Perilipin-1 specific t cells for selective fat targeting

Abstract

Targeting perilipin 1 (PLIN1)-expressing fat cells with T-cells is provided. Various T-cell receptors or chimeric antigen receptors (CARs) can be expressed in T-cell, e.g., cytotoxic or regulatory T-cells, to either reduce fat cells in a subject or regulate autoimmune responses to fat cells in a subject, respectively.

Description

Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PCPERILIPIN-1 SPECIFIC T CELLS FOR SELECTIVE FAT TARGETINGCROSS-REFERENCE TO RELATED PATENT APPLICATIONS

[0001] This application claims benefit of priority to U. S. Provisional Patent Application No.63 / 754,302, filed February’ 5, 2025, the entire contents of which are incorporated herein by reference for all purposes.STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSORED RESEARCH AND DEVELOPMENT

[0002] This invention was made with government support under 1R01DK137904-01A1 awarded by the National Institute of Diabetes and Digestive and Kidney Diseases. The government has certain rights m the invention.REFERENCE TO AN ELECTRONIC SEQUENCE LISTING

[0003] The contents of the electronic sequence listing (110221-1543849-012110WO.xml; Size: is 75,503 bytes; and Date of Creation: February, 2, 2026) is herein incorporated by reference in its entirety.BACKGROUND

[0004] T cell tolerance is enforced in the thymus through the expression of a wide array of self proteins called tissue-specific antigens (TSAs) on the surface of specialized medullary thymic epithelial cells (mTECs), under the control of the transcriptional regulator Autoimmune Regulator (Aire) (Anderson MS, Su MA. Nat Rev Immunol. 2016;16(4):247-58.). CD4 T cells that strongly recognize these TSAs are eliminated from the repertoire by undergoing either clonal deletion or conversion to regulatory T cells (Tregs). Highlighting its critical role in T cell tolerance, loss of Aire function in humans leads to Autoimmune Polyglandular Syndrome Type I (APS1), a rare autosomal recessive disorder characterized by multi-organ autoimmunity.Similarly, Aire-deficient (Aire⁻ / ⁻) mice develop a phenotype that closely resembles human disease, including immune infiltrates in multiple organs and tissue-specific autoantibodies. For a number of organs, such as the retina and lung, autoimmunity in Aire⁻ / ⁻ mice has been directly1US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC linked to an absence of thymic expression of an Aire-regulated antigen and escape of tissue¬ specific T cells. Understanding how defects in thymic tolerance lead to autoimmunity and defining the relevant self-antigens is a key step in designing tissue-specific cellular therapies.

[0005] Visceral adipose tissue is an endocrine organ whose main functions are energy storage and metabolic regulation. Dysregulation of adipose tissue homeostasis in key in the pathogenesis of obesity and type 2 diabetes. We have recently begun to appreciate that the adipose tissue houses many immune cell types that are essential for establishing an anti-inflammatory environment and modulating its metabolic activity (Man, K., Kallies, A. & Vasanthakumar, A. Cell Mol Immunol 19, 421-431 (2022). Notably, adipose-resident Tregs are a unique population of Tregs that are distinct from their counterparts in secondary lymphoid organs and are essential for regulation of fat homeostasis (Li C. Spallanzani RG, Mathis D,. Immunol.Rev. 295, 114-125 (2020)). Adipose Tregs possess a restricted oligoclonal T-cell repertoire, but the antigens that, they are recognizing have not been elucidated.

[0006] Acquired lipodystrophies are a group of syndromes characterized by an autoimmune-mediated loss of adipose tissue. Two major subtypes are recognized: acquired generalized lipodystrophy (AGL) which involves extensive loss of subcutaneous fat from nearly all large regions of the body, and acquired partial lipodystrophy (APL) involving mostly the upper body. Loss of adipose tissue predisposes patients with lipodystrophy to major metabolic complications, including insulin resistance and diabetes, dyslipidemia, and fatty liver, presenting a major therapeutic challenge. The pathogenesis of acquired lipodystrophies is not well understood, but a strong association of AGL with other autoimmune disorders, such as juvenile dermatomyositis and systemic lupus erythematous (SLE) has been shown, suggesting that loss of adipocytes may be immune-mediated. In some patients with AGL, infiltration of adipose tissue with mononuclear cells and lymphocytes (panniculitis) has been demonstrated.

[0007] We have recently shown that a subset of patients with acquired generalized lipodystrophy AGL) harbored autoantibodies to an adipocyte-specific lipid droplet protein, perilipin 1 (PLIN1) which is involved in regulation of lipolysis and adipocyte homeostasis (Mandel-Brehm C et al Diabetes. 2023;72(l):59-70.). Notably, anti-perilipin 1 antibodies were also found in a patient with lipodystrophy and autoimmune poly endocrine syndrome type 12US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC (APS1), establishing a link between loss of thymic tolerance and autoimmune attack on the adipose tissue.BRIEF SUMMARY

[0008] Various aspects are described herein. In some embodiments, a cell expressing (i) a heterologous T-cell receptor (TCR) that binds to a perilipin 1 (PLIN1) peptide complexed with a major histocompatibility (MHC) molecule or (ii) a chimeric antigen receptor (CAR) comprising a PLIN1 -binding extracellular domain is provided. In some embodiments, the cell is a T-cell.

[0009] In some embodiments, the cell comprises the heterologous TCR. In some embodiments, the cell does not express an endogenous TCR. In some embodiments, the cell comprises a nucleic acid encoding the heterologous TCR inserted in a coding sequence for an endogenous TCR such that expression of the endogenous TCR is disrupted. In some embodiments, the PLIN1 peptide is an MHC class I or class II peptide and comprises QWGASAAMQVVSRRQSEVRV (SEQ ID NO: 55), PWLHNLAASQDESHDDQTDT (SEQ ID NO: 56), AMQWSRR (SEQ ID NO: 57), SAAMQWSRRQS (SEQ ID NO: 58), WLHNLAASQDE (SEQ ID NO: 59), YNSTKEAHPLVASVCNAYEK (SEQ ID NO:78), SSGRQRTQKAPKAKPSLVRR (SEQ ID NO: 79) or a contiguous fragment of at least 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acids and optionally having one or two amino acid substitutions. In some embodiments, the MHC is a human leukocyte antigen (HLA) molecule and the PLIN1 peptide is a human PLIN1 peptide. In some embodiments, the heterologous TCR comprises an alpha chain comprising a variable alpha (Va) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa3 and a beta chain comprising a variable beta (VP) region comprising a CDRβ1, CDRβ2 and CDRβ3, wherein: CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO: 51) and CDRp3 comprises CASSXDRGDQLYF (SEQ ID NO: 52); or wherein CDRa3 comprises CAMRGXQGGSAKLIF (SEQ ID NO: 53) and CDRp3 comprises CASXXXXGXXX (SEQ ID NO: 54), where X is any ammo acid; or wherein CDRa3 comprises CALRNRGSALGRLHF (SEQ ID NO: 65) and CDR 3 comprises CASSFRVEQYF (SEQ ID NO: 69), or wherein CDRα3 comprises CAASATGGADRLTF (SEQ ID NO: 73) and CDRβ3 comprises CASSLGGDTQYF (SEQ ID NO: 77).3US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0010] In some embodiments, the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO:12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO:20 respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 30, SEQ ID NO: 31 and SEQ ID NO: 32, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44, respectively and CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO:65, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:67, SEQ ID NO:68 and SEQ ID NO:69, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 75, SEQ ID NO: 76 and SEQ ID NO: 77, respectively.

[0011] In some embodiments, the cell comprises the chimeric antigen receptor (CAR) In some embodiments, the PLIN 1 -binding extracellular domain comprises a single chain TCR. In some embodiments, the single chain TCR comprises an alpha chain comprising a variable alpha (Va) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa3 and a beta chain comprising a variable beta (VP) region comprising a CDRpi, CDRP2 and CDRp3, wherein: CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO:51) and CDRP3 comprises CASSXDRGDQLYF (SEQ ID NO: 52); or wherein CDRa3 comprises CAMRGXQGGSAKLIF (SEQ ID NO: 53) and CDRp3 comprises CASXXXXGXXX (SEQ ID NO:54), where X is any amino acid; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO:65, respectively and CDRpi, CDRp2 and CDRp34US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC comprise SEQ ID NO: 67, SEQ ID NO: 68 and SEQ ID NO: 69, respectively; or the CDRal, CDRα2 and CDRα3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:75, SEQ ID NO:76 and SEQ ID NO:77, respectively.

[0012] In some embodiments, the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO:12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO:20, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:30, SEQ ID NO: 31 and SEQ ID NO: 32, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44, respectively and CDRpi, CDRp2 and CDRP3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO:65, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:67, SEQ ID NO:68 and SEQ ID NO:69, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 75, SEQ ID NO: 76 and SEQ ID NO: 77, respectively.

[0013] In some embodiments, the cell is a cytotoxic T-cell. In some embodiments, the cell is a regulatory T-cell. In some embodiments, the cell is a human cell (e.g., a human cytotoxic or regulator T-cell).

[0014] Also provided is a nucleic acid encoding a TCR alpha chain comprising a variable alpha (Va) region comprising a complementarity determining region (CDR) al, CDRa2 and5US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CDRa3 wherein CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO:51) or CAMRGXQGGSAKLIF (SEQ ID NO:53), wherein X is any amino acid; or wherein CDRa3 comprises CALRNRGSALGRLHF (SEQ ID NO: 65); or wherein CDRa3 comprises CAASATGGADRLTF (SEQ ID NO: 73). In some embodiments, the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO:12, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:18, SEQ ID NO:19 and SEQ ID NO:20, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO:65 respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73 respectively.

[0015] Also provided is a nucleic acid encoding a TCR beta chain comprising a variable beta (VP) region comprising a CDRβ1, CDRβ2 and CDRβ3, wherein CDRβ3 comprises CASSXDRGDQLYF (SEQ ID NO:52) or CASXXXXGXXX (SEQ ID NO:54), wherein X is any amino acid; or wherein CDRβ3 comprises CASSFRVEQYF (SEQ ID NO:69), or wherein CDRβ3 comprises CASSLGGDTQYF (SEQ ID NO:77). In some embodiments, the CDRpi, CDR 2 and CDRp3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; or the CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO: 46, SEQ ID NO: 47 and SEQ ID NO:48, respectively; or CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:67, SEQ ID NO:68 and SEQ ID NO:69, respectively; or CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:75, SEQ ID NO:76 and SEQ ID NO:77, respectively.

[0016] In some embodiments, the nucleic acid comprises RN A. In some embodiments, the nucleic acid comprises DNA. In some embodiments, the nucleic acid comprises one or more non-naturally-occurring ribonucleotide or nucleoside modification.

[0017] Also provided is a vector comprising the nucleic acid encoding a TCR alpha chain comprising a variable alpha (Voc) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa.3 wherein CDRa.3 comprises CATDXGGSXXKLXF (SEQ ID NO:51) or CAMRGXQGGSAKLIF (SEQ ID NO:53), wherein X is any amino acid; or a TCR beta chain comprising a variable beta (VP) region comprising a CDRβ1, CDRβ2 and CDRβ3,6US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC wherein CDRp3 comprises CASSXDRGDQLYF (SEQ ID NO: 52) or CASXXXXGXXX (SEQ ID NO: 54), wherein X is any amino acid; or both.

[0018] Also provided is a method of forming a cell that expresses a heterologous T-cell receptor (TCR) that binds to perilipin 1 (PLIN1). In some embodiments, the method comprises, introducing: (a) a nucleic acid encoding a TCR alpha chain comprising a variable alpha (Va) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa3 wherein CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO:51) or CAMRGXQGGSAKLIF (SEQ ID NO: 53), wherein X is any amino acid; and (b) nucleic acid encoding a TCR beta chain comprising a variable beta (VP) region comprising a CDRpi, CDRp2 and CDRp3, wherein CDRp3 comprises CASSXDRGDQLYF (SEQ ID NO: 52) or CASXXXXGXXX (SEQ ID NO: 54), wherein X is any amino acid; such that the cell expresses the TCR alpha and beta chains to form the heterologous TCR that binds to PLIN1.

[0019] In some embodiments, the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO:12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO:20, respectively and CDRpi, CDRP2 and CDRβ3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; or the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:30, SEQ ID NO: 31 and SEQ ID NO: 32, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; or the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44, respectively and CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively,

[0020] In some embodiments, the cell is a T-cell.7US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0021] In some embodiments, the nucleic acid encoding the alpha chain and the nucleic acid encoding the beta chain comprise RNA. In some embodiments, the introducing comprises inserting the nucleic acids into an endogenous TCR coding sequence such that the endogenous TCR is not expressed. In some embodiments, the cells are obtained from a human subject. In some embodiments, following the introducing the cells expressing the TCR alpha and beta chains are introduced into the same or a different human subject. In some embodiments, the introducing occurs in vitro, ex vivo or in vivo.

[0022] Also provided is a method of reducing fat in a subject. In some embodiments, the method comprises introducing a cell as described herein, e.g., a cytotoxic T-cell into the subject in a sufficient amount to reduce fat in the subject. In some embodiments, the subject is a human.

[0023] Also provided is a method of reducing fat in a subject, the method comprising introducing the nucleic acids as described above or herein, or the vector as described above or herein, into one or more cytotoxic T-cell of the subject such that the one or more T-cell expresses a heterologous TCR that binds to PLIN1 and wherein the T-cell expressing the heterologous TCR are in a sufficient amount to reduce fat in the subject. In some embodiments, the subject is a human.

[0024] Also provided is a method of regulating fat in a subject. In some embodiments, the method comprises introducing a cell as described above or elsewhere herein (e.g., a regulatory T-cell) into the subject in a sufficient to reduce fat in the subject. In some embodiments, the subject is a human.

[0025] Also provided is a method of regulating fat in a subject. In some embodiments, the method comprises introducing the nucleic acids as described above or elsewhere herein, or the vector as described above or elsewhere herein, into one or more regulatory T-cell of the subject such that the one or more regulatory T-cell expresses a heterologous TCR that binds to PLIN1 and wherein the regulatory T-cell expressing the heterologous TCR are in a sufficient amount to regulate fat in the subject. In some embodiments, the subject is a human.DEFINITIONS

[0026] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention 8US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC belongs. All publications mentioned herein are incorporated herein by reference for the purpose of describing and disclosing devices, compositions, formulations, and methodologies which are described in the publication and which might be used in connection with the presently described invention.

[0027] Unless otherwise dictated by context, singular terms include pluralities, and plural terms include the singular. Generally, nomenclatures used in connection with, and techniques of, cell and tissue culture, molecular biology, immunology, microbiology, genetics and protein and nucleic acid chemistry are those w’ell-known and commonly used. Known methods and techniques are generally performed according to conventional methods well-known and as described in various general and more specific references, unless otherwise indicated. The nomenclatures used in connection with the laboratory procedures and techniques described in the present disclosure are those well-known and commonly used.

[0028] As used herein, the terms “a”, “an”, and “the” can refer to one or more unless specifically noted otherwise.

[0029] The use of the term “or” is used to mean “and / or,” unless explicitly indicated to refer to alternatives only, or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or.” As used herein “another” can mean at least a second or more.

[0030] The terms “about” and “approximately” as used herein shall generally mean an acceptable degree of error for the quantity measured given the nature or precision of the measurements. Exemplary degrees of error are within 20% (%); preferably, within 10%; and more preferably, within 5% of a given value or range of values. Any reference to “about X” or “approximately X” specifically indicates at least the values X, 0.95X, 0.96X, 0.97X, 0.98X, 0.99X, 1.01X, 1.02X, 1.03X, 1.04X, and 1.05X. Thus, expressions “about X” or “approximately X” are intended to teach and provide written support for a claim limitation of, for example, “0.98X.” Alternatively, in biological systems, the terms “about” and “approximately” may mean values that are within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold of a given value. Numerical quantities given herein are approximate unless stated otherwise, meaning that the term “about” or “approximately” can be inferred when not expressly9US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC stated. When “about” is applied to the beginning of a numerical range, it applies to both ends of the range.

[0031] The term “subject” or “patient” refers to a human or an animal (particularly a mammal) that receive either prophylactic or therapeutic treatment. For example, a subject can be a human. Examples of subjects include, but are not limited to: humans and other primates, including nonhuman primates, such as chimpanzees and other apes and monkey species; farm animals, such as cattle, sheep, pigs, goats and horses; domestic mammals such as dogs and cats; laboratory animals including rodents, such as mice, rats and guinea pigs; birds, including domestic, wild and game birds, such as chickens, turkeys and other gallinaceous birds, ducks, geese, and the like. The terms individual, subject, and patient, by themselves, do not denote a particular age, sex, race, or clinical status. Thus, subjects of any age, whether male or female, are intended to be covered by the present disclosure. Likewise, the methods of the present invention can be applied to any human race, including, for example, Caucasian (white), African-American (black), Native American, Native Hawaiian, Hispanic, Latino, Asian, and European,

[0032] “Pharmaceutically acceptable carrier” and “pharmaceutically acceptable excipient” are used interchangeably and refer to a substance or compound that aids or facilitates preparation, storage, administration, delivery, effectiveness, absorption by a subject, or any other feature of the composition for its intended use or purpose. Such a pharmaceutically acceptable carrier is not biologically or otherwise undesirable and can be included in the compositions described herein without causing a significant adverse toxicological effect on the subject or interacting in a deleterious manner with the other components of the pharmaceutical composition.

[0033] As used herein, the term “administering”, “administration”, or “administer” means delivering a nucleic acid or cell as described herein to a subject. The pharmaceutical compositions described herein are designed for delivery to subjects in need thereof by any suitable route or a combination of different routes. In particular embodiments, the cells or nucleic acids are administered to fat in the subject, e.g., via injection.

[0034] Perilipin 1 (PLIN1) is a protein involved in lipid storage droplets in adipocytes and is found in for example humans and mice. An exemplary human PLIN1 polypeptide is:MAVNKGLTLLDGDLPEQENVLQRVLQLPVVSGTCECFQKTYTSTKEAHPLVASVCNAY EKGVQSASSLAAWSMEPWRRLSTQFTAANELACRGLDHLEEKIPALQYPPEKIASELK10US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC DTISTRLRSARNSISVPIASTSDKVLGAALAGCELAWGVARDTAEFAANTRAGRLASGG ADLALGSIEKVVEYLLPADKEESAPAPGHQQAQKSPKAKPSLLSRVGALTNTLSRYTVQ TMARALEQGHTVAMWIPGVVPLSSLAQWGASVAMQAVSRRRSEVRVPWLHSLAAAQ EEDHEDQTDTEGEDTEEEEELETEENKFSEVAALPGPRGLLGGVAHTLQKTLQTTISAVT WAPAAVLGMAGRVLHLTPAPAVSSTKGRAMSLSDALKGVTDNVVDTVVHYVPLPRLS LMEPESEFRDIDNPPAEVERREAERRASGAPSAGPEPAPRLAQPRRSLRSAQSPGAPPGPG LEDEVATPAAPRPGFPAVPREKPKRRVSDSFFRPSVMEPILGRTHYSQLRKKS (SEQ ID NO: 60).

[0035] An exemplary mouse PLIN1 polypeptide is:MSMNKGPTLLDGDLPEQENVLQRVLQLPVVSGTCECFQKTYNSTKEAHPLVASVCNAY EKGVQGASNLAAWSMETVAT RLSTQFTAA. NTJACRGLDHLEEKIPALQYPPEKIASELK GTISTRLRSARNSISVPIASTSDKVTGATLAGCELALGMAKETAEYAANTRVGRLASGG ADLALGSIEKWTTLLPPDKESAPSSGRQRTQKAPKAKPSLVRRVSTLANTLSRHTMQTT AWALKQGHSLAMWIPGVAPLSSLAQWGASAAMQVVSRRQSEVRVPWLHNLAASQDE SHDDQTDTEGEETDDEEEEEESEAEENVI. REVTALPNPRGLLGGVVHWQNTI. RNTISA VTWAPAAVLGTVGRILHLTPAQAVSSTKGRAMSLSDALKGVTDNVVDTVVHYVPLPRL SLMEPESEFRDIDNPSAEAERKGSGARPASPESTPRPGQPRGSLRSVRGLSAPSCPGLDDK TEASARPGFLAMPREKPARRVSDSFFRPSVMEPILGRAQYSQLRKKS (SEQ ID NO:61).

[0036] As used herein, the term "nucleic acid" and "polynucleotide" are used interchangeably and refer to a polymer of nucleotides, which can include deoxyribonucleic acids (DNA), ribonucleic acids (RNA), or any combination and polymers thereof in either single- or double¬ stranded form. The term encompasses nucleic acids containing modified nucleotides.

[0037] The terms “polypeptide” and “peptide” are used interchangeably herein to refer to a polymer of amino acid residues m a single chain, wherein each ammo acid residue is linked to another by a peptide bond unless otherwise specified. The terms apply to amino acid polymers in which one or more amino acid residue is an artificial chemical mimetic of a corresponding naturally occurring ammo acid, as well as to naturally occurring ammo acid polymers and non- naturally occurring amino acid polymers. Ammo acid polymers may comprise entirely L-amino acids, entirely D-amino acids, or a mixture of L- and D-amino acids. The term “protein” as used herein refers to either a polypeptide or a dimer (i.e., two) or multimer (i.e., three or more) of11US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC single chain polypeptides. The single chain polypeptides of a protein may be joined by a covalent bond, e.g., a disulfide bond, or non-covalent interactions. In embodiments, a polypeptide as described herein can be at least 80, 85, 90, 95, 98%, or 99% identical to a reference polypeptide (e.g., any polypeptide sequence described herein).

[0038] A “domain” of a protein or a polypeptide refers to a region of the protein or polypeptide defined by structural and / or functional properties.

[0039] A “binding site” of a protein or a polypeptide refers to a location of a protein or a polypeptide molecule at which another molecule (which can be referred to as a “target,” a “ligand,” or a “binding partner”) can bind with specificity. In proteins and polypeptides, binding sites are formed in tertiary structure, with some residues actually participating in binding the ligand and other residues acting as a framework to provide correct conformation and orientation.

[0040] The term “amino acid” refers to any monomeric unit that can be incorporated into a peptide, polypeptide, or protein. Amino acids include naturally occurring a-amino acids and their stereoisomers, as well as unnatural (non-naturally occurring) amino acids and their stereoisomers. “Stereoisomers” of a given ammo acid refer to isomers having the same molecular formula and intramolecular bonds but different three-dimensional arrangements of bonds and atoms (e.g., an L-amino acid and the corresponding D-amino acid).

[0041] Naturally occurring amino acids are those encoded by the genetic code, as well as those ammo acids that are later modified, e.g., hydroxyproline, y-carboxyglutamate, and O-phosphoserine. Naturally occurring a-amino acids include, without limitation, alanine (Ala), cysteine (Cys), aspartic acid (Asp), glutamic acid (Glu), phenylalanine (Phe), glycine (Gly), histidine (His), isoleucine (Ile), arginine (Arg), lysine (Lys), leucine (Leu), methionine (Met), asparagine (Asn), proline (Pro), glutamine (Gin), serine (Ser), threonine (Thr), valine (Vai), tryptophan (Trp), tyrosine (Tyr), and their combinations. Stereoisomers of a naturally occurring a-amino acids include, without limitation, D-alanine (D-Ala), D-cysteine (D-Cys), D-aspartic acid (D-Asp), D-glutamic acid (D-Glu), D-phenylalanine (D-Phe), D-histidine (D-His), D-isoleucine (D-Ile), D-arginine (D-Arg), D-lysine (D-Lys), D-leucine (D-Leu), D-methionine (D-Met), D-asparagine (D-Asn), D-proline (D-Pro), D-glutamine (D-Gln), D-serine (D-Ser), D-threonine (D-Thr), D-valine (D-Val), D-tryptophan (D-Trp), D-tyrosine (D-Tyr), and their combinations.12US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0042] Unnatural (non-naturally occurring) amino acids include, without limitation, amino acid analogs, amino acid mimetics, synthetic amino acids, N-substituted glycines, and N-methyl amino acids in either the L- or D-configuration that function in a manner similar to the naturally occurring amino acids. For example, “amino acid analogs” can be unnatural ammo acids that have the same basic chemical structure as naturally occurring amino acids (z.e., a carbon that is bonded to a hydrogen, a carboxyl group, an amino group) but have modified side-chain groups or modified peptide backbones, e.g., homoserine, norleucine, methionine sulfoxide, methionine methyl sulfonium. “Amino acid mimetics” refer to chemical compounds that have a structure that is different from the general chemical structure of an amino acid, but that functions in a manner similar to a naturally occurring amino acid. Amino acids may be referred to by either the commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission,

[0043] The expression “conservatively modified variant” and related expression may apply to amino acid sequences, as well to nucleic acid sequences encoding ammo acid sequence.Substitutions, deletions or additions to a nucleic acid, peptide, polypeptide, or protein sequence which alters, adds or deletes a single ammo acid or a small percentage of ammo acids in the encoded sequence is a “conservatively modified variant” where the alteration results in the substitution of an ammo acid with a chemically similar amino acid. Conservative substitution tables providing functionally similar ammo acids are well known in the art. Such conservatively modified variants are in addition to and do not exclude polymorphic variants, interspecies homologs, and alleles of the invention. The following eight groups each contain amino acids that are conservative substitutions for one another:1) Alanine (A), Glycine (G);2) Aspartic acid (D), Glutamic acid (E);3) Asparagine (N), Glutamine (Q);4) Arginine (R), Lysine (K);5) Isoleucine (I), Leucine (L), Methionine (M), Valine (V);6) Phenylalanine (F), Tyrosine (Y), Tryptophan (W);7) Serine (S), Threonine (T); and8) Cysteine (C), Methionine (M).13US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0044] The terms “identity,” “substantial identity,” “similarity,” “substantial similarity,” “homology” and the related terms and expressions used in the context of describing nucleic acid or ammo acid sequences refer to a sequence that has at least 60% sequence identity to a reference sequence. Examples include at least: 60%, 65%, 70%, 75%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%, sequence identity, as compared to a reference sequence using the programs for comparison of nucleic acid or amino acid sequences, such as BLAST using standard parameters. For sequence comparison, typically one sequence acts as a reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. Default (standard) program parameters can be used, or alternative parameters can be designated. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters. A “comparison window” includes reference to a segment of any one of the number of contiguous positions (from 20 to 600, usually about 50 to about 200, more commonly about 100 to about 150), in which a sequence may be compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. Methods of alignment of sequences for comparison are well-known. Optimal alignment of sequences for comparison may be conducted, for example, by the local homology algorithm of Smith and Waterman (Smith and Waterman “Identification of common molecular subsequences.” J Mol Biol. 147(1): 195-7 (1981)) by the homology alignment algorithm of Needleman and Wunsch (Needleman and Wunsch “A general method applicable to the search for similarities in the amino acid sequence of two proteins.” J Mol Biol. 48(3):443-53 (1970)), by the search for similarity method of Pearson and Lipman (Pearson and Lipman “Improved tools for biological sequence comparison.” Proc. Natl. Acad. Sci. USA 85(8):2444-8 (1988)), by computerized implementations of these algorithms (for example, BLAST), or by manual alignment and visual inspection.

[0045] Algorithms that are suitable for determining percent sequence identity and sequence similarity include the BLAST 2.0 algorithm, which is described in Altschul et al. (1990) J. Mol. Biol. 215: 403-10. For sequence comparison, typically one sequence acts as a reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are entered into a computer, subsequence coordinates are designated, if14US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC necessary, and sequence algorithm program parameters are designated. Default program parameters can be used, or alternative parameters can be designated. The sequence comparison algorithm then calculates the percent sequence identities for the test sequences relative to the reference sequence, based on the program parameters.

[0046] A “comparison window,” as used herein, includes reference to a segment of any one of the number of contiguous positions selected from the group consisting of from 20 to 600, usually about 50 to about 200, more usually about 100 to about 150 in which a sequence may be compared to a reference sequence of the same number of contiguous positions after the two sequences are optimally aligned. Methods of alignment of sequences for comparison are well-known in the art. Optimal alignment of sequences for comparison may be conducted by the local homology algorithm of Smith & Waterman Add. APL. Math. 2:482 (1981), by the homology alignment algorithm of Needleman & Wunsch J. Mol. Biol. 48:443 (1970), by the search for similarity method of Pearson & Lipman Proc. Natl. Acad. Sci. (U. S. A.) 85: 2444 (1988), by computerized implementations of these algorithms (e.g., BLAST), or by manual alignment and visual inspection.

[0047] As used herein, the term “heterologous” refers to a molecule (e.g., peptide or protein) that is not normally or naturally produced or expressed by a cell or organism, or that is fused to a second sequence to which it is not fused in a natural cell or organism.

[0048] “Endogenous” as used herein refers to a molecule normally or naturally produced by a cell or organism. An endogenous TCR is encoded by a naturally occurring T-cell whereas a heterologous TCR is introduced by a human into the T-cell, for example recombmantly.BRIEF DESCRIPTION OF THE. DRAWINGS

[0049] FIGs. 1A-1C depict results of epitope mapping of TCR 1309. FIG. 1A sets forth the epitope sequence of TCR 1309. The core epitope is shaded in gray (AMQVVSRR, SEQ ID NO: 57). FIG. IB is a chart depicting an alanine scan. The stimulation index on the y-axis is shown for each clone listed on the x-axis, where an alanine was substituted for a non-alanine residue at the specified position. FIG. 1C is a chart depicting a truncation scan. For each clone on the x axis, there were two ammo acids deleted at a time from either the C-terminus (C2, C4, C8, C10) or N-terminus (N2, N4, N6, N8, N10) of the peptides. The modified peptides were expressed in15US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC the presence of C3g7 antigen-presenting cells and TCR 1309-expressing hybridomas and the upregulation of NFAT-GFP was assessed.

[0050] FIGs.2A-2C depict results of epitope mapping of TCR 2991. FIG.2A sets forth the epitope sequence of TCR 2991. The core epitope is shaded in gray (SAAMQVVSRRQS; SEQ ID NO: 58). FIG.2B is a chart depicting an alanine scan. The %NFAT-GFP on the y-axis is shown for each clone listed on the x-axis, where an alanine was substituted for a non-alanine residue at the specified position. The modified peptides were expressed in the presence of C3g7 antigen-presenting cells and TCR 2991 -expressing hybridomas and the upregulation of NFAT- GFP was assessed. FIG.2C is a chart depicting a truncation scan. For each clone on the x axis, there were two ammo acids deleted at a time from either the C -terminus (C2, C4, C8, CIO) or N-terminus (N2, N4, N6, N8, N10) of the peptides. The modified peptides were expressed in the presence of C3g7 antigen-presenting cells and TCR 2991 -expressing hybridomas and the upregulation of NFAT-GFP was assessed,

[0051] FIGs.3A-3B depict results of epitope mapping of TCR 2886. FIG.3A sets forth the epitope sequence of TCR 2886. The core epitope is shaded in gray (WLHNLAASQDE; SEQ ID NO: 59). FIG. 3B is a chart depicting an alanine scan. The %NFAT-GFP on the y-axis is shown for each clone listed on the x-axis, where an alanine was substituted for a non-alanine residue at the specified position. The modified peptides were expressed in the presence of C3g7 antigen-presenting cells and TCR 2886-expressing hybridomas and the upregulation of NFAT-GFP was assessed. FIG.3C is a chart depicting a truncation scan. For each clone on the x axis, there were two amino acids deleted at a time from either the C-terminus (C2, C4, C8, CIO) or N-terminus (N2, N4, N6, N8, N10) of the peptides. The modified peptides were expressed in the presence of C3g7 antigen-presenting cells and TCR 2886-expressing hybridomas and the upregulation of NFAT-GFP was assessed.DETAILED DESCRIPTION

[0052] Acquired generalized lipodystrophy (AGE) is a disorder characterized by nearly complete loss of subcutaneous adipose tissue and is thought, at least in part, to be driven by autoimmune-mediated destruction of adipocytes. A challenge in the field has been defining the specificity of the autoimmune response against the fat m this disease. The inventors have16US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC discovered that in both patients and a mouse model with lipodystrophy, a large fraction of the autoimmune response is against a lipid droplet protein, perilipin 1 (PLIN1). In many autoimmune diseases, autoantibodies help guide investigators on to the specificity not only of the B cell response, but also the T cell response. In fact, most autoimmune diseases are driven by an autoreactive T cell response, but due to the nature of how T cells recognize peptide antigens in the context of polymorphic MHC (HLA), unraveling their specificity in isolation is extremely challenging. Having determined that lipodystrophy is linked to PLIN1 autoantibodies, the inventors have identified clonal expansion of autoreactive T cells through single cell RNA-sequencing. Notably, multiple individual T cell clones identified through this approach were indeed specific for peptides derived from PLIN1.

[0053] In view of the discoveries above, the disclosure provides for cells, e.g., T-cells, that have been modified to express a T-cell receptor (TCR) or chimeric antigen receptor (CAR) that binds to PLIN1. As described in detail herein, the cells can be formed in vitro, ex vivo or in vivo and can be used to reduce (e.g., ablate) or regulate (e.g., increase fat by down regulating immune responses that harm fat cells) fat. in a subject depending on whether the T-cells are for example, cytotoxic (CD8+) T-cells or regulatory (FoxP3+ and optionally CD4+, CD25+) T-cells, respectively.

[0054] In some embodiments, cells that comprise or express a heterologous T-cell receptor (TCR) that binds to an MHC perilipin 1 (PLIN1) peptide presented by (e g., on the surface of a cell, complexed with) a class I or class II MHC molecule are provided. As described herein, cells expressing a heterologous TCR that binds to the PLIN 1 peptide presented by the MHC molecule have many uses including targeting the TCR-expressing cells to fat cells, where the targeted cell can then kill or regulate the targeted fat cell depending on the cell targeted to the fat cell.

[0055] Exemplary TCRs that bind to a PLIN1 peptide as presented by an MHC or HLA molecule are described herein. The MHC molecule can be a class I or class II molecule. The data provided herein relates to class II MHC molecules, and thus the PLINl peptides in this context are class II MHC peptides. Class II MHC peptides as described in for example, Roche, et al., Nature Reviews Immunology volume 15, pages 203-216 (2015). Class II MHC peptides are characterized as having a length of 12-25. See, e.g., Gfeller et al., Seminars in Immunology’,17US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC Vol 66, 101708 (2023). In other embodiments, the MHC molecules is a class I MHC molecule and in these embodiments, the PLIN1 peptide is a MHC class I peptide. MHC class I peptides are characterized in general, but without limitation, as having a length of 8-14.

[0056] TCRs are formed from an alpha chain polypeptide and a beta chain polypeptide, which in some embodiments are separate polypeptides and in some embodiments are provided as a single chain polypeptide, optionally linked by a linker amino acid sequence. Binding specificity of a TCR is in general mediated by six variable loops referred to a complementarity determining regions (CDRs), wherein the alpha chain comprises CDRcxl, CDRa2 and CDRa3 and the beta chain comprises CDRβ1, CDRβ2 and CDRβ3. In general, for each of the alpha and beta chains, CDRls and CDR2s bind to the HLA conserved alpha-helices on antigen presenting cells (APCs) whereas the CDR3s plays the most significant role in binding to the peptide antigen presented by the APCs. See, e.g., Wong et al., Frontiers in Immunol. Vol. 10: pp. 1-11, 15 Oct. 2019. The CDRs are contained within a variable region, e.g., the variable alpha (Va) region of the alpha chain comprises CDRotl, CDRa.2 and CDRoc3 whereas the variable beta region ( Vb) of the beta chain comprises CDRβ1, CDRβ2 and CDRβ3.

[0057] Unless otherwise stated, the term “TCR” should be understood to encompass full TCRs as well as antigen- binding portions or fragments thereof. In some embodiments, the TCR is an intact or full-length TCR, such as a TCR containing the a chain and P chain. In some embodiments, the TCR is an antigen-binding portion that is less than a full-length TCR but that binds to a specific peptide bound in an MHC molecule, such as binds to an MHC-peptide complex. In some cases, an antigen-binding portion or fragment of a TCR can contain only a portion of the structural domains of a full-length or intact TCR, but yet is able to bind the peptide epitope, such as MHC-peptide complex, to which the full TCR binds. In some cases, an antigen¬ binding portion contains the variable domains of a TCR, such as variable a (Va) chain and variable β (Vβ) chain of a TCR, or antigen-binding fragments thereof sufficient to form a binding site for binding to a specific MHC-peptide complex.

[0058] In some embodiments, the variable domains of the TCR contain complementarity determining regions (CDRs), which generally are the primary contributors to antigen recognition and binding capabilities and specificity of the peptide, MHC and / or MHC-peptide complex. In some embodiments, a CDR of a TCR or combination thereof forms all or substantially all of the 18US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC antigen-binding site of a given TCR molecule. The various CDRs within a variable region of a TCR chain generally are separated by framework regions (FRs), which generally display less variability among TCR molecules as compared to the CDRs (see, e.g., Jores et al., Proc. Nat'l Acad. Sci. U. S. A. 87:9138, 1990; Chothia et al., EMBO J. 7:3745, 1988; see also Lefranc et al., Dev. Comp. Immunol. 27:55, 2003). In some embodiments, CDR3 is the main CDR responsible for antigen binding or specificity, or is the most important among the three CDRs on a given TCR variable region for antigen recognition, and / or for interaction with the processed peptide portion of the peptide-MHC compl ex. In some contexts, the CDR1 of the alpha chain can interact with the N-termmal part of certain antigenic peptides. In some contexts, CDR1 of the beta chain can interact with the C-terminal part of the peptide. In some contexts, CDR2 contributes most strongly to or is the primary CDR responsible for the interaction with or recognition of the MHC portion of the MHC-peptide complex. In some embodiments, the variable region of the p-chain can contain a further hypervariable region (CDR4 or HVR4), which generally is involved in superantigen binding and not antigen recognition (Kotb (1995) Clinical Microbiology Reviews, 8:411-426).

[0059] In some embodiments, the α-chain and / or β-chain of a TCR also can contain a constant domain, a transmembrane domain and / or a short cytoplasmic tail (see, e.g., Janeway et al., Immunobiology: The Immune System in Health and Disease, 3rdEd., Current Biology Publications, p. 4:33, 1997). In some aspects, each chain (e.g. alpha or beta) of the TCR can possess one N-tenninal immunoglobulin variable domain, one immunoglobulin constant domain, a transmembrane region, and a short cytoplasmic tail at the C-terminal end. In some embodiments, a TCR, for example via the cytoplasmic tail, is associated with invariant proteins of the CD 3 complex involved in mediating signal transduction. In some cases, the structure allows the TCR to associate with other molecules like CD3 and subunits thereof. For example, a TCR containing constant domains with a transmembrane region may anchor the protein in the cell membrane and associate with invariant subunits of the CD3 signaling apparatus or complex. The intracellular tails of CD3 signaling subunits (e.g. CD3y, CD35, CD3E and CD3^ chains) contain one or more immunoreceptor tyrosine- based activation motif or ITAM and generally are involved in the signaling capacity of the TCR complex.19US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0060] One of ordinary skill in the art can determine or identify the various domains or regions of a TCR. In some cases, the exact locus of a domain or region can vary depending on the particular structural or homology modeling or other features used to describe a particular domain. It is understood that reference to amino acids, including to a specific sequence set forth as a SEQ ID NO used to describe domain organization of a TCR are for illustrative purposes and are not meant to limit the scope of the embodiments provided. In some cases, the specific domain (e.g. variable or constant) can be several amino acids (such as one, two, three or four) longer or shorter. In some aspects, residues of a TCR are known or can be identified according to the International Immunogenetics Information System (IMGT) numbering system (see e g. www.imgt.org; see also, Lefranc et al, (2003) Developmental and Comparative Immunology, 27(1); 55-77; and The T Cell Factsbook 2nd Edition, Lefranc and LeFranc Academic Press 2001), In some embodiments, the Cell Ranger available from 10X Genomic Inc., can be used to identify TCR clonotypes and for example CDR3.

[0061] In some embodiments, the a chain and P chain of a TCR each further contain a constant domain. In some embodiments, the a chain constant domain (Ca) and p chain constant domain (CP) individually are mammalian, such as is a human or murine constant domain. In some embodiments, the constant domain is adjacent to the cell membrane. For example, in some cases, the extracellular portion of the TCR formed by the two chains contains two membrane-proximal constant domains, and two membrane-distal variable domains, which variable domains each contain CDRs

[0062] Exemplary TCR alpha and beta chain CDRs are provided but are not intended to limit the scope of the full invention described herein. In some embodiments, the TCRs described herein bind to a PLIN1 peptide presented on an MHC molecule. In some embodiments, the MHC is an I ILA molecule. In some embodiments, the PLIN 1 peptide is an MHC class I or class II peptide and comprises QWGASAAMQVVSRRQSEVRV (SEQ ID NO:55), PWLHNLAASQDESHDDQTDT (SEQ ID NO:56), AMQVVSRR (SEQ ID NO:57), SAAMQVVSRRQS (SEQ ID NO:58) or WLHNLAASQDE (SEQ ID NO:59) or YNSTKEAHPLVASVCNAYEK (SEQ ID NO: 78) or SSGRQRTQKAPKAKPSLVRR (SEQ ID NO:79) or a contiguous fragment thereof at least 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 1920US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC or 20 amino acids and optionally having one or two amino acid substitutions, optionally where the underlines amino acids are not substituted.

[0063] For any alpha or beta variable regions, the framework sequences, e.g., the sequences flanking the CDRs, or even the CDRs themselves, can be humanized, e.g., such that the resulting sequence is less immunogenic in humans. In some embodiments, the alpha and beta variable regions are linked to a human alpha and beta constant regions, respectively.

[0064] As shown in the examples, a number of TCR sequences were isolated from various TCR clones and in general a high degree of sequence identity was observed between CDRα3 sequences as well as between CDRβ3 sequences.

[0065] In some embodiments, the TCR alpha chain comprises a CDRα3 of CAMRGLQGGSAKLIF (SEQ ID NO: 4), optionally with one or two ammo acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSGQGGSDYTF (SEQ ID NO: 8), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRα3 of CAMRGLQGGSAKLIF (SEQ ID NO:4) and a TCR beta chain that comprises a CDRβ3 of CASSGQGGSDYTF (SEQ ID NO: 8), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of TRDSSYF (SEQ ID NO:2) and a CDRα2 sequence of QDSYKKEN (SEQ ID NO: 3) and the TCR beta chain further comprises a CDRβ1 sequence of NNHDY (SEQ ID NO: 6) and a CDRβ2 sequence of SYVADS (SEQ ID NO: 7). In some embodiments, the variable alpha region comprises MLILSLLGAAFGSICFATSMAQKVTQTQTSISVVEKTTVTMDCVYETRDSSYFLFWYKQ TASGEIVFLIRQDSYKKENATVGHYSLNFQKPKSSIGLIITATQIEDSAVYFCAMRGLQGG SAKLIFGEGTKLTVSS (SEQ ID NO: 1) and / or the variable beta region comprises MGSRLFFVVLILLCAKHMEAAVTQSPRSKVAVTGGKVTLSCHQTNNHDYMYWYRQDT GHGLRLIHYSYVADSTEKGDIPDGYKASRPSQENFSLILELASLS QTAVYFCASSGQGGS DYTFGSGTRLLVI (SEQ ID NO: 5).

[0066] In some embodiments, the TCR alpha chain comprises a CDRo.3 of CAMRGNQGGSAKLIF (SEQ ID NO: 12), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRP3 of CASSDQEGSDYTF (SEQ ID NO: 16),21US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRα3 of CAMRGNQGGSAKLIF (SEQ ID NO: 12) and a TCR beta chain that comprises a CDRβ3 of CASSDQEGSDYTF (SEQ ID NO: 16), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of TRDSSYF(SEQ ID NO: 10) and a CDRα2 sequence of QDSYKKEN (SEQ ID NO: 11) and the TCR beta chain further comprises a CDRβ1 sequence of NSHNY (SEQ ID NO: 14) and a CDRβ2 sequence of SYGAGN (SEQ ID NO: 15). In some embodiments, the variable alpha region comprises MLILSLLGAAFGSICFATSMAQKVTQTQTSISVVEKTTVTMDCVYETRDSSYFLFWYKQ TASGEIVFLIRQDSYKKENATVGHYSLNFQKPKSSIGLIIT ATQIEDSAVYFCAMRGNQGG SAKLIFGEGTKLTVSS (SEQ ID NO: 9) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto and / or the variable beta region comprises MGSRLFLVLSLLCTKHMEAAVTQSPRNKVTVTGGNVTLSCRQTNSHNYMYWYRQDTG HGLRLIHYSYGAGNLQIGDVPDGYKATRTTQEDFFLLLELASPSQTSLYFCASSDQEGSD YTFGSGTRLLVI (SEQ ID NO: 13) or an ammo acid sequence at least 90, 95, 98, or 99% identical thereto.

[0067] In some embodiments, the TCR alpha chain comprises a CDRα3 of CAMRGIQGGSAKLIF (SEQ ID NO: 20), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSEGTGGSYEQYF (SEQ ID NO:24), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRα3 of CAMRGIQGGSAKLIF (SEQ ID NO:20) and a TCR beta chain that comprises a CDRβ3 of CASSEGTGGSYEQYF (SEQ ID NO:24), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of TRDSSYF (SEQ ID NO: 18) and a CDRα2 sequence of QDSYKKEN (SEQ ID NO: 19) and the TCR beta chain further comprises a CDRβ1 sequence of NSHNY (SEQ ID NO:22) and a CDRβ2 sequence of SYGAGN (SEQ ID NO:23). In some embodiments, the variable alpha region comprisesMLILSLLGAAFGSICFATSMAQKVTQTQTSISVVEKTTVTMDCVYETRDSSYFLFWYKQ TASGEIVFLIRQDSYKKENATVGHYSLNFQKPKSSIGLIIT ATQIEDSAVYFCAMRGIQGG SAKLIFGEGTKLTVSS (SEQ ID NO: 17) or an amino acid sequence at least 90, 95, 98, or 99%22US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC identical thereto and / or the variable beta region comprises MGSRLFLVTSLLCTKHMEAAVTQSPRNKVTVTGGNy^TLSCRQTNSHNYMYWT’RQDTG HGLRLIHYSYGAGNLOIGDVPDGYKATRTTQEDFFLLLELASPSOTSLYFCASSEGTGGS YEQ YF GPGTRLTVL (SEQ ID NO:21) or an ammo acid sequence at least 90, 95, 98, or 99% identical thereto.

[0068] In some embodiments, the TCR alpha chain comprises a CDRα3 of CAFNYNQGKLIF (SEQ ID NO:28), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASRVGLGGDAETLYF (SEQ ID NO: 32), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRα3 of CAFNYNQGKLIF (SEQ ID NO:28) and a TCR beta chain that comprises a CDRβ3 of CASRVGLGGDAETLYF (SEQ ID NO: 32), optionally either or both comprising one or two ammo acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of ASGYPA (SEQ ID NO:26) and a CDRα2 sequence of ASRDKEK (SEQ ID NO:27) and the TCR beta chain further comprises a CDRβ1 sequence of FNHDT (SEQ ID NO:30) and a CDRβ2 sequence of SITEND (SEQ ID NO:31 ). In some embodiments, the variable alpha region comprises MNSSPGFMTVMLLIFTRAHGDSVTQTEGQVALSEEDFLTIHCNYSASGYPALFWYVQYP GEGPQFLFRASRDKEKGSSRGFEATYDKGTTSFHLRKASVQESDSAVYYCAFNYNQGK LIFGQGTKLSIKP (SEQ ID NO:25) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto and / or the variable beta region comprises MNKWVFCWVTLCLLTVETTHGDGGIITQTPKFLIGQEGQKLTLKCQQNFNHDTMYWY RODSGKGLRLIYYSITENDLOKGDLSEGYDASREKKSSFSLTVTSAOKNEMAVFLCASR VGLGGDAETLYFGSGTRLTVL (SEQ ID NO:29) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto.

[0069] A consensus of the four CDRo.3 and CDRP3 sequences provided directly above are: CDRα3: CAMRGXQGGSAKLIF (SEQ ID NO: 53), where X is any amino acid, optionally an amino acid at the same position from one of the four CDRα3 sequences above, i.e., CAMRGLQGGSAKLIF (SEQ ID NO:4), CAMRGNQGGSAKLIF ( SEQ ID NO: 12), CAMRGIQGGSAKLIF (SEQ ID NO:20), or CAFNYNQGKLIF (SEQ ID NO:28); and CDRβ3 CASXXXXGXXX (SEQ ID NO: 54), where X is any amino acid, optionally an amino acid at the 23US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC same position from one of the four CDRβ3 sequences above, i.e., CASSGQGGSDYTF (SEQ ID NO: 8), CASSDQEGSDYTF (SEQ ID NO: 16), CASSEGTGGSYEQYF (SEQ ID NO:24) or CASRVGLGGDAETLYF (SEQ ID NO: 32).

[0070] Accordingly in some embodiments the TCR alpha variable region comprises a CDRα3 comprising CAMRGXQGGSAKLIF (SEQ ID NO: 53) and the TCR beta variable region comprises a CDRβ3 comprising CASXXXXGXXX (SEQ ID NO: 54).

[0071] In some embodiments, the TCR alpha chain comprises a CDRα3 of CATDWGGSNAKLTF (SEQ ID NO: 36), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSIDRGDTGQLYF (SEQ ID NO:40), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRα3 of CATDWGGSNAKLTF (SEQ ID NO:36)and a TCR beta chain that comprises a CDRβ3 of CASSIDRGDTGQLYF (SEQ ID NO:40), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of TSITA (SEQ ID NO:34) and a CDRα2 sequence of IRSNERE (SEQ ID NO:35) and the TCR beta chain further comprises a CDRβ1 sequence of FNHDT (SEQ ID NO:38) and a CDRβ2 sequence of SITEND (SEQ ID NO: 39). In some embodiments, the variable alpha region comprisesMHSLLGLLMVSLWLQLTRVNSQLAEENLWALSVHEGESVTVNCSYKTSITALQWYRQ KSGEGPAQLILIRSNEREKRNGRLRATLDTSSQSSSLSITATRCEDTAVYFCATDWGGSN AKLTFGKGTKLSVKS (SEQ ID NO:33) or an ammo acid sequence at least 90, 95, 98, or 99% identical thereto and / or the variable beta region comprises MNKWVFCWVTLCLLTVETTHGDGGIITQTPKFLIGQEGQKLTLKCQQNFNHDTMYWY RQDSGKGLRLIYYSITENDLQKGDLSEGYDASREKKSSFSLTVTSAQKNEMAVFLCASSI DRGDTGQLYFGEGSKLTVL (SEQ ID NO: 37) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto.

[0072] In some embodiments, the TCR alpha chain comprises a CDRa3 of CATDYGGSGNKLIF (SEQ ID NO:44), optionally with one or two ammo acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSLDRGDTGQLYF (SEQ ID NO:48), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR24US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC alpha chain that comprises a CDRa3 of CATDYGGSGNKLIF (SEQ ID NO:44) and a TCR beta chain that comprises a CDRP3 of CASSLDRGDTGQLYF (SEQ ID NO:48), optionally either or both comprising one or two ammo acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of TSITA (SEQ ID NO:42) and a CDRα2 sequence of IRSNERE (SEQ ID NO:43) and the TCR beta chain further comprises a CDRβ1 sequence of KGHPV (SEQ ID NO:46) and a CDRβ2 sequence of FQNQEV (SEQ ID NO:47). In some embodiments, the variable alpha region comprises MHSI GI ] [VSLWI> QLTRVNSQI EENE> WALSXTIEGESVTVNCSYKTSITALQWYRQ KSGEGPAQLILIRSNEREKRNGRLRATLDTSSQSSSLSITATRCEDTAVYFCATDYGGSGN KLIF GIGTLLS VKP (SEQ ID NO:41) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto and / or the variable beta region comprises MATRLLCYTVLCLLGARILNSKVTQTPRYLVKGQGQKAKMRCIPEKGHPVFWYQQNK NNEFKFLINFQNQEVLQQIDMTEKRFSAECPSNSPC SLEIQSSEAGDSALYLCASSLDRGD TGQLYFGEGSKLTVL (SEQ ID NO:45) or an ammo acid sequence at least 90, 95, 98, or 99% identical thereto,

[0073] A consensus of the two CDRo.3 and CDRp3 sequences provided directly above are: CDRα3: CATDXGGSXXKLXF (SEQ ID NO:51), where X is any amino acid, optionally an amino acid at the same position from one of the two CDRα3 sequences above, i.e., CATDWGGSNAKLTF (SEQ ID NO: 36) or CATDYGGSGNKLIF (SEQ ID NO: 44); and CDRβ3 CASSXDRGDQLYF (SEQ ID NO: 52), where X is any amino acid, optionally an amino acid at the same position from one of the two CDRβ3 sequences above, i.e,, CASSIDRGDTGQLYF (SEQ ID NO:40) or CASSLDRGDTGQLYF (SEQ ID NO:48).

[0074] Accordingly, in some embodiments, the TCR alpha variable region comprises a CDRα3 comprising CATDXGGSXXKLXF (SEQ ID NO:51) and the TCR beta variable region comprises a CDRβ3 comprising CASSXDRGDQLYF (SEQ ID NO: 52).

[0075] In some embodiments, the TCR alpha chain comprises a CDRα3 of CALRNRGSALGRLHF (SEQ ID NO: 65), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSFRVEQYF (SEQ ID NO: 69), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR25US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC alpha chain that comprises a CDRα3 of CALRNRGSALGRLHF (SEQ ID NO: 65) and a TCR beta chain that comprises a CDRβ3 of CASSFRVEQYF (SEQ ID NO: 69), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of YSATPY (SEQ ID NO: 63) and a CDRα2 sequence of YYSGDPVV (SEQ ID NO: 64) and the TCR beta chain further comprises a CDRβ1 sequence of MSHET (SEQ ID NO: 67) and a CDRβ2 sequence of CASSFRVEQYF (SEQ ID NO: 69). In some embodiments, the variable alpha region comprises MLLALLPAT. GIHFYTJiDAQAQSVTQPDARVTVSEGASLQLRCKYSYS ’YLFWYVQY PROCiL. OEI.. EKYYSGDPVVOCjVNGFEAtEFSKSN;SSFHLRKASVHWSDSAVYF(iAL. RYRCj S ALGRLHF GAGTQLIVIPD (SEQ ID NO: 62) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto and / or the variable beta region comprises MRVRLISAVVLCFLGTGLVDMKVTQMPRYLIKRMGENVLLECGQDMSHETMYWYRQ DPGLGLQLIYISYDVDSNSEGDIPKGYRVSRKKREHFSLILDSAKTNQTSVYFCASSFRVE QYFGPGTRLTVL (SEQ ID NO: 66) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto. In some embodiments, a TCR comprising the above sequences binds to a Class II peptide YNSTKEAHPLVASVCNAYEK (SEQ ID NO:78).

[0076] In some embodiments, the TCR alpha chain comprises a CDRα3 of CAASATGGADRLTF (SEQ ID NO: 73), optionally with one or two amino acid substitutions therein, the TCR beta chain comprises a CDRβ3 of CASSLGGDTQYF (SEQ ID NO: 77), optionally with one or two amino acid substitutions therein, or the TCR comprises both a TCR alpha chain that comprises a CDRoc3 of CAASATGGADRLTF (SEQ ID NO: 73) and a TCR beta chain that comprises a CDRβ3 of CASSLGGDTQYF (SEQ ID NO: 77), optionally either or both comprising one or two amino acid substitutions therein. In some embodiments, the TCR alpha chain further comprises a CDRα1 sequence of ENSAFDY (SEQ ID NO: 71) and a CDRα2 sequence of ILSVSDK (SEQ ID NO: 72) and the TCR beta chain further comprises a CDRβ1 sequence of SGHDT (SEQ ID NO: 75) and a CDRβ2 sequence of FRDEAV (SEQ ID NO: 76). In some embodiments, the variable alpha region comprises MDKILTASFLLLGLHLAGVSGQQEKRDQQQVQSPQSLTAAVEGETAILNCSYENSAFDY FPWYQQFPGEGPALLISILSVSDKKEDGRFTIFFNKREKKLSLHIADSQPGDSATYFCAAS ATGGADRLTFGKGTQLIIQP (SEQ ID NO: 70) or an ammo acid sequence at least 90, 95, 98,26US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC or 99% identical thereto and / or the variable beta region comprises MGTRLLGWAVFCLLDTVLSEAGVTQSPRYAVLQEGQAVSFWCDPISSGHDTLYWYQQP RDQGPQLLVYFRDEAVIDNSQLPSDRFSAVRPKGTNSTLKIQSAKQGDTATYLCASSLG GDTQYFGPGTRLLVLE (SEQ ID NO: 74) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto. In some embodiments, a TCR comprising the above sequences binds to a Class II peptide SSGRQRTQKAPKAKPSLVRR (SEQ ID NO: 79).

[0077] Any of the embodiments described above can be a full-length TCR or a portion thereof comprising the antigen-binding portion of the TCR. In full-length embodiments, the constant region of the TCR can be any TCR constant region known in the art. In some embodiments, the alpha chain constant region comprises IQNPEPAVYQLKDPRSQDSTLCLFTDFDSQINVPKTMESGTFITDKTVLDMKAMDSKSN GAIAWSNQTSFTCQDIFKETNATYPSSDVPCDATLTEKSFETDMNLNFQNLSVMGLRILL LKVAGFNLLMTLRLWS (SEQ ID NO:49) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto and / or the beta constant region comprises EDLRNVTPPKVSLFEPSKAEIANKQKATLVCLARGFFPDHVELSWWVNGKEVHSGVST DPQAYKESNYSYCLSSRLRVSATFWHNPRNHFRCQVQFHGLSEEDKWPEGSPKPVTQNI SAEAWGRADCGITSASYHQGVLSATILYEILLGKATLYAVLVSGLVLMAMVKKKNS(SEQ ID NO: 50) or an amino acid sequence at least 90, 95, 98, or 99% identical thereto.

[0078] As described elsewhere herein, the cell expressing the heterologous TCR, e.g,, any TCR described above or elsewhere herein, can be any cell. Exemplary cells include, e.g., prokaryotic or eukaryotic cells. In some embodiments, the cells are mammalian (e.g., human or non-human) cells, e.g., mammalian immune cells. Exemplary immune cells can include but are not limited to, T-cells, e.g., CD4 or CD8-expressing T-cells, and can include for example, cytotoxic CD8+ T-cells or regulatory (e.g., FOXp3+) T-cells (Tregs), any of which can be human or non-human mammalian cells.

[0079] Also provided are nucleic acids encoding the TCR alpha chains and / or beta chain polypeptides described herein, vectors comprising the nucleic acids and cells comprising the nucleic acids or vectors. Such a vector can be chosen from viral vectors and non-viral vectors, plasmids, cosmids, and artificial chromosomes. By way of example, the vector can be a viral vector, such as a lentiviral vector, adenoviral vector, an AAV vector, or a retroviral vector. Non-27US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC viral vector examples include physical vectors such as electroporation and chemical vectors, such as a lipid nanoparticles. In some embodiments, one or more codon encoding the polypeptides is optimized for expression in a target cell.

[0080] TCR-expressing nucleic acids can be introduced into the cells as desired to express the TCR in the cell. In some embodiments, the expression in the cells is transient, e.g., a DNA sequence encoding the TCR is not integrated into the genome of the cells. In some embodiments, a DNA sequence encoding the TCR is integrated into the genome of the cells. In some embodiments, the cell is an immune cell encoding an endogenous TCR alpha and or beta chain polypeptide and the nucleic acid(s) encoding the heterologous alpha and beta TCR polypeptides is introduced to disrupt the endogenous sequences, such that the cells only or primarily express the heterologous TCR compared to the endogenous TCR. Examples of methods for introducing TCR-encoding nucleic acids can include methods comprising targeted double strand breaks and homologous recombination, e.g., CRISPR-based methods of inserting homology-dependent recombination (HDR) templates into target genomic sites. Examples of such methods can be found in, e.g,, Nyberg et al., Cell 186, 446–460.e19 (2023); Schmidt, et al., Science 2022 02 04; 375(6580); Shy et al., Nat Biotechnol. 2023 04; 41(4):521-531. Methods of introducing TCR-encoding nucleic acids can also include methods comprising transient expression such as, for example, through mRNA vaccine-like approaches.

[0081] In some embodiments, the TCR alpha and / or beta chain polypeptides are encoded on RNA that is delivered to the cells and expressed in the cells. In some embodiments, the RNA contains one or more modified nucleotide that can for example, enhance expression or lengthen the half-life of the molecule in a subject or within a cell. Examples of such modifications can be found in, for example, Emily P. English et al., Sci. Transl. Med. 16, 2084 (2024). In some aspects, the modified nucleotide comprises a sugar modification, a nucleic acid base modification, and / or a phosphate backbone modification. Modifications that are useful for optimizing inhibitory nucleic acids are described, e.g., in Freier & Altmann (1997), Nucl. Acid Res., 25, 4429-4443; Uhlmann (2000), Curr. Opinion in Drug Development, 3(2), 293-213; and Deleavey and Damha (2012), Chemistry and Biology, 19: 937-954, and U. S. Pat. Nos.5,684,143, 5,858,988 and 6,291,438; Filippova et al. (2019), “Guide RNA modification as a way to improve CRISPR / Cas9-based genome- editing systems”, Biochimie., 167:49-60. Exemplary28US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC modifications can include but are not limited to pseudouridine, 1-methylpseudouridine (i.e., N1-methyl pseudouridine), N1-ethyl pseudouridine, 2-thiouridine, 4′-thiouridine, 5-Methyluridine, 1-methyluridine, 2-thio-1-methyl-1-deaza-pseudouridine, 2-thio-1′-methyl-pseudouridine, 2-thio-5-aza-uridine glycosides, 2-thio-dihydropseudouridine, 2-sulfanyl-dihydrouridine, 2-sulfanyl-pseudouridme, 4-methoxy-2-sulfanyl-pseudouridine, 4-methyl oxy-pseudouridine, 4-thio-l-methyl-pseudouridine, 4-thio-pseudouridine, 5-aza-uridine, 5-carboxyhydroxyuridme, 5-hydroxyuridine, dihydropseudouridine, 5-carbamoylmethyluridine, or 5-methoxyuridine.

[0082] In other embodiments, a chimeric antigen receptor (CAR) comprising an extracellular domain that selectively binds to PLIN1 is provided. CARs can comprise, for example (1) an extracellular target-binding domain that binds to PLIN1 or a PLIN1 peptide complexed with an MHC molecule, (2) a transmembrane domain, (3) a hinge domain, and (4) an intracellular signaling domain. The transmembrane domain links the extracellular target-binding domain and the intracellular signaling domain and anchors the CAR to the plasma membrane of the host cell that is modified to express the CAR, e.g., the plasma membrane of a human or mouse immune cell, e g., a cytotoxic or regulatory T-cell. The optional hinge domain links the extracellular targetbinding domain and the transmembrane domain for positioning the extracellular target-binding domain.

[0083] In other embodiments, the extracellular target-binding domain is an antibody.

[0084] The term “antibody” and the related terms refer to an immunoglobulin or its fragment that binds to a particular spatial and polar organization of another molecule. Immunoglobulins include various classes and isotypes, such as IgA, IgD, IgE, IgGl, IgG2a, IgG2b and IgG3, IgG4, IgM, etc. An antibody can be monoclonal or recombinant, and can be prepared by laboratory techniques, such as by preparing continuous hybrid cell lines and collecting the secreted protein, or by cloning and expressing nucleotide sequences or their mutagenized versions coding at least for the ammo acid sequences required for binding. Antibodies as referenced herein may have sequences derived from non-human antibodies, human sequence, chimeric sequences, and wholly synthetic sequences. The term “antibody” encompasses natural, artificially modified, and artificially generated antibody forms, such as humanized, human, single-chain, chimeric, synthetic, recombinant, hybrid, mutated, grafted, and in vitro generated antibodies and their fragments. The term “antibody” also includes composite forms including but29US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC not limited to fusion proteins containing an immunoglobulin moiety. “Antibody” also refers to non-quaternary antibody structures (such as camelids and camelid derivatives) and antigen-binding fragments of antibodies, minibodies, bispecific antibodies, nanobodies (also referred to as VHH fragments), and diabodies. Antibodies and antigen-binding portions thereof include domain-specific antibodies, single domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetrabodies, minibodies, nanobodies (e.g. monovalent nanobodies, bivalent nanobodies, etc., small modular immunopharmaceuticals (SMIPs), and shark variable IgNAR domains. Antibody fragments may include Fab, Fv, F(ab’)2, Fab’, sc-Fv, dsFv, ds-scFv, Fd, dAb, Fc, and the like, A natural antibody digested by papain yields three fragments: two Fab fragments and one Fc fragment. The Fc fragment is dimeric and contains two CH2 and two CH3 heavy chain domains, CH3 domains interact to form a homodimer. Fc domains in antibodies may also be optimized to alter antibody characteristics of interest (e.g,, bioavailabilty, serum half-life). In addition, aggregates, polymers and conjugates of immunoglobulins or their fragments can be used where appropriate. Additional details of antibodies useful in the context of this disclosure are provided below.

[0085] As used herein, the term antibody encompasses, but is not limited to, whole immunoglobulin (i.e., an intact antibody) of any class. natural immunoglobulin G (IgG) antibody molecule is a tetramer that contains two identical light (L) chains and two identical heavy (H) chains. Typically, each light chain is linked to a heavy chain by one covalent disulfide bond, while the number of disulfide linkages varies between the heavy chains of different immunoglobulin isotypes. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has at one end a variable domain (VH) followed by a number of constant domains. Each light chain has a variable domain at one end (VL) and a constant domain at its other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the light chain variable domain is aligned with the variable domain of the heavy chain. Within light and heavy chains, the variable and constant regions are joined by a “J” region of about 12 or more ammo acids, with the heavy chain also including a “D” region of about 10 more amino acids. See generally, Fundamental Immunology’, Paul, W., ed., 3rd ed. Raven Press, NY (1993), SH. 9 (incorporated by reference in its entirety for all purposes). Antibody sequences and structural information is widely available. The light chains of antibodies from any vertebrate species can be assigned to one of two clearly distinct30US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC types, called kappa (K) and lambda (λ), based on the amino acid sequences of their constant domains. Depending on the amino acid sequence of the constant domain of their heavy chains, immunoglobulins can be assigned to different classes. There are five major classes of immunoglobulins: IgA, IgD, IgE, IgG and IgM, and several of these may be further divided into subclasses (isotypes), e.g., IgG-1, IgG-2, IgG-3, and IgG-4; IgA-1 and IgA-2. The heavy chain constant domains that correspond to the different classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively. As used herein, the term antibody also encompasses an antibody fragment, for example, an antigen-binding fragment. Antigen-binding fragments comprise at least one antigen-binding domain, which, in turn, comprises at least one antigenbinding site. One example of an antigen-binding domain is an antigen-binding domain formed by a VH-VL dimer. Antibodies and antigen-binding fragments can be described by the antigen to which they specifically bind.

[0086] In some embodiments, the extracellular target-binding domain is an antigen-binding portion or antigen-binding fragment. The terms “antigen-binding portion” and “antigen-binding fragment” are used interchangeably in the present disclosure and refer to one or more fragments of an antibody that retains the ability to specifically bind to an antigen. The fragments are not necessarily generated from contiguous antibody sequences, but can be engineered proteins containing antibody sequences needed to form an antigen-binding site. Examples of antigenbinding fragments include, but are not limited to, a Fab fragment (a monovalent fragment consisting of the VL, VH, CL, and CH1 domains), F(ab')2 fragment (a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region), a single chain Fv (scFv) fragment (a fusion protein of the VH and VL regions), a disulfide-linked Fv (dsFv), complementarity determining regions (CDRs), VL (light chain variable region), VH (heavy chain variable region), nanobodies, and any combination of those or any other functional portion of an immunoglobulin peptide capable of binding to target antigen.

[0087] Within each light or heavy chain variable region, there are three short segments (averaging 10 amino acids in length) called the complementarity determining regions (“CDRs”). The more highly conserved portions of the variable domains are called the framework (FR). The variable domains of native heavy and light chains each comprise four FR regions, largely adopting a P-sheet configuration, connected by three CDRs, which form loops connecting, and in31US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC some cases forming part of, the (3-sheet structure. The CDRs in each chain are held together in close proximity' by the FR regions and, with the CDRs from the other chain, contribute to the formation of the antigen-binding site of antibodies. The constant domains are not involved directly in binding an antibody to an antigen, but exhibit various effector functions, such as participation of the antibody in antibody-dependent cellular toxicity. Each VH and VL generally comprises three CDRs and four FRs, arranged in the following order (from N-terminus to C-terminus): FR1 - CDR1 - FR2 - CDR2 - FR3 - CDR3 - FR4. The CDRs are involved in antigen binding and confer antigen specificity and binding affinity to the antibody. SeeKabat et al. Sequences of Proteins of Immunological Interest 5th ed., Public Health Service, National Institutes of Health, Bethesda, Maryland (1991). CDR sequences on the heavy chain (VH) may be designated as CDRH1, 2, 3, while CDR sequences on the light chain (VL) may be designated as CDRL1, 2, 3.

[0088] The term “variable region” refers to a domain in an antibody heavy chain or light chain that gives an antibody its specificity for binding to an antigen. Typically, an antibody variable region comprises four conserved “framework” regions interspersed with three hypervariable “complementarity determining regions.”

[0089] The term “complementarity determining region” or “CDR” refers to the three hypervariable regions in each chain that interrupt the four framework regions established by the light and heavy chain variable regions. The CDRs are primarily responsible for binding to an epitope of an antigen. The CDRs of each chain are typically referred to as CDRl, CDR2, and CDR3, numbered sequentially starting from the N-terminus, and are also typically identified by the chain in which the particular CDR is located. Thus, a VH CDR3 is located in the variable domain of the heavy chain of the antibody in which it is found, whereas a VL CDRl is the CDRl from the variable domain of the light chain of the antibody in which it is found.

[0090] As noted, the part of a variable region not contained in the CDRs is called the framework. The “framework regions” of different light or heavy chains are relatively conserved within a species. The framework region of an antibody, that is the combined framework regions of the constituent light and heavy chains, serves to position and align the CDRs in three- dimensional space. Framework sequences can be obtained from public DNA databases or published references that include germline antibody gene sequences. For example, germlme32US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC DNA sequences for human heavy and light chain variable region genes can be found in the “VBASE2” germline variable gene sequence database for human and mouse sequences. The amino acid sequences of the CDRs and framework regions can be determined using various well- known methods, some of which are described elsewhere in the present disclosure.

[0091] The term “monoclonal antibody” refers to antibodies produced by a single clone of cells or a single cell line and consisting of or consisting essentially of antibody molecules that are identical in their primary amino acid sequence. In some embodiments, a monoclonal antibody preparation comprises a population of antibodies that are identical and bind to the same epitope of an antigen, except for mutations that arise during monoclonal antibody production. Unless otherwise specified or clear from context, the term “monoclonal antibody” includes synthetic antibodies and antigen-binding fragments thereof.

[0092] “Fc region,” “Fc domain,” or “Fc sequence” and the related terms and expressions can all refer the “tail” region of an antibody sequence. Fc region contains at least the amino acid sequences of the CH2 and CH 3 domains of an antibody (and, in some antibody isotypes, can also contain CH4 domain sequence).

[0093] An “Fc fragment” contains two heavy chain fragments comprising the CH2 and CH3 domains of an antibody. The two heavy chain fragments are held together by two or more disulfide bonds and by hydrophobic interactions of the CH3 domains. An Fc domain introduced into a fusion or other type of engineered protein may promote dimerization. In some instances, an Fc domain introduced into a fusion or other type of engineered protein may improve its stability in vitro or upon administration in vivo. For examples, engineered Fc polypeptides according to the present disclosure may be incorporated into therapeutic agents to increase their in vivo half-life.

[0094] A “Fab fragment” is comprised of one light chain, and the CHI and variable regions of one heavy chain and can specifically recognize a target epitope, such as an epitope of a spike protein. A Fab domain introduced into a fusion protein according to the present disclosure results in binding of the fusion protein to the target. Such a fusion protein may be referred to as “Fab fusion protein,” “Fab-based fusion protein,” and by other related expressions.33US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC

[0095] A “single-chain variable fragment” or “scFv fragment” is a fusion protein comprising the variable regions of a heavy chain and a light chain from an antibody. The heavy chain and light chain portions may be connected by a linker peptide. An scFv fragment may retain the binding specificity of the antibody from which it is derived. A scFv domain introduced into a fusion protein according to the present disclosure results in binding of the fusion protein to the target. Such a fusion protein may be referred to as “an scFv fusion protein,” “scFv-based fusion protein,” and by other related expressions.

[0096] In some embodiments, the extracellular target-binding domain can be a single chain antibody (scFv) that binds to PLIN1. In some embodiments, the framework regions of the scFv have been humanized to reduce immunogenicity in a human. In some embodiments, the extracellular target-binding domain comprises a T-cell receptor (TCR) as described herein in which case the extracellular domain binds to a PLIN1 peptide (e.g., as described above) in a complex with an MHC In some embodiments, the TCR is a single-chain TCR wherein the alpha chain and the beta chain are a translational fusion, optionally linked by an amino acid linker,

[0097] A CAR construct encoding a CAR may also comprise a sequence that encodes a signal peptide to target the extracellular domain to the cell surface.

[0098] Any transmembrane suitable for use m a CAR construct may be employed. A transmembrane domain incorporated into a CAR construct may be derived either from a natural, synthetic, semi-synthetic, or recombinant source. Such transmembrane domains, include, but are not limited to, all or part of the transmembrane domain of the alpha, beta or zeta chain of the T-cell receptor, CD28, CD27, CD3 epsilon, CD45, CD4, CDS, CDS, CD9, CD16, CD22, CD33, CD37, CD64, CD80, CD86, CD134, CD137, GDI 54. In some embodiments, a transmembrane domain may include at least the transmembrane region(s) of, e.g., KIRDS2, 0X40, CD2, CD27, LFA-1 (CD Ila, CD18), ICOS (CD278), 4-1BB (CD137), GITR, CD40, BAFFR, FIVEM (LIGHTR), SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD160, CD19, IL2R beta, IL2R gamma, IL7R a, ITGA1, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, GDI Id, ITGAE, CD103, ITGAL, GDI la, LFA-1, ITGAM, GDI lb, ITGAX, CD) 1c, ITGB 1, CD29, ITGB2, CD 18, LFA-1, ITGB7, TNFR2, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), CEACAM1, CRT AM, Ly9 (CD229), CD160 (BY55), PSGL1,34US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CD100, (SEMA4D), SLAMF6 (NTB-A, Lyl08), SLAM (SLAMF1, CD150, IPO-3), BLAME, (SLAMF8), SELPLG (CD 162), LTBR, PAG / Cbp, NKG2D, or NKG2C.

[0099] In some embodiments, the CAR may contain one or more hinge domains that link the extracellular target-binding domain and the transmembrane domain for positioning the extracellular target-binding domain. Such a hinge domain may be derived either from a natural, synthetic, semi-synthetic, or recombinant source. The hinge domain can include the amino acid sequence of a naturally occurring immunoglobulin hinge region, e.g., a naturally occurring human immunoglobulin hinge region, or an altered immunoglobulin hinge region. Illustrative hinge domains suitable for use in the CARs described herein include the hinge region derived from the extracel lular regions of type 1 membrane proteins such as CD8 alpha, CD4, CD28, PD1, CD 152, and CD7, which may be wildtype hinge regions from these molecules or may be altered.

[0100] In some embodiments, the hinge domain is based on the hinge region of a human immunoglobulin IgG1 or IgG4. In some embodiments, the hinge region includes the IgGl or IgG4’s CH2 region, which may comprise one or more mutations. In some embodiments, the mutation is L235E, N297Q, or both L235E and N297Q (which is known as the EQ mutation in the IgG4 hinge region).

[0101] A CAR construct of the present disclosure can include one or more intracellular signaling domains, also referred to herein as co-stimulatory domains, activation domains, or cytoplasmic domains that activate or otherwise modulate an immune cell, (e.g., a T lymphocyte). The intracellular signaling domain is generally responsible for activation of at least one of the normal effector functions of the immune cell in which the CAR has been introduced. For example, a “first-generation CAR” generally has a CD3 zeta (CD3ζ) signaling domain.Additional co-stimulatory intracellular domains may also be introduced (e.g., second and third generation CARs) and further domains including homing and suicide domains may be included in CAR constructs.

[0102] In some embodiments, a co-stimulatory domain is used that increases CAR immune T cell cytokine production. In another embodiment, a co-stimulatory domain is used that facilitates immune cell (e.g., T cell) replication. In still another embodiment, a co-stimulatory domain is used that prevents CAR immune cell (e.g., T cell) exhaustion. In another embodiment, a co- 35US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC stimulatory’ domain is used that increases immune cell (e.g., T cell) antitumor activity. In still a further embodiment, a co-stimulatory domain is used that enhances survival of CAR immune cells (e.g., T cells) (e.g, post-infusion into patients).

[0103] Examples of intracellular signaling domains for use in a CAR include the cytoplasmic sequences of the T cell receptor (TCR) and co-receptors that act in concert to initiate signal transduction following antigen receptor engagement, as well as any derivative or variant of these sequences and any recombinant sequence that has the same functional capability’.

[0104] A primary signaling domain regulates primary' activation of the TCR complex either in a stimulatory way, or in an inhibitory’ way. Primary intracellular signaling domains that act in a stimulatory manner may contain signaling motifs which are known as immunoreceptor tyrosine-based activation motifs or IT AMs.

[0105] Examples of ITAM containing primary intracellular signaling domains include those of CD3 zeta, common FcR gamma, Fc gamma Rlla, FcR beta (Fc Epsilon Rib), CD3 gamma, CD3 delta, CDS epsilon, CD79a, CD79b, DAP10, and DAP12. In one embodiment, a CAR comprises an intracellular signaling domain, e.g., a primary' signaling domain of CD3 zeta.

[0106] An intracellular signaling domain of a CAR can comprise a primary intracellular signaling domain only, or may also comprise additional desired intracellular signaling domain(s) useful in the context of a CAR of the disclosure. For example, the intracellular signaling domain of the CAR can comprise a CD3 zeta chain portion and a co-stimulatory signaling domain. The co-stimulatory signaling domain refers to a portion of the CAR comprising the intracellular domain of a co-stimulatory molecule. A co-stimulatory molecule is a cell surface molecule other than an antigen receptor or its ligands that is required for an efficient response of lymphocytes to an antigen. Examples of such molecules include CD27, CD28, 4-1BB (CD137), 0X40, CD30, CD40, PD-1, ICOS, lymphocyte function-associated antigen-1 (LFA-1), CD2, CD7, LIGHT, NKG2C, B7-H3, and a ligand that binds to CD83, and the like. For example, CD27 costimulation has been demonstrated to enhance expansion, effector function, and survival of human CART cells in vitro and augments human T cell persistence and antitumor activity in vivo (Song et al. Blood. 2012; 119(3):696-706). Further examples of such co-stimulatory molecules include CDS, ICAM-1, GITR, BAFFR, HVEM (LIGHTR), SLAMF7, NKp80 (KLRF1), NKp44, NKp30, NKp46, CD 160, CD 19, CD4, CD8alpha, CD8beta, IL2Rbeta, IL2R 36US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC gamma, IL7R alpha, ITGA4, VLA1, CD49a, ITGA4, IA4, CD49D, ITGA6, VLA-6, CD49f, ITGAD, CD1 Id, ITGAE, CD103, ITGAL, CD1 la, LFA-1, ITGAM, CD1 lb, ITGAX, CD11c, ITGB 1, CD29, ITGB2, CD 18, LFA-1, ITGB7, TNFR2, TRANCE / RANKL, DNAM1 (CD226), SLAMF4 (CD244, 2B4), CD84, CD96 (Tactile), NKG2D, CEACAM1, CRTAM, Ly9 (CD229), CD160 (BY55), PSGL1, CD100 (SEMA4D), CD69, SLAMF6 (NTB-A, LylO8), SLAM, (SLAMF1, GDI 50, IPO-3), BLAME (SLAMF8), SELPLG (CD162), LTBR, LAT, GADS, SLP-76, PAG / Cbp, and CD 19a.

[0107] Also provided are nucleic acids encoding the CAR polypeptides described herein, vectors comprising the nucleic acids and cells comprising the nucleic acids or vectors. Vectors as described above in the context of TCR polypeptides can also be used as vectors for CAR nucleic acid-encoding vectors.

[0108] Any method of introducing the heterologous TCR-encoding nucleic acid into a cell can equally be applied to introduce a CAR-encoding nucleic acid into a cell. In some embodiments, the expression in the cells is transient, e.g., a DNA sequence encoding the CAR is not integrated into the genome of the cells. In some embodiments, a DNA sequence encoding the CAR is integrated into the genome of the cells. In some embodiments, the cell is an immune cell. In some embodiments, the CAR polypeptides are encoded on RNA that is delivered to the cells and expressed in the cells. In some embodiments, the RNA contains one or more modified nucleotide that can for example, enhance expression or lengthen the half-life of the molecule in a subject or within a cell, e.g., as described above with regard to TCR delivery.

[0109] In any of the above-described methods, the cells receiving the nucleic acids encoding the alpha and / or beta chain TCR polypeptide(s) or CAR-encoding nucleic acids can be in vitro (e.g., isolated or in culture), ex vivo (isolated from an animal (e.g., a human), optionally primary cells, that will be returned to the same (autologous) or a different (allogeneic) animal (e.g., different human) following introduction of the nucleic acids) or in vivo (for example, wherein the nucleic acid is introduced into an animal (e.g., human) and expressed in a cell (e.g., a T-cell) in the animal.

[0110] Methods of reducing fat or fat cells in a subject are provided. In some embodiments, cytotoxic T-cells targeted to cells expressing PLIN1 are administered to the subject in a sufficient amount to reduce fat in the subject. In other embodiments, a nucleic acid (optionally 37US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC in a vector) encoding a CAR or TCR that binds to PLIN1 or PLIN1 in a complex with an MHC molecule, respectively, is administered to the subject and the nucleic acid enters cytotoxic T-cells in the subject and expresses the CAR or TCR, directing the T-cells to PLIN1 -expressing cells and killing them. In some embodiments, by targeting the cytotoxic T-cells to PLIN1-expressing cells, which are fat cells, fat cells are reduced in the subject.

[0111] In other embodiments, methods of reducing autoimmune targeting of fat cells in a subject are provided. In subjects experiencing autoimmune-mediated destruction of adipocytes, targeting of regulatory T-cells (Tregs) to fat cells can ameliorate over-active immune reactions that harm fat cells. Exemplary disorders that can be ameliorated by the methods described herein include, but are not limited to, acquired generalized lipodystrophy (AGL). In some embodiments, regulatory T-cells targeted to cells expressing PLIN1 are administered to the subject in a sufficient amount to reduce fat cell killing in the subject and optionally thereby allow for an increase in fat in the subject. In other embodiments, a nucleic acid (optionally in a vector) encoding a CAR or TCR that binds to PLIN1 or PLIN1 in a complex with an MHC molecule, respectively, is administered to the subject and the nucleic acid enters regulatory T-cells m the subject and expresses the CAR or TCR, redirecting the regulatory T-cells to PLIN 1 -expressing cells and reducing immune system-based killing of fat cells. In some embodiments, by targeting the regulatory T-cells to PLIN1 -expressing cells, fat cells are increased in the subject.

[0112] The effective amount of the compositions described herein can be determined by one of ordinary skill in the art and will depend in part on the nature of the composition (e.g., cells expressing a CAR or TCR as described herein or nucleic acids encoding the CAR or TCR). An effective amount of any of the compositions described herein will vary and can be determined by one of skill in the art through experimentation and / or clinical trials. In some embodiments, the nucleic acids, vectors comprising the nucleic acids, or cells, are formulated and / or administered with a pharmacally-acceptable carrier or excipient.

[0113] Administration of the nucleic acid, vector or cell compositions described herein can be by any route as desired. In some embodiments, the compositions are administered directly to fat in a subject so as to localize the compositions in fat of the subject. Any suitable route of administration or combination of different routes can be used, including, but not limited to,38US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC parenteral administration (e.g., intravenous, intramuscular, subcutaneous, or intradermal injection, local injection into fat, or oral administration (e.g., in the form of a tablet or capsule).EXAMPLES

[0114] Aire- / - mice were recently shown to develop anti-perilipin 1 antibodies accompanied by immune infiltration of the adipose tissue, making this a model to study immunological determinants of fat-specific T-cell responses (Rackaityte E et al. JCI Insight. 2023 Dec 8;8(23)). ECHO MRI body composition analysis of over 25 Aire- / - mice and controls revealed that both male and female Aire- / - mice had dramatically reduced total fat mass compared to their Aire-sufficient littermates as well as significantly decreased weights of perigonadal and inguinal fat pads, resembling generalized lipodystrophy phenotype in humans.

[0115] Because CD4 T cells are the main drivers of autoimmunity in the APS1 patients and Aire- / - mice, the CD4 T cell compartment and repertoires involved in adipose infiltrates were examined. Flow cytometry on the stromal vascular fraction (SVF) obtained from fat pads digested with collagenase was performed. The Aire- / - mice showed a dramatic increase in the CD45+ cell numbers when normalized for tissue weight. CD4 and CDS cells comprised over 20% of the CD45+ infiltrate each, with majority belonging to the CD44HICD62LLoweffector T cell subsets. These findings show that, the adipose tissue is a target of autoimmune attack in Aire- / - mice,

[0116] To better understand the perturbations in adipose immune populations in Aire- / - mice, 10x single-cell RNA sequencing (scRNA-seq) was performed. The CD45+ cells from pooled fat pads of Aire- / - (n=6)and Aire+ / + (n=7) mice were sorted and over 76,000 cells were obtained. Consistent with the observations from the initial flow' cytometry analysis, Aire- / - mice showed a significant expansion of the T cell cluster (25.8%) compared to controls (5,6%). Analyzing the T-cell compartment was examined next by way of using transcriptional signatures to identify distinct populations. The expansion of several T cell subsets consistent with an ongoing immune response in the Aire- / - mice was observed. The largest increases were seen for the Th 1 -like Tnfsj8+) and memory (Ccr7+) CD4 T cells and exhausted (7c / 7t) and memory (Il7r+, CD28+) CD8 T cell clusters. Lastly, T-cell receptor (TCR) sequencing data was obtained for over 8,000 individual T cells (2264 Aire+ / + and 6055 Aire- / -). Among these, over 20 expanded clonotypes were identified39US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC (constituting >0.25% of total TCRs m our dataset), suggesting an antigen-driven response (Table 1). Mapping these clonotypes back onto the T ceil clusters revealed that highly expanded clones in the Aire- / - sample largely mapped to the antigen-experienced clusters, such as the Thl and exhausted CD8 T cells, while those in the control group mapped to the Treg cluster, known to harbor oligoclonal Treg populations under homeostatic conditions (Munos-Rojas et al., PNAS 2024 Vol. 121 Issue 4). Six clonally expanded TCRs in the Thl cluster were identified (Table 1: TCRs 1309, 2886, 2888, 1290, 1304 and 2291). Remarkably, three out of these six TCRs (1309, 1290 and 1304) had identical alpha chains (TRAV 16N) and similar beta chains (TRBV13-1 and TRBV3-3) and had highly similar CDR3a and CDR3b regions, suggesting that they may be recognizing the same epitope. Two other TCRs (2886 and 2888) also shared a distinct TRAV chain (TRAV8D-1) and possessed highly homologous CDR3a and CDR3b regions.

[0117] It was hypothesized that the clonally expanded Th l TCRs in the Aire- / - mice were specific for epitopes of perilipin-1. To test this, an in-vitro co-culture system was used, in which candidate TCRs were cloned into a retroviral expression vector and transduced into a TCR-deficient 58- / - hybridoma cell line expressing an NFAT-GFP reporter which is activated upon TCR engagement. To test whether these TCR clones recognized perilipin-1, a library of 20-mer overlapping peptides tiling the perilipin- 1 protein was constructed, resulting in a library of 51 peptides. The TCR- expressing hybridomas were co-cultured with an antigen-presenting B cell lymphoma cell line (C3g7) expressing the cognate MHCII, I-Ag7, in the presence of individual perilipin peptides and assessed for NF AT upregulation by flow cytometry. The screen identified that TCRs 1309, 1304, 1290 and 2881 were specific for the peptide containing amino acids 261-280 of perilipin-1, while TCRs 2886 and 2888 were specific for the peptide containing amino acids 281-280 of perilipin-1. The TCR clones did not respond to other peptides in the panel, indicating high degree of specificity for the corresponding epitope. See Table 1 and TCR sequences described herein.Table 1: Perilipin-specific T cell clones from mouse adipose tissueTCR Cell Clonotype Peptide TRAV CDR3A TRBV CDR3BClone Subset Frequency Specificity CAMRGNQGGSA CASSDQEGSDYTF1309 Thl 32 TRAV16N KLIF (SEQ ID TRBV13-1 Plinl 261-280(SEQ ID NO: 16)NO: 12)40US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CATDWGGSNAK CASSIDRGDTGQL2886 Thl 14 TRAV8D-1 LTF (SEQ ID NO: TRBV19 YF (SEQ ID NO: Plinl 281-30036) 40)CATDYGGSGNK CASSLDRGDTGQ2888 Thl 11 TRAV8D-1 LIT (SEQ ID NO: TRBV26 LYF (SEQ ID NO: Plinl 281-30044) 48)CAMRGIQGGSA CASSEGTGGSYE1290 Thl 8 TRAV16N KLIF (SEQ ID TRBV13-1 QYF (SEQ ID Plinl 261-280NO: 20) NO: 24)CAMRGLQGGSA CASSGQGGSDYT1304 Thl 9 TRAV16N KLIF (SEQ ID TRBVI3-3 Plinl 261-280F_(SEQ ID NO: 8)NO: 4)CASRVGLGGDAE CAFNYNQGKLIF2291 Thl 6 TRAV6N-7 TRBV19 TLYF (SEQ ID Plinl 261-280(SEQ ID NO: 28)NO: 32)CALRNRGSALG CASSFRVEQYF3212 Treg 13 TRAV9-4 RLHF (SEQ ID TRBV29 Plinl 41-60(SEQ ID NO: 69)NO: 65)CAASATGGADR CASSLGGDTQYF872 Treg 2 TRAV14-3 LTF (SEQ ID NO: TRBV14 Plinl 201-220(SEQ ID NO: 77)73)

[0118] Next, the specificity of the two expanded Treg TCRs identified from Aire-sufficient control mice (TCRs 3212 and 872) were investigated. Co-cultures with perilipin- 1 peptide library revealed that these TCRs recognized unique perilipin-1 epitopes which were distinct from those recognized by the Thl clones from the Aire-deficient mice, with TCR 3212 recognizing amino acids 41-60 and TCR 872 recognizing the ammo acids 201-220 (Table 1). Together, these data suggest that a population of perihpin-specific adipose Tregs exist under homeostatic conditions, and that tolerance to perilipin is lost in Aire"1' mice resulting in expansion of perilipin-reactive effector CD4 T cells.

[0119] The peptide-binding groove of the I- Ag7 MHCII molecule is known to be a promiscuous peptide binder. To further define the core epitopes of the perilipin- 1 -specific TCRs, efforts focused on 3 TCRs: TCR1309, TCR 2991 and TCR 2888, recognizing amino acids 281-300 (TCR 1309 and TCR2991) and amino acids 261-280 (TCR2888). Alanine scanning libraries of peptide 281- 300 and 261-290 were created by substituting alanine for non-alanine residues one at a time (FIG. IB, FIG. 2B, and FIG. 3B), and truncation libraries, by deleting two amino acids at a time from either the C-terminus (C2, C4, C8, CIO) or N-terminus (N2, N4, N6, N8, N10) of the peptides41US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC (FIG. 1C, FIG.2C, and FIG. 3C). TCR 1309, TCR 2991 and TCR 2888-expressing hybndomas were co-cultured with C3g7 antigen-presenting cells in the presence of modified peptides, and NFAT-GFP upregulation was assessed (FIGs. 1B-1C, FIGs. 2B-2C, and FIGs. 3B-3C). Loss of the peptide ability to stimulate the hybridoma indicated that mutated or truncated amino acid residues were part of the core T-cell epitope (shaded in gray) (FIG. 1A, FIG. 2A, and FIG. 3A).There was significant overlap between the core epitope of TCR 1309 (AMQWSRR; SEQ ID NO: 57) and TCR 2991 (SAAMQWSRRQS; SEQ ID NO: 58), furthermore methionine and glutamine at positions 268 and 269 were essential for activation of both TCRs, suggesting that these may be key for presentation of the corresponding peptide. The core epitope of TCR 2886 was mapped to WLHNLAASQDE (SEQ ID NO: 59). This approach allows identification of the immunodominant regions of peril ipin-1 in this system.

[0120] It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes m light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference m their entirety for all purposes.SEQUENCES1304TRAV (CDRs underlined)?1LdL. SL. L.(}AAFCJSICFATS?vIAQKVTQTQTSIS 'rVEKTTVT 4D(: VYETRDSSYFL. FWYKQ TASGEIVFLIRQDSYKKENATVGHYSLNFOKPKSSIGLnTATOIEDSAVYFCAMRGLQGG S AKLIFGEGTKLTVS S (SEQ ID NO: 1)CDRccl: TRDSSYF ( SEQ ID NO: 2 )CDRcc2: QDSYKKEN (SEQ ID NO: 3)CDRo.3: C MRGLQGGS AKL1F (SEQ ID NO: 4)TRBV MGSRLFFVVLILLCAKHMEAAVTQSPRSKVAVrGGKVTLSCHQTNNHDYMYWYRQDT GHGLRLIHYSYVADSTEKGDIPDGYKASRPSOENFSLILELASLSOTAVYFCASSGQGGS DYTFGSGTRLLVI (SEQ ID NO: 5)CDRpl: NNHDY (SEO ID NO: 6)42US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CDRp2: SWAPS (SEP ID NO: 7)CDRp3: CASSGQGGSDYTF (SEO ID NO: 8)1309TRAVN1LILSLLGAAFGSICFATSMAQKVTQTQTSISVVEKTTVTMDCVYETRDSSYFLFWYKQ TASGEIVFLIRQDSYKKENATVGHYSLNFOKPKSSIGLUTATQIEDSAVYFCAMRGNQGG S AKLIFGEGTKLTV S S (SEQ ID NO: 9)CDRod: TRDSSYF (SEQ ID NO: 10)CDRa.2: QDSYKKEN (SEQ ID NO: 11)CDRa3: CAMRGNQGGSAKLIF (SEP ID NO: 12)TRBV MGSRLFLVLSLLCTKIIMEAAVTQSPRNKVTVTGGNVTLSCRQTNSHNYIMYWYRQDTG HGLRLIHYSYGAGNLOIGDVPDGYKATRTTOEDFFLLLELASPSOTSLYFCASSDOEGSD YTFGSGTRLLVI (SEQ ID NO: 13)CDRpl: NSHNY (SEO ID NO: 14)CDRp2: SYGAGN (SEO ID NO: 15)CDRp3: CASSDQEGSDYTF (SEQ ID NO: 16)1290TRAV MLILSLLGAAFGSICFATSMAQKVTQTQTSISVVEKTTVTMDCVYEIRDSSYFLFWYKQ TASGEIVFLIRODSYKKENATVGHYSLNFOKPKSSIGLIITATOIEDSAVYFCAMRGIQGG S AKLIFGEGTKLTVS S (SEQ ID NO: 17)CDRo; TRDSSYF (SEQ ID NO: 18)CDRoc2: QDSYKKEN (SEQ ID NO: 19)CDRa3: CAMRG1QGGSAKL1F (SEO ID NO: 20)TRBV MGSRLFLVLSLLCTKIIMEAAVTQSPRNKVTVTGGNVTESCRQTNSHNYMYWYRQDTG HGLRLIHYSYGAGNLOIGDVPDGYKAIRTTOEDFFLLLELASPSQTSLYFCASSEGTGGS YEQYFGPGTRLTVL (SEQ ID NO: 21)CDR l: NSHNY (SEO ID NO: 22)CDRp2: SYGAGN (SEO ID NO: 23)43US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CDRp3: CASSEGTGGSYEQYF (SEQ ID NO: 24)2291TRAV MNSSPGFMTV'MLLIFTRAHGDSVTQTEGQVALSEEDFLTIHCNYSASGYPALFWYVQYP GEGPOFLFRASRDKEKGSSRGFEATYDKGTTSFHLRKASVOESDSAVYYCAFNYNOGK LIFGOGTK. LSIKP (SEQ ID NO: 25)CDRal: ASGYPA (SEO ID NO: 26)CDRa2: ASRDKEK (SEQ ID NO: 27)CDRa3: CAFNYNQGKLIF (SEQ ID NO: 28)TRBV NINKWVFCWVrLCLLTVETTHGDGGirrQTPKF'LIGQEGQKLrLKCQQNFNHDTMYWY RODSGKGLRLIYYSITENDLOKGDLSEGYDASREKKSSFSLTVTSAQKNEMAVFLCASR VGLGGDAETLYFGSGTRLTVL (SEQ ID NO: 29)CDRpl: FNHDT (SEQ ID NO: 30)CDRp2: SITEND (SEO ID NO: 31)CDRp3: CASRVGLGGDAETLYF (SEQ ID NO: 32)2886TRAVMHSIXGIXJVIVSLWI> QLTRVNSQIAEENI> WALSVTIEGESVTVNCSYKTSITALQWYRQ KSGEGPAOIJIJRSNEREKRNGRLRATLDTSSOSSSLSITATRCEDTAVYFCATDWGGSN AKLTFGKGTKLS VK S (SEQ ID NO: 33)CDRal: TSITA (SEQ ID NO: 34)CDRa2: IRSNERE (SEO ID NO: 35)CDRa3: CATDWGGSNAKLTF (SEQ ID NO: 36)TRBV MNKWVFCWVTLCLLTVETTHGDGGIITQTPKFLIGQEGQKLTLKCQQNFNHDTMYWY RODSGKGLRLIYYSITENDLOKGDLSEGYDASREKKSSFSLTVTSAOKNEMAVFLCASSI DRGDTGQLYFGEGSKLTVL(SEQ ID NO: 37)CDRpl: FNHDT (SEO ID NO: 38)CDRp2: SITEND (SEO ID NO: 39)CDRp3: CASSIDRGDTGQLYF (SEO ID NO: 40)44US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC2888TRAV MHSLLGLLMVSLWLQLTR SQLAEENLWALSVHEGESVTVNCSYKTSITALQWYRQ KSGEGPAOLILIRSNEREKRNGRLRATLDTSSOSSSLSITATRCEDTAVYFCATDYGGSGN KLIFGIGTLLSVKPISEQ ID NO: 41)CDRal: TSITA (SEO ID NO: 42)CDRoc2: IRSNERE (SEQ ID NO: 43)CDRcc3: CATDYGGSGNKLIF (SEO ID NO: 44)TRBV MATRLLCYTVLCLLGARILNSKVIQTPRYLVKGQGQKAKMRCIPEKGHI VFWYQQNK NNEFKFLINFONOEVLOOIDMTEKRFSAECPSNSPCSLEIOSSEAGDSALYLCASSLDRGD TGQLYFGEGSKLTVLi SEQ ID NO: 45)CDRpl: KGHPV (SEP ID NO: 46)CDRp2: FQNQEV (SEO ID NO: 47)CDRp3: CASSLDRGDTGQLYF (SEP ID NO: 48)3212TRAV MLJ. ALLPVI, GIHFVIRDAQAQSVTQPDARVWSEGASLQLRCKYSYSATPYIFWYVQY PROGLOLLLKYYSGDPWOGXZNGFEAEFSKSNSSFHJ., RKASVHWSDSAVYFCALRNRG SAEGREHFGAGTQEIVIPD (SEQ ID NO: 62)CDRal: YSATPY (SEO ID NO: 63)CDRa2: YYSGDPW (SEQ ID NO: 64)CDRa3: CALRN;RGSALGRLHF (SEQ ID NO: 65)TRBV MRVRLISAVVLCFLGTGLVDMKVTQMPRYL1KRMGENVLLECGQDMSHETMYWYRQ DPGLGLOLIYISYDVDSNSEGDIPKGYRVSRKKREHFSLILDSAKTNOTSVYFCASSFRVE QYFGPGTRLTVL(SEQ ID NO: 66)CDRpl: MSHET (SEQ ID NO: 67)CDRp2: SYD VPS (SEQ ID NO: 68)CDRp3: CASSFRVEQYF (SEQ ID NO: 69)45US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC872TRAV MDKILTASFLLLGLHLAGVSGQQEKRDQQQV7RQSPQSLT\rWEGETAILNCSYENSAFDY FPWYQQFPGEGPALLISILSVSDKKEDGRFTIFFNKREKKLSLHIADSQPGDSATYFCAAS ATGGADRLTFGKGTQLIIQP (SEQ ID NO: 70)CDRal: ENSAFDY (SEQ ID NO: 71)CDRa2: ILSVSDK (SEQ ID NO: 72)CDRa3: CAASATGGADRLTF (SEP ID NO: 73)TRBV MGTRLLGWAVFCLLDTVLSEAGVTQSPRYAVLQEGQAVSFWCDPISGFIDTLYWYQQP RDOGPOLLVYFRDEAVIDNSOLPSDRFSAVRPKGTNSTLKIOSAKOGDTATYLCASSLG GDTQYFGPGTRLLVLE (SEQ ID NO: 74)CDRpl: SGHDT (SEQ ID NO: 75)CDRp2: FRDEAV (SEQ ID NO: 76)CDRp3: CASSLGGDTQYF (SEO ID NO: 77)3212 recognized Class II peptide: YNS1XEAHPLVASVCNAYEK (SEQ ID NO:78)872 recognized Class II peptide: SSGRQRTQKAPKAKPSLVRR (SEQ ID NO: 79) TCRalpha Mouse Constant Region - common to all TCRs IQNPEPAVYQI. KDPRSQDSTI. CIETDFDSQINYTKTMESGTFITDKTVl. DNIKAMDSKSN GAlAWSNQTSFTCQDIFKETNATYPSSDVPCDATLTEKSFETDMNLNFQNLSV?v4GERILL LKVAGFNILMTI. RLWS(SEQ ID NO: 49)TCRbeta Mouse Constant Region - common to all TCRs EDI NVTPPKVSLFEPSKAEIANKQKATIA DARGFFPDHVELSWWVNGKEVHSGVST DPQAYKESNYSYCLSSRLRVSATFWHNPRNHFRCQVQFHGLSEEDKWPEGSPKPVTQNl SAEAWGRADCGITSASYHQGVLSA'niYEILLGKATI. YAVIVSGLVIMAMVKKKNS(SE Q ID NO: 50)Consensus CDRa3: CATDXGGSXXKLXF (SEQ ID NO: 51)Consensus CDRp3 CASSXDRGDQLYF (SEQ ID NO: 52)Consensus CDRa3 CAMRGXQGGSAKLIF (SEQ ID NO: 53)Consensus CDRp3 CASXXXXGXXX (SEQ ID NO: 54)46US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC QWGASAAMQVVSRRQSEVRV (SEQ ID NO: 55)PWLHNLAASQDESHDDQTDT (SEQ ID NO: 56)AMQVVSRR (SEQ ID NO: 57)SAAMQVVSRRQS (SEQ ID NO: 58)WLHNLAASQDE (SEQ ID NO: 59)47US1 155099385 1

Claims

Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC WHAT IS CLAIMED IS:

1. A cell expressing (i) a heterologous T-cell receptor (TCR) that binds to a perilipin 1 (PLIN1) peptide complexed with a major histocompatibility (MHC) molecule or (ii) a chimeric antigen receptor (CAR) comprising a PLIN1 -binding extracellular domain.

2. The cell of claim 1, wherein the cell is a T-cell.

3. The cell of claim 2, wherein the cell comprises the heterologous TCR.

4. The cell of claim 3, wherein the cell does not express an endogenous TCR.

5. The cell of claim 4, wherein the cell comprises a nucleic acid encoding the heterologous TCR inserted in a coding sequence for an endogenous TCR such that expression of the endogenous TCR is disrupted.

6. The cell of any one of claims 1-5, wherein the PLIN1 peptide is an MHC class I or class II peptide and comprises QWGASAAMQVVSRRQSEVRV (SEQ ID NO: 55), PWLHNLAASQDESHDDQTDT (SEQ ID NO: 56), AMQVVSRR (SEQ ID NO: 57), SAAMQVVSRRQS (SEQ ID NO: 58), WLHNLAASQDE (SEQ ID NO: 59), YNSTKEAHPLVASVCNAYEK (SEQ ID NO: 78), SSGRQRTQKAPKAKPSLVRR (SEQ ID NO:79) or a contiguous fragment of at least 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 amino acids and optionally having one or two ammo acid substitutions.

7. The cell of any one of claims 1-5, wherein the MHC is a human leukocyte antigen (HLA) molecule and the PLIN1 peptide is a human PLIN1 peptide.

8. The cell of any one of claims 1-7, wherein the heterologous TCR comprises an alpha chain comprising a variable alpha (Voc) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa3 and a beta chain comprising a variable beta (VP) region comprising a CDR i, CDR 2 and CDR 3, and wherein:CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO: 51) and CDRp3 comprises CASSXDRGDQLYF (SEQ ID NO: 52); or48US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC CDRa3 comprises CAMRGXQGGSAKLIF (SEQ ID NO: 53) and CDRp3 comprises CASXXXXGXXX (SEQ ID NO: 54), where X is any ammo acid; orCDRa3 comprises CALRNRGSALGRLHF (SEQ ID NO: 65) and CDRp3 comprises CASSFRVEQYF (SEQ ID NO: 69), orCDRa3 comprises CAASATGGADRLTF (SEQ ID NO: 73) and CDRp3 comprises CASSLGGDTQYF (SEQ ID NO: 77).

9. The cell of claim 8, wherein:the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO: 4, respectively and CDR i, CDRp2 and CDRp3 comprise SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO: 12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO:15 and SEQ ID NO: 16, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO:20 respectively and CDRpi, CDRP2 and CDRp3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28, respectively and CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:30, SEQ ID NO:31 and SEQ ID NO:32, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDR i, CDR 2 and CDRp3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; orthe CDR l, CDRa2 and CDRa3 comprise SEQ ID NO: 42, SEQ ID NO: 43 and SEQ ID NO:44, respectively and CDRpi, CDRP2 and CDRp3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO:65, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:67, SEQ ID NO:68 and SEQ ID NO:69, respectively; or49US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73, respectively and CDRpl, CDRp2 and CDRp3 comprise SEQ ID NO:75, SEQ ID NO:76 and SEQ ID NO:77, respectively.

10. The cell of claim 2, wherein the cell comprises the CAR.

11. The cell of claim 10, wherein the PLIN 1 -binding extracellular domain comprises a single chain TCR.

12. The cell of claim 11, wherein the single chain TCR comprises an alpha chain comprising a variable alpha (Voc) region comprising a complementarity' determining region (CDR) al, CDRa2 and CDRa3 and a beta chain comprising a variable beta (VP) region comprising a CDRpi, CDRP2 and CDRp3, and wherein:CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO: 51) and CDRp3 comprises CASSXDRGDQLYF (SEQ ID NO: 52); orCDRa3 comprises CAMRGXQGGSAKLIF (SEQ ID NO: 53) and CDRp3 comprises CASXXXXGXXX (SEQ ID NO: 54), where X is any amino acid; orCDRa3 comprises CALRNRGSALGRLI IF (SEQ ID NO: 65) and CDRp3 comprises CASSFRVEQYF (SEQ ID NO: 69), orCDRa3 comprises CAASATGGADRLTF (SEQ ID NO: 73) and CDRP3 comprises CASSLGGDTQYF (SEQ ID NO: 77).

13. The cell of claim 12, wherein:the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO: 2, SEQ ID NO: 3 and SEQ ID NO:4, respectively and CDRpl, CDRp2 and CDRp3 comprise SEQ ID NO:6, SEQ ID NO: 7 and SEQ ID NO: 8, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO: 12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:18, SEQ ID NO:19 and SEQ ID NO:20, respectively and CDRpl, CDRP2 and CDRP3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; or50US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:2,8 respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:30, SEQ ID NO:31 and SEQ ID NO:32, respectively; orthe CDRal, CDRa2 and CDRot.3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO: 42, SEQ ID NO: 43 and SEQ ID NO:44, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:63, SEQ ID NO:64 and SEQ ID NO: 65, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 67, SEQ ID NO:68 and SEQ ID NO:69, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO:73, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:75, SEQ ID NO:76 and SEQ ID NO:77, respectively.

14. The cell of any one of claims 2-12, wherein the cell is a cytotoxic T-cell.

15. The cell of any one of claims 2-12, wherein the cell is a regulatory T-cell.

16. The cell of any one of claims 1-15, wherein the cell is a human cell,17. A nucleic acid encoding a TCR alpha chain comprising a variable alpha (Vα) region comprising a complementarity determining region (CDR) α1, CDRα2 and CDRα3, wherein:CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO: 51) or CAMRGXQGGSAKLIF (SEQ ID NO: 53), wherein X is any amino acid; orCDRa3 comprises CALRNRGSALGRLHF (SEQ ID NO: 65); or CDRa3 comprises CAASATGGADRLTF (SEQ ID NO: 73).

18. The nucleic acid of claim 17, wherein:51US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO: 10, SEQ ID NO:11 and SEQ ID NO: 12, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:18, SEQ ID NO:19 and SEQ ID NO:20, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO: 28 respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO: 63, SEQ ID NO: 64 and SEQ ID NO: 65 respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:71, SEQ ID NO:72 and SEQ ID NO: 73 respectively.

19. A nucleic acid encoding a TCR beta chain comprising a variable beta (VP) region comprising a CDRβ1, CDRβ2 and CDRβ3, wherein CDRP3 comprises CASSXDRGDQLYF (SEQ ID NO: 52) or CASXXXXGXXX (SEQ ID NO: 54), wherein X is any amino acid; orwherein CDRP3 comprises CASSFRVEQYF (SEQ ID NO: 69), or wherein CDRp3 comprises CASSLGGDTQYF (SEQ ID NO: 77).

20. The nucleic acid of claim 19, wherein:the CDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; orthe CDR i, CDRp2 and CDRp3 comprise SEQ ID NO: 46, SEQ ID NO: 47 and SEQ ID NO:48, respectively; orCDRpi, CDRp2 and CDRp3 comprise SEQ ID NO: 67, SEQ ID NO: 68 and SEQ ID NO: 69, respectively; orCDRpi, CDRp2 and CDRp3 comprise SEQ ID NO:75, SEQ ID NO:76 and SEQ ID NO: 77, respectively.52US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC 21. The nucleic acid of claim 17 or 19, wherein the nucleic acid comprises RNA.

22. The nucleic acid of claim 17 or 19, wherein the nucleic acid comprises DNA.

23. The nucleic acid of claim 21, wherein the nucleic acid comprises one or more non-naturally-occurring ribonucleotide or nucleoside modification.

24. A vector comprising the nucleic acid of claim 17 or 19 or both the nucleic acid of claim 17 and 19.

25. A method of forming a cell that expresses a heterologous T-cell receptor (TCR) that binds to perilipin 1 (PLIN1), the method comprising,introducing:a. a nucleic acid encoding a TCR alpha chain comprising a variable alpha (Va) region comprising a complementarity determining region (CDR) al, CDRa2 and CDRa3 wherein CDRa3 comprises CATDXGGSXXKLXF (SEQ ID NO:51) or CAMRGXQGGSAKLIF (SEQ ID NO:53), wherein X is any amino acid; andb. nucleic acid encoding a TCR beta chain comprising a variable beta (Vβ) region comprising a CDRβ1, CDRβ2 and CDRβ3, wherein CDRβ3 comprises CASSXDRGDQLYF (SEQ ID NO: 52) or CASXXXXGXXX (SEQ ID NO:54), wherein X is any amino acid;such that the cell expresses the TCR alpha and beta chains to form the heterologous TCR that binds to PLIN1,26. The method of claim 25, wherein:the CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:6, SEQ ID NO: 7 and SEQ ID NO: 8, respectively; or53US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC the CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:10, SEQ ID NO:11 and SEQ ID NO: 12, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO: 14, SEQ ID NO: 15 and SEQ ID NO: 16, respectively; orthe CDRal, CDRcx2 and CDRot.3 comprise SEQ ID NO: 18, SEQ ID NO: 19 and SEQ ID NO:20, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:22, SEQ ID NO:23 and SEQ ID NO:24, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO: 26, SEQ ID NO: 27 and SEQ ID NO:2,8 respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:30, SEQ ID NO:31 and SEQ ID NO:32, respectively; orthe CDRal, CDRa2 and CDRa3 comprise SEQ ID NO:34, SEQ ID NO:35 and SEQ ID NO:36, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40, respectively; orthe CDRα1, CDRα2 and CDRα3 comprise SEQ ID NO:42, SEQ ID NO:43 and SEQ ID NO:44, respectively and CDRβ1, CDRβ2 and CDRβ3 comprise SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48, respectively.

27. The method of claim 25, wherein the cell is a T-cell.

28. The method of claim 25 or 27, wherein the nucleic acid encoding the alpha chain and the nucleic acid encoding the beta chain comprise RNA29. The method of claim 25 or 27, wherein the introducing comprises inserting the nucleic acids into an endogenous TCR coding sequence such that the endogenous TCR is not expressed.

30. The method of any one of claims 25-29, wherein the cells are obtained from a human subject.

31. The method of claim 30, wherein following the introducing the cells expressing the TCR alpha and beta chains are introduced into the same or a different human subject.54US1 155099385 1Attorney Docket No. 110221-1543849-012110WO Client Ref. No. CZB-320F -PC 32. The method of any one of claims 25-29, wherein the introducing occurs in vitro, ex vivo or in vivo.

33. A method of reducing fat in a subject, the method comprising introducing the cell of claim 14 into the subject in a sufficient amount to reduce fat in the subject.

34. The method of claim 33, wherein the subject is a human.

35. A method of reducing fat in a subject, the method comprising introducing the nucleic acids of claims 17 and 19, or the vector of claim 24, into one or more cytotoxic T-cell of the subject such that the one or more T-cell expresses a heterologous TCR that binds to PLIN1 and wherein the T-cell expressing the heterologous TCR are in a sufficient amount to reduce fat in the subject.

36. The method of claim 37, wherein the subject is a human.

37. A method of regulating fat in a subject, the method comprising introducing the cell of claim 15 into the subject in a sufficient to reduce fat in the subject.

38. The method of claim 37, wherein the subject is a human.

39. A method of regulating fat in a subject, the method comprising introducing the nucleic acids of claims 17 and 19, or the vector of claim 24, into one or more regulatory T-cell of the subject such that the one or more regulatory T-cell expresses a heterologous TCR that binds to PL1N1 and wherein the regulatory T-cell expressing the heterologous TCR are in a sufficient amount to regulate fat in the subject.

40. The method of claim 39, wherein the subject is a human.55US1 155099385 1

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