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E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

Th e pr e v i ous DNA m e t hy ltr an sferase i nh i b it o rs no t on l y b l o c k t h e catal ytic acti v ities o f DNM T s, bu t a l so tri gg er d e g ra d ati on o f t h ese p r o tei n s , es p eciall y DNM T s 1 and 3B T h is latter acti v it y is po te n tiall y im po rt ant for t h ei r ac ti v iti es f o r gene r e e xp ressi on b eca u se eac h o f t h ese tw o p r o te ins, e xp e r im en t a ll y , possess tr ans cri p ti on al re p ressi on p r op erties i nd e p e nd e nt o f t h ei r DNA m e t hy l a ti on ca t a l yt ic sites Th e azacy t os i ne nuc l eos i de s e xh i b it c o m p le x do se – res pon se c h a r acteri s ti cs. A t l ow conce ntrati on s ( 0.2 t o 1 μ M) , t h e e p i g e n etic acti v ities o f t hese d r ugs p r edo mi n ate , wit h do se- d e p e nd e n t re v ersal o f D NA met hy lati o and i nduc ti on o f termi n al d i f fere n tiati on i n s o me s y stems. As concen tr a ti on s are i n crease d, DNA d ama g e a nd a pop t o si s b ec o me mo r e p r o mi nen t . Cell li n es wit h 30 -f o l d resista n ce t o t h e c y t o t ox i c e f f ec t s o f dox ifl u ridi n e , a d riam y ci n, c y cl opho s ph ami d e (DAC ) c on ti nue t o r eve r se m e t hy l a ti on i n res pon se t o t h is nu cle o si d e , s ugg esti ng

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

t h at t h e m e t hy l a ti on r eve r s i ng a nd c y t o t ox ic acti v ities o f t h is c o m pound can b e se p a ra t ed . T he ab ilit y o f th ese d r ug s t o i nh i b it t h e cell c y cle , at least in p a r t t hrough i nduc ti on o f p2 1 W AF1/CIP1 e xp ressi on, c o m p licates t h e go al o f r e v e r si ng DNA m e t hy l a ti on, b eca u se t h e latter re qu ires DNA re p licati on w it h t h e azacy t os i ne nuc l eos i d e i n c o r po rate d i n t o t h e DNA . Th e im po rt ance o f l ow dos es o f t h e tw o azac y t o si n e nu cle o si d es t o ac h ie v e a t a r ge t ed t he r apeu ti c e f fect h as b ee n rece n tl y e xp l o re d i n a seri es of la bor at o r y obse r va ti ons. T ra n sie n t e xpo s u re o f bo t h le uk emia a nd s o li d t u m or c e ll s t o sub mi c r o m o l a r do ses i ndu ce s u c h cells t o und er go cell u lar r e progr ammi ng, acco m pan i ed by d ecreases i n a b ilit y t o cl on e i n l ong -ter m sel f-r e newa l assays and t o g r o w as e xp la n ts i n imm un e-i n c o m p ete n t mic e .

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

Th ese e f f ec t s occu r w it h pa rtial g e no me-wi d e DNA d emet hy lati on a nd c h a ng es i n gene exp r ess i on i n m u lti p le p at h wa y s po te n tiall y k e y f o r d riv ing t u m or i g e nes i s. Th e pha rm acok i ne ti c p r ope rties o f t h e tw o azac y t o si n e nu cle o si d es ar e als o v e ry im po rt an t t o cons i d er f o r t h eir cli n ical u se . I n t h is re g ar d, a ma jo r po te n tial cha ll enge f o r t he ir us a g e is t h e fact t h at t h ese d r ug s are h i gh l y un sta b le i n an aqueous so l u ti on, res u lti ng i n t h eir ra p i d hyd r o l y sis a nd r es u lta n t i nac ti va ti on . I n c li n ical p ractice , t h e d r ug s m u st b e a d mi n ister ed s hor tl y a ft e r r econs tit u ti on. T h e d r ug s are als o meta bo lize d by c y ti d i n e d eami n as e , l ead i ng t o a sho rt h alf-life i n p lasma . W h e n i n jecte d s ub c u ta neous l y , 5AC r eaches a ma x imal p lasma c on ce n trati on at 30 mi nu tes , w it h a t e rmi na l ha lf-life o f 1.5 t o 2.3 hou rs . At t h e U . S . F ood a nd Drug Ad mi n i s tr a ti on (F D A) a pp r ov e d do se o f 5 AC ( 75 m g / m 2 a d mi n is te r ed subcu t aneous l y d ail y f o r 7 d a y s) , p ea k p lasma c on ce n trati ons w e r e 3 t o 5 μ M, wh i ch i s we ll wit h i n t h e ra ng e o f DNMT i nh i b it o r y c on ce n t ra ti ons I n tr avenou s (IV) a d mi n istrati on o f t h e same do se h as led t o h i gh er peak p l as m a concent rati on s ( 1 1 μ M) wit h a s ho rter h alf-life ( a pprox i ma t e l y 22 mi nu t es ) . DAC g i v e n ov er 1 hou r IV at 15 t o 20 m g / m 2 produ ce d p l as m a concen tr a ti on s o f 1.1 t o 1.6 μ M du ri ng t h e i n f u si on ,

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

m g /m 2 w it h neu tr open i a as t h e do se-limiti ng t ox icit y . Stea dy -state p lasm a c on ce n t ra ti ons r anged fr o m 2 5 t o 40 n M , w h ic h is less t h a n t ho se u s u all y u se d t o i nduce exp r ess i on o f met hy late d g e n es i n tiss u e c u lt u re m od els .

E P IG E NETIC ABNORMA L I T IES AND GE NE EXPR E SSION CHANGES IN CAN C ER

DNA Methyltransferase Inhibitors

nan nan

HIS T ONE DEACETY L ASE IN H I B I T ORS

nan nan

Th e i n c reas i ng r ecogn iti on o f t h e critical im po rta n ce o f h ist on e m od i f ic a ti ons i n r egu l a ti ng t h e tra n scri p ti on al p ermissi v el y o f c h r o mati n h as le d t o i n t ense i n t e r es t i n co m pound s t h at ca n i nh i b it t h e acti v it y o f HDA C p r o t e i ns, f ac ilit a ti ng t h e acet y lati on o f l y si n es ass o ciate d wit h t r a n sc r i p ti ona l ac ti va ti on o f g e n es . As wit h t h e DNMT i nh i b it o rs d isc u s sed pr e v i ous l y , t he r e a r e m u lti p l e , s o metimes do se- d e p e nd e n t , effects o f HDA Cis i n p r ec li n i ca l s t ud i es. S o me o f t h ese ma y tr u l y b e e p i g e n etic , o t h e r s st r i c tl y cy t o t ox i c, and o t h ers a c o m b i n ati on o f bo t h . S o me acti on s o f HDAC i s m ay r e l a te t o alteri ng ho w c h r o mati n is ce n tral t o t he r e p ai r o f DNA. T hus, a t espec iall y h i gh do ses , t h ese c o m pound s ca n b l unt e f f icie n t r epa ir and even i ndu ce DNA b rea k s . T h ese e f fects ma y und er lie cell c y cl e a rr es t and i nduc ti o n o f cell d eat h as is o fte n ob ser v e d i n pr ecli n i ca l s t ud i es o f HDAC i s Pe rhaps nove l uses o f t hes e d r ug s ma y b e i n ferre d by res u lts fr o m re cent st ud ies sugges ti ng t hey cou l d b e e x tremel y po werf u l e p i g e n etic t h era py

HIS T ONE DEACETY L ASE IN H I B I T ORS

nan nan

HIS T ONE DEACETY L ASE IN H I B I T ORS

T ypes of Histone Deacetylase Inhibitors

nan nan

T ypes of Histone Deacetylase Inhibitors

HIS T ONE DEACETY L ASE IN H I B I T ORS

T ypes of Histone Deacetylase Inhibitors

Small Chain Fatty Acids

nan nan

Th e ea r li es t r epo rt o f t he use o f a n HDACi t o treat le uk emia d escri b e d t he t r eatme n t o f a ch il d w it h r e fr a ct o r y AML wit h i n tra v e nou s s od i u m bu t y r ate, w it h a co nco mit an t c l ea r ance o f p eri ph eral b l ood b last cells a nd a d ec r eme n t i n bone m a rr ow b lasts . N o res pon ses d e v el op e d i n a s ub se q u ent st udy of n i ne AM L pa ti en t s w ho were treate d wit h i n tra v e nou s bu t y rate .

HIS T ONE DEACETY L ASE IN H I B I T ORS

T ypes of Histone Deacetylase Inhibitors

nan nan

P h ase 1 s t ud i es o f sod i u m ph e ny l bu t y rate (NaPB) i n MDS a nd AML e xp l or e d 7 - day con ti nuous i n f u si on s a d mi n istere d m on t h l y o r b iwee k l y , and 21-d a y con ti nuous i n f us i ons a d mi n istere d m on t h l y . At t h e ma x im u m t o le r ate d dose ( 375 m g pe r k i log ram p er d a y ) , t h e mea n stea dy -state p la sma c on ce n t ra ti on was 0.3 m M, wi t h i n t h e ra ng e o f HDAC i nh i b iti on .

HIS T ONE DEACETY L ASE IN H I B I T ORS

T ypes of Histone Deacetylase Inhibitors

nan nan

I s o late d pa ti en t s deve l oped he mat o l og ic im p r ov eme n t i n res pon se t o NaPB .

HIS T ONE DEACETY L ASE IN H I B I T ORS

T ypes of Histone Deacetylase Inhibitors

nan nan

Simi la r t o Na P B, va l p r o i c a ci d (V P A) re qu ires n ear millim o lar c on ce n t ra ti ons t o e f f ec ti ve l y i nh i b it HDACs . Of 18 p atie n ts wit h MDS o r A M L w it h trili neage dysp l as ia treate d wit h V P A t o ta r g et p lasma c on ce n t ra ti ons o f 0.3 t o 0.7 mM , 6 p atie n ts d e v el op e d h emat o l og ic im prov e men t O f 20 e l de rl y p atie n ts wit h AML treate d wit h V P A , on l y 1 1 c ou l d r em a i n i n con tr o l l ong enough t o b e c on si d ere d e v al u a b le f o r r es pon s e. Fi ve had im p r ove me n t i n p latelet c oun ts . V P A i ndu ce d h emat o l og i c im p r ove m en t i n c o m b i n ati on wit h all-tra n sreti no ic aci d i n t wo of ei gh t pa ti en t s tr ea t ed w it h AML; a fl uo resce n ce i n sit u hyb ri d izati on a n al y sis showed de fi n iti ve evid e n ce o f termi n al d iffere n tiati on o f t h e mali gn a n t ce ll s A l a r ge r s t udy o f t h is c o m b i n ati on i ndu ce d h emat o l ogic r es pon s e i n on l y 2 o f 26 e l de rl y p atie n ts wit h AML . It a pp ears un li k el y t h at t h e s m a ll cha i n f a tt y ac i ds will d e v el op a n im po rta n t r o le i n t h e t r eatme n t o f m a li gnancy g i ven t h e a v aila b ilit y o f HDACis wit h v astl y gr eate r po t enc y .

HIS T ONE DEACETY L ASE IN H I B I T ORS

Hyd r oxamic Acids

nan nan

Hyd r oxamic Acids

HIS T ONE DEACETY L ASE IN H I B I T ORS

Hyd r oxamic Acids

nan nan

Th e F D A app r oved vo ri nos t a t as t h e first c o mmerciall y a v aila b le HDA Ci. Th e a pp r ova l was based on a cti v it y o f t h is a g e n t i n c u ta n e ou s T -cell l y m pho m a ( C T C L) . T h irt y -t hr ee p atie n ts wit h a me d ia n nu m b er o f fi v e pr i or s ys t e mi c t he r apy r eg ime n s recei v e d on e o f t h ree do se sc h e du les o f vor i no st a t i n a s i ng l e i ns tit u ti on st ud y Ei gh t p atie n ts ac h ie v e d a p artial r es pon s e, w it h a m ed i an tim e t o res pon se o f 12 wee k s a nd a me d ia n dur ati on o f r esponse o f 15 week s . O v erall , 45 % o f p atie n ts h a d relief o f prur it u s . F a ti gue, d i a rr hea, na usea , a nd t h r o m bo c y t op e n ia were c o mm on t ox icitie s. I n a m u lti cen t e r pha se 2 trial , 74 p atie n ts wit h rela p se d o r r e fr act ory C T C L we r e tr ea t ed wit h 400 m g d ail y . Similar t o t h e p ri o r st ud y , 29 % o f pa ti en t s r espond e d, c on sisti ng alm o st e n tirel y o f p artial r es pon s es. Med i an tim e t o r espon se was 56 d a y s , a nd me d ia n du rati on o f r es pon s e was g r ea t e r t han 6 mon t h s . I n ph ase 1 trials , res pon ses t o vor i no st a t have deve l oped i n ot h er non -H odgk i n ’ s a nd H odgk i n ’ s l y m pho m a cases Mo r e r ecen tl y , i n a trial c o m b i n i ng vo ri no stat wit h

HIS T ONE DEACETY L ASE IN H I B I T ORS

Cyclic T etrapeptides

nan nan

R o mi d e ps i n i s F DA app r oved f o r t h e treatme n t o f CTC L a nd p eri ph er al T - cell l y m pho m a . An tit u m o r acti v it y , i n cl ud i ng t u m o r l y sis s ynd r ome, w as d em ons tr a t ed i n a phase 1 st udy t h at e n r o lle d p atie n ts wit h c h r on ic l y m phocy ti c l euke mi a and AM L , bu t no c o m p lete o r p artial remissi on s w e r e se en . T he ad mi n i s tr a ti on o f r o mi d e p si n i ndu ces electr o car d i og ra phic c h a ng es , i nc l ud i ng T - wave fl a tte n i ng a nd S T -T wa v e d e p ressi on i n g reat e r t h a n h alf o f t he pos ttr ea tm en t traci ng s; ho we v e r , no c h a ng es i n ser u m ca rd iac tr opon i n l eve l s o r l e ft v e n tric u lar ejecti on fracti on h a v e b ee n r e por te d

HIS T ONE DEACETY L ASE IN H I B I T ORS

Benzamides

nan nan

Benzamides

HIS T ONE DEACETY L ASE IN H I B I T ORS

Benzamides

nan nan

En ti no stat , f o rm e rl y known a s MS- 275, was a d mi n istere d wee k l y times four t o pa ti en t s w it h r e l apsed and refract o r y AML i n a ph ase 1 st ud y . Inf ecti o n s, uns t eady ga t e, and s o m no le n ce were do se-limiti ng t ox icities . No cli n ical responses deve l oped, alt hough im p r ov eme n ts i n n e u tr oph il c ounts w e r e ob s e r ved . E n ti nos t a t d i d no t i n crease t h e res pon se rate i n p atie n ts w it h h i ghe r ri sk MD S and A ML wit h MDS-relate d c h a ng es w h e n c o m bined w it h aza c iti d i ne co m pa r ed t o a zaciti d i n e al on e . M o st rece n tl y , ho we ve r ,

HIS T ONE DEACETY L ASE IN H I B I T ORS

Benzamides

nan nan

st ud ies N S C L C sugges t t ha t en ti no stat c ou l d b e a v al u a b le t h era p e u tic a gent i n s o li d t u m o r s when used w it h esta b lis h e d t h era p ies . W h e n c o m b i n e d w ith t h e e p i de rm a l g r ow t h f ac t o r i nh i b it o r erl o ti n i b, i n a ra ndo mize d ph ase 2 t r ial for pa ti en t s w it h r ecu rr e nt a dv a n ce d NSCLC , e n ti no stat was no t e f f icacio us a l one bu t appea r ed t o c o m b i n e wit h erl o ti n i b t o b e n efit a g r o u p of p atie n t s whose t u m o r s con tai n e d b aseli n e h i gh E-ca dh eri n le v els . O ve r all s urv i v al i n t hese l a tt e r pa ti en ts y iel d e d a n i n crease d s u r v i v al b e n efit o f 9.4 v e r s u s 5.4 m on t hs. Fi na ll y , e n ti no stat si gn ifica n tl y i n crease d s u r v i v al wh e n c omb i ned w it h an a r o matase i nh i b it o r i n a ph ase 2 trial f o r p atie n ts w it h br e as t cance r

HIS T ONE DEACETY L ASE IN H I B I T ORS

Pharmacodynamic P r operties

nan nan

Th e a d mi n i s tr a ti on o f o r a l vor i no stat was ass o ciate d wit h a tra n sie n t i n c r ease i n ace t y l a ti on o f h i s t on e H 3 i n p eri ph eral b l ood l y m pho c y tes , wh ic h p e aked a t 2 hou r s pos t do si ng a nd re v erte d t o b aseli n e by 8 hou rs ; simila r changes we r e obse r ved i n t h e l y m ph nod e o f a treate d p atie n t wi th l y m pho m a T r ea tm en t w it h vo ri no stat was ass o ciate d wit h tra n sl o cati on o f pho s phory l a t ed s i gna l tr ansd ucer a nd acti v at o r o f tra n scri p ti on 3 (S T A T - 3 ) fro m nuc l eus t o cy t op l as m i n res pond i ng p atie n ts a nd wit h re du ce d mic rov e sse l dens it y . Simi la r changes i n t he acety lati on o f h ist on es 2 B a nd 3 were ob ser v e d in p e r i ph e ra l b l ood ce ll s fr o m p atie n ts treate d wit h LBH 589 . R o mi d e p sin i ndu ce d ace t y l a ti on o f H3 and H 4 i n p eri ph eral b l ood t u m o r cells wit h i n 4 hour s of dos i n o f i n t e r es t , p2 1 W AF1/CIP1 p r o tei n le v els als o i n crease d, ass o ciate d w it h an i nc r ease i n acet y lati on o f H 4 at t h e p21 p r o m o ter ( u si ng c hro mati n imm unop r ec i p it a ti on ) . T reatme n t wit h e n ti no stat le d t o i n creas ed acet y lati on o f H3 and H4 i n bo t h p eri ph eral b l ood a nd bon e marr o w . T his i n c r ease was de t ec t ab l e w it h i n 8 hou rs a nd remai n e d a bov e b aseli n e t hroughou t t he tr ea tm en t cyc le . T hu s , t h is c o m pound ma y p r ov i d e t h e m ost pro l onged i nh i b iti on o f p r o t e i n d eacet y lati on o f HDACis a nd is und er c urr e n t in ves ti ga ti on . I nc r e ases i n p2 1 W AF1/CIP1 a nd acti v ati on o f cas p as e 3 w e r e als o de m ons tr a t ed i n t hes e sam p les .

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

nan nan

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

DNA Methyltransferase Inhibitors

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

Ep i g e n eti c t he r apy has seen t h e m o st wi d es p rea d u se t o d ate a nd ac h ie ved t h e gr ea tes t e f fi cacy i n he m a tol og ic mali gn a n cies . T h e t h era p e u tic e f fica cy of 5A C and DAC f o r pa ti en t s wit h t h e c h r on ic m y el o i d n e op lasm m y el odysp l as i a ( MD S) and A ML h as b ee n well re v iewe d . T h eir FDA a pprov a l f o r MD S/ AM L e m e r g e d on l y after do ses were re du ce d, wit h r es u lta n t d imi n i sh i ng t ox i c iti e s f o r p atie n ts . T h e s u ccessf u l d e v el op me n t o f 5A C for t he tr ea tm en t o f MDS ca n b e cre d ite d lar g el y t o Sil v erma n et al i n t he Cance r and Le uk emia Gr oup B (CALGB) . T h e i nh i b it o r 5A C h ad success f u ll y i nduced t h e e xp ressi on o f h em og l ob i n F i n p atie nts w it h sic k l e ce ll ane mi a V iewi ng t h is c o m pound as a po te n tial i ndu c e r of te r mi na l d i f f e r en ti a ti on, Sil v erma n et al . c ondu cte d a series o f ph ase 2 t r ials of 5AC ad mi n i s t e r ed as a c on ti nuou s i n tra v e nou s i n f u si on o r as s ub c u ta neous i n j ec ti ons f o r t h e treatme n t o f MDS Base d on si gn ifi cant h emat o l og i c r esponses, t he g r oup p erf o rme d a ph ase 2 trial (CALGB 92 2 1 ) i n wh ic h pa ti en t s w it h l ow - and h i gh -ris k MDS wit h si gn ifica n t h emat opo i e ti c co m p r o mi se w ere ra ndo ml y assi gn e d t o recei v e s ub c u ta neous 5AC ( 75 m g / m 2 p er d a y d ail y f o r 7 d a y s , re p eate d on a 28-d a y c y cle) o r obse r va ti on. P a tie n ts on t h e ob ser v ati on arm wit h p r og ress ive d isease c ou l d c r oss ove r t o r ec ei v e 5 AC . T h is st udy firml y esta b lis h e d t he a b ilit y o f 5AC t o i nduce he m a t o l og ic im p r ov eme n t , a nd, less fre qu e n tl y , c o m p let e and pa rti a l r esponses . T h e me d ia n time t o d e v el op me n t o f A M L (de fi ned by 30 % bone m arr o w b last cells) o r d eat h was g reater i n the 5A C a rm by 9 m on t hs ( 21 ve rs u s 12 m on t h s); o f no te , t h e ob ser v ati on a rm i n cl ud e d pa ti en t s who subsequ e n tl y cr o sse d ov er t o 5 AC treatme n t . In a subsequen t phase 3 trial (AZA 001 ) p atie n ts wit h h i gh er ris k m y el odysp l as ti c synd r o m es were ra ndo ml y assi gn e d on e-t o - on e t o recei ve 5A C (75 m g / m 2 pe r day f o r 7 d a y s e v er y 28 d a y s) o r c onv e n ti on al care

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

(b est s uppo rti ve ca r e, l ow - do se c y tara b i n e , o r i n te n si v e c h em o t h era py as selecte d by i nves ti ga t o r s be f or e ra ndo mizati on ) . T h ree hund re d fift y -ei ght p atie n ts w e r e r ando ml y ass i gn e d t o recei v e 5 AC ( n = 179 ) o r c onv e n ti onal ca r e r e g im ens ( n = 179 ) . A ft er a me d ia n f o ll o w- up o f 21.1 m on t h s ( i n te rqu artil e r ange [I QR ] 15.1 t o 26.9 ) , me d ia n ov erall s u r v i v al was 24.5 m on t h s ( 9.9 no t r eached ) f o r t h e azaciti d i n e g r oup v ers u s 15.0 m on t h s ( 5.6 t o 24.1) f o r t he conven ti ona l care g r oup ( h azar d rati o [HR] 0.58 ; 95 % c onf i d e nce i n t e r va l [ C I] , 0.43 t o 0.77 ; p = 0.0001 ) . At 2 y ears , on t h e b as is of K a p l an - Me i e r es tim a t es, 50.8 % ( 95 % CI , 42.1 t o 58.8 ) o f p atie n ts i n the 5A C group we r e a li ve co m par e d wit h 26.2 % ( 95 % CI , 18.7 t o 34.3 ) i n th e c onv e n t iona l ca r e g r oup ( p < 0.0001 ) . Me d ia n time t o AML tra n sf o rmat ion w as 17.8 m on t hs (I QR 8.6 t o 36.8 ; 95 % CI , 13.6 t o 23.6 ) i n t h e 5 AC g r oup c o m p a red w it h 1 1.5 m on t hs ( 4.9 no t reac h e d ; 8.3 t o 14.5 ) i n t h e c onv e n t iona l ca r e g r oup ( HR 0.50 ; 95 % CI , 0.35 t o 0.70 ; p < 0.0001 ) . S ub se quen t unp l anned ana l ys es o f AZA 001 i n cl ud e d a n e x ami n ati on o f el d e r l y pa ti en t s w it h wha t wo ul d no w b e classifie d as AML ( b last c oun t 20% t o 30 %) . I n t hese 1 13 pat ie n ts , t h ere remai n s a statisticall y si gn ifica nt im prov e men t i n su r v i va l o f 24.5 m on t h s v ers u s 16.0 m on t h s (HR 0.47 ; 95 % C I ; p = 0.0001 ) Th e ea rl y deve l op m en t o f d ecita b i n e i n MDS t ook p lace p rimaril y i n Europ e unde r t he l eade r sh i p o f W ijerma n s et al . , T h ese i nv esti g at o rs pur s u e d i n tr avenous schedu li ng o f d ecita b i n e a d mi n istere d t h ree times daily for 3 d a ys ( 45 m g /m 2 pe r day t o tal do se) . T h is c y cle was re p eate d e v er y 6 w ee k s . P hase 2 s t ud i es sugge ste d a res pon se rate o f a pp r ox imatel y 50 % in M DS p ati en t s. I n a r ando mi z e d trial o f DAC v ers u s ob ser v ati on, p atie n t s w it h In ter na ti ona l Pr ognos ti c Sc o re ris k cate go ries i n terme d iate 1 t o h i gh r ecei v e d t he p r ev i ous l y li s t ed sc h e du le o f d ecita b i n e o r ob ser v ati on. N o c ro ss ove r was a ll owed i n t h i s trial . Res pon se rates re po rte d were: c o m p le te r es pon s e : 9 % , pa rti a l r esponse : 8 % , a nd h emat o l og ic im p r ov eme n t: 13 % .

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

a nd w el l t o l e r a t ed. A da il y -ti m es-fi v e sc h e du le ( 20 m g / m 2 p er d a y ) h as been F DA a pp r ove 99 pa ti en t s wit h MDS ( d e novo o r sec ond ar y ) o f a ny F r e n c h- Am e ri can - B riti sh ( F AB ) s ub t yp e a nd a n I n ter n ati on al Pr ogno sti c Sc or i ng Sys t e m (IPSS) sco r e equ al t o o r g reater t h a n 0.5 were treate d, w ith a n ov e ra ll r esponse r a t e o f 32 % ( 17 c o m p lete res pon ses [CR] p l u s 15 ma rrow C Rs [m CR ]) . A m o n g p atie n ts w ho im p r ov e d, 82 % d em on strat ed r es pon s es by t he end o f cyc l e tw o. T h is well-t o lerate d re g ime n all o ws ou t p atie n t ad mi n i s tr a ti on and, as no te d p re v i ou sl y , p r ov i d es p lasma le v e ls of d ecit ab i ne t ha t i nh i b it DN M T s . Th e 3 - day i n tr avenous sch e du le o f DAC h as b ee n st ud ie d i n tw o r a ndo mi zed tri a l s co m pa r ed t o s uppo rti v e care i n p atie n ts wit h h i gh er ri sk M DS. The fir s t tri a l con firm ed t h e h emat o l og ic acti v it y o f d ecita b i n e i n this p atie n t popu l a ti on bu t f a il ed t o s ho w a n im p r ov eme n t i n s u r v i v al i n t h e DA C - t rea t ed pa ti en t s . S u r v i v al was als o no t i n crease d i n t h e s ub se qu e nt t r ial , p e rfo rm ed by t he E u r ope a n O r g a n izati on f o r Researc h a nd T reatm ent of Ca n c e r (E O R T C ) . T he fail u re o f t h e ra ndo mize d d ecita b i n e trials t o s how a su r v i va l bene fit m ay b e p artiall y du e t o st udy d esi gn. B o t h r a ndo mi zed tri a l s o f 5AC cont i nu e d treatme n t un til d isease p r og ressi on f o r p atie n ts w ho d i d no t ach i eve c o m p lete remissi on ; i n fact , t h is mea n t t h at m o st p a t i en t s r ece i ved m a i n t e na n ce t h era p y . I n c on trast , bo t h ra ndo mize d t r ials of dec it ab i ne a ll owed a ma x im u m o f ei gh t c y cles o f treatme n t . T he n ee d for m a i n t enance t he r apy i n p atie n ts treate d wit h DNMT i nh i b it o rs has no t b ee n t es t ed i n p r ospec ti ve ra ndo mize d trials . A n a dd iti on al d i f fere n c e in t h e c onduc t o f t he t wo se t s o f DNMT i nh i b it o r trials i nvo l v es t h e du rati on of t h e r a py ad mi n i s t e r ed. T he me d ia n nu m b er o f c y cles o f treatme n t a d mi n is te r ed i n t he t wo r ando mize d trials o f d ecita b i n e was t h ree , c o m p a red t o n i ne i n t he azacy ti d i n e trials . T h is ma y reflect g reater t ox ic ity of t h e orig i na ll y 3 - day schedu le o f d ecita b i n e c o m p are d t o t h at o f t h e a pprov e d schedu l e o f 5AC. A lt hough t h e d i f fere n ces i n s u r v i v al ma y ref lect d i f f e r e nces i n tri a l des i gn and trial c ondu ct , emer g i ng d ata s ugg ests t h at d es p ite s imil a riti es i n m e t hy l a ti on re v ersal , t h e tw o d r ug s d i f fer i n o t h er po te n tiall y im po rt an t b i o l og i c p arameters , w h ic h ma y c on tri bu te t o cli n i cal ou tc o m es

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

T wo r ando mi zed phase 3 t r ials h a v e b ee n pub lis h e d treati ng el d erl y A M L pa ti en t s ( g r ea t e r t han 20 % b lasts) wit h d ecita b i n e , bo t h d em on stra ting im prov e men t i n su r v i va l t ha t was no t statisticall y si gn ifica n t . I n t h e Europ ea n s t ud y , 233 pa ti en t s r ecei v e d eit h er DAC at 15 m g / m 2 × 9 do ses ov e r 3 days on 42 - day cyc l es o r b est s uppo rti v e care . T h e p atie n ts recei ved a me d ian o f 4 cyc l es ( 0 t o 9 ) , a nd t h e ov erall s u r v i v al was im p r ov e d i n t he d ecita b in e -tr ea t ed pa ti en t s, bu t d i d no t reac h statistical si gn ifica n ce (med ian ov e r all su r v i va l [ O S] , 10.1 ver s u s 8.5 m on t h s , res p ecti v el y ; HR , 0.88 ; 95 % C I, 0.66 t o 1.17 ; t wo - s i ded, l og -ra nk p = 0.38 ) I n t h e M . D . A nd ers on Ca n ce r Cen t e r – l ed m u lti cen t er trial , 485 p atie n ts 65 y ears o r o l d er wer e r a ndo ml y ass i gned t o r ece i ve d ecita b i n e 20 m g / m 2 p er d a y as a 1 - hou r i n t r a v e nous i n f us i on f o r 5 con sec u ti v e d a y s e v er y 4 wee k s o r b est s uppor ti ve ca r e o r l ow - dose cy tara b i n e ( 20 m g / m 2 p er d a y f o r 10 d a y s e ve r y 4 w ee k s) . T he r e was a s imil a r im p r ov eme n t i n OS wit h d ecita b i n e ( 7.7 m on t h s; 95 % C I , 6.2 t o 9.2 ) ve rs u s t h e c on tr o l g r oup ( 5.0 m on t h s; 95 % CI , 4.3 t o 6.3 ; p = 0.108 ; HR, 0.8 5; 95 % CI , 0.69 t o 1.04 ) . Th e azacy t os i ne nuc l eos i de s re qu ire p r o l ong e d a d mi n istrati on t o d em on st ra t e he m a t o l og i c im p r ov eme n t i n MDS . Me d ia n time t o d e v el opmen t o f fir s t c li n i ca l res pon se i n t h e CALGB st ud ies o f 5 AC was t hr ee c y cl es ; 90 % o f r esponses d e v el op e d by c y cle si x . I n t h e ph ase 3 t r ial of d ecit ab i ne, t he m ed i an tim e t o res pon se was tw o c y cles , as als o see n in t h e alte rna ti ve r eg im en o f dec ita b i n e . It is , t h eref o re , e x tremel y im po r tant wh e n t rea ti ng pa ti en t s w it h a zac y t o si n e nu cle o si d es t o c o mmit t o a d mi n is te ri ng be t ween f ou r and si x c y cles o f t h era py b ef o re d etermi n i n g wh et h e r a pa ti en t i s r espond in g t o treatme n t . F u rt h erm o re , s u r v i v al b e n ef it is see n e ven i n pa ti en t s no t sho wi ng bon e marr o w im p r ov eme n t f o r 5 AC , p e rh a p s re l a t ed t o dec r eased tra n sf u si on re qu ireme n ts o r d ela y e d progr ess ion t o AM L . Beca use AM L i n t he con t ex t o f MDS is ar b itraril y d efi n e d b ase d on ma rrow bl as t coun t , ac ti v it y o f t h e aza nu cle o si d e a n al og s i n AML s hou ld no t b e s u r p ri s i ng. I n CA L GB 9221, 20 p atie n ts were reclassifie d upon ce n t r al pa t ho l ogy r ev i ew as meeti ng criteria f o r AML ( g reater t h a n 30 % b lasts ). The ir ou t co m es we r e c o m p ara b le t o t h e ov erall popu lati on i n t h e

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

st ud y . I n a ll t h r ee CA L GB st ud ies am ong p atie n ts meeti ng c u rre n t W o r ld H ealt h O r gan i za ti on ( WHO ) c riteria f o r AML ( g reater t h a n 20 % b lasts) , a c o m p let e r esponse was ach i ev e d i n 9 % a nd h emat o l og ic im p r ov eme n t i n 26% A r e tr ospec ti ve r ev i e w o f 20 p atie n ts wit h AML , i n cl ud i ng 8 p atie n ts wit h bone m a rr ow b l a sts g reater t h a n 29 % treate d wit h 5 AC , r e por te d a co m p l e t e r e mi ss i on i n 4 p atie n ts , a p artial res pon se i n 5, a nd a h emat o l og i c im p r ove m en t i n 3. T h e me d ia n du rati on o f res pon se was 8 m on t h s (r ange : 3 t o 33 m on t h s) DAC i ndu ce d a c o m p lete h emat o l og i c r es pon s e i n 2 o f 20 pa ti en t s tr e ate d w ho h a d t h e b lastic ph ase o f c h r on ic m y el o i d l euke mi a . T hese s t ud ies s ugg est acti v it y o f t h e azac y t o si n e nu cle o si des i n t he tr ea tm en t o f a s ub set o f AML p atie n ts . C u rre n t st ud ie s do no t all ow f o r t he de t e rmi na ti on o f w h et h er t h is s ub set is limite d t o MDS-ass o ciate d AM L ( AM L w it h M DS-relate d c h a ng es) , w h ic h te nd s t o h a ve l ow wh it e b l ood ce ll coun t s and h a v e a l o w p r o liferati v e rate , o r w h et h er t h ese c o m pounds a r e a l so ac ti v e f o r t ho se wit h AML wit hou t a h ist o r y o f a n tece den t he m a t o l og i c d i so r d e r . Se v eral re po rts d escri b e t h e se n siti v it y M DS a nd AM L , cha r ac t e ri zed by a bno rmalities o f c h r o m o s o me 7 a nd ass o ciate d w it h poo r ou t co m es i n res pon se t o c y tara b i n e- b ase d t h era py t o aza nu cl eos i des. I n one non r ando mize d retr o s p ecti v e st ud y , s u r v i v al o f s uch p atie n ts fo ll ow i ng t he ad mi n istrati on o f DNMT i nh i b it o rs s u r p asse d s urv i v al i n r esponse t o conven ti on al c y t o t ox ic c h em o t h era p y , similar t o t he ou tc o m es o f A Z A001 . A lt hough t he m echan i s m s und erl y i ng t h e cli n ical acti v it y o f azac y t o s ine a n al og s m ay i nvo l ve r eve r sa l o f g e n e met hy lati on, o t h er acti on s n ee d t o be c on si d e red. T he ad mi n i s tr a ti o n o f DAC h as b ee n s ho w n t o i ndu ce tra n si ent d ec r eme n t s o f m e t hy l a ti on i n non c od i ng re g i on s , i n cl ud i ng l ong i n te r s p e rsed nuc l ea r e l e m en t (LINE) a nd ALU eleme n ts . Earl y st ud ies t h at e x a m i ned m e t hy l a ti on r e versal o f t h e tar g et g e n e p1 5 INK4B i n res pon s e t o DA C showed no co rr e l a ti on b etwee n met hy lati on re v ersal a nd cli n ica l r es pon s e. C li n i ca l r espond ers t o DAC d e v el op e d si gn ifica n tl y h i ghe r e xpr essi on o f t h i s gene f o ll ow i ng treatme n t , a nd certai n l y k e y b i o l og ic r oles for t h is gene and it s l ow basa l e xp ressi on are p r ob a b le . M o re ov e r , i n one st ud y , t he c li n i ca l r esponse wa s cl o sel y ass o ciate d wit h t h e re v ersal o f

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

DNA Methyltransferase Inhibitors

nan nan

met hy lati on o f p15 o r CDH - 1 du ri ng t h e first c y cle o f treatme n t wit h 5 AC fo ll ow e d by t he HDAC i Na PB I n t h at st ud y , it was no tew o rt hy t h at t he a d mi n is t r a ti on o f 5AC p ri o r t o t h e a dd iti on o f a n HDACi was ass o ciate d w it h t h e i nduc ti on o f h i s t one acet y lati on. Alt hough t h e mec h a n ism und e r l y i ng t h i s ac ti v it y i s unkno w n, h ist on e acet y lati on h as b ee n ob ser ved fo ll ow i ng DNA da m age due to g amma irra d iati on . S ub se qu e n t st ud ie s h a v e found de m e t hy l a ti on f o l lo wi ng treatme n t wit h eit h er DAC o r 5 A C – bu t no t cons i s t en tl y assoc iate d wit h res pon se . , M o re w o r k will be r e qu i r e d t o answe r t he im po rta n t mec h a n istic qu esti on und er p i nn i ng t h e cli n ical a c ti v it y o f azacy t os i n e a n al og s .

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Combining Inhibitors in the T r eatment of Hematologic Malignancies

nan nan

I t is alm os t ce rt a i n t ha t t he b i gg est p r o mise o f e p i g e n etic t h era py lies i n st r ate g ie s t o co m b i ne ex i s ti ng a nd n ewer d r ug s wit h eac h o t h er a nd wit h c urr e n t che m o t he r ap i es and ta r g ete d t h era p ies . T o d ate , t h e e x am p le f o r e x isti ng agen t s i s t he co m b i na ti on o f DNMT i nh i b it o rs a nd HDAC i nh i b it ors based on t he hypo t h esis fr o m t h e la bo rat o r y t h at t h is p ara d i g m lea d s t o op tim a l r eexp r ess i on o f tra n scri p ti on all y sile n ce d g e n es wit h pro m o ter m e t hy l a ti on . T h is i n v itr o treatme n t p ara d i g m h as le d t o a v a r iet y o f c li n i ca l s t ud i es t ha t h a v e attem p te d t o a pp l y t h is c on ce p t t o t h e t r eatme n t o f he m a t o l og i c m a li gn a n cies . M u c h remai n s t o b e d etermi n e d w it h r e ga r d t o it s e f fi cacy an d p recisel y w h at d etermi n es t h is . T h e first st udy of sequen ti a l DNM T/ H DAC i nh i b it o rs a d mi n istere d a v ariet y o f do ses of 5AC f o r 5 t o 14 days f o ll o we d by 7 d a y s o f NaPB by c on ti nuous i nfu si on a t it s m ax im u m t o l e rate d do se t o p atie n ts wit h MDS a nd AML .

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Combining Inhibitors in the T r eatment of Hematologic Malignancies

nan nan

Th e c o m b i na ti on was we ll t o lerate d, a nd cli n ical res pon ses were fre qu e nt in p atie n ts rece i v i ng 5AC a t 50 m g / m 2 p er d a y d ail y f o r 10 d a y s a nd 25 m g/ m 2 p e r d a y da il y f o r 14 days, w it h 5 o f 14 p atie n ts at t ho se do se sc h e du les ac h ie v i n g co m p l e t e o r pa rti a l res pon se . In a p il o t s t ud y , 10 pa ti en t s wit h MDS o r AML were treate d wit h 5 AC at 75 m g /m 2 pe r day da il y tim es se v e n f o ll o we d by 5 d a y s o f NaPB g i v e n at

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Epigenetically T argeted Therapy in Nonhematologic Malignancies

nan nan

Epigenetically T argeted Therapy in Nonhematologic Malignancies

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Epigenetically T argeted Therapy in Nonhematologic Malignancies

nan nan

Th e e f f ic ac i es t ha t have e m e r g e d i n t h e a pp licati on o f e p i g e n eticall y ta r g ete d d r ugs t o he m a t o l og i c mali gn a n cies h as s pu rre d i n terest i n u si ng

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Epigenetically T argeted Therapy in Nonhematologic Malignancies

nan nan

e p i g e n et ic t he r apy f o r o t he r ty p es o f ca n ce r . As ou tli n e d as f o ll o ws , la bor at o r y s t ud i es and c li n i cal trials s uppo rt t h is a pp r o ac h. St ud ies i n t he la b h a v e been d ir ec t ed by l esson s lear n e d fr o m t h era py i n h emat o l og ic mali gn a nc i es, sugges ti ng t ha t l o w do ses o f d r ug s li k e DAC a nd 5 AC , i n the n a no m o l a r r ange, m ay avo i d ex cess t ox icities du e t o o ff-ta r g et e f fects o f the drug s a nd m ay m ax imi ze ep i g e n etic effects o f t h e a g e n ts . T h e d esire d e f f ects m ay r equ ir e mi n imi z i ng i n itial cell u lar c y t o t ox icit y , g i v i ng t u m o r cells tim e t o acc r ue m ax im a l c ell u lar re p r og rammi ng res pon ses t o t h e i nh i b iti o n o f DNM T s DAC and 5 AC are e f fecti v e on l y w h e n t h e y h a ve b ee n i n cor po r a t ed i n t o DNA, a fter w h ic h t h e y irre v ersi b l y i nh i b it DNMT catal y tic ac ti v it y and t a r ge t t h ese p r o tei n s f o r d e g ra d ati on . I n cell c u l tu r e a nd m ouse exp l an t s, l ow nano m o lar do ses a pp ear t o i ndu ce bo t h hu ma n le uk emi c and so li d t u m o r ce lls t o e xh i b it b l un ti ng o f self-re n ewal a nd t u m or i g e n i c ac ti v it y o f t u m o r stem-li k e cells . T h ese p recli n ical res u lts s ugg est a key poss i b ilit y t ha t u se o f e p i g e n etic t h era p ies mi gh t i nh i b it t h e se latte r ce l l popu l a ti ons, wh i ch o fte n are d iffic u lt t o era d icate a nd are a fa cto r i n r esist ance t o m any s t anda r d ca n cer t h era p ies . E xh a u sti on o f s u c h ce lls ov e r tim e du ri ng t he r apy w it h DAC o r 5 AC mi gh t e xp lai n t h e ob ser v ati on t h at m os t pa ti en t s w it h MD S/AML ta k e se v eral m on t h s t o reac h b est r es pon s e. L euke mi c s t e m c ells were no t elimi n ate d i n on e st udy i n MD S a nd A ML pa ti en t s tr ea t ed w it h 5 AC i n c o m b i n ati on wit h V P A , alt hough t h ei r fr eq uency dec r eased i n cli n ical res pond ers . Cli n i ca l tri a l s f o r co mm on s o li d t u m o rs , i n f o rme d t h r ough t h e p re v i ous la bor at o r y s t ud i es, have been i n itiate d i n cl ud i ng ph ase 2 d esi gn s u si ng l ow - do se st ra t eg i es w it h 5AC o ft en c o m b i n e d wit h u se o f h ist on e d eacet y las e i nh i b it ors. Si x t y -fi ve pa ti en t s w it h a dv a n ce d, m u lti p l y treate d NSCLCs w e r e t r eat ed w it h 5AC p l us en ti no stat O n l y 3 % o f p atie n ts d e v el op e d Res pon se E va l ua ti on C rit e ri a (RECIST)-meas u rea b le res pon ses; ho we v e r , t h ese t wo pa ti en t s had du r ab l e res pon ses , wit h s u r v i v al o f 3 t o 4 y ears .

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Epigenetically T argeted Therapy in Nonhematologic Malignancies

nan nan

E P IG E NETIC THERAPY FO R H EM A T OL OG IC MA LI G NANCIES

Epigenetically T argeted Therapy in Nonhematologic Malignancies

nan nan

Upr e gu l a ti on o f imm unogen i c p at h wa y s i n NSCLC a nd o t h er s o li d t u m o r cells , obse r ved i n l abo r a t o r y st ud ies , s ugg est a po te n tial f o r se qu e n ci ng DN M T i nh i b it o r s w it h imm un e c h ec kpo i n t i nh i b it o rs . T h is d r ug is als o r e por te d t o i nduce an tit u m o r res pon ses a nd imm un e rec ogn iti on i n a m o d el

NEW APROACHES T O EPIGEN ET IC T HE RA P Y

nan nan

NEW APROACHES T O EPIGEN ET IC T HE RA P Y

NEW APROACHES T O EPIGEN ET IC T HE RA P Y

nan nan

A s w e have ou tli ned p r ev i ous l y , t h e eme r g i ng p r o mise f o r e p i g e n etic t h e r a py and t he f u t u r e o f t he a pp r o ac h es ma y lie i n c o m b i n at o rial d r ug st r ate g ie s. A lt hough t h i s i s a lrea dy b ei ng e xp l o re d wit h o l d er a g e n ts , n e w drug s fo r new t a r ge t s a r e now e n teri ng t h e p ict u re . I n t h ese e f f o rt s, se v e r al t he m es we have i n tr odu ce d i n t h is c h a p ter will li k el y do mi n ate . M o st ep i gene ti c t he r ap i es will no t i ndu ce , w h e n u se d at tr u l y ta r g eti ng do ses , imm ed i a t e cy t o t ox i c e f fects . T h era p e u tic e f ficac y b ase d on cell u l a r r e progr ammi ng m ay r equ ir e s i gn ifica n t time t o ma n ifest . Cli n ical trial d esi gn s m ay need adap t a ti on s o t h at e f fecti v e t h era p ies are no t d iscar d e d du e t o p r e m a t u r e r esponse ev al u ati on s . Fi n all y , t h e u ltimate p r o mise f o r e p i g e n et ic t he r apy m ay li e w i th n ewer d r ug s no w e n teri ng cli n ical trials . Ou tc o me s w it h DNM T i nh i b it o rs ma y b e im p r ov e d wit h alter n ati v e sc h e du li ng o f o r a l azac iti d i ne o r t h r ough p r o l ong e d ph armac ok i n etics o f the d ecita b in e p r od r ug S G I 1 10. Als o, d r ug s ta r g eti ng o t h er p r o tei n s i n cl ud i ng B ET f a m il y b r o m odo m a i n p r o tei n s are g e n erati ng m u c h e x citeme n t . , –

NEW APROACHES T O EPIGEN ET IC T HE RA P Y

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B ET inh i b it o r s m ay i n t e rf e re wit h l o calizati on o f t h e on c og e n e C-MYC t o acet yla t ed l ys i nes i n r egu l ato r y re g i on s o f tar g et g e n es . T h ese i nh i b it ors a r e now en t e ri ng c li n ical trials . Ot h er p r o misi ng a pp r o ac h es i n cl ud e t he use o f i nh i b it o r s o f EZH 2, t h e e n z y me i n t h e PcG s y stem , w hich catal y zes t he r ep r ess i ve h i s t on e mar k H 3 K 27 me 3 . , A no t h er cli n ic al t r ial unde r way e m p l oys t a r get i ng o f t h e tra n sl o cati on i n w h ic h t h e p r o tei n mi x e d li neage l euke mi a ( M LL) is f u se d wit h se v eral ta r g ets , s u c h as i n i nf a n t le uke mi as. T hese tr ans l o cati on s res u lt i n a bno rmal recr u itme n t o f the h ist on e me t hy ltr ans f e r ase, DO T 1 L , t o tar g et g e n es li k e HOXA 9 T h is fu si on i nduces hype rm e t hy l a ti on o f H 3 K 79 a nd a bno rmal acti v ati on o f M LL ta r ge t genes. , V e r y s electi v e i nh i b it o rs o f DOT 1 L are no w i n cli n ical t ri a l s. Ep i gene ti ca ll y t a r ge t ed t he ra p ies c on ti nu e t o ho l d g reat p r o mise t h at r e progr ammi ng o f m a li gnan t cells c ou l d alter a pp r o ac h es t o ca n cer ma n a g e men t . Str a t eg i es t o me r g e o l d er d r ug s , w h ic h we h a v e f o c u se d on in t h is c h a p t e r , w it h t he newe r ag e n ts b riefl y d isc u sse d i n t h is secti on, will und e rp i n f u t u r e tri a l s t o t es t t h is a pp r o ac h.

NEW APROACHES T O EPIGEN ET IC T HE RA P Y

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Xu GL, Bestor TH, Bourc’his D, et al. Chromosome instability and immunodeficiency syndrome caused by mutations in a DNA methyltransferase gene. Natu r e 1999;402:187–191. Duvic M, T alpur R, Ni X, et al. Phase 2 trial of oral vorinostat (suberoylanilide hydroxamic acid, SAHA) for refractory cutaneous T -cell lymphoma (CTCL). Blood 2007;109:31–39. Olsen EA, Kim YH, Kuzel TM, et al. Phase IIb multicenter trial of vorinostat in patients with persistent, progressive, or treatment refractory cutaneous T -cell lymphoma. J Clin Oncol 2007;25:3109–3 1 15. O’Connor OA, Heaney ML, Schwartz L, et al. Clinical experience with intravenous and oral formulations of the novel histone deacetylase inhibitor suberoylanilide hydroxamic acid in patients with advanced hematologic malignancies. J Clin Oncol 2006;24:166–173. Ramalingam SS, Maitland ML, Frankel P , et al. Carboplatin and Paclitaxel in combination with either vorinostat or placebo for first-line therapy of advanced non-small-cell lung cance r . J Clin Oncol 2010;28:56–62. Giles F , Fischer T , Cortes J, et al. A phase I study of intravenous LBH589, a novel cinnamic hydroxamic acid analogue histone deacetylase inhibito r , in patients with refractory hematologic malignancies. Clin Cancer Res 2006;12:4628–4635. Richardson PG, Hungria VTM, Y oon S-S, et al. Panorama 1: A randomized, double-blind, phase 3 study of panobinostat or placebo plus bortezomib and dexamethasone in relapsed or relapsed and refractory multiple myeloma. ASCO Meeting Abstracts 2014;32:8510. Piekarz RL, Robey R, Sandor V , et al. Inhibitor of histone deacetylation, depsipeptide (FR901228), in the treatment of peripheral and cutaneous T -cell lymphoma: a case report. Blood 2001;98:2865–2868. Coi f fier B, Pro B, Prince HM, et al. Romidepsin for the treatment of relapsed/refractory peripheral T -cell lymphoma: pivotal study update demonstrates durable responses. J Hematol Oncol 2014;7: 1 1. Coi f fier B, Pro B, Prince HM, et al. Results from a pivotal, open-label, phase II study of romidepsin in relapsed or refractory peripheral T -cell lymphoma after prior systemic therap y . J Clin Oncol 2012;30:631–636. Piekarz RL, Frye AR, W right JJ, et al. Cardiac studies in patients treated with depsipeptide, FK228, in a phase II trial for T -cell lymphoma. Clin Cancer Res 2006;12:3762–3773. Gojo I, Jiemjit A, T repel JB, et al. Phase 1 and pharmacologic study of MS-275, a histone deacetylase inhibito r , in adults with refractory and relapsed acute leukemias. Blood 2007;109:2781–2790. Prebet T , Sun Z, Figueroa ME, et al. Prolonged administration of azacitidine with or without entinostat for myelodysplastic syndrome and acute myeloid leukemia with myelodysplasia-related changes: results of the US Leukemia Intergroup T rial E1905. J Clin Oncol 2014;32:1242–1248. W itta SE, Jotte RM, Konduri K, et al. Randomized phase II trial of erlotinib with and without entinostat in patients with advanced non-small-cell lung cancer who progressed on prior chemotherap y . J Clin Oncol 2012;30:2248–2255. Y ardley DA, Ismail-Khan RR, Melichar B, et al. Randomized phase II, double-blind, placebo-controlled study of exemestane with or without entinostat in postmenopausal women with locally recurrent or metastatic estrogen receptor-positive breast cancer progressing on treatment with a nonsteroidal aromatase inhibito r . J Clin Oncol 2013;31:2128–2135. Byrd JC, Marcucci G, Parthun MR, et al. A phase 1 and pharmacodynamic study of depsipeptide (FK228) in chronic lymphocytic leukemia and acute myeloid leukemia. Blood 2005;105:959–967. Charache S, Dover G, Smith K, et al. T reatment of sickle cell anemia with 5-azacytidine results in increased fetal hemoglobin production and is associated with nonrandom hypomethylation of DNA around the gamma-delta-beta-globin gene complex. P r oc Natl Acad Sci U S A 1983;80:4842–4846.

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20 1 1; 1 17:2697–2702. Craddock C, Quek L, Goardon N, et al. Azacitidine fails to eradicate leukemic stem/progenitor cell populations in patients with acute myeloid leukemia and myelodysplasia. Leukemia 2013;27:1028– 1036. Jue r gens RA, W rangle J, V endetti F P , et al. Combination epigenetic therapy has efficacy in patients with refractory advanced non-small cell lung cance r . Cancer Discov 20 1 1;1: 598–607. W rangle J, W ang W , Koch A, et al. Alterations of immune response of Non-Small Cell Lung Cancer with Azacytidine. Oncota r get 2013;4:2067–2079. Shakya R, Gonda T A, Quante M, et al. Hypomethylating therapy in an aggressive stroma-rich model of pancreatic carcinoma. Cancer Res 2013;73:885–896. Humeniuk R, Menon LG, Mishra PJ, et al. Decreased levels of UMP kinase as a mechanism of fluoropyrimidine resistance. Mol Cancer Ther 2009;8:1037–1044. Meng F , Sun G, Zhong M, et al. Anticancer e f ficacy of the combination of low-dose cisplatin and trichostatin A or 5-aza-2′-deoxycytidine in ovarian cancer cells. Paper presented at: 2012 ASCO Annual Meeting; 2012; Chicago, IL. Matei D, Fang F , Shen C, et al. Epigenetic resensitization to platinum in ovarian cance r . Cancer Res 2012;72:2197–2205. Fu S, Hu W , Iyer R, et al. Phase 1b-2a study to reverse platinum resistance through use of a hypomethylating agent, azacitidine, in patients with platinum-resistant or platinum-refractory epithelial ovarian cance r . Cancer 20 1 1; 1 17:1661–1669. Chung C W , Coste H, White JH, et al. Discovery and characterization of small molecule inhibitors of the BET family bromodomains. J Med Chem 20 1 1;54:3827–3838. Bernt KM, Zhu N, Sinha AU, et al. MLL-rearranged leukemia is dependent on aberrant H3K79 methylation by DOT1L. Cancer Cell 20 1 1;20:66–78. Daigle SR, Olhava EJ, Therkelsen CA, et al. Selective killing of mixed lineage leukemia cells by a potent small-molecule DOT1L inhibito r . Cancer Cell 20 1 1;20:53–65. Spannho f f A, Hauser A T , Heinke R, et al. The eme r ging therapeutic potential of histone methyltransferase and demethylase inhibitors. ChemMedChem 2009;4:1568–1582. Okada Y , Feng Q, Lin Y , et al. hDOT1L links histone methylation to leukemogenesis. Cell 2005;121:167–178.

BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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Th e ub i qu iti n p r o t easo m e syst em is i nvo l v e d i n t h e d e g ra d ati on o f m o re t h a n 80 % o f ce ll u l a r p r o t e i ns, i n cl ud i ng t ho se t h at c on tr o l cell-c y cle progr ess ion, apop t os i s, DNA r e p ai r , a nd t h e stress res pon se A k e y ste p in t h is process i s t he t agg i ng o f p r o tei n s ta r g ete d f o r d e g ra d ati on wit h m u lt iple c op ies o f ub i qu iti n, a 76–a mi no aci d p r o tei n w ho se p rimar y se qu e n ce a nd st ru ct ure i s h i gh l y conse r ved in o r g a n isms ra ng i ng fr o m y easts t o mammal s Once po l yub i qu iti n ate d, p r o tei n s ta r g ete d f o r d e g ra d ati on bind t o t h e 26 S p r o t easo m e, a ho l o e n z y me c o m po se d o f tw o 19 S re gu lat o r y c o m p le xes capp i ng a cen tr a l 2 0 S p r o te o l y tic c o re . T h e 20 S c o re is a ho ll ow “ b a rr el” cons i s ti ng o f f ou r s t a c k e d h e p tameric ri ng s . T h e s ubun its o f t h e r i ng s a re c l ass ifi ed as e it he r β s ubun its ( ou ter tw o ri ng s) o r β s ubun its (i nne r t wo r i ng s) . T he 19 S r egu l a t o r y c o m p le x c on sists o f a li d t h at rec ogn izes ub i qu itin a t ed p r o t e i n subs tr a tes wit h h i gh fi d elit y , a nd a b ase t h at c on tain s si x a d e nos i ne tri phospha t ases, un f o l d s p r o tei n s ub strates , rem ov es t h e po l yub i qu iti n t ag, and t h r eads t h em i n t o t h e catal y tic c h am b er o f t h e 20 S p a r ticle i n an adenos i ne tri pho s ph ate –d e p e nd e n t ma nn e r . U n li k e t yp ica l pro teases , t he 20 S p r o t easo m e i n e uk ar yo tic cells c on tai n s m u lti p le pro te o l yt i c ac ti v iti es r esu lti ng i n t h e clea v a g e o f p r o tei n ta r g ets after ma ny d i f f e r e n t a mi no ac i ds. I n m os t cells , t h e 20 S c o re p article c on tai n s t h e

BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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catal y tic subun it s β 5 (PS MB5 ) , β 1 (PSMB 1 ) , a nd β 2 (PSMB 2 ) , acc oun t ing for c hymo tr yps i n -li ke ( C T -L) , cas p aseli k e (C-L) , a nd tr yp si n li k e ( T -L) acti v ities , r espec ti ve l y , each d i f feri ng i n t h eir s ub strate p refere n ce .

BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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How e v e r , i n ce ll s o f he m a t opo ietic o ri g i n, s u c h as l y m pho c y tes a nd m ono c y t es, t he p r o t easo m e ca tal y tic s ubun its are e n c od e d by ho m o l ogous g e n e produc t s : L M P 7 (PS MB8 ) , LMP 2 (PSMB 9 ) , a nd MECL- 1 (PS MB 10 ) T hese imm unop r o teas o me s ubun its are als o i ndu ce d i n nonh em a t opo i e ti c ce ll s f o ll o wi ng e xpo s u re t o i n flammat o r y c y t ok i n es s uch as i n te rfe r on - γ (IF N - γ ) and t u m o r n ecr o sis fact o r al ph a (TNF- α ) . I n t h e imm unop r o t easo m e, t he 19 S re gu lat o r y c o m p le x ca n b e re p lace d wit h pro teas ome ac ti va t o r s such as P A 28, w ho se e xp ressi on is als o i ndu ce d i n cells fo l low i ng exposu r e t o IFN-γ . H yb ri d p r o teas o mes , bo t h f o r t h e catal y tic subun it s and r egu l a t o r y p articles , h a v e b ee n d escri b e d . G i v e n it s key r o l e i n m a i n tai n i ng cell u lar ho me o stasis , t h e ub i qu iti n pro teas ome sys t e m appea r ed t o b e a n un li k el y tar g et f o r ph armace u tical i n te rv e n ti on. Howeve r , a va riet y o f g r oundb rea k i ng st ud ies i n t h e 1990 s s ugg est ed t ha t i nh i b it o r s o f p r o teas o me f un cti on mi gh t p r ov e t o b e v ia b l e t h e r a p euti c agen t s . I n iti a l s t ud ies u se d s ub strate-relate d p e p ti d e al d e hydes t o i nv es t i ga t e t he p r o t eo l y ti c fun cti on s a nd s p ecificit y o f t h e p r o teas o me .

BIOC HE MI S T R Y O F T HE UBIQUITIN-PROTEASOME P A TH W A Y

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PROTEASOME INHIBI T ORS

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PROTEASOME INHIBI T ORS

PROTEASOME INHIBI T ORS

Chemical Classes of P r oteasome Inhibitors in Clinical Development

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A s of t he w riti ng o f t h i s ove r v ie w , si x d i f fere n t p r o teas o me i nh i b it o rs c o m pr isi ng t h r ee d i s ti nc t che mical classes h a v e b ee n teste d i n cli n ical tr ials ) and i nc l ude : ( 1 ) d i p e p ti d e bo r on ic aci d s , ( 2 ) p e p ti d e e poxy

PROTEASOME INHIBI T ORS

Chemical Classes of P r oteasome Inhibitors in Clinical Development

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k et on es , and ( 3 ) β-l ac t ones B o rtez o mi b (PS- 341, V elca d e) , a d i p e p tid e boron ic ac i d, was deve l oped by Mille nn i u m P h armace u ticals (Cam b ri dge, M A) a nd was t he fir s t PI app r ov e d f o r cli n ical u se . T w o a dd iti on al d i p e p ti de bo r on i c ac i ds have e n tere d cli n ical d e v el op me n t , i x az o mi b /ML N 9708 (M ill enn i u m) , cu rr en tl y i n ph ase III st ud ies , a nd d ela n z o mi b /CEP- 18770 ( T eva P ha rm aceu ti ca l s ; Fraze r , P A) , t h e cli n ical d e v el op me n t o f wh ic h h a s been ha lt ed. Ca rfil zo mi b (O nyx P h armace u ticals; Sa n Fra n cis co, C A), a tetr apep ti de epoxy ke to n e , recei v e d U . S . F ood a nd Dr ug Ad mi n i s tr a ti on (F DA ) app r o val i n 2012 . A sec ond p e p ti d e e poxy k et one pro teas ome i nh i b it o r , op r ozom i b (O nyx ) , e n tere d cli n ical st udy i n 2010. Th e t h i rd c l ass o f p r o t easo m e i nh i b it o rs , β-lact on es , is re p rese n te d by NPI- 0052 ( s a li nospo r a mi de A [ M ariz o mi b ]) a nd is c u rre n tl y b ei ng d e v el op e d by N e r e u s P ha rm aceu ti ca l s, I nc. ( Sa n Die go, CA) . T h e i n itial a pp r ov als f o r bo t h bor t ezo mi b and ca rfil zom i b were i n m u lti p le m y el o ma (MM) , a p lasma ce ll neop l as m and t he sec ond m o st c o mm on h emat o l og ic ca n ce r . How e v e r , t he ac ti v it y o f PI s i n o t h er B-cell n e op lasms h as res u lte d i n a n e xp a n si on o f t he c li n i ca l u tili z ati on o f t h is d r ug class .

PROTEASOME INHIBI T ORS

P r eclinical Activity of P r oteasome Inhibitors

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P r eclinical Activity of P r oteasome Inhibitors

PROTEASOME INHIBI T ORS

P r eclinical Activity of P r oteasome Inhibitors

nan nan

Simila r l y , pep ti de epoxy ke t o nes f o rm irre v ersi b le c ov ale n t a ddu cts wit h t h e activ e s it e t h r eon i ne bu t do s o v ia a du al c ov ale n t a ddu cti on o f γ-OH group a nd t he fr ee a mi ne . T h is i n teracti on is h i gh l y s p ecific f o r N-termi nal t hr e on i n e- con t a i n i ng hyd r o l a ses a nd re nd ers p e p ti d e e poxy k et on es t h e m o st sel ec ti ve p r o t easo m e i nh i b it o rs y et d escri b e d , Th e p rim a r y t a r ge t s o f t he se PIs wit h i n t h e c on stit u ti v e a nd imm unop r o t easo m es a r e t he C T -L s ubun its , β 5 a nd LMP 7, res p ecti v el y . D es p ite accoun ti ng f o r l ess t han 50 % o f t o tal p r o tei n t u r nov er by t h e pro teas ome, t hese subun it s a r e esse n tial f o r cell s u r v i v al . I n MM cell li nes, i nh i b iti n g bo t h subun it s (β 5 a n d LMP 7 ) is n ecessar y a nd s u fficie n t f o r t u m or c e ll dea t h . Cy t o t ox i c i ty o f o t h er t u m o r cell t yp es re qu ires t h e i nh i b iti o n o f m u lti p l e ac ti ve s ites b e yond t h e C T -L acti v it y . T h e c o m b i na ti on o f i nh i b it o r s spe cific f o r eit h er t h e T -L o r C-L acti v ities , w hich h a v e no c y t o t ox i c ac ti v it y on t h eir o w n, a ug me n ts t h e c y t o t ox ic po te n tia l o f t h e C T -L–spec ifi c i nh i b it o r s . G i v e n it s s t a t us as t he fir s t p r o teas o me i nh i b it o r a pp r ov e d f o r mar k ete d u se , t h e an tit u m o r po t en ti a l a n d p recli n ical acti v it y o f o t h er p r o teas o me i nh i b it ors have gene r a ll y been c o m p are d t o bo rtez o mi b . Carfilz o mi b s how e d equ i va l en t an tit u m o r a cti v it y t o bo rtez o mi b i n v itr o a g ai n st a p a nel of t u m o r ce ll li nes unde r s t and ar d c u lt u re c ond iti on s bu t was >10 -f o l d mo r e po te n t at i nduc i ng t u m o r ce ll d eat h w h e n cells were e xpo se d t o d r ug f o r a 1 - hour pu ls e, wh i ch mimi cs t he ph armac ok i n etics o f bo t h c o m pound s .

PROTEASOME INHIBI T ORS

P r eclinical Activity of P r oteasome Inhibitors

nan nan

M LN2238 (t he ac ti ve agen t o f i x az o mi b ) was acti v e i n t h e same m ou se m od els o f hu m an t u m o r s as bo rtez o mi b, bu t d em on strate d g reater le v els o f pro teas ome i nh i b iti on i n t he tum o rs . I n b i o c h emical assa y s o f p r o teas ome acti v it y , de l anzo mi b had an ide n tical po te n c y a nd s ubun it acti v it y p r o file to bor tez o mi b, bu t i n t u m o r cy t o t ox icit y assa y s , po te n c y relati v e t o bor tez o mi b was 2 - t o 10 -f o l d l ess . I n a dd iti on, d ela n z o mi b a pp eare d t o be

PROTEASOME INHIBI T ORS

P r eclinical Activity of P r oteasome Inhibitors

nan nan

less c y t o t ox i c t han bo rt ezo mi b t o no rmal cells a nd h a d a d i f fere n tial e f fe ct on c y t ok i ne r e l ease i n bone marr o w str o mal cells , s ugg esti ng a d i f fere n t ph a r ma co l og i c ac ti v it y . Op r o z o mi b is 10 -f o l d less po te n t t h a n carfilz o mi b i n pro te aso m e ac ti v it y assays, bu t s ho we d similar a n tit u m o r acti v it y i n m ou se t u m o r m ode l s , Ma riz o mi b d is p la y e d g reater po te n c y a g ai n st t he non– C T -L ac ti ve s it es o f t he p r o teas o me t h a n bo rtez o mi b . I n teresti ng l y , t h is a g e n t syne r g i zed w it h bo rtez o mi b i n k illi ng t u m o r cells i n v itr o . A ll o f t h e sec ond - gene r a ti on i nh i b it o rs h a v e s ho w n acti v it y i n t u m o r cells ma de r esista n t t o bo rt ezo mi b and / or MM cells is o late d fr o m p atie n ts rela p se d fro m bo rt ezo mi b - based t he r ap ie s Th e i nh i b iti on o f t u m o r ce lls wit h p r o teas o me i nh i b it o rs i ndu ces cell d eat h v i a t he i nduc ti on o f apo pt o sis t h r ough d eat h effect o r cas p ase acti v atio n . A lt hough t he m e c h a n ism und erl y i ng t h e i ndu cti on o f cell death r emai n s t o be f u ll y e l uc i da t ed, e x te n si v e researc h s ugg ests a c o m p le x i n te rp la y o f m u lti p l e pa t hways. PIs h a v e b ee n s ho w n t o affect t h e h alf-lif e of t h e BH3 - on l y m e m be r s o f t h e Bcl- 2 famil y , s p ecificall y BH 3– i n te r acti ng - do m a i n dea t h ago nist (Bi d ) a nd Bcl- 2 i n teracti ng k iller (Bi k ) .

PROTEASOME INHIBI T ORS

P r eclinical Activity of P r oteasome Inhibitors

nan nan

M or e ove r t he BH3 - on l y p r o tei n NOXA is up re gu late d at t h e tra n scri p ti on le v el by PI s Pr o t easo m e i nh i b iti on als o up re gu lates t h e e xp ressi on of se v e r al key ce ll- cyc l e checkpo i n t p r o tei n s t h at i n cl ud e p53 (a n i ndu cer o f G0 / G1 ce ll- cyc l e a rr es t t h r ou g h acc u m u lati on o f t h e c y cli n - d e p e nd e n t k i n ase [CDK ] i nh i b it o r p27 ); th e CDK i nh i b it o r p21 ; mammalia n c y cli ns A, B , D, a n d E; and tr ansc ri p ti on fact o rs E 2 F a nd R b . T h e tra n scri p ti on f act or nuc l ea r f ac t o r kappa B (NF- κ B) , a n im po rta n t re gu lat o r o f cell s urv i v al and cy t ok i ne / g r ow t h fact o r p r odu cti on is als o a f fecte d by pro teas ome i nh i b iti on i n m u lti p le wa y s . T h e n et effect on NF- κ B si gn ali ng is no t c ons i s t en t ac r oss va ri ou s assa y s a nd cell li n es , a nd its relati v e im por ta nce i n t he an tit u m o r e f fects o f PIs remai n s un clea r . Alt hough it i s i n te r estin g t o no t e t ha t pa ti en ts w ho se m y el o ma h ar bo r NF- κ B – acti v ati ng m u tati ons ( ~20 %) r espond be tter t o bo rtez o mi b t h a n t ho se wit hou t NF- κB– acti v atin g m u t a ti ons – I n MM cell li n es , t h ere is g r o wi ng e v i d e n ce t hat t h e maj o r de t e rmi nan t o f sens iti v it y t o p r o teas o me i nh i b iti on is t h e relati ve l o a d of p r o t e i n fl ux t o t he p r o teas o me . T h ese d ata s ugg est t h at i ndu ct ion

PROTEASOME INHIBI T ORS

Pharmacokinetics and Pharmacodynamics of Proteasome Inhibitors in Animals

nan nan

Pharmacokinetics and Pharmacodynamics of Proteasome Inhibitors in Animals

PROTEASOME INHIBI T ORS

Pharmacokinetics and Pharmacodynamics of Proteasome Inhibitors in Animals

nan nan

F o ll ow i ng i n tr avenous (I V ) ad mi n istrati on t o a n imals a nd hu ma n s , pro teas ome ac ti v it y i s i nh i b ite d i n a do se- d e p e nd e n t fas h i on wit h i n mi nutes; how e v e r , PI s such as bo rt ezom i b a nd carfilz o mi b are als o ra p i d l y cleare d fro m ci rcu l a ti on . Recov er y o f p r o teas o me acti v it y i n a n imals o cc u r s i n tiss u e s w it h a ha lf-lif e o f app r ox imatel y 24 hou rs , mirr o ri ng t h e rec ove r y time of ce ll s exposed t o sub l e t h al c on ce n trati on s o f PIs i n v itr o a nd li k el y r e f lecti ng new p r o t e i n syn t hes is .

PROTEASOME INHIBI T ORS IN CANC E R

Clinical Activity of Bortezomib

nan nan

i n wh ic h 202 pa ti en t s w it h he a v il y p retreate d d isease were treate d wit h bor tez o mi b a t 1.3 m g / m 2 . I n t h is trial , t h e ov erall res pon se rate (ORR) , d e f i n e d as pa ti en t s ach i ev i ng a t least a 50 % re du cti on i n ser u m o r u ri n e le v els o f t he m ye l o m a M p r o tei n, was 35 % . T h is cli n ical trial was s uppor t ed by t he CR EST tri a l, i n w h ic h t h e acti v it y o f 1.3 m g / m 2 do s e w as d ete r mi ned t o be supe ri o r t o a do se o f 1.0 m g / m 2 . B o rtez o mi b is als o acti v e a s a s i ng l e agen t i n ea rlier sta g e MM p atie n t popu lati on s . A si ng l e -a g e n t O R R o f 38 % , w it h a 6 % c o m p lete res pon se (CR) rate , was see n i n the ph ase III A PE X s t udy i n ea rl y rela p se d MM , wit h a time t o p r og ressi on (TT P ) of 6.2 m on t hs and a me d ia n du rati on o f res pon se o f 8 m on t h s . I n t h is st ud y , t he m a j o r g r ade 3 a nd 4 t ox icities were PN , 12 %; dy sest h esia and r elate d s y m p t o m s, 8 % t o 10 %; a n emia , 8 %; d iarr h ea , 8 %; n e u tr op e n ia , 14% ; a nd f a ti gue, 12 % . I n t h e fr on tli n e setti ng, bo rtez o mi b d em on strate d a si ng le - ag en t r esponse r a t e o f 41 % ( 5 % CR rate) B or t ezo mi b i s a l so app r ov e d f o r n ewl y d ia gno se d MM i n c o m b i n ati on w it h v el cade, m e l pha l an and p re dn is on e (VMP) . T h e ph ase III VIS T A tr ial e v al u ate d VM P i n pa ti en t s w it h un treate d MM w ho were i n eli g i b le f o r h i gh-dose t he r ap y T he add iti on o f bo rtez o mi b t o t h e mel ph ala n pr e dn is one ( M P) backbone s i gn ifica n tl y im p r ov e d res pon se rates i n t h is setti ng wit h an ORR o f 71 % f o r VMP (i n cl ud i ng 30 % CR) v ers u s 35 % (w it h only 4 % CR ) f o r M P . VMP was ass o ciate d wit h a TTP o f ~ 24 m on t h s , co m pa r ed w it h ~16.6 m on t h s wit h M P . After a 5 - y ear f o ll o w- up, t h e r e was a 31 % r educed ri sk o f d eat h f o r t h e VMP g r oup v ers u s MP-t r eate d pa ti en t s B or t ezo mi b has a l so shown p r o mise w h e n c o m b i n e d wit h o t h er a g e nts in r ela p se d and r e fr ac t o r y MM patie n ts . T h e c o m b i n ati on o f bo rtez o mi b w ith p e gy lated doxo r ub i c i n ( Dox il , Ce n t o c o r Ort ho Bi o tec h Pr odu cts , L . P . ; Hor s h a m, P A ) r esu lt ed i n an ORR o f 79 % i n rela p se d p atie n ts , a nd t ox icitie s we r e s imil a r t o t hos e ob ser v e d wit h eac h a g e n t a d mi n istere d se p a r atel y A phase III s t ud y i n 646 p atie n ts wit h rela p se d a nd refract ory MM c o m pa r ed t h i s tr ea tm en t w it h bo rtez o mi b al on e; t h e c o m b i n ati on produ ce d a 44 % ORR and ext e nd e d t h e TTP fr o m 6 t o 9.3 m on t h s . T he c o m b i na ti on o f bo rt ezo mi b wi t h re v limi d, le n ali do mi d e a nd d e x amet h as one

PROTEASOME INHIBI T ORS IN CANC E R

Clinical Activity of Bortezomib

nan nan

( R d), a s t anda r d o f ca r e i n t he treatme n t o f MM , res u lte d i n a n ORR o f 6 4 % a nd a me d i an du r a ti on o f r espon se o f 8.7 m on t h s . T h is acti v it y is stri k i ng g i v e n t ha t 53 % o f pa ti en t s had recei v e d p ri o r bo rtez o mi b a nd 75 % o f p atie n ts h ad r ece i ved p ri o r t h ali do mi d e , a cl o sel y relate d a n al og o f le n ali dom i de. O t he r agen t s t es te d i n c o m b i n ati on wit h bo rtez o mi b i n cl ude vor i no st a t , t he an ti- C S 1 m Ab, el o t u z u ma b, t h e Hs p90 i nh i b it o r ta n es p im yc i n, and t he Ak t i nh i b it o r p erif o si n e F ront li ne co m b i na ti ons w i th bo rtez o mi b i n MM p atie n ts h a v e s ho w n h i gh ORRs w it h a no t ab l e im p r ov eme n t i n CR rates . I n l ong er term st udies, CR r ate s w it h bo rt ezo mi b - bas e d c o m b i n ati on s h a v e b ee n s ho w n t o b e ass o ciate d w it h im p r oved c li n ical ou tc o mes . A c o mm un it y - b ase d ph a se IIIb st udy eva l ua ti ng bo rt ezom i b + d e x amet h as on e (VD) v ers u s bo rtez omib + t h ali do mi de + dexa m e t hason e (VTD) v ers u s VMP f ound similar ORR (60%, 7 0% , and 52 % , r espec t iv el y ) a nd CR rates ( 13 % , 18 % , a nd 15 % , r es p ecti ve l y ) . Bo rt ezo mi b + mel ph ala n + p re dn is on e + t h ali do mi d e (V MP T ) f o ll owed by bo rt ezom i b + t h ali do mi d e (VT) mai n te n a n ce res u l ted i n a s up eri o r CR r a t e co m pa r ed wit h VMP wit h no mai n te n a n ce ( 34 % v e r s u s 21 %) and im p r oved 2 - y ear p r og ressi on -free s u r v i v al ( 70 % v ers us 58.2%). A p r o t oco l m od ifi c ati on i n t h is trial i nvo l v e d c h a ng i ng fr o m tw ice w ee k l y t o week l y bo rt ezo mi b a d mi n istrati on, w h ic h y iel d e d similar TT P bu t r e duced t he i nc i dence ( 21 % v ers u s 43 %) a nd se v erit y o f PN ( 2 % g ra de 3 / 4 v e r s us 14 %) . T he bo rt ez omi b, le n ali do mi d e , a nd d e x amet h as on e c o m b i na ti on i n new l y d i agno se d MM res u lte d i n a ORR o f 100 % i n 66 p atie n ts , 29 % o f who m ach i ev e d a CR . B or t ezo mi b has a l so shown acti v it y i n o t h er h emat o l og ic ca n cers , mo st no ta b l y m an tl e ce ll l y m pho m a (MCL) . As a si ng le a g e n t i n 155 rela p s ed a nd r e frac t o r y MC L pa ti en t s, b o rtez o mi b y iel d e d a n ORR o f 33 % ( 8 % CR) , a me d ian du r a ti on o f r esponse o f 9.2 m on t h s , a nd a TTP o f 6.2 m on t h s .

PROTEASOME INHIBI T ORS IN CANC E R

Clinical Activity of Bortezomib

nan nan

T ox iciti es obse r ved we r e s imilar t o t ho se see n i n p atie n ts wit h MM a nd i n cl ud e d t h r o m bocy t open i a, PN , a nd fati gu e . W h e n bo rtez o mi b was u se d to t r eat bo th new l y d i agnosed a n d refract o r y MCL , a res pon se rate o f 46 % was ob se rv e d i n bo t h popu l a ti ons lea d i ng t o FDA a pp r ov al late i n 2006.

PROTEASOME INHIBI T ORS IN CANC E R

Clinical Activity of Bortezomib

nan nan

B or t ezo mi b has been t es t ed i n a v ariet y o f s o li d t u m o rs i n ph ase I a nd II st ud ies P a rti a l r esponses (PR) were re po rte d i n 8 % o f p atie n ts wit h r e fr act ory non–s m a ll- ce ll l ung ca n cer (NSCLC) , alt hough t h e TTP was 1.5 m on t h s . E xace r ba ti on o f P N was c o mm on. B o rtez o mi b was s ub se qu e n t ly teste d i n co m b i na ti on w it h pa clita x el , iri no teca n, a nd g emcitabi ne / ca r bop l a ti n ; how e v e r , res u lts h a v e no t b ee n e n c ou ra g i ng. B or tez o mi b con ti nues t o be t e ste d i n c o m b i n ati on wit h o t h er a g e n ts i n a v a r iet y o f t u m o r t ypes . Rece n t c li n i ca l ac ti v it y and p recli n ical d ata s ugg est t h at p r o teas o me i nh i b iti o n m ay ex t end t o nonon c o l ogy a pp licati on s . Si ng le-a g e n t bor tez o mi b t he r apy i n k i dney tra n s p la n t p atie n ts und er go i ng a n ti body -me d iated r e j ec ti on r esu lt ed i n a re du cti on o f dono r-s p ecific a n ti bod ies a nd im prov e d r ena l f unc ti on . I n m ou se m od els o f l upu s n e ph ritis , bo rtez omib r es u lte d i n a r educ ti on o f pa t hog e n ic p lasma cells a nd t h e p re v e n ti on o f d isease p r og r ess i on . T hese d ata s ugg est t h at PIs ma y b e u sef u l i n a wi de r a ng e of B - ce ll – m ed i a t ed d i se ases . H o we v e r , t ox icities wit h bo rtez o mi b, p a r tic u l a rl y P N, m ay p r even t w i d er a pp licati on o f t h is p artic u lar a g e n t .

PROTEASOME INHIBI T ORS IN CANC E R

Carfilzomib

nan nan

Carfilzomib

PROTEASOME INHIBI T ORS IN CANC E R

Carfilzomib

nan nan

Pa r allel phase I s t ud i es o f ca r f ilz o mi b h a v e b ee n c ondu cte d i n p atie n ts with m u lti p le t u m o r t ypes, and t wo ph ase I do se-fi nd i ng st ud ies tar g eti ng B-ce ll mali gn a nc i es have been co m p lete d. T h e first st udy u se d d ail y IV bo l u s do si ng wit h doses up t o 20 m g / m 2 f o r 5 c on sec u ti v e d a y s f o ll o we d by 9 d a y s of r es t and r esu lt ed i n sub sta n tial i nh i b iti on o f p r o teas o me acti v it y I n t h e sec ond s t ud y , ca rfil zo mi b was a d mi n istere d d ail y f o r 2 d a y s f o r 3 c on sec u ti ve weeks ( days 1, 2, 8, 9, 15, a nd 16 ) , f o ll o we d by 12 d a y s o f r ec ov e r y He m a t o l og i c t ox i c ities were t h e m o st fre qu e n t a dv erse e v e n t s, ob se rv e d a l ong w it h tr ans i en t, non c u m u lati v e ele v ati on s i n ser u m c r eati n i ne, usua ll y w it h i nc r e ases i n ser u m u rea n itr og e n a nd c on siste n t w ith a p r e r e na l e ti o l og y . New onse t PN was i n fre qu e n t . Am ong 20 e v al u a b le p atie n ts ( i nc l ud i ng bo rt ezo mi b -refract o r y p atie n ts) , 4 PRs a nd 1 mi no r r es pon s e we r e seen. Response s were als o du ra b le , lasti ng m o re t h a n 1 yea r

PROTEASOME INHIBI T ORS IN CANC E R

Carfilzomib

nan nan

i n s o me c ases. A lt hough t he ma x im u m t o lerate d do se o f carfilz o mi b wa s no t esta b li shed i n t h i s s t ud y , a do se o f 20 m g /m 2 was i n itiall y selecte d f o r t h e ph as e II s t ud i es. Based on t he phase I s t ud ies , a n op e n -la b el , si ng le-arm , ph ase II st udy o f si ng le - ag en t ca rfil zo mi b i n r e la p se d a nd refract o r y MM was i n itiate d i n 2007. Ca rfil zo mi b was ad mi n istere d as a n IV bo l u s on t h e twice-wee kly do se sch edu l e. P a ti en t s en r o lle d i n t h e i n itial ph ase o f t h e st udy ( 003 -A 0 ) h a d r ecei ved a m ed i an o f fi ve p ri o r t h era p ies , a nd 78 % o f p atie n ts h a d gr a d e 1 / 2 P N a t en tr y A m o n g 39 e v al u a b le p atie n ts i n 003 -A 0, 10 ( 26 %) ac h ie v e d a mi no r r esponse o r b ette r , i n cl ud i ng 5 PRs , a nd 16 a dd iti on al p atie n ts wit h s t ab l e d i sease. Base d on n ew safet y i n f o rmati on fr o m ph as e I st ud ies , t he p r o t oco l was a m end e d a nd t h e carfilz o mi b do se was escalat ed t o 27 m g /m 2 a ft e r t he fir s t cy cle ( 003 -A 1 ) . I n t h is trial , 266 p atie n ts we re e nro lle d and a ll pa ti en t s had p re v i ou sl y b ee n treate d wit h a n imm uno m odu l a t o r y agen t (IMiD) a nd bo rtez o mi b a nd were refract o r y t o t h ei r last t he r ap y . An ORR o f 24 % wit h a me d ia n du rati on o f res pon se of 8 m on t h s was r epo rt ed. Adve r s e e v e n ts were p re do mi n a n tl y h emat opo ieti c ( t hro m bocy t open i a, l y m phop e n ia , a nd a n emia) a nd t h ere was a <1 % rat e o f gr a d e 3 P N, desp it e 77 % hav i ng a h ist o r y o f PN . Base d on t h ese fi nd i ng s , ca rf ilz o mi b was g r an t ed cond iti on al a pp r ov al by t h e FDA i n 2012 f o r t he t r eatme n t o f pa ti en t s w it h r e la p se d a nd refract o r y m y el o ma w ho h a d r ecei v e d p ri o r bo rt ezo mi b and IMiD t h era p y . Th e pa r a ll e l P X - 171 - 004 trial e n r o lle d p atie n ts wit h rela p se d MM fo ll ow i ng one t o t h r ee p ri o r tr e atme n ts a nd w ho ma y h a v e b ee n refract o r y t o on e o r m o r e o f t hese t he r ap ies . Of t h e 155 p atie n ts e n r o lle d i n t h is t r ial , 120 had no t r ece i ved p rior bo rtez o mi b - b ase d t h era p y . I n p atie n ts w ith r ela p se d d i sease, nonhe m a t o l og ic a nd h emat o l og ic t ox icit y p r o files wer e simila r . Desp it e h i gh r a t es o f b aseli n e PN , re po rts o f w o rse n i ng n e u r op a thic s y m p t o m s we r e i n fr equen t ( 2 % i n ci d e n ce o f g ra d e 3 a nd no g ra d e 4 e v e nts ) . Ca rf ilz o mi b de m ons tr a t ed con si d era b le acti v it y i n bo rtez o mi b - n aï v e p atie n ts , i nduc i ng P R o r be tt e r i n 46 % o f 54 e v al u a b le p atie n ts at 20 m g / m 2 a nd 53 % o f pa ti en t s a t 27 m g/ m 2 . T h e res pon se rate i n p atie n ts p re v i ously

PROTEASOME INHIBI T ORS IN CANC E R

Ixazomib

nan nan

In itial cli n i ca l s t ud i es o f i xazo mi b i nvo l v e d do se escalati on st ud ies i n p atie n ts wit h he m a t o l og i c m al i gn a n cies a nd e xp l o re d bo t h wee k l y a nd t w ice w e ek l y dos i ng schedu l e s . Oral a d mi n istrati on res u lte d i n po te n t pro teas ome i nh i b iti on o f ~65 % . Cli n ical acti v it y i n p atie n ts wit h rela p se d MM w a s 16 % . I n pa ti en t s wi t h n ewl y d ia gno se d MM , i x az o mi b p l u s le n ali dom i de and l ow - dose d e x amet h as on e res u lte d i n a n ORR o f 93 % with 24% ac h i ev i ng a CR . T h i s c om b i n ati on is als o b ei ng i nv esti g ate d i n a ph ase III tri a l co m pa ri ng t h i s t o R d i n p atie n ts wit h rela p se d MM .

PROTEASOME INHIBI T ORS IN CANC E R

Op r ozomib

nan nan

In itial cli n i ca l t es ti ng o f op r o z o mi b i n p atie n ts wit h s o li d t u m o rs i nv esti g at ed a dos i ng schedu le c on sisti ng o f a 14 - d a y c y cle wit h on ce d a ily a d mi n is t r a ti on f o r 5 consecu t iv e d a y s . I n p atie n ts wit h rela p se d a nd / o r r e fr act ory B - ce ll neop l as m s, tw o do si ng sc h e du les are b ei ng u tilize d : t he sc h e du l e desc ri bed p r ev i ous l y a nd on e i nvo l v i ng 2 c on sec u ti v e d a y s o f do si ng repea t ed week l y Pr o teas o me i nh i b iti on f o ll o wi ng t h e a d mi n is t r a ti on o f op r ozo mi b reac h e d >80 % a nd cli n ical acti v it y was no t ed i n p atie n t s w it h MM and W M . I n p atie n ts recei v i ng t h e 5 c on sec u ti v e d a y sc h e du l e, 5 o f 19 MM pa ti en t s ( 26 %) a nd 8 o f 10 WM p atie n ts ( 80 %) ac h ie v e d a pa rti a l r esponse o r b ette r . E xp l o rati on o f t h e do se a nd sc h e du l e c on ti nues as a s i ng l e agen t and i n c o m b i n ati on wit h o t h er a n ti-MM t h e r a p ies .

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

nan nan

t h a n sta nda r d s t ag i ng sys t e m s . H o we v e r , t h is si gn at u re p r ov i d e d on l y a m od est inc r ease i n p r ed i c ti ve p o wer f o r treatme n t wit h bo rtez o mi b v ersu s d e x ameth asone. Mo r e r ecen tl y , Keats et al . rea n al y ze d t h is d ataset b as ed on a p at hway ana l ys i s o f N F- κ B a nd t h e realizati on t h at TRAF 3, a k e y r e gu lat o r y o f t he noncanon i c al NF- κ B p at h wa y , is a t u m o r s upp ress o r i n MM cel l li nes. T hey f ound a d ramatic e n ric h me n t f o r res pon se t o bor tez o mi b i n pa ti en t s w it h l o w le v els o f TRAF 3 e xp ressi on. H o we v e r , t h ese d at a r e m a i n t o be va li dat e d i n a se p arate sam p le set . A tra n scri p t omic a n al y sis o f sa m p l es de ri ved fr o m si ng le-a g e n t carfilz o mi b trials s ugg est that p atie n ts wit h t he h i ghes t l eve l o f imm unog l obu li n h ea vy c h ai n e xp ressi on w e r e t he m os t sens iti ve t o ca rfilz o mi b t h era p y Similar fi nd i ng s were no te d i n t he exp r ess i on da t a f ro m bo rtez o mi b -treate d p atie n ts d escri b e d pr e v i ous l y . T hese da t a a r e s uppo rte d by ph e no t yp ic d ata fr o m p atie n ts progr ess ing on bo rt ezo mi b - b ase d t h era p y , i n w h ic h resista n ce t o bo rtez omib w as ass oc i a t ed w it h a ded i f f er e n tiate d (a nd l o wer imm unog l obu li n e xpr essi ng ) B - ce ll pheno t ype . T a k e n t og et h e r , t h ese fi nd i ng s s ugg est t hat b i o ma rk er s, po t en ti a ll y t hose i nvo l v i ng a n a n al y sis o f p r o tei n l o a d o f imm unog l obu li n exp r ess i on, m a y b e d e v el op e d t o p re d ict t ho se p atie n ts m o st li ke l y t o r espond t o PI s.

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

nan nan

Ciechanover A. Intracellular protein degradation: from a vague idea thru the lysosome and the ubiquitin-proteasome system and onto human diseases and drug ta r geting. Biochim Biophys Acta 2012;1824:3–13. Kopp F , Hendil KB, Dahlmann B, et al. Subunit arrangement in the human 20S proteasome. P r oc Natl Acad Sci U S A 1997;94:2939–2944. W ilkinson KD. Ubiquitination and deubiquitination: ta r geting of proteins for degradation by the proteasome. Semin Cell Dev Biol 2000; 1 1:141–148. Braun BC, Glickman M, Kraft R, et al. The base of the proteasome regulatory particle exhibits chaperone-like activit y . Nat Cell Biol 1999;1:221–226. Groll M, Ditzel L, Lowe J, et al. Structure of 20S proteasome from yeast at 2.4 A resolution. Natu r e 1997;386:463–471. Borissenko L, Groll M. 20S proteasome and its inhibitors: crystallographic knowledge for drug development. Chem Rev 2007;107:687–717. Kloetzel PM, Ossendorp F . Proteasome and peptidase function in MHC-class-I-mediated antigen presentation. Curr Opin Immunol 2004;16:76–81.

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

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Gri f fin T A, Nandi D, Cruz M, et al. Immunoproteasome assembly: cooperative incorporation of interferon gamma (IFN-gamma)-inducible subunits. J Exp Med 1998;187:97–104. T anahashi N, Murakami Y , Minami Y , et al. Hybrid proteasomes. Induction by interferon-gamma and contribution to A TP-dependent proteolysis. J Biol Chem 2000;275:14336–14345. Adams J. The proteasome: a suitable antineoplastic ta r get. Nat Rev Cancer 2004;4:349–360. V initsky A, Michaud C, Powers JC, et al. Inhibition of the chymotrypsin-like activity of the pituitary multicatalytic proteinase complex. Biochemistry 1992;31:9421–9428. Orlowski RZ, Eswara JR, Lafond- W alker A, et al. T umor growth inhibition induced in a murine model of human Burkitt ’ s lymphoma by a proteasome inhibito r . Cancer Res 1998;58:4342–4348. Imajoh-Ohmi S, Kawaguchi T , Sugiyama S, et al. Lactacystin, a specific inhibitor of the proteasome, induces apoptosis in human monoblast U937 cells. Biochem Biophys Res Commun 1995;217:1070– 1077. Shinohara K, T omioka M, Nakano H, et al. Apoptosis induction resulting from proteasome inhibition. Biochem J 1996;317:385–388. Delic J, Masdehors P , Omura S, et al. The proteasome inhibitor lactacystin induces apoptosis and sensitizes chemo- and radioresistant human chronic lymphocytic leukaemia lymphocytes to TNF-alpha-initiated apoptosis. Br J Cancer 1998;77: 1 103– 1 107. Meng L, Mohan R, Kwok BH, et al. Epoxomicin, a potent and selective proteasome inhibito r , exhibits in vivo antiinflammatory activit y . P r oc Natl Acad Sci U S A 1999;96:10403–10408. Meng L, Kwok BH, Sin N, et al. Eponemycin exerts its antitumor e f fect through the inhibition of proteasome function. Cancer Res 1999;59:2798–2801. Dick LR, Fleming PE. Building on bortezomib: second-generation proteasome inhibitors as anti-cancer therap y . Drug Discov T oday 2010;15:243–249. Kirk CJ. Discovery and development of second-generation proteasome inhibitors. Semin Hematol 2012;49:207–214. Bross P F , Kane R, Farrell A T , et al. Approval summary for bortezomib for injection in the treatment of multiple myeloma. Clin Cancer Res 2004;10:3954–3964. Herndon TM, Deisseroth A, Kaminskas E, et al. U.S. Food and Drug Administration approval: carfilzomib for the treatment of multiple myeloma. Clin Cancer Res 2013;19:4559–4563. Bennett MK, Kirk CJ. Development of proteasome inhibitors in oncology and autoimmune diseases. Curr Opin Drug Discov Devel 2008; 1 1:616–625. Adams J, Behnke M, Chen S, et al. Potent and selective inhibitors of the proteasome: dipeptidyl boronic acids. Bioo r g Med Chem Lett 1998;8:333–338. Groll M, Berkers CR, Ploegh HL, et al. Crystal structure of the boronic acid-based proteasome inhibitor bortezomib in complex with the yeast 20S proteasome. Structu r e 2006;14:451–456. Groll M, Huber R, Potts BC. Crystal structures of Salinosporamide A (NPI-0052) and B (NPI-0047) in complex with the 20S proteasome reveal important consequences of beta-lactone ring opening and a mechanism for irreversible binding. J Am Chem Soc 2006;19:5136–5141. Groll M, Kim KB, Kairies N, et al. Crystal structure of epoxomicin: 20S proteasome reveals a molecular basis for selectivity of a’ b’-epoxyketone proteasome inhibitors. J Am Chem Soc 2000;122:1237–1238. Kisselev A F , van der Linden W A, Overkleeft HS. Proteasome inhibitors: an expanding army attacking a unique ta r get. Chem Biol 2012;19:99– 1 15. Arastu-Kapur S, Anderl JL, Kraus M, et al. Nonproteasomal targets of the proteasome inhibitors bortezomib and carfilzomib: a link to clinical adverse events. Clin Cancer Res 20 1 1;17:2734–2743.

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

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Kisselev A F , Callard A, Goldbe r g AL. Importance of the di f ferent proteolytic sites of the proteasome and the e f ficacy of inhibitors varies with the protein substrate. J Biol Chem 2006;281:8582–8590. Parlati F , Lee SJ, Aujay M, et al. Carfilzomib can induce tumor cell death through selective inhibition of the chymotrypsin-like activity of the proteasome. Blood 2009; 1 14:3439–3447. Britton M, Lucas MM, Downey SL, et al. Selective inhibitor of proteasome ’ s caspase-like sites sensitizes cells to specific inhibition of chymotrypsin-like sites. Chem Biol 2009;16:1278–1289. Mirabella AC, Pletnev AA, Downey SL, et al. Specific cell-permeable inhibitor of proteasome trypsin-like sites selectively sensitizes myeloma cells to bortezomib and carfilzomib. Chem Biol 20 1 1;18:608–618. Demo SD, Kirk CJ, Aujay MA, et al. Antitumor activity of PR-171, a novel irreversible inhibitor of the proteasome. Cancer Res 2007;67:6383–6391. Kupperman E, Lee EC, Cao Y , et al. Evaluation of the proteasome inhibitor MLN9708 in preclinical models of human cance r . Cancer Res 2010;70:1970–1980. Piva R, Ruggeri B, W illiams M, et al. CEP-18770: A novel, orally active proteasome inhibitor with a tumo r -selective pharmacologic profile competitive with bortezomib. Blood 2008; 1 1 1:2765–2775. Chauhan D, Singh A V , Aujay M, et al. A novel orally active proteasome inhibitor ONX 0912 triggers in vitro and in vivo cytotoxicity in multiple myeloma. Blood 2010; 1 16:4906–4915. Zhou HJ, Aujay MA, Bennett MK, et al. Design and synthesis of an orally bioavailable and selective peptide epoxyketone proteasome inhibitor (PR-047). J Med Chem 2009;52:3028–3038. Chauhan D, Catley L, Li G, et al. A novel orally active proteasome inhibitor induces apoptosis in multiple myeloma cells with mechanisms distinct from Bortezomib. Cancer Cell 2005;8:407–419. Chauhan D, Singh A, Brahmandam M, et al. Combination of proteasome inhibitors bortezomib and NPI-0052 trigger in vivo syne r gistic cytotoxicity in multiple myeloma. Blood 2008; 1 1 1:1654–1664. Chauhan D, T ian Z, Zhou B, et al. In vitro and in vivo selective antitumor activity of a novel orally bioavailable proteasome inhibitor MLN9708 against multiple myeloma cells. Clin Cancer Res 20 1 1;17:53 1 1–5321. Kuhn DJ, Chen Q, V oorhees PM, et al. Potent activity of carfilzomib, a novel, irreversible inhibitor of the ubiquitin-proteasome pathwa y , against preclinical models of multiple myeloma. Blood 2007; 1 10:3281–3290. Suzuki E, Demo S, Deu E, et al. Molecular mechanisms of bortezomib resistant adenocarcinoma cells. PLoS One 20 1 1;6:e27996. Zhang HG, W ang J, Y ang X, et al. Regulation of apoptosis proteins in cancer cells by ubiquitin. Oncogene 2004;23:2009–2015. Fernandez Y , V erhaegen M, Miller T P , et al. Di f ferential regulation of noxa in normal melanocytes and melanoma cells by proteasome inhibition: therapeutic implications. Cancer Res 2005;65:6294– 6304. Nikiforov MA, Riblett M, T ang WH, et al. T umor cell-selective regulation of NOXA by c-MYC in response to proteasome inhibition. P r oc Natl Acad Sci U S A 2007;104:19488–19493. Qin JZ, Zi f fra J, Stennett L, et al. Proteasome inhibitors trigger NOXA-mediated apoptosis in melanoma and myeloma cells. Cancer Res 2005;65:6282–6293. W ang Q, Mora-Jensen H, W eniger MA, et al. ERAD inhibitors integrate ER stress with an epigenetic mechanism to activate BH3-only protein NOXA in cancer cells. P r oc Natl Acad Sci U S A 2009;106:2200–2205. Mannava S, Zhuang D, Nair JR, et al. KLF9 is a novel transcriptional regulator of bortezomib- and LBH589-induced apoptosis in multiple myeloma cells. Blood 2012; 1 19:1450–1458.

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

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Koepp DM, Harper J W , Elledge SJ. How the cyclin became a cyclin: regulated proteolysis in the cell cycle. Cell 1999;97:431–434. Pagano M, T am S W , Theodoras AM, et al. Role of the ubiquitin-proteasome pathway in regulating abundance of the cyclin-dependent kinase inhibitor p27. Science 1995;269:682–685. W an F , Lenardo MJ. The nuclear signaling of NF-kappaB: current knowledge, new insights, and future perspectives. Cell Res 2010;20:24–33. Annunziata CM, Davis RE, Demchenko Y , et al. Frequent engagement of the classical and alternative NF-kappaB pathways by diverse genetic abnormalities in multiple myeloma. Cancer Cell 2007;12: 1 15–130. Chapman MA, Lawrence MS, Keats JJ, et al. Initial genome sequencing and analysis of multiple myeloma. Natu r e 20 1 1;471:467–472. Keats JJ, Fonseca R, Chesi M, et al. Promiscuous mutations activate the noncanonical NF-kappaB pathway in multiple myeloma. Cancer Cell 2007;12:131–144. Meister S, Schubert U, Neubert K, et al. Extensive immunoglobulin production sensitizes myeloma cells for proteasome inhibition. Cancer Res 2007;67:1783–1792. Obeng EA, Carlson LM, Gutman DM, et al. Proteasome inhibitors induce a terminal unfolded protein response in multiple myeloma cells. Blood 2006;107:4907–4916. Shabaneh TB, Downey SL, Goddard AL, et al. Molecular basis of differential sensitivity of myeloma cells to clinically relevant bolus treatment with bortezomib. PLoS One 2013;8:e56132. Papadopoulos K P , Burris HA III, Gordon M, et al. A phase I/II study of carfilzomib 2-10-min infusion in patients with advanced solid tumors. Cancer Chemother Pharmacol 2013;72:861–868. Papandreou CN, Daliani DD, Nix D, et al. Phase I trial of the proteasome inhibitor bortezomib in patients with advanced solid tumors with observations in androgen-independent prostate cance r . J Clin Oncol 2004;22:2108–2121. W ang Z, Y ang J, Kirk C, et al. Clinical pharmacokinetics, metabolism, and drug-drug interaction of carfilzomib. Drug Metab Dispos 2013;41:230–237. Y ang J, W ang Z, Fang Y , et al. Pharmacokinetics, pharmacodynamics, metabolism, distribution, and excretion of carfilzomib in rats. Drug Metab Dispos 20 1 1;39:1873–1882. Meiners S, Heyken D, W eller A, et al. Inhibition of proteasome activity induces concerted expression of proteasome genes and de novo formation of mammalian proteasomes. J Biol Chem 2003;278:21517–21525. Lonial S, W aller EK, Richardson PG, et al. Risk factors and kinetics of thrombocytopenia associated with bortezomib for relapsed, refractory multiple myeloma. Blood 2005;106:3777–3784. Richardson PG, Briembe r g H, Jagannath S, et al. Frequenc y , characteristics, and reversibility of peripheral neuropathy during treatment of advanced multiple myeloma with bortezomib. J Clin Oncol 2006;24:3 1 13–3120. Richardson PG, Barlogie B, Berenson J, et al. A phase 2 study of bortezomib in relapsed, refractory myeloma. N Engl J Med 2003;348:2609–2617. Jagannath S, Barlogie B, Berenson J, et al. A phase 2 study of two doses of bortezomib in relapsed or refractory myeloma. Br J Haematol 2004;127:165–172. Richardson PG, Sonneveld P , Schuster M W , et al. Bortezomib or high-dose dexamethasone for relapsed multiple myeloma. N Engl J Med 2005;352:2487–2498. Jagannath S, Brian D, W olf JL, et al. A phase 2 study of bortezomib as first-line therapy in patients with multiple myeloma. Blood 2004;104:333.

PROTEASOME INHIBI T ORS IN CANC E R

Biomarkers for P r oteasome Inhibitors

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myeloma or lymphoma. Clin Cancer Res 2012;18:4830–4840. Jagannath S, V ij R, Stewart AK, et al. An open-label single-arm pilot phase II study (PX-171-003-A0) of low-dose, single-agent carfilzomib in patients with relapsed and refractory multiple myeloma. Clin L ymphoma Myeloma Leuk 2012;12:310–318. Siegel DS, Martin T , W ang M, et al. A phase 2 study of single-agent carfilzomib (PX-171-003-A1) in patients with relapsed and refractory multiple myeloma. Blood 2012;120:2817–2825. Siegel DS, Martin T , W ang M, et al. Results of PX-171-003-A1, an open-label, single-arm, phase 2 (Ph 2) study of carfilzomib (CFZ) in patients (pts) with relapsed and refractory multiple myeloma (MM). Blood 2012;120:2817–2825. V ij R, Siegel DS, Jagannath S, et al. An open-label, single-arm, phase 2 study of single-agent carfilzomib in patients with relapsed and/or refractory multiple myeloma who have been previously treated with bortezomib. Br J Haematol 2012;158:739–748. V ij R, W ang M, Kaufman JL, et al. An open-label, single-arm, phase 2 (PX-171-004) study of single-agent carfilzomib in bortezomib-naive patients with relapsed and/or refractory multiple myeloma. Blood 2012; 1 19:5661–5670. Papadopoulos K, Capua Siegel DS, Singhal SB, et al. Phase 1b evaluation of the safety and e f ficacy of a 30-minute IV infusion of carfilzomib in patients with relapsed and/or refractory multiple myeloma. Blood 2010; 1 16:3024. Lee SJ, Levitsky K, Parlati F , et al. Clinical activity of carfilzomib correlates with inhibition of multiple proteasome subunits: application of a novel pharmacodynamics assa y . (In press). W ang M, Martin T , Bensinger W , et al. Phase 2 dose-expansion study (PX-171-006) of carfilzomib, lenalidomide, and low-dose dexamethasone in relapsed or progressive multiple myeloma. Blood 2013;122:3122–3128. Jakubowiak AJ, Dytfeld D, Gri f fith KA, et al. A phase 1/2 study of carfilzomib in combination with lenalidomide and low-dose dexamethasone as a frontline treatment for multiple myeloma. Blood 2012;120:1801–1809. Richardson PG, Baz R, W ang L, et al. Investigational agent MLN9708, an oral proteasome inhibito r , in patients (Pts) with relapsed and/or refractory multiple myeloma (MM): results from the expansion cohorts of a phase 1 dose-escalation stud y . Blood 20 1 1; 1 18:301. Richardson PG, Spencer A, Cannell P , et al. Phase 1 clinical evaluation of twice-weekly marizomib (NPI-0052), a novel proteasome inhibito r , in patients with relapsed/refractory multiple myeloma (MM). Blood 20 1 1; 1 18:302. Roy V , Reeder C, LaPlant BR, et al. Phase 2 trial Of single agent MLN9708 in patients with relapsed multiple myeloma not refractory to bortezomib. Blood 2013;122:1944. Hofmeister CC, Rosenbaum CA, Htut M, et al. T wice-weekly oral MLN9708 (ixazomib citrate), an investigational proteasome inhibito r , in combination with lenalidomide (len) and dexamethasone (dex) in patients (pts) with newly diagnosed multiple myeloma (MM): final phase 1 results and phase 2 data. Blood 2013;122:535. Papadopoulos K P , Mendelson DS, T olcher A W , et al. A phase I, open-label, dose-escalation study of the novel oral proteasome inhibitor (PI) ONX 0912 in patients with advanced refractory or recurrent solid tumors. Blood 20 1 1;29:3075. Kaufman JL, Siegel DS, V ij R, et al. Clinical profile of single-agent modified-release oprozomib tablets in patients (pts) with hematologic malignancies: updated results from a multicenter, open-label, dose escalation phase 1b/2 stud y . Blood 2013;122:3184.

IN T RODUCTION

nan nan

Ca n ce r ce ll s m ay ha r bo r de f e cts i n DNA re p air p at h wa y s lea d i ng t o g e no mi c i ns t ab ilit y . T h i s can f o ster t u m o ri g e n esis bu t als o p r ov i d es a w ea kn e ss t ha t can be exp l o it ed t h era p e u ticall y . T u m o rs wit h c o m p r o mis ed a b ilit y t o r epa ir doub l e - s tr and DNA b rea k s by ho m o l ogou s rec o m b i n ati on, i n cl ud i ng t hose w it h de f ec t s i n t h e BRCA 1 a nd BRCA 2 g e n es , are h i gh l y se n siti ve t o b l ockade o f t he re p air o f DNA si ng le-stra nd b rea k s , v ia t h e i nh i b iti o n o f t he enzy m e po l y( ADP-ri bo se) ( P ARP) . T h is p r ov i d es t h e basis for a sy n t he ti c l e t ha l app r oac h t o ca n cer t h era p y , w h ic h is s ho wi ng c on si d e rab l e p r o mi se i n t he c li n ic .

CELLU L AR DNA R E P AIR P A T H W A YS

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CELLU L AR DNA R E P AIR P A T H W A YS

CELLU L AR DNA R E P AIR P A T H W A YS

nan nan

DNA is con ti nua ll y da m aged by e nv ir on me n tal e xpo s u res a nd e ndog e n o us acti v ities , such as DNA r ep li c ati on a nd cell u lar free-ra d ical g e n erati on, wh ic h c ause d i ve r se l es i ons i n cl ud i ng b ase m od ificati on s , doub le-stra nd br ea k s ( DS B ) , s i ng l e - s tr and b rea k s (SSB) , a nd i n trastra nd a nd i n terstra nd c ro ss - lin ks T hese abe rr a ti ons are re p aire d by d isti n ct re p air p at h wa y s , wh ic h ar e coo r d i na t ed t o m a i n tai n t h e sta b ilit y a nd i n te g rit y o f t h e g e nome. Th is f aithf u l r epa ir o f DNA d ama g e is a n esse n tial p rere qu isite f o r t h e mai n te nance o f geno mi c i n t eg rit y a nd cell u lar a nd o r g a n ismal v ia b ilit y .

CELLU L AR DNA R E P AIR P A T H W A YS

nan nan

j o i n i ng of p r ev i ous l y un li nked DNA m o lec u les , po te n tiall y res u lti ng i n gro ss c h r o m oso m a l r ea rr ange me n ts s u c h as tra n sl o cati on s GC u ses a ho m o l ogous sequence, p r e f e r ab l y t h e sister c h r o mati d, as a tem p late t o r es yn t h e s i ze t he DNA su rr ound i ng t h e DSB , a nd t h eref o re g e n erall y resu lts i n acc ur at e r epa ir o f t he b r ea k . Re p air by GC is criticall y d e p e nd e n t on t he r ec o m b i nase f unc ti on o f RA D 51 a nd is facilitate d by a nu m b er o f o t h er pro tei n s . SS A a l so i nvo l ves t h e u se o f ho m o l ogou s se qu e n ces f o r t h e re pai r of D SB s, bu t un li ke GC, SS A is RAD 51 -i nd e p e nd e n t a nd i nvo l v es t h e a nn eali n g o f DNA s tr ands f o r m e d after resecti on at t h e DSB . T h e d etaile d mec h a n ism o f SS A i s s till ob sc u re bu t it fre qu e n tl y res u lts i n t h e l o ss o f one of t h e ho m o l ogous sequences a nd d eleti on o f t h e i n ter v e n i ng se qu e n ce

CELLU L AR DNA R E P AIR P A T H W A YS

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CELLU L AR DNA R E P AIR P A T H W A YS

nan nan

SS A is a po t en ti a ll y im po rt an t p at h wa y o f m u ta g e n esis b eca u se a si gn i f ic an t fr ac ti on o f m a mm a lia n g e no mes c on sist o f re p etiti v e eleme n ts . G C a nd S S A a r e ce ll- cyc l e r egu late d a nd are m o st acti v e i n S- G 2 ph ases o f t h e cell c yc l e .

T H E D EVE LOP MENT OF P ARP I N HI B I T ORS

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T H E D EVE LOP MENT OF P ARP I N HI B I T ORS

T H E D EVE LOP MENT OF P ARP I N HI B I T ORS

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T H E D EVE LOP MENT OF P ARP I N HI B I T ORS

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S ub se quen t s t ud i es w it h m o re po te n t P ARP i nh i b it o rs d em on strate d s yn e r gy w it h tem ozo l o mi de, an obse r v ati on t h at was ta k e n i n t o a cli n ical trial wi th AG014699, a P AR P i nh i b itor d e v el op e d by Pfize r . T h is a g e n t is no w b ei ng deve l oped by C l ov i s. A lt hough t h e maj o r f o c u s o f t h is c h a p ter is t he u se of P AR P i nh i b it o r s i n synth etic let h al t h era p e u tic strate g ies , t h eir u s e in c h em opo t en ti a ti on i n co m b i na ti on wit h c h em o t h era py remai n s und er act ive i nv esti g ati on, as desc ri bed l a t e r .

B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

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B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

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H ete ro z ygous ge rmli ne m u t a t ion s i n t h e BRCA 1 a nd BRCA 2 g e n es c on f e r a h i gh r is k o f b r eas t ( up t o 85 % lifetime ris k ) a nd ov aria n ( 10 % t o 40 %) ca n ce r i n add iti on t o a s i gn ifi c a n tl y i n crease d ris k o f p a n creatic , p r o state , a nd mal e b r eas t cance r T he g e n es h a v e b ee n classifie d as t u m o r s uppr es so r s, because t he w il d -t yp e BRCA allele is fre qu e n tl y l o st i n t u m o r s, a ph e nomenon t ha t occu r s by a v ariet y o f mec h a n isms . T h e BRCA 1 a nd BR C A2 genes encode l a r ge p r o tei n s t h at li k el y f un cti on i n m u lti p le cell ula r p at hw a y s , i nc l ud i ng tr ansc ri pt i on, cell-c y cle re gu lati on, a nd t h e mai n te nance o f geno m e i n t eg rit y . H o we v e r , t h e r o les o f BRCA 1 a nd BRC A2 i n DNA r epa ir have b ee n b est do c u me n te d . BRC A1 - and BRCA2 - de ficie n t cells are h i gh l y se n siti v e t o i on izi ng r a d iati on and d i sp l ay ch r o m o s o mal i n sta b ilit y , w h ic h is li k el y t o b e a d ire ct c on se quence o f un r epa ir ed DN A d ama g e . T h e similar g e no mic i n sta b il ity i n BRC A1 - and BRCA2 - de fi c ie n t cells a nd t h e i n teracti on o f bo t h BRC A1 a nd BR CA2 w it h RAD51 sugg este d a f un cti on al li nk b etwee n t h e t h ree pro tei n s i n t he RAD51 -m ed i a te d DNA d ama g e re p air p r o cess . H o we v e r , alt hough BRCA2 i s d ir ec tl y in vo l v e d i n RAD 51 -me d iate d re p ai r , a f fecti ng t h e c ho i ce be t ween GC and SSA , BRCA 1 acts up stream o f t h ese p at hw a y bo t h GC and SSA are re du ce d i n BRCA 1 - d eficie n t cells , p laci ng BRCA1 be f o r e t he b ra n c h po i n t o f GC a nd SSA . BRC A1 has a r o l e i n s i gn ali ng DNA d ama g e a nd cell-c y cle c h ec kpo i nt r e gu lati on , whe r eas BRC A 2 h as a m o re d irect r o le i n DNA re p air itse l f . BRC A2 i s t hough t t o p r o m o te g e no mic sta b ilit y t h r ough a r o le i n t h e err o r - fr ee r e pa ir o f D S Bs by GC v i a ass o ciati on wit h RAD 51. A b errati on s i n BR C A2 - d e fi c i en t ce ll s a ri se a t least i n p art by t h e u se o f t h e SSA p at h wa y . NHE J , howeve r , i s appa r en tl y un a f fecte d i n BRCA 2 - d eficie n t cells .

B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

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B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

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Lo ss of BRCA2, t he r e f o r e, r e s u lts i n t h e re p air o f DSBs by p refere n tial u tilizati on o f an e rr o r - p r one mec h a n ism , w h ic h po te n tiall y e xp lai n s t h e a pp a r e n t ch r o m oso m e i ns t ab i l it y ass o ciate d wit h BRCA 2 d eficie n c y .

B RCA 1 AND BRCA 2 MU T A TIONS AND DNA RE P AIR

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Th e phys i ca l i n t e r ac ti on b etwee n BRCA 2 a nd RAD 51 is esse n tial f o r e rro r -fr ee D S B r epa i r . BRCA2 is re qu ire d f o r t h e l o calizati on o f RAD 51 to sites of D NA da m age, whe r e RAD 51 f o rms t h e nu cle op r o tei n filame n t r e qu i r e d f o r r eco m b i na ti on. T h e f o ci o f t h e RAD 51 p r o tei n are a pp are n t in t h e nu cl eus a ft e r ce rt a i n f o rms o f DNA d ama g e a nd t h ese li k el y re p rese nt sites of repa ir by HR ; BRCA2 - d eficie n t cells do no t f o rm RAD 51 f o ci i n r es pon s e t o DNA da m age . T w o d i f fere n t do mai n s wit h i n BRCA 2 i n ter act w it h R AD51, t he e i gh t BRC re p eats i n t h e ce n tral p art o f t h e p r o tei n a nd a d isti n ct do m a i n, T R2, a t t he C -termi nu s .

P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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S yn t h eti c l e t ha lit y i s de fi ned a s t h e sit u ati on w h e n a m u tati on i n eit h er o f t wo g e n e s i nd i v i dua ll y has no e f fect , bu t c o m b i n i ng t h e m u tati on s lea d s to d eat h. T h i s e f f ec t was fir s t d escri b e d a nd st ud ie d i n g e n eticall y tracta b l e o r g a n is ms such as D r osoph il a a nd y east T h is e f fect ca n arise b eca u s e o f a nu m be r o f d i f f e r en t gene–g e n e i n teracti on s . E x am p les i n cl ud e tw o g e nes i n se p a ra t e se mir edundan t o r coop erati ng p at h wa y s , a nd tw o g e n es acti ng in t h e sam e pa t hway whe r e l oss of bo t h criticall y affects fl ux t h r ough t h e p at hw a y . T he im p li ca ti on i s t h at ta r g eti ng on e o f t h ese g e n es i n a ca n cer wh e r e t he o t he r i s de f ec ti ve shou l d b e selecti v el y let h al t o t h e t u m o r cell s bu t no t to x i c t o t he no rm a l ce lls . I n p ri n ci p le , t h is s hou l d lea d t o a la r g e t h e r a p euti c w i ndo w . T he o ri g i n al s ugg esti on t h at t h e c on ce p t o f s yn t h et ic let h alit y cou l d be used i n t he s electi on o r d e v el op me n t o f ca n cer t h e r a p euti cs ca m e fr o m Ha rt w ell et al . , a nd fr o m e xp erime n ts p erf o rme d i n y east . S yn t he ti c l e t ha l sc r een s h a v e no w b ee n p erf o rme d i n a nu m b er o f m od el o r gan i s m s and i n hu ma n cells , a nd t h ese h a v e re v eale d m u lti p l e po te n tial gene–gene i n t e r ac tio n s , s o me o f w h ic h c ou l d b e e xp l o ite d

P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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cli n icall y . Howeve r , syn t he ti c let h al t h era p ies h a v e no t b ee n cli n icall y used un til r ec en tl y , when ev i dence h as b ee n p r ov i d e d f o r P ARP- 1 i nh i b iti on a s a po te n tial syn t he ti c l e t ha l app roac h f o r t h e treatme n t o f BRCA -m u tati on– ass o ciate d cance r s. P A R P - 1 i nh i b iti on causes f ail u re o f t h e re p air o f SSB lesi on s bu t do e s no t a f f e c t D S B r epa i r . How e v e r , a p ersiste n t DNA SSB e n c oun tere d by a DNA r e p li ca ti on f o r k w ill cau se stalli ng o f t h e f o r k a nd ma y res u lt i n eit he r fork c o l lapse o r t he f o rm a ti on o f a DSB . T h eref o re , t h e l o ss o f P ARP- 1 i n c r ease s t he f o rm a ti on o f DN A lesi on s t h at mi gh t b e re p aire d by GC . A s a l o ss of func ti on o f e it he r BR CA 1 o r BRCA 2 im p airs GC , a l o ss o f P A R P-1 f unc ti on i n a BRCA1 - o r BRCA 2 - d efecti v e b ac kg r ound c ou l d r es u lt i n t he gene r a ti on o f r ep licati on -ass o ciate d DNA lesi on s no rmall y r e p ai r e d by s i s t e r ch r o m a ti d ex c h a ng e . If s o, t h is mi gh t lea d t o cell-c y cl e a rr est a nd / o r ce ll dea t h. T he ref o re , P ARP i nh i b it o rs c ou l d b e selecti v el y let h al t o ce ll s l ack i ng f unc ti on al BRCA 1 o r BRCA 2 bu t mi gh t b e mi n im ally t ox ic t o no rm a l ce ll s. T h i s wo ul d i nd icate a s yn t h etic let h al i n teracti on b et w ee n P AR P and BRCA1 o r BRCA 2. E x em p lif y i ng t h is p ri n ci p le , po t ent i nh i b it ors o f P AR P we r e app lie d t o cells d eficie n t i n eit h er BRCA 1 o r BRC A2. Ce ll su r v i va l assays sho we d t h at cell li n es lac k i ng wil d -t yp e BRC A1 o r BRCA2 we r e ex tremel y se n siti v e t o t h ese a g e n ts c o m p are d w ith h ete ro zy gous m u t an t o r w il d -t yp e cells . T o e xp l a i n t hese obse r va ti on s , a m od el was p r opo se d w h ere by p ersis tent si ng le - st rand gaps i n DNA cau se d by P ARP i nh i b iti on w h e n e n c oun tered by a r e p licati on f o r k mi gh t tri gg er f o r k arrest , c o lla p se , a nd / o r a DSB

P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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A lte rn ati ve l y , P AR P- 1 tr apped on DNA by t h e i nh i b iti on o f e n z y me acti vity mi gh t al so cause a f o r k co ll a pse . N o rmall y , t h ese DSBs w ou l d b e re p air ed by R AD 5 1 - dependen t GC . H o we v e r , i n t h e a b se n ce o f BRCA 1 o r BRC A2, t he r ep li ca ti on f o r k ca nno t b e restarte d a nd c o lla p ses , ca u si ng p e r siste n t ch r o m a ti d b r eaks. Wh e n re p aire d by t h e alter n ati v e err o r - p r on e D SB r e pa ir m echan i s m s o f S S A o r NHEJ , la r g e nu m b ers o f c h r o mati d a b e rr ati ons wou l d be i nduced, lea d i ng t o cell let h alit y T h e i d ea t h at t h e d e f ect in GC i s be i ng t a r ge t ed i n BRCA- d eficie n t cells is s uppo rte d by t he d em on st ra ti on t ha t de fi c i ency i n o t h er g e n es im p licate d i n HR als o c on fer s

P ARP-1 INHIBITI O N AS A SYNTHETIC L ETH AL THERAPE U TIC ST R A TEGY F O R T H E T RE A TMENT O F B RCA-DEFICIENT CANCERS

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se n siti v it y t o P AR P i nh i b it o r s T h is f u rt h er s ugg ests t h at t h is a pp r o ac h ma y b e mo r e w i de l y app li cab le i n t h e treatme n t o f s po ra d ic ca n cers wit h im p ai r me n t s o f t he HR pa t hw a y o r BRCA n es (see t h e f o ll o wi ng ) .

INITIAL C L INICAL R E SU L TS TES T ING SYN T HETIC LETHA L I T Y OF P ARP IN H IBI T ORS AND B RCA M U T A TION

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INITIAL C L INICAL R E SU L TS TES T ING SYN T HETIC LETHA L I T Y OF P ARP IN H IBI T ORS AND B RCA M U T A TION

INITIAL C L INICAL R E SU L TS TES T ING SYN T HETIC LETHA L I T Y OF P ARP IN H IBI T ORS AND B RCA M U T A TION

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P h ase I st ud i e s es t ab li shed t h at o la p ari b (AstraZe n eca , L ondon, UK; for me r l y KU - 0059436, KuD OS P h armace u ticals , Cam b ri dg e , UK) c ou ld be a d mi n is te r ed sa f e l y as a s i ng le a g e n t at a do se o f 400 m g twice p er d a y . Si d e e f f e c t s we r e c l ass ifi ed as mil d a nd were un li k e t ho se t yp icall y e xp e r ien ced w it h cy t o t ox i c ch em o t h era p y . Si gn ifica n t a nd du ra b le r es pon s es we r e obse r ved i n pa tie n ts wit h g erm-li n e BRCA 1 o r BRCA 2 m u tati ons and b r eas t ova r y o r p r o state ca n ce r . Of t h e 19 m u tati on carrier s e nro lle d, 9 had an ob j ec ti ve res pon se d efi n e d by Res pon se E v al u ati on C r ite r ia i n S o l d T u m o r s ( R ECIST) criteria a nd 12 h a d sta b le d isease f o r m or e t h a n 4 m on t hs i n du r a tio n. A similar ma gn it ud e o f cli n ical res pon s es w as ob s e r ved i n an expanded c oho rt . T h ese ob ser v ati on s are im p ressi v e b eca u se t he coho rt had been h ea v il y p retreate d a nd m o st were resista n t to a w i d e r a nge o f che m o t he r ap i e s . P h ase II s t ud i es we r e subsequ e n tl y p erf o rme d i n a dv a n ce d b reast a nd ov a r ia n cance r s a ri s i ng i n BRCA 1 a nd BRCA 2 m u tati on carriers T h e r e por te d r esponse r a t e was 41% i n t h e b reast st udy a nd 52 % i n t h e ov ari an group ; bo t h g r oups had been h ea v il y p retreate d. A g ai n, t h e d r ug was we ll t o le r ate d. Ano t he r s t udy o f BRCA 1 / 2 carriers wit h ov aria n ca n cer c o m pa r ed o la p a r i b w it h pegy l a t ed li poso mal doxo r ub ici n (PLD) . T h ere was no si gn i f ic an t d i f f e r ence i n t he r e s pon se rates , bu t t h ere were s o me d iffere n c es i n t h e pa ti en t cha r ac t e ri s ti cs and a n un e xp ecte d l y h i gh rate o f res pon se t o P LD. Th e r e a r e a l so r epo rt s o f r e s pon ses t o P ARP i nh i b it o rs i n BRCA 2 m u tati on ca rri e r s w it h p r os t a t e a nd p a n creati ca n ce r . A nu m b er o f o t he r

INITIAL C L INICAL R E SU L TS TES T ING SYN T HETIC LETHA L I T Y OF P ARP IN H IBI T ORS AND B RCA M U T A TION

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P A R P i nh i b it o r s a r e i n c li n i ca l d e v el op me n t ( ) , a nd s o me o f t hese h a v e s hown e f fi cacy i n t he tr e atme n t o f ca n cers arisi ng i n BRCA 1 o r BR C A2 m u t a ti on ca rri e r s .

T H E U SE OF P ARP INHI B I T ORS IN SPORAD I C CANCERS

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T H E U SE OF P ARP INHI B I T ORS IN SPORAD I C CANCERS

T H E U SE OF P ARP INHI B I T ORS IN SPORAD I C CANCERS

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G e r mli ne m u t a ti ons i n BR C A 1 o r BRCA 2 are relati v el y c o mm on i n h e r e d ita ry b r eas t and ova ri an ca n ce r . H o we v e r , i n acti v ati on o f BRCA g e nes by m u tati on i n spo r ad i c canc ers is rare , at least i n b reast ca n ce r , w h ic h m ay seem t o limit t he app li ca ti on o f P ARP i nh i b it o rs t o a wi d er ra ng e o f p atie n ts . Howeve r , m any t u m o rs d is p la y feat u res i n c o mm on wit h BRCA- d e f icie n t t u m o r s, i nc l ud i ng s imilar d efects i n DNA re p air du e t o eit h er e p i g e n et ic m u t a ti on o f BR C A 1 , s u c h as p r o m o ter met hy lati on, o r m u tati on of o t h e r c o m ponen t s o f BRC A-ass o ciate d p at h wa y s . T h is BRCA n ess may ma k e t hese t u m o r s a l so suscep ti b le t o P ARP i nh i b iti on F o r e x am p le , pho s ph at ase and t ens i n ho m olog ( PTEN) m u tati on s , w h ic h o cc u r wit h a fr e qu e ncy es tim a t ed a t 50 % t o 80 % i n s po ra d ic t u m o rs ma y ca u se P AR P