“
`
`map/ea! Na; mammals):
`ibposizim against E? Palm: {{3; 93 (1136.53 32563138}
`Patmtea. Mimi’me M3
`{hmitim by. (Jemima Richmr Ltd.
`{3m Rafi: £62363 Ema-1:2?
`
`WSSMS 3c PARYNER
`WWW * kaé
`swam-{gm 4%
`8'16???) MUNQHEN
`
`203
`
`Antiuterotrophic efiects of the pure antioestrogen ICI
`182,780 in adult female mankeys (Macaw nemestrina):
`quantitative magnetic resonance imaging
`
`£9510
`
`Mfi Dukes, J. C. Waterfall and A. E. Wakefing
`
`Cancer Research Departmem, 2ENBCA Pharmaccmicais, Meresidc, Aldcrley Faric, Mamlesfield,
`Cheshire gamma, UK.
`
`Rewsw MANUSCNFI’ RECEIVED 15 ngruary 1993
`
`
`AfiSTRACT
`
`size 01‘ the myometrinm deareaseé signifimntiy. by
`The antiutemtmphic cfiicacy of the pure antioestm—
`15% from cady In mid cycle and then rawwmi ta
`an K21 £82389 has been demonstrated pmiausly by
`( Egnetic resonance imaging (MR!) in ovariectomized
`near its initia! volume in the Kate cycle. Treatment
`with ICI 182380 beginning in the: aaxly part of the
`aestmgen»lreated monkeys
`(Macaw nemsxirina).
`menstruai cycie prevented the growth of the uterus.
`Fufiher charafiterizatim of ihe effects 0f {CI 182388
`in intact adult female monkeys with nomai menstrual
`The magnitude and duratinn at” the response was
`fiependent on whether a: not avalatian occurred
`cycles was underiaken to pmfide an indicatian 0f its
`during treatmcnt with It?! 382.1380; In animals rm»
`palatial aciions in prcmcnupausa} wumcn. Changes
`dared anevulamry, growth of the endametrium was
`in the volume of uterine tifisues wen: measured by
`blocked compietely by ICI 182380 and the wfiume of
`MR! in early, mid and law cycic. The wmme of the
`the tissue daclincd below tha: prcscm at th: siart 3f
`menu varied up to fivefold between individual
`the manstmai cycie. Antiuterozmphic eflficacy was
`moukcys but
`serial ubsawatinns
`in
`individuals
`signifiwmiy less in monkey whiuh mule:th during
`pmvidad suifinim: premisian ta afiaw accurate: asses?»
`treatment with iCI 182.?8fl. Th»: volume: caf mama
`ments to 33: made ef changes in tha endometrium and
`mairium was reduceé subsiantiafiy by amicastmgw
`mynmctrium timing the marge 3f the menstmai cycle
`treatmant, and the difierence in response between
`and follawing ICI 182,786 adminiatration, In cam‘
`ovulatory and anewiatozy mankeys was less marked
`parison with its initial size in unnamed mankeys, the
`than that of the endometrium.
`endwmetrium increased in valume by 60% and 125%
`3m:sz {af Enfiacrfnaiagy (i993) 3381 2&3—26‘?
`in the mid and late a}:ch respectively. In cont ran, the:
`
`KmTRGflUCTIGN
`
`ICI 182380 is a preteen: spacific pure: aminestmgen
`which may pm“: superiar m canvcntianai partial
`aganist anthestmgens in the ire-airman: of breast
`Cancer {Wakefing e: 31‘ 1993). Preview studies
`avail-431w the antisestmgenic potency and effiaacy of
`ICI 182386 in avariectomized Gestmgcmtreated
`monkeys (Macaw aemesfrina) by magnetic resonance
`imaging (MRI) of the uterus (Watertan at a}. 19:91;
`flukes e: 0!. 1992). A daily (Ease of {ii-1mg ICI
`ISRJSBEkg sample:er biwked the utermmphic
`actian Gf magenous aestradinE-«l’f3. Singie intra«
`museum:
`injectians
`at" a
`innyacting ail—based
`farmuiation of
`ICI
`182333 in wariectomézed
`menke'ys were 3150 shuwn t0 inhibit oastmgcn acticm
`in a dosevdepcndent sustaincd manner (Dukes er as}.
`
`1992}. MRI has been used to abscrve the 1::me
`during the Hanna) menstrual cyan: in women (mamas
`e: a!" 1986; Raynor er 4?. 1986; McCarthy er a}. 1986;
`Wiczyk e: :21. 3988; Earmfi er al‘ 1991) and the
`response :0 exogenous endocrine agents {Demas
`22 a}. $86; McCarthy a: £211 1936; Andreyko a; :23.
`1988; zawin e: :25. 1996), MRI has aim been 1133:}
`quantitatively {a measure the: changes in madame af
`ihe endometrium anti myometrium during the normal
`manslruai cycle in M. nemastrim (Watermn é’f a1.
`1992} The MRI studies repented here were performed
`is waluata {has antiutcmtwphic effects a? It?! 132.7780
`in admit {emaie monkeys with normal menstrual
`cycles{ Such gtudies are rciwant
`to the mammal
`appiicaiien of ICE £82333! in the treatment of bath
`malignant and benign oestmgen—responsive (£53me
`in pre- and perimmopausal women.
`
`mea! {3}“ Eadem’asfagy {I993} 138., 2036253
`m22w¥37§5f93ffl133~3283 “2.01333
`
`(E 29193 Jamaal 9f Emiocrinelngy L16
`
`Printed in Gram 3mm
`
`InnoPharma Exhibit 10570001
`
`

`

`1,.
`
`204 M. 1mm and others
`
`
`Efi‘eem ef 3C? 1’82, me in grief: memtmaffrrg menkefi
`
`MATERIALfi AN!) METBGDS
`
`Mimi handfieg are} matinee: Mules
`
`Mamie femaie pigtail monkeys (M. eemerrreze) of
`see kg body weight with regular menstrual eyeiee
`were used. The normai menstrual cycle in this. species.
`is 29-32 days in length; menstrual bieeding usuaily
`lasts 3—4 days. Daily vaginal swabs were taker: to
`detect the were: of meme-eel bleeding and the first
`day ef menstruation was riceignared day l ef the
`eyeie. Where required, bleed. samples {2 ml) fer eubw
`sequent humane anelyees were eelieeted daily. MRI
`examinatiens tee}: place before the animals received
`{heir returning feeri. in order m minimize artef‘aeteel
`neiee associated with get meiien. Anaesthesia was
`induced will: 8*? mg ketamineikg (Verifier: Parke
`
`Dares, Pentypoel. Sweet, LLK.) and riaintained
`with haleihene (Fluethene;
`1C1 Pharmaceuticals,
`Maeelesfielci, Cheshire, UK.)
`The timing 0!“ MR1, drug treatment and bleed
`sampling was echednled by reference to the firei tier
`of menstruatien. To erevide a baseline with which to
`compare the effects ef rimg treatment,
`three MRI
`measuremems ef‘ the uterus were made during each of
`two cycles in ten unireeteri menkeye (Warmer: er e3.
`19%). Images were recorded in early {days 2—6}. mid
`(days 12-18) and late (days 21.45} cycle am! sched-
`uled by reference to the elifi'erenoes in cycle length in
`individueis.
`images were reeerded at similar time-e
`paints in menkeys treated with R31 £82331 beginning
`early in the menstrual cyele (days 2—5). Tim ferme—
`laliens {if ICI 132383 were used: a. shortvaeting
`prapylene glycol-based evolution (F 1) administered
`once daily im. for 25 days at a dense ef {H (exS) or
`02 mg (a: 5) EC! ISQJflGikg, and a lengeaeting easier
`eii~baeed soiutiee (F2) given as a single im. inieetiee
`at” 2'5 (11:26) ar emg (anti) {CI L8238flflcg.
`
`Quantitative Mill
`
`The metth Fer measurement 0f endemeirium anti
`
`myemezriam eeiume is described in detail elsewhere
`{Water'th er 5;}. 1991, 1992.; Dukes er mi.
`l992).
`Eriefiy,
`the study empleyed a. Bieepee 40012.3
`instrument {Oxford Reeeareh Systems, Cerentry,
`Warwiekshire, UJC-) m reeerd a series (if eblique
`images (if the uterus. Fer eaeh animal on each
`oceasien, sagittal sighting images were used re deter»
`mine the: spatial {rev-ordinates 0f the uterine mrvix and
`famine. From these ee~erdinaies, MR! parameters
`were calculmed to allaw the acquisilian (if eight
`eemigeeue eeiiqne eliee images, the thickness, peeie
`tier: and Grieritatian {if which were famed m depend
`on the eeeter cementing the fundus and cervix. Thie
`pracedure allowe repmdueihie presematien {if the
`
`Jeureei e! Endeert‘neleg}? (29%) 133, 3333439
`
`uterus, independent ef its Sixth location and edema.
`tier! in the pelvis {Dukes e: mi. 19%; Waterten er :3}.
`1992}. Image matrices were 256 X 256 giving an in
`plane reseiuiiae ef 6-6 mm. The appearance el‘ the
`images (see the Flate} results from the choice eat” MR}
`parameters, in particular the magnetic: field strength
`of 2‘35 tesla, the eehe time ef 501115. the repetition
`time 3f 3 s. and euppressien {if {he signai fmm fat.
`To allew comparisons between imagee acquired on
`different making, iissue eelumes were measureni
`rather than enéometrium heighi er area, since the
`former avoids arlefaete fmrm repesitienieg errors
`or shape change-e. Endomeirium and myemetrium
`velumes were obtained by pixel wanting in the
`oblique images eeing the irregular regiemeflimereet
`facilities within the pregrems uxemr er DISNMR
`(Bruker Analytieehe Meestechnik Gmhfl. Karlsmhe,
`Germany). Only slices more feudal than the internal
`as were included in the analysis. The junction zone
`was includee with the myometrium. Any contribution
`in enciernetrium velume from the lumen was neglected
`since. except during menstruatien, this is irent—y narrow
`and insignificant in mmperisen with {he endenierrium
`(Dukes; e: a}, 1992; Waterien e: :33. 19?2}.
`Altheugh, in principle, the prescription 0f regien»
`ef~interesl can be subjeetive, the method empleyed
`minimizes errors, since the presentation cf the uterus
`was the same at each examination (Watenen er a3.
`1991. 3992; Dukes er e1. V392). Fer each sanitize]r the
`same eesewer analysed eli imagee. Weathers-week ami
`eyelevteeycle coefficients of variericn were as law as
`5% {Weterten er e}. 3992).
`
`Analysis of rewrite
`
`The volumes ef the endemetriem and myemetrium in
`treated and untreated greups were calculated as the
`arithmetic mean (i: 5.2M.) nemalized by reference
`to tissue velumes in individuals at
`the first MRI
`measurement. Differences between greup means were
`tested fer significance by the unpaired Student’s
`:--test.
`
`RESULTS
`
`Uterine volume in the natural menstrual cycle
`
`The velumee ef the endometrium and myemeu‘inm
`were meaeured by MRI in early. mid and late cycle
`in two cycles in each of ten different untreated
`mankeye {Text-fig. 3). The mean interval between
`repeat measurements was 4 menths (range 1-9
`mes-mile}. Tissue winmeg varied widely betwaen indi»
`eiduaie but less so in repeat measurements in the same
`animal Tyeical images reeerded in me maul-my at
`
`InnoPharma Exhibit 1057.0002
`
`

`

`
`
`' M. DUKES and othersEfll’cu oj’ICI l82.780 in adult meant/wring monkey-3 205
`
`
`
`4'0
`
`(“1
`
`3-5
`
`3.0
`2-5
`2'0
`
`\
`
`
`|
`
`I
`'
`
`\
`
`|
`
`w
`
`I
`
`'
`
`Q2—me:
`olume[cm) O
`
`
`
`M O
`
`‘ V
`
`:3 m
`
`mN0‘
`
`N JJ Lil
`
`Ng» on
`
`.. \o
`
`NLo)9.:
`
`N N m
`
`N \D
`
`0'5a
`
`N a:
`
`A.a“ V
`
`'
`
`I
`
`18
`
`[6
`
`l4
`
`I2
`10
`
`[38
`
`226
`
`235
`
`238
`
`233
`19]
`Animalnumbcr
`
`228
`
`2|9
`
`169
`
`217
`
`TEXT‘FIGURE l. (a) Endometrium and (b) myomeuium volumes in untreated
`monkeys during the menstrual cycle. For each of ten animals the bars represent
`(From left) tissue volumes measured by magnctic resonance imaging in two
`separate menstrual cycles, during the early (days 2—5}, mid (days 12-48) and
`lalc (days 21—25) stages of the cycle.
`
`Journal of Endocrinology (1993) 133. 203-209
`
`InnoPharma Exhibit 1057.0003
`
`

`

`206 M. DUK and others
`
`Ejects of [CI 182.780 in adult menstruating monkeys
`
`
`
`
`
`
`
`Normalizedvolumes(We)
`
`Untreated
`(n = 20)
`
`Pl (0-1)
`01:30
`
`F1 (02)
`(n = 4)
`
`F2 (2-5)
`in = 3)
`
`F2 {4-0)
`01 = 5)
`
`TEXT—FIGURE 2. (a) Endometrium and (b) myometrium volumes in untreated
`and ICI {BAND-treated anov'ulatory monkeys. Tissue volumes at mid and late
`cycle were normalized to those recorded at the beginning of the cycle in each
`individual. From the left, values are means i 5.8M. for untreated controls;
`monkeys receiving once daily injections of 1C] 182,780 (O-l 01' 0'2 mglkg) (Fl)
`for 25 days. beginning on cycle day 3; monkeys receiving a single injection of
`1C] 132.780 (25 or 4 mg/kg) (F2) on day 3.
`
`days 3 (early), 16 (mid) and 24 (late) of the cycle are
`illustrated in the Plate, figs 1—3 and in another case at
`days 2, 16 and 23 in the Plate, figs 7—9. Analysis of the
`data showed that >75% of the variance arises from
`
`Efl'ect of [Cl 182,780 on the uterus
`The effects of [CI 182,780 on the appearance of the
`uterus in MR images is shown in the Plate. Plate, figs
`4—6 and 10—12 illustrate the mid corpus of the uterus
`at early. mid and late cycle, in individuals treated
`with the rapid— (F1) or slow— (F2) release Formulations
`of KS] 182,780 respectively, compared with images
`recorded in control cycles in the same individuals
`(Plate,
`figs 1—3 and 7-9). The dramatic shrinkage
`of the endometrium (central high~signal zone)
`is
`immediately apparent;
`the myomctrial changes are
`less obvious from visual inspection of the images but
`emerge clearly from quantitative image analysis.
`In monkeys treated with [Cl 182,780, the magni-
`tude of changes in the volume of the endomctrium
`and myomctrium was dependent on whether or not
`individual monkeys were rendered anovulatory by the
`treatment. Where serum progesterone concentration
`(not shown) remained below 2 nmolfl in the second
`half of the menstrual cycle. animals were judged to be
`anovulatory; similarly where progesterone exceeded
`this value and remained elevated for at least ll] days,
`ovulation was assumed to have occurred. By this
`criterion, treatment with 0-] or 0-2 mg ICI 132,780fkg
`daily (Fl) blocked ovulation in three or four of the
`
`InnoPharma Exhibit 1057.0004
`
`interanimal variability. Average coefficients of varia—
`tion for volumes measured in difl‘erent cycles were in
`the range l4—19% for the endomctrium and 63—10%
`for the myometrium.
`The
`endometriurn increased
`
`in
`
`the
`
`in
`
`size
`
`follicular phase of the menstrual cycle in 17 of 20
`cycles (mean increase 60%, P<D-001; early vs mid
`cycle volume) and this growth continued in the
`luteal phase in 18 of 20 cycles (mean increase 65%,
`P<0-001; mid vs late cycle volume). Changes in the
`volume of the myometrium were smaller but a
`consistent pattern emerged in which a decrease in
`size was observed in 18 of 20 cycles in the follicular
`phase (mean decrease 16%, P<0'001; early vs mid
`cycle).
`In the luteal phase the volume of the
`myometrium increased in 18 of 20 cycles (mean
`10%. P<0-02; mid vs late cycle volume). The net
`effect of the follicular phase shrinkage and luteal
`phase growth of the myometrium restored mean
`myometrium volume to near that recorded initially
`(94%, P>0~05 compared with early cycle).
`
`Journal of Endocrinology (1993} 138. 203—209
`
`

`

`Eject: of [Cl [82.780 in adult menstruating monkeys
`
`M. DUKES and others
`
`20?
`
`
`
`
`200
`
`A100
`
`g3
`
`3’
`
`Ea E1
`
`0
`
`, E
`
`"5
`E0
`Z
`
`(“100
`
`50
`
`o
`
`Untreated
`
`Fzr4-0)
`1:20-53
`r295;
`Fzrz—s)
`Firs-2]
`rim-1)
`Fl (0-1)
`169
`233
`219
`217
`174
`225
`235
`(a) Endometrium and (b) myometrium volumes in untreated and lCl I82,780-treated ovulatory monkeys,
`TEXT-FIGURE 3.
`Tissue volumes at mid and late cycle. normalized to those recorded at the beginning of the cycle in each individual
`monkey. are compared with mean values (it starts. n=20) for untreated controls. From the left. values are
`means i 5.514. for untreated controls; monkeys receiving once daily injections of [CI 182.780 (0-! or 0-2 mgikg) {F1} for
`25 days beginning on cycle day 3; monkeys receiving a single injection of [CI 182.780 (25 or 4 mg/kg) {F2} on day 3,
`
`five monkeys respectively, in each dose group. Simi-
`larly. ovulation was blocked in three or five of the six
`monkeys receiving a single injection of 2-5 or 4mg
`ICI 182.7801kg (F2). There was a tendency for the
`her-dose groups to include fewer ovulatory cycles
`“in this was not statistically significant with the small
`group sizes employed.
`The mean changes in the volume of the endo-
`metrium and myometrium in [CI 132,780-treated
`animals are presented in Text-figs 2 and 3. Since
`absolute tissue volumes vary widely between indi-
`viduals
`(Text-fig.
`l),
`the effects of 1C] 182,780
`were assessed by normalizing the volume of tissue(s)
`recorded in each animal at
`the mid and late cycle
`observations to those recorded at the beginning of the
`cycle. prior to treatment, and by comparison with
`similarly normalized observations in the untreated
`controls.
`in monkeys rendered anovulatory by lCl
`[82.780 (Text-fig. 2). the normal follicular and luteal
`phase growth of the endometrium was blocked
`completely and tissue volume at mid and late cycle
`was significantly (P<0-001)
`less
`than that before
`treatment began in each of the treatment regimens.
`
`Although shrinkage of the endometrium was most
`apparent during the first half of the cycle {mean
`decrease — 61% compared with +60% control.
`P<0AOOL at mid cycle). blockade of growth was
`sustained throughout (mean — 67% compared with
`+i25"/u control. P<0'00l. at
`late cycle). When the
`occurrence of ovulatiOn was accounted for. no sig-
`nificant differences emerged between the effects of the
`different formulations and doses of ICI 182,780. with
`the exception that
`the 2-5 mg dose (F2) appeared
`slightly less efiective (P<O-05) than the 4-0 mg dose in
`the second half of the cycle The volume of the
`myometrium in [CI 182,780-treated anovulatory
`monkeys (Text-fig. 2) decreased more rapidly than in
`controls in each treatment group from early to mid
`cycle (mean - 35% compared with - 16% control.
`P<0'001. at mid cycle) and this decline continued in
`the second half of the cycle (mean - 54% compared
`with — 6% control, P<0-001, at late cycle).
`In monkeys in which treatment did not block
`ovulation (Text-fig. 3), the mean effect of 1C1 182.?80
`on the endometrium in the first half of the cycle
`( — 27% compared with +60% control. P<D-0l] was
`
`Journal of Endocrinology (1993} 138, 203—7209
`
`InnoPharma Exhibit 1057.0005
`
`

`

`y
`
`298
`
`M. DUKES and withers
`
`Efiecm of EC! 182,780 in Mull menstruating manicey;
`
`seen in the anawlawry mankcys
`less than that
`but not significantly {1333305} 50* Haw-wan that:
`mean respmse cancealccl mid: diffcrcnces between
`inéividual monkeys with five {3f the seven animals
`shnwing shrinkage (mean « 353:5%} whilst
`the:
`ramaining two mankeys experienced net growth at"
`the endametrium (+103% and +28%). In the latter
`animals, the gmmh 0f endamemum resumed in the
`luteal phase of the cycle 12+?O%) at a rate similar :0
`mat
`in animated mankeys {+55%}. Mean lissae
`valume remained, hawever, legs than that in animated
`canimls at the: and 0f the cycle: (Pvillfil), but grater
`than that
`in anomlatery monkeys (P<O-DOI). A
`similar pattern at" change was observed in the
`myamelrium (Texbfig. 3). At mid cycle the mean
`valume {3f
`the tissue was
`reduced significantly
`in 33%: P<0m compared with central) and this
`alike: was mt different {mm that
`in amvulawry
`animals (w 35%} However, as was the case for the
`endomelrium,‘ the: myometrium did recover in the
`luteal phase (+18%) but m: m the volume Seen in
`cwtmls. Al
`the late cycle ubservatiun, meets (in?
`treatment an this mynmstrium were significantly lesg
`{was}; in ovulaiary than in anavulawry mankeys.
`
`DISCUSSIGN
`
`MRI pmvidess an accurate: and rapmducihle means 0f
`measuring the changes in vellum: 0f the uterus in
`rcspoma in mmmgen trcalmem in ovaficcmmizgd
`moakeys (Watertight; e: 01. 199E) and has been aged in
`evaluate the antimemlwphic antigen 0:“ lC‘l 132330
`(Dukes e: m“. 1992} This technical}: has hem applied
`previously 10 determine the changes in volmm of
`the endometrlam and myometrium during the can ms
`of the namai mensirual cycle {If
`314'. 53332935er
`(Waive-“110:3 s: all. 1992) and in this {apart to measure
`the flats of treatmsnt with ICI 182388 {m mass
`changcsl
`At the time of menstruation, uterine aim, in partic-
`ular that of the endemetrium, would be expecmd m
`reach a minimum and 10 increase timing the failicular
`plus: {3f the cycle and, m a lesser Extent, in the lutgal
`phase. "Fa aédress this question, MRI measuremems
`were made at 53:13, mici am! lat: Cyclt: in each of ma
`manual cyclas in ten monkeys. In the early part :32“ the
`cycle (days 2-4), the valuma if she endamctrium and
`myomatrium in individuals varied ever a wiée range
`{up in fival‘alcl), but tissue volumes were: much lass
`variable in repetitive measurements in the same
`indiviclual (Taxi—fig. I), A cigar pattern 0? change:
`emerged during {he mmstruai cycle in which signifh
`cant gmmh cf the: endomelrium was $3311 in both {ha
`fallicular and luteal phases. Perhaps surprisingly, ths
`mean inflame in six: 0f that: mdnmeu‘ium was similar
`
`Jamaal q! Macriwlugy (19%} 138. 2133-209
`
`comparing the first and second half of {he cycle.
`Gastradial is cansidered m be flat: majar mitagan for
`the uterus and it was thus anticipated that :13: laxgest
`change in this andnmmrium would occur in the falli-
`cular phase at” the cycle. Hewever, a propcrtionately
`similar increase in the valume at” the andametrium
`
`was seen in the luteal phase ef the cycle, MRI cannot
`be: used to dflemine whether the effects 0f aestrogen
`and progestcmne on the size of the endometxium
`can be attrihuted mainly m epithelial hyperplasia as
`oppased to hyperfinphy. Changes; in the size of the
`myamctrium during {ha menstrual cycle were quantii
`tatively much smaller than these 0f the andnmeirium
`and! in the follicular phase, qualitatively difi‘erent
`{Ten-fig. l}. Remarkably, 11m mycmetrinm (increased
`in size when the endometrium was gmwing, mean
`myomclrmm mlmne being smaller in mid cycle than
`at menstruatlen. This may indicate that that: lhreslmld
`0f sensitivity of the myomctrium m oeslmgcns is
`higher than that 01‘ the endumcmum andior thal the
`rate at which {he myemetrium responds :0 changes
`in homanal status is lesa than that far the enciw
`metrium. Hawcwr mile:
`lrryltvcflzlanimus1
`such as
`harmoneénduced changcs;
`is: muscle: mm, may
`influfince the apparent size. cf the uterus and are mm
`amaumed for in these MRI images.
`Initial slutiies (11‘ the efiects cf ICI 182380 em-
`playecl a daily {lasing regimen with the antioestrogen
`laminated in propylene glycnl vehiflle {F3} to provide
`rapid release in vfivo. ll was Shawn previously thal the
`chasm (11:35:35 (if K11 132,?3l} {0-1—82 mglkg per day}
`achieve complete and rapidly reversible blackadc uf
`exogeneus assuage!) stimulation at” the uterus in the
`ovariectamized mankeys {Dukes a: all. 1992). Daily
`treatment timing the menstrual cycle pmvided c183:
`tfidence that 1C1 £32,780 also has a substantial
`antieestmgenic efi’act {>11 the uterus in intafl monkeys
`{Taxi-figs 2 and 3}. The magnitude of the anti
`uterotmphic cfi'ect was strangly influenced by the
`censurrent cndocrlna effects of
`ICI 182330,
`in
`particular by whether the munkeys ovulated during
`the: treatment pflfifld. In anovulatory monkeys, am
`only was the manual
`increase in volume 01‘ the
`endamclrium in this: fallicular phas: abs:an the: render»
`metrium actually dccrcasad in size to: velume mm-
`parable to these 3:611 in avariecmmizcd munkays afth
`oestrogen withdrawal (Wanner: at al. 1991; Dukes
`er al. 1992}, indicating that treatment achievecl mm—
`pletc blackade 0f 1hr: trophic acticn of endegenaus
`
`matrugcnsl This blockacic of“ nastragrzn actian (m that:
`endametrium was snslained in the mortal half of
`
`the cycle. An fiesta-agar: withdrawalwlike amiss-n an the
`myameitium was also recorded, where the normal
`small damage in volume: at" the mynmetrium in the
`fallicular phase was significantly amplified by ICI
`l82,?8fi, and 3150 became similar in magnimdz: {a than
`
`InnoPharma Exhibit 1057.0006
`
`

`

`
`
`m. DUKES and others 209
`Efi‘eem af EC! 182. 381i} is: ads}: mensrmring mankeys
`
`seen follawmg cestrogen withdrawal in ovariectm
`might} mankeys. This shrinkage. :1? {he myamirium
`ceatinuad in the luteal phase.
`The alternative oil-«based mug-acting {emulating
`(F2) of ICE 182,780, shown previausly to sustain
`blockade al' the ulcrolmphic actiorl a? assimilin in
`ovarieclomimé monkeys far appmximately 1 month
`(Dukes er 3!. 3992), graduced cfl'acls 0n the uterus
`similar to these: a? the short-acting Formulation.
`'I'heSc previws phamacnlagieal findings are cmircly
`cansistem with the findings in the present study with
`respect ta the duratian at” actien, the apparent dese-
`response, and the longer sustained blockade ni‘
`myamctrial than enclamelrial gmwzh. In mankeys
`rendared anevulatery, the antinlerotmphic effml of
`the 4H) mgfkg dase was inclistinguishable {mm that
`(if the shawaming fannulalion. Hawever, the law:
`2-5 mgflcg time dizl net fully black the lmphie acliun
`pandagcnuus assuagcns on the enéumeitium in the
`mam! half at" the cyclé, but did cam pragrcsslve
`myomctrial shrinkagfi. This prevides further evidence
`that the threshold sf sensitivity 0f the myemttrimn to
`nestrugens is higher Khan that of the endamelrium.
`With hath fowulations, the antiutemtmphic all?»
`navy of ICI 182380 in monkeys that maimed during
`treatment was mole variable: and generally smaller
`than in anovulamry animals, but mill substantial in
`the majority {five of aeven} of individuals {Text-figs 2
`and 3}.
`Overall, 91‘ the total :32" 22 treated ‘cycles’, 15 were
`anmulamry. With both fonnuiations there was same
`:vidence:
`that
`inhibilion of ovulalmn was éasm
`
`related, but than: were insufficient. data m efitablish
`slalistimlly aigm’ficanl clifi‘erences. The reascms {at the
`variability between individuals in their susceptibility
`{0 blockade a!“ ovulation are rm: understand.
`
`These studies warm mmplated usng nan-invasive
`:11 methods will: fewer than 20 animals. In practice
`slick infamatian is unabtainablc in terminal experi»
`meats. Far example, it is estimated that nitration of
`myometrial shrinkage at the sama timz as cndamelrial
`gmwth by terminal methads at the same significant:
`lcvel weuld have required we: 3’30 animals. Thus,
`MRI may be a particularly sensitive and pewarful
`100} for clinical studies in endearine phannacalggy.
`The clinical usefulness of ICI 182380 remains in be
`dfitennined, but its efi‘cat in a case m“ spontanwus
`adenomyosis
`in a mankey has been described
`(Waterten e: :23. 1993}.
`
`ACKNGWLEEGEMEN?S
`
`We {hank S. A. Brew, M. Herrecks, D. Miller,
`J. S. W. Morrell. 3. B. Latcumbe-Mcbauall, l). Priest
`and .l. Tattershall far expert technical assistance.
`
`REFERENCES
`
`Aadwyku. I. 3.... Blank-mad, 21., Mambafl. L. 5.. Mama. 5. 13.,
`Erica}; H. .9. 133:. R. B. (1933}. Us: cf an aganlxfis analng
`nf gonadmropiwrelcaaing llama“: {naiarelin} w twat
`leiomyamas: mama“! by ma.ng rmnancc imaging.
`Amarimn £2an :9} Qészerrirx and Gymmlogy 1.58, 903M910.
`Bartuli, 3. 1%. Maulin. (i, Balsam-33+. l... Chagnaud, C. S:
`Kasharian, M. {1991}. This normal Bums an magnctit
`mum imaging and variatlans amaze-z! with the:
`mammal 3:31;. Surgical-W&$&g§€ Assaimy l3, 213420.
`9333133.. B. E... Hamil, PI. & Jaffc. R. B. (19823}. Uterine:
`imaging: efi'wlfi (If harmonal stimulation. Radialagy 1159,
`323-1213.
`Dukes. M. Miller. 3)., Wakeling, A. E. & Waterion. I. (‘3.
`(I992). Améulemiwphic 1:3me 9f a pure amimlmgm, ICI
`182380: magnetic ram-nan“ imaging of the stems in
`avariestomimd monktys. Jamaal {If Endaamlagy 135,
`339-341
`Rayner, 3}. IL. Mack, L. A... Sauna. M. 1L, Shumaa, W. R.
`Mamana. M. A. a: Man. a. A. (1936}. Changing appearauw
`0f the rim-mall uterus timing the menstrual cycle; Radiafagy
`16!, 459-462.
`McCarthy, 8.. Taubcr. C. & Gert. I. (1986). Fatal-naive pelvis
`anammy: MR assessment at” vadatiom during the menslmal
`cycle and will: use at“ am} wntraccpiivts. Raéiaiagy 1&8}.
`l 194 23.
`
`Wakeling. A. E... flakes. M. & fiawler. 1. {I993}. A patent
`specific: part; antimslmgcit with clinical mmnfial. Cancel!
`Remam‘: 51, 3367-38?3.
`Watwon. .l. (2., 3mm. 8. A... flakes. 1%.. Harm-sits. M. 3:
`Wadsworth. P. {3993}. A $381: of aficnomyasis diagnmtd by
`magmatic remancc imaging and treated with the navel pure
`mtmlregen ICI NEW). Lawlvawzy Animal Scimw {In
`Press-.3
`Waleflon, .l. {1. mmmlwficbouall. 3. E. 8: Mill::'. l}. {3992}.
`Quantitative MR! of" the washing and Mann in monkeys.
`Magnesia Remnants in .3696er 28. 3&9}.
`Watering, J. fl, Miller. 1)., Bukas, M. & Harrell, J. S. W.
`(W913. leique NMR imaging sf the mama in macaques:
`marina teaponxe: w manger: stimulatian. Magattiicé Rename
`in Medicine 20. 223439.
`Wimyk, Fl. 13.. 3311623, C. 1... Richards, (3. 1., (3131‘. M. 3.,
`Geadal, E. 3.. Rabimwitz. J. G. & Laura's", N. (£983).
`Comparison {if magmatic manna: imaging and ultrasonnd in
`evalualing willme and mtlmmtrial dwclnpmcm {Maugham
`the 33mm? 63ml: Fag-way we? Sig-ti!in 48. 96§~9?S.
`Kama. Ni, McCarthy, 8., Scam. L, Langc, R, Levy. (3.. Valle.
`3. 8: Camilla. F”. (39%). Munimring therapy with a
`gunadotropm—rcleaszng humane analag: utiliiy 0f MR
`imaging. Rafiaiagy ['35. 503606.
`
`Jamaal a)!” Emtrmafagy {1993} 338,. 233.209
`
`InnoPharma Exhibit 1057.0007
`
`

`

`a
`
`Efacm 9f 16.? 182.789 m Md: mmsruwmg mankeys
`2m 51. mums and mhars
`n
`
`fiESCRIPTiON 0F PLATE
`
`Magnetic remnants: (MR) images a: the monkey uterus.
`Each illustraticm is a den-ail, represgming 3% X 3-4 cm,
`{mm 031: 3311:: at" 111:: eight racorded at cash maasurcmem.
`The: appearanca a: mid-cat?” utcri is Shawn as faklows.
`Tap raw: images abtained a: day 3 (Fig. 1), day 16 (Fig.
`2} and flay 24 (Fig. 3} if an animated menstrual cycle.
`Swami raw; images ubtained at day 3 (Fig. 4}, day 15
`(Fig. 5} and day M (Fig. 6) in the same animai during a
`cycle: is: which 6'} mg 1C1 18233433“ was injamd wily
`fram day 3. Third row: imagea obtained at day 2 (Fig. 3'),
`day E6 (Fig. 3) and day 23 (Fig. 9} at“ an untreated
`menstrual cycle. Batmm row: imagw obtained at day 2
`
`(Fig. 10}, day 16 (Fig. 11) and day 23 (Fig. 12} in the
`same animal éuring a cycle in which 4 mg 131 182,?8flikg
`sustained-Eckmse farmuiatien was injected an day 2. The
`cycies ahown in the: mp three mm were owiatarjr. Fm:
`at five mnmtfi: macs sf MR signal annuity an scan:
`{33:11:31 sigma} wig}, menstmai hammrhagc (0913: during
`menstruation); high siwal. endamctfium; mediums—19w
`signal, myommial junctinn mac; mediumhigh sigma}.
`myomtrium; and no signal. peripharal adipose tisaue.
`The: area (after: messenmhapefi} a!“ high signal infirmity
`secs: 3: the left; in Figs ‘2“«12 is mine in thx: bladder. At:
`nvary is partiafly visible {hottum right) in Figs “3-H.
`
`Jaw-Ra! {if Endae‘finafagy (19%} 13$. 233%”
`
`InnoPharma Exhibit 1057.0008
`
`

`

`I'l \l'l
`
`Ifflm-H u] If] ).\'._‘_ 735’“ m mfny nwrnn'mumg Hmull't‘lw
`
`I
`
`\I m Kl N‘ {ind others
`
`
`
`.|.u~n:.- r Iizu
`
`Jumnm' a»: Urdu. mud-«:1 ci‘MH HR. III? 3'”
`
`12
`
`InnoPharma Exhibit 1057.0009
`
`