9354685275.0578722055

160.98303472334.3602346599

170.85619153324.0272712859

180.77691453943.1076581576

191.03060091963.1393689551

201.26843190112.4575868083

211.04645631841.8709370541

220.91961312831.5538290788

231.07816711591.0781671159

241.03060091961.1574441097

251.2050103060.6500713493

260.99889012210.7134929443

270.9354685270.5866497542

280.98303472330.3805295703

290.95132392580.2378309814

300.96717932460.1744093864

311.17329950850.0634215951

320.91961312830.0951323926

331.33185349610.0951323926

341.25257650230.0158553988

350.98303472330.0634215951

361.18915490720.0475661963

371.1415887110.0951323926

381.45869668620.0158553988

391.45869668620.0158553988

401.17329950850.0317107975

411.26843190110.0158553988

421.09402251470

431.17329950850

441.45869668620

451.50626288250

461.39527509120

471.63310607260

481.33185349610

491.44284128750

501.33185349610

511.55382907880

521.66481687010

531.41113048990

541.52211828130

551.60139527510

561.63310607260

571.52211828130

581.44284128750

591.66481687010

601.26843190110

611.4745520850

621.25257650230

631.30014269860

641.41113048990

651.31599809740

661.36356429360

671.36356429360

681.2050103060

691.41113048990

701.33185349610

711.17329950850

721.07816711590

730.99889012210

741.1415887110

751.09402251470

761.06231171710

771.10987791340

780.88790233070

790.96717932460

800.80862533690

810.96717932460

820.71349294430

830.6025051530

840.58664975420

850.50737276040

860.52322815920

870.25368638020

880.36467417160

890.22197558270

900.2695417790

910.22197558270

920.20612018390

930.06342159510

940.04756619630

950.03171079750

9600

9700

980.01585539880

9900

10000

Sheet3

std

Variation of Geometric Potential

Rank 1Rank 1-3

>0.10.72325846970.9074990483

>0.20.6663494480.8698134754

>0.30.6166730110.8454510849

>0.40.57175485340.8083365055

>0.50.529120670.7611343738

>0.60.4503235630.6606395128

>0.70.36886181960.536733917

>0.80.2613247050.3818043396

>0.90.13323182340.1912828321

10.0702322040.1046821469

Rank 1

Rank 1-3

Sensitivity

Frequency

sensitivityspecificity

rank 1all ranksrank 1all ranks

1012.2363670911.72905187161000

2010.54531635951.40604218132000

306.97320919632.75508265253000.03800114

404.78814364434.00912027364000.1330039901

505.643169295110.2223066692500.019000570.2090062702

604.883146494412.9203876116600.07600228010.6080182405

705.130153904616.7395021851700.57001710054.446133384

806.004180125423.3897016911808.455253657628.1398441953

902.071062131910.20330609929090.404712141465.9889796694

1002.964088922714.78244347331000.47501425040.4370131104

rank 1all ranksrank 1all ranks

>1098.156944708398.1569447083>1099.828994869899.8289948698

>2096.427892836896.4278928368>2099.828994869899.8289948698

>3095.021850655595.0218506555>3099.828994869899.8289948698

>4092.26676800392.266768003>4099.828994869899.8289948698

>5088.257647729488.2576477294>5099.828994869899.8289948698

>6078.035341060278.0353410602>6099.619988599799.6199885997

>7065.114953448665.1149534486>7099.011970359199.0119703591

>8048.375451263548.3754512635>8094.565836975194.5658369751

>9024.985749572524.9857495725>9066.425992779866.4259927798

10014.782443473314.78244347331000.43701311040.4370131104

Sensitivity (%)

Distribution (%)

all ranks

Specificity (%)

Distribution (%)

all ranks

Sensitivity (%)

Frequency (%)

all ranks

Specificity (%)

Frequency (%)

Local Sequence-order Independent Alignment with Maximum-Weight Sub-Graph AlgorithmL E RV K D LL E RV K D LStructure AStructure B

Build an associated graph from the graph representations of two structures being compared. Each of the nodes is assigned with a weight from the similarity matrixThe maximum-weight clique corresponds to the optimum alignment of the two structuresXie and Bourne 2008 PNAS, 105(14) 5441

Similarity Matrix of AlignmentChemical SimilarityAmino acid grouping: (LVIMC), (AGSTP), (FYW), and (EDNQKRH)Amino acid chemical similarity matrix

Evolutionary CorrelationAmino acid substitution matrix such as BLOSUM45Similarity score between two sequence profiles

fa, fb are the 20 amino acid target frequencies of profile a and b, respectivelySa, Sb are the PSSM of profile a and b, respectively Computational MethodologyXie and Bourne 2008 PNAS, 105(14) 5441

Nothing in Biology {Including Drug Discovery} Makes Sense Except in the Light of EvolutionTheodosius Dobzhansky (1900-1975)

Lead Discovery from Fragment AssemblyPrivileged molecular moieties in medicinal chemistry

Structural genomics and high throughput screening generate a large number of protein-fragment complexes

Similar sub-site detection enhances the application of fragment assembly strategies in drug discovery1HQC: Holliday junction migration motor protein from Thermus thermophilus1ZEF: Rio1 atypical serine protein kinase from A. fulgidusComputational Methodology

Lead Optimization from Conformational ConstraintsSame ligand can bind to different proteins, but with different conformations

By recognizing the conformational changes in the binding site, it is possible to improve the binding specificity with conformational constraints placed on the ligand1ECJ: amido-phosphoribosyltransferase from E. Coli1H3D: ATP-phosphoribosyltransferase from E. Coli Computational Methodology

ScoringThe Point is this Approach Can Now be Applied on a Proteome-wide ScaleScores for binding site matching by SOIPPA follow an extreme value distribution (EVD). Benchmark studies show that the EVD model performs at least two-orders faster and is more accurate than the non-parametric statistical method in the previous SOIPPA version Xie, Xie and Bourne 2009 Bioinformatics 25(12) 305-312Blosum45 and b) McLachlan substitution matrices. Computational Methodology

AgendaComputational Methodology

Side Effects - The Tamoxifen Story

Repositioning an Existing Drug - The TB Story

Salvaging $800M The Torcetrapib Story

The Future? - The TB Drugome

Found..Evolutionary linkage between: NAD-binding Rossmann foldS-adenosylmethionine (SAM)-binding domain of SAM-dependent methyltransferasesCatechol-O-methyl transferase (COMT) is SAM-dependent methyltransferaseEntacapone and tolcapone are used as COMT inhibitors in Parkinsons disease treatmentHypothesis:Further investigation of NAD-binding proteins may uncover a potential new drug target for entacapone and tolcaponeRepositioning an Existing Drug - The TB Story

Functional Site Similarity between COMT and InhAEntacapone and tolcapone docked onto 215 NAD-binding proteins from different speciesM.tuberculosis Enoyl-acyl carrier protein reductase ENR (InhA) discovered as potential new drug targetInhA is the primary target of many existing anti-TB drugs but all are very toxicInhA catalyses the final, rate-determining step in the fatty acid elongation cycleAlignment of the COMT and InhA binding sites revealed similarities ...

Kinnings et al. 2009 PLoS Comp Biol 5(7) e1000423Repositioning an Existing Drug - The TB Story

Binding Site Similarity between COMT and InhARepositioning an Existing Drug - The TB Story

Summary of the TB StoryEntacapone and tolcapone shown to have potential for repositioningDirect mechanism of action avoids M. tuberculosis resistance mechanismsPossess excellent safety profiles with few side effects already on the marketIn vivo supportAssay of direct binding of entacapone and tolcapone to InhA reveals a possible lead with no chemical relationship to existing drugsKinnings et al. 2009 PLoS Comp Biol 5(7) e1000423Repositioning an Existing Drug - The TB Story

Summary from the TB Alliance Medicinal ChemistryThe minimal inhibitory concentration (MIC) of 260 uM is higher than usually consideredMIC is 65x the estimated plasma concentrationHave other InhA inhibitors in the pipelineRepositioning an Existing Drug - The TB Story

AgendaComputational Methodology

Side Effects - The Tamoxifen Story

Repositioning an Existing Drug - The TB Story

Salvaging $800M The Torcetrapib Story

The Future? - The TB Drugome

Selective Estrogen Receptor Modulators (SERM)One of the largest classes of drugsBreast cancer, osteoporosis, birth control etc.Amine and benzine moietySide Effects - The Tamoxifen StoryPLoS Comp. Biol., 2007 3(11) e217

Adverse Effects of SERMscardiac abnormalities thromboembolic disordersocular toxicities loss of calcium homeostatis ?????PLoS Comp. Biol., 3(11) e217Side Effects - The Tamoxifen Story

Structure and Function of SERCASacroplasmic Reticulum (SR) Ca2+ ion channel ATPase Regulating cytosolic calcium levels in cardiac and skeletal muscle

Cytosolic and transmembrane domains

Predicted SERM binding site locates in the TM, inhibiting Ca2+ uptakePLoS Comp. Biol., 3(11) e217Side Effects - The Tamoxifen Story

Binding Poses of SERMs in SERCA from Docking StudiesSalt bridge interaction between amine group and GLU

Aromatic interactions for both N-, and C-moiety6 SERMS A-F (red) PLoS Comp. Biol., 3(11) e217Side Effects - The Tamoxifen Story

The ChallengeDesign modified SERMs that bind as strongly to estrogen receptors but do not have strong binding to SERCA, yet maintain other characteristics of the activity profilePLoS Comp. Biol., 3(11) e217Side Effects - The Tamoxifen Story

AgendaComputational Methodology

Side Effects - The Tamoxifen Story

Repositioning an Existing Drug - The TB Story

Salvaging $800M The Torcetrapib Story

The Future? - The TB Drugome

The Torcetrapib StoryPLoS Comp Biol 2009 5(5) e1000387

Cholesteryl Ester Transfer Protein (CETP) collects triglycerides from very low density or low density lipoproteins (VLDL or LDL) and exchanges them for cholesteryl esters from high density lipoproteins (and vice versa)A long tunnel with two major binding sites. Docking studies suggest that it possible that torcetrapib binds to both of them.The torcetrapib binding site is unknown. Docking studies show that both sites can bind to torcetrapib with the docking score around -8.0.HDLLDLCETPCETP inhibitorXBad CholesterolGood CholesterolPLoS Comp Biol 2009 5(5) e1000387The Torcetrapib Story

Docking Scores eHits/AutodockPLoS Comp Biol 2009 5(5) e1000387The Torcetrapib Story

Off-targetPDB IdsTorcetrapibAnacetrapibJTT705Complex ligandCETP2OBD-11.675 / -5.72-11.375 / -8.15-7.563 / -6.65-8.324 (PCW)Retinoid X receptor1YOW1ZDT-11.420 / -6.600 -6.74-8.696 / -7.68 -7.35-6.276 / -7.28 -6.95-9.113 (POE)PPAR delta1Y0S-10.203 / -8.22-10.595 / -7.91-7.581 / -8.36-10.691(331)PPAR alpha2P54-11.036 / -6.67-0.835 / -7.27 -9.599 / -7.78-11.404(735)PPAR gamma1ZEO-9.515 / -7.31 > 0.0 / -8.25-7.204 / -8.11-8.075 (C01)Vitamin D receptor1IE8>0.0/ -4.73>0.0 / -6.25-6.628 / -9.70-8.354 (KH1) -7.35Glucocorticoid Receptor1NHZ1P93 /-4.43 /-5.63 /-7.08 /-0.58 /-7.09 /-9.42Fatty acid binding protein2F732PY12NNQ>0.0/ -4.33>0.0/-6.13 /-6.40>0.0/ -7.81>0.0/ -6.98 /-7.64-7.191 / -8.49 /-6.33 /6.35???T-Cell CD1B1GZP-8.815 / -7.02-13.515 / -7.15-7.590 / -8.02 -6.519 (GM2)IL-10 receptor1LQS / -4.59 / -6.77 / -5.95???GM-2 activator2AG9-9.345 / -6.26-9.674 / -6.98-8.617 / -6.17??? (MYR) -4.16(3CA2+) CARDIAC TROPONIN C1DTL /-5.83 /-6.71 /-5.79cytochrome bc1 complex1PP9 (PEG) /-6.97 /-9.07 /-6.641PP9 (HEM) /-7.21 /8.79 /-8.94human cytoglobin1V5H /-4.89 /-7.00 /-4.94

RASPPARRXRVDR+High blood pressureFABPFA+Anti-inflammatory function?Torcetrapib Anacetrapib JTT705JNK/IKK pathwayJNK/NF-KB pathway?Immune response to infection JTT705PPARPPAR?PLoS Comp Biol 2009 5(5) e1000387The Torcetrapib Story

AgendaComputational Methodology

Side Effects - The Tamoxifen Story

Repositioning an Existing Drug - The TB Story

Salvaging $800M The Torcetrapib Story

The Future? - The TB Drugome

1. StructuralDetermination& Modeling2. Binding site Similarity 3. Protein-ligandDockingGenomeStructuralProteomeProtein-drugInteractomeMetabolome4.1 Network Reconstruction Drugome4.2 Network Integration Existing DrugsTarget identificationDrug resistance mechanismDrug repurposingSide effect predictionNew therapeutics

Bioinformatics 2009 25(12) 305-312The Future

1. StructuralDetermination& Modeling2. Binding site Similarity 3. Protein-ligandDockingTB GenomeTB StructuralProteomeTB Protein-drugInteractomeTB Metabolome4.1 Network Reconstruction Drugome/TB4.2 Network Integration Existing DrugsTarget identificationDrug resistance mechanismDrug repurposingSide effect predictionNew therapeutics for MDR and XDR-TBThe TB DruggomeBioinformatics 2009 25(12) 305-312

Predicted protein-ligand interaction network of M.tuberculosis. Proteins that are predicted to have similar binding sites are connected. Squares represent the top 18 most connected proteins.

The TB DruggomeBioinformatics 2009 25(12) 305-312

The TB DruggomeBioinformatics 2009 25(12) 305-312

Some LimitationsStructural coverage of the given proteomeFalse hits / poor docking scoresLiterature searchingIts a hypothesis need experimental validationMoney Limitations

SummaryWe have established a protocol to look for off-targets for existing therapeutics and NCEs

Understanding these in the context of pathways would seem to be the next step towards a new understanding cheminfomatics meets systems biology

Lots of other opportunities to examine existing drugs DrugX and the Recovery Act

Bioinformatics Final Examples..Donepezil for treating Alzheimers shows positive effects against other neurological disordersOrlistat used to treat obesity has proven effective against certain cancer typesRitonavir used to treat AIDS effective against TBNelfinavir used to treat AIDS effective against different types of cancers

Lots of Opportunities

Acknowledgements

Sarah Kinnings

Lei XieLi XieJian Wanghttp://funsite.sdsc.edu

***Absorption, distribution, metabolism and excretion**P distance to environmental boundary; Pi Di and alphai D distance to central atom alpha direction to central atom *******Superimposition of the binding sites of COMT and ENRCOMT is show in green, its SAM co-factor is shown in yellow, and its BIE substrate is shown in purple. ENR is shown in blue, its NAD co-factor is shown in orange, and its 641 substrate is shown in red. Protein sequences were aligned according to the NAD and SAM co-factors.

Similarities in electrostatic potential were also observed in the substrate binding pockets of COMT and ENR. ****