Compositions of engineered exosomes and methods of loading luminal exosomes pay-loads

A polypeptide for modifying an exosome, comprising a sequence of (i) (M)(G)(G/A/S)(K/Q)(L/F/S/Q)(S/A)(K)(K) (SEQ ID NO:

• 118);
• (ii) (M)(G)(π)(X)(Φ/π)(π)(+)(+), wherein each parenthetical position represents an amino acid, and wherein π is any amino acid selected from the group consisting of (Pro, Gly, Ala, Ser), X is any amino acid, Φ is any amino acid selected from the group consisting of (Val, Ile, Leu, Phe, Trp, Tyr, Met), and (+) is any amino acid selected from the group consisting of (Lys, Arg, His);
• and wherein position five is not (+) and position six is neither (+) nor (Asp or Glu);
• or (ii) (M)(G)(π)(ξ)(Φ/π)(S/A/G/N)(+)(+), wherein each parenthetical position represents an amino acid, and wherein π is any amino acid selected from the group consisting of (Pro, Gly, Ala, Ser), ξ is any amino acid selected from the group consisting of (Asn, Gln, Ser, Thr, Asp, Glu, Lys, His, Arg), Φ is any amino acid selected from the group consisting of (Val, Ile, Leu, Phe, Trp, Tyr, Met), and (+) is any amino acid selected from the group consisting of (Lys, Arg, His);
• and wherein position five is not (+) and position six is neither (+) nor (Asp or Glu).

A method of making an engineered exosome, comprising the steps of:

• a. introducing into a cell a nucleic acid construct encoding a fusion polypeptide comprising (i) a first sequence encoding MARCKS, MARCKSL1, BASP1 or a fragment or a modification thereof, and (ii) a second sequence encoding a cargo peptide;
• b. maintaining the cell under conditions allowing the cell to express the fusion polypeptide;
• and
• c. obtaining the engineered exosome comprising the fusion polypeptide from said cell.