Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`0022-3565/87/2412-0401$OO.OO/0
`Jouamt(cid:1)i.
`or
`THE
`PHAassAcoL0GY
`CopyrightO
`1987 by The American
`
`AND
`Society
`
`EXPERIMENTAL
`for Pharmacology
`
`$02.OO/O
`THERAPEUTICS
`and Experimental
`
`Therapeutics
`
`Vol.
`Printed
`
`241, No.
`in U.S.A.
`
`2
`
`Pharmacokinetic
`Antiprogesterone
`
`of the Antiglucocorticoid
`Properties
`Steroid RU 486 in Man1
`
`and
`
`SHINICHI
`GEORGE
`
`LYNNETTE
`KAWAI,2
`P. CHROUSOS
`
`K. NIEMAN,
`
`DAVID
`
`0. BRANDON,
`
`ROBERT
`
`UDELSMAN,
`
`D. LYNN
`
`LORIAUX
`
`and
`
`Developmental
`
`Accepted
`
`for publication
`
`Endocrinology
`January
`
`National
`Branch,
`21 , 1987
`
`Institute
`
`of Child Health and Human Development,
`
`National
`
`Institutes
`
`of Health,
`
`Bethesda,
`
`Maryland
`
`-pro-
`
`ABSTRACT
`1 (cid:1)-[4-dimethyIamino
`RU 486
`7a-[1
`phenylj-1
`(1 7fl-hydroxy-1
`pynyljestra-4,9-dien-3-one)
`a clinically
`useful
`glucocorticoid
`iS
`The authors
`and progesterone
`studied
`the
`pharma-
`antagonist.
`of this drug
`in normal
`volunteers
`and
`patisnts
`cokinetic
`pro(cid:1)es
`a rat progesterone
`syndrome
`using
`radioreceptor
`Cushing’s
`with
`to those
`Obtained
`with
`gave
`This
`similar
`assay.
`assay
`values
`rat
`a
`assay.
`After
`single
`oral
`dose
`glucocorticoid
`radioreceptor
`1 1) to normal
`of 25 mg/kg
`1 1) or 1 0 mg/kg
`volunteers,
`(n =
`(n =
`plasma
`of progesterone
`COnCentratiOnS
`receptor-reactive
`mate-
`maximal
`hal
`± 288
`(mean
`and
`± S.D.)
`517
`reached
`levels
`of 754
`183
`1Lg/dI. This
`occurred
`at
`3.1±
`1.9
`and
`2.5
`±
`±
`1.0
`h,
`respectively.
`The
`respective
`apparent
`plasma
`half-lives
`were
`
`a
`
`7.0
`
`7.7
`
`20.6
`19.2
`Cushing’s
`with
`patients
`and
`Four
`h.
`±
`±
`relatively
`had
`mg/kg/day)
`(1 0-20
`chronically
`syndrome
`treated
`of
`from 506
`ranging
`reactivity
`receptor
`levels
`plasma
`constant
`circulating
`of
`Chromatographic
`(cid:1)g/dI.
`to
`I 1 84
`characterization
`corresponded
`the active
`fraction
`that
`showed
`reactivity
`receptor
`to RU 486
`N-mono-
`and
`N-didemethylated
`hydrophilic
`and
`the
`delty
`dose
`0.5% of
`in
`was
`excreted
`than
`metabolites.
`Less
`of
`these
`the
`The
`patients
`receptor
`reactivity.
`unne
`two
`as
`competed
`extensively
`drug
`bound
`to
`circulating
`albumin,
`which
`of
`with
`the
`glucocorticoid
`receptor
`intact
`human
`mononuclear
`for
`in a concentration-dependent
`manner.
`[3HJRU 486
`leukocytes
`
`its
`
`of
`
`(Phi-
`and
`
`antagonist
`and progesterone
`glucocorticoid
`A synthetic
`libert
`1981;
`1983; Moguilewsky
`Schreiber
`et
`at.,
`et
`at.,
`RU 486
`(17(cid:1)-hydroxy-11(cid:1)9-[4-dimethylamino
`Philibert,
`1984),
`cm-
`phenyl]-17a-[1-propynyl]estra-4,9-dien-3-one),
`has major
`ical
`in the
`treatment
`of hypercortisolism
`(Nieman
`a!.,
`uses
`et
`1982;
`and
`in the
`regulation
`of
`fertility
`(Herrman
`a!.,
`et
`1985a)
`et a!., 1985b).
`Kovacs
`et a!., 1985; Nieman
`et a!.,
`1984;
`Schaison
`suggested
`a long
`plasma
`half-
`Preliminary
`limited
`studies
`have
`this
`for
`1985;
`Shoupe
`1985).
`a!.,
`a!.,
`life
`drug
`et
`et
`(Deraedt
`studies
`were
`in a small
`number
`of
`subjects
`These
`performed
`used
`either
`and
`tracer
`methods
`or measurement
`of
`im-
`drug
`munoreactivity.
`pharmacoki-
`the
`examine
`to
`was
`study
`this
`of
`The
`purpose
`and
`glucocorti-
`progesterone
`using
`of RU 486
`netic
`properties
`of receptor-reactive
`RU
`plasma
`concentrations
`coid RRAs.
`The
`in normal
`subjects
`and
`its metabolites
`were
`determined
`486
`in four
`patients
`with
`single
`oral
`doses
`the
`and
`after
`drug
`with
`RU 486.
`The
`Cushing’s
`syndrome
`chronically
`in
`plasma
`proteins
`and
`the
`with
`interactions
`vitro
`glucocorticoid
`human
`leukocytes
`were
`also
`studied.
`
`of
`treated
`of RU
`receptor
`of
`
`486
`intact
`
`1 Presented
`
`in part
`
`at
`
`the American
`
`Federation
`
`for Clinical
`
`Research
`
`National
`
`Washington,
`Meeting,
`S Supported
`in
`Therapeutics
`and
`
`part
`the
`
`1986.
`D.C.,
`the
`Japanese
`by
`Rouseel-UCLAF
`
`Society
`Institute.
`
`of Clinical
`
`Pharmacology
`
`and
`
`Methods
`
`from the U.S. Na-
`study was
`for the
`obtained
`Subjects.
`Approval
`and Biologics
`the Human
`Investigation
`for Drugs
`and
`Bureau
`tional
`for the National
`Institutes
`of Health.
`The nature
`and
`aims
`Committee
`of
`the
`study were
`the
`subjects who gave
`their
`consent.
`explained
`to all
`The
`clinical
`at
`the Clinical
`Center
`of
`the
`were
`performed
`studies
`National
`Institutes
`of Health.
`ages 20 to 49 years
`five normal women,
`men and
`Seventeen
`normal
`(table
`RU 486
`(mean
`S.D.,
`8.6),
`1). The
`drug,
`received
`±
`30.7
`±
`from
`Roussel-UCLAF
`formulated
`into
`50-mg
`tablets,
`was
`a
`gift
`It was
`orally
`at 8:00
`or 8:00
`administered
`(Paris,
`France).
`AM.
`P.M.
`fasting. Half
`the
`at
`least
`8 h of
`of
`25 mg/kg
`after
`volunteers
`received
`RU 486,
`half
`received
`10 mg/kg.
`Blood
`and
`were
`drawn
`from
`samples
`vein immediately
`at 0.5,
`antecubital
`1, 1.5,
`2, 2.5,
`3, 4, 5, 6,
`before
`and
`24 h after
`plasma
`samples were
`16 and
`7, 8,
`administration.
`Separated
`kept
`at
`-70#{176}Cuntil
`assay.
`frozen
`with RU
`chronically
`treated
`syndrome
`Cushing’s
`Four
`patients
`with
`RU 486 measured
`of receptor-reactive
`plasma
`concentrations
`had
`486
`every
`6 h from 8:00
`on several
`occasions.
`They
`received RU 486 tablets
`of these
`patients
`The
`age,
`sex, diagnosis
`RU 486 doses
`and
`daily
`A.M.
`with
`serum
`given
`in table
`2. Patient
`3 had chronic
`are
`failure
`renal
`excretion
`of RU 486
`of 6 mg/dl.
`Twenty-four-hour
`urinary
`creatinine
`2 and
`3 and
`of
`the drug
`in patients
`c(cid:1)rebrospinal
`concentration
`fluid
`was
`Sampling
`4 were
`also mdasured
`reactivity.
`in patient
`as
`receptor
`at
`least
`2 weeks
`treatment
`with RU 486.
`of
`the onset
`performed
`after
`Steroids.
`[‘H)R5020
`([17a-methyl-’HJ17,21-dimethyl-19-nor-
`pregna-4,9-diene-3,
`20-dione;
`[‘H]promegestone;
`87.0
`Ci/mmol),
`
`unla-
`
`ABBREVIATIONS:
`
`RRA,
`
`radioreceptor
`
`assay;
`
`PBS,
`
`phosphate-buffered
`
`saline.
`
`401
`
`

`

`Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`402
`
`Kawal
`
`et al
`
`Vol.241
`
`TABLE 1
`Pharmacokinetic
`
`properties
`
`of RU 486 after
`
`single
`
`oral
`
`administration
`
`In normal
`
`volunteers
`
`Case
`
`1
`2
`3
`4
`5
`6
`7
`8
`9
`10
`11
`
`Mean
`
`± S.D.
`
`12
`13
`14
`15
`16
`17
`18
`19
`20
`21
`22
`± S.D.
`
`Mean
`
`(cid:1)se48(cid:1)
`
`mg/kg
`25
`25
`25
`25
`25
`25
`25
`25
`25
`25
`25
`
`10
`10
`10
`10
`10
`10
`10
`10
`10
`10
`10
`
`Actniristration
`
`A.M.
`
`P.M.
`
`P.M.
`
`P.M.
`AM.
`P.M.
`A.M.
`
`P.M.
`AM.
`A.M.
`A.M.
`
`AM.
`AM.
`A.M.
`A.M.
`
`P.M.
`
`P.M.
`
`P.M.
`
`P.M.
`
`P.M.
`A.M.
`AM.
`
`A9e
`
`yr
`26
`27
`34
`38
`23
`20
`36
`45
`25
`30
`28
`± 7.4
`20
`25
`29
`27
`39
`22
`49
`43
`41
`27
`22
`± 9.9
`
`30.2
`
`31.3
`
`a Maxvnum
`S (cid:1)
`
`plasma
`concentration
`
`of receptor-reactive
`concentration
`of receptor-reactive
`RU 486
`
`RU 486.
`24 h after
`
`adminiStratiOn.
`
`Sex
`
`Appaint T,(cid:1)
`
`Pe(cid:1)
`
`Tine
`
`M
`M
`M
`F
`F
`M
`M
`F
`M
`M
`M
`
`F
`M
`M
`M
`M
`M
`M
`M
`F
`M
`M
`
`19.2
`
`20.6
`
`h
`9.5
`22.6
`23.6
`33.7
`25.8
`13.8
`20.0
`13.5
`13.4
`20.1
`15.5
`± 7.0
`10.6
`25.7
`25.9
`32.8
`12.8
`12.5
`31.5
`21.2
`16.3
`21.6
`15.2
`± 7.7
`
`h
`3
`2.5
`2
`1.5
`4
`2
`4
`2
`8
`4
`1.5
`3.1 ± 1.9
`3
`2.5
`2
`2.5
`4
`0.5
`1.5
`2.5
`4
`2
`2.5
`2.5 ± 1.0
`
`C max
`
`(cid:1)zg/dl
`326
`877
`533
`877
`815
`846
`583
`595
`589
`1441
`815
`
`754
`
`± 288
`420
`714
`162
`249
`602
`489
`761
`536
`517
`639
`602
`517 ± 183
`
`C 24 h’
`
`308
`
`(cid:1)g/dl
`75
`376
`483
`420
`370
`244
`219
`188
`257
`539
`219
`± 140
`86
`271
`90
`146
`197
`135
`423
`203
`160
`214
`158
`189 ± 100
`
`2
`TABLE
`Concentrations
`Pt., patient; Conc.,
`
`of receptor-reactive
`concentration;
`CSF,
`
`fluids
`RU 486 in biologic
`cerebrospinal
`fluid; CA, carcinoma;
`
`with Cushing’s
`from patients
`ACTh,
`adrenocorticotropin.
`
`syndrome
`
`treated
`
`chronically
`
`with RU 486
`
`Pt
`
`Age Sex
`
`D(cid:1)gno(cid:1)s
`
`Doseof RU486
`
`PlasmaConc.
`
`yr
`25 M
`36 M
`45 F
`40 F
`
`1
`2
`3
`4
`#{149}Mean ± S.D.
`of samples.
`S Number
`
`ACTH
`ectopic
`ectopic ACTH
`adrenal CA,
`renal
`ectopic
`ACTH
`
`failure
`
`mg/kg/day
`20
`15
`10
`10
`
`(cid:1)g/dl
`506 ±
`(n =
`85’
`22)b
`± 27 (n = 4)
`910
`± 152 (n = 4)
`629
`1 184 ± 259 (n = 3)
`
`gig/kg/day
`
`pg/dI
`
`49
`29
`
`-
`
`-
`
`-
`42
`
`[3HJ
`and
`Ci/mmol)
`40.6
`([1,2-3H]cortisol;
`[3H]cortisol
`R5020,
`beled
`35.0 Ci/mmol)
`([6,7-3H]dexamethasone;
`dexamethasone
`obtained
`were
`from New England
`Nuclear
`(Bedford, MA).
`[3HJRU 486 ([6,7-3H]RU
`RU 486, RU 42 633
`50.6 Ci/mmol),
`486;
`unlabeled
`(N-monodemeth-
`ylated RU 486)
`and RU 42 848 (N-didemethylated
`RU 486) were gifts
`from Roussel-UCLAF.
`All other
`Sigma
`were
`steroids
`purchased
`from
`Louis, MO).
`Chemical
`Co.
`(St.
`preparation.
`cytosol
`Rat
`uterus
`and
`thymus
`Ovariectomized
`and adrenalectomized
`estrogen-treated
`rats
`(140-160
`female
`Holtzman
`of
`used
`as
`a source
`Co., Madison, WI) were
`g; Holtzman
`uterine
`4th
`On the 3rd and
`thymus
`glucocorticoid
`progesterone
`and
`receptors.
`oil was
`administered
`in sesame
`surgery,
`estradiol
`(1.5
`gig)
`days
`after
`i.m. Uteri
`and
`obtained
`on
`the
`5th
`day.
`They
`were
`thymuses
`were
`in ice-cold
`homogenization
`trimmed,
`individually
`placed
`rinsed
`and
`0.5 mM dithiothreitol,
`[10 mM Tris-HC1,
`1.5 mM EDTA,
`buffer
`glycerol
`(v/v),
`pH 7.4J. After
`homogenize-
`20 mM sodium
`molybdate,
`(OMNI-MIXER
`tion with a motor-driven
`homogenizer
`17105;
`E.
`I. du
`Pont de Nemours & Co., Newton,
`CT),
`cytosol
`prepared
`fractions
`were
`et a!., 1976).
`(Rifla
`by ultracentrifugation
`as previously
`These
`described
`within
`at
`kept
`-70C.
`The
`cytosol
`were
`frozen
`fractions
`were
`used
`month.
`The protein
`concentrations
`of
`the
`and
`thymus
`uterus
`cytosol
`and 5.0 ± 0.2 mg/ml,
`were
`4.1 ± 0.1 (mean
`± S.D.)
`respectively
`fractions
`(Bradford,
`1976).
`
`3%
`
`1
`
`in
`
`the
`of
`[3H]R5020
`and
`[‘HI
`
`or
`
`assays were performed
`These
`RRAS.
`extraction
`ethanol
`after
`fraction
`and
`Uterus
`as mentioned
`below.
`cytosol
`sample,
`were used for the progesterone
`RRA.
`fraction
`cytosol
`Thymus
`RRA.
`were
`employed
`for the glucocorticoid
`dexamethasone
`added
`to denature
`Four milliliters
`to 100 (cid:1)d
`ofplasma
`ofethanol
`were
`± 2% (mean
`plasma
`proteins.
`The
`recovery
`[‘H]RU
`486 was
`102
`of
`±
`with
`20 (cid:1)il of
`S.D.).
`Three
`to 10 (cid:1)il of ethanol
`solution
`were
`then mixed
`was
`propylene
`glycol
`in a 12 x 75-mm
`Ethanol
`evaporated
`glass
`tube.
`[‘HJR5020
`a Buckler
`vortex-evaporator,
`(cid:1)l of either
`and
`100
`using
`concentration
`in the homogenization
`buffer
`(final
`[3H]dexamethasone
`100 (cid:1)il of
`cytosol
`or thymus
`uterus
`6 nM) with
`either
`in assay
`tube,
`were added
`to the
`tube. The mixture
`was
`for 18 h at 4#{176}C.
`incubated
`The
`receptor-bound
`3H-steroid
`was
`from the
`free with
`0.5 ml
`separated
`of dextran-coated
`solution
`(0.2% dextran,
`Pharmacia
`T 70,
`charcoal
`were
`and
`samples
`2% charcoal,
`Sigma
`Norit
`A,
`in the
`buffer).
`The
`for
`then
`(4#{176}C)for 10 main at 3000
`and
`15 rain
`incubated
`centrifuged
`the
`supernatant
`was COUnted
`10 ml of counting
`fluid
`rpm.
`Half
`New England
`Nuclear).
`(Aquasol;
`the proges-
`of
`The
`RRAs were
`sensitivities
`performed
`in
`duplicate.
`2.5 x
`10’#{176}M,
`5 x
`glucocorticoid
`RRAs were
`10’#{176}and
`and
`terone
`containing
`The
`normal
`plasma
`samples
`different
`respectively.
`three
`concentrations
`of RU 486 were measured
`by these
`known
`assay
`pro-
`The
`values
`in both
`the
`cedures.
`RRAs
`were
`almost
`identical
`with
`
`(cid:1)
`

`

`1987
`
`Pharmacokinetics
`
`of RU 486
`
`in Man
`
`403
`
`in
`
`with
`
`of vans-
`coefficients
`intra-assay
`The
`and
`values.
`interassay
`expected
`9 and
`respectively.
`Plasma
`concentrations
`of
`17%,
`less
`tion were
`than
`RU 486 were undetectable
`before
`administration
`in normal
`volunteers
`and
`patients
`with
`Cushing’s
`syndrome.
`Thin-layer
`chromatography.
`ethanol,
`with
`extraction
`After
`a patient
`with
`(case
`2) and
`volunteer
`plasma
`samples
`of a normal
`on thin-layer
`chromatographed
`1) were
`Cushing’s
`syndrome
`(patient
`Separation
`Inc.,
`plate; Whatman
`Chemical
`chromatography
`(LK6DF
`NJ)
`a
`cyclohexane-ethylacetate
`Clifton,
`system
`at
`22#{176}C.
`(1:1)
`for
`were
`and
`assessed
`Scraped
`1.1-cm
`fractions
`extracted
`ethanol
`Recovery
`was
`reactivity.
`progesterone
`and
`receptor
`glucocorticoid
`The
`was used
`checked
`by addition
`of 10,000
`cpm oftracer.
`system
`same
`for examination
`or purification
`of
`RU 486, unlabeled
`RU 486
`tracer
`ethanol-extracted
`RU 486 tablets.
`The R(cid:1) values
`of RU
`and
`standard
`of
`these
`were
`identical.
`preparations
`486
`from each
`three
`The
`of
`properties
`Equilibrium
`dialysis.
`protein-binding
`plasma
`dialysis
`at 37#{176}C
`RU 486 and cortisol were
`evaluated
`by an equilibrium
`as described
`by Kley et al.
`plasma
`(obtained
`(1977)
`using
`normal
`human
`of
`and 4.5 g/dl
`from the National
`Institutes
`of Health
`Blood
`Bank)
`albumin
`(Sigma)
`solution
`PBS,
`containing
`0.9% sodium chlo-
`human
`pH 7.4. The
`0.2 nM.
`final
`ride,
`concentration
`of
`‘H-steroid
`was
`of albumin
`on the binding
`The
`cell-binding
`study.
`Intact
`effect
`[‘H]RU
`486 and [3H)dexamethasone
`to intact
`human mononuclear
`of
`was
`examined
`using
`a previously
`described
`system
`(Murak-
`leukocytes
`coat was obtained
`from the National
`ami
`1979).
`The
`et a!.,
`human
`buffy
`of Health
`Blood Bank. Mononuclear
`were
`sepa-
`Institutes
`leukocytes
`Bethesda, MD).
`rated
`lymphocyte
`separation
`using
`medium
`(Bionetics,
`PBS,
`cells were
`suspended
`in RPMI-1640
`washing
`with
`After
`media
`or without
`human
`albumin
`and adjusted
`to 1.0 x iO(cid:1) per milliliter.
`with
`They were
`then
`incubated
`with
`0.1
`to
`20
`nM [3HIRU
`486
`or
`[‘H]
`5% CO2 and 95% 02 for 18 h at 22#{176}Cin a shaking
`dexa.methasone
`under
`was determined
`by the
`addition
`of a
`incubator.
`Nonspecific
`binding
`of respective
`three wash-
`unlabeled
`steroid. After
`500-fold
`molar
`excess
`ings with
`ice-cold
`PBS,
`the
`pellets
`were
`resuspended
`in PBS
`and
`the
`The
`cell Viability
`by trypan
`blue dye
`counted.
`determined
`radioactivity
`was more
`95% in various
`conditions
`of media.
`The
`than
`equilibrium
`constant
`of RU 486 for the human mononuclear
`dissociation
`leukocyte
`was determined
`by this method
`(with no added
`glucocorticoid
`receptor
`with
`that
`of dexamethasone,
`albumin)
`compared
`to
`0.32
`nM,
`as
`whichwasl.1
`nM.
`Analyses
`of
`data
`curves
`elimination
`to the
`24-h
`sample
`of the drug.
`half-life
`Student’s
`paired
`
`be
`
`linear
`Least-squares
`of plasma RU 486 levels
`was used
`for determination
`
`regression
`(logarithm)
`of
`the
`
`for the
`analysis
`from the peak
`apparent
`plasma
`
`t
`
`test was used for comparison
`
`of RRAs.
`
`Results
`
`that
`
`thymus
`x
`10(cid:1);
`
`Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`dose
`oral
`table
`3 and
`who
`subjects
`S.D.)
`7.0
`(±
`±
`plasma
`concen-
`of 25 and
`10 mgI
`183
`and
`517
`±
`RU
`486
`levels
`RU 486 was
`in these
`or be-
`
`of
`total
`
`reactivities
`fractions
`we recovered
`5,
`receptor
`R(cid:1)
`4). The
`0.26
`0.33,
`5,
`fractions
`the
`between
`The
`fractions.
`from a normal
`had
`receptor
`
`fraction
`dialysis
`of unla-
`(cid:1)g/&)
`bound
`human
`
`This
`94.0%.
`of unlabeled
`
`RU 486
`a single
`after
`receptor-reactive
`progesterone
`in figure
`are
`shown
`in normal
`subjects
`of
`the
`drug
`values
`in normal
`apparent
`half-life
`1. The mean
`25 and
`10 mg/kg
`of RU 486 were
`19.2
`received
`and
`20.6
`± 7.7
`h,
`respectively.
`The
`maximum
`trations
`of RU 486
`administration
`after
`oral
`kg of
`were
`754
`(mean
`the
`288
`S.D.)
`±
`±
`drug
`g(cid:1)g/dl,
`respectively.
`In
`both
`groups,
`plasma
`reached
`the maximum
`about
`3 h after
`ingestion.
`still measurable
`24 h after
`ingestion.
`No
`differences
`were
`observed
`between
`men
`and women
`measurements
`tween morning
`and
`evening
`administration.
`in
`RU 486
`The
`plasma
`concentrations
`of
`receptor-reactive
`2.
`in table
`patients
`with
`Cushing’s
`syndrome
`are
`shown
`four
`12 h (8:00
`Patient
`1 had RU 486 measured
`every
`30 mm over
`was
`506
`(cid:1)g/
`The mean
`plasma
`concentration
`A.M.-8:00
`P.M.).
`24-h
`The
`dl, with
`a coefficient
`of
`variation
`of
`16.8%.
`urinary
`excretion
`of RU 486
`receptor
`reactivity
`was measured
`in pa-
`tients
`2 and
`3. Less
`0.5% of
`the
`daily
`dose was
`excreted
`than
`in the
`urine.
`In patient
`4, cerebrospinal
`fluid
`concentration
`RU 486
`receptor
`reactivity
`was
`approximately
`4% of
`the
`plasma
`receptor-reactive
`RU 486
`concentration.
`of
`in
`plasma.
`Receptor
`Metabolites
`RU
`486
`486
`and
`[3H]RU
`recoveries
`in the
`chromatographed
`shown
`ofplasma
`samples
`are
`in figure
`4. Although
`the
`highest
`amount
`of
`[3HJRU
`486
`in
`fraction
`reactivity
`also
`found
`in
`fractions
`4 and
`2 (fig.
`was
`of RU 486, RU 42
`and RU 42
`848 were
`values
`633
`and
`0.14,
`which
`corresponded
`to
`chromatographic
`4 and
`2,
`respectively.
`There
`was
`no
`difference
`progesterone
`and
`glucocorticoid
`RRAs
`of
`these
`chromatographed
`pretreatment
`plasma
`samples
`subject
`a patient
`with
`Cushing’s
`syndrome
`and
`reactivities
`less
`than
`the
`blank.
`bound
`The
`Plasma-bound
`fraction
`of RU
`by equilibrium
`[3H]RU
`486
`in plasma
`as determined
`of
`concentrations
`94.3
`± 1.8% (mean
`± S.D.).
`High
`was
`cortisol
`RU 486
`(up to 3000
`(cid:1)zg/dl
`ofplasma),
`beled
`(700
`on
`percent
`or progesterone
`had
`no
`effect
`(400
`(cid:1)g/&)
`The
`[3HJRU
`486.
`bound
`fraction
`of
`[3H]RU
`486
`to
`albumin
`solution
`(4.5
`g/dl)
`was
`91.8
`± 0.7%.
`The mean
`of bound
`[3H]cortisol
`in plasma
`was
`was
`reduced
`to 64.5% by addition
`oflarge
`amounts
`cortisol
`(700
`(cid:1)g/dl).
`albumin
`to mononuclear
`binding
`on RU 486
`of
`Effect
`the
`albumin
`on
`of
`effect
`shows
`the
`5
`leukocytes.
`Figure
`binding
`486
`or
`[3H]dexamethasone
`to human
`of 20 nM [3H]RU
`Total
`and
`specific
`binding
`of
`[3H]RU
`mononuclear
`leukocytes.
`with
`increasing
`albumin
`concentra-
`486
`significantly
`decreased
`tions.
`Compared
`with
`control
`(0% albumin),
`only
`8.1% of RU
`486
`bound
`specifically
`to cells when
`the media
`contained
`4.5%
`albumin.
`In contrast,
`the
`specific
`binding
`of
`[3Hjdexametha-
`sone
`to
`intact
`cells
`was minimally
`by
`the
`albumin
`affected
`concentration.
`The
`were
`observed
`using
`the
`lower
`same
`results
`concentrations
`of 3H-steroids.
`
`486.
`
`steroids
`various
`by
`[3H]R5020
`of
`displacement
`The
`RRAS.
`concentra-
`1. The
`in figure
`progesterone
`in the
`shown
`is
`RRA
`the
`[3H]R5020
`binding
`to
`that
`inhibited
`50% of
`tions
`uterus
`x
`2.9
`10(cid:1);
`(in molar)
`RU
`486,
`progesterone
`receptor
`were
`estradiol,
`8.5
`1.1
`x
`i0(cid:1);
`10(cid:1);
`progesterone,
`and
`x
`R5020,
`7.5
`cross-reactivity
`in this
`system.
`x
`iO-(cid:1).
`Cortisol
`had
`negligible
`[3Hjdexamethasone
`various
`by
`of
`The
`displacement
`curves
`concentrations
`The
`steroids
`were
`inhibited
`also
`obtained.
`binding
`to the
`glucocorti-
`of
`[3H]dexamethasone
`50%
`RU 486,
`2.0
`dexametha-
`coid
`receptor
`were
`(in molar)
`1.0 x iO(cid:1).
`sone,
`1.2 x 10_a;
`and
`cortisol,
`includ-
`samples
`The
`concentrations
`of RU 486
`in 20 plasma
`two
`from a patient
`with
`Cushing’s
`syndrome
`(patient
`ing
`were measured
`by
`both
`the
`progesterone
`and
`glucocorticoid
`2). No
`significant
`difference
`between
`these
`RRAs
`RRAs
`(fig.
`found
`(P > .2).
`was
`Pharmacokinetic
`(mean
`±
`S.E.)
`
`study.
`individual
`
`and
`
`plasma
`The
`pharmacokinetic
`
`concentrations
`properties
`
`1)
`
`of
`
`Discussion
`
`The mean
`reactive
`RU
`Because
`the
`of
`sampling,
`
`apparent
`486
`in
`terminal
`however,
`
`receptor-
`ofprogesterone
`half-life
`plasma
`h.
`about
`20
`our
`normal
`volunteers
`was
`log-linear
`phase
`was
`reached
`at 24 h
`not
`the
`actual
`half-life
`value may
`be
`consid-
`
`

`

`Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`404
`
`awa(cid:1)
`
`et al
`
`Vol.241
`
`of various
`1. CompetItion
`Fig.
`curves
`(displacement)
`receptor.
`uterus
`progesterone
`rat
`steroids
`for
`the
`as percentage
`of ccntrd
`bind-
`Results
`are expressed
`Ing in the absence
`ofcompetftors.
`Values
`shown were
`determined
`in duplicate.
`
`STEROID CONCENTRATION
`
`(M)
`
`et
`
`to
`
`at.,
`
`-..
`
`0 l
`
`U
`
`0 O
`
`:D
`
`Oj::
`
`04
`
`-I
`
`I
`
`200
`
`400
`
`600
`
`800
`
`PROGESTERONE
`REACTIVE
`-RU
`
`RECEPTOR
`486
`((cid:1)g/dI)
`
`of RU 486 measured
`concentrations
`plasma
`between
`Fig. 2. Correlation
`and glucocorticold
`and triangles
`RRAs. Cwcles
`by progesterone
`k(cid:1)dlcate
`a patient with Cushlng’s
`from normal
`volunteers
`and
`plasma
`samples
`RRAs was
`syndrome.
`respectively.
`No
`significant
`difference
`between
`.2).
`found
`(P >
`
`0
`
`2
`
`4
`
`6
`
`8
`
`10
`
`12
`
`TIME
`
`16
`
`IS
`
`20
`
`22
`
`24
`
`14
`
`(h)
`
`Fig.
`concentrations
`3. Plasma
`RU 486 after oral administration
`volunteers.
`
`(PRR-)
`receptor-reactive
`of progesterone
`of 25 or 10 mg/kg
`of the drug to normal
`
`longer.
`erably
`to that
`reported
`(1985).
`et a!.
`with
`compared
`
`is similar
`value
`this
`caveat,
`potential
`this
`Despite
`by Shoupe
`and implied
`by Deraedt
`(1985)
`et a!.
`of RU 486
`is
`long
`plasma
`apparent
`The
`half-life
`those
`of other
`steroids,
`which
`range
`from mini-
`
`3 to 5 h (dexameth-
`(progesterone)
`ute8
`a!.,
`1966)
`(Little
`the
`approximation
`of
`closest
`The
`(Kawai
`et
`asone)
`1985).
`we
`can make
`in the
`is
`of RU 486
`rate
`that
`clearance
`metabolic
`the
`ab-
`If we
`assume
`that
`with
`Cushing’s
`syndrome.
`patients
`tract
`from the
`was
`100%,
`of RU 486
`gastrointestinal
`sorption
`constant
`infusion
`the metabolic
`clearance
`rate
`computed
`by the
`and
`4.0
`model
`(Tait
`1961)
`would
`range
`between
`0.84
`aL,
`et
`2.0)
`liters/kg/day.
`the
`of
`receptor-
`(mean,
`Because
`amount
`RU 486
`excreted
`in the
`of our
`less
`urine
`patients
`was
`reactive
`0.5% of
`the
`we
`suggest
`that
`than
`total
`daily
`dose,
`kidneys
`in the
`clearance
`and
`play
`an
`insignificant
`role
`this
`receptor-reactive
`metabolites.
`of RU 486 may
`half-life
`The
`long
`apparent
`plasma
`explain
`studies
`using
`other
`long
`biologic
`effect
`the
`single
`oral
`in
`seen
`of RU 486
`in man.
`fi-lipotropin
`(Gaillard
`cortisol,
`Plasma
`doses
`adrenocorticotropin
`and
`1984;
`Bert,agna
`et
`at.,
`et
`at.,
`1984)
`starting
`increased,
`but
`only
`levels
`clearly
`(Gaillard
`et a!.,
`1984)
`and
`only
`at hours
`the
`drug
`5 to
`6 h after
`of
`administration
`endogenous
`corticotropin-
`of presumably
`elevated
`(morning)
`of
`releasing
`hormone
`secretion.
`In addition,
`the
`concentrations
`5th-
`these
`hormones
`in plasma
`and
`24-h
`urinary
`17-ketogenic
`mid
`excretion
`higher
`than
`control
`in the
`2 days
`after
`remained
`a 6-mg/kg
`of RU 486
`(Gaillard
`dose
`a!.,
`1984).
`In nonhuman
`et
`primates,
`antagonist
`activity
`early
`RU
`486
`was
`observed
`as
`0.5 h after
`i.m.
`injection
`ofthe
`drug
`in the
`early morning
`(Healy
`The
`different
`time
`and mode
`of administration
`1985).
`et at.,
`486 may
`account
`for
`the
`observed
`differences
`RU
`in
`its
`of action
`in man
`and monkey.
`half-life
`plasma
`apparent
`long
`of
`One
`the
`reasons
`for
`the
`(and
`possibly
`binding
`of
`RU 486 may
`be the
`extensive
`drug
`this
`proteins.
`previously
`shown
`We
`have
`its metabolites)
`to plasma
`globulin
`to corticosteroid-binding
`bind
`that
`RU 486
`does
`not
`globulin
`(Nieman
`or
`testosterone-estradiol-binding
`et
`at.,
`suggested
`that
`a-i-acid
`have
`1985a).
`et
`at.
`(1986)
`Philibert
`of
`the
`plasma
`RU
`486
`some
`for
`account
`may
`glycoprotein
`binding.
`Because
`[3HJRU
`486
`to
`of
`binding
`specific
`intact
`the
`leukocytes
`mononuclear
`by
`severely
`inhibited
`human
`presence
`(4.5%)
`albumin
`concentration
`ofphysiologic
`ofhuman
`was much
`that
`of
`[3Hjdexamethasone
`less
`affected),
`(whereas
`we believe
`that
`the
`binding
`of RU 486
`to plasma
`albumin
`The
`concentration
`of
`receptor-
`pharmacologically
`important.
`the
`in the
`cerebrospinal
`fluid
`of
`patient
`with
`reactive
`486
`RU
`4% of her
`concurrent
`Cushing’s
`syndrome
`was
`approximately
`plasma
`concentration,
`in agreement
`the
`notion
`that
`only
`
`the
`drug
`
`of
`
`its
`
`onset
`
`as
`
`of
`
`of
`
`is
`
`was
`
`with
`
`

`

`Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`PROGESTERONE
`
`RADSORECEPTOR
`
`ASSAY
`
`Gl.UCOCORT1COID
`
`RADIORECEPTOR
`
`ASSAY
`
`Pharmacokin#{149}tIcs
`
`of RU 486
`
`in Man
`
`405
`
`1987
`
`Do
`
`00
`40‘law
`
`column)
`(open
`reactlvfties
`4. Receptor
`Fig.
`of plasma
`in the chromatographed
`frections
`(at peak
`samples
`from a normal
`volunteer
`24 h after
`of
`the
`level
`and
`admInIStratiOn
`drug)w(cid:1)d
`a patient with Cushing’s
`syndrome
`RU 486 orally. Shaded
`column
`who
`received
`represents
`percent
`recovery
`of
`[‘H]RU
`the
`486 in eech
`fraction.
`Frections
`are numbered
`from the origin
`of
`the
`thin-layer
`chromatog-
`RU 486 (RU 42
`rephy. N-rnonodemethylated
`RU 486
`(RU 42
`633) w(cid:1)d N-didemethylated
`4 and 2,
`848)
`correspond
`to
`frections
`i’s-
`
`ww
`
`Uw
`
`w
`
`1
`
`2
`
`3
`
`4
`
`5
`
`6
`
`7
`
`S
`
`FRACTION
`
`101112131415
`S
`NUMBER
`
`123
`
`4
`
`7
`6
`5
`FRACTiON
`
`5
`
`101112131415
`9
`NUNSER
`
`and
`the
`progesterone
`tivity. We
`suggest
`that
`in man
`is
`due
`primarily
`that
`hydrophilic
`products
`i1(cid:1)-phenyl
`chain.
`
`glucocorticoid
`circulating
`to RU
`lack
`one
`
`and
`receptor
`receptor
`RU 486
`secondarily
`486
`and
`or two methyl
`groups
`
`bioac-
`some
`reactivity
`to
`its
`in the
`
`Acknowledgments
`
`authors
`The
`RU 486
`this
`for
`Terry M. Wood
`
`for generously
`Rouseel-UCLAF
`like
`would
`thank
`to
`study. We are also grateful
`to Mr. Gerald
`W.
`Peden
`for
`their
`superb
`technical
`assistance.
`
`providing
`and Ms.
`
`References
`
`J. P., HussoN,
`glucocorticoid
`
`J. M. AND GmAiw,
`F.:
`in man.
`J. Clin.
`action
`
`quantitation
`the
`for
`ofprotein-dye
`
`of micro-
`binding.
`Anal.
`
`C., LuT0N,
`X., BSRTAGNA,
`BERTAGNA,
`RU 486
`new steroid
`inhibits
`analog
`The
`EndocrinoL
`Metab.
`25-28,
`1984.
`59:
`BatDrolw,
`M. M.: A rapid and sensitive
`method
`of protein
`utilizing
`the principle
`gram quantities
`Biochem.
`72:
`248-254,
`1976.
`C., BusIGNv, M., Ciumizr,
`C., MOUREN,
`P., Cous’ry,
`Dmt*zivr,
`R., BONNAT,
`of RU
`J.: Pharmacokinetics
`M.,
`PmLmnRr,
`D.,
`PorTrait,
`J. AND
`S(cid:1)usoN,
`In The Antiprogestin
`Steroid
`RU 486 and Human
`ad.
`Control,
`Fertility
`486.
`S. J. Segal,
`New York,
`Press,
`1985.
`pp.
`by E. E. Baulieu
`and
`103-122,
`Plenum
`R. C., RIONDEL,
`W. AND BAULIEU,
`A., MUu.na,
`A. F., HERRMANN,
`GMLt(cid:1).RD,
`E. E.: RU 486: A steroid
`with
`antiglucocorticosteroid
`activity
`that
`only
`disin-
`at a specific
`time ofday.
`Proc. NatL
`hibits
`the
`human
`pituitary-adrenal
`system
`81:
`3879-3882,
`1984.
`Aced.
`Sd.
`U.S.A.
`P. W. AND HoDcnN,
`H. M., GOLD,
`D. L, CHRousos,
`G. P., SCHULTE,
`HEALY,
`G. D.: Increased
`vasopressin
`aecre-
`adrenocorticotropin,
`cortisol,
`and
`arginine
`RU 486. Dose
`in primates
`tion
`after
`the
`antiglucocorticoid
`steroid
`response
`J. Clin. EndocrinoL
`60:
`relationships.
`Metab.
`1-4,
`1985.
`D., TguTscH,
`W., Wysa,
`R., RIONDEL,
`A.,
`PHiuanRT,
`HERRMAN,
`E. E.: The
`effects
`of an antiprogesterone
`steroid
`BAULIRU,
`AND
`the menstrual
`cycle and of early
`Interruption
`pregnancy.
`Comptes
`294:
`933-940,
`1982.
`Y. AND HoMsu, M.: Differences
`S.,
`ICHIKAWA,
`KAWAI,
`among
`cortisol,
`prednisolone,
`and
`deza.methaaone
`Accelerated
`metabolism
`ofdexamethasone
`in renal
`80:
`MetabL
`848-854,
`1985.
`Ki.ny, H. K., BARTMANN,
`E. AND
`non-protein-bound
`method
`to measure
`oestradiol
`by equilibrium
`dialysis:
`1977.
`Endocrinol.
`85:
`209-219,
`B. A., Uoocsei,
`KovAcs,
`L.,
`SA5, M.,
`RE5CH,
`M.
`ROWE,
`P.
`J.:
`Termination
`of very
`AND
`sntiprogestational
`compound.
`Contraception
`Ln’FLE,
`B.,
`Tiur,
`F., T(cid:1)rr,
`S. A.
`S.
`J.
`(cid:1)D
`progesterone
`of
`in males
`clearance
`rate
`Invest.
`901-912,
`1966.
`45:
`D.: RU 38486:
`PHu.manT,
`MOGUILEWSKY,
`M.
`AND
`to the
`cytosolic
`activity
`correlated
`with
`strong
`binding
`J. Steroid
`Biochem.
`followed
`by an
`impaired
`activation.
`
`of
`
`G., SAiuz, E.
`on women
`Rendus
`
`properties
`in metabolic
`diseases:
`liver
`and
`renal
`in
`J. Clin. EndocrinoL
`failure.
`
`KRUSKEMPER,
`fractions
`Comparison
`
`and rapid
`H. L: A simple
`of
`cortisol,
`testosterone
`end
`with centrifugal
`Acts
`filtration.
`
`G., SWAHN,
`early
`pregnancy
`29:
`399-410,
`ERnzNumn,
`ovariectomized
`
`M. L., BYGDEMAN,
`by RU 486-an
`1984.
`F.: The
`females.
`
`metabolic
`J. Cliii.
`
`and
`
`Potent
`antiglucocorticoid
`glucocorticoid
`receptor
`20:
`271-276,
`1984.
`
`-a
`
`z C E 1
`
`=-
`
`(cid:1) h:
`
`90a
`
`(1)
`
`(g/dI)
`Albumin Concentration
`and specific
`column)
`of albumin
`effect
`5. The
`on the
`total
`(open
`Fig.
`(sheded
`(cid:1)Iumn)
`binding
`of 20
`nM [‘HIRU 486
`or [‘HIdeXamethaSOne
`to
`(MNL). Results
`intect
`human mononuclear
`leukocytes
`we
`expressed
`as
`percentage
`of
`those
`in media without
`human
`albumin.
`Values
`shown
`were
`determined
`in duplicate.
`
`of
`
`this
`
`drug
`
`crosses
`
`plasma
`
`membranes
`
`and
`
`is
`
`fraction
`free
`the
`active.
`biologically
`concentrations
`plasma
`The
`effective
`were
`syndrome
`Cushing’s
`tients
`with
`cortisol
`plasma
`the mean
`greater
`than
`patients.
`This
`from
`was
`unexpected
`mononuclear
`the
`human
`leukocytes
`3 times
`greater
`which
`is approximately
`approximately
`[or, by deduction,
`asone
`1979)].
`that
`of cortisol
`(Murakami
`et
`at.,
`may
`the
`to plasma
`explain
`drug
`albumin
`fractions
`of
`the
`The
`chromatographic
`tive material(s)
`corresponded
`to RU 486
`N-didemethylated
`metabolites.
`Deraedt
`that
`metabolites
`have
`a high
`shown
`
`pa-
`the
`of RU 486
`times
`20
`more
`than
`these
`of
`concentrations
`of RU 486
`to
`the
`affinity
`receptor,
`glucocorticoid
`of dexameth-
`that
`than
`than
`18 times
`greater
`of
`The
`binding
`tight
`this
`discrepancy.
`plasma
`receptor-reac-
`and
`its N-mono-
`(1985)
`et
`at.
`binding
`affinity
`
`in
`
`and
`have
`for
`
`these
`
`

`

`406
`
`Kawal
`
`at al
`
`Vol.241
`
`with estradiolbinding
`
`sites
`
`in rat uteri.
`
`Endocrinology
`
`99:
`
`1091-1096,
`
`E. E.:
`in
`phase
`
`digitalis
`1976.
`SCHAISON,
`Effects
`normal
`SCHREIBER,
`progestin
`1983.
`D., MISHELL,
`JR., L(cid:1)c(cid:1)aR(cid:1),
`E., LAHTEENMAKI,
`M., Gu’rinminz,
`D. R.,
`SHOUPE,
`ofthe
`P. AND SpiTz,
`in
`antiprogesterone
`RU 486
`I. M.: Endocrunologic
`effects
`Steroid RU 486
`of normal women.
`In The
`AntiprOgestin
`and
`the
`luteal
`phase
`S. J. Segal, pp.
`Human
`Fertility
`Control,
`ad.
`by E. E. Baulieu
`and
`285-293,
`New York,
`Plenum
`Press,
`1985.
`J. F., T(cid:1)iT,
`S. A. S.,
`ThiT,
`LI1’FLE,
`in the plasma
`of7-W-d-aldosterone
`80,1961.
`
`A. AND
`N., REINBERO,
`LE8TRAT,
`M.,
`G., Ggoiws
`BAULIRU,
`RU 486
`during
`IDidlUteaI
`steroid
`antiprogeeterone
`the
`of
`J. Clin. EndocrinoL Metab.
`61:
`women.
`484-489,
`1985.
`J.
`it, Hsugs,
`A. J. W.
`E. E.: Binding
`(cid:1)tiD
`BAUUEU,
`RU 486
`steroid receptors.
`rat
`ovary
`to
`Contraception
`
`of the
`28:
`
`anti-
`77-85,
`
`B. AND LAUMAS,
`ofnormal
`subjects.
`
`K.
`
`It: The disappearance
`J. Clin.
`Invest
`40:
`72-
`
`Downloaded from
`
`jpet.aspetjournals.org
`
` at ASPET Journals on January 19, 2016
`
`Send
`Bethesd(cid:1)
`
`requests
`reprint
`MD 20893.
`
`to:
`
`Shunichi
`
`Kawai,
`
`M.D.,
`
`NIH,
`
`Bldg.
`
`10, Ran.
`
`10N262,
`
`RODSARD,
`in circulating
`
`D., LoRi(cid:1)ux,
`mononuclear
`
`D. L
`AND LIPS(cid:1)F,
`leukocytes.
`Endo-
`
`D. D.,
`T., BsAt(cid:1)DoN,
`MuiuKAMI,
`M. B.: Glucocorticoid
`receptor
`104:
`1979.
`500-505,
`crunology
`B. C.,
`I. M., Nisui.A,
`C., SpiTz,
`G. P., KELLNER,
`NIEMAN,
`L K.,
`CHRousos,
`D. L:
`C. W. AND
`G.
`R,
`CUTLER,
`G.
`B., MERRIAM,
`LORIAUX,
`BARmN,
`with the glucocorticoid
`of Cushing’s
`syndrome
`antagonist
`Successful
`treatment
`RU 486. J. Clin. EndocrunoL
`Metab.
`61:
`536-540,
`1985a.
`BARDIN, C. W.,
`L K., Hasiy,
`D. L,
`It,
`G.
`SpiTz,
`I. M., MnwuAu,
`NIEMAN,
`of the antiproges-
`G. P.: Use
`of single doses
`LORIAUX,
`D. L AND CHRousos,
`In The
`women.
`terone
`steroid
`RU 486
`for
`induction
`of menstruation
`in normal
`Steroid RU 486
`ad.
`E. E.
`and
`Antiprogestin
`Human
`Fertility
`Control,
`and S. J. Segal,
`New York,
`1985b.
`Baulieu
`pp.
`279-283,
`Plenum
`Press,
`D., Dga(cid:1)givr,
`R. AND TEUTSCH,
`G.: RU 38486-a
`PHIUSERT,
`potent
`antigluco.
`in vivo
`the VIII
`corticoid
`(Abstract
`1463,
`p. 668).
`Presented
`International
`to
`of Pharmacology,
`Tokyo,
`Japan,
`Congress
`1981.
`D., MOGUILEw5KY,
`M. AND PO’I’FIER,
`M.,
`BONNAT,
`C., BU5IGNY,
`PHILIBERT,
`of human
`on pharmacokinetics
`Influence
`alpha
`1-acid glycoprotein
`(AAG)
`biological
`activityofRU
`486(Abstract
`1006,p.
`282).
`Presentedtothe
`CA, 1986.
`Society, Anaheim,
`S. M., PiTA,
`J. C., JR. AND LORIAUX,
`RJFKA,
`
`by
`
`J.:
`and
`Endocrine
`
`D. L: Mechanism
`
`of interaction
`
`of
`
`