(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
`(19) World Intellectual Property
`Organization
`International Bureau
`
`(43) International Publication Date
`4 May 2017 (04.05.2017)
`
`WIPOI PCT
`
`\9
`
`(10) International Publication Number
`
`WO 2017/075537 A1
`
`(51)
`
`International Patent Classification:
`C07K 14/705 (2006.01)
`C07K 19/00 (2006.01)
`C07K 16/28 (2006.01)
`
`(21)
`
`International Application Number:
`
`PCT/US2016/059582
`
`(22)
`
`International Filing Date:
`
`28 October 2016 (28.10.2016)
`
`Filing Language:
`
`Publication Language:
`
`English
`
`English
`
`(81) Designated States (unless otherwise indicated, for every
`kind of national protection available): AE, AG, AL, AM,
`AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY,
`BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM,
`DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT,
`HN, HR, HU, ID, IL, IN, IR, IS, JP, KE, KG, KN, KP, KR,
`KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME,
`MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ,
`OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA,
`SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM,
`TN, TR, TT, TZ, UA, UG, us, UZ, VC, VN, ZA, ZM,
`ZW.
`
`Priority Data:
`62/249,144
`62/331,010
`62/396,783
`
`30 October 2015 (30.10.2015)
`3 May 2016 (03.05.2016)
`19 September 2016 (19.09.2016)
`
`(84)
`
`US
`US
`US
`
`INC.
`Applicant: ALETA BIOTHERAPEUTICS
`[US/US]; 2 Mercer Road, Natick, Massachusetts 01760
`(US).
`
`Designated States (unless otherwise indicated, for every
`kind of regional protection available): ARIPO (BW, GH,
`GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ,
`TZ, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU,
`TJ, TM), European (AL, AT, BE, BG, CH, CY, CZ, DE,
`DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU,
`LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK,
`SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ,
`Inventors: LOBB, Roy; 580 Washington Street, Apt. 304,
`GW, KM, ML, MR, NE, SN, TD, TG).
`Wellesley, Massachusetts 02482 (US). RENNERT, Paul;
`205 Underwood Street, Holliston, Massachusetts 01746 Declarations under Rule 4.17 :
`(US).
`
`Agents: VETTER, Michael L. et a1.; Choate, Hall &
`Stewart LLP, Two International Place, Boston, Massachu-
`setts 02110 (US).
`
`as to applicant’s entitlement to applyfor and be granted a
`patent (Rule 4.1 7(ii))
`Published:
`
`(25)
`
`(26)
`
`(30)
`
`(71)
`
`(72)
`
`(74)
`
`with international search report (Art. 21(3))
`
`(54) Title: COMPOSITIONS AND METIIODS FOR TREATMENT OF CANCER
`
`Figure 1: Novel Inducible Genes
`
`® antigen binding domain /l
`
`cell membrane
`
`gene product
`,1 Q)
`
`I
`
`
`
`
`(D signaling domain
`
`\
` , signaling cascade
`
`(SD inserted inducible gene
`
`(57) Abstract: Compositions, e. g., compositions comprising cellular therapeutics and/or protein therapeutics, and methods of using
`such compositions for treating cancer are described.
`
`
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`WO 2017/075537
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`PCT/US2016/059582
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`COMPOSITIONS AND METHODS FOR TREATMENT OF CANCER
`
`Cross Reference To Related Applications
`
`[0001]
`
`This application claims priority to each of US. Provisional Patent Application
`
`Nos. 62/249,144 filed October 30, 2015; 62/331,010 filed May 3, 2016; and 62/396,783 filed
`
`September 19, 2016, the entire contents of each of which are hereby incorporated by reference.
`
`Background
`
`[0002]
`
`Adoptive cell therapy (ACT) is a treatment method in which cells are removed
`
`from a donor, cultured and/or manipulated in vitro, and then administered to a patient for the
`
`treatment of a disease. A variety of cell types have been used in ACT in an attempt to treat
`
`several classes of disorders. For the treatment of cancer, ACT generally involves the transfer of
`
`lymphocytes, such as chimeric antigen receptor (CAR) T cells. Use of such CAR T cells
`
`involves identifying an antigen on a tumor cell to which a CAR T cell can bind, but tumor
`
`heterogeneity can make antigen identification challenging. Accordingly, there remains a need
`
`for improved methods for treating cancer using adoptive cell therapy.
`
`Summary
`
`[0003]
`
`The present invention provides methods and compositions useful for treatment of
`
`cancer and/or for initiating or modulating immune responses. In some embodiments, the present
`
`invention provides cellular therapeutics (e.g., immune cells) comprising a constitutive expression
`
`construct, which comprises a promoter operably linked to a gene of interest. In some
`
`embodiments, the present invention provides cellular therapeutics (e.g., immune cells)
`
`comprising (i) an antigen binding receptor, wherein the antigen binding receptor comprises an
`
`antigen-binding domain, a transmembrane domain, and a cytosolic signaling domain, and (ii) an
`
`inducible expression construct, which comprises a promoter operably linked to a gene of interest.
`
`Among other things, the present invention encompasses the recognition that a combination of a
`
`

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`WO 2017/075537
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`PCT/US2016/059582
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`cellular therapeutic described herein and one or more additional therapies (e.g., one or more
`
`additional cellular therapeutics (e.g., CAR-T cell, CAR-NK cell, TCR-T cell, TIL cell, allogenic
`
`NK cell, and autologous NK cell), antibody-drug conjugate, an antibody, and/or a polypeptide
`
`described herein), can lead to improved induction of beneficial immune responses, for example a
`
`cellular response (e.g., T-cell activation).
`
`[0004]
`
`In some embodiments, the present disclosure provides methods of treating a
`
`subject having a tumor, comprising administering to the subject a cellular therapeutic described
`
`herein and/or a protein therapeutic described herein. In some embodiments, methods further
`
`comprise administration of one or more additional therapies (e. g, a second cellular therapeutic
`
`(e. g., CAR—T cell, CAR—NK cell, TCR—T cell, TIL cell, allogenic NK cell, and autologous NK
`
`cell), an antibody—drug conjugate, an antibody, and/or a polypeptide described herein).
`
`[0005]
`
`Other features, objects, and advantages of the present invention are apparent in
`
`the detailed description that follows. It should be understood, however, that the detailed
`
`description, while indicating embodiments of the present invention, is given by way of
`
`illustration only, not limitation. Various changes and modifications within the scope of the
`
`invention will become apparent to those skilled in the art from the detailed description.
`
`Brief Description of the Drawings
`
`[0006]
`
`The figures of the drawing are for illustration purposes only, not for limitation.
`
`[0007]
`
`Figure 1 is a schematic depicting an exemplary cellular therapeutic.
`
`[0008]
`
`Figure 2 is a schematic depicting an exemplary cellular therapeutic encoding an
`
`inducible scFv-CD19 fusion protein.
`
`[0009]
`
`Figure 3 is a schematic depicting an exemplary cellular therapeutic encoding an
`
`inducible scFv-EGFR fusion protein.
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`[0010]
`
`Figure 4 is a schematic depicting an exemplary “self amplifying” cellular
`
`therapeutic encoding an inducible scFv-CD19 fusion protein and an inducible CAR that targets
`
`CD19.
`
`[0011]
`
`Figure 5 is a schematic depicting an exemplary “self amplifying” cellular
`
`therapeutic encoding an inducible scFV—CD 19 fusion protein and a constitutively expressed CAR
`
`that targets CD 19.
`
`[0012]
`
`Figure 6 is a schematic depicting an exemplary “self amplifying” cellular
`
`therapeutic expressing an antigen binding receptor that does not include a signaling domain
`
`leading to induction of killing, and does include a signaling domain sufficient to induce gene
`
`transcription, and also encoding an inducible scFV-CD19 fusion protein and an inducible CAR
`
`(left) or a constitutively expressed CAR (right) that targets CD19.
`
`[0013]
`
`Figure 7 is a schematic depicting an exemplary cellular therapeutic encoding
`
`various inducible genes.
`
`[0014]
`
`Figure 8 is a schematic depicting an exemplary cellular therapeutic encoding an
`
`inducible cytokine.
`
`[0015]
`
`Figure 9 is a schematic depicting an exemplary cellular therapeutic encoding an
`
`inducible scFV-CD3O fusion protein.
`
`[0016]
`
`Figure 10 is a schematic depicting an exemplary cellular therapeutic encoding an
`
`inducible toxin.
`
`[0017]
`
`Figure 11 is a schematic depicting an exemplary cellular therapeutic encoding
`
`various inducible genes.
`
`[0018]
`
`Figures 12A, 12B, and 12C are schematics depicting exemplary CD19 variants.
`
`[0019]
`
`Figure 13 is a schematic depicting exemplary antibody fusion proteins in which a
`
`polypeptide antigen is fused to the C terminus of a light chain (LC) of an antibody, a polypeptide
`
`antigen is fused to the N terminus of a LC of an antibody, a polypeptide antigen is fused to the C
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`terminus of a heavy chain (HC) of an antibody, or a polypeptide antigen is fused to the N
`
`terminus of a HC of an antibody.
`
`[0020]
`
`Figure 14A and 14B show expression levels of various polypeptide antigen—
`
`antibody fusion constructs.
`
`[0021]
`
`Figure 15 is a schematic depicting exemplary antibody fusion proteins in which a
`
`polypeptide antigen is fused in various orientations to an scFv.
`
`[0022]
`
`Figure 16 shows expression levels of various polypeptide antigen—scFv fusion
`
`constructs.
`
`[0023]
`
`Figures 17A, 17B, 17C, and 17D show binding of panitumumab-CDl 9 fusion
`
`proteins to an anti-CD19 antibody (FMC63).
`
`[0024]
`
`Figure 18 shows binding of panitumumab-CD19 fusion proteins to an anti-CD19
`
`antibody (FMC63) relative to negative controls.
`
`[0025]
`
`Figures 19A, 19B, 19C, and 19D show binding ofLY2875358-CD19 fusion
`
`proteins to an anti-CD19 antibody (FMC63).
`
`[0026]
`
`Figure 20 shows binding of LY2875358-CD19 fusion proteins to an anti-CD19
`
`antibody (FMC63) relative to negative controls.
`
`[0027]
`
`Figure 21 shows a summary of expression of, and FMC63 binding to, various
`
`antibody-CD 19 fusion proteins.
`
`[0028]
`
`Figure 22 shows binding of trastuzumab scFv-CD19 fusion proteins to an anti-
`
`CD19 antibody (FMC63).
`
`[0029]
`
`Figures 23A, 23B, and 23C show binding of LY2875358—CD19 fusion proteins to
`
`c—Met expressing cells and to an anti—CD19 antibody (FMC63).
`
`[0030]
`
`Figures 24A and 24B show binding of trastuzumab scFv-CD19 fusion proteins to
`
`an anti-CD19 antibody (FMC63) and to Her-2 protein.
`
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`[0031]
`
`Figures 25A and 25B show binding of trastuzumab scFv-CD19 fusion proteins to
`
`an anti-CD19 antibody (FMC63) relative to negative controls.
`
`[0032]
`
`Figure 26 shows binding of CD19—scFv fusion proteins captured on anti—His
`
`antibody—coated ELISA plates.
`
`[0033]
`
`Figure 27 shows binding of CD19-scFv fusion proteins captured on anti-His
`
`antibody-coated ELISA plates.
`
`[0034]
`
`Figure 28 shows binding of CD19—scFv fusion proteins captured on anti—FMC63
`
`(anti—CD19)—coated plates, then detected with anti—His—HRP.
`
`[0035]
`
`Figure 29 shows detection of CD1 9-anti-Her2 trastuzumab scFv-human Fc fusion
`
`proteins in a "sandwich ELISA" format.
`
`[0036]
`
`Figure 30 shows the capture of multiple fusion proteins by anti-CD19 monoclonal
`
`antibody FMC63 and their detection by anti-His antibody coupled to HRP.
`
`[0037]
`
`Figure 31 shows the capture of CD19 full-length extracellular domain-anti-CD2O
`
`Leu16 scFv Vh-Vl-His fusion protein by the C-terminal His tag and then detected by mouse
`
`monoclonal antibody FMC63 anti-CD19 and then anti-mouse lgG-HRP.
`
`[0038]
`
`Figure 32 shows results for fusion proteins that incorporate CD22 protein
`
`domains, or anti-EGFRVIII scFv (#64: CD22-FMC63 scFv-His; #65: CD22-anti-CD2O scFv-His;
`
`#67: CD19 full ECD-anti-EGFRVIII scFv-his; #68: CD22-anti-EGFRVIII scFv-His).
`
`[0039]
`
`Figure 33 shows results for protein-antibody fusion proteins and protein-scFv
`
`fusion proteins derived from the same antibody, panitumumab (#57: Her2 extracellular domain-
`
`Panitumumab scFv Vh-Vl-His; #58 Her2 extracellular D4- Panitumumab scFv Vh-Vl-His;
`
`#3 3+4 (cotransfection of heavy and light chains; one chain carries the CD19 fusion): CD19
`
`extracellular D 1+2 Panitumumab antibody — His).
`
`[0040]
`
`Figure 34 shows binding affinity of purified CD19-anti-Her2 scFv-His fusion
`
`protein for the FMC63 antibody.
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`[0041]
`
`Figure 35 shows the binding affinity of the FMC63-bound CDl9-anti-Her2 scFV-
`
`His fusion protein to Her2.
`
`[0042]
`
`Figure 36 shows the binding affinity of the FMC63—bound CDl9—anti—Her2 scFV—
`
`His fusion protein to anti—Her2 scFV.
`
`[0043]
`
`Figure 37 shows a flow cytometry profile of fusion protein CDl9-ECD-Leul6
`
`scFV (VH/VL)(#63) bound to CD20 expressing 293 cells and labeled with anti-CD19
`
`monoclonal antibody FMC63-PE-conjugated.
`
`[0044]
`
`Figure 38 shows a flow cytometry profile of fusion protein CDl9—Dl+2—Leul6
`
`scFV (VH/VL) (#83) bound to CD20 expressing 293 cells and labeled with anti—CD19
`
`monoclonal antibody FMC63—PE—conjugated.
`
`[0045]
`
`Figure 39 shows a flow cytometry profile of fusion protein CDl9-Dl+2-Leul6
`
`scFV (VL/VH) (#85) bound to CD20 expressing 293 cells and labeled with anti-CD19
`
`monoclonal antibody FMC63-PE-conjugated.
`
`[0046]
`
`Figure 40 shows a flow cytometry profile of fusion protein CDl9-Dl+2-Leu16
`
`scFV (VH/VL)-hngGFc (#82) bound to CD20 expressing 293 cells + a-hngG—FITC.
`
`[0047]
`
`Figure 41 shows analysis of anti-hngG-FITC negative control: 293-CD20 + 0t-
`
`hulgG—FITC.
`
`[0048]
`
`Figure 42 shows a flow cytometry profile of fusion protein CDl9-Dl+2-Leu16
`
`scFV (VL/VH)-hngGFc (#84) bound to CD20 expressing 293 cells + a-hngG—FITC.
`
`[0049]
`
`Figure 43 shows a flow cytometry profile of fusion protein CD22-D123 -Leul6
`
`scFV (VH/VL) (#65) bound to CD20 expressing 293 cells + OL-HlS-PE.
`
`[0050]
`
`Figure 44 shows detection control for Her2 - A431 cells + Trastuzumab-PE,
`
`showing the background level of binding (A431 cells are Her2-negative).
`
`[0051]
`
`Figure 45 shows analysis of A431 + fusion protein Her2—ECD—Paniturnumab scFV
`
`(VH/VL) (#57) + Trastuzumab—PE—conjugated.
`
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`[0052]
`
`Figure 46 shows analysis of A431 + fusion protein Her2—D4-Panitumumab scFv
`
`(VH/VL) (#58) + Trastuzumab-PE-conj ugated.
`
`[0053]
`
`Figure 47 shows IF Ny ELISA results for BT474 cells coated with indicated
`
`peptide and incubated with CD19 specific CAR-T at effector target ratio of 10: 1.
`
`[0054]
`
`Figure 48 shows IFNy ELISA results for BT474 cells coated with indicated
`
`peptide and incubated with CD19 specific CAR-T at effector target ratio of 1:1.
`
`[0055]
`
`Figure 49 shows summary XTT-cytotoxicity results for BT474 cells coated with
`
`indicated peptide and incubated with CD19 specific CAR-T at effector target ratio of 10: 1.
`
`[0056]
`
`Figure 50 shows IF Ny ELISA results for BT474 cells coated with indicated
`
`peptide and incubated with CD19 specific CAR-T at effector target ratio of 10: 1.
`
`[0057]
`
`Figure 51 shows IFNy ELISA results for BT474 cells coated with indicated
`
`peptide and incubated with CD19 specific CAR-T at effector target ratio of 1:1.
`
`[0058]
`
`Figures 52A—52C show exemplary Fc-based constructs.
`
`[0059]
`
`Figures 53A—53C show exemplary Fc—based bi—specific constructs.
`
`[0060]
`
`Figures 54A and 54B show exemplary Fc-based constructs that include an Fc Ig
`
`“swap”.
`
`[0061]
`
`Figures 55A and 55B show exemplary constructs in which a loops in one or both
`
`Fc CH3 domains is replaced.
`
`[0062]
`
`Figure 56 shows an exemplary construct with fusion of a masking moiety to
`
`constructs described in Figures 52B and 52C with a masking moiety fused to the N-terminus of
`
`the scFV.
`
`[0063]
`
`Figure 57 shows an exemplary construct with fusion of a masking moiety to
`
`constructs described in Figures 53B and 53C with the masking moiety fused to the N-terminus of
`
`the VH and/or VL on the VH/VL arm.
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`[0064]
`
`Figure 58 shows an exemplary construct with fusion of a masking moiety to
`
`construct described in Figure 54B with a masking moiety fused to the N—terminus of each heavy
`
`chain.
`
`[0065]
`
`Figure 59 shows an exemplary construct with fusion of a masking moiety to
`
`constructs described in Figures 55A and 55B with a masking moiety fused to the N—terminus of a
`
`heavy chain and/or scFv VH.
`
`[0066]
`
`Figures 60A—6OD show analysis of GFP expression from the CMV promoter-
`
`tGFP construct (#66) under resting or activated conditions.
`
`[0067]
`
`Figures 6lA—61D show analysis of GFP expression from the human CD69
`
`promoter—tGFP (#46) under resting or activated conditions.
`
`[0068]
`
`Figures 62A—62D show analysis of GFP expression from the human TNFalpha
`
`promoter-tGFP (#47) under resting or activated conditions.
`
`[0069]
`
`Figures 63A—63D show analysis of GFP expression from the human NFAT
`
`element x 6 promoter-tGFP (#49) under resting or activated conditions.
`
`[0070]
`
`Figures 64A—64D show analysis of expression of CD69 on the surface of cells
`
`under resting or activated conditions.
`
`[0071]
`
`Figures 65A—65C depict binding of CDl9-containing fusion proteins (#42, #43,
`
`#56, #82, #83, #91, #92, #93, #94) to an FMC63 -coated plate. Figure 65D shows titer
`
`determiniations for fusion proteins #82, #83, #91, and #92.
`
`[0072]
`
`Figures 66A— 66D show the capture of multiple fusion proteins by plate bound
`
`antigen and their detection by anti-His antibody coupled to HRP.
`
`[0073]
`
`Figures 67A and 67B show flow cytometry results of fusion protein CDl9-Dl+2-
`
`Leul6 scFv (VH/VL) (#83) bound to CD20 expressing 293 cells and labeled with anti-His-PE
`
`(67A) or anti-CD19 monoclonal antibody FMC63-PE (67B).
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`[0074]
`
`Figures 68A and 68B show flow cytometry results of fusion protein CDl9-Dl+2-
`
`Leul6 scFv (VH/VL)—hngGFc (#82) bound to CD20 expressing 293 cells and labeled with (1-
`
`hngG—FITC (68A) or FMC63-PE or anti-CD19 monoclonal antibody FMC63-PE (68B).
`
`[0075]
`
`Figures 69A—69D show results of IFNy ELISA for construct #83 fusion protein.
`
`Figure 69A: 24 hrs, 10:1 effector:target ratio; Figure 69B: 24 hrs, 2:1 effector:target ratio; Figure
`
`69C: 48 hrs, 10:1 effector:target ratio; Figure 69D: 48 hrs, 2:1 effector:target ratio.
`
`[007 6]
`
`Figure 70 show results of IFNy ELISA for fusion protein derived from the
`
`cotransfection of construct #33+ construct #4 at 24 hrs, 2:1 effector:target ratio.
`
`[007 7]
`
`Figures 71A and 71B show summary XTT—cytotoxicity results for fusion protein
`
`#83 and 293—CD20 cells. Figure 71A: 48 hrs, 10:1 effector:target ratio; Figure 71B 48 hrs, 2:1
`
`effector:target ratio.
`
`[0078]
`
`Figures 72A and 72B show summary XTT-cytotoxi city results for fusion protein
`
`derived from the cotransfecti on of construct #33 + construct #4 and A432] cells. Figure72A: 24
`
`hrs, 10:] effector:target ratio. Figure 72B: 24 hrs, 2:] effector:target ratio.
`
`[0079]
`
`Figures 73A and 73B show expression of HER2 and EGFR in transiently
`
`transfected 293T cells.
`
`[0080]
`
`Figures 74A—74D show fusion protein #43 binding to 293T-Her2 expressing cells.
`
`[0081]
`
`Figures 75A—75D show binding of fusion proteins #94, and #95 to 293T-Her2
`
`expressing cells.
`
`[0082]
`
`Figures 76A and 76B show binding of fusion protein #94 to 293T-EGFR
`
`expressing cells.
`
`[0083]
`
`Figures 77A and 77B show CAR19-mediated cytotoxicity redirected to HER2+
`
`cells by CAR19 T cell secretion of fusion protein encoded by construct #42.
`
`[0084]
`
`Figure 78 shows binding of a heteromeric fusion protein comprised of fusion
`
`proteins #29 and #103 to anti—CD19 antibody FMC63 detected by HRP—conjugated mouse IgG
`
`antibody.
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`[0085]
`
`domain.
`
`Figures 79A and 79B shows yeast surface display of wild-type CD19 extracellular
`
`[0086]
`
`Figure 80 shows antibody binding to yeast—displayed CD19 extracellular domain.
`
`[0087]
`
`Figure 81 shows diversified regions of the extracellular domain.
`
`[0088]
`
`Figure 82 demonstrates combinatorial CD19 libraries are effectively displayed on
`
`yeast surface and maintain antibody binding.
`
`[0089]
`
`Figures 83A and 83B demonstrate combinatorial CD19 libraries can be enriched
`
`for binding ligands to EGFR and HERZ.
`
`Definitions
`
`[0090]
`
`In order for the present invention to be more readily understood, certain terms are
`
`first defined below. Additional definitions for the following terms and other terms are set forth
`
`throughout the specification.
`
`[0091]
`
`Administration: As used herein, the term “administration” refers to the
`
`administration of a composition to a subject or system. Administration to an animal subject
`
`(e. g., to a human) may be by any appropriate route. For example, in some embodiments,
`
`administration may be bronchial (including by bronchial instillation), buccal, enteral,
`
`interdermal, intra-arterial, intradermal, intragastric, intramedullary, intramuscular, intranasal,
`
`intraperitoneal, intrathecal, intravenous, intraventricular, within a specific organ (e.g.,
`
`intrahepatic), mucosal, nasal, oral, rectal, subcutaneous, sublingual, topical, tracheal (including
`
`by intratracheal instillation), transdermal, vaginal and vitreal. In some embodiments,
`
`administration may be intratumoral or peritumoral. In some embodiments, administration may
`
`involve intermittent dosing. In some embodiments, administration may involve continuous
`
`dosing (e.g., perfusion) for at least a selected period of time.
`
`[0092]
`
`Adoptive cell therapy: As used herein, “adoptive cell therapy” or “ACT” involves
`
`the transfer of immune cells with antitumour activity into cancer patients. In some embodiments,
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`ACT is a treatment approach that involves the use of lymphocytes with antitumour activity, the
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`in vitro expansion of these cells to large numbers and their infusion into a cancer-bearing host.
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`[0093]
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`Agent: The term “agent” as used herein may refer to a compound or entity of any
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`chemical class including, for example, polypeptides, nucleic acids, saccharides, lipids, small
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`molecules, metals, or combinations thereof. As will be clear from context, in some
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`embodiments, an agent can be or comprise a cell or organism, or a fraction, extract, or
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`component thereof. In some embodiments, an agent is or comprises a natural product in that it is
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`found in and/or is obtained from nature.
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`In some embodiments, an agent is or comprises one or
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`more entities that is man—made in that it is designed, engineered, and/or produced through action
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`of the hand of man and/or is not found in nature.
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`In some embodiments, an agent may be
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`utilized in isolated or pure form; in some embodiments, an agent may be utilized in crude form.
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`In some embodiments, potential agents are provided as collections or libraries, for example that
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`may be screened to identify or characterize active agents within them. Some particular
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`embodiments of agents that may be utilized in accordance with the present invention include
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`small molecules, antibodies, antibody fragments, aptamers, nucleic acids (e. g, siRNAs, shRNAs,
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`DNA/RNA hybrids, antisense oligonucleotides, ribozymes), peptides, peptide mimetics, etc. In
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`some embodiments, an agent is or comprises a polymer.
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`In some embodiments, an agent is not a
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`polymer and/or is substantially free of any polymer.
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`In some embodiments, an agent contains at
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`least one polymeric moiety. In some embodiments, an agent lacks or is substantially free of any
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`polymeric moiety.
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`[0094]
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`Amelioration: As used herein, “amelioration” refers to prevention, reduction
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`and/or palliation of a state, or improvement of the state of a subject. Amelioration includes, but
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`does not require, complete recovery or complete prevention of a disease, disorder or condition.
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`[0095]
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`Amino acid: As used herein, term “amino acid,” in its broadest sense, refers to any
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`compound and/or substance that can be incorporated into a polypeptide chain.
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`In some
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`embodiments, an amino acid has the general structure H2N—C(H)(R)—COOH. In some
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`embodiments, an amino acid is a naturally occurring amino acid. In some embodiments, an
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`amino acid is a synthetic amino acid; in some embodiments, an amino acid is a d—amino acid; in
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`some embodiments, an amino acid is an l-amino acid. “Standard amino acid” refers to any of the
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`twenty standard l-amino acids commonly found in naturally occurring peptides. “Nonstandard
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`amino acid” refers to any amino acid, other than the standard amino acids, regardless of whether
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`it is prepared synthetically or obtained from a natural source. As used herein, “synthetic amino
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`acid” encompasses chemically modified amino acids, including but not limited to salts, amino
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`acid derivatives (such as amides), and/or substitutions. Amino acids, including carboxy- and/or
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`amino-terminal amino acids in peptides, can be modified by methylation, amidation, acetylation,
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`protecting groups, and/or substitution with other chemical groups that can change the peptide’s
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`circulating half-life without adversely affecting their activity. Amino acids may participate in a
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`disulfide bond. Amino acids may comprise one or posttranslational modifications, such as
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`association with one or more chemical entities (e. g, methyl groups, acetate groups, acetyl
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`groups, phosphate groups, formyl moieties, isoprenoid groups, sulfate groups, polyethylene
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`glycol moieties, lipid moieties, carbohydrate moieties, biotin moieties, etc.) The term “amino
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`acid” is used interchangeably with “amino acid residue,” and may refer to a free amino acid
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`and/or to an amino acid residue of a peptide. It will be apparent from the context in which the
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`term is used whether it refers to a free amino acid or a residue of a peptide.
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`[0096]
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`Antibody: As used herein, the term “antibody” refers to a polypeptide that
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`includes canonical immunoglobulin sequence elements sufficient to confer specific binding to a
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`particular target antigen. As is known in the art, intact antibodies as produced in nature are
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`approximately 150 kD tetrameric agents comprised of two identical heavy chain polypeptides
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`(about 50 kD each) and two identical light chain polypeptides (about 25 kD each) that associate
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`with each other into What is commonly referred to as a “Y—shaped” structure. Each heavy chain
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`is comprised of at least four domains (each about 110 amino acids long)— an amino—terminal
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`variable (VH) domain (located at the tips of the Y structure), followed by three constant
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`domains: CH1, CH2, and the carboxy—terminal CH3 (located at the base of the Y’s stem). A
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`short region, known as the “switch”, connects the heavy chain variable and constant regions.
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`The “hinge” connects CH2 and CH3 domains to the rest of the antibody. Two disulfide bonds in
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`this hinge region connect the two heavy chain polypeptides to one another in an intact antibody.
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`Each light chain is comprised of two domains — an amino-terminal variable (VL) domain,
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`followed by a carboxy-terminal constant (CL) domain, separated from one another by another
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`“switch”. Intact antibody tetramers are composed of two heavy chain-light chain dimers in
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`which the heavy and light chains are linked to one another by a single disulfide bond, two other
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`disulfrde bonds connect the heavy chain hinge regions to one another, so that the dimers are
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`connected to one another and the tetramer is formed. Naturally-produced antibodies are also
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`glycosylated, typically on the CH2 domain. Each domain in a natural antibody has a structure
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`characterized by an “immunoglobulin fold” formed from two beta sheets (e.g., 3-, 4-, or 5-
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`stranded sheets) packed against each other in a compressed antiparallel beta barrel. Each
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`variable domain contains three hypervariable loops known as “complement determining regions”
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`(CDRl, CDR2, and CDR3) and four somewhat invariant “framework” regions (FRl, FRZ, FR3,
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`and FR4). When natural antibodies fold, the FR regions form the beta sheets that provide the
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`structural framework for the domains, and the CDR loop regions from both the heavy and light
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`chains are brought together in three-dimensional space so that they create a single hypervariable
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`antigen binding site located at the tip of the Y structure. The Fc region of naturally-occurring
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`antibodies binds to elements of the complement system, and also to receptors on effector cells,
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`including for example effector cells that mediate cytotoxicity. As is known in the art, aff1nity
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`and/or other binding attributes of PC regions for Fc receptors can be modulated through
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`glycosylation or other modification. In some embodiments, antibodies produced and/or utilized
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`in accordance with the present disclosure include glycosylated Fc domains, including Fc
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`domains with modified or engineered such glycosylation. For purposes of the present disclosure,
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`in certain embodiments, any polypeptide or complex of polypeptides that includes suffrcient
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`immunoglobulin domain sequences as found in natural antibodies can be referred to and/or used
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`as an “antibody”, whether such polypeptide is naturally produced (e. g., generated by an organism
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`reacting to an antigen), or produced by recombinant engineering, chemical synthesis, or other
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`artifrcial system or methodology.
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`In some embodiments, an antibody is polyclonal, in some
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`embodiments, an antibody is monoclonal. In some embodiments, an antibody has constant
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`region sequences that are characteristic of mouse, rabbit, primate, or human antibodies. In some
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`embodiments, antibody sequence elements are fully human, or are humanized, primatized,
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`chimeric, etc, as is known in the art. Moreover, the term “antibody” as used herein, can refer in
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`appropriate embodiments (unless otherwise stated or clear from context) to any of the art-known
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`or developed constructs or formats for utilizing antibody structural and functional features in
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`alternative presentation. For example, in some embodiments, an antibody utilized in accordance
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`with the present disclosure is in a format selected from, but not limited to, intact IgG, IgE and
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`IgM, bi- or multi- specific antibodies (e.g., Zybodies®, etc), single chain Fvs, polypeptide-Fc
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`fusions, Fabs, cameloid antibodies, masked antibodies (e.g., Probodies®), Small Modular
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`ImmunoPharmaceuticals (“SMIPsTM”), single chain or Tandem diabodies (TandAb®), VHHs,
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`Anticalins®, Nanobodies®, minibodies, BiTE®s, ankyrin repeat proteins or DARPINs®,
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`Avimers®, a DART, a TCR-like antibody, Adnectins®, Affilins®, Trans-bodies®, Afflbodies®,
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`a TrimerX®, MicroProteins, Fynomers®, Centyrins®, and a KALBITOR®.
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`In some
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`embodiments, an antibody may lack a covalent modif1cation (e.g., attachment of a glycan) that it
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`would have if produced naturally. In some embodiments, an antibody may contain a covalent
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`modification (e.g., attachment of a glycan, a payload (e.g., a detectable moiety, a therapeutic
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`moiety, a catalytic moiety, etc.), or other pendant group (e.g., poly-ethylene glycol, etc.)).
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`[0097]
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`Antibody-Dependent Cellular Cytotoxicily: As used herein, the term “antibody—
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`dependent cellular cytotoxicity” or “ADCC” refers to a phenomenon in which target cells bound
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`by antibody are killed by immune effector cells. Without wishing to be bound by any particular
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`theory, ADCC is typically understood to involve Fc receptor (FcR)—bearing effector cells can
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`recognizing and subsequently killing antibody—coated target cells (e. g, cells that express on their
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`surface specific antigens to which an antibody is bound). Effector cells that mediate ADCC can
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`include immune cells, including but not limited to one or more of natural killer (NK) cells,
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`macrophage, neutrophils, eosinophils.
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`[0098]
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`Antibody Fragment: As used herein, an “antibody fragment” includes a portion of
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`an intact antibody, such as, for example, the antigen-binding or variable region of an antibody.
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`Examples of antibody fragments include Fab, Fab’, F(ab’)2, and Fv fragments; triabodies;
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`tetrabodies; linear antibodies; single-chain antibody molecules; and multi-specific antibodies
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`formed from antibody fragments. For example, antibody fragments include isolated fragments,
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`“Fv” fragments (consisting of the variable regions of the heavy and light chains), recombinant
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`single chain polypeptide molecules in which light and heavy chain variable regions are
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`connected by a peptide linker (“scFv proteins”), recombinant single domain antibodies consisting
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`of a variable region of an antibody heavy chain (e. g., VHH), and minimal recognition units
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`consisting of the amino acid residues that mimic a hypervariable region (e.g., a hypervariable
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`region of a heavy chain variable region (VH), a hypervariable region of a light chain variable
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`region (VL), one or more CDR domains within the VH, and/or one or more CDR domains within
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`the VL). In many embodiments, an antibody fragment contains sufficient sequence of the parent
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`antibody of which it is a fragment that it binds to the same antigen as does the parent antibody, in
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`some embodiments, a fragment binds to the antig