CN111848806A - Egfr-cd3双功能抗体及其应用 - Google Patents

Egfr-cd3双功能抗体及其应用 Download PDF

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CN111848806A
CN111848806A CN202010559393.5A CN202010559393A CN111848806A CN 111848806 A CN111848806 A CN 111848806A CN 202010559393 A CN202010559393 A CN 202010559393A CN 111848806 A CN111848806 A CN 111848806A
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叶才果
王笑非
汪波
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Abstract

本发明提供了一种EGFR‑CD3双功能抗体，所述双功能抗体包括2条相同的轻链和2条相同的重组重链，其中每条所述轻链的氨基酸序列如SEQ ID NO.8所示，每条所述重组重链由EGFR抗体重链与CD3蛋白的单链抗体scFv通过接头序列连接组成，所述EGFR抗体重链的氨基酸序列如SEQ ID NO.7所示，所述接头序列linker的氨基酸序列如SEQ ID NO.9所示，所述CD3蛋白的单链抗体scFv的氨基酸序列如SEQ ID NO.1、SEQ ID NO.2、SEQ ID NO.3、SEQ ID NO.4、SEQ ID NO.5或SEQ ID NO.6的任一项所示。本发明的EGFR‑CD3双功能抗体能够与人CD3蛋白和EGFR特异性结合，降低鼠源抗体的免疫原性，同时有效激活T细胞，并阻断人EGF与EGFR的结合，可以通过结合EGFR锚定在肿瘤细胞表面，并通过与CD3结合激活T细胞，起到杀伤肿瘤细胞的目的。

Description

EGFR-CD3双功能抗体及其应用
技术领域
本发明涉及抗体，具体涉及能够阻断人EGF与EGFR结合并激活T细胞的双功能抗体。
背景技术
CD3蛋白是T细胞表面表达从刺激信号分子，其在T细胞激活起着非常重要的作用。CD3分子与TCR组成复合受体分子，激活酪氨酸激酶，促使CD3分子免疫受体酪氨酸活化基序中的酪氨酸(Y)磷酸化。磷酸化的酪氨酸(pY)进一步磷酸化下游含酪氨酸的蛋白，从而引起激酶活化的级联反应，调控T细胞增殖和活化的靶基因，引起基因的表达和转录，T细胞由静止状态转为增殖和活化状态。CD3分子具有稳定TCR结构和传递活化信号的作用。针对CD3分子的单克隆抗体能够激发或阻断T细胞活化信号转导，清除效应T细胞或诱导调节T细胞的产生，为治疗器官移植排斥和自身免疫性疾病提供了新的方法。
鼠源性单克隆抗体Muromonab-CD3(莫罗单抗，克隆号OKT3)能够显著激活T细胞，引起细胞细胞增殖。莫罗单抗是一种鼠单克隆抗体，用于治疗肾、心、肝移植后的急性排异反应，其主要机理是通过诱导调节性T细胞(Treg)增殖，并抑制杀伤性T(Teff)功能活性。其临床应用会引起宿主排斥反应，儿童患者使用该药可能增加致严重的神经系统并发症的风险，尤其是脑水肿和脑疝。借助抗体人源化的技术，可以降低鼠源性抗体的免疫原性。抗体人源化技术主要是指通过对鼠源(或其他动物种属)的单克隆抗体的氨基酸序列进行替换，使替换后的蛋白序列为人源序列或与之相似，同时保留单克隆抗体的亲和力和特异性，又降低了其对人体的异源性。抗体人源化通常对鼠源(或其他动物种属)抗体的可变区(CDR)进行氨基酸改变，而对抗体的恒定区部分(即CH和CL区)序列进行人类抗体基因序列的改变或替换。人源化抗体可以大大减少抗体药物对人类机体造成的例如，宿主免疫排斥，抗药抗体产生等毒副作用。
EGFR单抗药物，西妥昔单抗(Cetuximab)是第一个在全球多个国家获准上市的靶向作用于表皮生长因子受体(EGFR)的IgG1亚型人-鼠嵌合单克隆抗体，被批准用于治疗转移性结直肠癌和难治性头颈癌。Cetuximab通过靶向结合EGFR阻断EGFR与其天然配体EGF结合，从而打断细胞自泌性无限增殖的恶性循环，抑制肿瘤细胞增殖，诱导细胞凋亡，抑制内皮细胞增生和新生血管形成，抑制癌细胞侵袭、转移，增强细胞毒药物、电离辐射的抗癌作用。
双功能抗体是指具有双特异性的抗体，具有结合两种不同抗原的能力，是一种非天然抗体。通常采用基因工程法构建多种功能、多种用途的双功能抗体。双功能抗体作为一种新的药物在临床治疗中具有非常大的应用价值。T细胞重定向双功能抗体(TRBAs)是一种基于CD3抗体招募T细胞至靶细胞周围的一种双功能抗体。该双功能抗体的两臂能同时结合CD蛋白和肿瘤相关抗原，一方面通过结合CD3蛋白招募和激活T细胞介导肿瘤免疫，另一方面通过结合肿瘤相关抗原(TAA)把T细胞和肿瘤细胞拉近，这样可以更高效、更特异地杀伤肿瘤细胞，显著降低免疫毒副作用。
发明内容
本发明的目的在于结合西妥昔单抗和CD3单抗两种抗体的优势，提供了靶向肿瘤相关抗原和T细胞的双功能抗体，该双功能抗体既能阻断人EGF与EGFR的结合，还能招募免疫T细胞到肿瘤组织周围并激活T细胞，分泌IFNγ细胞因子，同时克服鼠源性CD3抗体的缺陷，降低免疫原性。
为实现上述目的，本发明提供了一种EGFR-CD3双功能抗体，所述双功能抗体包括2条相同的轻链和2条相同的重组重链，其中每条所述轻链的氨基酸序列如SEQ ID NO.8所示，每条所述重组重链由EGFR抗体重链与CD3蛋白的单链抗体scFv通过接头序列连接组成，所述EGFR抗体重链的氨基酸序列如SEQ ID NO.7所示，所述接头序列linker的氨基酸序列如SEQ ID NO.9所示，所述CD3蛋白的单链抗体scFv的氨基酸序列如SEQ ID NO.1、SEQ IDNO.2、SEQ ID NO.3、SEQ ID NO.4、SEQ ID NO.5或SEQ ID NO.6所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.10所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.11所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.12所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.13所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.14所示。
作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.15所示。
另一方面，本发明还提供了一种多核苷酸序列，所述多核苷酸序列编码所述的EGFR-CD3双功能抗体。
另一方面，本发明还提供了一种药物组合物，所述药物组合物包含所述的EGFR-CD3双功能抗体。
另一方面，本发明还提供了所述EGFR-CD3双功能抗体在制备抗肿瘤药物中的应用。
优选地，所述肿瘤包括但不限于结直肠癌和/或头颈肿瘤。
本发明提供的EGFR-CD3双功能抗体对鼠源人鼠嵌合抗体OKT3的高度可变区(包括VH和VL)进行了人源化改造，并用特定接头序列进行融合连接，成为单链抗体(scFv)，同时具有特异性结合人CD3蛋白与特异性结合人EGFR蛋白的作用，既能阻断人EGF与EGFR的结合，还能招募免疫T细胞到肿瘤组织周围并激活T细胞，诱导分泌IFNγ细胞因子，促进T细胞分化增殖用，对EGFR表达的上皮癌细胞A431具有杀伤作用，同时克服鼠源性CD3抗体的缺陷，降低免疫原性，能够用于治疗EGFR阳性表达的实体瘤，特别是结直肠肿瘤和头颈肿瘤。
附图说明
图1是本发明的EGFR-CD3双功能抗体的结构示意图。
图2是采用流式细胞方法检测本发明的EGFR-CD3双功能抗体对细胞表面CD3蛋白的结合能力的结果。
图3为采用流式细胞方法检测本发明的EGFR-CD3双功能抗体对细胞表面EGFR蛋白的结合能力的结果。
图4示出了EGFR-CD3双功能抗体对肿瘤细胞的杀伤作用。
具体实施方式
为更好地说明本发明的目的、技术方案和优点，下面将结合具体实施例对本发明作进一步说明。
值得注意的是，出于简要清楚的目的，以下实施例中对一些常规的技术操作步骤、试剂、仪器并未进行细致的描述，但应理解，如未特别说明，这些常规技术操作步骤、试剂、仪器对本领域普通技术人员而言是显而易见的。
下面结合具体实例来进一步描述本发明，本发明的优点和特点将会随着描述而更为清楚。但实施例仅是范例性的，并不对本发明的范围构成任何限制。本领域技术人员应该理解的是，在不偏离本发明的精神和范围下可以对本发明技术方案的细节和形式进行修改或替换，但这些修改和替换均落入本发明的保护范围内。
以下实施例内容包括CD3的单链抗体scFv的人源化改造和双功能抗体的构建。
本发明的EGFR-CD3双功能抗体中的CD3单链抗体的人源化通过分别进行重链可变区的框架区(Framwork)和抗原互补决定簇区域(CDR)进行人源氨基酸的逐步替换来实现。首先进行人鼠嵌合抗体scFv的构建，将克隆号为OKT3的VH和VL用连接序列(GGGGSGGGGSGGGGSGGGS，SEQ ID NO.16)进行连接。先对鼠源的scFv的框架区进行人源序列的替换，替换后的序列称为OKT-H1L1，见SEQ ID NO.1。随后，分别对鼠源的序列进行人源序列的逐步替换，分别形成人源化scFv，分别命名OKT-H2L2(SEQ ID NO.2)、OKT-H3L3(SEQ IDNO.3)、OKT-H4L1(SEQ ID NO.4)、OKT-H3L1(SEQ ID NO.5)和OKT-H3L2(SEQ ID NO.6)，人源化程度不断提高，接近全人源化水平。
本发明的EGFR-CD3双功能抗体的结构示意图如图1所示，其中重组重链由以下部分构成：Cetuximab抗体的完整重链-linker序列-OKT的scFv序列。EGFR-CD3双功能抗体的轻链由Cetuximab的轻链Cetuxi-L构成，序列如SEQ ID NO.8所示。Cetuximab抗体的完整重链Cetuxi-H的氨基酸序列如SEQ ID NO.7所示。
本发明的一种优选EGFR-CD3双功能抗体Cetuxi-OKT11，由重组重链Cetuxi-H1L1和轻链Cetuxi-L构成，重组重链Cetuxi-H1L1由Cetuximab抗体的完整重链、linker序列、OKT-H1L1连接而成，其氨基酸序列如SEQ ID NO.10所示。
本发明的另一种优选EGFR-CD3双功能抗体Cetuxi-OKT22，由重组重链Cetuxi-H2L2和轻链Cetuxi-L构成，重组重链Cetuxi-H2L2由Cetuximab抗体的完整重链、linker序列、OKT-H2L2连接而成，其氨基酸序列如SEQ ID NO.11所示。
本发明的另一种优选EGFR-CD3双功能抗体Cetuxi-OKT33，由重组重链Cetuxi-H3L3和轻链Cetuxi-L构成，重组重链Cetuxi-H3L3由Cetuximab抗体的完整重链、linker序列、OKT-H3L3连接而成，其氨基酸序列如SEQ ID NO.12所示。
本发明的另一种优选EGFR-CD3双功能抗体Cetuxi-OKT41，由重链Cetuxi-H4L1和轻链Cetuxi-L构成，重组重链Cetuxi-H4L1由Cetuximab抗体的完整重链、linker序列、OKT-H4L1连接而成，其氨基酸序列如SEQ ID NO.13所示。
本发明的另一种优选EGFR-CD3双功能抗体Cetuxi-OKT21，由重链Cetuxi-H2L1和轻链Cetuxi-L构成，重组重链Cetuxi-H2L1由Cetuximab抗体的完整重链、linker序列、OKT-H2L1连接而成，其氨基酸序列如SEQ ID NO.14所示。
本发明的另一种优选EGFR-CD3双功能抗体Cetuxi-OKT32，由重链Cetuxi-H3L2和轻链Cetuxi-L构成，重组重链Cetuxi-H3L2由Cetuximab抗体的完整重链、linker序列、OKT-H3L2连接而成，其氨基酸序列如SEQ ID NO.15所示。
实施例1：EGFR-CD3双功能抗体的抗体制备
根据EGFR-CD3双功能抗体的重链和轻链氨基酸序列匹配情况，分别人工合成双功能抗体的重链Cetuxi-H1L1、Cetuxi-H2L2、Cetuxi-H3L3、Cetuxi-H4L1、Cetuxi-H2L1、Cetuxi-H3L2和轻链Cetuxi-L的cDNA并构建到pcDNA3.1载体中。将测序鉴定正确的双功能抗体的轻链的质粒和重链的质粒按照1：1的重量比，共转染到293F细胞中。
电转条件：4mm点击杯，电压250v，5mm，细胞1×108。电转完成的细胞置于37℃，5％CO2，130rpm/min培养7天。细胞培养7天后，离心收集上清，将上清6000rpm离心10min，加入终浓度为200mM NaCl和50mM Tris-HCl(pH7.2)，并用0.45μm滤膜过滤收集滤液。样品用HiTrap MabSelect柱(GE公司)进行纯化。由此得到双功能抗体Cetuxi-OKT11、Cetuxi-OKT22、Cetuxi-OKT33、Cetuxi-OKT41、Cetuxi-OKT21和Cetuxi-OKT32。
实施例2：EGFR-CD3双功能抗体对Jurkat细胞表面CD3蛋白结合活性检测
用流式细胞技术对各EGFR-CD3双功能抗体(Cetuxi-OKT11、Cetuxi-OKT22、Cetuxi-OKT33、Cetuxi-OKT41、Cetuxi-OKT21和Cetuxi-OKT32)进行细胞表面蛋白结合力检测。检测EGFR-CD3双功能抗体对Jurkat细胞表面CD3蛋白的结合力。设置实验的阴性对照为人IgG抗体，阳性对照为人源化的OKT克隆抗体(义翘神州产品)，实验组为各表达纯化的EGFR-CD3双功能抗体。设置检测细胞为1×106/管，加入各抗体2μg/ml，于冰上孵育30min。离心洗涤细胞后，加入Alexa 488荧光素标记的鼠抗人二抗(1：200稀释)，于冰上孵育30min。孵育完成后，离心洗涤细胞。用流式细胞仪检测细胞的荧光标记情况。
各EGFR-CD3双功能抗体对CD3蛋白的亲和力如图2所示。如图所示，各人源化改造的双功能抗体(Cetuxi-OKT11、Cetuxi-OKT22、Cetuxi-OKT33、Cetuxi-OKT41、Cetuxi-OKT21和Cetuxi-OKT32)与Jurkat细胞表面CD3蛋白的结合情况显示，随着人源化程度的增加，其结合CD3蛋白的能力有差别，其中以Cetuxi-OKT22，Cetuxi-OKT33和Cetuxi-OKT32最强，Cetuxi-11最弱，但这些人源化改造的双功能抗体均能有效结合CD3蛋白。
实施例3：EGFR-CD3双功能抗体对上皮瘤细胞A431细胞表面EGFR蛋白结合活性检测
用流式细胞技术对各双功能抗体进行细胞表面蛋白结合力检测。检测双功能抗体对A431细胞表面EGFR蛋白的结合力。设置实验的阴性对照为人IgG抗体，阳性对照为人鼠嵌合抗体(爱必妥(Erbitux)单抗，即西妥昔单抗)，实验组为各表达纯化的双功能抗体。设置检测细胞为1×106/管，加入各抗体2μg/ml，于冰上孵育30min。离心洗涤细胞后，加入Alexa488荧光素标记的鼠抗人二抗(1：200稀释)，于冰上℃孵育30min。孵育完成后，离心洗涤细胞。用流式细胞仪检测细胞的荧光标记情况。
各EGFR-CD3双功能抗体对EGFR蛋白的亲和力如图3所示。随着人源化程度的增加，各人源化改造的双功能抗体(Cetuxi-OKT11、Cetuxi-OKT22、Cetuxi-OKT33、Cetuxi-OKT41、Cetuxi-OKT21和Cetuxi-OKT32)结合EGFR蛋白的能力各不相同，其中以Cetuxi-OKT21和Cetuxi-OKT32最强，Cetuxi-OKT22和Cetuxi-11最弱。
实施例4：EGFR-CD3双功能抗体对T细胞介导的肿瘤细胞杀伤能力检测
检测EGFR-CD3双功能抗体在肿瘤细胞周围募集T细胞并激活T细胞对肿瘤细胞的杀伤作用。具体地，中96孔细胞培养板中，贴壁培养上皮瘤细胞A431细胞，按照5000/孔加入细胞。细胞培养24h后，按上皮瘤细胞A431与T细胞Jurkat的数量比例为1：10，加入Jurkat细胞。
设置实验的阴性对照，加入人IgG抗体；阳性对照为人源化的OKT克隆抗体(#Cat:10977-H001，义翘神州)组，和OKT+CD28蛋白抗体(#GMP-11524-H001)联用组；实验组为各表达纯化的EGFR-CD3双功能抗体(Cetuxi-OKT11、Cetuxi-OKT22、Cetuxi-OKT33、Cetuxi-OKT41、Cetuxi-OKT21和Cetuxi-OKT32)。
实验结果显示(图4)，各人源化双功能抗体(Cetuxi-OKT22、Cetuxi-OKT41和Cetuxi-OKT32)对A431肿瘤细胞和HEK293细胞的杀伤力有所区别，对A431肿瘤细胞杀伤力比单独的OKT3单抗和OKT+CD28单抗联用的效果更好，而对HEK293细胞的杀伤力没有显著差异。
序列表
<110> 佛山安普泽生物医药股份有限公司
<120> EGFR-CD3双功能抗体及其应用
<160> 16
<170> SIPOSequenceListing 1.0
<210> 1
<211> 244
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 1
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Arg Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Leu Thr Thr Asp Lys Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Leu Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys
165 170 175
Pro Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln
180 185 190
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
195 200 205
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
210 215 220
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
225 230 235 240
Leu Glu Ile Lys
<210> 2
<211> 244
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 2
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Leu Thr Arg Asp Lys Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys
165 170 175
Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu Gln
180 185 190
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
195 200 205
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
210 215 220
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
225 230 235 240
Leu Glu Ile Lys
<210> 3
<211> 245
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 3
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met
35 40 45
Gly Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp Tyr Gln Gln
165 170 175
Lys Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu
180 185 190
Gln Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp
195 200 205
Phe Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr
210 215 220
Tyr Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr
225 230 235 240
Lys Leu Glu Ile Lys
245
<210> 4
<211> 244
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 4
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met
35 40 45
Gly Tyr Ile Asn Pro Ser Asn Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys
165 170 175
Pro Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln
180 185 190
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
195 200 205
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
210 215 220
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
225 230 235 240
Leu Glu Ile Lys
<210> 5
<211> 244
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 5
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Leu Thr Arg Asp Lys Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys
165 170 175
Pro Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln
180 185 190
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
195 200 205
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
210 215 220
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
225 230 235 240
Leu Glu Ile Lys
<210> 6
<211> 244
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 6
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Thr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met
35 40 45
Gly Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly
100 105 110
Thr Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
115 120 125
Ser Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln
130 135 140
Ser Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr
145 150 155 160
Cys Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys
165 170 175
Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu Gln
180 185 190
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
195 200 205
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
210 215 220
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
225 230 235 240
Leu Glu Ile Lys
<210> 7
<211> 468
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 7
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys
465
<210> 8
<211> 234
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 8
Met Arg Val Pro Ala Gln Leu Leu Gly Leu Leu Leu Leu Trp Phe Pro
1 5 10 15
Gly Ala Arg Cys Asp Ile Leu Leu Thr Gln Ser Pro Val Ile Leu Ser
20 25 30
Val Ser Pro Gly Glu Arg Val Ser Phe Ser Cys Arg Ala Ser Gln Ser
35 40 45
Ile Gly Thr Asn Ile His Trp Tyr Gln Gln Arg Thr Asn Gly Ser Pro
50 55 60
Arg Leu Leu Ile Lys Tyr Ala Ser Glu Ser Ile Ser Gly Ile Pro Ser
65 70 75 80
Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Ser Ile Asn
85 90 95
Ser Val Glu Ser Glu Asp Ile Ala Asp Tyr Tyr Cys Gln Gln Asn Asn
100 105 110
Asn Trp Pro Thr Thr Phe Gly Ala Gly Thr Lys Leu Glu Leu Lys Arg
115 120 125
Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln
130 135 140
Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr
145 150 155 160
Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser
165 170 175
Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr
180 185 190
Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys
195 200 205
His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro
210 215 220
Val Thr Lys Ser Phe Asn Arg Gly Glu Cys
225 230
<210> 9
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 9
Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser
1 5 10
<210> 10
<211> 723
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 10
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Arg Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile Gly
515 520 525
Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Leu Thr Thr Asp Lys Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Leu Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys Pro
645 650 655
Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln Ser
660 665 670
Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
675 680 685
Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys
690 695 700
Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys Leu
705 710 715 720
Glu Ile Lys
<210> 11
<211> 723
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 11
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile Gly
515 520 525
Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Leu Thr Arg Asp Lys Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys Pro
645 650 655
Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu Gln Ser
660 665 670
Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
675 680 685
Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys
690 695 700
Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys Leu
705 710 715 720
Glu Ile Lys
<210> 12
<211> 724
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 12
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met Gly
515 520 525
Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Ile Ser Ser Tyr Leu Asn Trp Tyr Gln Gln Lys
645 650 655
Pro Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu Gln
660 665 670
Ser Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe
675 680 685
Thr Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr
690 695 700
Cys Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys
705 710 715 720
Leu Glu Ile Lys
<210> 13
<211> 723
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 13
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met Gly
515 520 525
Tyr Ile Asn Pro Ser Asn Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys Pro
645 650 655
Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln Ser
660 665 670
Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
675 680 685
Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys
690 695 700
Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys Leu
705 710 715 720
Glu Ile Lys
<210> 14
<211> 723
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 14
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile Gly
515 520 525
Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Leu Thr Arg Asp Lys Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys Pro
645 650 655
Gly Lys Ala Pro Lys Arg Trp Ile Tyr Asp Thr Ser Ser Leu Gln Ser
660 665 670
Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
675 680 685
Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys
690 695 700
Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys Leu
705 710 715 720
Glu Ile Lys
<210> 15
<211> 723
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 15
Met Glu Phe Leu Gly Leu Phe Leu Cys Leu Val Thr Phe Pro Ser Cys
1 5 10 15
Val Leu Ser Gln Val Gln Leu Lys Gln Ser Gly Pro Gly Leu Val Gln
20 25 30
Pro Ser Gln Ser Leu Ser Ile Thr Cys Thr Val Ser Gly Phe Ser Leu
35 40 45
Thr Asn Tyr Gly Val His Trp Val Arg Gln Ser Pro Gly Lys Gly Leu
50 55 60
Glu Trp Leu Gly Val Ile Trp Ser Gly Gly Asn Thr Asp Tyr Asn Thr
65 70 75 80
Pro Phe Thr Ser Arg Leu Ser Ile Asn Lys Asp Asn Ser Lys Ser Gln
85 90 95
Val Phe Phe Lys Met Asn Ser Leu Gln Ser Asn Asp Thr Ala Ile Tyr
100 105 110
Tyr Cys Ala Arg Ala Leu Thr Tyr Tyr Asp Tyr Glu Phe Ala Tyr Trp
115 120 125
Gly Gln Gly Thr Leu Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro
130 135 140
Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr
145 150 155 160
Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr
165 170 175
Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro
180 185 190
Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr
195 200 205
Val Pro Ser Ser Ser Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn
210 215 220
His Lys Pro Ser Asn Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser
225 230 235 240
Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu
245 250 255
Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu
260 265 270
Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
275 280 285
His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
290 295 300
Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr
305 310 315 320
Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu Asn
325 330 335
Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro
340 345 350
Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln
355 360 365
Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val
370 375 380
Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val
385 390 395 400
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro
405 410 415
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr
420 425 430
Val Asp Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val
435 440 445
Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu
450 455 460
Ser Pro Gly Lys Gly Gly Ser Gly Gly Ser Ser Gly Gly Ser Ser Gln
465 470 475 480
Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala Ser
485 490 495
Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr Thr
500 505 510
Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met Gly
515 520 525
Tyr Ile Asn Pro Ser Arg Gly Tyr Thr Asn Tyr Ala Gln Lys Phe Gln
530 535 540
Gly Arg Val Thr Met Thr Arg Asp Thr Ser Thr Ser Thr Ala Tyr Met
545 550 555 560
Glu Leu Ser Ser Leu Arg Ser Glu Asp Thr Ala Val Tyr Tyr Cys Ala
565 570 575
Arg Tyr Tyr Asp Asp His Tyr Cys Leu Asp Tyr Trp Gly Gln Gly Thr
580 585 590
Thr Val Thr Val Ser Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
595 600 605
Gly Gly Gly Gly Ser Gly Gly Gly Ser Asp Ile Gln Met Thr Gln Ser
610 615 620
Pro Ser Ser Leu Ser Ala Ser Val Gly Asp Arg Val Thr Ile Thr Cys
625 630 635 640
Arg Ala Ser Gln Ser Val Ser Tyr Met Asn Trp Tyr Gln Gln Lys Pro
645 650 655
Gly Lys Ala Pro Lys Leu Leu Ile Tyr Asp Ala Ser Ser Leu Gln Ser
660 665 670
Gly Val Pro Ser Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
675 680 685
Leu Thr Ile Ser Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys
690 695 700
Gln Gln Trp Ser Ser Asn Pro Phe Thr Phe Gly Gln Gly Thr Lys Leu
705 710 715 720
Glu Ile Lys
<210> 16
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 16
Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly
1 5 10 15
Gly Gly Ser

Claims (10)

1.一种EGFR-CD3双功能抗体，所述双功能抗体包括2条相同的轻链和2条相同的重组重链，其中每条所述轻链的氨基酸序列如SEQ ID NO.8所示，每条所述重组重链由EGFR抗体重链与CD3蛋白的单链抗体scFv通过接头序列连接组成，所述EGFR抗体重链的氨基酸序列如SEQ ID NO.7所示，所述接头序列linker的氨基酸序列如SEQ ID NO.9所示，所述CD3蛋白的单链抗体scFv的氨基酸序列如SEQ ID NO.1、SEQ ID NO.2、SEQ ID NO.3、SEQ ID NO.4、SEQID NO.5或SEQ ID NO.6的任一项所示。
2.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，所述重组重链的氨基酸序列如SEQ ID NO.10所示。
3.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，所述重组重链的氨基酸序列如SEQ ID NO.11所示。
4.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，所述重组重链的氨基酸序列如SEQ ID NO.12所示。
5.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，作为一种优选的实施方案，所述重组重链的氨基酸序列如SEQ ID NO.13所示。
6.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，所述重组重链的氨基酸序列如SEQ ID NO.14所示。
7.根据权利要求1所述的EGFR-CD3双功能抗体，其特征在于，所述重组重链的氨基酸序列如SEQ ID NO.15所示。
8.一种多核苷酸序列，其特征在于，所述多核苷酸序列编码如权利要求1至7任一项所述的EGFR-CD3双功能抗体。
9.一种药物组合物，其特征在于，所述药物组合物包含如权利要求1至7任一项所述的EGFR-CD3双功能抗体。
10.根据权利要求1或7任一项所述的EGFR-CD3双功能抗体在制备抗肿瘤药物中的应用。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115029318A (zh) * 2022-05-12 2022-09-09 广州明征生物科技有限公司 携带多特异性抗体基因的间充质干细胞及其制药用途

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Publication number Priority date Publication date Assignee Title
US20170247476A1 (en) * 2014-09-25 2017-08-31 Amgen Inc. Protease-activatable bispecific proteins
US20180222989A1 (en) * 2015-08-04 2018-08-09 Glaxosmithkline Intellectual Property Development Limited Combination treatments and uses and methods thereof
CN109423495A (zh) * 2017-08-24 2019-03-05 上海恒润达生生物科技有限公司 一种双靶向嵌合抗原受体及其用途
CN109912716A (zh) * 2017-12-13 2019-06-21 凯惠科技发展(上海)有限公司 一种egfr抗体及其制备方法和应用

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US20170247476A1 (en) * 2014-09-25 2017-08-31 Amgen Inc. Protease-activatable bispecific proteins
US20180222989A1 (en) * 2015-08-04 2018-08-09 Glaxosmithkline Intellectual Property Development Limited Combination treatments and uses and methods thereof
CN109423495A (zh) * 2017-08-24 2019-03-05 上海恒润达生生物科技有限公司 一种双靶向嵌合抗原受体及其用途
CN109912716A (zh) * 2017-12-13 2019-06-21 凯惠科技发展(上海)有限公司 一种egfr抗体及其制备方法和应用

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115029318A (zh) * 2022-05-12 2022-09-09 广州明征生物科技有限公司 携带多特异性抗体基因的间充质干细胞及其制药用途

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