Dr. Wallace Rowe 
Page 2 
In order to obtain a reasonable assessment of the actual immunologic 
risks entailed by development of EL_ coli K12 strains expressing 
eukaryotic proteins, a series of issues need to be considered. These 
include : 
(1) What is the likelihood that a human protein or a protein 
cross-reactive with a human protein, if associated with E_^ coli K12 
could actually induce autoantibodies or auto-reactive T lymphocytes? 
(2) If such induction is possible, how large an antigenic load 
is required and for what period of time is exposure necessary? Is 
exposure through the GI tract likely to be an effective stimulant of 
autoantibodies? 
(3) How likely is it that the induction of such autoantibodies 
or auto-reactive T cells would have pathologic consequences? 
(A) Is it likely that such immunopathology , once initiated, 
would perpetuate itself beyond the time the stimulatory antigen 
was present? 
(5) If a risk of autoantibody production and/or immunopathology 
exists, is it one which would be limited to those who work directly 
with the agent or would it be a risk to the general population? 
We will attempt to develop discussions on these issues in the following 
paragraphs. 
The fact that autoantibodies (i.e. antibodies reactive with undenatured 
antigenic determinants of the individual producing the antibodies) can 
develop after immunization with native, denatured, or cross-reactive 
antigens is well established, as already noted earlier in this letter. 
Among the best studied systems are responses to thyroglobulin and to the 
acetylcholine receptor. Immunization of rabbits with homologous thyro- 
globulin incorporated into complete Freund's adjuvant has been shown to 
stimulate production of anti-thyroglobulin and to induce thyroiditis 
(1) . Similarly denatured, haptenated or heterologous thyroglobulin 
administered in aqueous form can cause the production of anti-thyro- 
globulin antibodies and the development of thyroid lesions. In general, 
immunization with native thyroglobulins in saline fails to initiate 
either antibody production or thyroiditis. In the bulk of the experi- 
mental systems reported, the doses of thyroglobulin used to produce 
autoimmunity were quite high, of the order of 6 mg/animal. Whether 
the autoimmune state persists after the initial treatment is terminated 
is not entirely clear. However, rabbits in which autoimmunity was 
induced by injection of denatured thyroglobulin will often reenter the 
tolerant or nonimmune state upon subsequent treatment with native thyro- 
globulin. 
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