Natural killer t (nkt) cells comprising engineered t cell receptors

A method for increasing the cell surface expression of a T cell receptor (TCR) in a NKT cell which comprises the steps of:

• (i) providing a TCR α chain and/or β chain sequence;
• (ii) determining an amino acid residue of the TCR α chain and/or β chain at one or more positions selected from:
• 96 of the α chain;
• 9 of the β chain;
• 10 of the β chain;
• 24 of the α chain;
• 19 of the α chain;
• 20 of the α chain;
• 50 of the α chain;
• 5 of the α chain;
• 8 of the α chain;
• 86 of the α chain;
• 39 of the α chain;
• 55 of the α chain;
• 43 of the β chain;
• 66 of the α chain;
• 19 of the β chain;
• 21 of the β chain;
• 103 of the β chain;
• 3 of the α chain;
• 7 of the α chain;
• 9 of the α chain;
• 11 of the α chain;
• 16 of the α chain;
• 18 of the α chain;
• 21 of the α chain;
• 22 of the α chain;
• 26 of the α chain;
• 40 of the α chain;
• 47 of the α chain;
• 48 of the α chain;
• 49 of the α chain;
• 51 of the α chain;
• 52 of the α chain;
• 53 of the α chain;
• 67 of the α chain;
• 68 of the α chain;
• 74 of the α chain;
• 76 of the α chain;
• 79 of the α chain;
• 81 of the α chain;
• 83 of the α chain;
• 90 of the α chain;
• 92 of the α chain;
• 93 of the α chain;
• and 101 of the α chain;
• (iii) altering the amino acid residue at one or more of the positions listed in step (ii) to an amino acid residue selected from:
• an aliphatic, non-polar amino acid or a methionine at position 96 of the α chain;
• an aliphatic, polar charged amino acid or a glutamine at position 9 of the β chain;
• an aromatic amino acid at position 10 of the β chain;
• an aliphatic, polar uncharged amino acid at position 24 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 19 of the α chain;
• an aliphatic, polar uncharged amino acid at position 20 of the α chain;
• an aliphatic, polar uncharged amino acid or a isoleucine, leucine or valine at position 50 of the α chain;
• an aliphatic, polar uncharged amino acid at position 5 of the α chain;
• an aliphatic, polar uncharged amino acid, a glutamic acid, lysine or arginine at position 8 of the α chain;
• an aliphatic, polar uncharged amino acid or alanine at position 86 of the α chain;
• an aromatic amino acid at position 39 of the α chain;
• an aliphatic, polar charged amino acid or asparagine at position 55 of the α chain;
• an aliphatic, polar charged amino acid or a glutamine at position 43 of the β chain;
• an aliphatic, non-polar amino acid or a serine at position 66 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 19 of the β chain;
• an aliphatic, non-polar amino acid or a methionine at position 21 of the β chain;
• an aliphatic, non-polar amino acid or a methionine at position 103 of the β chain;
• an aliphatic, polar uncharged amino acid at position 3 of the α chain;
• an aliphatic, polar uncharged amino acid or alanine at position 7 of the α chain;
• an aliphatic, non-polar amino acid at position 9 of the α chain;
• an aliphatic, non-polar amino acid or a isoleucine, leucine or valine at position 11 of the α chain;
• an aliphatic, non-polar amino acid or a serine at position 16 of the α chain;
• an aliphatic, polar uncharged amino acid at position 18 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 21 of the α chain;
• an aliphatic, polar uncharged amino acid or alanine at position 22 of the α chain;
• an aliphatic, polar charged amino acid or asparagine at position 26 of the α chain;
• an aromatic amino acid at position 40 of the α chain;
• an aliphatic, polar uncharged amino acid or alanine at position 47 of the α chain;
• an aliphatic, polar charged amino acid or a glutamine at position 48 of the α chain;
• an aliphatic, polar uncharged amino acid, glutamic acid, lysine or arginine at position 49 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 51 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 52 of the α chain;
• an aliphatic, non-polar amino acid or a methionine at position 53 of the α chain;
• an aliphatic, polar uncharged amino acid at position 67 of the α chain;
• an aliphatic, polar charged amino acid or a glutamine at position 68 of the α chain;
• an aliphatic, polar uncharged amino acid or an aspartic acid or histidine at position 74 of the α chain;
• an aromatic amino acid at position 76 of the α chain;
• an aliphatic, polar uncharged amino acid or an aspartic acid or histidine at position 79 of the α chain;
• an aliphatic, polar uncharged amino acid, glutamic acid, lysine or arginine at position 81 of the α chain;
• an aliphatic, non-polar amino acid or a serine at position 83 of the α chain;
• an aliphatic, polar charged amino acid or a glutamine at position 90 of the α chain;
• an aliphatic, polar uncharged amino acid or alanine at position 92 of the α chain;
• an aliphatic, polar charged amino acid or asparagine at position 93 of the α chain;
• an aliphatic, non-polar amino acid or a isoleucine, leucine or valine at position 101 of the α chain;
• and
• (iv) expressing a polynucleotide encoding the engineered TCR produced in step (iii) in a NKT cell.