Regulatory t cells expressing chimeric antigen receptors

Abstract

Some aspects relate to engineered cells comprising a nucleotide sequence encoding a chimeric antigen receptor (CAR) and at least at first coding exon of a FOXP3 gene, where expression of the CAR and FOXP3 are linked, such that CAR+ cells have a regulatory T cell (Treg) phenotype. Some aspects relate to methods of producing engineered cells in which expression of a CAR and FOXP3 are linked, thereby reducing the incidence of contaminating CAR+ T effector cells. Further aspects relate to methods of screening CARs suitable for expression by a Treg. Some aspects relate to engineered cells expressing an anti-CD19 chimeric antigen receptor (CAR). Some aspects relate to methods of producing engineered cells expressing an anti-CD19 CAR. Some aspects relate to uses of cells expressing anti- CD19 CARs.

Description

[0001]REGULATORY T CELLS EXPRESSING CHIMERIC ANTIGEN RECEPTORS RELATED APPLICATIONS This application claims the benefit under 35 U.S.C. § 119(e) of U.S. Provisional Application No.63 / 680,488, filed August 7, 2024; U.S. Provisional Application No. 63 / 680513, filed August 7, 2024; U.S. Provisional Application No.63 / 709,395, filed October 18, 2024; U.S. Provisional Application No.63 / 752,568, filed January 31, 2025; U.S. Provisional Application No.63 / 752,603, filed January 31, 2025; U.S. Provisional Application No.63 / 761,230, filed February 21, 2025; U.S. Provisional Application No.63 / 779,140, filed March 27, 2025; U.S. Provisional Application No.63 / 779,145, filed March 27, 2025; U.S. Provisional Application No.63 / 802,176, filed May 8, 2025; U.S. Provisional Application No. 63 / 802,220, filed May 8, 2025; U.S. Provisional Application No.63 / 805,085, filed May 13, 2025; and U.S. Provisional Application No.63 / 805,111, filed May 13, 2025, the contents of each of which are incorporated by reference herein in their entirety. REFERENCE TO AN ELECTRONIC SEQUENCE LISTING The contents of the electronic sequence listing (G097170039WO00-SEQ-NTJ.xml; Size: 223,678 bytes; and Date of Creation: August 7, 2025) is herein incorporated by reference in its entirety. BACKGROUND Undesired proliferation and / or activation of B cells are associated with multiple diseases and conditions, including cancer, autoimmune disease, inflammatory disease, allergic disease, and adverse consequences of transplantation. Existing therapies, such as B cell depletion, treat B cell-associated conditions by seeking to eliminate pathologic B cells. Autoimmune diseases are driven by aberrant response of the host immune system to its own antigens (autoantigens). T regulatory cells engineered to recognize autoantigens may be harnessed to prevent autoimmune disease or ameliorate the symptoms thereof. SUMMARY Some aspects relate to engineered T regulatory cells (Tregs) having, in the forkhead box protein 3 (FOXP3) locus, a nucleotide sequence encoding a chimeric antigen receptor (CAR), a nucleotide sequence encoding a first chemically-inducible signaling complex component (CISC), and a first coding exon (exon 2) of the FOXP3 gene, all under the control of a heterologous promoter; and, in a second locus (e.g., TRAC), a nucleotide sequence encoding a second CISC component (CISC). Such engineered Tregs have robust, sustainable expression of both FOXP3, imparting a stable Treg phenotype, and a CAR providing antigen- specificity, and are therefore of high therapeutic potential for treatment and improvement of autoimmune, allergic, and / or inflammatory diseases. Such engineered cells, populations thereof, and methods, systems, and kits for producing engineered cells and populations thereof are based, at least in part, on the recognition that expression of a CAR in a manner unlinked from FOXP3 expression may yield contaminating CAR+ T effector cells. Insertion of a donor template encoding the CAR into the FOXP3 locus, such that a promoter controls expression of both the CAR and FOXP3, reduces both (i) the probability that a given cell will express the CAR without also expressing FOXP3, thereby exhibiting a CAR+ T effector phenotype, and (ii) the prevalence of contaminating CAR+ T effector cells in a population of engineered cells. Such linkage of CAR and FOXP3 expression thus improves the purity of engineered cell populations and production efficiency of CAR+Tregs. In addition, inclusion on the same donor template of a sequence encoding a soluble FRB domain, separated from mTOR, such that cells expressing the CAR and FOXP3 also express the soluble FRB domain, allows rapamycin-mediated negative selection against cells that do not express the CAR, FOXP3, and soluble FRB domain. Moreover, where cells are edited to express two or more additional components that allow inducible proliferation of edited cells (e.g., CISC components), coding sequences for the additional components may be separated across multiple donor templates due to limitations on donor template size (e.g., AAV packaging limits). In such instances, linking expression of a coding sequence for at least one of the additional components to expression of the CAR and FOXP3 limits the potential for induced proliferation (e.g., by provision of a ligand for chemically induced dimerization) to select for cells that do not express both the CAR and FOXP3. Expression of a first CISC component, CAR, and first coding exon of FOXP3 from the FOXP3 locus, combined with expression of another CISC component in a different locus, therefore allows positive selection for CAR+Tregs (expressing both CAR and FOXP3) by chemically induced dimerization and e.g., IL-2R signal transduction. Such linkage of CAR and FOXP3 expression to produce CAR-expressing Tregs, optionally with positive and / or negative selection through chemically induced IL-2R signaling or rapamycin exposure, respectively, allows production of CAR-Treg populations with improved purity for therapeutic use. Also provided are methods of identifying CARs that are suitable for expression by an engineered regulatory T cell (Treg), by screening a library (plurality) of CARs for features including: (i) antigen-independent CD137 expression by CD4+ T cells, and (ii) antigen- dependent induction of CD69 and CD137 expression by Tregs. Accordingly, some aspects relate to an engineered cell comprising: (i) in a FOXP3 gene locus of a genomic nucleic acid, a first heterologous promoter, wherein the first heterologous promoter is operably linked to: (a) a nucleotide sequence encoding a chimeric antigen receptor (CAR); and (b) a nucleotide sequence encoding a first component of a chemically inducible signaling complex (CISC); and (c) a first coding exon of the FOXP3 gene, wherein the nucleotide sequences of (i)(a) and (i)(b) are 5′ to the first coding exon and in frame with the start codon of the FOXP3 gene; and (ii) in a second gene locus of a genomic nucleic acid, a second heterologous promoter, wherein the second heterologous promoter is operably linked to: (a) a nucleotide sequence encoding a second component of the CISC, wherein each of the first and second CISC components comprises an extracellular binding domain, a transmembrane domain, and a cytoplasmic domain, wherein one CISC component comprises an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain and an IL-2Rβ cytoplasmic domain or portion thereof, and wherein the other CISC component comprises an interleukin-2 receptor gamma IL-2Rγ transmembrane domain and an IL-2Rγ cytoplasmic domain or portion thereof, wherein the first and second extracellular binding domains dimerize in the presence of a ligand to transduce an IL-2 signal in the engineered cell. In some embodiments, the second gene locus is a TRAC gene locus. In some embodiments: the nucleotide sequences of (i)(a) and (i)(b) are separated by a first 2A motif-encoding nucleotide sequence, and the nucleotide sequence of either (i)(a) or (i)(b) is separated from the first coding exon of (i)(c) by a second 2A motif-encoding nucleotide sequence; and the second heterologous promoter is operably linked to (ii)(b) a nucleotide sequence encoding a soluble FRB domain, wherein the nucleotide sequences of (ii)(a) and (ii)(b) are separated by a third 2A motif-encoding nucleotide sequence. In some embodiments: the first heterologous promoter is operably linked to (i)(d) a nucleotide sequence encoding a soluble FRB domain, wherein the nucleotide sequence of (i)(d) is 5′ to the first coding exon, wherein: (1) the nucleotide sequence of (i)(a), (2) the nucleotide sequence of (i)(b), (3) the nucleotide sequence of (i)(d), and (4) the first coding exon are arranged in frame and separated by a combination of a first 2A motif-encoding nucleotide sequence, a second 2A motif-encoding nucleotide sequence, and a third 2A motif- encoding nucleotide sequence. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differ from each other by at least 1 insertion, deletion, or substitution. In some embodiments, the engineered cell is: (i) a hematopoietic stem cell or stem cell precursor; (ii) a CD3+ cell; and / or (iii) a CD4+ cell or a CD8+ cell. In some embodiments, the engineered cell has a regulatory T (Treg) cell phenotype. Some aspects relate to a cell population comprising a plurality of the engineered cell. In some embodiments, fewer than 10% of cells of the cell population are CAR+FOXP3–. In some embodiments, at least 80% of cells of the cell population are CAR+FOXP3+ and express the first and second CISC components. Some aspects relate to a method of making an engineered cell, the method comprising contacting a cell with: (i) a first donor template comprising: (a) a first 5′ homology arm having homology to a first nucleotide sequence within a FOXP3 gene locus of the cell genome; (b) a first 3′ homology arm having homology to a second nucleotide sequence within the FOXP3 gene locus; (c) a first heterologous promoter operably linked to: (d) a nucleotide sequence encoding a chimeric antigen receptor (CAR); and (e) a nucleotide sequence encoding a first component of a chemically inducible signaling complex (CISC), wherein the first heterologous promoter is inserted into the FOXP3 gene locus such that it is operably linked to a first coding exon of the FOXP3 gene and the nucleotide sequences of (i)(d) and (i)(e) are 5′ to the first coding exon and in frame with the start codon of the FOXP3 gene, wherein the first heterologous promoter of (i)(c), nucleotide sequence of (i)(d), and nucleotide sequence of (i)(e) are between the first 5′ homology arm and the first 3′ homology arm; and (ii) a second donor template comprising: (a) a second 5′ homology arm having homology to a first nucleotide sequence within a second gene locus of the cell genome; (b) a second 3′ homology arm having homology to a second nucleotide sequence within the second gene locus; (c) a second heterologous promoter operably linked to: (d) a nucleotide sequence encoding a second component of the CISC, wherein each of the first and second CISC components comprises an extracellular binding domain, a transmembrane domain, and a cytoplasmic domain, wherein one CISC component comprises an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain and an IL-2Rβ cytoplasmic domain or portion thereof, and wherein the other CISC component comprises an interleukin-2 receptor gamma IL-2Rγ transmembrane domain and an IL-2Rγ cytoplasmic domain or portion thereof, wherein the first and second extracellular binding domains dimerize in the presence of a ligand to transduce an IL-2 signal in the engineered cell. In some embodiments, the second gene locus is a TRAC gene locus. In some embodiments: the nucleotide sequences of (i)(d) and (i)(e) are separated by a first 2A motif-encoding nucleotide sequence, wherein the nucleotide sequence of either (i)(d) or (i)(e) is separated from the first coding exon by a second 2A motif-encoding nucleotide sequence; and the second heterologous promoter is operably linked to (ii)(e) a nucleotide sequence encoding a soluble FRB domain, wherein the nucleotide sequences of (ii)(d) and (ii)(e) are separated by a third 2A motif-encoding nucleotide sequence. In some embodiments: the first heterologous promoter is operably linked to (i)(f) a nucleotide sequence encoding a soluble FRB domain, wherein the nucleotide sequence of (i)(f) is inserted 5′ to the first coding exon, wherein after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f): (1) the nucleotide sequence of (i)(d), (2) the nucleotide sequence of (i)(e), (3) the nucleotide sequence of (i)(f), and (4) the first coding exon are arranged in frame and separated by a combination of a first 2A motif-encoding nucleotide sequence, a second 2A motif-encoding nucleotide sequence, and a third 2A motif-encoding nucleotide sequence. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other by at least 1 insertion, deletion, or substitution. In some embodiments, the cell is: (i) a hematopoietic stem cell or stem cell precursor; (ii) a CD3+ cell; and / or (iii) a CD4+ cell or a CD8+ cell In some embodiments, the engineered cell has a regulatory T (Treg) cell phenotype. In some embodiments, the first donor template and the second donor template are comprised in separate viral vectors. In some embodiments, the viral vectors are adeno-associated viral (AAV) vectors. In some embodiments, each of the AAV vectors is an AAV5 or AAV6 vector. In some embodiments, the method comprises contacting the cell with: (i) a first DNA endonuclease capable of cleaving at the FOXP3 gene locus or a nucleic acid encoding the first DNA endonuclease; and (ii) a second DNA endonuclease capable of cleaving at the second gene locus or a nucleic acid encoding the second DNA endonuclease. In some embodiments, the method comprises contacting the cell with: (i) an RNA-guided DNA endonuclease or a nucleic acid encoding the RNA- guided DNA endonuclease; (ii) a first guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the FOXP3 gene locus, or a nucleic acid encoding the first gRNA; and (iii) a second guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the second gene locus, or a nucleic acid encoding the second gRNA. Some aspects relate to a method of producing a population of genetically modified regulatory T (Treg) cells expressing a chimeric antigen receptor (CAR), the method comprising contacting a cell population with a plurality of the first donor template and a plurality of the second donor template, thereby producing a cell population comprising a plurality of the engineered cell. In some embodiments, the method comprises contacting the cell population with: (i) a plurality of an RNA-guided DNA endonuclease, or of a nucleic acid encoding the RNA-guided DNA endonuclease; (ii) a plurality of a first guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the FOXP3 gene locus, or of a nucleic acid encoding the first gRNA; and (iii) a plurality of a second guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the second gene locus, or of a nucleic acid encoding the second gRNA. In some embodiments, the method comprises contacting the cell population with: (i) a plurality of a first DNA endonuclease capable of cleaving at the FOXP3 gene locus, or of a nucleic acid encoding the first DNA endonuclease; and (ii) a plurality of a second DNA endonuclease capable of cleaving at the second gene locus, or of a nucleic acid encoding the second DNA endonuclease. In some embodiments, fewer than 10% of cells of the cell population are CAR+FOXP3–. In some embodiments, the extracellular binding domain of one CISC component comprises an FK506-binding protein 12 (FKBP) domain and the extracellular binding domain of the other CISC component comprises an FKBP-rapamycin-binding (FRB) domain, wherein the ligand is rapamycin, and wherein the method comprises contacting the cell population with rapamycin. In some embodiments, the method further comprises contacting the cell population with the ligand. Some aspects relate to a method for identifying a chimeric antigen receptor (CAR) suitable for expression by an engineered regulatory T cell (Treg), comprising: (a) for a plurality of chimeric antigen receptors (CARs) specific to the same target antigen, expressing each CAR of the plurality in a unique population of CD4+ T cells of a plurality of CD4+ T cell populations in the absence of the target antigen, wherein each CAR of the plurality comprises the same intracellular domain, the same transmembrane domain, the same hinge, and unique antigen-binding domain that binds to the target antigen, each unique antigen-binding domain having a different amino acid sequence of each other antigen-binding domain of other CARs of the plurality; (b) measuring the frequency of CD137 surface expression in each CD4+ T cell population; (c) identifying a first subset of the plurality of CARs, the first subset consisting of CARs of the plurality that, when expressed by a CD4+ T cell population in the absence of the target antigen, do not induce more than 10% of CD4+ T cells of the population to express CD137. In some embodiments, the method further comprises manufacturing a Treg expressing a CAR of the first subset. In some embodiments, the method further comprises: (d) expressing each CAR of the first subset in a different population of Tregs of a plurality of Treg populations in the presence of the target antigen; (e) measuring the frequency of CD137, CD69, and CAR surface expression in each Treg population; and (f) identifying a second subset of CARs of the plurality, the second subset consisting of CARs of the first subset that, when expressed by a Treg population in the presence of the target antigen, induce at least 40% of CAR+ Tregs of the population to express CD69 and CD137 on their surfaces. In some embodiments, the method further comprises manufacturing a Treg expressing a CAR of the second subset. In some embodiments, each Treg population comprises CD4+ T cells, each CD4+ T cell of the Treg population comprising, in a FOXP3 locus of the cell genome, a heterologous promoter inserted downstream from a TSDR, such that the heterologous promoter is operably linked to the first coding exon of the FOXP3 locus. In some embodiments, the Treg population comprises CD4+ T cells, each CD4+ T cell of the Treg population comprising, in a nucleic acid of the cell genome, a heterologous promoter operably linked to a nucleotide sequence encoding FOXP3. In some embodiments, each Treg population is a population of sorted Tregs. In some embodiments, manufacturing a Treg expressing the CAR of the first and / or second comprises a method as described in this section. Some aspects relate to engineered T regulatory cells (Tregs) comprising a chimeric antigen receptor (CAR) comprising a CD19-binding domain. Such engineered Tregs are useful for the treatment of diseases mediated by CD19-expressing cells, such as B cells. B cells can drive pathology in a number of diseases, such as multiple sclerosis (MS) and systemic lupus erythematosus (SLE), by producing autoantibodies, activating T cells via antigen presentation, and producing inflammatory cytokines. Treatments using T effector cells expressing anti-CD19 CARs, which have achieved success as cancer therapies, result in adverse side effects when used for diseases like MS or SLE, such as cytokine release syndrome and / or neurotoxicity. It was discovered, quite surprisingly, that the balance of cytotoxicity toward target cells and non-cytotoxic suppression (e.g., inhibition of target cell proliferation and / or activation) may be modulated by altering the hinge and / or transmembrane domain of a CAR. As described in the Examples, Tregs expressing an anti-CD19 CAR having a CD28 hinge and CD28 transmembrane domain were more cytotoxic towards CD19+ target cells, while Tregs expressing a CAR that differed only by containing an IgG4 hinge and CD8α transmembrane domain were more suppressive and less cytotoxic. Without wishing to be bound by a particular theory, it is posited that interaction between a target cell surface marker and a hinge or transmembrane domain may (i) stabilize the CAR’s interaction with a target antigen on the target cell and consequently promote heightened signal transduction and activation in the CAR-expressing cell. Additionally, or alternatively, target cell surface markers may bind to costimulatory receptors on an engineered cell (e.g., Treg), with these costimulatory receptors on the engineered cell exhibiting differential interactions with CARs having different hinges and / or transmembrane domains. See, e.g., Muller et al., Front Immunol.2021.12:639818. Conversely, limiting such interactions via a CAR hinge and / or transmembrane domain may reduce the extent of signal transduction in the CAR-expressing cell, biasing it towards a distinct phenotype. Modulation of this balance may be useful, for example, to control the extent of cytotoxicity and / or suppression as desired in a particular clinical context. These and other benefits of different hinge and transmembrane domains are described, for example, in Example 2. Accordingly, some aspects relate to an engineered T regulatory cell (Treg), further comprising a chimeric antigen receptor (CAR) comprising a CD19-binding domain. In some embodiments, the CD19-binding domain comprises: (i) a heavy chain variable domain (VH) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 126; and (ii) a light chain variable domain (VL) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 125. In some embodiments, the CD19-binding domain comprises: (i) a VH comprising an amino acid sequence of SEQ ID NO: 126; and (ii) a VL comprising an amino acid sequence of SEQ ID NO: 125. In some embodiments, the CD19-binding domain comprises an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 124. In some embodiments, the CD19-binding domain comprises an amino acid sequence of SEQ ID NO: 124. In some embodiments, the CD19-binding domain comprises: (i) a heavy chain variable domain (VH) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 35, SEQ ID NO: 44, SEQ ID NO: 53, SEQ ID NO: 62, SEQ ID NO: 75, SEQ ID NO: 86, SEQ ID NO: 96, SEQ ID NO: 106, SEQ ID NO: 116, 148, or 158; and (ii) a light chain variable domain (VL) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 34, SEQ ID NO: 43, SEQ ID NO: 52, SEQ ID NO: 61, SEQ ID NO: 74, SEQ ID NO: 85, SEQ ID NO: 95, SEQ ID NO: 105, SEQ ID NO: 115, 147, or 157. In some embodiments, the CD19-binding domain comprises: (i) a VH comprising an amino acid sequence of SEQ ID NO: 35, SEQ ID NO: 44, SEQ ID NO: 53, SEQ ID NO: 62, SEQ ID NO: 75, SEQ ID NO: 86, SEQ ID NO: 96, SEQ ID NO: 106, SEQ ID NO: 116, 148, or 158; and (ii) a VL comprising an amino acid sequence of SEQ ID NO: 34, SEQ ID NO: 43, SEQ ID NO: 52, SEQ ID NO: 61, SEQ ID NO: 74, SEQ ID NO: 85, SEQ ID NO: 95, SEQ ID NO: 105, SEQ ID NO: 115, 147, or 157. In some embodiments, the CD19-binding domain is a Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’)2 fragment, an IgG, a camelid heavy chain antibody, a single domain VHH, or a bivalent VHH. In some embodiments, the CAR comprises a hinge, a transmembrane domain, a first costimulatory domain, a second costimulatory domain, and an intracellular signaling domain. In some embodiments, the hinge is an IgG4 hinge or a CD8 hinge. In some embodiments, the IgG4 hinge comprises an amino acid sequence that differs by no more than two amino acids from an amino acid sequence of ESKYGPPCPSCP (SEQ ID NO: 137). In some embodiments, the IgG4 hinge comprises an amino acid sequence of ESKYGPPCPSCP (SEQ ID NO: 137). In some embodiments, the IgG4 hinge comprises an amino acid sequence that differs by no more than two amino acids from an amino acid sequence of ESKYGPPCPSCPA (SEQ ID NO: 145). In some embodiments, the IgG4 hinge comprises an amino acid sequence of ESKYGPPCPSCPA (SEQ ID NO: 145). In some embodiments, the hinge is a CD8 hinge. In some embodiments, the transmembrane domain is a transmembrane domain derived from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, the transmembrane domain is a CD28 transmembrane domain. In some embodiments, the first costimulatory domain is derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, or CD137 (4-1BB), or a Toll-like receptor (TLR). In some embodiments, the first costimulatory domain is a CD28 costimulatory domain. In some embodiments, the second costimulatory domain is derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, or CD137 (4-1BB), or a TLR. In some embodiments, the second costimulatory domain is a CD3ζ costimulatory domain. In some embodiments, the intracellular signaling domain is derived from OX40, CD2, CD3ζ, CD3γ, CD3δ, CD3ε, FCεRI, CD7, CD27, CD28, CD30, CD40, CD79a, CD79b, CD137 (4-1BB), ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, or a TLR. In some embodiments, the intracellular signaling domain is a CD3ζ intracellular signaling domain. In some embodiments, the CAR comprises the CD19-binding domain, an IgG4 hinge, a transmembrane domain, a CD28 costimulatory domain, and a CD3ζ intracellular signaling domain. In some embodiments, the transmembrane domain is selected from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, the transmembrane domain is a CD8α transmembrane domain. In some embodiments, the CAR comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 70. In some embodiments, the CAR comprises the CD19-binding domain, a CD28 hinge, a transmembrane domain, a CD28 costimulatory domain, and a CD3ζ intracellular signaling domain. In some embodiments, the transmembrane domain is selected from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, the transmembrane domain is a CD28 transmembrane domain. In some embodiments, the CAR comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO: 69. In some embodiments, the CAR further comprises a second costimulatory domain derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, or CD137 (4- 1BB), or a TLR, wherein the second costimulatory domain is either: (i) between the transmembrane domain and the CD28 costimulatory domain; or (ii) between the CD28 costimulatory domain and the CD3ζ intracellular signaling domain. In some embodiments, the engineered Treg further comprises: (i) a nucleic acid encoding a first component of a chemically induced signaling complex (CISC), the first CISC component comprising: (a) a first extracellular domain comprising an FK506-binding protein (FKBP) domain that binds rapamycin; (b) a first transmembrane domain; and (c) a first cytoplasmic domain comprising an intracellular signaling domain of a first cytokine receptor; and (ii) a nucleic acid encoding a second CISC component, the second CISC component comprising: (a) a second extracellular domain comprising an FKBP-rapamycin- binding (FRB) domain; (b) a second transmembrane domain; and (c) a second cytoplasmic domain comprising an intracellular signaling domain of a second cytokine receptor, wherein the first and second CISC components dimerize in the presence of rapamycin. In some embodiments, the first CISC component comprises, in N-to-C-terminal order, an FKBP domain, an interleukin-2 receptor gamma (IL-2Rγ) transmembrane domain, and an IL-2Rγ cytoplasmic domain, and wherein the second CISC component comprises, in N-to-C-terminal order, an FRB domain, an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain, and an IL-2Rβ cytoplasmic domain. In some embodiments, the engineered Treg comprises a nucleic acid encoding a soluble FRB domain. In some embodiments, the soluble FRB domain comprises an amino acid sequence having at least 90% sequence identity to an amino acid sequence of SEQ ID NO: 13. In some embodiments, the soluble FRB domain comprises an amino acid sequence of SEQ ID NO: 13. In some embodiments, the engineered Treg comprises a heterologous promoter located in a nucleic acid of the cell genome: (a) downstream from a regulatory T cell (Treg)-specific demethylated region (TSDR); and (b) upstream from the first coding exon of an endogenous FOXP3 gene. In some embodiments, the heterologous promoter located downstream from the TSDR is an MND promoter. In some embodiments, the engineered Treg is FOXP3-positive, CD3-positive, CD4- positive, and CD25-positive. In some embodiments, the engineered Treg is a sorted Treg. Some aspects relate to a method comprising administering the engineered Treg to a subject in need thereof. Some aspects relate to a method comprising administering an engineered Treg to a subject in need thereof, wherein the engineered Treg comprises a CAR comprising a CD19- binding domain. In some embodiments, 1 x 107to 1 x 1010engineered Tregs are administered. In some embodiments, 5 x 108to 2 x 109engineered Tregs are administered. In some embodiments, 8 x 108to 1.2 x 109engineered Tregs are administered. In some embodiments, 1 x 109engineered Tregs are administered. In some embodiments, wherein the engineered Tregs are administered intravenously. In some embodiments, the method further comprises administering rapamycin to the subject. In some embodiments, the engineered Tregs are administered in combination with rapamycin. In some embodiments, the method further comprises administering cyclophosphamide and / or fludarabine to the subject. In some embodiments, the engineered Tregs are administered in combination with cyclophosphamide and / or fludarabine. In some embodiments, administering the engineered Treg induces B cell suppression in the subject. In some embodiments, the B cell suppression comprises: (i) decreasing antibody production by plasmablasts and / or plasma cells; (ii) decreasing cytokine production by B cells; (iii) decreasing antigen-presenting capacity of B cells; (iv) reduced B cell differentiation; and / or (v) decreased B cell proliferation. In some embodiments, the B cells are CD19-positive B cells. In some embodiments, administering the engineered Treg does not induce B cell aplasia. In some embodiments, administering the engineered Treg reduces the abundance of circulating B cells that are activated, proliferating, and / or autoreactive in the subject. In some embodiments, administering the engineered Treg increases the abundance of naïve B cells in the subject. In some embodiments, administering the engineered Treg reduces the serum concentration of one or more antibodies that preferentially bind double-stranded DNA or Sm / RNP in the subject. In some embodiments, the subject is a human. In some embodiments, the subject has or is at risk of developing an autoimmune disease and / or an inflammatory disorder. In some embodiments, the autoimmune disease is systemic lupus erythematosus (SLE) or Sjögren’s disease (SjD). Some aspects relate to a method of manufacturing an engineered Treg, the method comprising introducing into a Treg a nucleic acid comprising a promoter operably linked to one or more of: (i) a nucleotide sequence encoding FOXP3 or a functional fragment thereof; (ii)(a) a nucleotide sequence encoding a first component of a chemically inducible signaling complex (CISC), the first component comprising: (1) a first extracellular domain comprising an FK506-binding protein (FKBP) domain that binds rapamycin, (2) a first transmembrane domain, and (3) a first cytoplasmic domain comprising an intracellular signaling domain of a first cytokine receptor; (ii)(b) a nucleotide sequence encoding a second component of the CISC, the second component comprising: (1) a second extracellular domain comprising an FKBP-rapamycin-binding (FRB) domain, (2) a second transmembrane domain, and (3) a second cytoplasmic domain comprising an intracellular signaling domain of a cytokine receptor; and / or (iii) a nucleotide sequence encoding a CAR comprising a CD19-binding domain. Some aspects relate to a chimeric antigen receptor (CAR) polypeptide comprising an amino acid sequence having at least 90% identity to an amino acid sequence of SEQ ID NO: 134. Some aspects relate to a CAR polypeptide comprising an amino acid sequence of SEQ ID NO: 134. Some aspects relate to a cell comprising the CAR polypeptide. Some aspects relate to a method of manufacturing an engineered Treg, the method comprising contacting a T cell with a lipid delivery vehicle comprising a T cell-targeting moiety and one or more nucleic acids that collectively encode: (i) a nucleic acid encoding a chimeric antigen receptor (CAR) polypeptide comprising a CD19-binding domain; and (ii) FOXP3 or a functional fragment thereof. In some embodiments, the one or more nucleic acids are messenger ribonucleic acids (mRNAs). In some embodiments, the lipid delivery vehicle is a lipid nanoparticle (LNP). In some embodiments, the T cell is a CD4+ T cell and the T cell-targeting moiety is a CD4-targeting moiety. In some embodiments, the CD4- targeting moiety is an anti-CD4 antibody or antigen-binding fragment thereof. In some embodiments, the anti-CD4 antibody is selected from the group consisting of zanolimumab, keliximab, clenoliximab, tregalizumab, and ibalizumab. BRIEF DESCRIPTION OF THE DRAWINGS FIGs.1A–1H show secretion of cytokines and chemokines by engineered cells expressing different CARs, including IFN-γ (FIG.1A), IL-6 (FIG.1B), IL-17A (FIG.1C), TNF-α (FIG.1D), IL-10 (FIG.1E), IL-2 (FIG.1F), IL-8 (FIG.1G), and CXCL10 (FIG.1H). FIG.2 shows secretion of IFN-γ by CAR-EngTregs engineered by dual-AAV insertion (FOXP3 and TRAC loci) compared to CAR-Teffs engineered by LV insertion. FIG.3 shows an experimental design for assaying cytotoxicity of anti-CD19 CAR-T cells against B cells or cell lines. FIG.4A shows engineered cell activation (as measured by CD69+ cells) after 24 hours of co-culture with primary B cells, NALM-6 cells, or negative control K562 cells, which do not express CD19. FIG.4B shows CAR expression (as measured by Whitlow+ cells) after 24 hours of co-culture with primary B cells, NALM-6 cells, or negative control K562 cells, which do not express CD19. FIGs.5A–5D show cytotoxicity following co-culture as described in FIG.3. FIG.5A shows representative staining at 24 hours. FIG.5C shows representative staining at 48 hours. FIGs.5B and 5D summarize target cell death (normalized to target cells alone, without co- culture of engineered cells) at 24 hours (FIG.5B) and 48 hours (FIG.5D). FIG.6 shows target cell death occurring in the presence of engineered cells made using a different donor compared to the cells assayed in FIGs.5A–5D. FIG.7 shows an experimental design for measuring T effector (Teff or Tresp) cell suppression by CAR-EngTregs (Tsupp). FIG.8A shows suppression of Teff cells by dual-AAV-edited anti-CD19 CAR- EngTregs. FIG.8B shows proliferation by Teff cells co-cultured with anti-CD19 CAR- EngTregs. FIG.8C shows activation of CAR-EngTregs (as measured by CD69 expression) 24 hours after activation with B cells. FIG.8D shows suppression of Teff cell proliferation by anti-CD19 CAR-EngTregs made with cells from two different donors. FIG.8E shows suppression by anti-CD19 CAR-EngTregs, observed by assaying cells from three different donors. FIG.8F shows suppression by polyclonal EngTregs with and without bead-mediated activation, as compared to antigen-responsive suppression by anti-CD19 CAR-EngTregs. FIG.9 shows an experimental design for measuring B cell suppression. FIGs.10A and 10B show proliferation by Tsupp and B cells, in assays where Tsupp cells were prepared using cells from two different cell source donors. FIG.10C shows CD38 and CD20 expression among B cells. FIGs.10D and 10E show % suppression. FIG.11A shows a study design testing the feasibility of dual AAV engineering of CD19 CAR-EngTregs, to evaluate expansion, viability, and enrichment of the dual-edited population at day 7, 10, 14, and 18. FIG.11B shows a representative schematic of EngTregs with AAVs inserted into the FOXP3 locus and TRAC locus, and architectures of two CARs having the same scFv and intracellular signaling domains, but one having a CD28 hinge and CD28 transmembrane domain and another having an IgG4 hinge and CD8α transmembrane domain. FIG.12 demonstrates that both CAR-EngTreg types show CAR+ and FOXP3+ enrichment at the end of the process of FIG.11A. FIGs.13A-13C show comparable viability (FIG.13C), expansion (FIG.13A), and enrichment (FIG.13B) of CAR-EngTreg populations to Dual AAV control EngTregs. FIG.14A shows a study design to phenotype CAR-EngTreg populations at end of manufacturing process. CAR-EngTregs generated were stained at the end of process, either day 18 or 21 (without resting), for FOXP3, CD25, CD127, CTLA-4, and ICOS to confirm Treg phenotype. FIG.14B shows strong enrichment of Treg population, and higher MFIs for Treg phenotype markers. *Mock cells retained activation markers since cells were not rested. FIG.15A shows a study design to phenotype CAR-EngTreg populations after cryopreservation, thawing, and resting for three days. FIG.15B shows strong enrichment of Treg population, and higher MFIs for Treg phenotype markers. Plots show live CD4+ cells. FIG.16A shows a study design to assess expression of activation markers upon antigen-specific stimulation of CAR19-EngTregs and an illustration of a CAR19-EngTreg (CD19 CAR EngTreg). CAR19-EngTregs generated were rested for three days in low IL-2 (5 ng / ml) and stimulated with allogeneic B cells prior to phenotyping. Cells were stained at 24 hours for CD69, TNFRII and GARP as a proxy for TGF-β to confirm antigen driven activation. FIGs.16B–17E show phenotypes of cells treated and assayed as in FIG.16A. FIGs.16B–16C show representative data from one donor, with cells gated on live, CD4+, and FOXP3+ cells prior to analysis. FIGs.17A–17E show data from three donors. Results show increases in activation markers and upregulation of Treg markers upon antigen-specific stimulation. FIG.16A shows upregulation of LAP and GARP. FIG.16B shows upregulation in CD19, 4-1BB, and TNFRII. FIG.17A shows antigen-driven activation as measured by CD69. FIGs.17B–17D show antigen-driven upregulation of Treg-associated markers CTLA- 4, ICOS, and TNFRII. FIG.17E shows antigen-driven Treg cytokine production as measured by GARP. FIG.18A shows a study design to assess cytokine and cytotoxic enzyme secretion of CAR19-EngTregs upon antigen-specific stimulation and an illustration of a CAR19-EngTreg (CD19 CAR EngTreg). CAR19-EngTregs were thawed, rested for 3 days in low-dose IL-2 (5 ng / ml), and the stimulated with B cells for one (all cytokines) or four days (perforin & granzyme B) and supernatants analyzed by cytometric bead array (CBA). FIGs.18B–21B show cytokine cytotoxic enzyme production from cells treated and assayed as in FIG.18A. FIGs.18B–18E and 20 show data from three donors. FIGs.19B and 21B show data from two donors, with CAR-Teff gated on CD4+Whitlow+ and CAR19- EngTregs gated on CD4+FOXP3+Whitlow+. FIGs.18B–19B show that CAR19-EngTregs secreted lower inflammatory cytokines than CAR19-Teffs upon target engagement. FIGs.20–21B shows that CAR19-EngTregs secrete perforin and granzyme B after antigen-specific stimulation, but production is lower than CAR19-Teffs. FIG.22A shows a study design to assess the ability of CAR19-EngTregs to kill CD19-expressing cells and an illustration of a CAR19-EngTreg (CD19 CAR EngTreg). CAR19-EngTregs were rested for three days in low IL-2 (5 ng / ml), co-cultured with CD19+Nalm-6 or CD19– K562 prior to killing assessment. Cells were stained at 48 hours with Annexin V and live / dead stain to determine cell killing. In a separate assay, CAR19- EngTregs were co-cultured with autologous, activated and cell trace violet (CTV)-labeled B cells prior to suppression assessment. Four days post co-culture, CTV dilution was assessed to determine suppression of B cell proliferation. Results are shown in FIGs.22B–25C. FIG.22B shows representative data from one donor at 48 hours, gated on CTV+ cells. Staurosporine was a positive control for cell killing. CAR19-EngTregs exhibited varying levels of cytotoxicity towards CD19+ cell lines. FIG.23 shows data from 3 donors, at 48 hours, with a 1:1 Effector:Target ratio. CAR19-EngTregs exhibited antigen-specific killing in this assay. FIG.24 shows data from 2 donors, at 48 hours, with either a 2.5:1 or 1:1 Effector:Target ratio. As in FIG.23, CAR19-EngTregs exhibited antigen-specific killing in this assay, at both Effector:Target ratios. FIG.25A shows killing of activated B cells by CAR19-EngTregs expressing the IgG4hinge.CD8αTM CAR compared to controls and CAR19-EngTregs expressing the CD28hinge.CD28TM CAR. FIG.25B shows activation phenotypes, as measured by CD69, of CAR19-EngTregs cultured with activated or un-activated B cells. FIG.25C shows CD19 and CD38 expression on activated and un-activated B cells. B cell activation was achieved using CpG DNA, rCD40L, and IL-21. FIG.26A shows a study design to assess the ability of CAR19-EngTregs to suppress polyclonal Teffector (Tresp) when activated with B cells. CAR19-EngTregs were rested for three days in low IL-2 (5 ng / ml), labeled with CFSE and preactivated with CTV labeled B- cells for one day. Tresp were preactivated with CD3 / CD28 beads for one day and labeled with eFluor670. Four days post co-culture, eFluor670 dilution was assessed to determine suppression of Tresp proliferation. FIGs.26B–26C show polyclonal T responder (Tresp) cell suppression by cells treated and assayed as in FIG.26A. Both CAR19-EngTreg populations displayed antigen-specific Tresp suppression at varying levels. FIG.26B shows representative data from one donor. FIG.26C shows data from 3 donors. FIG.27 shows a manufacturing process to generate CAR19-EngTregs for adoptive transfer into NSG mice. FIG.28A shows manufacturing an enrichment of CAR19-EngTregs as described in FIG.27. FIG.28B shows cell phenotypes at the time of freezing, indicating integration of transgenes and expression of the encoded CAR. FIG.29A shows cell phenotypes after thawing, without stimulation. FIG.29B shows cell phenotypes after thawing and stimulation with anti-CD3 / CD28 beads. FIG.30A shows a study design to evaluate CAR19-EngTregs in a pristane-induced humanized lupus model. FIG.30B shows B cell depletion by CAR19 Teffs, an anti-CD19 monoclonal antibody (mAb) Ineblizumab, and CAR19-EngTregs. FIG.30C shows T cell depletion by only CAR19-EngTregs. FIGs.30D and 30E show percentages of B cells and T cells, respectively, at interim necropsy, 4 weeks after cell dosing. FIGs.31A–31I show pathology at 4 and 8 weeks after cell dosing. FIGs.31A, 31D, and 31G show slides of lungs, liver, spleen, kidney, bone marrow, and skin. FIG.31B shows incidence of pathology signs in lungs. FIG.31C shows incidence of pathology signs in livers. FIG.31E shows incidence of pathology signs in spleens. FIG.31F shows incidence of pathology signs in kidneys. FIG.31H shows incidence of pathology signs in bone marrow of femurs and sternums. FIG.31I shows incidence of pathological signs in skin. FIG.32A shows total IgM in sera at baseline, 4 weeks after cell dosing, and 8 weeks after cell dosing. FIG.32B shows total IgG in sera at 8 weeks after cell dosing. FIG.33A shows pro-inflammatory cytokines in sera at 1 week after cell dosing. FIG. 33B shows production of cytotoxic enzymes in sera at 1 week after cell dosing. FIG.34 shows a study design to evaluate CAR19-EngTregs in a mouse model of systemic lupus erythematosus (SLE). NBSGW mice were humanized with CD34+ cells from a mobilized PBMC donor, pre-dosed with rapamycin (or vehicle control), and infused with CAR19-EngTregs (or CAR19-Teff controls) as adoptive cell transfer (ACT). Mice were administered rapamycin for the duration of the study. Mice in the indicated groups were euthanized at the indicated timepoints for analysis of serum cytokine concentrations, cell populations, and pathology. FIG.35 shows changes in body weight after adoptive cell transfer (ACT) with either autologous CAR19-Teffs or autologous CAR19-EngTregs. There was a significant separation between body weight change observed in the two treatment groups. FIG.36 shows B cell populations in blood at 1 week post-ACT, indicating B cell ablation by CAR19-EngTregs. M5, M10, and M30 indicate individual mice that had low weight and were likely anemic at the start of the study. FIG.37 shows T cell populations in spleens at 1 week post-ACT, indicating expansion of CAR19-EngTregs. FIG.38 show pro-inflammatory cytokine concentrations in sera at 1 week post-ACT, indicating CAR19-EngTregs produced lower levels of pro-inflammatory cytokines than CAR19-Teffs. FIG.39 shows B cell populations (in blood) and T cell populations (in spleens) of mice administered allogeneic CAR19-EngTregs at 2 weeks post-ACT, alongside the data of FIGs.36 and 37 at 1 week post-ACT. FIG.40 shows a study design to evaluate CAR19-EngTregs’ impact on B cells and downstream maturation. FIG.41 shows representative gating measuring B cell proliferation, plasmablast formation, and plasma cell formation. FIG.42 shows summary data at Effector:Target ratios of 1:5 or 1:1. At a higher Effector:Target ratio, CAR19-EngTregs decreased B cell proliferation and plasmablast formation, while plasma cell formation was decreased at both Effector:Target ratios. FIG.43A shows representative gating measuring IgM and IgG production. FIG.43B shows summary data at Effector:Target ratios of 1:5 or 1:1. At both Effector:Target ratios, CAR19-EngTregs prevented both IgM and IgG production from B cells. FIG.44 shows a study design to evaluate CAR19-EngTregs’ impact on plasmablasts and downstream maturation. FIG.45A shows representative gating measuring plasma cell formation and IgG production by plasma cells. FIG.45B shows summary data at an Effector:Target ratio of 1:5, indicating that CAR19-EngTregs prevented both plasma cell formation and IgG production. FIG.46A shows representative gating measuring plasmablast formation and IgG production. FIG.46B shows summary data at Effector:Target ratios of 1:1 and 1:5. CAR19- EngTregs prevented plasmablast formation, IgG production, and IgG secretion at both ratios. FIG.47 shows a study design to evaluate CAR19-EngTregs in a mouse model of systemic lupus erythematosus (SLE). NBSGW mice are humanized with CD34+ cells from a mobilized PBMC donor, pre-dosed with rapamycin (or vehicle control), and infused with CAR19-EngTregs (or CAR19-Teff controls) as adoptive cell transfer (ACT). Mice in rapamycin groups are administered rapamycin for the duration of the study, starting with the day before cell administration. Mice are euthanized at the indicated timepoint for analysis of serum cytokine concentrations, cell populations, and pathology. FIG.48A shows an AAV donor template for insertion into the FOXP3 locus. FIG.48B shows an AAV donor template for insertion into the TRAC locus. FIG.48C shows a timeline for production of engineered cells using AAV donor templates or lentiviral vectors. FIGs.49A–49D show phenotypes of engineered cells made using CD4+ or CD3+ cells from two different donors. FIGs.49A and 49B show cells made from donor 0586, and FIGs.49C–49D show cells made from donor 0523. FIG.49E shows cell growth kinetics of engineered cells made from both donors. FIG.49F shows CD25 and CTLA-4 of mock-edited or dual-edited cells made from both donors. FIGs.50A shows an experimental design for evaluating CAR19-EngTregs and CAR19-T effector cells in an imiquimod-induced mouse model of systemic erythematosus (SLE), as described in Example 10. FIG.50B shows a design process for producing CAR19- EngTregs and CAR19-T effector cells used in Example 10. FIG.51A shows ear thickening over time. FIG.51B shows spleen weights at day 12, as measures of splenomegaly. FIG.52A shows splenic engraftment of administered T cells at day 12. FIG.52B shows splenic B cell abundance (and reduction relative to naïve mice) at day 12. FIGs.53A–53E show engraftment of administered T cells in blood (FIG.53A), kidneys (FIG.53B), bone marrow (FIG.53C), lungs (FIG.53D), and mesenteric lymph nodes (FIG.53E) at day 12. FIGs.54A–54G show results of intracellular cytokine staining (ICS) analysis of splenocytes obtained from mice at day 12. FIGs.54A–54C show IFN-γ production frequency among CD3+ T cells (FIG.54A), CD4+ T cells (FIG.54B), and CD8+ T cells (FIG.54C). FIG.54D shows IL-21 production frequency among CD4+ T cells. FIG.54E shows Tfh cell frequency (top graph) and abundance (bottom graph). FIG.54F shows IL-21 production frequency among Tfh cells. FIG.55 shows results of cytometric bead array (CBA) analysis of cytokines produced by splenocytes obtained from mice at day 12. FIGs.56A–56C show splenic infiltration of neutrophils (FIG.56A), inflammatory monocytes (FIG.56B), and dendritic cells (FIG.56C) at day 12. FIGs.57A–57B demonstrate variability in lentiviral and adeno-associated viral transduction in generating chimeric antigen receptor-positive (CAR+) regulatory T cells (forkhead box protein 3-positive, FOXP3+). FIG.57A shows that 7.3% of transduced cells were FMC63-CAR+ effector T cells (FOXP3–) and 73.6% of transduced cells were CAR+ regulatory T cells (FOXP3+) (Tregs). FIG.57B shows that 19.8% of transduced cells were Hu19-CAR+ effector T cells and 49% are CAR+ Tregs. FIGs.58A–58D relate to improved constructs to limit incidence of contaminating CAR+ T effector cells following transduction. FIG.58A shows a construct comprising an MND promoter, a sequence encoding a CAR of interest, a coding sequence of a portion of a FOXP3 gene, and a sequence encoding a first chemically-inducible signaling complex (CISC) component (top) and a construct comprising a coding sequence of a T cell receptor alpha constant (TRAC) gene, an MND promoter, a sequence encoding a second CISC component, and a sequence encoding a naked FRB domain (bottom). FIG.58B shows the percentage of FOXP3+ versus FOXP3– T cells generated following transduction with a construct targeted to the TRAC locus. FIG.58C shows the percentage of rapamycin-enriched Hu19-CAR+ Tregs generated following transduction with a construct encoding a first CISC component targeted to the TRAC locus and a construct encoding a Hu19-CAR, a portion of a FOXP3 gene, and a second CISC component targeted to the FOXP3 locus (“dual edited” cells). FIG.58D shows increased numbers of Hu19-CAR Tregs compared to a positive control at 18 days and 21 days post-transduction. FIGs.59A–59B relate to decreased CAR+ T effector cells following transduction with a construct encoding a first CISC component targeted to the TRAC locus and a construct encoding a second CISC component, a portion of a FOXP3 gene, a Hu19-CAR targeted to the FOXP3 locus. FIG.59A shows mock-transfected cells and cells transfected with a negative control of ribonucleoprotein (RNP) alone. FIG.59B shows CAR+ Tregs (Whitlow+ FOXP3+) cells following editing with minimal presence of CAR+ T effector cells (Whitlow+ FOXP3–). FIGs.60A–60B demonstrate sustainable CAR and FOXP3 expression in cells transduced as in FIGs.60A–60B and enrichment for dual edited CAR+FOXP3+ cells over 21 days of culture. FIG.60A shows CAR+ Tregs with minimal contamination from CAR+ T effector cells at days 7, 10, 18, and 21 of culture. FIG.60B shows populations of cells that were mock transfected or transfected with positive or negative (RNP) controls at day 21 of culture. FIGs.61A–61B show TCRαβ and FOXP3 expression of cells edited as in FIGs.60A– 60B. FIG.61A shows Tregs (FOXP3+ TCRαβ+) cells at days 7, 10, 18, and 21 of culture. FIG.61B shows populations of cells that were mock transfected or transfected with positive or negative (RNP) controls at day 21 of culture. FIG.62 demonstrates that dual edited cells have comparable enrichment (left), expansion (middle), and survival (right) compared to positive controls. FIG.63 shows that the results of transduction as shown in FIG.59B are reproducible with minimal variability (“Round 2”). FIG.64 shows a multistep screening strategy for high throughput discovery and characterization of CAR-EngTregs expressing CARs with different scFvs. FIGs.65A–65B show results of a phage panning screen which identified a diverse panel of scFvs specific to Target X. FIG.65A shows negative deselection using an irrelevant antigen-overexpressing cell line, and positive selection using a cell line overexpressing Target X, and corresponding enrichment of phage clones over four rounds of panning. FIG.65B shows 180 phage clones selected for further characterization, based on Target X binding, and the hCDR3 diversity within this panel. FIGs.65C–65D show results of parallel immunization that generated antibodies (scFv identification) against Target X and its murine ortholog. FIG. 65C shows an outline of immunization campaigns, and FIG.65D shows sera responses to human Target X (top) or its murine ortholog (bottom). FIG.66A shows assessment and ranking of CAR surface expression, with the majority of CARs demonstrating robust surface expression. FIG.66B shows assessment of antigen-independent activation (tonic signaling), as evidenced by CD137 expression frequency in the absence of antigen, which correlated with the amount of IFN-γ secretion. FIG.66C shows antigen-dependent activation, as evidenced by CD137 and CD69 expression in the presence of antigen. FIG.67 shows rank ordering of CARs with different CAR expression, activity, and biophysical profiles. FIG.68 shows a CAR-EngTreg production process, and comparison of FOXP3, CD25, and CAR expression intensity in mock-edited cells, EngTregs (without CAR), CAR- EngTregs, and CAR-Teffs. FIG.69A shows antigen- and CAR-dependent activation in EngTregs, as evidenced by CD69+CD137+ frequency (top) and LAP+GARP+ frequency (bottom). Squares = irrelevant antigen; Circles = Target X. FIG.69B shows the correlation between CD69+CD137+ frequency and LAP+GARP+ frequency, among CAR+Tregs. FIG.70 shows antigen- and CAR-dependent suppression of polyclonal CD4+ Teff cell proliferation by CAR-EngTregs. CAR-EngTregs (Tsupp) were activated with HEK293 cells expressing irrelevant antigen or Target X. Simultaneously Teffector (Tresp) were activated via anti-CD3 / CD28 beads. Beads were washed out 16 hours post activation. Tresp were then co-cultured with Tsupp for the subsequent 72 hours, and proliferation was measured. Suppression was calculated as follows: % suppression = ((ab) / a)x100, where “a” is the percentage of Tresp proliferation in the absence of Tsupp and “b” is the percentage of Tresp proliferation in the presence of Tsupp. % suppression when Tsupp were activated by irrelevant antigen was also subtracted. FIGs.71A-71G relate to mouse CAR19 EngTreg therapy in a murine SLE model. FIG.71A is a schematic illustrating the experimental timeline. FIG.71B shows the result of a flow cytometry assay, demonstrating that CAR19 EngTreg are detectable in the spleen 4.5 months post-adoptive transfer based on expression of CD45.1 donor cell maker (left panel) and co-expression of FOXP3 and LNGFR (right 2 panels). FIG.71C shows reduced renal injury based upon NGAL serum levels (left) and renal histopathology (right) in EngTreg treated mice. FIG.71D shows the reduced lymphoid inflammatory response in spleen based on the number of white pulp foci in EngTreg treated mice. FIG.71E shows reduced dsDNA (left) and Sm / RNP (right) in serum in EngTreg treated mice at the endpoint of the experiment. FIGs.71F and 71G show graphical representations of flow cytometry results showing reduced proportions of: GC B cells and age-associated B cells (FIG.71F) and T follicular helper (Tfh), T peripheral helper cells (Tph) and memory CD4 T cells (FIG.71G) in spleen at the study endpoint. FIG.72A provides exemplary donor templates for production of EngTregs expressing a first CISC component (FKBP-IL2Rγ), soluble FRB domain polypeptide, and anti-CD19 CAR from the FOXP3 locus, and a second CISC component (FRB-IL2Rβ) and marker (LNGFR) from the TRAC locus. FIG.72B provides exemplary spacer sequences of guide RNAs for RNA-directed cleavage of FOXP3 and TRAC loci. FIG.72C shows a production timeline for generation and enrichment of dual-edited (DE) CD19CAR EngTreg from CD4+ T cells (top panel) and schematics for individual HDR events at TRAC and FOXP3 loci (bottom panels). FIG.72D shows representative flow cytometry of DE CD19CAR EngTreg at Day 3 (Pre-Enrichment) and endpoint (Post-Enrichment in rapamycin). FIG.72E shows immunophenotype of DE CD19CAR EngTreg, polyclonal EngTreg, and mock-edited controls. At endpoint, cells were assessed by flow cytometry for Treg markers. FIG.72F shows the % suppression of T effector proliferation at Day 4 in culture following CD3 / CD28 activation and using indicated ratio of DE CD19CAR EngTreg, polyclonal EngTreg, or mock-edited controls. Percentage suppression was calculated as [(%) Teff dividing without Treg – (%) Teff dividing with Treg) / (%) Teff dividing without Treg] x 100. FIG.73 shows an experimental schematic for assessing the impact of CAR19 EngTreg vs. Teff in the MRL / lpr model.8–12-week-old MRL / lpr mice will be pre- conditioned with 100mg / kg cyclophosphamide 24h prior to adoptive cell transfer. Mice will receive either CAR19 EngTreg or CAR19 Teff cells via the retro-orbital sinus (r.o.). A subset of cohorts will also receive IL-2 support as indicated. Biweekly peripheral bleeds will be performed to assess engraftment and impact on circulating B and T cells. Mice will be monitored for 20 weeks or ≥12 months prior to detailed endpoint analysis. FIG.74 shows an experimental schematic for development of and efficacy study in a PBMC-induced humanized Sjögren’s model. DETAILED DESCRIPTION Chimeric antigen receptors (CARs) are synthetic receptors that contain a binding domain specific for a particular protein, as well as additional structural and signaling domains. Cells, especially immune cells, can be engineered to express CARs, thus conferring a particular antigen specificity. Moreover, the structure of the CAR allows cells that require costimulation or antigen presentation on major histocompatibility complex (MHC) molecules, such as T cells, to engage cognate antigen and subsequently become activated in the absence of typically necessary co-stimulatory signals provided by other cells. Thus, expressing a CAR in a cell redirects that cell to recognize and, typically, induces cytotoxicity of cells expressing a specific protein. T cells engineered to constitutively express forkhead box protein 3 (FOXP3), the T regulatory cell (Treg) lineage transcription factor, and CARs against self-antigens have the potential to treat autoimmune disease and / or improve the associated symptoms. Autoimmune diseases are driven by the host’s immune response against self-antigens (autoantigens), and Tregs expressing receptors that recognize the same antigen may be used to suppress the aberrant immune activity. It is thought that the engineered CD-19 CAR+ Tregs described herein may cause an immune system reset in subjects having autoimmune disease(s). As used herein, “immune system reset” refers to a substantial depletion of pathogenic B cells by the anti-CD19 CAR+ Tregs, followed by a return of naïve and non-pathogenic B cell subsets in vivo. It is noted that, in most models of systemic lupus erythematosus (SLE), there are high levels of pathogenic B cells and T cells, as well as high levels of auto-antibodies (anti- double-strand DNA, anti-dsDNA); however, it has been shown that, in a mouse model of SLE, treatment with CAR19 Tregs restores immune cell composition (T cells and B cells) in the spleen and bone marrow and reduces anti-dsDNA antibody secretion (Doglio et al., Nat Commun.2024.15(1):2542). As shown herein, administration of CAR19+ Tregs in mouse models of SLE led to similar restoration (see, e.g., Examples 6 and 7 and FIGs.31-39). In addition, the CAR+ Tregs, in some embodiments, preferentially deplete (e.g., kill) activated B cells (see, e.g., Example 5; FIGs.25A-25C). In contrast to cytotoxic CD8+ T effector cells, Tregs do not necessarily kill cells following antigen recognition, but instead may exert suppressive activity through non-cytotoxic mechanisms including secretion of immunoregulatory cytokines. Accordingly, both killing of target cells by CAR+ Tregs, together with suppression of B cell activity, are expected to contribute to depletion of B cells and restoration of a non-pathogenic B cell compartment. Different approaches for producing engineered CAR+ Tregs (cells expressing FOXP3 and a CAR of interest), can result in a heterogenous T cell population wherein some cells are CAR+ Tregs and some cells are CAR+ T effector cells, which can exacerbate disease states or compromise the efficacy of CAR+ Treg therapy. Accordingly, some aspects relate to engineered cells comprising, in the FOXP3 gene locus, sequences encoding a CAR and a portion of a FOXP3 gene, as well as a first chemically inducible signaling complex (CISC) component, and, in another locus, a second CISC component. The genetic linkage of FOXP3 CAR expression, combined with the dimerization of CISCγ and CISCβ, for example, allows for the enrichment of CAR+ Tregs with minimal contamination by CAR+ T effector cells. Some aspects relate to methods of screening a plurality of CARs to identify a CAR suitable for expression in an engineered Treg. These methods are based, at least in part, on data showing that antigen-independent CD137 expression correlates with IFN-γ secretion, with such antigen-independent secretion of a pro-inflammatory cytokine being undesirable in a Treg that is otherwise useful for alleviating inflammatory responses. Additionally, surface expression of CD69 and CD137 by CAR-Tregs exposed to target antigen correlated with expression of LAP and GARP functional markers. Selection of CARs based on these features therefore allows more efficient and accurate identification of CARs suitable for manufacturing Tregs for therapeutic use. Cells expressing a CAR that binds CD19 (an anti-CD19 CAR), especially T cells, have been successfully used in treatments for cancers driven by malignant B cells. In such therapies, anti-CD19 CARs are typically expressed in T effector cells to direct the T effector cell’s cytotoxic effects against CD19-expressing cells. The use of T effector cells expressing an anti-CD19 CAR has been contemplated in the treatment of autoimmune conditions such as systemic lupus erythematosus (SLE), multiple sclerosis (MS), rheumatoid arthritis (RA), and others. However, use of such cells carries a risk of numerous adverse effects, such as inducing B cell aplasia (marked decrease of B cells), cytokine release syndrome, neurotoxicity, neutropenia (marked decrease in neutrophils), and infection. Accordingly, some aspects relate to engineered T regulatory cells comprising a CAR comprising a CD19-binding domain. Such cells can suppress B cell activity and thus ameliorate or eliminate symptoms associated with B cell-mediated diseases without inducing adverse side effects associated with the use of T effector cells expressing an anti-CD19 CAR. Some aspects relate to an engineered Treg expressing (i) an anti-CD19 CAR, (ii) a first CISC component comprising (a) an extracellular FKBP domain, (b) an IL-2Rγ transmembrane domain, and (c) an IL-2Rγ cytoplasmic domain or portion thereof, and (iii) a second CISC component comprising (a) an extracellular FRB domain, (b) an IL-2Rβ transmembrane domain, and (c) an IL-2Rβ cytoplasmic domain or portion thereof, and having an MND promoter inserted upstream from a first coding exon of a FOXP3 gene in the cell genome. In some embodiments, the engineered cell is an allogenic cell. In some embodiments, the engineered cell is an autologous cell. In some embodiments, the engineered cell is an allogeneic cell that is modified to be hypoimmunogenic. In some embodiments, the second CISC component comprises a truncated IL-2Rβ cytoplasmic domain. In some embodiments, the engineered cell expresses a TCR. In some embodiments, the TRAC locus is unmodified in the engineered cell. In some embodiments, the engineered cell comprises a modified TRAC locus. In some embodiments, the engineered cell expresses an exogenous TCR. Some aspects relate to a viral vector comprising an MND promoter operably linked to nucleotide sequences encoding (i) FOXP3, (ii) a first CISC component comprising (a) an extracellular FKBP domain, (b) an IL-2Rγ transmembrane domain, and (c) an IL-2Rγ cytoplasmic domain or portion thereof, (iii) a second CISC component comprising (a) an extracellular FRB domain, (b) an IL-2Rβ transmembrane domain, and (c) an IL-2Rβ cytoplasmic domain or portion thereof, and (iv) an anti-CD19 CAR. Some aspects relate to methods of administering, to a subject in need thereof, a viral vector comprising an MND promoter operably linked to nucleotide sequences encoding (i) FOXP3, (ii) a first CISC component comprising (a) an extracellular FKBP domain, (b) an IL-2Rγ transmembrane domain, and (c) an IL-2Rγ cytoplasmic domain or portion thereof, (iii) a second CISC component comprising (a) an extracellular FRB domain, (b) an IL-2Rβ transmembrane domain, and (c) an IL-2Rβ cytoplasmic domain or portion thereof, and (iv) an anti-CD19 CAR. In some embodiments, the viral vector is a lentiviral vector. In some embodiments, the second CISC component comprises a truncated IL-2Rβ cytoplasmic domain. Cells expressing chimeric antigen receptors (CARs) Some aspects relate to engineered cells comprising: (i) in a FOXP3 gene locus of a genomic nucleic acid, a first heterologous promoter operably linked to: (a) a nucleotide sequence encoding a chimeric antigen receptor (CAR); (b) a nucleotide sequence encoding a first CISC component; and (c) a first coding exon (exon 2) of the FOXP3 gene, where the nucleotide sequences of (i)(a) and (b) are 5′ to the first coding exon and in frame with the start codon of the FOXP3 gene; and (ii) in a second gene locus of a genomic nucleic acid, a second heterologous promoter operably linked to: (a) a nucleotide encoding a second CISC component. In such engineered cells, one CISC component comprises an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain and an IL-2Rβ cytoplasmic domain or portion thereof, and wherein the other CISC component comprises an interleukin-2 receptor gamma IL-2Rγ transmembrane domain and an IL-2Rγ cytoplasmic domain or portion thereof, wherein the first and second extracellular binding domains dimerize in the presence of a ligand to transduce an IL-2 signal in the genetically modified cell. CISC components are described below in the section entitled “Chemically induced signaling complexes (CISC).” In some embodiments, the second gene locus is selected from the group consisting of TRAC, TRBC, and CD3Z. In some embodiments, the second gene locus is a TRAC gene. In some embodiments, the second gene locus is a TRBC gene locus. In some embodiments, the second gene locus is a CD3Z gene locus. In some embodiments, the engineered cell expresses a T cell receptor (TCR) on its surface. In some embodiments, binding of CD3 (e.g., by anti- CD3 antibody or anti-CD3 / CD28 beads) on the engineered cell transduces a TCR signal in the cell. In some embodiments, the second gene locus is not a TRAC, TRBC, or CD3Z gene locus. Other non-limiting examples of targeted loci are described below in the sections entitled “Targeted loci”. In some embodiments, the engineered cell is modified as described below in the section entitled “Selection by Essential Gene Exon Knock-In (SLEEK).” In the FOXP3 gene locus of engineered cells, sequences (i)(a) and (i)(b), encoding a CAR and CISC component, respectively, may be present in any orientation upstream of the first coding exon. For instance, in some embodiments, the nucleotide sequence of (i)(a), encoding the CAR, is upstream from (5′ to) the nucleotide sequence of (i)(b), encoding the CISC component. In some embodiments, the nucleotide sequence of (i)(b), encoding the CISC component is upstream from (5′ to) the nucleotide sequence of (i)(a), encoding the CAR. In some embodiments, for the set of the (1) nucleotide sequence of (i)(a), (2) the nucleotide sequence of (i)(b), and (3) the first coding exon of the FOXP3 gene, each member (1)–(3) is separated from its closest other member(s) by a 2A motif-encoding nucleotide sequence. For instance, where the nucleotide sequence of (i)(a) is 5′ to the nucleotide sequence of (i)(b), a first 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(a) and (i)(b), and a second 2A motif-encoding nucleotide sequence separates the nucleotide sequence of (i)(b) and the first coding exon of the FOXP3 gene. In some embodiments, the FOXP3 gene locus further comprises (i)(d) a nucleotide sequence encoding a soluble FRB domain, to which the first heterologous promoter is operably linked, where the nucleotide sequence of (i)(d) is 5′ to and in frame with the first coding exon of the FOXP3 gene. In some embodiments, for the set of the (1) nucleotide sequence of (i)(a), (2) the nucleotide sequence of (i)(b), (3) the nucleotide sequence of (i)(d), and (4) the first coding exon of the FOXP3 gene, each member (1)–(4) is separated from its closest other member(s) by a 2A motif-encoding nucleotide sequence. For instance, where the FOXP3 gene locus comprises, in 5′-to-3′ order, the nucleotide sequence of (i)(a), the nucleotide sequence of (i)(b), the nucleotide sequence of (i)(d), and the first coding exon of the FOXP3 gene, a first 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(a) and (i)(b), a second 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(b) and (i)(d), and a third 2A motif-encoding nucleotide sequence separates the nucleotide sequenced of (i)(d) and the first coding exon of the FOXP3 gene. In some embodiments, the second gene locus further comprises (ii)(b) a nucleotide sequence encoding a soluble FRB domain, to which the second heterologous promoter is operably linked. The nucleotide sequence of (ii)(b) may be 5′ to or 3′ to the nucleotide sequence of (ii)(a). In some embodiments, the nucleotide sequences of (ii)(a) and (ii)(b) are separated by a third 2A motif-encoding nucleotide sequence, where the FOXP3 gene locus comprises first and second 2A motif-encoding nucleotide sequences. Non-limiting examples of arrangements of 2A motif-encoding nucleotide sequences, the nucleotide sequences of (i)(a), (i)(b), the first coding exon of the FOXP3 gene, and, where present, the nucleotide sequence of (i)(d) or (ii)(b) encoding the soluble FRB domain, are provided in the table below: 2A motifs are amino acid sequences that, when present in a longer amino acid sequence encoded by an mRNA, cause ribosomal skipping, resulting in release of an elongating polypeptide upstream from the 2A motif in the amino acid sequence (encoded by a nucleotide sequence upstream from the nucleotide sequence encoding the 2A motif). Following release, the ribosome continues translation of the nucleotide sequence downstream from the 2A motif, initiating production of a second polypeptide chain beginning with the first amino acid downstream from the 2A motif. Inclusion of n 2A motifs in a nucleotide sequence therefore allows production of n+1 encoded polypeptides from translation of the nucleotide sequence. Non-limiting examples of 2A motifs include F2A, E2A, P2A, and T2A, after foot-and-mouth disease virus, equine rhinitis A virus, porcine teschovirus-1, and Thosea asigna virus, respectively. See, e.g., Liu et al., Sci Rep.2017.7(1):2193. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences has at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mismatches (insertions, substitutions, and / or deletions) relative to each other 2A motif-encoding nucleotide sequence. Mismatches between nucleotide sequences reduce the probability of homologous recombination, which may have undesired consequences in engineered cells. For example, internal recombination between two 2A motif-encoding nucleotide sequences on the same nucleic acid may excise the nucleotide sequence between the recombining 2A motif-encoding nucleotide sequences, resulting in lost expression of gene products encoded by the excised nucleotide sequence. In addition, recombination between two nucleotide sequences on different chromosomes may result in a chromosomal translocation. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif-encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 substitutions. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif- encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 insertions. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif-encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 deletions. Each 2A motif-encoding nucleotide sequence may be any suitable 2A motif-encoding nucleotide sequence. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a 2A motif independently selected from the group consisting of F2A, P2A, T2A, and E2A. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a different 2A motif independently selected from the group consisting of F2A, P2A, T2A, and E2A. The skilled artisan will appreciate that two nucleotide sequences may encode the same 2A motif while differing by one or more nucleotides. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode an F2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode a P2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode a T2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode an E2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes an F2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a P2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a T2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes an E2A motif. In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(a), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(a), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(b). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(a), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(b), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(b), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(a), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(b). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(b), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(b), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(a), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(a), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(b), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(b), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). In some embodiments, the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(b), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(a), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(a). Some aspects relate to populations of cells comprising genetically engineered cells as described in this section. In some embodiments, at least 80% of cells of the population are FOXP3+CAR+. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+. In some embodiments, at least 80% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, no more than 10% of cells of the population are CAR+FOXP3–. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3–. In some embodiments, no more than 10% of cells of the population are CAR+FOXP3– and express both first and second CISC components. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3– and express both first and second CISC components. Chimeric antigen receptors (CARs) Some aspects relate to cells expressing a chimeric antigen receptor (CAR) comprising an antigen-binding domain specific for a target antigen or a portion thereof. CARs comprise multiple elements that allow for signal transduction upon recognition and binding of the antigen(s) for which they are specific (e.g., CD19). The antigen-binding domain is also called the binding domain or the extracellular binding domain and comprises an extracellular polypeptide that facilitates binding to the target antigen, which can subsequently induce intracellular signaling and a physiological response in the cell engineered to express the CAR. CARs may also comprise a hinge region (also called a stalk or spacer region), which is an that connects the antigen-binding domain to the transmembrane domain. As used herein, a “hinge” refers to a flexible joint that connects two regions (e.g., domains) of a protein. See, e.g., Wriggers & Schulten, Proteins.1997.29(1):1–14. The skilled artisan will appreciate that protein regions tethered by a hinge are more constrained in movement than regions connected by a flexible linker such as a glycine linker (e.g., having the amino acid sequence GGG). The transmembrane domain of a CAR is a conduit for signal transduction from the antigen-binding domain to the cytoplasmic domain(s). CARs have at least one cytoplasmic domain, also called endodomains, which can be costimulatory domains and / or intracellular signaling domains. The source(s) from which each domain is derived contribute to the overall functionality of the CAR. Cells engineered to express a chimeric antigen receptor (CAR) specific for a particular antigen are competent to recognize and bind that antigen. If the target antigen is expressed on a cell, the engineered cell comprising the CAR may modulate or kill the antigen-expressing cell. Accordingly, some aspects relate to cells expressing a chimeric antigen receptor (CAR) comprising an antigen-binding domain that specifically binds to CD19 (anti-CD19 CAR). In some embodiments, the cell comprises a nucleic acid comprising a nucleotide sequence encoding the CAR. Some aspects relate to engineered T regulatory cells comprising an anti-CD19 CAR, which can selectively deplete active B cells without inducing the negative effects observed in the use of T effector cells comprising an anti-CD19 CAR. Antigen-binding domains Any antigen can be targeted by a CAR. A CAR antigen-binding domain may be derived from any suitable source that allows binding of a target antigen, such as an antibody reagent. The term “antibody reagent” encompasses any complete antibody, as well as any antigen-binding fragment of an antibody, including a camelid heavy chain antibody fragment, an Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’) fragment, an IgG fragment, a single domain VHH, a bivalent VHH, complementarity determining regions (CDRs), or a domain antibody (dAb) fragment. Complete antibodies typically contain one variable heavy chain region (VH) and one variable light chain region (VL), which typically comprise three CDRs that confer specificity for an antigen. CDRs are linked together by framework regions (typically four framework regions), which help maintain the structure of the variable heavy and light chains but do not affect the antigen specificity. The amino acid residues that make up the CDRs and framework regions can be identified using established numbering conventions, including Kabat, Chothia, or IMGT. Antibodies can be from any source, such as rodents, primates, or humans, and include midibodies, chimeric antibodies, and humanized antibodies. Suitable methods of humanizing antibodies to improve their efficacy in humans include, for example, grafting non-human CDRs onto human antibody frameworks. Any suitable CAR that binds a target antigen may be used in engineered cells (e.g., in an engineered T regulatory cell). A chimeric antigen receptor may comprise an antigen- binding domain, a hinge domain, a transmembrane domain, a first costimulatory domain, a second costimulatory domain, and / or an intracellular signaling domain. In some embodiments, a CAR comprises in N terminal to C terminal order: an antigen-binding domain, a hinge domain, a transmembrane domain, and one or more signaling domains. In some embodiments, the antigen-binding domain is a Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’)2 fragment, an IgG, a camelid heavy chain antibody, a single domain VHH, or a bivalent VHH. In some embodiments, a CAR comprises an Fv that binds a target antigen. In some embodiments, a CAR comprises an scFv that binds a target antigen. In some embodiments, a CAR comprises a Fab fragment that binds a target antigen. In some embodiments, a CAR comprises a F(ab’) fragment that binds a target antigen. In some embodiments, a CAR comprises a F(ab’)2 fragment that binds a target antigen. In some embodiments, a CAR comprises an IgG that binds a target antigen. In some embodiments, a CAR comprises a camelid heavy chain antibody that binds a target antigen. In some embodiments, a CAR comprises a single domain VHH that binds a target antigen. In some embodiments, a CAR comprises a bivalent VHH that binds a target antigen. CD19-binding domains Any extracellular protein can be targeted by a CAR. Some aspects relate to a CAR comprising a CD19-binding domain. CD19 (also known as B-Lymphocyte Surface Antigen B4 or T-Cell Surface Antigen Leu-12, or CVID3) is a surface glycoprotein belonging to the immunoglobulin (Ig) superfamily that is expressed on B cells, beginning at the earliest stages of development until plasma cell terminal differentiation. CD19 is also expressed on follicular dendritic cells (FDCs), antigen-presenting cells resident to lymphoid follicles. Expression of anti-CD19 CARs in cells (e.g., in T regulatory cells), can redirect said cells to target CD19-expressing cells. An antibody, antigen-binding fragment thereof, or antigen-binding portion of a chimeric receptor is said to “recognize” a target antigen if it binds that antigen in a mixture of different proteins. Generally, “binding” refers to “preferential binding,” i.e., reaction or association with that antigen more frequently, more rapidly, with a greater duration, elicits higher biological responses, and / or with a greater affinity for the target antigen compared to other proteins. It should be understood that “preferential binding” to a particular antigen does not require exclusive binding (i.e., that the antibody, antigen-binding fragment thereof, or antigen-binding portion of a chimeric antigen receptor bind no proteins other than the target antigen). In some embodiments, the antibody, antigen-binding fragment thereof, or antigen- binding portion of a chimeric antigen receptor binds a target antigen with a KD of 0.1 to 100 nM, 0.3 to 30 nM, or 1 to 10 nM. In some embodiments, the KD is 2 to 10 nM, 2 to 8 nM, or 2 to 6 nM. In some embodiments, a CAR comprises a CD19-binding domain, which may be derived from any suitable source that allows specific binding of CD19, such as an antibody reagent. The term “antibody reagent” encompasses any complete antibody, as well as any antigen-binding fragment of an antibody, including a camelid heavy chain antibody fragment, an Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’)2 fragment, an IgG fragment, a single domain VHH, a bivalent VHH, complementarity determining regions (CDRs), or a domain antibody (dAb) fragment. Complete antibodies typically contain one variable heavy chain region (VH) and one variable light chain region (VL), which typically comprise three CDRs that confer specificity for an antigen. CDRs are linked together by framework regions (typically four framework regions), which help maintain the structure of the variable heavy and light chains but do not affect the antigen specificity. The amino acid residues that make up the CDRs and framework regions can be identified using established numbering conventions, including Kabat, Chothia, or IMGT. Antibodies can be from any source, such as rodents, primates, or humans, and include midbodies, chimeric antibodies, and humanized antibodies. Suitable methods of humanizing antibodies to improve their efficacy in humans include, for example, grafting non-human CDRs onto human antibody frameworks. Any suitable CAR that binds CD19 may be used in engineered cells (e.g., in an engineered T regulatory cell). A chimeric antigen receptor may comprise a CD19-binding domain, a hinge, a transmembrane domain, a first costimulatory domain, a second costimulatory domain, and / or an intracellular signaling domain. In some embodiments, a CAR comprises in N terminal to C terminal order: a CD19-binding domain, a hinge, a transmembrane domain, and one or more signaling domains. In some embodiments, a CAR comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 69, 70, 71, 72, 82, 83, 93, 103, 113, 123, 134, or 135. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 69. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 70. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 71. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 72. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 82. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 83. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 134. In some embodiments, a CAR comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 135. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 69. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 70. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 71. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 72. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 82. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 83. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 134. In some embodiments, a CAR comprises an amino acid sequence having at least 95%, 96%, 97%, 98%, 99%, or 100% identity to the amino acid sequence of SEQ ID NO: 135. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 69, 70, 71, 72, 82, 83, 93, 103, 113, 123, 134, or 135. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 69. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 70. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 71. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 72. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 82. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 83. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 134. In some embodiments, a CAR comprises an amino acid sequence of SEQ ID NO: 135. The skilled artisan will appreciate that SEQ ID NOs: 69, 70, 71, 72, 82, 83, 93, 103, 113, 123, 134, or 135 begin with a CD8α signal peptide, but other signal peptides are suitable for expression of CARs having the same combination of CD19- binding domain, hinge, transmembrane domain, and signaling domain(s). Accordingly, in some embodiments, percent identity is calculated using the portion of the amino acid sequence downstream from the CD8α signal peptide. In some embodiments, the CD19-binding domain is a Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’)2 fragment, an IgG, a camelid heavy chain antibody, a single domain VHH, or a bivalent VHH. In some embodiments, a CAR comprises an Fv that binds CD19. In some embodiments, a CAR comprises an scFv that binds CD19. In some embodiments, a CAR comprises a Fab fragment that binds CD19. In some embodiments, a CAR comprises a F(ab’) fragment that binds CD19. In some embodiments, a CAR comprises a F(ab’)2 fragment that binds CD19. In some embodiments, a CAR comprises an IgG that binds CD19. In some embodiments, a CAR comprises a camelid heavy chain antibody that binds CD19. In some embodiments, a CAR comprises a single domain VHH that binds CD19. In some embodiments, a CAR comprises a bivalent VHH that binds CD19. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 33. In some embodiments, a CAR comprises a CD19- binding domain comprising a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 35 and / or a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 34. In some embodiments, a CAR comprises a CD19-binding domain comprising a heavy chain complementarity determining region 1 (HCDR1) comprising the amino acid sequence of SEQ ID NO: 39, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 40, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 41, a light chain complementarity determining region 1 (LCDR1) comprising the amino acid sequence of SEQ ID NO: 36, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 37, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 38. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 42. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 44 and / or a VL comprising the amino acid sequence of SEQ ID NO: 43. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 48, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 49, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 50, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 45, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 46, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 47. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 51. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 53 and / or a VL comprising the amino acid sequence of SEQ ID NO: 52. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 57, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 58, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 59, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 54, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 55, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 56. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 60. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 62 and / or a VL comprising the amino acid sequence of SEQ ID NO: 61. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 66, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 67, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 68, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 63, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 64, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 65. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 73. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 75 and / or a VL comprising the amino acid sequence of SEQ ID NO: 74. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 79, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 80, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 81, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 76, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 77, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 78. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 84. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 86 and / or a VL comprising the amino acid sequence of SEQ ID NO: 85. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 90, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 91, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 92, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 87, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 88, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 89. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 94. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 96 and / or a VL comprising the amino acid sequence of SEQ ID NO: 97. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 100, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 101, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 102, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 97, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 98, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 99. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 104. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 106 and / or a VL comprising the amino acid sequence of SEQ ID NO: 105. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 110, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 111, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 112, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 107, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 108, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 109. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 114. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 116 and / or a VL comprising the amino acid sequence of SEQ ID NO: 115. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 120, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 121, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 122, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 117, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 118, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 119. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 124. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 126 and / or a VL comprising the amino acid sequence of SEQ ID NO: 125. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 130, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 131, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 132, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 127, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 128, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 129. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 147. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 149 and / or a VL comprising the amino acid sequence of SEQ ID NO: 148. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 152, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 153, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 154, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 149, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 150, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 151. In some embodiments, a CAR comprises a CD19-binding domain comprising the amino acid sequence of SEQ ID NO: 157. In some embodiments, a CAR comprises a CD19- binding domain comprising a VH comprising the amino acid sequence of SEQ ID NO: 159 and / or a VL comprising the amino acid sequence of SEQ ID NO: 158. In some embodiments, a CAR comprises a CD19-binding domain comprising a HCDR1 comprising the amino acid sequence of SEQ ID NO: 162, a HCDR2 comprising the amino acid sequence of SEQ ID NO: 163, a HCDR3 comprising the amino acid sequence of SEQ ID NO: 164, a LCDR1 comprising the amino acid sequence of SEQ ID NO: 159, a LCDR2 comprising the amino acid sequence of SEQ ID NO: 160, and a LCDR3 comprising the amino acid sequence of SEQ ID NO: 161. Hinges In some embodiments, a CAR comprises a hinge (also called a stalk or spacer). In some embodiments, a CAR comprises a hinge derived from CD8α, CD28, CD137 (4-1BB), OX40, CD3ζ, CD45, CD4, CD5, CD9, CD16, CD22, CD33, CD37, CD64, CD80, CD86, CD134, CD154, or ICOS. In some embodiments, a CAR comprises a CD28 hinge. In some embodiments, a CAR comprises a CD137 (4-1BB) hinge. In some embodiments, a CAR comprises an OX40 hinge. In some embodiments, a CAR comprises a CD3ζ hinge. In some embodiments, a CAR comprises a CD45 hinge. In some embodiments, a CAR comprises a CD4 hinge. In some embodiments, a CAR comprises a CD5 hinge. In some embodiments, a CAR comprises a CD9 hinge. In some embodiments, a CAR comprises a CD9 hinge. In some embodiments, a CAR comprises a CD16 hinge. In some embodiments, a CAR comprises a CD22 hinge. In some embodiments, a CAR comprises a CD33 hinge. In some embodiments, a CAR comprises a CD37 hinge. In some embodiments, a CAR comprises a CD64 hinge. In some embodiments, a CAR comprises a CD80 hinge. In some embodiments, a CAR comprises a CD86 hinge. In some embodiments, a CAR comprises a CD134 hinge. In some embodiments, a CAR comprises a CD154 hinge. In some embodiments, a CAR comprises an ICOS hinge. In some embodiments, a CAR comprises a CD8α hinge. In some embodiments, the CD8α hinge comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 136. In some embodiments, the CD8α hinge comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: 136. In some embodiments, the CD8α hinge comprises an amino acid sequence of SEQ ID NO: 136. In some embodiments, a CAR comprises an IgG4 hinge. In some embodiments, the IgG4 hinge comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 137. In some embodiments, the IgG4 hinge comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: 137. In some embodiments, the IgG4 hinge comprises an amino acid sequence of SEQ ID NO: 137. In some embodiments, the IgG4 hinge comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 145. In some embodiments, the IgG4 hinge comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: 145. In some embodiments, the IgG4 hinge comprises an amino acid sequence of SEQ ID NO: 145. In some embodiments, a CAR comprises a CD28 hinge. In some embodiments, the CD28 hinge comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: 138. In some embodiments, the CD28 hinge comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 138. In some embodiments, the CD28 hinge comprises an amino acid sequence of SEQ ID NO: 138. Transmembrane domains In some embodiments, a CAR comprises a transmembrane domain. In some embodiments, a CAR comprises a transmembrane domain derived from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, a CAR comprises a TCRα transmembrane domain. In some embodiments, a CAR comprises a TCRβ transmembrane domain. In some embodiments, a CAR comprises a TCRζ transmembrane domain. In some embodiments, a CAR comprises a CD3ε transmembrane domain. In some embodiments, a CAR comprises a CD3ζ transmembrane domain. In some embodiments, a CAR comprises a CD28 transmembrane domain. In some embodiments, a CAR comprises a CD45 transmembrane domain. In some embodiments, a CAR comprises a CD4 transmembrane domain. In some embodiments, a CAR comprises a CD5 transmembrane domain. In some embodiments, a CAR comprises a CD7 transmembrane domain. In some embodiments, a CAR comprises a CD9 transmembrane domain. In some embodiments, a CAR comprises a CD16 transmembrane domain. In some embodiments, a CAR comprises a CD22 transmembrane domain. In some embodiments, a CAR comprises a CD33 transmembrane domain. In some embodiments, a CAR comprises a CD37 transmembrane domain. In some embodiments, a CAR comprises a CD41 transmembrane domain. In some embodiments, a CAR comprises a CD64 transmembrane domain. In some embodiments, a CAR comprises a CD68 transmembrane domain. In some embodiments, a CAR comprises a CD134 transmembrane domain. In some embodiments, a CAR comprises a CD137 (4-1BB) transmembrane domain. In some embodiments, a CAR comprises a CD154 transmembrane domain. In some embodiments, a CAR comprises an ICOS domain. In some embodiments, a CAR comprises a CD8 transmembrane domain. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 139. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: 139. In some embodiments, the CD8 transmembrane domain comprises an amino acid sequence of SEQ ID NO: 139. In some embodiments, a CAR comprises a CD28 transmembrane domain. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 140. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to SEQ ID NO: CD28. In some embodiments, the CD28 transmembrane domain comprises an amino acid sequence of SEQ ID NO: 140. Signaling (costimulatory) domains In some embodiments, a CAR comprises a first costimulatory (or signaling) domain and / or a second costimulatory (or signaling) domain. In some embodiments, a CAR comprises a single costimulatory domain. In some embodiments, the single costimulatory domain is derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, CD137 (4-1BB), or a Toll-like receptor (TLR), In some embodiments, a CAR comprises a first costimulatory domain and a second costimulatory domain. In some embodiments, the first costimulatory domain and / or the second costimulatory domain are independently derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, CD137 (4-1BB), or a Toll-like receptor (TLR). In some embodiments, a CAR comprises an OX40 costimulatory domain. In some embodiments, a CAR comprises a CD2 costimulatory domain. In some embodiments, a CAR comprises a CD3ζ costimulatory domain. In some embodiments, a CAR comprises a CD7 costimulatory domain. In some embodiments, a CAR comprises a CD27 costimulatory domain. In some embodiments, a CAR comprises a CD28 costimulatory domain. In some embodiments, a CAR comprises a CD30 costimulatory domain. In some embodiments, a CAR comprises a CD40 costimulatory domain. In some embodiments, a CAR comprises a CD258 domain. In some embodiments, a CAR comprises an ICOS costimulatory domain. In some embodiments, a CAR comprises a GITR costimulatory domain. In some embodiments, a CAR comprises an IL-2Rβ costimulatory domain. In some embodiments, a CAR comprises an IL-2Rγ costimulatory domain. In some embodiments, a CAR comprises an IL-7Rα domain. In some embodiments, a CAR comprises a CTLA4 costimulatory domain. In some embodiments, a CAR comprises a PD-1 costimulatory domain. In some embodiments, a CAR comprises a B7-H3 costimulatory domain. In some embodiments, a CAR comprises a FCεRI costimulatory domain. In some embodiments, a CAR comprises a NKG2C costimulatory domain. In some embodiments, a CAR comprises a NKG2D costimulatory domain. In some embodiments, a CAR comprises a CD137 (4-1BB) costimulatory domain. In some embodiments, a CAR comprises a TLR costimulatory domain. In some embodiments, a CAR comprises a first costimulatory domain and / or a second costimulatory domain that is a CD3ζ signaling domain. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 141. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to an amino acid sequence of SEQ ID NO: 141. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence of SEQ ID NO: 141. In some embodiments, a CAR comprises a first costimulatory domain and / or a second costimulatory domain that is a 4-1BB signaling domain. In some embodiments, the 4-1BB signaling domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 142. In some embodiments, the 4-1BB signaling domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to an amino acid sequence of SEQ ID NO: 142. In some embodiments, the 4-1BB signaling domain comprises an amino acid sequence of SEQ ID NO: 142. In some embodiments, a CAR comprises a first costimulatory domain and / or a second costimulatory domain that is a CD28 signaling domain. In some embodiments, the CD28 signaling domain comprises an amino acid sequence having at least 95% sequence identity to SEQ ID NO: 143. In some embodiments, the CD28 signaling domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to an amino acid sequence of SEQ ID NO: 143. In some embodiments, the CD28 signaling domain comprises an amino acid sequence of SEQ ID NO: 143. In some embodiments, a CAR comprises a first costimulatory domain and / or a second costimulatory domain that is a FCεRI signaling domain. In some embodiments, the FCεRI signaling domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 144. In some embodiments, the FCεRI signaling domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to an amino acid sequence of SEQ ID NO: 144. In some embodiments, the FCεRI signaling domain comprises an amino acid sequence of SEQ ID NO: 144. In some embodiments, a CAR comprises a first costimulatory domain that is a CD3ζ signaling domain and a second costimulatory domain that is a CD28 signaling domain. In some embodiments, a CAR comprises a first costimulatory domain that is a CD3ζ signaling domain and a second costimulatory domain that is a FCεRI signaling domain. In some embodiments, a CAR comprises a first costimulatory domain that is a CD3ζ signaling domain and a second costimulatory domain that is a 4-1BB signaling domain. In some embodiments, a CAR comprises a CD3ζ signaling domain and two or more of: a CD28 signaling domain, a 4-1BB signaling domain, and a FCεRI signaling domain. In some embodiments, a CAR comprises a CD3ζ signaling domain, a CD28 signaling domain, a 4-1BB signaling domain, and a FCεRI signaling domain. In some embodiments, a CAR comprises two or more of: a CD3ζ signaling domain, a CD28 signaling domain, a 4-1BB signaling domain, and a FCεRI signaling domain. In some embodiments, a CAR comprises an intracellular signaling domain. In some embodiments, a CAR comprises an intracellular signaling domain derived from OX40, CD2, CD3ζ, CD3γ, CD3δ, CD3ε, FCεRI, CD7, CD27, CD28, CD30, CD40, CD79a, CD79b, CD137 (4-1BB), ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, or a TLR. In some embodiments, a CAR comprises an OX40 intracellular signaling domain. In some embodiments, a CAR comprises a CD2 intracellular signaling domain. In some embodiments, a CAR comprises a CD3γ intracellular signaling domain. In some embodiments, a CAR comprises a CD3δ intracellular signaling domain. In some embodiments, a CAR comprises a CD3ε intracellular signaling domain. In some embodiments, a CAR comprises a FCεRI intracellular signaling domain. In some embodiments, a CAR comprises a CD7 intracellular signaling domain. In some embodiments, a CAR comprises a CD27 intracellular signaling domain. In some embodiments, a CAR comprises a CD28 intracellular signaling domain. In some embodiments, a CAR comprises a CD30 intracellular signaling domain. In some embodiments, a CAR comprises a CD40 intracellular signaling domain. In some embodiments, a CAR comprises a CD79a intracellular signaling domain. In some embodiments, a CAR comprises a CD79b intracellular signaling domain. In some embodiments, a CAR comprises a CD137 (4-1BB) intracellular signaling domain. In some embodiments, a CAR comprises an ICOS intracellular signaling domain. In some embodiments, a CAR comprises a GITR intracellular signaling domain. In some embodiments, a CAR comprises an IL-2Rβ intracellular signaling domain. In some embodiments, a CAR comprises an IL-2Rγ intracellular signaling domain. In some embodiments, a CAR comprises an IL-7Rα intracellular signaling domain. In some embodiments, a CAR comprises a CTLA4 intracellular signaling domain. In some embodiments, a CAR comprises a TLR intracellular signaling domain. In some embodiments, a CAR comprises an intracellular domain that is a CD3ζ signaling domain. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence having at least 95% identity to SEQ ID NO: 141. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence having no more than 1, no more than 2, or no more than 3 mismatches relative to an amino acid sequence of SEQ ID NO: 141. In some embodiments, the CD3ζ signaling domain comprises an amino acid sequence of SEQ ID NO: 141. Signal peptides In some embodiments, a CAR comprises a signal peptide (also called a leader peptide). In some embodiments, the signal peptide is an IgK signal peptide, a CD8α signal peptide, a CD8β signal peptide, an interleukin (IL)-2 signal peptide, an IL-15 signal peptide, a CD45 signal peptide, or a GM-CSFRα signal peptide. In some embodiments, a CAR comprises an IgK signal peptide. In some embodiments, a CAR comprises a CD8α signal peptide. In some embodiments, a CAR comprises a CD8β signal peptide. In some embodiments, a CAR comprises an IL-2 signal peptide. In some embodiments, a CAR comprises an IL-15 signal peptide. In some embodiments, a CAR comprises a CD45 signal peptide. In some embodiments, a CAR comprises a GM-CSFRα signal peptide. In some embodiments, the signal peptide comprises an amino acid sequence of SEQ ID NO: 20. Domain combinations In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19- binding domain), a CD8α hinge, a first costimulatory domain comprising a 4-1BB signaling domain, a second costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), a CD8α hinge, a first costimulatory domain comprising a CD28 signaling domain, a second costimulatory domain comprising a 4-1BB signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), a CD8α hinge, a first costimulatory domain comprising a 4-1BB signaling domain, a second costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19- binding domain), an IgG4 hinge, a first costimulatory domain comprising a 4-1BB signaling domain, a second costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), an IgG4 hinge, a first costimulatory domain comprising a CD28 signaling domain, a second costimulatory domain comprising a 4-1BB signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), an IgG4 hinge, a first costimulatory domain comprising a 4-1BB signaling domain, a second costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19- binding domain), a CD28 hinge, a first costimulatory domain comprising a 4-1BB signaling domain, a second costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), a CD28 hinge, a first costimulatory domain comprising a CD28 signaling domain, a second costimulatory domain comprising a 4-1BB signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19- binding domain), an IgG4 hinge, a transmembrane domain, a costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, the transmembrane domain is derived from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, a CAR comprises a CD19-binding domain, an IgG4 hinge, a CD8α transmembrane domain, a costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19- binding domain), a CD28 hinge, a transmembrane domain, a costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. In some embodiments, the transmembrane domain is derived from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS. In some embodiments, a CAR comprises an antigen-binding domain (e.g., a CD19-binding domain), a CD28 hinge, a CD28 transmembrane domain, a costimulatory domain comprising a CD28 signaling domain, and a CD3ζ signaling domain. Engineered cells Engineered cells include cells that have been modified to express particular proteins that may confer a particular functionality to the cell. Cells engineered to express a chimeric antigen receptor (CAR) specific for a particular antigen are competent to recognize and bind that antigen. If the target antigen is expressed on a cell, the engineered cell comprising the CAR may modulate or kill the antigen-expressing cell. Some aspects relate to engineered cells (e.g., engineered T regulatory cells) comprising a chimeric antigen receptor (CAR) comprising a CD19-binding domain (an anti- CD19 CAR). Engineered cells expressing an anti-CD19 CAR can target and modulate CD19- expressing cells. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits production of one or more cytokines. In some embodiments, an engineered cell inhibits production of one or more cytokines by CD19+ cells. In some embodiments, an engineered cell inhibits production of one or more cytokines by B cells. In some embodiments, an engineered cell inhibits production of one or more cytokines by T cells. Non-limiting examples of cytokines whose production may be inhibited include interleukin (IL)-6, IL-10, tumor necrosis factor alpha (TNFα), B cell activating factor (BAFF), lymphotoxin alpha (LTα), interferon gamma (IFNγ), IL-4, and IL-13. In some embodiments, an engineered cell inhibits production of IL-6. In some embodiments, an engineered cell inhibits production of IL-10. In some embodiments, an engineered cell inhibits production of TNFα. In some embodiments, an engineered cell inhibits production of BAFF. In some embodiments, an engineered cell inhibits production of LTα. In some embodiments, an engineered cell inhibits production of IFNγ. In some embodiments, an engineered cell inhibits production of IL-4. In some embodiments, an engineered cell inhibits production of IL-13. The skilled artisan will appreciate that any suitable method may be used to determine the extent of inhibition of cytokine production by B or T cells, such as performing ELISAs to quantify B or T cell-derived cytokines or measuring intracellular cytokine production in B or T cells via flow cytometry. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell cytokine production by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits B cell cytokine production by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104- fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell cytokine production by at least 1.1-fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits B cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits T cell cytokine production by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits T cell cytokine production by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104- fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits T cell cytokine production by at least 1.1-fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits T cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits T cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits T cell cytokine production by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits production of antibodies. In some embodiments, an engineered cell inhibits production of antibodies by CD19+cells. In some embodiments, an engineered cell inhibits production of antibodies by B cells. In some embodiments, an engineered cell inhibits production of IgM. In some embodiments, an engineered cell inhibits production of IgG. In some embodiments, an engineered cell inhibits production of IgE. In some embodiments, an engineered cell inhibits production of IgA. In some embodiments, an engineered cell inhibits production of IgD. The skilled artisan will appreciate that any suitable method may be used to determine the extent of inhibition of antibody production, such as performing ELISAs to quantify target antibodies or complement-binding assays. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits production of antibodies by B cells by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits production of antibodies by B cells by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100- fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits production of antibodies by B cells by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits production of antibodies by B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits production of antibodies by B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits production of antibodies by B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits class-switch recombination. In some embodiments, an engineered cell inhibits class- switching from IgM to IgG. In some embodiments, an engineered cell inhibits class- switching from IgM to IgA. In some embodiments, an engineered cell inhibits class- switching from IgM to IgE. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits affinity maturation. In some embodiments, an engineered cell inhibits somatic hypermutation. In some embodiments, an engineered cell inhibits class-switch recombination. In some embodiments, an engineered cell inhibits somatic hypermutation and class-switch recombination. The skilled artisan will appreciate that any suitable method may be used to determine the extent of inhibition of class-switch recombination or somatic hypermutation, such as measuring expression of immunoglobulin isotypes on B cells, performing ELISAs to quantify the immunoglobulin isotypes produced by B cells, and sequencing of the B cell receptor (BCR) coding sequences. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits class-switch recombination and / or somatic hypermutation by at least 1.1-fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits class-switch recombination and / or somatic hypermutation by at least 1.1-fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits class-switch recombination and / or somatic hypermutation by at least 1.1-fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits class-switch recombination and / or somatic hypermutation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express a CAR. In some embodiments, an engineered cell inhibits class-switch recombination and / or somatic hypermutation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits class-switch recombination and / or somatic hypermutation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) induces anergy. As used herein, “anergy” refers to functional inactivation or unresponsiveness. In some embodiments, an engineered cell induces anergy in CD19+cells. In some embodiments, an engineered cell induces anergy in B cells. In some embodiments, an engineered cell reduces B cell proliferation. In some embodiments, an engineered cell abrogates B cell proliferation. In some embodiments, an engineered cell reduces antibody production by B cells. In some embodiments, an engineered cell abrogates B antibody production by B cells. In some embodiments, an engineered cell induces decreased signaling in B cells. In some embodiments, an engineered cell abrogates signaling in B cells. In some embodiments, an engineered cell induces upregulation of inhibitory molecules in or on B cells. In some embodiments, an engineered cell induces failure to internalize or recycle the B cell receptor (BCR) on B cells. The skilled artisan will appreciate that any suitable method may be used to measure B cell anergy, such as B cell receptor (BCR) internalization or recycling assays, quantification of B cell proliferation or antibody production, measuring intracellular signaling in B cells or expression of inhibitory molecules by B cells using flow cytometry. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) increases B cell anergy by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express a CAR. In some embodiments, an engineered cell increases B cell anergy by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell increases B cell anergy by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing a CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) increases B cell anergy by at least 90%, 100%, 125%, 150%, 175%, 200%, 250%, 300%, 350%, 400%, 450%, or 500% relative to a control cell that does not express a CAR. In some embodiments, an engineered cell increases B cell anergy by at least 90%, 100%, 125%, 150%, 175%, 200%, 250%, 300%, 350%, 400%, 450%, or 500% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell increases B cell anergy by at least 90%, 100%, 125%, 150%, 175%, 200%, 250%, 300%, 350%, 400%, 450%, or 500% relative to a control cell that is not an engineered Treg expressing a CAR. The abundance of B cells (e.g., total B cells or a specific subset of B cells) can be assessed by any suitable method, for example by detecting a cellular marker indicative of B cells (e.g., CD19), assessing a direct or indirect activity of B cells (e.g., antibody production), direct cell counting of B cells using a hemacytometer or automated cell counter, and / or by measuring the production of one or more cytokines produced by B cells (e.g., IL-10). In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits proliferation of CD19+cells. In some embodiments, an engineered cell inhibits proliferation of B cells. The skilled artisan will appreciate that any suitable method may be used to determine the extent of suppression of B cell proliferation, including but not limited to measuring frequency of B cell division by flow cytometry, bromodeoxyuridine (BrDU) incorporation assays, DNA quantification assays, and direct counting of cells using a hemacytometer or automated cell counter. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) depletes (e.g., kills) active B cells. As used herein, “active B cell” refers to a mature B cell that has encountered an antigen it recognizes and has initiated a response to the same. In some embodiments, an engineered anti-CD19 Treg kills B cells. Suitable methods of assessing T cell-mediated killing of other cells are known in the art (e.g., Piccinini et al., Cancer Immunol Immunother.2024.73(9):168) and described in the Examples. In some embodiments, an engineered anti-CD19 Treg preferentially kills B cells that are activated, proliferating, and / or autoreactive. “Activated” B cells are those that have recently encountered a cognate antigen and initiated a response to the same, and express one or more surface markers selected from CD80, CD86, CD27, IgM, IgD, CD38, CD138, CD10, and CD24. “Proliferating” B cells are those in the process of cell division, which may be identified, e.g., by surface presence of CD107a, increased intracellular abundance of Ki-67 or dilution of cell trace dye (e.g., CFSE or CTV). “Autoreactive” B cells, as used herein, are B cells that express a B cell receptor or antibody that preferentially binds a self-antigen. In some embodiments, an engineered anti-CD19 Treg preferentially kills activated B cells. In some embodiments, an engineered anti-CD19 Treg preferentially kills proliferating B cells. In some embodiments, an engineered anti-CD19 Treg kills autoreactive B cells. In some embodiments, an engineered anti-CD19 Treg preferentially kills B cells that are activated and / or proliferating and does not kill naïve B cells. An engineered cell that “preferentially kills” a specified subset of a cell type (e.g., activated B cells, as a subset of B cells) kills a statistically significantly higher proportion of cells of that subset, compared to the proportion of cells of that cell type that do not belong to the specified subset. For example, preferential killing of activated B cells may be identified by comparing the proportion of activated B cells killed to the proportion of naïve or resting B cells killed. As another example, preferential killing of proliferating B cells may be identified by comparing the proportion of proliferating B cells killed to the proportion of non- proliferating B cells killed. In some embodiments, an engineered anti-CD19 Treg substantially depletes B cells that are activated, proliferating, and / or autoreactive, and does not substantially deplete naïve B cells, in a subject. In some embodiments, administration of an engineered anti-CD19 Treg substantially reduces the proportion of circulating B cells in a subject that are activated, proliferating, or autoreactive. In some embodiments, administration of an engineered anti-CD19 Treg substantially reduces the number of circulating B cells in a subject. In some embodiments, administration of an engineered anti-CD19 Treg substantially reduces the serum concentration of antibodies that preferentially bind one or more self- antigens. In some embodiments, administration of an engineered anti-CD19 Treg substantially reduces the serum concentration of antibodies that preferentially bind double- stranded DNA. In some embodiments, administration of an engineered anti-CD19 Treg substantially reduces the serum concentration of antibodies that preferentially bind SmRNP. “SmRNP” refers to motifs shared between Smith and ribonucleoprotein (RNP), as anti-Sm and anti-RNP antibodies are prevalent among subjects with systemic lupus erythematosus (SLE). In some embodiments, administration of an engineered anti-CD19 Treg substantially increases the proportion of circulating B cells in a subject that are naïve. In some embodiments, administration of an engineered anti-CD19 Treg substantially increases the number of circulating B cells in a subject that are non-activated, non-proliferating, and / or non-autoreactive. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell proliferation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express an anti-CD19 CAR. In some embodiments, an engineered cell inhibits B cell proliferation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell proliferation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell proliferation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express an anti-CD19 CAR. In some embodiments, an engineered cell inhibits B cell proliferation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell proliferation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) depletes active B cells at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100- fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express an anti- CD19 CAR. In some embodiments, an engineered cell depletes active B cells by at least 1.1- fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell depletes active B cells by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) depletes active B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express an anti-CD19 CAR. In some embodiments, an engineered cell depletes active B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell depletes active B cells by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits activation of CD19+cells. In some embodiments, an engineered cell inhibits activation of B cells. In some embodiments, an engineered cell inhibits the recognition of antigen by B cells. In some embodiments, an engineered cell inhibits internalization and processing of antigen by B cells. In some embodiments, an engineered cell inhibits costimulatory signaling on B cells. In some embodiments, an engineered cell inhibits activation of signaling pathways in B cells. In some embodiments, an engineered cell inhibits expansion of B cell clones. In some embodiments, an engineered cell inhibits differentiation of B cells into plasmablasts, plasma cells, or memory B cells. In some embodiments, an engineered cell inhibits antibody production by B cells. In some embodiments, an engineered cell inhibits upregulation of activation markers in or on the surface of B cells, such as CD69, CD25, CD40, and major histocompatibility complex type II (MHC-II). The skilled artisan will appreciate that any suitable method may be used to determine the extent of inhibition of activation of B cells, such as enzyme-linked immunosorbent assays (ELISAs) to quantify target antibodies or cytokines; antigen presentation assays; or measurement of intracellular signaling, activation markers, or B cell maturity via flow cytometry. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell activation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that does not express an anti-CD19 CAR. In some embodiments, an engineered cell inhibits B cell activation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell activation by at least 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5- fold, 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, 100-fold, 1,000-fold, 104-fold, or 105-fold relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. In some embodiments, an engineered cell (e.g., an engineered T regulatory cell) inhibits B cell activation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that does not express an anti-CD19 CAR. In some embodiments, an engineered cell inhibits B cell activation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg. In some embodiments, an engineered cell inhibits B cell activation by at least 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% relative to a control cell that is not an engineered Treg expressing an anti-CD19 CAR. Cell types Embodiments of engineered cells may be any suitable cell type. In some embodiments, the cell is a T cell, a precursor T cell, or a hematopoietic stem cell. In some embodiments, the cell is an NK-T cell (e.g., a FoxP3– NK-T cell or a FoxP3+ NK-T cell). In some embodiments, the cell is a regulatory B (Breg) cell (e.g., a FoxP3– B cell or a FoxP3+ B cell). In some embodiments, the cell is a CD4+ T cell (e.g., a FoxP3–CD4+ T cell or a FoxP3+CD4+ T cell) or a CD8+ T cell (e.g., a FoxP3–CD8+ T cell or a FoxP3+CD8+ T cell). In some embodiments, the cell is a CD25- T cell. In some embodiments, the cell is a regulatory T (Treg) cell. In some embodiments, the Treg cell is a sorted Treg cell. Non- limiting examples of Treg cells are Tr1, Th3, CD8+CD28-, and Qa-1-restricted T cells. In some embodiments, the Treg cell is a FoxP3+ Treg cell. In some embodiments, the Treg cell expresses CTLA-4, LAG-3, CD25, CD39, CD27, CD70, CD357 (GITR), neuropilin-1, galectin-1, and / or IL-2Rα on its surface. Methods for producing engineered cells Some aspects relate to methods of preparing engineered cells comprising contacting a cell with: (i) a first donor template comprising (a) a first 5′ homology arm having homology to a first nucleotide sequence in a FOXP3 gene locus of a genomic nucleic acid, (b) a first 3′ homology arm having homology to a second nucleotide sequence in the FOXP3 gene locus, and (c) a first heterologous promoter operably linked to: (d) a nucleotide sequence encoding a chimeric antigen receptor (CAR); and (e) a nucleotide sequence encoding a first CISC component, where the first heterologous promoter is inserted into the FOXP3 gene such that the first heterologous promoter is operably linked to a first coding exon (exon 2) of the FOXP3 gene and the nucleotide sequences of (i)(d) and (e) are 5′ to the first coding exon and in frame with the start codon of the FOXP3 gene; and (ii) a second donor template comprising (a) a first 5′ homology arm having homology to a first nucleotide sequence in a second gene locus of a genomic nucleic acid, (b) a second 3′ homology arm having homology to a second nucleotide sequence in the second gene, and (c) a second heterologous promoter operably linked to: (d) a nucleotide encoding a second CISC component. Some aspects relate to systems, vectors, and / or lipid nanoparticles comprising one or both of the donor templates. In such donor templates, one CISC component comprises an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain and an IL-2Rβ cytoplasmic domain or portion thereof, and wherein the other CISC component comprises an interleukin-2 receptor gamma IL-2Rγ transmembrane domain and an IL-2Rγ cytoplasmic domain or portion thereof, wherein the first and second extracellular binding domains dimerize in the presence of a ligand to transduce an IL-2 signal in the genetically modified cell. CISC components are described below in the section entitled “Chemically induced signaling complexes (CISC).” In some embodiments, the second gene locus is selected from the group consisting of TRAC, TRBC, and CD3Z gene loci. In some embodiments, the second gene locus is a TRAC gene locus. In some embodiments, the second gene locus is a TRBC gene locus. In some embodiments, the second gene locus is a CD3Z gene locus. In some embodiments, the engineered cell expresses a T cell receptor (TCR) on its surface. In some embodiments, binding of CD3 (e.g., by anti-CD3 antibody or anti-CD3 / CD28 beads) on the engineered cell transduces a TCR signal in the cell. In some embodiments, the second gene locus is not a TRAC, TRBC, or CD3Z gene locus. Other non-limiting examples of targeted loci are described below in the sections entitled “Targeted loci”. In some embodiments, the engineered cell is modified as described below in the section entitled “Selection by Essential Gene Exon Knock-In (SLEEK).” In the FOXP3 gene locus of engineered cells following insertion of the first donor template, sequences (i)(d) and (i)(e), encoding a CAR and CISC component, respectively, may be present in any orientation upstream of the first coding exon. For instance, in some embodiments, the nucleotide sequence of (i)(d), encoding the CAR, is upstream from (5′ to) the nucleotide sequence of (i)(e), encoding the CISC component. In some embodiments, the nucleotide sequence of (i)(e), encoding the CISC component is upstream from (5′ to) the nucleotide sequence of (i)(d), encoding the CAR. In some embodiments, following insertion of the first donor template, for the set of the (1) nucleotide sequence of (i)(d), (2) the nucleotide sequence of (i)(e), and (3) the first coding exon of the FOXP3 gene, each member (1)–(3) is separated from its closest other member(s) by a 2A motif-encoding nucleotide sequence. For instance, where the nucleotide sequence of (i)(d) is 5′ to the nucleotide sequence of (i)(e), a first 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(d) and (i)(e), and a second 2A motif-encoding nucleotide sequence separates the nucleotide sequence of (i)(e) and the first coding exon of the FOXP3 gene. In some embodiments, the first donor template further comprises (i)(f)a nucleotide sequence encoding a soluble FRB domain, to which the first heterologous promoter is operably linked, where following insertion of the first donor template, the nucleotide sequence of (i)(f) is 5′ to and in frame with the first coding exon of the FOXP3 gene. In some embodiments, following insertion of the first donor template, for the set of the (1) nucleotide sequence of (i)(d), (2) the nucleotide sequence of (i)(e), (3) the nucleotide sequence of (i)(f), and (4) the first coding exon of the FOXP3 gene, each member (1)–(4) is separated from its closest other member(s) by a 2A motif-encoding nucleotide sequence. For instance, where the FOXP3 gene locus comprises, in 5′-to-3′ order, the nucleotide sequence of (i)(d), the nucleotide sequence of (i)(e), the nucleotide sequence of (i)(f), and the first coding exon of the FOXP3 gene, a first 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(d) and (i)(e), a second 2A motif-encoding nucleotide sequence separates the nucleotide sequences of (i)(e) and (i)(f), and a third 2A motif-encoding nucleotide sequence separates the nucleotide sequenced of (i)(f) and the first coding exon of the FOXP3 gene. In some embodiments, following insertion of the second donor template, the second gene locus further comprises (ii)(e) a nucleotide sequence encoding a soluble FRB domain, to which the second heterologous promoter is operably linked. The nucleotide sequence of (ii)(e) may be 5′ to or 3′ to the nucleotide sequence of (ii)(d). In some embodiments, the nucleotide sequences of (ii)(d) and (ii)(e) are separated by a third 2A motif-encoding nucleotide sequence, where the FOXP3 gene locus comprises first and second 2A motif- encoding nucleotide sequences. Non-limiting examples of arrangements of 2A motif-encoding nucleotide sequences, the nucleotide sequences of (i)(d), (i)(e), the first coding exon of the FOXP3 gene, and, where present, the nucleotide sequence of (i)(d) or (ii)(e) encoding the soluble FRB domain, following insertion of donor templates, are provided in the table below: In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences has at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 mismatches (insertions, substitutions, and / or deletions) relative to each other 2A motif-encoding nucleotide sequence. Mismatches between nucleotide sequences reduce the probability of homologous recombination, which may have undesired consequences in engineered cells. For example, internal recombination between two 2A motif-encoding nucleotide sequences on the same nucleic acid may excise the nucleotide sequence between the recombining 2A motif-encoding nucleotide sequences, resulting in lost expression of gene products encoded by the excised nucleotide sequence. In addition, recombination between two nucleotide sequences on different chromosomes may result in a chromosomal translocation. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif-encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 substitutions. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif- encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 insertions. In some embodiments, each of the first, second, and third 2A motif-encoding nucleotide sequences differs from each other 2A motif-encoding nucleotide sequence by at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 deletions. Each 2A motif-encoding nucleotide sequence may be any suitable 2A motif-encoding nucleotide sequence. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a 2A motif independently selected from the group consisting of F2A, P2A, T2A, and E2A. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a different 2A motif independently selected from the group consisting of F2A, P2A, T2A, and E2A. The skilled artisan will appreciate that two nucleotide sequences may encode the same 2A motif while differing by one or more nucleotides. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode an F2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode a P2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode a T2A motif. In some embodiments, two or more 2A motif-encoding nucleotide sequences of a set each encode an E2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes an F2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a P2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes a T2A motif. In some embodiments, each 2A motif-encoding nucleotide sequence of a set encodes an E2A motif. In some embodiments, after insertion of the nucleotide sequences of (i)(f) and (i)(e), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(d), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(e). In some embodiments, after insertion of the nucleotide sequences of (i)(f) and (i)(e), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(e), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(f) and (i)(e), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(e), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(d), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(e). In some embodiments, after insertion of the nucleotide sequences of (i)(f) and (i)(e), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(e), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, the nucleotide sequence of (ii)(e), the third 2A motif-encoding nucleotide sequence, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(f), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(d), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(f), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(e), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(f), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(e), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(f), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(f), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, after insertion of the nucleotide sequences of (i)(d), (i)(e), and (i)(f), the FOXP3 gene locus comprises, in 5′-to-3′ order: the first heterologous promoter, the nucleotide sequence of (i)(f), the first 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(e), the second 2A motif-encoding nucleotide sequence, the nucleotide sequence of (i)(d), the third 2A motif-encoding nucleotide sequence, and the first coding exon, wherein the second gene locus comprises, in 5′-to-3′ order: the second heterologous promoter, and the nucleotide sequence of (ii)(d). In some embodiments, the method comprises contacting the cell with a first DNA endonuclease capable of cleaving at the FOXP3 gene locus (or a nucleic acid encoding the first DNA endonuclease), and a second DNA endonuclease capable of cleaving at the second gene locus. The first and second DNA endonucleases may be the same type of DNA endonuclease (e.g., TALEN) or different DNA endonucleases. In some embodiments, the method comprises contacting the cell population with an RNA-guided DNA endonuclease (or a nucleic acid encoding the RNA-guided DNA endonuclease), a first guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the FOXP3 gene locus (or a nucleic acid encoding the first gRNA), and a second gRNA comprising a spacer sequence that is complementary to a nucleotide sequence in the second gene locus. In some embodiments, the method comprises contacting the cell with a first guide ribonucleoprotein (gRNP) and a second gRNP, each gRNP comprising the RNA-guided DNA endonuclease and the first or second gRNA, respectively. In some embodiments, the method comprises contacting the cell with the ligand. In some embodiments, the cell is contacted with the ligand for at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, or 21 days. In some embodiments, the cell is contacted with the ligand for 1 to 21, 2 to 21, 3 to 21, 4 to 21, 5 to 21, 6 to 21, 7 to 21, 8 to 21, 9 to 21, 10 to 21, 11 to 21, 12 to 21, 13 to 21, 14 to 21, 15 to 21, 16 to 21, 17 to 21, 18 to 21, 19 to 21, or 20 to 21 days. In some embodiments, the cell is contacted with the ligand for 3 to 4, 3 to 5, 3 to 6, 3 to 7, 3 to 8, 3 to 9, 3 to 10, 3 to 11, 3 to 12, 3 to 13, 3 to 14, 3 to 15, 3 to 16, 3 to 17, 3 to 18, 3 to 19, 3 to 20, or 3 to 21 days. In some embodiments, the extracellular binding domain of one CISC component comprises an FKBP domain and the extracellular binding domain of the other CISC component comprises an FRB domain, and the method comprises contacting the cell with rapamycin or a rapalog. In some embodiments, the extracellular binding domain of one CISC component comprises an FKBP domain and the extracellular binding domain of the other CISC component comprises an FRB domain, and the method comprises contacting the cell with rapamycin. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 3 days. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 7 days. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 14 days. Some aspects relate to methods of producing a population of genetically engineered Tregs expressing CARs, comprising contacting a cell population with a plurality of the first donor template and a plurality of the second donor template, thereby producing a cell population comprising a plurality of the genetically modified cell. In some embodiments, the method comprises contacting the cell population with a plurality of a first DNA endonuclease capable of cleaving at the FOXP3 gene locus (or a plurality of a nucleic acid encoding the first DNA endonuclease), and a plurality of a second DNA endonuclease capable of cleaving at the second gene locus. The first and second DNA endonucleases may be the same type of DNA endonuclease (e.g., TALEN) or different DNA endonucleases. In some embodiments, the method comprises contacting the cell population with a plurality of an RNA-guided DNA endonuclease (or a plurality of a nucleic acid encoding the RNA-guided DNA endonuclease), a plurality of a first guide RNA (gRNA) comprising a spacer sequence that is complementary to a nucleotide sequence in the FOXP3 gene locus (or a plurality of a nucleic acid encoding the first gRNA), and a plurality of a second gRNA comprising a spacer sequence that is complementary to a nucleotide sequence in the second gene locus. In some embodiments, the plurality of the RNA-guided DNA endonuclease comprises a plurality of a first guide ribonucleoprotein (gRNP) and a plurality of a second gRNP, each gRNP comprising the RNA-guided DNA endonuclease and the first or second gRNA, respectively. In some embodiments, the method comprises contacting the cell population with the ligand. In some embodiments, the cell population is contacted with the ligand for 1 to 21, 2 to 21, 3 to 21, 4 to 21, 5 to 21, 6 to 21, 7 to 21, 8 to 21, 9 to 21, 10 to 21, 11 to 21, 12 to 21, 13 to 21, 14 to 21, 15 to 21, 16 to 21, 17 to 21, 18 to 21, 19 to 21, or 20 to 21 days. In some embodiments, the cell population is contacted with the ligand for 3 to 4, 3 to 5, 3 to 6, 3 to 7, 3 to 8, 3 to 9, 3 to 10, 3 to 11, 3 to 12, 3 to 13, 3 to 14, 3 to 15, 3 to 16, 3 to 17, 3 to 18, 3 to 19, 3 to 20, or 3 to 21 days. In some embodiments, the extracellular binding domain of one CISC component comprises an FKBP domain and the extracellular binding domain of the other CISC component comprises an FRB domain, and the method comprises contacting the cell population with rapamycin or a rapalog. In some embodiments, the extracellular binding domain of one CISC component comprises an FKBP domain and the extracellular binding domain of the other CISC component comprises an FRB domain, and the method comprises contacting the cell population with rapamycin. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 3 days. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 7 days. In some embodiments, rapamycin is provided at a concentration of at least 0.1 mM, 0.3 mM, 1 mM, 3 mM, 10 mM, 30 mM, 100 mM, or 300 mM for at least 14 days. In some embodiments, at least 24 hours after contacting the cell population with the donor templates, at least 80% of cells of the population are FOXP3+CAR+. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+. In some embodiments, at least 24 hours after contacting the cell population with the donor templates, at least 80% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, at least 24 hours after contacting the cell population with the donor templates, no more than 10% of cells of the population are CAR+FOXP3–. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3–. In some embodiments, at least 24 hours after contacting the cell population with the donor templates, no more than 10% of cells of the population are CAR+FOXP3– and express both first and second CISC components. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3– and express both first and second CISC components. In some embodiments, after at least 7 days of contact with the ligand, at least 80% of cells of the population are FOXP3+CAR+. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+. In some embodiments, after at least 7 days of contact with the ligand, at least 80% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of cells of the population are FOXP3+CAR+ and express both first and second CISC components. In some embodiments, after at least 7 days of contact with the ligand, no more than 10% of cells of the population are CAR+FOXP3–. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3–. In some embodiments, after at least 7 days of contact with the ligand, no more than 10% of cells of the population are CAR+FOXP3– and express both first and second CISC components. In some embodiments, no more than 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of cells of the population are CAR+FOXP3– and express both first and second CISC components. Some aspects relate to methods of producing an engineered cell by introducing into the cell a nucleic acid comprising a promoter that is operably linked to a nucleotide sequence encoding an anti-CD19 CAR. In some embodiments, the nucleic acid is inserted into the genome of the cell (e.g., by homologous recombination or integration), such that the encoded CAR is expressed from the genome. In some embodiments, the CAR is expressed episomally. In some embodiments, the nucleotide sequence encoding the CAR does not comprise an intron. Transcription of mammalian genes generally yields RNA containing multiple exons of a coding sequence that are separated by intervening regions (introns), and processing of this RNA includes RNA splicing to remove introns, yielding an RNA comprising an open reading frame that is capable of being translated by ribosomes and tRNAs to produce the encoded polypeptide. Expression of a gene product from a coding sequence without introns abrogates the need for splicing, thereby allowing more rapid and efficient gene expression from an intron-deficient coding sequence relative to a coding sequence that must be spliced between transcription and translation. In some embodiments, a method comprises contacting a cell with one or more nucleic acids to produce a genetically modified cell. In some embodiments, the cell is contacted with a nucleic acid comprising a nucleotide sequence encoding an anti-CD19 CAR. In some embodiments, the nucleic acid is comprised in a vector (e.g., a viral vector). In some embodiments, the nucleic acid is present in a lipid delivery vehicle. In some embodiments, the nucleic acid is present in a lipid nanoparticle. In some embodiments, the cell is contacted with the anti-CD19 CAR. In some embodiments, the cell is in vitro. In some embodiments, the cell is ex vivo. In some embodiments, the cell is in vivo. In some embodiments, a cell is isolated from a subject, contacted with one or more nucleic acids, and administered to the same subject. In some embodiments, a cell is isolated from a first subject, contacted with one or more nucleic acids, and administered to a second, different subject. In some embodiments, a method comprises contacting a cell with one or more nucleic acids to produce an engineered cell. In some embodiments, contacting the cell with the nucleic acid, protein, or vector, introduces a nucleic acid, protein, or vector into a cell, respectively. In some embodiments, the cell is in vitro. In some embodiments, the cell is ex vivo. In some embodiments, a cell is isolated from a subject, contacted with one or more nucleic acids, and administered to the same subject. In some embodiments, the cell is ex vivo. In some embodiments, a cell is isolated from a first subject, contacted with one or more nucleic acids, and administered to a second, different subject. In some embodiments, the cell is in vivo. In some embodiments, the cell is modified extracorporeally. In some embodiments, the cell is a T cell. In some embodiments, the cell is a CD4+ T cell. In some embodiments, the cell is a T regulatory cell. In some embodiments, an engineered cell is modified to reduce risk of transplant rejection and improve engraftment (e.g., modified to be hypoimmunogenic). An engineered cell modified to be hypoimmunogenic may be less prone to immune rejection (e.g., less prone to immune rejection by about 2.5%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99%) relative to an engineered cell that is not modified to be hypoimmunogenic. Such hypoimmunogenic engineered cells can be used as “off-the-shelf” therapeutics in the treatment or amelioration of autoimmune diseases, allergic diseases, inflammatory diseases, or cancer. Modifications to confer hypoimmunogenicity of an engineered cell may include modifications to the genome of the engineered cell to knock out or reduce expression of genes that promote an inflammatory immune response compared to an engineered cell that has not been modified to be hypoimmunogenic. Such genes may include genes encoding proteins that regulate the expression of major histocompatibility complex (MHC) molecules or human leukocyte antigen (HLA), such as β2-microglobulin (B2M), NLR family CARD domain containing 5 (NLRC5), class II transactivator (CIITA), regulatory factor X5 (RFX5), RFXAP, RFXANK, nuclear transcription factor Y (NF-Y) complex proteins, and / or interferon regulatory factor 1 (IRF-1). An engineered cell may also be modified to have reduced or eliminated expression of one or more HLAs corresponding to MHC class I (MHC- I) or MHC class II (MHC-II), such as HLA-A, HLA-B, and / or HLA-C compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of B2M, NLRC5, CIITA, RFX5, RFXAP, RFXANK, NF-Y complex proteins, IRF-1, HLA-A, HLA-B, and / or HLA-C relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of B2M relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of NLRC5 relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of CIITA relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of RFX5 relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of RFXAP relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of NF-Y complex proteins relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of IRF-1 relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of HLA-A relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of HLA-B relative to an engineered cell that is not modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has reduced expression of HLA-C relative to an engineered cell that is not modified to be hypoimmunogenic. Modifications to confer hypoimmunogenicity of an engineered cell may include modifications to the genome of the engineered cell to knock in or increase expression of genes that promote immune tolerance compared to an engineered cell that has not been modified to be hypoimmunogenic. Such genes may include genes encoding proteins that are tolerogenic or inhibitory factors, such as programmed death-ligand 1 (PD-L1), cytotoxic T- lymphocyte associated protein 4 (CTLA-4), CD47, interleukin (IL)-35, HLA-G, or HLA-E. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of PD-L1, CTLA-4, CD47, IL-35, HLA-G, and / or HLA-E compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of PD-L1 compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of CTLA-4 compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of CD47 compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of IL-35 compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of HLA-G compared to an engineered cell that has not been modified to be hypoimmunogenic. In some embodiments, an engineered cell that has been modified to be hypoimmunogenic has increased expression of HLA-E compared to an engineered cell that has not been modified to be hypoimmunogenic. The skilled artisan will recognize that any modification that reduces or abolishes the expression of pro-inflammatory genes or increases the expression of tolerogenic or inhibitory genes may be made to an engineered cell to promote cell engraftment after transplantation. Nucleic acid molecules may be introduced into a cell using any suitable method to generate an engineered cell. For example, viral or non-viral systems may be used to deliver transgenes into a cell. The non-viral approaches include, but are not limited to, polymer nanoparticles, lipids, calcium phosphate, electroporation / nucleofection or biolistic delivery of DNA-coated microparticles (Matuskova and Durinikova, 2016. Advances in Molecular Retrovirology, InTech). Multiple vectors can be used in accordance with the nucleic acids and methods, depending on whether the DNA is integrated into chromatin of the host cell or not. Retroviral vectors such as those derived from gammaretroviruses or lentiviruses persist in the nucleus as integrated provirus and reproduce with cell division. Other types of vectors (e.g., those derived from herpesviruses or adenoviruses) remain in the cell in the episomal form. In some embodiments, the viral vector is selected from a modified virus derived from a virus selected from the group consisting of a retrovirus, lentivirus, gammavirus, adenovirus, adeno-associated virus, pox virus, alphavirus, and herpes virus. In some embodiments, the vector is a retrovirus, such as a modified gammavirus, lentivirus, murine stem cell virus, moloney murine leukemia virus, bovine leukaemia virus, Rous sarcoma virus, or spumavirus. In some embodiments, the viral vector is a retrovirus. In some embodiments, the viral vector is a lentiviral vector. In some embodiments, the viral vector is a gamma-retroviral vector. In some embodiments, one or more vectors are adeno-associated viral (AAV) vectors. In some embodiments, one or more AAV vectors is an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 vector. In some embodiments, one or more AAV vectors are AAV5 vectors. In some embodiments, one or more AAV vectors are AAV6 vectors. In some embodiments, the polypeptide is the sole polypeptide encoded by the nucleotide sequence, i.e., the nucleic acid molecule of the viral vector does not encode additional different proteins but may comprise additional control elements such as promoters and terminators. Some aspects relate to a composition comprising the viral vector. Some aspects relate to a regulatory T cell comprising any one of the nucleic acid molecules, or the viral vector. In some embodiments, the T cell is a T regulatory cell (Treg), such as a mammalian Treg. In some embodiments, the mammalian Treg expresses any one of the polypeptides. In some embodiments, the mammalian Treg is a human Treg. Some aspects relate to a method of preparing allogeneic (derived from a donor other than the intended recipient) or autologous (derived from the recipient) Tregs (e.g., with a stable Treg phenotype). In some aspects, the method comprises contacting T cells with the first and second donor templates or viral vectors comprising them, thereby editing said T cells and preparing Tregs expressing CARs. In some aspects, the method comprises contacting T cells with the nucleic acid molecule comprising a nucleotide sequence encoding any one of the polypeptides, or a viral vector comprising it, thereby endowing said T cells with a stable repair Treg phenotype, and thus preparing Tregs with a stable repair phenotype. In some embodiments, the T cell is a CD4+ T cell. In some embodiments, the T cell is a CD8+ T cell. In some embodiments, the T cell is an engineered Treg comprising a chimeric antigen receptor (CAR) comprising a CD19-binding domain. Preparation of CD4+ T cells may be performed using any suitable method (e.g., separation of human CD4+ T cells followed by preparation of recombinant retroviral stock and retroviral transduction of human CD4+ T cells). Recombinant retroviral and lentiviral vectors may be prepared and used to transduce cells using any suitable method, such as use of a commercial kit including packaging cells, plasmids and transfection reagents, which are offered by many companies, including Invitrogen®, Sigma®, Clontech®, Cell Biolabs®, SBI®, Genecopoeia® and many others. The methods are thus performed along with the guidelines supplied with the commercial kits. In short, according to a non-limiting example taught by the γ-Retrovirus Guide of Addgene, the following components are used: (a) γ-Retroviral transfer plasmid encoding a transgene of interest: The transgene sequence is flanked by long terminal repeat (LTR) sequences, which facilitate integration of the transfer plasmid sequences into the host genome. Typically, it is the sequences between and including the LTRs that is integrated into the host genome upon viral transduction; (b) Packaging genes (viral Gag-Pol): Gag is a structural precursor protein, and Pol is a polymerase; and (c) Envelope gene (may be pseudotyped to alter infectivity). As a non-limiting example, the three components described above (envelope, packaging, and transfer) are supplied by three types of plasmids, which are co-transfected into a 293T packaging cell line. This system allows for flexibility to pseudotype γ-retrovirus using different envelopes to modify tropism. Different envelope plasmids can direct the production of virus with various tropisms. As an example, recombinant retroviral stock may be prepared by cloning, taking the resulting plasmid as well as a plasmid carrying gag / pol and a plasmid carrying env, and transfecting HEK293T cells. In some embodiments, the plasmid may comprise any combination of a nucleotide sequence encoding a protein (mem-IL-33, ST2, IL1RAP, IL- 18R1, and / or IL18RAP), Gag / Pol, and Env. A further non-limiting example of methods for transducing human cells (e.g., CD4+ T cells), includes combining the viral supernatant with a transfection reagent (e.g., Polybrene (Merck®)), adding the composition to RetroNectin®- (Takara®) coated wells and spinning down. The resulting supernatant is then removed and CD4+ T cells are added and placed in an incubator for transfection. In some embodiments, the cells are T cells. In some embodiments, the cells are CD4+ T cells. In some embodiments, the cells are Tregs. In some embodiments, the cells are engineered Tregs comprising a CAR comprising a CD19-binding domain. In some embodiments, the cells are ex vivo. In some embodiments, the cells are in vivo. In some embodiments, the cells are extracorporeal. Homology arms Donor templates for insertion into FOXP3 gene and second gene loci comprise 5′ and 3′ homology arms, to target insertion of the nucleic acid into the FOXP3 and second gene loci, respectively, by homology-directed repair following introduction of a double-stranded break. Typically, the 5′ homology arm refers to a homology arm at the 5′ end of the nucleic acid, and 3′ homology arm refers to another homology arm at the 3′ end of the nucleic acid, when considering the coding strand of the nucleic acid. The 5′ homology arm will have homology to a first sequence in the targeted locus, and the 3′ homology arm will have homology to a second sequence in the targeted locus that is downstream from the first sequence in the targeted locus, such that the nucleic acid is inserted into the locus in a targeted manner. Following insertion, the modified locus will comprise the homology arms, in place of the first and second sequences in the targeted locus, and the sequence between the homology arms on the nucleic acid, in place of the sequence that was previously present between the first and second sequences in the targeted locus. The homology arms may be the same length, have similar lengths (within 100 bp of each other), or different lengths. In some embodiments, one or both homology arms have a length of 100–2,000 bp, 200–2,000 bp, 400–1,500 bp, 500–1,000 bp. In some embodiments, one or both homology arms are about 100 bp, about 200 bp, about 300 bp, about 400 bp, about 500 bp, about 600 bp, about 700 bp, about 800 bp, about 900 bp, about 1,000 bp, about 1,100 bp, about 1,200 bp, about 1,300 bp, about 1,400 bp, about 1,500 bp, about 1,600 bp, about 1,700 bp, about 1,800 bp, about 1,900 bp, or about 2,000 bp. In some embodiments, both homology arms are 100-2,000 nucleotides in length. In some embodiments, both homology arms are 300–1,000 nucleotides in length. In some embodiments, both homology arms are 300–700 nucleotides in length. In some embodiments, both homology arms are 300–500 nucleotides in length. In some embodiments, both homology arms are 500–700 nucleotides in length. In some embodiments, both homology arms are 700–1,000 nucleotides in length. Homology arms of a nucleic acid for insertion at a targeted genomic locus may be chosen based on homologous sequences in the targeted locus that are upstream and / or downstream from a site targeted for cleavage by a DNA endonuclease. For example, in some embodiments for insertion by homology-directed repair following cleavage at a given position (cleavage site) in the targeted locus, the 5′ homology arm of a nucleic acid for insertion has homology to a sequence upstream of the cleavage site, and the 3′ homology arm of the nucleic acid has homology to a sequence downstream of the cleavage site. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75–2,000, 100–1,000, 150–500 nucleotides upstream from the cleavage site. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75–2,000, 100– 1,000, 150–500 nucleotides upstream from a PAM sequence cleaved by an RNA-guided DNA endonuclease. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75–2,000, 100–1,000, 150– 500 nucleotides upstream from a sequence in the genome that is complementary to a spacer sequence of a gRNA. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a cleavage site. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a PAM sequence cleaved by an RNA-guided DNA endonuclease. In some embodiments, the 5′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a sequence in the genome that is complementary to a spacer sequence of a gRNA. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75–2,000, 100–1,000, 150–500 nucleotides upstream from the cleavage site. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75– 2,000, 100–1,000, 150–500 nucleotides upstream from a PAM sequence cleaved by an RNA- guided DNA endonuclease. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends 25–5,000, 50–3,000, 75–2,000, 100– 1,000, 150–500 nucleotides upstream from a sequence in the genome that is complementary to a spacer sequence of a gRNA. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a cleavage site. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a PAM sequence cleaved by an RNA-guided DNA endonuclease. In some embodiments, the 3′ homology arm has homology to a sequence 100–2,000 nucleotides in length that ends at a position 150–500 nucleotides upstream from a sequence in the genome that is complementary to a spacer sequence of a gRNA. In some embodiments, where a method includes a gRNA comprising a spacer sequence, neither the 5′ nor the 3′ homology arm of a nucleic acid for genomic insertion comprises a sequence that is complementary to the spacer sequence. In such embodiments, lack of a complementary sequence on the donor template reduces the chance of the gRNA binding to the donor template and mediating cleavage, which can reduce the efficiency of genomic insertion. In some embodiments, the donor template does not comprise a sequence that is complementary to the spacer sequence. In embodiments where a different DNA endonuclease that does not require a gRNA for targeted cleavage is used, the donor template does not comprise a sequence that is cleaved by the DNA endonuclease. Cell types Embodiments of methods for producing engineered cells (e.g., by in vitro or ex vivo gene editing, and / or administration of compositions, vectors, or nucleic acids to a subject for in vivo editing) may use any suitable cell type as a material for, e.g., introduction of nucleic acids, vectors, and / or compositions. It is to be understood that methods that comprise manipulation of CD4+ cells can be applied to other types of cells (e.g., CD8+ cells). In some embodiments, the method comprises editing an immune cell. Non-limiting examples of immune cells include B cells, T cells, and NK cells. In some embodiments, the method comprises editing a CD3+ T cell. In some embodiments, the method comprises editing a CD4+ T cell. In some embodiments, the method comprises editing a CD8+ T cell. In some embodiments, the method comprises editing an NK1.1+ T cell. In some embodiments, the method comprises editing a stem cell. In some embodiments, the method comprises editing a pluripotent stem cell. In some embodiments, the method comprises editing CD34+ hematopoietic stem cells (HSCs). In some embodiments, the method comprises editing induced pluripotent stem cells (iPSCs). Edited stem cells may be matured in vitro to produce Treg cells or administered to a subject to allow in vivo development into Treg cells. Edited stem cells may be matured into CD3+ Treg cells, CD4+ Treg cells, CD8+ Treg cells, NK1.1+ Treg cells, or a combination thereof. In some embodiments, a method comprises editing a T cell. A T cell or T lymphocyte is an immune system cell that matures in the thymus and produces a T cell receptor (TCR), e.g., an antigen-specific heterodimeric cell surface receptor typically comprised of an α-β heterodimer or a γ-δ heterodimer. T cells of a given clonality typically express only a single TCR clonotype that recognizes a specific antigenic epitope presented by a syngeneic antigen- presenting cell in the context of a major histocompatibility complex-encoded determinant. T cells can be naïve ("TN"; not exposed to antigen; increased expression of CD62L, CCR7, CD28, CD3, CD127, and CD45RA, and decreased or no expression of CD45RO as compared to TCM (described herein)), memory T cells (TM) (antigen experienced and long-lived), including stem cell memory T cells, and effector cells (antigen-experienced, cytotoxic). TM can be further divided into subsets of central memory T cells (TCM, expresses CD62L, CCR7, CD28, CD95, CD45RO, and CD127) and effector memory T cells (TEM, express CD45RO, decreased expression of CD62L, CCR7, CD28, and CD45RA). Effector T cells (TE) refer to antigen-experienced CD8+ cytotoxic T lymphocytes that express CD45RA, have decreased expression of CD62L, CCR7, and CD28 as compared to TCM, and are positive for granzyme and perforin. Helper T cells (TH) are CD4+ cells that influence the activity of other immune cells by releasing cytokines. CD4+ T cells can activate and suppress an adaptive immune response, and which of those two functions is induced will depend on the presence of other cells and signals. T cells can be collected using known techniques, and the various subpopulations or combinations thereof can be enriched or depleted by known techniques, for example, using antibodies that specifically recognize one or more T cell surface phenotypic markers, by affinity binding to antibodies, flow cytometry, fluorescence activated cell sorting (FACS), or immunomagnetic bead selection. Other exemplary T cells include regulatory T cells (Tregs, also known as suppressor T cells), such as CD4+ CD25+ (FoxP3+) regulatory T cells and Treg17 cells, as well as Tr1, Th3, CD8+CD28-, or Qa-1-restricted T cells. In some embodiments, the cell is a CD3+, CD4+, and / or CD8+ T cell. In some embodiments, the cell is a CD3+ T cell. In some embodiments, the cell is a CD4+CD8– T cell. In some embodiments, the Treg cell is a FoxP3+ Treg cell. In some embodiments, the Treg cell expresses CTLA-4, LAG-3, CD25, CD39, CD27, CD70, CD357 (GITR), neuropilin-1, galectin-1, and / or IL-2Rα on its surface. In some embodiments, the Treg is an engineered Treg; that is, a Treg that has been manipulated so that its genetic, epigenetic, and / or phenotypic identity is altered relative to an appropriate reference cell (such as an otherwise identical cell that has not been so manipulated). For example, an engineered Treg may be one that is engineered for constitutive expression of FOXP3. In some embodiments, the engineered Treg is a sorted engineered Treg. In some embodiments, a sorted T cell is engineered to express a CAR. In some embodiments, the Treg is not a sorted Treg, wherein a sorted Treg is isolated from a human sample based on the following cell surface markers: CD4+CD25hiCD127lo or CD4+CD25hiCD127loCD45RA+. In some embodiments, the Treg is an induced Treg (iTreg) or a peripheral Treg (pTreg); as used herein, an iTreg and a pTreg refer to a stem cell that has been stimulated (e.g., in the presence of TGF-β and IL-2), in vitro and in vivo, respectively. In some embodiments, the engineered Treg comprises a Treg engineered to express FOXP3, first and second CISC components, and a CAR (e.g., an anti-CD19 CAR) constitutively. In some embodiments, the cell is a human cell. In some embodiments, the cell is isolated from a biological sample. A biological sample may be a sample from a subject (e.g., a human subject) or a composition produced in a lab (e.g., a culture of cells). A biological sample obtained from a subject may be a liquid sample (e.g., blood or a fraction thereof, a bronchial lavage, cerebrospinal fluid, or urine), or a solid sample (e.g., a piece of tissue). In some embodiments, the cell is obtained from peripheral blood. In some embodiments, the cell is obtained from umbilical cord blood. In some embodiments, the cell is obtained by sorting cells of peripheral blood to obtain a desired cell population (e.g., CD3+ cells), and one or more cells of the sorted population are engineered. In some embodiments, the cell is in a subject. In some embodiments, the cell is in vivo. Targeted loci Nucleic acids comprising a coding sequence (e.g., a nucleotide sequence encoding a CAR) may be inserted into a targeted locus, such that a population of genetically modified cells contain the inserted sequences at a consistent location of the genome. Such consistency is useful, for example, in screening cells and cell populations by analyzing the targeted locus (e.g., by PCR amplification of genomic DNA using primers flanking the insertion site). In some embodiments, the nucleic acid comprising a heterologous promoter that is introduced into the cell is inserted at a targeted locus. In some embodiments, the sequence inserted into the targeted locus may replace all or part of the endogenous coding sequence in the targeted locus. In some embodiments, one or more mutations (e.g., nonsense mutation) is introduced into the endogenous coding sequence to prevent translation of a full-length polypeptide from the endogenous coding sequence. In some embodiments, all or part of the endogenous coding sequence is removed from the genome by insertion of the coding sequence on the inserted nucleic acid. In some embodiments, the targeted locus is a safe harbor locus. In some embodiments, the safe harbor locus is a HIPP11 locus. In some embodiments, the safe harbor locus is a ROSA26 locus. In some embodiments, the safe harbor locus is an AAVS1 locus. In some embodiments, the targeted locus is a T cell receptor locus. In some embodiments, the T cell receptor locus is a TRAC locus. In some embodiments, the T cell receptor locus is a TRBC locus. In some embodiments, the targeted locus is a CD3Z locus. In some embodiments, nucleic acids may be integrated in a non-targeted manner (e.g., by use of a lentiviral vector), such that a population of genetically modified cells contains diverse integration sites. In some embodiments, the nucleic acid, or vector comprising such a nucleic acid, is not integrated into the genome of the cell. For example, a plasmid or artificial chromosome (e.g., human artificial chromosome) may be introduced into the cell, with the heterologous promoter driving transcription of the operably linked coding sequence from the plasmid or artificial chromosome, without integration of the vector into the chromosome. In some embodiments, the introduced vector or nucleic acid replicates independently of endogenous chromosomes. In some embodiments, more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 copies of the vector are present in a genetically modified cell. In some embodiments, the number of copies of the vector in a cell exceeds the copy number of an individual chromosome in a cell. Delivery of nucleic acids Some aspects relate to methods of engineering cells to express a CAR (e.g., comprising a CD19-binding domain) and FOXP3 (or a functional fragment thereof) by contacting cells with a lipid delivery vehicle comprising one or more nucleic acids that collectively encode the CAR and FOXP3. Expression of both FOXP3 and CAR components is expected to provide a regulatory T cell phenotype and allow targeting of CD19+cells by such engineered regulatory T cells. In some embodiments, the one or more nucleic acids further collectively encode a first CISC component and a second CISC component. In some embodiments, the cells are contacted with the lipid delivery vehicle ex vivo. In some embodiments, the cells are contacted with the lipid delivery vehicle in vivo. Some aspects relate to lipid delivery vehicles comprising nucleic acids that collectively encode FOXP3 (or a functional fragment thereof) and a CAR (e.g., comprising a CD19-binding domain). Some embodiments relate to a lipid delivery vehicle conjugated to a T cell targeting moiety, wherein the lipid delivery vehicle comprises one or more nucleic acids that collectively encode (i) FOXP3; a nucleotide sequence encoding a first CISC component; a nucleotide sequence encoding a second CISC component; (ii) a CAR (e.g., comprising a CD19-binding domain); (iii) a first CISC component; and (iv) a second CISC component. Any suitable nucleic acid or nucleic acids may be used to encode FOXP3 and the CAR. In some embodiments, FOXP3 and the CAR are encoded by the same nucleic acid. In some embodiments, a lipid delivery vehicle comprises two or more nucleic acids, where each of the CAR and FOXP3 are encoded by different nucleic acids. In some embodiments, a first lipid delivery vehicle comprises a nucleic acid encoding the CAR, and a second lipid delivery vehicle comprises a nucleic acid encoding FOXP3. In some embodiments, a lipid delivery vehicle further comprises one or more nucleic acids encoding a first CISC component and a second CISC component. In some embodiments, the nucleic acid or nucleic acids are DNA. In some embodiments, the nucleic acid or nucleic acids are RNA. In preferred embodiments, an RNA is a messenger RNA (mRNA). mRNA is RNA that encodes at least one protein or a fragment thereof and can be translated to produce an encoded protein (e.g., FOXP3) or fragment in vitro, in vivo, in situ, or ex vivo. mRNA comprises an open reading frame (ORF) encoding a protein or fragment thereof. In some embodiments, the mRNA further comprises a 5^ untranslated region (UTR), 3^ UTR, a polyA tail, and / or a 5^ cap analog. In some embodiments, an RNA is a self-amplifying RNA. A self-amplifying RNA is an RNA encoding one or more proteins that, individually or in conjunction, are capable of replicating the self-amplifying RNA. In some embodiments, the proteins encoded by the self- amplifying RNA are non-structural proteins nsP1, nsP2, nsP3, and nsP4, which form an RNA-dependent RNA polymerase (RdRp), or replicase, that is capable of replicating the self- amplifying RNA. By encoding proteins that are capable of replicating the RNA, a self- amplifying RNA is capable of self-amplification in a cell. A self-amplifying RNA is also known as an RNA replicon. In some embodiments, an RNA is a circular RNA. A circular RNA is an RNA with no 5′ terminal nucleotide or 3′ terminal nucleotide. Every nucleotide in a circular RNA is covalently bonded to both (1) a 5′ adjacent nucleotide; and (2) a 3′ adjacent nucleotide. In a circular RNA with a nucleotide sequence comprising every nucleotide of the circular RNA in 5′-to-3′ order, the last nucleotide of the nucleotide sequence is covalently bound to the first nucleotide of the nucleotide sequence. In some embodiments, the lipid delivery vehicle is an exosome or extracellular vesicle. In some embodiments, a lipid delivery vehicle is a liposome. A liposome is a lipid particle comprising lipids arranged in one or more concentric lipid bilayers around a central region (e.g., an aqueous solution, suspension, or other aqueous composition). In some embodiments, a lipid delivery vehicle is a lipoplex. A lipoplex is a lipid particle comprising a cationic liposome and a nucleic acid (e.g., mRNA). Lipoplexes can be formed by contacting a liposome comprising a cationic lipid with a nucleic acid. In some embodiments, a lipoplex comprises multiple concentric lipid bilayers, wherein each concentric bilayer is separated by one or more nucleic acids. In some embodiments, the central region of the lipoplex is an aqueous solution, suspension, or other aqueous composition. In some embodiments, a lipid delivery vehicle is a lipid nanoparticle (LNP). LNPs typically comprise ionizable lipid (e.g., ionizable amino lipid), non-cationic lipid (e.g., phospholipid), structural lipid (e.g., sterol), and PEG-modified lipid components, along with the nucleic acid cargo of interest. Any suitable T cell targeting moiety can be used to direct a lipid delivery vehicle to a T cell for nucleic acid delivery. Non-limiting examples of T cell targeting moieties include antibodies against proteins expressed by T cells and natural ligands of proteins expressed by T cells. Antibodies targeting a specific T cell antigen (e.g., CD4 or CD8) will be understood to target the form expressed on T cells of the species of interest (e.g., human CD4 in the context of engineering human cells). In some embodiments, a T cell targeting moiety is an antibody or antigen-binding fragment thereof. In some embodiments, a T cell targeting moiety is an anti-CD3 antibody (e.g., an anti-CD3ε antibody, an anti-CD3γ antibody, an anti- CD3δ antibody, an anti-CD3ε / γ antibody, or an anti-CD3ε / δ antibody) (e.g., OKT3), an anti- CD4 antibody (e.g., ibalizumab), and anti-CD8 antibody, or an anti-CD25 antibody (e.g., daclizumab), or antigen-binding fragment of any aforementioned antibody. In some embodiments, a T cell targeting moiety is an anti-CD3 antibody or antigen-binding fragment thereof. In some embodiments, a T cell targeting moiety is an anti-CD4 antibody or antigen- binding fragment thereof. In some embodiments, a T cell targeting moiety is an anti-CD8 antibody or antigen-binding fragment thereof. Non-limiting examples of anti-CD3 antibodies include OKT3, muromonab, teplizumab, otelixizumab, visilizumab, and foralumab. Non- limiting examples of anti-CD4 antibodies include zanolimumab, keliximab, clenoliximab, tregalizumab, and ibalizumab. Non-limiting examples of anti-CD28 antibodies include theralizumab and FR104. Non-limiting examples of anti-CD25 antibodies include daclizumab and basiliximab. Non-antibody moieties that interact with T cells or their surface markers may be used to target lipid delivery vehicles to T cells. In some embodiments, a T cell targeting moiety is an HIV glycoprotein that binds to CD4. In some embodiments, a T cell targeting moiety is HIV glycoprotein 120 (gp120), which binds to CD4. In some embodiments, a T cell targeting moiety is IL-2, which binds to CD25 (also called IL-2R) (e.g., IL-2Rα, IL-2Rβ, and / or IL- 2Rγ). In some embodiments, a T cell targeting moiety is a CD25-binding fragment of IL-2. In some embodiments, a T cell targeting moiety is a peptide-major histocompatibility complex (MHC) molecule, which can be bound by T cell receptor (TCR) alpha (TCRα) and beta (TCRβ) of the CD3 / TCR complex. In some embodiments, a T cell targeting moiety is an MHC Class I molecule, which can be bound by CD8 (e.g., CD8α and / or CD8β). In some embodiments, a T cell targeting moiety is IL-16, which binds to CD4 (e.g., the D4 domain of CD4). In some embodiments, a T cell targeting moiety is a CD4-binding fragment of IL-16. T cell targeting moieties may be attached to a lipid delivery vehicle (e.g., an LNP) in any suitable manner (see, e.g., Tombácz et al. Mol Ther, 2021.29(11): 3293-3304 and Marques et al. Pharmaceutics, 2023.15(1): 216). Chemical processes by which a T cell targeting moiety (e.g., an antibody) can be conjugated to a lipid delivery vehicle (e.g., an LNP) include adsorption, covalent conjugation (e.g., carbodiimide, maleimide, or “click” chemistry), and avidin / streptavidin-biotin interactions. In some embodiments, a lipid delivery vehicle comprises structural modifications that facilitate conjugation to a T cell targeting moiety. In some embodiments, a T cell targeting moiety is conjugated to a lipid delivery vehicle via a thioether linkage (e.g., a T cell targeting antibody is reduced or thiolated to produce free sulfhydryl groups and a lipid delivery vehicle comprises maleimide-activated amino groups with which the sulfhydryl groups bond). In some embodiments, a T cell targeting moiety comprises avidin or streptavidin and a lipid delivery vehicle comprises biotin. In some embodiments, a T cell targeting moiety comprises biotin and a lipid delivery vehicle comprises avidin or streptavidin. In some embodiments, the Fc region of an antibody (e.g., an anti-CD3 antibody) is conjugated to a lipid delivery vehicle. In some embodiments, a T cell targeting moiety is covalently bound to a lipid delivery vehicle. In some embodiments, a T cell targeting moiety is conjugated to a lipid delivery vehicle (e.g., an LNP) via a linker. In some embodiments, a linker is a glycine-serine linker. In some embodiments, a linker is a non-cleavable linker. In some embodiments, a linker is a cleavable linker. In some embodiments, the linker comprises 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids, such as glycines, or a number of amino acids, such as glycine, within a range defined by any two of the aforementioned numbers. In some embodiments, the glycine spacer comprises at least 3 glycines. In some embodiments, the glycine spacer comprises a sequence set forth as GGGS (SEQ ID NO: 24), GGGSGGG (SEQ ID NO: 25) or GGG. Nucleases and guide RNAs Some aspects relate to the use of nucleases (e.g., DNA endonucleases) to introduce a double-stranded break into nucleic acid of a cell genome and edit the genome at a desired locus (e.g., to promote integration of a donor template at the locus by homology-directed repair and / or inactivate a targeted gene). Any one of multiple gene- or genome- editing methods can used to accomplish editing of one or more loci (e.g., FOXP3, TRAC, TRBC, CD3Z, AAVS1, and / or HIPP11). Non-limiting examples of gene editing methods include use of a DNA endonuclease such as an RNA-guided DNA endonuclease (e.g., Cas (e.g., Cas9)), zinc finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), or meganuclease; transposon-mediated gene editing; serine integrase-mediated gene editing; and lentivirus-mediated gene editing. In some embodiments, a gene editing method comprises knocking out or inactivating an endogenous gene, such as by producing a chromosomal gene knockout in the genome. As used herein, the term "chromosomal gene knockout" refers to a genetic alteration, inactivation, or introduced inhibitory agent in a host cell that prevents (e.g., reduces, delays, suppresses, or abrogates) production, by the host cell, of a functionally active endogenous polypeptide product. Alterations resulting in a chromosomal gene knockout or inactivation can include, for example, introduced nonsense mutations (including the formation of premature stop codons), missense mutations, gene deletion, or strand breaks. In certain embodiments, a chromosomal gene knock-out or gene knock-in (e.g., insertion) is made by chromosomal editing of a host cell. Chromosomal editing can be performed using, for example, endonucleases. As used herein "endonuclease" refers to an enzyme capable of catalyzing cleavage of a phosphodiester bond within a polynucleotide chain. A DNA endonuclease refers to an endonuclease that is capable of catalyzing cleavage of a phosphodiester bond within a DNA polynucleotide. In certain embodiments, an endonuclease is capable of cleaving a nucleic acid sequence in a targeted gene, thereby inactivating or "knocking out" the targeted gene. In some embodiments, an endonuclease is capable of cleaving a nucleic acid sequence in a targeted locus, promoting insertion of an exogenous nucleic acid sequence into the targeted locus by homologous recombination. An endonuclease may be a naturally occurring, recombinant, genetically modified, or fusion endonuclease. Examples of endonucleases for use in gene editing include zinc finger nucleases (ZFN), TALE-nucleases (TALEN), RNA-guided nucleases, CRISPR-Cas nucleases, meganucleases, or megaTALs. The nucleic acid strand breaks caused by DNA endonucleases are typically double- strand breaks (DSB), which may be commonly repaired through the distinct mechanisms of homology directed repair (HDR) by homologous recombination, or by non-homologous end joining (NHEJ). (NHEJ: Ghezraoui et al., 2014 Mol Cell 55(6):829-842; HDR: Jasin and Rothstein, 2013 Cold Spring Harb Perspect Biol 5(11):a012740, PMID 24097900). During HDR / homologous recombination, a donor nucleic acid molecule may be used for a donor gene "knock-in", for target gene "knock-out", and optionally to inactivate a target gene through a donor gene knock in or target gene knock out event. NHEJ is an error-prone repair process that often results in changes to the DNA sequence at the site of the cleavage, e.g., a substitution, deletion, or addition of at least one nucleotide. NHEJ may be used to "knock- out" a target gene. HDR is favored by the presence of a donor template at the time of DSB formation. As used herein, a "zinc finger nuclease" (ZFN) refers to a fusion protein comprising a zinc finger DNA-binding domain fused to a non-specific DNA cleavage domain, such as a Fokl endonuclease. Each zinc finger motif of about 30 amino acids binds to about 3 base pairs of DNA, and amino acids at certain residues can be changed to alter triplet sequence specificity (see, e.g., Desjarlais et al., Proc Natl Acad Sci U S A.90:2256-2260, 1993; Wolfe et al., J Mol Biol.285:1917-1934, 1999). Multiple zinc finger motifs can be linked in tandem to create binding specificity to desired DNA sequences, such as regions having a length ranging from about 9 to about 18 base pairs. By way of background, ZFNs mediate genome editing by catalyzing the formation of a site-specific DNA double strand break (DSB) in the genome, and targeted integration of a transgene comprising flanking sequences homologous to the genome at the site of DSB is facilitated by homology directed repair (HDR). Alternatively, a DSB generated by a ZFN can result in knock out of target gene via repair by non-homologous end joining (NHEJ), which is an error-prone cellular repair pathway that results in the insertion or deletion of nucleotides at the cleavage site. In certain embodiments, a gene knockout or inactivation comprises an insertion, a deletion, a mutation or a combination thereof, made using a ZFN molecule. As used herein, a "transcription activator-like effector nuclease" (TALEN) refers to a fusion protein comprising a TALE DNA-binding domain and a DNA cleavage domain, such as a FokI endonuclease. A "TALE DNA binding domain" or "TALE" is composed of one or more TALE repeat domains / units, each generally having a highly conserved 33-35 amino acid sequence with divergent 12th and 13th amino acids. The TALE repeat domains are involved in binding of the TALE to a target DNA sequence. The divergent amino acid residues, referred to as the Repeat Variable Diresidue (RVD), correlate with specific nucleotide recognition. The natural (canonical) code for DNA recognition of these TALEs has been determined such that an HD (histidine-aspartic acid) sequence at positions 12 and 13 of the TALE leads to the TALE binding to cytosine (C), NG (asparagine-glycine) binds to a T nucleotide, NI (asparagine-isoleucine) to A, NN (asparagine-asparagine) binds to a G or A nucleotide, and NG (asparagine-glycine) binds to a T nucleotide. Non-canonical (atypical) RVDs are also known (see, e.g., U.S. Patent Publication No. US 2011 / 0301073, which atypical RVDs are incorporated by reference herein for this purpose). TALENs can be used to direct site-specific double-strand breaks (DSB) in the genome of T cells. Non- homologous end joining (NHEJ) ligates DNA from both sides of a double-strand break in which there is little or no sequence overlap for annealing, thereby introducing errors that knock out gene expression. Alternatively, homology directed repair (HDR) can introduce a transgene at the site of DSB providing homologous flanking sequences are present in the donor template containing the transgene. In certain embodiments, a gene knockout comprises an insertion, a deletion, a mutation or a combination thereof, and made using a TALEN molecule. Gene-editing systems and methods may make use of viral or non-viral vectors or cassettes, as well as nucleases that allow site-specific or locus-specific gene-editing, such as RNA-guided nucleases, Cas nucleases (e.g., Cpf1 or Cas9 nucleases), meganucleases, TALENs, or ZFNs. Certain RNA-guided nucleases useful with some embodiments are disclosed in U.S. Patent No.11,162,114, which is expressly incorporated by reference herein for this purpose. Non-limiting examples of Cas nucleases include SpCas9, SaCas9, CjCas9, xCas9, C2c1, Cas13a / C2c2, C2c3, Cas13b, Cpf1, and variants thereof. Certain features useful with some embodiments are disclosed in WO 2019 / 210057, which is expressly incorporated by reference in its entirety. As used herein, a "clustered regularly interspaced short palindromic repeats / Cas" (CRISPR / Cas, or Cas) nuclease system refers to a system that employs a CRISPR RNA (crRNA)-guided Cas nuclease to recognize target sites within a genome (known as protospacers) via base-pairing complementarity and then to cleave the DNA if a short, conserved protospacer associated motif (PAM) immediately follows 3′ of the complementary target sequence. CRISPR / Cas systems are classified into types (e.g., type I, type II, type III, and type V) based on the sequence and structure of the Cas nucleases. The crRNA-guided surveillance complexes in types I and III need multiple Cas subunits. The Type II system, the most studied, comprises at least three components: an RNA-guided Cas9 nuclease, a crRNA, and a trans-acting crRNA (tracrRNA). The tracrRNA comprises a duplex forming region. A crRNA and a tracrRNA form a duplex that is capable of interacting with a Cas9 nuclease and guiding the Cas9 / crRNA:tracrRNA complex to a specific site on the target DNA via Watson- Crick base-pairing between the spacer on the crRNA and the protospacer on the target DNA upstream from a PAM. Cas9 nuclease cleaves a double-stranded break within a region defined by the crRNA spacer. Repair by NHEJ results in insertions and / or deletions which disrupt expression of the targeted locus. Alternatively, a donor template transgene with homologous flanking sequences can be introduced at the site of DSB via homology directed repair (HDR). The crRNA and tracrRNA can be engineered into a single guide RNA (sgRNA or gRNA) (see, e.g., Jinek et al., Science 337:816-21, 2012). Further, the region of the guide RNA complementary to the target site can be altered or programed to target a desired sequence (Xie et al., PLOS One 9:e100448, 2014; U.S. Pat. Appl. Pub. No. US 2014 / 0068797, U.S. Pat. Appl. Pub. No. US 2014 / 0186843; U.S. Pat. No.8,697,359, and PCT Publication No. WO 2015 / 071474; each of which is incorporated by reference). Non- limiting examples of CRISPR / Cas nucleases include Cas9, SaCas9, CjCas9, xCas9, C2C1, Cas13a / C2c2, C2c3, Cas13b, Cpf1, and variants thereof. In some embodiments, a gene knockout or inactivation comprises an insertion, a deletion, a mutation or a combination thereof, and made using an RNA-guided nuclease. Exemplary gRNA sequences and methods of using the same to knock out endogenous genes that encode immune cell proteins include those described in Ren et al., Clin Cancer Res. 2017.23(9):2255-2266, the gRNAs, Cas9 DNAs, vectors, and gene knockout techniques of which are hereby expressly incorporated by reference for this purpose. In some embodiments, a gene modification comprises an insertion of an exogenous nucleic acid sequence (e.g., promoter, transgene, and / or combinations thereof) into the genome of a cell, where an RNA-guided nuclease introduces a double-stranded break in the genome and the exogenous nucleic acid sequence is introduced into the genome by homology-directed repair. In some embodiments, a gRNA comprises a spacer sequence that is complementary to a nucleotide sequence in a FOXP3 locus. In some embodiments, a gRNA comprises a spacer sequence that is complementary to a nucleotide sequence in a TRAC locus. Examples of spacer sequences targeting the FOXP3 locus are provided as SEQ ID NOs.251–259, and examples of spacer sequences targeting the TRAC locus are provided as SEQ ID NOs.261– 263. Cas9 is an example of an RNA-guided DNA endonuclease that cleaves at an NGG PAM. Examples of RNA-guided DNA endonucleases that cleave at an NNNNCC PAM are described, e.g., in International Application No. WO 2019 / 236566, incorporated by reference herein for this purpose. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NGG protospacer adjacent motif (e.g., Cas9) comprises a spacer sequence of SEQ ID NO.251, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.251. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NGG protospacer adjacent motif (e.g., Cas9) comprises a spacer sequence of SEQ ID NO.252, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.252. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NGG protospacer adjacent motif (e.g., Cas9) comprises a spacer sequence of SEQ ID NO.253, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.253. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NGG protospacer adjacent motif (e.g., Cas9) comprises a spacer sequence of SEQ ID NO.254, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.254. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.255, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.255. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.256, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.256. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.257, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.257. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.258, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.258. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.259, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.259. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NGG protospacer adjacent motif (e.g., Cas9) comprises a spacer sequence of SEQ ID NO.261, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.261. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.262, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.262. In some embodiments, a gRNA for use with an RNA-guided DNA endonuclease that cleaves at an NNNNCC protospacer adjacent motif (e.g., APG07433.1 or APG08290.1) comprises a spacer sequence of SEQ ID NO.263, or a spacer sequence having no more than 1, 2, or 3 mismatches thereto. In some embodiments, the gRNA comprises a spacer sequence of SEQ ID NO.263. Selection by Essential Gene Exon Knock-In (SLEEK) Some embodiments of engineered cells comprise an inserted nucleic acid encoding at least a portion of an essential gene inserted into a locus comprising the essential gene. Some embodiments of methods of editing an engineered cell comprise inserting a nucleic acid encoding a portion of an essential gene into a locus comprising the essential gene. Some embodiments of nucleic acids comprise a nucleotide sequence encoding at least a portion of an essential gene and homology arms that direct insertion of the nucleic acid into a locus comprising the essential gene. Selection by Essential-gene Exon Knock-in (SLEEK) is described, for example, in Allen et al., Nat Biotechnol.2023. doi: 10.1038 / s41587-023- 01779-8. Insertion of at least a portion of an essential gene (e.g., an exon of an essential gene) allows selection to address the preference of some cells for non-homologous end joining (NHEJ) over homology-directed repair (HDR). As used herein, an “essential gene” refers to a gene with no alternatives in the cell, that could be expressed to compensate for loss of expression by the essential gene, where failure to express the product of the essential gene (i) inhibits cellular replication, (ii) results in cell death, and / or (iii) in a Treg cell, reduces in loss of the Treg cell phenotype (e.g., loss of FOXP3 expression). Essential genes may be those involved in key cellular processes such as central metabolism or transcription. In some embodiments, an essential gene is associated with central metabolism. In some embodiments, an essential gene is associated with transcription. In some embodiments, an essential gene is associated with protein folding. In some embodiments, an essential gene is associated with translocation. In some embodiments, an essential gene is associated with quality control. In some embodiments, an essential gene encodes a chaperone protein. In some embodiments, an essential gene encodes a cytoskeletal protein. In some embodiments, an essential gene encodes a cytoskeletal factor. In some embodiments, an essential gene encodes a protein involved in DNA repair. In some embodiments, an essential gene is GAPDH. In some embodiments, an essential gene is KIF11. In some embodiments, an essential gene is TBP. The cells, methods, and / or nucleic acids described in this section involve targeted genome editing, such as editing using a targeted nuclease (e.g., RNA-guided nuclease (e.g., Cas9), TALEN, ZFN, and / or meganuclease). Targeted chromosomal cleavage by a nuclease (e.g., a DNA endonuclease) at an essential gene locus allows disruption of the essential gene, if a corresponding correction (e.g., insertion of a donor nucleic acid compensating for the disruption) does not occur. For example, cleavage at a glyceraldehyde 3-phosphate dehydrogenase (GAPDH) locus, and consequent indel introduction by DNA repair (e.g., by NHEJ) may prevent production of functional GAPDH, leading to death of cells that fail to express this essential protein. But, where a donor nucleic acid targeting an essential gene locus, and comprising a nucleotide sequence encoding at least a portion of the essential gene, is inserted into the chromosome at the cleaved locus by homology-directed repair, restored expression of the essential gene avoids such cell death. This editing approach thus allows for highly efficient introduction of one or more transgenes into a cell, by insertion of a nucleic acid at an essential gene locus, where the nucleic acid comprises (i) the transgene(s), and (ii) at least a portion of the essential gene, such that expression of the essential gene occurs in cells comprising the inserted nucleic acid, and does not occur in cells lacking the inserted nucleic acid. Accordingly, some embodiments of cells comprise an inserted nucleic acid at an essential gene locus, where the inserted nucleic acid comprises (i) a heterologous protein- encoding nucleotide sequence, and (ii) at least a portion of the essential gene. Some embodiments of methods of editing a cell comprise contacting a cell with a nucleic acid comprising (i) a first homology arm, (ii) a second homology arm, each homology arm having homology to an essential gene locus, (iii) a heterologous protein-encoding nucleotide sequence, and (iv) at least a portion of the essential gene. Some embodiments of nucleic acids comprise (i) a first homology arm, (ii) a second homology arm, each homology arm having homology to an essential gene locus, (iii) a heterologous protein-encoding nucleotide sequence, and (iv) at least a portion of the essential gene. An inserted nucleic acid in a cell genome, or nucleic acid for insertion into a cell genome, may comprise any suitable portion of an essential gene. In some embodiments, the nucleic acid comprises a portion of an exon of the essential gene. In some embodiments, the nucleic acid comprises an exon of the essential gene. In some embodiments, the nucleic acid comprises two or more exons of the essential gene. In some embodiments, the nucleic acid comprises 2, 3, 4, 5, 6, 7, 8, 9, or 10 exons of the essential gene. A nucleic acid comprising multiple exons of the essential gene may comprise the exons in series without intervening nucleotides (e.g., without introns). The nucleic acid may comprise the last exon of the essential gene, with a stop codon downstream of the last codon encoding an amino acid of the protein encoded by the essential gene. The nucleic acid may comprise the first coding exon of the essential gene. The first coding exon of the essential gene may be the first exon present on mRNA encoded by the essential gene, or a different exon, depending on the location of the start codon of the open reading frame encoding the protein encoded by the essential gene. In some embodiments, the nucleic acid comprises a mutated exon, or mutated portion of an exon, of the essential gene. The mutated exon or exon portion may be codon-optimized for expression in a cell. The mutated exon or exon portion may comprise fewer codons than the naturally occurring form of the exon (e.g., encoding a shorter amino acid sequence). The mutated exon or exon portion may lack a stop codon that is present in the natural exon. The mutated exon or exon portion may lack a start codon that is present in the natural exon. Whereas a start codon is necessary in the wild-type gene to initiate translation, the encoded methionine may be dispensable to the essential gene product. In such instances, an inserted nucleic acid may comprise the heterologous protein-encoding nucleotide sequence in-frame with a mutant form of the first coding exon of the essential gene, where the start codon and one or more subsequent codons present in the naturally occurring first coding exon are not present in the mutant form of the first coding exon. Similarly, one or more C-terminal amino acids may be dispensable to the function of the essential gene product. In such instances, an inserted nucleic acid may comprise the heterologous protein-encoding nucleotide sequence in-frame with a mutant form of the last exon of the essential gene, and optionally mutant forms of one or more upstream exons of the essential gene, where one or more codons encoding C-terminal amino acids are absent from the mutant forms of the exons. In some embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 C-terminal amino acids of the essential gene product are not encoded by a cell genome comprising the inserted nucleic acid. In some embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 N- terminal amino acids of the essential gene product are not encoded by a cell genome comprising the inserted nucleic acid. An inserted nucleic acid in a cell genome, or nucleic acid for insertion into a cell genome, may comprise the heterologous protein-encoding nucleotide sequence upstream from the first coding exon of the essential gene. An inserted nucleic acid in a cell genome, or nucleic acid for insertion into a cell genome, may comprise the heterologous protein- encoding nucleotide sequence downstream from the last coding exon of the essential gene. In some embodiments, the heterologous protein-encoding nucleotide sequence, when inserted into a cell genome, is in-frame with the essential gene, such that a single transcribed RNA encodes the essential gene product and heterologous protein(s). In some embodiments, a nucleotide sequence encoding a 2A motif is present in-frame between the nucleotide sequence encoding the essential gene product and the heterologous protein-encoding nucleotide sequence. In some embodiments, the 2A motif is 3’ to the nucleotide sequence encoding the essential gene product, and 5’ to the heterologous protein-encoding nucleotide sequence. In some embodiments, the 2A motif is 3’ to the heterologous protein-encoding nucleotide sequence, and 5’ to the nucleotide sequence encoding the essential gene product. In some embodiments, the 2A motif is a P2A motif. In some embodiments, the 2A motif is a T2A motif. In some embodiments, the 2A motif is an E2A motif. In some embodiments, the 2A motif is an F2A motif. In some embodiments, multiple heterologous coding sequences are present in an inserted nucleic acid, or nucleic acid for insertion into a cell genome. In some embodiments, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 heterologous coding sequences are present in the nucleic acid. In some embodiments, two or more heterologous coding sequences are separated by nucleotide sequences encoding 2A motifs. In some embodiments, separate pairs of heterologous coding sequences are separated by different nucleotide sequences encoding 2A motifs. In some embodiments, each heterologous coding sequence is separated by a different nucleotide sequence encoding a 2A motif. Separation of multiple heterologous coding sequences by different nucleotide sequences encoding 2A motifs reduces the likelihood of recombination that could occur between identical nucleotide sequences encoding 2A motifs, leading to excision of a heterologous coding sequence from the cell genome. The 2A motifs separating different proteins encoded by different heterologous coding sequences may be the same or different 2A motifs. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding FOXP3. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a first CISC component as described in the “Chemically Induced Signaling Complex” section. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a second CISC component as described in the “Chemically Induced Signaling Complex” section. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a first CISC component, and a nucleotide sequence encoding a second CISC component. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a third CISC component (i.e., soluble FRB domain) as described in the “Chemically Induced Signaling Complex” section. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a chimeric antigen receptor (CAR). An inserted nucleic acid in a cell genome, or nucleic acid for insertion into a cell genome, may comprise an exogenous stop codon, a nucleotide sequence encoding a 5’ untranslated region (UTR), a nucleotide sequence encoding a 3’ untranslated region (UTR), and / or a nucleotide sequence encoding a polyA tail. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a 5’ UTR. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a 3’ UTR. In some embodiments, the nucleic acid comprises a nucleotide sequence encoding a 3’ UTR and a polyA tail. In some embodiments, a transcribed polyA tail comprises at least 100 consecutive adenosine nucleotides. A nucleic acid may be inserted at any suitable essential gene locus. In some embodiments, the essential gene locus is a GAPDH locus. In some embodiments, the essential gene locus is a KIF11 locus. In some embodiments, the essential gene locus is a TBP locus. In some embodiments, the gene locus is essential for Treg cell survival. In some embodiments, the gene locus is essential for maintenance of a Treg cell phenotype. Any suitable nuclease may be used for targeted cleavage. In some embodiments, a cell is edited by an RNA-guided nuclease. In some embodiments, the RNA-guided nuclease is a Cas endonuclease. In some embodiments, the RNA-guided nuclease is a Cas9 endonuclease. In some embodiments, the RNA-guided nuclease is a Cpf1 endonuclease. In some embodiments, the nuclease is a meganuclease. In some embodiments, the nuclease is a TALEN. In some embodiments, the nuclease is a MegaTAL. In some embodiments, the nuclease is a zinc finger nuclease (ZFN). The homology arms may be the same length, have similar lengths (within 100 bp of each other), or different lengths. In some embodiments, one or both homology arms have a length of 100–2,000 bp, 200–2,000 bp, 400–1,500 bp, or 500–1,000 bp. In some embodiments, one or both homology arms are about 100 bp, about 200 bp, about 300 bp, about 400 bp, about 500 bp, about 600 bp, about 700 bp, about 800 bp, about 900 bp, about 1,000 bp, about 1,100 bp, about 1,200 bp, about 1,300 bp, about 1,400 bp, about 1,500 bp, about 1,600 bp, about 1,700 bp, about 1,800 bp, about 1,900 bp, or about 2,000 bp. In some embodiments, both homology arms are 100-2,000 nucleotides in length. In some embodiments, both homology arms are 300–1,000 nucleotides in length. In some embodiments, both homology arms are 300–700 nucleotides in length. In some embodiments, both homology arms are 300–500 nucleotides in length. In some embodiments, both homology arms are 500–700 nucleotides in length. In some embodiments, both homology arms are 700–1,000 nucleotides in length. In some embodiments, the nucleic acid is a single-stranded DNA (ssDNA). In some embodiments, the nucleic acid is a closed-ended DNA (ceDNA). In some embodiments, the nucleic acid is present in a vector. Any suitable vector may comprise the nucleic acid. In some embodiments, the vector is a viral vector. In some embodiments, the vector is an integrase-deficient lentiviral vector (IDLV). In some embodiments, the vector is an adeno- associated viral (AAV) vector. In some embodiments, the AAV vector is an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 vector. In some embodiments, the AAV vector is an AAV5 vector. In some embodiments, the AAV vector is an AAV6 vector. In some embodiments, the nucleic acid is present in a lipid delivery vehicle. In some embodiments, the nucleic acid is present in a lipid nanoparticle. In some embodiments, the lipid nanoparticle comprises the nuclease or a nucleic acid encoding the nuclease. In some embodiments, the lipid nanoparticle comprises a guide RNA or a nucleic acid encoding the guide RNA. In some embodiments, the nucleic acid encoding the nuclease is an mRNA. Promoters Embodiments of the compositions, cells, nucleic acids, vectors, and methods that contemplate use of a promoter (e.g., heterologous promoter) may use any suitable promoter. In some embodiments, the promoter is active, promoting transcription of RNA even under pro-inflammatory conditions. In some embodiments, the promoter is operably linked to an endogenous nucleotide sequence encoding FOXP3. In some embodiments, the promoter is operably linked to an exogenous nucleotide sequence encoding FOXP3. In some embodiments, the promoter is operably linked to an exogenous nucleotide sequence encoding a chimeric antigen receptor (CAR). In some embodiments, the promoter is a heterologous promoter. A heterologous promoter, when operably linked to a coding sequence, refers to a promoter that is not operably linked to that coding sequence in nature. A heterologous promoter may also refer to a promoter that controls episomal expression of an introduced nucleic acid sequence. A heterologous promoter may be derived from a different location in the cell genome. A heterologous promoter may be a synthetic promoter that is not found in nature in any organism. A heterologous promoter may be obtained from another species (e.g., virus, bacterium). In some embodiments, the heterologous promoter is operably linked to an endogenous nucleotide sequence encoding FOXP3. In some embodiments, the heterologous promoter is operably linked to an exogenous nucleotide sequence encoding FOXP3. In some embodiments, the heterologous promoter is operably linked to an exogenous nucleotide sequence encoding a chimeric antigen receptor (CAR). In some embodiments, the promoter is a constitutive promoter, which promotes transcription of an operably linked sequence (e.g., a CAR and / or FOXP3) at a consistent rate. In some embodiments, a constitutively active promoter promotes transcription of an operably linked sequence (e.g., a CAR and / or FOXP3) at a supraphysiological rate. Constitutive promoters may be strong promoters, which promote transcription at a higher rate than an endogenous promoter, or weak promoters, which promote transcription at a lower rate than a strong or endogenous promoter. In some embodiments, the constitutive promoter is a strong promoter. In some embodiments, the constitutive promoter is a weak promoter. In some embodiments, the constitutive promoter is an EF-1α, PGK, or MND promoter. In some embodiments, another suitable promoter, such as an SV40, CMV, UBC, or CAGG promoter, is used. In some embodiments, the constitutive promoter is an MND promoter. In some embodiments, the promoter is an inducible promoter. Inducible promoters promote transcription of an operably linked sequence in response to the presence of an activating signal, or the absence of a repressor signal. In some embodiments, the inducible promoter is inducible by a drug or steroid. In some embodiments, a promoter is a ubiquitous promoter. In some embodiments, the ubiquitous promoter is Reverse Orientation Splice Acceptor 26 (ROSA26) promoter. In some embodiments, the promoter is a unidirectional promoter. In some embodiments, the unidirectional promoter drives expression in either a forward (e.g., directing transcription downstream from the promoter) or reverse direction (e.g., directing transcription upstream from the promoter), but not both. In some embodiments, the unidirectional promoter drives transcription downstream of the promoter. In some embodiments, the unidirectional promoter drives transcription upstream of the promoter. In some embodiments, the unidirectional promoter is a unidirectional constitutive promoter. In some embodiments, the unidirectional promoter is operably linked to an endogenous nucleotide sequence encoding FOXP3. In some embodiments, the unidirectional promoter is operably linked to an exogenous nucleotide sequence encoding FOXP3. In some embodiments, the unidirectional promoter is operably linked to an exogenous nucleotide sequence encoding a chimeric antigen receptor (CAR). In embodiments contemplating multiple promoters, the promoters may be the same or different promoters. In some embodiments, each of multiple promoters is a constitutive promoter. In some embodiments, each of multiple promoters is an MND promoter. Chemically induced signaling complexes (CISC) Some embodiments of methods of modifying cells comprise introducing into the cell one or more nucleic acids that collectively comprise (1) a first nucleic acid sequence encoding a first chemically inducible signaling complex (CISC) component, and (2) a second nucleic acid sequence encoding a second chemically inducible signaling complex (CISC) component, each CISC component comprising (a) an extracellular binding domain that is capable of binding to a CISC inducer molecule, (b) a transmembrane domain, and (c) an intracellular signaling domain, such that binding of the first and second CISC components to the CISC inducer molecule results in dimerization of the CISC components and a signal transduction event in the cell. Similarly, some embodiments of cells comprise (1) a first nucleic acid sequence encoding a first chemically inducible signaling complex (CISC) component, and (2) a second nucleic acid sequence encoding a second chemically inducible signaling complex (CISC) component, each CISC component comprising (a) an extracellular binding domain that is capable of binding to a CISC inducer molecule, (b) a transmembrane domain, and (c) an intracellular signaling domain, such that binding of the first and second CISC components to the CISC inducer molecule results in dimerization of the CISC components and a signal transduction event in the cell. Additionally, some nucleic acids and vectors comprise (1) a first nucleic acid sequence encoding a first chemically inducible signaling complex (CISC) component, and / or (2) a second nucleic acid sequence encoding a second chemically inducible signaling complex (CISC) component, each CISC component comprising (a) an extracellular binding domain that is capable of binding to a CISC inducer molecule, (b) a transmembrane domain, and (c) an intracellular signaling domain, such that binding of the first and second CISC components to the CISC inducer molecule results in dimerization of the CISC components and a signal transduction event in a cell. Expression of CISC components in a cell allows selective induction of signaling in a cell by manipulation of the presence and / or concentration of the CISC inducer molecule. Such controllable induction of signaling allows, for example, selective expansion of cells expressing both CISC components, where the signal transduction event results in proliferation of the cell. In some embodiments, where two nucleic acids, each encoding a different CISC component, are introduced into the cell, such selective expansion allows for selection of cells that contain both nucleic acids, as contacting a cell comprising only one CISC component would not induce dimerization with the absent second CISC component. Non-limiting examples of intracellular signaling domains include IL-2Rβ and IL-2Rγ cytoplasmic domains and functional derivatives thereof. In some embodiments, an intracellular signaling domain of one CISC component comprises an IL-2Rβ cytoplasmic domain or a functional derivative thereof, and an intracellular signaling domain of the other CISC component comprises an IL-2Rγ cytoplasmic signaling domain or a functional derivative thereof. In some embodiments, dimerization of the CISC components transduces an IL-2 signal in the cell. In some embodiments, dimerization of the CISC components induces phosphorylation of JAK1, JAK3, and / or STAT5 in the cell. In some embodiments, dimerization of the CISC components induces proliferation of the cell. In some embodiments, the IL-2Rβ cytoplasmic domain comprises the amino acid sequence of NCRNTGPWLKKVLKCNTPDPSKFFSQLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEIS PLEVLERDKVTQLLLQQDKVPEPASLSSNHSLTSCFTNQGYFFFHLPDALEIEACQVY FTYDPYSEEDPDEGVAGAPTGSSPQPLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSP PSTAPGGSGAGEERMPPSLQERVPRDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGE EVPDAGPREGVSFPWSRPPGQGEFRALNARLPLNTDAYLSLQELQGQDPTHLV (SEQ ID NO: 6). In some embodiments, the IL-2Rβ cytoplasmic domain is truncated, relative to a wild-type IL-2Rβ cytoplasmic domain. An exemplary truncated IL-2Rβ cytoplasmic domain that retains signal transduction activity is described, for example, in Cook et al., Mol Ther. 2023. S1525-0016(23)00255-1. In some embodiments, the truncated IL-2Rβ domain comprises the amino acid sequence of PAALGKDTIPWLGHLLVGLSGAFGFIILVYLLINCRNTGPWLKKVLKCNTPDPSKFFS QLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLERDKVTQLLLQQDKVPEPASL SLNTDAYLSLQELQ (SEQ ID NO: 23). In some embodiments, the IL-2Rγ cytoplasmic domain comprises the amino acid sequence of ERTMPRIPTLKNLEDLVTEYHGNFSAWSGVSKGLAESLQPDYSERLCLVSEIPPKGGA LGEGPGASPCNQHSPYWAPPCYTLKPET (SEQ ID NO: 7). Non-limiting examples of transmembrane domains include IL-2Rβ, IL-2Rγ, erythropoietin (Epo), and thrombopoietin (Tpo) transmembrane domains. In some embodiments, the transmembrane domain of a CISC component is derived from the same protein as the intracellular signaling domain of the CISC component (e.g., a CISC component comprising an IL-2Rβ intracellular domain comprises an IL-2Rβ transmembrane domain). In some embodiments, one CISC component comprises an IL-2Rβ transmembrane domain, and the other CISC component comprises an IL-2Rγ transmembrane domain. In some embodiments, an IL-2Rβ transmembrane domain comprises the amino acid sequence of IPWLGHLLVGLSGAFGFIILVYLLI (SEQ ID NO: 8). In some embodiments, an IL-2Rγ transmembrane domain comprises the amino acid sequence of VVISVGSMGLIISLLCVYFWL (SEQ ID NO: 9). Non-limiting examples of extracellular binding domains capable of binding a CISC inducer molecule include an FK506-binding protein (FKBP) domain and an FKBP- rapamycin-binding (FRB) domain. FKBP and FRB domains are capable of binding to rapamycin or rapalogs, such as those described below. In some embodiments, an extracellular binding domain of one CISC component comprises an FKBP domain, and an extracellular binding domain of the other CISC component comprises an FRB domain. In some embodiments, the CISC components form a heterodimer in the presence of the CISC inducer molecule. In some embodiments, an FKBP domain comprises an amino acid sequence of GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKPFKFMLGKQEVI RGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELLKLGE (SEQ ID NO: 10). In some embodiments, an FKBP domain comprises an amino acid sequence of GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKPFKFMLGKQEVI RGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELLKLE (SEQ ID NO: 4) (Gly107). In some embodiments, an FKBP domain comprises an F36V substitution, at a position corresponding to F36 of SEQ ID NO: 10 or SEQ ID NO: 4. Such a substitution allows the FKBP domain to interact with derivatives of rapamycin, such as AP1903. Clackson et al., Proc Natl Acad Sci U S A.1998.95(18):10437–10442. In some embodiments, an FRB domain comprises an amino acid sequence of EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETSFNQAYGRDLM EAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK (SEQ ID NO: 5). In some embodiments, an FRB domain comprises a T74L substitution, at a position corresponding to T74 of SEQ ID NO: 12. This T74L substitution corresponds to a T2098L substitution in the FRB domain as present on full-length mTOR. In some embodiments, an FRB domain comprises a K71P substitution, at a position corresponding to K71 of SEQ ID NO: 12. In some embodiments, an FRB domain comprises a K71T or K71A substitution, at a position corresponding to K71 of SEQ ID NO: 12. In some embodiments, an FRB domain comprises a W77F substitution, at a position corresponding to W77 of SEQ ID NO: 12. In some embodiments, an FRB domain comprises a K71P substitution, a T74L substitution, and a W77F substitution. Such substitutions allow the FRB domain to interact with derivatives of rapamycin, such as AP21967. Stankunas et al., Mol Cell.2003.12(6):1615–1624; Bayle et al., Chem Biol.2006.13(1):99–107. In some embodiments, a soluble FRB domain comprises an amino acid sequence of EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETSFNQAYGRDLM EAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK (SEQ ID NO: 13). In some embodiments, the soluble FRB domain consists of the amino acid sequence of EMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETSFNQAYGRDLM EAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISK (SEQ ID NO: 13). Each of the extracellular binding domains, transmembrane domains, and intracellular signaling domains of the CISC components may be connected to another domain of the same CISC component by a linker. In some embodiments, the linker comprises 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acids, such as glycines, or a number of amino acids, such as glycine, within a range defined by any two of the aforementioned numbers. In some embodiments, the glycine spacer comprises at least 3 glycines. In some embodiments, the glycine spacer comprises a sequence set forth as GGGS (SEQ ID NO: 24), GGGSGGG (SEQ ID NO: 25) or GGG. An extracellular binding domain may be connected to a transmembrane domain by a hinge. A hinge refers to a region that links the extracellular binding domain to the transmembrane domain, and may confer flexibility to the extracellular binding domain relative to the transmembrane domain. In some embodiments, the hinge positions the extracellular domain close to the plasma membrane to reduce the potential for recognition of the CISC by antibodies or binding fragments thereof. In some embodiments, the extracellular binding domain is located N-terminal to the hinge. In some embodiments, the hinge may be natural or synthetic. In some embodiments, a CISC component comprises a portion of an IL- 2Rγ extracellular domain between the extracellular binding domain and transmembrane domain. In some embodiments, the portion of the IL-2Rγ extracellular domain comprises the amino acid sequence of GSNTSKENPFLFALEA (SEQ ID NO: 15). In some embodiments, a CISC component comprises a portion of an IL-2Rβ extracellular domain between the extracellular binding domain and transmembrane domain. In some embodiments, the portion of the IL-2Rβ extracellular domain comprises the amino acid sequence of GKDT (SEQ ID NO: 14). In some embodiments, the portion of the IL-2Rγ extracellular domain comprises the amino acid sequence of QNLVIPWAPENLTLHKLSESQLELNWNNRFLNHCLEHLVQYRTDWDHSWTEQSVDY RHKFSLPSVDGQKRYTFRVRSRFNPLCGSAQHWSEWSHPIHWGSNTSKENPFLFALE A (SEQ ID NO: 16). In some embodiments, a CISC component comprises a portion of an IL- 2Rβ extracellular domain between the extracellular binding domain and transmembrane domain. In some embodiments, the portion of the IL-2Rβ extracellular domain comprises the amino acid sequence of KPFENLRLMAPISLQVVHVETHRCNISWEISQASHYFERHLEFEARTLSPGHTWEEAP LLTLKQKQEWICLETLTPDTQYEFQVRVKPLQGEFTTWSPWSQPLAFRTKPAALGKD T (SEQ ID NO: 17). In some embodiments, the CISC inducer molecule is rapamycin or a rapalog. In some embodiments, the CISC inducer molecule is rapamycin. Non-limiting examples of rapalogs include everolimus, CCI-779, C20-methallylrapamycin, C16-(S)-3-methylindolerapamycin, C16-iRap, C16-(S)-7-methylindolerapamycin, AP21967, C16- (S)Butylsulfonamidorapamycin, AP23050, sodium mycophenolic acid, benidipine hydrochloride, AP1903, and AP23573, and metabolites or derivatives thereof. In some embodiments, a method comprises introducing into a cell a nucleic acid encoding a third CISC component that is capable of binding to the CISC inducer molecule. Such CISC components are useful, for example, for binding to the intracellular CISC inducer molecules (e.g., intracellular rapamycin), thereby preventing the bound CISC inducer molecule from interacting with other intracellular molecules or structures (e.g., preventing rapamycin from interacting with mTOR). In some embodiments, the third CISC component is a soluble protein that does not comprise a transmembrane domain. In some embodiments, the third CISC component comprises an intracellular FRB domain. In some embodiments, a third CISC component is a soluble protein comprising an FRB domain and lacking a transmembrane domain. Nucleic acids encoding a first, second, and / or third CISC component may be comprised in one or more vectors. In some embodiments, a nucleic acid encoding a first CISC component is present on a separate vector from a nucleic acid encoding the second CISC component. In some embodiments, a nucleic acid encoding the third CISC component is present on the same vector as a nucleic acid encoding the first or second CISC component. In some embodiments, a nucleic acid encoding the third CISC component is present on a distinct vector from nucleic acids encoding the first and / or second CISC components. In some embodiments, one or more vectors are viral vectors. In some embodiments, one or more vectors are lentiviral vectors. In some embodiments, one or more vectors are adeno- associated viral (AAV) vectors. In some embodiments, one or more AAV vectors is an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 vector. In some embodiments, one or more AAV vectors are AAV5 vectors. In some embodiments, one or more AAV vectors are AAV6 vectors. In some embodiments, a CISC component comprises an amino acid sequence with at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% sequence identity to the amino acid sequence set forth as SEQ ID NO: 1, 2, 21, or 22. In some embodiments, one or more CISC components further comprise a signal peptide. The signal peptide may be any suitable signal peptide directs the translated CISC component to the cell membrane. Non- limiting examples of signal peptides include an LCN2 signal peptide (MPLGLLWLGLALLGALHAQA (SEQ ID NO: 18)), a CD8α signal peptide (MALPVTALLLPLALLLHAARPILWH (SEQ ID NO: 19)), and GM-CSFRα signal peptide (MLLLVTSLLLCELPHPAFLLI (SEQ ID NO: 20)). In some embodiments, one CISC component comprises an amino acid sequence with at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% sequence identity to the amino acid sequence set forth as SEQ ID NO: 1, and the other CISC component comprises an amino acid sequence with at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% sequence identity to the amino acid sequence set forth as SEQ ID NO: 2. In some embodiments, each CISC component further comprises a signal peptide, which may have the same or different amino acid sequences. The signal peptides may be any suitable signal peptide that directs the translated CISC component to the cell membrane. In some embodiments, one or more CISC components comprise an LCN2 signal peptide. In some embodiments, one or more CISC components comprise a CD8α signal peptide. In some embodiments, one or more CISC components comprise a GM-CSFRα signal peptide. In some embodiments, both CISC components comprise an LCN2 signal peptide. In some embodiments, both CISC components comprise a CD8α signal peptide. In some embodiments, both CISC components comprise a GM-CSFRα signal peptide. In some embodiments, a third CISC component comprises an amino acid sequence with at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% sequence identity to the amino acid sequence set forth as SEQ ID NO: 3. In some embodiments, a third CISC component consists of an amino acid sequence with at least 80%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or up to 100% sequence identity to the amino acid sequence set forth as SEQ ID NO: 3. In some embodiments, the third CISC component does not comprise a signal peptide. In some embodiments, the third CISC component does not comprise a transmembrane domain. Stable FOXP3 expression Some embodiments of methods of modifying cells comprise introducing a genetic modification in a cell that stabilizes expression of FoxP3. Similarly, some embodiments of cells comprise a genetic modification that stabilizes or increases FoxP3expression, relative to an unmodified cell. Additionally, some embodiments of nucleic acids and vectors stabilize FoxP3 expression in a cell. In some embodiments, an endogenous FOXP3 locus is modified in a cell, resulting in stabilized expression. For example, in some embodiments, a heterologous promoter is inserted within or downstream from a Treg-specific demethylated region (TSDR) in the genome, and upstream from a first coding exon of an endogenous FOXP3 coding sequence. In some embodiments, a promoter is inserted downstream from the TSDR, and within or upstream from the first coding exon of FOXP3. Insertion of a heterologous promoter in this manner bypasses endogenous regulation of FOXP3 by the TSDR, which can become methylated in inflammatory conditions, inhibiting transcription of the endogenous FOXP3 coding sequence from the endogenous FOXP3 promoter located upstream from the TSDR. Thus, such stabilized FoxP3 expression by heterologous promoter insertion allows stable FoxP3 expression even in inflammatory conditions, preventing transdifferentiation into a T effector cell. The heterologous promoter may be inserted at any position between the endogenous promoter and the first coding exon of the FOXP3 coding sequence. In some embodiments, the heterologous promoter is inserted 1–10,000, 10–1,000, 10–100, 10–5,000, 20–4,000, 30– 3,000, 40–2,000, 50–1,000, 60–750, 70–500, 80–400, 90–300, 100–200, 1–1,000, 1,000– 2,000, 2,000–3,000, 3,000–4,000, 4,000–5,000, 5,000–6,000, 6,000–7,000, 7,000–8,000, 8,000–9,000, or 9,000–10,000 nucleotides downstream from the TSDR of FOXP3. In some embodiments, the heterologous promoter is inserted 1–10,000, 10–1,000, 10–100, 10–5,000, 20–4,000, 30–3,000, 40–2,000, 50–1,000, 60–750, 70–500, 80–400, 90–300, 100–200, 1– 1,000, 1,000–2,000, 2,000–3,000, 3,000–4,000, 4,000–5,000, 5,000–6,000, 6,000–7,000, 7,000–8,000, 8,000–9,000, or 9,000–10,000 nucleotides upstream from the first coding exon of the FOXP3 coding sequence. In some embodiments, the heterologous promoter is inserted into the first coding exon, such that a synthetic first coding exon is created, where the synthetic first coding exon differs from the endogenous first coding exon but still comprises a start codon that is in-frame with the FOXP3 coding sequence of downstream FOXP3 exons. In some embodiments, the heterologous promoter is inserted into the TSDR, such that the TSDR is modified and does not inhibit transcription of the endogenous FOXP3 coding sequence in inflammatory conditions. In some embodiments, the nucleic acid comprising a heterologous promoter is comprised on a vector. In some embodiments, the vector is a viral vector. In some embodiments, the viral vector is an adeno-associated virus (AAV) vector. In some embodiments, the AAV vector is an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 vector. In some embodiments, the AAV vector is an AAV5 vector. In some embodiments, the AAV vector is an AAV6 vector. In some embodiments, a nucleic acid comprising a promoter operably linked to a nucleic acid sequence encoding FoxP3 or a functional fragment thereof is introduced into the cell. Expression of a heterologous promoter and sequence encoding FoxP3 is useful, for example, for expressing functional FoxP3 in cells containing genomic mutations in the FOXP3 coding sequence (e.g., cells from subjects having IPEX syndrome). Additionally, additional coding sequences (e.g., encoding a CAR) may be included in a nucleic acid, such that the heterologous promoter controls transcription of RNA encoding FoxP3 and RNA encoding one or more other proteins (e.g., CAR). In some embodiments, the sequence encoding FoxP3 is a cDNA sequence that does not comprise an intron. The introduced nucleic acid may be integrated into the genome at a targeted locus (e.g., by homologous recombination), integrated in a non-targeted manner (e.g., by delivery on a lentiviral vector), or not integrated. In some embodiments, the nucleic acid comprises a 5′ homology arm that is upstream from the promoter, and a 3′ homology arm that is downstream from the nucleic acid sequence encoding FoxP3, and both homology arms have homology to a targeted locus in a genome. Such homology arms promote insertion of the nucleic acid into the genome at the targeted locus by homologous recombination. The homology arms may be the same length, have similar lengths (within 100 bp of each other), or different lengths. In some embodiments, one or both homology arms have a length of 200– 2,000 bp, 400–1,500 bp, 500–1,000 bp. In some embodiments, one or both homology arms are about 100 bp, about 200 bp, about 300 bp, about 400 bp, about 500 bp, about 600 bp, about 700 bp, about 800 bp, about 900 bp, about 1,000 bp, about 1,100 bp, about 1,200 bp, about 1,300 bp, about 1,400 bp, about 1,500 bp, about 1,600 bp, about 1,700 bp, about 1,800 bp, about 1,900 bp, or about 2,000 bp. In some embodiments, the nucleic acid is inserted at a FOXP3 locus in the genome. In some embodiments, the nucleic acid is inserted at a non-FOXP3 locus. In some embodiments, the targeted locus is a safe harbor locus. In some embodiments, the safe harbor locus is an AAVS1 locus, a HIPP11 locus, or a ROSA26 locus. In some embodiments, the nucleic acid is inserted at a TCRα (TRAC) locus. In some embodiments, the nucleic acid is inserted at a TCRβ (TRBC) locus. In some embodiments, the nucleic acid is inserted at a CD3ζ (CD3Z) locus. In some embodiments, a nuclease capable of cleaving the genome at a targeted locus, or a nucleic acid encoding the nuclease (e.g., an mRNA) is introduced into the cell. Following delivery of the nuclease or transcription of the nuclease inside the cell, the nuclease introduces a double-stranded break at the targeted locus, thereby promoting integration of a donor template (e.g., nucleic acid comprising a promoter and sequence encoding FoxP3, or nucleic acid comprising a heterologous promoter for promoting transcription of an endogenous FOXP3 coding sequence) into the genome at the targeted locus by homology-directed repair. The nuclease may be any suitable nuclease, including a meganuclease, zinc finger nuclease, TALEN, or RNA-guided nuclease. In embodiments where an RNA-guided nuclease (or nucleic acid encoding an RNA-guided nuclease) is delivered, a guide RNA (or nucleic acid encoding a guide RNA) comprising a spacer sequence complementary to a genomic sequence at the targeted locus is introduced into the cell. A gRNA or nucleic acid encoding a gRNA may be introduced into the cell with the nuclease or nucleic acid encoding the nuclease, or introduced separately (e.g., in a separate vector or delivery vehicle). The RNA-guided nuclease may be any suitable RNA-guided nuclease, such as those described in the section entitled “Nucleases.” In some embodiments, a nucleic acid comprising a heterologous promoter operably linked to a sequence encoding FoxP3 or a functional fragment thereof is present on a vector. In some embodiments, the vector is a viral vector. In some embodiments, the vector is a lentiviral vector. In some embodiments, the vector is an adeno-associated virus (AAV) vector. In some embodiments, the AAV vector is an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, or AAV11 vector. In some embodiments, the AAV vector is an AAV5 vector. In some embodiments, the AAV vector is an AAV6 vector. In some embodiments, the vector is a plasmid. In some embodiments, the vector is bacterial artificial chromosome. In some embodiments, the vector is human artificial chromosome. In some embodiments, the vector integrates into a chromosome of the genome, and RNA encoding FoxP3 is transcribed from the genome of the cell. In other embodiments, the vector does not integrate into a chromosome, and the sequence encoding FoxP3 is expressed episomally. The heterologous promoter inserted into the FOXP3 locus or operably linked to the FOXP3 coding sequence may be any suitable promoter. In some embodiments, the promoter is a constitutive promoter. In some embodiments, the promoter is an MND, PGK, or EF-1α promoter. In some embodiments, the promoter is an MND promoter. In some embodiments, the promoter is an inducible promoter. In some embodiments, the promoter is inducible by a drug or steroid. Pharmaceutical compositions Some aspects relate to a pharmaceutical composition comprising a cell, vector, or nucleic acid, and a pharmaceutically acceptable excipient or carrier. Such pharmaceutical compositions are formulated, for example, for systemic administration, or administration to target tissues. “Acceptable” means that the excipient (carrier) must be compatible with the active ingredient of the composition (and preferably, capable of stabilizing the active ingredient) and not deleterious to the subject to be treated. Pharmaceutically acceptable excipients, carriers, buffers, stabilizers, isotonicizing agents, preservatives or antioxidants, or other materials well known to those skilled in the art. Such materials should be non-toxic and should not interfere with the efficacy of the active ingredient. The precise nature of the carrier or other material may depend on the route of administration, e.g., parenteral, intramuscular, intradermal, sublingual, buccal, ocular, intranasal, subcutaneous, intrathecal, intratumoral, oral, vaginal, or rectal. See, e.g., Remington: The Science and Practice of Pharmacy 20th Ed. (2000) Lippincott Williams and Wilkins, Ed. K. E. Hoover. The pharmaceutical compositions to be used for in vivo administration must be sterile, with the exception of any cells, viruses, and / or viral vectors being used to achieve a biological effect (e.g., immunosuppression). This is readily accomplished by, for example, filtration through sterile filtration membranes. The pharmaceutical compositions may be placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle. In some embodiments, a pharmaceutical composition is a biological product. A biological product includes: a virus, therapeutic serum, toxin, antitoxin, vaccine, blood, blood component or derivative, allergenic product, protein, or analogous product, or arsphenamine or derivative of arsphenamine (or any other trivalent organic arsenic compound), applicable to the prevention, treatment, or cure of a disease or condition of human beings. In some embodiments, the pharmaceutical compositions can be formulated for intramuscular injection, intravenous injection, intradermal injection, or subcutaneous injection. In some embodiments, the pharmaceutical compositions are formulated for infusion. The pharmaceutical compositions can comprise pharmaceutically acceptable carriers, buffer agents, excipients, salts, or stabilizers in the form of lyophilized formulations or aqueous solutions. See, e.g., Remington: The Science and Practice of Pharmacy 20th Ed. (2000) Lippincott Williams and Wilkins, Ed. K. E. Hoover). Acceptable carriers, excipients, or stabilizers are nontoxic to recipients at the dosages and concentrations used, and may comprise buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrans; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or non-ionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG). In some embodiments, the pharmaceutical composition comprises a nanoparticle or a lipid delivery vehicle (e.g., lipid nanoparticle or liposome). In some embodiments, the pharmaceutical composition comprises a lipid nanoparticle, which can be prepared by any suitable method, such as described in Epstein et al., Proc Natl Acad Sci USA .1985.82:3688; Hwang et al. Proc Natl Acad Sci USA.1980.77:4030; and U.S. Pat. Nos.4,485,045 and 4,544,545. Liposomes with enhanced circulation time are disclosed in U.S. Pat. No. 5,013,556. Particularly useful liposomes can be generated by the reverse phase evaporation method with a lipid composition comprising phosphatidylcholine, cholesterol and PEG- derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through filters of defined pore size to yield liposomes with the desired diameter. Lipids used in the formulation of lipid nanoparticles for delivering nucleic acids include ionizable amino lipids, non-cationic lipids, sterols, and polyethylene glycol-modified lipids. See, e.g., Buschmann et al., Vaccines. 2021.9(1):65. In some embodiments, the nucleic acid is surrounded by the lipids of the lipid nanoparticle and present in the interior of the lipid nanoparticle. In some embodiments, the nucleic acid is dispersed throughout the lipids of the lipid nanoparticle. In some embodiments, the lipid nanoparticle comprises an ionizable amino lipid, a non-cationic lipid, a sterol, and / or a polyethylene glycol (PEG)-modified lipid. The pharmaceutical compositions can be in unit dosage forms such as tablets, pills, capsules, powders, granules, solutions or suspensions, or suppositories, for oral, parenteral or rectal administration, or administration by inhalation or insufflation. For preparing solid compositions such as tablets, the principal active ingredient can be mixed with a pharmaceutical carrier, e.g., conventional tableting ingredients such as corn starch, lactose, sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalcium phosphate or gums, and other pharmaceutical diluents, e.g., water, to form a solid preformulation composition containing a homogeneous mixture, or a non-toxic pharmaceutically acceptable salt thereof. When referring to these preformulation compositions as homogeneous, it is meant that the active ingredient is dispersed evenly throughout the composition so that the composition may be readily subdivided into equally effective unit dosage forms such as tablets, pills and capsules. The tablets or pills of the novel composition can be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action. For example, the tablet or pill can comprise an inner dosage and an outer dosage component, the latter being in the form of an envelope over the former. The two components can be separated by an enteric layer that serves to resist disintegration in the stomach and permits the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymeric acids and mixtures of polymeric acids with such materials as shellac, cetyl alcohol and cellulose acetate. Pharmaceutical compositions for inhalation or insufflation include solutions and suspensions in pharmaceutically acceptable, aqueous or organic solvents, or mixtures thereof, and powders. The liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as set out above. In some embodiments, the compositions are administered by the oral or nasal respiratory route for local or systemic effect. Compositions in preferably sterile pharmaceutically acceptable solvents may be nebulized by use of gases. Nebulized solutions may be breathed directly from the nebulizing device or the nebulizing device may be attached to a face mask, tent or intermittent positive pressure breathing machine. Solution, suspension, or powder compositions may be administered, preferably orally or nasally, from devices which deliver the formulation in an appropriate manner. Pharmaceutical compositions may be useful for treating a subject that has or is at risk of developing an inflammatory, autoimmune, or allergic condition or disease. A subject having or at risk of developing an inflammatory, autoimmune, or allergic condition or disease may be identified by ascertaining the presence and / or absence of one or more risk factors, diagnostic indicators, or prognostic indications. The determination may be made based on clinical, cellular, or serologic findings, including flow cytometry, serology, and / or DNA analyses. The pharmaceutical compositions can include a therapeutically effective amount of any cell, vector, and / or nucleic acid. For example, in some embodiments, the pharmaceutical composition includes a cell, vector, or nucleic acid at any of the doses. A “therapeutically effective amount” refers to an amount effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result. The therapeutically effective amount may vary according to factors such as the age, sex, and weight of the individual, and the ability of the cell, nucleic acid, or vector to cause a desired response in the subject. In some embodiments, a therapeutically effective amount reduces the severity of a disease. In some embodiments, a therapeutically effective amount reduces the duration of a disease. In some embodiments, a therapeutically effective amount delays the progression of a disease. In some embodiments, a therapeutically effective amount alleviates one or more symptoms of a disease. In some embodiments, a therapeutically effective amount prevents recurrence of a disease. In some embodiments, a therapeutically effective amount delays recurrence of a disease. Pharmaceutical compositions can be prepared in accordance with known techniques. See, e.g., Remington, The Science And Practice of Pharmacy (21st ed., Philadelphia, Lippincott, Williams & Wilkins, 2005). For example, cells, vectors, or nucleic acids may be admixed with a pharmaceutically acceptable excipient, and the resulting composition is administered to a subject. The carrier must be acceptable in the sense of being compatible with any other ingredients in the formulation and must not be deleterious to the subject. The carrier can be a solid or a liquid, or both, and can be formulated with the compound as a unit- dose formulation. In some embodiments, a pharmaceutical composition comprises cells at a dose of about 104to about 1010cells / kg. In some embodiments, the pharmaceutical composition comprises cells at a dose of about: 104to 105, 105to 106, 106to 107, 107to 108, 108to 109, or 109to 1010cells / kg. In some embodiments, a pharmaceutical composition comprises cells at a dose of about 0.1 x 106, 0.2 x 106, 0.3 x 106, 0.4 x 106, 0.5 x 106, 0.6 x 106, 0.7 x 106, 0.8 x 106, 0.9 x 106, 1.0 x 106, 1.1 x 106, 1.2 x 106, 1.3 x 106, 1.4 x 106, 1.5 x 106, 1.6 x 106, 1.7 x 106, 1.8 x 106, 1.9 x 106, 2.0 x 106, 2.1 x 106, 2.2 x 106, 2.3 x 106, 2.4 x 106, 2.5 x 106, 2.6 x 106, 2.7 x 106, 2.8 x 106, 2.9 x 106, 3.0 x 106, 3.1 x 106, 3.2 x 106, 3.3 x 106, 3.4 x 106, 3.5 x 106, 3.6 x 106, 3.7 x 106, 3.8 x 106, 3.9 x 106, 4.0 x 106, 4.1 x 106, 4.2 x 106, 4.3 x 106, 4.4 x 106, 4.5 x 106, 4.6 x 106, 4.7 x 106, 4.8 x 106, 4.9 x 106, 5.0 x 106, 5.1 x 106, 5.2 x 106, 5.3 x 106, 5.4 x 106, 5.5 x 106, 5.6 x 106, 5.7 x 106, 5.8 x 106, 5.9 x 106, 6.0 x 106, 6.1 x 106, 6.2 x 106, 6.3 x 106, 6.4 x 106, 6.5 x 106, 6.6 x 106, 6.7 x 106, 6.8 x 106, 6.9 x 106, 7.0 x 106, 7.1 x 106, 7.2 x 106, 7.3 x 106, 7.4 x 106, 7.5 x 106, 7.6 x 106, 7.7 x 106, 7.8 x 106, 7.9 x 106, 8.0 x 106, 8.1 x 106, 8.2 x 106, 8.3 x 106, 8.4 x 106, 8.5 x 106, 8.6 x 106, 8.7 x 106, 8.8 x 106, 8.9 x 106, 9.0 x 106, 9.1 x 106, 9.2 x 106, 9.3 x 106, 9.4 x 106, 9.5 x 106, 9.6 x 106, 9.7 x 106, 9.8 x 106, 9.9 x 106, 1.0 x 107, 1.1 x 107, 1.2 x 107, 1.3 x 107, 1.4 x 107, 1.5 x 107, 1.6 x 107, 1.7 x 107, 1.8 x 107, 1.9 x 107, 2.0 x 107, 2.1 x 107, 2.2 x 107, 2.3 x 107, 2.4 x 107, 2.5 x 107, 2.6 x 107, 2.7 x 107, 2.8 x 107, 2.9 x 107, 3.0 x 107, 3.1 x 107, 3.2 x 107, 3.3 x 107, 3.4 x 107, 3.5 x 107, 3.6 x 107, 3.7 x 107, 3.8 x 107, 3.9 x 107, 4.0 x 107, 4.1 x 107, 4.2 x 107, 4.3 x 107, 4.4 x 107, 4.5 x 107, 4.6 x 107, 4.7 x 107, 4.8 x 107, 4.9 x 107, 5.0 x 107, 5.1 x 107, 5.2 x 107, 5.3 x 107, 5.4 x 107, 5.5 x 107, 5.6 x 107, 5.7 x 107, 5.8 x 107, 5.9 x 107, 6.0 x 107, 6.1 x 107, 6.2 x 107, 6.3 x 107, 6.4 x 107, 6.5 x 107, 6.6 x 107, 6.7 x 107, 6.8 x 107, 6.9 x 107, 7.0 x 107, 7.1 x 107, 7.2 x 107, 7.3 x 107, 7.4 x 107, 7.5 x 107, 7.6 x 107, 7.7 x 107, 7.8 x 107, 7.9 x 107, 8.0 x 107, 8.1 x 107, 8.2 x 107, 8.3 x 107, 8.4 x 107, 8.5 x 107, 8.6 x 107, 8.7 x 107, 8.8 x 107, 8.9 x 107, 9.0 x 107, 9.1 x 107, 9.2 x 107, 9.3 x 107, 9.4 x 107, 9.5 x 107, 9.6 x 107, 9.7 x 107, 9.8 x 107, 9.9 x 107, or 1.0 x 108cells / kg. In some embodiments, a pharmaceutical composition comprises an effective amount of a vector or nucleic acid. In some examples, the pharmaceutical composition comprises about 0.1 mg / kg to about 3 mg / kg of the vector or nucleic acid. In some embodiments, the pharmaceutical composition comprises about 0.1 mg / kg, about 0.25 mg / kg, about 0.5 mg / kg, about 0.75 mg / kg, about 1.0 mg / kg, about 1.5 mg / kg, about 2.0 mg / kg, about 2.5 mg / kg, or about 3.0 mg / kg of the vector or nucleic acid. In some embodiments, pharmaceutical composition comprises about 0.1 mg / kg to about 0.25 mg / kg, about 0.25 mg / kg to about 0.5 mg / kg, about 0.5 mg / kg to about 0.75 mg / kg, about 0.75 mg / kg to about 1.0 mg / kg, about 1.0 mg / kg to about 1.5 mg / kg, about 1.5 mg / kg to about 2.0 mg / kg, about 2.0 mg / kg to about 2.5 mg / kg, or about 2.5 mg / kg to about 3.0 mg / kg of the vector or nucleic acid. In some embodiments, the pharmaceutical composition comprises a vector or nucleic acid in a lipid delivery vehicle. In some embodiments, the pharmaceutical composition comprises a vector or nucleic acid in a lipid nanoparticle. Some embodiments of lipid nanoparticles comprise at least one cationic lipid, at least one non-cationic lipid, and at least one conjugated lipid. In more particular examples, lipid nanoparticles can comprise from about 50 mol % to about 85 mol % of a cationic lipid, from about 13 mol % to about 49.5 mol % of a non-cationic lipid, and from about 0.5 mol % to about 10 mol % of a lipid conjugate, and are produced in such a manner as to have a non- lamellar (e.g., non-bilayer) morphology. In other examples, lipid nanoparticles can comprise from about 40 mol % to about 85 mol % of a cationic lipid, from about 13 mol % to about 49.5 mol % of a non-cationic lipid, and from about 0.5 mol % to about 10 mol % of a lipid conjugate and are produced in such a manner as to have a non-lamellar (e.g., non-bilayer) morphology. Cationic lipids can include, for example, one or more of the following: palmitoyi- oleoyl-nor-arginine (PONA), MPDACA, GUADACA, ((6Z,9Z,28Z,31Z)-heptatriaconta- 6,9,28,31-tetraen-19-yl 4-(dimethylamino)butanoate) (MC3), LenMC3, CP-LenMC3, γ- LenMC3, CP-γ-LenMC3, MC3MC, MC2MC, MC3 Ether, MC4 Ether, MC3 Amide, Pan- MC3, Pan-MC4 and Pan MC5, 1,2-dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 1,2-dilinolenyloxy-N,N-dimethylaminopropane (DLenDMA), 2,2-dilinoleyl-4-(2- dimethylaminoethyl)-[1,3]-dioxolane (DLin-K-C2-DMA; “XTC2”), 2,2-dilinoleyl-4-(3- dimethylaminopropyl)-[1,3]-dioxolane (DLin-K-C3-DMA), 2,2-dilinoleyl-4-(4- dimethylaminobutyl)-[1,3]-dioxolane (DLin-K-C4-DMA), 2,2-dilinoleyl-5- dimethylaminomethyl-[1,3]-dioxane (DLin-K6-DMA), 2,2-dilinoleyl-4-N-methylpepiazino- [1,3]-dioxolane (DLin-K-MPZ), 2,2-dilinoleyl-4-dimethylaminomethyl-[1,3]-dioxolane (DLin-K-DMA), 1,2-dilinoleylcarbamoyloxy-3-dimethylaminopropane (DLin-C-DAP), 1,2- dilinoleyoxy-3-(dimethylamino)acetoxypropane (DLin-DAC), 1,2-dilinoleyoxy-3- morpholinopropane (DLin-MA), 1,2-dilinoleoyl-3-dimethylaminopropane (DLinDAP), 1,2- dilinoleylthio-3-dimethylaminopropane (DLin-S-DMA), 1-linoleoyl-2-linoleyloxy-3- dimethylaminopropane (DLin-2-DMAP), 1,2-dilinoleyloxy-3-trimethylaminopropane chloride salt (DLin-TMA.Cl), 1,2-dilinoleoyl-3-trimethylaminopropane chloride salt (DLin- TAP.Cl), 1,2-dilinoleyloxy-3-(N-methylpiperazino)propane (DLin-MPZ), 3-(N,N- dilinoleylamino)-1,2-propanediol (DLinAP), 3-(N,N-dioleylamino)-1,2-propanedio (DOAP), 1,2-dilinoleyloxo-3-(2-N,N-dimethylamino)ethoxypropane (DLin-EG-DMA), N,N-dioleyl- N,N-dimethylammonium chloride (DODAC), 1,2-dioleyloxy-N,N-dimethylaminopropane (DODMA), 1,2-distearyloxy-N,N-dimethylaminopropane (DSDMA), N-(1-(2,3- dioleyloxy)propyl)-N,N,N-trimethylammonium chloride (DOTMA), N,N-distearyl-N,N- dimethylammonium bromide (DDAB), N-(1-(2,3-dioleoyloxy)propyl)-N,N,N- trimethylammonium chloride (DOTAP), 3-(N-(N′,N′-dimethylaminoethane)- carbamoyl)cholesterol (DC-Chol), N-(1,2-dimyristyloxyprop-3-yl)-N,N-dimethyl-N- hydroxyethyl ammonium bromide (DMRIE), 2,3-dioleyloxy-N-[2(spermine- carboxamido)ethyl]-N,N-dimethyl-1-propanaminiumtrifluoroacetate (DOSPA), dioctadecylamidoglycyl spermine (DOGS), 3-dimethylamino-2-(cholest-5-en-3-beta- oxybutan-4-oxy)-1-(cis,cis-9,12-octadecadienoxy)propane (CLinDMA), 2-[5′-(cholest-5-en- 3-beta-oxy)-3′-oxapentoxy)-3-dimethy-1-(cis,cis-9′,1-2′-octadecadienoxy)propane (CpLinDMA), N,N-dimethyl-3,4-dioleyloxybenzylamine (DMOBA), 1,2-N,N′- dioleylcarbamyl-3-dimethylaminopropane (DOcarbDAP), 1,2-N,N′-dilinoleylcarbamyl-3- dimethylaminopropane (DLincarbDAP), or mixtures thereof. The cationic lipid can also be DLinDMA, DLin-K-C2-DMA (“XTC2”), MC3, LenMC3, CP-LenMC3, γ-LenMC3, CP-γ- LenMC3, MC3MC, MC2MC, MC3 Ether, MC4 Ether, MC3 Amide, Pan-MC3, Pan-MC4, Pan MC5, or mixtures thereof. In some embodiments, the cationic lipid may comprise from about 50 mol % to about 90 mol %, from about 50 mol % to about 85 mol %, from about 50 mol % to about 80 mol %, from about 50 mol % to about 75 mol %, from about 50 mol % to about 70 mol %, from about 50 mol % to about 65 mol %, or from about 50 mol % to about 60 mol % of the total lipid present in the particle. In some embodiments, the cationic lipid may comprise from about 40 mol % to about 90 mol %, from about 40 mol % to about 85 mol %, from about 40 mol % to about 80 mol %, from about 40 mol % to about 75 mol %, from about 40 mol % to about 70 mol %, from about 40 mol % to about 65 mol %, or from about 40 mol % to about 60 mol % of the total lipid present in the particle. The non-cationic lipid may comprise, e.g., one or more anionic lipids and / or neutral lipids. In some embodiments, the non-cationic lipid comprises one of the following neutral lipid components: (1) cholesterol or a derivative thereof; (2) a phospholipid; or (3) a mixture of a phospholipid and cholesterol or a derivative thereof. Examples of cholesterol derivatives include, but are not limited to, cholestanol, cholestanone, cholestenone, coprostanol, cholesteryl-2′-hydroxyethyl ether, cholesteryl-4′-hydroxybutyl ether, and mixtures thereof. The phospholipid may be a neutral lipid including, but not limited to, dipalmitoylphosphatidylcholine (DPPC), distearoylphosphatidylcholine (DSPC), dioleoylphosphatidylethanolamine (DOPE), palmitoyloleoyl-phosphatidylcholine (POPC), palmitoyloleoyl-phosphatidylethanolamine (POPE), palmitoyloleyol-phosphatidylglycerol (POPG), dipalmitoyl-phosphatidylethanolamine (DPPE), dimyristoyl- phosphatidylethanolamine (DMPE), distearoyl-phosphatidylethanolamine (DSPE), monomethyl-phosphatidylethanolamine, dimethyl-phosphatidylethanolamine, dielaidoyl- phosphatidylethanolamine (DEPE), stearoyloleoyl-phosphatidylethanolamine (SOPE), egg phosphatidylcholine (EPC), and mixtures thereof. In certain embodiments, the phospholipid is DPPC, DSPC, or mixtures thereof. In some embodiments, the non-cationic lipid (e.g., one or more phospholipids and / or cholesterol) may comprise from about 10 mol % to about 60 mol %, from about 15 mol % to about 60 mol %, from about 20 mol % to about 60 mol %, from about 25 mol % to about 60 mol %, from about 30 mol % to about 60 mol %, from about 10 mol % to about 55 mol %, from about 15 mol % to about 55 mol %, from about 20 mol % to about 55 mol %, from about 25 mol % to about 55 mol %, from about 30 mol % to about 55 mol %, from about 13 mol % to about 50 mol %, from about 15 mol % to about 50 mol % or from about 20 mol % to about 50 mol % of the total lipid present in the particle. When the non-cationic lipid is a mixture of a phospholipid and cholesterol or a cholesterol derivative, the mixture may comprise up to about 40, 50, or 60 mol % of the total lipid present in the particle. The conjugated lipid that inhibits aggregation of particles may comprise, e.g., one or more of the following: a polyethyleneglycol (PEG)-lipid conjugate, a polyamide (ATTA)- lipid conjugate, a cationic-polymer-lipid conjugates (CPLs), or mixtures thereof. In one particular embodiment, the nucleic acid-lipid particles comprise either a PEG-lipid conjugate or an ATTA-lipid conjugate. In certain embodiments, the PEG-lipid conjugate or ATTA-lipid conjugate is used together with a CPL. The conjugated lipid that inhibits aggregation of particles may comprise a PEG-lipid including, e.g., a PEG-diacylglycerol (DAG), a PEG dialkyloxypropyl (DAA), a PEG-phospholipid, a PEG-ceramide (Cer), or mixtures thereof. The PEG-DAA conjugate may be PEG-di lauryloxypropyl (C12), a PEG-dimyristyloxypropyl (C14), a PEG-dipalmityloxypropyl (C16), a PEG-distearyloxypropyl (C18), or mixtures thereof. Additional PEG-lipid conjugates suitable for use include, but are not limited to, mPEG2000-1,2-di-O-alkyl-sn3-carbomoylglyceride (PEG-C-DOMG). The synthesis of PEG- C-DOMG is described in PCT Application No. PCT / US08 / 88676. Yet additional PEG-lipid conjugates suitable for use include, without limitation, 1-[8′-(1,2-dimyristoyl-3-propanoxy)- carboxamido-3′,6′-dioxaoctanyl]carbamoyl-ω-methyl-poly(ethylene glycol) (2KPEG-DMG). The synthesis of 2KPEG-DMG is described in U.S. Pat. No.7,404,969. In some cases, the conjugated lipid that inhibits aggregation of particles (e.g., PEG- lipid conjugate) may comprise from about 0.1 mol % to about 2 mol %, from about 0.5 mol % to about 2 mol %, from about 1 mol % to about 2 mol %, from about 0.6 mol % to about 1.9 mol %, from about 0.7 mol % to about 1.8 mol %, from about 0.8 mol % to about 1.7 mol %, from about 1 mol % to about 1.8 mol %, from about 1.2 mol % to about 1.8 mol %, from about 1.2 mol % to about 1.7 mol %, from about 1.3 mol % to about 1.6 mol %, from about 1.4 mol % to about 1.5 mol %, or about 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, or 2 mol % (or any fraction thereof or range therein) of the total lipid present in the particle. Typically, in such instances, the PEG moiety has an average molecular weight of about 2,000 Daltons. In other cases, the conjugated lipid that inhibits aggregation of particles (e.g., PEG-lipid conjugate) may comprise from about 5.0 mol % to about 10 mol %, from about 5 mol % to about 9 mol %, from about 5 mol % to about 8 mol %, from about 6 mol % to about 9 mol %, from about 6 mol % to about 8 mol %, or about 5 mol %, 6 mol %, 7 mol %, 8 mol %, 9 mol %, or 10 mol % (or any fraction thereof or range therein) of the total lipid present in the particle. Typically, in such instances, the PEG moiety has an average molecular weight of about 750 Daltons. In some embodiments, the composition may comprise amphoteric liposomes, which contain at least one positive and at least one negative charge carrier, which differs from the positive one, the isoelectric point of the liposomes being between 4 and 8. This objective is accomplished owing to the fact that liposomes are prepared with a pH-dependent, changing charge. Liposomal structures with the desired properties are formed, for example, when the amount of membrane-forming or membrane-based cationic charge carriers exceeds that of the anionic charge carriers at a low pH and the ratio is reversed at a higher pH. This is always the case when the ionizable components have a pKa value between 4 and 9. As the pH of the medium drops, all cationic charge carriers are more charged and all anionic charge carriers lose their charge. Cationic compounds are useful in amphoteric liposomes. Without limitation, strongly cationic compounds can include, for example: DC-Chol 3-β-[N-(N′,N′-dimethylmethane) carbamoyl] cholesterol, TC-Chol 3-β-[N-(N′, N′, N...

Claims

CLAIMS What is claimed is:

1. An engineered T regulatory cell (Treg), further comprising a chimeric antigen receptor (CAR) comprising a CD19-binding domain.

2. The engineered Treg of claim 1, wherein the CD19-binding domain comprises: (i) a heavy chain variable domain (VH) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 126; and (ii) a light chain variable domain (VL) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO:

125.

3. The engineered Treg of claim 1 or claim 2, wherein the CD19-binding domain comprises: (i) a VH comprising an amino acid sequence of SEQ ID NO: 126; and (ii) a VL comprising an amino acid sequence of SEQ ID NO:

125.

4. The engineered Treg of claim 1, wherein the CD19-binding domain comprises an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO:

124.

5. The engineered Treg of claim 1 or claim 4, wherein the CD19-binding domain comprises an amino acid sequence of SEQ ID NO:

124.

6. The engineered Treg of claim 1, wherein the CD19-binding domain comprises: (i) a heavy chain variable domain (VH) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 35, SEQ ID NO: 44, SEQ ID NO: 53, SEQ ID NO: 62, SEQ ID NO: 75, SEQ ID NO: 86, SEQ ID NO: 96, SEQ ID NO: 106, SEQ ID NO: 116, 148, or 158; and (ii) a light chain variable domain (VL) comprising an amino acid sequence having at least 95% sequence identity to an amino acid sequence of SEQ ID NO: 34, SEQ ID NO: 43, SEQ ID NO: 52, SEQ ID NO: 61, SEQ ID NO: 74, SEQ ID NO: 85, SEQ ID NO: 95, SEQ ID NO: 105, SEQ ID NO: 115, 147, or 157.

7. The engineered Treg of claim 1, wherein the CD19-binding domain comprises: (i) a VH comprising an amino acid sequence of SEQ ID NO: 35, SEQ ID NO: 44, SEQ ID NO: 53, SEQ ID NO: 62, SEQ ID NO: 75, SEQ ID NO: 86, SEQ ID NO: 96, SEQ ID NO: 106, SEQ ID NO: 116, 148, or 158; and (ii) a VL comprising an amino acid sequence of SEQ ID NO: 34, SEQ ID NO: 43, SEQ ID NO: 52, SEQ ID NO: 61, SEQ ID NO: 74, SEQ ID NO: 85, SEQ ID NO: 95, SEQ ID NO: 105, SEQ ID NO: 115, 147, or 157.

8. The engineered Treg of claim 1, wherein the CD19-binding domain is a Fv, a scFv, a Fab fragment, a F(ab’) fragment, a F(ab’)2 fragment, an IgG, a camelid heavy chain antibody, a single domain VHH, or a bivalent VHH.

9. The engineered Treg of claim 1, wherein the CAR comprises a hinge, a transmembrane domain, a first costimulatory domain, a second costimulatory domain, and an intracellular signaling domain.

10. The engineered Treg of claim 9, wherein the hinge is an IgG4 hinge or a CD8 hinge.

11. The engineered Treg of claim 10, wherein the IgG4 hinge comprises an amino acid sequence that differs by no more than two amino acids from an amino acid sequence of ESKYGPPCPSCP (SEQ ID NO: 137).

12. The engineered Treg of claim 10, wherein the IgG4 hinge comprises an amino acid sequence of ESKYGPPCPSCP (SEQ ID NO: 137).

13. The engineered Treg of claim 10, wherein the IgG4 hinge comprises an amino acid sequence that differs by no more than two amino acids from an amino acid sequence of ESKYGPPCPSCPA (SEQ ID NO: 145).

14. The engineered Treg of claim 10, wherein the IgG4 hinge comprises an amino acid sequence of ESKYGPPCPSCPA (SEQ ID NO: 145).

15. The engineered Treg of claim 10, wherein the hinge is a CD8 hinge.

16. The engineered Treg of any one of claims 9-15, wherein the transmembrane domain is a transmembrane domain derived from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS.

17. The engineered Treg of claim 16, wherein the transmembrane domain is a CD28 transmembrane domain.

18. The engineered Treg of any one of claims 9-17, wherein the first costimulatory domain is derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, or CD137 (4-1BB), or a Toll-like receptor (TLR).

19. The engineered Treg of claim 18, wherein the first costimulatory domain is a CD28 costimulatory domain.

20. The engineered Treg of any one of claims 9-19, wherein the second costimulatory domain is derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI, NKG2C, NKG2D, or CD137 (4-1BB), or a TLR.

21. The engineered Treg of claim 20, wherein the second costimulatory domain is a CD3ζ costimulatory domain.

22. The engineered Treg of any one of claims 9-21, wherein the intracellular signaling domain is derived from OX40, CD2, CD3ζ, CD3γ, CD3δ, CD3ε, FCεRI, CD7, CD27, CD28, CD30, CD40, CD79a, CD79b, CD137 (4-1BB), ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, or a TLR.

23. The engineered Treg of claim 22, wherein the intracellular signaling domain is a CD3ζ intracellular signaling domain.

24. The engineered Treg of any one of claims 1-10, wherein the CAR comprises the CD19-binding domain, an IgG4 hinge, a transmembrane domain, a CD28 costimulatory domain, and a CD3ζ intracellular signaling domain.

25. The engineered Treg of claim 24, wherein the transmembrane domain is selected from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS.

26. The engineered Treg of claim 24 or 25, wherein the transmembrane domain is a CD8α transmembrane domain.

27. The engineered Treg of claim 26, wherein the CAR comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO:

70.

28. The engineered Treg of any one of claims 1-10, wherein the CAR comprises the CD19-binding domain, a CD28 hinge, a transmembrane domain, a CD28 costimulatory domain, and a CD3ζ intracellular signaling domain.

29. The engineered Treg of claim 28, wherein the transmembrane domain is selected from TCRα, TCRβ, TCRζ, CD3ε, CD3ζ, CD28, CD45, CD4, CD5, CD7, CD8, CD9, CD16, CD22, CD33, CD37, CD41, CD64, CD68, CD80, CD86, CD134, CD137 (4-1BB), CD154, or ICOS.

30. The engineered Treg of claim 28 or 29, wherein the transmembrane domain is a CD28 transmembrane domain.

31. The engineered Treg of claim 30, wherein the CAR comprises an amino acid sequence with at least 90% sequence identity to the amino acid sequence of SEQ ID NO:

69.

32. The engineered Treg of any one of claims 24-31, wherein the CAR further comprises a second costimulatory domain derived from OX40, CD2, CD3ζ, CD7, CD27, CD28, CD30, CD40, CD258, ICOS, GITR, IL-2Rβ, IL-2Rγ, IL-7Rα, CTLA4, PD-1, B7-H3, FCεRI,NKG2C, NKG2D, or CD137 (4-1BB), or a TLR, wherein the second costimulatory domain is either: (i) between the transmembrane domain and the CD28 costimulatory domain; or (ii) between the CD28 costimulatory domain and the CD3ζ intracellular signaling domain.

33. The engineered Treg of any one of claims 1-32, wherein the engineered Treg further comprises: (i) a nucleic acid encoding a first component of a chemically induced signaling complex (CISC), the first CISC component comprising: (a) a first extracellular domain comprising an FK506-binding protein (FKBP) domain that binds rapamycin; (b) a first transmembrane domain; and (c) a first cytoplasmic domain comprising an intracellular signaling domain of a first cytokine receptor; and (ii) a nucleic acid encoding a second CISC component, the second CISC component comprising: (a) a second extracellular domain comprising an FKBP-rapamycin-binding (FRB) domain; (b) a second transmembrane domain; and (c) a second cytoplasmic domain comprising an intracellular signaling domain of a second cytokine receptor, wherein the first and second CISC components dimerize in the presence of rapamycin.

34. The engineered Treg of claim 31, wherein the first CISC component comprises, in N- to-C-terminal order, an FKBP domain, an interleukin-2 receptor gamma (IL-2Rγ) transmembrane domain, and an IL-2Rγ cytoplasmic domain, and wherein the second CISC component comprises, in N-to-C-terminal order, an FRB domain, an interleukin-2 receptor beta (IL-2Rβ) transmembrane domain, and an IL-2Rβ cytoplasmic domain.

35. The engineered Treg of claim 34, wherein the engineered Treg comprises a nucleic acid encoding a soluble FRB domain.

36. The engineered Treg of claim 35, wherein the soluble FRB domain comprises an amino acid sequence having at least 90% sequence identity to an amino acid sequence of SEQ ID NO:

13.

37. The engineered Treg of claim 35 or 36, wherein the soluble FRB domain comprises an amino acid sequence of SEQ ID NO:

13.

38. The engineered Treg of claim 37, wherein the engineered Treg comprises a heterologous promoter located in a nucleic acid of the cell genome: (a) downstream from a regulatory T cell (Treg)-specific demethylated region (TSDR); and (b) upstream from a first coding exon of an endogenous FOXP3 gene.

39. The engineered Treg of claim 38, wherein the heterologous promoter located downstream from the TSDR is an MND promoter.

40. The engineered Treg of any one of claims 1-39, wherein the engineered Treg is FOXP3-positive, CD3-positive, CD4-positive, and CD25-positive.

41. The engineered Treg of any one of claims 1-40, wherein the engineered Treg is a sorted Treg.

42. A method comprising administering an engineered Treg to a subject in need thereof, wherein the engineered Treg comprises a CAR comprising a CD19-binding domain.

43. A method comprising administering the engineered Treg of any one of claims 1-41 to a subject in need thereof.

44. The method of claim 42 or 43, wherein 1 x 107to 1 x 1010engineered Tregs are administered.

45. The method of claim 44, wherein 5 x 108to 2 x 109engineered Tregs are administered.

46. The method of claim 45, wherein 8 x 108to 1.2 x 109engineered Tregs are administered.

47. The method of claim 46, wherein 1 x 109engineered Tregs are administered.

48. The method of any one of claims 42-47, wherein the engineered Tregs are administered intravenously.

49. The method of any one of claims 42-48, further comprising administering rapamycin to the subject.

50. The method of any one of claims 42-49, wherein the engineered Tregs are administered in combination with rapamycin.

51. The method of any one of claims 42-50, further comprising administering cyclophosphamide and / or fludarabine to the subject.

52. The method of any one of claims 42-51, wherein the engineered Tregs are administered in combination with cyclophosphamide and / or fludarabine.

53. The method of any one of claims 42-52, wherein administering the engineered Treg induces B cell suppression or depletion of active B cells in the subject.

54. The method of claim 53, wherein the B cell suppression comprises: (i) decreasing antibody production by plasmablasts and / or plasma cells; (ii) decreasing cytokine production by B cells; (iii) decreasing antigen-presenting capacity of B cells; (iv) reducing B cell differentiation; and / or (v) decreasing B cell proliferation.

55. The method of claim 53 or 54, wherein the B cells are CD19-positive B cells.

56. The method of any one of claims 53-55, wherein administering the engineered Treg does not induce B cell aplasia.

57. The method of any one of claims 53–56, wherein administering the engineered Treg reduces the abundance of circulating B cells that are activated, proliferating, and / or autoreactive in the subject.

58. The method of any one of claims 53–57, wherein administering the engineered Treg increases the abundance of naïve B cells in the subject.

59. The method of any one of claims 53–58, wherein administering the engineered Treg reduces the serum concentration of one or more antibodies that preferentially bind double- stranded DNA or Sm / RNP in the subject.

60. The method of any one of claims 53-59, wherein the subject is a human.

61. The method of any one of claims 53-60, wherein the subject has or is at risk of developing an autoimmune disease and / or an inflammatory disorder.

62. The method of claim 61, wherein the autoimmune disease is systemic lupus erythematosus (SLE) or Sjögren’s disease (SjD).

63. A method of manufacturing an engineered Treg, the method comprising introducing into a Treg a nucleic acid comprising a promoter operably linked to one or more of: (i) a nucleotide sequence encoding FOXP3 or a functional fragment thereof; (ii)(a) a nucleotide sequence encoding a first component of a chemically inducible signaling complex (CISC), the first component comprising: (1) a first extracellular domain comprising an FK506-binding protein (FKBP) domain that binds rapamycin, (2) a first transmembrane domain, and (3) a first cytoplasmic domain comprising an intracellular signaling domain of a first cytokine receptor; (ii)(b) a nucleotide sequence encoding a second component of the CISC, the second component comprising:(1) a second extracellular domain comprising an FKBP-rapamycin-binding (FRB) domain, (2) a second transmembrane domain, and (3) a second cytoplasmic domain comprising an intracellular signaling domain of a cytokine receptor; and / or (iii) a nucleotide sequence encoding a CAR comprising a CD19-binding domain.

64. A chimeric antigen receptor (CAR) polypeptide comprising an amino acid sequence having at least 90% identity to an amino acid sequence of SEQ ID NO: 69 or 70.

65. A CAR polypeptide comprising an amino acid sequence of SEQ ID NO: 69 or 70.

66. A cell comprising the CAR polypeptide of claim 64 or 65.

67. A method of manufacturing an engineered Treg, the method comprising contacting a T cell with a lipid delivery vehicle comprising a T cell-targeting moiety and one or more nucleic acids that collectively encode: (i) a nucleic acid encoding a chimeric antigen receptor (CAR) polypeptide comprising a CD19-binding domain; and (ii) FOXP3 or a functional fragment thereof.

68. The method of claim 67, wherein the one or more nucleic acids are messenger ribonucleic acids (mRNAs).

69. The method of claim 67 or 68, wherein the lipid delivery vehicle is a lipid nanoparticle (LNP).

70. The method of any one of claims 67–69, wherein the T cell is a CD4+ T cell and the T cell-targeting moiety is a CD4-targeting moiety.

71. The method of claim 70, wherein the CD4-targeting moiety is an anti-CD4 antibody or antigen-binding fragment thereof.

72. The method of claim 71, wherein the anti-CD4 antibody is selected from the group consisting of zanolimumab, keliximab, clenoliximab, tregalizumab, and ibalizumab.

73. A method of selectively depleting active B cells in a subject in need thereof, wherein the method comprises administering an engineered Treg to the subject, wherein the engineered Treg comprises a CAR comprising a CD19-binding domain.

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