NDA 21-272
`
`"
`
`46
`
`[Note: A 90-day continuous subcutaneous infusion study of UT-15 (lot numbers LRX-
`97101 and LRX-98A01)in rats (conducted atM
`Study No. 585D-103434—97), using osmotic pumps, at 0 (vehicle control), 0 (saline
`control), 50, 150 and 450 ng/kg/min, revealed no significant toxicological findings except
`for implantation site lesions (swelling, edema and inflammation) and significantly
`increased absolute and relative spleen weights in all
`treated female groups (dose
`dependent) and in the high dose male group. Increased absolute and relative liver (high
`dose females) and heart (high dose males) weights were also observed.
`
`A 28-day continuous subcutaneous infusion study in rats (conducted at M
`
`study number 1458), at targeted doses up to 450 ng/kg/min, compared
`the toxicological effects of a batch of UT-lS (lot number UTlSMIX-99GOOI) that had a
`different impurity profile to the lot that was previously tested (lot number UTISRP-
`981001) in the 26-week rat toxicity study. No significant differences in the toxicological
`profiles of these lots were noted]
`
`APPréJS THIS WAY
`0N fiRiGiNAL
`
`

`

`NDA 21-272
`
`'
`
`47
`
`Twenty-six Week Subcutaneous Continuous Infusion Toxicity Study in Dogs
`Followed by a 4-Week Recovefl Period '
`
`Testing Facility:W
`
`Study Number: 2129-98 .— Lab.’s No.)
`
`Study Dates: Initiation of Treatment — November 2, 1998
`'
`Terminal Sacrifice — May 3 to June 4, 1999
`
`GLP Compliance: The study was conducted in compliance with GLP regulations.
`
`Anim___a__ls: Beagle dogstM 4/sex/
`group, housed individually1n stainless steel cages and fed once daily with 400 g of
`w 25% Lab Dog Diet #8727C, were about 8 months old (males weighed 7.1 to 10.6
`kg and females 7.0 to 10.5 kg) at
`the onset of treatment. An additional
`three
`dogs/sex/group were assigned to the vehicle control and high dose groups for the
`recovery phase of the study.
`
`Dose Levels and Mode of Administration: The target dose levels and the concentrations
`of dosing solutions are given below.
`
`Low dose
`
`Vehicle control3
`
`' The test and control articles were administered at a constant rate of 0.05 ml/kg/hour to achieve target dose levels?"
`Saline control animals received 09% sodium chloride forinjection USP.Vehicle control animals received the vehicle
`containing sodium citrate, citric acid, sodium chloride and meta-crew! dissolved1n water for1njection USP (pH 6.8 -
`7.4).
`
`UT-lS (Lot No. UT15-98H01) solutions were prepared weekly by dilution of appropriate
`volumes of UT-15 stock solution (10 mg/ml) with the vehicle to achieve the desired
`concentrations. The dosing solutions were filtered through a .2;— filter to assure
`sterility.
`
`Analyses of test article concentrations at three different time points during the study
`showed that the concenuations of the dosing solutions were about —--—'"" (values
`corrected for volatiles and other mpurities) of the nominal values.
`
`

`

`NDA 21-272
`
`43
`
`It is stated that “the dose levels were selected by the sponsor together with the study
`director based on available data.”
`
`The test and control articles were infused subcutaneously, 24 hours/day, at a rate of 0.05
`ml/kg/hour. The infusion site on the dorsal side was surgiCally prepared, and the catheter
`was inserted under local anesthesia and secured in place. The catheter was connected to
`medical grade tubing that was passed through a jacket and tether system to the outside of
`the cage d00r. The infusion line was then connected to the reservoir containing the test
`article solution. The infusion rate was controlled using an infusion pump.
`
`The infusion site was changed approximately every 7 days (to another dorsal site). A
`topical antiseptic (4% chlorhexidine gluconate) was applied to the surgical sites
`throughout the treatment period. During the treatment period, animals (both control and
`treated) exhibiting skin lesions (wound, dermatitis, ulceration, edema and/or erythema)
`were treated with topical applications of 10% iodine and/or 0.05 or 4% chlorhexidine
`gluconate.
`
`signs and
`Observations/Measurements: Animals were observed daily for clinical
`mortality. A detailed clinical examination was performed one week prior to initiation of
`dosing and once weekly thereafier, with particular attention paid to surgical sites. Body
`weights were recorded for all animals one week prior to initiation of treatment, weekly
`during treatment and recovery periods and at study termination. Food consumption was
`recorded daily. Indirect ophthalmoscopy and slit lamp examinations were performed for
`all animals during the pretreatment period and also during weeks 13 and 26.
`Electrocardiograms (limb leads I, II and III; and augmented leads aVR, aVL and aVF)
`were recorded for all animals during the pretreatment period and also during treatment
`weeks 13 and 26. Hematology [RBC, WBC (total and differential), platelet and
`reticulocyte counts, hemoglobin, hematocrit, MCV, MCH, MCHC, prothrombin time,
`activated partial thromboplastin time, and blood cell morphology] and serum chemistry
`(urea nitrogen, creatinine, glucose, cholesterol,
`triglycerides,
`total protein, albumin,
`globulin, ALT, AST, alkaline phosphatase, bilirubin, calcium, sodium, potassium,
`phosphorus and chloride) evaluations, and urinalysis were performed on all animals
`pretest and during treatment weeks 13 and 26, and on all recovery animals at the end of
`the recovery period .
`
`Blood samples were collected from all dogs pretest and on days 1, 7, 14, 28, 42, 56, 70,
`84, 98, 112, 126, 140, 154, 168 and 182 for toxicokinetic evaluations. Blood samples
`were collected 3 hours post start of infusion on day l, and at the same hour on each of the
`remaining occasions. (Only blood samples from the first three dogs/sex/group were
`analyzed; samples collected from the remaining animals were discarded.)
`
`All main phase animals were euthanized after 26 weeks of treatment, and all recovery
`phase animals were kept for an additional 4 weeks (without dosing) before sacrifice.
`
`Complete necropsies were performed on all animals. Adrenals, brain, epididymides,
`heart, kidneys, liver, lungs, ovaries, prostate, spleen, testes, thymus, thyroids (with para-
`
`
`
`

`

`NDA 21-272
`
`‘
`
`'
`
`49
`
`.
`
`thyroids) and uterus were weighed. Representative sections of adrenals, aorta (thoracic),
`brain, cecum, cervix, colon, epididymides, esophagus, eyes, femoral bone, gallbladder,
`heart, last infusion site, kidneys, lacrimal glands, liver, lungs, lymph nodes (mesenteric
`and mandibular), mammary gland, optic nerves, ovaries, pancreas, pituitary, prostate,
`rectum, salivary gland (mandibular), sciatic nerve, skeletal muscle, skin and subcutis,
`small intestine (duodenum, ileum and jejunum), spinal cord (cervical), spleen, sternum
`and marrow, stomach, testes, thymus, thyroids with parathyroids, tongue, trachea, urinary
`bladder, uterus and vagina, and all abnormal tissues were fixed in neutral buffered 10%
`formalin (except epididymides, eyes, optic nerves and testes which were fixed in
`Zenker’s fluid).
`
`Three femoral bone marrow smears were prepared fiom each animal, but were not
`evaluated.
`
`Tissues were processed and stained with hematoxylin and eosin/phloxine for microscopic
`examination. Histopathological examination was performed on all above listed tissues
`from all animals.
`
`Data were analyzed for homogeneity of variance using Levene Median and for normality
`using Kolmorogov—Smimov tests. Homogeneous data were analyzed using the analysis of
`variance and the significance of intergroup differences was tested using Dunnett’s t test.
`Heterogeneous data were analyzed using Kruskal-Wallis test, and the significance was
`tested by Dunn’s test.
`
`Results: There were no deaths in this study.
`
`The most frequently observed clinical signs in animals of all dose groups, including
`controls, included the presence of lumps (firm), swellings (soft) and/or swellings around
`the lumps at or around the infusion site. The incidence and severity of these clinical signs
`were higher in drug-treated groups (especially in the high dose group) than in saline or
`vehicle controls. Generally, the lumps/swellings appeared l to 3 days following the start
`of infusion or the change to a new infusion site and they gradually disappeared 2 to 6
`days later.
`
`Dose-dependent erytherna around the infusion site was observed in females of all dose
`groups, including a few saline control animals. In males, erythema was frequently seen in
`mid and high dose animals.
`
`Animals from all drug-treated groups exhibited pain (sensitivity to touch) when palpated
`at the infusion site, the incidence being higher1n the high dose group than in other
`groups.
`
`Other clinical signs, including decreased activity, prostration, loose feces and wounds at
`the infusion sites, were observed sporadically in a few animals of one or more dose
`groups.
`
`

`

`NDA 21-272
`
`'
`
`so
`
`No treatment-related clinical signs were seen at the end of the recovery period.
`
`Dose-dependent decreases in mean body weights (2 to 10%), compared to body weights
`on Day 1, were noted during the first 1 to 4 weeks of treatment in all dose groups (both
`sexes). Reductions in body weights (1-3%) were also noted for saline and vehicle
`controls during this period. No further significant decreases in body weights were noted
`in any dose groups by about Day 29, and the animals started to gain weight thereafter. By
`the end of the treatment period (Day 183), the mean body weights of treated and control
`groups were 2-7% and 5-10% higher, respectively, than corresponding values on day 1.
`
`At the end of the recovery period, the body weight gains were comparable in the high
`dose and vehicle control groups.
`
`During the first week of treatment, the food consumption of the high dose group (both
`sexes) was
`significantly lower
`than control. The food consumption improved
`considerably during the second and third weeks of treatment, and by the fourth or fifih
`week, the food consumption of the high dose group was comparable to that of controls.
`There were no significant treatment-related effects on food consumption at other dose
`levels.
`
`During the recovery period, food intake was comparable in treated and control groups.
`
`There were no treatment-related ocular findings in the study.
`
`including the
`The test-drug had no effects on electrocardiographic parameters
`~ morphology of the P, QRS and T complexes, PR and QT intervals, ST segment, mean
`electrical axis, and heart rate.
`
`Reversible, non dose-related increases in mean white cell counts (9 — 113%; mainly due
`to an increase in neutrophils), compared to controls, were noted in the mid and high dose
`groups of both sexes during treatment weeks 13 and 26.
`
`treatment-related findings were seen in the clinical chemistry and
`No significant
`urinalysis parameters.
`
`Although not statistically significant, increases in the absolute (14 - 35%) and relative
`(31%) mean spleen weight, compared to vehicle control, were seen in both sexes of the
`high dose group. Afier the recovery period, the mean spleen weight was still higher in the
`high dose male group than in control.
`
`Macroscopically, thickening of the infusion site and masses around the infiision site were
`seen in some of the control dogs and in the majority of the drug-treated dogs.
`
`No treatment-related macroscopic findings were noted after 4 weeks of recovery.
`
`'
`
`
`
`
`
`

`

`NDA 21-272
`
`51
`
`the infusion site included
`Treatment-related histopathological findings observed at
`cellulitis, edema, fibrosis and hemorrhage.‘ Although the incidence of cellulitis and edema
`in the treated groups was comparable to that in saline or vehicle controls, the incidence
`and/or severity of hemorrhage and/or fibrosis was higher in drug-treated groups (not
`dose-dependent) than in controls.
`
`There were no other significant or otherwise remarkable histopathological findings.
`
`The mean plasma drug concentration (pooled data from males and females) versus study
`day for each of the three targeted doses is shown in Figure 11. Individual animal data
`showed that steady-state plasma concentrations were achieved in most animals by three
`hours on Day 1, and in all animals by Day 7. Moderate to wide variations were observed
`in steady-state plasma UT-lS concentrations over the 182-day monitoring period. The
`overall mean plasma steady-state concentrations and the clearance rates are presented
`below.
`
`The mean Css values increased proportionally with dose. Linear regression analysis of
`the individual animal Css versus dose data yielded a straight line with a coefficient of
`determination (r2) of 0.69. The coefficient increased to 0.93 when one high dose animal
`(showing aberrant pharmacokinetic pattern with non-detectable drug levels at many time
`points) was excluded, indicating dose proportional kinetics over the 50 to 200 ng/kg/min
`dose range.
`
`STEADY—STATE CONCENTRATIONS AND CLEARANCE
`RATES IN DOGS ADMINISTRRED UT-IS IN A ZGWEEK
`TOXICOKINETIC STUDY
`
`Parameter
`
`Dose
`
`FemaleDog MaleDog
`
`
`
`
`(ng/kg/min)nN=3
`N=3
`“gm” “Eu—
`
`
`
`
`
`
`(mnglmin)
`
`'
`‘
`‘
`mum-m
`‘ One male beagle dog at the ZOO-nykg/min dose showed non-detectable III-15 levels at many
`timcpointsoverthe lSO-daysmdyandthemforebwueddiememqmueswhfleatdtsann
`timeraisedfliemeanCUPvaluea
`
`

`

`
`
`
`
`MeanUT~1$Concentration(nymL)
`
`Figure 11.
`Plots of Mean UT-15 Concentration (tSBM) versus Study Day by ’Ihrget Does
`
`
`
`20
`
`40
`
`60
`
`80
`
`100
`
`120
`
`140
`
`160
`
`180
`
`200
`
`ThrgatDoso
`
`{—H son H—O mans/hymn
`
`l~l~l
`
`zoong/kg/min
`
`Study Day
`
`ZLZ‘IZV(IN
`
`ZS
`
`

`

`NDA 21-272
`
`-
`
`V
`
`53
`
`The steady—state concentrations achieved after continuous subcutaneous infusion of UT-
`15 for 26 weeks in rats and dogs or 12 weeks in human volunteers are compared below.
`The highest
`infusion rates achieved in rats and dogs (450 and 200 ng/kg/min,
`respectively) represent UT-lS doses that produced minimal to moderate toxicological
`effects, whereas in humans, the highest infusion rate (15 ng/kg/min) was at the upper end
`of the tolerated dose range.
`
`COMPARISON OF SHADY-STATE CONCENTRATIONS
`
`(no/ml)
`
`(us/ml.)
`
`(no/kwmin) c:
`
`(us/ml.)
`
`
`
`
`
`FOR UT-IS IN RAT, DOG, AND MAN ADMINISTERED UT-lS BY
`SUBCUTANEOUS INFUSION
`Dose
`Rat
`Dose
`
`("Silks/min)
`
`(ng/kg/min)
`
`
`“Ella—In”
`mulmmlm=
`
`
`
`
`m-Iln-mlm 099
`
`—_I_-lmm
`
`Human data are obtained fi'om Protocol 901119 (Reference 8)
`
`[Note: In a 13-week continuous subcutaneous infusion study of UT-15 (lot number LRX-
`
`98A01) in dogs (conducted at
`,
`l study number 2131-98)
`at target doses of 50, 150 and 300 ng/kg/min, a high dose female dog was sacrificed on
`day 13 due to moribund condition with a rectal prolapse. Pathology findings in this dog
`included multiple intestinal intussusceptions, intestinal hemorrhage, and a severe wound
`and ulceration at the infusion site. Because of these adverse findings, the high dose was
`lowered to 200 ng/kg/min on day 15, and to allow a wider spread between mid and high
`dose levels, the mid dose was lowered to 100 ng/kg/day. Other treatment-related findings
`included edema and/or erythema at the infusion site, dose—related reduction in body
`weights during the first 3 weeks of treatment, and dose-related increases in WBC counts
`in treated female groups during treatment weeks 6 and 13 (except in low dose females
`during week 6). There were no treatment-related findings at the end of a 2-week recovery
`period]
`
`APPEARS THlS WAY
`0N ORlGl‘lAL
`
`

`

`NDA 21-272
`
`’
`
`55
`
`/""\.
`
`—-'-—'"" osmotic pump was inserted into the subcutaneous pocket. Theincision was
`closed with wound clips.
`.
`
`Males were treated for 10 weeks prior to mating and continued through the 2 weeks
`mating period. Females were dosed for 2 weeks prior to mating and continued through
`the mating period till gestational day (gd) 6. (The confirmed day of copulation, evidenced
`by the presence of vaginal sperm or vaginal copulation plug, was designated as gd 0.)
`
`[According to the manufacturer’s specifications, the osmotic pump used in this study
`delivers 2.5 til/hour (nominal flow rate) for a nominal duration of 28 days. For F0 males,
`the pumps were implanted four times during the study (at about 3-week intervals, in
`weeks 0, 3, 6 and 9, for a total 12 weelm of exposure). Females that were found sperm or
`plug positive during the first week of mating did not receive a second pump for the
`remainder of the exposure period. For majority of the F0 females (that successfully mated
`during the first week of mating), the original osmotic pump implanted at the start of the
`study remained in place till its removal on gd 6. A few F0 females, that failed to mate
`within the first week of mating, received a second osmotic pump.]
`
`It is stated that “the highest dose level (450 ng/kg/min) administered was selected to
`induce maternal toxicity or low levels of lethality (S 10%), and was based on the results
`from a previous Segment II study performed” at _. The lower doses (50 and 150
`ng/kg/min) were chosen as fractions of the high dose.
`
`Observations and Measurements: Animals were checked daily for changes in general
`condition and behavior. Observations for mortality were made twice daily. Body weights
`of F0 males were recorded prior to the beginning of treatment and weekly thereafter. F0
`females were weighed weekly until confirmation of mating, and during gestation on gd 0,
`3, 6, 9, 12 and 15. Food consumption was determined weekly for all animals during the
`pre-mating period. During pregnancy, food consumption was recorded for gd 0-3, 3-6, 6-
`9, 9-12, and 12-15. (Food consumption was not determined during the mating period.)
`
`For the females, during the two-week premating exposure period, estrous cyclicity was
`assessed by daily vaginal lavage. The slides of the 14-day vaginal lavage were fixed,
`stained with toluidine blue, and read for the stages of the estrous cycle.
`
`After mating, all F0 males were sacrificed by C02 asphyxiation, and subjected to a
`complete gross necropsy. Testes and epididyrnides were weighed. Seminal fluid from the
`right cauda epididymis was evaluated for sperm number, motility and morphology.
`Sperm motility was evaluated immediately after necropsy; sperm number was determined
`using fixed, stained sperm specimen. The motility and number of sperm were evaluated
`
`using an
`Automated Sperm Analysis System, while sperm morphology was
`evaluated manually. The testes were fixed in Bouin’s solution and changed into 70%
`ethanol after about 24 hours. The lefi epididymis, and any gross lesions were fixed in
`neutral buffered 10% formalin for possible histopathologic examination. (Histopathologic
`examination was not performed.)
`
`-. —.—~
`
`......_» .-'-~,._....__' .,T~_~..._V.__. M- ...-,,.._.-_.
`
`.
`
`.. .
`
`.
`
`.
`
`.
`
`
`
`

`

`NDA 21-272
`
`.
`
`56
`
`On gd 15, all surviving F0 females were sacrificed, a complete necropsy was performed,
`and the pregnancy status was confirmedfby uterine examination. Numbers of corpora
`lutea,
`implantation sites, resorptions, and dead and live conceptuses were recorded.
`Ovaries and uterus were weighed, and these organs and any gross lesions were fixed in
`buffered neutral 10% formalin. For confirmation of pregnancy status, fixed uteri, that
`showed no visible implantation sites, were stained with potassium ferricyanide to
`visualize any implantation sites which may have undergone very early resorption. (Histo-
`pathologic examination of fixed tissues was not performed.)
`
`The indices for reproduction (mating, fertility and pregnancy indices) and gestational
`parameters (gestational index and pre-and post~implantation losses) were calculated.
`
`Quantitative continuous data were statistically analyzed using Bartlett’s test for homo-
`geneity of variances. If Bartlett’s test indicated lack of homogeneity of variances, then
`nonparametric statistical tests (Kruskal-Wallis Test follOwed by Mann-Whitney U test)
`were employed. If Bartlett’s test
`indicated homogeneous variances, then parametric
`statistical
`tests [General Linear Models procedures for the Analysis of Variance
`(ANOVA)] were used. All indices were analyzed by Chi-Square test and by the Cochran-
`Armitage test for linear trend on proportions.
`
`Results: There was no F0 parental mortality or morbidity.
`
`Males
`
`Treatment-related clinical signs for males were limited to swelling at the pump site and
`flushed appearance of cars, paws, and/or tail. Swelling at the pump site was noted in both
`control and drug—treated animals, with higher incidences seen in treated groups (no dose
`relationship). Flushed ears, paws, and/or tail were seen mostly in high dose animals, with
`only few animals affected in the mid dose group and none in the low dose group.
`
`Body weights of high dose male animals were significantly lower (5 to 6%) than
`concurrent control weights during the first 3 weeks of the study. No significant
`differences in body weights were noted between treated (any group) and concurrent
`control animals thereafter. During the first week of treatment, the food consumption for
`the mid and high dose groups was significantly lower than control. There were no
`significant treatment-related effects on food consumption thereafter.
`
`The mating, fertility and pregnancy indices were generally similar across control and
`treatment groups.
`
`There were no treatment-related effects on absolute and relative testes and epididymides
`weights and sperm evaluation parameters (epididymal sperm concentration, percent
`motile sperm, percent progressively motile sperm, and percent abnormal sperm).
`
`

`

`NDA 21—272
`
`'
`
`57
`
`Females
`
`Consistent with the findings in males, treatment-related clinical signs in females were
`limited to swelling at the pump site and flushed ears and paws. Swelling at the pump site
`was observed both in control and treated rats, with higher incidences seen in treated
`groups (no dose relationship). Flushed ears and paws were seen mostly at the high dose.
`
`Estrous cyclicity during the 14-day premating period showed no differences across
`control and treatment groups.
`
`There were no treatment~related effects on body weights during the premating period and
`throughout the gestation period. During the premating period, the food consumption in
`treated groups was lower than control (no dose relationship). There were no treatment-
`related effects on food consumption during the gestation period.
`
`female
`The F0 female terminal body weights, gravid uterine and ovary weights,
`reproductive indices, numbers of corpora lutea and implantation sites and litter incidences
`of resorptions,
`live and nonlive (resorbed plus dead) implants, and pre- .and post-
`implantation losses are presented in Table 16.
`
`UT-15 treatment had no effect on mating, fertility and gestational indices. There were no
`significant differences in maternal body weights and organ weights between control and
`treated groups. There were no treatment-related effects on the numbers of corpora lutea
`or total implantation sites per darn, resorptions per litter,
`litters with resorptions or
`nonlive implants, live and nonlive implants per litter, and pre- and post-implantation
`losses per litter.
`
`Under the conditions of this study, the reproductive and developmental NOAELs are at or
`above 450 ng/kg/min.
`
`APPEARS THIS WAY
`0N GREGEHAL
`
`APPEARS THIS WAY
`Oli fiRiGINAL
`
`

`

`NDA 21-272
`
`Table 16.
`
`58
`
`Summary and Statistical Analysis of the Femaie Reproductive indexes. Necropsy Wetghts and
`Uterine Contents
`(page 1 01 3)
`
`W L
`
`IT-15 (Mm, subcutaneous}
` O 50 150 450
`
`
`
`
`
`N0. of Males Paired
`
`No. of Fernates Paired
`
`No. of Femates that Mated
`
`25
`
`25
`
`25
`
`25
`
`25
`
`24'
`
`25
`
`25
`
`_
`
`24
`
`Mating Index (96) (no. funates that mated/no. females pared)
`100.0
`96.0
`
`No. ofPregnant Femates
`
`20
`
`23
`
`Fertility Index (96) (no. pregnant femaleslno. femates that mated)
`80.0
`95.8
`
`No. of Females with Live Littets
`
`19
`
`23
`
`96.0
`
`'
`20
`
`83.3
`
`’
`20
`
`Gestational Index (96) (no. females with live litters/no. pregnant females)
`95.0
`100.0
`
`100.0
`
`25
`
`25
`
`23
`
`92.0
`
`22
`
`95.7
`
`22
`
`100.0
`
`Pregnancy Index (96) (no. pregnant (armies/no. mam that mated)
`~80.0
`95.8
`
`83.3
`
`95.7
`
`Days Unb1 Sperm Positive (daysF-b
`'
`
`2.5
`2.4
`2.1
`2.0
`3 0.5
`3; 0.5
`1 0.5
`3; 0.2
`N=22
`N=23
`N820
`N=21
`
`
`Maternal Body Weight at Sacrifice (9P3
`347.93
`3, 6.25
`mm“
`
`Gravid Uterine Weight (9)va
`
`332.46
`1 6.41
`N=19
`
`333.47
`3 5.13
`N=19
`
`19270
`
`11.480
`N=16
`
`18.644
`
`11.087
`N=19
`
`17.356
`
`11.111
`=19
`
`343.46
`1 5.75
`=21
`
`10.015
`
`. 11.088
`=21
`
`Paired Ovary Weight (gtbvc
`
`02115
`02273
`0.2087
`02444
`1 0.0088
`1 0.0114
`1 0.0062
`i 0.0129
`
`
`
`N=19 =19N=16 N=21
`
`(continued)
`
`

`

`NDA 21-272
`
`59
`
`Table 16. (continued)
`
`Summary and Statistim! Analysis of the Female Reproductive ktdexes, Necropsy Weights and
`Uterine Contents
`(0090 2 of 3)
`
`W U
`
`T-15 gag/3911001, subwtaneous)
` 0 50 150- 450
`
`
`
`
`
`N0. Corpom Lutea per Damb
`
`\
`
`N0. unptantatton Sites per Litterb
`
`10.30
`10.57
`N=20
`
`14.95
`1 0.92
`N=20
`
`15.30
`10.49
`N=23
`
`13.43
`1 0.34
`N=23
`
`1525
`10.56
`N=20
`
`13.15
`1 000
`N320
`
`Pm Pretuptantation Loss per Limb
`
`11.54
`1 424
`N=20
`
`1.50
`10.51
`me
`
`12.51
`1 4.93
`11:20
`
`1437
`14.51
`N223
`
`'
`
`~ 15.31
`'1 421
`N=20
`
`0.03
`1 029
`11:23
`
`623
`1 2.33
`N=
`
`1.05
`1028
`11:20
`
`7.36
`1 125
`N=20
`
`14
`
`' a
`
`11
`
`12
`
`70.00
`
`34.70
`
`5500
`
`54.55
`
`0.00
`1 0.00
`11-20
`
`0.00
`10.00
`N=
`
`0.00
`1 0.00
`N=23
`
`0.00
`1 0.00
`N=20
`
`0.00
`10.00
`N=23
`
`‘
`
`0.00
`10.00
`=20
`
`0.00
`1 0.00
`N=
`
`0.00
`10.00
`N:
`
`0
`
`o
`
`0
`
`0
`
`No. Resorptlons per Uttetb
`
`Percent Rmptions per Littetb
`
`N0. Litters with Resorpfions
`
`'
`’
`.
`9t» Litters with Rmpfions _
`
`No. Late Fetal Deaths per Utterb
`
`Percent Late Fetal Deaths per Utterb
`
`1!
`
`N0. Litters with Late Fetal Deaths
`
`11. Litters with Late Fetal Deaths
`
`15.41
`10.42
`N=22
`
`14.55
`1 021
`N=
`
`'
`
`10.51
`1 4.55
`N=22
`
`0.77
`10.17
`N:
`
`5.67
`11.41
`N=22
`
`
`
`0.00 0.000.00 0.00W
`
`
`
`
`
`(continued)
`
`

`

`NDA 21-272
`
`'
`
`60
`
`Table 16. (continued)
`
`Summary and Statistics! Analysis of the Female Reproductive indexes, Necropsy Weights and
`,
`Uterine Contents
`(page 3 0(3)
`
`W U
`
`T~15 {Mam subcutaneous)
`50
`150
`
`450
`
`0
`
`No. Nontive Implants per Litterb-e
`
`Percent Nonlive Implants per unsbn"
`
`Na. Litters with Nontive nuance
`
`96 Litters with Martin lrhplants’
`
`No. Live implants per Limb
`
`-
`Percent Live Implants per Litter”
`
`1.50
`1 0.51
`N=20
`
`12.51
`14.93
`N=20
`
`0.83
`1', 0.29
`N=23
`
`6.23
`32.33
`N=23
`
`1.05
`1 0.20
`N=20
`
`7.36
`11.95
`N=—20
`
`0.77
`1 0.17
`N=22
`
`5.67
`4:141
`N=
`
`14
`
`s
`
`11
`
`‘
`
`-
`
`12
`
`70.00
`
`34.78
`
`13.45
`1 1.16
`. N=20
`
`87.49
`:0; 4.93
`. N=20
`
`12.61
`1 0.88
`N323
`
`93.77
`1 2.33
`N=23
`
`55.00
`
`‘
`12.10
`1 0.80
`N=2o
`
`92.64
`1 1.95
`N320
`
`54.55
`
`13.77
`:l; 0.88
`N=22
`
`94.33
`1 1.41
`N=22
`
`Percent Posfimplantation Loss per Utah
`5.37
`7.36
`6.23
`1251
`1 1.41
`1 1.95
`' 1 2.33
`1 4.93
`
`
`
`N823 N=20N320 N=22
`
`Magnum.
`hmdudesaflmmmwepoflodasmemeanzsfiu.
`eDoesnotincludehepregnantdamsforwhichspennwereheverdetectedsinoetheactualgestatioflat
`dayOWasnottmovm.
`Wmuummmmmmmmmymoemm.
`WWW.
`WsmwmdmmsiWWDOflmMmtbemmm
`mmvaflmoehomwmms.mbmmnpammemcstafisfidpmm
`anployed.
`
`

`

`
`
`NDA 21-272
`
`'
`
`61
`
`Developmental Toxicity Evaluation of UT-lS (Administered by Continuous
`Subcutaneous Infusion) in Rats
`'
`
`Testing Facility ”WWW
`
`4’”:
`
`(W
`
`Study Number: 65C-7019-200( "” No.)
`
`Study Dates: Initiation Date —- January 20, 1998
`Completion Date — April 8, 1998
`
`GLP Compliance: The study was conducted in compliance with GLP regulations.
`
`Animals: One hundred seventy female Sprague-Dawley rats, 56 days old (201-225 g
`
`body weight), were obtained from
`'
`_
`. Afier a week
`of quarantine, females were mated with male Sprague-Dawley rats that were obtained
`earlier from the same supplier and kept for breeding purposes. One hundred twenty~five
`sperm-positive females [about nine weeks of age and 216-275 g body weight on
`gestational day (gd) 0}, were assigned to five groups of 25 rats each. (The confirmed day
`of copulation, evidenced by the presence of vaginal sperm or vaginal plug, was
`designated as gd 0.)
`
`Non-mated females were group housed (maximum 3 per cage), and mated females were
`individually housed in solid bottom polycarbonate cages with stainless steel wire lids and
`.——~
`cage bedding. Males were housed individually. Food (—J'certifiedrrodent
`chow — No. 5002) and tap water were available ad Iibitum.
`
`Dose Levels, Mode of Administration and Treatment Regu_r_i'en: The target doses were 0,
`50, 150, 450 and 900 ng/kg/min.
`
`Stock solutions (10 mg/ml) of UT—lS (Lot No.LRX-98A01), formulated in vehicle
`(containing sodium citrate, citric acid, sodium chloride dissolved in sterile water for
`injection) and adjusted to pH 7.4, were diluted (with vehicle) to achieve the desired final
`concentrations.
`
`Analyses of the test solutions indicated that dosing formulations were 92-96% of target
`concentrations and the stock solution was 95% of target. It was shown that dosing
`g formulations were stable at 25 and 40° C for about four weeks.
`
`The test and vehicle solutions were administered by continuous subcutaneous infusion
`using subcutaneously implanted '
`’ osmotic pumps
`_._..—.-.___‘_‘
`at a
`nominal flow rate of . -f’-- to achieve the target dose levels.
`
`On the morning of gd 5 (between 0700 and 1100 hours), each study female rat was
`anesthetized by isoflurane inhalation. The dorsal subscapular area was surgically
`
`

`

`NDA 21-272
`
`’
`
`62
`
`prepared and an incision about 1.5 cm in diameter was made. The osmotic pump,
`preloaded with appropriate dosing solution, was inserted into the subcutaneous pocket
`and the incision was closed with wound clips.
`
`According to the manufacturer’s specifications, the osmotic pump model used in the
`study requires ”ref: to reach the steady state infusion rate once it is implanted.
`Therefore, the pump that was implanted on the morning of gd 5 would be at steady state
`by the morning of gd 6. The pump was removed at scheduled necropsy on gd 20 at about
`the same time that the pump was inserted on gd 5 (for a duration of 14 days of exposure
`at steady state).
`
`[It is stated that the doses were selected based on the results of a dose range-fmding study
`(continuous sc infusion of UT-15 at 0, 50, 150 and 450 ng/kg/min on days 6 through 20
`of gestation) in rats. In that Study, there was only minimal maternal toxicity and no
`evidence of developmental toxicity at the high dose. Therefore, 900 ng/kg/min was
`selected as the top dose for the definitive study to provide maternal and/or developmental
`toxicity. The remaining doses were the same as that used in the range-finding study.]
`
`Observations and Measurements: Animals were checked at least once daily for clinical
`signs during gestational days 0—4 (prior to pump implant), and twice daily during gd 5-20
`(pump implant period). Body weights were recorded on gd 0, 5, 6, 9, 12, 15, 18 and 20.
`Food consumption was determined for gd 0-5, 5-6, 6-9, 9-12, 12-15, 15-18 and 18-20.
`
`Blood samples from five dams per dose group were collected fi’om the lateral tail vein on
`gd 15 at time 0 (at about the same time the osmotic pump was inserted on gd 5), and 5
`different dams per group were bled at 4 hours after time 0, for maternal toxicokinetic
`evaluations.
`-
`
`On gestational day 20, all surviving females were killed by C02 asphyxiation at the
`approximate time (within 2 to 3 hours) the osmotic pump was implanted on gd 5. The
`wound clips were removed, the incision opened and the pump was removed, examined
`grossly and discarded. Animals were weighed and a complete necropsy was performed.
`Liver and uterus were weighed. Pregnancy status was confirmed by uterine examination.
`Uteri which. showed no visible implantation sites were stained with ammonium sulfide
`(10%) in order to visualize any implantation sites which may have undergone very early
`resorption. Numbers of corpora lutea, implantation sites, early and late resorptions, and
`dead and live fetuses were recorded. Live fetuses were dissected out, counted, weighed,
`sexed and examined for external morphological abnormalities, including clefi palate.
`Approximately one-half of the live fetuses in each litter were examined for visceral
`malformations, and their sex confirmed. These fetuses were decapitated, the heads fixed
`in Bouin’s solution and serial free-hand sections of the head were examined for sofi
`
`.
`
`tissue craniofacial malformations and variations. Although all fetuses in each litter were
`evicerated, fixed in ethanol, cleaned of tissues with KOH and stained with alizarin red
`S/alcian blue, only intact fetuses in each litter (not decapitated) were examined for
`skeletal malformations and variations. All fetal carcasses were stored in glycerin: 70%
`
`

`

`NDA 21-272
`
`"
`
`63
`
`ethanol (1:1; including those examined and not examined for skeletal malformations).
`Fetal head sections were stored in 79% ethanol after examination.
`
`Developmental toxicity parameters were calculated for each litter (dam) and then the
`mean was calculated using the litter (dam) values.
`
`for
`Quantitative continuous data were statistically analyzed using Bartlett’s test
`homogeneity of variances. If Bartlett’s test indicated lack of homogeneity of variances,
`then nonparametric statistical tests (Kruskal-Wallis test followed by Mann-Whitne