Engineered immune cells with receptor signal strength modulated by a hinge
The present disclosure provides engineered immune cells and methods for their creation and use. The immune cells comprise activating and blocking receptors. The signals of the activating and blocking receptors can be modulated via the identity of a hinge.
1. A method of producing an engineered immune cell, the method comprising:
determining a strength of a blocking signal sent from a cell surface blocking receptor caused to be expressed on an engineered immune cell from an introduced vector encoding said blocking receptor, said strength determined at least in-part due to a length of a hinge of the cell surface blocking receptor, wherein the length of the hinge is known, wherein a longer hinge lengths impart an increased blocking signal strength of the cell surface blocking receptor,
wherein the hinge comprises a peptide derived from leukocyte immunoglobulin-like receptor subfamily B member 1 (LILRB1) of between 10 and 64 amino acids in length; and
producing an engineered immune cell that expresses the cell surface blocking receptor having the hinge with a known length and a cell surface activating receptor that sends an activating signal that promotes a cytotoxic response and that is inhibited by the blocking signal,
said blocking and activating receptor each expressed from an introduced vector encoding the receptor and each receptor comprising an extracellular binding domain, a hinge, a transmembrane domain, and an intracellular domain.
2. The method of claim 1 , wherein cross-talk and/or structure function interactions between the hinge of the blocking receptor and the activating receptor further impart different signal strengths for the blocking receptor.
3. The method of claim 1 , wherein the increased blocking signal strength increases a half maximal effective concentration (EC 50 ) of an activating ligand for the activating receptors, wherein binding of the activating ligand to the activating receptor causes the receptor to send the activating signal to promote the cytotoxic response.
4. The method of claim 3 , wherein the engineered immune cell is caused to express the blocking and activating receptors at a ratio, and the ratio blocking receptors to activating receptors expressed is decreased as the length of the peptide of the hinge is increased.
5. The method of claim 3 , wherein the peptide of the hinge has a degree of flexibility, and the peptide's length and degree of flexibility modulate the blocking signal strength of the blocking receptor on the activating signal.
6. The method of claim 5 , wherein the hinge is selected from hinges that each have a known effect on the EC 50 of the activating ligand for the activating receptors to promote the cytotoxic response.
7. The method of claim 6 , wherein the length of the peptide of each hinge that has a known effect on the EC 50 of the activating ligand for the activating receptor is between 10 and 64 amino acids in length.
8. The method of claim 6 , wherein the peptide of the selected hinge is at least 24 amino acids in length.
9. The method of claim 6 , wherein the peptide of the selected hinge is at least 64 amino acids in length.
10. The method of claim 7 , wherein a hinge having a peptide of 64 amino acids in length causes at least a fifty-fold increase in the EC 50 relative to a hinge having a peptide of 10 amino acid in length.
11. The method of claim 8 , wherein the hinges that each have a known effect on the EC50 comprise hinges 2B1 and 2B1 truncated.
12. The method of claim 5 , wherein the flexible peptide comprises glycine-glutamine repeats and/or glycine-serine repeats.
13. The method of claim 1 , wherein the hinge comprises a rigid peptide that modulates the effect of the blocking signal on the activating signal, and the effect is a reduced inhibition of the activating signal.