Document LXnGJe4GNRdwn9gprk52jMdq

M 2.3.6. 0663 N-EtFOSA Pharmacokinetic - / 000475 HAZLETON WISCONSIN P O S T OFFICE B O X 7545 M A D IS O N. /v i 5 3 7 0 7- 7 5 4 5 Sponsor: 3M St. Paul, Minnesota rt C O R N IN G CcmiJ.'iny Phone F 'PRF S S ' 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 Laboratory: Hazleton Wisconsin, Inc. 3301 Kinsman Boulevard Madison, Wisconsin 53704 Laboratory Project Identification: HWI 6329-117 Page 1 of 24 p,i 7 r; F a - h r; -, 2 4 1 7 ?.' m a n i'Io . b ' 70 4 000476 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 monthly according 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 Nancy Diedrich Representtative, Quality Assurance Unit !0' f4- eTti Date 000477 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.0. Box 7545 Madison, WI 53707-7545 (608) 241-7292 Hazleton Wisconsin, Building No. 3 3802 Packers Avenue Madison, WI 53704 Inc. March 7, 1994 March 9, 1994 000478 Acute Toxicology Steven M. Glaza Study Director Manager Steven R. Sorenson Study Coordinator Patricia Padgham In-life Supervisor Rose M. Bridge Report Supervisor Quality Assurance Sherry R. W. Petsel Manager Page 4 of 24 HWI 6329-117 KEY PERSONNEL Laboratory Animal Medicine Cindy J. Cary, DVM Dipl ornate, ACLAM Supervisor Anatomical Pathology Jack Serfort/ Deborah L. Pirkel Supervisors Necropsy Anne Mosher Supervisor Pathology Data 000479 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 Page 32 4 6 7 7 7 98 11111110 1132 1154 16 000480 Page 6 of 24 HWI 6329-117 SUMMARY 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)] 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. 000481 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's Good Laboratory Practice Regulations for Nonclinical Laboratory Studies, 21 CFR 58. All procedures used in this study are in compliance with the Animals Welfare Act Regulations. In the opinion of the Sponsor and study director, the study did not unnecessarily duplicate 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. 53H0634; Exp. March 4, 1995) and described as a clear, colorless liquid. Purity and Stability The Sponsor assumes responsibility for test material purity and stability determinations (including under test conditions). A sample of the test material/vehicle mixtures for concentration, solubility, homogeneity, and stability analyses was not taken before administration as this was not requested by the Sponsor. The certificate of analysis, obtained from the supplier, documents the purity of the control material as 99.9%. The stability of 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). 000482 Page 8 of 24 HWI 6329-117 Safety Precautions 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".1 The animals were individually housed in screenbottom stainless steel cages in temperature- and humidity-controlled quarters, provided access to water ad libitum and a measured amount of Laboratory Rabbit Diet HF #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: Grouo Dose Level Dose Volume Number Treatment (mq/kq) (mL/kq) of Animals 1 (Control) 2 3 4 5 DMSO T-5904 T-5904 T-5904 T-5904 0.0 1.0 10.0 50.0 100.0 0.5 0.5 0.5 0.5 0.5 1 1 1 1 1 000483 Page 9 of 24 HWI 6329-117 Justification for Species Selection 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 10*C) until shipped to the Sponsor. 000484 Page 10 of 24 KWI 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 +10*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). 000485 Page 11 of 24 i I Pathology HWI 6329-117 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 DMS0 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 Steven M. Glaza Study Director Acute Toxicology REFERENCE 1. NIH Publication No. 86-23 (revised 1985). 000486 Page 12 of 24 Group 1 2 3 4 5 Table 1 Individual Body Weights (g) Dose Level (mq/kq) Sex Animal Number 0.0 Female F50169 1.0 Female F50170 10.0 Female F50171 50.0 Female F50097 100.0 Female F50098 Dav 0 2,231 2,386 2,176 2,173 2,300 HWI 6329-117 000487 GrouD 1 2 3 4 5 Page 13 of 24 HWI 6329-117 Table 2 Individual Clinical Signs Dose Level fma/ka) Sex Animal Number Observation Hour 0.5 2 4 24 48 0.0 Female F50169 Appeared normal / - - Decreased food consumption - / 1.0 Female F50170 Appeared normal / - - Decreased food consumption -- - 10.0 50.0 Female Female F50171 F50097 Appeared normal Decreased food consumption Appeared normal Hypoactivity Decreased food consumption - - - - / __ - */ / - -/ 100.0 Female F50098 Appeared normal Hypoactivity -/ Decreased food consumption - / Found dead (=7.5 hours post-injection) / / Indicates condition exists. - Not applicable. 000488 Page 14 of 24 APPENDIX A Protocol Deviations Protocol TP8084.PK HWI 6329-117 000489 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. 000490 HAZLETON WpMoA DsIItS OSNo WfCfI SiCcOOeN SNI Na So x I 7 SN4 S 53707-7545 Page 16 of 24 COINING l.fiK t++'+C*r><fVfl'tt'ty Sponsor 3M St. Paul, Minnesota PROTOCOL TP8084.PK Study Title: Single-Dose Intravenous Pharmacokinetic Study of T-5904 in Rabbits Date: March 4, 1994 Performing Laboratory: Hazleton Wisconsin, Inc. 3301 Kinsman Boulevard Madison, Wisconsin 53704 Laboratory Project Identification: HWI 6329-117 F Of t f X **H f S S M A I l t ) f l i v f y J 3 O I * I Nr. M A N K O U I [ VAIMI M A U <S O N Wl',< O M '.iN 000491 Page 17 of 24 TP8084.PK Page 2 --, STUDY IDENTIFICATION 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, WI 53707-7545 (608) 241-7292 Hazleton Wisconsin, Inc. Building No. 3 3802 Packers Avenue Madison, WI 53704 Week of March 7, 1994 Week of March 7, 1994 Week of April 11, 1994 000492 Page 18 of 24 TP8084.PK Page 3 -i. Study Single-Dose Intravenous Pharmacokinetic Study in Rabbits 2. Purpose To assess the level of systemic exposure to the test material when it is administered as a single intravenous injection to rabbits 3. Regulatory Compliance This study will be conducted in accordance with the following Good 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) [ ) Notification No. 3850, August 10, 1984 (Japanese MAFF) [ ] Notification No. 313, March 31, 1982, and as amended by Notification No. 870, October 5, 1988 (Japanese MOHW) All procedures in this protocol are in compliance with the Animal Welfare Act Regulations. In the opinion of the Sponsor and study director, the study does not unnecessarily duplicate any previous work. Hazleton does not accept any responsibility for the analysis of the samples or tissues collected in this study nor will these results be presented in the report of this study. 4. Quality Assurance The protocol, study conduct, and the final report will be audited by the Quality Assurance Unit in accordance with Hazleton Wisconsin (HWI) Standard Operating Procedures (SOPs) and policies. 5. Test Material A. Identification T-5904 B. Physical Description Yellow solid C. Purity and Stability The Sponsor assumes responsibility for purity and stability determinations (including under test conditions). Samples of test material/vehicle mixture(s) (if applicable) for concentration, solubility, homogeneity, and stability analyses will be taken before administration if requested by the Sponsor. These samples (if taken) will be sent to the Sponsor after experimental termination for possible analysis. D. Storage Room temperature 000493 Page 19 of 24 TP8084.PK Page 4 E. Reserve Samples Reserve samples will not be required for this study. F. Retention Any unused test material will be discarded after issuance of the final report, unless directed 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 recorded in the data file and included in the final report. B. Physical Description (To be documented in the raw data) C. Purity and Stability To be documented by HWI (information from the supplier). D. Storage Room temperature E. Reserve Samples See Section, 5. E. Reserve Samples F. Retention Any remaining vehicle will be discarded after issuance of the final report. G. Safety Precautions As required by HWI SOPs and policies 7. Experimental Design 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 000494 Page 20 of 24 TP8084.PK Page 5 (5) Number and Sex 5 females (6) Identification Individual numbered ear tag (7) Husbandry (a) Housing Individually, in screen-bottom stainless steel 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 manufacturer for nutritional components and environmental contaminants. (c) Water Ad libitum from an automatic system. Samples of the water are analyzed by HWI for total dissolved solids, hardness, and specified microbiological content and for selected elements, heavy metals, organophosphates, and chlorinated hydrocarbons. (d) Contaminants There are no known contaminants in the food or water that would interfere with this study. (e) Environment Environmental controls for the animal room will be set to maintain a temperature of 19 to 23C, a relative humidity of 50X 20%, and a 12-hour 1ight/12-hour dark cycle. (f) Acclimation 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 obviously poor health is placed on test. (9) Justification for Soecies Selection Historically, the New Zealand White albino rabbit has been the animal of choice because of the large amount of background information on this species. 00Q495 Page 21 of 24 TP8084.PK Page 6 B. Dose Administration (1) Test Groups Group Dose Level (mg/MU Number of Females 21 3 0 (Control) 110..00 1 1 1 4 5 15000..00 1 1 a The dose volume will be 0.5 mL/kg of body weight C. Dosino Procedures (1) Dosino Route Intravenous injection into a marginal ear vein over approximately 30 seconds. (2) Reason for Dosino Route Intravenous injection is an acceptable route to assess systemic exposure. (3) Dosino Duration Single dose (4) Dose Preparation The test material will be diluted with dimethyl sulfoxide (DMS0) to achieve a specific concentration for each dose level. Individual doses will be calculated based on the animal's body weight taken just before test material administration. The dose volume will be equal between all groups. The prepared test mixtures will be stored at room temperature until administration. After administration, any remaining test mixtures will be discarded. D. Observation of Animals (1) Clinical Observations The animals will be observed for clinical signs of toxicity at approximately 0.5, 2.0, 4.0, 24, and 48 hours after treatment. (2) Body Weights Just before test material administration. 000496 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 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. The samples will be stored at room temperature and then centrifuged, and the separate serum and cellular fractions stored at -10C to -30C. The separated serum and cellular fractions will be sent frozen to the Sponsor after experimental termination for 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) 778-5294 prior to the shipment of the samples. E. Termination (1) Unscheduled Sacrifices and Deaths An abbreviated gross necropsy will be done. Animals in a moribund condition will be anesthetized with sodium pentobarbital and exsanguinated without necropsy. (2) Scheduled Sacrifice At approximately 48 hours post-injection, animals will be anesthetized with sodium pentobarbital and exsanguinated without necropsy. 000497 Page 23 of 24 TP8084.PK Page 8 ** (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 -20C 10C. The bile (frozen) will be sent on dry ice to the Sponsor after experimental termination for possible future analysis. The samples will be shipped to the person listed in Section 7.D.(3). (c). F. Statistical Analyses No statistical analyses are required. 8. Report A final report including those items listed below will be submitted. Description of the test material Description of the test system Procedures Dates of experimental initiation and termination Description of any toxic effects 9. Location of Raw Data. Records, and Final Report Original data, or copies thereof, will be available at HWI to facilitate auditing the study during its progress and before acceptance of the final report. When the final report is completed, all original paper data, including those item listed below will be retained in the archives of 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 (original signed copy) The following supporting records will be retained at HWI but will not be archived with the study data. Animal receipt/acclimation records Water analysis records Animal room temperature and humidity records Refrigerator and freezer temperature records Instrument calibration and maintenance records 000498 Page 24 of 24 PROTOCOL APPROVAL TP8084.PK Page 9 Johft L. Butenhoff, P h D ' Sponsor's Representative 3M Steven M. Glaza Study Director Acute Toxicology Hazleton Wisconsin, Inc. O Representative Quality Assurance Unit Hazleton Wisconsin, Inc. (6329-117.PK,dsk2) Date Date Date 000499