Disturbances of Growth ControlDisturbances of Growth Control--& how to target them for therapy& how to target them for therapy
June 9, 2006June 9, 2006
Nancy HynesNancy HynesFriedrich Friedrich MiescherMiescher InstituteInstitute
SS07 #13704SS07 #13704
In the past 25 yrs cancer research has generated a largebody of knowledge on molecular mechanisms controlling allaspects of normal cellular physiology/signaling pathways.
These studies have also told us a lot about the molecularmechanisms underlying cancer development.
Goal – to design rational/molecularly targeted therapeutics for cancer treatment.
The signaling circuitry in a normal cell is integrated,The signaling circuitry in a normal cell is integrated,finely regulated and in tune with the external milieufinely regulated and in tune with the external milieu
Hanahan & Weinberg, Cell, 2000
Hanahan & Weinberg (2000) Cell
Normal cells are strictly dependent upon external cuesNormal cells are strictly dependent upon external cues
DifferentiateDifferentiate
DieDie
Divide/restDivide/rest
Hanahan & Weinberg (2000) Cell
The signaling circuitry in a cancer cell has undergoneThe signaling circuitry in a cancer cell has undergonemany changes many changes --
Red proteins:Mutated, lost oroverexpressed in cancer cells.
The circuitry in a tumor cell is The circuitry in a tumor cell is integrated, but no longer responsive to integrated, but no longer responsive to normal external cuesnormal external cues
Hanahan & Weinberg (2000) Cell
However, However, underlying principlesunderlying principles have begun to emerge and thesehave begun to emerge and theseprovide an important framework for understanding cancer andprovide an important framework for understanding cancer andfor designing therapies. for designing therapies.
In the past 25 yrs cancer research has generated a largeIn the past 25 yrs cancer research has generated a largebody of knowledge on molecular mechanisms controlling allbody of knowledge on molecular mechanisms controlling allaspects of normal cellular physiology. These studies have alsoaspects of normal cellular physiology. These studies have alsotold us a lot about the molecular mechanisms underlying told us a lot about the molecular mechanisms underlying cancer development. cancer development.
The apparent complexity makes the problem seem almostThe apparent complexity makes the problem seem almostunmanagableunmanagable! !
Hanahan & Weinberg (2000) Cell
Acquired Capabilities of Cancer CellsAcquired Capabilities of Cancer Cells
Coussens & Werb (2002) Nature 420:860
Cancer cells are influenced by their environment Cancer cells are influenced by their environment
Multiple cell types in the tumor environment Multiple cell types in the tumor environment
FibroblastsFibroblasts –– extracellularextracellular matrixmatrixMacrophagesMacrophages –– ligandligand production, e.g. VEGFproduction, e.g. VEGFEndothelial cellsEndothelial cells –– provide oxygen & growth factorsprovide oxygen & growth factors
--attract tumor cells to the blood & lymphattract tumor cells to the blood & lymphvesselsvessels
StressStressDNA damageDNA damageoncogenesoncogenes
Gain of function & loss of function mutations Gain of function & loss of function mutations contribute to the cancer phenotypecontribute to the cancer phenotype
Gain of function = Gain of function = oncogenesoncogenes
Loss of function = Loss of function = tumor suppressorstumor suppressors
Growth control pathways & cancerGrowth control pathways & cancer
The The RasRas/ERK & PI3K pathways are altered in most cancers. /ERK & PI3K pathways are altered in most cancers.
Shaw & Cantley (2006) Nature 441:424
Tumor Suppressors/Loss of Function MutationsTumor Suppressors/Loss of Function Mutations
Some tumor suppressors are lost in many cancer types:Some tumor suppressors are lost in many cancer types:PTEN, p53, PTEN, p53, RbRb
Loss of some tumor suppressors leads to cancer in specific organLoss of some tumor suppressors leads to cancer in specific organs:s:BRCA, APC, hMSH2, hMLH1, LKB1, VHLBRCA, APC, hMSH2, hMLH1, LKB1, VHL
Control functions/pathways important in many cell typesControl functions/pathways important in many cell types
Control cell specific functions Control cell specific functions
BRCA = breast cancer associated – breast/ovary cancerAPC = adenomatous polyposis coli – colon cancerMSH2 & MLH1 = DNA repair proteins – colon cancerLKB1 = ser/thr protein kinase 11 – GI tract polyps VHL = Von Hipple-Lindau – kidney cancer
Tumor SuppressorsTumor Suppressors
Tumor suppressor genes can be inherited:Tumor suppressor genes can be inherited:
1 mutant allele predisposes to specific tumor types1 mutant allele predisposes to specific tumor types22ndnd allele is lost during tumor developmentallele is lost during tumor development
Loss of tumor suppressor genes occurs in sporadic tumors:Loss of tumor suppressor genes occurs in sporadic tumors:
both copies are lost/mutated during cancer developmentboth copies are lost/mutated during cancer developmentusuallyusually inherited genes are also involved in sporadic cancerinherited genes are also involved in sporadic cancer
Loss of tumor suppressor genes relieve pathwaysLoss of tumor suppressor genes relieve pathwaysthat blocks cancer development.that blocks cancer development.
Retroviruses & CancerRetroviruses & Cancer
Proviral DNAProviral DNA: : -- encodes an oncogene encodes an oncogene -- activates an activates an oncogeneoncogene upon integration upon integration
RNARNA--containing virusescontaining virusesRSV MMTVMC29, SSV, Ha-MSVAEVEtc.
Rous Sarcoma VirusRous Sarcoma Virus
RSV is a replication competent virus encoding the v-src oncogene.
1911 - Isolated from a chicken fibrosarcoma by Peyton Rous.
1966 - Rous received the Nobel Prize for his discovery.
1977 - M.Bishop & H.Varmus - v-src related to c-src –our DNA encodes oncogenes! Nobel Prize in 1989.
c-src was “captured” by a retrovirus
SrcSrc characteristics characteristics
First member of the c-src family of cytoplasmic proteintyrosine kinases.
Domains promoting protein-protein interaction.
Src Homology domains = SH
SH2
SH3 Bind PBind P--TyrTyr Bind ProBind Pro--rich domainsrich domains
SH1 – kinasedomain
PP--Tyr 527 binds the SH2 domain of Tyr 527 binds the SH2 domain of SrcSrcmaintaining the maintaining the kinasekinase in an inactive state. in an inactive state.
AutoinhibitionAutoinhibition of of kinasekinase activityactivity
SH2 domains SH2 domains bind Pbind P--Tyr Tyr
SH3 domains SH3 domains bind Probind Pro--rich regions rich regions
CC--terminal terminal srcsrc kinasekinase = = CskCskPhosphorylatesPhosphorylates TyrTyr--527527
cc--SrcSrc is activated by various mechanismsis activated by various mechanisms
cc--SrcSrc activity is higher activity is higher insomeinsome human cancers (breast & colon). human cancers (breast & colon).
(STAT lecture (STAT lecture –– B. B. GronerGroner June 30June 30thth) )
Mouse Mammary Tumor Virus Mouse Mammary Tumor Virus -- MMTVMMTV
Transmitted in the milk of certain mouse strains (C3H).
Female mice have a high incidence of mammary tumors.
MMTV acts via insertional mutagenesis.
The first identified MMTV integration site was upstream of the int-1 gene (Wnt-1).
IntInt--1 = Wnt1 = Wnt--1, 1, an activator of the an activator of the ββ--catenin/Tcf pathwaycatenin/Tcf pathway
This pathway is activated in human colorectal tumorsmainly via mutation of APC. (Discussed in Summer 2005)
APC
HH--rasras -- the first the first oncogeneoncogenediscovered in humandiscovered in humantumor DNAtumor DNA
BarbacidBarbacid, Weinberg,, Weinberg,WiglerWigler grpsgrps ~1980~1980
T24 bladder T24 bladder cancer cell linecancer cell line
11stst slideslide
RasRas is mutated in manyis mutated in manyhuman tumors. human tumors.
KK--rasras or Nor N--rasras
2nd Slide2nd Slide
HH--rasras -- the first the first oncogeneoncogenediscovered in humandiscovered in humantumor DNAtumor DNA
Harvey Harvey MurineMurine Sarcoma Virus Sarcoma Virus –– HaHa--MSVMSV
Encodes the HEncodes the H--rasras oncogeneoncogene
RasRas encoding retroviruses have been isolated from mice, encoding retroviruses have been isolated from mice, rats and cats.rats and cats.
HH--rasras isolated from bladder tumor DNA had the same isolated from bladder tumor DNA had the same mutation found in viral Hmutation found in viral H--RasRas. .
RasRas proteins are proteins are ““G proteinsG proteins””that cycle between an inactive that cycle between an inactive GDPGDP--bound form and an activebound form and an activeGTPGTP--bound form.bound form.
OncogenicOncogenic RasRas proteins (viral or human) have an activating mutationproteins (viral or human) have an activating mutationthat prevents GAP from hydrolyzing GTP. that prevents GAP from hydrolyzing GTP.
Active Active RasRas is always is always ““onon”” &&signaling to its downstreamsignaling to its downstreameffectoreffector proteins.proteins.
Many classes of Many classes of RTKsRTKs are aberrantly activated in are aberrantly activated in human cancerhuman cancer
ErbB1ErbB1EGFREGFR
ErbB2ErbB2NeuNeuHER2HER2
ErbB3ErbB3HER3HER3
ErbB4ErbB4HER4HER4
Epidermal growth factor receptor & human cancer Epidermal growth factor receptor & human cancer
Avian Avian erythroblastosiserythroblastosis virus (AEV) virus (AEV)
Avian retrovirus which encodes two Avian retrovirus which encodes two oncogenesoncogenes::ErbAErbA and and ErbBErbB..
ErbAErbA is a mutated is a mutated ligandligand--independent version ofindependent version ofthe thyroid hormone receptor (T3).the thyroid hormone receptor (T3).
ErbBErbB is a mutated is a mutated ligandligand--independent version ofindependent version ofthe EGF receptor.the EGF receptor.
Concept of cooperating Concept of cooperating oncogenesoncogenes –– few if any few if any examples of a tumor arising from a single alteration.examples of a tumor arising from a single alteration.
Mechanisms leading to RTK activation in cancer:Mechanisms leading to RTK activation in cancer:using the ERBB receptors as an exampleusing the ERBB receptors as an example
Gene amplification & receptor Gene amplification & receptor overexpressionoverexpression
Mutations Mutations ––KinaseKinase Domain or ECDDomain or ECD
NRGARTGFα
Stromal cells
AutocrineAutocrine or or paracrineparacrine activation due to activation due to ligandligand availabilityavailability
Paul Ehrlich (1854Paul Ehrlich (1854--1915)1915)
1908 Nobel Prize1908 Nobel Prize
Targeting Cancer with Targeting Cancer with ““Magic BulletsMagic Bullets””
Ehrlich’s work on aniline dyes led to the concept that chemical substances might have special affinities for pathogenic organisms & cancer cells.
These dyes, referred to as “Magic Bullets”would go straight to organisms or cells &kill them.
AlkylatingAlkylating drugs were the 1drugs were the 1stst antianti--cancer agents usedcancer agents usedin the clinic & ~1940s in the clinic & ~1940s –– some success in lymphoma treatment.some success in lymphoma treatment.
Other types of chemotherapeutic drugs were developedOther types of chemotherapeutic drugs were developedin the 1950in the 1950--1960s 1960s –– drugs designed to block drugs designed to block purinepurine & & pyrimidinepyrimidine metabolism.metabolism.
Targeted antiTargeted anti--cancer drugs that block a cellular process, cancer drugs that block a cellular process, for example, DNA synthesis. for example, DNA synthesis.
Targeting Cancer with Targeting Cancer with ““Magic BulletsMagic Bullets””
Today antiToday anti--cancer drugs target specific proteins/pathwayscancer drugs target specific proteins/pathwaysknown to be altered in cancer cells. known to be altered in cancer cells.
(More history in – H.Varmus 2006 Science 312: 1162)
Inhibitors targeting different classes of Inhibitors targeting different classes of RTKsRTKs are are in the clinic or in advanced stages of developmentin the clinic or in advanced stages of development
EGFREGFR;;IressaIressaTarcevaTarceva
PDGFR/KitPDGFR/KitGleevecGleevec
FLT3FLT3::PKC412PKC412
VEGFRVEGFR::AvastinAvastinPTK787PTK787ZD6474ZD6474SU6668SU6668
FGFRFGFR::PD173074PD173074
Ret etc.Ret etc.
Sebolt-Leopold & English (2006) Nature 441: 457
KinaseKinase targeted antitargeted anti--cancer therapeuticscancer therapeutics
FDA Approved FDA Approved
KinaseKinase targeted antitargeted anti--cancer therapeutics in cancer therapeutics in development development
Sebolt-Leopold & English (2006) Nature 441: 457
mTORmTORCCICCI--779; RAD001779; RAD001PhosphoinositidePhosphoinositide kinasekinase--related related kinasekinase (PIKK) (PIKK)
Hernandez et al (2004) Trends in Call Biology 14:36
BCRBCR--AblAbl is constitutively active in CML; many experiments is constitutively active in CML; many experiments suggested that blocking suggested that blocking AblAbl kinasekinase activity would kill activity would kill leukemicleukemic cells. cells.
CML patients show a balanced translocation CML patients show a balanced translocation ––Philadelphia Chromosome described by Nowell & Hungerfordin the 1960s - encodes the BCR-Abl oncoprotein
Targeting BCRTargeting BCR--ABL in Chronic ABL in Chronic MyelogenousMyelogenous Leukemia Leukemia
The TKI The TKI imatinib/Glivecimatinib/Glivec was found to reverse the was found to reverse the cllinicalcllinicalsymptoms of CML symptoms of CML –– the Philadelphia Chromosome+ cells were lost. the Philadelphia Chromosome+ cells were lost.
First cancer in which a TKI was found effective. First cancer in which a TKI was found effective.
PDGFR Family Members & Cancer PDGFR Family Members & Cancer
PDGFR PDGFR –– ligandligand activated (lung tumors)activated (lung tumors)Kit Kit –– mutationsmutationsFlt3 Flt3 –– mutations (mutations (leukemiasleukemias) (PKC412) ) (PKC412)
Gleevec/imatinibGleevec/imatinib also targets the PDGF & Kit receptoralso targets the PDGF & Kit receptor
Kit Receptor & CancerKit Receptor & Cancer
Gastrointestinal Gastrointestinal stromalstromal tumors (GIST) are rare, tumors (GIST) are rare, however, they are the most frequent however, they are the most frequent mesenchymalmesenchymaltumors of the GI tract. tumors of the GI tract.
The majority of The majority of GISTsGISTs have activating mutations in thehave activating mutations in thecc--Kit receptor. Kit receptor. (Rubin et al 2001 Cancer Res., 61: 8118)(Rubin et al 2001 Cancer Res., 61: 8118)
GleevecGleevec blocks cblocks c--Kit Kit kinasekinase activity. activity. ((BuchdungerBuchdunger et al., 2000 J. et al., 2000 J. PharmacolPharmacol. Exp. . Exp. TherTher., 295: 139., 295: 139--145)145)
Complete Response to Complete Response to GleevecGleevec in a GIST patientin a GIST patient
StroobantsStroobants et al., 2003 et al., 2003 EurEur. J.Cancer 39: 2012. J.Cancer 39: 2012--20202020
GIST patient w abdominal metsbefore Glivec treatment
48 hr after Glivec 8 days after Glivec
Positron Emission Tomography (PET) with Positron Emission Tomography (PET) with 1818F F fluorodeoxyglucosefluorodeoxyglucose..
FGFR & CancerFGFR & Cancer
Gene amplification and/or Gene amplification and/or overexpressionoverexpression ofofreceptors occur in a high % of breast cancers.receptors occur in a high % of breast cancers.
20% FGFR20% FGFR--1155--10% FGFR10% FGFR--22
30% FGFR30% FGFR--44
Other cancers with high expression of Other cancers with high expression of FGFRsFGFRs::prostate cancerprostate cancergastric cancergastric cancercolorectal cancercolorectal cancerovarian cancerovarian cancer
FGFR
PPPPP P
PD
P
signaling
PD173074 is a selective FGFR PD173074 is a selective FGFR kinasekinase inhibitorinhibitor
PTyr
FGFR-4
SUM 52
PTyr
FGFR-2
SUM 52MDA-MB-453
- + - + - +- +MDA-MB-415
IP FGFR-4 IP FGFR-2
PD
PD173074 blocks FGFR phosphorylation in breast cancer cellsPD173074 blocks FGFR phosphorylation in breast cancer cells
KoziczakKoziczak et al., 2004 et al., 2004 OncogeneOncogene 23: 350123: 3501
MDA-MB-453
MCF10A
MDA-MB-415
SUM 52
day 0 day 4
day 0 day 4
day 0 day 4 day 8
day 0 day 4 day 8
day 8
day 8
DMSOPD173074
PD173074 blocks proliferation of cancer cells PD173074 blocks proliferation of cancer cells with active FGFR with active FGFR
MCF10A MCF10A –– normalnormalbreast cellsbreast cells
FGFRsFGFRs appear to be good therapeutic targets in breast cancer.appear to be good therapeutic targets in breast cancer.
KoziczakKoziczak et al., 2004 et al., 2004 OncogeneOncogene 23: 350123: 3501
proliferation, survival, migration & metastasisproliferation, survival, migration & metastasis
PPPPP
PPPPP
PPPPP
PPPPP
MAPK
AKT
STAT
SRC
mTOR
PPPPPPPPPP
PPPPP
PPPPP
PPPPP
PPPPPPPPPP
Nuclear CompartmentNuclear Compartment
Cytoplasmic DomainCytoplasmic DomainPPPPP
PPPPPPPPPPPPPPP
PPPPP PPPPPPI3K
ExtracellularExtracellular DomainDomain
Plasma MembranePlasma Membrane
ERBB Receptors
X
Z
ERBB targeted antibodies:HerceptinCetuximab
TKI
TKI
ERBB targeted tyrosine kinaseinhibitors (TKIs):GefitinibErlotinib
ERBB ERBB targetedtargeted inhibitorsinhibitors
Important conceptImportant concept
Despite the fact that individual tumors displayDespite the fact that individual tumors displaydifferent mutations, the pathways causing transformation different mutations, the pathways causing transformation of a normal cell into a cancer cell are often similar.of a normal cell into a cancer cell are often similar.
PI3K & MAPK pathwayPI3K & MAPK pathwayare active in most humanare active in most humantumors.tumors.
Growth control pathways & cancerGrowth control pathways & cancer
The PI3K/AKT pathway is altered in most cancers. The PI3K/AKT pathway is altered in most cancers.
Shaw & Cantley (2006) Nature 441:424
PI3K/PKB pathwayPI3K/PKB pathway
PI3K catalyzes the production of PI3K catalyzes the production of phosphatidylinositolphosphatidylinositol 3,4,53,4,5--triphosphatetriphosphate(PIP3) at the cell membrane.(PIP3) at the cell membrane.
PKB/Akt has two upstream PKB/Akt has two upstream kinaseskinasesrequired for its activation.required for its activation.
Ser308Ser308
Ser473Ser473
PI3K/PKB pathwayPI3K/PKB pathway
PIP3 recruits Akt to the membrane via its PH domainPIP3 recruits Akt to the membrane via its PH domain
PKB/Akt has many effectorsPKB/Akt has many effectors
Shaw & Cantley (2006) Nature 441:424
BadBad--PPBcl2 is free Bcl2 is free to promote to promote survivalsurvival
FOXOFOXO--PPp27 p27 cyclinE/Cdk2cyclinE/Cdk2
GSKGSK--33--PPCyclin D Cyclin D proliferationproliferation
AKT8 retrovirus isolated from AKT8 retrovirus isolated from thymomasthymomas in mice. in mice.
Retroviruses & the Retroviruses & the phosphoinositidephosphoinositide 33’’--kinase pathwaykinase pathway
The The oncogeneoncogene vv--Akt Akt -- encoded as a gagencoded as a gag--fusion proteinfusion proteinwith serinewith serine--threoninethreonine kinasekinase activity.activity.BellacosaBellacosa et al (1991) Science 254: 274et al (1991) Science 254: 274--277.277.
Stall,S (1987) PNAS 84: 5034Stall,S (1987) PNAS 84: 5034--50375037
The avian sarcoma virus 16 (ASV16) encodes an activeThe avian sarcoma virus 16 (ASV16) encodes an activeversion of the catalytic subversion of the catalytic sub--unit of PI 3unit of PI 3--kinase.kinase.
Chang et al (1997) Science 276: 1848.Chang et al (1997) Science 276: 1848.
The PI3K/PKB pathwayThe PI3K/PKB pathway
Somatic mutations in Somatic mutations in PIK3CAPIK3CA, which encodes the p110, which encodes the p110αα catalyticcatalyticsubsub--unit are found in many human tumors.unit are found in many human tumors.
Samuels et al., April 23, 2004, Science 304, 554.Samuels et al., April 23, 2004, Science 304, 554.
These mutations are predicted to lead to constitutiveThese mutations are predicted to lead to constitutiveactivity of the p110activity of the p110αα catalytic subcatalytic sub--unit.unit.
Tumor Suppressors/Loss of Function MutationsTumor Suppressors/Loss of Function Mutations
Some tumor suppressors are lost in many cancer types:Some tumor suppressors are lost in many cancer types:PTEN, p53, PTEN, p53, RbRb
Loss of some tumor suppressors leads to cancer in specific organLoss of some tumor suppressors leads to cancer in specific organs:s:BRCA, APC, hMSH2, hMLH1, LKB1, VHLBRCA, APC, hMSH2, hMLH1, LKB1, VHL
Control functions/pathways important in many cell typesControl functions/pathways important in many cell types
Control cell specific functions Control cell specific functions
BRCA = breast cancer associated – breast/ovary cancerAPC = adenomatous polyposis coli – colon cancerMSH2 & MLH1 = DNA repair proteins – colon cancerLKB1 = ser/thr protein kinase 11 – GI tract polyps VHL = Von Hipple-Lindau – kidney cancer
The PI3K pathway & PTENThe PI3K pathway & PTEN
PTEN was identified as a tumor suppressor gene frequently PTEN was identified as a tumor suppressor gene frequently deleted in advanced tumors.deleted in advanced tumors.Li et al. 1997 Science 275: 1943; Li et al. 1997 Science 275: 1943; SteckSteck et al 1997 Nat Genet 15: 356et al 1997 Nat Genet 15: 356
~ 50% of advanced human tumors carry inactivating mutations ~ 50% of advanced human tumors carry inactivating mutations in PTEN.in PTEN.
PTEN mutations are rare in breast cancer; LOH & promoterPTEN mutations are rare in breast cancer; LOH & promotermethylationmethylation leading to low expression of PTEN are common.leading to low expression of PTEN are common.
SHIP = SHIP = SHSH2 containing 2 containing iinositolnositol 55’’ pphosphatasehosphatase
PTEN = PTEN = pphosphatasehosphatase & & tentensinsin
The PI3K pathway & PTENThe PI3K pathway & PTEN
PTEN is a lipid PTEN is a lipid phosphatasephosphatase that antagonizes the action of PI3K by that antagonizes the action of PI3K by removing the 3removing the 3--phosphate from PtdIns(3,4,5)Pphosphate from PtdIns(3,4,5)P3.3.
PIP3PIP3
The PI3K pathway & PTENThe PI3K pathway & PTEN
PTEN PTEN downregulatesdownregulates a a kinasekinase cascade with important cellularcascade with important cellularmediators. mediators.
VivancoVivanco & Sawyers 2002 & Sawyers 2002 volvol 2, 4892, 489
PIP3PIP3 PIP2PIP2
The PI3K/PKB pathwayThe PI3K/PKB pathway
Active in most human tumorsActive in most human tumors
due to loss of negative regulatorsdue to loss of negative regulators
inactivating mutations in PTENinactivating mutations in PTEN
due to activating mutationsdue to activating mutations
catalytic subcatalytic sub--unit p110unit p110
LST8
LST8
RaptorRaptor
Alessi et al 2006 Ann Rev Biochem 75:137
mTORmTOR is controlled by growth factors & energy is controlled by growth factors & energy
Energy/stress/hypoxiaEnergy/stress/hypoxia
mTORmTOR = = mammalianmammalian targettarget of of rapamycinrapamycin
mTORmTOR is a member of the is a member of the phosphoinositidephosphoinositide--kinasekinase--related related kinasekinase family (PIKK). family (PIKK).
mTORmTOR acts as a central sensor for nutrientsacts as a central sensor for nutrients& energy.& energy.
LST8
LST8
RaptorRaptor
Alessi et al 2006 Ann Rev Biochem 75:137
mTORmTOR is controlled by growth factors & energy is controlled by growth factors & energy
PI3K activating mutations (p110PI3K activating mutations (p110αα))found in ~25% primary breast found in ~25% primary breast tumors. tumors.
Loss of PTEN (LOH) orLoss of PTEN (LOH) orpromoter promoter methylationmethylation--~30% breast tumors~30% breast tumors
mTOR
S65’
3’
40S
60S
40S
60S
S65’
3’P
4E-BP1eIF-4E
P P
eIF-4E4E-BP1P P P P
+
S6K1P P
ControlControl translationtranslation of of growthgrowth--relatedrelated mRNAsmRNAs
mTORmTOR controls translation of growthcontrols translation of growth--related mRNAsrelated mRNAs
Ribosomal proteins, cell cycle regulators (cyclin D) etc Ribosomal proteins, cell cycle regulators (cyclin D) etc ––proteins needed for tumor cell growth & proliferationproteins needed for tumor cell growth & proliferation
RapamycinRapamycin
FKBP12FKBP12RapamycinRapamycin blocks blocks mTORmTOR/Raptor /Raptor complex activitycomplex activity
40SS6
5’3’
60S
5’3’
40SS6
60S
5’3’
40SS6
Monosome
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
Disome
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
Trisome
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6
60S
40SS6 Polysome
60S
40SS6
40SS6
40SS6
60S
60S
60S
RibosomalSubunits
40SS6
5’3’
RapamycinRapamycin activity can be activity can be examined on examined on polysomepolysome profilesprofiles
RNA extraction
Northern Blot
QuantificationPolysome profile
Polysomefractions
80S
60S
40S
0
5
10
15
20
25
1 2 3 4 5 6 7 8 9 10 11
polysomal fractionsnon-polysomal
Relative
amou
nt
of c
yclin
D1
mRN
A (%)
polysomal fractionsnon-polysomal
Examine Examine polysomalpolysomal distribution of mRNA of interestdistribution of mRNA of interest
Cyclin D1 mRNA translation is sensitive to Cyclin D1 mRNA translation is sensitive to rapamycinrapamycin
KoziczakKoziczak & Hynes 2004 JBC 279:50004& Hynes 2004 JBC 279:50004
RapamycinRapamycin treated tumor cells have very low levels of cyclin D1 & treated tumor cells have very low levels of cyclin D1 & block in the G1 phase of the cell cycle.block in the G1 phase of the cell cycle.
Tumor Suppressors/Loss of Function MutationsTumor Suppressors/Loss of Function Mutations
Some tumor suppressors are lost in many cancer types:Some tumor suppressors are lost in many cancer types:PTEN, p53, PTEN, p53, RbRb
Loss of some tumor suppressors leads to cancer in specific organLoss of some tumor suppressors leads to cancer in specific organs:s:BRCA, APC, hMSH2, hMLH1, LKB1, VHLBRCA, APC, hMSH2, hMLH1, LKB1, VHL
Control functions/pathways important in many cell typesControl functions/pathways important in many cell types
Control cell specific functions Control cell specific functions
BRCA = breast cancer associated – breast/ovary cancerAPC = adenomatous polyposis coli – colon cancerMSH2 & MLH1 = DNA repair proteins – colon cancerLKB1 = ser/thr protein kinase 11 – GI tract polypsVHL = Von Hipple-Lindau – kidney cancer
LKB1 LKB1 kinasekinase activates AMPK & its family membersactivates AMPK & its family members
Alessi et al 2006 Ann Rev Biochem 75:137
LST8
LST8
RaptorRaptor
Alessi et al 2006 Ann Rev Biochem 75:137
Multiple Multiple mTORmTOR regulators are mutated in cancer regulators are mutated in cancer
PI3K activating mutations (p110PI3K activating mutations (p110αα))found in ~25% primary breast found in ~25% primary breast tumors. tumors.
Loss of PTEN (LOH) orLoss of PTEN (LOH) orpromoter promoter methylationmethylation--~30% breast tumors~30% breast tumors
~30% breast ~30% breast tumors have chromosomal tumors have chromosomal loss in 19p13.2loss in 19p13.2--13.313.3LKB1 locusLKB1 locus
Growth control pathways & cancerGrowth control pathways & cancer
The The RasRas/ERK & PI3K pathways /ERK & PI3K pathways are altered in most cancers. are altered in most cancers.
Sebolt-Leopold & English (2006) Nature 441: 457
Concentrated on PI3K/PKB pathway Concentrated on PI3K/PKB pathway
Tumor Suppressors/Loss of Function MutationsTumor Suppressors/Loss of Function Mutations
Some tumor suppressors are lost in many cancer types:Some tumor suppressors are lost in many cancer types:PTEN, p53, PTEN, p53, RbRb
Loss of some tumor suppressors leads to cancer in specific organLoss of some tumor suppressors leads to cancer in specific organs:s:BRCA, APC, hMSH2, hMLH1, LKB1, VHLBRCA, APC, hMSH2, hMLH1, LKB1, VHL
Control functions/pathways important in many cell typesControl functions/pathways important in many cell types
Control cell specific functions Control cell specific functions
BRCA = breast cancer associated – breast/ovary cancerAPC = adenomatous polyposis coli – colon cancerMSH2 & MLH1 = DNA repair proteins – colon cancerLKB1 = ser/thr protein kinase 11 – GI tract polyps VHL = Von Hipple-Lindau – kidney cancer
Tumor cells appropriate a blood supply from the host tissue by Tumor cells appropriate a blood supply from the host tissue by stimulating stimulating neoangiogenesisneoangiogenesis;;this process is required for tumor growth. this process is required for tumor growth.
Tumors and Tumors and neoangiogenesisneoangiogenesis
RTKsRTKs of the VEGF family and their ligands are involvedof the VEGF family and their ligands are involvedin tumorin tumor--induced induced neoangiogenesisneoangiogenesis..
VEGF = vascular endothelial cell growth factorVEGF = vascular endothelial cell growth factor
Ongoing angiogenesis & the increase in Ongoing angiogenesis & the increase in microvesselmicrovessel densitydensityaids invasive tumor cells to disseminate through the bloodstreamaids invasive tumor cells to disseminate through the bloodstreamto distant to distant metastaticmetastatic sites. sites.
VEGF receptor/VEGF Network VEGF receptor/VEGF Network
Lymphangiogenesis is controlled by VEGF-C/VEGF-D & theirreceptor VEGFR-3 present on lymphatic endothelium.
Tumor-associated angiogenesis is controlled by VEGF-A & itsreceptor VEGFR-2 present on vascular endothelium.
TammelaTammela et al 2004 et al 2004 Cardio.ResCardio.Res 65:55065:550
Tumor Tumor neoangiogenesisneoangiogenesis, VEGF & HIF, VEGF & HIF--11
Tumor cells produce VEGF,Tumor cells produce VEGF,and/or stimulate neighboring cells in the and/or stimulate neighboring cells in the stromastroma to synthesis it.to synthesis it.
Hypoxia stabilizes the TF HIFHypoxia stabilizes the TF HIF--11α, α, the VEGFthe VEGF promoter has a promoter has a HIFHIF--11αα binding site. binding site.
VEGF expression is tightly controlled; in the tumor settingVEGF expression is tightly controlled; in the tumor settingmultiple pathways contribute to its multiple pathways contribute to its upregulationupregulation, an important, an importantone being HIFone being HIF--11αα. . HIFHIF--11αα = hypoxia inducible factor= hypoxia inducible factor
PouyssegurPouyssegur et al 2006 Nature 441: 437et al 2006 Nature 441: 437
The VHL Tumor SuppressorThe VHL Tumor Suppressor
GermlineGermline mutations in VHL are responsible for von mutations in VHL are responsible for von HippleHipple--LindauLindaudisease, characterized by a predisposition to disease, characterized by a predisposition to tumorigenesistumorigenesisin the CNS, pancreas & kidney. in the CNS, pancreas & kidney.
VHL is inactivated in ~80% of sporadic,VHL is inactivated in ~80% of sporadic,clearclear--cell renal cancer. cell renal cancer.
VHL targets HIFVHL targets HIF--11αα for oxygenfor oxygen--dependent dependent ubiquitinationubiquitination & & degradation. degradation.
KrekKrek 2000, Nature Cell 2000, Nature Cell BiolBiol
OxygenOxygen--dependent degradation domain (ODD)dependent degradation domain (ODD)
VCBVCB--Cul2 complexCul2 complex
VHL, VHL, ElonginElongin B & C B & C complexedcomplexed withwithCul2 Cul2 -- form the VCBform the VCB--Cul2 complexCul2 complexwhich targets Hifwhich targets Hif--11α α for oxygenfor oxygen--dependent degradation. dependent degradation.
E1 activates E1 activates ubiquitinubiquitin
Activated Activated ubiquitinubiquitin is transferredis transferredto the E2 to the E2 ubiquitinubiquitin--conjugating enzymeconjugating enzyme
The E3 The E3 ubiquitinubiquitin ligaseligase confersconfersspecificity in targeting specificity in targeting UbUb totoappropriate substrates appropriate substrates --promoting degradation promoting degradation
VHL is an E3 VHL is an E3 ubiquitinubiquitin ligaseligase
Under conditions of Under conditions of normoxianormoxiaHIFHIF--11αα is continually degraded. is continually degraded.
Pro residues in the ODD Pro residues in the ODD domain are domain are ––OHOH& Hif& Hif--1 is degraded . 1 is degraded .
Hypoxia blocks Hypoxia blocks ubub--mediated mediated HIFHIF--11αα degradation. degradation.
HIFHIF--11α α degradationdegradation
The tumor suppressor VHLThe tumor suppressor VHL
VHL mutations in kidney cancer promote stabilization ofVHL mutations in kidney cancer promote stabilization ofHIFHIF--11αα, an important regulator of , an important regulator of proangiogenicproangiogenic factors factors including VEGF. including VEGF.
Mutations in VHL mimic hypoxia Mutations in VHL mimic hypoxia ––
HIFHIF--11αα degradation is preventeddegradation is prevented
PouyssegurPouyssegur et al 2006 Nature 441: 437et al 2006 Nature 441: 437
HIFHIF--11α α controlcontrol
* To read more on how hypoxia impacts on * To read more on how hypoxia impacts on mTORmTOR see:see:J. J. PouyssegurPouyssegur et al 2006 Nature 441:437 et al 2006 Nature 441:437
Tumor angiogenesis Tumor angiogenesis -- a therapeutic target a therapeutic target
Therapeutics that target the vascular endothelial cells, Therapeutics that target the vascular endothelial cells, which supply tumors with oxygen & nutrients, are predictedwhich supply tumors with oxygen & nutrients, are predictedto spare normal tissue to spare normal tissue -- leading to hypoxia & tumor cell death. leading to hypoxia & tumor cell death.
In normal adults angiogenesis is basically arrested;In normal adults angiogenesis is basically arrested;tumors are very dependent upon tumors are very dependent upon neoangiogenesisneoangiogenesis..
Tumors also induce Tumors also induce lymphangiogenesislymphangiogenesis; the ; the lymphaticslymphatics carry carry tumor cells to the lymph nodes & to distant organs.tumor cells to the lymph nodes & to distant organs.
Compounds targeting the VEGFR/VEGF network are used Compounds targeting the VEGFR/VEGF network are used in cancer treatment. in cancer treatment.
PTK787 PTK787
Targeting the VEGFR/VEGF Network Targeting the VEGFR/VEGF Network
AntiAnti--VEGFVEGF
VEGFVEGF--AA
VEGF blocking antibodies
Bevacizumab/AvastinBevacizumab/Avastin: humanized : humanized mAbmAbtargeting human VEGFtargeting human VEGF--AA
VEGFR TK inhibitors
PTK787: PTK787: VEGFRVEGFR--1, 1, --2 & 2 & --3 3
ERBB Receptors & CancerERBB Receptors & Cancer
Specific alterations in ERBB receptor expression Specific alterations in ERBB receptor expression and/or activation have been found in many human and/or activation have been found in many human tumors.tumors.
Breast, ovarian, NSCLC, Breast, ovarian, NSCLC, glioblastomaglioblastoma, SCC head , SCC head & neck, colon, bladder, etc.& neck, colon, bladder, etc.
ERBB Receptors and CancerERBB Receptors and Cancer
Cancer patients whose tumors have ERBB receptorCancer patients whose tumors have ERBB receptoralterations tend to have a more aggressive diseasealterations tend to have a more aggressive diseasethat is associated with factors predicting a poorthat is associated with factors predicting a poorclinical outcome. clinical outcome.
ERBB receptors have been intensely pursued as ERBB receptors have been intensely pursued as therapeutic targets. therapeutic targets.
antibodiesantibodiessmall molecule tyrosine small molecule tyrosine kinasekinase inhibitors (inhibitors (TKIsTKIs))
proliferation, survival, migration & metastasisproliferation, survival, migration & metastasis
PPPPP
PPPPP
PPPPP
PPPPP
MAPK
AKT
STAT
SRC
mTOR
PPPPPPPPPP
PPPPP
PPPPP
PPPPP
PPPPPPPPPP
Nuclear CompartmentNuclear Compartment
Cytoplasmic DomainCytoplasmic DomainPPPPP
PPPPPPPPPPPPPPP
PPPPP PPPPPPI3K
ExtracellularExtracellular DomainDomain
Plasma MembranePlasma Membrane
ERBB Receptors
X
Z
ERBB targeted antibodies:HerceptinCetuximab
TKI
TKI
ERBB targeted tyrosine kinaseinhibitors (TKIs):GefitinibErlotinib
ERBB ERBB targetedtargeted inhibitorsinhibitors
Trastuzumab(Herceptin)
Pertuzumab(Omnitarg)
Cetuximab(Erbitux)
Matuzumab
Panitumumab
ErbB2
ErbB2
EGFR
EGFR
EGFR
Genentech/Roche
Genentech
ImClone/Merck KGaABristol-Myers Squibb
Merck KGaA Phase II – NSCLC, gynecologic, pancreatic, Esophageal.
Abgenix
Approved ErbB2-overexpressing breast cancer; trials in combination with various drugs ongoing.
Phase II - ovarian, breast, prostate, NSCLC; inhibits ErbB2 dimerization; trials ongoing in low ErbB2expressors.
Approved CRC; trials in combination with various drugs ongoing- pancreatic, HNSCC & NSCLC.
Pivotal trial ongoing – 3rd
line CRC, other indications-RCC and NSCLC.
ErbBErbB targeted recombinant antibodies*targeted recombinant antibodies*
*humanized or *humanized or chimericchimeric
TargetTarget CompanyCompany
Hynes & Lane 2005 Nature Rev Cancer 5: 341Hynes & Lane 2005 Nature Rev Cancer 5: 341
ErbBErbB targeted targeted TKIsTKIs
TargetTarget CompanyCompany
Gefitinib(Iressa)
Erlotinib(Tarceva)
Lapatinib
AEE788
CI-1033
EKB-569
EXEL 7647/EXEL 0999
EGFR
EGFR
EGFR/ErbB2
EGFR/ErbB2/VEGFR
EGFR/ErbB2*
EGFR/ErbB2*
EGFR/ErbB2/KDR
AstraZeneca
Genentech/OSI Pharma
Approved 3rd line NSCLC, broad focus development.
GlaxoSmithKline
Novartis
Pfizer
Wyeth-Ayerst
EXELIXIS
Phase II - NSCLC
Approved 3rd line NSCLC, ongoing trials in other indications - HNSCC, GI & breast.
Phase III – Herceptin- & chemo-refractory breast cancer, other ongoing trials - refractory CRC
Phase I - First multi-function EGFR/ErbB2/VEGFR inhibitor; multiple potential Phase II indications.
Phase II – Breast, NSCLC
Phase I
*irreversible Hynes & Lane 2005 Nature Rev Cancer 5: 341Hynes & Lane 2005 Nature Rev Cancer 5: 341
ERBB Receptors and CancerERBB Receptors and Cancer
Understand mechanisms underlying activity of Understand mechanisms underlying activity of ERBB targeted inhibitors.ERBB targeted inhibitors.
How are patients chosen for treatment with a How are patients chosen for treatment with a specific inhibitor? specific inhibitor?
ERBB inhibitors in combination with other signal ERBB inhibitors in combination with other signal transduction inhibitors (transduction inhibitors (STIsSTIs).).
What mechanisms contribute to resistance?What mechanisms contribute to resistance?
EGFR EGFR kinasekinase domain mutations & NSCLCdomain mutations & NSCLC
In the initial clinical trials on In the initial clinical trials on gefitinibgefitinib & & erlotiniberlotinib,,most nonmost non--small cell lung cancer patients who respondedsmall cell lung cancer patients who respondedhad mutations in the had mutations in the kinasekinase domain of EGFR. domain of EGFR.
Lynch et al 2004 N Lynch et al 2004 N EnglEngl J Med 350 J Med 350 PaezPaez et al., 2004 Science 304et al., 2004 Science 304PaoPao et al 2004 Proc et al 2004 Proc NatlNatl AcadAcad SciSci 101101
First report on EGFR KD mutations.First report on EGFR KD mutations.
Clinical results suggested that KD mutations would Clinical results suggested that KD mutations would make a tumor more sensitive to make a tumor more sensitive to TKIsTKIs. .
Shigematsu & Gazdar 2006 Int J Cancer 118:257
Mutations target key structures around Mutations target key structures around the ATP binding cleft of the the ATP binding cleft of the kinasekinase domain domain
Mutations are predicted to keep the Mutations are predicted to keep the kinasekinasein the active conformation.in the active conformation.
Shigematsu & Gazdar 2006 Int J Cancer 118:257
EGFR EGFR kinasekinase domain mutations are domain mutations are oncogeniconcogenic
All EGFR KD mutations are All EGFR KD mutations are oncogeniconcogenic --mutant receptors transform* fibroblasts & epithelial cells.mutant receptors transform* fibroblasts & epithelial cells.((GreulichGreulich et al 2005 et al 2005 PLoSPLoS Medicine 2:e313)Medicine 2:e313)
(*soft agar growth, focus formation & tumor formation)(*soft agar growth, focus formation & tumor formation)
Frequency of Frequency of kinasekinase domain mutations domain mutations in EGFR/ErbB2 in NSCLC in EGFR/ErbB2 in NSCLC
5 studies exons 18-21 sequencedn = 1,108 - 350 mutations detected(31.6%)
EGFR EGFR
ErbB2ErbB2
3 studies sequenced KDn = 857 - 16 mutations detected(1.8%)
Shigematsu & Gazdar 2006 Int J Cancer 118:257
GermlineGermline mutation in EGFR mutation in EGFR kinasekinase domaindomain
A family with multiple cases of NSCLC associated with A family with multiple cases of NSCLC associated with germlinegermlinetransmission of an EGFR T790M mutation was described by D. Habertransmission of an EGFR T790M mutation was described by D. Haber& colleagues & colleagues (Bell et al 2005 December, Nature Genetics)(Bell et al 2005 December, Nature Genetics)
Gatekeeper residue in the ATP binding site
EGFR mutations & EGFR mutations & gefitinibgefitinib response in NSCLCresponse in NSCLC
6565--83% response rate for patients with mutant EGFR83% response rate for patients with mutant EGFR1010--14% 14% ““ WT EGFRWT EGFR
Why are the mutant receptors more sensitive to Why are the mutant receptors more sensitive to gefitinibgefitinib? ?
(Johnson & Janne 2005 JCO 23:6813)
Gazdar et al 2004 Trends Mol Med vol 10
Gefitinib/erlotinibGefitinib/erlotinib might bind the mutant might bind the mutant kinasekinase domain more readily domain more readily --In vitro evidence suggesting that mutant EGFR is more sensitive In vitro evidence suggesting that mutant EGFR is more sensitive to to TKIsTKIs
Predictions from EGFR Predictions from EGFR kinasekinase domain structure domain structure
Resistance to ERBB targeted Resistance to ERBB targeted TKIsTKIs
Resistance to Resistance to TKIsTKIs has emerged as a significant clinical problem, initiallyhas emerged as a significant clinical problem, initiallyin the context of CML & mutations in BCRin the context of CML & mutations in BCR--ABL, leading to ABL, leading to GleevecGleevec resistance. resistance. In clinical studies, T790M mutations were reported in cases of NIn clinical studies, T790M mutations were reported in cases of NSCLC thatSCLC thatrecur after initial TKI response. recur after initial TKI response.
The irreversible EGFR TKI CLThe irreversible EGFR TKI CL--387,785 blocks activity 387,785 blocks activity of receptors with T790M mutations. of receptors with T790M mutations.
TrastuzumabTrastuzumab & Breast Cancer& Breast Cancer--Questions that need to be addressed Questions that need to be addressed
How does How does tratuzumabtratuzumab function? function?
What contributes to What contributes to tratuzumabtratuzumab resistance / insensitivity? resistance / insensitivity?
% Cell Cycle Stage% Cell Cycle Stage
G1G1 SS G2/MG2/M
PBSPBS 6767 1212 2121
FRP5FRP5 6565 1616 2020
TrastuzumabTrastuzumab 9696 11 33
TrastuzumabTrastuzumab blocks proliferation of ErbB2blocks proliferation of ErbB2--ovexpressing breast cancer cellsovexpressing breast cancer cells
Lane et al 2000 MCB 20:3210Lane et al 2000 MCB 20:3210
This is accompanied by downThis is accompanied by down--regulation of pathway activity regulation of pathway activity ––PI3K/Akt. PI3K/Akt.
EGFR ErbB3 ErbB4Ferguson et al. (2003) Mol Cell Cho & Leahy (2002) Science Bouyain & Leahy (2005) PNAS
ErbB2Cho et al. (2003) Nature
ERBB receptor structures
In the absence of ligands EGFR, ErbB3 & ErbB4 are In the absence of ligands EGFR, ErbB3 & ErbB4 are ““closedclosed””..
ErbB2 has an extended structure.ErbB2 has an extended structure.
IIIIIIIV
Provided by D. LeahyProvided by D. Leahy
ErbB2 structure similar to ErbB2 structure similar to ligandligand--activated EGFRactivated EGFR
sErbB2sEGFR/EGF
EGF
IIIIIIIV
Provided by D. LeahyProvided by D. Leahy
ErbB2
Cho et al. (2003) NatureCho et al. (2003) Nature
TrastuzumabTrastuzumab
ErbB2/trastuzumab structure has been solvedErbB2/trastuzumab structure has been solved
IIIIIIIV
TrastuzumabTrastuzumab binding binding neither blocks ErbB2 neither blocks ErbB2 dimerizationdimerization with the other with the other ligandligand activated ERBB receptors, activated ERBB receptors, nor does it prevent ErbB2 activationnor does it prevent ErbB2 activation. .
Provided by D. LeahyProvided by D. Leahy
+BTC+HRG+EGF+PBS
93%
56% 61%
72% 60% 62%
57% 59%
+4D
5N
o tr
eat.
+BTC+HRG+EGF+PBS
93%
56% 61%
72% 60% 62%
57% 59%
+4D
5N
o tr
eat.
MotoyamaMotoyama et al., 2002 Cancer Res 62: 3151et al., 2002 Cancer Res 62: 3151
TrastuzumabTrastuzumab induced G1 block can be overcome by induced G1 block can be overcome by treatment with treatment with ErbBErbB ligandsligands
BT474 breast tumor cellsBT474 breast tumor cells
tras
tuzu
mab
tras
tuzu
mab
Cont
rol
Cont
rol
TrastuzumabTrastuzumab ResistanceResistance
Tumors might escape from trastuzumab if ERBB ligandsare present in high concentrations in the tumor environment.
One potential mechanism of trastuzumab resistance.
Provides a rationale for developing pan-ERBB inhibitors.
proliferation, survival, migration & metastasisproliferation, survival, migration & metastasis
PPPPP
PPPPP
PPPPP
PPPPP
MAPK
AKT
STAT
SRC
mTOR
PPPPPPPPPP
PPPPP
PPPPP
PPPPP
PPPPPPPPPP
Nuclear CompartmentNuclear Compartment
Cytoplasmic DomainCytoplasmic DomainPPPPP
PPPPPPPPPPPPPPP
PPPPP PPPPPPI3K
ExtracellularExtracellular DomainDomain
Plasma MembranePlasma Membrane
ERBB Receptors
X
Z
ERBB targeted antibodies:HerceptinCetuximab
TKI
TKI
ERBB targeted tyrosine kinaseinhibitors (TKIs):GefitinibErlotinib
ERBB ERBB targetedtargeted inhibitorsinhibitors
Mutations in signaling proteins downstream of ERBB receptorsmay play a role in sensitivity/resistance to ERBB targeted inhibitors.
Protein Production
GeneTranscription
Cell GrowthProliferation
Integrins
ILK
4E-BP1
eIF-4E P 70 S6k
PTEN
TSC1/2
mTOR
PI3K
AKT/PKB
FKBP12
Growth Factors
Rationale for combination of Rationale for combination of mTORmTOR + ERBB inhibitors+ ERBB inhibitorsDeregulated PI3K/Akt signalingDeregulated PI3K/Akt signalingactivates activates mTORmTOR, but pathway mutations , but pathway mutations can uncouple can uncouple mTORmTOR from the receptor.from the receptor.
Low PTEN levelsLow PTEN levels--implicated in resistance to implicated in resistance to TrastuzumabTrastuzumab..(Nagata et al., 2004 Cancer Cell)(Nagata et al., 2004 Cancer Cell)
Loss of negative regulators Loss of negative regulators --PTEN PTEN –– often found in often found in Breast cancer.Breast cancer.
PIK3CAPIK3CA mutations have beenmutations have beenfound in ~ 25% breast cancers.found in ~ 25% breast cancers.
Protein Production
GeneTranscription
Cell GrowthProliferation
Integrins
ILK
4E-BP1
eIF-4E P 70 S6k
PTEN
TSC1/2
mTOR
PI3K
AKT/PKB
FKBP12
Growth Factors
ErbBErbB inhibitor AEE788 inhibitor AEE788 ((TraxlerTraxler et al., 2004 Cancer Res 64)et al., 2004 Cancer Res 64)
RAD001
RAD001 binds the RAD001 binds the immunophilinimmunophilin FKBP12 FKBP12 & blocks & blocks mTORmTOR activityactivity((BoulayBoulay et al., 2004 Cancer Res 64)et al., 2004 Cancer Res 64)
Rationale for combination of Rationale for combination of mTORmTOR + + ErbBErbB inhibitorsinhibitors
Despite the multiple alterations in a cancer cell Despite the multiple alterations in a cancer cell --
Our increasing knowledge of the Our increasing knowledge of the mechanisms & pathways underlying mechanisms & pathways underlying cancer development has led to the cancer development has led to the development of new therapeutic development of new therapeutic strategies ! strategies !
Major goals for the future: Major goals for the future: -- choose the patient population to treat with a targeted inhibitochoose the patient population to treat with a targeted inhibitorr-- choose the optimal combination of inhibitorschoose the optimal combination of inhibitors-- understand mechanisms of resistance understand mechanisms of resistance