PROTEINS COMPRISING BINDING REGIONS, SHIGA TOXIN A SUBUNIT EFFECTOR REGIONS, AND CARBOXY-TERMINAL, ENDOPLASMIC RETICULUM LOCALIZATION SIGNAL MOTIFS
The present invention provides proteins comprising binding regions for cell-type specific targeting, Shiga toxin effector regions derived from A Subunits of members of the Shiga toxin family for providing Shiga toxin effector functions (e.g. cellular internalization and cytotoxicity), and carboxy-terminal endoplasmic reticulum localization signal motifs. The presently disclosed proteins can comprise additional exogenous materials, such as, e.g., antigens, cytotoxic agents, and detection-promoting agents, and are capable of targeted delivery of these additional exogenous materials into the interiors of target cells. The proteins of the present invention have uses in methods such as, e.g., methods involving targeted killing of target cells, delivering exogenous materials into target cells, labeling subcellular compartments of target cells, and diagnosing and/or treating a variety of conditions including cancers, tumors, other growth abnormalities, immune disorders, and microbial infections.
1 . A cytotoxic protein comprising:
a) a binding region comprising a single-chain variable fragment that binds at least one extracellular target molecule;
b) a Shiga toxin effector region comprising a sequence that is at least 95% identical to amino acids 1 to 251 of SEQ ID NO:1;
wherein the amino acid residue corresponding to position 75 of SEQ ID NO: 1 is asparagine, the amino acid residue corresponding to position 77 of SEQ ID NO: 1 is tyrosine, the amino acid residue corresponding to position 167 of SEQ ID NO: 1 is glutamate, the amino acid residue corresponding to position 170 of SEQ ID NO: 1 is arginine, and the amino acid residue corresponding to position 176 of SEQ ID NO: 1 is arginine; and
c) a carboxy-terminal endoplasmic reticulum retention/retrieval signal motif of a member of the KDEL family.