INDUS 20190321403A1 IN (19 ) United States (12 ) Patent Application Publication ( 10 ) Pub . No .: US 2019/0321403 A1 LEVITSKY (43 ) Pub . Date : Oct. 24 , 2019 ( 54 ) ENGINEERED B CELLS AND RELATED Publication Classification COMPOSITIONS AND METHODS ( 51 ) Int. Ci. A61K 35/17 (2006.01 ) (71 ) Applicant: Juno Therapeutics , Inc., Seattle, WA CI2N 15/85 (2006.01 ) (US ) C12N 15/62 ( 2006.01 ) A61 ) 1/10 (2006.01 ) ( 72 ) Inventor : Hyam I. LEVITSKY , Seattle , WA C07K 16/28 ( 2006.01 ) (US ) (52 ) U.S. CI. CPC A61K 35/17 (2013.01 ) ; C12N 15/85 ( 73 ) Assignee: Juno Therapeutics, Inc. , Seattle , WA ( 2013.01) ; C12N 15/62 (2013.01 ) ; A61K 45/06 (US ) ( 2013.01 ) ; CO7K 16/28 ( 2013.01) ; C12N 2015/8518 ( 2013.01 ) ; A61J 1/10 ( 2013.01) (21 ) Appl. No .: 16 /465,109 (57 ) ABSTRACT Provided herein are engineered B cells , such as for adoptive ( 22 ) PCT Filed : Nov. 30 , 2017 cell therapy. In some aspects , also provided are methods and compositions for engineering and producing the cells , com ( 86 ) PCT No.: PCT /US2017 / 064075 positions containing the cells , and methods for their admin $ 371 ( c ) ( 1 ) , istration to subjects . In some embodiments , the cells are (2 ) Date : May 29 , 2019 engineered to produce and /or secrete an exogenous protein , such as a therapeutic protein , including antibodies and antigen - binding fragments thereof. In some aspects , features Related U.S. Application Data of the cells and methods provide for increased or improved (60 ) Provisional application No. 62/ 429,709, filed on Dec. activity , efficacy and /or persistence of the cells . 2 , 2016 . Specification includes a Sequence Listing . US 2019/0321403 A1 Oct. 24 , 2019 1 ENGINEERED B CELLS AND RELATED effect secretion of the exogenous protein from the cell, COMPOSITIONS AND METHODS wherein the exogenous protein is not an antibody. [0007 ] In some embodiments , provided are engineered B CROSS -REFERENCE TO RELATED cells comprising one or more nucleic acid molecules com APPLICATIONS prising one or more coding sequences encoding an exog enous protein , wherein expression of the exogenous protein [0001 ] This application claims priority from U.S. provi in the engineered B cell is conditional. sional application No. 62 /429,709 filed Dec. 2 , 2016 , [ 0008 ] In some embodiments , provided are engineered B entitled “ Engineered B Cells and Related Compositions and cells comprising one or more nucleic acid molecules com Methods, ” the contents of which are incorporated by refer prising one or more coding sequences encoding an exog ence in their entirety . enous protein , wherein the engineered B cell expresses an endogenous antibody and comprises a modification that INCORPORATION BY REFERENCE OF prevents class - switching of the endogenous antibody and /or SEQUENCE LISTING prevents switching of the endogenous antibody from a [0002 ] The present application is being filed with a membrane -associated form to a secreted form . Sequence Listing in electronic format. The Sequence Listing [ 0009 ] In some embodiments , provided are engineered B is provided as a file entitled 735042006640seqlist.txt, cre cells comprising one or more nucleic acid molecules com ated Nov. 30 , 2017 , which is 64,032 bytes in size . The prising one or more coding sequences encoding an exog information in electronic format of the Sequence Listing is enous protein , wherein at least one of the one or more incorporated by reference in its entirety . nucleic acid molecules is integrated into or replaces all or a portion of a heavy chain immunoglobulin locus or a light FIELD chain immunoglobulin locus of the B cell. [ 0003] The present disclosure relates in some aspects to [0010 ] In some embodiments , provided are engineered B engineered B cells , such as for adoptive cell therapy. In some cells comprising one or more nucleic acid molecules com aspects , the disclosure further relates to methods and com prising one or more coding sequences encoding an exog positions for engineering and producing the cells , compo enous protein , wherein the engineered B cell comprises one sitions containing the cells , and methods for their adminis or more modifications resulting in a greater capacity for the tration to subjects . In some embodiments , the cells are engineered B cell to produce and/ or secrete the exogenous engineered to produce and /or secrete an exogenous protein , protein . such as a therapeutic protein , including antibodies and [ 0011 ] In some embodiments , provided are engineered B antigen - binding fragments thereof. In some aspects , features cells comprising : one or more nucleic acid molecules com of the cells and methods provide for increased or improved prising one or more coding sequences encoding an exog activity , efficacy and /or persistence of the cells . enous protein ; and a chimeric receptor comprising a ligand binding domain , wherein , upon ligand binding , the receptor is capable of inducing (i ) a mitogenic or proliferative signal; BACKGROUND and / or ( ii ) a signal that is capable of modulating the differ [0004 ] Various methods are available for treating diseases , entiation of the engineered B cell. including infectious diseases , cancers, and autoimmune dis [0012 ] In some embodiments , provided are engineered B eases , using therapeutic proteins, such as antibodies or cells comprising : one or more nucleic acid molecules com antigen -binding fragments thereof. Such approaches gener prising one or more coding sequences encoding an exog ally involve repeated injections of recombinantly -produced enous protein ; and a recombinant receptor comprising a proteins, which can provide various therapeutic effects via ligand binding domain , wherein , upon ligand binding , the one or more mechanisms. The presence of the therapeutic receptor is capable of inducing (i ) a mitogenic or prolifera proteins in the body following administration is generally tive signal; and /or ( ii) a signal that is capable of modulating transient. Improved compositions and methods are needed , the differentiation of the engineered B cell, wherein the for example , to improve efficacy of such therapies, for exogenous protein does not bind to the target of the ligand example , by increasing the duration of action of the thera binding domain of the receptor and/ or the exogenous protein pies . Provided are products , compositions, methods and does not contain a ligand binding site contained in the ligand articles of manufacture that meet such needs . binding domain of the receptor. [ 0013 ] In some of any such embodiments , the exogenous SUMMARY protein is secreted by the B cell or is capable of being [ 0005 ] Provided are engineered B cells capable of produc secreted by the B cell. In some embodiments , the one or ing and / or secreting an exogenous protein , such as a thera more coding sequences comprises a nucleotide sequence peutic protein , such as for use in adoptive cell therapy, for encoding a secretory signal peptide . In some embodiments , example to treat diseases and / or conditions in a subject in the secretory signal peptide comprises an amino acid need thereof. Also provided are compositions comprising sequence selected from among SEQ ID NOs: 76-202 . the cells , methods of producing and using the cells , such as [ 0014 ] In some of any such embodiments , the exogenous for treating a disease and /or condition , and articles of protein is a dimer . In some embodiments , the one or more manufacture comprising the cells or for use in a method nucleic acid molecules comprises a single nucleic acid described herein . molecule comprising a first coding sequence encoding a first [0006 ] In some embodiments , provided are engineered B domain or subunit of the dimer and a second coding cells comprising one or more nucleic acid molecules com sequence encoding a second domain or subunit of the dimer . prising one or more coding sequences encoding an exog [0015 ] In some of any such embodiments , the exogenous enous protein under the control of one or more elements to protein is a therapeutic protein . US 2019/0321403 A1 Oct. 24 , 2019 2 [ 0016 ] In some of any such embodiments , the exogenous nucleic acid molecule comprising a first coding sequence protein binds to a target molecule associated with a disease encoding the heavy chain and a second coding sequence or condition , wherein the molecule is optionally a protein , encoding the light chain of the antibody or antigen -binding wherein the molecule or protein is expressed on the surface fragment thereof. In some embodiments , the antibody or of a cell . In some embodiments , the disease or condition is antigen -binding fragment thereof comprises one or more selected from among a tumor or cancer, an autoimmune modifications in the heavy chain and /or light chain such that disease , an infectious disease or condition , and an inflam when the exogenous antibody or antigen - binding fragment is matory disease. In some embodiments , the disease or con expressed in a cell, the frequency of mispairing with a heavy dition is a tumor or cancer . In some embodiments , the chain and / or light chain of an endogenous antibody is disease or condition is a viral infection . In some embodi reduced . In some embodiments, the one or more modifica ments , the viral infection is human immunodeficiency virus tions are in the CH2 and / or CH3 region of the constant (HIV ) infection . chain . In some embodiments , the one or more modifications [0017 ] In some of any such embodiments , the exogenous comprise a knob - into -hole (KiH ) modification or a dock and protein binds to a molecule selected from ROR1 , Her2 , lock (DNL ) modification . In some embodiments , the anti L1- CAM , CD19 , CD20 , CD22, mesothelin , CEA , hepatitis body or antigen -binding fragment thereof is a full- length B surface antigen , anti- folate receptor, CD23 , CD24 , CD30 , antibody . In some embodiments , the antibody or antigen CD33 , CD38 , CD44 , EGFR , EGP - 2 , EGP - 4 , EPHa2 , binding fragment thereof is a single chain antibody frag ErbB2 , ErbB3 , ErbB4 , FBP , fetal acetylcholine receptor, ment. In some embodiments , the antibody or antigen -bind GD2, GD3 , HMW -MAA , IL - 22R -alpha , IL -13R - alpha2 , ing fragment thereof is an scFv.