USOO8877913B2
(12) United States Patent (10) Patent No.: US 8,877,913 B2 Nakagawa et a1. (45) Date of Patent: Nov. 4, 2014
(54) ANTI-CD4 ANTIBODY OTHER PUBLICATIONS (71) Applicants:Tomoaki Nakagawa, Tokyo (JP); Darrell Anderson et al., “A Primatized MAb to Human CD4 Causes Sayaka Hori, Tokyo (JP); Rinpei Niwa, Receptor Modulation, without Marked Reduction in CD4+T Cells in Tokyo (JP); Tsuguo Kubota, Tokyo (JP); Chimpanzees: In Vitro and in Vivo Characterization of a MAb Kazuhiro Masuda, Tokyo (JP); (IDEC-CE9.1) to Human CD4”, Clinical Immunology and Kazuyasu Nakamura, Tokyo (JP) Immunopathology, 1997, 84(1): 73-84. Paul J. Carter, “Potent antibody therapeutics by design”, Nature (72) Inventors: Tomoaki Nakagawa, Tokyo (JP); Reviews, Immunology, 2006, 6: 343-357. Sayaka Hori, Tokyo (JP); Rinpei Niwa, Youn Kim et al., “Clinical ef?cacy of zanolimumab (HuMax-CD4): two phase 2 studies in refractory cutaneous T-cell lymphoma”, Tokyo (JP); Tsuguo Kubota, Tokyo (JP); Blood, 2007, 109: 4655-4662. Kazuhiro Masuda, Tokyo (JP); Onn M. Kon et al., “Randomised, dose-ranging, placebo-controlled Kazuyasu Nakamura, Tokyo (JP) study of chimeric antibody to CD4 (keliximab) in chronic severe asthma”, The Lancet, 1998, 352: 1109-1113. (73) Assignee: Kyowa Hakko Kirin Co., Ltd, Tokyo Matthias Merkenschlarger et al., “Functional Epitope Analysis of the (J P) Human CD4 Molecule: The MHC Class II-Dependent Activation of Resting T Cells is Inhibited by Monoclonal Antibodies to CD4 Notice: Subject to any disclaimer, the term of this Regardless whether or Not They Recognize Epitopes Involved in the patent is extended or adjusted under 35 Binding of MHC Class II or HIV gp120”, The Journal of Immunol U.S.C. 154(b) by 86 days. ogy, 1990, 145(9): 2839-2845. Akito Natsume et al., “Engineered Antibodies 0f IgGl/IgG3 Mixed (21) Appl. No.: 13/755,472 Isotype with Enhanced Cytotoxic Activities”, Cancer Research, 2008,68: 3863-3872. Roland Newman et al., “‘Primatization’” of recombinant antibodies (22) Filed: Jan. 31, 2013 for immunotherapy of human diseases: A macaque/human chimeric antibody against human CD4, Biotechnology, 1992, 10: 1455-1460. (65) Prior Publication Data Govind Ragupathi et al., “Antibodies against Tumor Cell Glycolipids and Proteins, but Not Mucins, Mediate Complement-Dependent US 2013/0177944 A1 Jul. 11,2013 Cytotoxicity”, The Journal of Immunology, 2005, 174: 5706-5712. Manjula P. Reddy et al., “Elimination of Fe Receptor-Dependent Effector Functions of a Modi?ed IgG4 Monoclonal Antibody to Related US. Application Data Human CD4”, The Journal of Immunology, 2000, 164: 1925-1933. Ellis L. Reinherz et al., “Separation of functional subsets of human T (62) Division of application No. 12/647,698, ?led on Dec. cells by amonoclonal antibody”, Immunology, Proc. Natl. Acad. Sci. 28, 2009, now Pat. No. 8,399,621. USA, 1979, 76(8): 4061-4065. David Rider et al., “A Human CD4 Monoclonal Antibody for the (60) Provisional application No. 61/141,393, ?led on Dec. Treatment of T-Cell Lymphoma Combines Inhibition of T-Cell Sig 30, 2008. naling by a Dual Mechanism with Potent Fc-Dependent Effector Activity”, Cancer Research, 2007, 67(20): 9945-9953. (30) Foreign Application Priority Data (Continued) Dec. 26, 2008 (JP) ...... 2008-331904 Primary Examiner * Laura B Goddard Assistant Examiner * Meera Natarajan (51) Int. Cl. (74) Attorney, Agent, or Firm * Sughrue Mion, PLLC C07H 21/04 (2006.01) (52) US. Cl. (57) ABSTRACT USPC ...... 536/2353; 530/387.1 An anti-CD4 antibody which binds to CD4, has a high a?inity Field of Classi?cation Search (58) and has a high effector activity, such as an antibody-depen None dent cellular cytotoxicity (ADCC activity) or complement See application ?le for complete search history. dependent cellular cytotoxicity (CDC activity), is required for a disease relating to a CD4-expressing cell. (56) References Cited The present invention can provide a monoclonal antibody or an antibody fragment thereof, which binds to a CD4 extra U.S. PATENT DOCUMENTS cellular region with high a?inity and also exhibits a high 7,037,496 B2* 5/2006 Ghrayeb et al...... 424/1331 ADCC activity or a high CDC activity; a hybridoma which 7,214,775 B2 5/2007 Hanai et al. produces the antibody; a DNA which encodes the antibody; a 2006/0183195 A1 8/2006 Lonberg et al. vector which contains the DNA; a transforrnant obtainable by 2007/0148165 A1 6/2007 Shitara et al. introducing the vector; a process for producing an antibody or an antibody fragment thereof using the hybridoma or the FOREIGN PATENT DOCUMENTS transformant; and a therapeutic agent using the antibody or WO 9709351 A1 3/1997 the antibody fragment thereof or a diagnostic agent using the WO 97/13852 A1 4/1997 antibody or the antibody fragment thereof. WO 2007/011041 A1 1/2007 WO 2007009469 A2 1/2007 4 Claims, 15 Drawing Sheets US 8,877,913 B2 Page 2
(56) References Cited pincott Williams &Wilkins, Hagertown, MD, US, Feb. I, 2007, vol. 30, No. 2, pp. 190-202. European Patent Of?ce, Search Report dated Dec. 19, 2012 issued in OTHER PUBLICATIONS counterpart European Application No. 098350622. Choy, Ernest H. S., et al., “Chimaeric anti-CD4 monoclonal antibody Samuel Troadec et al., “Biological activities on T lymphocytes of a cross-linked by monocyte Fc gamma receptor mediates apoptosis of baculovirus-expressed chimeric recombinant IgGl antibody With human CD4 lymphocytes,” European Journal of Immunology, Oct. speci?city for the CDR3-like loop on the D1 domain of the CD4 1993, vol. 23, No. 10, pp. 2676-2681. molecule”, Clinical Immunology, 2006, 119: 38-50. Japanese Patent Of?ce, Communication dated Aug. 13, 2013 issued Troadec, Samuel, et al., “In Vitro Antitumoral Activity of in counterpart Japanese Patent Application No. 2010-544182. Baculovirus-expressed Chimeric Recombinant Ant-CD4 Antibody l3B8.2 on T-cell Lymphomas”, Journal of Immunotherapy, Lip * cited by examiner
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Patent NOV. 4, 2014 Sheet 15 0f 15 US 8,877,913 B2 mmv2.owmmoncwmw 2.91 39:32v6w>m0BEE.coszm?wcm;6am |<|38%.328Sa- |X|2.3me35:00 lo]38522:325:. Ill#852com.323:. cocv ocom cccw ocov (sww) ewn|0/\ Joumi US 8,877,913 B2 1 2 ANTI-CD4 ANTIBODY Patent Literature 5), and the like. These antibodies are anti bodies which exert their medicinal ef?cacy by speci?cally CROSS-REFERENCE TO RELATED attacking CD4-expressing cells which are target cells, and it is APPLICATIONS considered that the mechanism of medicinal ef?cacy is mainly due to an ADCC activity (Non-Patent Literatures 6 This is a divisional of Us. patent application Ser. No. and 7). Meanwhile, these antibodies are shown to be devoid of 12/647,698 (allowed), ?led Dec. 28, 2009, which claims pri a CDC activity which is generally known as one of the main ority to Us. Provisional Patent Application No. 61/141,393, mechanism of medicinal ef?cacy of therapeutic antibodies, ?led Dec. 30, 2008, and Japanese Patent Application No. like ADCC (Non-Patent Literatures 6 and 7). 2008-331904 ?led Dec. 26, 2008, the contents of which are incorporated herein by reference in their entirety. The potency of CDC activity is different depending on the subclass of antibodies, and human IgG1 and IgG3 subclasses BACKGROUND OF THE INVENTION have a high CDC activity. It is known that the intensity of CDC activity among the subclasses is generally in the order of 1. Field of the Invention IgG3 ngG1>>IgG2ngG4. In addition, there is a case where The present invention relates to a monoclonal antibody or the CDC activity of an antibody is exerted or is not exerted an antibody fragment thereof, which binds to human CD4 depending on an antigen to which the antibody binds (Non extracellular region with a high af?nity and also exhibits a Patent Literature 11). Accordingly, every antibody cannot high antibody-dependent cellular cytotoxicity (hereinafter, always exhibit the CDC activity. referred to as “ADCC activity”) or a high complement-depen 20 Among the anti-CD4 mAbs con?rmed hitherto, little is dent cellular cytotoxicity (hereinafter, referred to as “CDC activity”); a hybridoma which produces the antibody; a DNA known of the antibodies having a CDC activity. Uniquely, which encodes the antibody; a vector which contains the even though it is known that OKT4 exhibits a CDC activity DNA; a transformant obtainable by introducing the vector; a against a CD4-positive human cell line only when a rabbit process for producing an antibody or an antibody fragment 25 serum which is high in complement number is used (Non thereof using the hybridoma or the transformant; and a thera Patent Literature 8), no examples of an antibody which exhib peutic agent and a diagnostic agent using the antibody or the its a CDC activity against a CD4-positive human cell line antibody fragment thereof. when a human complement is used have been reported. 2. Brief Description of the Background Art A dissociation constant KD of an antibody for several Cluster of differentiation 4 (hereinafter, referred to as 30 known anti-CD4 mAbs has been reported. For example, anti “CD4”) is a glycoprotein having a molecular weight of about bodies shown in the following table have been reported to 55 kDa, which is expressed on the cell surface of most of have a dissociation constant KB of about 7 to 0.01 nM (Table thymic cells, about 2/3 of peripheral blood T cells, monocytes, 1). Upon the calculation of dissociation constants, there is a and macrophage. CD4 is a type I transmembrane protein in possibility that measurement values vary depending on assay which four immunoglobulin superfamily domains (desig 35 equipments, assay methods, and analysis methods. There nated in order as D1 to D4 from the N terminal to the cell fore, when comparing dissociation constants, the values membrane side) are present on the outside of the cells, and which were measured and analyzed under the same condi two N-linked sugar chains in total are bound to the domains tions are required. D3 to D4. CD4 binds to a major histocompatibility complex (MHC) class II molecule through D1 and D2 domains, and 40 TABLE 1 then activates the T cells. Further, it is also known that CD4 polymerizes through D3 and D4 domains. D1 domain of CD4 Clone Name Dissociation Constant [M] is known to serve as a receptor for a human immunode? 6G5* 7.1 X 10*9 ciency virus (hereinafter, referred to as “HIV”) (N on-Patent CE9.1* 3.2><10’ll Literature 1). 45 Leu—3a** 1.0 ><10*ll CD4 is also known as T4, and the gene has been cloned in 13B8.2*** 5.0><10’9 1985 (Non-Patent Literature 2), and the DNA sequence, the *WO97/13852, amino acid sequence and the three-dimensional structure of **Non-Patent Literature 9, CD4 are publicly available from a known database. For ** *Non-Patent Literature 10 example, these can be obtained by reference to Accession 50 Nos. P01730 (SWISSPROT), M12807 (EMBL), and the like. Non-Patent Literature 1: Leucocyle Typing VI, 49 (1997) The anti-CD4 monoclonal antibody OKT4 was ?rst con Non-Patent Literature 2: Cell, 42, 93 (1985) ?rmed as a monoclonal antibody (hereinafter, referred to as “mAb”) which binds to CD4 (Non-Patent Literature 3). Since Non-Patent Literature 3: Proc. Natl. Acad. Sci. USA, 76, 4061 then, a large number of monoclonal antibodies against CD4 55 (1979) (hereinafter, referred to as “anti-CD4 mAbs”) have been Non-Patent Literature 4: Blood, 109, 4655 (2007) reported. Most of anti-CD4 mAbs reported hitherto are Non-Patent Literature 5: Lancet, 352, 1109 (1998) known to recognize D1 domain (Non-Patent Literature 1). Non-Patent Literature 6: Clin Immunollmmunopalhol, 84, 73 Some of anti-CD4 mAbs are under clinical development for the purpose of treating cancers, immune diseases, and infec 60 (1997) tions. For example, based on the fact that the binding between Non-Patent Literature 7: Cancer Res, 67, 9945 (2007) CD4 and HIV is essential for the infection of HIV, an antibody Non-Patent Literature 8: J Immunol, 164, 1925 (2000) which recognizes D1 domain of CD4 can inhibit the infection Non-Patent Literature 9: J Immunol, 145, 2839 (1990) of HIV, under the development as an HIV therapeutic agent. Examples of the anti-CD4 mAb developed as a therapeutic 65 Non-Patent Literature 10: Clinical Immunology, 119, 38 agent for cancers or immune diseases include zanolimumab (2006) (6G5) (Non-Patent Literature 4), keliximab (CE9.1) (Non Non-Patent Literature 11: J. Immunol, 174, 5706 (2005) US 8,877,913 B2 3 4 SUMMARY OF THE INVENTION FIGS. 9(A) to 9(D) show an antibody-dependent cellular cytotoxicity (ADCC activity) of various anti-CD4 antibodies An object of the present invention is to provide a mono on various human T cell lines. The ordinate represents a clonal antibody or an antibody fragment thereof, which binds cellular cytotoxicity rate (%), and the abscissa represents an to a CD4 extracellular region with high af?nity and also antibody concentration of each antibody. A in the broken line exhibits a high ADCC activity or CDC activity; a hybridoma represents a monoclonal antibody 6G5-l, A in the solid line which produces the antibody; a DNA which encodes the represents a monoclonal antibody 6G5-P, A represents a chi antibody; a vector which contains the DNA; a transformant meric antibody 6G5-113F, . represents a chimeric antibody obtainable by introducing the vector; a process for producing KM4045, I represents a chimeric antibody KM4046, B rep an antibody or an antibody fragment thereof using the hybri resents a chimeric antibody KM4047, x in the broken line represents a chimeric antibody KM4048, and x in the solid doma or the transformant; and a therapeutic agent or a diag line represents a chimeric antibody KM4049. FIG. 9(A) nostic agent using the antibody or the antibody fragment shows anADCC activity when HPB-ALL was used as a target thereof. cell. FIG. 9(B) shows an ADCC activity when HUT78 was used as a target cell. FIG. 9(C) shows anADCC activity when BRIEF DESCRIPTION OF THE DRAWINGS SUP-Tl is used as a target cell. FIG. 9(D) shows an ADCC activity when HPB-ALL was used as a target cell. FIG. 1 shows a construction process of a CD4 expression FIGS. 10(A) to 10(B) show a complement-dependent cel vector. lular cytotoxicity (CDC activity) of various anti-CD4 anti FIG. 2 shows a construction process of a CD4-Fc expres 20 bodies on a CD4-expressing transfectant or a human lym sion vector. phoma cell line. The ordinate represents a cellular FIGS. 3(A) to 3(D) show the reactivity of an anti-CD4 rat cytotoxicity rate (%), and the abscissa represents an antibody monoclonal antibody and an anti-CD4 mouse antibody concentration of each antibody. FIG. 10(A) shows a CDC OKT4 for the human T cell lymphoma cell line HPB-ALL in activity on a CD4-expressing transfectant. FIG. 10(B) shows the FCM analysis. The graph is divided into (A) to (D) 25 a CDC activity on the human lymphoma cell line HPB-ALL. according to the subclass of antibodies. The abscissa repre Q in the broken line represents an anti-CD4 human antibody sents a concentration of each of the antibodies, and the ordi 6G5-l, Q in the solid line represents an anti-CD4 human nate represents the mean ?uorescence intensity, MFI value. In antibody 6G5-113F, A represents an anti-CD4 chimeric anti FIG. 3(A), . represents an anti-CD4 rat monoclonal anti body KM4045, . represents an anti-CD4 chimeric antibody body KM4065, and x represents an anti-CD4 rat monoclonal 30 KM4046, I represents an anti-CD4 chimeric antibody antibody KM4069. In FIG. 3(B), x represents an anti-CD4 rat KM4047, - represents an anti-CD4 chimeric antibody monoclonal antibody KM4068. In FIG. 3(C), I represents an KM4048, I represents an anti-CD4 chimeric antibody anti-CD4 rat monoclonal antibody KM4066, and B repre KM4049, and I in the broken line represents an anti-CD4 sents an anti-CD4 rat monoclonal antibody KM4067. In FIG. mouse antibody OKT4. 3(D), . represents an anti-CD4 mouse monoclonal antibody 35 FIG. 11 shows the reactivity of various anti-CD4 antibod OKT4. ies for a CD4-positive T cell lymphoma cell line HPB-ALLin FIG. 4 shows a complement-dependent cellular cytotoxic the use of a ?ow cytometer. The abscissa represents an anti ity (CDC activity) of an anti-CD4 rat monoclonal antibody body concentration (ug/mL), and the ordinate represents a and an anti-CD4 mouse monoclonal antibody OKT4 on a mean ?uorescence intensity, MFI value. . represents a chi CD4-expressing transfectant. The ordinate represents a cel 40 meric antibody KM4045, B represents a humanized antibody lular cytotoxicity rate (%), and the abscissa represents an HVOLVO, 0 represents a humanized antibody HVZLVO, <> antibody concentration of an anti-CD4 monoclonal antibody. represents a humanized antibody HV3LVO, - represents a . represents an anti-CD4 rat monoclonal antibody KM4066, humanized antibody HV4LVO, x represents a humanized I represents an anti-CD4 rat monoclonal antibody KM4067, antibody HV4LV6, A in the broken line represents a human irepresents an anti-CD4 rat monoclonal antibody KM4068, 45 antibody 6G5-l, and A in the solid line represents a human and 0 represents an anti-CD4 mouse monoclonal antibody antibody 6G5-P. OKT4. FIGS. 12(A) to 12(C) show an antibody-dependent cellular FIG. 5 shows a cloning process of an anti-CD4 rat mono cytotoxicity (ADCC activity) of various anti-CD4 antibodies clonal antibody. on various human T cell lymphoma lines. The ordinate rep FIG. 6 shows a construction process of an anti-CD4 chi 50 resents a cellular cytotoxicity rate (%), and the abscissa rep meric antibody expression vector. resents an antibody concentration (ng/mL). . represents a FIG. 7 shows a construction process of expression vectors chimeric antibody KM4045, 0 represents a humanized anti of anti-CD4 human antibodies 6G5-l and 6G5-113F. body KM8045, A in the broken line represents a human FIG. 8 shows the reactivity of various anti-CD4 antibodies antibody 6G5-l, and A in the solid line represents a human in the binding ELISA. The abscissa represents a concentra 55 antibody 6G5-P. FIG. 12(A) shows an ADCC activity when tion of each of the antibodies, and the ordinate represents a HPB-ALL was used as a target cell. FIG. 12(B) shows an binding activity of each of the antibodies. The reactivity of ADCC activity when HUT78 was used as a target cell. FIG. each antibody against recombinant human CD4 is shown. A 12(C) shows an ADCC activity when CCRF-CEM was used in the broken line represents a monoclonal antibody 6G5- l, A as a target cell. in the solid line represents an anti-CD4 human antibody 6G5 60 FIG. 13 shows anti-tumor effect (early stage cancer model) P, A represents an anti-CD4 human antibody 6G5-113F, . of each antibody on the human T cell lymphoma cell line HH represents an anti-CD4 chimeric antibody KM4045, I rep which had been transplanted into a SCID mouse. The abscissa resents an anti-CD4 chimeric antibody KM4046, B repre represents the number of days after tumor transplantation, sents an anti-CD4 chimeric antibody KM4047, x in the bro and the ordinate represents a tumor volume. x represents a ken line represents an anti-CD4 chimeric antibody KM4048, 65 negative control group, A represents a KM8045-adminis and x in the solid line represents an anti-CD4 chimeric anti tered group, and . represents a 6G5-P-administered group. body KM4049. The bar represents a standard deviation. US 8,877,913 B2 5 6 FIGS. 14(A) and 14(B) show anti-tumor effects (metasta wherein the monoclonal antibody is a monoclonal anti sis model) of individual antibodies on CD4/EL4 cells which body which binds to the same epitope in an extracellular had been transplanted into a C57BL/6 mouse. In FIG. 14(A), region of CD4 as an epitope to which an antibody in which the ordinate represents a liver weight ratio. In FIG. 14(B), the CDRs 1 to 3 of an H chain of the antibody comprise the ordinate represents a kidney weight ratio. The bar represents amino acid sequences of SEQ ID NOs:27 to 29, respec a standard deviation. tively, and CDRs 1 to 3 of an L chain of the antibody FIG. 15 shows anti-tumor effects (progressive cancer comprise the amino acid sequences of SEQ ID NOsz30 to model) of each antibody on an HH cell line which had been 32, respectively, binds; transplanted into a SCID mouse. The abscissa represents the (8) The antibody or the antibody fragment thereof described number of days after tumor transplantation, and the ordinate in any one of the above (1) to (7), wherein the monoclonal represents a tumor volume. x represents a negative control antibody is a recombinant antibody; group, A represents a group to which 20 ug/head of KM8045 (9) The recombinant antibody or the antibody fragment was administered, . represents a group to which 100 ug/head thereof described in the above (8), wherein the recombi of KM8045 was administered, and I represents a group to nant antibody is a recombinant antibody selected from a which 200 ug/head of KM8045 was administered. The bar represents a standard deviation. human chimeric antibody, a humanized antibody and a human antibody; DETAILED DESCRIPTION OF THE INVENTION (10) The recombinant antibody or the antibody fragment thereof described in the above (9), wherein an H chain The present invention relates to the following (1) to (35): 20 variable region (hereinafter, referred to as “VH”) of the (1) A monoclonal antibody against human CD4 or an anti antibody and an L chain variable region (hereinafter, body fragment thereof, which has a dissociation constant referred to as “VL”) of the antibody comprise the amino (hereinafter, referred to as “KD”) less than 1><10_9M of an acid sequences of any one group selected from SEQ ID antibody to an antigen, binds to an extracellular region of NO:16 and SEQ ID NO:26, respectively; and SEQ ID human CD4 with high af?nity and has a high antibody 25 NO: 12 and SEQ ID NO:22, respectively; dependent cellular cytotoxicity (hereinafter, referred to as (11) The recombinant antibody or the antibody fragment “ADCC activity”); thereof described in the above (9), wherein CDRs 1 to 3 of (2) The monoclonal antibody or the antibody fragment an H chain of the antibody and CDRs 1 to 3 of the L chain thereof described in the above (1 ), wherein the antibody is of an antibody comprise the amino acid sequences of any an antibody having a high complement-dependent cellular 30 one group selected from SEQ ID NOs:51 to 53 and SEQ ID cytotoxicity (CDC activity); NOs:54 to 56, respectively; and SEQ ID NOs:27 to 29 and (3) The monoclonal antibody or the antibody fragment thereof described in the above (2), wherein the antibody is SEQ ID NOsz30 to 32, respectively; (12) The humanized antibody or the antibody fragment an antibody which has a CDC activity on a human cancer thereof described in the above (9), wherein the recombi cell line expressing human CD4; 35 (4) The monoclonal antibody or the antibody fragment nant antibody is a humanized antibody, and VH and VL of thereof described in any one of the above (1 ) to (3), wherein the antibody comprise the amino acid sequences of any one the monoclonal antibody is a monoclonal antibody which group selected from SEQ ID NO:77 and SEQ ID NO:78, competes with an antibody in which complementarity respectively; SEQ ID NO:96 and SEQ ID NO:78, respec determining regions (hereinafter, referred to as “CDRs”) 1 40 tively; SEQ ID NO:98 and SEQ ID NO:78, respectively; to 3 of a heavy chain (hereinafter, referred to as “H chain”) SEQ ID NO:100 and SEQ ID NO:78, respectively; and of the antibody comprise the amino acid sequences of SEQ SEQ ID NO: 100 and SEQ ID NO: 102, respectively; ID NOs:51 to 53, respectively, and CDRs 1 to 3 of a light (13) The antibody or the antibody fragment thereof described chain (hereinafter, referred to as “L chain”) of the antibody in any one of the above (1) to (13), wherein the antibody comprise the amino acid sequences of SEQ ID NOs:54 to 45 fragment is an antibody fragment selected from Fab, Fab', 56, respectively, to bind to an extracellular region of CD4; F(ab')2, a single chain antibody (scFv), a dimerized V (5) The monoclonal antibody or the antibody fragment region (diabody), a disul?de stabilizedV region (dst) and thereof described in any one of the above (1) to (4), the a peptide comprising CDR; monoclonal antibody is a monoclonal antibody which (14) A DNA which encodes the antibody or the antibody binds to the same epitope as an epitope in an extracellular 50 fragment thereof described in any one of the above (1) to region of CD4 to which an antibody in which CDRs 1 to 3 (13); of an H chain of the antibody comprise the amino acid (15) A recombinant vector which comprises the DNA sequences of SEQ ID NOs:51 to 53, respectively, and described in the above (14); CDRs 1 to 3 of an L chain of the antibody comprise the (16) A transformant obtainable by introducing the recombi amino acid sequences of SEQ ID NOs:54 to 56, respec 55 nant vector described in the above (15) into a host cell; tively, binds; (17) A process for producing the monoclonal antibody or the (6) The monoclonal antibody or the antibody fragment antibody fragment thereof described in any one of the thereof described in any one of the above (1 ) to (2), wherein above (1) to (13), comprising culturing the transformant the monoclonal antibody is a monoclonal antibody which described in the above (16) in a medium to produce and competes with an antibody in which CDRs 1 to 3 of an H 60 accumulate the antibody or the antibody fragment thereof chain of the antibody comprise the amino acid sequences described in any one of the above (1) to (14) in the culture, of SEQ ID NOs:27 to 29, respectively, and CDRs 1 to 3 of and collecting the antibody or the antibody fragment an L chain of the antibody comprise the amino acid thereof from the culture; sequences of SEQ ID NOsz30 to 32, respectively, to bind to (18) A method for immunologically detecting or measuring an extracellular region of CD4. 65 human CD4, comprising using the antibody or the antibody (7) The monoclonal antibody or the antibody fragment fragment thereof described in any one of the above (1) to thereof described in any one of the above (1) to (2) and (6), (13);