Document b5kVRg0e8OVkpoGoK56g7E3Vg

HAZLETON W W IS C 0 N S IN PC S - 0 iC 6 C) x 7 54 5 M;,D!Sor,, 5 --7 0 7 - 7 5 4 5 Sponsor: 3M St. Paul, Minnesota CORNING (@oiiii-iny P one FPF FINAL REPORT Study Title: Single-Dose Intravenous Pharmacokinetic Study of T-5904 in Rabbits Author: Steven M. Glaza Study Completion Date: October 14, 1994 Performing Laborator Hazleton Wisconsin, Inc. 3301 Kinsman Boulevard Madison, Wisconsin 53704 LaboratoryProject Identification: HWI 6329-117 Page I of 24 41 7 Page 2 of 24 HWI 6329-117 QUALITY ASSURANCE STATEMENT This report has been reviewed by the Quality Assurance Unit of Hazleton Wisconsin, Inc., in accordance with the Food and Drug Administration (FDA) Good Laboratory Practice Regulations,21 CFR 58.35 (b) (6) (7). The following inspections were conducted and findings reported to the Study Director and management. Written status reports of inspections and findings are issued to Hazleton management mont.hlyaccording to standard operating procedures. Inspection Dates From To- Phase Date Reported to Study Director Date to Management 03/01/94 03/07/94 04/26/94 10/11/94 03/01/94 03/07/94 04/26/94 10/11/94 Protocol Review Body Weight Data/Report Review Report Re-Review 03/02/94 03/07/94 04/29/94 10/11/94 04/10/94 04/10/94 05/10/94 11/10/94 &:- - NancyDie ich v Represent@tive, Quality Assurance Unit Date /0 -14-q4 Page 3 of 24 STUDY IDENTIFICATION Single-Dose Intravenous Pharmacokinetic Study of T-5904 in Rabbits HWI 6329-117 Test Material Sponsor Sponsor's Representative Study Director Study Location Study Timetable Experimental Start Date Experimental Termination Date T-5904 3M Toxicology Services 220-2E-02 3M Center St. Paul, MN 55144 John L. Butenhoff, PhD 3M Toxicology Services 220-2E-02 3M Center St. Paul, MN 55144 (612) 733-1962 Steven M. Glaza Hazleton Wisconsin, Inc. P.O. Box 7545 Madison, WI 53707-7545 (608) 241-7292 Hazleton Wisconsin, Inc. Building No. 3 3802 Packers Avenue Madison, WI 53704 March 7, 1994 March 9, 1994 Acute Toxicolog Steven M. Glaza Study Director Manager Steven R. Sorenson Study Coordinator Patricia Padgham In-lifeSupervisor Rose M. Bridge Report Supervisor Quality Assurance Sherry R. W. Petsel Manager Page 4 of 24 KEY PERSONNEL HWI 6329-117 LaboratoryAnimal Medicine Cindy J. Cary, DVM Diplomate, ACLAM Supervisor Anatomical Patholoov Jack Serfort/ DeborahL. Pirkel Supervisors Necropsy Anne Mosher Supervisor Pathology Data Page 5 of 24 CONTENTS Quality Assurance Statement Study Identification Key Personnel Summary Objective Regulatory Compliance Test and Control Material Test System Procedures Results Discussion Signature Reference Table 1 Individual Body Weights (g) 2 Individual Clinical Signs Appendix A Protocol Deviations Protocol TP8084.PK HWI 6329-117 Paqe 2 3 4 6 7 7 7 8 9 10 11 11 11 12 13 14 15 16 Page 6 of 24 SUMMARY HWI 6329-117 This study was done to assess the level of systemic exposure of T-5904 when administered by intravenous injection to rabbits. Female Hra:(NZW)SPF rabbits were assigned at random to five groups (one/group). On Day 0, the animals received a single intravenous injection of the vehicle [Dimethyl Sulfoxide (DMSO)J at a dose volume of 0.5 mL/kg or 1.0, 10.0, 50.0, or 100.0 mg of T-5904/kg of body weight. Clinical observations were conducted at approximately 0.5, 2, 4, 24, and 48 hours after intravenous injection. Body weights were determined just before test material administration (Day 0). Blood samples were collected from the marginal ear vein of the animals at 2-, 4-, 6-, 12-, and 24-hours post-injection. In addition, at the time of experimental termination (48-hours post-injection) a large volume of blood was obtained from each surviving animal. All samples were centrifuged and separated into serum and cellular fractions. The animal found dead during the study was necropsied. Animals surviving to the end of the 48-hour post-injection period were anesthetized with sodium pentobarbital and exsanguinated without necropsy. The bile from each animal surviving to termination was collected and sent frozen to the Sponsor after termination of the in-life phase. Administration of T-5904 resulted in a decrease in food consumption at all dose levels, hypoactivity in the animals treated at the 50.0 and 100.0 mg/kg dose levels, and death of the animal treated at the 100.0 mg/kg dose level. Page 7 of 24 HWI 6329-117 OBJECTIVE The objective of this study was to assess the level of systemic exposure to the test material, T-5904, when administered as a single intravenous injection to rabbits. REGULATORY COMPLIANCE This study was conducted in accordance with the U.S. Food and Drug Administration'sGood Laboratory Practice Regulations for Nonclinical Laboratory Studies, 21 CFR 58. All procedures used in this study are in compliancewith the Animals Welfare Act Regulations. In the opinion of the Sponsor and study director, the study did not unnecessarilyduplicate any previous work. Hazleton Wisconsin (HWI) does not accept any responsibility for the analysis of the samples collected in this study nor are these results presented in this report. TEST AND CONTROL MATERIAL Identification The test material was identified as T-5904 and described as a yellow solid. The control material was Dimethyl Sulfoxide (DMSO) (Sigma Chemical Company, St. Louis, Missouri; Lot No. 53HO634; Exp. March 4, 1995) and described as a clear, colorlessliquid. Purity and Stability The Sponsor assumes responsibilityfor test material purity and stability determinations(includingunder test conditions). A sample of the test material/vehiclemixtures for concentration,solubility,homogeneity,and stability analyses was not taken before administrationas this was not requested by the Sponsor. The certificateof analysis, obtained from the supplier, documents the purity of the control material as 99.9%. The stabilityof the control material was considered to be adequate for the purposes of this study. Storage and Retention The test and control material were stored at room temperature. Any unused test or control material will be discarded after issuance of the final report according to HWI Standard Operating Procedure (SOP). Page 8 of 24 HWI 6329-117 Safety PrecauligfLs The test and control material handling procedures were according to HWI SOPS and policies. TEST SYSTEM Test Animal Adult albino rabbits of the Hra:(NZW)SPF strain were received from HRP, Inc., Kalamazoo, MI, on February 23, 1994 and maintained at the Hazleton Wisconsin facility at 3802 Packers Avenue, Madison, Wisconsin. Animal husbandry and housing at HWI comply with standards outlined in the "Guide for the Care and Use of Laboratory Animals".' The animals were individually housed in screenbottom stainless steel cages in temperature- and humidity-controlled quarters, provided access to water ad 7ibitum and a measured amount of Laboratory Rabbit Diet HFO #5326, PMI Feeds, Inc., and held for an acclimation period of at least 7 days. The feed is routinely analyzed by the manufacturer for nutritional components and environmental contaminants. Samples of water are periodically analyzed by HWI. There were no known contaminants in the feed or water that would have interfered with or affected the results of the study. All animals were identified by animal number and corresponding ear tag and maintained during the observation period as specified for the acclimation period. During the conduct of the study, the temperature of the animal room ranged from 20 to 23*C and the relative humidity ranged from 48 to 60%. The animals were housed under the required temperature and humidity conditions. Selection of Test Animals The animals were selected at random based on health and body weight requirements. Study Design Female animals weighing from 2,173 to 2,386 g at initiation of treatment were placed into the following study groups: Group Treatment Dose Level Dose Volume Number (mg/kg) (mL/kq) of Animals 1 (Control) DMSO 0.0 0.5 1 2 T-5904 1.0 0.5 1 3 T-5904 10.0 0.5 1 4 T-5904 50.0 0.5 1 5 T-5904 100.0 0.5 1 Page 9 of 24 HWI 6329-117 Justificati ion Historically, the New Zealand White albino rabbit has been the animal of choice because of the large amount of background information on this species. PROCEDURES Dose Preparation and Administration The test material was diluted with DMSO to achieve a specific concentration for each dose level. An individual dose of each respective test solution or control was calculated for each animal based on its body weight on the day of treatment. The respective test solution was administered by intravenous injection into a marginal ear vein. The dose was given as a slow push (duration ranging from 35 to 63 seconds). The prepared test solutions were stored at room temperature until administered. After administration, any remaining test solutions were discarded. Reason for Route of Administration Intravenous injection is an acceptable route to assess systemic exposure. Observations of Animals Clinical observations were conducted at approximately 0.5, 2, 4, 24 and 48 hours after intravenous injection. Body weights were determined just before test material administration (Day 0). Sample Collections Blood samples (approximately 2 mL) were collected from the marginal ear vein of all animals at 2, 4, and 6 hours post-injection. Subsequent collection of blood from surviving animals was conducted at 12 and 24 hours post-injection. At the time of necropsy (approximately 48-hours post-injection) a large volume of blood (approximately 45 to 60 mL) was obtained from each surviving animal. All samples were centrifuged and separated into serum and cellular fractions. The blood samples were then stored frozen (-20'C tlO*C) until shipped to the Sponsor. Page 10 of 24 HWI 6329-117 Pathology The animal found dead during the study was subjected to an abbreviated gross necropsy examination and any abnormalities were recorded. At termination of the experimental phase, surviving animals were anesthetized with sodium pentobarbital and exsanguinated without necropsy. The bile from each animal surviving to termination was collected and immediately placed on dry ice, then frozen by placing in a freezer set to maintain a temperature of -70*C tlO*C. After necropsy or bile collection, the animals were discarded. Shipment of Blood and Bile The blood samples and bile were sent frozen (on dry ice) to the Sponsor (James D. Johnson, 3M E.E. & P.C., Bldg. 2-3E-09, 935 Bush Avenue, St. Paul, MN, 55106) after completion of the in-life phase. Statistical Analyses No statistical analyses were required by the protocol. Location of Raw Data, Records, and Final Report The raw data, records, and a copy of the final report will be retained in the archives of HWI in accordance with HWI SOP. RESULTS Body Weights Individual and mean body weights are in Table 1. Clinical Observations Individual clinical signs are in Table 2. The animals treated at 0.0, 1.0, 10.0, and 50.0 mg/kg appeared normal at 0.5 and 2 hours post-injection. The animal treated at 0.0 mg/kg exhibited a decrease in food consumption at 4, 24, and 48 hours.- The animals treated at 1.0 and 10.0 mg/kg exhibited a decrease in food consumption at 4 and 24 hours, then returned to a normal appearance at the 48-hour observation. The animal treated at 50.0 mg/kg exhibited hypoactivity at 4 and 24 hours and a decrease in food consumption at 4, 24, and 48 hours. The animal treated at 100.0 mg/kg was found dead approximately 7.5 hours post-injection following signs of hypoactivity (at 2 and 4 hours) and decreased food consumption (at 4 hours). Page 11 of 24 HWI 6329-117 Pathology The animals treated at 0.0, 1.0, 10.0, and 50.0 mg/kg survived to termination of the experimental phase and were not necropsied. The animal treated at 100 mg/kg died on the day of treatment and was necropsied. There were no visible lesions in this animal. DISCUSSION The level of systemic exposure of T-5904 was evaluated in female albino rabbits when administered as a single intravenous injection at levels of 0.0, 1.0, 10.0, 50.0, and 100.0 mg/kg using DMSO as the vehicle. Administration of this material resulted in a decrease in food consumption at all dose levels, hypoactivity at the 50.0 and 100.0 mg/kg dose levels, and death of the animal treated at the 100.0 mg/kg dose level. SIGNATURE @teven M. Glaza Study Director Acute Toxicology Date REFERENCE 1. NIH Publication No. 86-23 (revised 1985). Page 12 of 24 Group 1 2 3 4 5 Table I Individual Body Weights (g) Dose Level (mg/kq) Sex Animal Number 'Day 0 0.0 Female F50169 2,231 1.0 Female F50170 2,386 10.0 Female F50171 2,176 50.0 Female F50097 2,173 100.0 Female F50098 2,300 HWI 6329-117 GrouD 1 2 3 4 5 Dose Level (mg/kq) 0.0 Sex Female 1.0 Female 10.0 Female 50.0 Female 100.0 Female Page 13 of 24 Table 2 Individual Clinical Signs Animal Number Observation 0.5 F50169 Appeared normal Decreased food consumption F50170 Appeared normal Decreased food consumption F50171 Appeared normal Decreased food consumption F50097 Appeared normal Hypoactivity Decreased food consumption F50098 Appeared normal Hypoactivity Decreased food consumption Found dead (=7.5 hours post-injection) HWI 6329-117 Hour 2 4 24 48 - - - Indicates condition exists. Not applicable. Page 14 of 24 APPENDIX A Protocol Deviations Protocol TP8084.PK HWI 6329-117 Page 15 of 24 Protocol Deviations HWI 6329-117 Protocol Page 6, 7. Experimental Design, C. Dosing Procedures, (1) Dosing Route. Intravenous injection into a marginal ear vein over approximately 30 seconds. Actual Procedure The rate of injection ranged from 35 to 63 seconds. Page 7, 7. Experimental Design, D. Observation of Animals, (3) Sample Collections, (c) Method of Collection. Blood samples (2 mL) will be collected from the marginal ear vein. In addition, at the time of necropsy approximately 50 70 mL of blood will be obtained from each animal. A separate 2-mL sample of blood was not taken at the time of the schedule sacrifice (48 hours post-injection) since as much blood volume as possible (approximately 45 to 60 mL) was obtained from each animal. Page 8, 7. Experimental Design, E. Termination, (2) Schedule Sacrifice, (a) Sample Collection. The bile from each animal surviving to termination will be collected and immediately placed on dry ice then frozen by placing in a freezer set to maintain a temperature of -20*C 10*C. The bile samples were placed in a freezer set to maintain a temperature of -70*C 10*C. These deviations are not considered to have had an adverse effect on the outcome of the study. VIAZLMOCC:N W W I S,C 0 N S I N F'0 S T 0 F I C E 9 0 x 7 5 A 5 MAOISON WISCONSIN 53707-7545 Page 16 of 24 CC)ftP4tP4r- Sponsor: 3M St. Paul, Minnesota PROTOCOL TP8084.PK Study Title: Single-Dose Intravenous PharmacokineticStudy of T-5904 in Rabbits Date: March 4, 1994 Performing Laboratory: Hazleton Wisconsin, Inc. 3301 Kinsman Boulevard Madison, Wisconsin 53704 LaboratoryPro.i?ctIdentification: HWI 6329-117 Phon e 609 2 1 44 1 F0 P i ';S k4Aik I I , v F F4 10 v..,(, NIAL)- SON Page 17 of 24 STUDY IDENTIFICATION TP8084.PK Page 2 Single-Dose Intravenous Pharmacokinetic Study of T-5904 in Rabbits HWI No. Test Material Sponsor Sponsor's Representative Study Director Study Location Proposed Study Timetable Experimental Start Date Experimental Termination Date Draft Report Date 6329-117 T-5904 3M Toxicology Services 220-2E-02 3M Center St. Paul, MN 55144 John L. Butenhoff, PhD 3M Toxicology Services 220-2E-02 3M Center St. Paul, MN 55144 (612) 733-1962 Steven M. Glaza Hazleton Wisconsin, Inc. P.O. Box 7545 Madison, Wl 53707-7545 (608) 241-7292 Hazleton Wisconsin, Inc. Building No. 3 3802 Packers Avenue Madison, Wl 53704 Week of March 7, 1994 Week of March 7, 1994 Week of April 11, 1994 Page 18 of 24 TP8084.PK Page 3 -1. StuLd y Single-Dose Intravenous Pharmacokinetic Study in Rabbits 2. Purpose To assess the level of systemicexposure to the test material when it is administeredas a single intravenousinjectionto rabbits 3. Regulatory Compliance This study will be conducted in accordancewith the followingGood Laboratory Practice Regulations/Standards/Guidelines: [ ] Conduct as a Nonregulated Study [X] 21 CFR 58 (FDA) 40 CFR 160 (EPA-FIFRA) 40 CFR 792 (EPA-TSCA) C(81)30(Final)(OECD) NotificationNo. 3850, August 10, 1984 (JapaneseMAFF) Notification No. 313, March 31, 1982, and as amended by Notification No. 870, October 5, 1988 (JapaneseMOHW) All procedures in this protocol are in compliancewith the Animal Welfare Act Regulations. In the opinion of the Sponsor and study director, the study does not unnecessarilyduplicateany previous work. Hazletondoes not accept any responsibilityfor the analysis of the samplesor tissuescollectedin this study nor will these resultsbe presentedin the report of this study. 4. Quality Assurance The protocol,study conduct, and the final reportwill be audited by the Quality Assurance Unit in accordance with Hazleton Wisconsin (HWI) Standard Operating Procedures (SOPS) and policies. S. Test Material A. Identification T-5904 B. PhysicalDes iption Yellow solid C. Purityand Stability The Sponsor assumes responsibilityfor purity and stability determinations(includingunder test conditions). Samples of test material/vehiclmeixture(s)(if applicable)for concentration,solubility,homogeneity,and stabilityanalyses will be taken before administrationif requestedby the Sponsor. These samples (if taken) will be sent to the Sponsor after experimentalterminationfor possibleanalysis. D. Storage Room temperature Page 19 of 24 TP8084.PK Page 4 E. Reserve Samples Reserve sampleswill not be required for this study. F. Retention Any unused test materialwill be discarded after issuance of the final report, unlessdirected otherwise by the Sponsor. G. Safety Precautions As required by HWI SOPs and policies 6. Vehicle A. Identification Dimethyl Sulfoxide (DMSO); Lot number, source, and expiration date to be recordedin the data file and includedin the final report. B. Physical Description (To be documented in the raw data) C. Purity and Stability To be documentedby HWI (informationfrom the supplier). D. Storage Room temperature E. Reserve Samples See Section, 5. E. Reserve Samples F. Retention Any remainingvehiclewill be discarded after issuance of the final report. G. Safety Precautions As required by HWI SOPs and policies 7. Experimental Design A. Animals (1) Species Rabbit (2) Strain/Source Hra:(NZW)SPF/HRP, Inc. (3) Age at Initiation Adult (4) Weight at Initiation 2.0 to 3.0 kg Page 20 of 24 TP8084.PK Page 5 (5) Number and Sex 5 females (6) Identification Individualnumbered ear tag (7) Husbandry (a) Housing Individually,in screen-bottomstainlesssteel cages (heavy gauge) (b) Food A measured amount of Laboratory Rabbit Diet HF #5326 (PMI Feeds, Inc.). Animals will not be fasted before dose administration. The food is routinely analyzed by the manufacturerfor nutritionalcomponents and environmentalcontaminants. (c) Water Ad libitum from an automatic system. Samples of the water are analyzedby HWI for total dissolved solids, hardness, and specified microbiologicalcontent and for selectedelements, heavy metals, organophosphates,and chlorinated hydrocarbons. (d) Contaminants There are no known contaminants in the food or water that would interferewith this study. (e) Environment Environmentalcontrols for the animal room will be set to maintain a temperatureof 19 to 230C, a relative humidity of 50% +20%, and a 12-hour light/12-hourdark cycle. (f)@Lcclimation At least 7 days (8.)Selection of Test Animals Based on health and body weight according to HWI SOPS. An adequate number of extra animals will be purchased so that no animal in obviouslypoor health is placed on test. (9) Justificationfor SpeciesSelection Historically,the New Zealand White albino rabbit has been the animal of choice because of the large amount of background informationon this species. Page 21 of 24 TP8084.PK Page 6 B. Dose Administration (1) Test Grou-p-s- Group Dose Level (mg/kq)"- Number of Females 1 0 (Control) 1 2 1.0 1 3 10.0 1 4 50.0 1 5 100.0 1 a The dose volume will be 0.5 mL/kg of body weight C. Dosing Procedures (1) Dosing Route Intravenousinjectioninto a marginal ear vein over approximately 30 seconds. (2) Reason for Dosing Route Intravenousinjectionis an acceptable route to assess systemic exposure. (3) Dosing Duration Single dose (4) Dose Preparation The test materialwill be dilutedwith dimethyl sulfoxide (DMSO) to achievea specific concentrationfor each dose level. Individualdoses will be calculatedbased on the animal's body weight taken just before test material administration. The dose volume will be equal between all groups. The preparedtest mixtures will be stored at room temperatureuntil administration.After administration, any remaining test mixtures will be discarded. D. Observation of Animals (1) Clinical Observations The animalswill be observedfor clinical signs of toxicity at approximately0.5, 2.0, 4.0, 24, and 48 hours after treatment. (2) Body Weights Just before test materialadministration. Page 22 of 24 TP8084.PK Page 7 (3) Sample Collections (a) Frequency 2, 4, 6, 8, 12, 24, and 48 hours post-injection (b) Number of Animals All (c) Method of Collection Blood samples(2 mL) will be collectedfrom the marginalear vein. In addition,at the time of necropsy,approximately50 - 70 mL of blood will be obtained from each animal. The sampleswill be stored at room temperature and then centrifuged, and the separate serum and cellular fractionsstored at -10*C to -30'C. The separatedserum and cellular fractionswill be sent frozen to the Sponsor after experimentalterminationfor possible future analysis. Samples will be shipped to: James D. Johnson 3M E.E. & P.C. Bldg. 2-3E-09 935 Bush Avenue St. Paul, MN 55106 James D. Johnson will be notified by telephone at (612) 778-5294prior to the shipment of the samples. E. Termination (1) Unscheduled Sacrifices and Deaths An abbreviatedgross necropsywill be done. Animals in a moribund conditionwill be anesthetizedwith sodium pentobarbital and exsanguinatedwithout necropsy. (2) Scheduled Sacrifice At approximately48 hours post-injection,animals will be anesthetized with sodium pentobarbital and exsanguinated without necropsy. Page 23 of 24 TP8084.PK Page 8 (a) Saml)leCollection The bile from each animal survivingto termination will be collectedand immediatelyplaced on dry ice then frozen by placing in a freezerset to maintain a temperature of -200C 100C. The bile (frozen)will be sent on dry ice to the Sponsor after experimentalterminationfor possible future analysis. The samples will be shipped to the person listed in Section 7.D.(3).(c). F. StatisticalAnalyses No statisticalanalysesare required. B. Report A final report includingthose items listedbelow will be submitted. Descriptionof the test material Description of the test system Procedures Dates of experimentalinitiationand termination Descriptionof any toxic effects 9. Location of Raw Data, Records, and Final Report Originaldata, or copiesthereof,will be availableat HWI to facilitateauditingthe studyduring its progressand before acceptanceof the final report. When the final report is completed, all original paper data, includingthose item listed below will be retained in the archivesof HWI according to HWI SOP. Protocol and protocol amendments Dose preparation records In-life records Body weights Dose administration Observations Sample collection records Study correspondence Final report (originalsignedcopy) The following supportingrecordswill be retainedat HWI but will not be archived with the study data. Animal receipt/acclimationrecords Water analysis records Animal room temperature and humidity records Refrigerator and freezer temperaturerecords Instrument calibration and maintenance records Page 24 of 24 PROTOCOL APPROVAL ol Joh6 L. Butenhoff,PhD Sponsor's Representative 3M Steven M. Glaza Study Director Acute Toxicology Hazleton Wisconsin,Inc. /Fpr@@entative Quality Assurance Unit Hazleton Wisconsin,Inc. (6329-117.PK,dsk2) Date Date Date TP8084.PK Page 9