EP0378666A1 - Analogues de l'interleukine ii - Google Patents

Analogues de l'interleukine ii

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Publication number
EP0378666A1
EP0378666A1 EP89908361A EP89908361A EP0378666A1 EP 0378666 A1 EP0378666 A1 EP 0378666A1 EP 89908361 A EP89908361 A EP 89908361A EP 89908361 A EP89908361 A EP 89908361A EP 0378666 A1 EP0378666 A1 EP 0378666A1
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EP
European Patent Office
Prior art keywords
amino acid
amino acids
leu
lys
substitution
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP89908361A
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German (de)
English (en)
Other versions
EP0378666A4 (en
Inventor
Bruce W. Altrock
Thomas C. Boone
Robert A. Goldman
William C. Kenny
Yitzhak Stabinsky
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Amgen Inc
Original Assignee
Amgen Inc
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Publication date
Application filed by Amgen Inc filed Critical Amgen Inc
Publication of EP0378666A1 publication Critical patent/EP0378666A1/fr
Publication of EP0378666A4 publication Critical patent/EP0378666A4/en
Withdrawn legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/52—Cytokines; Lymphokines; Interferons
    • C07K14/54—Interleukins [IL]
    • C07K14/55—IL-2
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates generally to the manipulation of genetic materials and, more
  • Interleukin II a glycoprotein with a molecular weight of approximately 15,000, is a member of a group of proteins, called lymphokines, that control the body's immune response.
  • IL-2 is produced by certain white blood cells, lectin- or antigen-activated T cells, and plays a central role in the body's immune system as a lymphocyte regulating molecule.
  • IL-2 has been reported to enhance thymocyte mitogenesis, to stimulate long-term in vitro growth of activated T-cell clones, to induce cytotoxic T-cell reactivity, to modulate immunological effects on
  • activated B cells and lymphokine activated cells to induce plaque-forming cell responses in cultures of nude mouse spleen cells, and to regulate production of gamma interferon. It also augments natural killer cell activity and mediates the recovery of the immune
  • lymphocytes in selected immunodeficient states.
  • IL-2 is used to maintain cultures of functional monoclonal T-cells to study the molecular nature of T-cell differentiation, and to help elicit the mechanism of differentiated
  • IL-2 has application in both research and the treatment of neoplastic and
  • IL-2 asserts its effect by binding to a
  • the IL-2 molecule has become a focal point for studying receptor-effector interactions that modulate cell proliferation in the immune response.
  • the high affinity (K D ⁇ 10 -11 M) receptor reponsible for mediating the effect of IL-2 on target cells consists of two distinct membrane-bound proteins of size 55 kD (p55 or Tac) and 75 kD (p75); each of these two proteins can act by itself as an apparent low affinity (K D ⁇ 10 -8 M) receptor for IL-2, and both are required for IL-2 activity. This suggests that IL-2 must bind both p55 and p75 to form a trimeric complex for activity, and by inference, that IL-2 must have two separate receptor binding sites.
  • Taniguchi, T., et al., Nature, 302: 305-310 (1983) described the sequence analysis, cloning, and expression of a complementary DNA coding for human IL-2, cloned from a cDNA library prepared from partially purified IL-2 mRNA from the Jurkat leukemia cell line.
  • IL-2 was proposed to comprise 133 amino acid residues and to have a calculated molecular weight of about
  • Taniguchi described the cloning procedures and the expression of the cDNA for IL-2 in cultured monkey COS cells. The publication states that expression of the IL-2 cDNA in E. coli had not yet been
  • the subject invention relates to IL-2 analogs having modified receptor domains, and analogs having stabilized IL-2 structure.
  • the subject invention also relates to IL-2 analogs which have been modified to permit the attachment of a ligand. More particularly the subject invention relates to a polypeptide product of the expression in a host cell of a manufactured gene, the polypeptide having an amino acid sequence
  • Leu Gin Met lie Leu Asn Gly lie Asn Asn
  • the invention also encompasses: IL-2 analogs wherein one, two, three or more original amino acids in any helix or in any helices of IL-2 (advantageously helix A and/or helix F) have been replaced by substitution amino acids which maintain or reduce the amphiphilicity of the helix or helices; analogs wherein one, two, three or more original amino acids in helix A, B, B' and/or E have been replaced by substitution amino acids, each having a different charge from the original amino acid it replaces; and analogs wherein one, two, three or more original amino acids in helix A, B, B', C, D, E and/or F have been replaced by substitution amino acids, each having a greater preference for alpha-helical structure then the original amino acid it replaces.
  • the invention also relates to manufactured DNA sequences encoding such polypeptides. Further, the invention relates to
  • Figure 1(a) represents the alpha carbon backbone of IL-2.
  • Figure 1(b) is a schematic stereo drawing of IL-2; helices are represented as cylinders.
  • Figure 2 is a schematic drawing showing a possible mode of interaction of IL-2 with its receptors.
  • Figure 3 shows the IL-2 structure and the positions of relevant amino acids.
  • IL-2 Interleukin II
  • site specific modifications of the proposed receptor binding domains of naturally occurring IL-2 are made, and alterations which stabilize IL-2 helix structure and the overall IL-2 structure, are made.
  • an amino acid such as an odd cysteine is incorporated into IL-2 at a location far removed from the proposed receptor binding domains but accessible to chemical reaction with other molecules.
  • the base sequence includes one or more codons selected from among alternative codons specifying the same amino acid on the basis of preferential expression characteristics for the codon in a projected host microorganism, e.g., E. coli (see Alton et al., PCT application WO 83/04053).
  • manufactured genes include those wherein there is provided the nucleotide bases for a codon specifying an additional amino acid residue in the polypeptide coded for, which facilitates the direct expression in E. coli organisms (e.g., an initial Met residue).
  • the base sequence of codons include those wherein there is provided the nucleotide bases for a codon specifying an additional amino acid residue in the polypeptide coded for, which facilitates the direct expression in E. coli organisms (e.g., an initial Met residue).
  • specifying the desired polypeptide is preceded by and/or followed by and/or includes one or more sequences of bases facilitating formation of expression vectors or generation of new structural genes for polypeptide analogs, i.e., sequences of bases providing for selected restriction endonuclease cleavage sites on one or both ends of the structural gene or at intermediate positions therein, and sequences providing a site for ribosome binding, e.g. CAA GGA GGT.
  • Also provided by the present invention are manufactured genes capable of directing the microbial expression of IL-2 analogs which differ from the
  • manufactured DNA sequences are inserted into viral or circular plasmid DNA vectors to form hybrid vectors and the hybrid vectors are employed to transform microbial hosts such as bacteria (e.g., E. coli), yeast cells, or mammalian cells in culture.
  • bacteria e.g., E. coli
  • yeast cells e.g., yeast cells
  • mammalian cells e.g., mammalian cells
  • microorganisms are thereafter grown under appropriate nutrient conditions and express the polypeptide products of the invention.
  • compositions comprising effective amounts of polypeptide products of the invention together with suitable diluents, adjuvants and/or carriers useful in IL-2 therapy.
  • the term "manufactured" as applied to a DNA sequence or gene shall designate a product chemically synthesized by assembly of nucleotide bases, synthesized by site-directed mutagenesis, or derived from the biological replication of a product thus synthesized.
  • the term is exclusive of products "synthesized” by cDNA methods or genomic cloning methodologies which involve materials which are of biological origin.
  • substitution amino acid means an amino acid which replaces the naturally occurring (“original”) amino acid, and which is different from the original amino acid.
  • antibodies are provided which specifically bind the polypeptides of the subject invention but which do not cross-react with naturally occurring IL-2. These antibodies can be tagged using methods known to those skilled in the art.
  • Arginine Arg
  • Asparagine Asn
  • Aspartic acid Asp
  • Cysteine Cys
  • Glutamine Gin
  • Glutamic acid Glu
  • Glycine Gly; Histidine, His; Isoleucine, He; Leucine, Leu; Lysine, Lys; Methionine, Met; Phenylalanine, Phe; Proline, Pro; Serine, Ser; Threonine, Thr; Tryptophan, Trp; Tyrosine, Tyr; Valine, Val.
  • nucleotide bases A for adenine; G for guanine; T for thymine; U for uracil; and C for cytosine.
  • IL-2 is an alpha-helical protein (Fig. 1), Brandhuber et al.,
  • Cys 58 of the disulfide are helix C, residues 66 to 78, and D, residues 83 to 101; following Cys 105 is a short, apparently helical stretch E, residues 106 to 113, which leads into the carboxyl-terminal helix F, residues 117 to 133. There are no apparent segments of 8-secondary structure in the molecule. The overall helical content of about 65 percent is in good agreement with estimates based on circular dichroism.
  • disulfide between Cys 58 and Cys 105 links two extending loops that connect the helices across the "top” (in the orientation of Fig. 1) of the molecule.
  • Helices B, C, D, and F form an antiparallel alpha helical bundle which differs significantly from the classical four-helix bundle represented by
  • cytochrome c ' cytochrome b 562 , and myohemerythrin.
  • the packing regions of the helices are shorter, involving only three to four turns of helix, while classical four- helix bundles usually have at least five turns in each helix. Further, the packing angles all fall in the range of 25o to 30o, and hence are somewhat larger than the average of approximately 18o found in classical four-helix bundles.
  • Murine IL-2 is expected to have a similar structure to recombinant human IL-2 beginning with helix A and including the proline-induced bend in helix
  • the murine and human IL-2 sequences have 64 percent overall homology.
  • the amino acid sequence of the mature murine protein is identical to the human sequence for the first seven residues, and then has one or more insertions, a total of 15 amino acids, relative to human, including a
  • IL-2 receptor binding do not discriminate between those involving a p55-(IL-2) interaction, a p75-(IL-2)
  • Antibodies to peptides that cross-react with IL-2 have been used to map global regions in the IL-2 sequence likely to be important in receptor binding.
  • Kuo and Robb have presented evidence
  • deletion of residues 1 to 10 of human IL-2 reduces induction of proliferation of murine CTLL-2 cells by only 30 to 50 percent, whereas deletion of residues 1 to 20 (the amino terminus including helix A) abolishes activity
  • Helices B, C, D, and F form a structural scaffold, and that helices A, B' and part of B, and E form the receptor binding sites of IL-2 (Fig. 2).
  • the involvement of helix E is suggested primarily by its spatial accessibility and its proximity to regions of the molecule probably involved in receptor interactions.
  • IL-2 binds, through a high affinity receptor, to T-cells but will also bind and activate other immune system cells through lower affinity receptors. It is believed that activation of these other cells
  • IL-2 analogs wherein amino acids, advantageously hydrophilic amino acids in helices in the receptor binding domain, are replaced by amino acids having a different charge (e.g. replacing an amino acid having a positively charged side chain by an amino acid having a negatively charged side chain, or by an amino acid having an uncharged side chain, see below). Also encompassed by the subject invention are analogs wherein one or more amino acids which have a preference for ⁇ -helical structure (see Chou and Fasman, Annu. Rev. Biochem. 47, 251(1978).
  • the helices of IL-2 are amphiphilic helices (see Kaiser et al., PNAS, 80, 1137-1143(1983) and Kaiser et al., Science, 223, 249-255 ( 1984).
  • This amphiphilic helical structure is shared by several cytotoxic peptides such as mellitin, pardoxin, and maganins. More specifically, the F helix is very amphiphilic and some of the amino acids in the F helix do not have a strong preference for the ⁇ -helical structure, and it is believed that the interaction of the hydrophobic face of the F helix with the C and D helices provides the energy required to maintain the F helix sequence in its helical form.
  • IL-2 analogs have been constructed containing altered helix sequences in which the amino acid replacements were selected to contain residues with a greater preference for a ⁇ -helical structure (e.g. Asn ⁇ Gin, Trp ⁇ Phe, Ser ⁇ Gin) and consequently the helices do not require strong
  • amphiphilic interactions to maintain their helical structure and thus the amphiphilicity of the helix can be maintained (for example an amino acid having a
  • hydrophilic side chain being changed to a different amino acid having a hydrophilic side chain - the three
  • substitutions noted above maintain amphiphilicity) or altered (for example by replacement of an amino acid having a hydrophilic side chain with one having a
  • Analogs have been constructed to which other molecules can be covalently attached without damaging activity. These analogs are used to attach toxins, reporter groups, or antiviral or other therapeutic compounds which lead to the development of IL-2
  • the subject invention includes alterations where an additional amino acid is inserted between existing amino acids as an alternative to replacement of an existing amino acid.
  • Hydrophilic amino acids in alpha helical regions (designated A, B', B and E in Figures 1, 2 and 3), implicated in high affinity receptor interactions, are among the candidate locations for amino acid
  • stimulation process allows for a greater therapeutic index for the material (e.g. by a reduction in
  • IL-2 undesirable side effects which results from stimulation of one cell type while at the same time retaining the ability of IL-2 to stimulate an appropriate effector cell type which, in turn, limits disease).
  • certain alterations in helices in the receptor binding domain produce IL-2 analogs having a reduced capacity to induce induction of lymphokines such as INF- ⁇ , IL-1 and TNF, but having equivalent biological activity, relative to (Ala 125 ) IL-2 or natural IL-2.
  • the short E helix is involved in receptor binding. Therefore, the E helix is an excellent target for mutations that alter receptor binding.
  • FIG. 3 shows the IL-2 structure, the proposed receptor binding domains, and positions of the relevant amino acids.
  • Receptor - effector e.g. IL-2
  • the E helix has no positively charged amino acid side chains and three negatively charged side chains (Glu 106, Asp 109 and Glu 110) .Alterations of the type of charge in the E helix of IL-2 thus alters binding efficiency.
  • An Asp 109 ⁇ Lys 109 substitution alters receptor binding with a single mutation. Asp 109 protrudes from the center of the E helix and is a conserved amino acid.
  • the B' helix has one negatively charged amino acid side chain (Glu 52), and four positively charged side chains (Lys 48, Lys 49, Lys 54, and
  • the A helix is involved in receptor binding.
  • the A helix has one negatively charged amino acid side chain (Glu 115), and one positively charged side chain (His 16).
  • Glu 115 amino acid side chain
  • His 16 positively charged side chain
  • the F helix is an amphiphilic helix. Some of the amino acids in the F helix do not have a strong preference for the ⁇ -helical structure and it is
  • IL-2 analogs have been constructed containing altered F helix sequences in which the amino acid replacements were selected to contain residues with a higher
  • attachment of a ligand include the carboxyl terminus as well as portions of the molecule with surface exposure chosen such as to minimally perturb the structure of active IL-2.
  • a cysteine residue provides a sulphydryl group which can be chemically conjugated to:
  • toxins for selective cell killing, in vitro or in vivo.
  • Conjugation of IL-2 to cytotoxic agents should direct the toxins to cells which present IL-2 receptors. These conjugations are useful in treating certain leukemias, transplant rejections, autoimmune or altered immune states, or other cell populations of pathological significance;
  • drugs directed therapeutic delivery, e.g.
  • AZT for AIDS.
  • Cells that are susceptible to HIV infection are, for the most part, the cells that carry IL-2 receptors.
  • antibodies or mitogens selective cell targeting, e.g. helper T cells using OKT 4 or
  • recombinant IL-2 gene allows for the specific chemical conjugation of other molecules to IL-2, by reaction with the free sulphydryl group(s), in a manner that does not affect the binding of IL-2 to its cell surface receptors.
  • the carboxy terminus region of IL-2 is not involved in receptor binding and is a good location for the incorporation of an odd cysteine.
  • Leu 132 was chosen because it is close to the C-terminus (next to last amino acid) and it is an unconserved residue when comparing human, bovine, and murine IL-2 sequences.
  • An analog in which cysteine is simply added to the carboxy end of IL-2 (Cys 134 ) IL-2) also accomplishes the goal of incorporating an odd cysteine at the C-terminus.
  • the bovine and murine sequences contain an insertion between amino acids 80 and 81.
  • the amino acids are part of a four amino acid loop (amino acids 79 to 82) that connects the C and D helices of IL-2. This observation coupled with the fact that this region of the molecule is far removed from the proposed receptor binding domains and other Cys residues in the molecule, makes this an ideal location for an insertion of an odd cysteine residue. Substitutions such as Leu 80 ⁇ Cys 80 are also encompassed by the invention.
  • a Lys 8 ⁇ Cys 8 substitution at a nonconserved residue provides a reactive group near the amino
  • IL-2 Some structural component or combination of components of IL-2 have retained or lost biological properties of intact IL-2 together with the ability to bind IL-2 receptor (s). Such a peptide is useful in place of intact IL-2 or as an antagonist of its action(s).
  • Peptides for this application include at least one of the A, B, B' and E helices, for example, the invention includes A and E helical regions which preserve their own internal symmetry as species isolated and apart from the conformational constraints of the intact parent
  • Such isolated structures bind to components of the IL-2 receptor and either do or do not have IL-2 biological activity. Such structures retain activity on only a subset of IL-2 responsive cells allowing for greater precision in manipulating the IL-2 response. Such isolated structures have lost biological activity but function as competitive inhibitors of IL-2 binding and are useful in antagonizing physiological states involving a stimulant effect of IL-2. Such structures function to up or down regulate IL-2 receptors and, thereby influence cellular receptivity to IL-2. V. Additional Alterations
  • Additional analogs which are encompassed by the present invention include the analogs of IL-2 noted above further characterized by the presence of one or more of the following alterations in the amino acid sequence of naturally-occurring IL-2.
  • IL-2 and IL-2 analogs are very hydrophobic proteins and as such have a propensity to bind pyrogens which are also hydrophobic.
  • These peptides can be formulated in a stable, monomeric form at acidic pH values. Under these conditions, pyrogens tend to form higher molecular weight aggregates, even though the monomeric molecular weight of pyrogens is comparable to that of IL-2.
  • manufacturing procedures which fractionate proteins on the basis of size are used to separate monomeric IL-2 from aggregated forms of
  • pyrogens A suitable procedure for carrying out this step is by ultrafiltration through YM-30 (Amicon) membranes. Repeated dilution and ultrafiltration can be used to enhance the yield of IL-2. Glucose, mannitol, or another bulking agent, can be added as a toxicity modifier and the desired concentration of the IL-2 can be obtained by concentration by ultrafiltration or by dilution with an appropriate buffer. Pyrogens can also be separated from monomeric IL-2 by size exclusion chromatography, e.g. using Sephadex G-75. Detergents
  • the method of the subject invention is easy to scale up and is very cost effective.
  • the following examples illustrate practice of the invention in the manufacture of the DNA sequences coding for microbial expression of IL-2 and polypeptide analogs thereof. Also illustrated is the construction of expression vectors for microbial expression of desired polypeptides.
  • This example is directed to the procedure employed in the synthesis of oligonucleotide sequences employed to manufacture the IL-2 analog genes according to the invention.
  • Oligonucleotide sequences were synthesized using a four-step procedure with several intermediate washes. Syntheses were performed on Applied Biosystems (ABI) Model 380 automated synthesizers using ABI
  • the final oligonucleotide chain was treated with fresh concentrated ammonia at room temperature for 2.0 hours. After decanting the solution from the
  • the concentrated ammonia solution was heated at 60oC for 16 hours in a sealed tube.
  • Each oligonucleotide solution was extracted with 1-butanol and ethyl ether and the concentration of each solution was determined with a spectrophotometer (260nm).
  • 5.0 OD units of each oligonucleotide were dried down for preparative electrophoresis and loaded into a 15% p ⁇ lyacrylamide, 7 molar urea gel. After electrophoresis, the product band was visualized by UV shadowing, cut from the gel, extracted and then desalted on a G-50 Sephadex column to yield the purified oligonucleotide.
  • This example relates to the use of recombinant methods to generate analogs of IL-2.
  • Site directed mutagenesis procedures according to Souza, et al., published PCT Application No. WO 85/00817, published
  • Oligonucleotide site-directed mutagenesis was performed by cloning the IL-2 region from Xbal to BamHl, from expression vector pCFM 536 IL-2 into both M13mp10 and M13mp11, and the single-stranded phage DNA was isolated as for DNA sequencing.
  • pCFM536 see U.S. Patent 4,710,473 hereby incorporated by reference
  • any suitable expression vector could have been used. This DNA was mixed with the synthetic
  • plaques contained the IL-2 sequence with the desired base changes. These plaques were identified by lifting plaques onto nitrocellulose filters, and then
  • IL-2 analogs essentially free of pyrogenic substances and endotoxins. This was accomplished by further purification of the molecule.
  • the protein was oxidized in the presence of Cu 2+ , concentrated and chromatographed on Sephadex G-75 equilibrated with 1% laurate/25 mM Tris/ 5% ethanol, pH 8.7. Those fractions containing monomeric forms of IL-2 were pooled and the protein was precipitated by addition of an equal volume of ethanol. The pellet was collected by centrifugation, washed with 50% ethanol, and then solubilized in
  • This example relates to the activity of analogs generated in Example 2.
  • IL-2 analogs were generated that differ from the native sequence of IL-2 by two amino acids. All analogs tested have Ala at position 125.
  • (Ala 125 ) IL-2 (which differs from the native sequence of IL-2 by one amino acid) was used as a positive control for these experiments.
  • (Asp 51 )IL-2 and (Glu 48 ) IL-2 are molecules with amino acid changes in the putative receptor-binding B' domain of IL-2. Specifically, in (Asp 51 )IL-2 the neutral threonine at position 51 was replaced by a negatively charged aspartic acid, and in (Glu 48 ) IL-2 the positively charged lysine was replaced by the negatively charged glutamic acid.
  • (Ala 125 )IL-2 as T cell mitogens and as inducers of LAK cells.
  • (Glu 48 ) IL-2 consistently induced less IFN-gamma production from hPBL cultures than did an equal concentration of (Ala 125 ) IL-2.
  • (Asp 51 ) IL-2 was less effective than
  • (Asp 51 ) IL-2 was equal to that of (Ala 125 ) IL-2 while induction by (Glu 48 ) IL-2 was approximately 50% as effective.
  • (Gly 47 ) IL-2 had only 2% to 20% of the biological activity of (Ala 125 ) IL-2 as predicted.
  • the first step involved the introduction of an Eco Rl restriction site at a region in the gene corresponding to the Glu 116 Phe 117 sequence. This was accomplished by site directed mutagenesis using the primer shown below which codes for the desired change in DNA sequence while leaving the encoded amino acid sequence intact. Mutagenesis Primer for the Introduction of Eco Rl Site

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Abstract

L'invention concerne des analogues de l'interleukine II et des séquences d'ADN comprenant des gènes structuraux codant pour de tels analogues qui diffèrent des formes naturelles en terme d'identité et/ou de position d'un ou de plusieurs acides aminés.
EP19890908361 1988-07-05 1989-07-05 Interleukin ii analogs Withdrawn EP0378666A4 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US21499888A 1988-07-05 1988-07-05
US214998 1988-07-05

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EP0378666A1 true EP0378666A1 (fr) 1990-07-25
EP0378666A4 EP0378666A4 (en) 1992-01-22

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JP (1) JPH03500415A (fr)
AU (1) AU627477B2 (fr)
WO (1) WO1990000565A1 (fr)

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EA034925B1 (ru) * 2014-08-11 2020-04-07 Делиниа, Инк. Модифицированные варианты il-2, которые селективно активируют регуляторные т-клетки, для лечения аутоиммунных заболеваний
BR112017021696A2 (pt) 2015-04-10 2018-07-10 Amgen Inc muteínas de interleucina-2 para a expansão de células t regulatórias
US20170204154A1 (en) 2016-01-20 2017-07-20 Delinia, Inc. Molecules that selectively activate regulatory t cells for the treatment of autoimmune diseases
CN110167957A (zh) 2016-11-08 2019-08-23 德里尼亚公司 用于治疗自身免疫疾病的il-2变体
KR102545250B1 (ko) * 2020-03-31 2023-06-21 한미약품 주식회사 신규한 면역 활성 인터루킨 2 아날로그

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See also references of WO9000565A1 *

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EP0378666A4 (en) 1992-01-22
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AU627477B2 (en) 1992-08-27
JPH03500415A (ja) 1991-01-31

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