Synthetic pathway activators
Provided herein are novel synthetic pathway activators comprising multimerization regions, transmembrane domains, and intracellular signaling domains.
1 . A nucleic acid encoding a synthetic pathway activator (SPA) peptide comprising a chimeric polypeptide comprising:
a. an extracellular domain comprising a CD34 epitope or a CD34 ectodomain;
b. a lipid anchor or a transmembrane domain;
c. an intracellular signaling domain, wherein the intracellular signaling domain comprises a type I cytokine receptor superfamily box2 peptide motif comprising VSVVEIEANDKKP (SEQ ID NO: 107), and a tyrosine phosphorylation motif comprising YXPQ; and
d. a multimerization region.
2 . The nucleic acid of claim 1 , wherein the intracellular signaling domain comprises a sequence having at least 80%, 85%, 90%, 95%, or 99% identity to the sequence as set forth in SEQ ID NO: 60, or comprises the sequence as set forth in SEQ ID NO: 60.
3 . The nucleic acid of claim 1 , wherein the extracellular domain comprises a CD34 epitope, the multimerization region comprises a multimerization region comprising an unpaired cysteine residue, the lipid anchor or transmembrane domain comprises a gp130 transmembrane domain, and the intracellular signaling domain comprises a gp130 intracellular signaling domain.
4 . The nucleic acid of claim 1 , wherein the SPA peptide comprises a sequence with at least 90% or 95% sequence identity to a sequence as set forth in SEQ ID NO: 81.
5 . The nucleic acid of claim 1 , wherein the SPA peptide comprises a sequence as set forth in SEQ ID NO: 81.
6 . A synthetic pathway activator (SPA) peptide comprising a chimeric polypeptide comprising:
a. an extracellular domain comprising a CD34 epitope or a CD34 ectodomain;
b. a lipid anchor or a transmembrane domain;
c. an intracellular signaling domain, wherein the intracellular signaling domain comprises a type I cytokine receptor superfamily box2 peptide motif comprising VSVVEIEANDKKP (SEQ ID NO: 107), and a tyrosine phosphorylation motif comprising YXPQ; and
d. a multimerization region.
7 . The SPA peptide of claim 6 , wherein the extracellular domain comprises a CD34 epitope, the multimerization region comprises a multimerization region comprising an unpaired cysteine residue, the lipid anchor or transmembrane domain comprises a gp130 transmembrane domain, and the intracellular signaling domain comprises a gp130 intracellular signaling domain.
8 . The SPA peptide of claim 6 , comprising a sequence with at least 90% or 95% sequence identity to a sequence as set forth in SEQ ID NO: 81.
9 . The SPA peptide of claim 6 , comprising a sequence as set forth in SEQ ID NO: 81.
10 . A multimer comprising the SPA peptide of claim 6 .
11 . A multimer comprising the SPA peptide of claim 8 .
12 . A multimer comprising the SPA peptide of claim 9 .
13 . A vector comprising the nucleic acid of claim 1 .
14 . A vector comprising the nucleic acid of claim 4 .
15 . A vector comprising the nucleic acid of claim 5 .
16 . A system comprising:
a. optionally, a nucleic acid encoding a first chimeric polypeptide comprising a priming receptor;
b. a nucleic acid encoding a second chimeric polypeptide, optionally comprising a chimeric antigen receptor (CAR); and
c. the nucleic acid of claim 1 .
17 . A cell or population of cells comprising the nucleic acid of claim 1 .
18 . A cell or population of cells comprising the nucleic acid of claim 4 .
19 . A cell or population of cells comprising the nucleic acid of claim 5 .
20 . A pharmaceutical composition comprising the cell or population of cells of 17 , and a pharmaceutically acceptable excipient.
21 . A pharmaceutical composition comprising the cell or population of cells of 18 , and a pharmaceutically acceptable excipient.
22 . A pharmaceutical composition comprising the cell or population of cells of 19 , and a pharmaceutically acceptable excipient.
23 . The nucleic acid of claim 1 , wherein the CD34 epitope comprises a sequence as set forth in SEQ ID NO: 238.
24 . The nucleic acid of claim 1 , further comprising an inducible promoter or a constitutive promoter operably linked to the nucleic acid encoding the SPA, optionally wherein the inducible promoter is an HNF1α promoter and/or wherein the constitutive promoter is an EF1α promoter.
25 . The SPA peptide of claim 6 , wherein the intracellular signaling domain comprises a sequence having at least 80%, 85%, 90%, 95%, or 99% identity to a sequence as set forth in SEQ ID NO: 60, or comprises the sequence as set forth in SEQ ID NO: 60.
26 . The SPA peptide of claim 6 , wherein
a. the CD34 epitope comprises a sequence as set forth in SEQ ID NO: 238; and/or
b. the multimerization region comprises
i. at least an unpaired cysteine residue, and/or
ii. the multimerization region results in constitutive activity of the intracellular signaling domain.
27 . The cell or population of cells of claim 17 , wherein the cell is an immune cell, a human immune cell, a primary human immune cell, a natural killer (NK) cell, a T cell, a CD8+ T cell, a CD4+ T cell, a primary T cell, a primary human T cell or a T cell progenitor cell.
28 . The nucleic acid of claim 1 , wherein
a. the multimerization region comprises at least an unpaired cysteine residue; and/or
b. the multimerization region results in constitutive activity of the intracellular signaling domain.
29 . The nucleic acid of claim 1 , wherein the transmembrane domain comprises a sequence at least 90% or 95% identical to the sequence as set forth in SEQ ID NO: 61 or wherein the transmembrane domain comprises the sequence as set forth in SEQ ID NO: 61.
30 . The SPA peptide of claim 6 , wherein the transmembrane domain comprises a sequence at least 90% or 95% identical to the sequence as set forth in SEQ ID NO: 61 or wherein the transmembrane domain comprises the sequence as set forth in SEQ ID NO: 61.