Anti-IL-27 antibodies and uses thereof

Abstract

The present disclosure relates to anti-IL-27 antibodies and antigen binding portions thereof. The disclosure also relates to methods of treating or ameliorating one or more symptoms of a disease, such as cancer, by administering an antibody or antigen-binding portion thereof. The present disclosure also relates to methods for detecting IL-27, e.g., in a subject or sample.

Description

[0001] This application is a divisional application of the invention application with an application date of March 22, 2019, a Chinese application number of 201980027625.8, and an invention title of "Anti-IL-27 Antibodies and Their Uses".

[0002] Cross-reference to related applications

[0003] This application claims the benefit of U.S. Provisional Application No. 62 / 646,496, filed on March 22, 2018, the entire content of which is incorporated herein by reference. Technical field

[0004] The present disclosure generally relates to compositions and methods for modulating IL-27 signaling. More specifically, the present disclosure relates to immunogenic compositions (e.g., antibodies, antibody fragments, etc.) that bind to IL-27 and modulate IL-27 signaling. Background art

[0005] In recent years, increasing evidence has shown that the immune system is an important barrier to tumor formation and development. The principle that there are naturally occurring T cells with anti-tumor potential or activity in cancer patients rationalizes the development of immunotherapeutic methods in oncology. Immune cells, such as T cells, macrophages, and natural killer cells, can exhibit anti-tumor activity and effectively control the occurrence and growth of malignant tumors. Tumor-specific or related antigens can induce immune cells to recognize and eliminate malignant tumors (Chen & Mellman, (2013) Immunity 39(1): 1-10). Despite the presence of tumor-specific immune responses, malignant tumors often evade or avoid immune attack through various immune regulatory mechanisms, resulting in the inability to control the occurrence and progression of tumors (Motz & Coukos, (2013) Immunity 39(1): 61-730). In fact, an emerging hallmark of cancer is the utilization of these immune regulatory mechanisms and the failure of anti-tumor immune responses, leading to tumor escape and evasion of immune killing (Hanahan and Weinberg (2011) Cell 144(5): 646-674).

[0006] IL-27 is a heterodimeric cytokine composed of two subunits (EBI3 and IL-27p28). IL-27 is structurally related to the IL-12 and IL-6 cytokine families. IL-27 binds to a heterodimeric receptor composed of IL-27Rα (WSX1) and the gp130 chain and mediates signaling through said heterodimeric receptor, which mainly mediates signal transduction through STAT1 and STAT3. Initial reports described IL-27 as an immune-enhancing cytokine that supports CD4+ T cell proliferation, helper T (Th)1 cell differentiation, and IFN-γ production, usually acting in concert with IL-12. Subsequent studies have shown that IL-27 exhibits complex immunomodulatory functions, producing pro-inflammatory or anti-inflammatory effects depending on the biological context and experimental model used. IL-27 can drive the expression of different immunomodulatory molecules in human cancer cells, which can support local perturbations in the immune response in vivo (Fabbi et al., (2017) Mediators Inflamm 3958069. Published online February 1, 2017 doi:10.1155 / 2017 / 3958069, and references therein).

[0007] Despite significant advances in cancer treatment and management, there remains a continuing need for new and effective therapies for treating and managing cancer. SUMMARY OF THE INVENTION

[0008] Disclosed herein are antibodies or antigen-binding portions thereof that specifically bind to and antagonize human IL-27 (interleukin 27) with high affinity and specificity. Also provided are nucleic acid molecules, expression vectors, host cells, and methods for preparing the antibody molecules that encode the antibody molecules. Also provided are pharmaceutical compositions comprising the antibody molecules. The anti-IL-27 antibodies or antigen-binding portions thereof disclosed herein can be used (either alone or in combination with other therapeutic agents or procedures) to treat, prevent, and / or diagnose conditions, including immune disorders and cancer. Accordingly, disclosed herein are compositions and methods for using anti-IL-27 antibody molecules to treat and / or diagnose various diseases, including cancer and immune disorders.

[0009] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof exhibits at least one or more of the following characteristics: (i) having an equilibrium dissociation constant (K D(i) binds to human IL-27; (ii) blocks the binding of IL-27 to the IL-27 receptor; (iii) inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells; (iv) inhibits or reduces the inhibition of CD161 expression in cells; (v) inhibits or reduces the expression of PD-L1 and / or TIM-3 in cells; (vi) induces or increases the secretion of one or more cytokines mediated by PD-1 from cells; and (vii) a combination of (i) to (vi).

[0010] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof has an equilibrium dissociation constant (K D ) that binds to human IL-27.

[0011] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof binds to recombinant human IL-27 or murine IL-27.

[0012] In one aspect, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof has heavy and light chain CDRs, and the heavy and light chain CDRs are: (i) heavy chain CDR1 is N-GFTFXXXX-C (SEQ ID NO:408), heavy chain CDR2 is N-ISSSXXYI-C (SEQ ID NO:409), and heavy chain CDR3 sequence is SEQ ID NO:163; light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:169, 170, and 171, respectively; or (ii) heavy chain CDR1 is N-FTFXXXXMN-C (SEQ ID NO:410), heavy chain CDR2 is N-XISSSXXYIXYADSVKG-C (SEQ ID NO:411), and heavy chain CDR3 sequence is SEQ ID NO:166; light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:172, 173, and 174, respectively.

[0013] In one embodiment, heavy chain CDR1 is N-GFTF[S / A / R][S / R][T / Y][G / S]-C (SEQ ID NO:412) and heavy chain CDR2 is N-ISSS[S / G][S / A]YI-C (SEQ ID NO:413); or heavy chain CDR1 is N-FTF[S / A / R][S / R][T / Y][G / S]MN-C (SEQ ID NO:414) and heavy chain CDR2 is N-[G / S]ISSS[S / G][S / A]YI[L / Y]YADSVKG-C (SEQ ID NO:415).

[0014] In another embodiment, the corresponding heavy and light chain CDRs are: (i) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 161, 162, and 163, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 169, 170, and 171, respectively; (ii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 164, 165, and 166, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 172, 173, and 174, respectively; (iii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 73, 74, and 75, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 81, 82, and 83, respectively; (iv) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 76, 77, and 78, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 84, 85, and 86, respectively; (v) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 95, 96, and 97, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 103, 104, and 105, respectively; (vi) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 98, 99, and 100, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 106, 107, and 108, respectively; (vii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 117, 118, and 119, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 125, 126, and 127, respectively; (viii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 120, 121, and 122, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 128, 129, and 130, respectively; (ix) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 139, 140, and 141, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 147, 148, and 149, respectively; (x) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 142, 143, and 144, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 150, 151, and 152, respectively;(xi) The heavy chain CDR1, CDR2 and CDR3 sequences are SEQ ID NO:51, 52 and 53 respectively, and the light chain CDR1, CDR2 and CDR3 sequences are SEQ ID NO:59, 60 and 61 respectively; or (xii) The heavy chain CDR1, CDR2 and CDR3 sequences are SEQ ID NO:54, 55 and 56 respectively, and the light chain CDR1, CDR2 and CDR3 sequences are SEQ ID NO:62, 63 and 64 respectively.;

[0015] Another aspect of the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, and the heavy and light chain CDRs are respectively: (i) The heavy chain CDR1 is N-GGSFSXYX-C (SEQ ID NO:416), the heavy chain CDR2 is N-IDXSGXT-C (SEQ ID NO:417) and the heavy chain CDR3 is N-ARXXXYY[X] n＝0-1 DSS[X] n＝4-6 DX-C (SEQ ID NO:418); and the light chain CDR1 is N-QXXSXY-C (SEQ ID NO:419), the light chain CDR2 is N-DXS-C (SEQ ID NO:420) and the light chain CDR3 is N-QQXXDXPIT-C (SEQ ID NO:421); or (ii) The heavy chain CDR1 is N-GSFSXYXWS-C (SEQ ID NO:422), the heavy chain CDR2 is N-SIDXSGXTXYNPSLKS-C (SEQ ID NO:423) and the heavy chain CDR3 is N-ARXXXYY[X] n＝0-1 DSS[X] n＝4-6 DX-C (SEQ ID NO:418); and the light chain CDR1 is N-XASQXXSXYLX-C (SEQ ID NO:424), the light chain CDR2 is N-DXSNXXT-C (SEQ ID NO:425) and the light chain CDR3 is N-QQXXDXPIT-C (SEQ ID NO:421).

[0016] In certain embodiments, (i) separately, heavy chain CDR1 is N-GGSFS[R / D]Y[E / Y]-C (SEQ ID NO:426), heavy chain CDR2 is N-ID[W / Y]SG[I / S]T-C (SEQ ID NO:427), and heavy chain CDR3 is N-AR[D / L][P / G][M / V]YY[- / Y]DSS[VSTGSV / DLGF]D[V / I]-C (SEQ ID NO:428); and light chain CDR1 is N-Q[S / D][V / I]S[S / N]Y-C (SEQ ID NO:429), light chain CDR2 is N-D[S / A]S-C (SEQ ID NO:430), and light chain CDR3 is N-QQ[D / Y][S / D]D[H / L]PIT-C (SEQ ID NO:431); or (ii) separately, heavy chain CDR1 is N-GSFS[R / D]Y[E / Y]WS-C (SEQ ID NO:432), heavy chain CDR2 is N-SID[W / Y]SG[I / S]T[N / E]YNPSLKS-C (SEQ ID NO:433), and heavy chain CDR3 is N-AR[D / L][P / G][M / V]YY[- / Y]DSS[VSTGSV / DLGF]D[V / I]-C (SEQ ID NO:428); and light chain CDR1 is N-[Q / R]ASQ[S / D][V / I]S[S / N]YL[N / A]-C (SEQ ID NO:434), light chain CDR2 is N-D[S / A]SN[R / L][A / E]T-C (SEQ ID NO:435), and light chain CDR3 is N-QQ[D / Y][S / D]D[H / L]PIT-C (SEQ ID NO:431).

[0017] In some embodiments, (i) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:229, 230, and 231, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:237, 238, and 239, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:232, 233, and 234, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:240, 241, and 242, respectively.

[0018] In certain embodiments, (i) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 251, 252, and 253, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 259, 260, and 261, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 254, 255, and 256, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO: 262, 263, and 264, respectively.

[0019] Another aspect of the disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs that are the heavy chain CDR1, CDR2, and CDR3 sequences of SEQ ID NO: 23, 24, and 25, respectively, and the CDR1, CDR2, and CDR3 sequences of SEQ ID NO: 26, 27, and 28, respectively.

[0020] Another aspect of the disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs that are: (i) the heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 339, 340, and 341, respectively, and the light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 347, 348, and 349, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 342, 343, and 344, respectively, and the light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 350, 351, and 352, respectively.

[0021] Another aspect of the disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs that are: (i) the heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 185, 186, and 187, respectively, and the light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 193, 194, and 195, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 188, 189, and 190, respectively, and the light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 196, 197, and 198, respectively.

[0022] One aspect of the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, and the heavy and light chain CDRs are: (i) heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 207, 208, and 209, respectively, and light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 215, 216, and 217, respectively; or (ii) heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 210, 211, and 212, respectively, and light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 218, 219, and 220, respectively.

[0023] Another aspect of the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, and the heavy and light chain CDRs are: (i) heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 273, 274, and 275, respectively, and light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 281, 282, and 283, respectively; or (ii) heavy chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 276, 277, and 278, respectively, and light chain CDR1, CDR2, and CDR3 sequences SEQ ID NO: 284, 285, and 286, respectively.

[0024] One aspect of the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, and the heavy and light chain CDRs are: (i) respectively, heavy chain CDR1 is N-GFTFSSYG-C (SEQ ID NO: 361), heavy chain CDR2 is N-IXXDGSXK-C (SEQ ID NO: 436) and heavy chain CDR3 is N-ARXAP[X] n＝3-8 DV-C (SEQ ID NO: 437); and light chain CDR1 is N-QSXSSY-C (SEQ ID NO: 438), light chain CDR2 is N-XXS-C (SEQ ID NO: 439) and light chain CDR3 is N-QQXXXXP[X] n＝0-1T-C (SEQ ID NO:440); or (ii) separately, heavy chain CDR1 is N-FTFSSYGMX-C (SEQ ID NO:441), heavy chain CDR2 is N-XIXXDGSXKYYXDSVKG-C (SEQ ID NO:442) and heavy chain CDR3 is N-ARXAP[X] n＝3-8 DV-C (SEQ ID NO:437); and light chain CDR1 is N-RASQSXSSYLX-C (SEQ ID NO:443), light chain CDR2 is N-[X] n＝1-2 SS[X] n＝3-4 -C (SEQ ID NO:444) and light chain CDR3 is N-QQXXXXP[X] n＝0-1 T-C (SEQ ID NO:440).

[0025] In certain embodiments, (i) separately, heavy chain CDR1 is N-GFTFSSYG-C (SEQ ID NO:361), heavy chain CDR2 is N-I[K / W][Q / Y]DGS[E / N]K-C (SEQ ID NO:445) and heavy chain CDR3 is N-AR[D / G]AP[WDIYDYYM / EY V]DV-C (SEQ ID NO:446); and light chain CDR1 is N-QS[I / V]SSY-C (SEQ ID NO:447), light chain CDR2 is N-[A / D][A / S]S-C (SEQ ID NO:448) and light chain CDR3 is N-QQ[S / Y][Y / S][V / L][P / Y]P[W / -]T-C (SEQ ID NO:449); or (ii) heavy chain CDR1 is N-FTFSSYGM[S / H]-C (SEQ IDNO:450); heavy chain CDR2 is N-[N / V]I[K / W][Q / Y]DGS[E / N]KYY[V / A]DSVKG-C (SEQ ID NO:451) and heavy chain CDR3 is N-AR[D / G]AP[WDI YDYYM / EYV]DV-C (SEQ ID NO:446); and light chain CDR1 is N-RASQS[I / V]SSYL[N / A]-C (SEQ ID NO:452), light chain CDR2 is N-[AA / D]SS[LQS / NRAT]-C (SEQ ID NO:453) and light chain CDR3 is N-QQ[S / Y][Y / S][V / L][P / Y]P[W / -]T-C (SEQ ID NO:449).

[0026] In some embodiments, (i) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:361, 362, and 363, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:369, 370, and 371, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:364, 365, and 366, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:372, 373, and 374, respectively.

[0027] In one embodiment, (i) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:383, 384, and 385, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:391, 392, and 393, respectively; or (ii) the heavy chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:386, 387, and 388, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are SEQ ID NO:394, 395, and 396, respectively.

[0028] Another aspect of the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, and the heavy and light chain CDRs are: (i) respectively, heavy chain CDR1 is N-GFTFXXXX-C (SEQ ID NO:408), heavy chain CDR2 is N-IXXXXXXX-C (SEQ ID NO:456), and heavy chain CDR3 is N-AR[X] n＝6-15 DX-C (SEQ ID NO:458); and light chain CDR1 is N-QS[X] n＝1-3 SS[X] n＝0-4 Y-C (SEQ ID NO:460), light chain CDR2 is N-XXS-C (SEQ ID NO:462), and light chain CDR3 is N-QQXXXXP[X] n＝0-1 T-C (SEQ ID NO:464); or (ii) respectively, heavy chain CDR1 is N-FTFXXXXMX-C (SEQ ID NO:466), heavy chain CDR2 is N-XIXXXXXXXXYXDSVKG-C (SEQ ID NO:468), and heavy chain CDR3 is N-AR[X] n＝6-15 DX-C (SEQ ID NO:470); and light chain CDR1 is N-RASQSXSSYLX-C (SEQ IDNO:472), light chain CDR2 is N-[X] n＝1-2 S[X]n＝4-5 -C (SEQ ID NO:474) and the light chain CDR3 is N-QQXXXXP[X] n＝0-1 T-C (SEQ ID NO:476).

[0029] In certain embodiments, (i) separately, the heavy chain CDR1 is N-GFTF[S / A / R][S / R][T / Y][G / S]-C (SEQ ID NO:454), the heavy chain CDR2 is N-I[S / K / W][S / Q / Y][S / D][S / G][S / A][Y / E / N][I / K]-C (SEQ ID NO:455) and the heavy chain CDR3 is N-AR[DGGRTSYTATAHNWF / DAPWDIYDYYM / GAPEYV]D[P / V]-C (SEQ ID NO:457); and the light chain CDR1 is N-QS[VLF / I / V]SS[NNKN / -]Y-C (SEQ ID NO:459), the light chain CDR2 is N-[W / A / D][A / S]S-C (SEQ ID NO:461) and the light chain CDR3 is N-QQ[H / S / Y][A / Y / S][S / V / L][A / P / Y]P[P / W / -]T-C (SEQ ID NO:463); or (ii) separately, the heavy chain CDR1 is N-FTF[S / A / R][S / R][T / Y][G / S]M[N / S / H]-C (SEQ ID NO:465), the heavy chain CDR2 is N-[G / S / N / V]I[S / K / W][S / Q / Y][S / D][S / G][S / A][Y / E / N][I / K][L / Y]Y[V / A]DSVKG-C (SEQ ID NO:467) and the heavy chain CDR3 is N-AR[D / G][GGRTSYTATAHNWF / APWDIYDYYM / APEYV]D[P / V]-C (SEQ ID NO:469); and the light chain CDR1 is N-RASQS[I / V]SSYL[N / A]-C (SEQ ID NO:471), the light chain CDR2 is N-[WA / AA / D]S[TRES / SLQS / SNRAT]-C (SEQ ID NO:473) and the light chain CDR3 is N-QQ[H / S / Y][A / Y / S][S / V / L][A / P / Y]P[P / W / -]T-C (SEQ ID NO:475).

[0030] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells. In some embodiments, the cells are immune cells. In some embodiments, the cells are cancer cells.

[0031] In some embodiments, the isolated monoclonal antibody or antigen binding portion thereof inhibits or reduces the inhibition of CD161 expression in a cell (eg, improves or alleviates the inhibition of CD161 expression in a cell). In some embodiments, the cell is an immune cell.

[0032] In some embodiments, the isolated monoclonal antibody or its antigen binding portion inhibits or reduces IL-27-mediated PD-L1 and / or TIM-3 expression in cells. In some embodiments, PD-L1 expression is inhibited or reduced. In some embodiments, TIM-3 expression is inhibited or reduced. In some embodiments, both PD-L1 expression and TIM-3 expression are reduced. In some embodiments, the cell is an immune cell.

[0033] In some embodiments, the isolated monoclonal antibody or antigen binding portion thereof induces or increases PD-1 mediated secretion of one or more cytokines from cells. In some embodiments, the one or more cytokines are TNFα. In some embodiments, the one or more cytokines are IL-6. In some embodiments, the one or more cytokines are TNFα and IL-6. In some embodiments, the cells are immune cells.

[0034] In some embodiments, the isolated monoclonal antibody or its antigen-binding portion is selected from the group consisting of: IgG1, IgG2, IgG3, IgG4, IgM, IgA1, IgA2, IgD and IgE antibodies. In some embodiments, the antibody is an IgG1 antibody or an IgG4 antibody. In some embodiments, the antibody comprises a wild-type IgG1 heavy chain constant region. In some embodiments, the antibody comprises a wild-type IgG4 heavy chain constant region. In some embodiments, the antibody comprises an Fc domain containing at least one mutation. In some embodiments, the antibody comprises a mutant IgG1 heavy chain constant region. In some embodiments, the antibody comprises a mutant IgG4 heavy chain constant region. In some embodiments, the mutant IgG4 heavy chain constant region comprises any one or a combination of substitutions S228P, L235E, L235A according to EU numbering.

[0035] In some embodiments, the disclosure provides an isolated monoclonal antibody, or antigen binding portion thereof, that binds to substantially the same epitope on IL-27 as the antibody, or antigen binding portion thereof, according to any of the preceding embodiments.

[0036] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that binds to at least one of the amino acid residues of IL-27 that is bound by an antibody or antigen-binding portion thereof according to any one of the foregoing embodiments.

[0037] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof, wherein mutation of the epitope on IL-27 that is bound by the antibody or antigen-binding portion thereof inhibits, reduces, or blocks binding to the antibody or antigen-binding portion thereof and to an antibody or antigen-binding portion thereof according to any one of the foregoing embodiments.

[0038] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that binds to an epitope on IL-27, wherein the epitope is the same as or similar to the epitope bound by the antibody molecule described in Table 12.

[0039] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs selected from the group consisting of:

[0040] (i) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 51, 52, and 53, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 59, 60, and 61, respectively;

[0041] (ii) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 73, 74, and 75, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 81, 82, and 83, respectively;

[0042] (iii) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 95, 96, and 97, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 103, 104, and 105, respectively;

[0043] (iv) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 117, 118, and 119, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 125, 126, and 127, respectively;

[0044] (v) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 139, 140, and 141 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 147, 148, and 149 respectively;

[0045] (vi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 161, 162, and 163 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 169, 170, and 171 respectively;

[0046] (vii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 185, 186, and 187 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 193, 194, and 195 respectively;

[0047] (viii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 207, 208, and 209 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 215, 216, and 217 respectively;

[0048] (ix) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 229, 230, and 231 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 237, 238, and 239 respectively;

[0049] (x) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 251, 252, and 253 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 259, 260, and 261 respectively;

[0050] (xi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 273, 274, and 275 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 281, 282, and 283 respectively;

[0051] (xii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 295, 296, and 297 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 303, 304, and 305 respectively;

[0052] (xiii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 317, 318, and 319, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 325, 326, and 327, respectively;

[0053] (xiv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 339, 340, and 341, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 347, 348, and 349, respectively;

[0054] (xv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 361, 362, and 363, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 369, 370, and 371, respectively; and

[0055] (xvi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 383, 384, and 385, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 391, 392, and 393, respectively.

[0056] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or its antigen-binding portion comprises heavy and light chain CDRs, wherein the heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 161, 162, and 163, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 169, 170, and 171, respectively.

[0057] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or its antigen-binding portion comprises heavy and light chain CDRs selected from the group consisting of:

[0058] (i) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 54, 55, and 56, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 62, 63, and 64, respectively;

[0059] (ii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:76, 77, and 78 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:84, 85, and 86 respectively;

[0060] (iii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:98, 99, and 100 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:106, 107, and 108 respectively;

[0061] (iv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:120, 121, and 122 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:128, 129, and 130 respectively;

[0062] (v) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:142, 143, and 144 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:150, 151, and 152 respectively;

[0063] (vi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:164, 165, and 166 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:172, 173, and 174 respectively;

[0064] (vii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:188, 189, and 190 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:196, 197, and 198 respectively;

[0065] (viii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:210, 211, and 212 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:218, 219, and 220 respectively;

[0066] (ix) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:232, 233, and 234 respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO:240, 241, and 242 respectively;

[0067] (x) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 254, 255, and 256, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 262, 263, and 264, respectively;

[0068] (xi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 276, 277, and 278, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 284, 285, and 286, respectively;

[0069] (xii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 298, 299, and 300, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 306, 307, and 308, respectively;

[0070] (xiii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 320, 321, and 322, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 328, 329, and 330, respectively;

[0071] (xiv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 342, 343, and 344, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 350, 351, and 352, respectively;

[0072] (xv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 364, 365, and 366, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 372, 373, and 374, respectively; and

[0073] (xvi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 386, 387, and 388, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 394, 395, and 396, respectively.

[0074] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, wherein the heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NOs: 164, 165, and 166, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NOs: 172, 173, and 174, respectively.

[0075] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, wherein the amino acid sequences of the heavy chain CDR1, CDR2, and CDR3 are SYSMS (SEQ ID NO: 23), YISYDGGSAYYPDT VKG (SEQ ID NO: 24), and HGDYDDDDAMDY (SEQ ID NO: 25), respectively, and wherein the amino acid sequences of the light chain CDR1, CDR2, and CDR3 are RA SENIYSYLA (SEQ ID NO: 26), NAETLTE (SEQ ID NO: 27), and QHHYGTPLT (SEQ ID NO: 28), respectively.

[0076] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 57, 79, 101, 123, 145, 167, 191, 213, 235, 257, 279, 301, 323, 345, 367, and 389; and wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 65, 87, 109, 131, 153, 175, 199, 221, 243, 265, 287, 309, 331, 353, 375, and 397.

[0077] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, the heavy and light chain variable regions comprising amino acid sequences selected from the groups consisting of:

[0078] (i) SEQ ID NO: 57 and 65;

[0079] (ii) SEQ ID NO: 79 and 87;

[0080] (iii) SEQ ID NO: 101 and 109;

[0081] (iv) SEQ ID NO: 123 and 131;

[0082] (v) SEQ ID NO: 145 and 153;

[0083] (vi) SEQ ID NO: 167 and 175;

[0084] (vii) SEQ ID NO: 191 and 199;

[0085] (viii) SEQ ID NO: 213 and 221;

[0086] (ix) SEQ ID NO: 235 and 243;

[0087] (x) SEQ ID NO: 257 and 265;

[0088] (xi) SEQ ID NO: 279 and 287;

[0089] (xii) SEQ ID NO: 301 and 309;

[0090] (xiii) SEQ ID NO: 323 and 331;

[0091] (xiv) SEQ ID NO: 345 and 353;

[0092] (xv) SEQ ID NO: 367 and 375; and

[0093] (xvi) SEQ ID NO: 389 and 397.

[0094] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, wherein the heavy chain variable region comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 57, 79, 101, 123, 145, 167, 191, 213, 235, 257, 279, 301, 323, 345, 367, and 389; and wherein the light chain variable region comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 65, 87, 109, 131, 153, 175, 199, 221, 243, 265, 287, 309, 331, 353, 375, and 397.

[0095] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions that respectively comprise amino acid sequences having at least 90% identity to amino acid sequences selected from the group consisting of:

[0096] (i) SEQ ID NOs: 57 and 65;

[0097] (ii) SEQ ID NOs: 79 and 87;

[0098] (iii) SEQ ID NOs: 101 and 109;

[0099] (iv) SEQ ID NOs: 123 and 131;

[0100] (v) SEQ ID NOs: 145 and 153;

[0101] (vi) SEQ ID NOs: 167 and 175;

[0102] (vii) SEQ ID NOs: 191 and 199;

[0103] (viii) SEQ ID NOs: 213 and 221;

[0104] (ix) SEQ ID NOs: 235 and 243;

[0105] (x) SEQ ID NOs: 257 and 265;

[0106] (xi) SEQ ID NOs: 279 and 287;

[0107] (xii) SEQ ID NO: 301 and 309;

[0108] (xiii) SEQ ID NO: 323 and 331;

[0109] (xiv) SEQ ID NO: 345 and 353;

[0110] (xv) SEQ ID NO: 367 and 375; and

[0111] (xvi) SEQ ID NO: 389 and 397.

[0112] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, and the heavy and light chain variable regions comprise the amino acid sequences set forth in SEQ ID NOs: 167 and 175, respectively.

[0113] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chains, and the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0114] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chains, wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and wherein the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0115] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and wherein the light chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0116] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 71, 93, 115, 137, 159, 181, 205, 227, 249, 271, 293, 315, 337, 359, 381, and 403; and wherein the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0117] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 71, 93, 115, 137, 159, 181, 205, 227, 249, 271, 293, 315, 337, 359, 381, and 403; and wherein the light chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0118] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain each comprise an amino acid sequence selected from the group consisting of:

[0119] (i) SEQ ID NO: 67 and 69;

[0120] (ii) SEQ ID NO: 89 and 91;

[0121] (iii) SEQ ID NO: 111 and 113;

[0122] (iv) SEQ ID NO: 133 and 135;

[0123] (v) SEQ ID NO: 155 and 157;

[0124] (vi) SEQ ID NO: 177 and 179;

[0125] (vii) SEQ ID NO: 201 and 203;

[0126] (viii) SEQ ID NO: 223 and 225;

[0127] (ix) SEQ ID NO: 245 and 247;

[0128] (x) SEQ ID NO: 267 and 269;

[0129] (xi) SEQ ID NO: 289 and 291;

[0130] (xii) SEQ ID NO: 311 and 313;

[0131] (xiii) SEQ ID NO: 333 and 335;

[0132] (xiv) SEQ ID NO: 355 and 357;

[0133] (xv) SEQ ID NO: 377 and 379; and

[0134] (xvi) SEQ ID NO: 399 and 401.

[0135] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain each comprise an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of:

[0136] (i) SEQ ID NO: 67 and 69;

[0137] (ii) SEQ ID NO: 89 and 91;

[0138] (iii) SEQ ID NO: 111 and 113;

[0139] (iv) SEQ ID NO: 133 and 135;

[0140] (v) SEQ ID NO: 155 and 157;

[0141] (vi) SEQ ID NO: 177 and 179;

[0142] (vii) SEQ ID NO: 201 and 203;

[0143] (viii) SEQ ID NO: 223 and 225;

[0144] (ix) SEQ ID NO: 245 and 247;

[0145] (x) SEQ ID NO: 267 and 269;

[0146] (xi) SEQ ID NO: 289 and 291;

[0147] (xii) SEQ ID NO: 311 and 313;

[0148] (xiii) SEQ ID NO: 333 and 335;

[0149] (xiv) SEQ ID NO: 355 and 357;

[0150] (xv) SEQ ID NO: 377 and 379; and

[0151] (xvi) SEQ ID NO: 399 and 401.

[0152] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain respectively comprise an amino acid sequence selected from the group consisting of:

[0153] (i) SEQ ID NO:71 and 69;

[0154] (ii) SEQ ID NO:93 and 91;

[0155] (iii) SEQ ID NO:115 and 113;

[0156] (iv) SEQ ID NO:137 and 135;

[0157] (v) SEQ ID NO:159 and 157;

[0158] (vi) SEQ ID NO:181 and 179;

[0159] (vii) SEQ ID NO:205 and 203;

[0160] (viii) SEQ ID NO:227 and 225;

[0161] (ix) SEQ ID NO:249 and 247;

[0162] (x) SEQ ID NO:271 and 269;

[0163] (xi) SEQ ID NO:293 and 291;

[0164] (xii) SEQ ID NO:315 and 313;

[0165] (xiii) SEQ ID NO:337 and 335;

[0166] (xiv) SEQ ID NO:359 and 357;

[0167] (xv) SEQ ID NO:381 and 379; and

[0168] (xvi) SEQ ID NO:403 and 401.

[0169] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain each comprise an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of:

[0170] (i) SEQ ID NO:71 and 69;

[0171] (ii) SEQ ID NO:93 and 91;

[0172] (iii) SEQ ID NO:115 and 113;

[0173] (iv) SEQ ID NO:137 and 135;

[0174] (v) SEQ ID NO:159 and 157;

[0175] (vi) SEQ ID NO:181 and 179;

[0176] (vii) SEQ ID NO:205 and 203;

[0177] (viii) SEQ ID NO:227 and 225;

[0178] (ix) SEQ ID NO:249 and 247;

[0179] (x) SEQ ID NO:271 and 269;

[0180] (xi) SEQ ID NO:293 and 291;

[0181] (xii) SEQ ID NO:315 and 313;

[0182] (xiii) SEQ ID NO:337 and 335;

[0183] (xiv) SEQ ID NO:359 and 357;

[0184] (xv) SEQ ID NO:381 and 379; and

[0185] (xvi) SEQ ID NO:403 and 401.

[0186] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise the amino acid sequences set forth in SEQ ID NOs: 177 and 179, respectively.

[0187] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise amino acid sequences that have at least 90% identity to the amino acid sequences set forth in SEQ ID NOs: 177 and 179, respectively.

[0188] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise the amino acid sequences set forth in SEQ ID NOs: 181 and 179, respectively.

[0189] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise amino acid sequences that have at least 90% identity to the amino acid sequences set forth in SEQ ID NOs: 181 and 179, respectively.

[0190] In some embodiments, the present disclosure provides a method of inhibiting or reducing STAT1 and / or STAT3 phosphorylation in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or antigen-binding portion thereof inhibits or reduces STAT1 and / or STAT3 phosphorylation in the cell.

[0191] In some embodiments, the present disclosure provides a method of inhibiting or reducing IL-27-mediated inhibition of CD161 expression in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided herein, wherein the antibody or antigen-binding portion thereof inhibits or reduces the inhibition of CD161 expression in the cell.

[0192] In some embodiments, the present disclosure provides a method of inhibiting or reducing IL-27-mediated PD-L1 and / or TIM-3 expression in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or antigen-binding portion thereof inhibits or reduces the expression of PD-L1 and / or TIM-3 in the cell.

[0193] In some embodiments, the present disclosure provides a method of inducing or enhancing the secretion of one or more cytokines from a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or its antigen-binding portion induces or enhances the secretion of one or more PD-1-mediated cytokines from the cell.

[0194] In some embodiments, the present disclosure provides a method of stimulating an immune response in a subject, the method comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure that specifically binds to and antagonizes human IL-27, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier.

[0195] In some embodiments, the present disclosure provides a method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an isolated monoclonal antibody or its antigen-binding portion provided by the present disclosure that specifically binds to and antagonizes IL-27, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier.

[0196] In some embodiments, the present disclosure provides a method of stimulating an immune response in a subject or treating cancer in a subject, the method comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces STAT1 and / or STAT3 phosphorylation in a cell, thereby stimulating the immune response or treating the cancer.

[0197] In some embodiments, the present disclosure provides a method of stimulating an immune response in a subject or treating cancer in a subject, the method comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces IL-27-mediated inhibition of CD161 expression in a cell, thereby stimulating the immune response or treating the cancer.

[0198] In some embodiments, the present disclosure provides methods of stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces IL-27-mediated PD-L1 expression and / or TIM-3 expression in cells, thereby stimulating the immune response or treating the cancer.

[0199] In some embodiments, the present disclosure provides methods of stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition induces or increases the secretion of one or more cytokines from cells mediated by PD-1, thereby stimulating the immune response or treating the cancer.

[0200] In some embodiments, the cancer is selected from the group consisting of: lung cancer (e.g., non-small cell lung cancer), sarcoma, testicular cancer, ovarian cancer, pancreatic cancer, breast cancer (e.g., triple negative breast cancer), melanoma (including, e.g., uveal melanoma, etc.), head and neck cancer (e.g., squamous head and neck cancer), colorectal cancer, bladder cancer, endometrial cancer, prostate cancer, thyroid cancer, hepatocellular carcinoma, gastric cancer, brain cancer, lymphoma (e.g., DL-BCL), leukemia (e.g., AML) or kidney cancer (e.g., renal cell carcinoma, e.g., renal clear cell carcinoma).

[0201] In some embodiments, the present disclosure provides methods of treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or its antigen-binding portion that specifically binds to and antagonizes IL-27 and is provided by the present disclosure, in combination with one or more additional therapeutic agents or procedures, wherein the second therapeutic agent or procedure is selected from the group consisting of: chemotherapy, targeted anti-cancer therapy (including, e.g., tyrosine kinase inhibitors (TKIs)), oncolytic agents, cytotoxic agents, immune-based therapies, cytokines, surgical procedures, radiation procedures, activators of co-stimulatory molecules, inhibitors of inhibitory molecules, vaccines or cellular immunotherapy or combinations thereof.

[0202] In some embodiments, the one or more additional therapeutic agents are PD-1 antagonists, PD-L1 inhibitors, TIM-3 inhibitors, LAG-3 inhibitors, TIGIT inhibitors, CD112R inhibitors, TAM inhibitors, STING agonists, 4-1BB agonists or combinations thereof.

[0203] In some embodiments, one or more additional therapeutic agents are PD-1 antagonists. In some embodiments, the PD-1 antagonists are selected from the group consisting of: PDR001, nivolumab, pembrolizumab, pidilizumab, MEDI0680, REGN2810, TSR-042, PF-06801591, and AMP-224.

[0204] In certain embodiments, one or more additional therapeutic agents are PD-L1 inhibitors. In some embodiments, the PD-L1 inhibitors are selected from the group consisting of: FAZ053, atezolizumab, avelumab, durvalumab, and BMS-936559. In some embodiments, the present disclosure provides a method of enhancing one or more activities of an anti-PD-1 antibody (e.g., enhancing PD-1-mediated cytokine secretion; enhancing anti-PD-1-mediated TNFα secretion; enhancing anti-PD-1-mediated IL-6 secretion from cells exposed to an anti-PD-1 antibody), the method comprising exposing cells to an antibody or an antigen-binding portion thereof provided by the present disclosure, and simultaneously or sequentially exposing the cells to an anti-PD-1 antibody, thereby enhancing one or more activities of the anti-PD1 antibody.

[0205] In certain embodiments, the present disclosure provides a pharmaceutical composition comprising an anti-PD-1 antibody and / or an anti-PD-L1 antibody, and an antibody or an antigen-binding portion thereof disclosed herein (e.g., an anti-IL-27 antibody) and a pharmaceutically acceptable carrier.

[0206] In related embodiments, the present disclosure provides a kit comprising an anti-PD-1 antibody and / or an anti-PD-L1 antibody and an antibody or an antigen-binding portion thereof disclosed herein (e.g., an anti-IL-27 antibody) for simultaneous or sequential administration, and instructions for use thereof.

[0207] In some embodiments, one or more additional therapeutic agents are TIM-3 inhibitors, optionally wherein the TIM-3 inhibitor is MGB453 or TSR-022.

[0208] In some embodiments, one or more additional therapeutic agents are LAG-3 inhibitors, optionally wherein the LAG-3 inhibitors are selected from the group consisting of LAG525, BMS-986016, and TSR-033.

[0209] In some embodiments, one or more additional therapeutic agents are TIGIT inhibitors. In some embodiments, one or more additional therapeutic agents are CD112R inhibitors. In some embodiments, one or more additional therapeutic agents are TAM (Axl, Mer, Tyro) inhibitors. In some embodiments, one or more additional therapeutic agents are STING agonists. In some embodiments, one or more additional therapeutic agents are 4-1BB agonists.

[0210] In some embodiments, the present disclosure provides a method for detecting IL-27 in a sample from a subject, the method comprising (a) contacting the sample with a detection antibody under conditions that permit the formation of a detection antibody-IL-27 complex if IL-27 is present in the sample from the subject, wherein the detection antibody is an antibody or an antigen-binding fragment thereof provided by the present disclosure; and (b) detecting the presence of the complex (if any) produced in step (a).

[0211] In some embodiments, the present disclosure provides a method for detecting an IL-27-related cancer in a subject, the method comprising the steps of: (a) contacting the sample with a detection antibody under conditions that permit the formation of a detection antibody-IL-27 complex if IL-27 is present in the sample from a subject suspected of having an IL-27-related cancer, wherein the detection antibody is an antibody or an antigen-binding portion thereof provided by the present disclosure; and (b) detecting the presence of the complex (if any) produced in step (a). In some embodiments, the detection antibody is conjugated to a detectable label. In some embodiments, the method further comprises contacting the sample with a capture antibody to produce a complex comprising IL-27 and the capture antibody if IL-27 is present in the sample, wherein the capture antibody is an antibody or an antigen-binding portion thereof provided by the present disclosure.

[0212] In some embodiments, the detection antibody or the capture antibody comprises heavy and light chain CDRs, wherein the amino acid sequences of the heavy chain CDR1, CDR2, and CDR3 are SYSMS (SEQ ID NO:23), YISYDGGSAYYPDTVKG (SEQ ID NO:24), and HGDYDDDDAMDY (SEQ ID NO:25), respectively, and wherein the amino acid sequences of the light chain CDR1, CDR2, and CDR3 are RASENIYSYLA (SEQ ID NO:26), NAETLTE (SEQ ID NO:27), and QHHYGTPLT (SEQ ID NO:28), respectively.

[0213] In some embodiments, the detection antibody or capture antibody comprises heavy and light chain variable regions that comprise the amino acid sequences set forth in SEQ ID NO:1 and SEQ ID NO:3, respectively.

[0214] In some embodiments, the capture antibody is immobilized on a solid support. In some embodiments, the sample is contacted with the capture antibody and then with the detection antibody. In some embodiments, the sample is a body fluid sample. In some embodiments, the fluid sample is blood, serum, plasma, cell lysate, or tissue lysate.

[0215] In some embodiments, the cancer is selected from renal cell carcinoma (RCC), hepatocellular carcinoma (HCC), lung cancer, gastroesophageal cancer, ovarian cancer, endometrial cancer, melanoma, leukemia, and lymphoma. In some embodiments, the cancer is renal cell carcinoma (RCC). In other embodiments, the cancer is hepatocellular carcinoma (HCC). In some embodiments, the cancer is selected from leukemia and lymphoma. In some embodiments, the cancer is acute myeloid leukemia (AML).

[0216] Definitions

[0217] Unless otherwise noted, the terms used in the claims and the specification are defined as follows.

[0218] It must be noted that, unless the context clearly requires otherwise, as used in the specification and the appended claims, the singular forms "a", "an", and "the" include plural referents.

[0219] As used herein, "about" will be understood by one of ordinary skill in the art and will vary to some extent depending on the context in which it is used. If the use of the term is not clear to one of ordinary skill in the art, given the context in which it is used, "about" will mean plus or minus 10% of a particular value.

[0220] As used herein, the term "agonist" refers to any molecule that partially or fully promotes, induces, increases, and / or activates the biological activity of a native polypeptide disclosed herein. Suitable agonist molecules specifically include agonist antibodies or antibody fragments, native polypeptides, fragments of peptides or proteins, or amino acid sequence variants. In some embodiments, activation in the presence of an agonist is observed in a dose-dependent manner. In some embodiments, the measured signal (e.g., biological activity) is at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% higher than the signal measured using a negative control under comparable conditions. Also disclosed herein are methods for identifying agonists suitable for the methods of the present disclosure. For example, these methods include, but are not limited to, binding assays such as enzyme-linked immunosorbent assay (ELISA), Forte systems and radioimmunoassay (RIA). These assays determine the ability of an agonist to bind to a target polypeptide (e.g., a receptor or ligand) and thus indicate the ability of the agonist to promote, increase, or activate polypeptide activity. Functional assays can also be used to determine the efficacy of an agonist, such as the ability of an agonist to activate or promote polypeptide function. For example, a functional assay can include contacting a polypeptide with a candidate agonist molecule and measuring a detectable change in one or more biological activities normally associated with the polypeptide. The potency of an agonist is typically defined by its EC 50 value (the concentration required to activate 50% of the agonist response). The lower the EC 50 value, the greater the potency of the agonist and the lower the concentration required to activate the maximal biological response.

[0221] As used herein, the term "alanine scanning" refers to a technique for determining the contribution of a specific wild-type residue to the stability or one or more functions (e.g., binding affinity) of a given protein or polypeptide. The technique involves replacing the wild-type residue in a polypeptide with an alanine residue and then evaluating the stability or one or more functions (e.g., binding affinity) of the alanine-substituted derivative or mutant polypeptide and comparing it to the wild-type polypeptide. Techniques for replacing wild-type residues in a polypeptide with alanine residues are known in the art.

[0222] The term "ameliorate" refers to any therapeutically beneficial outcome in the treatment of a disease state (e.g., cancer), including prevention, reduction in severity or progression, remission, or cure.

[0223] As used herein, the term "amino acid" refers to naturally occurring and synthetic amino acids, as well as amino acid analogs and amino acid mimetics that act in a manner similar to naturally occurring amino acids. Naturally occurring amino acids are those encoded by the genetic code, as well as those that have been subsequently modified, such as hydroxyproline, γ-carboxyglutamic acid, and O-phosphoserine. Amino acid analogs are compounds that have the same basic chemical structure as a naturally occurring amino acid (i.e., a carbon bonded to a hydrogen, a carboxyl group, an amino group, and an R group), such as homoserine, norleucine, methionine sulfoxide, and methionine methyl sulfonium. Such analogs have a modified R group (e.g., norleucine) or a modified peptide backbone, but retain the same basic chemical structure as the naturally occurring amino acid. Amino acid mimetics are compounds that have a structure different from the general chemical structure of an amino acid, but that function similarly to a naturally occurring amino acid.

[0224] Amino acids can be referred to herein by their commonly known three-letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Similarly, nucleotides can be referred to by their commonly accepted single-letter codes.

[0225] As used herein, "amino acid substitution" refers to the replacement of at least one existing amino acid residue in a predetermined amino acid sequence (the amino acid sequence of the starting polypeptide) with a second, different "substituting" amino acid residue. "Amino acid insertion" refers to the incorporation of at least one additional amino acid into a predetermined amino acid sequence. Although an insertion generally consists of the insertion of one or two amino acid residues, larger "peptide insertions" can also be made, such as the insertion of about 3 to about 5 or even up to about 10, 15, or 20 amino acid residues. As disclosed above, one or more of the inserted residues can be naturally occurring or non-naturally occurring. "Amino acid deletion" refers to the removal of at least one amino acid residue from a predetermined amino acid sequence.

[0226] As used herein, the terms "amount" or "level" are used in the broadest sense and refer to the amount, concentration, or abundance of a substance (e.g., metabolite, small molecule, protein, mRNA, marker). When referring to a metabolite or small molecule (e.g., a drug), the terms "amount", "level", and "concentration" are often used interchangeably and generally refer to the detectable amount in a biological sample. An "elevated level" or "increased level" refers to an increase in the amount, concentration, or abundance of a substance in a sample relative to a control sample (such as from one or more individuals not suffering from a disease or disorder (e.g., cancer)) or an internal control. In some embodiments, an elevated level of a substance (e.g., a drug) in a sample refers to an increase in the amount of the substance by about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% relative to the amount of the substance in a control sample, as determined by techniques known in the art (e.g., HPLC). A "decreased level" refers to a decrease in the amount, concentration, or abundance of a substance (e.g., a drug) in an individual relative to a control (such as from one or more individuals not suffering from a disease or disorder (e.g., cancer)) or an internal control. In some embodiments, a decreased level is a negligible or undetectable amount, concentration, or abundance. In some embodiments, a decreased level of a substance (e.g., a drug) in a sample refers to a decrease in the amount of the substance by about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% relative to the amount of the substance in a control sample, as determined by techniques known in the art (e.g., HPLC).

[0227] When referring to a protein, mRNA, or marker (such as those described herein), the terms "level of expression" or "expression level" are generally used interchangeably and typically refer to the detectable amount of the protein, mRNA, or marker in a biological sample. In some aspects, the detectable amount or detectable level of a protein, mRNA, or marker is related to the likelihood of response to an agent (such as those described herein). "Expression" generally refers to the process by which the information contained in a gene is transformed into a structure that is present and functional in a cell (e.g., a protein marker such as PD-L1). Thus, as used herein, "expression" can refer to transcription into a polynucleotide, translation into a polypeptide, or even modification of the polynucleotide and / or polypeptide (e.g., post-translational modification of a polypeptide). Fragments of the transcribed polynucleotide, translated polypeptide, or polynucleotide and / or polypeptide modification (e.g., post-translational modification of a polypeptide) should also be considered expressed, whether they are derived from transcripts generated by alternative splicing or degradation of transcripts, or from post-translational processing of a polypeptide (e.g., by proteolysis). "Expressed gene" includes genes that are transcribed into a polynucleotide (as mRNA) and then translated into a polypeptide, as well as genes that are transcribed into ribonucleic acid but not translated into a polypeptide (e.g., transfer RNA and ribosomal RNA). "Elevated expression", "elevated expression level", or "elevated level" refers to an increase in the expression or level of a substance in a sample relative to a control sample (such as one or more individuals without a disease or disorder (e.g., cancer)) or an internal control. In some embodiments, elevated expression of a substance (e.g., a protein marker such as PD-L1) in a sample refers to an increase in the amount of the substance by about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% relative to the amount of the substance in a control sample, as determined by techniques known in the art (e.g., FACS). "Reduced expression", "reduced expression level", or "reduced level" refers to a decrease in the expression or level of a substance (e.g., a protein marker) in an individual relative to a control (such as one or more individuals without a disease or disorder (e.g., cancer)) or an internal control. In some embodiments, reduced expression is little or no expression. In some embodiments, reduced expression of a substance (e.g., a protein marker) in a sample refers to a decrease in the amount of the substance by about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% relative to the amount of the substance in a control sample, as determined by techniques known in the art (e.g., FACS).

[0228] As used herein, the terms "angiogenesis" or "neovascularization" refer to the process by which new blood vessels develop from pre-existing blood vessels (Varner et al., (1999) Angiogen. 3:53-60; Mousa et al., (2000) Angiogen. Stim. Inhib. 35:42-44; Kim et al., (2000) Amer. J. Path. 156:1345-1362; Kim et al., (2000) J. Biol. Chem. 275:33920-33928; Kumar et al (2000) Angiogenesis: From Molec ularto Integrative Pharm. 169-180). Endothelial cells from pre-existing blood vessels or circulating endothelial stem cells (Takahashi et al., (1995) Nat. Med. 5:434-438; Isn er et al., (1999) J. Clin. Invest. 103:1231-1236) are activated in response to growth factor or hormone signals, or hypoxia or ischemia conditions and migrate, proliferate and differentiate into structures with lumens to form new blood vessels. During ischemia, such as that which occurs in cancer, the need to increase oxygenation and nutrient delivery will apparently induce the affected tissue to secrete angiogenic factors; these factors stimulate the formation of new blood vessels. Several other terms are related to angiogenesis.

[0229] As used herein, the term "antagonist" refers to any molecule that partially or completely blocks, inhibits or neutralizes the biological activity of the native polypeptides disclosed herein. Suitable antagonistic molecules specifically include antagonistic antibodies or antibody fragments, fragments of native polypeptides, peptides or proteins, or amino acid sequence variants. In some embodiments, inhibition in the presence of an antagonist is observed in a dose-dependent manner. In some embodiments, the measured signal (e.g., biological activity) is at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95% or at least about 100% lower than the signal measured using a negative control under comparable conditions. Methods for identifying antagonists suitable for the methods disclosed herein are also disclosed herein. For example, these methods include, but are not limited to, binding assays such as enzyme-linked immunosorbent assay (ELISA), Forte Systems, radioimmunoassay (RIA), Meso Scale Discovery assays (e.g., Meso Scale Discovery electrochemiluminescence (MSD-ECL) and bead-based assays). These assays determine the ability of an antagonist to bind to a target polypeptide (e.g., a receptor or ligand) and thus indicate the ability of the antagonist to inhibit, neutralize, or block polypeptide activity. Functional assays can also be used to determine the efficacy of an antagonist, such as the ability of the antagonist to inhibit polypeptide or agonist function. For example, a functional assay can include contacting a polypeptide with a candidate agonist molecule and measuring a detectable change in one or more biological activities normally associated with the polypeptide. The potency of an antagonist is generally defined by its EC 50 value (the concentration required to inhibit 50% of the agonist response). The lower the IC 50 value, the greater the potency of the antagonist and the lower the concentration required to inhibit the maximal biological response.

[0230] As used herein, the phrase "antibody or antigen-binding portion thereof that antagonizes human IL-27" refers to an antibody that antagonizes at least one activity of human IL-27 that is recognized in the art (e.g., IL-27 biological activity and / or one or more downstream pathways or other IL-27-mediated functions mediated by IL-27 signaling), e.g., an antibody associated with a reduction (or decrease) of at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more of human IL-27 activity. Additional examples of IL-27 biological activity and / or one or more downstream pathways or other IL-27-mediated functions mediated by IL-27 signaling are described in more detail below and elsewhere herein.

[0231] As used herein, the term "anti-IL-27 antagonist antibody" (which may be interchangeably referred to as "anti-IL-27 antibody") refers to an antibody that specifically binds to and antagonizes IL-27 biological activity and / or one or more downstream pathways or other IL-27-mediated functions mediated by IL-27 signaling. Anti-IL-27 antagonist antibodies include antibodies that block, antagonize, inhibit, suppress, or reduce IL-27 biological activity (e.g., ligand binding, enzymatic activity) (including downstream pathways mediated by IL-27 signaling or function, such as receptor binding and / or eliciting a cellular response to IL-27 or its metabolites). In some embodiments, the anti-IL-27 antagonistic antibodies provided by the present disclosure bind to human IL-27 and prevent, block, or inhibit the binding of human IL-27 to its cognate or normal receptor (e.g., the IL-27 receptor) or one or more receptor subunits (e.g., gp130 and / or IL-27Rα (also known as WSX1 / TCCR)). In some embodiments, the anti-IL-27 antagonistic antibody prevents, blocks, or inhibits the binding of human IL-27 to gp130. In some embodiments, the anti-IL-27 antagonistic antibody prevents, blocks, or inhibits the binding of human IL-27 to IL-27Rα. In some embodiments, the anti-IL-27 antagonistic antibody prevents, blocks, or inhibits the dimerization of IL-27 monomers. In some embodiments, the anti-IL-27 antagonistic antibody specifically binds to an EBI3 monomer. In some embodiments, the anti-IL-27 antagonistic antibody specifically binds to an IL-27p28 monomer. In some embodiments, the anti-IL-27 antagonistic antibody specifically binds to two IL-27 monomers. In some embodiments, the anti-IL-27 antagonistic antibody specifically binds to a discontinuous epitope comprising both EBI3 and P28. In some embodiments, the anti-IL-27 antagonistic antibody inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells. In some embodiments, the anti-IL-27 antagonistic antibody inhibits or reduces the inhibition of CD161 expression in cells (e.g., improves or alleviates IL-27-mediated inhibition of CD161 expression in cells). In some embodiments, the anti-IL-27 antagonistic antibody inhibits or reduces the expression of PD-L1 and / or TIM-3 in cells. In some embodiments, the anti-IL-27 antagonistic antibody induces or increases the secretion of one or more cytokines mediated by PD-1 from cells. In some embodiments, the anti-IL-27 antagonistic antibody binds to human IL-27 and stimulates or enhances an anti-tumor response. In some embodiments, the anti-IL-27 antagonistic antibody binds to human IL-27 with an affinity of 15 nM or less. In some embodiments, the anti-IL-27 antagonistic antibody binds to human IL-27 and comprises a wild-type or mutant IgG1 heavy chain constant region or a wild-type or mutant IgG4 heavy chain constant region. Examples of anti-IL-27 antagonistic antibodies are provided herein.

[0232] As used herein, the term "antibody" refers to a complete antibody comprising two light chain polypeptides and two heavy chain polypeptides. Complete antibodies include different antibody isotypes, including IgM, IgG, IgA, IgD, and IgE antibodies. The term "antibody" includes polyclonal antibodies, monoclonal antibodies, chimerized or chimeric antibodies, humanized antibodies, primatized antibodies, deimmunized antibodies, and fully human antibodies. Antibodies can be prepared in or derived from any of a variety of species, such species being, for example, mammals such as humans, non-human primates (e.g., orangutans, baboons, or chimpanzees), horses, cows, pigs, sheep, goats, dogs, cats, rabbits, guinea pigs, gerbils, hamsters, rats, and mice. Antibodies can be purified or recombinant antibodies. As used herein, the term "antibody fragment", "antigen-binding fragment", or similar terms refers to a fragment that retains the ability to bind to a target antigen (e.g., IL-27) and inhibit the activity of the target antigen. Such fragments include, for example, single-chain antibodies, single-chain Fv fragments (scFv), Fd fragments, Fab fragments, Fab' fragments, or F(ab')2 fragments. An scFv fragment is a single polypeptide chain that includes both the heavy and light chain variable regions of the antibody from which the scFv is derived. Additionally, the definition of an antibody also includes intracellular antibodies, minibodies, triabodies, and diabodies, which are applicable to the methods described herein. See, e.g., Todorovska et al., (2001) J. Immunol. Methods 248(1):47-66; Hudson and Kortt, (1999) J. Immunol. Methods 231(1):177-189; Poljak, (1994) Structure 2(12):1121-1123; Rondon and Marasco, (1997) Annu. Rev. Microbiol. 51:257-283, the disclosures of each of which are incorporated herein by reference in their entirety.

[0233] As used herein, the term "antibody fragment" also includes, for example, single domain antibodies, such as camelized single domain antibodies. See, for example, Muyldermans et al., (2001) Trends Biochem. Sci. 26:230-235; Nuttall et al., (2000) Curr. Pharm. Biotech. 1:253-263; Reichmann et al., (1999) J. Immunol. Meth. 231:25-38; PCT Application Publication Nos. WO 94 / 04678 and WO 94 / 25591, and U.S. Patent No. 6,005,079, which are hereby incorporated by reference in their entirety. In some embodiments, the present disclosure provides single domain antibodies comprising two single domain antibodies having modified VH domains (such that a single domain antibody can be formed).

[0234] In some embodiments, the antigen-binding fragment comprises the variable region of a heavy chain polypeptide and the variable region of a light chain polypeptide. In some embodiments, the antigen-binding fragments described herein comprise the CDRs of the light and heavy chain polypeptides of an antibody.

[0235] The term "antigen-presenting cell" or "APC" is a cell that displays a foreign antigen complexed with MHC on its surface. T cells recognize such a complex via the T cell receptor (TCR). Examples of APC cells include, but are not limited to: B cells, dendritic cells (DCs), peripheral blood mononuclear cells (PBMCs), monocytes (such as THP-1), B lymphoblastoid cells (such as C1R.A2,1518B-LCL), and monocyte-derived dendritic cells (DCs). Some APCs internalize antigens by phagocytosis or receptor-mediated endocytosis.

[0236] The term "antigen presentation" refers to the process by which an APC captures an antigen and makes it recognizable by T cells (e.g., as a component of an MHC-I and / or MHC-II conjugate).

[0237] As used herein, the term "apoptosis" refers to the process of programmed cell death that occurs in multicellular organisms (such as humans). The highly regulated biochemical and molecular events that lead to apoptosis can result in observable and characteristic morphological changes to the cell, including membrane blebbing, cell volume shrinkage, chromosomal DNA condensation and fragmentation, and mRNA decay. A common method for identifying cells (including T cells) undergoing apoptosis is to expose the cells to a protein (Annexin V) conjugated to a fluorophore. Annexin V is commonly used to detect apoptotic cells by its ability to bind to phosphatidylserine on the outer leaflet of the plasma membrane, which binding is an early indicator that the cell is undergoing the process of apoptosis.

[0238] As used herein, the term "B cell" (or "B lymphocyte") refers to a type of white blood cell that is a subtype of lymphocyte. B cells function in the humoral immune component of the adaptive immune system by secreting antibodies. B cells also present antigens and secrete cytokines. B cells, unlike the other two types of lymphocytes (T cells and natural killer cells), express B cell receptors (BCRs) on their cell membranes. The BCR allows B cells to bind to specific antigens, which will initiate an antibody response against that antigen.

[0239] As used herein, the term "binding to immobilized IL-27" refers to the ability of the antibodies of the present disclosure to bind to IL-27 (e.g., IL-27 expressed on the cell surface or attached to a solid support).

[0240] As used herein, the term "bispecific" or "bifunctional antibody" refers to an artificial hybrid antibody that has two different heavy / light chain pairs and two different binding sites. Bispecific antibodies can be produced by a variety of methods, including fusion of hybridomas or ligation of Fab' fragments. See, e.g., Songsivilai & Lachmann, (1990) Clin. Exp. Immunol. 79:315-321; Kostelny et al., (1992) J. Immunol. 148:1547-1553.

[0241] Conventionally, the recombinant production of bispecific antibodies is based on the co-expression of two immunoglobulin heavy chain / light chain pairs, where the two heavy chain / light chain pairs have different specificities (Milstein and Cuello, (1983) Nature 305:537-539). Antibody variable domains with the desired binding specificities (antibody-antigen binding sites) can be fused to immunoglobulin constant domain sequences. Preferably, the heavy chain variable region is fused to an immunoglobulin heavy chain constant region (including at least a portion of the hinge region, CH2 region, and CH3 region). For further details on currently known illustrative methods for generating bispecific antibodies, see, for example, Suresh et al., (1986) Methods Enzymol. 121:210; PCT Publication No. WO 96 / 27011; Brennan et al., (1985) Science 229:81; Shalaby et al., J. Exp. Med. (1992) 175:217-225; Kostelny et al., (1992) J. Immunol. 148(5):1547-1553; Hollinger et al., (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Gruber et al., (1994) J. Immunol. 152:5368, and Tutt et al., (1991) J. Immunol. 147:60. Bispecific antibodies also include cross-linked or heteroconjugate antibodies. Heteroconjugate antibodies can be prepared using any convenient cross-linking method. Suitable cross-linking agents are well known in the art and are disclosed, along with many cross-linking techniques, in U.S. Patent No. 4,676,980.

[0242] Also described are various techniques for preparing and isolating bispecific antibody fragments directly from recombinant cell cultures. For example, bispecific antibodies have been produced using leucine zippers. See, e.g., Kostelny et al. (1992) J Immunol 148(5):1547-1553. The leucine zipper peptides from Fos and Jun proteins can be linked to the Fab’ portions of two different antibodies by gene fusion. Antibody homodimers can be reduced to monomers in the hinge region and then re-oxidized to form antibody heterodimers. The method can also be used to produce antibody homodimers. The “diabody” technology described by Hollinger et al. (1993) Proc Natl Acad Sci USA 90:6444-6448 provides an alternative mechanism for preparing bispecific antibody fragments. The fragments contain a heavy chain variable domain (VH) linked to a light chain variable domain (VL) by a linker that is too short to allow pairing between the two domains on the same chain. Thus, the VH and VL domains of one fragment are forced to pair with the complementary VL and VH domains of another fragment, thereby forming two antigen-binding sites. Another strategy for preparing bispecific antibody fragments by using single-chain Fv (scFv) dimers has also been reported. See, e.g., Gruber et al. (1994) J Immunol 152:5368. Alternatively, the antibodies can be “linear antibodies” as described in Zapata et al. (1995) Protein Eng. 8(10):1057-1062. Briefly, these antibodies contain a pair of tandem Fd segments (VH-CH1-VH-CH1), which form a pair of antigen-binding regions. Linear antibodies can be bispecific or monospecific.

[0243] Antibodies with more than two valencies (e.g., trispecific antibodies) have been envisioned and described, for example, in Tutt et al. (1991) J Immunol 147:60.

[0244] The present disclosure also includes variant forms of multispecific antibodies, such as the dual variable domain immunoglobulin (DVD-Ig) molecules described in Wu et al. (2007) Nat Biotechnol 25(11): 1290-1297. DVD-Ig molecules are designed such that two different light chain variable domains (VL) from two different parental antibodies are directly tandemly linked or linked via a short linker by recombinant DNA techniques, followed by the light chain constant domain. Similarly, the heavy chain contains two tandemly linked different heavy chain variable domains (VH), followed by the constant regions CH1 and the Fc region. Methods for preparing DVD-Ig molecules from two parental antibodies are further described, for example, in PCT Publication Nos. WO 08 / 024188 and WO 07 / 024715. In some embodiments, the bispecific antibody is a Fab tandem immunoglobulin, in which the light chain variable region having a second specificity is fused to the heavy chain variable region of a full antibody. Such antibodies are described, for example, in International Patent Application Publication No. WO 2015 / 103072.

[0245] As used herein, "cancer antigen" or "tumor antigen" refers to (i) a tumor-specific antigen, (ii) a tumor-associated antigen, (iii) a cell expressing a tumor-specific antigen, (iv) a cell expressing a tumor-associated antigen, (v) an embryonic antigen on a tumor, (vi) an autologous tumor cell, (vii) a tumor-specific membrane antigen, (viii) a tumor-associated membrane antigen, (ix) a growth factor receptor, (x) a growth factor ligand, and (xi) any other type of antigen or antigen-presenting cell or material associated with cancer.

[0246] As used herein, the term "cancer-specific immune response" refers to an immune response induced by the presence of a tumor, cancer cell, or cancer antigen. In certain embodiments, the response includes the proliferation of cancer antigen-specific lymphocytes. In certain embodiments, the response includes the expression and upregulation of antibodies and T cell receptors and the formation and release of lymphokines, chemokines, and cytokines. The innate immune system and the acquired immune system interact to initiate an antigen response against a tumor, cancer cell, or cancer antigen. In certain embodiments, the cancer-specific immune response is a T cell response.

[0247] The term "cancer" is recognized in the art and refers to a malignant tumor of epithelial or endocrine tissue, including cancers of the respiratory system, gastrointestinal system, urogenital system, testicular cancer, breast cancer, prostate cancer, endocrine system cancer, and melanoma. The anti-IL-27 antibodies described herein can be used to treat patients suffering from, suspected of having any type of cancer (including renal cancer or melanoma) or any viral disease or at high risk of developing said cancer or viral disease. Exemplary cancers include those formed from tissues of the cervix, lung, prostate, breast, head and neck, colon, and ovary. The term also includes carcinosarcoma, which includes a malignant tumor composed of carcinomatous tissue and sarcomatous tissue. "Adenocarcinoma" refers to a cancer derived from glandular tissue or in which the tumor cells form recognizable glandular structures.

[0248] As used herein, the term "CD112R" refers to a member of the poliovirus receptor-like protein family and is a co-inhibitory receptor of human T cells. CD112R is preferentially expressed on T cells and inhibits T cell receptor-mediated signaling. CD112 is widely expressed on antigen-presenting cells and tumor cells and is the ligand of CD112R. CD112R competes with CD226 for binding to CD112. Disrupting the CD112R-CD112 interaction enhances the response of human T cells. CD112R serves as a novel checkpoint of human T cells through its interaction with CD112. As used herein, the term "CD112R inhibitor" refers to an agent that disrupts, blocks, or inhibits the biological function or activity of CD112R.

[0249] As used herein, the term "CD137" (or "4-1BB") refers to a member of the tumor necrosis factor (TNF) receptor superfamily. 4-1BB is a co-stimulatory immune checkpoint molecule, mainly targeting activated T cells. Crosslinking of CD137 enhances T cell proliferation, IL-2 secretion, survival, and cytolytic activity. As used herein, the term "4-1BB agonist" refers to an agent that stimulates, induces, or increases one or more functions of 4-1BB. An exemplary 4-1BB agonist is Utomilumab (PF-05082566), a fully human IgG2 monoclonal antibody that targets this 4-1BB to stimulate T cells.

[0250] As used herein, the term "CD161" (also known as killer cell lectin-like receptor subfamily B, member 1 (KLRB1); NK1.1, or NKR-P1A) refers to a member of the C-type lectin superfamily. CD161 is a marker of T cells, and the expression of CD161 is associated with the infiltration of T cells into the tumor microenvironment of many different cancer types. CD161 is further described in Fergusson et al., ((2014) Cell Reports 9(3): 1075-1088, which is incorporated herein by reference in its entirety).

[0251] As used herein, the term "IL-27" or "interleukin 27" refers to the IL-27 cytokine. IL-27 is related to the IL-6 / IL-12 cytokine family and is a heterodimeric cytokine that comprises a first subunit called Epstein-Barr virus-induced gene 3 (EBI3; also referred to as IL-27 subunit beta and IL-27B) and a second subunit called IL-27p28 (also referred to as IL30, IL-27 subunit alpha and IL-27A). IL-27 is mainly synthesized by activated antigen-presenting cells including monocytes, endothelial cells and dendritic cells (Jankowski et al. (2010) Arch Immunol. Ther. Exp. 58:417-425, Diakowski et al. (2013) Adv. Clin. Exp. Med. (2013) 22(5):683-691). Although IL-27 can have pro-inflammatory effects, many studies have shown an important role of IL-27 as an immunosuppressant (Shimizu et al. (2006) J. Immunol. 176:7317-7324, Hisada et al. (2004) Cancer Res. 64:1152-1156, Diakowski (2013) ibid.). Although it was initially described as a factor promoting Th1 response initiation, it was later found that IL-27 exerts a major T cell inhibitory function by restricting Th1 responses, inhibiting Th2 and Th17 cell differentiation, and regulating the development of Tr1 and other regulatory T cell populations (Dietrich et al. (2014) J. Immunol. 192:5382-5389). In addition to its role as an immunomodulator, IL-27 also regulates angiogenesis, hematopoiesis and osteoclastogenesis (ibid.).

[0252] IL-27 signals through a heterodimeric type I cytokine receptor (IL-27 receptor or IL-27R) that comprises a first subunit called WSX1 (also referred to as IL-27 receptor subunit alpha, IL-27RA, T cell cytokine receptor type 1 (TCCR) and cytokine receptor-like 1 (CRL1)) and a second subunit called gp130 (also referred to as interleukin-6 signal transducer (IL6ST), interleukin-6 receptor subunit beta (IL-6RB) and oncostatin M receptor). p130 is also a receptor subunit for cytokines of the IL-6 family (Liu et al. (2008) Scan. J. Immunol. 68:22-299, Diakowski (2013) ibid.). Signaling of IL-27 through IL-27R activates multiple signaling cascades, including the JAK-STAT and p38 MAPK pathways.

[0253] EBI3 is also thought to have biological functions independent of the p28 or IL-27 heterodimers. For example, EBI3 also interacts with p35 to form the heterodimeric cytokine IL-35 (Yoshida et al. (2015) Annu. Rev Immunol. 33:417-43) and has been shown to be selectively overexpressed in certain cell types without a corresponding increase in p28 or IL-27 (Larousserie et al. (2005) Am. J. Pathol. 166(4):1217-28).

[0254] The amino acid sequences of exemplary human EBI3 protein are provided in SEQ ID NO:698 (NCBI reference sequence: NP_005746.2; N-MTPQLLLALVLWASCPPCSGRKGPPAALTLPRVQCRASRYPIAVDCSWTLPPAPNSTSPVSFIATYRLGMAARGHSWPCLQQTPTSTSCTITDVQLFSMAPYVLNVTAVHPWGSSSSFVPFITEHIIKPDPPEGVRLSPLAERQLQVQWEPPGSWPFPEIFSLKYWIRYKRQGAARFHRVGPIEATSFILRAVRPRARYYVQVAAQDLTDYGELSDWSLPATATMSLGK-C). The amino acid sequences of exemplary human p28 protein are provided in SEQ ID NO:699 (NCBI reference sequence: NP_663634.2; N-MGQTAGDLGWRLSLLLLPLLLVQAGVWGFPRPPGRPQLSLQELRREFTVSLHLARKLLSEVRGQAHRFAESHLPGVNLYLLPLGEQLPDVSLTFQAWRRLSDPERLCFISTTLQPFHALLGGLGTQGRWTNMERMQLWAMRLDLRDLQRHLRFQVLAAGFNLPEEEEEEEEEEEEERKGLLPGALGSALQGPAQVSWPQLLSTYRLLHSLELVLSRAVRELLLLSKAGHSVWPLGFPTLSPQP-C).The amino acid sequence of an exemplary human WSX1 protein is provided in SEQ ID NO: 700 (NCBI reference sequence: NP_004834.1; N-MRGGRGAPFWLWPLPKLALLPLLWVLFQRTRPQGSAGPLQCYGVGPLGDLNCSWEPLGDLGAPSELHLQSQKYRSNKTQTVAVAAGRSWVAIPREQLTMSDKLLVWGTKAGQPLWPPVFVNLETQMKPNAPRLGPDVDFSEDDPLEATVHWAPPTWPSHKVLICQFHYRRCQEAAWTLLEPELKTIPLTPVEIQDLELATGYKVYGRCRMEKEEDLWGEWSPILSFQTPPSAPKDVWVSGNLCGTPGGEEPLLLWKAPGPCVQVSYKVWFWVGGRELSPEGITCCCSLIPSGAEWARVSAVNATSWEPLTNLSLVCLDSASAPRSVAVSSIAGSTELLVTWQPGPGEPLEHVVDWARDGDPLEKLNWVRLPPGNLSALLPGNFTVGVPYRITVTAVSASGLASASSVWGFREELAPLVGPTLWRLQDAPPGTPAIAWGEVPRHQLRGHLTHYTLCAQSGTSPSVCMNVSGNTQSVTLPDLPWGPCELWVTASTIAGQGPPGPILRLHLPDNTLRWKVLPGILFLWGLFLLGCGLSLATSGRCYHLRHKVLPRWVWEKVPDPANSSSGQPHMEQVPEAQPLGDLPILEVEEMEPPPVMESSQPAQATAPLDSGYEKHFLPTPEELGLLGPPRPQVLA-C).The amino acid sequence of an exemplary human gp130 protein is provided in SEQ ID NO:701 (NCBI Reference Sequence: NP_002175.2; N-MLTLQTWLVQ ALFIFLTTESTGELLDPCGYISPESPVVQLHSNFTAVCVLKEKCMDYFHVNANYIVWKTNHFTIPKEQYTIINRTASSVTFTDIASLNIQLTCNILTFGQLEQNVYGITIISGLPPEKPKNLSCIVNEGKKMRCEWDGGRETHLETNFTLKSEWATHKFADCKAKRDTPTSCTVDYSTVYFVNIEVWVEAENALGKVTSDHINFDPVYKVKPNPPHNLSVINSEELSSILKLTWTNPSIKSVIILKYNIQYRTKDASTWSQIPPEDTASTRSSFTVQDLKPFTEYVFRIRCMKEDGKGYWSDWSEEASGITYEDRPSKAPSFWYKIDPSHTQGYRTVQLVWKTLPPFEANGKILDYEVTLTRWKSHLQNYTVNATKLTVNLTNDRYLATLTVRNLVGKSDAAVLTIPACDFQATHPVMDLKAFPKDNMLWVEWTTPRESVKKYILEWCVLSDKAPCITDWQQEDGTVHRTYLRGNLAESKCYLITVTPVYADGPGSPESIKAYLKQAPPSKGPTVRTKKVGKNEAVLEWDQLPVDVQNGFIRNYTIFYRTIIGNETAVNVDSSHTEYTLSSLTSDTLYMVRMAAYTDEGGKDGPEFTFTTPKFAQGEIEAIVVPVCLAFLLTTLLGVLFCFNKRDLIKKHIWPNVPDPSKSHIAQWSPHTPPRHNFNSKDQMYSDGNFTDVSVVEIEANDKKPFPEDLKSLDLFKKEKINTEGHSSGIGGSSCMSSSRPSISSSDENESSQNTSSTVQYSTVVHSGYRHQVPSVQVFSRSESTQPLLDSEERPEDLQLVDHVDGGDGILPRQQYFKQNCSQHESSPDISHFERSKQVSSVNEEDFVRLKQQISDHISQSCGSGQMKMFQEVSAADAFGPGTEGQVERFETVGMEAATDEGMPKSYLPQTVRQGGYMPQ-C).

[0255] As used herein, the term "competition", when used in the context of antigen-binding proteins (e.g., immunoglobulins, antibodies, or antigen-binding fragments thereof) competing for binding to the same epitope, refers to the interaction between antigen-binding proteins determined by an assay (e.g., a competitive binding assay; a cross-blocking assay), wherein a test antigen-binding protein (e.g., a test antibody) inhibits (e.g., reduces or blocks) the specific binding of a reference antigen-binding protein (e.g., a reference antibody) to a common antigen (e.g., IL-27 or a fragment thereof).

[0256] A polypeptide or amino acid sequence "derived from" a designated polypeptide or protein refers to the source of the polypeptide. Preferably, a polypeptide or amino acid sequence derived from a particular sequence has an amino acid sequence that is substantially the same as that sequence or a portion thereof (wherein the portion consists of at least 10-20 amino acids, preferably at least 20-30 amino acids, more preferably at least 30-50 amino acids), or alternatively can be recognized by one of ordinary skill in the art as being derived from that sequence. A polypeptide derived from another peptide may have one or more mutations relative to the starting polypeptide, e.g., one or more amino acid residues that have been replaced by another amino acid residue or that have had one or more amino acid residues inserted or deleted.

[0257] A polypeptide may comprise a non-naturally occurring amino acid sequence. Such variants must have less than 100% sequence identity or similarity to the starting molecule. In certain embodiments, the variant will have, for example, an amino acid sequence having about 75% to less than 100%, more preferably about 80% to less than 100%, more preferably about 85% to less than 100%, more preferably about 90% to less than 100% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%), most preferably about 95% to less than 100% amino acid sequence identity or similarity to the amino acid sequence of the starting polypeptide over the length of the variant molecule.

[0258] In certain embodiments, there is one amino acid difference between the starting polypeptide sequence and the sequence derived therefrom. Identity or similarity to the sequence is defined herein as the percentage of amino acid residues in the candidate sequence that are identical (i.e., the same residue) to the initial sequence after aligning the sequences and introducing gaps (if necessary) to achieve the maximum percentage of sequence identity. In certain embodiments, the polypeptide consists of, consists essentially of, or comprises an amino acid sequence selected from the sequences shown in Table 12. In certain embodiments, the polypeptide comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to an amino acid sequence selected from the sequences shown in Table 12. In certain embodiments, the polypeptide comprises a contiguous amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to a contiguous amino acid sequence selected from the sequences shown in Table 12. In certain embodiments, the polypeptide comprises an amino acid sequence having at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, or 500 (or any integer among these numbers) contiguous amino acids of an amino acid sequence selected from the sequences shown in Table 12.

[0259] In certain embodiments, the antibodies of the present disclosure are encoded by nucleotide sequences. The nucleotide sequences of the invention can be used in a variety of applications, including cloning, gene therapy, protein expression and purification, mutagenesis introduction, DNA vaccination of hosts in need thereof, antibody generation (for, e.g., passive immunization), PCR, primer and probe generation, and the like. In certain embodiments, the nucleotide sequences of the invention comprise a nucleotide sequence selected from the sequences shown in Table 12, consist of, or consist essentially of, the nucleotide sequences. In certain embodiments, the nucleotide sequence comprises a nucleotide sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to a nucleotide sequence selected from the sequences shown in Table 12. In certain embodiments, the nucleotide sequence comprises a continuous nucleotide sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identity to a continuous nucleotide sequence selected from the sequences shown in Table 12. In certain embodiments, the nucleotide sequence comprises a nucleotide sequence having at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 200, 300, 400, or 500 (or any integer among these numbers) consecutive nucleotides of a nucleotide sequence selected from the sequences shown in Table 12.

[0260] One of ordinary skill in the art will also understand that the antibodies suitable for the methods disclosed herein can be altered such that they differ in sequence from the naturally occurring or native sequences from which they are derived while retaining the desired activity of the native sequence. For example, nucleotide or amino acid substitutions can be made that result in conservative substitutions or alterations at "non-essential" amino acid residues. Mutations can be introduced by standard techniques such as site-directed mutagenesis and PCR-mediated mutagenesis.

[0261] Antibodies suitable for the methods disclosed herein may contain conservative amino acid substitutions at one or more amino acid residues, e.g., at essential or non-essential amino acid residues. A "conservative amino acid substitution" is an amino acid substitution in which the amino acid residue is replaced with an amino acid residue having a similar side chain. Families of amino acid residues having similar side chains have been defined in the art and include basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), β-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Thus, non-essential amino acid residues in the binding polypeptide are preferably replaced with another amino acid residue from the same side chain family. In certain embodiments, a stretch of amino acids may be replaced with a structurally similar stretch that differs in the sequence and / or composition of side chain family members. Alternatively, in certain embodiments, mutations may be introduced randomly along all or part of the coding sequence, e.g., by saturation mutagenesis, and the resulting mutants may be incorporated into the binding polypeptides of the invention and screened for their ability to bind to the desired target.

[0262] As used herein, the term antigen "cross-presentation" refers to the presentation of exogenous protein antigens to T cells by MHC class I and II molecules on APCs.

[0263] As used herein, the term "cross-reactivity" refers to the ability of the antibodies of the present disclosure to bind to IL-27 from different species. For example, an antibody of the present disclosure that binds human IL-27 may also bind IL-27 of another species. As used herein, cross-reactivity is measured by detecting specific reactivity with a purified antigen in a binding assay (e.g., SPR, ELISA), or by detecting binding to cells that physiologically express IL-27 or otherwise functionally interacting with said cells. Methods for determining cross-reactivity include the standard binding assays described herein, e.g., by Biacore TM Surface plasmon resonance (SPR) analysis, using Biacore TM 2000 SPR instrument (Biacore AB, Uppsala, Sweden) or flow cytometry.

[0264] As used herein, the term "cytotoxic T lymphocyte (CTL) response" refers to an immune response induced by cytotoxic T cells. The CTL response is mainly mediated by CD8 + T cells.

[0265] As used herein, the term "dendritic cell" or "DC" refers to an antigen-presenting cell type that is a bone marrow (BM)-derived leukocyte and is the most potent type of antigen-presenting cell. DCs capture and process antigens, converting proteins into peptides that are presented on major histocompatibility complex (MHC) molecules recognized by T cells. DCs are heterogeneous, such as myeloid and plasmacytoid DCs; although all DCs are capable of taking up antigens, processing antigens, and presenting them to naive T cells, DC subtypes have different markers and differ in terms of their localization, migratory pathways, detailed immune functions, and dependence on infectious or inflammatory stimuli (for which they are generated). During the development of the adaptive immune response, the phenotype and function of DCs play a role in initiating tolerance, memory, and polarization of T-helper cell 1 (Th1), Th2, and Th17 differentiation.

[0266] As used herein, the term "dendritic cell activation" refers to the transition from an immature dendritic cell to a mature dendritic cell; and activated dendritic cells include mature dendritic cells and dendritic cells during the transition process, in which the expression of CD80 and CD86, which induce co-stimulatory signals, is elevated by activation stimuli. Mature human dendritic cells are cells that are positive for the expression of CD40, CD80, CD86, and HLA class II (e.g., HLA-DR). For example, based on markers selected from the group consisting of CD80 and CD86, immature dendritic cells can be distinguished from mature dendritic cells. Immature dendritic cells are weakly positive, preferably negative, for these markers, while mature dendritic cells are positive. The distinction of mature dendritic cells is routinely performed by those skilled in the art, and the respective markers and methods for measuring their expression are also well known to those skilled in the art.

[0267] As used herein, the term "EC 50 " refers to the concentration of an antibody or its antigen-binding portion that induces a response that is 50% of the maximum response (i.e., halfway between the maximum response and the baseline) in an in vitro or in vivo assay.

[0268] As used herein, the term "effective dose" or "effective dosage" is defined as an amount sufficient to achieve or at least partially achieve the desired effect. The term "therapeutically effective dose" is defined as an amount sufficient to cure or at least partially arrest a disease and its complications in a patient already suffering from the disease. The amount effective for such use depends on the severity of the condition being treated and the general state of the patient's own immune system.

[0269] As used herein, the term "epitope" or "antigenic determinant" refers to the site on an antigen to which an immunoglobulin or antibody specifically binds. The term "epitope mapping" refers to the process or method of identifying the binding site or epitope of an antibody or its antigen-binding fragment on its target protein antigen. Methods and techniques for epitope mapping are provided herein. Epitopes can be formed either by contiguous amino acids or by non-contiguous amino acids that are brought into close proximity by the tertiary folding of the protein. Epitopes formed by contiguous amino acids are generally retained upon exposure to denaturing solvents, whereas epitopes formed by tertiary folding are generally lost upon treatment with denaturing solvents. Epitopes typically comprise at least 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation. Methods for determining which epitopes are bound by a given antibody (i.e., epitope mapping) are well known in the art and include, for example, immunoblotting and immunoprecipitation assays in which the reactivity of overlapping or contiguous peptides of IL-27 with a given anti-IL-27 antibody is tested. Methods for determining the spatial conformation of epitopes include techniques known in the art and techniques described herein, for example, x-ray crystallography and two-dimensional nuclear magnetic resonance (see, for example, Epitope Mapping Protocols in Methods in Molecular Biology, Vol. 66, G.E. Morris, ed. (1996)).

[0270] The present disclosure also includes antibodies that bind to epitopes on IL-27 that comprise all or part of the epitopes recognized by the specific antibodies described herein (e.g., identical or overlapping regions or regions between or spanning such regions).

[0271] The present disclosure also includes antibodies that bind to the same epitope and / or antibodies that compete with the antibodies described herein for binding to human IL-27. Antibodies that recognize the same epitope or compete for binding can be identified using conventional techniques. Such techniques include, for example, immunoassays that show the ability of one antibody to block the binding of another antibody to a target antigen, i.e., competitive binding assays. Competitive binding is assayed in a determination in which the immunoglobulin being tested inhibits the specific binding of a reference antibody to a common antigen, such as IL-27. Many types of competitive binding assays are known, e.g.: solid-phase direct or indirect radioimmunoassay (RIA), solid-phase direct or indirect enzyme immunoassay (EIA), sandwich competitive assay (see Stahli et al., Methods in Enzymology 9:242 (1983)); solid-phase direct biotin-avidin EIA (see Kirkland et al., J. Immunol. 137:3614 (1986)); solid-phase direct labeled assay, solid-phase direct labeled sandwich assay (see Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Press (1988)); solid-phase direct labeled RIA using an I-125 label (see Morel et al., Mol. Immunol. 25(1):7 (1988)); solid-phase direct biotin-avidin protein EIA (Cheung et al., Virology 176:546 (1990)); and direct labeled RIA. (Moldenhauer et al., Scand. J. Immunol. 32:77 (1990)). Typically, such an assay involves the use of purified antigen bound to a solid surface or cells bearing one of the two, an unlabeled test immunoglobulin, and a labeled reference immunoglobulin. Competitive inhibition is measured by determining the amount of label bound to the solid surface or cells in the presence of the test immunoglobulin. The test immunoglobulin is typically present in excess. Generally, when the competing antibody is present in excess, it will inhibit the specific binding of the reference antibody to the common antigen by at least 50 - 55%, 55 - 60%, 60 - 65%, 65 - 70%, 70 - 75% or more.

[0272] Other techniques include, for example, epitope mapping methods such as x-ray analysis of antigen:antibody complex crystals, which provide atomic resolution of epitopes, and mass spectrometry combined with hydrogen / deuterium (H / D) exchange, which studies the conformation and dynamics of antigen:antibody interactions. Other methods monitor the binding of an antibody to an antigen fragment or a mutant form of the antigen, where loss of binding due to modification of an amino acid residue in the antigen sequence is generally considered an indication of an epitope component. Additionally, computational combinatorial methods for epitope mapping can also be used. These methods rely on the ability of a target antibody to affinity isolate specific short peptides from a combinatorial phage display peptide library. The peptides are then considered to be leads corresponding to the epitope definition of the antibody used to screen the peptide library. Computational algorithms have also been developed for epitope mapping, and these algorithms have been shown to be able to map conformationally discontinuous epitopes.

[0273] As used herein, the term "Fc-mediated effector function" or "Fc effector function" refers to the biological activity of an antibody other than the primary function and purpose of the antibody. For example, the effector function of a therapeutic agnostic antibody is the biological activity other than the activation of a target protein or pathway. Examples of antibody effector functions include C1q binding and complement-dependent cytotoxicity; Fc receptor binding; antibody-dependent cell-mediated cytotoxicity (ADCC); phagocytosis; downregulation of cell surface receptors (e.g., B cell receptors); lack of activation of platelets expressing Fc receptors; and B cell activation. Many effector functions begin with the binding of Fc to an Fcγ receptor. In some embodiments, tumor antigen-targeting antibodies have effector functions, such as ADCC activity. In some embodiments, the tumor antigen-targeting antibodies described herein comprise variant constant regions that have enhanced effector functions (e.g., enhanced ability to mediate ADCC) relative to the unmodified form of the constant region.

[0274] As used herein, the term "Fc receptor" refers to a polypeptide found on the surface of immune effector cells that binds to the Fc region of an antibody. In some embodiments, the Fc receptor is an Fcγ receptor. There are three subclasses of Fcγ receptors: FcγRI (CD64), FcγRII (CD32), and FcγRIII (CD16). All four IgG isotypes (IgG1, IgG2, IgG3, and IgG4) bind to and activate the Fc receptors FcγRI, FcγRIIA, and FcγRIIIA. FcγRIIB is an inhibitory receptor, and thus antibodies that bind to this receptor do not activate complement and cellular responses. FcγRI is a high-affinity receptor that binds IgG as a monomer, while FcγRIIA and FcγRIIA are low-affinity receptors that bind IgG only as multimers and with a slightly lower affinity. The binding of an antibody to an Fc receptor and / or C1q is controlled by specific residues or domains within the Fc region. Binding also depends on residues located within the hinge region and within the CH2 portion of the antibody. In some embodiments, the agonistic and / or therapeutic activity of the antibodies described herein depends on the binding of the Fc region to an Fc receptor (e.g., FcγR). In some embodiments, the agonistic and / or therapeutic activity of the antibodies described herein is enhanced by the binding of the Fc region to an Fc receptor (e.g., FcγR).

[0275] A list of certain Fc receptor sequences used in this disclosure is shown in Table 13 below.

[0276] As used herein, the term "glycosylation pattern" is defined as the pattern of carbohydrate units that are covalently linked to a protein, and more specifically, to an immunoglobulin. The glycosylation pattern of a heterologous antibody can be characterized as being substantially similar to the glycosylation pattern that is naturally present on antibodies produced by a non-human transgenic animal species, where one of ordinary skill in the art would recognize that the glycosylation pattern of the heterologous antibody is more similar to the glycosylation pattern in the non-human transgenic animal species than to the species from which the transgenic CH genes are derived.

[0277] As used herein, the term "human antibody" includes antibodies having variable and constant regions (if any) with human germline immunoglobulin sequences. The human antibodies of the disclosure may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by in vitro random or site-specific mutagenesis or by in vivo somatic mutation) (see, e.g., Lonberg et al., (1994) Nature 368(6474):856-859); Lonberg, (1994) Handbook of Experimental Pharmacology 113:49-101; Lonberg & Huszar, (1995) Intern. Rev. Immunol. 13:65-93, and Harding & Lonberg, (1995) Ann. N.Y. Acad. Sci. 764:536-546). However, the term "human antibody" does not include antibodies in which CDR sequences from the germline of another mammalian species, such as a mouse, have been grafted onto human framework sequences (i.e., humanized antibodies).

[0278] As used herein, the term "xenogeneic antibody" is defined relative to a transgenic non-human organism that produces such an antibody. The term refers to an antibody having an amino acid sequence or an encoding nucleic acid sequence corresponding to an amino acid sequence or an encoding nucleic acid sequence found in an organism that does not consist of the transgenic non-human animal and is typically from a species other than the transgenic non-human animal.

[0279] The terms "induce an immune response" and "enhance an immune response" are used interchangeably and refer to the stimulation of an immune response (i.e., passive or adaptive) to a particular antigen. The term "induce" as used with respect to inducing CDC or ADCC refers to the stimulation of a particular direct cell killing mechanism.

[0280] As used herein, the term "immunogenic cell death" (or "immunogenic apoptosis") refers to a form of cell death that is associated with the activation of one or more signaling pathways that induce the pre-death expression and release of damage-associated molecular pattern (DAMP) molecules (e.g., adenosine triphosphate, ATP) from tumor cells, resulting in enhanced immunogenicity of the tumor cells and the death of the tumor cells in an immunogenic manner (e.g., by phagocytosis). As used herein, the term "immunogenic cell death inducer" refers to a chemical, biological, or pharmaceutical agent that induces the process, pathway, or manner of immunogenic cell death.

[0281] As used herein, the terms "inhibit", "reduce" or "block" (e.g., referring to inhibiting or reducing human IL-27-mediated STAT1 and / or STAT3 phosphorylation in cells) are used interchangeably and include partial and complete inhibition / blockade. Inhibition / blockade of IL-27 reduces or alters the normal level or type of activity that would occur in the absence of inhibition or blockade. Inhibition and blockade are also intended to include any measurable decrease in the binding affinity of IL-27 when contacted with an anti-IL-27 antibody as compared to IL-27 not contacted with the anti-IL-27 antibody, e.g., inhibiting the binding of IL-27 by at least about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100%.

[0282] As used herein, the terms "inhibit angiogenesis", "reduce angiogenesis" and "decrease angiogenesis" refer to reducing the level of angiogenesis in a tissue to an amount that is at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99% or less lower than the amount in the corresponding control tissue, and most preferably the same level as observed in the control tissue.

[0283] As used herein, the term "inhibit growth" (e.g., as it pertains to cells) is intended to include any measurable decrease in cell growth, e.g., inhibiting cell growth by at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% or 100%.

[0284] As used herein, a subject "in need of prevention", "in need of treatment" or "in need thereof" is a subject who would reasonably benefit from a given treatment (such as treatment with a composition comprising an anti-IL-27 antibody) as determined by a suitable practicing physician (e.g., in the case of a human, a doctor, nurse or caregiver; in the case of a non-human mammal, a veterinarian).

[0285] The term "in vivo" refers to a process that occurs in a living organism.

[0286] As used herein, the term "isolated antibody" refers to an antibody that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to and antagonizes human IL-27 is substantially free of antibodies that specifically bind to antigens other than IL-27). However, an isolated antibody that specifically binds to an epitope may cross-react with other IL-27 proteins from different species. However, in the specific binding assays described herein, the antibody continues to show specific binding to human IL-27. Additionally, an isolated antibody is generally substantially free of other cellular materials and / or chemicals. In some embodiments, combinations of "isolated" antibodies having different IL-27 specificities are combined in a well-defined composition.

[0287] As used herein, the term "isolated nucleic acid molecule" refers to a nucleic acid encoding an antibody or antibody portion that binds to IL-27 (e.g., V H 、V L 、CDR3), and is intended to mean a nucleic acid molecule in which the nucleotide sequence encoding the antibody or antibody portion does not contain other nucleotide sequences encoding antibodies or antibody portions that bind to antigens other than IL-27, and such other sequences may naturally flank the nucleic acid in the human genomic DNA. For example, a sequence selected from the sequences shown in Table 12 corresponds to the nucleotide sequences of the heavy chain (V H ) and light chain (V L ) variable regions of the anti-IL-27 antibody monoclonal antibodies described herein.

[0288] As used herein, "isotype" refers to the class of antibody (such as IgM or IgG1) encoded by the heavy chain constant region gene. In some embodiments, the human monoclonal antibodies of the present disclosure are of the IgG1 isotype. In some embodiments, the human monoclonal antibodies of the present disclosure are of the IgG2 isotype. In some embodiments, the human monoclonal antibodies of the present disclosure are of the IgG3 isotype. In some embodiments, the human monoclonal antibodies of the present disclosure are of the IgG4 isotype. As will be apparent to those skilled in the art, the identification of antibody isotypes (e.g., IgG1, IgG2, IgG3, IgG4, IgM, IgA1 IgA2, IgD, and IgE) is routine in the art and generally involves a combination of sequence alignments with known antibodies, published Fc variant sequences, and conserved sequences.

[0289] As used herein, the term "isotype switching" refers to the phenomenon in which the class or isotype of an antibody changes from one Ig class to another.

[0290] As used herein, the term "KD" or "K D " refers to the equilibrium dissociation constant of the binding reaction between an antibody and an antigen. K DThe value is a numerical representation of the ratio of the antibody dissociation rate constant (kd) to the antibody association rate constant (ka). K D The value is inversely proportional to the binding affinity of the antibody for the antigen. K D The smaller the value, the greater the affinity of the antibody for its antigen. Affinity is the strength with which a single molecule binds to its ligand and is typically measured and reported by the equilibrium dissociation constant (K D ), which is used to evaluate and rank the strength of bimolecular interactions.

[0291] As used herein, the term "kd" or "k d " (or "koff" or "k off ") refers to the dissociation rate constant at which an antibody dissociates from an antibody / antigen complex. The value of kd is a numerical representation of the fraction of complexes that decay or dissociate per second, expressed in seconds -1 as the unit.

[0292] As used herein, the term "ka" or "k a " (or "kon" or "k on ") is intended to refer to the association rate constant at which an antibody associates with an antigen. The value of ka is a numerical representation of the number of antibody / antigen complexes formed per second in a 1 molar (1M) solution of antibody and antigen, expressed in M -1 seconds -1 as the unit.

[0293] As used herein, the term "leukocyte" refers to a type of white blood cell that participates in protecting the body against infectious organisms and foreign substances. Leukocytes are produced in the bone marrow. There are 5 main types of leukocytes, divided into two major categories: polymorphonuclear leukocytes (neutrophils, eosinophils, basophils) and mononuclear leukocytes (monocytes and lymphocytes).

[0294] As used herein, the term "lymphocyte" refers to a type of white blood cell or leukocyte that participates in the immune defense of the body. There are two main types of lymphocytes: B cells and T cells.

[0295] As used herein, the terms "linked", "fused", or "fusion" are used interchangeably. These terms refer to joining together two or more elements or components or domains by any means, including chemical conjugation or recombinant means. Methods of chemical conjugation (e.g., using heterobifunctional crosslinkers) are known in the art.

[0296] As used herein, "local administration" or "local delivery" refers to the delivery of a composition or agent that does not rely on transport of the composition or agent through the vascular system to its intended target tissue or site. For example, a composition can be delivered by injection or implantation of the composition or agent or by injection or implantation of a device containing the composition or agent. After local administration near the target tissue or site, the composition or agent or one or more of its components can diffuse into the intended target tissue or site.

[0297] As used herein, "MHC molecule" refers to two types of molecules: MHC class I and MHC class II. MHC class I molecules present antigen to specific CD8+ T cells, and MHC class II molecules present antigen to specific CD4+ T cells. Antigens delivered exogenously to an APC are primarily processed for association with MHC class II of the APC. In contrast, antigens delivered endogenously to an APC are primarily processed for association with MHC class I.

[0298] As used herein, the term "monoclonal antibody" refers to an antibody that exhibits a single binding specificity and affinity for a particular epitope. Thus, the term "human monoclonal antibody" refers to an antibody that exhibits a single binding specificity and has variable regions and optionally constant regions derived from human germline immunoglobulin sequences. In some embodiments, a human monoclonal antibody is produced by a hybridoma that includes a B cell obtained from a transgenic non-human animal (e.g., a transgenic mouse) fused to an immortalized cell, the animal having a genome that includes a human heavy chain transgene and a light chain transgene.

[0299] As used herein, the term "monocyte" refers to a type of white blood cell that can differentiate into macrophages and dendritic cells to effect an immune response.

[0300] As used herein, the term "natural killer (NK) cell" refers to a type of cytotoxic lymphocyte. These are large, usually granular, non-T, non-B lymphocytes that kill certain tumor cells and play important roles in innate immunity against viruses and other intracellular pathogens and in antibody-dependent cell-mediated cytotoxicity (ADCC).

[0301] As used herein, the term "naturally occurring" as applied to an object refers to the fact that the object can exist in nature. For example, a polypeptide or polynucleotide sequence that exists in a living organism (including a virus) is naturally occurring, and the polypeptide or polynucleotide sequence can be isolated from a natural source and has not been intentionally modified by a human in the laboratory.

[0302] As used herein, the term "non-switched isotype" refers to the isotype class of the heavy chain that is produced when isotype switching has not occurred; the CH gene encoding the non-switched isotype is typically the first CH gene immediately downstream of the functionally rearranged VDJ gene. Isotype switching is divided into classical isotype switching and non-classical isotype switching. Classical isotype switching occurs through a recombination event involving at least one switch sequence region in the transgenic. Non-classical isotype switching can occur, for example, through homologous recombination (delta-related deletion) between human sigma μ and human sigma μ . Alternative non-classical switching mechanisms, such as intergenic and / or interchromosomal recombination, can occur and effect isotype switching.

[0303] As used herein, the term "nucleic acid" refers to deoxyribonucleotides or ribonucleotides and polymers thereof in single-stranded or double-stranded form. Unless specifically limited otherwise, the term includes nucleic acids containing known analogs of natural nucleotides which have similar binding properties as the reference nucleic acid and are metabolized in a manner similar to naturally occurring nucleotides. Unless otherwise indicated, a particular nucleic acid sequence also implicitly encompasses conservatively modified variants thereof (e.g., degenerate codon substitutions) and complementary sequences as well as the explicitly recited sequence. Specifically, degenerate codon substitutions may be achieved by generating sequences in which the third position of one or more selected (or all) codons is substituted with a mixture of bases and / or deoxyinosine residues (Batzer et al., Nucleic Acid Res. 19:5081, 1991; Ohtsuka et al., Biol. Chem. 260:2605-2608, 1985; and Cassol et al., 1992; Rossolini et al., Mol. Cell. Probes 8:91-98, 1994). For arginine and leucine, modifications at the second base can also be conservative. The term nucleic acid may be used interchangeably with gene, cDNA, and mRNA encoded by a gene.

[0304] The polynucleotides used herein can be composed of any polyribonucleotides or polydeoxyribonucleotides, which can be unmodified RNA or DNA or modified RNA or DNA. For example, polynucleotides can be composed of single-stranded and double-stranded DNA, DNA that is a mixture of single-stranded and double-stranded regions, single-stranded and double-stranded RNA, and RNA that is a mixture of single-stranded and double-stranded regions, hybrid molecules comprising DNA and RNA that can be single-stranded regions or more commonly double-stranded regions or mixtures of single-stranded and double-stranded regions. Additionally, polynucleotides can be composed of triple-stranded regions comprising RNA or DNA or RNA and DNA. Polynucleotides can also comprise one or more modified bases or a DNA or RNA backbone modified for stability or other reasons. "Modified" bases include, for example, tritylated bases and rare bases such as inosine. A variety of modifications can be made to DNA and RNA; thus, "polynucleotide" includes chemically, enzymatically, or metabolically modified forms.

[0305] When a nucleic acid is in a functional relationship with another nucleic acid sequence, it is "operably linked". For example, if a promoter or enhancer affects the transcription of a sequence, it is operably linked to the coding sequence. In the context of transcriptional regulatory sequences, operably linked means that the DNA sequences being linked are contiguous and, where necessary to join two protein-coding regions (the two protein-coding regions being contiguous and in-frame). For transposable sequences, operably linked means that the sequence is capable of effecting transpositional recombination.

[0306] As used herein, "parenteral administration", "parenterally administered", and other grammatically equivalent phrases refer to modes of administration other than enteral and topical administration, typically by injection, including but not limited to intravenous, intranasal, intraocular, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, intratracheal, subcutaneous, subepidermal, intraarticular, subcapsular, subarachnoid, intraspinal, epidural, intracerebral, intracranial, intracarotid, and intrasternal injection and infusion.

[0307] As used herein, the term "patient" includes humans and other mammalian subjects receiving prophylactic or therapeutic treatment.

[0308] As used herein, the term "PD-1 antagonist" refers to any compound or biomolecule that inhibits the PD-1 signaling pathway or otherwise inhibits the function of PD-1 in a cell (e.g., an immune cell). In some embodiments, the PD-1 antagonist blocks the binding of PD-L1 to PD-1 and / or the binding of PD-L2 to PD-1. In some embodiments, the PD-1 antagonist specifically binds to PD-1. In some embodiments, the PD-1 antagonist specifically binds to PD-L1.

[0309] In the context of two or more nucleic acid or polypeptide sequences, the term "percent identity" refers to two or more sequences or subsequences having a specified percentage of identical nucleotides or amino acids, when compared and aligned for maximum correspondence, as measured using one of the following sequence comparison algorithms (e.g., BLASTP and BLASTN, or other algorithms available to those of skill in the art) or by visual inspection. Depending on the application, the "percent identity" may exist over the region of the sequences being compared, e.g., over a functional domain, or over the full length of the two sequences being compared. For sequence comparison, typically one sequence acts as a reference sequence to which the test sequence is compared. When using a sequence comparison algorithm, the test and reference sequences are input into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. The sequence comparison algorithm then calculates the percent sequence identity for one or more test sequences relative to the reference sequence, based on the designated program parameters.

[0310] Optimal alignment of sequences for comparison can be conducted, e.g., by the local homology algorithm of Smith & Waterman, Adv. Appl. Math. 2:482 (1981), by the homology alignment algorithm of Needleman & Wunsch, J. Mol. Biol. 48:443 (1970), by the similarity search method of Pearson & Lipman, Proc. Nat'l Acad. Sci. USA 85:2444 (1988), by computerized implementations of these algorithms (GAP, BESTFIT, FASTA, and TFASTA in the Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Dr., Madison, Wis.), or by visual inspection (generally see Ausubel et al., infra).

[0311] An example of an algorithm suitable for determining percent sequence identity and sequence similarity is the BLAST algorithm, which is described in Altschul et al., J. Mol. Biol. 215:403-410 (1990). Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information website.

[0312] As used herein generally, "pharmaceutically acceptable" means those compounds, materials, compositions, and / or dosage forms that, within the scope of sound medical judgment, are suitable for use in contact with the tissues, organs, and / or bodily fluids of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit / risk ratio.

[0313] As used herein, "pharmaceutically acceptable carrier" means and includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. The compositions can include pharmaceutically acceptable salts, such as acid addition salts or base addition salts (see, e.g., Berge et al. (1977) J Pharma Sci 66 : 1-19).

[0314] As used herein, the terms "polypeptide", "peptide" and "protein" are used interchangeably and refer to polymers of amino acid residues. The terms apply to amino acid polymers in which one or more amino acid residues are artificial chemical mimics of the corresponding naturally occurring amino acids, as well as to naturally occurring amino acid polymers and non-naturally occurring amino acid polymers.

[0315] As used herein, the term "prevent", when used in connection with a disorder, refers to the administration of a composition that reduces the frequency of symptoms of the disorder or delays its onset in a subject relative to a subject who has not received the composition.

[0316] As used herein, the terms "purified" or "isolated" for any protein (antibody or fragment) described herein refer to a polypeptide that has been separated or purified from the components that naturally accompany it (e.g., proteins or other naturally occurring biological or organic molecules) (e.g., other proteins, lipids, and nucleic acids in a prokaryote that expresses the protein). Generally, a polypeptide is purified when it constitutes at least 60% by weight (e.g., at least 65%, 70%, 75%, 80%, 85%, 90%, 92%, 95%, 97% or 99%) of the total protein in a sample.

[0317] As used herein, the term "programmed cell death protein 1" or "PD-1" refers to the programmed cell death protein 1 polypeptide (an immunosuppressive receptor belonging to the CD28 family) and is encoded by the PDCD1 gene in humans. Alternative names or synonyms for PD-1 include: PDCD1, PD1, CD279, and SLEB2. PD-1 is mainly expressed in vivo on previously activated T cells, B cells, and myeloid cells and binds to two ligands, PD-L1 and PD-L2. As used herein, the term "PD-1" includes human PD-1 (hPD-1), variants, isotypes, and species homologs of hPD-1, and analogs that share at least one common epitope with hPD-1. The complete hPD-1 sequence can be found under GenBank accession number AAC51773.

[0318] As used herein, the term "programmed death ligand-1" or "PD-L1" is one of two cell surface glycoprotein ligands of PD-1 (the other being PD-L2), which downregulates T cell activation and cytokine secretion upon binding to PD-1. Alternative names and synonyms for PD-L1 include: PDCD1L1, PDL1, B7H1, B7-4, CD274, and B7-H. The term "PD-L1" as used herein includes human PD-L1 (hPD-L1), variants, isotypes, and species homologs of hPD-L1, as well as analogs having at least one common epitope with hPD-L1. The complete hPD-L1 sequence can be found under GenBank accession number Q9NZQ7.

[0319] PD-1 is referred to as an immunosuppressive protein that negatively regulates TCR signaling (Ishida, Y. et al. (1992) EMBO J. 11:3887-3895; Blank, C. et al. (Epub Dec 29, 2006) Immunol. Immunother. 56(5):739-745). The interaction between PD-1 and PD-L1 can act as an immune checkpoint, which can lead to reduced T cell receptor-mediated proliferation (Dong et al. (2003) J. Mol. Med. 81:281-7; Blank et al. (2005) Cancer Immunol. Immunother. 54:307-314; Konishi et al. (2004) Clin. Cancer Res. 10:5094-100). Immunosuppression can be reversed by inhibiting the local interaction of PD-1 with PD-L1 or PD-L2; the effect is additive when the interaction of PD-1 with PD-L2 is also blocked (Iwai et al. (2002) Proc. Nat'l. Acad. Sci. USA 99:12293-7; Brown et al. (2003) J. Immunol. 170:1257-66).

[0320] For several cancers, the survival and proliferation of tumors are maintained by tumor-mediated immune checkpoint regulation. This regulation can lead to disruption of the function of the anti-cancer immune system. For example, recent studies have shown that tumor cells expressing immune checkpoint receptor ligands, such as PD-L1 or PD-L2, can specifically downregulate immune system activity in the tumor microenvironment and promote cancer immune escape by inhibiting T cells. PD-L1 is highly expressed in a variety of human cancers (Dong et al., (2002) Nat Med 8:787-789). The PD-L1 receptor PD-1 is expressed on lymphocytes (e.g., activated T cells) and is normally involved in downregulating (specifically by inhibiting T cells) the immune system and promoting self-tolerance. However, when the PD-1 receptor expressed on T cells binds to the cognate PD-L1 ligand on tumor cells, the resulting T cell inhibition leads to impaired immune responses against the tumor (e.g., reduced tumor-infiltrating lymphocytes or cancer cells establishing immune escape).

[0321] In large cohorts of, for example, ovarian, renal, colorectal, pancreatic, hepatic, and melanoma cancers, it has been shown that PD-L1 expression is associated with poor prognosis and reduced overall survival regardless of subsequent treatment (see, e.g., Dong et al., (2002) Nat Med 8(8):793-800; Yang et al., (2008) Invest Ophthalmol Vis Sci 49(6):2518-2525; Ghebeh et al., (2006) Neoplasia 8:190-198; Hamanishi et al., (2007) Proc NatAcad Sci USA 104:3360-3365; Thompson et al., (2006) Clin Genitourin Cancer 5:206-211; Nomi et al., (2005) Clin Cancer Res 11:2947-2953; Inman et al., (2007) Cancer 109:1499-1505; Shimauchi et al., (2007) Int J Cancer 121:2585-2590; Gao et al., (2009) ClinCancer Res 15:971-979; Nakanishi et al., (2007) Cancer Immunol Immunother 56:1173-1182; Hino et al., (2010) Cancer 116(7):1757-1766). Similarly, PD-1 expression on tumor lymphocytes has been found to mark dysfunctional T cells in breast cancer (Kitano et al., (2017) ESMO Open2(2):e000150) and melanoma (Kleffel et al., (2015) Cell 162(6):1242-1256). PD-1 antagonists, such as those that affect the function of the PD-1 / PD-L1 / PD-L2 signaling axis and / or disrupt the interaction between PD-1 and PD-L1 and / or PD-L2, have been developed and represent a novel class of anti-tumor inhibitors that act by modulating immune cell-tumor cell interactions.

[0322] As used herein, the term "rearranged" refers to the configuration of a heavy or light chain immunoglobulin locus in which the V segments are in a conformation encoding substantially intact V H or V L domain immediately adjacent to the D-J or J segments. Rearranged immunoglobulin loci can be identified by comparison to germline DNA; a rearranged locus will have at least one recombined heptamer / nonamer homology element.

[0323] As used herein, the term "recombinant host cell" (or simply "host cell") is intended to refer to a cell into which a recombinant expression vector has been introduced. It should be understood that such terms refer not only to the particular subject cell but also to the progeny of such a cell. Because certain modifications may occur in succeeding generations due to either mutation or environmental influences, such progeny may not, in fact, be identical to the parental cell, but are still included within the scope of the term "host cell" as used herein.

[0324] As used herein, the term "recombinant human antibody" includes all human antibodies prepared, expressed, produced or isolated by recombinant means, such as (a) antibodies isolated from an animal (e.g., a mouse) or a hybridoma made therefrom that is transgenic or chromosomally modified for human immunoglobulin genes, (b) antibodies isolated from a host cell transformed to express the antibody, e.g., from a transfectoma, (c) antibodies isolated from a recombinant combinatorial human antibody library, and (d) antibodies prepared, expressed, produced or isolated by any other means that involve splicing human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies comprise variable and constant regions utilizing specific human germline immunoglobulin sequences, the variable and constant region sequences being encoded by germline genes, but including subsequent rearrangements and mutations that occur, for example, during antibody maturation. As is known in the art (see, e.g., Lonberg (2005) Nature Biotech. 23(9):1117-1125), the variable region contains the antigen-binding domain, which is encoded by various genes that rearrange to form an antibody specific for a foreign antigen. In addition to rearrangement, the variable region may be further modified by multiple single amino acid changes (referred to as somatic mutation or hypermutation) to increase the affinity of the antibody for the foreign antigen. The constant region will change in response to further antigenic challenge (i.e., isotype switching). Thus, the nucleic acid molecules encoding the light and heavy chain immunoglobulin polypeptides that rearrange and are somatically mutated in response to antigen may not have exactly the same sequence as the original nucleic acid molecules, but rather have substantially the same or similar (i.e., having at least 80% identity).

[0325] As used herein, the term "reference antibody" (which may be used interchangeably with "reference mAb") or "reference antigen-binding protein" is an antibody or antigen-binding fragment thereof that binds to a specific epitope on IL-27 and is used to establish a relationship between itself and one or more different antibodies, wherein the relationship is the binding of the reference antibody and one or more different antibodies to the same epitope on IL-27. As used herein, the term means an anti-IL-27 antibody that can be used in a test or assay, such as those described herein (e.g., a competitive binding assay), as a competitor, wherein the assay can be used to discover, identify or develop one or more different antibodies that bind to the same epitope.

[0326] As used herein, the terms "specific binding", "selective binding", "selectively binds" and "specifically binds" refer to the binding of an antibody to an epitope on a predetermined antigen. Generally speaking, when using recombinant human IL-27 as an analyte and an antibody as a ligand measured by surface plasmon resonance (SPR) technology in a BIACORE 2000 instrument, the antibody has a specific binding affinity of about less than 10 -6 M, such as less than about 10 -7 , 10 -8 M, 10 -9 M or 10 -10 M or even lower equilibrium dissociation constant (K D ) and its affinity for binding to a predetermined antigen is at least twice that of its affinity for binding to a non-specific antigen (e.g., BSA, casein) other than the predetermined antigen or a closely related antigen. In certain embodiments, when using recombinant human IL-27 as an analyte and the antibody as a ligand in a BIACORE 2000 instrument by surface plasmon resonance (SPR) technology, the antibody specifically binds to IL-27 at a concentration of less than about 100 nM (10 -7 M), optionally less than about 50 nM (5 x 10 -8 M), optionally less than about 15 nM (1.5 x 10 -8 M), optionally less than about 10 nM (10 -8 M), optionally less than about 5 nM (5 x 10 -9 M), optionally less than about 1 nM (10 -9 M), optionally less than about 0.1 nM (10 -10 M), optionally less than about 0.01 nM (10 -11 M) or an even smaller equilibrium dissociation constant (K D ), wherein the binding to the predetermined antigen occurs with an affinity at least 2 times greater than the affinity of the antibody for binding to a non-specific antigen (e.g., BSA, casein) other than the predetermined antigen or a closely related antigen. The phrases "antibodies that recognize antigens" and "antibodies specific for antigens" are used interchangeably herein with the term "antibodies that specifically bind to antigens".

[0327] As used herein, the term "STAT1 phosphorylation" refers to the phosphorylation of signal transducer and activator of transcription 1 (STAT1) polypeptide, a transcription factor encoded by the human STAT1 gene. STAT molecules are phosphorylated by receptor-associated kinases, leading to activation and dimerization by forming homodimers or heterodimers, which translocate to the nucleus and function as transcription factors. STAT1 can be activated (i.e., phosphorylated) in response to signal transduction by several ligands, including IL-27. IL-27 signals through the IL-27R, resulting in STAT1 phosphorylation (pSTAT1). STAT1 plays a key role in gene expression involved in cell survival, viability, or pathogen response. Methods for determining STAT1 phosphorylation as a result of IL-27 signaling include, but are not limited to, flow cytometric analysis of cells labeled with an antibody that specifically recognizes phosphorylated STAT1 (see, e.g., Tochizawa et al., (2006) J Immunol Methods 313(1-2):29-37).

[0328] As used herein, the term "STAT3 phosphorylation" refers to the phosphorylation of signal transducer and activator of transcription 3 (STAT3) polypeptide, a transcription factor encoded by the human STAT3 gene. STAT3 mediates the expression of multiple genes in response to cellular stimuli and thus plays a key role in many cellular processes such as cell growth and apoptosis. Methods for determining STAT3 phosphorylation as a result of IL-27 signaling include, but are not limited to, analysis of cells or cell extracts labeled with an antibody that specifically recognizes phosphorylated STAT3 (see, e.g., Fursov et al., (2011) Assay Drug Dev Technol 9(4):420-429).

[0329] As used herein, the term "switch sequence" refers to those DNA sequences responsible for switch recombination. The "switch donor" sequence, typically the μ switch region, will be 5' (i.e., upstream) of the construct region that will be deleted during switch recombination. The "switch acceptor" region will be between the construct region to be deleted and the replacement constant region (e.g., γ, ε, etc.). Since there is no specific site where recombination always occurs, the final gene sequence usually cannot be predicted from the construct.

[0330] As used herein, the term "subject" includes any human or non-human animal. For example, the methods and compositions of the present invention can be used to treat a subject suffering from an immune disorder. The term "non-human animal" includes all vertebrates, such as mammals and non-mammals, such as non-human primates, sheep, dogs, cows, chickens, amphibians, reptiles, etc.

[0331] For nucleic acids, the term "substantial homology" means that two nucleic acids or their designated sequences, when optimally aligned and compared, are identical in at least about 80% of the nucleotides, usually at least about 90% to 95%, more preferably at least about 98% to 99.5% of the nucleotides, with appropriate nucleotide insertions or deletions. Alternatively, substantial homology exists when a segment hybridizes to the complementary sequence of a strand under selective hybridization conditions.

[0332] The percent identity between two sequences is a function of the number of identical positions shared by the sequences (i.e., % homology = number of identical positions / total number of positions × 100), where account is taken of the number of gaps that need to be introduced to optimally align the two sequences and the length of each gap. The comparison of sequences and the determination of the percent identity between two sequences can be accomplished using a mathematical algorithm as described in the following non-limiting examples.

[0333] The percent identity between two nucleotide sequences can be determined using the GAP program in the GCG software package (available at http: / / www.gcg.com), using the NWSgapdna.CMP matrix and gap weights of 40, 50, 60, 70, or 80 and length weights of 1, 2, 3, 4, 5, or 6. The percent identity between two nucleotide or amino acid sequences can also be determined using the algorithm of E. Meyers and W. Miller (CABIOS, 4:11-17 (1989)) incorporated into the ALIGN program (version 2.0), using the PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4. Additionally, the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol. (48):444-453 (1970)) algorithm incorporated into the GAP program in the GCG software package (available at http: / / www.gcg.com), using the Blossum62 matrix or the PAM250 matrix, gap weights of 16, 14, 12, 10, 8, 6, or 4, and length weights of 1, 2, 3, 4, 5, or 6.

[0334] The nucleic acid and protein sequences of the present disclosure can be further used as "query sequences" to perform searches against public databases to identify, for example, related sequences. Such searches can be carried out using the NBLAST and XBLAST programs (version 2.0) of Altschul et al. (1990) J. Mol. Biol. 215: 403-10. BLAST nucleotide searches can be performed using the NBLAST program, score = 100, word length = 12, to obtain nucleotide sequences homologous to the nucleic acid molecules of the present invention. BLAST protein searches can be performed using the XBLAST program, score = 50, word length = 3, to obtain amino acid sequences homologous to the protein molecules of the present invention. To obtain gapped alignments for comparison purposes, gapped BLAST can be used as described in Altschul et al., (1997) Nucleic Acids Res. 25(17): 3389-3402. When using the BLAST and gapped BLAST programs, the default parameters of the corresponding programs (e.g., XBLAST and NBLAST) can be used. See http: / / www.ncbi.nlm.nih.gov.

[0335] The nucleic acid can exist in the form of whole cells, cell lysates, or partially purified or substantially pure forms. The nucleic acid is "isolated" or "substantially purified" when purified from other cellular components or other contaminants (e.g., other cellular nucleic acids or proteins) by standard techniques including alkaline / SDS treatment, CsCl banding, column chromatography, agarose gel electrophoresis, and other techniques known in the art. See, F. Ausubel et al., eds Current Protocols in Molecular Biology, Greene Publishing and Wiley Interscience, New York (1987).

[0336] The nucleic acid compositions of the present disclosure (although usually natural sequences except for modified restriction sites, etc.) from cDNA, genomic, or mixtures thereof can be mutated according to standard techniques for providing gene sequences. For coding sequences, these mutations can affect the amino acid sequence as desired. In particular, DNA sequences that are substantially homologous to or derived from the native V sequences, D sequences, J sequences, constant sequences, switch sequences, and other such sequences described herein are contemplated (where "derived from" means that the sequence is the same as or modified from another sequence).

[0337] As used herein, the term "STING" (or TMEM173) refers to the stimulator of interferon genes, which is a protein that functions both as a direct cytosolic DNA sensor and as an adaptor protein. In humans, STING is encoded by the TMEM173 gene. STING plays an important role in innate immunity. When cells are infected with intracellular pathogens such as viruses, mycobacteria, and intracellular parasites, STING induces the production of type I interferons. Type I interferons, mediated by STING, protect infected cells and neighboring cells from local infection by binding to the same cell that secreted them and neighboring cells. The NCBI Genbank database provides an exemplary amino acid sequence of STING under accession number NP_001288667.

[0338] The term "T cell" refers to a type of white blood cell that is distinguished from other white blood cells by the presence of T cell receptors on its cell surface. There are several T cell subsets, including but not limited to T helper cells (also known as T H cells or CD4 + T cells) and subtypes, including T H 1, T H 2, T H 3, T H 17, T H 9 and T FH cells, cytotoxic T cells (also known as T C cells, CD8 + T cells, cytotoxic T lymphocytes, T-killer cells, killer T cells), memory T cells and subtypes, including central memory T cells (T CM cells), effector memory T cells (T EM and T ERMA cells) and resident memory T cells (T RM cells), regulatory T cells (also known as T reg cells or inhibitory T cells) and their subtypes, including CD4 + FOXP3 + T reg cells, CD4 + FOXP3 - T reg cells, Tr1 cells, Th3 cells and T reg 17 cells, natural killer T cells (also known as NKT cells), mucosal associated invariant T cells (MAIT), and γδ T cells (γδ T cells), including Vγ9 / Vδ2 T cells. Any one or more of the foregoing or unmentioned T cells can be the target cell type for the methods of use of the present invention.

[0339] As used herein, the term "T cell-mediated response" refers to any response mediated by T cells (including but not limited to effector T cells (e.g., CD8 + cells) and helper T cells (e.g., CD4 + cells)). T cell-mediated responses include, for example, the cytotoxicity and proliferation of T cells.

[0340] As used herein, the term "therapeutically effective amount" or "therapeutically effective dose" or similar terms used herein are intended to mean an amount of an agent (e.g., an anti-IL-27 antibody or an antigen-binding fragment thereof) that will elicit a desired biological or medical response (e.g., improvement of one or more symptoms of cancer).

[0341] As used herein, the term "TAM receptor" refers to the TAM receptor protein tyrosine kinases (TYRO3, AXL, and MER). TAM receptors are involved in the regulation of immune system homeostasis. In the context of cancer, TAM receptors have a dual regulatory role, controlling the initiation and progression of tumor development while controlling the associated anti-tumor responses of different immune cells. Further description of TAM receptors can be found in Paolino and Penninger (2016) Cancers 8(97): doi:10.3390 / cancers8100097). As used herein, the term "TAM receptor inhibitor" or "TAM inhibitor" refers to an agent that inhibits, blocks, or reduces the function or activity of a TAM receptor.

[0342] As used herein, unless otherwise specified, the term "TIGIT" or "T cell immunoreceptor with Ig and ITIM domains" refers to any native TIGIT from any vertebrate source, including mammals such as primates (e.g., humans) and rodents (e.g., mice and rats). TIGIT is also known in the art as DKFZp667A205, FLJ39873, protein containing V-set and immunoglobulin domains 9, protein containing V-set and transmembrane domains 3, VSIG9, VSTM3, and WUCAM. The term also includes naturally occurring variants of TIGIT, such as splice variants or allelic variants. The amino acid sequence of an exemplary human TIGIT can be found under UniProt accession number Q495A1.

[0343] As used herein, the terms “treat,” “treating,” and “treatment” refer to therapeutic or prophylactic measures. A method of “treatment” involves administering a human antibody of the present disclosure to a subject in need thereof (e.g., a subject in need of an enhanced immune response against a specific antigen or a subject that may ultimately develop such a disorder) to prevent, cure, delay, reduce the severity of, or ameliorate one or more symptoms of the disorder or recurrence of the disorder, or to prolong the survival of the subject beyond that expected in the absence of such treatment.

[0344] As used herein, the term “tumor microenvironment” (or “cancer microenvironment”; abbreviated as TME) refers to the cellular environment or context in which a tumor or neoplasm resides, including the surrounding vasculature as well as non-cancerous cells, including but not limited to immune cells, fibroblasts, bone marrow-derived inflammatory cells, and lymphocytes. Signal molecules and the extracellular matrix also constitute the TME. The tumor and the surrounding microenvironment are closely related and constantly interact. The tumor can affect the microenvironment by releasing extracellular signals, promoting tumor angiogenesis, and inducing peripheral immune tolerance, while immune cells in the microenvironment can affect the growth and evolution of tumor cells.

[0345] As used herein, the term “unrearranged” or “germline configuration” refers to the configuration of a V segment in which the V segment does not recombine to be in close proximity to a D or J segment.

[0346] As used herein, the term “vector” is intended to refer to a nucleic acid molecule capable of transporting another nucleic acid molecule to which it is linked. One type of vector is a “plasmid,” which refers to a circular double-stranded DNA loop into which additional DNA segments can be ligated. Another type of vector is a viral vector, into which additional DNA segments can be ligated into the viral genome. Certain vectors are capable of autonomous replication in a host cell into which they are introduced (e.g., bacterial vectors having a bacterial origin of replication and episomal mammalian vectors). Other vectors (e.g., non-episomal mammalian vectors) can integrate into the genome of the host cell upon introduction into the host cell and thereby replicate with the host genome. In addition, certain vectors are capable of directing the expression of genes operably linked to them. Such vectors are referred to herein as “recombinant expression vectors” (or simply “expression vectors”). Typically, expression vectors useful in recombinant DNA techniques are in the form of plasmids. In the present specification, “plasmid” and “vector” may be used interchangeably because plasmids are the most commonly used form of vector. However, the present invention is intended to include such other forms of expression vectors, such as viral vectors (e.g., replication-defective retroviruses, adenoviruses, and adeno-associated viruses), which have equivalent functions.

[0347] Unless otherwise defined, 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 disclosure belongs. Preferred methods and materials are described below, although methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the presently disclosed methods and compositions. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. BRIEF DESCRIPTION OF THE DRAWINGS

[0348] Figure 1 is a table providing affinity data for anti-IL-27 antibodies, as indicated. Affinity measurements were performed using ForteBio and MesoScale Discovery methods.

[0349] Figure 2 is a graph depicting the binding of an anti-IL-27 antibody to recombinant IL-27 bound to a plate, as measured by ELISA, as indicated.

[0350] Figures 3A - 3E are a bar graph, two graphs, a bar graph, and a line graph, respectively. Figure 3A is a graph depicting the inhibition of IL-27-mediated STAT1 phosphorylation in human whole blood by an anti-IL-27 antibody, as measured by flow cytometry, as indicated. Figure 3B is a graph depicting the inhibition of IL-27-mediated STAT1 phosphorylation in human PBMCs by an anti-IL-27 antibody, as measured by flow cytometry, as indicated. Figure 3C is a graph depicting the inhibition of IL-27-mediated STAT1 phosphorylation in U937 cells by an anti-IL-27 antibody, as measured by flow cytometry, as indicated. Figure 3D is a graph depicting the inhibition of IL-27-mediated STAT1 phosphorylation in HUT-78 cells by an anti-IL-27 antibody, as measured by flow cytometry, as indicated. Figure 3E is a graph showing the inhibition of IL-27-mediated pSTAT1 in human whole blood T cells by SRF388.

[0351] Figure 4 is a graph depicting the reversal of the inhibition of IL-27-mediated CD161 expression in T cells by a series of concentrations of an anti-IL-27 antibody, as indicated. CD161 expression was determined by flow cytometry.

[0352] Figure 5A is a graph depicting the extent to which an anti-IL-27 antibody enhances PD-1-mediated TNFα secretion in human PBMCs, as measured by ELISA. Figure 5BA graph depicting the extent to which an anti-IL-27 antibody enhances PD-1-mediated IL-6 secretion in human PBMCs as measured by ELISA. Figure 5C A dot plot showing that the combination of SRF388 and PD-1 blockade results in increased cytokine production in PBMCs from healthy donors and RCC patients (Abbreviations: CBA = Cytometric Bead Array, IFNγ = interferon γ, MSD = MesoScale Discovery, PBMC = peripheral blood mononuclear cells, PD-1 = programmed death receptor-1, RCC = renal cell carcinoma, TNFα = tumor necrosis factor α). Figure 5D Showing that IL-27 inhibits cytokine production after PD-1 blockade and is restored when combined with SRF388 (Abbreviations: Ctrl = control, ns = not significant, PBMC = peripheral blood mononuclear cells, rhIL-27 = recombinant human IL-27). Figure 5E Outlining the observed cytokine induction in PBMC cultures of such cells (specifically, TNFα, IFNγ, IL-6, and IL-17A) when various specified types of cells are contacted with SRF388 antibody, αPD-1 antibody, or a combination of SRF388 and αPD-1 antibody. Figure 5F Showing the effect of different concentrations of the specified individual antibodies (SRF405, SRF410, SRF411, SRF414, SRF416, SRF536, SRF543, SRF529, SRF381, SRF388, and Ab7) on pSTAT1 signaling in U937 (lymphoma) cells. Figure 5G Showing the effect of different concentrations of these individual antibodies on pSTAT1 signaling in PBMCs (peripheral blood mononuclear cells). Figure 5H Showing the effect of different concentrations of these individual antibodies on CD161 signaling in PBMCs (peripheral blood mononuclear cells). Figure 5I Showing the effect of different concentrations of these individual antibodies (excluding SRF414) on PD-L1 signaling in CD4 T lymphocytes (CD4 cells). Figure 5J Showing the effect of different concentrations of these individual antibodies (excluding SRF529) on PD-L1 signaling in monocytes. Figure 5K Showing the effect of different concentrations of these individual antibodies (excluding SRF529) on TIM-3 signaling in monocytes.

[0353] Figure 6A A graph depicting the inhibition of IL-27-mediated PD-L1 expression resulting from treatment of human monocytes with an anti-IL-27 antibody as determined by flow cytometry. Figure 6BA graph depicting the dose-dependent inhibition of IL-27-mediated PD-L1 expression resulting from treating human monocytes with a series of concentrations of an anti-IL-27 antibody that specifically binds to the EBI3 monomer, as measured by flow cytometry. Figure 6C A graph depicting the inhibition of IL-27-mediated TIM3 expression resulting from treating human monocytes with an anti-IL-27 antibody, as measured by flow cytometry. Figure 6D A graph depicting the inhibition of IL-27-mediated PD-L1 expression resulting from treating resting human T cells with an anti-IL-27 antibody, as measured by flow cytometry.

[0354] Figure 7A A dot plot depicting the number of surface lung B16F10 metastatic nodules (lung nodules) from B16F10 tumor-bearing mice treated with an anti-IL27 antibody (SRF388), an isotype control antibody, an αWSX-1 antibody, or a combination of αPD-1 and αCTLA-4 antibodies, as determined by visual counting of nodules from lungs isolated from mice, as indicated. Figure 7B A graph providing the growth kinetics of bioluminescent B16-Luc tumors in mice treated with an anti-IL-27 antibody (SRF388) or an isotype control antibody, as measured by bioluminescence imaging analysis. Figure 7C A series of images of fixed, sectioned lung tissue stained with hematoxylin and eosin are shown, as indicated, which were isolated from B16F10 tumor-bearing mice treated with an anti-IL27 antibody (SRF388), an isotype control antibody, an αWSX-1 antibody, or a combination of αPD-1 and αCTLA-4 antibodies. Figure 7D A dot plot depicting the total tumor area, as indicated, expressed as a percentage of the total tissue area of fixed, sectioned lung tissue stained with hematoxylin and eosin that is occupied by B16F10 tumor tissue, as determined by image analysis software. The B16F10 tumor tissue in the lung tissue was isolated from B16F10 tumor-bearing mice treated with an anti-IL27 antibody (SRF388), an isotype control antibody, an αWSX-1 antibody, or a combination of αPD-1 and αCTLA-4 antibodies. Similar decreases in the number of surface lung metastases and total tumor area were observed for IL-27RA (WSX-1)-mediated antibody blockade and for the anti-PD-1 + anti-CTLA-4 combination therapy.

[0355] Figure 8A A scatter plot providing microarray data of genes with an expression change greater than >1.0 log2 fold change (black dots) in splenocytes isolated from mice overexpressing IL-27 after treatment with an IL-27 micro-ring. Figure 8B A graph is provided depicting as indicated, as Figure 8AGraph of the expression levels of selected immunomodulatory genes in splenocytes. Figure 8C Ectopic expression of human IL-27 was shown to induce inhibitory receptor expression on murine T cells in vivo, and SRF388 decreased inhibitory receptor expression on T cells in vivo after IL-27 micro-ring treatment. Six-week-old female Balb / c mice were injected with empty vector (control) or hIL-27 micro-ring. Five days after transfection, peripheral blood mononuclear cells (PBMCs) (upper left and upper right panels) and total splenocytes (lower left and lower right panels) were collected, stained, and analyzed by flow cytometry. Expression of the indicated markers was analyzed on CD4+ T cells (upper left and lower left panels) and CD8+ T cells (upper right and lower right panels). Analysis was performed using FlowJo software. Figure 8D SRF388 was shown to inhibit detection of micro-ring-derived human IL-27 in murine plasma.

[0356] Figure 9 An overview list of the characteristics of selected monoclonal antibodies is presented.

[0357] Figure 10A A chart of the antibody sequences is presented, where the sequence partitions reflect NT numbering. Figure 10B A chart of the antibody sequences (corresponding to the sequence chart of Figure 10A ) is presented, where the sequence partitions reflect ImMunoGeneTics (IMGT) numbering. In both Figure 10A and Figure 10B , the amino acids highlighted in the CDR sequences show mutations from the germline-encoded sequences. As will be apparent to those skilled in the art, determination of antibody numbering, including CDR sequences, framework sequences, etc., can be performed in a variety of well-recognized ways in the art, including by the NT and IMGT numbering systems presented in Figure 10A and Figure 10B and employed elsewhere in this document. Detailed Description

[0358] The present disclosure provides, at least in part, antibody molecules that bind to human IL-27 with high affinity and specificity. In one embodiment, human antibodies that bind to IL-27 are disclosed herein. As used herein, the terms “IL-27” and “IL27” may be used interchangeably to refer to the heterodimeric cytokine IL-27, which is composed of two different subunits encoded by two different genes: Epstein-Barr virus-induced gene 3 (EBI3) and IL-27p28. IL-27 has both pro-inflammatory and anti-inflammatory properties and has different effects on hematopoietic and non-hematopoietic cells.

[0359] Accordingly, in one aspect, the present disclosure provides a monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or the antigen-binding portion thereof exhibits at least one or more of the following characteristics:

[0360] (i) binds to human IL-27 with an equilibrium dissociation constant (K D ) of 15 nM or less;

[0361] (ii) blocks the binding of IL-27 to the IL-27 receptor;

[0362] (iii) inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells;

[0363] (iv) inhibits or reduces the IL-27-mediated inhibition of CD161 expression in cells;

[0364] (v) inhibits or reduces the IL-27-mediated expression of PD-L1 and / or TIM-3 in cells;

[0365] (vi) induces or increases the secretion of one or more cytokines mediated by PD-1 from cells; and

[0366] (vii) a combination of (i) to (vi).

[0367] In other aspects, the present disclosure provides a monoclonal antibody or an antigen-binding portion thereof that specifically binds to human IL-27 and inhibits or reduces IL-27 biological activity or IL-27 signaling.

[0368] Other aspects of the invention include nucleic acid molecules, expression vectors, host cells, and methods for preparing antibody molecules encoding the antibody molecules. Immunoconjugates, multispecific or bispecific molecules, and pharmaceutical compositions comprising the antibody molecules are also provided. The anti-IL-27 antibody molecules disclosed herein can be used for the treatment, prevention, and / or diagnosis of cancerous or malignant disorders, such as solid tumors and liquid tumors (e.g., leukemia, e.g., lymphoma, e.g., AML), lung cancer (e.g., non-small cell lung cancer), pancreatic cancer, breast cancer (e.g., triple-negative breast cancer), melanoma, testicular cancer, sarcoma, head and neck cancer (e.g., squamous head and neck cancer), liver cancer (e.g., hepatocellular carcinoma (HCC)), colorectal cancer, ovarian cancer, brain cancer (e.g., glioblastoma multiforme), or kidney cancer (e.g., renal cell carcinoma, e.g., renal clear cell carcinoma).

[0369] Anti-IL-27 antibodies and antigen-binding fragments thereof

[0370] The present disclosure provides antibodies and antigen-binding portions thereof that specifically bind to and antagonize IL-27, particularly human IL-27. Isolated monoclonal antibodies or antigen-binding portions thereof that specifically bind human IL-27 are provided herein, which comprise heavy and light chain CDRs and variable sequences as shown in Table 12.

[0371] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof exhibits at least one or more of the following properties: (i) binds to human IL-27 with an equilibrium dissociation constant (K D ) of 15 nM or less; (ii) blocks the binding of IL-27 to the IL-27 receptor; (iii) inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells; (iv) inhibits or reduces the inhibition of CD161 expression in cells; (v) inhibits or reduces the expression of PD-L1 and / or TIM-3 in cells; (vi) induces or increases the secretion of one or more cytokines mediated by PD-1 from cells; and (vii) a combination of (i) to (vi).

[0372] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof binds to human IL-27 with an equilibrium dissociation constant (K D ) of 15 nM or less.

[0373] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof binds to recombinant human IL-27 or murine IL-27.

[0374] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells. In some embodiments, the cells are immune cells. In some embodiments, the cells are cancer cells.

[0375] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof inhibits or reduces the inhibition of CD161 expression in cells (e.g., improves or alleviates the inhibition of CD161 expression in cells). In some embodiments, the cells are immune cells.

[0376] In some embodiments, the isolated monoclonal antibody or antigen-binding portion thereof inhibits or reduces the expression of PD-L1 and / or TIM-3 in cells. In some embodiments, PD-L1 expression is inhibited or reduced. In some embodiments, TIM-3 expression is inhibited or reduced. In some embodiments, both PD-L1 expression and TIM-3 expression are reduced. In some embodiments, the cells are immune cells.

[0377] In some embodiments, the isolated monoclonal antibody or antigen binding portion thereof induces or increases PD-1 mediated secretion of one or more cytokines from cells. In some embodiments, the one or more cytokines are TNFα. In some embodiments, the one or more cytokines are IL-6. In some embodiments, the one or more cytokines are TNFα and IL-6. In some embodiments, the cells are immune cells.

[0378] In some embodiments, the isolated monoclonal antibody or its antigen-binding portion is selected from the group consisting of: IgG1, IgG2, IgG3, IgG4, IgM, IgA1 IgA2, IgD and IgE antibodies. In some embodiments, the antibody is an IgG1 antibody or an IgG4 antibody. In some embodiments, the antibody comprises a wild-type IgG1 heavy chain constant region. In some embodiments, the antibody comprises a wild-type IgG4 heavy chain constant region. In some embodiments, the antibody comprises an Fc domain containing at least one mutation. In some embodiments, the antibody comprises a mutant IgG1 heavy chain constant region. In some embodiments, the antibody comprises a mutant IgG4 heavy chain constant region. In some embodiments, the mutant IgG4 heavy chain constant region comprises any one or a combination of substitutions S228P, L235E, L235A according to EU numbering.

[0379] In some embodiments, the disclosure provides an isolated monoclonal antibody, or antigen binding portion thereof, that binds to substantially the same epitope on IL-27 as the antibody, or antigen binding portion thereof, according to any of the preceding embodiments.

[0380] In some embodiments, the present disclosure provides an isolated monoclonal antibody, or antigen binding portion thereof, that binds to at least one of the amino acid residues comprising IL-27 bound by the antibody, or antigen binding portion thereof, according to any of the preceding embodiments.

[0381] In some embodiments, the present disclosure provides an isolated monoclonal antibody, or antigen binding portion thereof, wherein mutation of an epitope on IL-27 bound by the antibody, or antigen binding portion thereof, inhibits, reduces, or blocks binding to the antibody, or antigen binding portion thereof, and to the antibody, or antigen binding portion thereof, according to any of the preceding embodiments.

[0382] In some embodiments, the disclosure provides an isolated monoclonal antibody, or antigen binding portion thereof, that binds to an epitope on IL-27, wherein the epitope is the same as or similar to the epitope bound by the antibody molecules described in Table 12.

[0383] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs selected from the group consisting of:

[0384] (i) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 51, 52, and 53, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 59, 60, and 61, respectively;

[0385] (ii) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 73, 74, and 75, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 81, 82, and 83, respectively;

[0386] (iii) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 95, 96, and 97, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 103, 104, and 105, respectively;

[0387] (iv) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 117, 118, and 119, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 125, 126, and 127, respectively;

[0388] (v) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 139, 140, and 141, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 147, 148, and 149, respectively;

[0389] (vi) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 161, 162, and 163, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 169, 170, and 171, respectively;

[0390] (vii) heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 185, 186, and 187, respectively, and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs: 193, 194, and 195, respectively;

[0391] (viii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:207, 208 and 209 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:215, 216 and 217 respectively;

[0392] (ix) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:229, 230 and 231 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:237, 238 and 239 respectively;

[0393] (x) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:251, 252 and 253 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:259, 260 and 261 respectively;

[0394] (xi) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:273, 274 and 275 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:281, 282 and 283 respectively;

[0395] (xii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:295, 296 and 297 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:303, 304 and 305 respectively;

[0396] (xiii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:317, 318 and 319 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:325, 326 and 327 respectively;

[0397] (xiv) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:339, 340 and 341 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:347, 348 and 349 respectively;

[0398] (xv) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:361, 362 and 363 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO:369, 370 and 371 respectively; and

[0399] (xvi) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 383, 384, and 385, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 391, 392, and 393, respectively.

[0400] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, wherein the heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 161, 162, and 163, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 169, 170, and 171, respectively.

[0401] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs selected from the group consisting of:

[0402] (i) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 54, 55, and 56, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 62, 63, and 64, respectively;

[0403] (ii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 76, 77, and 78, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 84, 85, and 86, respectively;

[0404] (iii) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 98, 99, and 100, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 106, 107, and 108, respectively;

[0405] (iv) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 120, 121, and 122, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 128, 129, and 130, respectively;

[0406] (v) The heavy chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 142, 143, and 144, respectively, and the light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NO: 150, 151, and 152, respectively;

[0407] (vi) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 164, 165 and 166 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 172, 173 and 174 respectively;

[0408] (vii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 188, 189 and 190 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 196, 197 and 198 respectively;

[0409] (viii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 210, 211 and 212 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 218, 219 and 220 respectively;

[0410] (ix) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 232, 233 and 234 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 240, 241 and 242 respectively;

[0411] (x) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 254, 255 and 256 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 262, 263 and 264 respectively;

[0412] (xi) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 276, 277 and 278 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 284, 285 and 286 respectively;

[0413] (xii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 298, 299 and 300 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 306, 307 and 308 respectively;

[0414] (xiii) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 320, 321 and 322 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 328, 329 and 330 respectively;

[0415] (xiv) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 342, 343 and 344 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 350, 351 and 352 respectively;

[0416] (xv) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 364, 365 and 366 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 372, 373 and 374 respectively; and

[0417] (xvi) The heavy chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 386, 387 and 388 respectively, and the light chain CDR1, CDR2 and CDR3 sequences shown in SEQ ID NO: 394, 395 and 396 respectively.

[0418] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, wherein the heavy chain CDR1, CDR2 and CDR3 sequences are shown in SEQ ID NO: 164, 165 and 166 respectively, and the light chain CDR1, CDR2 and CDR3 sequences are shown in SEQ ID NO: 172, 173 and 174 respectively.

[0419] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs, wherein the amino acid sequences of the heavy chain CDR1, CDR2 and CDR3 are SYSMS (SEQ ID NO: 23), YISYDGGSAYYPDTVKG (SEQ ID NO: 24) and HGDYDDDDAMDY (SEQ ID NO: 25) respectively, and wherein the amino acid sequences of the light chain CDR1, CDR2 and CDR3 are RASENIYSYLA (SEQ ID NO: 26), NAETLTE (SEQ ID NO: 27) and QHHYGTPLT (SEQ ID NO: 28) respectively.

[0420] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 57, 79, 101, 123, 145, 167, 191, 213, 235, 257, 279, 301, 323, 345, 367, and 389; and wherein the light chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 65, 87, 109, 131, 153, 175, 199, 221, 243, 265, 287, 309, 331, 353, 375, and 397.

[0421] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, and the heavy and light chain variable regions comprise amino acid sequences selected from the groups consisting of:

[0422] (i) SEQ ID NOs: 57 and 65;

[0423] (ii) SEQ ID NOs: 79 and 87;

[0424] (iii) SEQ ID NOs: 101 and 109;

[0425] (iv) SEQ ID NOs: 123 and 131;

[0426] (v) SEQ ID NOs: 145 and 153;

[0427] (vi) SEQ ID NOs: 167 and 175;

[0428] (vii) SEQ ID NOs: 191 and 199;

[0429] (viii) SEQ ID NOs: 213 and 221;

[0430] (ix) SEQ ID NOs: 235 and 243;

[0431] (x) SEQ ID NOs: 257 and 265;

[0432] (xi) SEQ ID NOs: 279 and 287;

[0433] (xii) SEQ ID NOs: 301 and 309;

[0434] (xiii) SEQ ID NO: 323 and 331;

[0435] (xiv) SEQ ID NO: 345 and 353;

[0436] (xv) SEQ ID NO: 367 and 375; and

[0437] (xvi) SEQ ID NO: 389 and 397.

[0438] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions, wherein the heavy chain variable region comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 57, 79, 101, 123, 145, 167, 191, 213, 235, 257, 279, 301, 323, 345, 367, and 389; and wherein the light chain variable region comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NO: 65, 87, 109, 131, 153, 175, 199, 221, 243, 265, 287, 309, 331, 353, 375, and 397.

[0439] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain variable regions that respectively comprise an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of:

[0440] (i) SEQ ID NO: 57 and 65;

[0441] (ii) SEQ ID NO: 79 and 87;

[0442] (iii) SEQ ID NO: 101 and 109;

[0443] (iv) SEQ ID NO: 123 and 131;

[0444] (v) SEQ ID NO: 145 and 153;

[0445] (vi) SEQ ID NO: 167 and 175;

[0446] (vii) SEQ ID NO: 191 and 199;

[0447] (viii) SEQ ID NO: 213 and 221;

[0448] (ix) SEQ ID NO: 235 and 243;

[0449] (x) SEQ ID NO: 257 and 265;

[0450] (xi) SEQ ID NO: 279 and 287;

[0451] (xii) SEQ ID NO: 301 and 309;

[0452] (xiii) SEQ ID NO: 323 and 331;

[0453] (xiv) SEQ ID NO: 345 and 353;

[0454] (xv) SEQ ID NO: 367 and 375; and

[0455] (xvi) SEQ ID NO: 389 and 397.

[0456] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or the antigen-binding portion thereof comprises heavy and light chain variable regions, and the heavy and light chain variable regions comprise the amino acid sequences set forth in SEQ ID NO: 167 and 175, respectively.

[0457] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or the antigen-binding portion thereof comprises heavy and light chains, and the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0458] In some embodiments, the present disclosure provides an isolated monoclonal antibody or an antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or the antigen-binding portion thereof comprises heavy and light chains, wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and wherein the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0459] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 67, 89, 111, 133, 155, 177, 201, 223, 245, 267, 289, 311, 333, 355, 377, and 399; and wherein the light chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0460] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 71, 93, 115, 137, 159, 181, 205, 227, 249, 271, 293, 315, 337, 359, 381, and 403; and wherein the light chain comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0461] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, wherein the heavy chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 71, 93, 115, 137, 159, 181, 205, 227, 249, 271, 293, 315, 337, 359, 381, and 403; and wherein the light chain comprises an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of SEQ ID NOs: 69, 91, 113, 135, 157, 179, 203, 225, 247, 269, 291, 313, 335, 357, 379, and 401.

[0462] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise amino acid sequences selected from the group consisting of:

[0463] (i) SEQ ID NO:67 and 69;

[0464] (ii) SEQ ID NO:89 and 91;

[0465] (iii) SEQ ID NO:111 and 113;

[0466] (iv) SEQ ID NO:133 and 135;

[0467] (v) SEQ ID NO:155 and 157;

[0468] (vi) SEQ ID NO:177 and 179;

[0469] (vii) SEQ ID NO:201 and 203;

[0470] (viii) SEQ ID NO:223 and 225;

[0471] (ix) SEQ ID NO:245 and 247;

[0472] (x) SEQ ID NO:267 and 269;

[0473] (xi) SEQ ID NO:289 and 291;

[0474] (xii) SEQ ID NO:311 and 313;

[0475] (xiii) SEQ ID NO:333 and 335;

[0476] (xiv) SEQ ID NO:355 and 357;

[0477] (xv) SEQ ID NO:377 and 379; and

[0478] (xvi) SEQ ID NO:399 and 401.

[0479] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain each comprise an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of:

[0480] (i) SEQ ID NO: 67 and 69;

[0481] (ii) SEQ ID NO: 89 and 91;

[0482] (iii) SEQ ID NO: 111 and 113;

[0483] (iv) SEQ ID NO: 133 and 135;

[0484] (v) SEQ ID NO: 155 and 157;

[0485] (vi) SEQ ID NO: 177 and 179;

[0486] (vii) SEQ ID NO: 201 and 203;

[0487] (viii) SEQ ID NO: 223 and 225;

[0488] (ix) SEQ ID NO: 245 and 247;

[0489] (x) SEQ ID NO: 267 and 269;

[0490] (xi) SEQ ID NO: 289 and 291;

[0491] (xii) SEQ ID NO: 311 and 313;

[0492] (xiii) SEQ ID NO: 333 and 335;

[0493] (xiv) SEQ ID NO: 355 and 357;

[0494] (xv) SEQ ID NO: 377 and 379; and

[0495] (xvi) SEQ ID NO: 399 and 401.

[0496] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain respectively comprise amino acid sequences selected from the group consisting of:

[0497] (i) SEQ ID NO:71 and 69;

[0498] (ii) SEQ ID NO:93 and 91;

[0499] (iii) SEQ ID NO:115 and 113;

[0500] (iv) SEQ ID NO:137 and 135;

[0501] (v) SEQ ID NO:159 and 157;

[0502] (vi) SEQ ID NO:181 and 179;

[0503] (vii) SEQ ID NO:205 and 203;

[0504] (viii) SEQ ID NO:227 and 225;

[0505] (ix) SEQ ID NO:249 and 247;

[0506] (x) SEQ ID NO:271 and 269;

[0507] (xi) SEQ ID NO:293 and 291;

[0508] (xii) SEQ ID NO:315 and 313;

[0509] (xiii) SEQ ID NO:337 and 335;

[0510] (xiv) SEQ ID NO:359 and 357;

[0511] (xv) SEQ ID NO:381 and 379; and

[0512] (xvi) SEQ ID NO:403 and 401.

[0513] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain each comprise an amino acid sequence having at least 90% identity to an amino acid sequence selected from the group consisting of:

[0514] (i) SEQ ID NO:71 and 69;

[0515] (ii) SEQ ID NO:93 and 91;

[0516] (iii) SEQ ID NO:115 and 113;

[0517] (iv) SEQ ID NO:137 and 135;

[0518] (v) SEQ ID NO:159 and 157;

[0519] (vi) SEQ ID NO:181 and 179;

[0520] (vii) SEQ ID NO:205 and 203;

[0521] (viii) SEQ ID NO:227 and 225;

[0522] (ix) SEQ ID NO:249 and 247;

[0523] (x) SEQ ID NO:271 and 269;

[0524] (xi) SEQ ID NO:293 and 291;

[0525] (xii) SEQ ID NO:315 and 313;

[0526] (xiii) SEQ ID NO:337 and 335;

[0527] (xiv) SEQ ID NO:359 and 357;

[0528] (xv) SEQ ID NO:381 and 379; and

[0529] (xvi) SEQ ID NO:403 and 401.

[0530] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise the amino acid sequences set forth in SEQ ID NO: 177 and 179, respectively.

[0531] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise amino acid sequences having at least 90% identity to the amino acid sequences set forth in SEQ ID NO: 177 and 179, respectively.

[0532] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise the amino acid sequences set forth in SEQ ID NO: 181 and 179, respectively.

[0533] In some embodiments, the present disclosure provides an isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof comprises a heavy chain and a light chain, and the heavy chain and the light chain comprise amino acid sequences having at least 90% identity to the amino acid sequences set forth in SEQ ID NO: 181 and 179, respectively.

[0534] Methods for generating anti-IL-27 antibodies and antigen-binding fragments thereof

[0535] The present disclosure also describes methods for generating any of the anti-IL-27 antibodies or antigen-binding fragments thereof described herein. In some embodiments, methods for preparing the antibodies described herein may include immunizing a subject (e.g., a non-human mammal) with a suitable immunogen. Suitable immunogens for generating any of the antibodies described herein are set forth herein. For example, to generate an antibody that binds IL-27, one of skill in the art may immunize a suitable subject (e.g., a non-human mammal such as a rat, mouse, gerbil, hamster, dog, cat, pig, goat, horse, or non-human primate) with IL-27. In some embodiments, the full-length human IL-27 EBI3 monomer polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 97 is used as the immunogen. In some embodiments, the full-length human IL-27 p28 monomer polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 98 is used as the immunogen.

[0536] A suitable subject (e.g., a non-human mammal) can be immunized with a suitable antigen, followed by multiple booster immunizations sufficient to elicit antibody production in the mammal. The immunogen can be administered to the subject (e.g., a non-human mammal) together with an adjuvant. Adjuvants useful for generating antibodies in a subject include, but are not limited to, protein adjuvants; bacterial adjuvants such as whole bacteria (BCG, Corynebacterium parvum, or Salmonella minnesota) and bacterial components including cell wall skeletons, trehalose dimycolate, monophosphoryl lipid A, methanol extractable residues (MER) of the tubercle bacillus, complete or incomplete Freund's adjuvant; viral adjuvants; chemical adjuvants such as aluminum hydroxide and iodoacetate and cholesterol hemisuccinate. Other adjuvants useful in methods for inducing an immune response include, for example, cholera toxin and parapoxvirus proteins. See also Bieg et al. (1999) Autoimmunity 31(1) : 15-24. See also, for example, Lodmell et al. (2000) Vaccine 18 : 1059-1066; Johnson et al. (1999) J Med Chem 42 : 4640-4649; Baldridge et al. (1999) Methods 19 : 103-107 and Gupta et al. (1995) Vaccine 13(14) : 1263-1276.

[0537] In some embodiments, the method includes preparing a hybridoma cell line that secretes a monoclonal antibody that binds to an immunogen. For example, a suitable mammal such as a laboratory mouse is immunized with the above-described IL-27 polypeptide. Antibody-producing cells (e.g., B cells of the spleen) of the immunized mammal can be isolated 2 to 4 days after at least one booster immunization with the immunogen and then grown briefly in culture before being fused with cells of a suitable myeloma cell line. The cells can be fused in the presence of a fusion promoter such as vaccinia virus or polyethylene glycol. The hybrid cells obtained in the fusion are cloned, and cell clones that secrete the desired antibody are selected. For example, spleen cells of Balb / c mice immunized with a suitable immunogen can be fused with cells of the myeloma cell line PAI or the myeloma cell line Sp2 / 0-Ag 14. After fusion, the cells are amplified in a suitable medium, and the medium is supplemented with a selection medium, such as HAT medium, at regular intervals to prevent the growth of normal myeloma cells beyond the desired hybridoma cells. Then, the secretion of the desired antibody (e.g., an antibody that binds to human IL-27) by the obtained hybrid cells is screened, and in some embodiments, a person skilled in the art can identify anti-IL-27 antibodies from the non-immune-biased libraries described in U.S. Patent No. 6,300,064 (to Knappik et al.; Morphosys AG) and Schoonbroodt et al. (2005) Nucleic Acids Res 33(9) : anti-IL-27 antibodies can be identified from the non-immune-biased libraries described in e81.

[0538] In some embodiments, the methods described herein can involve, for example, phage display technology, bacterial display, yeast surface display, eukaryotic virus display, mammalian cell display, and cell-free (e.g., ribosome display) antibody screening techniques or be used in combination with such techniques (see, for example, Etz et al. (2001) J Bacteriol 183 : 6924-6935; Cornelis (2000) Curr Opin Biotechnol 11 : 450-454; Klemm et al. (2000) Microbiology 146 : 3025-3032; Kieke et al. (1997) Protein Eng 10 : 1303-1310; Yeung et al. (2002) BiotechnolProg 18 : 212-220; Boder et al. (2000) Methods Enzymology 328 : 430-444; Grabherr et al. (2001) Comb Chem High Throughput Screen 4: 185 - 192; Michael et al. (1995) Gene Ther 2 : 660 - 668; Pereboev et al. (2001) J Virol 75 : 7107 - 7113; Schaffitzel et al. (1999) J Immunol Methods 231 : 119 - 135 and Hanes et al. (2000) Nat Biotechnol 18 : 1287 - 1292).

[0539] Methods for identifying antibodies using various phage display methods are known in the art. In phage display methods, functional antibody domains are displayed on the surface of phage particles carrying the polynucleotide sequences encoding them. Such phages can be used to display the antigen - binding domains of antibodies, such as Fab, Fv, or disulfide - stabilized Fv antibody fragments, which are expressed from libraries or combinatorial antibody libraries (e.g., human or murine). The phages used in these methods are typically filamentous phages, such as fd and M13. The antigen - binding domains are expressed as proteins recombinantly fused to any one of the phage coat proteins pIII, pVIII, or pIX. See, e.g., Shi et al. (2010) JMB 397 : 385 - 396. Examples of phage display methods useful for preparing immunoglobulins or fragments thereof described herein include those disclosed in the following references: Brinkman et al. (1995) J Immunol Methods 182 : 41 - 50; Ames et al. (1995) J Immunol Methods 184 : 177 - 186; Kettleborough et al. (1994) Eur J Immunol 24 : 952 - 958; Persic et al. (1997) Gene 187 : 9 - 18; Burton et al. (1994) Advances in Immunology 57: 191 - 280; and PCT Publication Nos. WO 90 / 02809, WO 91 / 10737, WO 92 / 01047, WO 92 / 18619, WO 93 / 11236, WO 95 / 15982, and WO 95 / 20401. Suitable methods are also described, for example, in U.S. Patent Nos. 5,698,426, 5,223,409, 5,403,484, 5,580,717, 5,427,908, 5,750,753, 5,821,047, 5,571,698, 5,427,908, 5,516,637, 5,780,225, 5,658,727, 5,733,743, and 5,969,108.

[0540] In some embodiments, a phage display antibody library can be generated using mRNA collected from B cells of an immunized mammal. For example, a spleen cell sample containing B cells can be isolated from a mouse immunized with the above IL-27 polypeptide. mRNA can be isolated from the cells and converted to cDNA using standard molecular biology techniques. See, for example, Sambrook et al. (1989) "Molecular Cloning: A Laboratory Manual, 2nd ed.," Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.; Harlow and Lane (1988), supra; Benny K.C. Lo (2004), supra; and Borrebaek (1995), supra. The cDNA encoding the variable regions of the immunoglobulin heavy and light chain polypeptides is used to construct a phage display library. Methods for generating such libraries are described, for example, in Merz et al. (1995) J Neurosci Methods 62(1-2) : 213 - 9; Di Niro et al. (2005) Biochem J 388 (Pt 3) : 889–894 and Engberg et al. (1995) Methods Mol Biol 51 : 355 - 376.

[0541] In some embodiments, a combination of selection and screening can be used to identify a target antibody from, for example, a population of antibodies produced by hybridomas or a phage display antibody library. Suitable methods are known in the art and are described, for example, in Hoogenboom (1997) Trends in Biotechnology 15: 62 - 70; Brinkman et al. (1995), ibid.; Ames et al. (1995), ibid.; Kettleborough et al. (1994), ibid.; Persic, et al. (1997), ibid. and Burton et al. (1994), ibid. For example, using standard molecular biology techniques, a variety of phagemid vectors are generated (each phagemid vector encoding a fusion protein of a phage coat protein (e.g., pIII, pVIII, or pIX of M13 phage) and different antigen-binding regions), and then introduced into a bacterial population (e.g., Escherichia coli). In some embodiments, the expression of phage in bacteria may require the use of helper phage. In some embodiments, helper phage is not required (see, e.g., Chasteen et al., (2006) Nucleic Acids Res 34(21): e145). The phage produced from the bacteria is recovered and then contacted with, for example, a (fixed) target antigen bound to a solid support. The phage can also be contacted with an antigen in solution, and subsequently the complex is bound to a solid support.

[0542] Any immunology- or biochemistry-based method known in the art can be used to characterize the specificity and binding affinity of a subset of antibodies screened using the above methods for a specific antigen (e.g., human IL-27). For example, the specific binding of an antibody to IL-27 can be determined, for example, using an immunology- or biochemistry-based method such as, but not limited to, the ELISA assay, SPR assay, immunoprecipitation assay, affinity chromatography, and equilibrium dialysis as described above. Immunoassays that can be used to analyze the immunospecific binding and cross-reactivity of antibodies include, but are not limited to, competitive and non-competitive assay systems that use techniques such as Western blotting, RIA, ELISA (enzyme-linked immunosorbent assay), "sandwich" immunoassays, immunoprecipitation assays, immunodiffusion assays, agglutination assays, complement fixation assays, immunoradiometric assays, fluorescence immunoassays, and protein A immunoassays. Such assays are routine and well-known in the art.

[0543] It should be understood that the above methods can also be used to determine, for example, whether an anti-IL-27 antibody does not bind to full-length human IL-27 and / or IL-27 protein.

[0544] In embodiments where the selected CDR amino acid sequences are short sequences (e.g., less than 10 - 15 amino acids in length), it can be as described, for example, in Shiraishi et al. (2007) Nucleic Acids Symposium Series 51(1):As described in U.S. Patent No. 6,995,259 and 129-130, nucleic acids encoding CDRs are chemically synthesized. For a given nucleic acid sequence encoding a recipient antibody, the region of the nucleic acid sequence encoding the CDR can be replaced with chemically synthesized nucleic acids using standard molecular biology techniques. The 5' and 3' ends of the chemically synthesized nucleic acids can be synthesized to contain sticky end restriction enzyme sites for cloning the nucleic acids into the nucleic acid encoding the variable region of the donor antibody.

[0545] In some embodiments, the anti-IL-27 antibodies described herein comprise an altered heavy chain constant region that has reduced (or no) effector function relative to its corresponding unaltered constant region. The effector function associated with the constant region of the anti-IL-27 antibody can be adjusted by altering the properties of the constant region or the Fc region. Altered effector functions include, for example, modulation of one or more of the following activities: antibody-dependent cell cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), apoptosis, binding to one or more Fc receptors, and pro-inflammatory responses. Modulation refers to an increase, decrease, or elimination of the effector function activity exhibited by a test antibody comprising the altered constant region compared to the activity of the unaltered form of the constant region. In certain embodiments, modulation includes cases where the activity is eliminated or completely absent.

[0546] In one embodiment, the anti-IL-27 antibodies described herein comprise an IgG4 heavy chain constant region. In one embodiment, the IgG4 heavy chain constant region is a wild-type IgG4 heavy chain constant region. In another embodiment, for example, according to EU numbering (Kabat, E.A. et al., supra), the IgG4 constant region contains mutations, such as one or both of S228P and L235E or L235A. Representative sequences of wild-type and mutant IgG4 constant regions for the antibodies of the present disclosure are shown in Table 12. In one embodiment, the anti-IL-27 antibodies described herein comprise an IgG1 constant region. In one embodiment, the IgG1 heavy chain constant region is a wild-type IgG1 heavy chain constant region. In another embodiment, the IgG1 heavy chain constant region contains mutations. Representative sequences of wild-type and mutant IgG4 constant regions for the antibodies of the present disclosure are shown in Table 12.

[0547] An altered constant region having an altered FcR binding affinity and / or ADCC activity and / or altered CDC activity is a polypeptide that has an increased or decreased FcR binding activity and / or ADCC activity and / or CDC activity as compared to the unaltered form of the constant region. An altered constant region that shows enhanced binding to FcR binds at least one FcR with a greater affinity than the unaltered polypeptide. An altered constant region that shows decreased binding to FcR binds at least one FcR with a lower affinity than the unaltered form of the constant region. Such variants that show decreased binding to FcR may have little or no appreciable binding to FcR, e.g., 0 to 50% (e.g., less than 50%, 49%, 48%, 47%, 46%, 45%, 44%, 43%, 42%, 41%, 40%, 39%, 38%, 37%, 36%, 35%, 34%, 33%, 32%, 31%, 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1%) binding to FcR as compared to the binding level of the native sequence immunoglobulin constant region or Fc region to FcR. Similarly, an altered constant region that shows adjusted ADCC and / or CDC activity may show enhanced or decreased ADCC and / or CDC activity as compared to the unaltered constant region. For example, in some embodiments, an anti-IL-27 antibody comprising an altered constant region may exhibit from about 0 to 50% (e.g., less than 50%, 49%, 48%, 47%, 46%, 45%, 44%, 43%, 42%, 41%, 40%, 39%, 38%, 37%, 36%, 35%, 34%, 33%, 32%, 31%, 30%, 29%, 28%, 27%, 26%, 25%, 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1%) of the ADCC and / or CDC activity of the unaltered form of the constant region. An anti-IL-27 antibody described herein that exhibits decreased ADCC and / or CDC and that comprises an altered constant region may exhibit decreased ADCC and / or CDC activity or no ADCC and / or CDC activity.

[0548] In some embodiments, the anti-IL-27 antibodies described herein exhibit decreased effector function or no effector function. In some embodiments, the anti-IL-27 antibody comprises a hybrid constant region or a portion thereof, such as a G2 / G4 hybrid constant region (see, e.g., Burton et al. (1992) Adv Immun51 : 1-18; Canfield et al. (1991) J Exp Med 173 : 1483-1491 and Mueller et al. (1997) Mol Immunol 34(6) : 441-452). See above.

[0549] In some embodiments, the anti-IL-27 antibody may comprise an altered constant region that exhibits enhanced or reduced complement-dependent cytotoxicity (CDC). The CDC activity can be adjusted by introducing one or more amino acid substitutions, insertions, or deletions in the Fc region of the antibody. See, for example, U.S. Patent No. 6,194,551. Optionally or additionally, one or more cysteine residues can be introduced into the Fc region, thereby allowing the formation of interchain disulfide bonds in this region. The resulting homodimeric antibody may have increased or decreased internalization capacity and / or enhanced or reduced complement-mediated cell killing. See, for example, Caron et al. (1992) J Exp Med 176 : 1191-1195 and Shopes (1992) Immunol 148 : 2918-2922; PCT Publication Nos. WO 99 / 51642 and WO 94 / 29351; Duncan and Winter (1988) Nature 322 : 738-40; and U.S. Patent Nos. 5,648,260 and 5,624,821.

[0550] Expression and purification of recombinant antibodies

[0551] The antibodies or antigen-binding fragments thereof described herein can be produced using a variety of techniques known in the fields of molecular biology and protein chemistry. For example, nucleic acids encoding one or both of the heavy and light chain polypeptides of the antibody can be inserted into an expression vector comprising transcriptional and translational regulatory sequences, including, for example, promoter sequences, ribosome binding sites, transcriptional start and stop sequences, translational start and stop sequences, transcriptional terminator signals, polyadenylation signals, and enhancer or activator sequences. The regulatory sequences include a promoter and transcriptional start and stop sequences. Additionally, the expression vector can include more than one replication system such that it can be maintained in two different organisms, for example, for expression in mammalian or insect cells and for cloning and amplification in a prokaryotic host.

[0552] Several possible vector systems can be used to express heavy and light chain polypeptides cloned from nucleic acids in mammalian cells. One class of vectors relies on integration of the desired gene sequences into the host cell genome. Cells with stably integrated DNA can be selected by co-introducing a drug resistance gene such as Escherichia coli gpt (Mulligan and Berg (1981) Proc Natl Acad Sci USA 78 :2072) or Tn5 neo (Southern and Berg (1982) Mol Appl Genet 1 :327). The selectable marker gene can be ligated to the DNA sequence to be expressed, or the selectable marker gene can be introduced into the same cell by co-transfection (Wigler et al. (1979) Cell 16 :77). A second class of vectors utilizes DNA elements that confer the ability to replicate autonomously as episomal plasmids. These vectors can be derived from animal viruses such as bovine papillomavirus (Sarver et al. (1982) Proc Natl Acad Sci USA, 79 :7147), cytomegalovirus, polyomavirus (Deans et al. (1984) Proc Natl Acad Sci USA 81 :1292) or SV40 virus (Lusky and Botchan (1981) Nature 293 :79).

[0553] The expression vector can be introduced into cells in a manner suitable for subsequent expression of the nucleic acid. As discussed below, the method of introduction is mainly determined by the target cell type. Exemplary methods include CaPO4 precipitation, liposome fusion, cationic liposomes, electroporation, viral infection, dextran-mediated transfection, polyamine-mediated transfection, protoplast fusion and direct microinjection.

[0554] Suitable host cells for expressing antibodies or antigen-binding fragments thereof include yeast, bacteria, insects, plants and mammalian cells. Of particular interest are bacteria such as Escherichia coli, fungi such as Saccharomyces cerevisiae and Pichia pastoris, insect cells such as SF9, mammalian cell lines (e.g., human cell lines) and primary cell lines.

[0555] In some embodiments, the antibody or fragment thereof can be expressed in a transgenic animal (e.g., a transgenic mammal) and purified therefrom. For example, as in, e.g., Houdebine (2002) Curr Opin Biotechnol 13(6): 625 - 629; as described in van Kuik - Romeijn et al. (2000) Transgenic Res 9(2): 155 - 159 and Pollock et al. (1999) J Immunol Methods 231(1 - 2): 147 - 157, antibodies can be produced in transgenic non - human mammals (e.g., rodents) and the antibodies can be isolated from milk.

[0556] Antibodies and their fragments can be produced from cells by culturing host cells transformed with an expression vector containing nucleic acid encoding the antibody or fragment for a period of time under conditions sufficient to permit protein expression. Such conditions for protein expression will vary with the choice of expression vector and host cell and will be readily determined by those skilled in the art through routine experimentation. For example, antibodies expressed in E. coli can be refolded from inclusion bodies (e.g., see Hou et al. (1998) Cytokine 10 : 319 - 30). Bacterial expression systems and methods of using them are well known in the art (see Current Protocols in Molecular Biology, Wiley & Sons, and Molecular Cloning - A Laboratory Manual - 3rd Edition, Cold Spring Harbor Laboratory Press, New York (2001)). The choice of codons, suitable expression vectors, and suitable host cells will vary depending on many factors and can be readily optimized as needed. The antibodies (or fragments thereof) described herein can be expressed in mammalian cells or other expression systems (including but not limited to yeast, baculovirus) and in vitro expression systems (see, e.g., Kaszubska et al. (2000) Protein Expression and Purification 18 : 213 - 220).

[0557] After expression, the antibody and its fragments can be separated. The antibody or its fragments can be separated or purified by a variety of methods known to those skilled in the art, depending on the other components present in the sample. Standard purification methods include electrophoretic techniques, molecular techniques, immunological techniques, and chromatographic techniques, including ion exchange chromatography, hydrophobic chromatography, affinity chromatography, and reverse phase HPLC chromatography. For example, an antibody can be purified using a standard anti-antibody column (e.g., a protein-A or protein-G column). Ultrafiltration and diafiltration techniques in combination with protein concentration are also useful. See, e.g., Scopes (1994) "Protein Purification," 3rd ed., Springer-Verlag, New York City, New York. The degree of purification required will vary depending on the intended use. In some cases, purification of the expressed antibody or its fragments is not necessary.

[0558] Methods for determining the yield or purity of a purified antibody or its fragments are known in the art and include, for example, the Bradford assay, ultraviolet spectroscopy, the biuret protein assay, the Lowry protein assay, the amido black protein assay, high pressure liquid chromatography (HPLC), mass spectrometry (MS), and gel electrophoresis methods (e.g., using protein stains such as Coomassie blue or colloidal silver staining).

[0559] Modification of an antibody or its antigen-binding fragment

[0560] An antibody or its antigen-binding fragment can be modified after expression and purification. The modification can be a covalent or non-covalent modification. Such modifications can be introduced into the antibody or fragment by reacting, for example, a targeted amino acid residue of the polypeptide with an organic derivatizing agent capable of reacting with a selected side chain or terminal residue. Any of a number of criteria, including, for example, structural analysis or amino acid sequence analysis of the antibody or fragment, can be used to select a suitable site for modification.

[0561] In some embodiments, an antibody or an antigen-binding fragment thereof can be conjugated to a heterologous moiety. The heterologous moiety can be, for example, a heterologous polypeptide, a therapeutic agent (e.g., a toxin or a drug), or a detectable label such as, but not limited to, a radiolabel, an enzymatic label, a fluorescent label, a heavy metal label, a luminescent label, or an affinity tag such as biotin or streptavidin. Suitable heterologous polypeptides include, for example, antigen tags (FLAG (DYKDDDDK (SEQ ID NO: 405)), polyhistidine (6-His; HHHHHH (SEQ ID NO: 406)), hemagglutinin (HA; YPYDVPDYA (SEQ ID NO: 407)), glutathione-S-transferase (GST), or maltose-binding protein (MBP) for purifying the antibody or fragment. Heterologous polypeptides also include polypeptides (e.g., enzymes) used as diagnostic or detectable markers such as luciferase, fluorescent proteins (e.g., green fluorescent protein (GFP)), or chloramphenicol acetyltransferase (CAT). Suitable radiolabels include, for example 32 P, 33 P, 14 C, 125 I, 131 I, 35 S and 3 H. Suitable fluorescent labels include, but are not limited to, fluorescein, fluorescein isothiocyanate (FITC), green fluorescent protein (GFP), DyLight TM 488, phycoerythrin (PE), propidium iodide (PI), PerCP, PE-Alexa 700, Cy5, allophycocyanin, and Cy7. Luminescent labels include, for example, any of a variety of luminescent lanthanide element (e.g., europium or terbium) chelates. For example, suitable europium chelates include europium chelates of diethylenetriaminepentaacetic acid (DTPA) or 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). Enzymatic labels include, for example, alkaline phosphatase, CAT, luciferase, and horseradish peroxidase.

[0562] Two proteins (e.g., an antibody and a heterologous moiety) can be cross-linked using any of a number of known chemical cross-linking agents. Examples of such cross-linking agents are those that link two amino acid residues via a linkage containing a "hindered" disulfide bond. In these linkages, the disulfide bond within the cross-linking unit is protected (by hindering the groups on either side of the disulfide bond) from reduction by, for example, the action of reduced glutathione or disulfide reductase. A suitable reagent, 4-succinimidyloxycarbonyl-α-methyl-α(2-pyridyldithio)toluene (SMPT), forms such a linkage between two proteins using a terminal lysine on one protein and a terminal cysteine on the other protein. Heterobifunctional reagents that cross-link via different coupling moieties on each protein can also be used. Other useful cross-linking agents include, but are not limited to, those that link two amino groups (e.g., N-5-azido-2-nitrobenzoyloxysuccinimide), two sulfhydryl groups (e.g., 1,4-bismaleimidobutane), an amino group and a sulfhydryl group (e.g., m-maleimidobenzoyl-N-hydroxysuccinimide ester), an amino group and a carboxyl group (e.g., 4-[p-azidosalicylamido]butylamine), and an amino group and the guanidyl group present in the side chain of arginine (e.g., p-azidophenylglyoxal monohydrate).

[0563] In some embodiments, a radiolabel can be conjugated directly to the amino acid backbone of an antibody. Alternatively, the radiolabel can be part of a larger molecule (e.g., m- 125 I]iodophenyl-N-hydroxysuccinimide ( 125 I]mIPNHS) that contains 125 I and binds to a free amino group to form a meta-iodophenyl (mIP) derivative of the relevant protein (see, e.g., Rogers et al. (1997) J Nucl Med 38 :1221-1229) or a chelate (e.g., a chelate with DOTA or DTPA), which is then conjugated to the protein backbone. Methods for conjugating radiolabels or larger molecules / chelates containing them to the antibodies or antigen-binding fragments described herein are known in the art. Such methods involve incubating the protein with the radiolabel under conditions (e.g., pH, salt concentration, and / or temperature) that are favorable for binding of the radiolabel or chelator to the protein (see, e.g., U.S. Patent No. 6,001,329).

[0564] Methods for conjugating a fluorescent label (sometimes referred to as a "fluorophore") to a protein (e.g., an antibody) are known in the field of protein chemistry. For example, a succinimidyl (NHS) ester or tetrafluorophenyl (TFP) ester moiety attached to a fluorophore can be used to conjugate the fluorophore to a free amino group (e.g., of lysine) or a thiol group (e.g., of cysteine) of a protein. In some embodiments, the fluorophore can be conjugated to a heterobifunctional crosslinker moiety such as sulfo-SMCC. Suitable conjugation methods involve incubating the antibody protein or a fragment thereof with the fluorophore under conditions that promote binding of the fluorophore to the protein. See, e.g., Welch and Redvanly (2003) "Handbook of Radiopharmaceuticals: Radiochemistry and Applications," John Wiley and Sons (ISBN 0471495603).

[0565] In some embodiments, the antibody or fragment can be modified, for example, with moieties that improve the stability and / or retention of the antibody in circulation (e.g., in blood, serum, or other tissues). For example, the antibody or fragment can be PEGylated as described, e.g., in Lee et al. (1999) Bioconjug Chem 10(6) : 973-8; Kinstler et al. (2002) Advanced Drug Deliveries Reviews 54 : 477-485; and Roberts et al. (2002) Advanced Drug Delivery Reviews 54 : 459-476, or the antibody or fragment can be HESylated (Fresenius Kabi, Germany; see, e.g., et al. (2010) Int J Pharm 387(1-2) : 110-119). The stabilizing moiety can increase the stability or retention of the antibody (or fragment) by at least 1.5-fold (e.g., at least 2-fold, 5-fold, 10-fold, 15-fold, 20-fold, 25-fold, 30-fold, 40-fold, or 50-fold or more).

[0566] In some embodiments, the antibodies or antigen-binding fragments thereof described herein can be glycosylated. In some embodiments, the antibodies or antigen-binding fragments thereof described herein can be enzymatically or chemically treated, or the antibodies or antigen-binding fragments can be produced from cells such that the antibodies or fragments have reduced glycosylation or no glycosylation. Methods for producing antibodies with reduced glycosylation are known in the art and are described, for example, in U.S. Patent No. 6,933,368; Wright et al. (1991) EMBO J 10(10):2717-2723; and Co et al. (1993) Mol Immunol 30:1361.

[0567] Pharmaceutical Compositions and Formulations

[0568] In certain embodiments, the invention provides pharmaceutical compositions comprising an anti-IL-27 antibody and a pharmaceutically acceptable diluent, carrier, solubilizer, emulsifier, preservative, and / or adjuvant.

[0569] In certain embodiments, acceptable formulation materials are preferably non-toxic to the recipient at the dosages and concentrations employed. In certain embodiments, one or more formulation materials are used for subcutaneous and / or intravenous administration. In certain embodiments, the pharmaceutical composition may comprise formulation materials for altering, maintaining, or preserving, for example, the pH, osmolarity, viscosity, clarity, color, isotonicity, odor, sterility, stability, dissolution, or release rate, adsorption, or permeability of the composition. In certain embodiments, suitable formulation materials include, but are not limited to, amino acids (such as glycine, glutamine, asparagine, arginine, or lysine); antimicrobial agents; antioxidants (such as ascorbic acid, sodium sulfite, or sodium bisulfite); buffers (such as borate, bicarbonate, Tris-HCl, citrate, phosphate, or other organic acids); fillers (such as mannitol or glycine); chelating agents (such as ethylenediaminetetraacetic acid (EDTA)); complexing agents (such as caffeine, polyvinylpyrrolidone, β-cyclodextrin, or hydroxypropyl-β-cyclodextrin); fillers; monosaccharides; disaccharides; and other carbohydrates (such as glucose, mannose, or dextrin); proteins (such as serum albumin, gelatin, or immunoglobulins); coloring agents, flavoring agents, and diluents; emulsifying agents; hydrophilic polymers (such as polyvinylpyrrolidone); low molecular weight polypeptides; salt-forming counterions (such as sodium); preservatives (such as benzalkonium chloride, benzoic acid, salicylic acid, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid, or hydrogen peroxide); solvents (such as glycerol, propylene glycol, or polyethylene glycol); sugar alcohols (such as mannitol or sorbitol); suspending agents; surfactants or wetting agents (such as pluronics, PEG, sorbitan esters, polysorbates such as polysorbate 20, polysorbate 80, triton, tromethamine, lecithin, cholesterol, tyloxapol); stability enhancers (such as sucrose or sorbitol); tonicity enhancers (such as alkali metal halides, preferably sodium chloride or potassium chloride, mannitol, sorbitol); delivery vehicles; diluents; excipients; and / or pharmaceutical adjuvants. (Remington's Pharmaceutical Sciences, 18th Edition, A.R. Gennaro, Editor, Mack Publishing Company (1995). In certain embodiments, the formulation comprises PBS; 20 mM NaOAc (pH 5.2), 50 mM NaCl; and / or 10 mM NaOAc (pH 5.2), 9% sucrose. In certain embodiments, the optimal pharmaceutical composition will be determined by one of ordinary skill in the art based on, for example, the intended route of administration, delivery format, and desired dosage. See, e.g., Remington's Pharmaceutical Sciences, supra.)In certain embodiments, such compositions can affect the physical state, stability, in vivo release rate, and / or in vivo clearance rate of the anti-IL-27 antibody.

[0570] In certain embodiments, the primary vehicle or carrier in the pharmaceutical composition can be essentially aqueous or non-aqueous. For example, in certain embodiments, suitable vehicles or carriers can be water for injection, saline solution, or artificial cerebrospinal fluid, which may be supplemented with other materials commonly used in compositions for parenteral administration. In certain embodiments, the saline comprises isotonic phosphate buffered saline. In certain embodiments, neutral buffered saline or saline mixed with serum albumin are further exemplary vehicles. In certain embodiments, the pharmaceutical composition comprises a Tris buffer at a pH of about 7.0 - 8.5, or an acetate buffer at a pH of about 4.0 - 5.5, which may also comprise sorbitol or a suitable alternative thereof. In certain embodiments, a composition comprising an anti-IL-27 antibody can be prepared for storage by mixing the selected composition of desired purity with an optional formulation (Remington's Pharmaceutical Sciences, supra) in the form of a lyophilized cake or an aqueous solution. Additionally, in certain embodiments, a composition comprising an anti-IL-27 antibody can be formulated into a lyophilizate using a suitable excipient such as sucrose.

[0571] In certain embodiments, the pharmaceutical composition can be selected for parenteral delivery. In certain embodiments, the composition can be selected for delivery by inhalation or through the digestive tract (such as orally). The preparation of such pharmaceutically acceptable compositions is within the capabilities of those skilled in the art.

[0572] In certain embodiments, the formulation components are present at concentrations acceptable to the site of administration. In certain embodiments, buffers are used to maintain the composition at physiological pH or slightly lower pH, typically in the pH range of about 5 to about 8.

[0573] In certain embodiments, when parenteral administration is contemplated, the therapeutic composition may be in the form of a pyrogen-free, parenterally acceptable aqueous solution in a pharmaceutically acceptable vehicle, the aqueous solution comprising an anti-IL-27 antibody. In certain embodiments, the vehicle for parenteral injection is sterile distilled water, wherein the anti-IL-27 antibody is formulated as a sterile isotonic solution and appropriately preserved. In certain embodiments, the preparation may involve formulating the desired molecule with an agent such as an injectable microsphere, a bioerodible particle, a polymeric compound such as polylactic acid or polyglycolic acid, a bead, or a liposome, the agent providing controlled or sustained release of the product, which can then be delivered by depot injection. In certain embodiments, hyaluronic acid may also be used, which may have the effect of promoting the duration in circulation. In certain embodiments, an implantable drug delivery device may be used to introduce the desired molecule.

[0574] In certain embodiments, the pharmaceutical composition may be formulated for inhalation. In certain embodiments, the anti-IL-27 antibody may be formulated as a dry powder for inhalation. In certain embodiments, an inhalation solution comprising the anti-IL-27 antibody may be formulated with a propellant for aerosol delivery. In certain embodiments, the solution may be nebulized. Pulmonary administration is further described in PCT Application No. PCT / US94 / 001875, which describes the pulmonary delivery of chemically modified proteins.

[0575] In certain embodiments, oral administration of the formulation is contemplated. In certain embodiments, the anti-IL-27 antibody administered in this manner may or may not be formulated with the carriers commonly used for formulating solid dosage forms such as tablets and capsules. In certain embodiments, the capsule may be designed to release the active portion of the formulation at a point in the gastrointestinal tract where bioavailability is maximized and pre-systemic degradation is minimized. In certain embodiments, at least one additional agent may be included to facilitate the absorption of the anti-IL-27 antibody. In certain embodiments, diluents, flavoring agents, low melting waxes, vegetable oils, lubricants, suspending agents, tablet disintegrants, and binders may also be used.

[0576] In certain embodiments, the pharmaceutical composition may comprise a mixture of an effective amount of an anti-IL-27 antibody with a non-toxic excipient suitable for tablet production. In certain embodiments, a solution may be prepared in unit dose form by dissolving the tablet in sterile water or another suitable vehicle. In certain embodiments, suitable excipients include, but are not limited to, inert diluents such as calcium carbonate, sodium carbonate or bicarbonate, lactose, or calcium phosphate; or binders such as starch, gelatin, or gum arabic; or lubricants such as magnesium stearate, stearic acid, or talc.

[0577] Other pharmaceutical compositions will be apparent to those skilled in the art, including those that comprise an anti-IL-27 antibody in a sustained or controlled release formulation. In certain embodiments, techniques for formulating a variety of other sustained or controlled delivery components, such as liposomal carriers, bioerodible microparticles or porous beads, and depot injections, are also known to those skilled in the art. See, for example, PCT Application No. PCT / US93 / 00829, which describes the controlled release of porous polymer microparticles for delivering pharmaceutical compositions. In certain embodiments, a sustained release formulation may comprise a semipermeable polymer matrix in the form of a shaped article (e.g., a film or a microcapsule). Sustained release matrices may include polyesters, hydrogels, poly(lactide) (U.S. Patent No. 3,773,919 and EP 058,481), copolymers of L-glutamic acid and γ-ethyl-L-glutamic acid (Sidman et al., Biopolymers, 22:547-556 (1983)), poly(2-hydroxyethyl methacrylate) (Langer et al., J. Biomed. Mater. Res., 15:167-277 (1981) and Langer, Chem. Tech., 12:98-105 (1982)), ethylene vinyl acetate (Langer et al., supra), or poly-D(-)-3-hydroxybutyric acid (EP 133,988). In certain embodiments, a sustained release composition may also comprise liposomes, which may be prepared by any of several methods known in the art. See, for example, Eppstein et al., Proc. Natl. Acad. Sci. USA, 82:3688-3692 (1985); EP 036,676; EP088,046, and EP 143,949.

[0578] Pharmaceutical compositions for in vivo administration are generally sterile. In certain embodiments, this can be accomplished by filtration through sterile filtration membranes. In certain embodiments, when the composition is lyophilized, sterilization using this method can be carried out either before or after lyophilization and reconstitution. In certain embodiments, compositions for parenteral administration may be stored in lyophilized form or in solution form. In certain embodiments, parenteral compositions are generally placed in a container having a sterile access port, e.g., an intravenous injection solution bag or vial having a stopper that can be pierced by a subcutaneous injection needle.

[0579] In certain embodiments, once the pharmaceutical composition is formulated, it may be stored in a sterile vial in the form of a solution, suspension, gel, emulsion, solid, or as a dehydrated or lyophilized powder. In certain embodiments, such formulations may be stored in ready-to-use or reconstituted-for-administration form (e.g., lyophilized).

[0580] In certain embodiments, a kit for generating a single-dose administration unit is provided. In certain embodiments, the kit may include a first container containing a dry protein and a second container containing an aqueous formulation. In certain embodiments, kits including single-chamber and multi-chamber pre-filled syringes (e.g., liquid syringes and lyophilized syringes) are included.

[0581] In certain embodiments, an effective amount of a pharmaceutical composition comprising an anti-IL-27 antibody for therapeutic use will depend, for example, on the therapeutic context and the target. Those skilled in the art will understand that, according to certain embodiments, a suitable dosage level for treatment will thus vary in part depending on the molecule being delivered, the indication for which the anti-IL-27 antibody is used, the route of administration, and the size (body weight, body surface area, or organ size) and / or condition (age and general health status) of the patient. In certain embodiments, the clinician may titrate the dose and modify the route of administration to obtain an optimal therapeutic effect.

[0582] In certain embodiments, the frequency of administration will take into account the pharmacokinetic parameters of the anti-IL-27 antibody in the formulation used. In certain embodiments, the clinician will administer the composition until a dose that achieves the desired effect is reached. In certain embodiments, the composition may thus be administered over time as a single dose or two or more doses (which may or may not contain the same amount of the desired molecule), or as a continuous infusion via an implant device or catheter. Further refinement of the appropriate dose is routinely performed by those of ordinary skill in the art and within the scope of their routine tasks. In certain embodiments, the appropriate dose can be determined by using appropriate dose-response data.

[0583] In certain embodiments, the route of administration of the pharmaceutical composition follows known methods, such as oral, by injection via intravenous, intraperitoneal, intracerebral (intraparenchymal), intraventricular, intramuscular, subcutaneous, intraocular, intraarterial, intraportal, or intralesional routes; by sustained release systems or implant devices. In certain embodiments, the composition can be administered by bolus injection or continuously by infusion, or by an implant device. In certain embodiments, the individual components of a combination therapy can be administered by different routes.

[0584] In certain embodiments, the composition may be administered locally by implanting a membrane, sponge, or another suitable material on which the desired molecule has been adsorbed or encapsulated. In certain embodiments, in the case of using an implant device, the device may be implanted into any suitable tissue or organ, and delivery of the desired molecule may be effected by diffusion, timed-release bolus, or continuous infusion. In certain embodiments, it may be desirable to use the pharmaceutical composition comprising the anti-IL-27 antibody ex vivo. In such cases, cells, tissues, and / or organs removed from a patient are exposed to the pharmaceutical composition comprising the anti-IL-27 antibody, and thereafter the cells, tissues, and / or organs are implanted back into the patient.

[0585] In certain embodiments, the anti-IL-27 antibody may be delivered by implanting certain genetically engineered cells to express and secrete a polypeptide using methods such as those described herein. In certain embodiments, such cells may be animal or human cells, and may be autologous, allogeneic, or xenogeneic. In certain embodiments, the cells may be immortalized. In certain embodiments, to reduce the chance of an immune response, the cells may be encapsulated to avoid infiltration of the surrounding tissue. In certain embodiments, the encapsulating material is generally a biocompatible, semipermeable polymeric shell or membrane that allows release of one or more protein products but prevents the cells from being destroyed by the patient's immune system or other harmful factors from the surrounding tissue.

[0586] Applications

[0587] The compositions described herein may be used in a variety of diagnostic and therapeutic applications. For example, a labeled antigen-binding molecule may be used to detect the presence or amount of a target antigen in a sample (e.g., a biological sample). The compositions may be used in in vitro assays to study the inhibition of target antigen function. In some embodiments, such as where the composition binds to a complement protein and inhibits the complement protein, the composition may be used as a positive control in an assay designed to identify other novel compounds that inhibit complement activity or otherwise are useful in treating complement-related disorders. For example, an IL-27 inhibitory composition may be used as a positive control in an assay to identify other compounds (e.g., small molecules, aptamers, or antibodies) that reduce or eliminate IL-27 production. The compositions may also be used in the therapeutic methods described in detail below.

[0588] In some embodiments, the present disclosure provides a method for detecting IL-27 in a biological sample or a subject, comprising (i) contacting the sample or subject (and optionally a reference sample or subject) with any antibody described herein under conditions that permit interaction between the antibody molecule and IL-27, and (ii) detecting the formation of a complex between the antibody molecule and the sample or subject (and optionally the reference sample or subject).

[0589] Kits

[0590] The pharmaceutical kit may include the anti-IL-27 antibody disclosed herein and instructions for use. The pharmaceutical kit may contain, in a suitable container, the anti-IL-27 antibody, one or more controls, and various buffers, reagents, enzymes, and other standard components known in the art. In some aspects, the present disclosure provides a pharmaceutical kit that contains the anti-IL-27 antibody or antigen-binding portion disclosed herein, and instructions for use for stimulating an immune response in a subject or treating cancer in a subject, optionally with instructions for combination with one or more additional therapeutic agents or procedures described herein.

[0591] The container may include at least one vial, well, test tube, flask, bottle, syringe, or other container device into which the anti-IL-27 antibody can be placed and, in some cases, appropriately aliquoted therein. If additional components are provided, the pharmaceutical kit may contain additional containers into which the components can be placed. The pharmaceutical kit may also include means for commercially selling the anti-IL-27 antibody and any other reagent containers in a sealed manner. Such containers may include injection-molded or blow-molded plastic containers that hold the required vials therein. The container and / or the pharmaceutical kit may include a label with instructions for use and / or warnings.

[0592] Method of Use

[0593] The compositions of the present invention have many in vitro and in vivo uses, including the detection and / or quantification of IL-27 and / or antagonism of IL-27 function.

[0594] In some embodiments, the present disclosure provides a method of inhibiting or reducing STAT1 and / or STAT3 phosphorylation in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or its antigen-binding portion inhibits or reduces STAT1 and / or STAT3 phosphorylation in the cell.

[0595] In some embodiments, the present disclosure provides a method of inhibiting or reducing the inhibition of CD161 expression in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or its antigen-binding portion inhibits or reduces the inhibition of CD161 expression in the cell.

[0596] In some embodiments, the present disclosure provides a method of inhibiting or reducing PD-L1 and / or TIM-3 expression in a cell, the method comprising contacting the cell with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or its antigen-binding portion inhibits the expression of PD-L1 and / or TIM-3 in the cell.

[0597] In some embodiments, the present disclosure provides methods for inducing or enhancing the secretion of one or more cytokines from cells, the methods comprising contacting the cells with an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, wherein the antibody or its antigen-binding portion induces or enhances the secretion of one or more PD-1-mediated cytokines from the cells.

[0598] In some embodiments, the present disclosure provides methods for stimulating an immune response in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure that specifically binds to and antagonizes human IL-27, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier.

[0599] In some embodiments, the present disclosure provides methods for treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or its antigen-binding portion provided by the present disclosure that specifically binds to and antagonizes IL-27, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier.

[0600] In some embodiments, the present disclosure provides methods for stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells, thereby stimulating the immune response or treating the cancer.

[0601] In some embodiments, the present disclosure provides methods for stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces the inhibition of CD161 expression in cells, thereby stimulating the immune response or treating the cancer.

[0602] In some embodiments, the present disclosure provides methods for stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition inhibits or reduces the expression of PD-L1 and / or TIM-3 in cells, thereby stimulating the immune response or treating the cancer.

[0603] In some embodiments, the present disclosure provides methods of stimulating an immune response in a subject or treating cancer in a subject, the methods comprising administering to the subject an effective amount of an isolated monoclonal antibody or antigen-binding fragment provided by the present disclosure, or a pharmaceutical composition comprising the antibody or its antigen-binding portion and a pharmaceutically acceptable carrier, wherein the antibody or its antigen-binding portion or the pharmaceutical composition induces or enhances the secretion of one or more cytokines mediated by PD-1 from cells, thereby stimulating the immune response or treating the cancer.

[0604] In some embodiments, the cancer is selected from: lung cancer (e.g., non-small cell lung cancer), sarcoma, testicular cancer, ovarian cancer, pancreatic cancer, breast cancer (e.g., triple-negative breast cancer), melanoma, head and neck cancer (e.g., squamous head and neck cancer), colorectal cancer, bladder cancer, endometrial cancer, prostate cancer, thyroid cancer, hepatocellular carcinoma, gastric cancer, brain cancer, lymphoma (e.g., DL-BCL), leukemia (e.g., AML), or kidney cancer (e.g., renal cell carcinoma, e.g., renal clear cell carcinoma).

[0605] The above compositions can be used in particular for methods of treating or preventing various cancers in a subject. The compositions can be administered to a subject (e.g., a human subject) using a variety of methods that depend in part on the route of administration. The route can be, for example, intravenous injection or infusion (IV), subcutaneous injection (SC), intraperitoneal (IP) injection, intramuscular injection (IM), or intrathecal injection (IT). The injection can be a bolus or a continuous infusion.

[0606] Administration can be achieved, for example, by local infusion, injection, or by implantation. The implant can be a porous, non-porous, or gel-like material, including membranes, such as a salivary elastomeric membrane or fibers. The implant can be configured to release the composition to the subject continuously or periodically. See, for example, U.S. Patent Application Publication No. 20080241223; U.S. Patents Nos. 5,501,856, 4,863,457, and 3,710,795; EP488401; and EP 430539, each of which is incorporated herein by reference in its entirety. The composition can be delivered to the subject by an implantable device (e.g., an osmotic pump, a biodegradable implant, an electro-diffusion system, an electro-osmotic system, a vapor pressure pump, an electrolytic pump, a bubbling pump, a piezoelectric pump, an erosion-based system, or an electromechanical system) based on, for example, a diffusion system, an erodible system, or a convection system.

[0607] In some embodiments, the anti-IL-27 antibody or its antigen-binding fragment is therapeutically delivered to the subject by local administration.

[0608] The appropriate dosage of the antibody or fragment thereof described herein (a dosage capable of treating or preventing cancer in a subject) can depend on a variety of factors, including, for example, the age, sex, and weight of the subject to be treated and the particular inhibitor compound used. For example, different dosages of the intact anti-IL-27 antibody may be required to treat a subject with cancer compared to the dosage of the IL-27-binding Fab’ antibody fragment required to treat the same subject. Other factors that affect the dosage administered to a subject include, for example, the type or severity of the cancer. For example, a subject with metastatic melanoma may require a different dosage of the anti-IL-27 antibody compared to a subject with glioblastoma. Other factors can include, for example, other medical conditions that affect the subject concurrently or previously, the general health of the subject, the subject's genetic predisposition, diet, time of administration, rate of excretion, drug combinations, and any other additional therapeutic agents administered to the subject. It should also be understood that the specific dosage and treatment regimen for any particular subject will also depend on the judgment of the practicing physician (e.g., a doctor or nurse). Appropriate dosages are described herein.

[0609] The pharmaceutical composition can comprise a therapeutically effective amount of the anti-IL-27 antibody or antigen-binding fragment thereof described herein. If more than one agent is used, one of ordinary skill in the art can readily determine such an effective amount, in part, based on the effect of the antibody administered or the combined effect of the antibody with one or more additional active agents. The therapeutically effective amount of the antibody or fragment thereof described herein can also vary depending on factors such as the individual's disease state, age, sex, and weight, and the ability of the antibody (and one or more additional active agents) to elicit a desired response (e.g., reduction in tumor growth) in the individual. For example, a therapeutically effective amount of the anti-IL-27 antibody can inhibit (reduce its severity or eliminate its occurrence) and / or prevent a particular disease condition and / or any symptom of a particular disease condition known in the art or described herein. A therapeutically effective amount is also an amount in which any toxic or detrimental effects of the composition are outweighed by the therapeutic beneficial effects.

[0610] The appropriate human dosage of any antibody or fragment thereof described herein can be further evaluated, for example, in a Phase I dose escalation study. See, for example, van Gurp et al. (2008) Am J Transplantation 8(8):1711-1718; Hanouska et al. (2007) Clin Cancer Res 13(2, part 1):523-531; and Hetherington et al. (2006) Antimicrobial Agents and Chemotherapy 50(10):3499-3500.

[0611] In some embodiments, the composition comprises any of the antibodies or antigen-binding fragments thereof described herein and one or more (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven or more) additional therapeutic agents such that the composition is therapeutically effective overall. For example, the composition may comprise an anti-IL-27 antibody and an alkylating agent described herein, wherein the concentrations of the antibody and the agent are each therapeutically effective for treating or preventing cancer (e.g., melanoma) in a subject when combined.

[0612] The toxicity and therapeutic efficacy of such compositions can be determined by known pharmaceutical procedures in cell cultures or experimental animals (e.g., animal models of any of the cancers described herein). These procedures can be used, for example, to determine the LD 50 (lethal dose for 50% of the population) and the ED 50 (therapeutically effective dose for 50% of the population). The dose ratio between toxicity and therapeutic efficacy is the therapeutic index, which can be expressed as the ratio of LD 50 / ED 50 . Antibodies or antigen-binding fragments thereof that exhibit a high therapeutic index are preferred. Although compositions that exhibit toxic side effects can be used, care should be taken to design a delivery system that targets such compounds to the site of the affected tissue and minimizes potential damage to normal cells, thereby reducing side effects.

[0613] Data obtained from cell culture assays and animal studies can be used to formulate a range of doses for use in humans. The dose of such an antibody or antigen-binding fragment is generally within the range of circulating concentrations of the antibody or fragment that includes the ED 50 and has little or no toxicity. The dose can vary within this range depending on the dosage form used and the route of administration employed. For the anti-IL-27 antibody described herein, the therapeutically effective dose can initially be estimated from cell culture assays. Doses can be formulated in animal models to achieve a range of circulating plasma concentrations that includes the IC 50 determined in cell culture (i.e., the antibody concentration that achieves half-maximal inhibition of symptoms). Such information can be used to more accurately determine the useful dose in humans. The levels in plasma can be measured, for example, by high performance liquid chromatography. In some embodiments, for example, in cases where local administration (e.g., to the eye or joint) is desired, cell culture or animal modeling can be used to determine the dose required to achieve a therapeutically effective concentration within the local site.

[0614] In some embodiments, the method can be combined with other therapeutic agents for cancer. For example, the composition can be administered to a subject simultaneously with, prior to, or following radiation, surgery, targeted or cytotoxic chemotherapy, chemoradiation, hormone therapy, immunotherapy, gene therapy, cell transplantation therapy, precision medicine, genome editing therapy, or other pharmaceutical therapies.

[0615] As described above, the compositions described herein (e.g., anti-IL-27 compositions) can be used to treat a variety of cancers, such as but not limited to: Kaposi's sarcoma, leukemia, acute lymphoblastic leukemia, acute myeloid leukemia, myeloblasts promyelocyte myelomonocytic monocyticerythroleukemia, chronic leukemia, chronic myelogenous (granulocytic) leukemia, chronic lymphocytic leukemia, mantle cell lymphoma, primary central nervous system lymphoma, Burkitt lymphoma and marginal zone B-cell lymphoma, polycythemia vera lymphoma, Hodgkin's disease, non-Hodgkin's disease, multiple myeloma, Waldenström macroglobulinemia, heavy chain disease, solid tumors, sarcomas and carcinomas, fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon sarcoma, colorectal cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchial carcinoma, renal cell carcinoma, hepatocellular carcinoma (HCC), hepatoma, cholangiocarcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, uterine cancer, testicular tumors, lung cancer, small cell lung cancer, non-small cell lung cancer, bladder cancer, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, retinoblastoma, nasopharyngeal carcinoma, esophageal cancer, basal cell carcinoma, biliary tract cancer, bladder cancer, bone cancer, brain and central nervous system (CNS) cancer, cervical cancer, choriocarcinoma, colorectal cancer, connective tissue cancer, digestive system cancer, endometrial cancer, esophageal cancer, eye cancer, head and neck cancer, gastric cancer, intraepithelial neoplasia, kidney cancer, laryngeal cancer, liver cancer, lung cancer (small cell, large cell), melanoma, neuroblastoma; oral cancer (e.g., lip, tongue, mouth and pharynx), ovarian cancer, pancreatic cancer, retinoblastoma, rhabdomyosarcoma, rectal cancer; respiratory system cancers, sarcomas, skin cancer, gastric cancer, testicular cancer, thyroid cancer, uterine cancer and urinary system cancer.

[0616] Combination therapy

[0617] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof provided by the present disclosure can be combined with one or more additional therapeutic agents or treatments (e.g., another therapeutic agent or treatment for cancer). For example, the anti-IL-27 antibody or antigen-binding portion thereof can be administered to a subject (e.g., a human patient) in combination with one or more additional therapeutic agents, wherein the combination provides a therapeutic benefit to a subject having cancer or at risk of developing cancer.

[0618] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof and one or more additional therapeutic agents are administered at the same time (e.g., simultaneously). In other embodiments, the anti-IL-27 antibody or antigen-binding portion thereof is administered at a first time, and one or more additional therapeutic agents are administered at a second time (e.g., sequentially). In some embodiments, one or more additional therapeutic agents are administered at a first time, and the anti-IL-27 antibody is administered at a second time.

[0619] The anti-IL-27 antibody or antigen-binding fragment thereof described herein can replace or enhance a previously or currently administered therapy. For example, after treatment with the anti-IL-27 antibody or antigen-binding fragment thereof, the administration of one or more additional therapeutic agents can be stopped or reduced, e.g., one or more additional therapeutic agents are administered at a lower level. In some embodiments, the administration of the previous therapy can be maintained. In some embodiments, the previous therapy will be maintained until the level of the anti-IL-27 antibody reaches a level sufficient to provide a therapeutic effect.

[0620] In some embodiments, the present disclosure provides a method of treating cancer in a subject, the method comprising administering to the subject an effective amount of a monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes IL-27 isolated by the present disclosure in combination with one or more additional therapeutic agents or procedures, wherein the second therapeutic agent or procedure is selected from the group consisting of chemotherapy, targeted anti-cancer therapy, oncolytic drugs, cytotoxic agents, immune-based therapies, cytokines, surgical procedures, radiation procedures, activators of costimulatory molecules, inhibitors of inhibitory molecules, vaccines, or cellular immunotherapies or combinations thereof.

[0621] In some embodiments, one or more additional therapeutic agents are PD-1 antagonists, TIM-3 inhibitors, LAG-3 inhibitors, TIGIT inhibitors, CD112R inhibitors, TAM inhibitors, STING agonists, 4-1BB agonists, or combinations thereof.

[0622] In some embodiments, one or more additional therapeutic agents are PD-1 antagonists. In some embodiments, the PD-1 antagonists are selected from the group consisting of: PDR001, nivolumab, pembrolizumab, pidilizumab, MEDI0680, REGN2810, TSR-042, PF-06801591, and AMP-224. In certain embodiments, one or more additional therapeutic agents are PD-L1 inhibitors. In some embodiments, the PD-L1 inhibitors are selected from the group consisting of: FAZ053, atezolizumab, avelumab, durvalumab, and BMS-936559. In some embodiments, the present disclosure provides a method of enhancing one or more activities of an anti-PD-1 antibody (e.g., enhancing PD-1-mediated cytokine secretion; enhancing anti-PD-1-mediated TNFα secretion; enhancing anti-PD-1-mediated IL-6 secretion from cells exposed to an anti-PD-1 antibody), the method comprising exposing cells to an antibody or an antigen-binding portion thereof provided by the present disclosure, and simultaneously or sequentially exposing to an anti-PD-1 antibody, thereby enhancing one or more activities of the anti-PD1 antibody.

[0623] In some embodiments, one or more additional therapeutic agents are sunitinib cabozantinib axitinib lenvatinib everolimus bevacizumab epalrestat, NKTR-214 (CD-122-biased agonist), tivozanib abexinostat, ipilimumab tremelimumab, pazopanib sorafenib temsirolimus ramucirumab niraparib, savolitinib, vorolanib (X-82), regorafenib donafenib (multi-kinase inhibitor), camrelizumab (SHR-1210), pexastimogene devacirepvec (JX-594), ramucirumab apatinib (YN968D1), doxorubicin capsules tivantinib (ARQ197), ADI-PEG 20, binimetinib, apatinib mesylate, nintedanib, leridumab, nivolumab pembrolizumab pidilizumab avelumab Durvalumab cemiplimab-rwlc Tislelizumab and Spartalizumab.

[0624] In some embodiments, one or more additional therapeutic agents are TIM-3 inhibitors, optionally wherein the TIM-3 inhibitor is MGB453 or TSR-022.

[0625] In some embodiments, one or more additional therapeutic agents are LAG-3 inhibitors, optionally wherein the LAG-3 inhibitor is selected from the group consisting of LAG525, BMS-986016, and TSR-033.

[0626] In some embodiments, one or more additional therapeutic agents are TIGIT inhibitors. In some embodiments, one or more additional therapeutic agents are CD112R inhibitors. In some embodiments, one or more additional therapeutic agents are TAM (Axl, Mer, Tyro) inhibitors. In some embodiments, one or more additional therapeutic agents are STING agonists. In some embodiments, one or more additional therapeutic agents are 4-1BB agonists.

[0627] Combinations with chemotherapeutic agents

[0628] Chemotherapeutic agents suitable for combination and / or co-administration with the compositions of the invention include, for example: paclitaxel, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, teniposide, vincristine, vinblastine, colchicine, doxorubicin, daunorubicin, dihydroxyanthrancindione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and puromycin and their analogs or homologs. Other agents include, for example, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil, dacarbazine), alkylating agents (e.g., mechlorethamine, thiotepa, chlorambucil, melphalan, carmustine (BSNU), lomustine (CCNU), cyclophosphamide, busulfan, dibromomannitol, streptozocin, mitomycin C, cis-dichlorodiammineplatinum (II) (DDP), procarbazine, hexamethylmelamine, cisplatin, carboplatin, oxaliplatin, nedaplatin, satraplatin, or triplatin tetranitrate), anthracyclines (e.g., daunorubicin (formerly called daunomycin) and doxorubicin), antibiotics (e.g., dactinomycin (formerly called actinomycin), bleomycin, mithramycin, and anthramycin (AMC)), and antimitotic agents (e.g., vincristine and vinblastine) and temozolomide.

[0629] Combination with a PD-1 / PD-L1 antagonist

[0630] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof provided by the present disclosure is combined (e.g., co-administered) with one or more PD-1 antagonists that specifically bind to human PD-1 or PD-L1 and inhibit PD-1 / PD-L1 biological activity and / or one or more downstream pathways and / or cellular processes mediated by human PD-1 / PD-L1 signaling or other human PD-1 / PD-L1-mediated functions.

[0631] Accordingly, provided herein are PD-1 antagonists that can directly or allosterically block, antagonize, inhibit, or reduce PD-1 / PD-L1 biological activity, including downstream pathways and / or cellular processes mediated by PD-1 / PD-L1 signaling, such as receptor binding and / or eliciting a cellular response to PD-1 / PD-L1. Also provided herein are PD-1 antagonists that reduce the amount or quantity of human PD-1 or PD-L1 produced by a cell or a subject.

[0632] In some embodiments, the present disclosure provides PD-1 antagonists that bind to human PD-1 and prevent, inhibit, or reduce the binding of PD-L1 to PD-1. In some aspects, the PD-1 antagonist binds to the mRNA encoding PD-1 or PD-L1 and prevents translation. In some embodiments, the PD-1 antagonist binds to the mRNA encoding PD-1 or PD-L1 and causes degradation and / or turnover.

[0633] In some embodiments, the PD-1 antagonist inhibits PD-1 signaling or function. In some embodiments, the PD-1 antagonist blocks the binding of PD-1 to PD-L1, PD-L2, or PD-L1 and PD-L2. In some embodiments, the PD-1 antagonist blocks the binding of PD-1 to PD-L1. In some embodiments, the PD-1 antagonist blocks the binding of PD-1 to PD-L2. In some embodiments, the PD-1 antagonist blocks the binding of PD-1 to PD-L1 and PD-L2. In some embodiments, the PD-1 antagonist specifically binds to PD-1. In some embodiments, the PD-1 antagonist specifically binds to PD-L1. In some embodiments, the PD-1 antagonist specifically binds to PD-L2.

[0634] In some embodiments, the PD-1 antagonist inhibits the binding of PD-1 to its cognate ligand. In some embodiments, the PD-1 antagonist inhibits the binding of PD-1 to PD-L1, PD-1 to PD-L2, or PD-1 to PD-L1 and PD-L2. In some embodiments, the PD-1 antagonist does not inhibit the binding of PD-1 to its cognate ligand.

[0635] In some embodiments, the PD-1 antagonist is a isolated monoclonal antibody (mAb) or an antigen-binding fragment thereof that specifically binds to PD-1 or PD-L1. In some embodiments, the PD-1 antagonist is an antibody or an antigen-binding fragment thereof that specifically binds to human PD-1. In some embodiments, the PD-1 antagonist is an antibody or an antigen-binding fragment thereof that specifically binds to human PD-L1. In some embodiments, the PD-1 antagonist is an antibody or antigen-binding fragment that binds to human PD-L1 and inhibits the binding of PD-L1 to PD-1. In some embodiments, the PD-1 antagonist is an antibody or antigen-binding fragment that binds to human PD-1 and inhibits the binding of PD-L1 to PD-1.

[0636] Several immune checkpoint antagonists that inhibit or disrupt the interaction between PD-1 and one or both of its ligands, PD-L1 and PD-L2, are in clinical development or are currently available to clinicians for the treatment of cancer.

[0637] Examples of anti-human PD-1 monoclonal antibodies or antigen-binding fragments thereof that may comprise a PD-1 antagonist in any of the compositions, methods, and uses provided by the present disclosure include, but are not limited to: (pembrolizumab, MK-3475, h409A11; see US8952136, US8354509, US8900587, and EP2170959, which are hereby incorporated by reference in their entireties; Merck), (nivolumab, BMS-936558, MDX-1106, ONO-4538; see US7595048, US8728474, US9073994, US9067999, EP1537878, US8008449, US8779105, and EP2161336, which are hereby incorporated by reference in their entireties; Bristol Myers Squibb), MEDI0680 (AMP-514), BGB-A317, and BGB-108 (BeiGene), 244C8 and 388D4 (see WO2016106159, which is hereby incorporated by reference; Enumeral Biomedical), PDR001 (Novartis), and REGN2810 (Regeneron). Thus, in some embodiments, the PD-1 antagonist is pembrolizumab. In some embodiments, the PD-1 antagonist is nivolumab.

[0638] Examples of anti-human PD-L1 monoclonal antibodies or antigen-binding fragments thereof that may comprise a PD-1 antagonist in any of the compositions, methods, and uses provided by the present disclosure include, but are not limited to: (Avelumab, MSB0010718C, see WO2013 / 79174, which is incorporated herein by reference in its entirety; Merck / Pfizer), (Durvalumab, MEDI4736), (Atezolizumab, MPDL3280A, RG7446; see WO2010 / 077634, which is incorporated herein by reference in its entirety; Roche), MDX-1105 (BMS-936559, 12A4; see US7943743 and WO2013 / 173223, both of which are incorporated herein by reference in their entirety; Medarex / BMS) and FAZ053 (Novartis). Thus, in some embodiments, the PD-1 antagonist is avelumab. In some embodiments, the PD-1 antagonist is durvalumab. In some embodiments, the PD-1 antagonist is atezolizumab.

[0639] In some embodiments, the PD-1 antagonist is an immunoadhesin that specifically binds to human PD-1 or human PD-L1, e.g., a fusion protein comprising an extracellular portion or a PD-1-binding portion of PD-L1 or PD-L2 fused to a constant region of an immunoglobulin molecule (such as the Fc region). Examples of immunoadhesins that specifically bind to PD-1 are described in WO2010 / 027827 and WO2011 / 066342, both of which are incorporated herein by reference in their entirety. In some embodiments, the PD-1 antagonist is AMP-224 (also known as B7-DCIg), which is a PD-L2-Fc fusion protein that specifically binds to human PD-1.

[0640] One of ordinary skill in the art will understand that any PD-1 antagonist that binds to PD-1 or PD-L1 and disrupts the PD-1 / PD-L1 signaling pathway is suitable for the compositions, methods, and uses disclosed herein.

[0641] In some embodiments, the PD-1 / PD-L1 antagonist is a small molecule, nucleic acid, peptide, peptidomimetic, protein, carbohydrate, carbohydrate derivative, or glycopolymer. Exemplary small molecule PD-1 inhibitors are described in Zhan et al., (2016) Drug Discov Today 21(6):1027-1036.

[0642] Combination with TIM-3 inhibitors

[0643] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof provided by the present disclosure is combined (e.g., co-administered) with a TIM-3 inhibitor. The TIM-3 inhibitor can be an antibody, an antigen-binding fragment thereof, an immunoadhesin, a fusion protein, or an oligopeptide. In some embodiments, the TIM-3 inhibitor is selected from MGB453 (Novartis), TSR-022 (Tesaro), or LY3321367 (Eli Lilly). In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof is co-administered with MGB453. In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof is co-administered with TSR-022.

[0644] Combination with a LAG-3 inhibitor

[0645] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof provided by the present disclosure is combined (e.g., co-administered) with a LAG-3 inhibitor. The LAG-3 inhibitor can be an antibody, an antigen-binding fragment thereof, an immunoadhesin, a fusion protein, or an oligopeptide. In some embodiments, the LAG-3 inhibitor is selected from LAG525 (Novartis), BMS-986016 (Bristol-Myers Squibb), TSR-033 (Tesaro), MK-4280 (Merck & Co), or REGN3767 (Regeneron).

[0646] Other combinations

[0647] In some embodiments, the anti-IL-27 antibody or antigen-binding portion thereof provided by the present disclosure is combined (e.g., co-administered) with a TIGIT inhibitor, a kinase inhibitor (e.g., a tyrosine kinase inhibitor (TKI)), a CD112R inhibitor, a TAM receptor inhibitor, a STING agonist, and / or a 4-1BB agonist or a combination thereof.

[0648] Detection methods

[0649] In some embodiments, the anti-IL-27 antibody or antigen-binding fragment thereof described herein can be used in a method for detecting and / or quantifying human IL-27 in a biological sample. Accordingly, the anti-IL-27 antibody or antigen-binding fragment thereof described herein can be used for diagnosing, predicting, and / or determining the progression of a patient's disease (e.g., cancer).

[0650] As defined herein, monitoring the improvement of cancer in a subject (e.g., a human patient) means assessing changes in the disease parameters of the subject, such as a reduction in tumor growth. In some embodiments, the assessment is performed at least one (1) hour after administration, such as at least 2, 4, 6, 8, 12, 24, or 48 hours, or at least 1 day, 2 days, 4 days, 10 days, 13 days, 20 days or longer, or at least 1 week, 2 weeks, 4 weeks, 10 weeks, 13 weeks, 20 weeks or longer. The subject can be evaluated at one or more of the following times: before starting treatment; during treatment; or after one or more elements of the treatment have been administered. The evaluation can include assessing the need for further treatment, e.g., assessing whether the dose, administration frequency, or treatment duration should be changed. It can also include assessing the need to add or withdraw a selected treatment modality, e.g., adding or withdrawing any cancer treatment described herein.

[0651] In some embodiments, the present disclosure provides a method for detecting IL-27 in a sample from a subject, the method comprising (a) contacting the sample with a detection antibody under conditions that permit the detection antibody to form a detection antibody-IL-27 complex if IL-27 is present in the sample from the subject, wherein the detection antibody is an antibody provided by the present disclosure or an antigen-binding fragment thereof; and (b) detecting the presence of the complex (if any) produced in step (a).

[0652] In some embodiments, the present disclosure provides a method for detecting an IL-27-related cancer in a subject, the method comprising the steps of: (a) contacting a sample from a subject suspected of having an IL-27-related cancer with a detection antibody under conditions that permit the detection antibody to form a detection antibody-IL-27 complex if IL-27 is present in the sample, wherein the detection antibody is an antibody provided by the present disclosure or an antigen-binding portion thereof; and (b) detecting the presence of the complex (if any) produced in step (a). In some embodiments, the detection antibody is conjugated to a detectable label. In some embodiments, the method further comprises contacting the sample with a capture antibody to produce a complex comprising IL-27 and the capture antibody if IL-27 is present in the sample, wherein the capture antibody is an antibody provided by the present disclosure or an antigen-binding portion thereof.

[0653] In some embodiments, the capture antibody is immobilized on a solid support. In some embodiments, the sample is contacted with the capture antibody before the detection antibody. In some embodiments, the sample is a body fluid sample. In some embodiments, the fluid sample is blood, serum, plasma, cell lysate, or tissue lysate.

[0654] In some embodiments, the cancer is selected from renal cell carcinoma (RCC), hepatocellular carcinoma, lung cancer, gastroesophageal cancer, ovarian cancer, endometrial cancer, melanoma, leukemia, and lymphoma. In some embodiments, the cancer is renal cell carcinoma (RCC). In other embodiments, the cancer is hepatocellular carcinoma (HCC). In some embodiments, the cancer is selected from leukemia and lymphoma. In some embodiments, the cancer is acute myeloid leukemia (AML).

[0655] Example

[0656] Although the present disclosure has been described with reference to its specific embodiments, those skilled in the art should understand that various changes and equivalent substitutions can be made without departing from the true spirit and scope of the present disclosure. Additionally, many modifications can be made to adapt a particular situation, material, composition of matter, process, process step to the objective, spirit, and scope of the present disclosure. All such modifications are intended to be within the scope of the present disclosure.

[0657] Example 1: Generation and Characterization of Anti-IL-27 Antibodies That Specifically Bind to the IL-27 EBI3 Monomer

[0658] This example describes the generation of anti-IL-27 antibodies that specifically bind to the EBI3 subunit of human IL-27. Briefly, BALB / c mice were immunized with a human EBI3 immunogenic carrier (Aldevron) and used to generate and isolate hybridomas expressing anti-EBI3 monoclonal antibodies. The isolated hybridomas included hybridomas expressing anti-IL-27 antibody molecules (referred to herein as Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, Ab7, and Ab8). The hybridoma supernatants were analyzed by flow cytometry of mammalian cells expressing surface-targeted human EBI3. All of the tested hybridoma clone supernatants bound to the cells expressing EBI3 (data not shown).

[0659] An exemplary isolated anti-EBI3 antibody, Ab7 (hereinafter referred to as "Ab7", which comprises an immunoglobulin heavy chain variable region (hereinafter referred to as "Ab7-V H0 ") and an immunoglobulin light chain variable region (hereinafter referred to as "Ab7-V L0 ")) was sequenced and further characterized below (the amino-terminal signal peptide sequence is not shown).

[0660] The heavy chain variable region of the isolated Ab7 antibody (Ab7-V H0 ) has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0661] EVKLVESGGGLVQPGGSLKLFCAASGFTFTSYSMSWVRQTPEKRLEWVAYISYDGGSAYYPDTVKGRFSISRDNAKKTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTSVTVSS(SEQ ID NO:1).

[0662] The nucleic acid sequence encoding the heavy chain variable region Ab7-V H0 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0663] GAGGTGAAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAAACTCTTCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGACTCCAGAGAAGAGGCTGGAGTGGGTCGCATACATTAGTTATGATGGTGGTAGCGCCTACTACCCTGACACTGTGAAGGGCCGGTTCTCCATCTCCAGAGACAATGCCAAGAAAACCCTGTATCTGCAAATGAGCAGCCTGAAGTCTGAGGACACGGCCATGTATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCTCAGTCACCGTCTCCTCA(SEQ ID NO:2).

[0664] The light chain variable region of the isolated Ab7 antibody (Ab7-V L0 ) has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0665] DIQMTQSPASLSASVGETVTITCRASENIYSYLAWYQQKQGKSPQLLVYNAETLTEGVPSRFSGSGSGTQFSLKINSLQPEDFGNYYCQHHYGTPLTFGAGTKLDLK(SEQ ID NO:3).

[0666] The heavy chain of the isolated Ab7 antibody (Ab7-V H0 -mIgG2a) has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0667] EVKLVESGGGLVQPGGSLKLFCAASGFTFTSYSMSWVRQTPEKRLEWVAYISYDGGSAYYPDTVKGRFSISRDNAKKTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTSVTVSSAKTTAPSVYPLAPVCGDTTGSSVTLGCLVKGYFPEPVTLTWNSGSLSSGVHTFPAVLQSDLYTLSSSVTVTSSTWPSQSITCNVAHPASSTKVDKKIEPRGPTIKPCPPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKVNNKDLPAPIERTISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIYVEWTNNGKTELNYKNTEPVLDSDGSYFMYSKLRVEKKNWVERNSYSCSVVHEGLHNHHTTKSFSRTPGK (SEQ ID NO:183).

[0668] The nucleic acid sequence encoding the light chain variable region of Ab7-V L0 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0669] GACATCCAGATGACCCAGTCTCCAGCCTCCCTGTCTGCATCTGTAGGAGAAACTGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAGCATGGTATCAGCAGAAACAGGGGAAATCTCCTCAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACACAATTCTCTCTCAAGATCAACAGTCTGCAACCTGAAGATTTTGGGAATTACTACTGTCAACATCATTACGGTACCCCGCTCACATTCGGCGCTGGGACCAAGCTGGATCTGAAA (SEQ ID NO:4).

[0670] The light chain of the isolated Ab7 antibody (Ab7-V L0-mκ) has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0671] DIQMTQSPASLSASVGETVTITCRASENIYSYLAWYQQKQGKSPQLLVYNAETLTEGVPSRFSGSGSGTQFSLKINSLQPEDFGNYYCQHHYGTPLTFGAGTKLDLKADAAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC (SEQ ID NO:184).

[0672] Humanized Ab7 antibodies were designed using methods known in the art. Briefly, the V-region gene sequences encoding murine monoclonal Ab7 antibodies were used to construct a series of fully humanized antibodies. The variable region genes were cloned into vectors encoding the human IgG1 heavy chain constant domain and the human κ light chain constant domain. The chimeric and humanized antibodies were transiently expressed in mammalian cells. Humanization of the isolated Ab7 heavy chain variable region resulted in 5 humanized heavy chain variable region variants (hereinafter referred to as "Ab7-V H1 ", "Ab7-V H2 ", "Ab7-V H3 ", "Ab7-V H4 ", and "Ab7-V H5 "). Humanization of the isolated Ab7 light chain variable region resulted in 4 humanized light chain variable region variants (hereinafter referred to as "Ab7-V L1 ", "Ab7-V L2 ", "Ab7-V L3 ", and "Ab7-V L4 ").

[0673] The protein sequences defining the humanized Ab7 variant variable regions and the nucleotide sequences encoding the humanized Ab7 variant variable regions are summarized below (the amino-terminal signal peptide sequences are not shown).

[0674] The heavy chain variable region Ab7-V H1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0675] EVKLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKKTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTSVTVSS(SE Q ID NO:5).

[0676] The nucleic acid sequence encoding the heavy chain variable region Ab7-V H1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0677] GAGGTGAAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTCGCATACATTAGTTATGATGGTGGTAGCGCCTACTACCCTGACACTGTGAAGGGCCGGTTCACCATCTCCAGAGACAATTCCAAGAAAACCCTGTATCTGCAAATGAGCAGCCTGAAGTCTGAGGACACGGCCATGTATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCTCAGTCACCGTCTCCTCA(SEQ IDNO:6).

[0678] The heavy chain variable region Ab7-V H2 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0679] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTLVTVSS(SEQ ID NO:7).

[0680] The nucleic acid sequence encoding the heavy chain variable region Ab7-V H2 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0681] GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTCGCATACATTAGTTATGATGGTGGTAGCGCCTACTACCCTGACACTGTGAAGGGCCGGTTCACCATCTCCAGAGACAATTCCAAGAACACCCTGTATCTGCAAATGAGCAGCCTGAAGTCTGAGGACACGGCCATGTATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCCTGGTCACCGTCTCCTCA(SEQ IDNO:8).

[0682] Heavy chain variable region Ab7-V H3 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0683] EVKLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKKTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSS(SEQ ID NO:9).

[0684] Nucleic acid sequence encoding the heavy chain variable region Ab7-V H3 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0685] GAGGTGAAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTCGCATACATTAGTTATGATGGTGGTAGCGCCTACTATCCTGACACTGTGAAGGGCCGGTTCACCATCTCCAGAGACAATTCCAAGAAAACCCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTATATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCCTGGTCACCGTCTCCTCA(SEQ IDNO:10).

[0686] Heavy chain variable region Ab7-V H4 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0687] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSS(SEQ ID NO:11).

[0688] The nucleic acid sequence encoding the heavy chain variable region Ab7-V H4 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0689] GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTCGCATACATTAGTTATGATGGTGGTAGCGCCTACTATCCTGACACTGTGAAGGGCCGGTTCACCATCTCCAGAGACAATTCCAAGAACACCCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTATATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCCTGGTCACCGTCTCCTCA(SEQ IDNO:12).

[0690] Heavy chain variable region Ab7-V H5 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0691] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVSYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSS(SEQ ID NO:13).

[0692] Nucleic acid sequence encoding the heavy chain variable region Ab7-V H5 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0693] GAGGTGCAGCTGGTGGAGTCTGGGGGAGGCTTGGTACAGCCTGGGGGGTCCCTGAGACTCTCCTGTGCAGCCTCTGGATTCACCTTTACCAGCTATTCCATGAGCTGGGTCCGCCAGGCTCCAGGGAAGGGGCTGGAGTGGGTGTCTTACATTAGTTATGATGGTGGTAGCGCCTACTATCCTGACACTGTGAAGGGCCGGTTCACCATCTCCAGAGACAATTCCAAGAACACCCTGTATCTGCAAATGAACAGCCTGAGAGCCGAGGACACGGCCGTATATTACTGTGCAAGACATGGAGACTATGACGACGACGACGCGATGGACTACTGGGGCCAAGGAACCCTGGTCACCGTCTCCTCA(SEQ IDNO:14).

[0694] Light chain variable region Ab7-V L1 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0695] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLAWYQQKQGKAPKLLVYNAETLTEGVPSRFSGSGSGTDFTLTISSLQPEDFANYYCQHHYGTPLTFGQGTKLDIK(SEQ ID NO:15).

[0696] Encoding the light chain variable region Ab7-V L1 The nucleic acid sequence has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0697] GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGACAGAGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAGCATGGTATCAGCAGAAACAGGGGAAAGCCCCTAAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTCTGCAACCTGAAGATTTTGCAAATTACTACTGTCAACATCATTACGGTACCCCGCTCACATTCGGCCAAGGGACCAAGCTGGATATCAAA(SEQ ID NO:16).

[0698] Light chain variable region Ab7-V L2 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0699] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLAWYQQKPGKAPKLLVYNAETLTEGVPSRFSGSGSGTDFTLTISSLQPEDFANYYCQHHYGTPLTFGQGTKLEIK(SEQ ID NO:17).

[0700] Encoding the light chain variable region Ab7-V L2 The nucleic acid sequence has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0701] GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGACAGAGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAGCATGGTATCAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTCTGCAACCTGAAGATTTTGCAAATTACTACTGTCAACATCATTACGGTACCCCGCTCACATTCGGCCAAGGGACCAAGCTGGAAATCAAA(SEQ ID NO:18).

[0702] Light chain variable region Ab7-V L3 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0703] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLAWYQQKPGKAPKLLVYNAETLTEGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQHHYGTPLTFGQGTKLEIK(SEQ ID NO:19).

[0704] Encoding the light chain variable region Ab7-V L3 The nucleic acid sequence has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0705] GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGACAGAGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAGCATGGTATCAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTCTGCAACCTGAAGATTTTGCAACTTACTACTGTCAACATCATTACGGTACCCCGCTCACATTCGGCCAAGGGACCAAGCTGGAAATCAAA(SEQ ID NO:20).

[0706] Light chain variable region Ab7-V L4 Has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0707] DIQMTQSPSSLSASVGDRVTITCRASENIYSYLNWYQQKPGKAPKLLVYNAETLTEGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQHHYGTPLTFGQGTKLEIK(SEQ ID NO:21).

[0708] Encoding the light chain variable region Ab7-V L4 The nucleic acid sequence has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4):

[0709] GACATCCAGATGACCCAGTCTCCATCCTCCCTGTCTGCATCTGTAGGAGACAGAGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAAATTGGTATCAGCAGAAACCAGGGAAAGCCCCTAAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTCTGCAACCTGAAGATTTTGCAACTTACTACTGTCAACATCATTACGGTACCCCGCTCACATTCGGCCAAGGGACCAAGCTGGAAATCAAA(SEQ ID NO:22).

[0710] The amino acid sequences of the heavy and light chain CDRs (Kabat definition) of the isolated parental Ab7 antibody are shown in Table 1 below.

[0711] Table 1

[0712]

[0713] To generate full-length chimeric and humanized heavy or light chain antibody sequences, each of the above heavy chain variable regions was combined with the human IgG1 constant region, and each of the above light chain variable regions was combined with the human kappa constant region.

[0714] The protein sequences defining the full-length heavy and light chains of the chimeric and humanized Ab7 variants are summarized below (the amino-terminal signal peptide sequences are not shown).

[0715] Heavy chain Ab7-V H0 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0716] EVKLVESGGGLVQPGGSLKLFCAASGFTFTSYSMSWVRQTPEKRLEWVAYISYDGGSAYYPDTVKGRFSISRDNAKKTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTSVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK(SEQ ID NO:29).

[0717] The nucleic acid sequence encoding the heavy chain Ab7-V H0 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0718]

[0719] Heavy chain Ab7-V H1 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0720] EVKLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKKTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTSVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:31).

[0721] The nucleic acid sequence encoding heavy chain Ab7-V H1 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0722]

[0723] Heavy chain Ab7-V H2 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0724] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMSSLKSEDTAMYYCARHGDYDDDDAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:33).

[0725] The nucleic acid sequence encoding heavy chain Ab7-V H2 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0726]

[0727] Heavy chain Ab7-V H3 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0728] EVKLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKKTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:35).

[0729] The nucleic acid sequence encoding heavy chain Ab7-V H3 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0730]

[0731] Heavy chain Ab7-V H4 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0732] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVAYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:37).

[0733] Encoding heavy chain Ab7-V H4 -The nucleic acid sequence of IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0734]

[0735] Heavy chain Ab7-V H5 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0736] EVQLVESGGGLVQPGGSLRLSCAASGFTFTSYSMSWVRQAPGKGLEWVSYISYDGGSAYYPDTVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARHGDYDDDDAMDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:39).

[0737] The nucleic acid sequence encoding heavy chain Ab7-V H5 -IgG1 has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0738]

[0739] Light chain Ab7-V L0 -κ has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR 3-FR4-constant region):

[0740] DIQMTQSPASLSASVGETVTITCRASENIYSYLAWYQQKQGKSPQLLVYNAETLTEGVPSRFSGSGSGTQFSLKINSLQPEDFGNYYCQHHYGTPLTFGAGTKLDLKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO:41).

[0741] The nucleic acid sequence encoding light chain Ab7-V L0 -κ has the following sequence (FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4-constant region):

[0742] GACATCCAGATGACCCAGTCTCCAGCCTCCCTGTCTGCATCTGTAGGAGAAACTGTCACCATCACTTGCCGGGCAAGTGAGAACATTTACAGCTATTTAGCATGGTATCAGCAGAAACAGGGGAAATCTCCTCAGCTCCTGGTCTATAATGCAGAAACCTTGACAGAAGGGGTCCCATCAAGGTTCAGCGGCAGTGGATCTGGGACACAATTCTCTCTCAAGATCAACAGTCTGCAACCTG...

Claims

1. An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to and antagonizes human IL-27, wherein the antibody or antigen-binding portion thereof exhibits at least one or more of the following characteristics: (i) Binds to human IL-27 with an equilibrium dissociation constant (K D ) of 15 nM or less; (ii) Blocks the binding of IL-27 to the IL-27 receptor; (iii) Inhibits or reduces STAT1 and / or STAT3 phosphorylation in cells; (iv) Inhibits or reduces IL-27-mediated inhibition of CD161 expression in cells; (v) Inhibits or reduces IL-27-mediated PD-L1 and / or TIM-3 expression in cells; and (vi) Induces or increases the secretion of one or more cytokines mediated by PD-1 from cells.

2. The isolated monoclonal antibody or antigen-binding portion thereof according to claim 1, wherein the antibody or antigen-binding portion thereof binds to human IL-27 with an equilibrium dissociation constant (K D ) of 15 nM or less.

3. The isolated monoclonal antibody or antigen-binding portion thereof according to claim 1 or 2, wherein the antibody or antigen-binding portion thereof binds to murine IL-27.

4. An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs selected from the group consisting of: (i) Heavy chain CDR1 consisting of N-GFTFXXXX-C (SEQ ID NO:408), heavy chain CDR2 consisting of N-ISSSXXYI-C (SEQ ID NO:409), and heavy chain CDR3 sequence shown in SEQ ID NO:163; and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs:169, 170, and 171, respectively; or (ii) Heavy chain CDR1 consisting of N-FTFXXXXMN-C (SEQ ID NO:410), heavy chain CDR2 consisting of N-XISSSXXYIXYADSVKG-C (SEQ ID NO:411), and heavy chain CDR3 sequence shown in SEQ ID NO:166; and light chain CDR1, CDR2, and CDR3 sequences shown in SEQ ID NOs:172, 173, and 174, respectively.

5. The isolated monoclonal antibody according to claim 4, wherein the heavy chain CDR1 consists of N-GFTF[S / A / R][S / R][T / Y][G / S]-C (SEQ ID NO: 412) and the heavy chain CDR2 consists of N-ISSS[S / G][S / A]YI-C (SEQ ID NO: 413); or the heavy chain CDR1 consists of N-FTF[S / A / R][S / R][T / Y][G / S]MN-C (SEQ ID NO: 414) and the heavy chain CDR2 consists of N-[G / S]ISSS[S / G][S / A]YI[L / Y]YADSVKG-C (SEQ ID NO: 415).

6. The isolated monoclonal antibody according to claim 4, wherein: (i) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:161, 162, and 163, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:169, 170, and 171, respectively; (ii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:164, 165, and 166, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:172, 173, and 174, respectively; (iii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:73, 74, and 75, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:81, 82, and 83, respectively; (iv) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:76, 77, and 78, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:84, 85, and 86, respectively; (v) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:95, 96, and 97, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:103, 104, and 105, respectively; (vi) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:98, 99, and 100, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:106, 107, and 108, respectively; (vii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:117, 118, and 119, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:125, 126, and 127, respectively; (viii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:120, 121, and 122, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:128, 129, and 130, respectively; (ix) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:139, 140, and 141, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:147, 148, and 149, respectively; (x) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:142, 143, and 144, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO:150, 151, and 152, respectively; (xi) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 51, 52, and 53, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 59, 60, and 61, respectively; or (xii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 54, 55, and 56, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 62, 63, and 64, respectively.

7. An isolated monoclonal antibody or antigen-binding portion thereof that specifically binds to human IL-27, wherein the antibody or antigen-binding portion thereof comprises heavy and light chain CDRs selected from the group consisting of: (i) Separately, heavy chain CDR1 consisting of N-GGSFSXYX-C (SEQ ID NO: 416), heavy chain CDR2 consisting of N-IDXSGXT-C (SEQ ID NO: 417), and heavy chain CDR3 sequence consisting of N-ARXXXYY[X] n＝0-1 DSS[X] n＝4-6 DX-C (SEQ ID NO: 418); and light chain CDR1 consisting of N-QXXSXY-C (SEQ ID NO: 419), light chain CDR2 consisting of N-DXS-C (SEQ ID NO: 420), and light chain CDR3 sequence consisting of N-QQXXDXPIT-C (SEQ ID NO: 421); or (ii) Separately, the heavy chain CDR1 consisting of N-GSFSXYXWS-C (SEQ ID NO:422), the heavy chain CDR2 consisting of N-SIDXSGXTXYNPSLKS-C (SEQ ID NO:423), and the heavy chain CDR3 sequence consisting of N-ARXXXYY[X] n＝0-1 DSS[X] n＝4- 6DX-C (SEQ ID NO:418); and the light chain CDR1 consisting of N-XASQXXSXYLX-C (SEQ ID NO:424), the light chain CDR2 consisting of N-DXSNXXT-C (SEQ ID NO:425), and the light chain CDR3 sequence consisting of N-QQXXDXPIT-C (SEQ ID NO:421).

8. The isolated monoclonal antibody according to claim 7, wherein: (i) Separately, the heavy chain CDR1 consists of N-GGSFS[R / D]Y[E / Y]-C (SEQ ID NO: 426), the heavy chain CDR2 consists of N-ID[W / Y]SG[I / S]T-C (SEQ ID NO: 427), and the heavy chain CDR3 sequence consists of N-AR[D / L][P / G][M / V]YY[- / Y]DSS[VSTGSV / DLGF]D[V / I]-C (SEQ ID NO: 428); and the light chain CDR1 consists of N-Q[S / D][V / I]S[S / N]Y-C (SEQ ID NO: 429), the light chain CDR2 consists of N-D[S / A]S-C (SEQ ID NO: 430), and the light chain CDR3 sequence consists of N-QQ[D / Y][S / D]D[H / L]PIT-C (SEQ ID NO: 431); or (ii) Separately, the heavy chain CDR1 consists of N-GSFS[R / D]Y[E / Y]WS-C (SEQ ID NO: 432), the heavy chain CDR2 consists of N-SID[W / Y]SG[I / S]T[N / E]YNPSLKS-C (SEQ ID NO: 433), and the heavy chain CDR3 sequence consists of N-AR[D / L][P / G][M / V]YY[- / Y]DSS[VSTGSV / DLGF]D[V / I]-C (SEQ ID NO: 428); and the light chain CDR1 consists of N-[Q / R]ASQ[S / D][V / I]S[S / N]YL[N / A]-C (SEQ ID NO: 434), the light chain CDR2 consists of N-D[S / A]SN[R / L][A / E]T-C (SEQ ID NO: 435), and the light chain CDR3 sequence consists of N-QQ[D / Y][S / D]D[H / L]PIT-C (SEQ ID NO: 431).

9. The isolated monoclonal antibody according to claim 7, wherein: (i) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 229, 230, and 231, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 237, 238, and 239, respectively; or (ii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 232, 233, and 234, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 240, 241, and 242, respectively.

10. The isolated monoclonal antibody according to claim 7, wherein: (i) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 251, 252, and 253, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 259, 260, and 261, respectively; or (ii) The heavy chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 254, 255, and 256, respectively, and the light chain CDR1, CDR2, and CDR3 sequences are shown in SEQ ID NO: 262, 263, and 264, respectively.

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