WO2025193972A1 - Methods of using il-15 signaling to enhance nk and nk-like t cell function against aml - Google Patents

Abstract

Provided herein are chimeric innate receptor (OR) fusion proteins that enhance the persistence of NK and NK-like T cells in response to AML antigens, such as CD33. The OR fusion proteins of the present technology are useful in methods for treating cancer, such as AML. The OR fusion proteins of the present technology are useful in methods for treating AML.

Description

Atty. Dkt. No.: 115872-3215 METHODS OF USING IL-15 SIGNALING TO ENHANCE NK AND NK-LIKE T CELL FUNCTION AGAINST AML CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims the benefit of and priority to U.S. Provisional Application No.63/564,900, filed March 13, 2024, the contents of which are incorporated herein by reference in their entireties. TECHNICAL FIELD [0002] The present disclosure provides chimeric innate receptor (CIR) fusion proteins that enhance the persistence of NK and NK-like CD8 T cells in response to acute myeloid leukemia (AML) antigens, such as CD33. The CIR fusion proteins of the present technology are useful in methods for treating AML. GOVERNMENT SUPPORT [0003] This invention was made with government support under AI150999 awarded by the National Institutes of Health. The government has certain rights in the invention. BACKGROUND [0004] The following description of the background of the present technology is provided simply as an aid in understanding the present technology and is not admitted to describe or constitute prior art to the present technology. [0005] While CAR-T therapies have shown great success with B-cell malignancies, toxicities such as graft-versus-host-disease (GVHD) and cytokine release syndrome are still prevalent. Natural killer (NK) cells have been shown to be a promising target for engineered cell therapies because of their innate ability to recognize a broad range of malignancy, however the lack of in vivo persistence and transducability are limiting. SUMMARY OF THE PRESENT TECHNOLOGY [0006] In one aspect, the present disclosure provides a chimeric innate receptor (CIR) fusion protein comprising (i) an extracellular antigen binding domain that specifically binds to an AML antigen and (ii) a mammalian cytokine receptor polypeptide comprising a transmembrane domain and an intracellular domain, wherein the mammalian cytokine -1- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 receptor polypeptide comprises a mammalian CD122 polypeptide, a mammalian CD132 polypeptide, or a mammalian IL-21R polypeptide, and wherein the extracellular antigen binding domain is operably linked to the mammalian cytokine receptor polypeptide. In some embodiments, the extracellular antigen binding domain and the mammalian cytokine receptor polypeptide are directly linked or linked via a peptide linker. Additionally or alternatively, in certain embodiments, the AML antigen is selected from the group consisting of CD33, FLT3, CD123, and CD371. [0007] Additionally or alternatively, in some embodiments of the CIR fusion protein of the present technology, the extracellular antigen binding domain is located at the N- terminus of the mammalian cytokine receptor polypeptide. In certain embodiments, the mammalian cytokine receptor polypeptide comprises the amino acid sequence of SEQ ID NO: 21, SEQ ID NO: 22, or SEQ ID NO: 23. [0008] Additionally or alternatively, in certain embodiments, the CIR fusion protein of the present technology further comprises a hinge domain between the extracellular antigen binding domain and the mammalian cytokine receptor polypeptide. The hinge domain may comprise a CD8 hinge domain, a CD28 hinge domain, a CD122 hinge domain, or a CD132 hinge domain. In some embodiments, the hinge domain comprises the amino acid sequence of SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, or SEQ ID NO: 27. [0009] In any of the preceding embodiments, the CIR fusion protein of the present technology, further comprises a signal peptide located at the N-terminus of the extracellular antigen binding domain. In some embodiments, the signal peptide comprises the amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, or SEQ ID NO: 19. [0010] Additionally or alternatively, in some embodiments of the CIR fusion protein described herein, the extracellular antigen binding domain comprises a scFv (e.g., a human scFv). In some embodiments of the CIR fusion protein described herein, the scFv comprises the immunoglobulin variable heavy (VH) domain sequence of SEQ ID NO: 5 and the immunoglobulin variable light (VL) domain sequence of SEQ ID NO: 6. In other embodiments, the scFv comprises the immunoglobulin variable heavy (V_H) domain sequence of SEQ ID NO: 7 and the immunoglobulin variable light (V_L) domain sequence of -2- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 SEQ ID NO: 8. In any of the foregoing embodiments, the CIR fusion protein of the present technology comprises an amino acid sequence selected from any one of SEQ ID NOs: 28- 35. [0011] In one aspect, the present disclosure provides a recombinant nucleic acid molecule encoding any and all embodiments of the CIR fusion protein disclosed herein. In another aspect, the present disclosure provides an expression vector comprising any and all embodiments of the recombinant nucleic acid molecule described herein. In yet another aspect, the present disclosure provides a host cell comprising any and all embodiments of the recombinant nucleic acid molecule described herein or any and all embodiments of the expression vector described herein. [0012] In another aspect, the present disclosure provides an engineered immune cell comprising any and all embodiments of the CIR fusion protein disclosed herein. In some embodiments, the engineered immune cell is a NK cell or a NK-like T cell. [0013] In one aspect, the present disclosure provides a composition comprising any and all embodiments of the CIR fusion protein disclosed herein or any and all embodiments of the engineered immune cell disclosed herein, and a pharmaceutically-acceptable carrier. [0014] In another aspect, the present disclosure provides a method for treating acute myelogenous leukemia (AML) in a subject in need thereof comprising administering to the subject an effective amount of any and all embodiments of the engineered immune cell disclosed herein. In some embodiments, the method further comprises administering to the subject an effective amount of a cytokine, wherein the cytokine is a cognate ligand of the mammalian cytokine receptor polypeptide. In certain embodiments, the cytokine is IL-15 or IL-21. Additionally or alternatively, in some embodiments, the method further comprises separately, sequentially or simultaneously administering an additional therapeutic agent to the subject. [0015] Also provided herein are kits comprising any and all embodiments of the recombinant nucleic acid molecule described herein or any and all embodiments of the expression vector described herein, and instructions for use. [0016] Also disclosed herein are methods for preparing immune cells for adoptive cell therapy (ACT) comprising: (a) isolating immune cells from a donor subject, (b) transducing -3- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 the immune cells with any and all embodiments of the recombinant nucleic acid molecule or the expression vector described herein, and (c) administering the transduced immune cells to a recipient subject. The donor subject and the recipient subject may be the same or different. In some embodiments, the immune cells are NK cells, or NK-like CD8 T cells. BRIEF DESCRIPTION OF THE DRAWINGS [0017] FIG.1. Schematic representation of classical NK receptors found on NKG2C+ CD8-T cells. [0018] FIG.2. In vivo survival of NALM6 tumor-bearing mice following treatment with CAR-engineered NKG2C+ CD8 T cells, NKG2C- CD8 T cells, or NK cells. [0019] FIG.3. Quantification of median fluorescence intensity of CD122 (left) and CD132 (right) by flow cytometry of whole PBMCs taken from three healthy donors with 10ng/mL of IL-15. [0020] FIG.4. Quantification of pSTAT5 levels expressed as a frequency (left) and median fluorescence intensity (right) by flow cytometry following in vitro stimulation of whole PBMCs taken from three healthy donors with 10ng/mL of IL-15. [0021] FIG.5. In vitro activation of whole PBMCs taken from three healthy donors following overnight stimulation with 10ng/mL IL-15. Frequencies of total NKG2D, 2B4 and CD16+ cells per cell subtype (left) and MFI (right) were quantified with flow cytometry. [0022] FIG.6. Anti-CD33-CD122 chimeric innate receptor designs (top) containing three different hinge domains, CD8h, CD28h or CD122h. Schematic representation (bottom) of STAT5 phosphorylation and downstream signaling in response to recognition of CD33 by engineered receptor. [0023] FIG.7. Representative dot plots of non-transduced cells (grey) and cells engineered with the CD8h construct (red), CD28h construct (orange) and CD122h construct (green). Cells were stained with an F(ab)2 AF647 antibody and expression was compared to transduction efficiency measured by an mCherry reporter. [0024] FIG.8. Quantification of pSTAT5+ levels in engineered NKG2C+ CD8 T cells (left) and NK cells (right) expressed as a frequency of live cells (top) following in vitro -4- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 stimulation with 10ng/mL IL-15 (pink) or 10μg recombinant CD33 (blue). In IL-15 stimulated cells MFI was quantified (bottom), and NT cells (grey) was compared to cells engineeered with CD8h (red), CD28h (orange) or CD122h (green). DETAILED DESCRIPTION [0025] It is to be appreciated that certain aspects, modes, embodiments, variations and features of the present methods are described below in various levels of detail in order to provide a substantial understanding of the present technology. [0026] In practicing the present methods, many conventional techniques in molecular biology, protein biochemistry, cell biology, immunology, microbiology and recombinant DNA are used. See, e.g., Sambrook and Russell eds. (2001) Molecular Cloning: A Laboratory Manual, 3rd edition; the series Ausubel et al. eds. (2007) Current Protocols in Molecular Biology; the series Methods in Enzymology (Academic Press, Inc., N.Y.); MacPherson et al. (1991) PCR 1: A Practical Approach (IRL Press at Oxford University Press); MacPherson et al. (1995) PCR 2: A Practical Approach; Harlow and Lane eds. (1999) Antibodies, A Laboratory Manual; Freshney (2005) Culture of Animal Cells: A Manual of Basic Technique, 5th edition; Gait ed. (1984) Oligonucleotide Synthesis; U.S. Patent No.4,683,195; Hames and Higgins eds. (1984) Nucleic Acid Hybridization; Anderson (1999) Nucleic Acid Hybridization; Hames and Higgins eds. (1984) Transcription and Translation; Immobilized Cells and Enzymes (IRL Press (1986)); Perbal (1984) A Practical Guide to Molecular Cloning; Miller and Calos eds. (1987) Gene Transfer Vectors for Mammalian Cells (Cold Spring Harbor Laboratory); Makrides ed. (2003) Gene Transfer and Expression in Mammalian Cells; Mayer and Walker eds. (1987) Immunochemical Methods in Cell and Molecular Biology (Academic Press, London); and Herzenberg et al. eds (1996) Weir’s Handbook of Experimental Immunology. Methods to detect and measure levels of polypeptide gene expression products (i.e., gene translation level) are well-known in the art and include the use of polypeptide detection methods such as antibody detection and quantification techniques. (See also, Strachan & Read, Human Molecular Genetics, Second Edition. (John Wiley and Sons, Inc., NY, 1999)). [0027] With the recent discovery of NK-like CD8 T cells that harbor both NK and T cell properties (Sottile, Rosa, et al., Science Immunology, vol.6, no.63, 24 (2021)), there is -5- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 new potential to engineer these cells in for adoptive therapies that takes advantage of the adaptive killing and innate recognition, while avoiding on-tumor off-target toxicities. Recent literature has described the existence of NK-like CD8 T cells that have a T cell-like transcriptional profile, including the expression of a TCR, but also express classical NK receptors. NKG2C+ CD8 Ts are one such NK-like cell (FIG.1), that arise and expand following human cytomegalovirus (HCMV) infection. [0028] CAR-engineered version of these cells exhibit a higher killing capacity against acute myeloid leukemia (AML) than their conventional counterparts, and this effect is enhanced in the presence of IL-15 (FIG.2). IL-15 is a cytokine that binds to CD122 and common gamma chain (CD132) and leads to the proliferation and survival of NK cells. IL- 15 has been shown to have adverse effects when administered in vivo. Thus, there is increasing interest in harnessing innate proliferative signals in these NKG2C+ NK-like CD8 T cells. [0029] We aim to utilize the innate-like properties of NKG2C+ and other NK-like CD8 T cells by generating an engineered cell capable of rapid expansion in response to IL-15 signaling, while relying on their natural cytotoxicity for AML killing. As described herein, the responsiveness of NK-like T cells to IL-15 stimulation was examined and chimeric innate receptors (CIRs) that induce the IL-15 signaling pathway in response to CD33 recognition, a common AML antigen, was designed and validated. Definitions [0030] Unless defined otherwise, all technical and scientific terms used herein have the meaning commonly understood by a person skilled in the art to which this disclosure belongs. The following references provide one of skill with a general definition of many of the terms used in the present disclosure. Singleton et al., Dictionary of Microbiology and Molecular Biology (2nd ed.1994); The Cambridge Dictionary of Science and Technology (Walker ed., 1988); The Glossary of Genetics, 5th Ed., R. Rieger et al. (eds.), Springer Verlag (1991); and Hale & Marham, The Harper Collins Dictionary of Biology (1991). As used herein, the following terms have the meanings ascribed to them below, unless specified otherwise. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the disclosure. -6- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [0031] All numerical designations, e.g., pH, temperature, time, concentration, and molecular weight, including ranges, are approximations which are varied (+) or (−) by increments of 1.0 or 0.1, as appropriate or alternatively by a variation of +/− 20% or +/− 15%, or alternatively 10% or alternatively 5% or alternatively 2%. As will be understood by one skilled in the art, for any and all purposes, all ranges disclosed herein also encompass any and all possible subranges and combinations of subranges thereof. Furthermore, as will be understood by one skilled in the art, a range includes each individual member. [0032] As used herein, the singular forms “a”, “an” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise. [0033] As used herein, the term “about” or “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, “about” can mean within 3 or more than 3 standard deviations, per the practice in the art. Alternatively, “about” can mean a range of up to 20%, up to 10%, up to 5%, or up to 1% of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, within 5-fold, or within 2-fold, of a value. [0034] As used herein, the term “administration” of an agent to a subject includes any route of introducing or delivering the agent to a subject to perform its intended function. Administration can be carried out by any suitable route, including, but not limited to, intravenously, intramuscularly, intraperitoneally, subcutaneously, and other suitable routes as described herein. Administration includes self-administration and the administration by another. “Administration” of a cell or vector or other agent and compositions containing same can be performed in one dose, continuously or intermittently throughout the course of treatment. Methods of determining the most effective means and dosage of administration are known to those of skill in the art and will vary with the composition used for therapy, the purpose of the therapy, the target cell being treated, and the subject being treated. Single or multiple administrations can be carried out with the dose level and pattern being selected by the treating physician or in the case of animals, by the treating veterinarian. In some embodiments, administering or a grammatical variation thereof also refers to more -7- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 than one doses with certain interval. In some embodiments, the interval is 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 10 days, 2 weeks, 3 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 1 year or longer. In some embodiments, one dose is repeated for once, twice, three times, four times, five times, six times, seven times, eight times, nine times, ten times or more. Suitable dosage formulations and methods of administering the agents are known in the art. Route of administration can also be determined and method of determining the most effective route of administration are known to those of skill in the art and will vary with the composition used for treatment, the purpose of the treatment, the health condition or disease stage of the subject being treated, and target cell or tissue. Non-limiting examples of route of administration include oral administration, intraperitoneal, infusion, nasal administration, inhalation, injection, and topical application. In some embodiments, the administration is an infusion (for example to peripheral blood of a subject) over a certain period of time, such as about 30 minutes, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, about 11 hours, about 12 hours, about 24 hours or longer. [0035] As used herein “adoptive cell therapeutic composition” refers to any composition comprising cells suitable for adoptive cell transfer. In exemplary embodiments, the adoptive cell therapeutic composition comprises peripheral blood mononuclear cells. In one embodiment, the adoptive cell therapeutic composition comprises NK cells and NK-like T cells. [0036] The term “amino acid” refers to naturally occurring and non-naturally occurring amino acids, as well as amino acid analogs and amino acid mimetics that function in a manner similar to the naturally occurring amino acids. Naturally encoded amino acids are the 20 common amino acids (alanine, arginine, asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine) and pyrolysine and selenocysteine. Amino acid analogs refer to agents that have the same basic chemical structure as a naturally occurring amino acid, i.e., an α carbon that is bound to a hydrogen, a carboxyl group, an amino group, and an R group, such as, homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. Such analogs have modified R groups -8- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 (such as, norleucine) or modified peptide backbones, but retain the same basic chemical structure as a naturally occurring amino acid. In some embodiments, amino acids forming a polypeptide are in the D form. In some embodiments, the amino acids forming a polypeptide are in the L form. In some embodiments, a first plurality of amino acids forming a polypeptide is in the D form and a second plurality is in the L form. [0037] Amino acids are referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides, likewise, are referred to by their commonly accepted single-letter code. [0038] As used herein, the term “analog” refers to a structurally related polypeptide or nucleic acid molecule having the function of a reference polypeptide or nucleic acid molecule. [0039] As used herein, the term “antibody” means not only intact antibody molecules, but also fragments of antibody molecules that retain immunogen-binding ability. Such fragments are also well known in the art and are regularly employed both in vitro and in vivo. Accordingly, as used herein, the term “antibody” means not only intact immunoglobulin molecules but also the well-known active fragments F(ab')2, and Fab. F(ab')2, and Fab fragments that lack the Fc fragment of intact antibody, clear more rapidly from the circulation, and may have less non-specific tissue binding of an intact antibody (Wahl et al., J. Nucl. Med.24:316-325 (1983)). Antibodies may comprise whole native antibodies, monoclonal antibodies, human antibodies, humanized antibodies, camelised antibodies, multispecific antibodies, bispecific antibodies, chimeric antibodies, Fab, Fab', single chain V region fragments (scFv), single domain antibodies (e.g., nanobodies and single domain camelid antibodies), VNAR fragments, Bi-specific T-cell engager (BiTE) antibodies, minibodies, disulfide-linked Fvs (sdFv), and anti-idiotypic (anti-Id) antibodies, intrabodies, fusion polypeptides, unconventional antibodies and antigen binding fragments of any of the above. In particular, antibodies include immunoglobulin molecules and immunologically active fragments of immunoglobulin molecules, i.e., molecules that contain an antigen binding site. Immunoglobulin molecules can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, and IgY), class (e.g., IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2), or subclass. -9- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [0040] In certain embodiments, an antibody is a glycoprotein comprising at least two heavy (H) chains and two light (L) chains inter-connected by disulfide bonds. Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as V_H) and a heavy chain constant (CH) region. The heavy chain constant region is comprised of three domains, CH1, CH2, and CH3. Each light chain is comprised of a light chain variable region (abbreviated herein as V_L) and a light chain constant C_L region. The light chain constant region is comprised of one domain, C_L. The V_H and V_L regions can be further subdivided into regions of hypervariability, termed complementarity determining regions (CDR), interspersed with regions that are more conserved, termed framework regions (FR). Each V_H and V_L is composed of three CDRs and four FRs arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The variable regions of the heavy and light chains contain a binding domain that interacts with an antigen. The constant regions of the antibodies may mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g., effector cells) and the first component (Cl q) of the classical complement system. As used herein interchangeably, the terms “antigen binding portion”, “antigen binding fragment”, or “antigen binding region” of an antibody, refer to the region or portion of an antibody that binds to the antigen and which confers antigen specificity to the antibody; fragments of antigen binding proteins, for example antibodies, include one or more fragments of an antibody that retain the ability to specifically bind to an antigen. It has been shown that the antigen binding function of an antibody can be performed by fragments of a full-length antibody. Examples of antigen binding portions encompassed within the term “antibody fragments” of an antibody include a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CH1 domains; a F(ab)2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; a Fd fragment consisting of the V_H and C_H1 domains; a Fv fragment consisting of the V_L and V_H domains of a single arm of an antibody; a dAb fragment (Ward et al., Nature 341 : 544-546 (1989)), which consists of a VH domain; and an isolated complementarity determining region (CDR). An “isolated antibody” or “isolated antigen binding protein” is one which has been identified and separated and/or recovered from a component of its natural environment. “Synthetic antibodies” or “recombinant antibodies” are generally generated -10- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 using recombinant technology or using peptide synthetic techniques known to those of skill in the art. [0041] Antibodies and antibody fragments can be wholly or partially derived from mammals (e.g., humans, non-human primates, goats, guinea pigs, hamsters, horses, mice, rats, rabbits and sheep) or non-mammalian antibody producing animals (e.g., chickens, ducks, geese, snakes, and urodele amphibians). The antibodies and antibody fragments can be produced in animals or produced outside of animals, such as from yeast or phage (e.g., as a single antibody or antibody fragment or as part of an antibody library). [0042] Furthermore, although the two domains of the Fv fragment, V_L and V_H, are coded for by separate genes, they can be joined, using recombinant methods, by a synthetic linker that enables them to be made as a single protein chain in which the VL and VH regions pair to form monovalent molecules. These are known as single chain Fv (scFv); see e.g., Bird et al., Science 242:423-426 (1988); and Huston et al., Proc. Natl. Acad. Sci.85 : 5879-5883 (1988). These antibody fragments are obtained using conventional techniques known to those of ordinary skill in the art, and the fragments are screened for utility in the same manner as are intact antibodies. [0043] As used herein, the term “single-chain variable fragment” or “scFv” is a fusion protein of the variable regions of the heavy (VH) and light chains (VL) of an immunoglobulin (e.g., mouse or human) covalently linked to form a V_H::V_L heterodimer. The heavy (V_H) and light chains (V_L) are either joined directly or joined by a peptide-encoding linker (e.g., about 10, 15, 20, 25 amino acids), which connects the N-terminus of the VH with the C-terminus of the V_L, or the C-terminus of the V_H with the N-terminus of the V_L. The linker is usually rich in glycine for flexibility, as well as serine or threonine for solubility. The linker can link the heavy chain variable region and the light chain variable region of the extracellular antigen binding domain. In certain embodiments, the linker comprises amino acids having GGGGSGGGGSGGGGS (SEQ ID NO: 1). In certain embodiments, the nucleic acid sequence encoding the amino acid sequence of SEQ ID NO: 1 is ggcggcggcggatctggaggtggtggctcaggtggcggaggctcc (SEQ ID NO: 2). [0044] Despite removal of the constant regions and the introduction of a linker, scFv proteins retain the specificity of the original immunoglobulin. Single chain Fv polypeptide -11- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 antibodies can be expressed from a nucleic acid comprising VH- and VL-encoding sequences as described by Huston, et al. (Proc. Nat. Acad. Sci. USA, 85:5879-5883 (1988)). See, also, U.S. Patent Nos. 5,091,513, 5,132,405 and 4,956,778; and U.S. Patent Publication Nos. 20050196754 and 20050196754. Antagonistic scFvs having inhibitory activity have been described (see, e.g., Zhao et al., Hybridoma (Larchmt) 27(6):455-51 (2008); Peter et al., J Cachexia Sarcopenia Muscle (2012); Shieh et al., J Imunol 183(4):2277-85 (2009); Giomarelli et al., Thromb Haemost 97(6):955-63 (2007); Fife eta., J Clin Invst 116(8):2252- 61 (2006); Brocks et al., Immunotechnology 3(3): 173-84 (1997); Moosmayer et al., Ther Immunol 2(10):31- 40 (1995). Agonistic scFvs having stimulatory activity have been described (see, e.g., Peter et al., J Biol Chem 25278(38):36740-7 (2003); Xie et al., Nat Biotech 15(8):768-71 (1997); Ledbetter et al., Crit Rev Immunol 17(5-6):427-55 (1997); Ho et al., Bio Chim Biophys Acta 1638(3):257-66 (2003)). [0045] As used herein, an “antigen” refers to a molecule to which an antibody can selectively bind. The target antigen may be a protein (e.g., an antigenic peptide), carbohydrate, nucleic acid, lipid, hapten, or other naturally occurring or synthetic compound. An antigen may also be administered to an animal subject to generate an immune response in the subject. [0046] As used herein, a “cancer” is a disease state characterized by the presence in a subject of cells demonstrating abnormal uncontrolled replication and in some aspects, the term may be used interchangeably with the term “tumor.” The term “cancer or tumor antigen” refers to an antigen known to be associated and expressed in a cancer cell or tumor cell (such as on the cell surface) or tissue, and the term “cancer or tumor targeting antibody” refers to an antibody that targets such an antigen. In some embodiments, the cancer or tumor antigen is not expressed in a non-cancer cell or tissue. In some embodiments, the cancer or tumor antigen is expressed in a non-cancer cell or tissue at a level significantly lower compared to a cancer cell or tissue. In some embodiments, the cancer is a solid tumor. In other embodiments, the cancer is not a solid tumor. In some embodiments, the cancer is from a carcinoma, a sarcoma, a myeloma, a leukemia, or a lymphoma. In some embodiments, the cancer is a primary cancer or a metastatic cancer. In some embodiments, the cancer is a relapsed cancer. In some embodiments, the cancer reaches a remission, but can relapse. In some embodiments, the cancer is unresectable. -12- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [0047] As used herein, the term “cell population” refers to a group of at least two cells expressing similar or different phenotypes. In non-limiting examples, a cell population can include at least about 10, at least about 100, at least about 200, at least about 300, at least about 400, at least about 500, at least about 600, at least about 700, at least about 800, at least about 900, at least about 1000 cells, at least about 10,000 cells, at least about 100,000 cells, at least about 1×10⁶ cells, at least about 1×10⁷ cells, at least about 1×10⁸ cells, at least about 1×10⁹ cells, at least about 1×10¹⁰ cells, at least about 1×10¹¹ cells, at least about 1×10¹² cells, or more cells expressing similar or different phenotypes. [0048] As used herein, a “cleavable peptide”, which is also referred to as a “cleavable linker,” means a peptide that can be cleaved, for example, by an enzyme. One translated polypeptide comprising such cleavable peptide can produce two final products, therefore, allowing expressing more than one polypeptides from one open reading frame. One example of cleavable peptides is a self-cleaving peptide, such as a 2A self-cleaving peptide. 2A self-cleaving peptides, is a class of 18-22 aa-long peptides, which can induce the cleaving of the recombinant protein in a cell. In some embodiments, the 2A self-cleaving peptide is selected from P2A, T2A, E2A, F2A and BmCPV2A. See, for example, Wang Y, et al. Sci Rep.2015;5:16273. Published 2015 Nov 5. As used herein, the terms “T2A” and “2A peptide” are used interchangeably to refer to any 2A peptide or fragment thereof, any 2A-like peptide or fragment thereof, or an artificial peptide comprising the requisite amino acids in a relatively short peptide sequence (on the order of 20 amino acids long depending on the virus of origin) containing the consensus polypeptide motif D-V/I-E-X-N-P-G-P (SEQ ID NO: 3), wherein X refers to any amino acid generally thought to be self-cleaving. [0049] As used herein, “complementary” sequences refer to two nucleotide sequences which, when aligned anti-parallel to each other, contain multiple individual nucleotide bases which pair with each other. Paring of nucleotide bases forms hydrogen bonds and thus stabilizes the double strand structure formed by the complementary sequences. It is not necessary for every nucleotide base in two sequences to pair with each other for sequences to be considered “complementary”. Sequences may be considered complementary, for example, if at least 30%, 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% of the nucleotide bases in two sequences pair with each other. In some embodiments, the term complementary refers to 100% of the nucleotide bases in two -13- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 sequences pair with each other. In addition, sequences may still be considered “complementary” when the total lengths of the two sequences are significantly different from each other. For example, a primer of 15 nucleotides may be considered “complementary” to a longer polynucleotide containing hundreds of nucleotides if multiple individual nucleotide bases of the primer pair with nucleotide bases in the longer polynucleotide when the primer is aligned anti-parallel to a particular region of the longer polynucleotide. Nucleotide bases paring is known in the field, such as in DNA, the purine adenine (A) pairs with the pyrimidine thymine (T) and the pyrimidine cytosine (C) always pairs with the purine guanine (G); while in RNA, adenine (A) pairs with uracil (U) and guanine (G) pairs with cytosine (C). Further, the nucleotide bases aligned anti-parallel to each other in two complementary sequences, but not a pair, are referred to herein as a mismatch. [0050] A “composition” is intended to mean a combination of active agent and another compound or composition, inert (for example, a nanoparticle, detectable agent or label) or active, such as an adjuvant, diluent, binder, stabilizer, buffers, salts, lipophilic solvents, preservative, adjuvant or the like and include carriers, such as pharmaceutically acceptable carriers. In some embodiments, the carrier (such as the pharmaceutically acceptable carrier) comprises, or consists essentially of, or yet further consists of a nanoparticle, such as an polymeric nanoparticle carrier or an lipid nanoparticle that can be used alone or in combination with another carrier, such as an adjuvant or solvent. Carriers also include pharmaceutical excipients and additives proteins, peptides, amino acids, lipids, and carbohydrates (e.g., sugars, including monosaccharides, di-, tri, tetra-oligosaccharides, and oligosaccharides; derivatized sugars such as alditols, aldonic acids, esterified sugars and the like; and polysaccharides or sugar polymers), which can be present singly or in combination, comprising alone or in combination 1-99.99% by weight or volume. Exemplary protein excipients include serum albumin such as human serum albumin (HSA), recombinant human albumin (rHA), gelatin, casein, and the like. Representative amino acid components, which can also function in a buffering capacity, include alanine, arginine, glycine, arginine, betaine, histidine, glutamic acid, aspartic acid, cysteine, lysine, leucine, isoleucine, valine, methionine, phenylalanine, aspartame, and the like. Carbohydrate excipients are also intended within the scope of this technology, examples of which include -14- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 but are not limited to monosaccharides such as fructose, maltose, galactose, glucose, D- mannose, sorbose, and the like; disaccharides, such as lactose, sucrose, trehalose, cellobiose, and the like; polysaccharides, such as raffinose, melezitose, maltodextrins, dextrans, starches, and the like; and alditols, such as mannitol, xylitol, maltitol, lactitol, xylitol sorbitol (glucitol) and myoinositol. A composition as disclosed herein can be a pharmaceutical composition. A “pharmaceutical composition” is intended to include the combination of an active agent with a carrier, inert or active, making the composition suitable for diagnostic or therapeutic use in vitro, in vivo or ex vivo. [0051] As used herein, a “control” is an alternative sample used in an experiment for comparison purpose. A control can be “positive” or “negative.” For example, where the purpose of the experiment is to determine a correlation of the efficacy of a therapeutic agent for the treatment for a particular type of disease, a positive control (a composition known to exhibit the desired therapeutic effect) and a negative control (a subject or a sample that does not receive the therapy or receives a placebo) are typically employed. [0052] As used herein, the term “effective amount” or “therapeutically effective amount” refers to a quantity of an agent sufficient to achieve a desired therapeutic effect. In the context of therapeutic applications, the amount of a therapeutic agent administered to the subject can depend on the type and severity of the infection and on the characteristics of the individual, such as general health, age, sex, body weight and tolerance to drugs. It can also depend on the degree, severity and type of disease. The skilled artisan will be able to determine appropriate dosages depending on these and other factors. [0053] As used herein, the term “engineered immune cell” refers to an immune cell that is genetically modified. [0054] As used herein, the term “expression” refers to the process by which polynucleotides are transcribed into mRNA and/or the process by which the transcribed mRNA is subsequently being translated into peptides, polypeptides, or proteins. If the polynucleotide is derived from genomic DNA, expression can include splicing of the mRNA in a eukaryotic cell. The expression level of a gene can be determined by measuring the amount of mRNA or protein in a cell or tissue sample. In one aspect, the expression level of a gene from one sample can be directly compared to the expression level of that -15- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 gene from a control or reference sample. In another aspect, the expression level of a gene from one sample can be directly compared to the expression level of that gene from the same sample following administration of the compositions disclosed herein. The term “expression” also refers to one or more of the following events: (1) production of an RNA template from a DNA sequence (e.g., by transcription) within a cell; (2) processing of an RNA transcript (e.g., by splicing, editing, 5’ cap formation, and/or 3’ end formation) within a cell; (3) translation of an RNA sequence into a polypeptide or protein within a cell; (4) post-translational modification of a polypeptide or protein within a cell; (5) presentation of a polypeptide or protein on the cell surface; and (6) secretion or presentation or release of a polypeptide or protein from a cell. [0055] As used herein, an "expression vector" includes vectors capable of expressing DNA that is operatively linked with regulatory sequences, such as promoter regions, that are capable of effecting expression of such DNA fragments. Such additional segments can include promoter and terminator sequences, and optionally can include one or more origins of replication, one or more selectable markers, an enhancer, a polyadenylation signal, and the like. Expression vectors are generally derived from plasmid or viral DNA, or can contain elements of both. Thus, an expression vector refers to a recombinant DNA or RNA construct, such as a plasmid, a phage, recombinant virus or other vector that, upon introduction into an appropriate host cell, results in expression of the cloned DNA. Appropriate expression vectors are well known to those of skill in the art and include those that are replicable in eukaryotic cells and/or prokaryotic cells and those that remain episomal or those which integrate into the host cell genome. [0056] As used herein, the term “heterologous nucleic acid molecule or polypeptide” refers to a nucleic acid molecule (e.g., a cDNA, DNA or RNA molecule) or polypeptide that is either not normally expressed or is expressed at an aberrant level in a cell or sample obtained from a cell. This nucleic acid can be from another organism, or it can be, for example, an mRNA molecule that is not normally expressed in a cell or sample. [0057] As used herein, the term “immune cell” refers to any cell that plays a role in the immune response of a subject. Immune cells are of hematopoietic origin, and include lymphocytes, such as B cells and T cells ; natural killer cells; myeloid cells, such as -16- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 monocytes, macrophages, dendritic cells, eosinophils, neutrophils, mast cells, basophils, and granulocytes. [0058] As used herein, the term “linker” refers to any amino acid sequence comprising from a total of 1 to 200 amino acid residues; or about 1 to 10 amino acid residues, or alternatively 8 amino acids, or alternatively 6 amino acids, or alternatively 5 amino acids that may be repeated from 1 to 10, or alternatively to about 8, or alternatively to about 6, or alternatively to about 5, or alternatively, to about 4, or alternatively to about 3, or alternatively to about 2 times. For example, the linker may comprise up to 15 amino acid residues consisting of a pentapeptide repeated three times. In one embodiment, the linker sequence is a (G4S)n (SEQ ID NO: 4), wherein n is 1, or 2, or 3, or 4, or 5, or 6, or 7, or 8, or 9, or 10, or 11, or 12, or 13, or 14, or 15. [0059] As used herein, the term “native immune cell” refers to an immune cell that naturally occurs in the immune system. [0060] As used herein, "operably linked" with reference to nucleic acid sequences, regions, elements or domains means that the nucleic acid regions are functionally related to each other. For example, nucleic acid encoding a leader peptide can be operably linked to nucleic acid encoding a polypeptide, whereby the nucleic acids can be transcribed and translated to express a functional fusion protein, wherein the leader peptide effects secretion of the fusion polypeptide. In some instances, the nucleic acid encoding a first polypeptide (e.g., a leader peptide) is operably linked to nucleic acid encoding a second polypeptide and the nucleic acids are transcribed as a single mRNA transcript, but translation of the mRNA transcript can result in one of two polypeptides being expressed. For example, an amber stop codon can be located between the nucleic acid encoding the first polypeptide and the nucleic acid encoding the second polypeptide, such that, when introduced into a partial amber suppressor cell, the resulting single mRNA transcript can be translated to produce either a fusion protein containing the first and second polypeptides, or can be translated to produce only the first polypeptide. In another example, a promoter can be operably linked to nucleic acid encoding a polypeptide, whereby the promoter regulates or mediates the transcription of the nucleic acid. -17- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [0061] As used herein, the “percent homology” between two amino acid sequences is equivalent to the percent identity between the two sequences. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology = # of identical positions/total # of positions × 100), taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. [0062] The percent homology between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4: 11-17 (1988)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In addition, the percent homology between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol.48:444-453 (1970)) algorithm which has been incorporated into the GAP program in the GCG software package (available at www.gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6. [0063] Additionally or alternatively, the amino acids sequences of the presently disclosed subject matter can further be used as a “query sequence” to perform a search against public databases to, for example, identify related sequences. Such searches can be performed using the XBLAST program (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215 :403-10. BLAST protein searches can be performed with the XBLAST program, score = 50, wordlength = 3 to obtain amino acid sequences homologous to the specified sequences disclosed herein. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al., (1997) Nucleic Acids Res. 25(17):3389-3402. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used. [0064] “Pharmaceutically acceptable carriers” refers to any diluents, excipients, or carriers that may be used in the compositions disclosed herein. In some embodiments, a pharmaceutically acceptable carrier comprises, or consists essentially of, or yet further consists of a nanoparticle, such as an polymeric nanoparticle carrier or an lipid nanoparticle (LNP). Additionally or alternatively, pharmaceutically acceptable carriers include ion -18- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances, such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose- based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. Suitable pharmaceutical carriers are described in Remington's Pharmaceutical Sciences, Mack Publishing Company, a standard reference text in this field. They can be selected with respect to the intended form of administration, that is, oral tablets, capsules, elixirs, syrups and the like, and consistent with conventional pharmaceutical practices. [0065] The terms “polynucleotide”, “nucleic acid” and “oligonucleotide” are used interchangeably and refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides or analogs thereof. Polynucleotides can have any three-dimensional structure and may perform any function, known or unknown. The following are non-limiting examples of polynucleotides: a gene or gene fragment (for example, a probe, primer, EST or SAGE tag), exons, introns, messenger RNA (mRNA), transfer RNA, ribosomal RNA, ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, nucleic acid probes and primers. A polynucleotide can comprise modified nucleotides, such as methylated nucleotides and nucleotide analogs. If present, modifications to the nucleotide structure can be imparted before or after assembly of the polynucleotide. The sequence of nucleotides can be interrupted by non-nucleotide components. A polynucleotide can be further modified after polymerization, such as by conjugation with a labeling component. The term also refers to both double- and single-stranded molecules. Unless otherwise specified or required, any embodiment of this disclosure that is a polynucleotide encompasses both the double-stranded form and each of two complementary single-stranded forms known or predicted to make up the double-stranded form. A polynucleotide is composed of a specific sequence of four nucleotide bases: adenine (A); cytosine (C); guanine (G); thymine (T); and uracil (U) for thymine when the polynucleotide is RNA. Thus, the term “polynucleotide sequence” is the -19- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 alphabetical representation of a polynucleotide molecule. This alphabetical representation can be input into databases in a computer having a central processing unit and used for bioinformatics applications such as functional genomics and homology searching. [0066] The terms “polypeptide,” “peptide,” and “protein” are used interchangeably herein to refer to a polymer of amino acid residues. The terms apply to naturally occurring amino acid polymers as well as amino acid polymers in which one or more amino acid residues is a non-naturally occurring amino acid, e.g., an amino acid analog. The terms encompass amino acid chains of any length, including full length proteins, wherein the amino acid residues are linked by covalent peptide bonds. [0067] As used herein, “regulatory sequence” or “regulatory region” or “expression control sequence” of a nucleic acid molecule means a cis- acting nucleotide sequence that influences expression, positively or negatively, of an operably linked gene. Regulatory regions include sequences of nucleotides that confer inducible (i.e., require a substance or stimulus for increased transcription) expression of a gene. When an inducer is present or at increased concentration, gene expression can be increased. Regulatory regions also include sequences that confer repression of gene expression (i.e., a substance or stimulus decreases transcription). When a repressor is present or at increased concentration, gene expression can be decreased. Regulatory regions are known to influence, modulate or control many in vivo biological activities including cell proliferation, cell growth and death, cell differentiation and immune modulation. Regulatory regions typically bind to one or more trans-acting proteins, which results in either increased or decreased transcription of the gene. [0068] Particular examples of gene regulatory regions are promoters and enhancers. Promoters are sequences located around the transcription or translation start site, typically positioned 5' of the translation start site. Promoters usually are located within 1 Kb of the translation start site, but can be located further away, for example, 2 Kb, 3 Kb, 4 Kb, 5 Kb or more, up to and including 10 Kb. Polymerase II and III are examples of promoters. A polymerase II or “pol II” promoter catalyzes the transcription of DNA to synthesize precursors of mRNA, and most shRNA and microRNA. Examples of pol II promoters are known in the art and include without limitation, the phosphoglycerate kinase (“PGK”) promoter; EF1-alpha; CMV (minimal cytomegalovirus promoter); and LTRs from retroviral -20- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 and lentiviral vectors. In some embodiments, the promoter is a constitutive promoter. As used herein, the term “constitutive promoter” refers to a promoter that allows for continual transcription of the coding sequence or gene under its control in all or most tissues of a subject at all or most developing stages. Non-limiting examples of the constitutive promoters include a CMV promoter, a simian virus 40 (SV40) promoter, a polyubiquitin C (UBC) promoter, an EF1-alpha promoter, a PGK promoter and a CAG promoter. In some embodiments, the promoter is a conditional promoter, which allows for continual transcription of the coding sequence or gene under certain conditions. In further embodiments, the conditional promoter is an immune cell specific promoter, which allows for continual transcription of the coding sequence or gene in an immune cell. Non-limiting examples of the immune cell specific promoters include a promoter of a B29 gene promoter, a CD14 gene promoter, a CD43 gene promoter, a CD45 gene promoter, a CD68 gene promoter, a IFN-β gene promoter, a WASP gene promoter, a T-cell receptor β-chain gene promoter, a V9 γ (TRGV9) gene promoter, a V2 δ (TRDV2) gene promoter, and the like. [0069] Enhancers are known to influence gene expression when positioned 5' or 3' of the gene, or when positioned in or a part of an exon or an intron. Enhancers also can function at a significant distance from the gene, for example, at a distance from about 3 Kb, 5 Kb, 7 Kb, 10 Kb, 15 Kb or more. [0070] Regulatory regions also include, but are not limited to, in addition to promoter regions, sequences that facilitate translation, splicing signals for introns, maintenance of the correct reading frame of the gene to permit in-frame translation of mRNA and, stop codons, leader sequences and fusion partner sequences, internal ribosome binding site (IRES) elements for the creation of multigene, or polycistronic, messages, polyadenylation signals to provide proper polyadenylation of the transcript of a gene of interest and stop codons, and can be optionally included in an expression vector. [0071] As used herein, the term “separate” therapeutic use refers to an administration of at least two active ingredients at the same time or at substantially the same time by different routes. [0072] As used herein, the term “sequential” therapeutic use refers to administration of at least two active ingredients at different times, the administration route being identical or -21- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 different. More particularly, sequential use refers to the whole administration of one of the active ingredients before administration of the other or others commences. It is thus possible to administer one of the active ingredients over several minutes, hours, or days before administering the other active ingredient or ingredients. There is no simultaneous treatment in this case. [0073] As used herein, the term “simultaneous” therapeutic use refers to the administration of at least two active ingredients by the same route and at the same time or at substantially the same time. [0074] As used herein, the terms “subject,” “individual,” or “patient” are used interchangeably and refer to an individual organism, a vertebrate, or a mammal and may include humans, non-human primates, rodents, and the like (e.g., which is to be the recipient of a particular treatment, or from whom cells are harvested). In certain embodiments, the individual, patient or subject is a human. [0075] As used herein, "synthetic," with reference to, for example, a synthetic nucleic acid molecule or a synthetic gene or a synthetic peptide refers to a nucleic acid molecule or polypeptide molecule that is produced by recombinant methods and/or by chemical synthesis methods. As used herein, production by recombinant means by using recombinant DNA methods means the use of the well-known methods of molecular biology for expressing proteins encoded by cloned DNA. [0076] “Treating” or “treatment” as used herein covers the treatment of a disease or disorder described herein, in a subject, such as a human, and includes: (i) inhibiting a disease or disorder, i.e., arresting its development; (ii) relieving a disease or disorder, i.e., causing regression of the disorder; (iii) slowing progression of the disorder; and/or (iv) inhibiting, relieving, or slowing progression of one or more symptoms of the disease or disorder. In some embodiments, treatment means that the symptoms associated with the disease are, e.g., alleviated, reduced, cured, or placed in a state of remission. [0077] It is also to be appreciated that the various modes of treatment of disorders as described herein are intended to mean “substantial,” which includes total but also less than total treatment, and wherein some biologically or medically relevant result is achieved. The -22- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 treatment may be a continuous prolonged treatment for a chronic disease or a single, or few time administrations for the treatment of an acute condition. [0078] The compositions used in accordance with the disclosure can be packaged in dosage unit form for ease of administration and uniformity of dosage. The term "unit dose" or "dosage" refers to physically discrete units suitable for use in a subject, each unit containing a predetermined quantity of the composition calculated to produce the desired responses in association with its administration, i.e., the appropriate route and regimen. The quantity to be administered, both according to number of treatments and unit dose, depends on the result and/or protection desired. Precise amounts of the composition also depend on the judgment of the practitioner and are peculiar to each individual. Factors affecting dose include physical and clinical state of the subject, route of administration, intended goal of treatment (alleviation of symptoms versus cure), and potency, stability, and toxicity of the particular composition. Upon formulation, solutions are administered in a manner compatible with the dosage formulation and in such amount as is therapeutically effective. The formulations are easily administered in a variety of dosage forms, such as the type of injectable solutions described herein. CIR Fusion Proteins of the Present Technology [0079] In one aspect, the present disclosure provides a chimeric innate receptor (CIR) fusion protein that comprises (i) an extracellular antigen binding domain that specifically binds to an AML antigen and (ii) a mammalian cytokine receptor polypeptide comprising a transmembrane domain and an intracellular domain, wherein the mammalian cytokine receptor polypeptide comprises a mammalian CD122 polypeptide, a mammalian CD132 polypeptide, or a mammalian IL-21R polypeptide and wherein the extracellular antigen binding domain is operably linked to the mammalian cytokine receptor polypeptide. In some embodiments, the extracellular antigen binding domain is located at the N-terminus of the cytokine receptor polypeptide. The extracellular antigen binding domain and the mammalian cytokine receptor polypeptide may be directly linked, or linked via a linker. In certain embodiments, the extracellular antigen-binding domain of the expressed CIR fusion protein binds to an AML antigen that is expressed by a tumor cell (e.g., at the surface of a tumor cell). -23- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [0080] Additionally or alternatively, in some embodiments, the CIR fusion protein of the present technology further comprises a fluorescent marker. Non-limiting examples of fluorescent markers include green fluorescent protein (GFP), blue fluorescent protein (e.g., EBFP, EBFP2, Azurite, and mKalamal), cyan fluorescent protein (e.g., ECFP, Cerulean, and CyPet), and yellow fluorescent protein (e.g., YFP, Citrine, Venus, and YPet). [0081] Extracellular Antigen-Binding Domain of a CIR. In certain embodiments, the extracellular antigen-binding domain of a CIR fusion protein specifically binds to an AML antigen selected from among CD33, FLT3, CD123, and CD371. In certain embodiments, the extracellular antigen-binding domain is derived from a monoclonal antibody (mAb) that binds to an AML antigen. In some embodiments, the extracellular antigen-binding domain comprises, or consists essentially of, or yet further consists of an scFv. In some embodiments, the extracellular antigen-binding domain comprises, or consists essentially of, or yet further consists of a Fab, which is optionally crosslinked. In some embodiments, the extracellular binding domain comprises, or consists essentially of, or yet further consists of a F(ab)2. In certain embodiments, the extracellular antigen-binding domain comprises, or consists essentially of, or yet further consists of a human scFv that binds specifically to an AML antigen. In certain embodiments, the scFv is identified by screening scFv phage library with an AML antigen-Fc fusion protein. In some embodiments, any of the foregoing molecules are included in a CIR fusion protein with a heterologous sequence to form the extracellular antigen-binding domain. [0082] In certain embodiments, the extracellular antigen-binding domain of a presently disclosed CIR fusion protein has a high binding specificity and high binding affinity to an AML antigen. For example, in some embodiments, the extracellular antigen-binding domain of the CIR fusion protein (embodied, for example, in a human scFv or an analog thereof) binds to a particular AML antigen with a dissociation constant (K_d) of about 1 × 10^-5 M or less. In certain embodiments, the Kd is about 5 × 10^-6 M or less, about 1 × 10^-6 M or less, about 5 × 10^-7 M or less, about 1 × 10^-7 M or less, about 5 × 10^-8 M or less, about 1 × 10^-8 M or less, about 5 × 10^-9 or less, about 4 × 10^-9 or less, about 3 × 10^-9 or less, about 2 × 10^-9 or less, or about 1 × 10^-9 M or less. In certain non-limiting embodiments, the Kd is from about 3 × 10^-9 M or less. In certain non-limiting embodiments, the Kd is from about 3 -24- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 × 10^-9 to about 2 × 10^-7. [0083] Binding of the extracellular antigen-binding domain (embodiment, for example, in an scFv or an analog thereof) of a presently disclosed AML-antigen-specific CIR fusion protein can be confirmed by, for example, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), FACS analysis, bioassay (e.g., growth inhibition), or Western Blot assay. Each of these assays generally detect the presence of protein-antibody complexes of particular interest by employing a labeled reagent (e.g., an antibody, or an scFv) specific for the complex of interest. For example, the scFv can be radioactively labeled and used in a radioimmunoassay (RIA) (see, for example, Weintraub, B., Principles of Radioimmunoassays, Seventh Training Course on Radioligand Assay Techniques, The Endocrine Society, March, 1986, which is incorporated by reference herein). The radioactive isotope can be detected by such means as the use of a γ counter or a scintillation counter or by autoradiography. [0084] In certain embodiments, the extracellular antigen-binding domain (e.g., human scFv) comprises a heavy chain variable (VH) region and a light chain variable (VL) region, optionally linked with a linker sequence, for example a linker peptide (e.g., SEQ ID NO: 1 or SEQ ID NO: 4), between the heavy chain variable (V_H) region and the light chain variable (V_L) region. In certain embodiments, the extracellular antigen-binding domain is a human scFv-Fc fusion protein or full length human IgG with VH and VL regions. [0085] In certain non-limiting embodiments, the extracellular antigen-binding domain of the presently disclosed CIR fusion protein comprises a linker connecting the heavy chain variable (VH) region and light chain variable (VL) region of the extracellular antigen- binding domain. As used herein, the term “linker” refers to a functional group (e.g., chemical or polypeptide) that covalently attaches two or more polypeptides or nucleic acids so that they are connected to one another. As used herein, a “peptide linker” refers to one or more amino acids used to couple two proteins together (e.g., to couple VH and VL domains). In certain embodiments, the linker comprises amino acids having the sequence set forth in SEQ ID NO: 1 or SEQ ID NO: 4. In certain embodiments, the nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 1 is set forth in SEQ ID NO: 2. [0086] Exemplary amino acid sequences of the heavy chain variable (V_H) domain and -25- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 light chain variable (VL) domain present in the extracellular antigen-binding domain of a CIR fusion protein described herein include, but are not limited to: [0087] CD33 V_H domain (SEQ ID NO: 5): EVQLLESGGGLVQPGGSLRLSCAASGFTFSTYAMSWVRQAPGKGLEWVSAISGRGGSTY YTDSVKGRFTISRDNSKNTVSLQMNSLRAEDTAVYYCAGRGDYYYYYGMDVWGQGTTVT VSA [0088] CD33 V_L domain (SEQ ID NO: 6): DIVMTQSPLSSPVTLGQPASFSCRSSQSLVYSDGNTYLSWLQQRPGQPPRLLIYKISNRFSG VPDRFSGSGAGTDFTLKISRVEAEDVGVYYCMQSTQFPHTFGQGTKLEIK [0089] CD33 V_H domain (SEQ ID NO: 7): QVQLVQSGAEVKKPGSSVKVSCKASGYTFTDYNMHWVRQAPGQGLEWIGYIYPYNGGT GYNQKFKSKATITADESTNTAYMELSSLRSEDTAVYYCARGRPAMDYWGQGTLVTVSS [0090] CD33 V_L domain (SEQ ID NO: 8): DIQMTQSPSSLSASVGDRVTITCRASESVDNYGISFMNWFQQKPGGAPKLLIYAASNQGS GVPSRFSGSGSGTDFTLTISSLQPDDFATYYCQQSKEVPWTFGQGTKVEIK [0091] Examples of other heavy chain variable (V_H) region and light chain variable (V_L) region amino acid sequences that bind other AML antigens are known in the art, and are described in US Patent Nos.9,023,996 and 8,071,099 (anti-FLT3 antibodies), US Patent Nos.10,077,313 and 10,912,842 (anti-CD123 antibodies) and US Patent Publication No. 20220195064 (anti-CD371 antibodies), the contents of which are incorporated by reference in their entireties. [0092] Additionally or alternatively, in some embodiments, the extracellular antigen- binding domain can comprise a leader or a signal peptide sequence that directs the nascent protein into the endoplasmic reticulum. The signal peptide or leader can be essential if the CIR fusion protein is to be glycosylated and anchored in the cell membrane. The signal sequence or leader sequence can be a peptide sequence (about 5, about 10, about 15, about 20, about 25, or about 30 amino acids long) present at the N-terminus of the newly synthesized proteins that direct their entry to the secretory pathway. [0093] In certain embodiments, the signal peptide is covalently joined to the N-terminus -26- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 of the extracellular antigen-binding domain. In certain embodiments, the signal peptide comprises a human CD8 signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 9 as provided below: MALPVTALLLPLALLLHA (SEQ ID NO: 9). [0094] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 9 is set forth in SEQ ID NO: 10, which is provided below: ATGGCTCTCCCAGTGACTGCCCTACTGCTTCCCCTAGCGCTTCTCCTGCATGCA (SEQ ID NO: 10). [0095] In certain embodiments, the signal peptide comprises a human CD8 signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 11 as provided below: MALPVTALLLPLALLLHAARP (SEQ ID NO: 11). [0096] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 11 is set forth in SEQ ID NO: 12, which is provided below: ATGGCCCTGCCAGTAACGGCTCTGCTGCTGCCACTTGCTCTGCTCCTCCATGCAG CCAGGCCT (SEQ ID NO: 12). [0097] In certain embodiments, the signal peptide comprises a mouse CD8 signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 13 as provided below: MASPLTRFLSLNLLLLGESII (SEQ ID NO: 13). [0098] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 13 is set forth in SEQ ID NO: 14, which is provided below: ATGGCCAGCCCCCTGACCAGGTTCCTGAGCCTGAACCTGCTGCTGCTGGGCGAG AGCATCATC (SEQ ID NO: 14). [0099] In certain embodiments, the signal peptide comprises a mouse CD8 signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 15 as provided below: MASPLTRFLSLNLLLLGE (SEQ ID NO: 15). [00100] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 15 is set forth in SEQ ID NO: 16, which is provided below: ATGGCCAGCCCCCTGACCAGGTTCCTGAGCCTGAACCTGCTGCTGCTGGGCGAG (SEQ ID NO: 16). [00101] In certain embodiments, the signal peptide comprises an IL2RB signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 17 as provided below: -27- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00102] MAAPALSWRLPLLILLLPLATSWASA (SEQ ID NO: 17). [00103] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 17 is set forth in SEQ ID NO: 18, which is provided below: [00104] ATGGCGGCCCCTGCTCTGTCCTGGCGTCTGCCCCTCCTCATCCTCCTC CTGCCCCTGGCTACCTCTTGGGCATCTGCA (SEQ ID NO: 18). [00105] In certain embodiments, the signal peptide comprises an Ig signal polypeptide comprising amino acids having the sequence set forth in SEQ ID NO: 19 as provided below: [00106] MDWIWRILFLVGAATGAHS (SEQ ID NO: 19) [00107] The nucleotide sequence encoding the amino acid sequence of SEQ ID NO: 19 is set forth in SEQ ID NO: 20, which is provided below: [00108] ATGGATTGGATTTGGCGCATTCTGTTTCTGGTGGGAGCCGCAACCGG AGCACATAGT (SEQ ID NO: 20) [00109] Exemplary amino acid sequences of the mammalian cytokine receptor polypeptides of the CIR fusion protein described herein include, but are not limited to: [00110] CD122 transmembrane domain-intracellular domain (TM-ICD) [00111] IPWLGHLLVGLSGAFGFIILVYLLINCRNTGPWLKKVLKCNTPDPSKFFSQ LSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLERDKVTQLLLQQDKVPEPASL SSNHSLTSCFTNQGYFFFHLPDALEIEACQVYFTYDPYSEEDPDEGVAGAPTGSSPQ PLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSPPSTAPGGSGAGEERMPPSLQERVP RDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGEEVPDAGPREGVSFPWSRPPGQG EFRALNARLPLNTDAYLSLQELQGQDPTHLV (SEQ ID NO: 21) [00112] CD132 transmembrane domain-intracellular domain (TM-ICD) [00113] VVISVGSMGLIISLLCVYFWLERTMPRIPTLKNLEDLVTEYHGNFSAWSG VSKGLAESLQPDYSERLCLVSEIPPKGGALGEGPGASPCNQHSPYWAPPCYTLKPET (SEQ ID NO: 22) [00114] IL-21 transmembrane domain-intracellular domain (TM-ICD) [00115] GWNPHLLLLLLLVIVFIPAFWSLKTHPLWRLWKKIWAVPSPERFFMPLY KGCSGDFKKWVGAPFTGSSLELGPWSPEVPSTLEVYSCHPPRSPAKRLQLTELQEPA ELVESDGVPKPSFWPTAQNSGGSAYSEERDRPYGLVSIDTVTVLDAEGPCTWPCSC EDDGYPALDLDAGLEPSPGLEDPLLDAGTTVLSCGCVSAGSPGLGGPLGSLLDRLK -28- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 PPLADGEDWAGGLPWGGRSPGGVSESEAGSPLAGLDMDTFDSGFVGSDCSSPVEC DFTSPGDEGPPRSYLRQWVVIPPPLSSPGPQAS (SEQ ID NO: 23) [00116] Additionally or alternatively, in some embodiments, the CIR fusion protein further comprises a hinge domain between the extracellular antigen binding domain and the cytokine receptor polypeptide. In certain embodiments, the hinge domain comprises a CD8 hinge domain, a CD28 hinge domain, CD122 hinge domain or a CD132 hinge domain. [00117] Exemplary amino acid sequences of the hinge domain of the CIR fusion protein described herein include, but are not limited to: [00118] CD8 hinge domain [00119] TTTPAPRPPTPAPTIASQPLSLRPEACRPAAGGAVHTRGLDFACD (SEQ ID NO: 24) [00120] CD28 hinge domain [00121] IEVMYPPPYLDNEKSNGTIIHVKGKHLCPSPLFPGPSKP (SEQ ID NO: 25) [00122] CD122 hinge domain [00123] YEFQVRVKPLQGEFTTWSPWSQPLAFRTKPAALGKDT (SEQ ID NO: 26) [00124] CD132 hinge domain [00125] YTFRVRSRFNPLCGSAQHWSEWSHPIHWGSNTSKENPFLFALEA (SEQ ID NO: 27) [00126] Additionally or alternatively, in some embodiments, the CIR fusion proteins of the present technology comprise an amino acid sequence selected from any one of: [00127] CIR with CD8 hinge (CD8 signal peptide-CD33scFv-CD8hinge-CD122TM- CD122ICD-P2A-mcherry) (SEQ ID NO: 28) [00128] MALPVTALLLPLALLLHAEVQLLESGGGLVQPGGSLRLSCAASGFTFSTYA MSWVRQAPGKGLEWVSAISGRGGSTYYTDSVKGRFTISRDNSKNTVSLQMNSLRAEDTAV YYCAGRGDYYYYYGMDVWGQGTTVTVSAGGGGSGGGGSGGGGSDIVMTQSPLSSPVTL GQPASFSCRSSQSLVYSDGNTYLSWLQQRPGQPPRLLIYKISNRFSGVPDRFSGSGAGTD FTLKISRVEAEDVGVYYCMQSTQFPHTFGQGTKLEIKTTTPAPRPPTPAPTIASQPLSL RPEACRPAAGGAVHTRGLDFACDIPWLGHLLVGLSGAFGFIILVYLLINCRNTGP -29- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 WLKKVLKCNTPDPSKFFSQLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLER DKVTQLLLQQDKVPEPASLSSNHSLTSCFTNQGYFFFHLPDALEIEACQVYFTYDPY SEEDPDEGVAGAPTGSSPQPLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSPPSTAP GGSGAGEERMPPSLQERVPRDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGEEVP DAGPREGVSFPWSRPPGQGEFRALNARLPLNTDAYLSLQELQGQDPTHLVGSGATN FSLLKQAGDVEENPGPMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEG RPYEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGF KWERVMNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEA SSERMYPEDGALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLD ITSHNEDYTIVEQYERAEGRHSTGGMDELYK [00129] CIR with CD28 hinge (CD8 signal peptide-CD33scFv-CD28hinge-CD122TM- CD122ICD-P2A-mcherry) (SEQ ID NO: 29) [00130] MALPVTALLLPLALLLHAEVQLLESGGGLVQPGGSLRLSCAASGFTFSTYA MSWVRQAPGKGLEWVSAISGRGGSTYYTDSVKGRFTISRDNSKNTVSLQMNSLRAEDTAV YYCAGRGDYYYYYGMDVWGQGTTVTVSAGGGGSGGGGSGGGGSDIVMTQSPLSSPVTL GQPASFSCRSSQSLVYSDGNTYLSWLQQRPGQPPRLLIYKISNRFSGVPDRFSGSGAGTD FTLKISRVEAEDVGVYYCMQSTQFPHTFGQGTKLEIKIEVMYPPPYLDNEKSNGTIIH VKGKHLCPSPLFPGPSKPIPWLGHLLVGLSGAFGFIILVYLLINCRNTGPWLKKVL KCNTPDPSKFFSQLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLERDKVTQL LLQQDKVPEPASLSSNHSLTSCFTNQGYFFFHLPDALEIEACQVYFTYDPYSEEDPD EGVAGAPTGSSPQPLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSPPSTAPGGSGAG EERMPPSLQERVPRDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGEEVPDAGPRE GVSFPWSRPPGQGEFRALNARLPLNTDAYLSLQELQGQDPTHLVGSGATNFSLLKQ AGDVEENPGPMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERV MNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERM YPEDGALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHN EDYTIVEQYERAEGRHSTGGMDELYK [00131] CIR with CD122 hinge (CD8 signal peptide-CD33scFv-CD122hinge- CD122TM-CD122ICD-P2A-mcherry) (SEQ ID NO: 30) -30- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00132] MALPVTALLLPLALLLHAEVQLLESGGGLVQPGGSLRLSCAASGFTFSTYA MSWVRQAPGKGLEWVSAISGRGGSTYYTDSVKGRFTISRDNSKNTVSLQMNSLRAEDTAV YYCAGRGDYYYYYGMDVWGQGTTVTVSAGGGGSGGGGSGGGGSDIVMTQSPLSSPVTL GQPASFSCRSSQSLVYSDGNTYLSWLQQRPGQPPRLLIYKISNRFSGVPDRFSGSGAGTD FTLKISRVEAEDVGVYYCMQSTQFPHTFGQGTKLEIKYEFQVRVKPLQGEFTTWSP WSQPLAFRTKPAALGKDTIPWLGHLLVGLSGAFGFIILVYLLINCRNTGPWLKKV LKCNTPDPSKFFSQLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLERDKVTQ LLLQQDKVPEPASLSSNHSLTSCFTNQGYFFFHLPDALEIEACQVYFTYDPYSEEDP DEGVAGAPTGSSPQPLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSPPSTAPGGSGA GEERMPPSLQERVPRDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGEEVPDAGPR EGVSFPWSRPPGQGEFRALNARLPLNTDAYLSLQELQGQDPTHLVGSGATNFSLLK QAGDVEENPGPMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYE GTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWER VMNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSER MYPEDGALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSH NEDYTIVEQYERAEGRHSTGGMDELYK [00133] aCD33D-CD28h-CD132TMICD-P2A-BFP (SEQ ID NO: 31) [00134] MALPVTALLL PLALLLHAQV QLVQSGAEVK KPGSSVKVSC KASGYTFTDY NMHWVRQAPG QGLEWIGYIY PYNGGTGYNQ KFKSKATITA DESTNTAYME LSSLRSEDTA VYYCARGRPA MDYWGQGTLV TVSSGGGGSG GGGSGGGGSD IQMTQSPSSL SASVGDRVTI TCRASESVDN YGISFMNWFQ QKPGGAPKLL IYAASNQGSG VPSRFSGSGS GTDFTLTISS LQPDDFATYY CQQSKEVPWT FGQGTKVEIK IEVMYPPPYL DNEKSNGTII HVKGKHLCPS PLFPGPSKPV VISVGSMGLI ISLLCVYFWL ERTMPRIPTL KNLEDLVTEY HGNFSAWSGV SKGLAESLQP DYSERLCLVS EIPPKGGALG EGPGASPCNQ HSPYWAPPCY TLKPETGSGA TNFSLLKQAG DVEENPGPMV SKGEELFTGV VPILVELDGD VNGHKFSVSG EGEGDATYGK LTLKFICTTG KLPVPWPTLV TTLSHGVQCF SRYPDHMKQH DFFKSAMPEG YVQERTIFFK DDGNYKTRAE VKFEGDTLVN RIELKGIDFK EDGNILGHKL EYNFNSHNVY IMADKQKNGI KANFKIRHNI EDGSVQLADH YQQNTPIGDG PVLLPDSHYL STQSALSKDP NEKRDHMVLL EFVTAAGITL GMDELYK -31- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00135] aCD33D-CD8h-CD132TMICD-P2A-BFP (SEQ ID NO: 32) [00136] MALPVTALLL PLALLLHAQV QLVQSGAEVK KPGSSVKVSC KASGYTFTDY NMHWVRQAPG QGLEWIGYIY PYNGGTGYNQ KFKSKATITA DESTNTAYME LSSLRSEDTA VYYCARGRPA MDYWGQGTLV TVSSGGGGSG GGGSGGGGSD IQMTQSPSSL SASVGDRVTI TCRASESVDN YGISFMNWFQ QKPGGAPKLL IYAASNQGSG VPSRFSGSGS GTDFTLTISS LQPDDFATYY CQQSKEVPWT FGQGTKVEIK TTTPAPRPPT PAPTIASQPL SLRPEACRPA AGGAVHTRGL DFACDVVISV GSMGLIISLL CVYFWLERTM PRIPTLKNLE DLVTEYHGNF SAWSGVSKGL AESLQPDYSE RLCLVSEIPP KGGALGEGPG ASPCNQHSPY WAPPCYTLKP ETGSGATNFS LLKQAGDVEE NPGPMVSKGE ELFTGVVPIL VELDGDVNGH KFSVSGEGEG DATYGKLTLK FICTTGKLPV PWPTLVTTLS HGVQCFSRYP DHMKQHDFFK SAMPEGYVQE RTIFFKDDGN YKTRAEVKFE GDTLVNRIEL KGIDFKEDGN ILGHKLEYNF NSHNVYIMAD KQKNGIKANF KIRHNIEDGS VQLADHYQQN TPIGDGPVLL PDSHYLSTQS ALSKDPNEKR DHMVLLEFVT AAGITLGMDE LYK [00137] aCD33D-CD132h-CD132TMICD-P2A-BFP (SEQ ID NO: 33) [00138] MALPVTALLL PLALLLHAQV QLVQSGAEVK KPGSSVKVSC KASGYTFTDY NMHWVRQAPG QGLEWIGYIY PYNGGTGYNQ KFKSKATITA DESTNTAYME LSSLRSEDTA VYYCARGRPA MDYWGQGTLV TVSSGGGGSG GGGSGGGGSD IQMTQSPSSL SASVGDRVTI TCRASESVDN YGISFMNWFQ QKPGGAPKLL IYAASNQGSG VPSRFSGSGS GTDFTLTISS LQPDDFATYY CQQSKEVPWT FGQGTKVEIK YTFRVRSRFN PLCGSAQHWS EWSHPIHWGS NTSKENPFLF ALEAVVISVG SMGLIISLLC VYFWLERTMP RIPTLKNLED LVTEYHGNFS AWSGVSKGLA ESLQPDYSER LCLVSEIPPK GGALGEGPGA SPCNQHSPYW APPCYTLKPE TGSGATNFSL LKQAGDVEEN PGPMVSKGEE LFTGVVPILV ELDGDVNGHK FSVSGEGEGD ATYGKLTLKF ICTTGKLPVP WPTLVTTLSH GVQCFSRYPD HMKQHDFFKS AMPEGYVQER TIFFKDDGNY KTRAEVKFEG DTLVNRIELK GIDFKEDGNI LGHKLEYNFN SHNVYIMADK QKNGIKANFK IRHNIEDGSV QLADHYQQNT PIGDGPVLLP DSHYLSTQSA LSKDPNEKRD HMVLLEFVTA AGITLGMDEL YK -32- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00139] CD33scFv-CD8hinge-CD122TM-CD122ICD paired with a CD33scFv- CD8hinge-CD132TM-CD132ICD construct (SEQ ID NO: 34) MALPVTALLLPLALLLHAQVQLVQSGAEVKKPGSSVKVSCKASGYTFTDYNMHWVRQ APGQGLEWIGYIYPYNGGTGYNQKFKSKATITADESTNTAYMELSSLRSEDTAVYYCARG RPAMDYWGQGTLVTVSSGGGGSGGGGSGGGGSDIQMTQSPSSLSASVGDRVTITCRAS ESVDNYGISFMNWFQQKPGGAPKLLIYAASNQGSGVPSRFSGSGSGTDFTLTISSLQPDD FATYYCQQSKEVPWTFGQGTKVEIKTTTPAPRPPTPAPTIASQPLSLRPEACRPAAG GAVHTRGLDFACDIPWLGHLLVGLSGAFGFIILVYLLINCRNTGPWLKKVLKCNTP DPSKFFSQLSSEHGGDVQKWLSSPFPSSSFSPGGLAPEISPLEVLERDKVTQLLLQQD KVPEPASLSSNHSLTSCFTNQGYFFFHLPDALEIEACQVYFTYDPYSEEDPDEGVAG APTGSSPQPLQPLSGEDDAYCTFPSRDDLLLFSPSLLGGPSPPSTAPGGSGAGEERMP PSLQERVPRDWDPQPLGPPTPGVPDLVDFQPPPELVLREAGEEVPDAGPREGVSFP WSRPPGQGEFRALNARLPLNTDAYLSLQELQGQDPTHLVGSGATNFSLLKQAGDV EENPGPMALPVTALLLPLALLLHAEVQLLESGGGLVQPGGSLRLSCAASGFTFSTYAM SWVRQAPGKGLEWVSAISGRGGSTYYTDSVKGRFTISRDNSKNTVSLQMNSLRAEDTAVY YCAGRGDYYYYYGMDVWGQGTTVTVSAGGGGSGGGGSGGGGSDIVMTQSPLSSPVTL GQPASFSCRSSQSLVYSDGNTYLSWLQQRPGQPPRLLIYKISNRFSGVPDRFSGSGAGTD FTLKISRVEAEDVGVYYCMQSTQFPHTFGQGTKLEIKTTTPAPRPPTPAPTIASQPLSL RPEACRPAAGGAVHTRGLDFACDVVISVGSMGLIISLLCVYFWLERTMPRIPTLK NLEDLVTEYHGNFSAWSGVSKGLAESLQPDYSERLCLVSEIPPKGGALGEGPGASP CNQHSPYWAPPCYTLKPETGSGATNFSLLKQAGDVEENPGPMVSKGEEDNMAIIKE FMRFKVHMEGSVNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFAWDILSPQFM YGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVVTVTQDSSLQDGEFIYK VKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKGEIKQRLKLKDGGHYDA EVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAEGRHSTGGMDELY K [00140] aCD33D8hCD122-2AaCD33P8hCD132-2AmCh (SEQ ID NO: 35) [00141] MALPVTALLL PLALLLHAQV QLVQSGAEVK KPGSSVKVSC KASGYTFTDY NMHWVRQAPG QGLEWIGYIY PYNGGTGYNQ KFKSKATITA DESTNTAYME LSSLRSEDTA VYYCARGRPA MDYWGQGTLV TVSSGGGGSG GGGSGGGGSD IQMTQSPSSL SASVGDRVTI TCRASESVDN YGISFMNWFQ QKPGGAPKLL IYAASNQGSG VPSRFSGSGS GTDFTLTISS LQPDDFATYY CQQSKEVPWT FGQGTKVEIK TTTPAPRPPT PAPTIASQPL SLRPEACRPA AGGAVHTRGL DFACDIPWLG HLLVGLSGAF GFIILVYLLI NCRNTGPWLK KVLKCNTPDP SKFFSQLSSE HGGDVQKWLS SPFPSSSFSP GGLAPEISPL EVLERDKVTQ LLLQQDKVPE PASLSSNHSL TSCFTNQGYF FFHLPDALEI EACQVYFTYD PYSEEDPDEG VAGAPTGSSP QPLQPLSGED DAYCTFPSRD DLLLFSPSLL GGPSPPSTAP GGSGAGEERM -33- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 PPSLQERVPR DWDPQPLGPP TPGVPDLVDF QPPPELVLRE AGEEVPDAGP REGVSFPWSR PPGQGEFRAL NARLPLNTDA YLSLQELQGQ DPTHLVGSGA TNFSLLKQAG DVEENPGPMA LPVTALLLPL ALLLHAEVQL LESGGGLVQP GGSLRLSCAA SGFTFSTYAM SWVRQAPGKG LEWVSAISGR GGSTYYTDSV KGRFTISRDN SKNTVSLQMN SLRAEDTAVY YCAGRGDYYY YYGMDVWGQG TTVTVSAGGG GSGGGGSGGG GSDIVMTQSP LSSPVTLGQP ASFSCRSSQS LVYSDGNTYL SWLQQRPGQP PRLLIYKISN RFSGVPDRFS GSGAGTDFTL KISRVEAEDV GVYYCMQSTQ FPHTFGQGTK LEIKTTTPAP RPPTPAPTIA SQPLSLRPEA CRPAAGGAVH TRGLDFACDV VISVGSMGLI ISLLCVYFWL ERTMPRIPTL KNLEDLVTEY HGNFSAWSGV SKGLAESLQP DYSERLCLVS EIPPKGGALG EGPGASPCNQ HSPYWAPPCY TLKPETGSGA TNFSLLKQAG DVEENPGPMV SKGEEDNMAI IKEFMRFKVH MEGSVNGHEF EIEGEGEGRP YEGTQTAKLK VTKGGPLPFA WDILSPQFMY GSKAYVKHPA DIPDYLKLSF PEGFKWERVM NFEDGGVVTV TQDSSLQDGE FIYKVKLRGT NFPSDGPVMQ KKTMGWEASS ERMYPEDGAL KGEIKQRLKL KDGGHYDAEV KTTYKAKKPV QLPGAYNVNI KLDITSHNED YTIVEQYERA EGRHSTGGMD ELYK [00142] CD33 scFv sequences are italicized; hinge domains are bold; signal peptides are double underlined; CD122 TM-CD122 ICD and CD132 TM-CD132 ICD are underlined or dash underlined; Highlighted in gray is the fluorescent reporter Polynucleotides, Polypeptides and Analogs [00143] Also included in the presently disclosed subject matter are polynucleotides encoding any CIR fusion protein disclosed herein and their corresponding polypeptides or fragments that may be modified in ways that enhance their anti-tumor activity when expressed in an engineered immune cells (e.g., NK cells, NK-like CD8 T cells). The presently disclosed subject matter provides methods for optimizing an amino acid sequence or a nucleic acid sequence by producing an alteration in the sequence. Such alterations can comprise certain mutations, deletions, insertions, or post-translational modifications. The presently disclosed subject matter further comprises analogs of any naturally-occurring polypeptide of the presently disclosed subject matter. Analogs can differ from a naturally- occurring polypeptide of the presently disclosed subject matter by amino acid sequence -34- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 differences, by post-translational modifications, or by both. Analogs of the presently disclosed subject matter can generally exhibit at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%), about 98%, about 99% or more identity or homology with all or part of a naturally-occurring amino, acid sequence of the presently disclosed subject matter. The length of sequence comparison is at least about 5, about 10, about 15, about 20, about 25, about 50, about 75, about 100 or more amino acid residues. Again, in an exemplary approach to determining the degree of identity, a BLAST program can be used, with a probability score between e^-3 and e^-100 indicating a closely related sequence. Modifications comprise in vivo and in vitro chemical derivatization of polypeptides, e.g., acetylation, carboxylation, phosphorylation, or glycosylation; such modifications can occur during polypeptide synthesis or processing or following treatment with isolated modifying enzymes. Analogs can also differ from the naturally-occurring polypeptides of the presently disclosed subject matter by alterations in primary sequence. These include genetic variants, both natural and induced (for example, resulting from random mutagenesis by irradiation or exposure to ethanemethyl sulfate or by site-specific mutagenesis as described in Sambrook, Fritsch and Maniatis, Molecular Cloning: A Laboratory Manual (2nd ed.), CSH Press, 1989, or Ausubel et al., supra). Also included are cyclized peptides, molecules, and analogs which contain residues other than L- amino acids, e.g., D-amino acids or non-naturally occurring or synthetic amino acids, e.g., beta (β) or gamma (γ) amino acids. [00144] In addition to full-length polypeptides, the presently disclosed subject matter also provides fragments of any one of the polypeptides or peptide domains of the presently disclosed subject matter. A fragment can be at least about 5, about 10, about 13, or about 15 amino acids. In some embodiments, a fragment is at least about 20 contiguous amino acids, at least about 30 contiguous amino acids, or at least about 50 contiguous amino acids. In some embodiments, a fragment is at least about 60 to about 80, about 100, about 200, about 300 or more contiguous amino acids. Fragments of the presently disclosed subject matter can be generated by methods known to those of ordinary skill in the art or can result from normal protein processing (e.g., removal of amino acids from the nascent polypeptide that are not required for biological activity or removal of amino acids by alternative mRNA splicing or alternative protein processing events). -35- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00145] Non-protein analogs have a chemical structure designed to mimic the functional activity of a protein. Such analogs are administered according to methods of the presently disclosed subject matter. Such analogs can exceed the physiological activity of the original polypeptide. Methods of analog design are well known in the art, and synthesis of analogs can be carried out according to such methods by modifying the chemical structures such that the resultant analogs increase the antineoplastic activity of the original polypeptide when expressed in an engineered immune cells (e.g., NK cells, NK-like CD8 T cells). These chemical modifications include, but are not limited to, substituting alternative R groups and varying the degree of saturation at specific carbon atoms of a reference polypeptide. The protein analogs can be relatively resistant to in vivo degradation, resulting in a more prolonged therapeutic effect upon administration. Assays for measuring functional activity include, but are not limited to, those described in the Examples below. [00146] In accordance with the presently disclosed subject matter, the polynucleotides encoding any CIR fusion protein disclosed herein can be modified by codon optimization. Codon optimization can alter both naturally occurring and recombinant gene sequences to achieve the highest possible levels of productivity in any given expression system. Factors that are involved in different stages of protein expression include codon adaptability, mRNA structure, and various cis-elements in transcription and translation. Any suitable codon optimization methods or technologies that are known to ones skilled in the art can be used to modify the polynucleotides of the presently disclosed subject matter, including, but not limited to, OptimumGene™, Encor optimization, and Blue Heron. [00147] In some embodiments, a nucleic acid as disclosed herein further comprises a regulatory sequence directing the expression of any CIR fusion protein disclosed herein. In other embodiments, the nucleic acid comprises a first regulatory sequence directing the expression of a first CIR fusion protein described herein (e.g., CD33 scFv-CD8 hinge- CD122 TM-CD122 ICD) and a second regulatory sequence directing the expression of a second CIR fusion protein (e.g., CD33 scFv-CD8 hinge-CD132 TM-CD132 ICD) described herein. In other embodiments, the first regulatory sequence is the same as the second regulatory sequence. In some embodiments, the first regulatory sequence is different from the second regulatory sequence. -36- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 Vectors [00148] Many expression vectors are available and known to those of skill in the art and can be used for nonendogenous expression of any CIR fusion protein disclosed herein. The choice of expression vector will be influenced by the choice of host expression system. Such selection is well within the level of skill of the skilled artisan. In general, expression vectors can include transcriptional promoters and optionally enhancers, translational signals, and transcriptional and translational termination signals. Expression vectors that are used for stable transformation typically have a selectable marker which allows selection and maintenance of the transformed cells. In some cases, an origin of replication can be used to amplify the copy number of the vector in the cells. [00149] Vectors also can contain additional nucleotide sequences operably linked to the ligated nucleic acid molecule, such as, for example, an epitope tag such as for localization, e.g., a hexa-his tag or a myc tag, hemagglutinin tag or a tag for purification, for example, a GST fusion, and a sequence for directing protein secretion and/or membrane association. [00150] Heterologous expression of any CIR fusion protein disclosed herein can be controlled by any promoter/enhancer known in the art. Suitable bacterial promoters are well known in the art and described herein below. Other suitable promoters for mammalian cells, yeast cells and insect cells are well known in the art and some are exemplified below. Selection of the promoter used to direct expression of a heterologous nucleic acid depends on the particular application and is within the level of skill of the skilled artisan. Promoters which can be used include but are not limited to eukaryotic expression vectors containing the SV40 early promoter (Bernoist and Chambon, Nature 290:304-310(1981)), the promoter contained in the 3' long terminal repeat of Rous sarcoma virus (Yamamoto et al., Cell 22:787-797(1980)), the herpes thymidine kinase promoter (Wagner et al., Proc. Natl. Acad. Sci. USA 75: 1441-1445 (1981)), the regulatory sequences of the metallothionein gene (Brinster et al., Nature 296:39-42 (1982)); prokaryotic expression vectors such as the β- lactamase promoter (Jay et al., Proc. Natl. Acad. Sci. USA 75:5543 (1981)) or the tac promoter (DeBoer et al., Proc. Natl. Acad. Sci. USA 50:21-25(1983)); see also "Useful Proteins from Recombinant Bacteria": in Scientific American 242:79-94 (1980)); plant expression vectors containing the nopaline synthetase promoter (Herrera- Estrella et al., Nature 505:209-213(1984)) or the cauliflower mosaic virus 35S RNA promoter (Gardner et -37- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 al., Nucleic Acids Res.9:2871(1981)), and the promoter of the photosynthetic enzyme ribulose bisphosphate carboxylase (Herrera-Estrella et al., Nature 510: 115-120(1984)); promoter elements from yeast and other fungi such as the Gal4 promoter, the alcohol dehydrogenase promoter, the phosphoglycerol kinase promoter, the alkaline phosphatase promoter, and the following animal transcriptional control regions that exhibit tissue specificity and have been used in transgenic animals: elastase I gene control region which is active in pancreatic acinar cells (Swift et al., Cell 55:639-646 (1984); Ornitz et al., Cold Spring Harbor Symp. Quant. Biol.50:399-409(1986); MacDonald, Hepatology 7:425-515 (1987)); insulin gene control region which is active in pancreatic beta cells (Hanahan et al., Nature 515: 115-122 (1985)), immunoglobulin gene control region which is active in lymphoid cells (Grosschedl et al., Cell 55:647-658 (1984); Adams et al., Nature 515:533- 538 (1985); Alexander et al., Mol. Cell Biol.7: 1436-1444 (1987)), mouse mammary tumor virus control region which is active in testicular, breast, lymphoid and mast cells (Leder et al., Cell 15:485-495 (1986)), albumin gene control region which is active in liver (Pinckert et al., Genes and Devel.1:268-276 (1987)), alpha-fetoprotein gene control region which is active in liver (Krumlauf et al., Mol. Cell. Biol.5:1639-403 (1985)); Hammer et al., Science 255:53-58 (1987)), alpha-1 antitrypsin gene control region which is active in liver (Kelsey et al., Genes and Devel.7:161-171 (1987)), beta globin gene control region which is active in myeloid cells (Magram et al., Nature 515:338-340 (1985)); Kollias et al., Cell 5:89-94 (1986)), myelin basic protein gene control region which is active in oligodendrocyte cells of the brain (Readhead et al., Cell 15:703-712 (1987)), myosin light chain-2 gene control region which is active in skeletal muscle (Shani, Nature 514:283-286 (1985)), and gonadotrophic releasing hormone gene control region which is active in gonadotrophs of the hypothalamus (Mason et al., Science 254: 1372- 1378 (1986)). [00151] In addition to the promoter, the expression vector typically contains a transcription unit or expression cassette that contains all the additional elements required for the expression of a fusion protein of the present technology in host cells. A typical expression cassette contains a promoter operably linked to the nucleic acid sequence encoding the polypeptide chains of interest and signals required for efficient polyadenylation of the transcript, ribosome binding sites and translation termination. Additional elements of the cassette can include enhancers. In addition, the cassette typically -38- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 contains a transcription termination region downstream of the structural gene to provide for efficient termination. The termination region can be obtained from the same gene as the promoter sequence or can be obtained from different genes. [00152] Some expression systems have markers that provide gene amplification such as thymidine kinase and dihydrofolate reductase. Alternatively, high yield expression systems not involving gene amplification are also suitable, such as using a baculovirus vector in insect cells, with a nucleic acid sequence encoding a germline antibody chain under the direction of the polyhedron promoter or other strong baculovirus promoter. [00153] Any methods known to those of skill in the art for the insertion of DNA fragments into a vector can be used to construct expression vectors containing a nucleic acid encoding any of the polypeptides provided herein. These methods can include in vitro recombinant DNA and synthetic techniques and in vivo recombinants (genetic recombination). The insertion into a cloning vector can, for example, be accomplished by ligating the DNA fragment into a cloning vector which has complementary cohesive termini. If the complementary restriction sites used to fragment the DNA are not present in the cloning vector, the ends of the DNA molecules can be enzymatically modified. Alternatively, any site desired can be produced by ligating nucleotide sequences (linkers) onto the DNA termini; these ligated linkers can contain specific chemically synthesized nucleic acids encoding restriction endonuclease recognition sequences. [00154] In some embodiments, the expression vector is a plasmid, a cosmid, a bacmid, a bacterial artificial chromosome (BAC), a yeast artificial chromosome (YAC), a viral vector, or a retroviral vector. [00155] Exemplary plasmid vectors useful to produce the polypeptides provided herein contain a strong promoter, such as the HCMV immediate early enhancer/promoter or the MHC class I promoter, an intron to enhance processing of the transcript, such as the HCMV immediate early gene intron A, and a polyadenylation (poly A) signal, such as the late SV40 polyA signal. [00156] Genetic modification of engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can be accomplished by transducing a substantially homogeneous cell composition with a recombinant DNA or RNA construct. The vector can be a retroviral vector (e.g., gamma retroviral), which is employed for the introduction of the DNA or RNA construct -39- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 into the host cell genome. For example, a polynucleotide encoding any and all embodiments of the CIR fusion protein disclosed herein can be cloned into a retroviral vector and expression can be driven from its endogenous promoter, from the retroviral long terminal repeat, or from an alternative internal promoter. [00157] Non-viral vectors or RNA may be used as well. Random chromosomal integration, or targeted integration (e.g., using a nuclease, transcription activator-like effector nucleases (TALENs), Zinc-finger nucleases (ZFNs), and/or clustered regularly interspaced short palindromic repeats (CRISPRs), or transgene expression (e.g., using a natural or chemically modified RNA) can be used. [00158] For initial genetic modification of the cells to provide engineered immune cells (e.g., NK cells, NK-like CD8 T cells) expressing any CIR fusion protein disclosed herein, a retroviral vector can be employed for transduction. However, any other suitable viral vector or non-viral delivery system can be used for genetic modification of cells. For subsequent genetic modification of the cells to provide cells comprising an antigen presenting complex comprising at least two co-stimulatory ligands, retroviral gene transfer (transduction) likewise proves effective. Combinations of retroviral vector and an appropriate packaging line are also suitable, where the capsid proteins will be functional for infecting human cells. Various amphotropic virus-producing cell lines are known, including, but not limited to, PA12 (Miller et al. (1985) Mol. Cell. Biol.5:431-437); PA317 (Miller et al. (1986) Mol. Cell. Biol.6:2895-2902); and CRIP (Danos et al. (1988) Proc. Natl. Acad. Sci. USA 85:6460-6464). Non -amphotropic particles are suitable too, e.g., particles pseudotyped with VSVG, RD114 or GALV envelope and any other known in the art. [00159] Possible methods of transduction also include direct co-culture of the cells with producer cells, e.g., by the method of Bregni, et al., Blood 80: 1418-1422(1992), or culturing with viral supernatant alone or concentrated vector stocks with or without appropriate growth factors and polycations, e.g., by the method of Xu, et al., Exp. Hemat. 22:223-230 (1994); and Hughes, et al., J. Clin. Invest.89: 1817 (1992). [00160] Transducing viral vectors can be used to express a co-stimulatory ligand and/or secrete a cytokine (e.g., 4-1BBL and/or IL-12) in an engineered immune cell (e.g., NK cell, NK-like CD8 T cell). In some embodiments, the chosen vector exhibits high efficiency of infection and stable integration and expression (see, e.g., Cayouette et al., Human Gene -40- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 Therapy 8:423-430 (1997); Kido et al., Current Eye Research 15:833-844 (1996); Bloomer et al., Journal of Virology 71 :6641-6649, 1997; Naldini et al., Science 272:263267 (1996); and Miyoshi et al., Proc. Natl. Acad. Sci. U.S.A.94: 10319, (1997)). Other viral vectors that can be used include, for example, adenoviral, lentiviral, and adeno-associated viral vectors, vaccinia virus, a bovine papilloma virus, or a herpes virus, such as Epstein-Barr Virus (also see, for example, the vectors of Miller, Human Gene Therapy 15-14, (1990); Friedman, Science 244: 1275-1281 (1989); Eglitis et al., BioTechniques 6:608-614, (1988); Tolstoshev et al., Current Opinion in Biotechnology 1:55-61(1990); Sharp, The Lancet 337: 1277-1278 (1991); Cornetta et al., Nucleic Acid Research and Molecular Biology 36:311- 322 (1987); Anderson, Science 226:401-409 (1984); Moen, Blood Cells 17:407-416 (1991); Miller et al., Biotechnology 7:980-990 (1989); Le Gal La Salle et al., Science 259:988-990 (1993); and Johnson, Chest 107:77S-83S (1995)). Retroviral vectors are particularly well developed and have been used in clinical settings (Rosenberg et al., N. Engl. J. Med 323:370 (1990); Anderson et al., U.S. Pat. No.5,399,346). [00161] In certain non-limiting embodiments, the vector expressing any CIR fusion protein disclosed herein is a retroviral vector, e.g., an oncoretroviral vector. In some instances, the retroviral vector is a SFG retroviral vector or murine stem cell virus (MSCV) retroviral vector. In certain non-limiting embodiments, the vector expressing a nucleic acid encoding a CIR fusion protein is a lentiviral vector. In certain non-limiting embodiments, the vector expressing a nucleic acid encoding a CIR fusion protein is a transposon vector. [00162] Non-viral approaches can also be employed for the expression of a protein in a cell. For example, a nucleic acid molecule can be introduced into a cell by administering the nucleic acid in the presence of lipofection (Feigner et al., Proc. Nat'l. Acad. Sci. U.S.A. 84:7413, (1987); Ono et al., Neuroscience Letters 17:259 (1990); Brigham et al., Am. J. Med. Sci.298:278, (1989); Staubinger et al., Methods in Enzymology 101 :512 (1983)), asialoorosomucoid-polylysine conjugation (Wu et al., Journal of Biological Chemistry 263 : 14621 (1988); Wu et al., Journal of Biological Chemistry 264: 16985 (1989)), or by micro- injection under surgical conditions (Wolff et al., Science 247: 1465 (1990)). Other non- viral means for gene transfer include transfection in vitro using calcium phosphate, DEAE dextran, electroporation, and protoplast fusion. Liposomes can also be potentially beneficial for delivery of DNA into a cell. Transplantation of normal genes into the -41- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 affected tissues of a subject can also be accomplished by transferring a normal nucleic acid into a cultivatable cell type ex vivo (e.g., an autologous or heterologous primary cell or progeny thereof), after which the cell (or its descendants) are injected into a targeted tissue or are injected systemically. Recombinant receptors can also be derived or obtained using transposases or targeted nucleases (e.g., Zinc finger nucleases, meganucleases, or TALE nucleases). Transient expression may be obtained by RNA electroporation. [00163] cDNA expression for use in polynucleotide therapy methods can be directed from any suitable promoter (e.g., the human cytomegalovirus (CMV), simian virus 40 (SV40), or metallothionein promoters), and regulated by any appropriate mammalian regulatory element or intron (e.g., the elongation factor la enhancer/promoter/intron structure). For example, if desired, enhancers known to preferentially direct gene expression in specific cell types can be used to direct the expression of a nucleic acid. The enhancers used can include, without limitation, those that are characterized as tissue- or cell-specific enhancers. Alternatively, if a genomic clone is used as a therapeutic construct, regulation can be mediated by the cognate regulatory sequences or, if desired, by regulatory sequences derived from a heterologous source, including any of the promoters or regulatory elements described above. [00164] The resulting cells can be grown under conditions similar to those for unmodified cells, whereby the modified cells can be expanded and used for a variety of purposes. [00165] In some embodiments, a vector as disclosed herein further comprises a regulatory sequence directing the expression of any CIR fusion protein disclosed herein. In further embodiments, the vector comprises a single regulatory sequence directing the expression of any CIR fusion protein disclosed herein. [00166] In other embodiments, the vector comprises a first regulatory sequence directing the expression of a first CIR fusion protein described herein (e.g., CD33 scFv-CD8 hinge- CD122 TM-CD122 ICD) and a second regulatory sequence directing the expression of a second CIR fusion protein (e.g., CD33 scFv-CD8 hinge-CD132 TM-CD132 ICD) described herein. In other embodiments, the first regulatory sequence is the same as the second regulatory sequence. In some embodiments, the first regulatory sequence is different from the second regulatory sequence. -42- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 Engineered Immune Cells of the Present Technology [00167] The presently disclosed subject matter provides engineered immune cells (e.g., NK cells, NK-like CD8 T cells) that exhibit heterologous expression of any and all embodiments of the CIR fusion protein described herein. [00168] Additionally or alternatively, in some embodiments, the engineered immune cells may further comprise a CIR fusion protein that comprises (i) an extracellular antigen binding domain that specifically binds to an AML antigen and (ii) a cytokine receptor polypeptide comprising a transmembrane domain and an intracellular domain, wherein the extracellular antigen binding domain is located at the N-terminus of the cytokine receptor polypeptide. In certain embodiments, engineered immune cells can be transduced with a vector comprising nucleic acid sequences that encode any CIR fusion protein disclosed herein. Examples of AML antigens include, but are not limited to, CD33, FLT3, CD123, and CD371. [00169] The presently disclosed subject matter also provides methods of using such engineered immune cells for the treatment of cancer (e.g., AML). The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter can express non-endogenous levels of any CIR fusion protein disclosed herein for the treatment of cancer, e.g., for treatment of AML. Such engineered immune cells (e.g., NK cells, NK- like CD8 T cells) can be administered to a subject (e.g., a human subject) in need thereof for the treatment of cancer (e.g., AML). [00170] The presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) may further include at least one recombinant or exogenous co-stimulatory ligand. For example, the presently disclosed engineered immune cells (e.g., NK cells, NK- like CD8 T cells) can be further transduced with at least one co- stimulatory ligand, such that the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) co-expresses or is induced to co-express any CIR fusion protein disclosed herein and the at least one co- stimulatory ligand. Co-stimulatory ligands include, but are not limited to, members of the tumor necrosis factor (TNF) superfamily, and immunoglobulin (Ig) superfamily ligands. TNF is a cytokine involved in systemic inflammation and stimulates the acute phase reaction. Its primary role is in the regulation of immune cells. Members of TNF superfamily share a number of common features. The majority of TNF superfamily members are synthesized as type II transmembrane proteins (extracellular C-terminus) -43- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 containing a short cytoplasmic segment and a relatively long extracellular region. TNF superfamily members include, without limitation, nerve growth factor (NGF), CD40L (CD40L)/CD 154, CD137L/4-1BBL, TNF-a, CD134L/OX40L/CD252, CD27L/CD70, Fas ligand (FasL), CD30L/CD153, tumor necrosis factor beta (TNFP)/lymphotoxin-alpha (LTa), lymphotoxin-beta Ο-Τβ), CD257/B cell-activating factor (B AFF)/Bly s/THANK/Tall- 1, glucocorticoid-induced TNF Receptor ligand (GITRL), and TF-related apoptosis-inducing ligand (TRAIL), LIGHT (TNFSF14). The immunoglobulin (Ig) superfamily is a large group of cell surface and soluble proteins that are involved in the recognition, binding, or adhesion processes of cells. These proteins share structural features with immunoglobulins — they possess an immunoglobulin domain (fold). Immunoglobulin superfamily ligands include, but are not limited to, CD80 and CD86, both ligands for CD28, PD-L1/(B7-H1) that ligands for PD-1. In certain embodiments, the at least one co- stimulatory ligand is selected from the group consisting of 4-1BBL, CD80, CD86, CD70, OX40L, CD48, TNFRSF14, PD-L1, and combinations thereof. In certain embodiments, the engineered immune cell (e.g., NK cell, NK-like CD8 T cell) of the present technology comprises one recombinant co-stimulatory ligand (e.g., 4-1BBL). In certain embodiments, the engineered immune cell (e.g., NK cell, NK-like CD8 T cell) comprises two recombinant co-stimulatory ligands (e.g., 4-1BBL and CD80). [00171] Furthermore, the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can further comprise at least one exogenous cytokine. For example, a presently disclosed engineered immune cell (e.g., NK cell, NK-like CD8 T cell) can be further transduced with at least one cytokine, such that the engineered immune cell (e.g., NK cell, NK-like CD8 T cell) secrete the at least one cytokine as well as express any CIR fusion protein disclosed herein. Additionally or alternatively, in certain embodiments, the at least one cytokine is selected from the group consisting of IL-2, IL-4, IL-7, IL-12, IL- 15, IL-18, IL-21 and IL-23. [00172] The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can be generated from peripheral donor lymphocytes (see Examples described herein). The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can be autologous, non- autologous (e.g., allogeneic), or derived in vitro from engineered progenitor or stem cells. -44- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00173] In certain embodiments, the presently disclosed engineered immune cell (e.g., NK cell, NK-like CD8 T cell) expresses from about 1 to about 5, from about 1 to about 4, from about 2 to about 5, from about 2 to about 4, from about 3 to about 5, from about 3 to about 4, from about 4 to about 5, from about 1 to about 2, from about 2 to about 3, from about 3 to about 4, or from about 4 to about 5 vector copy numbers per cell of a heterologous nucleic acid encoding any CIR fusion protein disclosed herein. [00174] For example, the higher the non-endogenous levels of any CIR fusion protein disclosed herein in an engineered immune cell (e.g., NK cell, NK-like CD8 T cell), the greater cytotoxicity and/or cytokine production the engineered immune cell (e.g., NK cell, NK-like CD8 T cell) exhibits. [00175] Additionally, or alternatively, the cytotoxicity and cytokine production of a presently disclosed engineered immune cell (e.g., NK cell, NK-like CD8 T cell) are proportional to the expression level of any CIR fusion protein disclosed herein in the immune cell (e.g., NK cell, NK-like CD8 T cell). [00176] The unpurified source of immune cells (e.g., NK cells, NK-like CD8 T cells) can be any known in the art, such as the bone marrow, fetal, neonate or adult or other hematopoietic cell source, e.g., fetal liver, peripheral blood or umbilical cord blood. Various techniques can be employed to separate the cells. For instance, negative selection methods can remove non-immune cell initially. Monoclonal antibodies are particularly useful for identifying markers associated with particular cell lineages and/or stages of differentiation for both positive and negative selections. [00177] A large proportion of terminally differentiated cells can be initially removed by a relatively crude separation. For example, magnetic bead separations can be used initially to remove large numbers of irrelevant cells. In some embodiments, at least about 80%, usually at least 70% of the total hematopoietic cells will be removed prior to cell isolation. [00178] Procedures for separation include, but are not limited to, density gradient centrifugation; resetting; coupling to particles that modify cell density; magnetic separation with antibody-coated magnetic beads; affinity chromatography; cytotoxic agents joined to or used in conjunction with a mAb, including, but not limited to, complement and -45- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 cytotoxins; and panning with antibody attached to a solid matrix, e.g., plate, chip, elutriation or any other convenient technique. [00179] Techniques for separation and analysis include, but are not limited to, flow cytometry, which can have varying degrees of sophistication, e.g., a plurality of color channels, low angle and obtuse light scattering detecting channels, impedance channels. [00180] The cells can be selected against dead cells, by employing dyes associated with dead cells such as propidium iodide (PI). In some embodiments, the cells are collected in a medium comprising 2% fetal calf serum (FCS) or 0.2% bovine serum albumin (BSA) or any other suitable, preferably sterile, isotonic medium. [00181] In some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) comprise one or more additional modifications. For example, in some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) comprise and express (is transduced to express) a chimeric co-stimulatory receptor (CCR). CCR is described in Krause et al. (1998) J. Exp. Med.188(4):619-626, and US20020018783, the contents of which are incorporated by reference in their entireties. CCRs mimic co- stimulatory signals, but do not provide a T-cell activation signal, e.g., CCRs lack a CD3ζ polypeptide. CCRs provide co-stimulation, e.g., a CD28-like signal, in the absence of the natural co-stimulatory ligand on the antigen-presenting cell. [00182] In some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) are further modified to suppress expression of one or more genes. In some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) are further modified via genome editing. Various methods and compositions for targeted cleavage of genomic DNA have been described. Such targeted cleavage events can be used, for example, to induce targeted mutagenesis, induce targeted deletions of cellular DNA sequences, and facilitate targeted recombination at a predetermined chromosomal locus. See, for example, U.S. Patent Nos.7,888,121; 7,972,854; 7,914,796; 7,951,925; 8,110,379; 8,409,861; 8,586,526; U.S. Patent Publications 20030232410; 20050208489; 20050026157; 20050064474; 20060063231; 201000218264; 20120017290; 20110265198; 20130137104; 20130122591; 20130177983 and 20130177960, the disclosures of which are incorporated by reference in their entireties. These methods often involve the use of engineered cleavage systems to induce a double strand break (DSB) or a nick in a target DNA sequence such that -46- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 repair of the break by an error born process such as non-homologous end joining (NHEJ) or repair using a repair template (homology directed repair or HDR) can result in the knock out of a gene or the insertion of a sequence of interest (targeted integration). Cleavage can occur through the use of specific nucleases such as engineered zinc finger nucleases (ZFN), transcription-activator like effector nucleases (TALENs), or using the CRISPR/Cas system with an engineered crRNA/tracr RNA ('single guide RNA') to guide specific cleavage. In some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) are modified to result in disruption or inhibition of PD-1, PDL-1, Tim-3 or CTLA-4 (see, e.g. U.S. Patent Publication 20140120622), or other immunosuppressive factors known in the art (Wu et al. (2015) Oncoimmunology 4(7): e1016700, Mahoney et al. (2015) Nature Reviews Drug Discovery 14, 561–584). Formulations [00183] The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology and compositions comprising the same can be conveniently provided as sterile liquid preparations, e.g., isotonic aqueous solutions, suspensions, emulsions, dispersions, or viscous compositions, which may be buffered to a selected pH. Liquid preparations are normally easier to prepare than gels, other viscous compositions, and solid compositions. Additionally, liquid compositions are somewhat more convenient to administer, especially by injection. Viscous compositions, on the other hand, can be formulated within the appropriate viscosity range to provide longer contact periods with specific tissues. Liquid or viscous compositions can comprise carriers, which can be a solvent or dispersing medium containing, for example, water, saline, phosphate buffered saline, polyol (for example, glycerol, propylene glycol, liquid polyethylene glycol, and the like) and suitable mixtures thereof. [00184] Sterile injectable solutions can be prepared by incorporating the compositions of the presently disclosed subject matter, e.g., a composition comprising the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology, in the required amount of the appropriate solvent with various amounts of the other ingredients, as desired. Such compositions may be in admixture with a suitable carrier, diluent, or excipient such as sterile water, physiological saline, glucose, dextrose, or the like. The compositions can also be lyophilized. The compositions can contain auxiliary substances such as wetting, dispersing, or emulsifying agents (e.g., methylcellulose), pH buffering -47- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 agents, gelling or viscosity enhancing additives, preservatives, flavoring agents, colors, and the like, depending upon the route of administration and the preparation desired. Standard texts, such as “REMINGTON' S PHARMACEUTICAL SCIENCE”, 17th edition, 1985, incorporated herein by reference, may be consulted to prepare suitable preparations, without undue experimentation. [00185] Various additives which enhance the stability and sterility of the compositions, including antimicrobial preservatives, antioxidants, chelating agents, and buffers, can be added. Prevention of the action of microorganisms can be ensured by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. Prolonged absorption of the injectable pharmaceutical form can be brought about by the use of agents delaying absorption, for example, aluminum monostearate and gelatin. According to the presently disclosed subject matter, however, any vehicle, diluent, or additive used would have to be compatible with the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter. [00186] The compositions can be isotonic, i.e., they can have the same osmotic pressure as blood and lacrimal fluid. The desired isotonicity of the compositions of the presently disclosed subject matter may be accomplished using sodium chloride, or other pharmaceutically acceptable agents such as dextrose, boric acid, sodium tartrate, propylene glycol or other inorganic or organic solutes. Sodium chloride is suitable particularly for buffers containing sodium ions. [00187] Viscosity of the compositions, if desired, can be maintained at the selected level using a pharmaceutically acceptable thickening agent. Methylcellulose can be used because it is readily and economically available and is easy to work with. Other suitable thickening agents include, for example, xanthan gum, carboxymethyl cellulose, hydroxypropyl cellulose, carbomer, and the like. The concentration of the thickener can depend upon the agent selected. The important point is to use an amount that will achieve the selected viscosity. Obviously, the choice of suitable carriers and other additives will depend on the exact route of administration and the nature of the particular dosage form, e.g., liquid dosage form (e.g., whether the composition is to be formulated into a solution, a suspension, gel or another liquid form, such as a time release form or liquid-filled form). -48- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00188] Those skilled in the art will recognize that the components of the compositions should be selected to be chemically inert and will not affect the viability or efficacy of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) as described in the presently disclosed subject matter. This will present no problem to those skilled in chemical and pharmaceutical principles, or problems can be readily avoided by reference to standard texts or by simple experiments (not involving undue experimentation), from this disclosure and the documents cited herein. [00189] One consideration concerning the therapeutic use of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter is the quantity of cells necessary to achieve an optimal effect. The quantity of cells to be administered will vary for the subject being treated. In certain embodiments, from about 10² to about 10¹², from about 10³ to about 10¹¹, from about 10⁴ to about 10¹⁰, from about 10⁵ to about 10⁹, or from about 10⁶ to about 10⁸ engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter are administered to a subject. More effective cells may be administered in even smaller numbers. In some embodiments, at least about 1 × 10⁸, about 2 × 10⁸, about 3 × 10⁸, about 4 × 10⁸, about 5 × 10⁸, about 1 × 10⁹, about 5 × 10⁹, about 1 × 10¹⁰, about 5 × 10¹⁰, about 1 × 10¹¹, about 5 × 10¹¹, about 1 × 10¹² or more engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter are administered to a human subject. The precise determination of what would be considered an effective dose may be based on factors individual to each subject, including their size, age, sex, weight, and condition of the particular subject. Dosages can be readily ascertained by those skilled in the art from this disclosure and the knowledge in the art. Generally, engineered immune cells (e.g., NK cells, NK-like CD8 T cells) are administered at doses that are nontoxic or tolerable to the patient. [00190] The skilled artisan can readily determine the amount of cells and optional additives, vehicles, and/or carrier in compositions to be administered in methods of the presently disclosed subject matter. Typically, any additives (in addition to the active cell(s) and/or agent(s)) are present in an amount of from about 0.001% to about 50% by weight) solution in phosphate buffered saline, and the active ingredient is present in the order of micrograms to milligrams, such as from about 0.0001 wt % to about 5 wt %, from about 0.0001 wt% to about 1 wt %, from about 0.0001 wt% to about 0.05 wt%, from about 0.001 -49- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 wt% to about 20 wt %, from about 0.01 wt% to about 10 wt %, or from about 0.05 wt% to about 5 wt %. For any composition to be administered to an animal or human, and for any particular method of administration, toxicity should be determined, such as by determining the lethal dose (LD) and LD50 in a suitable animal model e.g., rodent such as mouse; and, the dosage of the composition(s), concentration of components therein and timing of administering the composition(s), which elicit a suitable response. Such determinations do not require undue experimentation from the knowledge of the skilled artisan, this disclosure and the documents cited herein. And, the time for sequential administrations can be ascertained without undue experimentation. Administration [00191] The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter can be provided systemically or directly to a subject for treating cancer (e.g., AML). In certain embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) described herein are directly injected into an organ of interest. Additionally or alternatively, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology are provided indirectly to the organ of interest, for example, by administration into the circulatory system (e.g., the tumor vasculature) or into the tissue of interest. Expansion and differentiation agents can be provided prior to, during or after administration of cells and compositions to increase production of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) either in vitro or in vivo. [00192] Engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter can be administered in any physiologically acceptable vehicle, systemically or regionally, normally intravascularly, intraperitoneally, intrathecally, or intrapleurally, although they may also be introduced into bone or other convenient site where the cells may find an appropriate site for regeneration and differentiation (e.g., thymus). In certain embodiments, at least 1 × 10⁵ cells can be administered, eventually reaching 1 × 10¹⁰ or more. In certain embodiments, at least 1 × 10⁶ cells can be administered. A cell population comprising engineered immune cells (e.g., NK cells, NK- like CD8 T cells) can comprise a purified population of cells. Those skilled in the art can readily determine the percentage of engineered immune cells (e.g., NK cells, NK-like CD8 T cells) in a cell population using various well-known methods, such as fluorescence -50- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 activated cell sorting (FACS). The ranges of purity in cell populations comprising engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can be from about 50% to about 55%, from about 55% to about 60%, about 60% to about 65%, from about 65% to about 70%, from about 70% to about 75%, from about 75% to about 80%, from about 80% to about 85%; from about 85% to about 90%, from about 90% to about 95%, or from about 95 to about 100%. Dosages can be readily adjusted by those skilled in the art (e.g., a decrease in purity may require an increase in dosage). The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can be introduced by injection, catheter, or the like. If desired, factors can also be included, including, but not limited to, interleukins, e.g., IL-2, IL-4, IL-7, IL-12, IL-15, IL-18, IL-21, IL-23, as well as the other interleukins, the colony stimulating factors, such as G-, M- and GM-CSF, interferons, e.g., γ- interferon. [00193] In certain embodiments, compositions of the presently disclosed subject matter comprise any and all embodiments of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology with a pharmaceutically acceptable carrier. Administration can be autologous or non-autologous. For example, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology and compositions comprising the same can be obtained from one subject, and administered to the same subject or a different, compatible subject. Peripheral blood derived immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter or their progeny (e.g., in vivo, ex vivo or in vitro derived) can be administered via localized injection, including catheter administration, systemic injection, localized injection, intravenous injection, or parenteral administration. When administering a pharmaceutical composition of the presently disclosed subject matter (e.g., a pharmaceutical composition comprising any and all embodiments of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) disclosed herein), it can be formulated in a unit dosage injectable form (solution, suspension, emulsion). Therapeutic Uses of the Engineered Immune Cells of the Present Technology [00194] For treatment, the amount of the engineered immune cells (e.g., NK cells, NK- like CD8 T cells) provided herein administered is an amount effective in producing the desired effect, for example, treatment of a cancer (e.g., AML) or one or more symptoms of a cancer (e.g., AML). An effective amount can be provided in one or a series of -51- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 administrations of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) provided herein. An effective amount can be provided in a bolus or by continuous perfusion. For adoptive immunotherapy using the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) disclosed herein, cell doses in the range of about 10⁶ to about 10¹⁰ may be infused. Lower doses of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology may be administered, e.g., about 10⁴ to about 10⁸. [00195] The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter can be administered by any methods known in the art, including, but not limited to, pleural administration, intravenous administration, subcutaneous administration, intranodal administration, intratumoral administration, intrathecal administration, intrapleural administration, intraperitoneal administration, and direct administration to the thymus. In certain embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology and the compositions comprising thereof are intravenously administered to the subject in need. Methods for administering cells for adoptive cell therapies, including, for example, donor lymphocyte infusion and engineered immune cell therapies, and regimens for administration are known in the art and can be employed for administration of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) provided herein. [00196] For example, the presently disclosed subject matter provides methods of reducing tumor burden in a subject. In one non-limiting example, the method of reducing tumor burden comprises administering an effective amount of the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to the subject and administering a suitable antibody targeted to the tumor, thereby inducing tumor cell death in the subject. In some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) and the antibody are administered at different times. For example, in some embodiments, the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) are administered and then the antibody is administered. In some embodiments, the antibody is administered 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 18 hours, 24 hours, 30 hours, 26 hours, 48 hours or more than 48 hours after the administration of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology. -52- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00197] The presently disclosed subject matter provides various methods of using any and all embodiments of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) provided herein. For example, the presently disclosed subject matter provides methods of reducing tumor burden in a subject. In one non-limiting example, the method of reducing tumor burden comprises administering an effective amount of the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to the subject, thereby inducing tumor cell death in the subject. [00198] The presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can reduce the number of tumor cells, reduce tumor size, and/or eradicate the tumor in the subject. In certain embodiments, the method of reducing tumor burden comprises administering an effective amount of engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology to the subject, thereby inducing tumor cell death in the subject. In some embodiments, the cancer (e.g., AML) is a relapsed or refractory cancer (e.g., AML). In some embodiments, the cancer (e.g., AML) is resistant to one or more cancer therapies, e.g., one or more chemotherapeutic drugs. [00199] The presently disclosed subject matter also provides methods of increasing or lengthening survival of a subject with cancer (e.g., AML). In one non-limiting example, the method of increasing or lengthening survival of a subject with cancer (e.g., AML) comprises administering an effective amount of the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to the subject, thereby increasing or lengthening survival of the subject. The presently disclosed subject matter further provides methods for treating cancer (e.g., AML) in a subject, comprising administering the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to the subject. Also provided herein are methods for treating of inhibiting tumor growth or metastasis in a subject comprising contacting a tumor cell with an effective amount of any of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) provided herein. [00200] Cancers whose growth may be inhibited using the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the presently disclosed subject matter include cancers typically responsive to immunotherapy (e.g., AML). [00201] Additionally, the presently disclosed subject matter provides methods of increasing immune-activating cytokine production in response to a cancer (e.g., AML) cell -53- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 in a subject in need thereof. In one non-limiting example, the method comprises administering the presently disclosed engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to the subject. The immune-activating cytokine (which is also referred to herein as a cytokine) can be granulocyte macrophage colony stimulating factor (GM-CSF), IFNα, IFN-β, IFN-γ, TNFα, IL-2, IL-4, IL-7, IL-12, IL-15, IL-18, IL-21, IL-23, and combinations thereof. [00202] Suitable human subjects for therapy typically comprise two treatment groups that can be distinguished by clinical criteria. Subjects with “advanced disease” or “high tumor burden” are those who bear a clinically measurable tumor. A clinically measurable tumor is one that can be detected on the basis of tumor mass (e.g., by palpation, CAT scan, sonogram, mammogram or X-ray; positive biochemical or histopathologic markers on their own are insufficient to identify this population). A pharmaceutical composition embodied in the presently disclosed subject matter is administered to these subjects to elicit an anti- tumor response, with the objective of palliating their condition. Ideally, reduction in tumor mass occurs as a result, but any clinical improvement constitutes a benefit. Clinical improvement comprises decreased risk or rate of progression or reduction in pathological consequences of the tumor. [00203] Another group of suitable subjects is known in the art as the “adjuvant group.” These are individuals who have had a history of neoplasia, but have been responsive to another mode of therapy. The prior therapy can have included, but is not restricted to, surgical resection, radiotherapy, and traditional chemotherapy. As a result, these individuals have no clinically measurable tumor. However, they are suspected of being at risk for progression of the disease, either near the original tumor site, or by metastases. This group can be further subdivided into high-risk and low-risk individuals. The subdivision is made on the basis of features observed before or after the initial treatment. These features are known in the clinical arts, and are suitably defined for each different neoplasia. Features typical of high-risk subgroups are those in which the tumor has invaded neighboring tissues, or who show involvement of lymph nodes. Another group has a genetic predisposition to neoplasia but has not yet evidenced clinical signs of neoplasia. [00204] The subjects can have an advanced form of disease, in which case the treatment objective can include mitigation or reversal of disease progression, and/or amelioration of -54- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 side effects. The subjects can have a history of the condition, for which they have already been treated, in which case the therapeutic objective will typically include a decrease or delay in the risk of recurrence. [00205] Further modification can be introduced to the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to avert or minimize the risks of immunological complications, or when healthy tissues express the same target antigens as the tumor cells, leading to outcomes similar to GvHD. Modification of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) can include engineering a suicide gene into the engineered immune cells (e.g., NK cells, NK-like CD8 T cells). Suitable suicide genes include, but are not limited to, Herpes simplex virus thymidine kinase (hsv- tk), inducible Caspase 9 Suicide gene (iCasp-9), and a truncated human epidermal growth factor receptor (EGFRt) polypeptide. In certain embodiments, the suicide gene is an EGFRt polypeptide. The EGFRt polypeptide can enable engineered immune cells (e.g., NK cells, NK-like CD8 T cells) elimination by administering anti-EGFR monoclonal antibody (e.g., cetuximab). The suicide gene can be included within the vector comprising nucleic acids encoding any CIR fusion protein disclosed herein. A presently disclosed engineered immune cell (e.g., NK cell, NK-like CD8 T cell) incorporated with a suicide gene can be pre-emptively eliminated at a given time point post engineered immune cell infusion, or eradicated at the earliest signs of toxicity. [00206] In another aspect, the present disclosure provides a method of preparing immune cells for adoptive cell therapy (ACT) comprising: (a) isolating immune cells (e.g., NK cells, NK-like CD8 T cells) from a donor subject, (b) transducing the immune cells with a nucleic acid encoding any and all embodiments of the CIR fusion protein described herein or an expression vector comprising said nucleic acid, and (c) administering the transduced immune cells to a recipient subject. In certain embodiments, the nucleic acid encodes the amino acid sequence of any one of SEQ ID NOs: 28-35. In some embodiments of the ACT methods described herein, the donor subject and the recipient subject are the same or different. Combination Therapy [00207] The engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology may be employed in conjunction with other therapeutic agents useful in -55- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 the treatment of cancers (e.g., AML). For example, any and all embodiments of the engineered immune cells (e.g., NK cells, NK-like CD8 T cells) described herein may be separately, sequentially or simultaneously administered with at least one additional cancer therapy. Examples of additional cancer therapy include chemotherapeutic agents, immune checkpoint inhibitors, monoclonal antibodies that specifically target tumor antigens, immune activating agents (e.g., interferons, interleukins, cytokines), oncolytic virus therapy and cancer vaccines. In some embodiments, the additional cancer therapy is selected from among a chemotherapy, a radiation therapy, an immunotherapy, a monoclonal antibody, an anti-cancer nucleic acid, an anti-cancer protein, an anti-cancer virus or microorganism, a cytokine, or any combination thereof. [00208] Radiation therapy includes, but is not limited to, exposure to radiation, e.g., ionizing radiation, UV radiation, as known in the art. Exemplary dosages include, but are not limited to, a dose of ionizing radiation at a range from at least about 2 Gy to not more than about 10 Gy or a dose of ultraviolet radiation at a range from at least about 5 J/m² to not more than about 50 J/m², usually about 10 J/m². [00209] In some embodiments, the methods further comprise sequentially, separately, or simultaneously administering an immunotherapy to the subject. In some embodiments, the immunotherapy regulates immune checkpoints. In further embodiments, the immunotherapy comprises, or consists essentially of, or yet further consists of an immune checkpoint inhibitor, such as a Cytotoxic T-Lymphocyte Associated Protein 4 (CTLA-4) inhibitor, or a Programmed Cell Death 1 (PD-1) inhibitor, or a Programmed Death Ligand 1 (PD-L1) inhibitor. In yet further embodiments, the immune checkpoint inhibitor comprises, or consists essentially of, or yet further consists of an antibody or an equivalent thereof recognizing and binding to an immune checkpoint protein, such as an antibody or an equivalent thereof recognizing and binding to CTLA-4 (for example, Yervoy (ipilimumab), CP-675,206 (tremelimumab), AK104 (cadonilimab), or AGEN1884 (zalifrelimab)), or an antibody or an equivalent thereof recognizing and binding to PD-1 (for example, Keytruda (pembrolizumab), Opdivo (nivolumab), Libtayo (cemiplimab), Tyvyt (sintilimab), BGB- A317 (tislelizumab), JS001 (toripalimab), SHR1210 (camrelizumab), GB226 (geptanolimab), JS001 (toripalimab), AB122 (zimberelimab), AK105 (penpulimab), HLX10 (serplulimab), BCD-100 (prolgolimab), AGEN2034 (balstilimab), MGA012 (retifanlimab), -56- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 AK104 (cadonilimab), HX008 (pucotenlimab), PF-06801591 (sasanlimab), JNJ-63723283 (cetrelimab), MGD013 (tebotelimab), CT-011 (pidilizumab), or Jemperli (dostarlimab)), or an antibody or an equivalent thereof recognizing and binding to PD-L1 (for example, Tecentriq (atezolizumab), Imfinzi (durvalumab), Bavencio (avelumab), CS1001 (sugemalimab), or KN035 (envafolimab)). [00210] In some embodiments, the methods further comprise sequentially, separately, or simultaneously administering a cytokine to the subject. In some embodiments, the cytokine is administered prior to, during, or subsequent to administration of the one or more engineered immune cells (e.g., NK cells, NK-like CD8 T cells). In some embodiments, the cytokine is selected from the group consisting of interferon α, interferon β, interferon γ, complement C5a, IL-2, TNFα, CD40L, IL-12, IL-23, IL-15, IL-18, IL-21, CCL1, CCL11, CCL12, CCL13, CCL14-1, CCL14-2, CCL14-3, CCL15-1, CCL15-2, CCL16, CCL17, CCL18, CCL19, CCL19, CCL2, CCL20, CCL21, CCL22, CCL23-1, CCL23-2, CCL24, CCL25-1, CCL25-2, CCL26, CCL27, CCL28, CCL3, CCL3L1, CCL4, CCL4L1, CCL5, CCL6, CCL7, CCL8, CCL9, CCR10, CCR2, CCR5, CCR6, CCR7, CCR8, CCRL1, CCRL2, CX3CL1, CX3CR, CXCL1, CXCL10, CXCL11, CXCL12, CXCL13, CXCL14, CXCL15, CXCL16, CXCL2, CXCL3, CXCL4, CXCL5, CXCL6, CXCL7, CXCL8, CXCL9, CXCL9, CXCR1, CXCR2, CXCR4, CXCR5, CXCR6, CXCR7 and XCL2. [00211] The methods for treating cancer may further comprise sequentially, separately, or simultaneously administering to the subject at least one chemotherapeutic agent, optionally selected from the group consisting of nitrogen mustards, ethylenimine derivatives, alkyl sulfonates, nitrosoureas, gemcitabine, triazenes, folic acid analogs, anthracyclines, taxanes, COX-2 inhibitors, pyrimidine analogs, purine analogs, antibiotics, enzyme inhibitors, epipodophyllotoxins, platinum coordination complexes, vinca alkaloids, substituted ureas, methyl hydrazine derivatives, adrenocortical suppressants, hormone antagonists, endostatin, taxols, camptothecins, SN-38, doxorubicin, doxorubicin analogs, antimetabolites, alkylating agents, antimitotics, anti-angiogenic agents, tyrosine kinase inhibitors, mTOR inhibitors, heat shock protein (HSP90) inhibitors, proteosome inhibitors, HDAC inhibitors, pro-apoptotic agents, methotrexate and CPT-11. -57- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 Kits [00212] The presently disclosed subject matter provides kits for the treatment of AML. In certain embodiments, the kit may be used in the manufacture of a therapeutic composition containing an effective amount of engineered immune cells (e.g., NK cells, NK-like CD8 T cells) described herein. In some embodiments, the kits include a non- endogenous expression vector comprising a heterologous nucleic acid encoding any and all embodiments of the CIR fusion proteins described herein. In certain embodiments, the CIR fusion protein comprises (i) an extracellular antigen binding domain that specifically binds to an AML antigen (e.g., CD33, FLT3, CD123, and CD371) and (ii) a cytokine receptor polypeptide comprising a transmembrane domain and an intracellular domain, wherein the extracellular antigen binding domain is located at the N-terminus of the cytokine receptor polypeptide. [00213] In some embodiments, the kit comprises a sterile container which contains the kit components; such containers can be boxes, ampules, bottles, vials, tubes, bags, pouches, blister-packs, or other suitable container forms known in the art. Such containers can be made of plastic, glass, laminated paper, metal foil, or other materials suitable for holding medicaments. [00214] If desired, the kit further comprises instructions for preparing engineered immune cells (e.g., NK cells, NK-like CD8 T cells) of the present technology for adoptive cell therapy, and methods of using said engineered immune cells (e.g., NK cells, NK-like CD8 T cells) to treat AML in a subject in need thereof. The instructions will generally include information about the use of the composition for the treatment of AML. In other embodiments, the instructions include at least one of the following: description of the therapeutic agent; dosage schedule and administration for treatment of AML or symptoms thereof; precautions; warnings; indications; counter-indications; overdose information; adverse reactions; animal pharmacology; clinical studies; and/or references. The instructions may be printed directly on the container (when present), or as a label applied to the container, or as a separate sheet, pamphlet, card, or folder supplied in or with the container. [00215] The at least one engineered immune cell (e.g., NK cells, NK-like CD8 T cells) of the present technology may be provided to the subject in the form of a syringe or -58- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 autoinjection pen containing a sterile, liquid formulation or lyophilized preparation (e.g., Kivitz et al., Clin. Ther.28:1619-29 (2006)). [00216] A device capable of delivering the engineered immune cells (e.g., NK cells, NK- like CD8 T cells) of the present technology through an administrative route may be included in the kit. Examples of such devices include syringes (for parenteral administration) or inhalation devices. [00217] The kit components may be packaged together or separated into two or more containers. In some embodiments, the containers may be vials that contain sterile, lyophilized formulations that are suitable for reconstitution. A kit may also contain one or more buffers suitable for reconstitution and/or dilution of other reagents. Other containers that may be used include, but are not limited to, a pouch, tray, box, tube, or the like. Kit components may be packaged and maintained sterilely within the containers. EXAMPLES [00218] The present technology is further illustrated by the following Examples, which should not be construed as limiting in any way. Example 1: Chimeric Innate Receptors of the Present Technology [00219] We designed a chimeric innate receptor that links an anti-CD33 scFv with the transmembrane and intracellular domain of a CD122 molecule. CD33 is a common antigen found on cells of myeloid lineages, including various AMLs, thus propagating IL-15 signaling and leading to increased proliferation, activation and effector molecule preparation in recognition of AML (FIG.6). The sequences of the generated CIR correspond to SEQ ID NOs: 28-30 and 34. [00220] Results. Baseline CD122 and CD132 expression is higher in NK-like CD8 T cells compared to conventional CD8 T cells (FIG.3). NK and other NK-like CD8 T cells are responsive to IL-15 stimulation (FIGs.4-5). Engineered NKG2C+ and NK cells did not signal in response to CD33 but had enhanced signaling with IL-15. See FIG.8. IL-21R can be used as an alternate cytokine receptor (instead of IL-15 signalling). Other common AML antigens could be considered such as FLT3, CD123, CD371. -59- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 EQUIVALENTS [00221] The present technology is not to be limited in terms of the particular embodiments described in this application, which are intended as single illustrations of individual aspects of the present technology. Many modifications and variations of this present technology can be made without departing from its spirit and scope, as will be apparent to those skilled in the art. Functionally equivalent methods and apparatuses within the scope of the present technology, in addition to those enumerated herein, will be apparent to those skilled in the art from the foregoing descriptions. Such modifications and variations are intended to fall within the scope of the present technology. It is to be understood that this present technology is not limited to particular methods, reagents, compounds compositions or biological systems, which can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting. [00222] In addition, where features or aspects of the disclosure are described in terms of Markush groups, those skilled in the art will recognize that the disclosure is also thereby described in terms of any individual member or subgroup of members of the Markush group. [00223] As will be understood by one skilled in the art, for any and all purposes, particularly in terms of providing a written description, all ranges disclosed herein also encompass any and all possible subranges and combinations of subranges thereof. Any listed range can be easily recognized as sufficiently describing and enabling the same range being broken down into at least equal halves, thirds, quarters, fifths, tenths, etc. As a non- limiting example, each range discussed herein can be readily broken down into a lower third, middle third and upper third, etc. As will also be understood by one skilled in the art all language such as “up to,” “at least,” “greater than,” “less than,” and the like, include the number recited and refer to ranges which can be subsequently broken down into subranges as discussed above. Finally, as will be understood by one skilled in the art, a range includes each individual member. Thus, for example, a group having 1-3 cells refers to groups having 1, 2, or 3 cells. Similarly, a group having 1-5 cells refers to groups having 1, 2, 3, 4, or 5 cells, and so forth. -60- 4880-6647-0061.1 Atty. Dkt. No.: 115872-3215 [00224] All patents, patent applications, provisional applications, and publications referred to or cited herein are incorporated by reference in their entirety, including all figures and tables, to the extent they are not inconsistent with the explicit teachings of this specification. -61- 4880-6647-0061.1

Claims

Atty. Dkt. No.: 115872-3215 WHAT IS CLAIMED IS 1. A chimeric innate receptor (CIR) fusion protein comprising (i) an extracellular antigen binding domain that specifically binds to an AML antigen and (ii) a mammalian cytokine receptor polypeptide comprising a transmembrane domain and an intracellular domain, wherein the mammalian cytokine receptor polypeptide comprises a mammalian CD122 polypeptide, a mammalian CD132 polypeptide, or a mammalian IL- 21R polypeptide, and wherein the extracellular antigen binding domain is operably linked to the mammalian cytokine receptor polypeptide. 2. The CIR fusion protein of claim 1, wherein the extracellular antigen binding domain and the mammalian cytokine receptor polypeptide are directly linked or linked via a peptide linker. 3. The CIR fusion protein of claim 1 or 2, wherein the extracellular antigen binding domain is located at the N-terminus of the mammalian cytokine receptor polypeptide. 4. The CIR fusion protein of any one of claims 1-3, wherein the mammalian cytokine receptor polypeptide comprises the amino acid sequence of SEQ ID NO: 21, SEQ ID NO: 22, or SEQ ID NO: 23. 5. The CIR fusion protein of any one of claims 1-4, further comprising a hinge domain between the extracellular antigen binding domain and the mammalian cytokine receptor polypeptide. 6. The CIR fusion protein of claim 5, wherein the hinge domain comprises a CD8 hinge domain, a CD28 hinge domain, a CD122 hinge domain, or a CD132 hinge domain. 7. The CIR fusion protein of claim 5 or 6, wherein the hinge domain comprises the amino acid sequence of SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, or SEQ ID NO: 27. 8. The CIR fusion protein of any one of claims 1-7, further comprising a signal peptide located at the N-terminus of the extracellular antigen binding domain. -62--6647-0061.1 Atty. Dkt. No.: 115872-3215 9. The CIR fusion protein of claim 8, wherein the signal peptide comprises the amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, or SEQ ID NO: 19. 10. The CIR fusion protein of any one of claims 1-9, wherein the AML antigen is selected from the group consisting of CD33, FLT3, CD123, and CD371. 11. The CIR fusion protein of any one of claims 1-10, wherein the extracellular antigen binding domain comprises a scFv, optionally wherein the scFv is a human scFv. 12. The CIR fusion protein of claim 11, wherein the scFv comprises the immunoglobulin variable heavy (VH) domain sequence of SEQ ID NO: 5 and the immunoglobulin variable light (V_L) domain sequence of SEQ ID NO: 6. 13. The CIR fusion protein of claim 11, wherein the scFv comprises the immunoglobulin variable heavy (VH) domain sequence of SEQ ID NO: 7 and the immunoglobulin variable light (VL) domain sequence of SEQ ID NO: 8. 14. The CIR fusion protein of any one of claims 1-13, comprising an amino acid sequence selected from any one of SEQ ID NOs: 28-35. 15. A recombinant nucleic acid molecule encoding the CIR fusion protein of any one of claims 1-14. 16. An expression vector comprising the recombinant nucleic acid molecule of claim 15. 17. A host cell comprising the recombinant nucleic acid molecule of claim 15 or the expression vector of claim 16. 18. An engineered immune cell comprising the CIR fusion protein of any one of claims 1-14. 19. The engineered immune cell of claim 18, wherein the immune cell is a NK cell or a NK-like T cell. 20. A composition comprising the CIR fusion protein of any one of claims 1-14 or the engineered immune cell of claim 18 or 19, and a pharmaceutically-acceptable carrier. -63--6647-0061.1 Atty. Dkt. No.: 115872-3215 21. A method for treating acute myelogenous leukemia (AML) in a subject in need thereof comprising administering to the subject an effective amount of the engineered immune cell of claim 18 or 19. 22. The method of claim 21, further comprising administering to the subject an effective amount of a cytokine, wherein the cytokine is a cognate ligand of the mammalian cytokine receptor polypeptide. 23. The method of claim 22, wherein the cytokine is IL-15 or IL-21. 24. The method of any one of claims 21-23, further comprising separately, sequentially or simultaneously administering an additional therapeutic agent to the subject. 25. A kit comprising the recombinant nucleic acid molecule of claim 15 or the expression vector of claim 16, and instructions for use. 26. A method of preparing immune cells for adoptive cell therapy (ACT) comprising: (a) isolating immune cells from a donor subject, (b) transducing the immune cells with the recombinant nucleic acid molecule of claim 15 or the expression vector of claim 16, and (c) administering the transduced immune cells to a recipient subject. 27. The method of claim 26, wherein the immune cells are NK cells, or NK-like CD8 T cells. 28. The method of claim 26 or 27, wherein the donor subject and the recipient subject are the same or different. -64--6647-0061.1