Amendment #1 dated Aug 20, l973 specified 4 additional clinical chemistry laboratory measurements: 1.) serum insulin, 2.)serum ornithine carbamyl transferase, 3.) serum protein electrophoresis, 4.) serum total protein.

Two of the above assays (serum insulin, and serum ornithine carbamyl transferase) were apparently not done, because no data for these two parameters was submitted to FDA, and we could find no raw data or other evidence that they were done.

Amendment #2 dated Aug 20, l973, specified 8 coronal sections of brain to be examined microscopically, and also described the procedure for sectioning the urinary bladder.  Four transverse sections from each urinary bladder were to be examined microscopically.

Amendment #3 dated Sept. 6, l973 extended the study until it reached a point where mortality reduced the control group to 20 animals per sex, provided survival of treated groups was not less than 10 per sex per group.  (This represented a survival of approximately 30%).

Amendment #4 dated Jan 9, l974 added serum cholesterol to the clinical chemistry measurements to be made at terminal sacrifice, and terminated the study after 114 weeks of treatment.  Terminal sacrifice was to begin on 1-24-74 and continue through 2-1-74.

Our examination of the original data showed that serum cholesterol determinations were done at day 796 and 798 (terminal bleeding) as specified in the above amendment, but the data was not included in the submission to FDA.  The submission to FDA (Vol. 1 p. 286) reported a significant decrease in serum cholesterol that was more perceptible towards the end of the study, and may have been related to compound administration.  Therefore, the omitted data may have been important.

Serum cholesterol determinations were also done at day 546 (78 weeks) and not reported in the submission to FDA.

The protocol for Clinical Chemistry procedures specified that BUN determinations were to be done at 78 weeks (546 days).  The submission to FDA contained no BUN data for day 546, but our review of the raw data indicated that BUN's had been done at day 546.  Some BUN's were also done at day 735 (105 weeks) and not reported in the submission to FDA, but this data was not complete for all animals.

Attached to the protocol is a memo dated Oct. 31, l972 which describes an acute infection spreading in the rat colony, and the administration of penicillin to combat the infection, and a memo dated May 8, l973 listing scheduled dates to be added to Body and Feeder Weights of housing groups A & B.

The final Histology Lab Protocol, dated 1-21-74, specifies 24 organs to be embedded for control and high dose animals, and 19 organs to be embedded for low and mid dose groups.  The organs which were to be embedded for the control and high dose groups but to be omitted in the low and mid dose groups include:  lymph node, nerve, bone, eye, and salivary glands.

Pathology sheets (blank forms) to be used at terminal sacrifice were reproduced (xeroxed) with check marks, time (death to tissue fix), fixative, study, and project number already entered.  Twenty-seven (27) organs were checked off, to be embedded.  However, as stated above, the control and high dose animals were to have 24 organs embedded, according to the protocol, and the mid and low dose 19.  Therefore, all pathology sheets for animals killed by design have incorrectly identified the specific organs and tissues to be embedded.