Multi-chain chimeric polypeptides and uses thereof
The present disclosure relates to methods that include the use of a first multi-chain chimeric polypeptide and a second multi-chain chimeric polypeptide for stimulating the NK cells, inducing or increasing proliferation of the NK cells, inducing differentiation of the NK cells, and treating a subject in need thereof using activated NK cells.
1. A method of promoting the activation and proliferation of a natural killer cell, the method comprising:
(a) contacting a natural killer cell in a liquid culture medium comprising an effective amount of a first multi-chain chimeric polypeptide for 15 minutes to one day under conditions that allow for differentiation of the natural killer (NK) cell; and
(b) contacting the natural killer cell in a liquid culture medium comprising an effective amount of (i) a second multi-chain chimeric polypeptide, and (ii) an IgG1 antibody construct, for 1 day to 30 days under conditions that allow for the activation and proliferation of the natural killer cell,
wherein the first multi-chain chimeric polypeptide comprises:
a first chimeric polypeptide comprising a first target-binding domain, a soluble tissue factor domain comprising a sequence that is at least 90% identical to SEQ ID NO: 5, and a first domain of a pair of affinity domains comprising a sequence that is at least 90% identical to SEQ ID NO: 26, and
a second chimeric polypeptide comprising a second domain of a pair of affinity domains comprising a sequence that is at least 90% identical to SEQ ID NO: 24 and a second target-binding domain,
wherein the first chimeric polypeptide and the second chimeric polypeptide in the first multi-chain chimeric polypeptide associate through the binding of the first domain and the second domain of the pair of affinity domains in the first multi-chain chimeric polypeptide;
wherein (A) the first target-binding domain in the first multi-chain chimeric polypeptide comprises a sequence at least 90% identical to SEQ ID NO: 20 and the second target-binding domain in the first multi-chain chimeric polypeptide comprises a first sequence that is at least 90% identical to SEQ ID NO: 14 and a second sequence that is at least 90% identical to SEQ ID NO: 16, or (B) the first target-binding domain in the first multi-chain chimeric polypeptide comprises a first sequence that is at least 90% identical to SEQ ID NO: 14 and a second sequence that is at least 90% identical to SEQ ID NO: 16, and the second target-binding domain in the first multi-chain chimeric polypeptide comprises a sequence at least 90% identical to SEQ ID NO: 20; and
wherein the second multi-chain chimeric polypeptide comprises:
a first chimeric polypeptide comprising a first target-binding domain, a soluble tissue factor domain comprising a sequence that is at least 90% identical to SEQ ID NO: 5, a first domain of a pair of affinity domains comprising a sequence that is at least 90% identical to SEQ ID NO: 26, and
a second chimeric polypeptide comprising a second domain of a pair of affinity domains comprising a sequence that is at least 90% identical to SEQ ID NO: 24, and a second target-binding domain,
wherein the first chimeric polypeptide and the second chimeric polypeptide in the second multi-chain chimeric polypeptide associate through the binding of the first domain and the second domain of the pair of affinity domains in the second multi-chain chimeric polypeptide;
wherein (A) the first target-binding domain in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 23 and the second target-binding domain in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 22, or (B) the first target-binding domain in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 22 and the second target-binding domain in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 23; and
wherein the IgG1 antibody construct comprises at least one antigen-binding domain that binds specifically to the soluble tissue factor domain.
2. The method of claim 1 , wherein the first target-binding domain and the soluble tissue factor domain directly abut each other in the first chimeric polypeptide in the first multi-chain chimeric polypeptide.
3. The method of claim 1 , wherein the first chimeric polypeptide in the first multi-chain chimeric polypeptide further comprises a linker sequence between the first target-binding domain and the soluble tissue factor domain in the first chimeric polypeptide.
4. The method of claim 1 , wherein the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide in the first multi-chain chimeric polypeptide directly abut each other in the first chimeric polypeptide.
5. The method of claim 1 , wherein the first chimeric polypeptide in the first multi-chain chimeric polypeptide further comprises a linker sequence between the soluble tissue factor domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
6. The method of claim 1 , wherein the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide in the first multi-chain chimeric polypeptide directly abut each other in the second chimeric polypeptide.
7. The method of claim 1 , wherein the second chimeric polypeptide in the first multi-chain chimeric polypeptide further comprises a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
8. The method of claim 1 , wherein the soluble tissue factor domain is a soluble human tissue factor domain.
9. The method of claim 8 , wherein the soluble human tissue factor domain does not comprise one or more of:
a lysine at an amino acid position that corresponds to amino acid position 20 of mature wildtype human tissue factor protein;
an isoleucine at an amino acid position that corresponds to amino acid position 22 of mature wildtype human tissue factor protein;
a tryptophan at an amino acid position that corresponds to amino acid position 45 of mature wildtype human tissue factor protein;
an aspartic acid at an amino acid position that corresponds to amino acid position 58 of mature wildtype human tissue factor protein;
a tyrosine at an amino acid position that corresponds to amino acid position 94 of mature wildtype human tissue factor protein;
an arginine at an amino acid position that corresponds to amino acid position 135 of mature wildtype human tissue factor protein; and
a phenylalanine at an amino acid position that corresponds to amino acid position 140 of mature wildtype human tissue factor protein.
10. The method of claim 1 , wherein the first multi-chain chimeric polypeptide and the second multi-chain chimeric polypeptide do not stimulate coagulation in a mammal.
11. The method of claim 1 , wherein the first target-binding domain and the linker domain in the first chimeric polypeptide in the second multi-chain chimeric polypeptide directly abut each other in the first chimeric polypeptide.
12. The method of claim 1 , wherein the first chimeric polypeptide in the second multi-chain chimeric polypeptide further comprises a linker sequence between the first target-binding domain and the linker domain in the first chimeric polypeptide.
13. The method of claim 1 , wherein the linker domain and the first domain of the pair of affinity domains directly abut each other in the first chimeric polypeptide in the second multi-chain chimeric polypeptide.
14. The method of claim 1 , wherein the first chimeric polypeptide in the second multi-chain chimeric polypeptide further comprises a linker sequence between the linker domain and the first domain of the pair of affinity domains in the first chimeric polypeptide.
15. The method of claim 1 , wherein the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide of the second multi-chain chimeric polypeptide directly abut each other in the second chimeric polypeptide.
16. The method of claim 1 , wherein the second chimeric polypeptide in the second multi-chain chimeric polypeptide further comprises a linker sequence between the second domain of the pair of affinity domains and the second target-binding domain in the second chimeric polypeptide.
17. The method of claim 1 , wherein the IgG1 antibody construct is a monoclonal IgG1 antibody, where both antigen-binding domains in the monoclonal IgG1 antibody bind specifically to the soluble tissue factor domain.
18. The method of claim 1 , wherein the first period of time is 15 minutes to 4 hours.
19. The method of claim 1 , wherein the liquid culture medium in step (b) comprises the second multi-chain chimeric polypeptide and the IgG1 antibody construct at a molar ratio of about 0.5:1 to about 2:1.
20. The method of claim 1 , wherein the NK cell was previously obtained from a subject.
21. The method of claim 20 , wherein the method further comprises obtaining the NK cell from the subject prior to step (a).
22. The method of claim 1 , wherein the NK cell has previously been genetically modified to express a chimeric antigen receptor or a recombinant T-cell receptor.
23. The method of claim 1 , wherein the method further comprises, after step (b), isolating the NK cell.
24. The method of claim 1 , wherein the method further comprises, after the contacting step, administering the NK cell to a subject identified or diagnosed as having a cancer.
25. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 67; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 69.
26. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 67; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 69.
27. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 67; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 69.
28. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 64;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 66;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 68; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 70.
29. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 71; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 73.
30. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 71; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 73.
31. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 63;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 65;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 71; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 73.
32. The method of claim 1 , wherein:
the first chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 64;
the second chimeric polypeptide in the first multi-chain chimeric polypeptide comprises SEQ ID NO: 66;
the first chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 72; and
the second chimeric polypeptide in the second multi-chain chimeric polypeptide comprises SEQ ID NO: 74.
33. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 20;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence that is at least 90% identical to SEQ ID NO: 14 and a second sequence that is at least 90% identical to SEQ ID NO: 16;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 23; and
the second target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 22.
34. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 20:
the soluble tissue factor domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 5;
the first domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 26;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence that is at least 95% identical to SEQ ID NO: 14 and a second sequence that is at least 95% identical to SEQ ID NO: 16;
the second domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 24;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 23:
the soluble tissue factor domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 5;
the first domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 26;
the second target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 22; and
the second domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 24.
35. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 20:
the soluble tissue factor domain of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 5;
the first domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 26;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence of SEQ ID NO: 14 and a second sequence of SEQ ID NO: 16;
the second domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 24;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 23:
the soluble tissue factor domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 5;
the first domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 26;
the second target-binding domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 22; and
the second domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 24.
36. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 20;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence that is at least 90% identical to SEQ ID NO: 14 and a second sequence that is at least 90% identical to SEQ ID NO: 16;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 22; and
the second target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 90% identical to SEQ ID NO: 23.
37. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 20:
the soluble tissue factor domain of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 5;
the first domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 26;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence that is at least 95% identical to SEQ ID NO: 14 and a second sequence that is at least 95% identical to SEQ ID NO: 16;
the second domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 24;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 22:
the soluble tissue factor domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 5;
the first domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 26;
the second target-binding domain of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 23; and
the second domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises a sequence that is at least 95% identical to SEQ ID NO: 24.
38. The method of claim 1 , wherein:
the first target-binding domain of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 20:
the soluble tissue factor domain of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 5;
the first domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 26;
the second target-binding domain of the first multi-chain chimeric polypeptide comprises a first sequence of SEQ ID NO: 14 and a second sequence of SEQ ID NO: 16;
the second domain of the pair of affinity domains of the first multi-chain chimeric polypeptide comprises SEQ ID NO: 24;
the first target-binding domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 22:
the soluble tissue factor domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 5;
the first domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 26;
the second target-binding domain of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 23; and
the second domain of the pair of affinity domains of the second multi-chain chimeric polypeptide comprises SEQ ID NO: 24.