A FURTHER ATTENUATED DERIVATIVE OF A COLD-PASSAGED TEMPERATURE-SENSITIVE MUTANT OF HUMAN RESPIRATORY SYNCYTIAL VIRUS RETAINS IMMUNOGENICITYAND PROTECTIVE EFFICACY AGAINST WILD-TYPE CHALLENGE IN SERONEGATIVE CHIMPANZEES

A cold-passaged (cp), temperature-sensitive (ts) RSV mutant designated RSV cpts-248 (shut-off temperature 38 degrees C), which possesses hos t-range mutations acquired during 52 passages at low temperature in bo vine tissue culture and a ts phenotype introduced by subsequent chemic al mutagenesis, was found previously to be attenuated, immunogenic, an d protective against wild-type challenge in seronegative chimpanzees. We sought to introduce additional attenuating mutations such as small- plaque (sp) and ts mutations into RSV cpts-248 by chemical mutagenesis with 5-fluorouracil with the intent of obtaining cpts-248 derivatives that are more attenuated in mice or chimpanzees and that are move gen etically stable following replication in vivo. Ten mutants of RSV cpts -248 which had acquired a sp phenotype or a second ts mutation were ge nerated by chemical mutagenesis. Five cpts-248 derivatives which had a cquired mutations that specified a 36 degrees C shut-off temperature f or plaque formation and one which had acquired only a sp phenotype wer e more restricted in replication in Balb/c mice than the cpts-248 pare ntal strain. One mutant, designated RSV cpts-248/404 (shut-off tempera ture 36 degrees C), was 100 times more restricted in replication in th e nasal turbinates of mice and 1000 times more restricted in the nasop harynx of seronegative chimpanzees than its cpts-248 parent. The cpts- 248/404 mutant was completely restricted in replication in the lower r espiratory tract of chimpanzees even following direct intratracheal ad ministration. The ts phenotype of the cpts-248/404 mutant was stable d uring replication in vivo in mice and chimpanzees. Chimpanzees immuniz ed with cpts-248/404 were fully protected against upper respiratory tr act disease and lower respiratory tract virus replication upon subsequ ent challenge with wild-type virus. The cpts-248/404 virus and related mutants exhibit many desirable characteristics which make them promis ing vaccine candidates.