ER ER andand PRPR

progesteron

estradiol

-- BeforeBefore menopausemenopauseestrogensestrogensareare producedproduced by by enzymeenzymearomatasearomatasemostlymostly inin::** ovaryovary ((grangranulosaulosacellscells) )

andand inin otherother tissuestissues, , suchsuchas:as:** subcatenoussubcatenousadipocytesadipocytes((especiallyespeciallyestradiol)estradiol)* * skeletalskeletalmusclemuscle((especiallyespeciallyestradiol)estradiol)* stroma * stroma cellscells inin breastbreast ((especiallyespeciallyestroneestrone))* * osteocytesosteocytes((especiallyespeciallyestroneestrone))* placenta (* placenta (especiallyespeciallyestriol)estriol)

-- AfterAfter menopausemenopauseestrogensestrogensareare not not producedproduced by by ovaryovary but but aromatasearomataseisis stillstill activeactive ininotherother tissuestissues..

SitesSitesofof estrogen estrogen synthesessyntheses

androstenodiontestosterone

estroneestradiol

aromatasearomatase

EstradiolEstradiol

•• In In thethe femalefemalereproductivereproductive system system itit actsactsmostlymostly on on endometriumendometrium..

•• StimulatesStimulatesthethe epithelialepithelial cellscellsofof thethe basalbasallayerlayer ofof thethe endometriumendometrium to to proliferateproliferate , , forming a forming a thickthick mucosamucosaas as wellwell as as numerousnumerousendometrialendometrial glandsglands((proliferativeproliferative phasephaseofofendometrialendometrial cyclecycle, 6, 6--14 14 daysdays).).

•• Increases Increases thethe expressionexpressionofof progesterone progesterone receptors in receptors in endometriumendometrium..

uterusuterus

endometrium

vaginavagina

EstradiolEstradiol•• StimulatesStimulatesthethe developmentdevelopmentofof extensiveextensivemucosalmucosalfoldsfolds ofof thethe oviductoviduct as as wellwell as as thethe formationformation ofof ciliaciliaon on thesetheseepithelialepithelial cellscells..

•• PromotesPromotesgrowthgrowth ofof uterusuterus, , vaginavagina, , andand oviductsoviducts,,as as wellwell as as mammarymammary glandsglands..

•• TriggersTriggers estrus estrus inin femalesfemalesofof differentdifferent mammalsmammals((supportedsupported by progesteron).by progesteron).

oviduct

...All you needis love...

EstradiolEstradiol

•• Estradiol iEstradiol increasesncreases development of secondary sex development of secondary sex characteristicscharacteristics: : promotespromotes growthgrowth ofof widerwider pelvicpelvic bonesbonesas as wellwell as as thethe closureclosure ofof epiphysalepiphysal platesplates inin longlong bonesbones. . HoweverHowever many many ofof femalefemale secondarysecondarysex sex characteristicscharacteristics areareduedue to to thethe absenceabsenceofof androgensandrogens. .

•• AllowsAllows developmentdevelopment ofof softersofter skin skin andand promotespromotesdepositiondeposition ofof fatfat inin subcatenoussubcatenous zoneszones, , particularlyparticularly ininbreastbreast andand buttocksbuttocks, , leadingleading to to maturemature femalefemaleshapeshape..

•• Estradiol Estradiol enhancesenhances calciumcalcium depositiondeposition inin bonebone andandstimulatesstimulatesbonebonetissuetissueformationformation ..

•• Functions as a Functions as a neuroprotectantneuroprotectant andand cardioprotectantcardioprotectant..

ProgesteroneProgesterone

•• IsIs involvedinvolved inin maintenancemaintenanceofof pregnancypregnancy..

•• Progesteron Progesteron isis absentabsentinin thethe bloodblood duringduring thethe follicularfollicularphasephaseandand appearsappearsonlyonly afterafter ovulationovulation..

•• In In thethe femalefemale reproductivereproductive system system itit actsacts mostlymostly on on thethe glandsglands inin thethe endometriumendometrium to to promotepromote theirtheir secretorysecretoryactivityactivity ((secretorysecretoryphasephaseofofendometrialendometrial cyclecycle) ) andand prepareprepare endometriumendometrium for for pregnancypregnancy..

•• ProducedProduced by by thethe ovariesovaries andand placenta placenta (i(in n nonpregnantnonpregnant womenwomen–– inin corpuscorpus luteumluteum))

•• Increases mammary gland alveolarIncreases mammary gland alveolar--lobular formationlobular formation andand growthgrowth ofof breastbreast. .

•• II nhibitnhibit ss new follicular developmentnew follicular development..

•• ActivatesActivates thethe endometrialendometrial glandgland to to secrete fluidssecrete fluids..

•• Maintains the functions of the placenta. Maintains the functions of the placenta.

•• Keeps the Keeps the endometriumendometrium in a thickened conditionin a thickened condition..

•• Stops the uterus Stops the uterus contractionscontractions..

•• Prevents lactation until after the birth (with estrogen) Prevents lactation until after the birth (with estrogen)

•• StrengthensStrengthensthe mucus plug covering the cervix the mucus plug covering the cervix

to prevent infection. to prevent infection.

•• Strengthens the pelvic walls in preparation for Strengthens the pelvic walls in preparation for labourlabour. .

At the end of the pregnancyAt the end of the pregnancy, the levels of progesterone secreted by the placenta drop , the levels of progesterone secreted by the placenta drop off.off. It is this action that stimulates the beginning of the contractIt is this action that stimulates the beginning of the contractions that will lead to ions that will lead to birth.birth.

ProgesteroneProgesteroneduringduring thethe pregnancypregnancy::

•• From single gene by alternate transcriptionFrom single gene by alternate transcriptioninitiation (different promoters)initiation (different promoters) ..

•• IsoformsIsoforms ddifferiffer s s inin activitiesactivities..

•• PRPR--B B isis highlyhighly expressedexpressedinin endometriumendometrium duringduring proliferativeproliferative phasephase, but not , but not duringduring

secretorysecretoryphasephase..

•• In In humanshumansPRPR--A A actsactsas a as a repressorrepressorofof activityactivity ofof PRPR--B, GR, ER, AR, B, GR, ER, AR, andand MR.MR.

hPRhPR--BB

hPRhPR--AA

AF-3 DBD LBD

DBD LBD

11 933933

933933165165

AF-2

AF-2

AF-1

AF-1

H

H

Progesterone Receptor:Progesterone Receptor:A and B FormsA and B Forms

Progesterone Receptor:Progesterone Receptor:A and B FormsA and B Forms

LigandsLigands for Progesterone Receptorfor Progesterone Receptor

O

H3CO

O

H3CO

O

O

H

O

H3CO

O

OHCH3

NCH3H3C

O

OH

NCH3H3C

Et

Et

Et

OAc

CH3

OH

Progestins Antiprogestins

R5020(Promegestone)

ORG2058

Progesterone[natural]

RU486 (Mifepristone)

ZK98299(Onapristone)

Medroxyprogesterone Acetate (MPA)

[HRT]

O

OH

HH

Norethindrone[oral contraceptives]

Ru486 as a ligand Ru486 as a ligand ofof PR:PR:

-- RU486 bRU486 bindindss to to PRPR 3 times strong3 times stronger er than progesteronethan progesterone

-- RRU486 promotes a high affinity interaction of PR withU486 promotes a high affinity interaction of PR withDNADNA, , thusthus antagonistsantagonists--bound PR bound PR can effectivelycan effectivelycompete with binding of agonistcompete with binding of agonist--bound PR to bound PR to PREsPREs..

-- II n the presence of RU486n the presence of RU486PR recruitPR recruitss corepressorscorepressorstoto promoterspromoters..

-- PR PR ligandedliganded withwith RU486 RU486 hashas the abilitythe ability to to heterodimerizeheterodimerize with PR boundwith PR bound agonistagonist. . SuchSuchheterodimersheterodimers had a significantly reducedhad a significantly reducedability to bind to ability to bind to PREsPREs. . HeterodimerizationHeterodimerizationcould potentially sequester acould potentially sequester aportion of cellular PR bound to agonist in an inactive portion of cellular PR bound to agonist in an inactive form,form, without requiring direct binding of RU486 to PR.without requiring direct binding of RU486 to PR.

RU486 actionRU486 action::

1) initiates breakdown of 1) initiates breakdown of endometriumendometrium -- prevents embryo implantation prevents embryo implantation

2) promotes contraction of uterine wall and 2) promotes contraction of uterine wall and diladilatatationtion of cervix, increaseof cervix, increasess sensitivity sensitivity to PGFto PGF22αααααααα -- expulsion of embryo (early pregnancy termination)expulsion of embryo (early pregnancy termination)

RU 486 RU 486 –– PR PR antagonistantagonist

Note: Mifepristone = RU486

pharmacologicalabortion

(chorionic gonadotropin)

"morning after" pill

Estrogen Receptors Estrogen Receptors αααααααα and and ββββββββ genesgenes

••

DDifferentifferent tissuetissue//cellcell distributionsdistributions

AF1 AF1 isis veryvery activeactive inin ERERαααααααα but not but not inin ERERββββββββ..

••

TwoTwo separateseparategenesgenes

••

SeveralSeveralsplicingsplicing isoformsisoforms

1 144 227 255NN--

504 530--CCEERRββ

1 180 263 302NN--

552 595--CCEERRαα

AFAF--11 DNADNA LigandLigand / / AFAF--22A/BA/B CC DD EE FF

1188%% 9797%% 3300%% 5959%% 1818%%

••

-- ERERαααααααα isis much much betterbetter characterizedcharacterized. Role . Role ofof ERERββββββββ isisnot not fullyfully recognizedrecognized.. One One ofof itsits splicingsplicing formsforms (503 aa)(503 aa)was was reportedreported to to actact as a dominant as a dominant negativenegativeregulator.regulator.

-- ERERββββββββ isis stronglystrongly expressedexpressedinin malemale reproductivereproductive systemsystem..

-- ERERαααααααα andand ERERββββββββ havehave thethe same DNA same DNA bindingbinding domaindomain ((identidenticalical PP--boxbox), ), thusthus theythey bind to bind to thethe same consensus same consensus sequencesequence

-- ER ER form form homodimershomodimers, , bothboth ERERαααααααα//ERERαααααααα, , , , , , , , ERERββββββββ////////ERERββββββββ,,,,,,,, oror ERERαααααααα/ER/ERββββββββ..

-- ThereThere areare no no drugsdrugs whichwhich actact as a as a selectiveselectiveligandsligands for one for one typetype ofof ER receptorER receptor. . SomeSomecompoundscompounds, , howeverhowever, bind , bind withwith much much higherhigher affinityaffinity (120x) to one (120x) to one ofof themthem. . SomeSome areareagonistsagonistsofof one receptor one receptor andand antagonistsantagonistsofof thethe secondsecondreceptor.receptor.

ERERαααααααα andand ERERββββββββ::

Estrogen Estrogen receptorsreceptors ((ERsERs):):* * ERERss areare localizedlocalized inin cellcell nucleusnucleusandand areare boundbound to to heatheat shockshockproteinsproteins. . AfterAfter ligand ligand bindingbinding theythey form form homodimerhomodimer andand bind to consensus bind to consensus sequencessequencesinin thethe promoterspromoters ofof targettargetgenesgenes..

* * ERERss afterafter bindingbinding to consensus to consensus sequencesequenceareareubiquitinatedubiquitinated .. PossiblyPossiblyER ER transducestransducessignalsignalonlyonly onceonceandand thenthen isis degradeddegradedby by proteasomesproteasomes..

EstrogenEstrogen

Cell membraneCell membrane

NucleusNucleus

+ or + or -- GeneGene

expressionexpression

* * ClassicalClassicalwayway ofof ER ER activationactivation isis bindingbinding ofof ligand (e.g. estradiol).ligand (e.g. estradiol).

* * ER ER maymay be be activatedactivated withoutwithout bindingbinding thethe ligandsligands e.g. as a e.g. as a resultresult ofofphosphorylationphosphorylation.. ER ER cancan be be phosphorylatedphosphorylated e.g. by e.g. by PKAPKA (protein (protein kinasekinase--AA)), PKC, PKC (protein (protein kinasekinase--CC), ), cyclinecycline--dependentdependent kinaseskinases, MAP , MAP kinaseskinases.. PhosphorylationPhosphorylation cancan taketake place place bothboth atat AFAF--1 1 andand AFAF--2.2.

* * PhosphorylationPhosphorylation maymay influence influence thethe ligand ligand bindingbinding, , dimerizationdimerization, , bindingbinding to DNA to DNA andand interactioninteraction withwith coco--factorsfactors.. PossiblyPossibly, , eveneven ififphosphorylationphosphorylation maymay occuroccur withoutwithout ligand ligand bindingbinding, , thethe bindingbinding ofofligand ligand increasesincreasesitit . .

ActivationActivation ofof ER:ER:

Estrogen Estrogen actionaction::

i) i) classicalclassicalactivationactivation ofof transcriptiontranscriptionthroughthrough nuclearnuclear ERER

ii) ii) activationactivation ofof membranemembraneER ER andandstimulationstimulation ofof kinasekinasepathwayspathways

iii) iii) activationactivation ofof nonnon--ER ER membranemembraneassociatedassociatedestrogen estrogen bindingbinding proteinsproteins((EBPsEBPs) ) andand activationactivation ofof kinasekinasepathwayspathways

ER ER maymay affectaffect genegenetranscriptiontranscription by:by:

* * DirectDirect bindingbinding to consensus to consensus sequencesequence..

* * Influence on Influence on NFNFκκκκκκκκBB ((formationformation ofof complexcomplex withwith cc--relrel subunitsubunit) ) –– itit inhibitsinhibits activityactivity ofofNFNFκκκκκκκκBB. . ThereforeTherefore, , estrogensestrogensmaymay havehave antiinflammatoryantiinflammatory activityactivity , , inhibitinginhibiting e.g. ILe.g. IL--6, 6, ILIL --11ββββββββ, , TNFTNFαααααααα, M, M--CSF .CSF .

* * Influence on Influence on Sp1Sp1 ((formationformation ofof complexcomplex withwith Sp1) Sp1) –– increasesincreasesactivityactivity ofof Sp1Sp1,, leadingleadinge.g. to e.g. to inductioninduction ofof RARRARαααααααα andand eNOSeNOSandand nNOSnNOS expressionexpression..

* Influence on * Influence on APAP--11 ((throughthrough interactioninteraction withwith p160 p160 coactivatorscoactivators) ) –– usuallyusually increasesincreasesactivityactivity ofof APAP--1, 1, but but itit maymay dependdependon on typetype ofof ER ER receptorsreceptorsandand typetype ofof ligand.ligand.

* * Influence on Influence on Nrf2Nrf2 ((whichwhich bind to bind to thethe antioxidantantioxidant responseresponseelement) element) –– inin most most casescasesititmaymay increaseincreasethethe activityactivity ofof promoterspromoters ofof genesgenesinvolvedinvolved inin protectionprotection againstagainst oxidativeoxidativestressstress..

Influence Influence ofof ER on ER on genegenetranscriptiontranscription ::

Influence Influence ofof ER on ER on genegenetranscriptiontranscription ::

Nrf2 - mode

ERERαααααααα andand ERERββββββββ inin thethe prostateprostate

MMajorityajority ofof epithelial cell nuclei epithelial cell nuclei ininthetheprostateprostateexpress express ERERββ (ER(ERαα arearepresentpresentonlyonly on on somesomestromalstromalcellscells))

control ERαααα−−−−////−−−−

ERββββ−−−−////−−−− ERββββ−−−−////−−−−, , , , ERαααα−−−−////−−−−

prostateprostate acinaeacinae

Nilsson et al. 2001

andand finallyfinally stopsstops��������

MenopauseMenopause

•• MenopauseMenopause((cessationcessationofof ovulationovulation) ) isis uniqueunique to to humanhuman. . FemalesFemalesofof otherother speciesspeciesshow a show a declinedecline inin but not but not completecompleteterminationtermination ofof reproductivereproductive functionsfunctions..

•• MenopauseMenopauseoccursoccurs usuallyusually betweenbetweenthethe agesagesofof 4848--52. 52. ThisThis hashas not not changedchangedconsiderablyconsiderablythroughoutthroughout historyhistory, , althoughalthough menarchemenarche occursoccurs atat yangeryanger agesagestodaytoday thanthan 100 100 yearsyears ago ago andand lifelife expectancyexpectancyhashas increasedincreased..

•• MenopauseMenopauseisis associatedassociatedwithwith decreasedecreaseinin estradiol estradiol productionproduction andand inin consequencyconsequencyananelevationelevation inin plasmaplasma gonadotropingonadotropin levelslevels. . ItIt stimulatesstimulates ovarianovarian stroma stroma cellscells to to continuecontinueproducingproducing androstenedioneandrostenedione. .

•• EstroneEstrone, , derivedderived almostalmost entirelyentirely fromfrom peripheralperipheral conversionconversion ofof adrenaladrenal andand ovarianovarianandrostenedionandrostenedion, , becomesbecomesthethe dominant estrogen.dominant estrogen.

•• BecauseBecausethethe ratioratio ofof estrogensestrogensto to androgensandrogensdecreasesdecreases, , somesomewomenwomen havehave hirsuitismhirsuitism duedueto androgen to androgen accessaccess(e.g. (e.g. muostachemuostache...)....).

0

10

20

30

40

50

60 EstroneEstronepgp

g /m

l/m

l

earlyearly latelatefollicularfollicular phasephase

postmenopausalpostmenopausal0

10

20

30

40

50

60

70 EstradiolEstradiol

pgpg /

ml

/ml

earlyearly latelatefollicularfollicular phasephase

postmenopausalpostmenopausal

0

200

400

600

800

1000

1200

1400

1600

1800 AndrostenedioneAndrostenedione

pgpg /

ml

/ml

earlyearly latelatefollicularfollicular phasephase

postmenopausalpostmenopausal

0

50

100

150

200

250

300 TestosteroneTestosterone

pgpg /

ml

/ml

earlyearly latelatefollicularfollicular phasephase

postmenopausalpostmenopausal

HormonalHormonal changeschangesduringduring menopausemenopause

Osteoporosis

OsteoporoOsteoporosissis

-- One One ofof thethe most most frequentfrequent chronicchronic diseasesdiseases. 80% . 80% ofof patientspatients areare womenwomen. .

-- DevelopsDevelopsbothboth inin malemale andand femalefemale, but , but inin malemale thisthis processprocessisis slowerslower (as (as theythey havehave moremoredensedensebonesbones, , smallersmaller decreasedecreaseinin sex sex hormoneshormonesandand areare moremore activeactive)). .

-- InIn USA USA ~~90% 90% fracturesfractures ((hiphip andand backbonebackbone)) inin oldold womenwomen andand 70% 70% inin oldold men men resultsresultsfromfrom osteoporosisosteoporosis..

BoneBonedensitydensity ((hiphip)) inin womenwomenolderolder thanthan 6565::>833 mg/cm>833 mg/cm22 -- normnorm -- 0%0%833833--648 mg/cm648 mg/cm22 -- osteopeniaosteopenia-- 40%40%<648 mg/cm<648 mg/cm22 -- osteoporoosteoporosissis -- 13%13% (i(in n olderolder thanthan > 80 > 80 yearsyears-- 27%)27%)<648 mg/cm<648 mg/cm22 andand fracturesfractures –– strongstrong osteoporosisosteoporosis-- 7%, 7%, (i(in n olderolder thanthan 80 80 -- 27%)27%)

-- SimilaritySimilarity inin bonebonedensitydensity isis higherhigher inin monozygoticmonozygoticthanthan inin dizygoticdizygotic twinstwins. . ThusThus therethere isisa a significantsignificant influence influence ofof geneticgeneticfactor(sfactor(s) controlling ) controlling bonebonemetabolismmetabolism..

EstrogenEstrogenss andand bonesbones

* * DevelopmentDevelopment ofof bonesbones duringduring maturitionmaturition dependsdependsmostlymostly on on estrogensestrogensbothboth inin malesmalesandand femalesfemales..

* In * In bothboth sexessexes, estrogen , estrogen deficiencydeficiency leadsleads to to osteoporosisosteoporosis: : inin thethe onlyonly knownknown casecase ofofcompletecompleteinsensivityinsensivity to to estrogensestrogens((point point mutationmutation ofof ER receptor ER receptor resultingresulting inin formationformation ofofstopstop--codoncodon) ) thethe illill manman hadhad increasedincreasedbonebonemetabolismmetabolism,, osteoporosisosteoporosis, , immatureimmature epiphysalepiphysalplatesplates andand continouscontinous growthgrowth ofof bonesbonesalsoalso duringduring adulthoodadulthood. . SimilarSimilar effectseffectsareare observedobservedinin men men withwith unfuctionalunfuctional aromatasearomatase..

* * Estradiol Estradiol increasesincreasesproliferaprolifera tiontion, , migramigrationtion, , maturitionmaturition andand secretorysecretoryactivityactivity ofof chondrocyteschondrocytes,,leadingleading to to maturitionmaturition ofof epiphysalepiphysalplatesplatesandandceasesceasesthethe growthgrowth ofof longlong bonesbones..

RiskRisk ofof osteoporosisosteoporosis

sexsex

nutritionnutrition

J. Z. Ilich, and J. E. Kerstetter (2000).

OsteoporoOsteoporosissis

-- HigherHigher riskrisk ofof osteoporosisosteoporosisisis noticednoticed inin womenwomen withwith familialfamilial historyhistory ofof osteoporosisosteoporosis. . FrequencyFrequencyofof osteoporosisosteoporosisisis increasedincreasedby:by:

** inactiveinactive lifelife stylestyle, , * * lowlow calciumcalcium diet (diet (especiallyespeciallyduringduring maturitionmaturition ),),** high high alcoholalcohol intakeintake,,* * slimnessslimness

-- RiskRisk isis increasedincreasedinin patientspatients treatedtreated withwith corticosteroidscorticosteroidsandand thyroidthyroid hormoneshormones..

NormalNormal trabeculartrabecular bonebone OsteoporoticOsteoporoticbonebone

OsteoporoOsteoporosissis

-- FracturesFractures areare usuallyusually a a consequencesconsequencesofof a a fallfall , but , but sometimessometimestheythey occuroccur spontaneouslyspontaneously((~~ 5%).5%).

-- TheyThey cancan concernconcernallall bonesbones, but , but usuallyusually theythey occuroccur inin::* * backbonesbackbones, , * * upperupper part part ofof femurfemur ((andand hiphip))* * wristwrist (as a (as a consequenceconsequenceofof fallfall ))

-- FracturesFractures inin backbonesbackbonesareare painfulpainful , , but (as but (as oftenoften theythey do not do not resultresult fromfrom strongstrong trauma) trauma) areare oftenoften ignoredignored. . TheyThey areare alsoalso commonlycommonly diagnoseddiagnosedas as osteoarthritisosteoarthritis. . UsuallyUsually theythey arearerecognizedrecognizedduringduring routineroutine XX--rayray ofof lungslungs. .

-- IfIf therethere areare severalseveral fracturesfractures, , theythey maymay leadlead to to kypkyphosishosis ((”” widowwidow humphump”” ), ), resultingresulting inindisturbeddisturbed functionsfunctions ofof lungslungs andand heartheart andand difficultiesdifficulties inin deeperdeeperbreathingbreathing..

Model Model ofof vertebraevertebrae

University of Arizona Extension Center

a a normalnormal bonebone; ;

a a boneboneaffectedaffectedby by osteopeniaosteopeniashowingshowingthethebeginningbeginning ofof a a lacklack ofofcalciumcalcium andand lowlow bonebonedensitydensity, ,

a a boneboneshowingshowinga a severeseverecasecaseofof osteoporosisosteoporosis

kyphosiskyphosis

EstrogenEstrogenss aandnd bonesbones

* * PerhapsPerhaps, , inin bonebone cellscells bothboth ERERαααααααα andand ERERββββββββ areare expressedexpressed. . TheThe levellevel ofof expressionexpressionisis, , howeverhowever, , relativelyrelatively lowlow..

* * EstrogEstrogensens actact bothboth on on osteoblastsosteoblastsandand osteoclastsosteoclasts.. In In osteoblastsosteoblaststheythey upregulateupregulate thetheexpressionexpressionofof alalkkalicalic phosphatasephosphatase, , osteoponosteopontintin , , osteoosteocalcincalcin, , osteoneosteonectinctin.. ItIt isis associatedassociatedwithwith increasedincreased cellcell differentiationdifferentiation , , productionproduction ofof extracellularextracellular matrixmatrix , , andand strongerstrongermineralizationmineralization ofof matrixmatrix ..

* * EstrogensEstrogensinhibitinhibit thethe activationactivation ofof osteoclastsosteoclastsandand increaseincreasetheirtheir apoptosisapoptosis. . ItIt increasesincreasesexpressionexpressionofof osteoprotegrinosteoprotegrin, , thethe decoydecoyreceptor for receptor for osteoclastsosteoclastsdifferentiationdifferentiation factorfactor..

* * PossiblyPossibly, , estrogensestrogensaffectaffect bonesbones mostlymostly throughthrough regulationregulation ofof cytokinecytokine andand growthgrowthfactorsfactors expressionsexpressions:: increasedincreased synthesissynthesisofof IGFIGF --II andand reducedreduced synthesissynthesisofof ILIL --66,, ILIL --11ββββββββ, , TNF, MTNF, M --CSF, CSF, TGFTGFββββββββ..

* * EstrogensEstrogensupregulateupregulate thethe expressionexpressionofof 11αααααααα--hydroxylase hydroxylase whichwhich producesproducesthethe activeactive form form ofof vitaminvitamin D.D.

ProgesteronProgesteronee andand bonesbones

* * ProgesteronProgesteronee protectsprotects againstagainstosteoporosisosteoporosis, but , but itsits efficacyefficacy isis lowerlower thanthan thatthat ofof estrogenestrogen..

* * ProgesteroneProgesteronereceptorsreceptorsareare presentpresentbothboth inin osteoblastsosteoblastsandand osteoclastsosteoclasts..

* * IncreasesIncreasesproliferationproliferation andand differentiationdifferentiation ofof osteoblastsosteoblastsinin vitrovitro ..

* * ProgesteroneProgesteroneincreasesincreasesexpressionexpressionofof 11αααααααα--hydroxylasehydroxylase

SomeSomenuclearnuclear receptorsreceptors((ERER, AR, , AR, PRPR) ) stimustimu--latelate expressionexpressionofof cyclincyclin D, D, whichwhich activatesactivatesCdk4. Cdk4. ItIt leadsleadsto to phosphorylationphosphorylation ofof pRBpRB, , andandincreasesincreasestranscriptiontranscription ofof genesgenesincreasingincreasingproliferationproliferation ..

OthersOthers receptorsreceptors(VDR, RAR) (VDR, RAR) increaseincreasep21 p21 expressionexpression, , thusthus blockblock CdkCdk activityactivity , , whichwhichkeepskeepscellscellsatat G1 G1 phasephase. .

EstrogensEstrogensareare necessarynecessaryfor for normalnormal breastbreast developmentdevelopment. .

Postnatal mammaryPostnatal mammarygland development involves two distinct gland development involves two distinct phasesphases::

At pubertyAt puberty, estrogen promotes , estrogen promotes ductalductal elongationelongation and and dichotomous branchingdichotomous branching. At adulthood, the virgin gland . At adulthood, the virgin gland becomesbecomesrelativelyrelatively qquiescentuiescentwith the exception of sidewith the exception of sidebranching and alveolar budding that occur as abranching and alveolar budding that occur as aresult of result of the cyclic rise of ovarian steroids. the cyclic rise of ovarian steroids.

At pregnancyAt pregnancy, exposure to progesterone and, exposure to progesterone andprolactinprolactin results results in extensive in extensive epithelial proliferationepithelial proliferation,, increased increased dichodicho--tomoustomoussideside branchingbranchingand and differentiationdifferentiationof milkof milk--filled filled alveolar lobules.alveolar lobules.

ER ER inin breastbreast

RRelativelyelativelysmallsmallproportion (under 10%) of luminal epithelial cellsproportion (under 10%) of luminal epithelial cellsexpress express ERERαα in the normal breast, in the normal breast, whereas whereas myoepithelialmyoepithelialand and stromalstromalcells do notcells do not. P. Proportionroportionof positive cells declined in of positive cells declined in luteallutealphase inphase innaturally cycling women. However, in women withnaturally cycling women. However, in women withbreastbreastcancer, this cancer, this luteallutealphase decrease was not phase decrease was not observedobserved.ExpressionExpressionofof ERERαα isis higherhigherinin womenwomenofof EuropeanEuropeanthanthanofof AsianAsianoror AfricanAfrican originorigin..

BREAST CANCERBREAST CANCER

•• One out One out ofof everyevery 12 12 womenwomenwill will developdevelopbreastbreast cancercancer..

•• BreastBreast cancercancer isis thethe secondsecondleadingleading causecauseofof deathdeath inin womenwomen ((commonestcommonestcausecauseofof cancercancerdeathdeath inin womenwomen, 6% , 6% ofof allall deathsdeaths))

•• WorldwideWorldwide , 600,000 , 600,000 casescasesofof breastbreast cancercancer areare diagnoseddiagnosedeacheachyearyear. In UK . In UK therethere areare 24 000 24 000 newnew casescasesandand 12 800 12 800 deathsdeathsannuallyannually..

•• AboutAbout 80% 80% ofof invasiveinvasivebreastbreast cancercanceroccursoccurs

WHO ClassificationWHO Classification ofof breastbreast cancerscancers

* * Epithelial Epithelial

* * DuctalDuctal (85%) (85%) * * Lobular (1%) Lobular (1%) * * Papillary (<5%) Papillary (<5%)

* * Mixed Connective tissue and EpithelialMixed Connective tissue and Epithelial

inin womenwomenoverover ageage50.50.

TypicalTypical breastbreast cancercancerpresentpresent as a as a painlesspainless bbumpump

PPrognosticrognostic factorsfactors inin breastbreast cancercancer

•• AgeAgeYounger women have poorer prognosis of Younger women have poorer prognosis of equivalent stageequivalent stage

•• TumourTumour sizesizeDiameter of Diameter of tumourtumour correlates directly with survival correlates directly with survival

•• Lymph node statusLymph node statusSingle best prognostic factor Single best prognostic factor Direct correlation between number and level of Direct correlation between number and level of nodes involved and survival nodes involved and survival

•• Metastases Metastases Distant metastases worsen survival Distant metastases worsen survival

LocallyLocally advancedadvancedbreastbreast carcinomacarcinoma

A tumor A tumor accompaniedaccompaniedby by ulcerationulceration

LL ymphoedemaymphoedema

-- EstrogenEstrogenss areare producedproduced inin many many typestypes ofof cancerscancers, , regardlessregardless ofof thethe presencepresence ororabsenceabsenceofof ER ER andand PR, PR, bothboth inin prepre-- andand postmenopausalpostmenopausalwomenwomen..

-- Estrogen Estrogen levellevel inin thethe breastbreast tumorstumors inin postmenopausalpostmenopausalwomenwomen cancan be ~20be ~20--fold fold higherhigherthanthan inin serumserum..

Estrogen Estrogen synthesissynthesisinin breastbreast cancercancer

-- AboutAbout 70% 70% ofof breastbreast cancerscancershavehavereceptorsreceptors for for estrogensestrogensandand progesteroneprogesterone. Estrogen . Estrogen isis a major a major mitogenmitogen for for thisthis typetype ofof cancercancer..

-- ThereThere isis a a strongstrong correlationcorrelation betweenbetween thethe levellevel ofof estrogenesestrogenesinin postpost--menopausalmenopausalwomenwomenandand thethe riskrisk ofof breastbreast cancercancer..

Estrogen in breast cancer:

↑↑↑↑ Proliferation ↓↓↓↓ Apoptosis

BreastBreast cancercancer–– antiestrogenicantiestrogenictherapytherapy-- ItIt was was notifiednotified inin 1896 1896 thatthat ovarectomyovarectomy leadsleadsto to significantsignificant decreasedecreaseinin breastbreast carcinoma carcinoma ((GG.. BeadsonBeadson, Lancet), Lancet)..

-- HoweverHowever, , alreadyalready inin 1900 1900 assessedassessedthatthat ovarectomyovarectomy helpedhelped onlyonly inin 30% 30% ofof casescases, , andandimprovementsimprovements lastedlasted onlyonly for ~2 for ~2 yearsyears. . DespiteDespite thesethese limitationslimitations endocrynicendocrynic therapytherapybecamebecamea standard a standard inin treatmenttreatment ofof breastbreast cancercancer. . ItIt was was surgicalsurgical antiestrogenicantiestrogenictherapytherapy::

* * ovarectomyovarectomy(i(in n premenopausalpremenopausalwomenwomen))* * adrenaleadrenalectomyctomy (i(in n postpost--menopausalmenopausalwomenwomen))

-- SurgicalSurgical ovarectomyovarectomyhashasbeenbeenreplacedreplacedby by ovaryovary irradiationirradiation

-- For ~20 For ~20 yearsyearspharmacologicalpharmacologicalantiestrogenicantiestrogenictherapytherapy isis appliedapplied usingusing::

•• ER ER antagonistsantagonists•• aromatasearomataseinhibitorsinhibitors

-- ItIt was was foundfound inin 11971 971 thatthat antiestrogenicantiestrogenic therapytherapy isis effectiveeffective inin treatmenttreatment ofof cancerscancerswithwith a high a high levellevel ofof ER ER expressionexpression((improvementimprovement was was observedobservedinin 60% 60% ofof patientspatients) )

-- ImmunohistochemistryImmunohistochemistry evidencedevidenced thatthat cellcell inin thethe same tumor same tumor maymay havehave differendifferen levellevel ofof ER ER expressionexpression. . ThereforeTherefore, , thethe antiestrogenicantiestrogenic therapytherapy isis less less effectiveeffective inin thethe latelate stagesstages ofof cancercancer((higherhigher diversitydiversity ofof cellscells))..

AntiestrogenicAntiestrogenic therapytherapy –– selectionselectionofof patientspatients

CancerCancer withwith differentdifferent expressionexpressionofof ERER

-- TamoTamoxxifenifen isis anan antiestrogenantiestrogen,, whichwhich blocksblocks bindingbinding ofof estrogen to ERestrogen to ER..

-- ItIt was was inventedinvented duringduring studystudy on on contraceptivecontraceptivepillspills inin 19601960’’ s.s.

-- In 1970In 1970’’ s s laboratorylaboratory strategiesstrategies ofof breastbreast cancercancer preventionprevention usingusing tamoxifentamoxifen was was elaboratedelaborated. . HoweverHowever, , itit diddid not not stirstir upup thethe interestinterest ofof cliniciansclinicians, , whowho believedbelieved ininchemotherapychemotherapy..

-- 25 25 yearsyears laterlater tamoxifentamoxifen was was foundfound to be to be effectiveeffective alsoalso inin clinicalclinical practicepractice –– underunderconditioncondition ofof selectionselectionofof appropriateappropriate groupgroup ofof patienspatiens((cancerscancerswithwith high high levellevel ofof ER) ER) andandlonglong--termterm applicationapplication ((moremore thanthan 5 5 yearsyears))

-- TamoTamoxxifenifen couldcould be be usefuluseful inin womenwomenatat high high riskrisk ofof breastbreast cancercancer..

SERM (SERM (selectiveselectiveestrogenestrogen--receptor receptor modulatorsmodulators))TamoxifenTamoxifen

ComparisonComparison ofof shortshort--termterm andand longlong--termterm treatmenttreatment withwith tamoxifentamoxifen on on thethegrowthgrowth ofof breastbreast cancercancer inin ratsrats

PharmacologicallyPharmacologicallyinducedinducedcancercancer..TreatmentTreatmentwithwith tamoxifentamoxifenbeganbegan30 30 daysdaysafterafter inductioninduction

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-- MoreMore thanthan 13 000 13 000 womenwomen atat high high riskrisk ofof breastbreast cancercancer ((beforebefore andand afterafter menopausemenopause) ) werewere treatedtreated for 5 for 5 yearsyearswithwith tamoxifentamoxifen oror placeboplacebo. .

-- TamoTamoxxifenifen decreaseddecreasedthethe riskrisk ofof breastbreast cancercancerby by 50%.50%.

ThereThere werewere, , howeverhowever, , seriousserioussideside--effectseffects::

-- TendencyTendencyto to increasedincreasedbonebonefracturefracture raterate

-- IncreaseIncrease(2.6 (2.6 -- 4 4 foldfold) ) frequencyfrequency ofof uterusuterus cancercancer inin postmenopausalpostmenopausalwomenwomen..

In In thethe latelate 1980s 1980s itit was was recommendedrecommendedas a as a drugdrug protectingprotecting thethe womenwomen withwith a high a high riskrisk ofofbreastbreast cancercancer, but not , but not inin generalgeneralpopulationpopulation becausebecauseofof thethe riskrisk ofof cervicalcervical cancercancer..

TamoxifenTamoxifen –– clinicalclinical trialstrials

TamoxifenTamoxifen –– clinicalclinical trialstrials

-- TamoxifenTamoxifen therapytherapy whenwhen startedstarted directlydirectly afterafter surgicalsurgical removalremoval ofof earlyearly stagestage ofofestrogeneestrogene--sensitivesensitive cancercancer ((withwith ER ER expressionexpression) ) decreaseddecreasedmortalitymortality afterafter 5 5 yearsyears by by 28%.28%.

-- AfterAfter employmentemployment ofof tamoxifentamoxifen inin UK, UK, numbernumber ofof deathsdeaths causedcaused by by breastbreast cancercancerdecreaseddecreasedfromfrom 16 000 16 000 toto 12 80012 800..

NolvadexNolvadex

-- TamoTamoxxifenifen isis usedusedas a as a competitivecompetitive antagonistantagonistofof ER.ER.

-- AfterAfter bindingbinding tamoxifentamoxifen, , ERER maymay dimerizedimerize andand bind to consensus bind to consensus sequencesequence. . ChangedChangedconformationconformation ofof suchsuch a a complexcomplex changeschangesinteractioninteraction withwith corepressorscorepressorsandand coactivatorscoactivators, , whenwhen comparedcomparedwithwith estrogenestrogen--ligatedligated receptor.receptor.

-- EffectsEffects ofof tamoxifentamoxifen cancan be be tissuetissue--specificspecific. In . In somesometissuestissues((uterusuterus) ) thethe drugdrug behavesbehavesas as partialpartial agonistagonist. . ThereforeTherefore itsits effectivityeffectivity isis limitedlimited ..

-- TamoTamoxifenxifen doesdoesnot not decreasedecreasethethe expressionexpressionofof PR (PR (andand evenevencancan increaseincreaseitit ))....

-- PatientsPatientssufferingsuffering fromfrom cancercancer lackinglacking ER do not ER do not respondrespondto to treatmenttreatment withwith tamoxifentamoxifen. .

-- Many Many cancerscancersafterafter temporarytemporary improvementimprovement acquireacquire resistancyresistancyto to tamoxifentamoxifen andand startsstarts to to growthgrowth againagain. .

TamoxifenTamoxifen -- characteristicscharacteristics

Tissue SelectivityTissue Selectivity

•• AntagonistAntagonist -- bbreastreast, , uuterusterus

•• AgonistAgonist -- bbone, one, bbrain, rain, ccolonolon,, neuronsneurons, , bloodblood vesselsvessels

-- Ideal Profile for Ideal Profile for SERM SERM ((selectiveselective estrogenestrogen--receptor receptor modulatorsmodulators))

-- DrugDrug similarsimilar to to tamoxifentamoxifen, but , but withwith weakerweaker proestrogenicproestrogenicactivityactivity inin thethe uterusuterus, , expectedexpectedto to maintainsmaintains appropriateappropriate bonebonedensitydensity inin thethe postmenopausalpostmenopausalwomenwomen

-- EfficientEfficient chemopreventivechemopreventivedrugdrug: 13 : 13 breastbreast cancercancercasescasesout out ofof 5129 5129 womenwomeninin raloxifeneraloxifenegroupgroup (0.25%)(0.25%) versusversus27 out 27 out ofof 2576 2576 inin placebo placebo groupgroup (1.05%)(1.05%) inin threethree yearsyears-- MORE MORE trialtrial ..

-- OngoingOngoing clinicalclinical trialtrial was was designeddesignedto test to test itsits effectivityeffectivityinin preventionprevention ofof ischemicischemicheartheart diseasediseaseinin postmenopausalpostmenopausalwomenwomen-- no no positivepositive but but alsoalsono no negativenegativeeffectseffects. .

-- BeneficialBeneficial ((appliedapplied withwith estrogensestrogens) ) inin protectionprotection againstagainstosteoporosisosteoporosis..

SimilarlySimilarly effectiveeffective inin chemopreventionchemoprevention, but , but safersafer --less less sidesideeffectseffectsobservedobservedinin randomizedrandomized clinicalclinical trialstrials ..

SERM SERM -- RaloxifeneRaloxifene

EvistaEvista

WhyWhy doesdoestamoxifentamoxifen actact as as proestrogenproestrogeninin uterusuterus, , whilewhile raloxifeneraloxifene notnot??

-- SideSide chainchain ofof raloxifeneraloxifene isis locatedlocated 1 1 ÅÅ closercloser toto asparticaspartic acidacid 351351 thanthan sideside chainchain ofoftamoxifentamoxifen inin LBD (LBD ( importantimportant for SRC for SRC bindingbinding). ).

-- IfIf ER ER undergoesundergoes thethe mutamutationtion (D351Y), (D351Y), whatwhat happenshappens inin somesome cancerscancers, , raloxifeneraloxifenestartsstarts to to actact as estrogen.as estrogen.

tamoxifenraloxifene

Johnson & Dowsett, Nature Rew. 2003

AromataAromatasese-- AromataAromatasese((microsomalmicrosomal cytochromecytochromeP450P450) ) isis anan enzymeenzymetransformingtransforming androgensandrogensto to estrogensestrogens

-- In In humanhuman populationpopulation aromatasearomataseisis polymorphicpolymorphic andand itit maymay influence influence thethe estrogen estrogen levellevelinin womenwomen..

-- IncreasedIncreasedexpressionexpressionofof aromatasearomataseinin cancerscancers, , resultsresults fromfrom usageusageofof differentdifferent promoterpromoter((insteadinstead ofof promoterpromoter whichwhich isis normallynormally activeactive inin adipocytesadipocytes, , activeactive becomesbecomespromoterpromoternormallynormally activeactive inin thethe ovaryovary).).

AAromataromataseseinhibitorsinhibitors

-- AromataseAromataseinhibitorsinhibitors maymay be steroid be steroid oror nonnon--steroid steroid compoundscompounds. .

* * Steroid Steroid inhibitorsinhibitors ((e.g.e.g. exemestaneexemestane) ) areare derivetivesderivetives ofof androstenedionandrostenedionee((aromatasearomatasesubstratesubstrate) ) andand actact as as competitvecompetitve inhibitorsinhibitors . .

* * NonNon--steroid steroid inhibitorsinhibitors ((npnp. . letrozoleletrozole, , anastrozoleanastrozole) ) bind bind throughthrough nitrogennitrogen atom atom to iron atom to iron atom inin hemehemegroupgroup inin aromatasearomatase..

-- FirstFirst drugsdrugs werewere not not fullyfully specificspecificandand becausebecauseofof sidesideeffectseffectscouldcould not be not be usedusedinin a a fullyfullyefficientefficient dosesdosesbecausebecauseofof seriousserioussideside effectseffects. Many . Many aromatasearomataseinhibitorsinhibitors ((drugsdrugs ofof thethe I I andand II II generationsgenerations) ) introducedintroduced to to thethe market market atat thethe beginningbeginning 19199090s s havehave beenbeenwithdrownwithdrown ..

-- DrugsDrugs ofof thethe thirdthird generationgeneration ((e.g. e.g. letrozolletrozolee)) cancan be be appliedapplied inin dosesdoses whichwhich ininpostmenopausalpostmenopausalwomenwomen almostalmost completelycompletely inhibitinhibit aromatasearomatase(99%)(99%) andand do not do not produceproducesidesideeffectseffectsapartapart ofof thosethoseresultingresulting fromfrom inhibitioninhibition ofof hormonehormone synthesissynthesis..

ATACATAC trialtrial ((anastrozoleanastrozole, , tamoxifentamoxifen, or combination , or combination tamoxifentamoxifen and and anastrozoleanastrozole))

-- 9366 9366 postmenopausalpostmenopausalpatientspatients, , withwith earlyearly breastbreast cancercancerwerewere divideddivided to 3 to 3 groupsgroups: :

a) a) tamotamoxifxif enen ((antagonistantagonistofof ER)ER), , b) b) anastanastrr ozoleozole((aromatasearomataseinhibitor)inhibitor)c) c) combinationcombination ofof bothboth drugsdrugs

-- comparingcomparing to to tamoxifentamoxifen, afer , afer 3333 monthsmonths inin patientspatients treatedtreated withwith anastrozoleanastrozole was was demonstrateddemonstrated::

* much * much higherhigher proportionproportion ofof patientspatients freefree ofof diseasedisease* * significantlysignificantly less less casescasesofof developmentdevelopmentofof newnew cancercancer inin thethe secondsecondbreastbreast* less * less significantsignificant sideside--effectseffects, , althoughalthough higherhigher fracturefracture raterateandand higherhigher levellevel ofof boneboneresorptionresorption markersmarkers

-- combinationcombination ofof bothboth drugsdrugs was less was less effectiveeffectivethanthananastrozolanastrozolalonealone..

AromataseAromatase inhibitorsinhibitors –– clinicalclinical applicationsapplications

protectionprotection againstagainst distantdistant metastasismetastasis -- tamoxifentamoxifen v. v. letrozoleletrozole

Johnson & Dowsett, Nature Rew. 2003

WhatWhat wouldwould be be profitableprofitable to to rememberremember inin JuneJune::

-- StructureStructure andand isoformsisoforms ofof ER ER andand PRPR

-- Role Role ofof ER ER inin osteoporosisosteoporosis

-- ER inhibitor ER inhibitor andand aromatasearomataseinhibitorsinhibitors inin treatmenttreatmentofof breastbreast cancercancer

ThankThank youyou andand seeseeyouyou nextnext weekweek......

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https://https://biotka.mol.uj.edu.pl/~hemeoxygenasebiotka.mol.uj.edu.pl/~hemeoxygenase

VenusVenusfromfrom SchelklingenSchelklingen(35,000(35,000--40,000 40,000 yearsyearsago)ago)