A pharmaceutical composition and its use

Abstract

The present invention provides a pharmaceutical composition, which includes a protein and an immune checkpoint inhibitor, wherein the protein includes a fusion protein, and the fusion protein includes cytokines IL12, IL2 and GMCSF. The present invention also provides a kit including the pharmaceutical composition and the use of the pharmaceutical composition or the kit in preparing a drug for treating tumors.

Description

Technical Field

[0001] This application relates to the field of biomedicine, specifically to a pharmaceutical composition and its use. Background Technology

[0002] Cancer is a serious threat to human health. In recent years, immunotherapy, as a novel treatment method, has shown great potential in cancer treatment. Cytokines are crucial immune signals in the body, and cytokine fusion protein technology is another hot topic in cancer immunotherapy today. This method is based on the fact that these cytokines have the same or related functional activities but different target sites, and uses genetic engineering technology to fuse two or more cytokines together. However, the efficacy of using cytokine fusion protein technology for cancer treatment is still unsatisfactory, and there are many areas that need improvement. Summary of the Invention

[0003] On one hand, this application provides a pharmaceutical composition comprising a protein and an immune checkpoint inhibitor, wherein the protein comprises a fusion protein, and the fusion protein contains the cytokines IL12, IL2, and GMCSF.

[0004] In some embodiments, the immune checkpoint inhibitors include inhibitors of PD1, PD-L1, and / or CTLA-4.

[0005] In some implementations, the cytokines are derived from mammals.

[0006] In some embodiments, the protein also includes a targeting portion.

[0007] In some implementations, the targeting portion is capable of specifically recognizing and / or binding to tumor-associated antigens.

[0008] In some embodiments, the tumor-associated antigen is selected from the group consisting of the EDB domain of fibronectin, the EDA domain of fibronectin, and necrotic regions.

[0009] In some embodiments, the targeting portion includes an antibody or an antigen-binding fragment thereof.

[0010] In some embodiments, the targeting portion comprises an amino acid sequence shown in any of the following groups: SEQ ID NO. 1-15.

[0011] In some embodiments, the protein is a single-chain protein.

[0012] In some embodiments, the single-chain protein comprises the amino acid sequence shown in any of the following groups: SEQ ID NO.27-52.

[0013] In some embodiments, the protein is a dimer composed of a first polypeptide chain and a second polypeptide chain, wherein the first polypeptide chain is different from the second polypeptide chain.

[0014] In some embodiments, the first polypeptide chain contains IL12a and the second polypeptide chain contains IL12b.

[0015] In some embodiments, the IL2 or a functional fragment thereof is located in the first polypeptide chain or the second polypeptide chain, and the GMCSF or a functional fragment thereof is located in the first polypeptide chain or the second polypeptide chain.

[0016] In some embodiments, the IL2 or a functional fragment thereof is located in the first polypeptide chain or the second polypeptide chain, the GMCSF or a functional fragment thereof is located in the first polypeptide chain or the second polypeptide chain, and one or more of the target moieties are each independently located in the first polypeptide chain or the second polypeptide chain.

[0017] In some embodiments, the first polypeptide chain contains, from the N-terminus to the C-terminus, the IL12a or a functional fragment thereof and the IL2 or a functional fragment thereof; or, the first polypeptide chain contains, from the N-terminus to the C-terminus, the IL2 or a functional fragment thereof and the IL12a or a functional fragment thereof; or, the first polypeptide chain contains, from the N-terminus to the C-terminus, the IL12a or a functional fragment thereof and the GMCSF or a functional fragment thereof.

[0018] In some embodiments, the second polypeptide chain comprises, from the N-terminus to the C-terminus, the IL12b or a functional fragment thereof and the GMCSF or a functional fragment thereof; or, the second polypeptide chain comprises, from the N-terminus to the C-terminus, the GMCSF or a functional fragment thereof and the IL12b or a functional fragment thereof; or, the second polypeptide chain comprises, from the N-terminus to the C-terminus, the IL12b or a functional fragment thereof and the IL2 or a functional fragment thereof.

[0019] In some embodiments, in the pharmaceutical composition,

[0020] a) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 53 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 57;

[0021] b) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 54 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 57;

[0022] c) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 53 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 58;

[0023] d) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 54 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 58;

[0024] e) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 55 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 59;

[0025] f) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO.56 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO.60.

[0026] In some embodiments, the first polypeptide chain may, from the N-terminus to the C-terminus, sequentially include the targeting portion, the IL12a or a functional fragment thereof, the IL2 or a functional fragment thereof, and the GMCSF or a functional fragment thereof; or, the first polypeptide chain may, from the N-terminus to the C-terminus, sequentially include the IL2 or a functional fragment thereof, the IL12a or a functional fragment thereof, and the GMCSF or a functional fragment thereof.

[0027] In some embodiments, the second polypeptide chain comprises, from the N-terminus to the C-terminus, the IL12b or a functional fragment thereof and the targeting portion.

[0028] In some embodiments, in the pharmaceutical composition,

[0029] 1) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 61;

[0030] 2) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 62;

[0031] 3) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 63;

[0032] 4) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 67 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 61;

[0033] 5) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 68 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 62;

[0034] 6) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 69 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 63;

[0035] 7) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 70 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO. 64;

[0036] 8) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO.71 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO.64;

[0037] 9) The first polypeptide chain contains the amino acid sequence shown in SEQ ID NO.72 and the second polypeptide chain contains the amino acid sequence shown in SEQ ID NO.65.

[0038] On the other hand, this application provides a kit comprising the aforementioned pharmaceutical composition.

[0039] On the other hand, this application also provides the use of the pharmaceutical composition or the kit in the preparation of a medicament for treating tumors.

[0040] In some implementations, the tumor includes a solid tumor.

[0041] In some implementations, the tumor includes melanoma.

[0042] In some implementations, the tumor includes breast cancer.

[0043] In some embodiments, the tumor includes lung cancer.

[0044] In some embodiments, the pharmaceutical composition or kit described in this application is used to treat tumors.

[0045] On the other hand, this application also provides a method for treating tumors, which includes administering the pharmaceutical composition or the kit to a subject in need.

[0046] In some embodiments, the administration includes administering the protein first, followed by administering the immune checkpoint inhibitor.

[0047] In some embodiments, the administration includes intratumoral injection, intravenous injection, or subcutaneous injection.

[0048] In some implementations, the tumor includes a solid tumor.

[0049] In some implementations, the tumor includes melanoma.

[0050] In some implementations, the tumor includes breast cancer.

[0051] In some embodiments, the tumor includes lung cancer.

[0052] Other aspects and advantages of this application will readily be apparent to those skilled in the art from the detailed description below. Only exemplary embodiments of this application are shown and described in the following detailed description. As will be appreciated by those skilled in the art, the content of this application enables them to make modifications to the disclosed specific embodiments without departing from the spirit and scope of the invention to which this application pertains. Accordingly, the descriptions in the accompanying drawings and specification of this application are merely exemplary and not restrictive. Attached Figure Description

[0053] The specific features of the invention involved in this application are shown in the appended claims. The features and advantages of the invention can be better understood by referring to the exemplary embodiments and drawings described in detail below. A brief description of the drawings is as follows:

[0054] Figure 1 A-1C shows the effect of inducing expression of mIL12aIL2-IL12bGMCSF heterodimer on PD1 expression in T cells.

[0055] Figure 2 The image shows the growth of melanoma in mice after treatment with a combination of mIL12bIL12aIL2GMCSF, mIL12bIL12aIL2GMCSF and PD1 antibody as described in this application.

[0056] Figure 3 The image shows the growth of melanoma in mice after treatment with a combination of mIL12bIL12aIL2DiaNHS76F8GMCSF, mIL12bIL12aIL2DiaNHS76F8GMCSF, and PD1 antibody as described in this application.

[0057] Figure 4 The image shows the growth of melanoma in mice after treatment with a combination of mIL12aIL2-IL12bGMCSF, mIL12aIL2-IL12bGMCSF and PD1 antibody as described in this application.

[0058] Figure 5The image shows the growth of breast cancer in mice after treatment with a combination of mIL12bIL12aIL2DiaNHS76F8GMCSF, mIL12bIL12aIL2DiaNHS76F8GMCSF, and PD1 antibody as described in this application.

[0059] Figure 6 The image shows the growth of lung cancer in mice after treatment with a combination of mIL12bIL12aIL2DiaNHS76F8GMCSF, mIL12bIL12aIL2DiaNHS76F8GMCSF, and PD1 antibody as described in this application. Detailed Implementation

[0060] The following specific embodiments illustrate the implementation of the invention. Those skilled in the art can easily understand other advantages and effects of the invention from the content disclosed in this specification.

[0061] Terminology Definition

[0062] In this application, the term "pharmaceutical composition" generally refers to a composition suitable for administration to a patient. The pharmaceutical compositions of this application include proteins and immune checkpoint inhibitors, wherein the proteins include fusion proteins, and the fusion proteins contain the cytokines IL12, IL2, and GMCSF. In some embodiments, the pharmaceutical composition may also contain suitable formulations of one or more (pharmaceutically effective) carriers, stabilizers, excipients, diluents, solubilizers, surfactants, emulsifiers, preservatives, and / or adjuvants. For example, the acceptable components of the composition are non-toxic to the recipient at the dose and concentration used. The pharmaceutical compositions of this application include, but are not limited to, liquid, freeze-dried, and lyophilized compositions.

[0063] In this application, the term "immune checkpoint inhibitor" generally refers to a molecule that, in whole or in part, reduces, inhibits, interferes with, or regulates one or more checkpoint proteins. Checkpoint proteins regulate T cell activation or function, and various checkpoint proteins are known, such as CTLA-4 and its ligands CD80 and CD86; and PD1 and its ligands PD-L1 and PD-L2 (Pardoll, Nature Reviews Cancer 12: 252-264, 2012). These proteins are responsible for co-stimulatory or inhibitory interactions in T cell responses. Immune checkpoint proteins regulate and maintain self-tolerance and the duration and magnitude of physiological immune responses. Immune checkpoint inhibitors include antibodies or antibody-derived antibodies. For example, in this application, the immune checkpoint inhibitor may include inhibitors of PD1, PD-L1, and / or CTLA-4.

[0064] In this application, the term "protein" can be considered as belonging to "cytokine fusion protein," which generally refers to a fusion protein obtained by fusing two or more cytokines together through gene recombination technology. It not only possesses the unique biological activities of its constituent factors or significantly enhances some of their activities, but may also exert complex biological functions that are not present in simple combinations of single cytokines through complementary and synergistic effects of biological activities, and may even generate some new structures and biological functions.

[0065] In this application, the terms "IL12," "IL12a," "IL12b," "IL2," and "GMCSF" can be considered as belonging to the category of "cytokines." The term "cytokines" generally refers to a class of small molecule proteins with broad biological activity synthesized and secreted by immune cells (such as monocytes, macrophages, T cells, B cells, NK cells, etc.) and certain non-immune cells (e.g., endothelial cells, epidermal cells, fibroblasts, etc.) upon stimulation. These cytokines play an important regulatory role in cell-cell interactions, cell growth, and differentiation. In this application, the cytokines may be selected from one or more of the following: interleukins (IL) and colony-stimulating factors (CSF). Interleukins generally refer to cytokines produced by lymphocytes, monocytes, or other non-monocytes. In this application, interleukins may be selected from one or more of the following: IL12 and IL2. In this application, colony-stimulating factors generally refer to cytokines that can stimulate different hematopoietic stem cells to form cell colonies in a semi-solid culture medium. In this application, the colony-stimulating factor may be granulocyte-macrophage colony-stimulating factor (GMCSF).

[0066] In this application, the term "IL12" generally refers to interleukin-12, which plays an important regulatory role in intercellular interactions, immune regulation, hematopoiesis, and inflammation. The IL12 molecule is typically a heterodimer, usually comprising two subunits: a p40 subunit (40 kDa) and a p35 subunit (35 kDa), linked together by disulfide bonds. In this application, IL12 containing the p35 subunit (35 kDa) can be represented as IL12a, and IL12 containing the p40 subunit (40 kDa) can be represented as IL12b. For example, the p35 subunit of mouse-derived IL12 (mIL12) may contain the amino acid sequence shown in SEQ ID NO. 16, and the p40 subunit may contain the amino acid sequence shown in SEQ ID NO. 17. For example, the p35 subunit of human IL12 (hIL12) may contain the amino acid sequence shown in SEQ ID NO.18, and the p40 subunit may contain the amino acid sequence shown in SEQ ID NO.19.

[0067] In this application, the term "IL2" generally refers to interleukin-2, which plays an important regulatory role in cell-cell interactions, immune regulation, hematopoiesis, and inflammation. For example, mouse-derived IL2 (mIL2) may contain the amino acid sequence shown in SEQ ID NO. 76, and human-derived IL2 (hIL2) may contain the amino acid sequence shown in SEQ ID NO. 77.

[0068] In this application, the term "GMCSF" generally refers to granulocyte-macrophage colony-stimulating factor. The GMCSF may have a four-alpha helical bundle structure. For example, mouse-derived GMCSF (mGMCSF) may contain the amino acid sequence shown in SEQ ID NO. 20. As another example, human-derived GMCSF (hGMCSF) may contain the amino acid sequence shown in SEQ ID NO. 21.

[0069] In this application, the term "antibody" generally refers to an immunoglobulin or a fragment thereof or a derivative thereof, encompassing any polypeptide that includes an antigen-binding site, whether it is produced in vitro or in vivo. This term includes, but is not limited to, polyclonal, monoclonal, single-specific, multi-specific, non-specific, humanized, single-chain, chimeric, synthetic, recombinant, hybrid, mutated, and transplanted antibodies. Unless otherwise modified by the term "complete," such as in "complete antibody," for the purposes of this invention, the term "antibody" also includes antibody fragments, such as Fab, F(ab′)2, Fv, scFv, Fd, dAb, and other antibody fragments that retain antigen-binding function. Typically, such fragments should include an antigen-binding domain.

[0070] A basic 4-chain antibody unit is a heterotetrameric glycoprotein composed of two identical light (L) chains and two identical heavy (H) chains. IgM antibodies consist of five basic heterotetrameric units plus a polypeptide chain called the J chain and contain 10 antigen-binding sites, while IgA antibodies consist of 2-5 basic 4-chain units that can polymerize with the J chain to form a multivalent combination. For IgG, a 4-chain unit is typically about 150,000 Daltons. Each L chain is linked to an H chain by a covalent disulfide bond, and two H chains are linked to each other by one or more disulfide bonds depending on the H chain isoform. Each H and L chain also has regularly spaced intrachain disulfide bridges. Each H chain has a variable domain (VH) at its N-terminus, followed by three constant domains (CH) for α and γ chains, and four CH domains for μ and ε isoforms. Each L chain has a variable domain (VL) at its N-terminus and a constant domain at its other end. VL corresponds to VH, and CL corresponds to the first constant domain (CH1) of the heavy chain. Specific amino acid residues are thought to form interfaces between the variable domains of the light and heavy chains. VH and VL pair together to form a single antigen-binding site. For the structure and properties of different classes of antibodies, see, for example, Basic and Clinical Immunology, 8th Edition, Daniel P. Sties, Abba I. Terr and Tristram G. Parsolw (eds), Appleton & Lange, Norwalk, Conn., 1994, p. 71 and Chapter 6. The L chain from any vertebrate species can be classified into one of two distinct types, called κ and λ, based on the amino acid sequence of its constant domain. Immunoglobulins can be classified into different classes or isotypes depending on the amino acid sequence of their heavy chain (CH) constant domain. There are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, with heavy chains named α, δ, ε, γ, and μ, respectively. Based on relatively small differences in CH sequence and function, γ and α classes are further divided into subclasses. For example, humans express the following subclasses: IgG1, IgG2A, IgG2B, IgG3, IgG4, IgA1, and IgK1.

[0071] In this application, the term "targeting portion" generally refers to a portion that acts against specific tissues or cells. For example, a targeting portion can specifically target tumor-associated antigens. In this application, the targeting portion includes an antibody or its antigen-binding fragment.

[0072] In this application, the term "specific recognition and / or binding" generally refers to a measurable and reproducible interaction, such as the binding between a target and an antibody. For example, an antibody that specifically binds to a target (which may be an epitope) binds to that target with greater affinity, strength, ease, and / or duration than it binds to other targets. In some embodiments, the antibody specifically binds to an epitope on a protein that is conserved across proteins of different species. In another embodiment, specific binding may include, but is not required to be, exclusive binding.

[0073] In this application, the term "tumor-associated antigen" (TAA) generally refers to an antigen molecule present on tumor cells or normal cells. The tumor-associated antigen may include embryonic proteins, glycoprotein antigens, and squamous cell antigens. The tumor-associated antigen may be selected from the group consisting of the EDB domain of fibronectin, the EDA domain of fibronectin, and necrotic regions.

[0074] In this application, the term "antigen-binding fragment" generally refers to a fragment having antigen-binding activity. In this application, the antigen-binding fragment may be selected from the group consisting of: Fab, Fab', F(ab')2, F(ab)2, dAb, separated complementarity-determining regions (CDRs), Fv, and scFv.

[0075] In this application, the term "single-chain protein" generally refers to a polypeptide with a primary structure consisting of an uninterrupted sequence of a single continuous amino acid residue. For example, in this application, the single-chain protein may comprise the amino acid sequence shown in any of the following groups: SEQ ID NO.27-52.

[0076] In this application, the term "dimer" generally refers to a polymeric complex formed by two monomeric units that are typically non-covalently bonded. Each monomeric unit can be a macromolecule, such as a polypeptide chain or a polynucleotide. For example, in this application, the protein can be a dimer composed of a first polypeptide chain and a second polypeptide chain.

[0077] In this application, the term "polypeptide chain" generally refers to a macromolecule comprising two or more covalently linked peptides. Peptides within a polypeptide chain can be linked to each other by a peptide bond. Each polypeptide chain may include an N-terminus or amino terminus and a C-terminus or carboxyl terminus.

[0078] In this application, the term "functional fragment" generally refers to a fragment that retains a specific function. For example, an IL12a functional fragment refers to a fragment that retains the function of IL12a. For instance, an IL12a functional fragment could be IL12a, fragment (GenBank: AIC49052.1). As another example, an IL12b functional fragment could be IL12b, fragment (GenBank: AIC54621.1).

[0079] In this application, the term "kit" generally refers to a packaged product containing the pharmaceutical composition of this application. The kit may contain a box or container holding the kit components. The box or container is labeled or labeled with a Food and Drug Administration-approved treatment regimen. The box or container holds the components of the pharmaceutical composition of this application, for example, the components may be contained in a plastic, polyethylene, polypropylene, ethylene, or propylene container. The container may be a capped tube or bottle. Furthermore, the kit may also contain instructions for use to administer the pharmaceutical composition of this application.

[0080] In this application, the term "tumor" generally refers to a growth or solid lesion formed by abnormal cell growth. In this application, a tumor can be a solid tumor or a hematologic malignancy. For example, a tumor can include melanoma.

[0081] In this application, the term "subject" generally refers to a human or non-human animal, including but not limited to cats, dogs, horses, pigs, cows, sheep, rabbits, mice, rats, or monkeys.

[0082] In this application, the term "administration" generally refers to a method of giving a dose of a liquid formulation or drug to a subject (e.g., a patient). Administration can be performed by any suitable means, including parenteral, intrapulmonary, and intranasal administration, as well as intralesional administration (if required for local treatment). Parenteral administration includes, for example, intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Administration can be carried out by any suitable route, such as by injection (e.g., intravenous or subcutaneous), depending in part on whether the administration is transient or prolonged. Various dosing schedules are covered herein, including but not limited to single administration or multiple administrations at various time points, bolus administration, and pulsatile infusion. For example, in this application, the administration may be intratumoral injection. "Intratumoral injection" generally refers to injecting a dose of a liquid formulation or drug into the tumor. In some cases, the administration may also be intravenous or subcutaneous injection.

[0083] In this application, the term "comprising" generally means including the explicitly specified features, but does not exclude other elements.

[0084] In this application, the term "about" generally refers to a variation within a range of 0.5% to 10% above or below a specified value, such as a variation within a range of 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10% above or below a specified value. Invention Details

[0086] Pharmaceutical Compositions and Kits

[0087] On one hand, this application provides a pharmaceutical composition comprising a protein and an immune checkpoint inhibitor, wherein the protein comprises a fusion protein, and the fusion protein contains the cytokines IL12, IL2, and GMCSF.

[0088] In this application, the immune checkpoint inhibitor may include inhibitors of PD1, PD-L1, and / or CTLA-4. For example, the immune checkpoint inhibitor may be an antibody against PD1, PD-L1, and / or CTLA-4.

[0089] In this application, the cytokines may be derived from mammals. For example, in some embodiments, the mammal may be human or mouse. For example, the amino acid sequence of mouse-derived IL12a (represented as mIL12a) may be as shown in SEQ ID NO. 16, the amino acid sequence of mouse-derived IL12b (represented as mIL12b) may be as shown in SEQ ID NO. 17, and the amino acid sequence of mouse-derived GMCSF (represented as mGMCSF) may be as shown in SEQ ID NO. 20. As another example, the amino acid sequence of human-derived IL12a (represented as hIL12a) may be as shown in SEQ ID NO. 18, the amino acid sequence of human-derived IL12b (represented as hIL12b) may be as shown in SEQ ID NO. 19, and the amino acid sequence of human-derived GMCSF (represented as hGMCSF) may be as shown in SEQ ID NO. 21. As yet another example, the amino acid sequence of mouse-derived IL2 may be as shown in SEQ ID NO. 76, and the amino acid sequence of human-derived IL2 may be as shown in SEQ ID NO. 77.

[0090] In this application, the protein may further include a targeting portion. The number of targeting portions may be one or more. The targeting portions may be the same or different. The targeting portions are capable of specifically recognizing and / or binding to tumor-associated antigens. The tumor-associated antigens may be selected from the group consisting of: the EDB domain of fibronectin, the EDA domain of fibronectin, and necrotic regions.

[0091] In this application, the targeting portion may include an antibody or an antigen-binding fragment thereof. The antigen-binding fragment may be selected from the group consisting of: Fab, Fab', F(ab')2, F(ab)2, dAb, isolated complementarity-determining regions (CDRs), Fv, and scFv. In some embodiments, the antigen-binding fragment may be scFv.

[0092] In this application, the targeting portion may comprise any of the amino acid sequences shown in the following group: SEQ ID NO. 1-15.

[0093] For example, the targeting portion of the protein can be selected from the following group: L19V L (Its amino acid sequence may be as shown in SEQ ID NO. 10), L19V H (Its amino acid sequence may be as shown in SEQ ID NO.11), F8V L (Its amino acid sequence may be as shown in SEQ ID NO.12), F8V H (Its amino acid sequence may be as shown in SEQ ID NO.13), NHS76V L (Its amino acid sequence may be as shown in SEQ ID NO.14) and NHS76V H (Its amino acid sequence may be as shown in SEQ ID NO.15).

[0094] In this application, the cytokines can be linked to each other or to the target region via linkers. The linker can be a linker peptide. In some embodiments, the linker may contain a thrombin cleavage site.

[0095] In this application, the linker may comprise any of the amino acid sequences shown in the following group: SEQ ID NO. 22-26.

[0096] For example, the cytokines can be linked together via the linker. In this application, IL12a, IL12b, IL2, and GMCSF can be linked together via the linker peptide. For example, the linker peptide may contain an amino acid sequence as shown in any one of SEQ ID NO. 22 and 24.

[0097] For example, the cytokines and the targeting moiety can be linked by the linker. In this application, the targeting moiety can be linked to IL12a, IL12b, IL2, and GMCSF via the linker peptide. For example, the linker peptide may contain the amino acid sequence shown in any of SEQ ID NO. 22-26.

[0098] In this application, the protein may be a single-chain protein, which may contain the amino acid sequence shown in any of the following groups: SEQ ID NO.27-52.

[0099] For example, the structure of the single-chain protein may be such that the C-terminus of mIL12b is fused with the N-terminus of mIL12a, the C-terminus of mIL12a is fused with the N-terminus of mIL2, and the C-terminus of mIL2 is fused with the N-terminus of mGMCSF, thereby forming the mIL12b-mIL12a-mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.27), and may be simply referred to as mIL12bIL12aIL2GMCSF protein.

[0100] For example, the structure of the single-chain protein may be such that the C-terminus of mIL12b is fused with the N-terminus of mIL12a, the C-terminus of mIL12a is fused with the N-terminus of mGMCSF, and the C-terminus of mGMCSF is fused with the N-terminus of mIL2, thereby forming a mIL12b-mIL12a-mGMCSF-mIL2 protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.28), and may be simply referred to as mIL12bIL12aGMCSFIL2 protein.

[0101] For example, the structure of the single-chain protein can be such that the C-terminus of hIL12b is fused with the N-terminus of hIL12a, the C-terminus of hIL12a is fused with the N-terminus of hIL2, and the C-terminus of hIL2 is fused with the N-terminus of hGMCSF, thereby forming the hIL12b-hIL12a-hIL2-hGMCSF protein molecule (whose amino acid sequence can be as shown in SEQ ID NO.29), and can be simply referred to as hIL12bIL12aIL2GMCSF protein.

[0102] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mL19V. H N-terminal fusion, mL19V H C-terminus and mL19V L N-terminal fusion, mL19V L C-terminus and mL19V H N-terminal fusion, mL19V H C-terminus and mL19V L N-terminal fusion, mL19V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mL19V. H -mL19V L -mL19VH -mL19V L The mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.30) may be simply referred to as mIL12bIL12aDiaL19IL2GMCSF protein.

[0103] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mF8V. H N-terminal integration, mF8V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-end and mF8V L N-terminal integration, mF8V L The C-terminus of mIL12 and the N-terminus of mIL2 fuse, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mF8V. H -mF8V L -mF8V H -mF8V L The -mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.31) may be simply referred to as mIL12bIL12aDiaF8IL2GMCSF protein.

[0104] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mNHS76V. H N-terminal fusion, mNHS76V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L C-terminal and mNHS76V H N-terminal fusion, mNHS76V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mNHS76V. H -mNHS76V L -mNHS76V H -mNHS76V LThe mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.32) can be simply referred to as mIL12bIL12aDiaNHS76IL2GMCSF protein.

[0105] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mNHS76V. H N-terminal fusion, mNHS76V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mNHS76V. H -mF8V L -mF8V H -mNHS76V L The mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.33) may be simply referred to as mIL12bIL12aDiaNHS76F8IL2GMCSF protein.

[0106] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mNHS76V. H N-terminal fusion, mNHS76V H C-terminus and mL19V L N-terminal fusion, mL19V L C-terminus and mL19V H N-terminal fusion, mL19V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mNHS76V. H -mL19V L -mL19V H -mNHS76V LThe -mIL2-mGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.34) can be simply referred to as mIL12bIL12aDiaNHS76L19IL2GMCSF protein.

[0107] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mF8V. H N-terminal integration, mF8V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L C-terminal and mNHS76V H N-terminal fusion, mNHS76V H C-end and mF8V L N-terminal integration, mF8V L The C-terminus of mIL12 and the N-terminus of mIL2 fuse, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mF8V. H -mNHS76V L -mNHS76V H -mF8V L The mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.35) can be abbreviated as mIL12bIL12aDiaF8NHS76IL2GMCSF protein.

[0108] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mF8V. H N-terminal integration, mF8V H C-terminus and mL19V L N-terminal fusion, mL19V L C-terminus and mL19V H N-terminal fusion, mL19V H C-end and mF8V L N-terminal integration, mF8V L The C-terminus of mIL12 and the N-terminus of mIL2 fuse, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mF8V. H -mL19V L -mL19V H -mF8V LThe mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.36) can be simply referred to as mIL12bIL12aDiaF8L19IL2GMCSF protein.

[0109] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mL19V. H N-terminal fusion, mL19V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L C-terminal and mNHS76V H N-terminal fusion, mNHS76V H C-terminus and mL19V L N-terminal fusion, mL19V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of GMCSF, thereby forming mIL12b-mIL12a-mL19V. H -mNHS76V L -mNHS76V H -mL19V L The -mIL2-mGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.37) can be simply referred to as mIL12bIL12aDiaL19NHS76IL2GMCSF protein.

[0110] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, and the C-terminus of mIL12a is fused to mL19V. H N-terminal fusion, mL19V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-terminus and mL19V L N-terminal fusion, mL19V L The C-terminus of mIL12 fuses with the N-terminus of mIL2, and the C-terminus of mIL2 fuses with the N-terminus of mGMCSF, thereby forming mIL12b-mIL12a-mL19V. H -mF8V L -mF8V H -mL19V LThe mIL2-mGMCSF protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.38) can be simply referred to as mIL12bIL12aDiaL19F8IL2GMCSF protein.

[0111] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, the C-terminus of mIL12a is fused to the N-terminus of mIL2, and the C-terminus of mIL2 is fused to mNHS76V. H N-terminal fusion, mNHS76V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-terminal and mNHS76V L N-terminal fusion, mNHS76V L The C-terminus and N-terminus of mGMCSF fuse to form mIL12b-mIL12a-mIL2-mNHS76V. H -mF8V L -mF8V H -mNHS76V L -mGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.39), and may be simply referred to as mIL12bIL12aIL2DiaNHS76F8GMCSF protein.

[0112] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, the C-terminus of mIL12a is fused to the N-terminus of mIL2, and the C-terminus of mIL2 is fused to mF8V. H N-terminal integration, mF8V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-end and mF8V L N-terminal integration, mF8V L The C-terminus and the N-terminus of mGMCSF are fused to form mIL12b-mIL12a-mIL2-mF8V. H -mF8V L -mF8V H -mF8V L -mGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.40), and may be simply referred to as mIL12bIL12aIL2DiaF8GMCSF protein.

[0113] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, the C-terminus of mIL12a is fused to the N-terminus of mIL2, the C-terminus of mIL2 is fused to the N-terminus of mGMCSF, and the C-terminus of mGMCSF is fused to mNHS76V. H N-terminal fusion, mNHS76V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-terminal and mNHS76V L The N-terminus is fused to form mIL12b-mIL12a-mIL2-mGMCSF-mNHS76V. H -mF8V L -mF8V H -mNHS76V L The protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.41) may be abbreviated as mIL12bIL12aIL2GMCSFDiaNHS76F8 protein.

[0114] For example, the structure of the single-chain protein could be such that the C-terminus of mIL12b is fused to the N-terminus of mIL12a, the C-terminus of mIL12a is fused to the N-terminus of mIL2, the C-terminus of mIL2 is fused to the N-terminus of mGMCSF, and the C-terminus of mGMCSF is fused to mF8V. H N-terminal integration, mF8V H C-end and mF8V L N-terminal integration, mF8V L C-end and mF8V H N-terminal integration, mF8V H C-end and mF8V L The N-terminus is fused to form mIL12b-mIL12a-mIL2-mGMCSF-mF8V H -mF8V L -mF8V H -mF8V L The protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.42) may be abbreviated as mIL12bIL12aIL2GMCSFDiaF8 protein.

[0115] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hL19V. H N-terminal fusion, hL19VH C-end and hL19V L N-terminal fusion, hL19V L C-end and hL19V H N-terminal fusion, hL19V H C-end and hL19V L N-terminal fusion, hL19V L The C-terminus of hIL12 fuses with the N-terminus of hIL2, and the C-terminus of hIL2 fuses with the N-terminus of hGMCSF, thus forming hIL12b-hIL12a-hL19V. H -hL19V L -hL19V H -hL19V L -hIL2-hGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.43), and may be simply referred to as hIL12bIL12aDiaL19IL2GMCSF protein.

[0116] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hNHS76V. H N-terminal fusion, hNHS76V H C-end and hNHS76V L N-terminal fusion, hNHS76V L C-end and hNHS76V H N-terminal fusion, hNHS76V H C-end and hNHS76V L N-terminal fusion, hNHS76V L The C-terminus of hIL12 fuses with the N-terminus of hIL2, and the C-terminus of hIL2 fuses with the N-terminus of hGMCSF, thus forming hIL12b-hIL12a-hNHS76V. H -hNHS76V L -hNHS76V H -hNHS76V L -hIL2-hGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.44), and may be simply referred to as hIL12bIL12aDiaNHS76IL2GMCSF protein.

[0117] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hNHS76V. H N-terminal fusion, hNHS76V H C-end and hF8V L N-terminal integration, hF8V LC-end and hF8V H N-terminal integration, hF8V H C-end and hNHS76V L N-terminal fusion, hNHS76V L The C-terminus of hIL12 fuses with the N-terminus of hIL2, and the C-terminus of hIL2 fuses with the N-terminus of hGMCSF, thus forming hIL12b-hIL12a-hNHS76V. H -hF8V L -hF8V H -hNHS76V L -hIL2-hGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.45), and may be simply referred to as hIL12bIL12aDiaNHS76F8IL2GMCSF protein.

[0118] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, the C-terminus of hIL12a is fused to the N-terminus of hIL2, and the C-terminus of hIL2 is fused to hNHS76V. H N-terminal fusion, hNHS76V H C-end and hF8V L N-terminal integration, hF8V L C-end and hF8V H N-terminal integration, hF8V H C-end and hNHS76V L N-terminal fusion, hNHS76V L The C-terminus of hIL12b fuses with the N-terminus of hGMCSF to form hIL12b-hIL12a-hIL2-hNHS76V. H -hF8V L -hF8V H -hNHS76V L -hGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.46), and may be simply referred to as hIL12bIL12aIL2DiaNHS76F8GMCSF protein.

[0119] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, the C-terminus of hIL12a is fused to the N-terminus of hIL2, and the C-terminus of hIL2 is fused to hF8V. H N-terminal integration, hF8V H C-end and hF8V L N-terminal integration, hF8V L C-end and hF8V H N-terminal integration, hF8V HC-end and hF8V L N-terminal integration, hF8V L The C-terminus and N-terminus of hGMCSF fuse to form hIL12b-hIL12a-hIL2-hF8V. H -hF8V L -hF8V H -hF8V L -hGMCSF protein molecule (its amino acid sequence may be as shown in SEQ ID NO.47) and may be simply referred to as hIL12bIL12aIL2DiaF8GMCSF protein.

[0120] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hL19V. H N-terminal fusion, hL19V H C-end and hL19V L N-terminal fusion, hL19V L C-end and hL19V H N-terminal fusion, hL19V H C-end and hL19V L N-terminal fusion, hL19V L The C-terminus of hIL12b fuses with the N-terminus of hIL-12a, and the C-terminus of hIL-12a fuses with the N-terminus of hIL-12b, thus forming hIL12b-hIL12a-hL19V. H -hL19V L -hL19V H -hL19V L The -hGMCSF-hIL2 protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.48) can be simply referred to as hIL12bIL12aDiaL19GMCSFIL2 protein.

[0121] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hNHS76V. H N-terminal fusion, hNHS76V H C-end and hNHS76V L N-terminal fusion, hNHS76V L C-end and hNHS76V H N-terminal fusion, hNHS76V H C-end and hNHS76V L N-terminal fusion, hNHS76V LThe C-terminus of hIL12b fuses with the N-terminus of hGMCSF, and the C-terminus of hGMCSF fuses with the N-terminus of hIL2, thus forming hIL12b-hIL12a-hNHS76V. H -hNHS76V L -hNHS76V H -hNHS76V L The -hGMCSF-hIL2 protein molecule (whose amino acid sequence may be as shown in SEQ ID NO.49) can be simply referred to as hIL12bIL12aDiaNHS76GMCSFIL2 protein.

[0122] For example, the structure of the single-chain protein could be such that the C-terminus of hIL12b is fused to the N-terminus of hIL12a, and the C-terminus of hIL12a is fused to hNHS76V. H N-terminal fusion, hNHS76V H C-end and hF8V L N-terminal integration, hF8V L C-end and hF8V H N-terminal integration, hF8V H C-end and hNHS76V L N-terminal fusion, hNHS76V L The C-terminus of hIL12b fuses with the N-terminus of hGMCSF, and the C-terminus of hGMCSF fuses with the N-terminus of hIL2, thus forming hIL12b-hIL12a-hNHS76V. H -hF8V L -hF8V H -hNHS76V L -hGMCSF-hIL2 protein molecule (its amino acid sequence may be as shown in SEQ ID NO.50), and may be simply referred to as hIL12bIL12aDiaNHS76F8GMCSFIL2 protein.

[0123] Furthermore, in some embodiments, the single-chain protein may also be mIL12bIL12aIL2DiaNHS76F8GMCSF-Thr (its amino acid sequence may be as shown in SEQ ID NO.51), which is structurally similar to the mIL12bIL12aIL2DiaNHS76F8GMCSF protein, differing only in the linker. The linker of mIL12bIL12aIL2DiaNHS76F8GMCSF-Thr contains a thrombin cleavage site.

[0124] Furthermore, in some embodiments, the single-chain protein may also be hIL12bIL12aIL2DiaNHS76F8GMCS F-Thr (its amino acid sequence may be as shown in SEQ ID NO.52), which is structurally similar to the hIL12bIL12aIL2DiaNHS76F8GM CSF protein, differing only in the linker. The linker of hIL12bIL12aIL2DiaNHS76F8GMCSF-Thr contains a thrombin cleavage site.

[0125] In this application, the protein may also be a dimer composed of a first polypeptide chain and a second polypeptide chain, wherein the first polypeptide chain is different from the second polypeptide chain.

[0126] In this application, the first polypeptide chain may contain IL12a, and the second polypeptide chain may contain IL12b.

[0127] In this application, the IL2 or its functional fragment may be located in the first polypeptide chain or the second polypeptide chain, and the GMCSF or its functional fragment may be located in the first polypeptide chain or the second polypeptide chain.

[0128] In this application, the first polypeptide chain may sequentially include IL12a or a functional fragment thereof and IL2 or a functional fragment thereof from the N-terminus to the C-terminus; or, the first polypeptide chain may sequentially include IL2 or a functional fragment thereof and IL12a or a functional fragment thereof from the N-terminus to the C-terminus; or, the first polypeptide chain may sequentially include IL12a or a functional fragment thereof and GMCSF or a functional fragment thereof from the N-terminus to the C-terminus.

[0129] In this application, the second polypeptide chain may sequentially include IL12b or a functional fragment thereof and GMCSF or a functional fragment thereof from the N-terminus to the C-terminus; or, the second polypeptide chain may sequentially include GMCSF or a functional fragment thereof and IL12b or a functional fragment thereof from the N-terminus to the C-terminus; or, the second polypeptide chain may sequentially include IL12b or a functional fragment thereof and IL2 or a functional fragment thereof from the N-terminus to the C-terminus.

[0130] In some embodiments, in the pharmaceutical composition, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 53 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 57; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 54 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 57; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 53 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 58; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 54 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 58; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 55 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 59; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 56 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 60.

[0131] In this application, the protein may be a dimer composed of a first polypeptide chain and a second polypeptide chain.

[0132] For example, in the dimer, the C-terminus of mIL12a and the N-terminus of mIL2 can fuse to form the first polypeptide chain mIL12a-mIL2 (sequence shown in SEQ ID NO. 53), and the C-terminus of mIL12b and the N-terminus of mGMCSF can fuse to form the second polypeptide chain mIL12b-mGMCSF (sequence shown in SEQ ID NO. 57), thus forming a mIL12a-mIL2-mIL12b-mGMCSF protein heterodimer, simply referred to as the mIL12aIL2-IL12bGMCSF heterodimer.

[0133] For example, in the dimer, the C-terminus of mIL2 and the N-terminus of mIL12a can fuse to form the first polypeptide chain mIL2-mIL12a (sequence shown in SEQ ID NO. 54), and the C-terminus of mIL12b and the N-terminus of mGMCSF can fuse to form the second polypeptide chain mIL12b-mGMCSF (sequence shown in SEQ ID NO. 57), thus forming the mIL2-mIL12a-mIL12b-mGMCSF protein heterodimer, abbreviated as mIL2IL12a-IL12bGMCSF heterodimer.

[0134] For example, in the dimer, the C-terminus of mIL12a and the N-terminus of mIL2 can fuse to form the first polypeptide chain mIL12a-mIL2 (sequence shown in SEQ ID NO. 53), and the C-terminus of mGMCSF and the N-terminus of mIL12b can fuse to form the second polypeptide chain mGMCSF-mIL12b (sequence shown in SEQ ID NO. 58), thus forming a protein heterodimer of mIL12a-mIL2-mGMCSF-mIL12b, simply referred to as the mIL12aIL2-GMCSFIL12b heterodimer.

[0135] For example, in the dimer, the C-terminus of mIL2 and the N-terminus of mIL12a can fuse to form the first polypeptide chain mIL2-mIL12a (sequence shown in SEQ ID NO. 54), and the C-terminus of mGMCSF and the N-terminus of mIL12b can fuse to form the second polypeptide chain mGMCSF-mIL12b (sequence shown in SEQ ID NO. 58), thus forming a protein heterodimer of mIL2-mIL12a-mGMCSF-rmIL12b, simply referred to as the mIL2IL12a-GMCSFIL12b heterodimer.

[0136] For example, in the dimer, the C-terminus of mIL12a and the N-terminus of mGMCSF can fuse to form the first polypeptide chain mIL12a-mGMCSF (sequence shown in SEQ ID NO. 55), and the C-terminus of mIL12b and the N-terminus of mIL2 can fuse to form the second polypeptide chain mIL12b-mIL2 (sequence shown in SEQ ID NO. 59), thus forming a protein heterodimer of mIL12a-mGMCSF-mIL12b-mIL2, simply referred to as the mIL12aGMCSF-IL12bIL2 heterodimer.

[0137] For example, in the dimer, the C-terminus of hIL12a and the N-terminus of hIL2 can fuse to form the first polypeptide chain hIL12a-hIL2 (sequence shown in SEQ ID NO. 56), and the C-terminus of hIL12b and the N-terminus of hGMCSF can fuse to form the second polypeptide chain hIL12b-hGMCSF (sequence shown in SEQ ID NO. 60), thus forming the hIL12a-hIL2-hIL12b-hGMCSF protein heterodimer, abbreviated as hIL12aIL2-IL12bGMCSF heterodimer.

[0138] In this application, the IL2 or its functional fragment may be located in the first polypeptide chain or the second polypeptide chain, the GMCSF or its functional fragment may be located in the first polypeptide chain or the second polypeptide chain, and one or more of the target portions may each be independently located in the first polypeptide chain or the second polypeptide chain.

[0139] In this application, the first polypeptide chain may sequentially include the targeting portion, the IL12a or its functional fragment, the IL2 or its functional fragment, and the GMCSF or its functional fragment from the N-terminus to the C-terminus; or, the first polypeptide chain may sequentially include the IL2 or its functional fragment, the IL12a or its functional fragment, and the GMCSF or its functional fragment from the N-terminus to the C-terminus.

[0140] In this application, the second polypeptide chain may sequentially include the IL12b or a functional fragment thereof and the targeting portion from the N-terminus to the C-terminus.

[0141] In some embodiments, in the pharmaceutical composition, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 61; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 62; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 66 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 63; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 67 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 61; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 68 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 62; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 69 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 63; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 70 and the second polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 64; or, the first polypeptide chain may comprise the amino acid sequence shown in SEQ ID NO. 64. The first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 71 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 64; or, the first polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 72 and the second polypeptide chain may contain the amino acid sequence shown in SEQ ID NO. 65.

[0142] For example, in the dimer, the C-terminus of mIL12b can interact with L19V. H N-terminal fusion, L19V H The C-end can be connected to L19V L The N-terminus of mIL2 can fuse with the N-terminus of mIL12a to form a second polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 61), and the C-terminus of mIL2 can fuse with the N-terminus of mIL12a, while the C-terminus of mIL12a can fuse with the N-terminus of mGMCSF to form a first polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 66), thereby forming mIL12b-L19V. H -L19V L -mIL2-mIL12a-mGMCSF protein heterodimer, abbreviated as mIL12bscL19-IL2IL12aGMCSF heterodimer.

[0143] For example, in the dimer, the C-terminus of mIL12b can interact with F8V. H N-terminal integration, F8V H The C-end can be used with F8V L The N-terminus of mIL2 can fuse with the N-terminus of mIL12a to form a second polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 62), and the C-terminus of mIL2 can fuse with the N-terminus of mIL12a, while the C-terminus of mIL12a can fuse with the N-terminus of mGMCSF to form a first polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 66), thereby forming mIL12b-F8V. H -F8V L -mIL2-mIL12a-mGMCSF protein heterodimer, abbreviated as mIL12bscF8-IL2IL12aGMCSF heterodimer.

[0144] For example, in the dimer, the C-terminus of mIL12b can interact with NHS76V. H N-terminal fusion, NHS76V H The C-end can be connected to NHS76V L The N-terminus of mIL2 can fuse with the N-terminus of mIL12a to form a second polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 63), and the C-terminus of mIL2 can fuse with the N-terminus of mIL12a, while the C-terminus of mIL12a can fuse with the N-terminus of mGMCSF to form a first polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 66), thereby forming mIL12b-NHS76V. H -NHS76V L -mIL2-mIL12a-mGMCSF protein heterodimer, abbreviated as mIL12bscNHS76-IL2IL12aGMCSF heterodimer.

[0145] For example, in the dimer, the C-terminus of mIL12b can interact with L19V. H N-terminal fusion, L19V H The C-end can be connected to L19V L The N-terminus is fused to form a second polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 61), and L19V H The C-end can be connected to L19V L N-terminal fusion, L19V L The C-terminus of mIL12 can fuse with the N-terminus of mIL12a, the C-terminus of mIL12a can fuse with the N-terminus of mIL2, and the C-terminus of mIL2 can fuse with the N-terminus of mGMCSF to form the first polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 67), thereby forming mIL12b-L19V. H -L19V L-L19V H -L19V L -mIL12a-mIL2-mGMCSF protein heterodimer, abbreviated as mIL12bscL19-scL19IL12aIL2GMCSF heterodimer.

[0146] For example, in the dimer, the C-terminus of mIL12b can interact with F8V. H N-terminal integration, F8V H The C-end can be used with F8V L The N-terminus is fused to form a second polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 62), and F8V H The C-end can be used with F8V L N-terminal integration, F8V L The C-terminus of mIL12 can fuse with the N-terminus of mIL12a, the C-terminus of mIL12a can fuse with the N-terminus of mIL2, and the C-terminus of mIL2 can fuse with the N-terminus of mGMCSF to form the first polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 68), thereby forming mIL12b-F8V. H -F8V L -F8V H -F8V L -mIL12a-mIL2-mGMCSF protein heterodimer, abbreviated as mIL12bscF8-scF8IL12aIL2GMCSF heterodimer.

[0147] For example, in the dimer, the C-terminus of mIL12b can interact with NHS76V. H N-terminal fusion, NHS76V H The C-end can be connected to NHS76V L The N-terminus is fused to form a second polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 63), and NHS76V H The C-end can be connected to NHS76V L N-terminal fusion, NHS76V L The C-terminus of mIL12b can fuse with the N-terminus of mIL12a, the C-terminus of mIL12a can fuse with the N-terminus of mIL2, and the C-terminus of mIL2 can fuse with the N-terminus of GMCSF to form the first polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 69), thereby forming mIL12b-NHS76V. H -NHS76V L -NHS76V H -NHS76V L-mIL12a-mIL2-mGMCSF protein heterodimer, abbreviated as mIL12bscNHS76-scNHS76IL12aIL2GMCSF heterodimer.

[0148] For example, in the dimer, the C-terminus of hIL12b can interact with F8V. H N-terminal integration, F8V H The C-end can be used with F8V L The N-terminus of hIL2 can be fused with the N-terminus of hIL12a to form a second polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 64), and the C-terminus of hIL2 can be fused with the N-terminus of hIL12a, and the C-terminus of hIL12a can be fused with the N-terminus of hGMCSF to form a first polypeptide chain (the amino acid sequence of which may be as shown in SEQ ID NO. 70), thereby forming hIL12b-F8V. H -F8V L -hIL2-hIL12a-hGMCSF protein heterodimer, abbreviated as hIL12bscF8-IL2IL12aGMCSF heterodimer.

[0149] For example, in the dimer, the C-terminus of hIL12b can interact with F8V. H N-terminal integration, F8V H The C-end can be used with F8V L The N-terminus is fused to form a second polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 64), and F8V H The C-end can be used with F8V L N-terminal integration, F8V L The C-terminus of hIL12 can fuse with the N-terminus of hIL12a, the C-terminus of hIL12a can fuse with the N-terminus of hIL2, and the C-terminus of hIL2 can fuse with the N-terminus of hGMCSF to form the first polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO.71), thereby forming hIL12b-F8V. H -F8V L -F8V H -F8V L -hIL12a-hIL2-hGMCSF protein heterodimer, abbreviated as hIL12bscF8-scF8IL12aIL2GMCSF heterodimer.

[0150] For example, in the dimer, the C-terminus of hIL12b can interact with NHS76V. H N-terminal fusion, NHS76V H The C-end can be connected to NHS76V LThe N-terminus is fused to form a second polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 65), and NHS76V H The C-end can be connected to NHS76V L N-terminal fusion, NHS76V L The C-terminus of hIL12b can fuse with the N-terminus of hIL12a, the C-terminus of hIL12a can fuse with the N-terminus of hIL2, and the C-terminus of hIL2 can fuse with the N-terminus of hGMCSF to form the first polypeptide chain (whose amino acid sequence may be as shown in SEQ ID NO. 72), thereby forming hIL12b-NHS76V. H -NHS76V L -NHS76V H -NHS76V L -hIL12a-hIL2-hGMCSF protein heterodimer, abbreviated as hIL12bscNHS76-scNHS76IL12aIL2GMCSF heterodimer.

[0151] On the other hand, this application provides a kit comprising the pharmaceutical composition described in this application. The kit may contain a box or container holding the kit components. The box or container is labeled or labeled with a Food and Drug Administration-approved treatment regimen. The box or container holds the components of the pharmaceutical composition of this application; for example, the components may be contained in a plastic, polyethylene, polypropylene, ethylene, or propylene container. The container may be a capped tube or bottle. Furthermore, the kit may also contain instructions for use to administer the pharmaceutical composition described in this application.

[0152] use

[0153] On the other hand, this application also provides the use of the pharmaceutical composition or the kit described herein in the preparation of a medicament that can be used to treat tumors.

[0154] In this application, the pharmaceutical composition or the kit described herein can be used to treat tumors.

[0155] On the other hand, this application also provides a method for treating tumors, which includes administering the pharmaceutical composition or the kit to a subject in need.

[0156] In this application, the tumor can include both solid tumors and non-solid tumors. For example, the tumor can include melanoma. For example, the tumor can include breast cancer. For example, the tumor can include lung cancer.

[0157] In this application, the application may include administering the protein first, followed by administering the immune checkpoint inhibitor. For example, the application may include administering the single-chain protein described in this application first, followed by administering the immune checkpoint inhibitor described in this application. As another example, the application may include administering the dimer described in this application first, followed by administering the immune checkpoint inhibitor described in this application.

[0158] In this application, the administration may include intratumoral injection. For example, injecting the pharmaceutical composition described in this application into the tumor. In some embodiments, the administration method may also be oral administration, intravenous administration, intramuscular administration, in situ administration at the tumor site, inhalation, rectal administration, vaginal administration, transdermal administration, or administration through a subcutaneous reservoir. In some embodiments, the administration may also include intravenous injection or subcutaneous injection.

[0159] In some embodiments, the pharmaceutical compositions described herein can be formulated for oral administration, intravenous administration, intramuscular administration, in situ administration at the tumor site, inhalation, rectal administration, vaginal administration, transdermal administration, or administration via a subcutaneous reservoir.

[0160] In some embodiments, the pharmaceutical composition described in this application may further comprise a pharmaceutically acceptable carrier. For example, the pharmaceutically acceptable carrier may include buffers, antioxidants, preservatives, low molecular weight peptides, proteins, hydrophilic polymers, amino acids, sugars, chelating agents, counterions, metal complexes, and / or nonionic surfactants. For example, the pharmaceutically acceptable carrier may include excipients, such as those selected from the group consisting of starch, dextrin, sucrose, lactose, magnesium stearate, calcium sulfate, carboxymethyl ether, talc, calcium alginate gel, chitosan, and nanospheres. For example, the pharmaceutically acceptable carrier may also be selected from the group consisting of pH adjusters, osmotic pressure regulators, solubilizers, and antibacterial agents.

[0161] The embodiments described below are not intended to be limited by any theory, but are merely for illustrating the pharmaceutical compositions and uses of this application, and are not intended to limit the scope of the invention.

[0162] Example

[0163] Reagents: DMEM medium, 1640 medium, and fetal bovine serum were purchased from Lifetechnologies; cell culture flasks and plates were purchased from Corning; doxycycline was purchased from Shanghai Sangon Biotech Co., Ltd.; puromycin and blasticidin were purchased from Chemicon; restriction endonucleases were purchased from Takara and NEB; ligases were purchased from NEB; DNA polymerase was purchased from Takara; plasmid extraction kits and gel extraction kits were purchased from Omega Biotech; primer synthesis was performed by Shanghai Sangon Biotech Co., Ltd.; gene synthesis and sequencing were performed by Lifetechnologies; flow cytometry antibodies were purchased from Ebioscience; PD1 blocking antibodies were purchased from BioXcell; and protein magnetic bead purification kits were purchased from Beaver Biotechnology.

[0164] Example 1. mIL12aIL2-IL12bGMCSF heterodimer induces PD1 expression in T cells.

[0165] 1.1 Construction of the first expression vector

[0166] The synthesized rtTA DNA sequence, with BamHI and EcoRI sites at both ends, was ligated into the vector pUC57. The vector was digested using the following enzyme digestion system: 6 μg plasmid, 4 μl digestion buffer, 1 μl BamHI, 1 μl EcoRI, and water to a total volume of 40 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 4.4 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel, and the rtTA fragment was recovered after electrophoresis for later use.

[0167] Digest the pLentis-CMV-IRES-Bsd vector in an EP tube using the following digestion system: 2 μg plasmid, 3 μl digestion buffer, 1 μl BamHI, 1 μl EcoRI, and water to a total volume of 30 μl. Incubate at 37°C for 12 hours. Remove the EP tube, add 3.3 μl of 10× loading buffer, and perform electrophoresis on a 1% agarose gel. After electrophoresis, recover the pLentis-CMV-IRES-Bsd vector fragment for later use.

[0168] The pLentis-CMV-IRES-Bsd and rtTA were ligated using the following system: 2 μl pLentis-CMV-IRES-Bsd, 2 μl rtTA, 1 μl ligase buffer, 0.5 μl T4 DNA ligase, and 4.5 μl water. The mixture was incubated at room temperature for 4 hours. The ligation system was then used to transform competent *E. coli* cells. The next day, colonies were picked from the transformed plates and cultured overnight in LB medium at 37°C using a shaker. Plasmids were extracted from the cultured bacteria using a plasmid extraction kit. Enzyme digestion was used to confirm successful ligation into the vector. The correct vector was then sequenced, confirming successful construction of the first expression vector, pLentis-CMV-rtTA-IRES-Bsd.

[0169] 1.2 Obtaining cells containing the first expression vector

[0170] First, the virus for the first expression vector was prepared using the following method:

[0171] 1. Digest and culture 293FT cells, count them, and then divide them into 3×10⁻⁶ cells. 6 Cells / wells were seeded into 10cm culture dishes with a culture medium volume of 10ml.

[0172] 2. On the second evening, observe the cell condition. If the cell condition is suitable and good, perform transfection. Add chloroquine to the culture plate to a final concentration of 25 μM. Take a test tube and add sterile water and the following plasmids (pMD2.G 5 μg + pSPAX2 15 μg + pLentis-CMV-rtTA-IRES-Bsd 20 μg), with a total volume of 1045 μl. Then add 155 μl of 2M CaCl2 and mix well. Finally, add 1200 μl of 2×HBS while vortexing. After the addition is complete, quickly add the mixture to the cell culture wells and gently shake to mix.

[0173] 3. On the morning of the third day, observe the cell status and replace the culture medium with 10 ml of fresh DMEM medium.

[0174] 4. On the morning of the fifth day, observe the cell status and collect the supernatant from the culture dish. Filter it through a 0.45μm filter and then place it in a high-speed centrifuge tube. Centrifuge at 50,000g for 2 hours. Carefully discard the supernatant and blot the liquid with absorbent paper as much as possible. Then resuspend the precipitate with 500μl HBSS. After dissolving for 2 hours, aliquot it into small tubes and store at -70℃.

[0175] Then, the tumor cells were transfected using the first expression vector virus, as follows:

[0176] Digested cultured mouse melanoma cells B16, at 10 5One cell per well was seeded into a 6-well plate with a culture volume of 1 ml. After 24 hours, 10 μl of the first expression vector virus was added, and the cells were cultured for another 24 hours. The supernatant was discarded, and fresh culture medium was added. Once the cells reached confluence, they were transferred to a culture flask, and blasticidin was added at a concentration suitable for this cell type. The cells were cultured again, with the culture medium changed every two days, maintaining a blasticidin concentration of 8 μg / ml. After one week of selection, the surviving cells were identified as cells that stably expressed the regulatory protein and were named B16(rtTA).

[0177] 1.3 Construction of a second expression vector encoding mIL12aIL2-IL12bGMCSF heterodimers

[0178] The mIL12aIL2-IL12bGMCSF heterodimer gene was synthesized, with BamHI and EcoRI restriction sites at both ends. The gene was then digested with BamHI and EcoRI using the following digestion system: 5 μg mIL12aIL2-IL12bGMCSF heterodimer plasmid, 4 μl digestion buffer, 1 μl BamHI, 1 μl EcoRI, and water to a total volume of 40 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 4.4 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel. The mIL12aIL2-IL12bGMCSF gene fragment was recovered after electrophoresis and is ready for use.

[0179] The amino acid sequence of the first polypeptide chain of the mIL12aIL2-IL12bGMCSF heterodimer is shown in SEQ ID NO.53, and the amino acid sequence of the second polypeptide chain is shown in SEQ ID NO.57; the nucleotide sequence encoding the mIL12aIL2-IL12bGMCSF is shown in SEQ ID NO.73.

[0180] The expression vector pLentis-PTRE-MCS-PGK-PURO was digested with the following enzyme digestion system: 2 μg pLentis-PTRE-MCS-PGK-PURO plasmid, 3 μl digestion buffer, 1 μl BamHI, 1 μl EcoRI, and water was added to a total volume of 30 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 3.3 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel. The pLentis-PTRE-MCS-PGK-PURO vector fragment was recovered after electrophoresis and is ready for use.

[0181] The pLentis-PTRE-MCS-PGK-PURO and IL2 were ligated using the following ligation system: 2 μl pLentis-PTRE-MCS-PGK-PURO, 2 μl mIL12aIL2-IL12bGMCSF, 1 μl ligase buffer, 0.5 μl T4 DNA ligase, and 4.5 μl water. The mixture was incubated at room temperature for 4 hours. The ligation system was then used to transform competent *E. coli* cells. The next day, colonies were picked from the transformed plates and cultured overnight in LB medium at 37°C using a shaker. Plasmids were extracted from the cultured bacteria using a plasmid extraction kit. Enzyme digestion was used to confirm successful ligation into the vector. The correct vector was then sequenced, confirming successful construction of the second expression vector pLentis-PTRE-mIL12aIL2-IL12bGMCSF-PGK-PURO.

[0182] 1.4 Preparation of cells expressing mIL12a, IL2-IL12b, GMCSF heterodimers

[0183] The virus for preparing the mIL12aIL2-IL12bGMCSF heterodimer expression vector was prepared using the same method as the virus for the first expression vector. Cultured B16(rtTA) tumor cells were digested and added at a ratio of 10... 5 One cell per well was seeded into a 6-well plate with a culture volume of 1 ml. After 24 hours, 10 μl of the regulatory expression vector virus (i.e., the virus of the mIL12aIL2-IL12bGMCSF heterodimer expression vector) was added. After culturing in an incubator for another 24 hours, the supernatant was discarded and replaced with fresh culture medium. After the cells reached confluence, they were transferred to a culture flask and puromycin was added at a final concentration of 3 μg / ml. After culturing for another three days, the surviving cells were the cells that could regulate the expression of mIL12aIL2-IL12bGMCSF. This cell was named B16(rtTA)-mIL12aIL2-IL12bGMCSF.

[0184] 1.5 Analysis of the effect of inducing expression of mIL12a, IL2-IL12b, and GMCSF heterodimers on PD1 expression in T cells

[0185] B16(rtTA)-mIL12aIL2-IL12b GMCSF cells in logarithmic growth phase were digested and diluted with HBSS to 2×10⁻⁶. 6Ten 8-10 week old female C57BL / 6 mice were injected with 50 μl / mouse using a 1 ml syringe into the right dorsal side of the tumor. After tumor growth, the mice were fed water containing 2 g / L doxycycline. Cells from the spleen and tumor tissue were isolated from mice on days 0 and 3 after doxycycline administration. Red blood cells were lysed with erythrocyte lysis buffer, filtered through a sieve, and a single-cell suspension was obtained. This suspension was then stained with CD45, CD3, CD4, CD8, and PD1 antibodies. After staining, the cells were washed twice with PBS and analyzed by flow cytometry to determine changes in PD1 expression in T cells. Results are as follows. Figure 1 As shown, where Figure 1 A, Figure 1 B and Figure 1 C shows the expression of PD1 in mouse spleen CD4T cells, spleen CD8T cells, and tumor-infiltrating CD3T cells, respectively. It can be seen that the expression of PD1 in mouse spleen CD4T cells, CD8T cells, and tumor-infiltrating CD3T cells is significantly increased.

[0186] Example 2. Expression of mIL12bIL12aIL2GMCSF single-chain proteins

[0187] 2.1 Constructing the expression vector

[0188] The single-stranded protein molecule mIL12bIL12aIL2GMCSF was synthesized by adding a 6*His terminus to facilitate purification. The corresponding DNA sequence was synthesized, with BamHI and XhoI restriction sites at the beginning and end, respectively. The resulting plasmid containing the target gene was digested using the following regimen: 5 μg plasmid, 4 μl digestion buffer, 1 μl BamHI, and 1 μl XhoI, with water added to a total volume of 40 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 4.4 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel. The mIL12bIL12aIL2GMCSF protein gene fragment was recovered after electrophoresis and is ready for use.

[0189] The amino acid sequence of the mIL12bIL12aIL2GMCSF single-chain protein is shown in SEQ ID NO.27, and the nucleotide sequence encoding the mIL12bIL12aIL2GMCSF is shown in SEQ ID NO.74.

[0190] Digest the pLentis-CMV-MCS-IRES-PURO vector in an EP tube using the following formulation: 2 μg pLentis-CMV-MCS-IRES-PURO vector plasmid, 3 μl digestion buffer, 1 μl BamHI, and 1 μl XhoI. Add water to a total volume of 30 μl and incubate at 37°C for 12 hours. Remove the EP tube, add 3.3 μl of 10× loading buffer, and perform electrophoresis on a 1% agarose gel. After electrophoresis, recover the pLentis-CMV-MCS-IRES-PURO vector fragment for later use.

[0191] The expression vector pLentis-CMV-MCS-IRES-PURO was ligated using the following system: 2 μl pLentis-CMV-MCS-IRES-PURO vector fragment, 2 μl mIL12bIL12aIL2GMCSF gene fragment, 1 μl ligase buffer, 0.5 μl T4 DNA ligase, and 4.5 μl water. The mixture was incubated at room temperature for 4 hours. The ligation system was then used to transform competent *E. coli* cells. The next day, colonies were picked from the transformed plates and cultured overnight in LB medium at 37°C using a shaker. Plasmids were extracted from the cultured bacteria using a plasmid extraction kit. Enzyme digestion was used to confirm successful ligation into the vector. Sequencing of the correct vector confirmed successful construction. The expression vector pLentis-CMV-mIL12bIL12aIL2GMCSF-IRES-PURO was obtained.

[0192] 2.2 Preparation of expression virus

[0193] 1) Digest and culture 293FT cells, count them, and then divide them into 3×10⁻⁶ cells. 6 One cell per well is seeded into a 10cm culture dish, with a culture medium volume of 10ml.

[0194] 2) On the second evening, observe the cell condition. If the cells are in good condition, proceed with transfection. Add chloroquine to the culture plate to a final concentration of 25 μM. Take a test tube and add sterile water and the following plasmids (pMD2.G 6 μg + pSPAX2 15 μg + expression vector obtained in Example 2.1 above 20 μg), for a total volume of 1045 μl. Then add 155 μl of 2M CaCl2 and mix well. Finally, add 1200 μl of 2×HBS while vortexing. After the addition is complete, quickly add the mixture to the cell culture wells and gently shake to mix.

[0195] 3) On the morning of the third day, observe the cell status and replace the culture medium with 10 ml of fresh DMEM medium.

[0196] 4) On the morning of the fifth day, observe the cell status and collect the supernatant from the culture dish. Filter it with a 0.45μm filter and then place it in a high-speed centrifuge tube. Centrifuge at 50,000g for 2 hours. Carefully discard the supernatant and blot the liquid with absorbent paper as much as possible. Then resuspend the precipitate with 200μl HBSS. After dissolving for 2 hours, aliquot it into small tubes and store at -70℃.

[0197] 2.3 Preparation of expression cells

[0198] Digested cultured 293A cells, at 10 5 Cells were seeded per well into 6-well plates, with a culture volume of 1 ml. After 24 hours, 10 μl of virus expressing the target gene (i.e., the virus obtained in Example 2.2) was added, and the cells were cultured for another 24 hours. The supernatant was then discarded, and fresh culture medium was added for further culture. Once the cells reached confluence, they were transferred to culture flasks, and puromycin was added to a final concentration of 3 μg / ml. The cells were cultured again, with the culture medium changed every two days while maintaining the puromycin concentration. After one week of selection, the surviving cells were identified as cells stably expressing the protein and named 293A-mIL12bIL12aIL2GMCSF.

[0199] 2.4 Protein Expression and Purification

[0200] The constructed cells expressing mIL12bIL12aIL2GMCSF (293A-mIL12bIL12aIL2GMCSF) were passaged into 15cm culture dishes. After the cells reached confluence, the culture medium was replaced with 30ml CDM4HEK293, and the cells were cultured for another 5 days. The supernatant was then collected, filtered through a 0.45μm filter, and concentrated by ultrafiltration using a 50kDa AMICON ULTRA-15. The concentrated protein solution was purified using nickel chelate magnetic beads (purchased from Beaver Biotechnology Co., Ltd.) according to the instructions. The purified protein solution was then ultrafiltered using an AMICON ULTRA-0.5 ultrafiltration tube, and the buffer was replaced with PBS. Finally, the protein concentration was detected using an IL12p70 ELISA kit. The protein concentration was adjusted to 2μg / μl with PBS, aliquoted, and stored at -20℃.

[0201] Example 3. Expression of mIL12bIL12aIL2DiaNHS76F8GMCSF single-chain proteins

[0202] 3.1 Constructing an expression carrier

[0203] The single-chain protein molecule mIL12bIL12aIL2DiaNHS76F8GMCSF was synthesized by adding a 6*His terminus to its end for purification. The corresponding DNA sequence was synthesized, with BamHI and XhoI restriction sites at the beginning and end, respectively. The resulting plasmid containing the target gene was digested using the following regimen: 5 μg plasmid, 4 μl digestion buffer, 1 μl BamHI, and 1 μl XhoI, with water added to a total volume of 40 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 4.4 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel. The mIL12bIL12aIL2DiaNHS76F8GMCSF protein gene fragment was recovered after electrophoresis and is ready for use.

[0204] The amino acid sequence of the mIL12bIL12aIL2DiaNHS76F8GMCSF single-chain protein is shown in SEQ ID NO.39, and the nucleotide sequence encoding the mIL12bIL12aIL2DiaNHS76F8GMCSF is shown in SEQ ID NO.75.

[0205] Digest the pLentis-CMV-MCS-IRES-PURO vector in an EP tube using the following formulation: 2 μg pLentis-CMV-MCS-IRES-PURO vector plasmid, 3 μl digestion buffer, 1 μl BamHI, and 1 μl XhoI. Add water to a total volume of 30 μl and incubate at 37°C for 12 hours. Remove the EP tube, add 3.3 μl of 10× loading buffer, and perform electrophoresis on a 1% agarose gel. After electrophoresis, recover the pLentis-CMV-MCS-IRES-PURO vector fragment for later use.

[0206] The expression vector pLentis-CMV-MCS-IRES-PURO was ligated using the following system: 2 μl pLentis-CMV-MCS-IRES-PURO vector fragment, 2 μl mIL12bIL12aIL2DiaNHS76F8GMCSF gene fragment, 1 μl ligase buffer, 0.5 μl T4 DNA ligase, and 4.5 μl water. The ligation was incubated at room temperature for 4 hours. The ligation system was then used to transform competent *E. coli* cells. The next day, colonies were picked from the transformed plates and cultured overnight in LB medium at 37°C using a shaker. Plasmids were extracted from the cultured bacteria using a plasmid extraction kit. Enzyme digestion was used to confirm successful ligation into the vector. Sequencing of the correct vector confirmed successful construction. The expression vector pLentis-CMV-mIL12bIL12aIL2DiaNHS76F8GMCSF-IRES-PURO was obtained.

[0207] 3.2 Preparation of expression virus

[0208] 1) Digest and culture 293FT cells, count them, and then divide them into 3×10⁻⁶ cells. 6 One cell per well is seeded into a 10cm culture dish, with a culture medium volume of 10ml.

[0209] 2) On the second evening, observe the cell condition. If the cell condition is good, proceed with transfection. Add chloroquine to the culture plate to a final concentration of 25 μM. Take a test tube and add sterile water and the following plasmids (pMD2.G 6 μg + pSPAX2 15 μg + expression vector obtained in Example 3.1 above 20 μg), with a total volume of 1045 μl. Then add 155 μl of 2M CaCl2 and mix well. Finally, add 1200 μl of 2×HBS while vortexing. After the addition is complete, quickly add the mixture to the cell culture wells and gently shake to mix.

[0210] 3) On the morning of the third day, observe the cell status and replace the culture medium with 10 ml of fresh DMEM medium.

[0211] 4) On the morning of the fifth day, observe the cell status and collect the supernatant from the culture dish. Filter it with a 0.45μm filter and then place it in a high-speed centrifuge tube. Centrifuge at 50,000g for 2 hours. Carefully discard the supernatant and blot the liquid with absorbent paper as much as possible. Then resuspend the precipitate with 200μl HBSS. After dissolving for 2 hours, aliquot it into small tubes and store at -70℃.

[0212] 3.3 Preparation of expression cells

[0213] Digested cultured 293A cells, at 10 5 Cells / well were seeded into 6-well plates with a culture volume of 1 ml. After 24 hours, 10 μl of virus expressing the target gene (i.e., the virus obtained in Example 3.2) was added, and the cells were cultured for another 24 hours. The supernatant was then discarded, and fresh culture medium was added for further culture. Once the cells reached confluence, they were transferred to culture flasks, and puromycin was added to a final concentration of 3 μg / ml. The cells were cultured again, with the culture medium changed every two days while maintaining the puromycin concentration. After one week of selection, the surviving cells were identified as cells stably expressing the protein and named 293A-mIL12bIL12aIL2DiaNHS76F8GMCSF.

[0214] 3.4 Protein Expression and Purification

[0215] The constructed cells expressing mIL12bIL12aIL2DiaNHS76F8GMCSF (293A-mIL12bIL12aIL2DiaNHS76F8GMCSF) were passaged into 15cm culture dishes. After the cells reached confluence, the culture medium was replaced with 30ml CDM4HEK293, and the cells were cultured for another 5 days. The supernatant was then collected, filtered through a 0.45μm filter, and concentrated by ultrafiltration using 50kDa AMICONULTRA-15. The concentrated protein solution was purified using nickel chelate magnetic beads (purchased from Beaver Biotechnology Co., Ltd.) according to the instructions. The purified protein solution was then ultrafiltered using AMICON ULTRA-0.5 ultrafiltration tubes, and the buffer was replaced with PBS. Finally, the protein concentration was detected using an IL12p70 ELISA kit. The protein concentration was adjusted to 2μg / μl with PBS, aliquoted, and stored at -20℃.

[0216] Example 4. Effects of combined treatment with mIL12bIL12aIL2GMCSF and PD1 antibody on melanoma growth in mice.

[0217] 2×10 5 Digested and cultured mouse melanoma cells (B16) were injected subcutaneously into the right side of C57BL / 6 mice (6-10 weeks old, female), and treatment began when the long diameter of the tumor reached 6-8 mm.

[0218] Take 25 μl of the protein solution prepared in Example 2, then add it to 50 μl of glycerol. Quickly mix by pipetting with a pipette tip to avoid generating air bubbles, obtaining the injection formulation. Use a 29G insulin syringe to draw up the prepared injection solution and slowly inject it into the tumor. After injection, leave the needle in place for a short time to reduce solution spillage. After injection, return the mice to their cages and record the growth of the tumors. This experiment was divided into 3 groups: 1. No injection group, 2. Protein solution injection group, and 3. Protein solution injection combined with PD1 antibody treatment group (i.e., combination therapy group). In the combination therapy group, starting from the second day after protein solution injection, mice were injected intraperitoneally with 200 μg of PD1 antibody (specifically BioXcell, InVivoMAb anti-mouse PD-1 (CD279), catalog number BE0146), once every 3 days, for a total of 4 injections. Finally, the growth of tumors in each group was compared. The results are as follows: Figure 2 As shown, the combined treatment of mIL12bIL12aIL2GMCSF and PD1 antibody significantly inhibited tumor growth.

[0219] Example 5. Effect of combined treatment with mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody on melanoma growth in mice.

[0220] 2×10 5 Digested and cultured mouse melanoma cells (B16) were injected subcutaneously into the right side of C57BL / 6 mice (6-10 weeks old, female), and treatment began when the long diameter of the tumor reached 6-8 mm.

[0221] Take 25 μl of the protein solution prepared in Example 3, adjust the total volume to 200 μl with PBS, and inject the protein solution into the tail vein of mice using a 29G insulin syringe. Inject once daily for a total of 5 times. After injection, return the mice to their cages and record tumor growth. This experiment was divided into 3 groups: 1. No injection group, 2. Protein solution injection group, and 3. Protein solution injection combined with PD1 antibody treatment group (i.e., combined treatment group). In the combined treatment group, starting from the second day after the completion of the 5 protein solution injections, 200 μg of PD1 antibody (specifically BioXcell, InVivoMAb anti-mouse PD-1 (CD279), catalog number BE0146) was injected intraperitoneally every 3 days for a total of 4 times. Finally, the tumor growth of each group was compared. Results are as follows: Figure 3 As shown, the combined treatment of mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody significantly inhibited tumor growth.

[0222] Example 6. Expression of mIL12aIL2-IL12bGMCSF heterodimer proteins

[0223] 6.1 Constructing expression vectors

[0224] The mIL12aIL2-IL12bGMCSF heterodimer gene was synthesized. A 6*His sequence was added to the C-terminus of the IL2 gene for subsequent protein purification. The synthesized gene contained BamHI and XhoI restriction sites at both ends, respectively. The gene was then digested with BamHI and XhoI using the following digestion system: 5 μg mIL12aIL2-IL12bGMCSF heterodimer plasmid, 4 μl digestion buffer, 1 μl BamHI, 1 μl XhoI, and water to a total volume of 40 μl. The mixture was incubated at 37°C for 12 hours. The EP tube was then removed, and 4.4 μl of 10× loading buffer was added. Electrophoresis was performed on a 1% agarose gel. The mIL12aIL2-IL12bGMCSF gene fragment was recovered after electrophoresis and is ready for use.

[0225] The amino acid sequence of the first polypeptide chain of the mIL12aIL2-IL12bGMCSF heterodimer is shown in SEQ ID NO.53, and the amino acid sequence of the second polypeptide chain is shown in SEQ ID NO.57; the nucleotide sequence encoding the mIL12aIL2-IL12bGMCSF is shown in SEQ ID NO.73.

[0226] Digest the pLentis-CMV-MCS-IRES-PURO vector in an EP tube using the following formulation: 2 μg pLentis-CMV-MCS-IRES-PURO vector plasmid, 3 μl digestion buffer, 1 μl BamHI, and 1 μl XhoI. Add water to a total volume of 30 μl and incubate at 37°C for 12 hours. Remove the EP tube, add 3.3 μl of 10× loading buffer, and perform electrophoresis on a 1% agarose gel. After electrophoresis, recover the pLentis-CMV-MCS-IRES-PURO vector fragment for later use.

[0227] The mIL12aIL2-IL12bGMCSF and pLentis-CMV-MCS-IRES-PURO gene fragments were ligated using the following system: 2 μl pLentis-CMV-MCS-IRES-PURO vector fragment, 2 μl mIL12aIL2-IL12bGMCSF gene fragment, 1 μl ligase buffer, 0.5 μl T4 DNA ligase, and 4.5 μl water. The mixture was incubated at room temperature for 4 hours. The ligation system was then used to transform competent *E. coli* cells. The next day, colonies were picked from the transformed plates and cultured overnight in LB medium at 37°C using a shaker. Plasmids were extracted from the cultured bacteria using a plasmid extraction kit. Enzyme digestion was used to confirm successful ligation into the vector. Sequencing of the correct vector confirmed successful construction. The expression vector pLentis-CMV-mIL12aIL2-IL12bGMCSF-IRES-PURO was obtained.

[0228] 6.2 Preparation of expression virus

[0229] 1) Digest and culture 293FT cells, count them, and then divide them into 3×10⁻⁶ cells. 6 One cell per well is seeded into a 10cm culture dish, with a culture medium volume of 10ml.

[0230] 2) On the second evening, observe the cell condition. If the cells are in good condition, proceed with transfection. Add chloroquine to the culture plate to a final concentration of 25 μM. Take a test tube and add sterile water and the following plasmids (pMD2.G 6 μg + pSPAX2 15 μg + expression vector obtained in Example 2.1 above 20 μg), for a total volume of 1045 μl. Then add 155 μl of 2M CaCl2 and mix well. Finally, add 1200 μl of 2×HBS while vortexing. After the addition is complete, quickly add the mixture to the cell culture wells and gently shake to mix.

[0231] 3) On the morning of the third day, observe the cell status and replace the culture medium with 10 ml of fresh DMEM medium.

[0232] 4) On the morning of the fifth day, observe the cell status and collect the supernatant from the culture dish. Filter it with a 0.45μm filter and then place it in a high-speed centrifuge tube. Centrifuge at 50,000g for 2 hours. Carefully discard the supernatant and blot the liquid with absorbent paper as much as possible. Then resuspend the precipitate with 200μl HBSS. After dissolving for 2 hours, aliquot it into small tubes and store at -70℃.

[0233] 6.3 Preparation of expression cells

[0234] Digested cultured 293A cells, at 10 5 Cells were seeded per well into 6-well plates, with a culture volume of 1 ml. After 24 hours, 10 μl of virus expressing the target gene (i.e., the virus obtained in Example 2.2) was added, and the cells were cultured for another 24 hours. The supernatant was then discarded, and fresh culture medium was added for further culture. Once the cells reached confluence, they were transferred to culture flasks, and puromycin was added to a final concentration of 3 μg / ml. The cells were cultured again, with the culture medium changed every two days while maintaining the puromycin concentration. After one week of selection, the surviving cells were identified as cells stably expressing the protein and named 293A-mIL12aIL2-IL12bGMCSF.

[0235] 6.4 Protein Expression and Purification

[0236] The constructed cells expressing mIL12aIL2-IL12bGMCSF (293A-mIL12aIL2-IL12bGMCSF) were passaged into 15cm culture dishes. After the cells reached confluence, the culture medium was replaced with 30ml CDM4HEK293, and the cells were cultured for another 5 days. The supernatant was then collected, filtered through a 0.45μm filter, and concentrated using a 50kd AMICON ULTRA-15 ultrafiltration tube. The concentrated protein solution was purified using nickel chelate magnetic beads (purchased from Beaver Biotechnology Co., Ltd.) according to the instructions. The purified protein solution was then ultrafiltered using an AMICON ULTRA-0.5 ultrafiltration tube, and the buffer was replaced with PBS. Finally, the protein concentration was detected using an IL12p70 ELISA kit. The protein concentration was adjusted to 2μg / μl with PBS, aliquoted, and stored at -20℃.

[0237] Example 7. Effects of combined treatment with mIL12a, IL2-IL12b, GMCSF, and PD1 antibody on melanoma growth in mice.

[0238] 2×10 5 Digested and cultured mouse melanoma cells (B16) were injected subcutaneously into the right side of C57BL / 6 mice (6-10 weeks old, female), and treatment began when the long diameter of the tumor reached 6-8 mm.

[0239] Take 25 μl of the protein solution prepared in Example 6, then add it to 50 μl of glycerol. Quickly mix by pipetting with a pipette tip to avoid generating air bubbles, obtaining the injection formulation. Use a 29G insulin syringe to draw up the prepared injection solution and slowly inject it into the tumor. After injection, leave the needle in place for a short time to reduce solution spillage. After injection, return the mice to their cages and record the growth of the tumors. This experiment was divided into 3 groups: 1. No injection group, 2. Protein solution injection group, and 3. Protein solution injection combined with PD1 antibody treatment group (i.e., combination therapy group). In the combination therapy group, starting from the second day after protein solution injection, mice were injected intraperitoneally with 200 μg of PD1 antibody (specifically BioXcell, InVivoMAb anti-mouse PD-1 (CD279), catalog number BE0146), once every 3 days, for a total of 4 injections. Finally, the growth of tumors in each group was compared. The results are as follows: Figure 4 As shown, the combined treatment of mIL12aIL2-IL12bGMCSF and PD1 antibody significantly inhibited tumor growth.

[0240] Example 8. Effect of combined treatment with mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody on breast cancer growth in mice.

[0241] 2×10 5 Digested and cultured mouse breast cancer cells (4T1) were injected subcutaneously into the right side of Balb / c mice (6-10 weeks old, female), and treatment began when the long diameter of the tumor reached 6-8 mm.

[0242] Take 25 μl of the protein solution prepared in Example 3, adjust the total volume to 200 μl with PBS, and inject the protein solution into the tail vein of mice using a 29G insulin syringe. Inject once daily for a total of 5 times. After injection, return the mice to their cages and record tumor growth. This experiment was divided into 3 groups: 1. No injection group, 2. Protein solution injection group, and 3. Protein solution injection combined with PD1 antibody treatment group (i.e., combined treatment group). In the combined treatment group, starting from the second day after the completion of the 5 protein solution injections, 200 μg of PD1 antibody (specifically BioXcell, InVivoMAb anti-mouse PD-1 (CD279), catalog number BE0146) was injected intraperitoneally every 3 days for a total of 4 times. Finally, the tumor growth of each group was compared. Results are as follows: Figure 5 As shown, the combined treatment of mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody significantly inhibited tumor growth.

[0243] Example 9. Effect of combined treatment with mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody on lung cancer growth in mice.

[0244] 2×10 5 Digested and cultured mouse lung cancer cells (LLC) were injected subcutaneously into the right side of C57BL / 6 mice (6-10 weeks old, female), and treatment began when the long diameter of the tumor reached 6-8 mm.

[0245] Take 25 μl of the protein solution prepared in Example 3, adjust the total volume to 200 μl with PBS, and inject the protein solution into the tail vein of mice using a 29G insulin syringe. Inject once daily for a total of 3 times. After injection, return the mice to their cages and record tumor growth. This experiment was divided into 3 groups: 1. No injection group, 2. Protein solution injection group, and 3. Protein solution injection combined with PD1 antibody treatment group (i.e., combined treatment group). In the combined treatment group, starting from the second day after the completion of 5 protein solution injections, 200 μg of PD1 antibody (specifically BioXcell, InVivoMAb anti-mouse PD-1 (CD279), catalog number BE0146) was injected intraperitoneally every 3 days for a total of 4 times. Finally, the tumor growth of each group was compared. Results are as follows: Figure 6As shown, the combined treatment of mIL12bIL12aIL2DiaNHS76F8GMCSF and PD1 antibody significantly inhibited tumor growth. sequence list <110> Zhang Jinyu <120> A pharmaceutical composition and its use <130> 0069-PA-013CN <160> 77 <170> PatentIn version 3.5 <210> 1 <211> 469 <212> PRT <213> Artificial Sequence <220> <223> DiaL19 <400> 1 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe 20 25 30 Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly 115 120 125 Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala 130 135 140 Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro 145 150 155 160 Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr 165 170 175 Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr 180 185 190 Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys 195 200 205 Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly Gln Gly Thr Lys Val 210 215 220 Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser 225 230 235 240 Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro 245 250 255 Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser 260 265 270 Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu 275 280 285 Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp 290 295 300 Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr 305 310 315 320 Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr 325 330 335 Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr 340 345 350 Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser 355 360 365 Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys 370 375 380 Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp Tyr Gln Gln 385 390 395 400 Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala Ser Ser Arg 405 410 415 Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp 420 425 430 Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr 435 440 445 Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly Gln Gly Thr 450 455 460 Lys Val Glu Ile Lys 465 <210> 2 <211> 477 <212> PRT <213> Artificial Sequence <220> <223> DiaF8 <400> 2 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe 20 25 30 Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr 115 120 125 Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu 130 135 140 Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr 145 150 155 160 Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser 165 170 175 Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly 180 185 190 Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala 195 200 205 Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln 210 215 220 Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser 225 230 235 240 Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly 245 250 255 Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly 260 265 270 Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly 275 280 285 Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr 290 295 300 Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 305 310 315 320 Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp 325 330 335 Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe Asp 340 345 350 Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly 355 360 365 Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro 370 375 380 Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met 385 390 395 400 Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu 405 410 415 Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe 420 425 430 Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu 435 440 445 Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg 450 455 460 Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 465 470 475 <210> 3 [[ID=Y]]<211> 475 <212> PRT <213> Artificial Sequence <220> <223> DiaNHS76 <400> 3 Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly 20 25 30 Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp 35 40 45 Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu 50 55 60 Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser 65 70 75 80 Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln 115 120 125 Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys 130 135 140 Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys 145 150 155 160 Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro 165 170 175 Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala 180 185 190 Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr 195 200 205 Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly Gly 210 215 220 Thr Lys Leu Thr Val Leu Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 225 230 235 240 Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 245 250 255 Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr 260 265 270 Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly 275 280 285 Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr 290 295 300 Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser 305 310 315 320 Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr 325 330 335 Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp 340 345 350 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser 355 360 365 Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr 370 375 380 Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser 385 390 395 400 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly 405 410 415 Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser 420 425 430 [[ID=2I]]Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp 435 440 445 Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val 450 455 460 Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 465 470 475 <210> 4 <211> 476 <212> PRT <213> Artificial Sequence <220> <223> DiaNHS76F8 <400> 4 Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly 20 25 30 Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp 35 40 45 Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu 50 55 60 Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser 65 70 75 80 Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln 115 120 125 Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser 130 135 140 Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln 145 150 155 160 Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser 165 170 175 Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr 180 185 190 Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val 195 200 205 Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly 210 215 220 Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 225 230 235 240 Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu 245 250 255 Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe 260 265 270 Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys 275 280 285 Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr 290 295 300 Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser 305 310 315 320 Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 325 330 335 Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr 340 345 350 Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly 355 360 365 Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln 370 375 380 Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala 385 390 395 400 Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 405 410 415 Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser 420 425 430 Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu 435 440 445 Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His 450 455 460 Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 465 470 475 <210> 5 <211> 472 <212> PRT <213> Artificial Sequence <220> <223> DiaNHS76L19 <400> 5 Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly 20 25 30 Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp 35 40 45 Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu 50 55 60 Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser 65 70 75 80 Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu[[ID=3*]] 100 105 110 Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln 115 120 125 Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser 130 135 140 Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp Tyr Gln 145 150 155 160 Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala Ser Ser 165 170 175 Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr 180 185 190 Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val 195 200 205 Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly Gln Gly 210 215 220 Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 225 230 235 240 Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu 245 250 255 Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe 260 265 270 Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys 275 280 285 Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr 290 295 300 Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser 305 310 315 320 Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 325 330 335 Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly 340 345 350 Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Ser Ser Glu Leu 355 360 365 Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile 370 375 380 Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln 385 390 395 400 Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn 405 410 415 Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn 420 425 430 Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp 435 440 445 Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val Val Phe Gly 450 455 460 Gly Gly Thr Lys Leu Thr Val Leu 465 470 <210> 6 <211> 476 <212> PRT <213> Artificial Sequence <220> <223> DiaF8NHS76 <400> 6 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe 20 25 30 Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr 115 120 125 Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile Thr 130 135 140 Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln 145 150 155 160 Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg 165 170 175 Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr 180 185 190 Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr 195 200 205 Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly 210 215 220 Gly Thr Lys Leu Thr Val Leu Ser Ser Ser Ser Gly Ser Ser Ser Ser 225 230 235 240 Gly Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly 245 250 255 Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly 260 265 270 Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro 275 280 285 Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr 290 295 300 Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr 305 310 315 320 Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp 325 330 335 Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr 340 345 350 Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly 355 360 365 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 370 375 380 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro 385 390 395 400 Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 405 410 415 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser 420 425 430 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu 435 440 445 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro 450 455 460 Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 465 470 475 <210> 7 <211> 473 <212> PRT <213> Artificial Sequence <220> <223> DiaF8L19 <400> 7 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe 20 25 30 Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35​​​​​Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr 115 120 125 Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu 130 135 140 Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp Tyr 145 150 155 160 Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala Ser 165 170 175 Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly 180 185 190 Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala 195 200 205 Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly Gln 210 215 220 Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser 225 230 235 240 Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly 245 250 255 Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly 260 265 270 Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro Gly 275 280 285 Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr 290 295 300 Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 305 310 315 320 Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp 325 330 335 Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp 340 345 350 Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile Val 355 360 365 Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala 370 375 380 Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe Leu Ala 385 390 395 400 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly 405 410 415 Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly 420 425 430 Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp 435 440 445 [[ID=!17]]Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe 450×455 460 Gly Gln Gly Thr Lys Val Glu Ile Lys 465 470 <210> 8 <211> 472 <212> PRT <213> Artificial Sequence <220> <223> DiaL19NHS76 <400> 8 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe 20 25 30 Note: There seems to be an error in the original text where "450×455" in line 19 might be a mistake. I've translated it as "450×455" as it is in the original, but it's possible it should be something else. Also, in line 17, there is an exclamation mark (!) added which seems to be a mistake in the original text input. I've removed it in the translation for clarity. If these are not mistakes in the original, please let me know for a more accurate translation.Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala 115 120 125 Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp 130 135 140 Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln 145 150 155 160 Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile 165 170 175 Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr 180 185 190 Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser 195 200 205 Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu 210 215 220 Thr Val Leu Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser 225 230 235 240 Ser Gly Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro 245 250 255 Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser 260 265 270 Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu 275 280 285 Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro 290 295 300 Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln 305 310 315 320 Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr 325 330 335 Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly 340 345 350 苏氨酸 亮氨酸 缬氨酸 苏氨酸 缬氨酸 丝氨酸 丝氨酸 甘氨酸 甘氨酸 丝氨酸 甘氨酸 甘氨酸 谷氨酸 异亮氨酸 缬氨酸 亮氨酸 355 360 365 苏氨酸 谷氨酰胺 丝氨酸 脯氨酸 甘氨酸 苏氨酸 亮氨酸 丝氨酸 亮氨酸 丝氨酸 脯氨酸 甘氨酸 谷氨酸 精氨酸 丙氨酸 苏氨酸 370 375 380 亮氨酸 丝氨酸 半胱氨酸 精氨酸 丙氨酸 丝氨酸 谷氨酰胺 丝氨酸 缬氨酸 丝氨酸 丝氨酸 丝氨酸 苯丙氨酸 亮氨酸 丙氨酸 色氨酸 385 390 395 400 酪氨酸 谷氨酰胺 谷氨酰胺 赖氨酸 脯氨酸 甘氨酸 谷氨酰胺 丙氨酸 脯氨酸 精氨酸 亮氨酸 亮氨酸 异亮氨酸 酪氨酸 酪氨酸 丙氨酸 405 410 415 丝氨酸 丝氨酸 精氨酸 丙氨酸 苏氨酸 甘氨酸 异亮氨酸 脯氨酸 天冬氨酸 精氨酸 苯丙氨酸 丝氨酸 甘氨酸 丝氨酸 甘氨酸 丝氨酸 420 425 430 甘氨酸 苏氨酸 天冬氨酸 苯丙氨酸 苏氨酸 亮氨酸 苏氨酸 异亮氨酸 丝氨酸 精氨酸 亮氨酸 谷氨酸 脯氨酸 谷氨酸 天冬氨酸 苯丙氨酸 435 440 445 丙氨酸 缬氨酸 酪氨酸 酪氨酸 半胱氨酸 谷氨酰胺 谷氨酰胺 苏氨酸 甘氨酸 精氨酸 异亮氨酸 脯氨酸 脯氨酸 苏氨酸 苯丙氨酸 甘氨酸 450 455 460 谷氨酰胺 甘氨酸 苏氨酸 赖氨酸 缬氨酸 谷氨酸 异亮氨酸 赖氨酸 465 470 <210> 9 <211> 473 <212> PRT <213> Artificial Sequence <220> <223> DiaL19F8 <400> 9 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe 20 25 30 Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly 115 120 125 Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala 130 135 140 Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro 145 150 155 160 Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr 165 170 175 Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr 180 185 190 Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys 195 200 205 Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val 210 215 220 Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser 225 230 235 240 Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro 245 250 255 Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser 260 265 270 Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu 275 280 285 Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp 290 295 300 Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr 305 310 315 320 Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr 325 330 335 Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln 340 345 350 Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val 355 360 365 Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala 370 375 380 Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala 385 390 395 400 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr 405 410 415 Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly 420 425 430 Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp 435 440 445 Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe 450 455 460 Gly Gln Gly Thr Lys Val Glu Ile Lys 465 470 <210> 10 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> L19VL <400> 10 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser 20 25 30 Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45 Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro 85 90 95 Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 11 <211> 114 <212> PRT <213> Artificial Sequence <220> <223> L19VH <400> 11 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe 20 25 30 Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 100 105 110 Thr Val <210> 12 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> F8VL <400> 12 Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly 1 5 10 15 Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro 20 25 30 Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 35 40 45 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser 50 55 60 Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu 65 70 75 80 Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro 85 90 95 Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 100 105 <210> 13 <211> 118 <212> PRT <213> Artificial Sequence <220> <223> F8VH <400> 13 Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly 1 5 10 15 Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe 20 25 30 Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val 50 55 60 Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr 65 70 75 80 Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr 100 105 110 Leu Val Thr Val Ser Ser 115 <210> 14 <211> 108 <212> PRT <213> Artificial Sequence <220> <223> NHS76VL <400> 14 Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln 1 5 10 15 Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala 20 25 30 Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr 35 40 45 Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser 50 55 60 Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu 65 70 75 80 Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His 85 90 95 Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 100 105 <210> 15 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> NHS76VH <400> 15 Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu 1 5 10 15 Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly 20 25 30 Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp 35 40 45 Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu 50 55 60 Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser 65 70 75 80 Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 16 <211> 193 <212> PRT <213> Mus musculus <400> 16 Arg Val Ile Pro Val Ser Gly Pro Ala Arg Cys Leu Ser Gln Ser Arg 1 5 10 15 Asn Leu Leu Lys Thr Thr Asp Asp Met Val Lys Thr Ala Arg Glu Lys 20 25 30 Leu Lys His Tyr Ser Cys Thr Ala Glu Asp Ile Asp His Glu Asp Ile 35 40 45 Thr Arg Asp Gln Thr Ser Thr Leu Lys Thr Cys Leu Pro Leu Glu Leu 50 55 60 His Lys Asn Glu Ser Cys Leu Ala Thr Arg Glu Thr Ser Ser Thr Thr 65 70 75 80 Arg Gly Ser Cys Leu Pro Pro Gln Lys Thr Ser Leu Met Met Thr Leu 85 90 95 Cys Leu Gly Ser Ile Tyr Glu Asp Leu Lys Met Tyr Gln Thr Glu Phe 100 105 110 Gln Ala Ile Asn Ala Ala Leu Gln Asn His Asn His Gln Gln Ile Ile 115 120 125 Leu Asp Lys Gly Met Leu Val Ala Ile Asp Glu Leu Met Gln Ser Leu 130 135 140 Asn His Asn Gly Glu Thr Leu Arg Gln Lys Pro Pro Val Gly Glu Ala 145 150 155 160 Asp Pro Tyr Arg Val Lys Met Lys Leu Cys Ile Leu Leu His Ala Phe 165 170 175 Ser Thr Arg Val Val Thr Ile Asn Arg Val Met Gly Tyr Leu Ser Ser 180 185 190 Ala <210> 17 <211> 313 <212> PRT <213> Mus musculus <400> 17 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser 305 310 <210> 18 <211> 197 <212> PRT <213> Homo sapiens <400> 18 Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys Leu 1 5 10 15 His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln Lys 20 25 30 Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile Asp 35 40 45 His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys Leu 50 55 60 Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu Thr 65 70 75 80 Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser Phe 85 90 95 Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met Tyr 100 105 110 Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro Lys 115 120 125 Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu Leu 130 135 140 Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser Ser 145 150 155 160 Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile Leu 165 170 175 Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met Ser 180 185 190 Tyr Leu Asn Ala Ser 195 <210> 19 <211> 306 <212> PRT <213> Homo sapiens <400> 19 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser 305 <210> 20 <211> 124 <212> PRT <213> Mus musculus <400> 20 Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val 1 5 10 15 Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val Thr 20 25 30 Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser Phe Lys Lys 35 40 45 Leu Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu Gln Gly Leu Arg 50 55 60 Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn Met Thr Ala Ser Tyr 65 70 75 80 Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln 85 90 95 Val Thr Thr Tyr Ala Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr 100 105 110 Asp Ile Pro Phe Glu Cys Lys Lys Pro Gly Gln Lys 115 120 <210> 21 <211> 127 <212> PRT <213> Homo sapiens <400> 21 Ala Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His Val 1 5 10 15 Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg Asp Thr 20 25 30 Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu Met Phe Asp 35 40 45 Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu Leu Tyr Lys Gln 50 55 60 Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly Pro Leu Thr Met Met 65 70 75 80 Ala Ser His Tyr Lys Gln His Cys Pro Pro Thr Pro Glu Thr Ser Cys 85 90 95 Ala Thr Gln Ile Ile Thr Phe Glu Ser Phe Lys Glu Asn Leu Lys Asp 100 105 110 Phe Leu Leu Val Ile Pro Phe Asp Cys Trp Glu Pro Val Gln Glu 115 120 125 <210> 22 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Linker 1 – A link between cytokines or between cytokines and their target molecules. <400> twenty two Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1 5 10 15 <210> twenty three <211> 16 <212> PRT <213> Artificial Sequence <220> <223> Linker 2 – The link between cytokines and targets <400> twenty three Gly Gly Gly Gly Ser Leu Val Pro Arg Gly Ser Gly Gly Gly Gly Ser 1 5 10 15 <210> twenty four <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Linker 3 – A link between cytokines or between cytokines and their target molecules. <400> twenty four Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 1 5 10 15 <210> 25 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> Linker 4 – The link between cytokines and targets <400> 25 Ser Ser Ser Ser Gly Leu Val Pro Arg Gly Ser Ser Ser Ser Gly 1 5 10 15 <210> 26 <211> 6 <212> PRT <213> Artificial Sequence <220> <223> Linker 5 – The link between cytokines and targets <400> 26 Gly Ser Ala Asp Gly Gly 1 5 <210> 27 <211> 824 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aIL2GMCSF <400> 27 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 580 585 590 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 595 600 605 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln 610 615 620 Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 625 630 635 640 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 645 650 655 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp 660 665 670 Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Gly Gly Gly 675 680 685 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Arg 690 695 700 Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val Glu Ala Ile Lys 705 710 715 720 Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val Thr Leu Asn Glu Glu 725 730 735 Val Glu Val Val Ser Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val 740 745 750 Gln Thr Arg Leu Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr 755 760 765 Lys Leu Lys Gly Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr 770 775 780 Cys Pro Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr 785 790 795 800 Ala Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe 805 810 815 Glu Cys Lys Lys Pro Gly Gln Lys 820 <210> 28 <211> 824 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aGMCSFIL2 <400> 28 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Arg Ser Pro Ile Thr 530 535 540 Val Thr Arg Pro Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn 545 550 555 560 Leu Leu Asp Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val 565 570 575 Ser Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu 580 585 590 Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly 595 600 605 Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr 610 615 620 Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile 625 630 635 640 Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys 645 650 655 Pro Gly Gln Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 660 665 670 Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu 675 680 685 Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu 690 695 700 Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn Tyr 705 710 715 720 Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr Leu Pro 725 730 735 Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu Asp Glu Leu 740 745 750 Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser Lys Ser Phe Gln 755 760 765 Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile Arg Val Thr Val Val 770 775 780 Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys Gln Phe Asp Asp Glu 785 790 795 800 Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp Ile Ala Phe Cys Gln 805 810 815 Ser Ile Ile Ser Thr Ser Pro Gln 820 <210> 29 <211> 808 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aIL2GMCSF <400> 29 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln 530 535 540 Leu Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly 545 550 555 560 Ile Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys 565 570 575 Phe Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu 580 585 590 Glu Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser 595 600 605 Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val 610 615 620 Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr 625 630 635 640 Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr 645 650 655 Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr Gly Gly Gly Gly Ser Gly 660 665 670 Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Ala Arg Ser Pro Ser 675 680 685 Pro Ser Thr Gln Pro Trp Glu His Val Asn Ala Ile Gln Glu Ala Arg 690 695 700 Arg Leu Leu Asn Leu Ser Arg Asp Thr Ala Ala Glu Met Asn Glu Thr 705 710 715 720 Val Glu Val Ile Ser Glu Met Phe Asp Leu Gln Glu Pro Thr Cys Leu 725 730 735 Gln Thr Arg Leu Glu Leu Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr 740 745 750 Lys Leu Lys Gly Pro Leu Thr Met Met Ala Ser His Tyr Lys Gln His 755 760 765 Cys Pro Pro Thr Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe 770 775 780 Glu Ser Phe Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe 785 790 795 800 Asp Cys Trp Glu Pro Val Gln Glu 805 <210> 30 <211> 1308 <212> PRT[[ID=?]] <213> Artificial Sequence <220> <223> mIL12bIL12aDiaL19IL2GMCSF <400> 30 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly 580 585 590 Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp 625 630 635 640 Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu 645 650 655 Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu 660 665 670 Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe 675 680 685 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 690 695 700 Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly 705 710 715 720 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro 725 730 735 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro 740 745 750 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu 770 775 780 Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys 785 790 795 800 Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg 805 810 815 Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser 820 825 830 Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile 835 840 845 Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu 850 855 860 Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr 865 870 875 880 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly 885 890 895 Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro 900 905 910 Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser 915 920 925 Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu 930 935 940 Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe 945 950 955 960 Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu 965 970 975 Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile 980 985 990 Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Gly Gly Gly 995 1000 1005 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr 1010 1015 1020 Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln 1025 1030 1035 Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu Gln Leu Leu Met 1040 1045 1050 Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn Tyr Arg Asn Leu 1055 1060 1065 Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr Leu Pro Lys Gln 1070 1075 1080 Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu Asp Glu Leu Gly 1085 1090 1095 Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser Lys Ser Phe Gln 1100 1105 1110 Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile Arg Val Thr Val 1115 1120 1125 Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys Gln Phe Asp 1130 1135 1140 Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp Ile Ala 1145 1150 1155 Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Ser Ser Ser Ser 1160 1165 1170 Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala Pro Thr Arg 1175 1180 1185 Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val Glu Ala Ile 1190 1195 1200 Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val Thr Leu Asn 1205 1210 1215 Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser Phe Lys Lys Leu 1220 1225 1230 Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu Gln Gly Leu Arg 1235 1240 1245 Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn Met Thr Ala Ser 1250 1255 1260 Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu Thr Asp Cys Glu 1265 1270 1275 Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile Asp Ser Leu Lys Thr 1280 1285 1290 Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys Pro Gly Gln Lys 1295 1300 1305 <210> 31 <211> 1316 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaF8IL2GMCSF [[ID=​​​​​​​​​​​​​​​​​​​​​Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly 580 585 590 Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu 625 630 635 640 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 645 650 655 Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu 660 665 670 Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val 675 680 685 Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro 690 695 700 Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp 705 710 715 720 Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser 725 730 735 Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg 740 745 750 Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser 755 760 765 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val 770 775 780 Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu 785 790 795 800 Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met 805 810 815 Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala 820 825 830 Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly 835 840 845 Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln 850 855 860 Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys 865 870 875 880 Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val 885 890 895 Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser 900 905 910 Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys 915 920 925 Arg Ala Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln 930 935 940 Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg 945 950 955 960 Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp 965 970 975 Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr 980 985 990 Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr 995 1000 1005 Lys Val Glu Ile Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1010 1015 1020 Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser 1025 1030 1035 Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 1040 1045 1050 Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 1055 1060 1065 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr 1070 1075 1080 Phe Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu 1085 1090 1095 Gln Cys Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp 1100 1105 1110 Leu Thr Gln Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe 1115 1120 1125 Ile Ser Asn Ile Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp 1130 1135 1140 Asn Thr Phe Glu Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val 1145 1150 1155 Asp Phe Leu Arg Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser 1160 1165 1170 Thr Ser Pro Gln Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser 1175 1180 1185 Ser Ser Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg 1190 1195 1200 Pro Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu 1205 1210 1215 Asp Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser 1220 1225 1230 Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu 1235 1240 1245 Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys 1250 1255 1260 Gly Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro 1265 1270 1275 Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala 1280 1285 1290 Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe 1295 1300 1305 Glu Cys Lys Lys Pro Gly Gln Lys 1310 1315 <210> 32 <211> 1314 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaNHS76IL2GMCSF <400> 32 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys<( 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly 530 535 540 Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val 545 550 555 560 Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln 565 570 575 Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly 580 585 590 Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val 595 600 605 Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala 610 615 620 Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe 625 630 635 640 Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser 645 650 655 Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu 660 665 670 Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr 675 680 685 Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val 690 695 700 Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser 705 710 715 720 Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln 725 730 735 Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly 740 745 750 Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser Ser 755 760 765 Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Gln Val Gln 770 775 780 Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser 785 790 795 800 Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp 805 810 815 Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser 820 825 830 Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg 835 840 845 Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu 850 855 860 Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly 865 870 875 880 Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val 885 890 895 Ser Ser Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala 900 905 910 Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp 915 920 925 Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln 930 935 940 Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile 945 950 955 960 Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr 965 970 975 Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser 980 985 990 Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu 995 1000 1005 Thr Val Leu Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 1010 1015 1020 Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala 1025 1030 1035 Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His 1040 1045 1050 Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met 1055 1060 1065 Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys 1070 1075 1080 Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 1085 1090 1095 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr 1100 1105 1110 Gln Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser 1115 1120 1125 Asn Ile Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr 1130 1135 1140 Phe Glu Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe 1145 1150 1155 Leu Arg Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser 1160 1165 1170 Pro Gln Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser 1175 1180 1185 Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp 1190 1195 1200 Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp 1205 1210 1215 Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu 1220 1225 1230 Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys Ile 1235 1240 1245 Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly Ala 1250 1255 1260 Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr 1265 1270 1275 Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe 1280 1285 1290 Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys 1295 1300 1305 Lys Lys Pro Gly Gln Lys 1310 <210> 33 <211> 1315 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaNHS76F8IL2GMCSF <400> 33 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly 530 535 540 Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val 545 550 555 560 Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln 565 570 575 Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly 580 585 590 Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val 595 600 605 Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala 610 615 620 Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe 625 630 635 640 Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser 645 650 655 Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser 660 665 670 Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser 675 680 685 Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg 690 695 700 Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg 705 710 715 720 Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg 725 730 735 Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly 740 745 750 Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser 755 760 765 Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln 770 775 780 Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg 785 790 795 800 Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser 805 810 815 Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile 820 825 830 Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg 835 840 845 Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met 850 855 860 Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser 865 870 875 880 Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr 885 890 895 Val Ser Ser Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro 900 905 910 Ala Val Ser Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly 915 920 925 Asp Ser Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly 930 935 940 Gln Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly 945 950 955 960 Ile Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu 965 970 975 Thr Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn 980 985 990 Ser Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys 995 1000 1005 Leu Thr Val Leu Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 1010 1015 1020 Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr 1025 1030 1035 Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 1040 1045 1050 His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg 1055 1060 1065 Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 1070 1075 1080 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln 1085 1090 1095 Cys Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu 1100 1105 1110 Thr Gln Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile 1115 1120 1125 Ser Asn Ile Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn 1130 1135 1140 Thr Phe Glu Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp 1145 1150 1155 Phe Leu Arg Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr 1160 1165 1170 Ser Pro Gln Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser 1175 1180 1185 Ser Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro 1190 1195 1200 Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp 1205 1210 1215 Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn 1220 1225 1230 Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys 1235 1240 1245 Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly 1250 1255 1260 Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro 1265 1270 1275 Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp 1280 1285 1290 Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu 1295 1300 1 305 Cys Lys Lys Pro Gly Gln Lys 1310 1315 <210> 34 <211> 1311 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaNHS76L19IL2GMCSF <400> 34 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly 530 535 540 Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val 545 550 555 560 Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln 565 570 575 Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly 580 585 590 Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val 595 600 605 Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala 610 615 620 Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe 625 630 635 640 Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser 645 650 655 Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser 660 665 670 Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser 675 680 685 Ser Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg 690 695 700 Leu Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg 705 710 715 720 Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg 725 730 735 Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg 740 745 750 Ile Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser 755 760 765 Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln 770 775 780 Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg 785 790 795 800 Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe Ser Met Ser 805 810 815 Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile 820 825 830 Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg 835 840 845 Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met 850 855 860 Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro 865 870 875 880 Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Gly 885 890 895 Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val 900 905 910 Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg 915 920 925 Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val 930 935 940 Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg 945 950 955 960 Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly 965 970 975 Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser 980 985 990 Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 995 1000 1005 Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1010 1015 1020 Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu Ala Gln 1025 1030 1035 Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu Gln 1040 1045 1050 Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn Tyr 1055 1060 1065 Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr Leu 1070 1075 1080 Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu Asp 1085 1090 1095 Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser Lys 1100 1105 1110 Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile Arg 1115 1120 1125 Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 1130 1135 1140 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg 1145 1150 1155 Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Ser 1160 1165 1170 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala 1175 1180 1185 Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val 1190 1195 1200 Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val 1205 1210 1215 Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser Phe 1220 1225 1230 Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu Gln 1235 1240 1245 Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn Met 1250 1255 1260 Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu Thr 1265 1270 1275 Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile Asp Ser 1280 1285 1290 Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys Pro 1295 1300 1305 Gly Gln Lys 1310 <210> 35 <211> 1315 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaF8NHS76IL2GMCSF <400> 35 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly 580 585 590 Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu 625 630 635 640 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 645 650 655 Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala 660 665 670 Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser 675 680 685 Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu 690 695 700 Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe 705 710 715 720 Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala 725 730 735 Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser 740 745 750 Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser 755 760 765 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Gln Val 770 775 780 Gln Leu Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu 785 790 795 800 Ser Leu Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr 805 810 815 Trp Gly Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly 820 825 830 Ser Ile Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser 835 840 845 Arg Val Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys 850 855 860 Leu Ser Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg 865 870 875 880 Gly Lys Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr 885 890 895 Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro 900 905 910 Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg 915 920 925 Ala Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys 930 935 940 Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala 945 950 955 960 Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe 965 970 975 Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr 980 985 990 Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys 995 1000 1005 Val Glu Ile Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly 1010 1015 1020 Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr 1025 1030 1035 Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 1040 1045 1050 His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg 1055 1060 1065 Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 1070 1075 1080 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln 1085 1090 1095 Cys Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu 1100 1105 1110 Thr Gln Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile 1115 1120 1125 Ser Asn Ile Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn 1130 1135 1140 Thr Phe Glu Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp 1145 1150 1155 Phe Leu Arg Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr 1160 1165 1170 Ser Pro Gln Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser 1175 1180 1185 Ser Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro 1190 1195 1200 Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp 1205 1210 1215 Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn 1220 1225 1230 Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys 1235 1240 1245 Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly 1250 1255 1260 Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro 1265 1270 1275 Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp 1280 1285 1290 Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu 1295 1300 1305 Cys Lys Lys Pro Gly Gln Lys 1310 1315 <210> 36 <211> 1312 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaF8L19IL2GMCSF <400> 36 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly 580 585 590 Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu 625 630 635 640 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 645 650 655 Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu 660 665 670 Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val 675 680 685 Ser Ser Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro 690 695 700 Arg Leu Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp 705 710 715 720 Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser 725 730 735 Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly 740 745 750 Arg Ile Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser 755 760 765 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val 770 775 780 Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu 785 790 795 800 Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Ser Phe Ser Met 805 810 815 Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ser 820 825 830 Ile Ser Gly Ser Ser Gly Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly 835 840 845 Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln 850 855 860 Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys 865 870 875 880 Pro Phe Pro Tyr Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val 885 890 895 Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu 900 905 910 Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln 915 920 925 Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 930 935 940 Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile 945 950 955 960 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 965 970 975 Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln 980 985 990 Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile 995 1000 1005 Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 1010 1015 1020 Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu Ala 1025 1030 1035 Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu 1040 1045 1050 Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn 1055 1060 1065 Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr 1070 1075 1080 Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu 1085 1090 1095 Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser 1100 1105 1110 Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 1115 1120 1125 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu 1130 1135 1140 Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg 1145 1150 1155 Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln 1160 1165 1170 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 1175 1180 1185 Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His 1190 1195 1200 Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro 1205 1210 1215 Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser 1220 1225 1230 Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu 1235 1240 1245 Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn 1250 1255 1260 Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu 1265 1270 1275 Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile Asp 1280 1285 1290 Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys 1295 1300 1305 Pro Gly Gln Lys 1310 <210> 37 <211> 1311 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaL19NHS76IL2GMCSF <400> 37 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly 580 585 590 Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp 625 630 635 640 Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Ser 645 650 655 Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr 660 665 670 Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser 675 680 685 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly 690 695 700 Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser 705 710 715 720 Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp 725 730 735 Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val 740 745 750 Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu 770 775 780 Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys 785 790 795 800 Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile 805 810 815 Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His 820 825 830 Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 835 840 845 Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val 850 855 860 Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser 865 870 875 880 Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 885 890 895 Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser 900 905 910 Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser 915 920 925 Val Ser Ser Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala 930 935 940 Pro Arg Leu Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro 945 950 955 960 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 965 970 975 Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr 980 985 990 Gly Arg Ile Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 995 1000 1005 Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1010 1015 1020 Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu Ala Gln 1025 1030 1035 Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu Gln 1040 1045 1050 Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn Tyr 1055 1060 1065 Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr Leu 1070 1075 1080 Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu Asp 1085 1090 1095 Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser Lys 1100 1105 1110 Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile Arg 1115 1120 1125 Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 1130 1135 1140 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg 1145 1150 1155 Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Ser 1160 1165 1170 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala 1175 1180 1185 Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val 1190 1195 1200 Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val 1205 1210 1215 Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser Phe 1220 1225 1230 Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu Gln 1235 1240 1245 Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn Met 1250 1255 1260 Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu Thr 1265 1270 1275 Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile Asp Ser 1280 1285 1290 Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys Pro 1295 1300 1305 Gly Gln Lys 1310 <210> 38 <211> 1312 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aDiaL19F8IL2GMCSF <400> 38 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly 530 535 540 Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala 545 550 555 560 Ser Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala 565 570 575 Pro Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly 580 585 590 Thr Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg 595 600 605 Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 610 615 620 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp 625 630 635 640 Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu 645 650 655 Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu 660 665 670 Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe 675 680 685 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 690 695 700 Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly 705 710 715 720 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro 725 730 735 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro 740 745 750 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu 770 775 780 Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys 785 790 795 800 Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg 805 810 815 Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser 820 825 830 Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile 835 840 845 Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu 850 855 860 Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu 865 870 875 880 Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 885 890 895 Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu 900 905 910 Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln 915 920 925 Ser Val Ser Ser Ser Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln 930 935 940 Ala Pro Arg Leu Leu Ile Tyr Tyr Ala Ser Ser Arg Ala Thr Gly Ile 945 950 955 960 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr 965 970 975 Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln 980 985 990 Thr Gly Arg Ile Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile 995 1000 1005 Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 1010 1015 1020 Ser Ala Pro Thr Ser Ser Ser Thr Ser Ser Ser Thr Ala Glu Ala 1025 1030 1035 Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln His Leu Glu 1040 1045 1050 Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser Arg Met Glu Asn 1055 1060 1065 Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe Lys Phe Tyr 1070 1075 1080 Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys Leu Glu 1085 1090 1095 Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln Ser 1100 1105 1110 Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 1115 1120 1125 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu 1130 1135 1140 Cys Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg 1145 1150 1155 Arg Trp Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln 1160 1165 1170 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 1175 1180 1185 Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His 1190 1195 1200 Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp Asp Met Pro 1205 1210 1215 Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn Glu Phe Ser 1220 1225 1230 Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys Ile Phe Glu 1235 1240 1245 Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly Ala Leu Asn 1250 1255 1260 Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro Thr Pro Glu 1265 1270 1275 Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp Phe Ile Asp 1280 1285 1290 Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu Cys Lys Lys 1295 1300 1305 Pro Gly Gln Lys 1310 <210> 39 <211> 1315 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aIL2DiaNHS76F8GMCSF <400> 39 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 580 585 590 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 595 600 605 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln 610 615 620 Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 625 630 635 640 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 645 650 655 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp 660 665 670 Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Gly Gly Gly 675 680 685 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln Val Gln Leu 690 695 700 Gln Glu Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu 705 710 715 720 Thr Cys Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly 725 730 735 Trp Ile Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile 740 745 750 Tyr His Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val 755 760 765 Thr Ile Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser 770 775 780 Ser Val Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys 785 790 795 800 Trp Ser Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser 805 810 815 Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr 820 825 830 Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser 835 840 845 Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly 850 855 860 Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly 865 870 875 880 Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu 885 890 895 Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln 900 905 910 Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu 915 920 925 Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser 930 935 940 Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly 945 950 955 960 Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu 965 970 975 Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp 980 985 990 Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser 995 1000 1005 Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr 1010 1015 1020 Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val 1025 1030 1035 Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp 1040 1045 1050 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly 1055 1060 1065 Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly 1070 1075 1080 Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr 1085 1090 1095 Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu 1100 1105 1110 Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg 1115 1120 1125 Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr 1130 1135 1140 Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg 1145 1150 1155 Asp Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu 1160 1165 1170 Thr Val Leu Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser 1175 1180 1185 Ser Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg Pro 1190 1195 1200 Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu Asp 1205 1210 1215 Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser Asn 1220 1225 1230 Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu Lys 1235 1240 1245 Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys Gly 1250 1255 1260 Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro Pro 1265 1270 1275 Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala Asp 1280 1285 1290 Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe Glu 1295 1300 1305 Cys Lys Lys Pro Gly Gln Lys 1310 1315 <210> 40​​​​​​​​​​​​​​​​​​​​​​​​​​​​​Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 580 585 590 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 595 600 605 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln 610 615 620 Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 625 630 635 640 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 645 650 655 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp 660 665 670 Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Gly Gly Gly 675 680 685 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu 690 695 700 Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu 705 710 715 720 Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp 725 730 735 Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser 740 745 750 Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe 755 760 765 Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn 770 775 780 Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr 785 790 795 800 His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val 805 810 815 Ser Ser Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly 820 825 830 Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala 835 840 845 Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro 850 855 860 Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr 865 870 875 880 Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr 885 890 895 Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys 900 905 910 Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val 915 920 925 Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Ser Gly Ser Ser Ser 930 935 940 Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro 945 950 955 960 Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser 965 970 975 Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu 980 985 990 Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp 995 1000 1005 Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn 1010 1015 1020 Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala 1025 1030 1035 Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr 1040 1045 1050 Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly 1055 1060 1065 Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser 1070 1075 1080 Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val 1085 1090 1095 Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala 1100 1105 1110 Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile 1115 1120 1125 Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu 1130 1135 1140 Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys 1145 1150 1155 Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys 1160 1165 1170 Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser 1175 1180 1185 Ser Ser Ser Gly Ala Pro Thr Arg Ser Pro Ile Thr Val Thr Arg 1190 1195 1200 Pro Trp Lys His Val Glu Ala Ile Lys Glu Ala Leu Asn Leu Leu 1205 1210 1215 Asp Asp Met Pro Val Thr Leu Asn Glu Glu Val Glu Val Val Ser 1220 1225 1230 Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val Gln Thr Arg Leu 1235 1240 1245 Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr Lys Leu Lys 1250 1255 1260 Gly Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr Cys Pro 1265 1270 1275 Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr Ala 1280 1285 1290 Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe 1295 1300 1305 Glu Cys Lys Lys Pro Gly Gln Lys 1310 1315 <210> 41 <211> 1315 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aIL2GMCSFDiaNHS76F8 <400> 41 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 580 585 590 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 595 600 605 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln 610 615 620 Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 625 630 635 640 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 645 650 655 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp 660 665 670 Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Gly Gly Gly 675 680 685 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Arg 690 695 700 Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val Glu Ala Ile Lys 705 710 715 720 Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val Thr Leu Asn Glu Glu 725 730 735 Val Glu Val Val Ser Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val 740 745 750 Gln Thr Arg Leu Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr 755 760 765 Lys Leu Lys Gly Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr 770 775 780 Cys Pro Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr 785 790 795 800 Ala Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe 805 810 815 Glu Cys Lys Lys Pro Gly Gln Lys Ser Ser Ser Ser Gly Ser Ser Ser 820 825 830 Ser Gly Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu Ser Gly Pro 835 840 845 Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser 850 855 860 Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro 865 870 875 880 Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser 885 890 895 Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp 900 905 910 Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala 915 920 925 Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp 930 935 940 Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly 945 950 955 960 Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro 965 970 975 Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met 980 985 990 Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu 995 1000 1005 Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg 1010 1015 1020 Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser 1025 1030 1035 Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met 1040 1045 1050 Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile 1055 1060 1065 Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser 1070 1075 1080 Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro 1085 1090 1095 Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe 1100 1105 1110 Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly 1115 1120 1125 Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr 1130 1135 1140 Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn 1145 1150 1155 Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu 1160 1165 1170 Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu 1175 1180 1185 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 1190 1195 1200 Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser 1205 1210 1215 Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser 1220 1225 1230 Leu Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln 1235 1240 1245 Ala Pro Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly 1250 1255 1260 Ile Pro Asp Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser 1265 1270 1275 Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr 1280 1285 1290 Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val Val Phe Gly Gly 1295 1300 1305 Gly Thr Lys Leu Thr Val Leu 1310 1315 <210> 42 <211> 1316 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aIL2GMCSFDiaF8 <400> 42 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110 Gly Arg Phe Thr Cys Ser Trp Leu Val Gln Arg Asn Met Asp Leu Lys 115 120 125 Phe Asn Ile Lys Ser Ser Ser Ser Ser Pro Asp Ser Arg Ala Val Thr 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met Glu Asn Tyr Arg Asn Leu Lys Leu Pro Arg Met Leu Thr Phe 580 585 590 Lys Phe Tyr Leu Pro Lys Gln Ala Thr Glu Leu Lys Asp Leu Gln Cys 595 600 605 Leu Glu Asp Glu Leu Gly Pro Leu Arg His Val Leu Asp Leu Thr Gln 610 615 620 Ser Lys Ser Phe Gln Leu Glu Asp Ala Glu Asn Phe Ile Ser Asn Ile 625 630 635 640 Arg Val Thr Val Val Lys Leu Lys Gly Ser Asp Asn Thr Phe Glu Cys 645 650 655 Gln Phe Asp Asp Glu Ser Ala Thr Val Val Asp Phe Leu Arg Arg Trp 660 665 670 Ile Ala Phe Cys Gln Ser Ile Ile Ser Thr Ser Pro Gln Gly Gly Gly 675 680 685 Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Arg 690 695 700 Ser Pro Ile Thr Val Thr Arg Pro Trp Lys His Val Glu Ala Ile Lys 705 710 715 720 Glu Ala Leu Asn Leu Leu Asp Asp Met Pro Val Thr Leu Asn Glu Glu 725 730 735 Val Glu Val Val Ser Asn Glu Phe Ser Phe Lys Lys Leu Thr Cys Val 740 745 750 Gln Thr Arg Leu Lys Ile Phe Glu Gln Gly Leu Arg Gly Asn Phe Thr 755 760 765 Lys Leu Lys Gly Ala Leu Asn Met Thr Ala Ser Tyr Tyr Gln Thr Tyr 770 775 780 Cys Pro Pro Thr Pro Glu Thr Asp Cys Glu Thr Gln Val Thr Thr Tyr 785 790 795 800 Ala Asp Phe Ile Asp Ser Leu Lys Thr Phe Leu Thr Asp Ile Pro Phe 805 810 815 Glu Cys Lys Lys Pro Gly Gln Lys Ser Ser Ser Ser Gly Ser Ser Ser 820 825 830 Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly 835 840 845 Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser 850 855 860 Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro 865 870 875 880 Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser 885 890 895 Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 900 905 910 Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu 915 920 925 Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr Leu Phe 930 935 940 Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser 945 950 955 960 Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser 965 970 975 Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser 980 985 990 Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg 995 1000 1005 Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp 1010 1015 1020 Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 1025 1030 1035 Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln 1040 1045 1050 Met Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu 1055 1060 1065 Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser 1070 1075 1080 Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln 1085 1090 1095 Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr 1100 1105 1110 Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln Ala Pro Gly Lys 1115 1120 1125 Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly Gly Ser Thr 1130 1135 1140 Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 1145 1150 1155 Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala 1160 1165 1170 Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr 1175 1180 1185 Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 1190 1195 1200 Gly Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr 1205 1210 1215 Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala 1220 1225 1230 Ser Gln Ser Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys 1235 1240 1245 Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg 1250 1255 1260 Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr 1265 1270 1275 Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe Ala 1280 1285 1290 Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro Thr Phe Gly 1295 1300 1305 Gln Gly Thr Lys Val Glu Ile Lys 1310 1315 <210> 43 <211> 1292 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaL19IL2GMCSF <400> 43 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu 530 535 540 Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe 545 550 555 560 Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys 565 570 575 Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr 580 585 590 Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser 595 600 605 Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly 625 630 635 640 Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile Val Leu 645 650 655 Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr 660 665 670 Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp 675 680 685 Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala 690 695 700 Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser 705 710 715 720 Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe 725 730 735 Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly 740 745 750 Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser 755 760 765 Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly 770 775 780 Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser 785 790 795 800 Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro 805 810 815 Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr 820 825 830 Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 835 840 845 Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu 850 855 860 Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr 865 870 875 880 Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile 885 890 895 Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg 900 905 910 Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu 915 920 925 Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr 930 935 940 Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser 945 950 955 960 Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu 965 970 975 Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr 980 985 990 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Gly Gly Gly Gly Ser Gly 995 1000 1005 Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser 1010 1015 1020 Thr Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp Leu 1025 1030 1035 Gln Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu 1040 1045 1050 Thr Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr 1055 1060 1065 Glu Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu 1070 1075 1080 Glu Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg 1085 1090 1095 Pro Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu 1100 1105 1110 Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr 1115 1120 1125 Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln 1130 1135 1140 Ser Ile Ile Ser Thr Leu Thr Ser Ser Ser Ser Gly Ser Ser Ser 1145 1150 1155 Ser Gly Ser Ser Ser Ser Gly Ala Pro Ala Arg Ser Pro Ser Pro 1160 1165 1170 Ser Thr Gln Pro Trp Glu His Val Asn Ala Ile Gln Glu Ala Arg 1175 1180 1185 Arg Leu Leu Asn Leu Ser Arg Asp Thr Ala Ala Glu Met Asn Glu 1190 1195 1200 Thr Val Glu Val Ile Ser Glu Met Phe Asp Leu Gln Glu Pro Thr 1205 1210 1215 Cys Leu Gln Thr Arg Leu Glu Leu Tyr Lys Gln Gly Leu Arg Gly 1220 1225 1230 Ser Leu Thr Lys Leu Lys Gly Pro Leu Thr Met Met Ala Ser His 1235 1240 1245 Tyr Lys Gln His Cys Pro Pro Thr Pro Glu Thr Ser Cys Ala Thr 1250 1255 1260 Gln Ile Ile Thr Phe Glu Ser Phe Lys Glu Asn Leu Lys Asp Phe 1265 1270 1275 Leu Leu Val Ile Pro Phe Asp Cys Trp Glu Pro Val Gln Glu 1280 1285 1290 <210> 44 <211> 1298 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaNHS76IL2GMCSF <400> 44 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 530 535 540 Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr 545 550 555 560 Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly 565 570 575 Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr 580 585 590 Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser 595 600 605 Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp 625 630 635 640 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser 645 650 655 Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr 660 665 670 Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser 675 680 685 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly 690 695 700 Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser 705 710 715 720 Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp 725 730 735 Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val 740 745 750 Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu 770 775 780 Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys 785 790 795 800 Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile 805 810 815 Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His 820 825 830 Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 835 840 845 Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val 850 855 860 Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser 865 870 875 880 Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 885 890 895 Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val 900 905 910 Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg 915 920 925 Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val 930 935 940 Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg 945 950 955 960 Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly 965 970 975 Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser 980 985 990 Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 995 1000 1005 Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1010 1015 1020 Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu Glu 1025 1030 1035 His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn Asn 1040 1045 1050 Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe Tyr 1055 1060 1065 Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu Glu 1070 1075 1080 Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser 1085 1090 1095 Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn 1100 1105 1110 Val Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys 1115 1120 1125 Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg 1130 1135 1140 Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr Ser Ser 1145 1150 1155 Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala Pro 1160 1165 1170 Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His Val Asn 1175 1180 1185 Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg Asp Thr 1190 1195 1200 Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu Met Phe 1205 1210 1215 Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu Leu Tyr 1220 1225 1230 Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly Pro Leu 1235 1240 1245 Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro Pro Thr Pro 1250 1255 1260 Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu Ser Phe Lys 1265 1270 1275 Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe Asp Cys Trp 1280 1285 1290 Glu Pro Val Gln Glu 1295 <210> 45 <211> 1299 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaNHS76F8IL2GMCSF <400> 45 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 530 535 540 Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr 545 550 555 560 Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly 565 570 575 Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr 580 585 590 Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser 595 600 605 Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp 625 630 635 640 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu 645 650 655 Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu 660 665 670 Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe 675 680 685 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 690 695 700 Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly 705 710 715 720 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro 725 730 735 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro 740 745 750 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu 770 775 780 Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys 785 790 795 800 Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg 805 810 815 Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser 820 825 830 Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile 835 840 845 Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu 850 855 860 Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu 865 870 875 880 Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 885 890 895 Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser 900 905 910 Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu 915 920 925 Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro 930 935 940 Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp 945 950 955 960 Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr 965 970 975 Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp 980 985 990 Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val 995 1000 1005 Leu Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 1010 1015 1020 Ser Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu 1025 1030 1035 Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn 1040 1045 1050 Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe 1055 1060 1065 Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu 1070 1075 1080 Glu Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln 1085 1090 1095 Ser Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile 1100 1105 1110 Asn Val Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met 1115 1120 1125 Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn 1130 1135 1140 Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr Ser 1145 1150 1155 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala 1160 1165 1170 Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His Val 1175 1180 1185 Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg Asp 1190 1195 1200 Thr Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu Met 1205 1210 1215 Phe Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu Leu 1220 1225 1230 Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly Pro 1235 1240 1245 Leu Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro Pro Thr 1250 1255 1260 Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu Ser Phe 1265 1270 1275 Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe Asp Cys 1280 1285 1290 Trp Glu Pro Val Gln Glu 1295 <210> 46 <211> 1299 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aIL2DiaNHS76F8GMCSF <400> 46 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln 530 535 540 Leu Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly 545 550 555 560 Ile Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys 565 570 575 Phe Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu 580 585 590 Glu Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser 595 600 605 Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val 610 615 620 Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr 625 630 635 640 Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr 645 650 655 Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr Gly Gly Gly Gly Ser Gly 660 665 670 Gly Gly Gly Ser Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser 675 680 685 Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala 690 695 700 Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg 705 710 715 720 Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser 725 730 735 Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser 740 745 750 Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr 755 760 765 Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys 770 775 780 Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly 785 790 795 800 Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu 805 810 815 Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val 820 825 830 Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro 835 840 845 Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp 850 855 860 Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser 865 870 875 880 Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg 885 890 895 Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser 900 905 910 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val 915 920 925 Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu 930 935 940 Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met 945 950 955 960 Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala 965 970 975 Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly 980 985 990 Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln 995 1000 1005 Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala 1010 1015 1020 Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr 1025 1030 1035 Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser Ser Glu Leu 1040 1045 1050 Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr Val Arg 1055 1060 1065 Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser Trp 1070 1075 1080 Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly 1085 1090 1095 Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser 1100 1105 1110 Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala 1115 1120 1125 Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly 1130 1135 1140 Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser 1145 1150 1155 Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ala 1160 1165 1170 Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His Val 1175 1180 1185 Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg Asp 1190 1195 1200 Thr Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu Met 1205 1210 1215 Phe Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu Leu 1220 1225 1230 Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly Pro 1235 1240 1245 Leu Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro Pro Thr 1250 1255 1260 Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu Ser Phe 1265 1270 1275 Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe Asp Cys 1280 1285 1290 Trp Glu Pro Val Gln Glu 1295 <210> 47 <211> 1300 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aIL2DiaF8GMCSF <400> 47 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln 530 535 540 Leu Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly 545 550 555 560 Ile Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys 565 570 575 Phe Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu 580 585 590 Glu Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln Ser 595 600 605 Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile Asn Val 610 615 620 Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr 625 630 635 640 Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr 645 650 655 Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr Gly Gly Gly Gly Ser Gly 660 665 670 Gly Gly Gly Ser Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser 675 680 685 Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala 690 695 700 Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg Gln 705 710 715 720 Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser Gly 725 730 735 Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser 740 745 750 Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg 755 760 765 Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu Tyr 770 775 780 Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 785 790 795 800 Gly Ser Gly Gly Glu Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser 805 810 815 Leu Ser Pro Gly Glu Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser 820 825 830 Val Ser Met Pro Phe Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala 835 840 845 Pro Arg Leu Leu Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro 850 855 860 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile 865 870 875 880 Ser Arg Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met 885 890 895 Arg Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 900 905 910 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu 915 920 925 Val Gln Leu Leu Glu Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser 930 935 940 Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr 945 950 955 960 Met Ser Trp Val Arg Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser 965 970 975 Ala Ile Ser Gly Ser Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys 980 985 990 Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu 995 1000 1005 Gln Met Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 1010 1015 1020 Ala Lys Ser Thr His Leu Tyr Leu Phe Asp Tyr Trp Gly Gln Gly 1025 1030 1035 Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu Ile Val 1040 1045 1050 Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg 1055 1060 1065 Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe 1070 1075 1080 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu 1085 1090 1095 Ile Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe 1100 1105 1110 Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg 1115 1120 1125 Leu Glu Pro Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg 1130 1135 1140 Gly Arg Pro Pro Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys 1145 1150 1155 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly 1160 1165 1170 Ala Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His 1175 1180 1185 Val Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg 1190 1195 1200 Asp Thr Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu 1205 1210 1215 Met Phe Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu 1220 1225 1230 Leu Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly 1235 1240 1245 Pro Leu Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro Pro 1250 1255 1260 Thr Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu Ser 1265 1270 1275 Phe Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe Asp 1280 1285 1290 Cys Trp Glu Pro Val Gln Glu 1295 1300 <210> 48 <211> 1292 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaL19GMCSFIL2 <400> 48 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu ​​​​​​​​​​​​​​Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Glu Val Gln Leu Leu Glu Ser Gly Gly Gly Leu 530 535 540 Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Phe 545 550 555 560 Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro Gly Lys 565 570 575 Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr Thr Tyr 580 585 590 Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp Asn Ser 595 600 605 Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr Trp Gly 625 630 635 640 Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile Val Leu 645 650 655 Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg Ala Thr 660 665 670 Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu Ala Trp 675 680 685 Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr Tyr Ala 690 695 700 Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser Gly Ser 705 710 715 720 Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu Asp Phe 725 730 735 Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr Phe Gly 740 745 750 Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly Ser Ser Ser 755 760 765 Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu Ser Gly Gly 770 775 780 Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys Ala Ala Ser 785 790 795 800 Gly Phe Thr Phe Ser Ser Phe Ser Met Ser Trp Val Arg Gln Ala Pro 805 810 815 Gly Lys Gly Leu Glu Trp Val Ser Ser Ile Ser Gly Ser Ser Gly Thr 820 825 830 Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile Ser Arg Asp 835 840 845 Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu Arg Ala Glu 850 855 860 Asp Thr Ala Val Tyr Tyr Cys Ala Lys Pro Phe Pro Tyr Phe Asp Tyr 865 870 875 880 Trp Gly Gln Gly Thr Leu Val Thr Val Gly Gly Ser Gly Gly Glu Ile 885 890 895 Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu Arg 900 905 910 Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Ser Ser Phe Leu 915 920 925 Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile Tyr 930 935 940 Tyr Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly Ser 945 950 955 960 Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro Glu 965 970 975 Asp Phe Ala Val Tyr Tyr Cys Gln Gln Thr Gly Arg Ile Pro Pro Thr 980 985 990 Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Gly Gly Gly Gly Ser Gly 995 1000 1005 Gly Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Ala Arg Ser Pro 1010 1015 1020 Ser Pro Ser Thr Gln Pro Trp Glu His Val Asn Ala Ile Gln Glu 1025 1030 1035 Ala Arg Arg Leu Leu Asn Leu Ser Arg Asp Thr Ala Ala Glu Met 1040 1045 1050 Asn Glu Thr Val Glu Val Ile Ser Glu Met Phe Asp Leu Gln Glu 1055 1060 1065 Pro Thr Cys Leu Gln Thr Arg Leu Glu Leu Tyr Lys Gln Gly Leu 1070 1075 1080 Arg Gly Ser Leu Thr Lys Leu Lys Gly Pro Leu Thr Met Met Ala 1085 1090 1095 Ser His Tyr Lys Gln His Cys Pro Pro Thr Pro Glu Thr Ser Cys 1100 1105 1110 Ala Thr Gln Ile Ile Thr Phe Glu Ser Phe Lys Glu Asn Leu Lys 1115 1120 1125 Asp Phe Leu Leu Val Ile Pro Phe Asp Cys Trp Glu Pro Val Gln 1130 1135 1140 Glu Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Ser Ser Ser Ser 1145 1150 1155 Gly Ala Pro Thr Ser Ser Ser Thr Lys Lys Thr Gln Leu Gln Leu 1160 1165 1170 Glu His Leu Leu Leu Asp Leu Gln Met Ile Leu Asn Gly Ile Asn 1175 1180 1185 Asn Tyr Lys Asn Pro Lys Leu Thr Arg Met Leu Thr Phe Lys Phe 1190 1195 1200 Tyr Met Pro Lys Lys Ala Thr Glu Leu Lys His Leu Gln Cys Leu 1205 1210 1215 Glu Glu Glu Leu Lys Pro Leu Glu Glu Val Leu Asn Leu Ala Gln 1220 1225 1230 Ser Lys Asn Phe His Leu Arg Pro Arg Asp Leu Ile Ser Asn Ile 1235 1240 1245 Asn Val Ile Val Leu Glu Leu Lys Gly Ser Glu Thr Thr Phe Met 1250 1255 1260 Cys Glu Tyr Ala Asp Glu Thr Ala Thr Ile Val Glu Phe Leu Asn 1265 1270 1275 Arg Trp Ile Thr Phe Cys Gln Ser Ile Ile Ser Thr Leu Thr 1280 1285 1290 <210> 49 <211> 1298 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaNHS76GMCSFIL2 <400> 49 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 530 535 540 Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr 545 550 555 560 Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly 565 570 575 Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr 580 585 590 Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser 595 600 605 Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp 625 630 635 640 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Ser 645 650 655 Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val Ala Leu Gly Gln Thr 660 665 670 Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg Ser Tyr Tyr Ala Ser 675 680 685 Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val Leu Val Ile Tyr Gly 690 695 700 Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg Phe Ser Gly Ser Ser 705 710 715 720 Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly Ala Gln Ala Glu Asp 725 730 735 Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser Ser Gly Asn His Val 740 745 750 Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Gln Val Gln Leu Gln Glu 770 775 780 Ser Gly Pro Gly Leu Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys 785 790 795 800 Ala Val Ser Gly Tyr Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile 805 810 815 Arg Gln Pro Pro Gly Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His 820 825 830 Ser Gly Ser Thr Tyr Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile 835 840 845 Ser Val Asp Thr Ser Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val 850 855 860 Thr Ala Ala Asp Thr Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser 865 870 875 880 Lys Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly 885 890 895 Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser Val 900 905 910 Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu Arg 915 920 925 Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Val 930 935 940 Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp Arg 945 950 955 960 Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr Gly 965 970 975 Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp Ser 980 985 990 Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val Leu 995 1000 1005 Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 1010 1015 1020 Ala Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu His 1025 1030 1035 Val Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser Arg 1040 1045 1050 Asp Thr Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser Glu 1055 1060 1065 Met Phe Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu Glu 1070 1075 1080 Leu Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys Gly 1085 1090 1095 Pro Leu Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro Pro 1100 1105 1110 Thr Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu Ser 1115 1120 1125 Phe Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe Asp 1130 1135 1140 Cys Trp Glu Pro Val Gln Glu Ser Ser Ser Ser Gly Ser Ser Ser 1145 1150 1155 Ser Gly Ser Ser Ser Ser Gly Ala Pro Thr Ser Ser Ser Thr Lys 1160 1165 1170 Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp Leu Gln Met 1175 1180 1185 Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu Thr Arg 1190 1195 1200 Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr Glu Leu 1205 1210 1215 Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu Glu Glu 1220 1225 1230 Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro Arg 1235 1240 1245 Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys Gly 1250 1255 1260 Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala Thr 1265 1270 1275 Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser Ile 1280 1285 1290 Ile Ser Thr Leu Thr 1295 <210> 50 <211> 1299 <212> PRT <213> Artificial Sequence <220> <223> hIL12bIL12aDiaNHS76F8GMCSFIL2 <400> 50 Ile Trp Glu Leu Lys Lys Asp Val Tyr Val Val Glu Leu Asp Trp Tyr 1 5 10 15 Pro Asp Ala Pro Gly Glu Met Val Val Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Gly Ile Thr Trp Thr Leu Asp Gln Ser Ser Glu Val Leu Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Gln Val Lys Glu Phe Gly Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Val Leu Ser His Ser Leu Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asp Gly Ile Trp Ser Thr Asp Ile Leu Lys 85 90 95 Asp Gln Lys Glu Pro Lys Asn Lys Thr Phe Leu Arg Cys Glu Ala Lys 100 105 110 Asn Tyr Ser Gly Arg Phe Thr Cys Trp Trp Leu Thr Thr Ile Ser Thr 115 120 125 Asp Leu Thr Phe Ser Val Lys Ser Ser Arg Gly Ser Ser Asp Pro Gln 130 135 140 Gly Val Thr Cys Gly Ala Ala Thr Leu Ser Ala Glu Arg Val Arg Gly 145 150 155 160 Asp Asn Lys Glu Tyr Glu Tyr Ser Val Glu Cys Gln Glu Asp Ser Ala 165 170 175 Cys Pro Ala Ala Glu Glu Ser Leu Pro Ile Glu Val Met Val Asp Ala 180 185 190 Val His Lys Leu Lys Tyr Glu Asn Tyr Thr Ser Ser Phe Phe Ile Arg 195 200 205 Asp Ile Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Leu Lys Pro Leu 210 215 220 Lys Asn Ser Arg Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Thr Trp 225 230 235 240 Ser Thr Pro His Ser Tyr Phe Ser Leu Thr Phe Cys Val Gln Val Gln 245 250 255 Gly Lys Ser Lys Arg Glu Lys Lys Asp Arg Val Phe Thr Asp Lys Thr 260 265 270 Ser Ala Thr Val Ile Cys Arg Lys Asn Ala Ser Ile Ser Val Arg Ala 275 280 285 Gln Asp Arg Tyr Tyr Ser Ser Ser Trp Ser Glu Trp Ala Ser Val Pro 290 295 300 Cys Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 305 310 315 320 Ser Arg Asn Leu Pro Val Ala Thr Pro Asp Pro Gly Met Phe Pro Cys 325 330 335 Leu His His Ser Gln Asn Leu Leu Arg Ala Val Ser Asn Met Leu Gln 340 345 350 Lys Ala Arg Gln Thr Leu Glu Phe Tyr Pro Cys Thr Ser Glu Glu Ile 355 360 365 Asp His Glu Asp Ile Thr Lys Asp Lys Thr Ser Thr Val Glu Ala Cys 370 375 380 Leu Pro Leu Glu Leu Thr Lys Asn Glu Ser Cys Leu Asn Ser Arg Glu 385 390 395 400 Thr Ser Phe Ile Thr Asn Gly Ser Cys Leu Ala Ser Arg Lys Thr Ser 405 410 415 Phe Met Met Ala Leu Cys Leu Ser Ser Ile Tyr Glu Asp Leu Lys Met 420 425 430 Tyr Gln Val Glu Phe Lys Thr Met Asn Ala Lys Leu Leu Met Asp Pro 435 440 445 Lys Arg Gln Ile Phe Leu Asp Gln Asn Met Leu Ala Val Ile Asp Glu 450 455 460 Leu Met Gln Ala Leu Asn Phe Asn Ser Glu Thr Val Pro Gln Lys Ser 465 470 475 480 Ser Leu Glu Glu Pro Asp Phe Tyr Lys Thr Lys Ile Lys Leu Cys Ile 485 490 495 Leu Leu His Ala Phe Arg Ile Arg Ala Val Thr Ile Asp Arg Val Met 500 505 510 Ser Tyr Leu Asn Ala Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 515 520 525 Gly Gly Gly Gly Ser Gln Val Gln Leu Gln Glu Ser Gly Pro Gly Leu 530 535 540 Val Lys Pro Ser Glu Thr Leu Ser Leu Thr Cys Ala Val Ser Gly Tyr 545 550 555 560 Ser Ile Ser Ser Gly Tyr Tyr Trp Gly Trp Ile Arg Gln Pro Pro Gly 565 570 575 Lys Gly Leu Glu Trp Ile Gly Ser Ile Tyr His Ser Gly Ser Thr Tyr 580 585 590 Tyr Asn Pro Ser Leu Lys Ser Arg Val Thr Ile Ser Val Asp Thr Ser 595 600 605 Lys Asn Gln Phe Ser Leu Lys Leu Ser Ser Val Thr Ala Ala Asp Thr 610 615 620 Ala Val Tyr Tyr Cys Ala Arg Gly Lys Trp Ser Lys Phe Asp Tyr Trp 625 630 635 640 Gly Gln Gly Thr Leu Val Thr Val Ser Ser Gly Gly Ser Gly Gly Glu 645 650 655 Ile Val Leu Thr Gln Ser Pro Gly Thr Leu Ser Leu Ser Pro Gly Glu 660 665 670 Arg Ala Thr Leu Ser Cys Arg Ala Ser Gln Ser Val Ser Met Pro Phe 675 680 685 Leu Ala Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro Arg Leu Leu Ile 690 695 700 Tyr Gly Ala Ser Ser Arg Ala Thr Gly Ile Pro Asp Arg Phe Ser Gly 705 710 715 720 Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser Arg Leu Glu Pro 725 730 735 Glu Asp Phe Ala Val Tyr Tyr Cys Gln Gln Met Arg Gly Arg Pro Pro 740 745 750 Thr Phe Gly Gln Gly Thr Lys Val Glu Ile Lys Ser Ser Ser Ser Gly 755 760 765 Ser Ser Ser Ser Gly Ser Ser Ser Ser Gly Glu Val Gln Leu Leu Glu 770 775 780 Ser Gly Gly Gly Leu Val Gln Pro Gly Gly Ser Leu Arg Leu Ser Cys 785 790 795 800 Ala Ala Ser Gly Phe Thr Phe Ser Leu Phe Thr Met Ser Trp Val Arg 805 810 815 Gln Ala Pro Gly Lys Gly Leu Glu Trp Val Ser Ala Ile Ser Gly Ser 820 825 830 Gly Gly Ser Thr Tyr Tyr Ala Asp Ser Val Lys Gly Arg Phe Thr Ile 835 840 845 Ser Arg Asp Asn Ser Lys Asn Thr Leu Tyr Leu Gln Met Asn Ser Leu 850 855 860 Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys Ala Lys Ser Thr His Leu 865 870 875 880 Tyr Leu Phe Asp Tyr Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 885 890 895 Gly Gly Ser Gly Gly Ser Ser Glu Leu Thr Gln Asp Pro Ala Val Ser 900 905 910 Val Ala Leu Gly Gln Thr Val Arg Ile Thr Cys Gln Gly Asp Ser Leu 915 920 925 Arg Ser Tyr Tyr Ala Ser Trp Tyr Gln Gln Lys Pro Gly Gln Ala Pro 930 935 940 Val Leu Val Ile Tyr Gly Lys Asn Asn Arg Pro Ser Gly Ile Pro Asp 945 950 955 960 Arg Phe Ser Gly Ser Ser Ser Gly Asn Thr Ala Ser Leu Thr Ile Thr 965 970 975 Gly Ala Gln Ala Glu Asp Glu Ala Asp Tyr Tyr Cys Asn Ser Arg Asp 980 985 990 Ser Ser Gly Asn His Val Val Phe Gly Gly Gly Thr Lys Leu Thr Val 995 1000 1005 Leu Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly 1010 1015 1020 Ser Ala Pro Ala Arg Ser Pro Ser Pro Ser Thr Gln Pro Trp Glu 1025 1030 1035 His Val Asn Ala Ile Gln Glu Ala Arg Arg Leu Leu Asn Leu Ser 1040 1045 1050 Arg Asp Thr Ala Ala Glu Met Asn Glu Thr Val Glu Val Ile Ser 1055 1060 1065 Glu Met Phe Asp Leu Gln Glu Pro Thr Cys Leu Gln Thr Arg Leu 1070 1075 1080 Glu Leu Tyr Lys Gln Gly Leu Arg Gly Ser Leu Thr Lys Leu Lys 1085 1090 1095 Gly Pro Leu Thr Met Met Ala Ser His Tyr Lys Gln His Cys Pro 1100 1105 1110 Pro Thr Pro Glu Thr Ser Cys Ala Thr Gln Ile Ile Thr Phe Glu 1115 1120 1125 Ser Phe Lys Glu Asn Leu Lys Asp Phe Leu Leu Val Ile Pro Phe 1130 1135 1140 Asp Cys Trp Glu Pro Val Gln Glu Ser Ser Ser Ser Gly Ser Ser 1145 1150 1155 Ser Ser Gly Ser Ser Ser Ser Gly Ala Pro Thr Ser Ser Ser Thr 1160 1165 1170 Lys Lys Thr Gln Leu Gln Leu Glu His Leu Leu Leu Asp Leu Gln 1175 1180 1185 Met Ile Leu Asn Gly Ile Asn Asn Tyr Lys Asn Pro Lys Leu Thr 1190 1195 1200 Arg Met Leu Thr Phe Lys Phe Tyr Met Pro Lys Lys Ala Thr Glu 1205 1210 1215 Leu Lys His Leu Gln Cys Leu Glu Glu Glu Leu Lys Pro Leu Glu 1220 1225 1230 Glu Val Leu Asn Leu Ala Gln Ser Lys Asn Phe His Leu Arg Pro 1235 1240 1245 Arg Asp Leu Ile Ser Asn Ile Asn Val Ile Val Leu Glu Leu Lys 1250 1255 1260 Gly Ser Glu Thr Thr Phe Met Cys Glu Tyr Ala Asp Glu Thr Ala 1265 1270 1275 Thr Ile Val Glu Phe Leu Asn Arg Trp Ile Thr Phe Cys Gln Ser 1280 1285 1290 Ile Ile Ser Thr Leu Thr 1295 <210> 51 <211> 1316 <212> PRT <213> Artificial Sequence <220> <223> mIL12bIL12aIL2DiaNHS76F8GMCSF-Thr <400> 51 Met Trp Glu Leu Glu Lys Asp Val Tyr Val Val Glu Val Asp Trp Thr 1 5 10 15 Pro Asp Ala Pro Gly Glu Thr Val Asn Leu Thr Cys Asp Thr Pro Glu 20 25 30 Glu Asp Asp Ile Thr Trp Thr Ser Asp Gln Arg His Gly Val Ile Gly 35 40 45 Ser Gly Lys Thr Leu Thr Ile Thr Val Lys Glu Phe Leu Asp Ala Gly 50 55 60 Gln Tyr Thr Cys His Lys Gly Gly Glu Thr Leu Ser His Ser His Leu 65 70 75 80 Leu Leu His Lys Lys Glu Asn Gly Ile Trp Ser Thr Glu Ile Leu Lys 85 90 95 Asn Phe Lys Asn Lys Thr Phe Leu Lys Cys Glu Ala Pro Asn Tyr Ser 100 105 110[[ID=​​​​​ 130 135 140 Cys Gly Met Ala Ser Leu Ser Ala Glu Lys Val Thr Leu Asp Gln Arg 145 150 155 160 Asp Tyr Glu Lys Tyr Ser Val Ser Cys Gln Glu Asp Val Thr Cys Pro 165 170 175 Thr Ala Glu Glu Thr Leu Pro Ile Glu Leu Ala Leu Glu Ala Arg Gln 180 185 190 Gln Asn Lys Tyr Glu Asn Tyr Ser Thr Ser Phe Phe Ile Arg Asp Ile 195 200 205 Ile Lys Pro Asp Pro Pro Lys Asn Leu Gln Met Lys Pro Leu Lys Asn 210 215 220 Ser Gln Val Glu Val Ser Trp Glu Tyr Pro Asp Ser Trp Ser Thr Pro 225 230 235 240 His Ser Tyr Phe Ser Leu Lys Phe Phe Val Arg Ile Gln Arg Lys Lys 245 250 255 Glu Lys Met Lys Glu Thr Glu Glu Gly Cys Asn Gln Lys Gly Ala Phe 260 265 270 Leu Val Glu Lys Thr Ser Thr Glu Val Gln Cys Lys Gly Gly Asn Val 275 280 285 Cys Val Gln Ala Gln Asp Arg Tyr Tyr Asn Ser Ser Cys Ser Lys Trp 290 295 300 Ala Cys Val Pro Cys Arg Val Arg Ser Gly Gly Gly Gly Ser Gly Gly 305 310 315 320 Gly Gly Ser Gly Gly Gly Gly Ser Arg Val Ile Pro Val Ser Gly Pro 325 330 335 Ala Arg Cys Leu Ser Gln Ser Arg Asn Leu Leu Lys Thr Thr Asp Asp 340 345 350 Met Val Lys Thr Ala Arg Glu Lys Leu Lys His Tyr Ser Cys Thr Ala 355 360 365 Glu Asp Ile Asp His Glu Asp Ile Thr Arg Asp Gln Thr Ser Thr Leu 370 375 380 Lys Thr Cys Leu Pro Leu Glu Leu His Lys Asn Glu Ser Cys Leu Ala 385 390 395 400 Thr Arg Glu Thr Ser Ser Thr Thr Arg Gly Ser Cys Leu Pro Pro Gln 405 410 415 Lys Thr Ser Leu Met Met Thr Leu Cys Leu Gly Ser Ile Tyr Glu Asp 420 425 430 Leu Lys Met Tyr Gln Thr Glu Phe Gln Ala Ile Asn Ala Ala Leu Gln 435 440 445 Asn His Asn His Gln Gln Ile Ile Leu Asp Lys Gly Met Leu Val Ala 450 455 460 Ile Asp Glu Leu Met Gln Ser Leu Asn His Asn Gly Glu Thr Leu Arg 465 470 475 480 Gln Lys Pro Pro Val Gly Glu Ala Asp Pro Tyr Arg Val Lys Met Lys 485 490 495 Leu Cys Ile Leu Leu His Ala Phe Ser Thr Arg Val Val Thr Ile Asn 500 505 510 Arg Val Met Gly Tyr Leu Ser Ser Ala Gly Gly Gly Gly Ser Gly Gly 515 520 525 Gly Gly Ser Gly Gly Gly Gly Ser Ala Pro Thr Ser Ser Ser Thr Ser 530 535 540 Ser Ser Thr Ala Glu Ala Gln Gln Gln Gln Gln Gln Gln Gln Gln Gln 545 550 555 560 Gln Gln His Leu Glu Gln Leu Leu Met Asp Leu Gln Glu Leu Leu Ser 565 570 575 Arg Met...

Claims

1. A pharmaceutical composition comprising a protein and an immune checkpoint inhibitor, wherein the protein is a fusion protein, and the fusion protein comprises cytokines IL12, IL2 and GMCSF; The immune checkpoint inhibitor is a PD1 antibody; The protein is a single-chain protein shown in SEQ ID NO:27, SEQ ID NO:39, SEQ ID NO:29, SEQ ID NO:46 or SEQ ID NO:52, or a dimer composed of a first polypeptide chain shown in SEQ ID NO:53 and a second polypeptide chain shown in SEQ ID NO:

57.

2. A kit comprising the pharmaceutical composition according to claim 1.

3. Use of the pharmaceutical composition according to claim 1 or the kit according to claim 2 in the preparation of a drug for treating melanoma, breast cancer and / or lung cancer.

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