Animal Models in Clinical and Translational Science

Keith C. Cheng, MD, PhDKeith C. Cheng, MD, PhDJake Gittlen Cancer Research Jake Gittlen Cancer Research

FoundationFoundationPenn State College of MedicinePenn State College of Medicine

kcheng76@gmail.comkcheng76@gmail.com

Navigating the Translational Researcher Through a Complex of

Animal and Biological ResourcesNCRR

March 6-7, 2006

TopicsTopics Audiences and functionalitiesAudiences and functionalities Animal Models: Model SystemsAnimal Models: Model Systems Biological Processes/DiseasesBiological Processes/Diseases Linking Concepts between web Linking Concepts between web sitessites

Systems MorphogeneticsSystems Morphogenetics Answering and Raising Answering and Raising QuestionsQuestions

Examples of Model System, Examples of Model System, Process, DiseaseProcess, Disease

Quality of InformationQuality of Information Accessibility of informationAccessibility of information

AudienceAudience Clinician ScientistsClinician Scientists Translational ResearchersTranslational Researchers Basic ScientistsBasic Scientists

Audience: Who’s at the Audience: Who’s at the bedside?bedside?

Clinician ScientistsClinician Scientists Translational ResearchersTranslational Researchers Basic ScientistsBasic Scientists Clinicians*Clinicians* PublicPublic

Patients*Patients* General PublicGeneral Public StudentsStudents

PhD traineesPhD trainees MD traineesMD trainees College StudentsCollege Students HS StudentsHS Students Middle School StudentsMiddle School Students

LegislatorsLegislators

Audience Audience FunctionalitiesFunctionalities

One-stop shoppingOne-stop shopping Engender thinking processes Engender thinking processes (genetics/reverse genetics)(genetics/reverse genetics)

FeedbackFeedback For new researchers:For new researchers:

““Nice model but how does that help the Nice model but how does that help the patient?”patient?”

““Nice model, but in what ways is the model Nice model, but in what ways is the model different from the range of human disease?different from the range of human disease?

““What are the limitations of the model?”What are the limitations of the model?” ““What would another ideal model do that What would another ideal model do that current ones don’t?”current ones don’t?”

Animal Models: The Animal Models: The obviousobvious

VertebratesVertebrates PrimatesPrimates MiceMice RatsRats ZebrafishZebrafish

Animal Models: OthersAnimal Models: Others InvertebratesInvertebrates

FliesFlies WormsWorms

Single Cell OrganismsSingle Cell Organisms PhagePhage BacteriaBacteria YeastYeast

Other modelsOther models XenopusXenopus DogsDogs TetrahymenaTetrahymena Other fish: Other fish: Fugu/Medaca/XiphophorusFugu/Medaca/Xiphophorus

HydraHydra

Animal Model: Man!Animal Model: Man! AnatomyAnatomy HistologyHistology PhysiologyPhysiology GeneticsGenetics Gene expressionGene expression DiseaseDisease Correlations with animal modelsCorrelations with animal models EnvironmentEnvironment Psychosocial issuesPsychosocial issues

Biological Biological Process/DiseaseProcess/Disease

AgingAging Normal VariationNormal Variation NeurophysiologyNeurophysiology CancerCancer PigmentationPigmentation MetabolismMetabolism Infectious DiseaseInfectious Disease

Linking ConceptsLinking Concepts Biochemical and Signal Biochemical and Signal Transduction PathwaysTransduction Pathways

GenesGenes Gene ExpressionGene Expression Anatomy Anatomy HistologyHistology Subcellular localizationSubcellular localization Life SpanLife Span PhysiologyPhysiology Disease Disease

Linking Concept: Linking Concept: ComparisonsComparisons

Wild type vs. mutant or morphantWild type vs. mutant or morphant Older vs. YoungerOlder vs. Younger Treated vs. untreatedTreated vs. untreated One organism vs. anotherOne organism vs. another MultilevelMultilevel

OrganOrgan TissueTissue CellCell SubcellularSubcellular MolecularMolecular

An example of gene comparisons: human An example of gene comparisons: human polymorphism rs1426654 affects a polymorphism rs1426654 affects a

conserved amino acidconserved amino acid

conserved C F P L DVA ATFMA PE conserved C F P L DVA ATFMA PE (NCKX)(NCKX)

conserved CD YFLPSLE I LGLSQDVAGATFMA GSSAPE VT FLG conserved CD YFLPSLE I LGLSQDVAGATFMA GSSAPE VT FLG (NCKX5)(NCKX5)

zf5 CDEYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLGzf5 CDEYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLG medaka5 CDDYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLGmedaka5 CDDYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLG fugu5 CDDYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLGfugu5 CDDYFLPSLEVISERLGLSQDVAGATFMAAGSSAPELVTAFLGXentrop5 CESYFIPSLEVISERLGLSQDVAGATFMAIGSSAPEFVTVFLGXentrop5 CESYFIPSLEVISERLGLSQDVAGATFMAIGSSAPEFVTVFLG chk5 CDDYFLPSLEIITECLGLSQDVAGATFMAAGSSAPELVTAFLGchk5 CDDYFLPSLEIITECLGLSQDVAGATFMAAGSSAPELVTAFLG dog5 CDEYFLPSLEIISETLGLSQDVAGATFMAAGSSAPELVTAFLGdog5 CDEYFLPSLEIISETLGLSQDVAGATFMAAGSSAPELVTAFLG cow5 CDEYFLPSLEIISESLGLSQDVAGATFMAAGSSAPELVTAFLGcow5 CDEYFLPSLEIISESLGLSQDVAGATFMAAGSSAPELVTAFLG mu5 CDKYFLPSLEIISDSLGLSQDVAGATFMAAGSSAPELVTAFLGmu5 CDKYFLPSLEIISDSLGLSQDVAGATFMAAGSSAPELVTAFLG rat5 CDKYFLPSLEIISDSLGLSQDVAGATFMAAGSSAPELVTAFLGrat5 CDKYFLPSLEIISDSLGLSQDVAGATFMAAGSSAPELVTAFLG hunckx5 CDEYFLPSLEIISESLGLSQDVAGATFMAAGSSAPELVTAFLG hunckx5 CDEYFLPSLEIISESLGLSQDVAGATFMAAGSSAPELVTAFLG

(YRI/HCB/JPT)(YRI/HCB/JPT) hunckx5 CDEYFLPSLEIISESLGLSQDVAGhunckx5 CDEYFLPSLEIISESLGLSQDVAGTTTFMAAGSSAPELVTAFLG (CEU) TFMAAGSSAPELVTAFLG (CEU)

Victor Canfields*

The zebrafish The zebrafish goldengolden phenotypephenotype

48 hpf

72 hpf

wild-type golb1/golb1

Becky Lamason

Diminished melanosome Diminished melanosome morphogenesis in morphogenesis in goldengolden

skinskin

Greg Ning

Wei Zhao, Adam Sidor, Joe Gershenson, Victor Canfield, Jason Mest

Comparison within a genome: Regional homozygosity in Europeans (CEU)

Comparison between genomes: Diminished regional heterozygosity near SLC24A5

Comparison between Comparison between populations:populations: SLC24A5 SLC24A5

rs1426654 allele rs1426654 allele frequenciesfrequenciesPopulation Thr (A) allele

frequencySample

sizeRef

European American, Utah (CEPH) 1.0 59 1European American, Baltimore 0.987 39 2European American, Chicago 0.987 39 2 European Total 0.992 127Japanese, Tokyo 0.011 44 1Cantonese, China 0.012 40 2Han Chinese, Beijing 0.011 45 1 East Asian Total 0.012 129Zapotec, Mexico 0.069 29 2 Indigenous American Total 0.069 29Yoruba, Ibadan, Nigeria 0.025 60 1Ghana, West Africa 0.000 33 2Cameroon, West Africa 0.000 20 2Senegal, West Africa 0.065 46 2Botswana, West Africa 0.048 21 2 African Total 0.030 180

1 www.HapMap.org2 M. W. Smith et al, 2004

Systems MorphogeneticsSystems Morphogenetics WhereWhere When When Why (function)Why (function) PhysiologyPhysiology DiseaseDisease Response to treatmentsResponse to treatments

TherapeuticTherapeutic Side-effectsSide-effects Rare adverse responsesRare adverse responses

Answering QuestionsAnswering Questions

Function*Function* MutantsMutants Knock-downsKnock-downs DiseaseDisease

PathwayPathway Tissue specificityTissue specificity Subcellular localizationSubcellular localization Age specificityAge specificity Downstream effectsDownstream effects

Raising QuestionsRaising Questions

What we don’t knowWhat we don’t know Important to direct questionsImportant to direct questions PrioritizationPrioritization Identification of information with Identification of information with degrees of uncertaintydegrees of uncertainty

Identify information of conflicting Identify information of conflicting opinionopinion

Highlighting of important unknowns, Highlighting of important unknowns, clinical problems and clinical goalsclinical problems and clinical goals

Animal Model Example: Animal Model Example: ZebrafishZebrafish

DevelopmentDevelopment Disease (cancer, infectious Disease (cancer, infectious disease, etc)disease, etc)

Drug developmentDrug development ToxicologyToxicology Bridge organism between Bridge organism between invertebrates and mammalsinvertebrates and mammals

Disease Example: CancerDisease Example: Cancer

Models span yeast to humansModels span yeast to humans Many diseasesMany diseases Many different pathophysiologiesMany different pathophysiologies Complicated geneticsComplicated genetics Primitive treatmentsPrimitive treatments Genomic instability creates Genomic instability creates moving targetmoving target

Cancer genome anatomy projectCancer genome anatomy project

Process Example: Process Example: PigmentationPigmentation

Different genes important in different Different genes important in different settingssettings

Most common variations affecting colorMost common variations affecting color Skin vs. hair vs. eyeSkin vs. hair vs. eye Two to three most importantTwo to three most important Many genes modulateMany genes modulate Other potential effects: Other potential effects:

Parkinson’s DiseaseParkinson’s Disease Age-related macular degenerationAge-related macular degeneration Frostbite susceptibilityFrostbite susceptibility

““Solvable” problemSolvable” problem

PigmentationPigmentation Variable anatomy/histologyVariable anatomy/histology

Distribution of pigmented cellsDistribution of pigmented cells Variable pigmentVariable pigment

Transfer of pigment between cellsTransfer of pigment between cells Variable biochemistry (eumelanin vs Variable biochemistry (eumelanin vs pheomelanin)pheomelanin)

Variations in melanosome morphologyVariations in melanosome morphology Proteomics coming onlineProteomics coming online Extensive geneticsExtensive genetics Extensive evolutionary conservation of Extensive evolutionary conservation of functionfunction

Diseases: cancers, normal and abnormal Diseases: cancers, normal and abnormal patterns, normal and abnormal pigmentationpatterns, normal and abnormal pigmentation

Quality of Information: Quality of Information: We must reach for the We must reach for the ideal, but much is…ideal, but much is…

InaccurateInaccurate IncompleteIncomplete MisleadingMisleading UncertainUncertain LimitedLimited“No similarity-to-human data found for SLC24A5 in HomoloGene for: Pan troglodytes, Sus scrofa, Bos taurus, Danio rerio . . .”

– source: <http://genome-www.stanford.edu/cgi-bin/genecards/carddisp?SLC24A5>

http://www.dsi.univ-paris5.fr/genatlas/fiche.php?symbol=SLC24A5 accessed – 3/2/2006

•Website updated on 1/20/2006

•Incorrectly cites expression and subcellular localization data

•States nervous and cone photoreceptor expression only

•States plasma membrane localization

•We have shown ubiquitous expression in adults (higher in pigmented tissues)

•We have shown subcellular localization to an internal organelle

•Cites SNP data from paper, but ignores expression data

Real Tissue distribution

Real subcellular localization

Metabolic Disease:Metabolic Disease:GA-1 (Glutaric Acidemia type 1) GA-1 (Glutaric Acidemia type 1)

Glutaryl CoA Dehydrogenase Glutaryl CoA Dehydrogenase (GCDH) deficiency(GCDH) deficiency

Lysine degradation pathwayLysine degradation pathway Causes accumulation of Glutaryl CoACauses accumulation of Glutaryl CoA ~ 90% of untreated suffer acute striatal ~ 90% of untreated suffer acute striatal necrosis associated with cerebral palsy for necrosis associated with cerebral palsy for life; ~30% still have this outcome after life; ~30% still have this outcome after treatmenttreatment

After age of 3, this no longer occursAfter age of 3, this no longer occurs New diet-induced mouse model now allows New diet-induced mouse model now allows mechanistic studies and drug developmentmechanistic studies and drug development

Key information: where is gene expressed, Key information: where is gene expressed, where do metabolites accumulate, when, why where do metabolites accumulate, when, why striatum and not other parts of brain, why striatum and not other parts of brain, why age-dependent resistance?age-dependent resistance?

Accessibility of Accessibility of informationinformation

Information accessibility despite Information accessibility despite variations invariations in PresentationPresentation PurposePurpose OrganizationOrganization Names and file formatsNames and file formats Relationships (ontologies) Relationships (ontologies) ResolutionResolution QualityQuality How up-to-date?How up-to-date?

Solution: Semantic WebSolution: Semantic Web CategorizationCategorization Computation-dependent extraction and Computation-dependent extraction and organization of informationorganization of information

ConclusionsConclusions Plan now for a wide audiencePlan now for a wide audience Clinician and patient involvementClinician and patient involvement User feedback will be critical to discuss and User feedback will be critical to discuss and respond torespond to

Integrate simpler models and humansIntegrate simpler models and humans The human model will deal with critical issues The human model will deal with critical issues that are not possible to study in other model that are not possible to study in other model organisms organisms

Functional (mutant, knock-down), developmental, Functional (mutant, knock-down), developmental, and physiological data require greater and physiological data require greater prominenceprominence

Highlight what we do NOT know for future Highlight what we do NOT know for future investigationinvestigation

It will be important to monitor limitations of It will be important to monitor limitations of models and datamodels and data