12 
proteins. Fte felt the fusion proteins would be unstable and rapidly degraded 
in the bacteria and in crude lysate preparations. 
Dr. Gill said this later possibility could be tested by spiking the preparation 
with vhole purified diphtheria toxin. This would give a limited answer, however, 
since fusion proteins may be degraded at different rates than the whole toxin. 
Dr. Habig said some toxicity questions could be answered by permitting the 
a-MSH-diphtheria toxin hybrid protein to be purified and tested in clinical 
trials . 
Dr. Gottesman said the working group must deal with two questions. The first 
is whether fusing the Sphl fragment with the a-MSH gene would confer on the 
hybrid molecule properties quite different from the prcperties of diphtheria 
toxin. The second is the toxicity of the Sph l fragment and any randomly 
generated fusion proteins. 
Dr. Formal said the crux of the matter is vhether the host- vector system 
carrying the hybrid molecule is safe. Che important question is vhether 
diphtheria toxin could be produced by Eb_ coll K-12 in the lumen of the bowel . 
Dr. Formal said there is no evidence K-12 produces diphtheria toxin in the 
bowel . 
Dr. Malcolm Martin of the National Institute of Allergy and Infectious Diseases 
(NIAID) said the working group should focus on an evaluation of the Eh coll 
host-vector system carrying the hybrid gene rather than on the toxicity of the 
hybrid toxin. 
Dr. Gottesman said Appendix F of the NIH Guidelines is based on the pharmacologi- 
cal activity of toxins. The working group, therefore, would consider toxin 
toxicity. Dr. Gill agreed. 
Dr. Gottesman said if the a-MSH -Sph l conjugate protein is not as toxic as diph- 
theria toxin, then containment lower than BL4 is indicated. She asked whether 
the working group is willing to lcwer containment to the RL1 level for procedures 
involving the a-MSH-Sphl conjugate protein. 
Dr. Gollier said a judgment call was made when diphtheria toxin was classified 
in Appendix F of the NIH Guidelines in 1980. Ch the basis of diphtheria toxin 
toxicity, this toxin might have been classified as requiring either BL3 or BL4 
containment. He thought the BL4 containment assigned at that time was inappro- 
priately high. He pointed out that Appendix F permits the cloning at BL1 t 
EK1 of Pseudomonas aerug inosa exotoxin A vhich is only ten to fifty times less 
toxic than diphtheria toxin. 
Dr. Habig said the pathogen elaborating diphtheria toxin, Gorynebactenum 
diphthenae , is grown at RL2 containment m clinical laboratories and large 
culture production. The NIH Guidelines also require BL2 containment for work 
with Corynebactenum diphthenae . 
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