Page 27 of Attachment H 
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A 14-day subacute toxicity study of crystallized asparaginase was conducted 
in the dog and monkey at doses of 200, 1000 and 5000 I.U./kg body weight, 
and in the rabbit at 50, 100 and 200 I.U./kg. The rabbit had been found by 
other groups to be very sensitive to asparaginase toxicity, and our study 
confirmed this. A one-month subacute toxicity study in the dog and monkey 
was then done at 1000 I.U./kg. Toxicity encountered in both of these 
studies with crystallized asparaginase was relatively mild, and no serious 
toxicity was observed which would contraindicate a cautious clinical trial 
of the enzyme. Subsequently, clinical evaluation of our crystallized 
asparaginase was done by groups at St. Jude Children's Hospital, Memphis, 
and at the Sloan-Kettering Memorial Hospital, New York. In brief, no 
significant differences in clinical toxicity or efficacy were observed for 
the crystallized enzyme over other purified asparaginase preparations. 
In summary, crude preparations of _E. col i l-asparaginase produced side 
effects such as chills, fever, nausea and vomiting which appeared to be 
associated with contamination by pyrogens and bacterial endotoxin. These 
particular side effects were minimal when highly purified preparations of 
the enzyme were administered. However, the principal toxic effects were 
observed with the purified enzyme as well as with the cruder preparations . 
The immunogenic nature of this large, foreign protein of 130,000 daltons and 
the enzymatic activity, per se , both asparaginase and glutaminase activity, 
comprise the chief reasons for the observed clinical and toxicologic effects. 
[ 463 ] 
