MHC class I epitope delivering polypeptides
The present invention is directed to T-cell epitope delivering polypeptides which deliver one or more CD8+ T-cell epitopes to the MHC class I presentation pathway of a cell, including toxin-derived polypeptides which comprise embedded T-cell epitopes and are de-immunized. The present invention provides cell-targeted, CD8+ T-cell epitope delivering molecules for the targeted delivery of cytotoxicity to certain cells, e.g., infected or malignant cells, for the targeted killing of specific cell types, and the treatment of a variety of diseases, disorders, and conditions, including cancers, immune disorders, and microbial infections. The present invention also provides methods of generating polypeptides capable of delivering one or more heterologous T-cell epitopes to the MHC class I presentation pathway, including polypeptides which are 1) B-cell and/or CD4+ T-cell de-immunized, 2) comprise embedded T-cell epitopes, and/or 3) comprises toxin effectors which retain toxin functions.
1. A Shiga toxin effector polypeptide comprising an amino acid sequence having at least 90% identity to amino acids 1 to 251 of SEQ ID NO: 1;
wherein the amino acid sequence comprises at least four endogenous B-cell epitope regions, and comprises:
amino acid substitutions V54I, R55L, I57F, P59F, E60T, and E61L in SEQ ID NO: 1,
wherein the endogenous B-cell epitope regions are natively positioned in a wild-type Shiga toxin A Subunit within amino acid residues: 39-48 of SEQ ID NO:1; 94-115 of SEQ ID NO:1; 179-190 of SEQ ID NO:1; and 243-251 of SEQ ID NO:1; and
wherein the amino acid sequence comprises an asparagine at the amino acid residue corresponding to position 75 of SEQ ID NO: 1, a tyrosine at the amino acid residue corresponding to position 77 of SEQ ID NO: 1, a tyrosine at the amino acid residue corresponding to position 114 of SEQ ID NO: 1, a glutamate at the amino acid residue corresponding to position 167 of SEQ ID NO: 1, an arginine at the amino acid residue corresponding to position 170 of SEQ ID NO: 1, an arginine at the amino acid residue corresponding to position 176 of SEQ ID NO: 1, and a tryptophan at the amino acid residue corresponding to position 203 of SEQ ID NO: 1.
2. The Shiga toxin effector polypeptide according to claim 1 , wherein the amino acid sequence has at least 95% sequence identity to amino acids 1 to 251 of SEQ ID NO: 1.
3. The Shiga toxin effector polypeptide according to claim 1 , wherein the amino acid sequence has at least 95% sequence identity to SEQ ID NO: 16.