Methods and compositions for stem cell differentiation

Abstract

The present disclosure provides methods and compositions for stem cell differentiation. In some examples, the stem cells are pluripotent stem cells (PSCs). The PSC may comprise a nucleic acid comprising a readable frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding a readable frame of one or more transcription factors.

Description

[0001] Cross-references

[0002] This application claims the benefit of UK patent application number 2209676.2 filed on June 30, 2022, the entirety of which is hereby incorporated by reference into this document.

[0003] Sequence Listing

[0004] This application is filed with a sequence listing in electronic format. The sequence listing is provided as a file named 57963-707_601_SL.XML, which was created on June 30, 2023 and is 2,513 bytes in size. The information in the electronic format of the sequence listing is incorporated by reference in its entirety. Background Art

[0005] Induced pluripotent stem cells (iPSCs) can be programmed to various phenotypes by introducing one or more transcription factors or one or more molecules that regulate transcription or transcription factors. For example, iPSCs can be programmed to be immune cells, which can be used, for example, in immunotherapeutic applications. Summary of the invention

[0006] On the one hand, the present disclosure provides a pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, immune cells express FOXP3. In some embodiments, immune cells express CD34. In some embodiments, immune cells express CD45. In some embodiments, immune cells express CD4. In some embodiments, immune cells are regulatory T cells. In some embodiments, immune cells are hematopoietic progenitor cells. In some embodiments, immune cells are leukocytes. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, forkhead box, protein containing GATA zinc finger domain, nuclear factor of activated T cells, protein containing IPT domain, microRNA protein encoding host gene, Notch receptor, RAR related lonely receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and protein containing BTB domain. In some embodiments, one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, PSC is provided in culture medium. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, the culture medium is not changed during the differentiation of PSC into immune cells.

[0007] On the other hand, the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0008] On the other hand, the present disclosure provides a pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces PSC to differentiate into immune cells, wherein PSC is provided in a culture medium, and the culture medium is not changed during the differentiation of PSC into immune cells. In some embodiments, the immune cells express FOXP3. In some embodiments, the immune cells express CD34. In some embodiments, the immune cells express CD45. In some embodiments, the immune cells express CD4. In some embodiments, the immune cells are regulatory T cells. In some embodiments, the immune cells are hematopoietic progenitor cells. In some embodiments, the immune cells are leukocytes. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, RAR-related orphan receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSC differentiation into an immune cell. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of PSCs into immune cells in 11 days or less, 5 days or less, 4 days or less, or 1 day or less.

[0009] On the other hand, the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0010] On the other hand, the present disclosure provides a method for generating an immune cell colony, the method comprising: providing one or more pluripotent stem cells (PSC); expressing a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors in one or more PSCs; and generating an immune cell colony from one or more PSCs. In some embodiments, at least one of the immune cells expresses FOXP3. In some embodiments, at least one of the immune cells expresses CD34. In some embodiments, at least one of the immune cells expresses CD45. In some embodiments, at least one of the immune cells expresses CD4. In some embodiments, at least one of the immune cells is a regulatory T cell. In some embodiments, at least one of the immune cells is a hematopoietic progenitor cell. In some embodiments, at least one of the immune cells is a leukocyte. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, protein containing GATA zinc finger domain, nuclear factor of activated T cells, protein containing IPT domain, microRNA protein encoding host gene, Notch receptor, RAR related lonely receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and protein containing BTB domain. In some embodiments, one or more PSCs are provided in culture medium. In some embodiments, the culture medium is not changed during the generation of immune cell colonies from one or more PSCs. In some embodiments, one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces one or more PSCs to differentiate into immune cell colonies. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of one or more PSCs into an immune cell population in 11 days or less, 5 days or less, 4 days or less, or 1 day or less.

[0011] Another aspect of the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0012] On the other hand, the present disclosure provides a method for generating an immune cell colony, comprising: providing one or more pluripotent stem cells (PSCs); inducing the expression of a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame of one or more transcription factors in one or more PSCs, wherein the nucleic acid, wherein the induction occurs in 28 days or less; and generating an immune cell colony from one or more PSCs. In some embodiments, at least one of the immune cells expresses FOXP3. In some embodiments, at least one of the immune cells expresses CD34. In some embodiments, at least one of the immune cells expresses CD45. In some embodiments, at least one of the immune cells expresses CD4. In some embodiments, at least one of the immune cells is a regulatory T cell. In some embodiments, at least one of the immune cells is a hematopoietic progenitor cell. In some embodiments, at least one of the immune cells is a leukocyte. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, protein containing GATA zinc finger domain, nuclear factor of activated T cells, protein containing IPT domain, microRNA protein encoding host gene, Notch receptor, RAR related lonely receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and protein containing BTB domain. In some embodiments, one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, one or more PSCs are provided in a culture medium. In some embodiments, the culture medium is not changed during the generation of an immune cell population from one or more PSCs. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of one or more PSCs into an immune cell population in 11 days or less, 5 days or less, 4 days or less, or 1 day or less.

[0013] Another aspect of the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0014] On the other hand, the present disclosure provides an immune cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD4. In some embodiments, the immune cell is a regulatory T cell. In some embodiments, the immune cell is a hematopoietic progenitor cell. In some embodiments, the immune cell is a leukocyte. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, protein containing GATA zinc finger domain, nuclear factor of activated T cells, protein containing IPT domain, microRNA protein encoding host gene, Notch receptor, RAR related lonely receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and protein containing BTB domain. In some embodiments, one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, PSC is provided in culture medium. In some embodiments, the culture medium is not changed during the differentiation of PSC into immune cells. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid encoding an open reading frame of one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells.

[0015] Another aspect of the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0016] On the other hand, the present disclosure provides an immune cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid, wherein the immune cell is provided in a culture medium, and the culture medium is not changed during the differentiation of PSC into immune cells. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD4. In some embodiments, the immune cell is a regulatory T cell. In some embodiments, the immune cell is a hematopoietic progenitor cell. In some embodiments, the immune cell is a leukocyte. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, RAR-related orphan receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSC differentiation into an immune cell. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of PSCs into immune cells in 11 days or less, 5 days or less, 4 days or less, or 1 day or less.

[0017] Another aspect of the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0018] Another aspect of the present disclosure provides a cell expressing FOXP3, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of the cell expressing FOXP3 from PSC. In some embodiments, the cell expressing FOXP3 further expresses CD34. In some embodiments, the cell expressing FOXP3 further expresses CD45. In some embodiments, the cell expressing FOXP3 further expresses CD4. In some embodiments, the cell expressing FOXP3 is a regulatory T cell. In some embodiments, the cell expressing FOXP3 is a hematopoietic progenitor cell. In some embodiments, the cell expressing FOXP3 is a leukocyte. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSCs to differentiate into cells expressing FOXP3 in 28 days or less. In some embodiments, the PSCs are provided in a culture medium. In some embodiments, the culture medium is not changed during differentiation of the PSCs into cells expressing FOXP3.

[0019] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing FOXP3 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, nutrients, growth factors, or microenvironment / matrix optimization are not performed.

[0020] Another aspect of the present disclosure provides a cell expressing FOXP3, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces the cell expressing FOXP3 to differentiate from a pluripotent stem cell (PSC), and the PSC is provided in a culture medium, and the culture medium is not changed during the differentiation of the PSC into a cell expressing FOXP3. In some embodiments, the cell expressing FOXP3 further expresses CD34. In some embodiments, the cell expressing FOXP3 further expresses CD45. In some embodiments, the cell expressing FOXP3 further expresses CD4. In some embodiments, the cell expressing FOXP3 is a regulatory T cell. In some embodiments, the cell expressing FOXP3 is a hematopoietic progenitor cell. In some embodiments, the cell expressing FOXP3 is a leukocyte. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing FOXP3 from PSC. In some embodiments, PSC is provided in a culture medium. In some embodiments, the culture medium is not changed during differentiation of PSC into cells expressing FOXP3. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3.In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0021] Another aspect of the present disclosure provides a cell expressing CD45, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD45 from PSCs. In some embodiments, cells expressing CD45 further express CD34. In some embodiments, cells expressing CD45 further express CD45. In some embodiments, cells expressing CD45 further express CD4. In some embodiments, cells expressing CD45 are regulatory T cells. In some embodiments, cells expressing CD45 are hematopoietic progenitor cells. In some embodiments, cells expressing CD45 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSC differentiation into cells expressing CD45 in 11 days or less, 5 days or less, 4 days or less, 1 day or less. In some embodiments, PSC is provided in a culture medium. In some embodiments, the culture medium is not changed during differentiation of PSC into cells expressing FOXP3.

[0022] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing CD45 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0023] Another aspect of the present disclosure provides a cell expressing CD45, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD45 from pluripotent stem cells (PSCs), and the PSC is provided in a culture medium, and the culture medium is not changed during the differentiation of the PSC into cells expressing CD45. In some embodiments, cells expressing CD45 further express CD34. In some embodiments, cells expressing CD45 further express CD45. In some embodiments, cells expressing CD45 further express CD4. In some embodiments, cells expressing CD45 are regulatory T cells. In some embodiments, cells expressing CD45 are NK cells. In some embodiments, cells expressing CD45 are hematopoietic progenitor cells. In some embodiments, cells expressing CD45 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of PSCs into cells expressing CD45 in 28 days or less. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of cells expressing CD45 from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0024] In another aspect, the present disclosure provides a cell population comprising two or more cells expressing CD45 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0025] Another aspect of the present disclosure provides a cell expressing CD34, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD34 from PSCs. In some embodiments, cells expressing CD34 further express FOXP3. In some embodiments, cells expressing CD34 further express CD45. In some embodiments, cells expressing CD34 further express CD4. In some embodiments, cells expressing CD34 are regulatory T cells. In some embodiments, cells expressing CD34 are hematopoietic progenitor cells. In some embodiments, cells expressing CD34 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSCs to differentiate into cells expressing CD34 in 28 days or less. In some embodiments, PSCs are provided in a culture medium. In some embodiments, the culture medium is not changed during differentiation of PSCs into cells expressing FOXP3.

[0026] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing CD34 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0027] Another aspect of the present disclosure provides a cell expressing CD34, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces differentiation of cells expressing CD34 from pluripotent stem cells (PSC), and the PSC is provided in a culture medium, and the culture medium is not changed during the differentiation of the PSC into cells expressing CD34. In some embodiments, cells expressing CD34 further express FOXP3. In some embodiments, cells expressing CD34 further express CD45. In some embodiments, cells expressing CD34 further express CD4. In some embodiments, cells expressing CD34 are regulatory T cells. In some embodiments, cells expressing CD34 are hematopoietic progenitor cells. In some embodiments, cells expressing CD34 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of PSCs into cells expressing CD34. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of cells expressing CD34 from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of cells expressing CD34 from PSCs in 4 days or less.

[0028] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing CD34 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0029] Another aspect of the present disclosure provides a cell expressing CD4, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD4 from PSC. In some embodiments, cells expressing CD4 further express FOXP3. In some embodiments, cells expressing CD4 further express CD45. In some embodiments, cells expressing CD4 are regulatory T cells. In some embodiments, cells expressing CD4 are hematopoietic progenitor cells. In some embodiments, cells expressing CD4 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces PSCs to differentiate into cells expressing CD4 in 28 days or less. In some embodiments, PSCs are provided in a culture medium. In some embodiments, the culture medium is not changed during differentiation of PSCs into cells expressing FOXP3.

[0030] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing CD4 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrients, growth factors or microenvironment / matrix optimization is performed.

[0031] Another aspect of the present disclosure provides a cell expressing CD4, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD4 from pluripotent stem cells (PSC), and the PSC is provided in a culture medium, and the culture medium is not changed during the differentiation of the PSC into cells expressing CD4. In some embodiments, cells expressing CD4 further express FOXP3. In some embodiments, cells expressing CD4 further express CD45. In some embodiments, cells expressing CD4 further express CD4. In some embodiments, cells expressing CD4 are regulatory T cells. In some embodiments, cells expressing CD4 are hematopoietic progenitor cells. In some embodiments, cells expressing CD4 are leukocytes. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of PSC into cells expressing CD4. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD4 from PSC. In some embodiments, in 4 days or less or 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD4 from PSC.

[0032] Another aspect of the present disclosure provides a cell population comprising two or more cells expressing CD4 disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrients, growth factors or microenvironment / matrix optimization is performed.

[0033] Another aspect of the present disclosure provides a pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors or a transcriptional activator encoding an open reading frame of one or more transcription factors, wherein the one or more transcription factors are selected from a transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into an immune cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD4. In some embodiments, the immune cell is a regulatory T cell. In some embodiments, the immune cell is a hematopoietic progenitor cell. In some embodiments, the immune cell is a leukocyte. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, the nucleic acid comprises three or more open reading frames encoding one or more transcription factors. In some embodiments, the nucleic acid comprises four or more open reading frames encoding one or more transcription factors. In some embodiments, the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the four or more transcription factors are selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factors, proteins containing BTB domains, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, RING finger proteins, forkhead boxes, Notch receptors, microRNA protein encoding host genes, proteins containing GATA zinc finger domains, nuclear factor of activated T cells, proteins containing IPT domains, TCF / LEF transcription factor family, Wnt enhancer complex, NF-κB complex subunits, proteins containing IPT domains, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, nuclear factor of activated T cells, IPT domain-containing protein, phosphatase 1 regulatory subunit, TCF / LEF transcription factor family, Wnt enhancer complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, phosphatase 1 regulatory subunit, interferon regulatory factor, Kruppel-like factor, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, IPT domain-containing protein, basic helix-loop-helix protein, and BTB domain-containing protein. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Kruppel-like factor, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, NF-κB complex subunit, IPT domain-containing protein, CUT-like homeobox and pseudogene, T-box transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, and Wnt enhanceosome complex. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2 type, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, Notch receptor, microRNA protein encoding host gene, RAR-related orphan receptor, NF-κB complex subunit, IPT domain-containing protein, Runt-related transcription factor, basic helix-loop-helix protein. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, Notch receptor, microRNA protein-encoding host gene, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: ring finger proteins, forkhead boxes, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, interferon regulatory factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, microRNA protein encoding host genes, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins. In some embodiments, one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, GATA zinc finger domain containing proteins, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Notch receptors, RAR-related orphan receptors, NF-κB complex subunits, proteins containing IPT domains, T box transcription factors, TCF / LEF transcription factor family, Wnt enhancer complex, proteins containing BTB domains, zinc finger C2H2 type. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, interferon regulatory factors, zinc finger C2H2 type, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, CUT-like homeoboxes and pseudogenes, basic helix-loop-helix proteins, TCF / LEF transcription factor family, Wnt enhanceosome complex, BTB domain-containing proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead box, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, Notch receptors, microRNA protein-encoding host genes, and T-box transcription factors. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of forkhead boxes, GATA zinc finger domain-containing proteins, zinc finger C2H2 type, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, RAR-related orphan receptors, and basic helix-loop-helix proteins.In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2 type, phosphatase 1 regulatory subunits, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors, basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: zinc finger C2H2 type, BAF complex, forkhead boxes, proteins containing GATA zinc finger domains, phosphatase 1 regulatory subunits, interferon regulatory factors, Kruppel-like factors, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, NF-κB complex subunits, Runt-related transcription factors, CUT-like homeoboxes and pseudogenes, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: RING finger proteins, forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, proteins containing IPT domains, microRNA protein encoding host genes, RAR-related orphan receptors, Runt-related transcription factors, T-box transcription factors, basic helix-loop-helix proteins, and proteins containing BTB domains. In some embodiments, the one or more transcription factors are selected from the family of transcription factors consisting of: forkhead boxes, proteins containing GATA zinc finger domains, zinc finger C2H2-type proteins, phosphatase 1 regulatory subunits, Kruppel-like factors, Notch receptors, microRNA protein encoding host genes, NF-κB complex subunits, proteins containing IPT domains, and basic helix-loop-helix proteins. In some embodiments, the one or more transcription factors are selected from the transcription factor family consisting of: forkhead box, GATA zinc finger domain-containing protein, zinc finger C2H2-type protein, phosphatase 1 regulatory subunit, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, Runt-related transcription factor, basic helix-loop-helix protein, TCF / LEF transcription factor family, Wnt enhancer complex, and BTB domain-containing protein.In some embodiments, one or more transcription factors are selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, forkhead box, protein containing GATA zinc finger domain, nuclear factor of activated T cells, protein containing IPT domain, microRNA protein encoding host gene, Notch receptor, RAR related lonely receptor, NF-κB complex subunit, T box transcription factor, TCF / LEF transcription factor family, Wnt enhancer complex, and protein containing BTB domain. In some embodiments, one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, PSC is provided in culture medium. In some embodiments, the culture medium is not changed during the differentiation of PSC into immune cells. In some embodiments, in 28 days or less, a nucleic acid encoding an open reading frame of one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells.

[0034] Another aspect of the present disclosure provides a cell population comprising two or more immune cells disclosed herein. In some embodiments, the cells are adherent cells. In some embodiments, the cells are suspension cells. In some embodiments, at least 5% of the cells express FOXP3. In some embodiments, at least 10% of the cells express FOXP3. In some embodiments, at least 20% of the cells express FOXP3. In some embodiments, at least 30% of the cells express FOXP3. In some embodiments, at least 40% of the cells express FOXP3. In some embodiments, at least 1% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 3% of the cells express CD45. In some embodiments, at least 4% of the cells express CD45. In some embodiments, at least 5% of the cells express CD45. In some embodiments, at least 6% of the cells express CD45. In some embodiments, at least 1% of the cells express CD34. In some embodiments, at least 2% of the cells express CD34. In some embodiments, at least 2.5% of the cells express CD34. In some embodiments, at least 2% of the cells express CD4. In some embodiments, at least 3% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, at least 4% of the cells express CD4. In some embodiments, no nutrient, growth factor, or microenvironment / matrix optimization is performed.

[0035] One aspect of the present disclosure is a pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors comprises NOTCH1 or a functional derivative thereof, and in 28 days or less, the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into an immune cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD4. In some embodiments, the immune cell is a regulatory T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a hematopoietic progenitor cell. In some embodiments, the immune cell is a leukocyte. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, in 5 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, in 4 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, in 3 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, in 2 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces PSC to differentiate into immune cells. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding one or more open reading frames encoding one or more transcription factors induces differentiation of a PSC into an immune cell in 1 day or less.In some embodiments, the one or more transcription factors further include a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the one or more transcription factors further include a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the one or more transcription factors further include a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the one or more transcription factors further include a transcription factor selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors further include transcription factors selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, Runt-related transcription factor, protein containing BTB domain, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, NOTCH1 or its functional derivatives comprise the intracellular domain ("ICD") of NOTCH1. In some embodiments, NOTCH1 or its functional derivatives consist of the intracellular domain ("ICD") of NOTCH1. The cell population comprises two or more of the PSCs of any embodiment of the present disclosure. In some embodiments, the cell population is an adherent cell. In some embodiments, the cell population is a suspension cell. In some embodiments, at least 5% of the cell population expresses FOXP3. In some embodiments, at least 10% of the cell population expresses FOXP3. In some embodiments, at least 20% of the cell population expresses FOXP3. In some embodiments, at least 30% of the cell population express FOXP3. In some embodiments, at least 40% of the cell population express FOXP3. In some embodiments, at least 1% of the cell population expresses CD45. In some embodiments, at least 2% of the cell population expresses CD45. In some embodiments, at least 3% of the cell population expresses CD45. In some embodiments, at least 4% of the cell population expresses CD45. In some embodiments, at least 5% of the cell population expresses CD45. In some embodiments, at least 6% of the cell population expresses CD45.In some embodiments, at least 10% of the cell population express CD45. In some embodiments, at least 15% of the cell population express CD45. In some embodiments, at least 20% of the cell population express CD45. In some embodiments, at least 30% of the cell population express CD45. In some embodiments, at least 40% of the cell population express CD45. In some embodiments, at least 50% of the cell population express CD45. In some embodiments, at least 1% of the cell population express CD8. In some embodiments, at least 2% of the cell population express CD8. In some embodiments, at least 2.5% of the cell population express CD8. In some embodiments, at least 3% of the cell population express CD8. In some embodiments, at least 1% of the cell population express CD34. In some embodiments, at least 2% of the cell population express CD34. In some embodiments, at least 2.5% of the cell population express CD34. In some embodiments, at least 2% of the cell population express CD4. In some embodiments, at least 3% of the cell population express CD4. In some embodiments, at least 4% of the cell population express CD4. In some embodiments, at least 5% of the cell population express CD4. In some embodiments, at least 10% of the cell population express CD4. In some embodiments, at least 15% of the cell population express CD4. In some embodiments, at least 1% of the cell population express CD45 and CD34. In some embodiments, at least 2% of the cell population express CD45 and CD34. In some embodiments, at least 2.5% of the cell population express CD45 and CD34. In some embodiments, at least 3% of the cell population express CD45 and CD34. In some embodiments, at least 5% of the cell population express CD45 and CD56. In some embodiments, at least 6% of the cell population express CD45 and CD56. In some embodiments, at least 10% of the cell population express CD45 and CD56. In some embodiments, at least 15% of the cell population express CD45 and CD56. In some embodiments, at least 20% of the cell population express CD45 and CD56. In some embodiments, at least 30% of the cell population express CD45 and CD56. In some embodiments, at least 40% of the cell population express CD45 and CD56. In some embodiments, at least 50% of the cell population express CD45 and CD56. In some embodiments, at least 60% of the cell population express CD45 and CD56.In some embodiments, at least 70% of the cell population express CD45 and CD56. In some embodiments, at least 80% of the cell population express CD45 and CD56. In some embodiments, at least 5% of the cell population express CD45 and CD34. In some embodiments, at least 6% of the cell population express CD45 and CD34. In some embodiments, at least 10% of the cell population express CD45 and CD34. In some embodiments, at least 15% of the cell population express CD45 and CD34. In some embodiments, at least 1% of the cell population express CD14 and CD11b. In some embodiments, at least 2% of the cell population express CD14 and CD11b. In some embodiments, at least 2.5% of the cell population express CD14 and CD11b. In some embodiments, at least 3% of the cell population express CD14 and CD11b.

[0036] Another aspect of the present disclosure is a method for generating an immune cell population, the method comprising: providing one or more pluripotent stem cells (PSCs); expressing a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors in one or more PSCs, wherein the one or more transcription factors comprise NOTCH1; and generating an immune cell population from one or more PSCs. In some embodiments, at least one of the immune cells expresses FOXP3. In some embodiments, at least one of the immune cells expresses CD34. In some embodiments, at least one of the immune cells expresses CD45. In some embodiments, at least one of the immune cells expresses CD4. In some embodiments, at least one of the immune cells is a regulatory T cell. In some embodiments, at least one of the immune cells is a hematopoietic stem / progenitor cell. In some embodiments, at least one of the immune cells is a hematopoietic progenitor cell. In some embodiments, at least one of the immune cells is a leukocyte. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors induces one or more PSCs to differentiate into an immune cell population. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces one or more PSCs to differentiate into immune cell colonies. In some embodiments, in 5 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces one or more PSCs to differentiate into immune cell colonies. In some embodiments, in 4 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces one or more PSCs to differentiate into immune cell colonies. In some embodiments, in 3 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces one or more PSCs to differentiate into immune cell colonies. In some embodiments, the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding one or more open reading frames encoding one or more transcription factors induces differentiation of one or more PSCs into an immune cell population in 2 days or less.In some embodiments, in 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces one or more PSCs to differentiate into an immune cell population. In some embodiments, the one or more transcription factors further include a transcription factor selected from a transcription factor family consisting of zinc finger C2H2 type, BAF complex, ring finger protein, forkhead box, protein containing a GATA zinc finger domain, Runt-related transcription factor, protein containing a BTB domain, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, protein containing an IPT domain, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors further include a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor. In some embodiments, the one or more transcription factors further include a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.In some embodiments, the one or more transcription factors further include a transcription factor selected from the transcription factor family consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhancer complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein encoding host gene, Notch receptor, microRNA protein encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T box transcription factor. In some embodiments, the one or more transcription factors include any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, the nucleic acid comprises two or more open reading frames encoding one or more transcription factors. In some embodiments, one or more transcription factors further include transcription factors selected from the transcription factor family consisting of: zinc finger C2H2 type, BAF complex, Runt-related transcription factor, protein containing BTB domain, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix. In some embodiments, NOTCH1 or its functional derivatives include the intracellular domain ("ICD") of NOTCH1. In some embodiments, NOTCH1 or its functional derivatives consist of the intracellular domain ("ICD") of NOTCH1. In some embodiments, the immune cell population is an adherent cell. In some embodiments, the immune cell population is a suspension cell. In some embodiments, at least 5% of the immune cell population expresses FOXP3. In some embodiments, at least 10% of the immune cell population expresses FOXP3. In some embodiments, at least 20% of the immune cell population expresses FOXP3. In some embodiments, at least 30% of the immune cell population express FOXP3. In some embodiments, at least 40% of the immune cell population express FOXP3. In some embodiments, at least 1% of the immune cell population expresses CD45. In some embodiments, at least 2% of the immune cell population expresses CD45. In some embodiments, at least 3% of the immune cell population expresses CD45. In some embodiments, at least 4% of the immune cell population expresses CD45. In some embodiments, at least 5% of the immune cell population expresses CD45. In some embodiments, at least 6% of the immune cell population expresses CD45.In some embodiments, at least 10% of the immune cell population express CD45. In some embodiments, at least 15% of the immune cell population express CD45. In some embodiments, at least 20% of the immune cell population express CD45. In some embodiments, at least 30% of the immune cell population express CD45. In some embodiments, at least 40% of the immune cell population express CD45. In some embodiments, at least 50% of the immune cell population express CD45. In some embodiments, at least 1% of the immune cell population express CD8. In some embodiments, at least 2% of the immune cell population express CD8. In some embodiments, at least 2.5% of the immune cell population express CD8. In some embodiments, at least 3% of the immune cell population express CD8. In some embodiments, at least 1% of the immune cell population express CD34. In some embodiments, at least 2% of the immune cell population express CD34. In some embodiments, at least 2.5% of the immune cell population express CD34. In some embodiments, at least 2% of the immune cell population express CD4. In some embodiments, at least 3% of the immune cell population expresses CD4. In some embodiments, at least 4% of the immune cell population expresses CD4. In some embodiments, at least 5% of the immune cell population expresses CD4. In some embodiments, at least 10% of the immune cell population expresses CD4. In some embodiments, at least 15% of the immune cell population expresses CD4. In some embodiments, at least 1% of the immune cell population expresses CD45 and CD34. In some embodiments, at least 2% of the immune cell population expresses CD45 and CD34. In some embodiments, at least 2.5% of the immune cell population expresses CD45 and CD34. In some embodiments, at least 3% of the immune cell population expresses CD45 and CD34. In some embodiments, at least 5% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 6% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 10% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 15% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 20% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 30% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 40% of the immune cell population expresses CD45 and CD56. In some embodiments, at least 50% of the immune cell population expresses CD45 and CD56.In some embodiments, at least 60% of the immune cell population express CD45 and CD56. In some embodiments, at least 70% of the immune cell population express CD45 and CD56. In some embodiments, at least 80% of the immune cell population express CD45 and CD56. In some embodiments, at least 5% of the immune cell population express CD45 and CD34. In some embodiments, at least 6% of the immune cell population express CD45 and CD34. In some embodiments, at least 10% of the immune cell population express CD45 and CD34. In some embodiments, at least 15% of the immune cell population express CD45 and CD34. In some embodiments, at least 1% of the cell population express CD14 and CD11b. In some embodiments, at least 2% of the cell population express CD14 and CD11b. In some embodiments, at least 2.5% of the cell population express CD14 and CD11b. In some embodiments, at least 3% of the cell population express CD14 and CD11b.

[0037] Another aspect of the present disclosure is an isolated immune cell population, wherein at least 5%, 6%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70%, or 80% of the immune cell population is engineered to express CD45 and CD56. In some embodiments, at least 1%, 2%, 2.5%, or 3% of the immune cell population is engineered to express CD14 and CD11b. In some embodiments, at least 1%, 2%, 2.5%, or 3% of the immune cell population is engineered to express CD45 and CD34.

[0038] Other aspects and advantages of the present disclosure will become readily apparent to those skilled in the art from the following detailed description, wherein only illustrative embodiments of the present disclosure are shown and described. As will be appreciated, the present disclosure is capable of other and different embodiments, and its several details are capable of modification in various obvious respects, all without departing from the present disclosure. Accordingly, the drawings and description should be regarded as illustrative rather than restrictive.

[0039] Incorporation by reference

[0040] All publications, patents and patent applications mentioned in this specification are incorporated herein by reference to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference. If the publications and patents or patent applications incorporated by reference conflict with the disclosure contained in this specification, this specification is intended to supersede and / or take precedence over any such conflicting material. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] The novel features of the present invention are particularly set forth in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description and accompanying drawings (also referred to herein as "Figures" and "FIG.") which set forth illustrative embodiments utilizing the principles of the present invention, in which:

[0042] Figure 1 Depicted is the expression of FOXP3 induction in 5C3 stem cells treated in ImmunoCult medium and mTeSR1 stem cell medium. Adherent cells and suspension cells are depicted.

[0043] Figure 2 Exemplary markers of immune cell development are depicted.

[0044] Figure 3 CD45 expression by cells to which various transcription factor formulations were delivered is depicted.

[0045] Figure 4 Depicted are CD45 expression of cells with and without transcription factor (TF) induction.

[0046] Figure 5 CD4 expression by cells to which various transcription factor formulations were delivered is depicted.

[0047] Figure 6 Depicted are CD34 expression of cells with and without transcription factor (TF) induction.

[0048] Figures 7A-7D CD45 expression in cells with or without transcription factor (TF) induction is demonstrated. Figures 7A-7C Representative flow cytometry and corresponding gating for TRA-1-60 (iPSC marker) and CD45 (hematopoietic cell lineage marker) are shown. Fig.7D Shown is the quantification of the percentage of cells with CD45 expression in uninduced cells (D2-uninduced) or induced cells transfected with unique combinations of TFs.

[0049] Figures 8A-8D CD4 expression in cells with or without transcription factor (TF) induction is illustrated. Figures 8A-8C Representative flow cytometry and corresponding gating for TRA-1-60 (iPSC marker) and CD4 (T helper marker) are shown. Fig.8D Shown is the quantification of the percentage of cells with CD4 expression in uninduced cells (D2-uninduced) or induced cells transfected with unique combinations of TFs.

[0050] Figures 9A-9D CD8 expression in cells with or without transcription factor (TF) induction is illustrated. Figures 9A-9C Representative flow cytometry and corresponding gating for markers TRA-1-60 (iPSC marker) and CD8 (cytotoxic T cell marker) are shown. Fig.9D Shown is the quantification of the percentage of cells with CD8 expression in uninduced cells (D2-uninduced) or induced cells transfected with unique combinations of TFs.

[0051] Figures 10A-10D CD45 expression in cells with or without transcription factor (TF) induction is demonstrated. Figures 10A-10C Representative flow cytometry and corresponding gating for markers TRA-1-60 (iPSC marker) and CD45 (hematopoietic cell lineage marker) are shown. Fig. 10D Shown is the quantification of the percentage of cells with CD45 expression in uninduced cells (D3-uninduced) or induced cells transfected with a combination of TFs.

[0052] Figures 11A-11D CD8 expression in cells with or without transcription factor (TF) induction is illustrated. Figures 11A-11C Representative flow cytometry and corresponding gating for markers TRA-1-60 (iPSC marker) and CD8 (cytotoxic T cell marker) are shown. Fig.11D Shown is the quantification of the percentage of cells with CD8 expression in uninduced cells (D3-uninduced) or induced cells transfected with unique combinations of TFs.

[0053] Figures 12A-12L The expression of a single CD45 marker or the expression of double positive markers CD45+CD56+, CD34+CD45+ and CD14 / CD11b+ in cells with or without transcription factor (TF) induction is illustrated. Figures 12A-12H TF-uninduced cells ( Figures 12A-12D ) and induced cells ( Figures 12E-12H ) and corresponding gates: TRA-1-60 (iPSC marker), CD45 (hematopoietic cell lineage marker), CD34 (hematopoietic stem / progenitor cell marker), CD56 (NK cell marker), CD14 / CD11b (myeloid marker). Figures 12I-12L The cell CD45 ( Fig.12I )、CD34+CD45+ double positive ( Fig.12J )、CD45+CD56+ double positive ( Figure 12K ), and CD14+ / CD11b+ double positive ( Figure 12L) as a percentage.

[0054] Figures 13A-13C CD45 expression in cells with or without transcription factor (TF) induction is demonstrated. Figures 13A-13B Representative flow cytometric plots and gating strategies are shown. Fig. 13C Quantification of CD45 expression in uninduced and induced cells is shown.

[0055] Figures 14A-14L The top performers of TF formula D2-21 are shown to induce expression of single CD45 marker or double positive markers CD45+CD56+, CD34+CD45+ and CD14 / CD11b+ in cells under transcription factor (TF) induction. Bar graph ( Fig.14A , 14D , 14G, 14J) show representative flow cytometry of the percentage of cells expressing the indicated markers. The corresponding gates for the indicated markers are shown ( Figures 14B-14C ; 14E-14F; 14H-14I; 14K-14L). DETAILED DESCRIPTION

[0056] Although various embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Various modifications, changes and substitutions may occur to those skilled in the art without departing from the present invention. It should be understood that various alternatives to the embodiments of the present invention described herein may be employed.

[0057] introduction

[0058] Pluripotent stem cells (PSC) are characterized by their self-renewal ability while maintaining efficacy for therapeutic applications. PSC also has the ability to differentiate into a large number of different phenotypes (e.g., immune cells). Various compositions, preparations, and methods for promoting the effective differentiation of PSC into immune cells are disclosed herein. In some embodiments, the immune cells are T cells. In some embodiments, the immune cells are hematopoietic stem / progenitor cells. In some embodiments, the immune cells are NK cells. In some embodiments, the T cells are regulatory T cells. In some embodiments, the immune cells are early hematopoietic progenitor cells that tend to be T cells. In some embodiments, the immune cells are directed progenitor cells. In some embodiments, the immune cells are mature T cells. In some embodiments, the immune cells express FOXP3. In some embodiments, the immune cells express CD45. In some embodiments, the immune cells express CD34. In some embodiments, the immune cells express CD4. In some embodiments, the immune cells express CD8.

[0059] Immune cells generated by utilizing the compositions, formulations, and methods disclosed herein can be used in therapeutic applications where classical cell lineage immune cells have been routinely used. Because the compositions, formulations, and methods disclosed herein enable efficient and reproducible differentiation of PSCs into potent immune cells, the various compositions, formulations, and methods disclosed herein provide robust improvements to therapeutic agents.

[0060] Pluripotent stem cells

[0061] Pluripotent stem cells (PSC) can be characterized by self-renewal and potent. PSC can divide indefinitely and produce identical daughter cells. When provided with a signal, PSC can differentiate into various phenotypes. In one example, PSC is an embryonic stem cell (ESC). In another example, PSC is an induced PSC (iPSC). PSC (e.g., ESC and iPSC) can express TRA-1-60. TRA-1-60 expression can indicate the differentiation ability of cells.

[0062] Induced pluripotent stem cells (iPSCs) are pluripotent stem cells derived from adult cells that have been genetically reprogrammed to an embryonic stem (ES) cell-like state by expressing genes and factors important for maintaining the defining properties of embryonic stem cells. iPSCs are similar to ES cells in many ways, including the expression of ES cell markers, chromatin methylation patterns, embryoid body formation, teratoma formation, viable chimera formation, pluripotency, and the ability to contribute to many different tissues in vitro. Studies have reported that iPSCs are directed to differentiate into different functional cell types in vitro, and the cell therapy effects of implanted iPSCs have been demonstrated in several animal models of disease. Directed differentiation is a bioengineering method that utilizes the potential of stem cells by limiting the differentiation of stem cells to specific cell types or target tissues in vitro. Directed differentiation may be mainly applied to PSCs of mammalian origin, such as mouse and human cells for biomedical research applications. Cell differentiation may involve a transition from a proliferation mode to a differentiation mode. Directed differentiation can include simulating developmental culture under controlled conditions, involving specific substrates or extracellular matrices that promote cell adhesion and differentiation, and determining the composition of the culture medium. Signal transduction factors, such as growth factors or small molecules, can be applied in different dosages and exposure time sequences or in combination to regulate differentiation. Direct reprogramming, also referred to as transdifferentiation or direct conversion, can include overexpression of one or more factors introduced into the cell. In one example, one or more factors can be transcription factors. By analyzing cell type specific markers, gene expression profiles, and functional assays, the appropriate differentiation of the target cell type can be verified.

[0063] The present disclosure provides pluripotent stem cells (PSC). PSC may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The transcription factors disclosed herein may be exogenous transcription factors. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. PSC may include one or more transcription factors. PSC may include a transcription activator encoding an open reading frame of one or more transcription factors. The activator may be a CRISPR activator of one or more transcription factors. The activator may be a small molecule activator that induces the expression of one or more transcription factors. One or more transcription factors may be exogenous. One or more transcription factors may be endogenous. A nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors may induce PSC to differentiate into immune cells. In some embodiments, the immune cells are T cells. In some embodiments, the immune cells are hematopoietic stem / progenitor cells. In some embodiments, the immune cells are NK cells. In some embodiments, T cells are regulatory T cells. In some embodiments, the immune cells are early hematopoietic progenitor cells that tend to T cells. In some embodiments, the immune cells are directed progenitor cells. In some embodiments, the immune cells are mature T cells. In some embodiments, the immune cells express FOXP3. In some embodiments, the immune cells express CD45. In some embodiments, the immune cells express CD34. In some embodiments, the immune cells express CD4. In some embodiments, the immune cells express CD8. Nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors can induce differentiation of PSCs into immune cells in about 30 days, about 29 days, about 28 days, about 27 days, about 26 days, about 25 days, about 24 days, about 23 days, about 22 days, about 21 days, about 20 days, about 19 days, about 18 days, about 17 days, about 16 days, about 15 days, about 14 days, about 13 days, about 12 days, about 11 days, about 10 days, about 9 days, about 8 days, about 7 days, about 6 days, about 5 days, about 4 days, about 3 days, about 2 days, about 1 day or less. Nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors can induce PSC differentiation into immune cells in 28 days or less. Nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors can induce PSC differentiation into immune cells in 11 days or less.The nucleic acid comprising the open reading frame encoding one or more transcription factors, one or more transcription factors or the transcription activator encoding the open reading frame of one or more transcription factors can induce PSC to differentiate into immune cells in 4 days or less. The nucleic acid comprising the open reading frame encoding one or more transcription factors, one or more transcription factors or the transcription activator encoding the open reading frame of one or more transcription factors can induce PSC to differentiate into immune cells in 3 days or less or 1 day or less. PSC can be provided in a culture medium. The culture medium does not need to be changed during the differentiation of immune cells. A cell colony comprising one or more immune cells can be generated. The cell colony may include adherent cells. The cell colony may include suspension cells. The cell colony may include adherent cells and suspension cells. The cell colony may be provided in a culture medium. The culture medium may not need to be changed during the differentiation of PSC into immune cells. The culture medium may not require any nutrients, growth factors or microenvironment or matrix optimization.

[0064] Immune cells

[0065] The ability of the immune system to control inflammatory responses and prevent reactions to self-antigens or harmless environmental molecules can be obtained during fetal life or early postnatal period. Immune cells such as T cells have various characteristics and functions that assist the development and maintenance of the immune system. T cells develop from hematopoietic stem cells (HSC) in the bone marrow. For T cells, the selection and acquisition of effector and regulatory functions may occur in the thymus. Various markers of T cells can be associated with different stages of T cell development. Two examples of markers of hematopoietic progenitor cells are proteins CD34 and CD45. CD34 is a cell surface marker that can contribute to the attachment of hematopoietic cells in the bone marrow and can identify early hematopoietic cell lineages. CD45 is a protein tyrosine phosphatase that can be expressed in leukocytes and can be a marker of directed progenitor cells. CD45 can be expressed throughout the progenitor cell stage and in mature T cells. CD4 is a co-receptor of the T cell receptor (TCR), which can play a role in antigen presenting cells that recognize and bind to major histocompatibility complex II (MHC class II) protein complexes. The CD8 antigen is a cell surface glycoprotein present on most cytotoxic T lymphocytes, mediating potent cell-cell interactions within the immune system. CD8 acts both as an adhesion molecule for MHC class I molecules and as a co-receptor for TCR for T cell activation.

[0066] The present disclosure provides an immune cell. The immune cell may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. The immune cell may include one or more transcription factors. The immune cell may include a transcription activator of an open reading frame encoding one or more transcription factors, wherein the nucleic acid. The activator may be a CRISPR activator of one or more transcription factors. The activator may be a small molecule activator that induces the expression of one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the immune cell is an early hematopoietic progenitor cell that tends to be a T cell. In some embodiments, the immune cell is a directed progenitor cell. In some embodiments, the immune cell is a mature T cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD4. In some embodiments, the immune cell expresses CD8. In some embodiments, nucleic acid, one or more transcription factors or transcription activators encoding the open reading frames of one or more transcription factors are expressed to induce PSC to differentiate into immune cells. In some embodiments, immune cells are provided in culture medium. Culture medium can be changed during the differentiation of PSC into immune cells. In some embodiments, in 28 days or less, nucleic acid, one or more transcription factors or transcription activators encoding the open reading frames of one or more transcription factors are induced to differentiate immune cells from PSC. In some embodiments, in 11 days or less, nucleic acid, one or more transcription factors or transcription activators encoding the open reading frames of one or more transcription factors are induced to differentiate immune cells from PSC. In some embodiments, in 4 days or less or 1 day or less, nucleic acid, one or more transcription factors or transcription activators encoding the open reading frames of one or more transcription factors are induced to differentiate immune cells from PSC.

[0067] The present disclosure provides a cell expressing FOXP3. Cells expressing FOXP3 may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. Transcription factors disclosed herein may be exogenous transcription factors. Nucleic acids may include two or more open reading frames encoding one or more transcription factors. Cells expressing FOXP3 may include one or more transcription factors. Cells expressing FOXP3 may include a transcription activator encoding an open reading frame of one or more transcription factors. In some embodiments, immune cells are T cells. In some embodiments, cells expressing FOXP3 are regulatory T cells. In some embodiments, nucleic acids comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors are expressed to induce PSC to differentiate into cells expressing FOXP3. In some embodiments, cells expressing FOXP3 are provided in a culture medium. The culture medium does not have to be changed during differentiation of PSC into cells expressing FOXP3. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing FOXP3 from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, in 4 days or less or 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0068] The present disclosure provides a cell expressing CD45. The cell expressing CD45 may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. The cell expressing CD45 may include one or more transcription factors. The cell expressing CD45 may include a transcription activator of an open reading frame encoding one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the cell expressing CD45 is a progenitor T cell. In some embodiments, the cell expressing CD45 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator of an open reading frame encoding one or more transcription factors is expressed to induce PSC to differentiate into a cell expressing CD45. In some embodiments, the cell expressing CD45 is provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into a cell expressing CD45. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD45 from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, in 4 days or less or 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0069] The present disclosure provides a cell expressing CD34. The cell expressing CD34 may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. The cell expressing CD34 may include one or more transcription factors. The cell expressing CD34 may include a transcription activator of an open reading frame encoding one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the cell expressing CD34 is a progenitor T cell. In some embodiments, the cell expressing CD34 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator of an open reading frame encoding one or more transcription factors is expressed to induce PSC to differentiate into a cell expressing CD34. In some embodiments, the cell expressing CD34 is provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into a cell expressing CD34. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD34 from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, in 4 days or less or 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0070] The present disclosure provides a cell expressing CD4. The cell expressing CD4 may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. The cell expressing CD4 may include one or more transcription factors. The cell expressing CD4 may include a transcription activator of an open reading frame encoding one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the cell expressing CD4 is a progenitor T cell. In some embodiments, the cell expressing CD4 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator of an open reading frame encoding one or more transcription factors is expressed to induce PSC to differentiate into a cell expressing CD4. In some embodiments, the cell expressing CD4 is provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into a cell expressing CD4. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD4 from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, in 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0071] The present disclosure provides a T cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the T cell is a progenitor T cell. In some embodiments, the T cell is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into T cells. In some embodiments, T cells are provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into T cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of T cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0072] The present disclosure provides a regulatory T cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into regulatory T cells. In some embodiments, regulatory T cells are provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into regulatory T cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of regulatory T cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0073] The present disclosure provides a T cell progenitor cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into T cell progenitor cells. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, T cell progenitor cells are provided in a culture medium. The culture medium may not need to be changed during the differentiation of PSC into T cell progenitor cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of T cell progenitor cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0074] The present disclosure provides a mature T cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into mature T cells. In some embodiments, mature T cells are provided in a culture medium. The culture medium may not need to be changed during the differentiation of PSC into mature T cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces the differentiation of mature T cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces the differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0075] The present disclosure provides a population of two or more immune cells described herein. In some embodiments, at least one immune cell is a T cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, at least one immune cell is an early hematopoietic progenitor cell that tends to be a T cell. In some embodiments, at least one immune cell is a directed progenitor cell. In some embodiments, at least one immune cell is a mature T cell. In some embodiments, at least one immune cell is a hematopoietic stem / progenitor cell. In some embodiments, at least one immune cell is a NK cell. In some embodiments, at least one immune cell expresses FOXP3. In some embodiments, at least one immune cell expresses CD45. In some embodiments, at least one immune cell expresses CD34. In some embodiments, at least one immune cell expresses CD4.

[0076] Regulatory T cells (Treg)

[0077] Throughout life, peripheral regulatory mechanisms control autoreactive cells that escape initial checkpoints and regulate undesirable immune responses to symbiotic and environmental antigens. Peripheral tolerance can include active mechanisms, in which regulatory T cells (Tregs) can play a major role. Tregs can play their functions in different tissues, at sites of inflammation, and in close contact with T effector (Teff) cells.

[0078] Treg can express protein forkhead box P3 (FOXP3). FOXP3 can be bound to a specific region of DNA and help control the activity of genes involved in immune system regulation. FOXP3 positive (FOXP3+) Treg can be propagated in a limited manner in vitro and can have problematic cytokines, except low transforming growth factor-β (TGF-β) and interleukin-35 (IL-35). However, they can respond very actively to interleukin-2 (IL-2), which can work through its receptor and through the activation of STAT5. IL-2 is an important factor for Treg survival and maintenance in vivo and may require higher doses for expansion in vitro. Other factors may affect Treg, such as TGF-β, thymic stromal lymphopoietin (TSLP), and costimulatory molecules such as CD28. Specific Treg colonies have been associated with the regulation of different T helper (Th) subclasses. FOXP3 can collaborate with cell lineage-specific transcription factors to bias Treg ability toward the regulation of immune responses of diverse types. Mouse models have shown that modulation of Treg cells can treat autoimmune diseases and cancer and can promote organ transplantation and wound healing.

[0079] The expression of FOXP3 may be required for Treg development and may control the genetic programming that specifies the fate of cells. FOXP3 may be a potent inhibitor of IL-2 production and may upregulate the expression of CD25 receptors and the expression of Treg marker CTLA4. FOXP3 may support the maintenance of an immunosuppressive environment. The lack of FOXP3 in specific KO mice or in natural mouse mutants, mice with dandruff may lead to massive lymphocyte proliferation and severe autoimmune syndrome. Mutations in FOXP3 may lead to similar phenotypes in men. However, the establishment of mouse strains carrying defective FoxP3 alleles has demonstrated that this gene may be necessary for the function of Treg, but not for the development of Treg. Specifically, FOXP3 may enhance the response to IL-2 of pre-established Treg features. In humans, Treg differentiation may be characterized by specific demethylation of more than one hundred loci, which become accessible for FOXP3 binding when FOXP3 is expressed. Importantly, the CpG methylation state of DNA can regulate FoxP3 expression and reach its target. Regulation of FOXP3 expression can occur at both transcriptional and translational levels. The present disclosure provides one or more Treg or FOXP3-expressing cells, which include one or more exogenous expression cassettes. One or more expression cassettes may include one or more transcription factors. One or more transcription factors may induce the differentiation of one or more PSCs. The exogenous expression cassette may induce one or more PSCs to differentiate into Treg or FOXP3-expressing cells. The exogenous expression cassette may induce one or more PSCs to differentiate into Treg or FOXP3-expressing cells in 28 days or less. One or more PSCs may be provided in a culture medium. The culture medium may not need to be changed during the differentiation of PSCs into Treg or FOXP3-expressing cells. The culture medium may not require any nutrients, growth factors, or microenvironment or matrix optimization.

[0080] The present disclosure provides a regulatory T cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into regulatory T cells. In some embodiments, regulatory T cells are provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into regulatory T cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of regulatory T cells from PSC. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0081] The present disclosure provides a cell colony comprising one or more regulatory T cells or cells expressing FOXP3 differentiated from one or more PSCs. The cell colony may comprise adherent cells. The cell colony may comprise suspension cells. The cell colony may comprise adherent cells and suspension cells. The cell colony may be provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSCs into Tregs or cells expressing FOXP3. The culture medium may not require any nutrients, growth factors, or microenvironment or matrix optimization. At least 5% of the cells may express FOXP3. At least about 5%, at least about 6%, at least about 7%, at least about 8%, at least about 9%, at least about 10%, at least about 11%, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least about 16%, at least about 17%, at least about 18%, at least about 19%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, or at least about 40% of the cells may express FOXP3.

[0082] Cells expressing CD45

[0083] CD45 is a molecule that may be present on the surface of nucleated hematopoietic cells and their precursors. Specifically, CD45 is a protein tyrosine phosphatase that may be expressed in leukocytes. CD45 may be a type I transmembrane protein. CD45 may be a marker for committed progenitor cells and may be expressed throughout the progenitor cell stage and in mature T cells.

[0084] The present disclosure provides a cell expressing CD45 or a cell expressing any other immune cell or immune-related marker disclosed herein, the marker is associated with non-natural exogenous induction of (TF family or TF or TF combination) of iPSC differentiation along the immune cell differentiation trajectory, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the cell expressing CD45 is a progenitor T cell. In some embodiments, the cell expressing CD45 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into a cell expressing CD45. In some embodiments, cells expressing CD45 are provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into cells expressing CD45. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of cells expressing CD45 from PSCs in 28 days or less. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0085] The present disclosure provides a cell colony, the cell colony comprising two or more cells expressing CD45, or two or more cells expressing any other immune cell or immune-related marker disclosed herein, the marker being associated with non-natural exogenous induction of (TF family or TF or TF combination) iPSC differentiation along the immune cell differentiation track disclosed herein. The cell colony may comprise adherent cells. The cell colony may comprise suspension cells. The cell colony may comprise adherent cells and suspension cells. The cell colony may be provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into immune cells or immune-like cells or cells expressing CD45, cells expressing CD34 or cells expressing CD4 or cells expressing CD8 or cells expressing immune cell lineage markers or immune cell-related markers disclosed herein, the marker being associated with non-natural exogenous induction of (TF formula) iPSC differentiation along the immune cell differentiation track. The cell colony may be provided in a culture medium. The culture medium may not require any nutrients, growth factors or microenvironment or matrix optimization. At least one cell in the cell colony may express CD45. At least about 0.5% to at least about 20% of the cell population may express CD45. At least about 0.5%, at least about 1%, at least about 2%, at least about 3%, at least about 4%, at least about 5%, at least about 6%, at least about 7%, at least about 8%, at least about 9%, at least about 10%, at least about 11%, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least about 16%, at least about 17%, at least about 18%, at least about 19%, at least about 20%, or more of the cell population may express CD45. In some embodiments, at least about 1% to at least about 6% of the cell population may express CD45. In some embodiments, at least about 3% to at least about 9% of the cell population may express CD45. In some embodiments, greater than 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80% of the cell population may express CD45.

[0086] In some embodiments, at least 0.01% of the immune cells express CD45 or may express any immune cell lineage marker or immune cell-associated marker disclosed herein, which is associated with non-natural exogenous induction of (TF recipe) iPSC differentiation along the immune cell differentiation trajectory. In some embodiments, at least 0.1% of the immune cells express CD45. In some embodiments, at least 0.5% of the immune cells express CD45. In some embodiments, at least 1% of the immune cells express CD45. In some embodiments, at least 1.5% of the immune cells express CD45. In some embodiments, at least 2% of the immune cells express CD45. In some embodiments, at least 3% of the immune cells express CD45. In some embodiments, at least 4% of the immune cells express CD45. In some embodiments, at least 5% of the immune cells express CD45. In some embodiments, at least 6% of the immune cells express CD45. In some embodiments, at least 7% of the immune cells express CD45. In some embodiments, at least 8% of the immune cells express CD45. In some embodiments, at least 9% of the immune cells express CD45. In some embodiments, at least 10% of the immune cells express CD45. In some embodiments, at least 12% of the immune cells express CD45. In some embodiments, at least 15% of the immune cells express CD45. In some embodiments, at least 17% of the immune cells express CD45. In some embodiments, at least 20% of the immune cells express CD45. In some embodiments, at least 25% of the immune cells express CD45. In some embodiments, at least 30% of the immune cells express CD45. In some embodiments, at least 35% of the immune cells express CD45. In some embodiments, at least 40% of the immune cells express CD45. In some embodiments, at least 45% of the immune cells express CD45. In some embodiments, at least 50% of the immune cells express CD45. In some embodiments, at least 60% of the immune cells express CD45. In some embodiments, at least 70% of the immune cells express CD45. In some embodiments, at least 80% of the immune cells express CD45. In some embodiments, at least 90% of the immune cells express CD45. In some embodiments, at least 95% of the immune cells express CD45. In some embodiments, at least 99% of the immune cells express CD45.

[0087] In some embodiments, the percentage of immune cells expressing CD45, or cells expressing any other immune cell marker or immune cell lineage-associated marker disclosed herein, associated with non-natural, exogenous induction of (a disclosed TF family, TF, TF combination) iPSC differentiation along an immune cell differentiation trajectory, is at least about 1% to about 50%, or at least about 1% to greater than 50%, 60%, 70%, or 80%. In some embodiments, the portion of the cell population expressing CD45 is at least about 1% to about 6.5%. In some embodiments, the portion of the cell population that expresses CD45 is at least about 1% to about 1.5%, about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1.5% to about 4%, about 1.5% to about 4.5%, about 1.5% to about 5%, about 1.5% to about 5.5%, about 1.5% to about 6%, about 1.5% to about 6.5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%, about 2% to about 6.5%, about 2% to about 3%, about 2.5% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%. 5% to about 4%, about 2.5% to about 4.5%, about 2.5% to about 5%, about 2.5% to about 5.5%, about 2.5% to about 6%, about 2.5% to about 6.5%, about 3% to about 3.5%, about 3% to about 4%, about 3% to about 4.5%, about 3% to about 5%, about 3% to about 5.5%, about 3% to about 6%, about 3% to about 6.5%, about 3.5% to about 4%, about 3.5% to about 4.5%, about 3.5% to about 5%, about 3.5% to about 5.5%, about In some embodiments, the portion of the cell population that expresses CD45 is at least about 1%, about 1.5%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%,In some embodiments, the portion of a cell population expressing CD45 is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%. In some embodiments, the portion of a cell population expressing CD45 is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, about 6.5%. In some embodiments, the portion of a cell population expressing CD45 is at least about 7% to about 50%. In some embodiments, the portion of the cell population that expresses CD45 is at least about 7% to about 7.5%, about 7% to about 8%, about 7% to about 8.5%, about 7% to about 9%, about 7% to about 9.5%, about 7% to about 10%, about 7% to about 15%, about 7% to about 20%, about 7% to about 30%, about 7% to about 40%, about 7% to about 50%, about 7.5% to about 8%, about 7.5% to about 8.5%, about 7.5% to about 9%, about 7.5% to about 9.5%, about 7% to about 10%, about 7% to about 15%, about 7% to about 20%, about 7% to about 30%, about 7% to about 40%, about 7% to about 50%, about 7.5% to about 8%, about 7.5% to about 8.5%, about 7.5% to about 9%, about 7.5% to about 9.5%, about 7.5% to about 10%, about 7.5% to about 15%, about 7.5% to about 20%, about 7.5% to about 30%, about 7.5% to about 40%, about 7.5% to about 50%, about 8% to about 8.5%, about 8% to about 9%, about 8% to about 9.5%, about 8% to about 10%, about 8% to about 15%, about 8% to about 20%, about 8% to about 30%, about 8% to about 40%, about 8% to about 50%, about 8.5% to about 9%, about 8.5% to about 9.5%, about 8% to about 10%, about 8% to about 15%, about 8% to about 20%, about 8% to about 30%, about 8% to about 40%, about 8% to about 50%, about 8.5% to about 9%, about 8.5% to about 9.5%, about 8. 5% to about 10%, about 8.5% to about 15%, about 8.5% to about 20%, about 8.5% to about 30%, about 8.5% to about 40%, about 8.5% to about 50%, about 9% to about 9.5%, about 9% to about 10%, about 9% to about 15%, about 9% to about 20%, about 9% to about 30%, about 9% to about 40%, about 9% to about 50%, about 9.5% to about 10%, about 9.5% to about 15%, about 9% to about 20%, about 9% to about 30%, about 9% to about 40%, about 9% to about 50%, about 9.5% to about 10%, about 9.5% to about 15%, about 9.5% to about 20%, about 9.5% to about 30%, about In some embodiments, the portion of the cell population that expresses CD45 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 15%, about 10%, about 20%, about 10%, about 30%, about 10%, about 40%, about 10%, about 50%, about 15%, about 20%, about 15%, about 30%, about 15%, about 40%, about 15%, about 50%, about 20%, about 30%, about 20%, about 40%, about 20%, about 50%, about 30%, about 40%, about 30%, about 50%, or about 40% to 50%. In some embodiments, the portion of the cell population that expresses CD45 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 15%, about 20%, about 30%, about 40%, or about 50%.In some embodiments, the portion of a cell population that expresses CD45 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 15%, about 20%, about 30%, or about 40%. In some embodiments, the portion of a cell population that expresses CD45 is at least about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 15%, about 20%, about 30%, about 40%, or about 50%. In some embodiments, the portion of a cell population that expresses CD45 is greater than 50%.

[0088] Cells expressing CD34

[0089] The present disclosure provides a cell expressing CD34, or a cell expressing any other immune cell marker or immune cell lineage-related marker disclosed herein, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be a non-natural or exogenous transcription factor. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cell expressing CD34 is a progenitor T cell. In some embodiments, the cell expressing CD34 is a mature T cell. In some embodiments, the cell expressing CD34 is a hematopoietic stem / progenitor cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into a cell expressing CD34. In some embodiments, cells expressing CD34 are provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into a cell expressing CD34. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of cells expressing CD34 from PSCs in 28 days or less. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0090] The present disclosure provides a cell population comprising two or more cells expressing CD34 disclosed herein. At least one cell in the cell population can express CD34. At least about 0.5% to at least about 20% of the cell population can express CD34. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more of the cell population can express CD34. In some embodiments, at least about 2% to at least about 2.5% of the cell population can express CD34. In some embodiments, the portion of the cell population expressing CD34 is at least about 1% to about 6.5%. In some embodiments, the portion of the cell population expressing CD34 can be greater than 6.5%. In some embodiments, the portion of the cell population that expresses CD34 is at least about 1% to about 1.5%, about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1.5% to about 4%, about 1.5% to about 4.5%, about 1.5% to about 5%, about 1.5% to about 5.5%, about 1.5% to about 6%, about 1.5% to about 6.5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%, about 2% to about 6.5%, about 2% to about 3%, about 2.5% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%. 5% to about 4%, about 2.5% to about 4.5%, about 2.5% to about 5%, about 2.5% to about 5.5%, about 2.5% to about 6%, about 2.5% to about 6.5%, about 3% to about 3.5%, about 3% to about 4%, about 3% to about 4.5%, about 3% to about 5%, about 3% to about 5.5%, about 3% to about 6%, about 3% to about 6.5%, about 3.5% to about 4%, about 3.5% to about 4.5%, about 3.5% to about 5%, about 3.5% to about 5.5%, about 3.5% to about 6%, about 3.5% to about 6.5%, about 4% to about 4.5%, about 4% to about 5%, about 4% to about 5.5%, about 4% to about 6%, about 4% to about 6.5%, about 4.5% to about 5%, about 4.5% to about 5.5%, about 4.5% to about 6%, about 4.5% to about 6%, about 5% to about 6.5%, about 5% to about 5.5%, about 5% to about 6%, about 5% to about 6.5%, about 5.5% to about 6%, about 5.5% to about 6%, or about 6% to about 6.5%.In some embodiments, the portion of a cell population expressing CD34 is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, or about 6.5%. In some embodiments, the portion of a cell population expressing CD34 is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%. In some embodiments, the portion of a cell population expressing CD34 is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%.

[0091] CD4-expressing cells

[0092] The present disclosure provides a cell expressing CD4, or a cell expressing any other immune cell marker disclosed herein or immune cell lineage-related marker, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cell expressing CD4 is a progenitor T cell. In some embodiments, the cell expressing CD4 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into cells expressing CD4. In some embodiments, cells expressing CD4 are provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into cells expressing CD4. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of cells expressing CD4 from PSCs in 28 days or less. In some embodiments, nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames encoding one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0093] The present disclosure provides a cell population comprising two or more cells expressing CD4 disclosed herein. At least one cell in the cell population can express CD4. At least about 0.5% to at least about 20% in the cell population can express CD4. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more in the cell population can express CD4. In some embodiments, at least about 2% to at least about 5% in the cell population can express CD4. In some embodiments, the portion expressing CD4 in the cell population is at least about 1% to about 6.5%. In some embodiments, the portion of the cell population that expresses CD4 is at least about 1% to about 1.5%, about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3 ... 1.5% to about 4%, about 1.5% to about 4.5%, about 1.5% to about 5%, about 1.5% to about 5.5%, about 1.5% to about 6%, about 1.5% to about 6.5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%, about 2% to about 6.5%, about 2% to about 3%, about 2.5% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%. 5% to about 4%, about 2.5% to about 4.5%, about 2.5% to about 5%, about 2.5% to about 5.5%, about 2.5% to about 6%, about 2.5% to about 6.5%, about 3% to about 3.5%, about 3% to about 4%, about 3% to about 4.5%, about 3% to about 5%, about 3% to about 5.5%, about 3% to about 6%, about 3% to about 6.5%, about 3.5% to about 4%, about 3.5% to about 4.5%, about 3.5% to about 5%, about 3.5% to about 5.5%, about 3.5% to about 6%, about 3.5% to about 6.5%, about 4% to about 4.5%, about 4% to about 5%, about 4% to about 5.5%, about 4% to about 6%, about 4% to about 6.5%, about 4.5% to about 5%, about 4.5% to about 5.5%, about 4.5% to about 6%, about 4.5% to about 6%, about 5% to about 6.5%, about 5% to about 5.5%, about 5% to about 6%, about 5% to about 6.5%, about 5.5% to about 6%, about 5.5% to about 6%, or about 6% to about 6.5%.In some embodiments, the portion of cells expressing CD4 in a population is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, or about 6.5%. In some embodiments, the portion of cells expressing CD4 in a population is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%. In some embodiments, the portion of cells expressing CD4 in a population is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%. In some embodiments, the portion of cells expressing CD4 in a population is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, or about 6.5%. In some embodiments, the portion of cells expressing CD4 in a population is at least about 7% to about 20%. In some embodiments, the portion of the cell population that expresses CD4 is at least about 7% to about 7.5%, about 7% to about 8%, about 7% to about 8.5%, about 7% to about 9%, about 7% to about 9.5%, about 7% to about 10%, about 7% to about 12%, about 7% to about 14%, about 7% to about 15%, about 7% to about 18%, about 7% to about 20%, about 7.5% to about 8%, about 7.5% to about 8.5%, about 7.5% to about 9%, about 7.5% to about 9.5%, about 7.5% to about 10%, about 7.5% to about 12%, about 7.5% to about 14%, about 7.5% to about 15%, about 7.5% to about 18%, about 7.5% to about 20%, about 8% to about 8.5%, about 8% to about 9%, about 8% to about 9.5%, about 8% to about 10%, about 8% to about 12%, about 8% to about 14%, about 8% to about 15%, about 8% to about 18%, about 8% to about 20%, about 8.5% to about 9%, about 8.5% to about 9.5%, about 8% to about 10%, about 8% to about 12%, about 8% to about 14%, about 8% to about 15%, about 8% to about 18%, about 8% to about 20%, about 8.5% to about 9%, about 8.5% to about 9.5%, about 8. 5% to about 10%, about 8.5% to about 12%, about 8.5% to about 14%, about 8.5% to about 15%, about 8.5% to about 18%, about 8.5% to about 20%, about 9% to about 9.5%, about 9% to about 10%, about 9% to about 12%, about 9% to about 14%, about 9% to about 15%, about 9% to about 18%, about 9% to about 20%, about 9.5% to about 10%, about 9.5% to about 12%, about 9.5% to about 14%, about 9.5% to about 15%, about 9.5% to about 18%, about 9.5% to about 20%, about 10% to about 12%, about 10% to about 14%, about 10% to about 15%, about 10% to about 18%, about 10% to about 20%, about 12% to about 14%, about 12% to about 15%, about 12% to about 18%, about 12% to about 20%, about 14% to about 15%, about 14% to about 18%, about 14% to about 20%, about 15% to about 18%, about 15% to about 20%, or about 18% to about 20%.In some embodiments, the portion of cells expressing CD4 in the population is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 12%, about 14%, about 15%, about 18%, or about 20%. In some embodiments, the portion of cells expressing CD4 in the population is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 12%, about 14%, about 15%, or about 18%. In some embodiments, the portion of cells expressing CD4 in the population is at least about 7.5%, about 8%, about 8.5%, about 9%, about 9.5%, about 10%, about 12%, about 14%, about 15%, about 18%, or about 20%. In some embodiments, the portion of cells expressing CD4 in the population may be greater than 20%.

[0094] CD8

[0095] The present disclosure provides a cell expressing CD8, or a cell expressing any other immune cell marker disclosed herein or immune cell lineage-related marker, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cell expressing CD8 is a progenitor T cell. In some embodiments, the cell expressing CD8 is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into cells expressing CD8. In some embodiments, cells expressing CD8 are provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into cells expressing CD8. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD8 from PSC in 28 days or less. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0096] The present disclosure provides a cell population comprising two or more cells expressing CD8 disclosed herein. At least one cell in the cell population can express CD8. At least about 0.5% to at least about 20% in the cell population can express CD8. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more in the cell population can express CD8. In some embodiments, at least about 2% to at least about 5% in the cell population can express CD8.

[0097] In some embodiments, the portion of the cell population expressing CD8 is at least about 1% to about 5%. In some embodiments, the portion of the cell population expressing CD8 is at least about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2.5 In some embodiments, the portion of the cell population expressing CD8 is at least about 1%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 2.5% to about 5%, about 3% to about 3.5%, about 3% to about 4%, about 3% to about 4.5%, about 3% to about 5%, about 3.5% to about 4%, about 3.5% to about 4.5%, about 3.5% to about 5%, about 4% to about 4.5%, about 4% to about 5%, or about 4.5% to about 5%. In some embodiments, the portion of the cell population expressing CD8 is at least about 1%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, or about 5%. In some embodiments, the portion of the cell population expressing CD8 is at least about 1%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, or about 4.5%. In some embodiments, the portion of the population of cells that express CD8 is at least about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, or about 5%.

[0098] Double positive

[0099] CD45+ and CD34+

[0100] The present disclosure provides a cell expressing CD45 and CD34, or a cell expressing any other immune cell marker disclosed herein or immune cell lineage-related marker, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cells expressing CD45 and CD34 are early hematopoietic stem / progenitor cells. In some embodiments, the cells expressing CD45 and CD34 are mature hematopoietic stem / progenitor cells. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC differentiation into cells expressing CD45 and CD34. In some embodiments, cells expressing CD45 and CD34 are provided in culture medium. The culture medium does not need to be changed during the differentiation of PSC into cells expressing CD45 and CD34. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD45 and CD34 from PSC. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0101] The present disclosure provides a cell population comprising two or more cells expressing CD45 and CD34 disclosed herein. At least one cell in the cell population can express CD45 and CD34. At least about 0.5% to at least about 20% of the cell population can express CD45 and CD34. In some embodiments, at least about 2% to at least about 5% of the cell population can express CD45 and CD34. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 1% to about 6.5%. In some embodiments, the portion of the cell population that expresses CD45 and CD34 is at least about 1% to about 1.5%, about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1% to about 5.5%, about 1% to about 6%, about 1% to about 6.5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3.5%. 5%, about 1.5% to about 4%, about 1.5% to about 4.5%, about 1.5% to about 5%, about 1.5% to about 5.5%, about 1.5% to about 6%, about 1.5% to about 6.5%, about 2% to about 2.5%, about 2% to about 3%, about 2% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%, about 2% to about 6.5%, about 2.5% to about 3%, about 2.5% to about 3.5%, about 2% to about 4%, about 2% to about 4.5%, about 2% to about 5%, about 2% to about 5.5%, about 2% to about 6%, about 2% to about 6.5%, about 2.5% to about 3%, about 2.5% to about 3.5%, From about 2.5% to about 4%, from about 2.5% to about 4.5%, from about 2.5% to about 5%, from about 2.5% to about 5.5%, from about 2.5% to about 6%, from about 2.5% to about 6.5%, from about 3% to about 3.5%, from about 3% to about 4%, from about 3% to about 4.5%, from about 3% to about 5%, from about 3% to about 5.5%, from about 3% to about 6%, from about 3% to about 6.5%, from about 3.5% to about 4%, from about 3.5% to about 4.5%, from about 3% to about 5%, from about 3% to about 5.5%, from about 3% to about 6%, from about 3% to about 6.5%, from about 3.5% to about 4%, from about 3.5% to about 4.5%, from about 3.5% to about 5%, from about 3.5% to about 5.5% %, about 3.5% to about 6%, about 3.5% to about 6%, about 4% to about 4.5%, about 4% to about 5%, about 4% to about 5.5%, about 4% to about 6%, about 4% to about 6.5%, about 4.5% to about 5%, about 4.5% to about 5.5%, about 4.5% to about 6%, about 4.5% to about 6%, about 5% to about 6.5%, about 5% to about 5.5%, about 5% to about 6%, about 5% to about 6.5%, about 5% to about 5.5%, about 5% to about 6%, about 5% to about 6.5%, about 5.5% to about 6%, about 5.5% to about 6%, or about 6% to about 6.5%. In some embodiments, the portion of the cell population that expresses CD45 and CD34 is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, or about 6.5%.In some embodiments, the portion of a cell population expressing CD45 and CD34 is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, or about 6%. In some embodiments, the portion of a cell population expressing CD45 and CD34 is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, about 5%, about 5.5%, about 6%, or about 6.5%.

[0102] In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7% to about 20%. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7% to about 7.5%, about 7% to about 8%, about 7% to about 8.5%, about 7% to about 9%, about 7% to about 10%, about 7% to about 15%, about 7% to about 20%, about 7.5% to about 8%, about 7.5% to about 8.5%, about 7.5% to about 9%, about 7.5% to about 10%, about 7.5% to about 15%, about 7.5% to about 20%. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 10%, about 15%, or about 20%. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 10%, about 15%, or about 20%. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 10%, about 15%, or about 20%. In some embodiments, the portion of the cell population expressing CD45 and CD34 is at least about 7%, about 7.5%, about 8%, about 8.5%, about 9%, about 10%, or about 15%. In some embodiments, the portion of the cell population that expresses CD45 and CD34 is at least about 7.5%, about 8%, about 8.5%, about 9%, about 10%, about 15%, or about 20%.

[0103] Double positive

[0104] CD45+ and CD56+

[0105] The present disclosure provides a cell expressing CD45 and CD56, or a cell expressing any other immune cell marker disclosed herein or immune cell lineage-related marker, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a NK (natural killer) cell. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cells expressing CD45 and CD56 are progenitor NK cells. In some embodiments, the cells expressing CD45 and CD56 are mature NK cells. In some embodiments, the nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into cells expressing CD45 and CD56. In some embodiments, cells expressing CD45 and CD56 are provided in culture medium. The culture medium does not need to be changed during the differentiation of PSC into cells expressing CD45 and CD56. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD45 and CD56 from PSC. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0106] The present disclosure provides a cell population comprising two or more cells expressing CD45 and CD56 disclosed herein. At least one cell in the cell population can express CD45 and CD56. At least about 0.5% to at least about 80% of the cell population can express CD45 and CD56. In some embodiments, greater than 80% of the cell population can express CD45 and CD56. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 1% to about 12%. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 1% to about 2%, about 1% to about 3%, about 1% to about 4%, about 1% to about 5%, about 1% to about 6%, about 1% to about 7%, about 1% to about 8%, about 1% to about 9%, about 1% to about 10%, about 1% to about 11%, about 1% to about 12%, about 2% to about 3%, about 2% to about 4%, about 2% to about 5%, about 2% to about 6%, about 2% to about 7%, about 2% to about 8%, about 2% to about 9%, about 2% to about 10%, about 2% to about 11%, about 2% to about 12%, about 3% to about 4%, about 3% to about 5%, about 3% to about 6%, about 3% to about 7%, about 3% to about 8%, about 3% to about 9%, about 3% to about 10%, about 3% to about 11%, about 3% to about 12%, about 4% to about 5%, about 4% to about 6%, From about 4% to about 7%, from about 4% to about 8%, from about 4% to about 9%, from about 4% to about 10%, from about 4% to about 11%, from about 4% to about 12%, from about 5% to about 6%, from about 5% to about 7%, from about 5% to about 8%, from about 5% to about 9%, from about 5% to about 10%, from about 5% to about 11%, from about 5% to about 12%, from about 6% to about 7%, from about 6% to about 8%, from about 6% to about 9%, from about 6% to about 10%, from about 6% to about In some embodiments, the portion of the cell population that expresses CD45 and CD56 is at least about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 8% to about 12%, about 9% to about 10%, about 8% to about 11%, about 8% to about 12%, about 9% to about 10%, about 9% to about 11%, about 9% to about 12%, about 10% to about 11%, about 10% to about 12%, or about 11% to about 12%. In some embodiments, the portion of the cell population that expresses CD45 and CD56 is at least about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, or about 12%. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, or about 11%.In some embodiments, the portion of the cell population that expresses CD45 and CD56 is at least about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, or about 12%.

[0107] In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 15% to about 80%. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 15% to about 20%, about 15% to about 30%, about 15% to about 40%, about 15% to about 45%, about 15% to about 50%, about 15% to about 55%, about 15% to about 60%, about 15% to about 65%, about 15% to about 70%, about 15% to about 75%, about 15% to about 80%, about 20% to about 30%, about 20% to about 40%, about 20% to about 45%, about 20% to about 50%, about 20% to about 55%, about 20% to about 60%, about 20% to about 65%, about 20% to about 70%, about 20% to about 75%, about 20% to about 80%, about 30% to about 40%, about 30% to about 45%, about 30% to about 50%, about 30% to about 55%, about 30% to about 60%, about 30% to about 65%, about 30% to about 70%, about 30% to about 75%, about 30% to about 80%, about 40% to about 45%, about 40% to about 50%, about 30% to about 55%, about 30% to about 60%, about 30% to about 65%, about 30% to about 70%, about 30% to about 75%, about 30% to about 80%, about 40% to about 45%, about 40% to about 50%, about 40% to about 55%, about 40% to about 60%, about 40% to about 65%, about 40% to about 70%, about 40% to about 75%, about 40% to about 80%, about 45% to about 50%, about 45% to about 55%, about 45% to about 60%, about 45% to about 65%, about 45% to about 70%, about 45% to about 75%, about 45% to about 80%, about 50% to about 55%, about 50% to about 60%, about 50% to about 65%, about 50% to about 70%. In some embodiments, the portion of the cell population that expresses CD45 and CD56 is at least about 15%, about 20%, about 30%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 60% to about 65%, about 60% to about 70%, about 60% to about 75%, about 60% to about 80%, about 65% to about 70%, about 65% to about 75%, about 65% to about 80%, about 70% to about 75%, about 70% to about 80%, or about 75% to about 80%. In some embodiments, the portion of the cell population that expresses CD45 and CD56 is at least about 15%, about 20%, about 30%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, or about 80%. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 15%, about 20%, about 30%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75%. In some embodiments, the portion of the cell population expressing CD45 and CD56 is at least about 20%, about 30%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, or about 80%.In some embodiments, the fraction of cells in the population that express CD45 and CD56 is greater than 80%. In some embodiments, the fraction of cells in the population that express CD45 and CD56 is anywhere in the range of 0.5% to greater than 90%.

[0108] Double positive

[0109] CD14 and CD11b

[0110] The present disclosure provides a cell expressing CD14 and CD11b, or a cell expressing any other immune cell marker disclosed herein or immune cell lineage-related marker, the cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a macrophage / granulocyte lineage cell, including macrophages and granulocytes. In some embodiments, the immune cell is any immune cell expressing an immune cell marker disclosed herein or an immune cell lineage-related marker. In some embodiments, the cell expressing CD14 and CD11b is a macrophage / granulocyte progenitor cell. In some embodiments, the cell expressing CD14 and CD11b is a mature macrophage or granulocyte. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into cells expressing CD14 and CD11b. In some embodiments, cells expressing CD14 and CD11b are provided in culture medium. Culture medium does not need to be changed during differentiation of PSC into cells expressing CD14 and CD11b. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of cells expressing CD14 and CD11b from PSC. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC.

[0111] The present disclosure provides a cell population comprising two or more cells expressing CD14 and CD11b disclosed herein. At least one cell in the cell population can express CD14 and CD11b. At least about 0.5% to at least about 5% of the cell population can express CD14 and CD11b. In some embodiments, greater than 5% of the cell population can express CD14 and CD11b. In some embodiments, the portion expressing CD14 and CD11b in the cell population is at least about 1% to about 5%. In some embodiments, the portion of the cell population that expresses CD14 and CD11b is at least about 1% to about 1.5%, about 1% to about 2%, about 1% to about 2.5%, about 1% to about 3%, about 1% to about 3.5%, about 1% to about 4%, about 1% to about 4.5%, about 1% to about 5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1.5% to about 4%, about 1.5% to about 4.5%, about 1.5% to about 5%, about 1.5% to about 2%, about 1.5% to about 2.5%, about 1.5% to about 3%, about 1.5% to about 3.5%, about 1.5% to about 4%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is at least about 1%, about 1.5%, about 2%, about 2%, about 3%, about 3.5%, about 4%, about 4.5%, about 2%, about 5%, about 2.5% to about 3%, about 2.5% to about 3.5%, about 2.5% to about 4%, about 2.5% to about 4.5%, about 2.5% to about 5%, about 3% to about 3.5%, about 3% to about 4%, about 3% to about 4.5%, about 3% to about 5%, about 3.5% to about 4%, about 3.5% to about 4.5%, about 3.5% to about 5%, about 4% to about 4.5%, about 4% to about 5%, or about 4.5% to about 5%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, or about 5%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is at least about 1%, about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, or about 4.5%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is at least about 1.5%, about 2%, about 2.5%, about 3%, about 3.5%, about 4%, about 4.5%, or about 5%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is less than 1%. In some embodiments, the portion of the cell population expressing CD14 and CD11b is greater than 5%.

[0112] T cells

[0113] The present disclosure provides a T cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the T cell is a progenitor T cell. In some embodiments, the T cell is a mature T cell. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into T cells. In some embodiments, T cells are provided in a culture medium. The culture medium may not have to be changed during the differentiation of PSC into T cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of T cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0114] T cell progenitors

[0115] The present disclosure provides a T cell progenitor cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into T cell progenitor cells. In some embodiments, T cell progenitor cells are provided in a culture medium. The culture medium may not need to be changed during the differentiation of PSC into T cell progenitor cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of T cell progenitor cells from PSC. In some embodiments, in 11 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces differentiation of immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0116] The present disclosure provides a cell population comprising two or more progenitor T cells disclosed herein. At least one cell in the cell population can express CD45. At least about 0.5% to at least about 20% of the cell population can express CD45. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more of the cell population can express CD45. In some embodiments, at least about 1% to at least about 6% of the cell population can express CD45. In some embodiments, at least about 3% to at least about 9% of the cell population can express CD45.

[0117] The present disclosure provides a cell population comprising two or more progenitor T cells disclosed herein. At least one cell in the cell population can express CD34. At least about 0.5% to at least about 20% of the cell population can express CD45. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more of the cell population can express CD34. In some embodiments, at least about 2% to at least about 2.5% of the cell population can express CD34.

[0118] Mature T cells

[0119] The present disclosure provides a kind of mature T cell, which comprises: comprising nucleic acid, one or more transcription factors or transcription activators encoding open reading frames of one or more transcription factors.Transcription factors disclosed herein may be exogenous transcription factors.In some embodiments, nucleic acid, one or more transcription factors or transcription activators encoding open reading frames of one or more transcription factors are expressed to induce PSC to differentiate into mature T cells.In some embodiments, mature T cells are provided in culture medium.The culture medium may not need to be changed during the differentiation of PSC into mature T cells.In some embodiments, in 28 days or less, nucleic acid, one or more transcription factors or transcription activators encoding open reading frames of one or more transcription factors are induced to differentiate mature T cells from PSC.In some embodiments, in 11 days or less, nucleic acid, one or more transcription factors or transcription activators encoding open reading frames of one or more transcription factors are induced to differentiate immune cells from PSC. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors induces differentiation of an immune cell from a PSC in 4 days or less, 1 day or less.

[0120] The present disclosure provides a cell population comprising two or more mature T cells disclosed herein. At least one cell in the cell population can express CD45. At least about 0.5% to at least about 20% of the cell population can express CD45. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more of the cell population can express CD45. In some embodiments, at least about 1% to at least about 6% of the cell population can express CD45. In some embodiments, at least about 3% to at least about 9% of the cell population can express CD45.

[0121] The present disclosure provides a cell population comprising two or more mature T cells disclosed herein. At least one cell in the cell population can express CD34. At least about 0.5% to at least about 20% of the cell population can express CD45. At least about 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20% or more of the cell population can express CD34. In some embodiments, at least about 2% to at least about 2.5% of the cell population can express CD34.

[0122] Transcription Factors

[0123] The present disclosure provides transcription factors and cells for inducing pluripotent stem cells (PSCs) to differentiate into immune cells.

[0124] Transcription factor (TF) can be used to trigger differentiation program.Some transcription factors can be able to induce stem cell differentiation into phenotype.Phenotype can be immune cell.Some transcription factors can be able to induce stem cell differentiation into specific cell lineage, such as hematopoietic stem / progenitor cell, NK cell, regulatory T cell, cell expressing FOXP3, progenitor T cell, cell expressing CD34, cell expressing CD45, cell expressing CD4, and mature T cell.In some aspects, the combination of transcription factors can be used to realize differentiation to specific cell lineage.Combination can realize cell type or cell subtype that can not be realized by any single transcription factor.Combination can realize the same cell type as one of single transcription factors, but can realize more effectively.

[0125] The present disclosure provides an immune cell. The immune cell may include a nucleic acid comprising an open reading frame encoding one or more transcription factors. The nucleic acid may include two or more open reading frames encoding one or more transcription factors. The immune cell may include one or more transcription factors. The immune cell may include a transcription activator of an open reading frame encoding one or more transcription factors. The transcription factor disclosed herein may be an exogenous transcription factor. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the immune cell is an early hematopoietic progenitor cell tending to a T cell. In some embodiments, the immune cell is a directed progenitor cell. In some embodiments, the immune cell is a mature T cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD4. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator of an open reading frame encoding one or more transcription factors is expressed to induce PSC to differentiate into an immune cell. In some embodiments, the nucleic acid may include two or more open reading frames encoding one or more transcription factors. In some embodiments, the nucleic acid may include three or more open reading frames encoding one or more transcription factors. In some embodiments, the nucleic acid may include four or more open reading frames encoding one or more transcription factors. In some embodiments, the nucleic acid may include five or more open reading frames encoding one or more transcription factors. In some embodiments, immune cells are provided in culture medium. The culture medium does not need to be changed during the differentiation of PSC into immune cells. In some embodiments, in 28 days or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces the differentiation of immune cells from PSC. In some embodiments, in 11 days or less, 5 days or less, 4 days or less, 1 day or less, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors induces the differentiation of immune cells from PSC. In some embodiments, the one or more transcription factors comprise any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10, or 11.

[0126] Some transcription factors may require a critical amount of expression to effectively induce differentiation, such as at least 5, 10, 15, 20, 25 or 50 copies / cell of an open reading frame (ORF). Other factors may need to be expressed below a certain threshold because of the high level of toxicity that may exist, such as less than 20, 10 or 5 copies / cell. Increased expression levels may also be achieved by increasing the copy number of the ORF, for example by using a higher copy number vector or by using a transposon.

[0127] Compared to PSCs that do not contain nucleic acids comprising open reading frames encoding one or more transcription factors disclosed herein, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors, PSCs that contain nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors disclosed herein can have higher levels of expression of one or more transcription factors.Compared to PSCs that do not contain nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors disclosed herein, PSCs that contain nucleic acids comprising open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors disclosed herein can have higher levels of mRNA expression of one or more transcription factors. A PSC comprising a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors disclosed herein can have at least about 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 40-fold, 50-fold, 60-fold, 70-fold, 80-fold, 90-fold, 100-fold or more expression of one or more transcription factors compared to a PSC without a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcriptional activator encoding an open reading frame encoding one or more transcription factors disclosed herein.

[0128] Zinc finger C2H2 type

[0129] The present disclosure provides an immune cell, comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame encoding one or more transcription factors, wherein the nucleic acid. The one or more transcription factors may be zinc finger C2H2 type transcription factors. The zinc finger C2H2 type transcription factor may be IKZF1, IKFZ4, KLF10, or ZBTB7B.

[0130] IKZF1

[0131] The present disclosure provides immune cells and PSCs. The present disclosure provides immune cells or PSCs comprising nucleic acids comprising open reading frames encoding one or more transcription factors. The present disclosure provides immune cells or PSCs comprising one or more transcription factors. The present disclosure provides immune cells or PSCs comprising transcriptional activators comprising open reading frames encoding one or more transcription factors, wherein the nucleic acids. The one or more transcription factors may be zinc finger C2H2 type transcription factors. The zinc finger C2H2 type transcription factor may be IKZF1.

[0132] IKZF1 may also be referred to as BAF chromatin remodeling complex subunit IKZF1, CTIP2, HRIT1-α, CTIP-2, radiation-induced tumor suppressor gene 1 protein, B-cell CLL / lymphoma 11B (zinc finger protein), IKZF1, BAF complex component, COUP-TF-interacting protein 2, B-cell lymphoma / leukemia 11B, B-cell CLL / lymphoma 11B, SMARCM2, ZNF856B, BCL-11B, HRit1, RIT1, B-cell CLL / lymphoma 11B / T-cell receptor delta constant region fusion protein, zinc finger protein HRit1α, B-cell lymphoma / leukemia 11B, B-cell CLL / lymphoma 11B, IKZF1 / TRDC fusion, ATL1-α, ATL1-δ, ATL1-γ, ATL1-β, IDDFSTA, IMD49, and ATL1.

[0133] The present disclosure provides an expression cassette comprising one or more transcription factors. The one or more transcription factors may comprise IKZF1. IKZF1 may be introduced into the expression cassette. IKZF1 may be expressed in the expression cassette. IKZF1 may be introduced into the expression cassette together with one or more other transcription factors. A combination comprising one or more transcription factors may be produced. A combination of transcription factors may be introduced into the expression cassette. A combination of transcription factors may be expressed in the expression cassette. The expression cassette may be introduced into PSC. An expression cassette comprising IKZF1 and one or more other transcription factors may induce PSC to differentiate into Treg or cells expressing FOXP3.

[0134] The present disclosure provides a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. One or more transcription factors may comprise IKZF1. An expression cassette comprising IKZF1 may be introduced into a PSC. An expression cassette comprising IKZF1 may be expressed in a PSC. An expression cassette comprising IKZF1 may induce PSC to differentiate into Treg or cells expressing FOXP3. Different amounts of IKZF1 may be introduced into a PSC. For IKZF1, at least 5, 10, 15, 20, 25 or 50 copies of an open reading frame (ORF) may be introduced per cell. An increase in expression levels may also be achieved by increasing the copy number of the ORF, for example, by using a vector with a higher copy number or by using a transposon.

[0135] The present disclosure provides an immune cell, comprising: an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the immune cell is an early hematopoietic progenitor cell tending to a T cell. In some embodiments, the immune cell is a directed progenitor cell. In some embodiments, the immune cell is a mature T cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD4. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into an immune cell. In some embodiments, the immune cell is provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into an immune cell. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 28 days or less. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0136] The present disclosure provides one or more Treg or FOXP3-expressing cells. One or more Treg or FOXP3-expressing cells may comprise nucleic acids. Nucleic acids may comprise open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors. One or more transcription factors may comprise IKZF1. One or more Treg or FOXP3-expressing cells may differentiate from one or more PSCs. One or more Treg or FOXP3-expressing cells may comprise one or more transcription factors, which may induce one or more PSCs to differentiate into one or more Treg or FOXP3-expressing cells.

[0137] The present disclosure provides one or more cells expressing CD45. One or more cells expressing CD45 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF1. One or more cells expressing CD45 may differentiate from one or more PSCs. One or more cells expressing CD45 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD45.

[0138] The present disclosure provides one or more cells expressing CD34. One or more cells expressing CD34 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF1. The one or more cells expressing CD34 may differentiate from one or more PSCs. The one or more cells expressing CD34 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD34.

[0139] The present disclosure provides one or more cells expressing CD4. One or more cells expressing CD4 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF1. The one or more cells expressing CD4 may differentiate from one or more PSCs. The one or more cells expressing CD4 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD4.

[0140] IKZF2

[0141] The present disclosure provides immune cells and PSCs comprising nucleic acids. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The present disclosure provides an immune cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. The one or more transcription factors may be zinc finger C2H2 type transcription factors. The zinc finger C2H2 type transcription factor may be, for example, IKZF2.

[0142] IKZF2 may also be referred to as IKAROS family zinc finger 2, ZNFN1A2, zinc finger protein subfamily 1A, 2 (Helio), Ikaros family zinc finger protein 2, zinc finger protein Helio, Helio, zinc finger DNA binding protein Helio, IKAROS family zinc finger 2 (Helio), ANF1A2 or ZNF1A2.

[0143] The present disclosure provides an expression cassette comprising one or more transcription factors. The one or more transcription factors may comprise IKZF2. IKZF2 may be introduced into the expression cassette. IKZF2 may be expressed in the expression cassette. IKZF2 may be introduced into the expression cassette together with one or more other transcription factors. A combination comprising one or more transcription factors may be produced. A combination of transcription factors may be introduced into the expression cassette. A combination of transcription factors may be expressed in the expression cassette. The expression cassette may be introduced into PSC. An expression cassette comprising IKZF2 and one or more other transcription factors may induce PSC to differentiate into Treg or cells expressing FOXP3.

[0144] The present disclosure provides a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. One or more transcription factors may comprise IKZF2. An expression cassette comprising IKZF2 may be introduced into PSC. An expression cassette comprising IKZF2 may be expressed in PSC. An expression cassette comprising IKZF2 may induce PSC to differentiate into Treg or cells expressing FOXP3. Different amounts of IKZF2 may be introduced into PSC. For IKZF2, at least 5, 10, 15, 20, 25 or 50 copies of open reading frames (ORFs) / cell may be introduced. By increasing the copy number of ORF, for example, by using a vector with a higher copy number or by using a transposon, an increase in expression level may also be achieved.

[0145] The present disclosure provides an immune cell, comprising: an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the immune cell is an early hematopoietic progenitor cell tending to a T cell. In some embodiments, the immune cell is a directed progenitor cell. In some embodiments, the immune cell is a mature T cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD4. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into an immune cell. In some embodiments, the immune cell is provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into an immune cell. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 28 days or less. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0146] The present disclosure provides one or more Treg or FOXP3-expressing cells. One or more Treg or FOXP3-expressing cells may comprise nucleic acids. Nucleic acids may comprise open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors. One or more transcription factors may comprise IKZF2. One or more Treg or FOXP3-expressing cells may differentiate from one or more PSCs. One or more Treg or FOXP3-expressing cells may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more Treg or FOXP3-expressing cells.

[0147] The present disclosure provides one or more cells expressing CD45. One or more cells expressing CD45 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF2. One or more cells expressing CD45 may differentiate from one or more PSCs. One or more cells expressing CD45 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD45.

[0148] The present disclosure provides one or more cells expressing CD34. One or more cells expressing CD34 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF2. One or more cells expressing CD34 may differentiate from one or more PSCs. One or more cells expressing CD34 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD34. In some embodiments, cells expressing immune cell markers such as CD4 or CD8 or CD56 or any other marker associated with an immune cell lineage or an immune cell marker disclosed herein may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing immune cell markers.

[0149] The present disclosure provides one or more cells expressing CD4. One or more cells expressing CD4 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF2. The one or more cells expressing CD4 may differentiate from one or more PSCs. The one or more cells expressing CD4 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD4.

[0150] IKZF4

[0151] The present disclosure provides immune cells and PSCs comprising nucleic acids. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The present disclosure provides an immune cell comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. The one or more transcription factors may be zinc finger C2H2 type transcription factors. The zinc finger C2H2 type transcription factor may be, for example, IKZF4.

[0152] IKZF4 may also be referred to as IKAROS family zinc finger 4, ZNFN1A4, zinc finger protein, subfamily 1A, 4 (Eos), Ikaros family zinc finger protein 4, zinc finger protein Eos, Eos, zinc finger transcription factor Eos, IKAROS family zinc finger 4 (Eos), KIAA1782 or EOS.

[0153] The present disclosure provides an expression cassette comprising one or more transcription factors. The one or more transcription factors may comprise IKZF4. IKZF4 may be introduced into the expression cassette. IKZF4 may be expressed in the expression cassette. IKZF4 may be introduced into the expression cassette together with one or more other transcription factors. A combination comprising one or more transcription factors may be produced. A combination of transcription factors may be introduced into the expression cassette. A combination of transcription factors may be expressed in the expression cassette. The expression cassette may be introduced into PSC. An expression cassette comprising IKZF4 and one or more other transcription factors may induce PSC to differentiate into Treg or cells expressing FOXP3.

[0154] The present disclosure provides a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors or a transcription activator encoding an open reading frame of one or more transcription factors, wherein the nucleic acid. One or more transcription factors may comprise IKZF4. An expression cassette comprising IKZF4 may be introduced into PSC. An expression cassette comprising IKZF4 may be expressed in PSC. An expression cassette comprising IKZF4 may induce PSC to differentiate into Treg or cells expressing FOXP3. Different amounts of IKZF4 may be introduced into PSC. For IKZF4, at least 5, 10, 15, 20, 25 or 50 copies of open reading frames (ORFs) / cell may be introduced. By increasing the copy number of ORF, for example, by using a vector with a higher copy number or by using a transposon, an increase in expression level may also be achieved.

[0155] The present disclosure provides an immune cell, comprising: an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. In some embodiments, the immune cell is a T cell. In some embodiments, the immune cell is a hematopoietic stem / progenitor cell. In some embodiments, the immune cell is a NK cell. In some embodiments, the T cell is a regulatory T cell. In some embodiments, the immune cell is an early hematopoietic progenitor cell tending to a T cell. In some embodiments, the immune cell is a directed progenitor cell. In some embodiments, the immune cell is a mature T cell. In some embodiments, the immune cell expresses FOXP3. In some embodiments, the immune cell expresses CD45. In some embodiments, the immune cell expresses CD34. In some embodiments, the immune cell expresses CD4. In some embodiments, a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors is expressed to induce PSC to differentiate into an immune cell. In some embodiments, the immune cell is provided in a culture medium. The culture medium does not have to be changed during the differentiation of PSC into an immune cell. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 28 days or less. In some embodiments, nucleic acids encoding open reading frames of one or more transcription factors, one or more transcription factors, or transcriptional activators encoding open reading frames of one or more transcription factors induce differentiation of immune cells from PSCs in 11 days or less, 5 days or less, 4 days or less, 1 day or less.

[0156] The present disclosure provides one or more Treg or FOXP3-expressing cells. One or more Treg or FOXP3-expressing cells may comprise nucleic acids. Nucleic acids may comprise open reading frames encoding one or more transcription factors, one or more transcription factors, or transcription activators encoding open reading frames of one or more transcription factors. One or more transcription factors may comprise IKZF4. One or more Treg or FOXP3-expressing cells may differentiate from one or more PSCs. One or more Treg or FOXP3-expressing cells may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more Treg or FOXP3-expressing cells.

[0157] The present disclosure provides one or more cells expressing CD45. One or more cells expressing CD45 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF4. One or more cells expressing CD45 may differentiate from one or more PSCs. One or more cells expressing CD45 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD45.

[0158] The present disclosure provides one or more cells expressing CD34. One or more cells expressing CD34 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading frame of one or more transcription factors. The one or more transcription factors may comprise IKZF4. One or more cells expressing CD34 may differentiate from one or more PSCs. One or more cells expressing CD34 may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing CD34. In some embodiments, cells expressing immune cell markers such as CD4 or CD8 or CD56 or any other marker associated with an immune cell lineage or an immune cell marker disclosed herein may comprise one or more transcription factors that may induce one or more PSCs to differentiate into one or more cells expressing immune cell markers.

[0159] The present disclosure provides one or more cells expressing CD4. One or more cells expressing CD4 may comprise a nucleic acid. The nucleic acid may comprise an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding an open reading fr...

Claims

1. A pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding the open reading frame of one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcription activator encoding the open reading frame of one or more transcription factors comprises one or more Notch receptors or functional derivatives thereof, and wherein in 28 days or less, the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcription activator encoding the open reading frame of one or more transcription factors induces the PSC to differentiate into an immune cell.

2. The PSC of claim 1, wherein the immune cell expresses FOXP3.

3. The PSC of claim 1, wherein the immune cell expresses CD34.

4. The PSC of claim 1, wherein the immune cell expresses CD45.

5. The PSC of claim 1, wherein the immune cell expresses CD4. The PSC of claim 1 , wherein the immune cell expresses CD8.

7. The PSC of claim 1, wherein the immune cell is a regulatory T cell.

8. The PSC of claim 1, wherein the immune cell is a hematopoietic progenitor cell.

9. The PSC of claim 1, wherein the immune cell is a leukocyte.

10. The PSC of claim 1, wherein the immune cell is a natural killer cell.

11. The PSC of claim 1, wherein the immune cell is a macrophage.

12. The PSC of claim 1, wherein the immune cell is a granulocyte.

13. The PSC of claim 1, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcriptional activator encoding the open reading frame encoding one or more transcription factors induces the PSC to differentiate into an immune cell in 11 days or less, 5 days or less, 4 days or less, 3 days or less, 2 days or less, or 1 day or less.

14. The PSC of claim 1, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

15. The PSC of claim 14, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

16. The PSC of claim 15, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

17. The PSC of claim 16, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

18. The PSC of claim 1, wherein the nucleic acid comprises two or more open reading frames encoding one or more transcription factors.

19. The PSC of claim 17, wherein the one or more transcription factors further comprise a transcription factor selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix.

20. The PSC of claim 1, wherein the one or more Notch receptors or the functional derivatives thereof comprise NOTCH1 or a functional derivative thereof.

21. The PSC of claim 20, wherein the NOTCH1 or the functional derivative thereof comprises an intracellular domain ("ICD") of NOTCH1.

22. The PSC of claim 21, wherein the NOTCH1 or the functional derivative thereof consists of the intracellular domain ("ICD") of NOTCH1.

23. A cell population comprising two or more of the PSCs of claim 1.

24. The cell population of claim 23, wherein the cell population is adherent cells.

25. The cell population of claim 23, wherein the cell population is suspension cells.

26. The cell population of claim 23, wherein at least 5%, 10%, 20%, 30% or 40% of the cells in the population express FOXP3.

27. The cell population of claim 23, wherein at least 1%, 2%, 3%, 4%, 5%, 6%, 10%, 15%, 20%, 30%, 40% or 50% of the cells in the population express CD45.

28. The cell population of claim 23, wherein at least 1%, 2%, 2.5% or 3% of the cells in the population express CD8.

29. The cell population of claim 23, wherein at least 1%, 2%, or 2.5% of the cells in the population express CD34.

30. The cell population of claim 23, wherein at least 2%, 3%, 4%, 5%, 10% or 15% of the cells in the population express CD4.

31. The cell population of claim 23, wherein at least 1%, 2%, 2.5% or 3% of the cells in the population express CD45 and CD34.

32. The cell population of claim 23, wherein at least 5%, 6%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70% or 80% of the cells in the population express CD45 and CD56.

33. The cell population of claim 23, wherein at least 5%, 6%, 10% or 15% of the cells in the population express CD45 and CD34.

34. The cell population of claim 23, wherein at least 1%, 2%, 2.5% or 3% of the cells in the population express CD14 and CD11b.

35. A method for generating an immune cell population, the method comprising: a. Providing one or more pluripotent stem cells (PSC); b. Expression in said one or more PSCs i. a nucleic acid comprising an open reading frame encoding one or more transcription factors, ii. one or more transcription factors, or iii. a transcriptional activator of said open reading frame encoding one or more transcription factors, wherein the one or more transcription factors comprise one or more Notch receptors or functional derivatives thereof; and c. generating the immune cell population from the one or more PSCs.

36. The method of claim 35, wherein the immune cell population comprises immune cells expressing FOXP3.

37. The method of claim 35, wherein the immune cell population comprises immune cells expressing CD34.

38. The method of claim 35, wherein the immune cell population comprises immune cells expressing CD45.

39. The method of claim 35, wherein the immune cell population comprises immune cells expressing CD4.

40. The method of claim 35, wherein the immune cell population comprises immune cells expressing CD8.

41. The method of claim 35, wherein the immune cell population comprises regulatory T cells.

42. The method of claim 35, wherein the immune cell population comprises hematopoietic progenitor cells.

43. The method of claim 35, wherein the immune cell population comprises leukocytes.

44. The method of claim 35, wherein the immune cell population comprises natural killer cells.

45. The method of claim 35, wherein the immune cell population comprises macrophages.

46. ​​The method of claim 35, wherein the immune cell population comprises granulocytes.

47. The method of claim 35, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcriptional activator encoding the open reading frame encoding one or more transcription factors induces differentiation of the one or more PSCs into the immune cell population in 28 days or less, 11 days or less, 5 days or less, 4 days or less, 3 days or less, 2 days or less, or 1 day or less.

48. The method of claim 35, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

49. The method of claim 48, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

50. The method of claim 49, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

51. The method of claim 50, wherein the one or more transcription factors further comprise a transcription factor in a transcription factor family selected from the group consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

52. The method of claim 45, wherein the nucleic acid comprises two or more open reading frames encoding one or more transcription factors.

53. The method of claim 51, wherein the one or more transcription factors further comprise a transcription factor selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix.

54. The method of claim 35, wherein the one or more Notch receptors or the functional derivatives thereof comprise NOTCH1 or a functional derivative thereof.

55. The method of claim 54, wherein the NOTCH1 or the functional derivative thereof comprises an intracellular domain ("ICD") of NOTCH1.

56. The method of claim 55, wherein the NOTCH1 or the functional derivative thereof consists of the intracellular domain ("ICD") of NOTCH1.

57. The method of claim 35, wherein the immune cell population is adherent cells.

58. The method of claim 35, wherein the immune cell population is suspension cells.

59. The method of claim 35, wherein at least 5%, 10%, 20%, 30% or 40% of the cell population express FOXP3.

60. The method of claim 35, wherein at least 1%, 2%, 2.5% or 3% of the cell population express CD8.

61. The method of claim 35, wherein at least 1%, 2%, 3%, 4%, 5%, 6%, 10%, 15%, 20%, 30%, 40% or 50% of the cell population express CD45.

62. The method of claim 35, wherein at least 1%, 2%, or 2.5% of the cell population express CD34.

63. The method of claim 35, wherein at least 2%, 3%, 4%, 5%, 10% or 15% of the cell population express CD4.

64. The method of claim 35, wherein at least 1%, 2%, 2.5% or 3% of the cell population express CD45 and CD34.

65. The method of claim 35, wherein at least 5%, 6%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70% or 80% of the cell population express CD45 and CD56.

66. The method of claim 35, wherein at least 5%, 6%, 10% or 15% of the cell population express CD45 and CD34.

67. The method of claim 35, wherein at least 1%, 2%, 2.5% or 3% of the cell population express CD14 and CD11b.

68. A pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator of the open reading frame encoding one or more transcription factors, wherein in 28 days or less, the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcription activator of the open reading frame encoding one or more transcription factors induce the PSC to differentiate into an immune cell.

69. The PSC of claim 68, wherein the immune cell expresses FOXP3.

70. The PSC of claim 68, wherein the immune cells express CD34.

71. The PSC of claim 68, wherein the immune cells express CD45.

72. The PSC of claim 68, wherein the immune cells express CD4.

73. The PSC of claim 68, wherein the immune cells express CD8.

74. The PSC of claim 68, wherein the immune cell is a regulatory T cell.

75. The PSC of claim 68, wherein the immune cell is a hematopoietic progenitor cell.

76. The PSC of claim 68, wherein the immune cells are leukocytes.

77. The PSC of claim 68, wherein the immune cell is a natural killer cell.

78. The PSC of claim 68, wherein the immune cell is a macrophage.

79. The PSC of claim 68, wherein the immune cell is a granulocyte.

80. The PSC of claim 68, wherein the immune cells express CD56.

81. The PSC of claim 68, wherein the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, Runt-related transcription factors, BTB domain-containing proteins, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, zinc finger C2H2 type, interferon regulatory factors, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, NF-κB complex subunits, RAR-related orphan receptors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors.

82. The PSC of claim 81, wherein the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger proteins, forkhead boxes, GATA zinc finger domain-containing proteins, Runt-related transcription factors, BTB domain-containing proteins, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type proteins, phosphatase 1 regulatory subunits, zinc finger C2H2 type, interferon regulatory factors, Kruppel-like factors, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing proteins, microRNA protein-encoding host genes, Notch receptors, microRNA protein-encoding host genes, NF-κB complex subunits, RAR-related orphan receptors, CUT-like homeoboxes and pseudogenes, and T-box transcription factors.

83. The PSC of claim 82, wherein the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

84. The PSC of claim 83, wherein the four or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

85. The PSC of claim 68, wherein the nucleic acid comprises two or more open reading frames encoding one or more transcription factors.

86. The PSC of claim 84, wherein the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix.

87. The PSC of claim 68, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcriptional activator encoding the open reading frame encoding one or more transcription factors induces the PSC to differentiate into an immune cell in 11 days or less, 5 days or less, 4 days or less, 3 days or less, 2 days or less, or 1 day or less.

88. The PSC of claim 68, wherein the one or more transcription factors comprise one or more Notch receptors or functional derivatives thereof.

89. The PSC of claim 88, wherein the one or more Notch receptors or the functional derivatives thereof comprise NOTCH1 or a functional derivative thereof.

90. The PSC of claim 89, wherein the NOTCH1 or the functional derivative thereof comprises an intracellular domain ("ICD") of NOTCH1.

91. The PSC of claim 90, wherein the NOTCH1 or the functional derivative thereof consists of the intracellular domain ("ICD") of NOTCH1.

92. A cell population comprising two or more of the PSCs of claim 68.

93. The cell colony of claim 92, wherein the cell colony is adherent cells.

94. The cell population of claim 92, wherein the cell population is suspension cells.

95. The cell population of claim 92, wherein at least 5% of the cells in the population express FOXP3.

96. The cell population of claim 92, wherein at least 1% of the cells in the population express CD45.

97. The cell population of claim 92, wherein at least 1% of the cells in the population express CD8.

98. The cell population of claim 92, wherein at least 1% of the cells in the population express CD34.

99. The cell population of claim 92, wherein at least 2% of the cells in the population express CD4.

100. The cell population of claim 92, wherein at least 1% of the cell population express CD45 and CD34.

101. The cell population of claim 92, wherein at least 1% of the cell population express CD14 and CD11b.

102. A method for generating an immune cell population, the method comprising: a. Providing one or more pluripotent stem cells (PSC); b. Expression in said one or more PSCs i. a nucleic acid comprising an open reading frame encoding one or more transcription factors, ii. one or more transcription factors, or iii. a transcriptional activator of said open reading frame encoding one or more transcription factors; and c. generating the immune cell population from the one or more PSCs.

103. The method of claim 102, wherein the immune cell population comprises immune cells expressing FOXP3.

104. The method of claim 102, wherein the immune cell population comprises immune cells expressing CD34.

105. The method of claim 102, wherein the immune cell population comprises immune cells expressing CD45.

106. The method of claim 102, wherein the immune cell population comprises immune cells expressing CD4.

107. The method of claim 102, wherein the immune cell population comprises immune cells expressing CD8.

108. The method of claim 102, wherein the immune cell population comprises regulatory T cells.

109. The method of claim 102, wherein the immune cell population comprises hematopoietic progenitor cells.

110. The method of claim 102, wherein the immune cell population comprises leukocytes.

111. The method of claim 102, wherein the immune cell population comprises natural killer cells.

112. The method of claim 102, wherein the immune cell population comprises macrophages.

113. The method of claim 102, wherein the immune cell population comprises granulocytes.

114. The method of claim 102, wherein the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

115. The method of claim 114, wherein the two or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT class homeobox and pseudogene, and T-box transcription factor.

116. The method of claim 115, wherein the three or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

117. The method of claim 116, wherein the four or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, RING finger protein, forkhead box, GATA zinc finger domain-containing protein, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, zinc finger C2H2 type protein, phosphatase 1 regulatory subunit, zinc finger C2H2 type, interferon regulatory factor, Kruppel-like factor, TCF / LEF transcription factor family, Wnt enhanceosome complex, nuclear factor of activated T cells, IPT domain-containing protein, microRNA protein-encoding host gene, Notch receptor, microRNA protein-encoding host gene, NF-κB complex subunit, RAR-related orphan receptor, CUT-like homeobox and pseudogene, and T-box transcription factor.

118. The method of claim 83, wherein the nucleic acid comprises two or more open reading frames encoding one or more transcription factors.

119. The method of claim 117, wherein the one or more transcription factors are selected from the family of transcription factors consisting of zinc finger C2H2 type, BAF complex, Runt-related transcription factor, BTB domain-containing protein, basic helix-loop-helix (bHLH), nuclear factor-κB, and basic helix-loop-helix.

120. The method of claim 102, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcriptional activator encoding the open reading frame encoding one or more transcription factors induces differentiation of the one or more PSCs into the immune cell population in 28 days or less, 11 days or less, 5 days or less, 4 days or less, 3 days or less, 2 days or less, or 1 day or less.

121. The method of claim 102, wherein the one or more transcription factors comprise one or more Notch receptors or functional derivatives thereof.

122. The method of claim 121, wherein the one or more Notch receptors or the functional derivatives thereof comprise NOTCH1 or a functional derivative thereof.

123. The method of claim 122, wherein the NOTCH1 or the functional derivative thereof comprises an intracellular domain ("ICD") of NOTCH1.

124. The method of claim 123, wherein the NOTCH1 or the functional derivative thereof consists of the intracellular domain ("ICD") of NOTCH1.

125. The method of claim 102, wherein at least 5% of the cell population express FOXP3.

126. The method of claim 102, wherein at least 1% of the cell population express CD45.

127. The method of claim 102, wherein at least 1% of the cell population express CD8.

128. The method of claim 102, wherein at least 1% of the cell population express CD34.

129. The method of claim 102, wherein at least 2% of the cell population express CD4.

130. The method of claim 102, wherein at least 1% of the cell population express CD45 and CD34.

131. The method of claim 102, wherein at least 5% of the cell population express CD45 and CD56.

132. The method of claim 102, wherein at least 5% of the cell population express CD45 and CD34.

133. The method of claim 102, wherein at least 1% of the cell population express CD14 and CD11b.

134. An isolated population of immune cells, wherein at least 5%, 6%, 10%, 15%, 20%, 30%, 40%, 50%, 60%, 70% or 80% of the immune cell population are engineered to express CD45 and CD56.

135. The immune cell population of claim 134, wherein at least 1%, 2%, 2.5% or 3% of the immune cell population are engineered to express CD14 and CD11b.

136. The immune cell population of claim 134, wherein at least 1%, 2%, 2.5% or 3% of the immune cell population are engineered to express CD45 and CD34.

137. A pluripotent stem cell (PSC), comprising: a nucleic acid comprising an open reading frame encoding one or more transcription factors, one or more transcription factors, or a transcription activator encoding the open reading frame of one or more transcription factors, wherein the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcription activator encoding the open reading frame of one or more transcription factors comprises any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10, or 11, wherein in 28 days or less, the nucleic acid comprising an open reading frame encoding one or more transcription factors, the one or more transcription factors, or the transcription activator encoding the open reading frame of one or more transcription factors induces the PSC to differentiate into an immune cell.

138. A method of generating an immune cell population, the method comprising: a. Providing one or more pluripotent stem cells (PSC); b. Expression in said one or more PSCs i. a nucleic acid comprising an open reading frame encoding one or more transcription factors, ii. one or more transcription factors, or iii. a transcriptional activator of said open reading frame encoding one or more transcription factors, wherein the one or more transcription factors comprise any one of the transcription factor combinations in Tables 1, 2, 3, 5, 6, 7, 8, 9, 10 or 11; and c. generating the immune cell population from the one or more PSCs.

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