采用定向分子进化方法构建改形抗CD28重链单域抗体及抗人卵巢癌三特异抗体的构建与表达
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摘要
本研究的第一个目的是验证一种具有实际意义，简化易行构建改形抗体的新方法。与以往其它方法不同的是：该方法采用实验定向分子进化方法，不需要对抗体序列进行空间模拟，以确定人源抗体受体的FRs以及受体FRs上的哪个氨基酸残基需要进一步突变；该改形方法将抗体的改形及亲和力成熟于同一步骤完成，从而节省构建的时间及劳动强度。利用该方法构建了改形抗CD28重链单域抗体。根据鼠源抗CD28VH氨基酸序列，从GenBank中得到两个与该序列最同源的人源抗体，选用其中之一作为改形抗体的主框架FRs区。在保留鼠源CDRs氨基酸残基的同时，对主框架FRs一些氨基酸残基进行可选择替换突变，可选择替换原则根据对原始鼠源序列；鼠源抗体所在Kabat分类中的种属序列；两个人源抗体序列及人源抗体所在Kabat分类中的种属序列的比较而确定。选择大肠杆菌喜好密码子，推导出其基因序列；合成大小不同的核苷酸片段，采用重叠PCR扩增得到改形抗体全长基因。在合成片段时，将突变位点要被替换氨基酸残基和替换氨基酸残基的核苷酸采用简并合成，使二者都有机会在完整基因中出现。为了进一步提高改形抗体基因数目，在重叠PCR扩增改形抗体全长基因时，采用高浓度Mg~(2+)和Taq DNA聚合酶，以便进一步随机引入突变。利用PCR产物构建了一个改形抗CD28VH抗体库，经过三轮淘选，获得了具有高结合活性的改形抗体基因。进一步将两个改形基因与c-myc及人IgG3'CL铰链区基因融合在大肠杆菌BL21(DE3)中表达，复性后的融合表达蛋白仍表现为具有较高与CD28分子的结合活性。这一结果表明，该构建改形抗体方法是有效和可行的，通过一步就可以获得满意的改形抗体。
本研究第二个目的是构建及表达一种新的三特异抗体：抗人卵巢癌scFv×改形抗CD3 scFv×改形抗CD28VH。构建的三特异抗体有两种类型：环形三特异抗体；非环形三特异抗体。在三特异抗体构建中使用了不同的连接肽序列(interlinker)，这些序列的使用，引入了一些不同的生物学功能。两种类型的三
博土论文 中文摘要
特异抗体的基因在大肠杆菌BLZI pE3）中表达后，测定了表达蛋白与CD28分子
的结合活性。
The first aim of this study was to demonstrate a new and practical method for constructing reshaping antibodies. Different from other methods, it was not necessary to carry out modeling the sequences for determining the human acceptor and which amino acid residues in human acceptor FRs should be substituted, and reshaping and enhancing the antigen binding affinity shared one procedure at the same time. The reshaping anti-CD28 single domain antibodies were constructed by using this method. According to the amino acid sequence of a mouse anti-human CD28 monoclonal antibody VH, two most homologous sequences of human antibodies were searched from GenBank and one of them was used as a main framework residues for constructing the reshaping antibody FRs. The mouse anti-CD28 VH CDRs were still kept, but some amino acid residues
in human FRs were substituted according to the comparison of searched human antibody sequences, original mouse antibody sequence, and their consensus sequences. In order to increase diversity of the gene, the amino acid residues planned to substitute were reserved in synthesizing the gene sequences by code degeneracy. Also, when the synthesized nucleotide fragments in different length were assembled and amplified to intact genes by overlap PCR, Taq DNA polymerase and high Mg2+ concentration were used for inducing more mutations randomly. A phage display library was constructed by using the PCR products. The genes of reshaping anti-CD28 VH that had high antigen binding activity were selected after three round panning selections. Two genes were fused with c-myc tag and the hinge region of human IgGS'CL gene, and expressed in E. coli BL21(DE3). The antigen binding affinity and specificity of renatured fusion protein were determined. The results suggested that the procedure we used was feasible and efficient, with which it could be gotten an ideal reshaping antibody in one step and so save the time and labor.
The second aim of this study was to construct and express a novel trispecific antibody fragment: anti-human Ovarian Carcinoma scFvX reshaping anti-CD3 scFvX reshaping anti-CD28VH. Two types of the trispecific antibodies were constructed: one was constructed in a cycle formation; another was in a broken cycle formation. Different interlinkers were used in the construction, and some interlinkers could induce some biological properties beside the connection function. The molecular masses of the trispecific antibodies were about 84 kDa, which were suitable for deliver and tumor targeting. The genes of the two type trispecific antibodies were expressed in E.coli BL21(DE3), and the CD28 binding affinity of the expression proteins were determined.
本研究第二个目的是构建及表达一种新的三特异抗体：抗人卵巢癌scFv×改形抗CD3 scFv×改形抗CD28VH。构建的三特异抗体有两种类型：环形三特异抗体；非环形三特异抗体。在三特异抗体构建中使用了不同的连接肽序列(interlinker)，这些序列的使用，引入了一些不同的生物学功能。两种类型的三
博土论文 中文摘要
特异抗体的基因在大肠杆菌BLZI pE3）中表达后，测定了表达蛋白与CD28分子
的结合活性。
The first aim of this study was to demonstrate a new and practical method for constructing reshaping antibodies. Different from other methods, it was not necessary to carry out modeling the sequences for determining the human acceptor and which amino acid residues in human acceptor FRs should be substituted, and reshaping and enhancing the antigen binding affinity shared one procedure at the same time. The reshaping anti-CD28 single domain antibodies were constructed by using this method. According to the amino acid sequence of a mouse anti-human CD28 monoclonal antibody VH, two most homologous sequences of human antibodies were searched from GenBank and one of them was used as a main framework residues for constructing the reshaping antibody FRs. The mouse anti-CD28 VH CDRs were still kept, but some amino acid residues
in human FRs were substituted according to the comparison of searched human antibody sequences, original mouse antibody sequence, and their consensus sequences. In order to increase diversity of the gene, the amino acid residues planned to substitute were reserved in synthesizing the gene sequences by code degeneracy. Also, when the synthesized nucleotide fragments in different length were assembled and amplified to intact genes by overlap PCR, Taq DNA polymerase and high Mg2+ concentration were used for inducing more mutations randomly. A phage display library was constructed by using the PCR products. The genes of reshaping anti-CD28 VH that had high antigen binding activity were selected after three round panning selections. Two genes were fused with c-myc tag and the hinge region of human IgGS'CL gene, and expressed in E. coli BL21(DE3). The antigen binding affinity and specificity of renatured fusion protein were determined. The results suggested that the procedure we used was feasible and efficient, with which it could be gotten an ideal reshaping antibody in one step and so save the time and labor.
The second aim of this study was to construct and express a novel trispecific antibody fragment: anti-human Ovarian Carcinoma scFvX reshaping anti-CD3 scFvX reshaping anti-CD28VH. Two types of the trispecific antibodies were constructed: one was constructed in a cycle formation; another was in a broken cycle formation. Different interlinkers were used in the construction, and some interlinkers could induce some biological properties beside the connection function. The molecular masses of the trispecific antibodies were about 84 kDa, which were suitable for deliver and tumor targeting. The genes of the two type trispecific antibodies were expressed in E.coli BL21(DE3), and the CD28 binding affinity of the expression proteins were determined.
引文
1．郭亚军主编，(2000)：“细胞生物学与肿瘤免疫学研究进展”2000年6月第1版，军事医学科学出版社，6ISBN 7-80121-228-2。
2．刘喜富，萧飒，顾征等．(1996)．抗人CD3单链抗体及改形单域抗体的表达．中国科学(C辑)．26：428-435．
3．蒋欣(1998)：抗人CD3改形单链抗体及抗人卵巢癌双特异单链抗体的构建与表达。博士论文，中国科学院遗传研究所。
4. Abraltmsen L, Tom Jeffrey, Burnier J, et al., (1991): Engineering subrilisin and its substrates for efficient ligafon of peptide bonds in aqueous solution. Biochemistry, 30:4151-4159.
5. Baca M, Scanlan TS, Stephenson RC, et al.(1997~a): Phage display of a catalytic antibody to optimize affinity for transition-state analog binding. Proc. Natl. Acad. Sci. USA, 94,10063-10068.
6. Baca M, Presta L, O'Connor SJ, et al. (1997~b): Antibody humanization using monovalent phage display. J. Biol. Chem. 272:10678-10684.
7. Bartel DP & Szostak JW (1993): Isolation of new ribozymes from a large pool of random sequences. Science, 261:1411-1418.
8. Baskar S, Ostrand-Rosenberg S, Nabavi N, et al. (1993). Constitutive expression of B7 restores immunogenicity of tumor cells expressing truncated major histocompatibility complex class Ⅱ molecules. Proc. Natl. Acad. Sci. USA 90:5687-5690.
9. Bock LC, Griffin LC, Latham JA, et al.(1992): Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Nature,355(6360):564-566.
10. Boder ET & Wittrup KD (1997): Yeast surface display for screening combinatorial polypeptide libraries. Nature Biotechnol. 15,553-557.
11. Boder ET, Midelfort KS & Wittrup KD (2000): Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity. Proc. Natl. Acad. Sci. USA, 97(20): 10701-10705.
12. Bohlen H, Hopff T, Manzke O, et al. (1993): Lysis of maligant B cells from patients with B-chronic lymphocytic leukemia by autologous T cells activated with CD3×CD19 bispceific antibodies in combination with bivalent CD28 antibodies. Blood, 82:1803-1812.
13. Bohlen H, Manzke O, Titzer S, et al. (1997): Prevention of Epstein-Barr virus-induced human B-cell lymphoma in severe combined immunodeficient mice treated with CD3×CD19 bispecific antibodies,CD28 monospecific antibodies, and autologous T cells. Cancer Res., 57:1704-1709.
14. Boulianne GL, Hozumi N & Shulman MJ (1984): Production of functional chimeric mouse/human antibody. Nature. 312, 643-646.
15. Bruggemann M and Taussig MJ (1997): Production of human antibody repertoires in transgenic mice. Curr. Opin. Biotechnol. 8:455-458.
16. Cai X and Green A (1995): Anti-melanoma antibodies from melanoma patients immunized with genetically modified autologus tumour cells: selection of specific antibodies from single-chain Fv fusion phage libraries. Proc. Natl. Acad. Sci. USA 92:6537-6541.
17. Carter,P.,Presta,L.,Gorman, C.M. et al.(1992): Humanization of an anti-p185HER2 antibody for human cancer therapy. Proc. Natl. Acad. Sci. USA, 89,4285-4289.
18. Cheetham MT, Hale G, Waldmann H, et al.(1998): Crystal structures of a rat anti-CD_(52) (CAMPATH-1) therapeutic antibody Fab fragment and its humanized counterpart. J. Mol. Biol. 284,85-99.
19. Chen K &Amold FH (1991): Enzyme engineering for nonaqueous solvents: random mutagenesis to enhance activity of subtilisin E in polar organic media. Biote-chnology, 1991,9(11): 1073-1077.
20. Chen K, & Amold FH (1993): Tuning the activity of an enzyme for unusual environments: sequential random mutagenesis of subtilisin E for catalysis in dimethylformamide. Proc. Natl. Acad. Sci. USA, 90(2):5618-5622.
21. Chen L, Linsley PS & Hellstrom KE (1993): Costimulation of T cells for tumor immunity.Immunol. Today. 14:483-486.
22. Clackson T, et al.(1991): making antibody fragments using phage display libraries. Nature,352,624-628.
23. Clark EA, Martin PJ, Hansen JA, et al. (1983): Evolution of epitopes on human and nonhuman primate lymphocyte cell surface antigens. Immunogenefics, 18,599-615.
24. Coulie PG, Weynants P, Lehmann F, et al. (1993): Genes coding for tumor antigens recognized by human cytolytic T lymphocytes. J. Immunother. 14:104-109.
25. Daugherty PS, Chen G, Iverson BL, et al.(2000): Quantitative analysis of the effect of the mutation frequency on the affinity maturation of single chain Fv antibodies. Proc. Natl. Acad.Sci. USA,97(5),2029-2034.
26. Deborah JL, Thersa LW, Jeffrey AB, et al. (1996): CD28/B7 system of T cell costimulation. Annu. Rev. Immunol. 14: 233-258.
27. Demanet C, Brissinck J, Leo O, et al.(1996): Bispecific antiboddy-mediated immunotherapy of the BCL1 lymphoma:increasd efficacy with multiple injections and CD28-induced costimulation. Blood, 87: 4390-4398.
28. Dong H, Zhu G, Tamada K, et a1.(1999): B7-H1, a third member of the B7 family, co-stimulates T-cell proliferation and interleukin-10 secretion. Nat. Med., 5:1365-1369.
29. Dunn IS (1996): Phage display of proteins. Curr. Opin. Biotechnol. 7, 547-553.
30. Efimov VP, Neplluev Ⅳ, Mesyanzhinov VV(1995): Bacteriophage T4 as a surface display vector. Virus Genes, 10:173-177.
31. Ekland EH, Szostak JW & Bartel DP(1995): Structurally complex and highly active RNA ligases derived from random RNA sequence. Science,269:364-370.
32. Ellington AD & Szostak JW (1990): In vitro selection of RNA molecules that bind specific ligands. Nature,346: 818-823.
33. Fay TN, Jacobs I, Teisner B, et al.(1988): Two fetal antigens (FA-1 and FA-2) and endometrial proteins (PP12 and PP14) isolated from amniotic fluid; preliminary observations in fetal and maternal tissues. Eur J Obstet Gynecol Reprod Biol ,29(1):73-85.
34. Fishwild DM, O'Donnell SL, Bengoechea T, et al. (1996): High-avidity human lgK monoclonal antibodies from a novel strain of minilocus transgenie mice. Nature Biotech.14:845-851.
35. Foote J & Winter G (1992): Antibody framework residues affecting the conformation of the hypervariable loops. J. Mol. Biol. 224, 487-499.
36. French RR,(1998): Production of bispecific and trispecific F(ab)2 and F(ab)3 antibody derivatives. Methods Mol Biol, 80:121-134.
37. Georgious G, Poetschke HL, et al.(1993): Practical applications of engineering Gram-negative bacterial cell surfaces. TIBTECH, 11 (1):6-10.
38. Georgious G, Stathopoulos C, Dangherty PS, et al.(1997): Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nature Biotechnol. 15,29-34.
39. Gorman SD, Clark MR, Routledge EC, et al.(1991): Reshaping a therapeutic CD4 antibody. Proc. Natl. Acad. Sci. USA,88: 4181-4185.
40. Graus YE, et a1.(1997): Human anti-nicotinic acetylcholine receptor recombinant Fab fragments isolated from thymus-derived phage display libraries from myasthenia gravis patients reflect predominant specificities in serium and block the action of pathogenic serium antibodies. J Immunol, 158,1919-1929.
41. Greener A, Callahan M, & Jerpseth B(1996): An efficient random mutagenesis technique using and E. coli mutator strain. In: Trower MK ed. Methods in Molecular Biology. New Jersey: Jumana Press, 375-385.
42. Griffiths AD, Malmqvist M, Marks JD, et al. (1993): Human anti-self antibodies with high specificity from phage display libraries. EMBO J. 12:725-734.
43. Grosse-Hovest L,Brandl M, Dohlsten M, et al.(1999): Tumor-growth inhibition with bispecific antibody fragments in a syngeneic mouse melanoma model: the role of targeted T-cell co-stimulation via CD28. Int. J. Cancer, 80 (1): 138-144.
44. Guex N & Peitsch MC (1997): SWISS-MODEL and the Swiss-Pdb Viewer: An environment for comparative protein modeling. Electrophoresis 18:2714-2723.
45. Gulick AM & Fahl WE (1995): Forced evolution of glutathione S-transferase to create a more efficient drug detoxicafion enzyme. PNAS, 92:8140-8144.
46. Gundram Jung, Uwe Freimann, Zofia Von Marschall, et al.(1991): Target cell-induced T cell activation with bi- and trispecific antibody fragments. Eur. J. Immunol. 21: 2431-2435.
47. Hahne M, Rimoldi D, Schroter M, et al.(1996): Melanoma cell expression of Fas(Apo-1/CD95)ligand: implication for tumor immune escape. Science 274:1363-1366.
48. Hall A & knowles JR (1976): Directed selective pressure on a beta-lactamase to analyze molecular changes involved in development of enzyme function. Nature,264,803-804.
49. Hanes J & Pluckthun A (1997): In viuo selection and diversification of functional proteins using ribosome display. Proc. Natl. Acad. Sci. USA, 94, 4937-4942.
50. Hansen JA, Martin PJ & Nowinski RC (1980): Monoclonal antibodies identifying a novel T-cell antigen and Ia antigens of human Lymphocytes. Immunogenetics,10, 247-260.
51. Hara T, Fu SM & Hansen JA (1985): Human T cell activation:Ⅱ. A new activation pathway used by a major T cell population Via a disulfide-bonded dimer of a 44 kilodalton polypcptide(9.3 antigen).J. Exp. Med. 161, 1513-1524.
52. Hayden MS, Grosmaire LS, Norris NA, et al.(1996): Costimulation by CD28 sFv expressed on the tumor cell surface or as a soluble bispecific molecule targeted to the L6 carcinoma antigen.Tissue antigen. 48: 242-254.
53. Holmes MA, Buss TN & Foote J (1998): Conformational correction mechanisms aiding a antigen recognition by a humanized antibody. J. Exp. Med., 187: 479-485.
54. Hombach A, Jung W, Pohl C,et al. (1993): A CD16/CD30 bispecific antibody induces lysis of Hodgkin cells by unstimulated Natural Killer cells in vitro and in vive. Int. J. Cancer.55:830-837.
55. Huse WD,Sastry L, Iverson SA, et al. (1989): Generation of a large combinatioorial library of the immunoglobulin repertoire in phage lambda. Science. 246:1275-1281.
56. Jakobovits A, (1995): Production of fully human antibodies by transgenic mice. Curr. Opin. Biotech. 6:561-566.
57. Jones PT, Dear PH, Foote J, et al. (1986): Replacing the complimentarity-determining regions in human antibody with those from a mouse. Nature 321:522-525.
58. Joyce GF(1989): Amplification, mutation and selection of catalytic RNA. Gene,82:83-87.
59. June CH, Bluestone JA, Nadler LM,Thompson CB(1994): The B7 and CD28 receptor families. Immunol Today, 15(7): 321-332.
60. Jung G, Freimann U, Von Marschall Z, et al.(1991): Target cell-induced T cell activation with bi-and trispecific antibody fragments. Eur J Immunol 21(10):2431-5:2435.
61. Kabat EA, Wu TT, Perry HM, et al. (1991): Sequence of proteins of immunological interest.
Fifth edition, NIH., Bethesda,MD., publication NO. 91-3242
62. Kaga S, Ragg S, Rogers KA, et al. (1998): Activation of p21-CDC42/Rac-activated kinases by CD28 signaling:p21-activated kinase (PAK) and MEK kinase(MEKK1) may mediate the therphy between CD3 and CD28 signals. J. Immunol., 4182-4189.
63. Kettleborough CA, Saldanha J, Heaaith VJ, et al. (1991): Humanization of a mouse monoclonal antibody by CDR-grafting: the importance of framework residues on loop conformation. Protein Engng. 4,773-783.
64. Kohler G & Milstein C (1975): Continous cultures of fused cells secreting antibody of predefined specificity. Nature 256:495-497.
65. Kruif J (1995): Selection and application of human single chain Fv antibody fragments from semi-synthetic phage antibody display library with designed CDR3 regions. J Mol Biol, 248:97-105.
66. Ledbetter JA, Martin PJ, Speoner CE, et al.(1985): Antibodies to Tp67 and Tp44 augment and sustain proliferative responses of activated T cells. J. of Immunology. 135,2331-2336.
67. Liao XC, Fournier S, Killeen N, et al(1997): Itk negatively regulates induction of T cell proliferation by CD28 costimulation. J. Exp. Med., 186(2):221-228.
68. Ling V, Wu PW, Finnerty HF, et al.(2000): Identification of GL50, a novel B7-like protein that functionally binding to ICOS receptor. J. Immunol., 164:1653-1657.
69. Liu K-Q, Bunnell SC, Gurniak CB, et al. (1998): T cell receptor-initiated calcium release is uncoupled from capacitative calcium entry in Itk-deficient T cells. J. Exp. Med., 187(10): 1721-1727.
70. Logtenberg T, Young FM, Van Es JH, et al. (1989): Autoantibodies encoded by the most Jh-proximal human immunoglobulin heavy chain variable region gene. J. Exp. Med. 170(4), 1347-1355.
71. Lonberg N, Taylor LD, Harding FA, et al. (1994): Antigen-specific human antibodies from mice comprising four distinct genetic modifications. Nature 368:856-859.
72. Low NM, Holliger P & Winter G (1996): Mimicking somatic hypermutation: affinity maturation of antibodies displayed on bacteriophage using a bacteria mutator strain. J. Mol.Biol. 250:359-368.
73. Manzke O, Berthold F, Huebe K, et al.(1999): CD3×CD19 bispecific antibodies and CD28 bivalent antibodies enhance T-cell reactivity against autologous leukemic cells in pediatric B-All bone marrow.Int. J. Cancer, 80:715-722.
74. Marks JD, Grittiths AD, Malmqvist M, et al. (1992): By-passing immunization: building high affinity human antibodies by chain shuffling. Bio/Tech. 10:779-783.
75. Mazumder A & Rosenberg AS (1984): Successful immunotherapy of NK-resistant established pulmonary melanoma metastases by the intravenous adoptive transfer of syngenic lymphocytes activated in vitro by interleukin-2. Science. 255:1487-1489.
76. Mazzoni A, Mezzanzanica D, Jung G, et al.(1996): CD3-CD28 costimulation as a means to avoiding T cell preactivation in bispecific monoelonal antibody-based treatment of ovarian carcinoma. Cancer Res., 56:5443-5449.
77. McCafferty J, Griffiths AD, Winter G, et al. (1990): Phage antibodies: Filamentous phage displaying antibodies variable domains. Nature 348:552-554.
78. Mendez M J, Green LL, Corvalan JRF, et al. (1997): Functional transplant of megabase human immunoglobulin loci recapitulates human antibody response in mice. Nature Genet. 15:146-156.
79. Mills DR, Peterson RL, Spiegelman S (1967): An extracellular darwinian experiment with a self-duplication nucleic acid molecule. PANS, 58(1) :217-224.
80. Moore JC, Jin HM, Kuchner O, et al.(1997): Strategies for the in vitro evolution of protein function: enzyme evolution by random recombination of improved sequence. J. Mol. Biol. 272,336-347.
81. Moretta A, Pantaleo G, Botet ML, et al.(1985): Involvement of T44 molecules in an antigen-independent pathway of T cell activation. J. Exp. Med. 162: 823-838.
82. Neuberger MS, Williams GT & Fox RO (1984): Recombinant antibodies possessing novel effector functions. Nature 312,604-608.
83. Oliver Manzke,Frank Berthold, Kai Huebel, et al. (1999): CD3×CD19 bispecific antibodies and CD28 bivalent antibodies enhance T-cell reactivity against autologous leukemic cells in pediatric B-All bone marrow. Int. J. Cancer, 80:715-722.
84. Ollson C,Michaelssoon E,Parra E.U.P. et al.(1998): Biased dependency of CD80 versus CD86
in the induction of transcription factors regulating the human IL-2 promoter. Int. Immunol.,10(4):499-506.
85. Pardoll DM (1993): Cancer vaccines. Immunol. Today. 14:310-316.
86. Peitsch MC(1995): Protein modeling by E-mail. Bio/Technology 13,658-660.
87. Peitsch MC(1996): ProMod and Swiss-Model: Internet-based tools for automated comparative protein modeling. Biochem Sec Trans 24: 274-279.
88. Person MA, et a1.(1991): Generation of diverse high -affinity human monoclonal antibodies by repertoire cloning. Proc Natl Acad Sci USA, 88,10134-10137.
89. Pluckthun A (1998): Protein Dynamics, Function and Design, edited by Jardetzky et al.,Plenum Press, New York. (http://www. unizh.ch/～pluckth/)
90. Popov AV, Butzler C, Frippiat J-P, et al. (1996): Assembly and extension of yeast artificial chromosomes to build up a large locus. Gene 177:195-205.
91. Portolano S, Mclachlan SM & Rapoport B (1993): High affinity thyroid-specific human autoantibodies display on the surface of filamentous phage use Ⅴ gene similar to other autoantibodies. J Immunol, 151:2839-2851.
92. Powis SJ, Deverson EV, Coadwell WJ, et al. (1992): Effect of polymorphism of an MHC-linked transporter on the peptides assembled in a class Ⅰ molecule. Nature. 357:211-215.
93. Rennet C, Jung W, Sahin U, et al. (1994a): Cure of xenografted human tumors by bispecific monoclonal antibodies and human T cells. Science. 264:833-835.
94. Renner C, Jung W, Sahin U, et al. (1994b): Mechanisms of T lymphocyte activation and cytotoxicity induced by bispecific CD3 and CD28 antibodies. Immunobiology. 192:132.
95. Restifo NP, Esquivel F, Kawamaki Y, et al. (1993): Identification of human cancers defecient in antigen processing. J. Exp. Med. 177:265-272.
96. Riechmann L, Clark M, Waldmann H, et al. (1988): Reshaping human antibodies for therapy.Nature,332,323-327.
97. Roberts BL, Markland W, Ley AC, et al.(1992): Directed evolution of a protein: Selection of protein neutrophil elastase inhibitors display on M13 fusion phage. PNAS. 1992, 89: 2429-2433.
98. Schoonjans R, Willems A, Schoonooghe S, et al.(2000a): Fab chains As an efficient
heterodimerization scaffold for the production of recombinant bispecific and trispecific antibody derivatives. J Immunol,165(12): 7050-7.
99. Schoonjans R, Willems A, Grooten J, et al. ,(2000b): Efficient heterodimcrization of recombinant bi-and trispecific antibodies. Bioseparation ,9(3): 179-83.
100. Schott ME, Frazier KA, Pollock DK, et al.,(1993): Preparation, characterization, and in vivo biodistribution properties of synthetically cross-linked multivalent antitumor antibody fragments. Bioconjug Chem ,4(2): 153-65.
101. Schwartz RH (1992): Costimulation of T lyphocytes: the role of CD28,CTLA-4, and B7/BB1 in interleukin-2 production and immunotherapy. Cell, 71: 1065-1068.
102. Slavik JM, Hutchcroft JE & Bierer BE (1999): CD28/CTLA-4 and CD80/CD86 families,signaling and function. Immunologic Research. 19/1:1-24.
103. Smith GP(1985): Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science,228,1315-1317.
104. Somasundaram C, Sundarapandiyan K, Keler T, et al. (1999): Development of a trispecific antibody conjugate that directs two distinct tumor-associated antigens to CD64 on myeloid effector cells. Hum Antibodies, 9(1):47-54.
105. Soumillion P, Jespers L, Bouchet M, et al.(1994): Identification and characterization of Src SH3 ligands from phage-displayed random peptide libraries. J Biol Chem, 269: 23853-23856.
106. Staerz UD & Bevan MJ (1986): Hybird hybridoma producing a bispecific monoclonal antibody that can focus effector T-cell activity. Proc. Natl. Acad Sci. USA 83:1453-1457.
107. Stemmer WP (1994~a): Rapid evolution of a protein in vitro by DNA. Nature,370,389-391.
108. Stemmer WP (1994~b): DNA shuffling by random fragmentation and reassembly: In vitro recombination for molecular evolution. Proc. Natl. Acad. Sci. USA, 91(22): 10747-10751.
109. Sternberg N & Hoess RH (1995): Display of peptides and proteins on the surface of bacgerriophage. Proc. Natl. Acad. Sci. USA 92:1609-1613.
110. Swallow MM,Wallin JJ,Sha WC (1999): B7h, a novel costimulatory homolog of B7.1 and B7.2, is induced by TNF alpha. Immunity, 11:423-432.
111. Townsend SE & Allison JP (1993): Tumor rejection after direct costimulation of CD8+ T cells by B7-transfected melanoma cells. Science. 259:368-372.
112. Tuerk C & Gold L(1990):Systemic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science, 1990,249:505-510.
113. Tutt A, Stevenson GT, Glennie MJ(1991): Trispecific F(ab')3 derivatives that use cooperative signaling via the TCR/CD3 complex and CD2 to activate and redirect resting cytotoxic T cells. J Immunol 147(1):60-9.
114. Van der Bruggen P, Traversari C, Chomez P, et al. (1991): A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science. 254:1643-1647.
115. Wang CI, Yang Q and Craik CS(1995): Isolation of a high affinity inhibitor of urokinase-type plasminogen activator by phage display of ecotin. J Biol Chem,270:12250-12256.
116. Weiss A, Manger B & Imboden J (1986): synergy between the T3/antigen receptor complex and Tp44 in the activation of human T cells. J. of Immunology. 137,0819-0825.
117. Wong WM, Vakis SA, Ayre KR, et al.,(2000): Rheumatoid arthritis T cells produce Th1 cytokines in response to stimulation with a novel trispecific antibody directed against CD2,CD3, and CD28. Scand J Rheumatol ,29(5):282-7.
118. Yang XF, Weber GF, Cantor H(1997): A novel Bcl-x isoform connected to the T cell receptor regulates apoptosis in T cells. Immunity, 7:629-639.
119. Yoshinaga SK,Whoriskey JS, Khare SD, et al.(1999): T-cell co-simulation through B7RP-1 and ICOS. Nature, 402: 837-832.
120. You L & Arnold FH (1996): Directed evolution of subtilisin E in Bacillus subfilis to enhance total activity in aqueous dimethlyformamide. Protein Eng., 9(1):77-83.
121. Zhang JH, Dawes G & Stemmer WP(1997): Directed evolution of a fucosidase from a glactosidase by DNA shuffling and sceening. PNAS, 94:4504-4509.
2．刘喜富，萧飒，顾征等．(1996)．抗人CD3单链抗体及改形单域抗体的表达．中国科学(C辑)．26：428-435．
3．蒋欣(1998)：抗人CD3改形单链抗体及抗人卵巢癌双特异单链抗体的构建与表达。博士论文，中国科学院遗传研究所。
4. Abraltmsen L, Tom Jeffrey, Burnier J, et al., (1991): Engineering subrilisin and its substrates for efficient ligafon of peptide bonds in aqueous solution. Biochemistry, 30:4151-4159.
5. Baca M, Scanlan TS, Stephenson RC, et al.(1997~a): Phage display of a catalytic antibody to optimize affinity for transition-state analog binding. Proc. Natl. Acad. Sci. USA, 94,10063-10068.
6. Baca M, Presta L, O'Connor SJ, et al. (1997~b): Antibody humanization using monovalent phage display. J. Biol. Chem. 272:10678-10684.
7. Bartel DP & Szostak JW (1993): Isolation of new ribozymes from a large pool of random sequences. Science, 261:1411-1418.
8. Baskar S, Ostrand-Rosenberg S, Nabavi N, et al. (1993). Constitutive expression of B7 restores immunogenicity of tumor cells expressing truncated major histocompatibility complex class Ⅱ molecules. Proc. Natl. Acad. Sci. USA 90:5687-5690.
9. Bock LC, Griffin LC, Latham JA, et al.(1992): Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Nature,355(6360):564-566.
10. Boder ET & Wittrup KD (1997): Yeast surface display for screening combinatorial polypeptide libraries. Nature Biotechnol. 15,553-557.
11. Boder ET, Midelfort KS & Wittrup KD (2000): Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity. Proc. Natl. Acad. Sci. USA, 97(20): 10701-10705.
12. Bohlen H, Hopff T, Manzke O, et al. (1993): Lysis of maligant B cells from patients with B-chronic lymphocytic leukemia by autologous T cells activated with CD3×CD19 bispceific antibodies in combination with bivalent CD28 antibodies. Blood, 82:1803-1812.
13. Bohlen H, Manzke O, Titzer S, et al. (1997): Prevention of Epstein-Barr virus-induced human B-cell lymphoma in severe combined immunodeficient mice treated with CD3×CD19 bispecific antibodies,CD28 monospecific antibodies, and autologous T cells. Cancer Res., 57:1704-1709.
14. Boulianne GL, Hozumi N & Shulman MJ (1984): Production of functional chimeric mouse/human antibody. Nature. 312, 643-646.
15. Bruggemann M and Taussig MJ (1997): Production of human antibody repertoires in transgenic mice. Curr. Opin. Biotechnol. 8:455-458.
16. Cai X and Green A (1995): Anti-melanoma antibodies from melanoma patients immunized with genetically modified autologus tumour cells: selection of specific antibodies from single-chain Fv fusion phage libraries. Proc. Natl. Acad. Sci. USA 92:6537-6541.
17. Carter,P.,Presta,L.,Gorman, C.M. et al.(1992): Humanization of an anti-p185HER2 antibody for human cancer therapy. Proc. Natl. Acad. Sci. USA, 89,4285-4289.
18. Cheetham MT, Hale G, Waldmann H, et al.(1998): Crystal structures of a rat anti-CD_(52) (CAMPATH-1) therapeutic antibody Fab fragment and its humanized counterpart. J. Mol. Biol. 284,85-99.
19. Chen K &Amold FH (1991): Enzyme engineering for nonaqueous solvents: random mutagenesis to enhance activity of subtilisin E in polar organic media. Biote-chnology, 1991,9(11): 1073-1077.
20. Chen K, & Amold FH (1993): Tuning the activity of an enzyme for unusual environments: sequential random mutagenesis of subtilisin E for catalysis in dimethylformamide. Proc. Natl. Acad. Sci. USA, 90(2):5618-5622.
21. Chen L, Linsley PS & Hellstrom KE (1993): Costimulation of T cells for tumor immunity.Immunol. Today. 14:483-486.
22. Clackson T, et al.(1991): making antibody fragments using phage display libraries. Nature,352,624-628.
23. Clark EA, Martin PJ, Hansen JA, et al. (1983): Evolution of epitopes on human and nonhuman primate lymphocyte cell surface antigens. Immunogenefics, 18,599-615.
24. Coulie PG, Weynants P, Lehmann F, et al. (1993): Genes coding for tumor antigens recognized by human cytolytic T lymphocytes. J. Immunother. 14:104-109.
25. Daugherty PS, Chen G, Iverson BL, et al.(2000): Quantitative analysis of the effect of the mutation frequency on the affinity maturation of single chain Fv antibodies. Proc. Natl. Acad.Sci. USA,97(5),2029-2034.
26. Deborah JL, Thersa LW, Jeffrey AB, et al. (1996): CD28/B7 system of T cell costimulation. Annu. Rev. Immunol. 14: 233-258.
27. Demanet C, Brissinck J, Leo O, et al.(1996): Bispecific antiboddy-mediated immunotherapy of the BCL1 lymphoma:increasd efficacy with multiple injections and CD28-induced costimulation. Blood, 87: 4390-4398.
28. Dong H, Zhu G, Tamada K, et a1.(1999): B7-H1, a third member of the B7 family, co-stimulates T-cell proliferation and interleukin-10 secretion. Nat. Med., 5:1365-1369.
29. Dunn IS (1996): Phage display of proteins. Curr. Opin. Biotechnol. 7, 547-553.
30. Efimov VP, Neplluev Ⅳ, Mesyanzhinov VV(1995): Bacteriophage T4 as a surface display vector. Virus Genes, 10:173-177.
31. Ekland EH, Szostak JW & Bartel DP(1995): Structurally complex and highly active RNA ligases derived from random RNA sequence. Science,269:364-370.
32. Ellington AD & Szostak JW (1990): In vitro selection of RNA molecules that bind specific ligands. Nature,346: 818-823.
33. Fay TN, Jacobs I, Teisner B, et al.(1988): Two fetal antigens (FA-1 and FA-2) and endometrial proteins (PP12 and PP14) isolated from amniotic fluid; preliminary observations in fetal and maternal tissues. Eur J Obstet Gynecol Reprod Biol ,29(1):73-85.
34. Fishwild DM, O'Donnell SL, Bengoechea T, et al. (1996): High-avidity human lgK monoclonal antibodies from a novel strain of minilocus transgenie mice. Nature Biotech.14:845-851.
35. Foote J & Winter G (1992): Antibody framework residues affecting the conformation of the hypervariable loops. J. Mol. Biol. 224, 487-499.
36. French RR,(1998): Production of bispecific and trispecific F(ab)2 and F(ab)3 antibody derivatives. Methods Mol Biol, 80:121-134.
37. Georgious G, Poetschke HL, et al.(1993): Practical applications of engineering Gram-negative bacterial cell surfaces. TIBTECH, 11 (1):6-10.
38. Georgious G, Stathopoulos C, Dangherty PS, et al.(1997): Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nature Biotechnol. 15,29-34.
39. Gorman SD, Clark MR, Routledge EC, et al.(1991): Reshaping a therapeutic CD4 antibody. Proc. Natl. Acad. Sci. USA,88: 4181-4185.
40. Graus YE, et a1.(1997): Human anti-nicotinic acetylcholine receptor recombinant Fab fragments isolated from thymus-derived phage display libraries from myasthenia gravis patients reflect predominant specificities in serium and block the action of pathogenic serium antibodies. J Immunol, 158,1919-1929.
41. Greener A, Callahan M, & Jerpseth B(1996): An efficient random mutagenesis technique using and E. coli mutator strain. In: Trower MK ed. Methods in Molecular Biology. New Jersey: Jumana Press, 375-385.
42. Griffiths AD, Malmqvist M, Marks JD, et al. (1993): Human anti-self antibodies with high specificity from phage display libraries. EMBO J. 12:725-734.
43. Grosse-Hovest L,Brandl M, Dohlsten M, et al.(1999): Tumor-growth inhibition with bispecific antibody fragments in a syngeneic mouse melanoma model: the role of targeted T-cell co-stimulation via CD28. Int. J. Cancer, 80 (1): 138-144.
44. Guex N & Peitsch MC (1997): SWISS-MODEL and the Swiss-Pdb Viewer: An environment for comparative protein modeling. Electrophoresis 18:2714-2723.
45. Gulick AM & Fahl WE (1995): Forced evolution of glutathione S-transferase to create a more efficient drug detoxicafion enzyme. PNAS, 92:8140-8144.
46. Gundram Jung, Uwe Freimann, Zofia Von Marschall, et al.(1991): Target cell-induced T cell activation with bi- and trispecific antibody fragments. Eur. J. Immunol. 21: 2431-2435.
47. Hahne M, Rimoldi D, Schroter M, et al.(1996): Melanoma cell expression of Fas(Apo-1/CD95)ligand: implication for tumor immune escape. Science 274:1363-1366.
48. Hall A & knowles JR (1976): Directed selective pressure on a beta-lactamase to analyze molecular changes involved in development of enzyme function. Nature,264,803-804.
49. Hanes J & Pluckthun A (1997): In viuo selection and diversification of functional proteins using ribosome display. Proc. Natl. Acad. Sci. USA, 94, 4937-4942.
50. Hansen JA, Martin PJ & Nowinski RC (1980): Monoclonal antibodies identifying a novel T-cell antigen and Ia antigens of human Lymphocytes. Immunogenetics,10, 247-260.
51. Hara T, Fu SM & Hansen JA (1985): Human T cell activation:Ⅱ. A new activation pathway used by a major T cell population Via a disulfide-bonded dimer of a 44 kilodalton polypcptide(9.3 antigen).J. Exp. Med. 161, 1513-1524.
52. Hayden MS, Grosmaire LS, Norris NA, et al.(1996): Costimulation by CD28 sFv expressed on the tumor cell surface or as a soluble bispecific molecule targeted to the L6 carcinoma antigen.Tissue antigen. 48: 242-254.
53. Holmes MA, Buss TN & Foote J (1998): Conformational correction mechanisms aiding a antigen recognition by a humanized antibody. J. Exp. Med., 187: 479-485.
54. Hombach A, Jung W, Pohl C,et al. (1993): A CD16/CD30 bispecific antibody induces lysis of Hodgkin cells by unstimulated Natural Killer cells in vitro and in vive. Int. J. Cancer.55:830-837.
55. Huse WD,Sastry L, Iverson SA, et al. (1989): Generation of a large combinatioorial library of the immunoglobulin repertoire in phage lambda. Science. 246:1275-1281.
56. Jakobovits A, (1995): Production of fully human antibodies by transgenic mice. Curr. Opin. Biotech. 6:561-566.
57. Jones PT, Dear PH, Foote J, et al. (1986): Replacing the complimentarity-determining regions in human antibody with those from a mouse. Nature 321:522-525.
58. Joyce GF(1989): Amplification, mutation and selection of catalytic RNA. Gene,82:83-87.
59. June CH, Bluestone JA, Nadler LM,Thompson CB(1994): The B7 and CD28 receptor families. Immunol Today, 15(7): 321-332.
60. Jung G, Freimann U, Von Marschall Z, et al.(1991): Target cell-induced T cell activation with bi-and trispecific antibody fragments. Eur J Immunol 21(10):2431-5:2435.
61. Kabat EA, Wu TT, Perry HM, et al. (1991): Sequence of proteins of immunological interest.
Fifth edition, NIH., Bethesda,MD., publication NO. 91-3242
62. Kaga S, Ragg S, Rogers KA, et al. (1998): Activation of p21-CDC42/Rac-activated kinases by CD28 signaling:p21-activated kinase (PAK) and MEK kinase(MEKK1) may mediate the therphy between CD3 and CD28 signals. J. Immunol., 4182-4189.
63. Kettleborough CA, Saldanha J, Heaaith VJ, et al. (1991): Humanization of a mouse monoclonal antibody by CDR-grafting: the importance of framework residues on loop conformation. Protein Engng. 4,773-783.
64. Kohler G & Milstein C (1975): Continous cultures of fused cells secreting antibody of predefined specificity. Nature 256:495-497.
65. Kruif J (1995): Selection and application of human single chain Fv antibody fragments from semi-synthetic phage antibody display library with designed CDR3 regions. J Mol Biol, 248:97-105.
66. Ledbetter JA, Martin PJ, Speoner CE, et al.(1985): Antibodies to Tp67 and Tp44 augment and sustain proliferative responses of activated T cells. J. of Immunology. 135,2331-2336.
67. Liao XC, Fournier S, Killeen N, et al(1997): Itk negatively regulates induction of T cell proliferation by CD28 costimulation. J. Exp. Med., 186(2):221-228.
68. Ling V, Wu PW, Finnerty HF, et al.(2000): Identification of GL50, a novel B7-like protein that functionally binding to ICOS receptor. J. Immunol., 164:1653-1657.
69. Liu K-Q, Bunnell SC, Gurniak CB, et al. (1998): T cell receptor-initiated calcium release is uncoupled from capacitative calcium entry in Itk-deficient T cells. J. Exp. Med., 187(10): 1721-1727.
70. Logtenberg T, Young FM, Van Es JH, et al. (1989): Autoantibodies encoded by the most Jh-proximal human immunoglobulin heavy chain variable region gene. J. Exp. Med. 170(4), 1347-1355.
71. Lonberg N, Taylor LD, Harding FA, et al. (1994): Antigen-specific human antibodies from mice comprising four distinct genetic modifications. Nature 368:856-859.
72. Low NM, Holliger P & Winter G (1996): Mimicking somatic hypermutation: affinity maturation of antibodies displayed on bacteriophage using a bacteria mutator strain. J. Mol.Biol. 250:359-368.
73. Manzke O, Berthold F, Huebe K, et al.(1999): CD3×CD19 bispecific antibodies and CD28 bivalent antibodies enhance T-cell reactivity against autologous leukemic cells in pediatric B-All bone marrow.Int. J. Cancer, 80:715-722.
74. Marks JD, Grittiths AD, Malmqvist M, et al. (1992): By-passing immunization: building high affinity human antibodies by chain shuffling. Bio/Tech. 10:779-783.
75. Mazumder A & Rosenberg AS (1984): Successful immunotherapy of NK-resistant established pulmonary melanoma metastases by the intravenous adoptive transfer of syngenic lymphocytes activated in vitro by interleukin-2. Science. 255:1487-1489.
76. Mazzoni A, Mezzanzanica D, Jung G, et al.(1996): CD3-CD28 costimulation as a means to avoiding T cell preactivation in bispecific monoelonal antibody-based treatment of ovarian carcinoma. Cancer Res., 56:5443-5449.
77. McCafferty J, Griffiths AD, Winter G, et al. (1990): Phage antibodies: Filamentous phage displaying antibodies variable domains. Nature 348:552-554.
78. Mendez M J, Green LL, Corvalan JRF, et al. (1997): Functional transplant of megabase human immunoglobulin loci recapitulates human antibody response in mice. Nature Genet. 15:146-156.
79. Mills DR, Peterson RL, Spiegelman S (1967): An extracellular darwinian experiment with a self-duplication nucleic acid molecule. PANS, 58(1) :217-224.
80. Moore JC, Jin HM, Kuchner O, et al.(1997): Strategies for the in vitro evolution of protein function: enzyme evolution by random recombination of improved sequence. J. Mol. Biol. 272,336-347.
81. Moretta A, Pantaleo G, Botet ML, et al.(1985): Involvement of T44 molecules in an antigen-independent pathway of T cell activation. J. Exp. Med. 162: 823-838.
82. Neuberger MS, Williams GT & Fox RO (1984): Recombinant antibodies possessing novel effector functions. Nature 312,604-608.
83. Oliver Manzke,Frank Berthold, Kai Huebel, et al. (1999): CD3×CD19 bispecific antibodies and CD28 bivalent antibodies enhance T-cell reactivity against autologous leukemic cells in pediatric B-All bone marrow. Int. J. Cancer, 80:715-722.
84. Ollson C,Michaelssoon E,Parra E.U.P. et al.(1998): Biased dependency of CD80 versus CD86
in the induction of transcription factors regulating the human IL-2 promoter. Int. Immunol.,10(4):499-506.
85. Pardoll DM (1993): Cancer vaccines. Immunol. Today. 14:310-316.
86. Peitsch MC(1995): Protein modeling by E-mail. Bio/Technology 13,658-660.
87. Peitsch MC(1996): ProMod and Swiss-Model: Internet-based tools for automated comparative protein modeling. Biochem Sec Trans 24: 274-279.
88. Person MA, et a1.(1991): Generation of diverse high -affinity human monoclonal antibodies by repertoire cloning. Proc Natl Acad Sci USA, 88,10134-10137.
89. Pluckthun A (1998): Protein Dynamics, Function and Design, edited by Jardetzky et al.,Plenum Press, New York. (http://www. unizh.ch/～pluckth/)
90. Popov AV, Butzler C, Frippiat J-P, et al. (1996): Assembly and extension of yeast artificial chromosomes to build up a large locus. Gene 177:195-205.
91. Portolano S, Mclachlan SM & Rapoport B (1993): High affinity thyroid-specific human autoantibodies display on the surface of filamentous phage use Ⅴ gene similar to other autoantibodies. J Immunol, 151:2839-2851.
92. Powis SJ, Deverson EV, Coadwell WJ, et al. (1992): Effect of polymorphism of an MHC-linked transporter on the peptides assembled in a class Ⅰ molecule. Nature. 357:211-215.
93. Rennet C, Jung W, Sahin U, et al. (1994a): Cure of xenografted human tumors by bispecific monoclonal antibodies and human T cells. Science. 264:833-835.
94. Renner C, Jung W, Sahin U, et al. (1994b): Mechanisms of T lymphocyte activation and cytotoxicity induced by bispecific CD3 and CD28 antibodies. Immunobiology. 192:132.
95. Restifo NP, Esquivel F, Kawamaki Y, et al. (1993): Identification of human cancers defecient in antigen processing. J. Exp. Med. 177:265-272.
96. Riechmann L, Clark M, Waldmann H, et al. (1988): Reshaping human antibodies for therapy.Nature,332,323-327.
97. Roberts BL, Markland W, Ley AC, et al.(1992): Directed evolution of a protein: Selection of protein neutrophil elastase inhibitors display on M13 fusion phage. PNAS. 1992, 89: 2429-2433.
98. Schoonjans R, Willems A, Schoonooghe S, et al.(2000a): Fab chains As an efficient
heterodimerization scaffold for the production of recombinant bispecific and trispecific antibody derivatives. J Immunol,165(12): 7050-7.
99. Schoonjans R, Willems A, Grooten J, et al. ,(2000b): Efficient heterodimcrization of recombinant bi-and trispecific antibodies. Bioseparation ,9(3): 179-83.
100. Schott ME, Frazier KA, Pollock DK, et al.,(1993): Preparation, characterization, and in vivo biodistribution properties of synthetically cross-linked multivalent antitumor antibody fragments. Bioconjug Chem ,4(2): 153-65.
101. Schwartz RH (1992): Costimulation of T lyphocytes: the role of CD28,CTLA-4, and B7/BB1 in interleukin-2 production and immunotherapy. Cell, 71: 1065-1068.
102. Slavik JM, Hutchcroft JE & Bierer BE (1999): CD28/CTLA-4 and CD80/CD86 families,signaling and function. Immunologic Research. 19/1:1-24.
103. Smith GP(1985): Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science,228,1315-1317.
104. Somasundaram C, Sundarapandiyan K, Keler T, et al. (1999): Development of a trispecific antibody conjugate that directs two distinct tumor-associated antigens to CD64 on myeloid effector cells. Hum Antibodies, 9(1):47-54.
105. Soumillion P, Jespers L, Bouchet M, et al.(1994): Identification and characterization of Src SH3 ligands from phage-displayed random peptide libraries. J Biol Chem, 269: 23853-23856.
106. Staerz UD & Bevan MJ (1986): Hybird hybridoma producing a bispecific monoclonal antibody that can focus effector T-cell activity. Proc. Natl. Acad Sci. USA 83:1453-1457.
107. Stemmer WP (1994~a): Rapid evolution of a protein in vitro by DNA. Nature,370,389-391.
108. Stemmer WP (1994~b): DNA shuffling by random fragmentation and reassembly: In vitro recombination for molecular evolution. Proc. Natl. Acad. Sci. USA, 91(22): 10747-10751.
109. Sternberg N & Hoess RH (1995): Display of peptides and proteins on the surface of bacgerriophage. Proc. Natl. Acad. Sci. USA 92:1609-1613.
110. Swallow MM,Wallin JJ,Sha WC (1999): B7h, a novel costimulatory homolog of B7.1 and B7.2, is induced by TNF alpha. Immunity, 11:423-432.
111. Townsend SE & Allison JP (1993): Tumor rejection after direct costimulation of CD8+ T cells by B7-transfected melanoma cells. Science. 259:368-372.
112. Tuerk C & Gold L(1990):Systemic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. Science, 1990,249:505-510.
113. Tutt A, Stevenson GT, Glennie MJ(1991): Trispecific F(ab')3 derivatives that use cooperative signaling via the TCR/CD3 complex and CD2 to activate and redirect resting cytotoxic T cells. J Immunol 147(1):60-9.
114. Van der Bruggen P, Traversari C, Chomez P, et al. (1991): A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science. 254:1643-1647.
115. Wang CI, Yang Q and Craik CS(1995): Isolation of a high affinity inhibitor of urokinase-type plasminogen activator by phage display of ecotin. J Biol Chem,270:12250-12256.
116. Weiss A, Manger B & Imboden J (1986): synergy between the T3/antigen receptor complex and Tp44 in the activation of human T cells. J. of Immunology. 137,0819-0825.
117. Wong WM, Vakis SA, Ayre KR, et al.,(2000): Rheumatoid arthritis T cells produce Th1 cytokines in response to stimulation with a novel trispecific antibody directed against CD2,CD3, and CD28. Scand J Rheumatol ,29(5):282-7.
118. Yang XF, Weber GF, Cantor H(1997): A novel Bcl-x isoform connected to the T cell receptor regulates apoptosis in T cells. Immunity, 7:629-639.
119. Yoshinaga SK,Whoriskey JS, Khare SD, et al.(1999): T-cell co-simulation through B7RP-1 and ICOS. Nature, 402: 837-832.
120. You L & Arnold FH (1996): Directed evolution of subtilisin E in Bacillus subfilis to enhance total activity in aqueous dimethlyformamide. Protein Eng., 9(1):77-83.
121. Zhang JH, Dawes G & Stemmer WP(1997): Directed evolution of a fucosidase from a glactosidase by DNA shuffling and sceening. PNAS, 94:4504-4509.