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CN111848798A - 可结合bcma的纳米抗体及其应用 - Google Patents

可结合bcma的纳米抗体及其应用 Download PDF

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CN111848798A
CN111848798A CN202010732288.7A CN202010732288A CN111848798A CN 111848798 A CN111848798 A CN 111848798A CN 202010732288 A CN202010732288 A CN 202010732288A CN 111848798 A CN111848798 A CN 111848798A
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吴喜林
吴稚伟
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Y Clone Medical Science Co ltd
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Nanjing Anrui Biotechnology Co ltd
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Abstract

本发明涉及一种可结合human BCMA的多肽，包括3个互补决定区CDR1‑3，CDR1序列为或包括SEQ ID NO:1‑104所示序列之一，CDR2序列为或包括SEQ ID NO:105‑207所示序列之一，CDR3序列为或包括SEQ ID NO:208‑311所示序列之一。BCMA是B细胞成熟抗原，本发明针对B细胞淋巴瘤进行纳米抗体药物开发和诊断试剂开发，通过制备BCMA蛋白、免疫羊驼、利用噬菌体库展示纳米单抗的平台技术等，筛选到特异性结合BCMA的纳米抗体VHH，鉴定了其CDR序列，并构建了人源化抗体BANB；同时利用流式细胞术检测方法鉴定人源化抗体与细胞表面BCMA蛋白的结合。本发明还为B细胞淋巴瘤的临床治疗提供潜在的纳米抗体新药,以及相应的基因治疗方法。

Description

可结合BCMA的纳米抗体及其应用
技术领域
本发明涉及生物医药领域。更特别地，涉及一种可结合人BCMA的多肽，所述的多肽在制备BCMA检测剂或B细胞淋巴瘤治疗药物中的应用。
背景技术
BCMA是B细胞成熟抗原，是一种跨膜糖蛋白，亦称CD269或TNFRSF17，是TNF受体超家族成员之一，它的配体是B细胞激活因子(BAFF)以及增殖诱导配体(APRIL)。BCMA的表达限于B细胞谱系，主要存在于浆细胞和浆母细胞上，研究表明，BCMA mRNA和蛋白在恶性浆细胞上的表达远高于正常浆细胞，而在初始B细胞和记忆B细胞及其他正常组织细胞上不表达。BAFFF及APRIL通过BCMA的信号传导促进恶性浆细胞的存活。此外，有研究表明，在供体淋巴细胞输注与移植物抗肿瘤反应后缓解的多发性骨髓瘤患者中检测到BCMA抗体，以上研究结果表明，BCMA是非常有前景的B细胞淋巴瘤的靶向疗法抗原。
1993年，一种来源于骆驼科的新型天然抗体被发现。该抗体天然缺失轻链而只由重链组成，其重链包含两个恒定区(CH2和CH3)、一个铰链区和一个重链可变区(Variableheavy chain domain，VHH，即抗原结合位点)，该重链可变区的相对分子质量约为13KDa，仅为常规抗体的1/10，且分子高度和直径均在纳米级别，是目前可获得的最小的功能性抗体片段，因此又被称为纳米单抗(Nanobody，Nb)。因纳米单抗稳定性高(90℃条件下仍不会降解)、亲和力高、与人源抗体同源性超过80％、毒性和免疫原性均较低等特点，最近纳米单抗被广泛用于免疫诊断试剂盒研发、影像学研发以及针对肿瘤、炎症、传染病和神经系统疾病等领域的抗体药物研发。
治疗性抗体药物发展的趋势从鼠源、人鼠嵌合、人源化再到全人源，再到近年得到广泛关注的纳米抗体，纳米抗体的研发得到高速发展。2018年9月首个纳米抗体药物获批。目前国内外有多个纳米抗体药物在临床研究阶段。我们期望通过免疫羊驼，获得高亲和力的靶向BCMA的治疗性纳米抗体。
发明内容
本发明通过用抗原免疫羊驼，获取羊驼源纳米单抗及其VHH，用于诊断和治疗B细胞淋巴瘤病人。基于这些研究，本发明提供了一种可结合BCMA的多肽，包括3个互补决定区CDR1-3，CDR1序列为或包括SEQ ID NO:1-104所示序列之一，CDR2序列为或包括SEQ ID NO:105-207所示序列之一，CDR3序列为或包括SEQ ID NO:208-311所示序列之一。
在一个具体实施方案中，所述多肽还包括4个框架区FR1-4，所述FR1-4与所述CDR1-3交错排列。例如，可将FR1-4序列设计为如SEQ ID NO:312-315所示(羊驼源)，但本发明的范围不限于此。抗体的特异性识别和结合能力主要由CDR区序列决定，FR序列影响不大，可根据物种来设计，这是本领域公知的。例如，可设计人源、鼠源或羊驼源的FR区序列来连接上述CDR，从而得到一个可结合人BCMA的多肽或结构域。例如，可将FR1-4序列设计为如SEQ ID NO:316-319所示(人源)，人源化后的多肽具有等同性质的结合活性。
在一个优选实施方案中，所述多肽为单克隆抗体。
在一个优选实施方案中，所述多肽为VHH。
在一个优选实施方案中，所述多肽为羊驼源的VHH或人源化的VHH。
在一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR1的序列为GRIEXTYV，第5位的X表示天冬酰胺或丝氨酸；
II)CDR2的序列如SEQ ID NO:153-156之一所示，并且
III)CD3的序列如SEQ ID NO:255、257-260之一所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:50，CDR2的序列为SEQ ID NO:153，并且CDR3的序列为SEQ ID NO:257；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:154，并且CDR3的序列为SEQ ID NO:258；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:155，并且CDR3的序列为SEQ ID NO:259；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:156，并且CDR3的序列为SEQ ID NO:255；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:153，并且CDR3的序列为SEQ ID NO:255；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:153，并且CDR3的序列为SEQ ID NO:260。
在另一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR1的序列为GRTFSSYX，第7位的X表示精氨酸、丝氨酸、酪氨酸或缬氨酸；
II)CDR2的序列如SEQ ID NO:167-170之一所示，并且
III)CD3的序列如SEQ ID NO:271-274之一所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:63，CDR2的序列为SEQ ID NO:167，并且CDR3的序列为SEQ ID NO:271；或
CDR1的序列为SEQ ID NO:64，CDR2的序列为SEQ ID NO:168，并且CDR3的序列为SEQ ID NO:272；或
CDR1的序列为SEQ ID NO:65，CDR2的序列为SEQ ID NO:169，并且CDR3的序列为SEQ ID NO:273；或
CDR1的序列为SEQ ID NO:66，CDR2的序列为SEQ ID NO:170，并且CDR3的序列为SEQ ID NO:274。
在另一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR1的序列如28或31所示；
II)CDR2的序列如SEQ ID NO:132所示，并且
III)CD3的序列如SEQ ID NO:234或235所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:28，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:234；或
CDR1的序列为SEQ ID NO:28，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:235；或
CDR1的序列为SEQ ID NO:31，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:234。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:49，CDR2的序列为SEQ ID NO:152，并且CDR3的序列为SEQ ID NO:256；或
CDR1的序列为SEQ ID NO:52，CDR2的序列为SEQ ID NO:152，并且CDR3的序列为SEQ ID NO:256。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:62，CDR2的序列为SEQ ID NO:155，并且CDR3的序列为SEQ ID NO:270；或
CDR1的序列为SEQ ID NO:74，CDR2的序列为SEQ ID NO:177，并且CDR3的序列为SEQ ID NO:270。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:68，CDR2的序列为SEQ ID NO:172，并且CDR3的序列为SEQ ID NO:220；或
CDR1的序列为SEQ ID NO:13，CDR2的序列为SEQ ID NO:117，并且CDR3的序列为SEQ ID NO:220。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:39，CDR2的序列为SEQ ID NO:142，并且CDR3的序列为SEQ ID NO:245；或
CDR1的序列为SEQ ID NO:39，CDR2的序列为SEQ ID NO:142，并且CDR3的序列为SEQ ID NO:246。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:57，CDR2的序列为SEQ ID NO:161，并且CDR3的序列为SEQ ID NO:265；或
CDR1的序列为SEQ ID NO:57，CDR2的序列为SEQ ID NO:162，并且CDR3的序列为SEQ ID NO:265。
在另一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR3的序列为HLYGAGGXLDA，其中，第8为的X表示异亮氨酸或精氨酸；
II)CDR1的序列如SEQ ID NO:98或100所示，并且
III)CD2的序列如SEQ ID NO:201、203或204所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:98，CDR2的序列为SEQ ID NO:201，并且CDR3的序列为SEQ ID NO:304；或
CDR1的序列为SEQ ID NO:100，CDR2的序列为SEQ ID NO:203，并且CDR3的序列为SEQ ID NO:307；或
CDR1的序列为SEQ ID NO:100，CDR2的序列为SEQ ID NO:204，并且CDR3的序列为SEQ ID NO:307。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:99，CDR2的序列为SEQ ID NO:202，并且CDR3的序列为SEQ ID NO:305；或
CDR1的序列为SEQ ID NO:99，CDR2的序列为SEQ ID NO:202，并且CDR3的序列为SEQ ID NO:306。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:10，CDR2的序列为SEQ ID NO:114，并且CDR3的序列为SEQ ID NO:217；或
CDR1的序列为SEQ ID NO:27，CDR2的序列为SEQ ID NO:131，并且CDR3的序列为SEQ ID NO:217。
在另一个具体实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:71，CDR2的序列为SEQ ID NO:137，并且CDR3的序列为SEQ ID NO:278；或
CDR1的序列为SEQ ID NO:34，CDR2的序列为SEQ ID NO:137，并且CDR3的序列为SEQ ID NO:240。
在另一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR2的序列为ITSDGXT，其中，第6位的X表示丝氨酸、缬氨酸或苏氨酸；
II)CDR1的序列如SEQ ID NO:36、70或94所示，并且
III)CD3的序列如SEQ ID NO:242、277或300所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:36，CDR2的序列为SEQ ID NO:139，并且CDR3的序列为SEQ ID NO:242；或
CDR1的序列为SEQ ID NO:70，CDR2的序列为SEQ ID NO:174，并且CDR3的序列为SEQ ID NO:277；或
CDR1的序列为SEQ ID NO:94，CDR2的序列为SEQ ID NO:197，并且CDR3的序列为SEQ ID NO:300。
在另一个具体实施方案中，所述多肽的CDR序列如下：
I)CDR2的序列为ITSGGXT，其中，第6位的X表示丝氨酸、天冬氨酸或苏氨酸；
II)CDR1的序列如SEQ ID NO:83、87或104所示，并且
III)CD3的序列如SEQ ID NO:289、293或311所示。
优选地，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:83，CDR2的序列为SEQ ID NO:186，并且CDR3的序列为SEQ ID NO:289；或
CDR1的序列为SEQ ID NO:87，CDR2的序列为SEQ ID NO:190，并且CDR3的序列为SEQ ID NO:293；或
CDR1的序列为SEQ ID NO:104，CDR2的序列为SEQ ID NO:207，并且CDR3的序列为SEQ ID NO:311。
本发明还提供了上述多肽在制备B细胞淋巴瘤治疗药物中的应用。
本发明还提供了上述多肽的核酸编码序列。
在一个实施方案中，所述核酸编码序列为DNA编码序列或RNA编码序列。
在一个具体实施方案中，所述核酸编码序列存在于基因表达框中。
本发明还提供了包含上述核酸编码序列的表达框的表达载体。
在一个优选实施方案中，所述表达载体为病毒载体。
在一个优选实施方案中，所述表达载体为腺相关病毒表达载体(AAV载体)。
本发明还提供了上述核酸编码序列和表达载体在B细胞淋巴瘤治疗药物中的应用。
本发明还提供了上述多肽在制备BCMA检测剂中的应用。可用于检测细胞表面BCMA的特异性表达情况。
在一个具体实施方案中，所述检测剂为细胞流式检测剂。
在一个优选实施方案中，所述多肽的CDR序列如下：
CDR1的序列为SEQ ID NO:17，CDR2的序列为SEQ ID NO:121，并且CDR3的序列为SEQ ID NO:224；或
CDR1的序列为SEQ ID NO:28，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:234；或
CDR1的序列为SEQ ID NO:28，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:235；或
CDR1的序列为SEQ ID NO:31，CDR2的序列为SEQ ID NO:132，并且CDR3的序列为SEQ ID NO:234；或
CDR1的序列为SEQ ID NO:33，CDR2的序列为SEQ ID NO:136，并且CDR3的序列为SEQ ID NO:239；或
CDR1的序列为SEQ ID NO:42，CDR2的序列为SEQ ID NO:145，并且CDR3的序列为SEQ ID NO:249；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:153，并且CDR3的序列为SEQ ID NO:255；或
CDR1的序列为SEQ ID NO:51，CDR2的序列为SEQ ID NO:153，并且CDR3的序列为SEQ ID NO:260；或
CDR1的序列为SEQ ID NO:53，CDR2的序列为SEQ ID NO:157，并且CDR3的序列为SEQ ID NO:261；或
CDR1的序列为SEQ ID NO:57，CDR2的序列为SEQ ID NO:162，并且CDR3的序列为SEQ ID NO:265；或
CDR1的序列为SEQ ID NO:77，CDR2的序列为SEQ ID NO:180，并且CDR3的序列为SEQ ID NO:283；或
CDR1的序列为SEQ ID NO:78，CDR2的序列为SEQ ID NO:181，并且CDR3的序列为SEQ ID NO:284；或
CDR1的序列为SEQ ID NO:96，CDR2的序列为SEQ ID NO:199，并且CDR3的序列为SEQ ID NO:302。
本发明针对B细胞淋巴瘤治疗药物进行纳米抗体药物开发和诊断试剂研发，通过制备sBCMA蛋白、免疫羊驼、利用噬菌体库展示纳米单抗的平台技术等，筛选到特异性结合sBCMA的纳米抗体VHH，鉴定了其CDR序列，并构建了人源化的VHH-huFc1(BANB)；同时利用流式细胞术评估人源化抗体与细胞表面BCMA蛋白的结合。本发明为B细胞淋巴瘤的临床治疗提供潜在的纳米抗体新药,同时为B细胞淋巴瘤患者的诊断提供相应的检测试剂。
附图说明
图1为sBCMA第3和4次免疫羊驼一周后的抗血清效价检测曲线；
图2为以sBCMA-VHH噬菌体抗体文库为模板扩增的PCR产物的电泳图；
图3为sBCMA-VHH噬菌体抗体文库的淘选鉴定，其中，A为噬菌体文库针对BCMA蛋白淘选后ELISA检测统计图；B为从第二轮(2nd)、第三轮(3rd)和第四轮(4th)淘选后的噬菌体抗体文库分别挑选48、46和47个克隆进行噬菌体ELISA检测统计图；
图4为原核表达的VHH抗体的ELISA检测统计图，每个点代表一个克隆，纵坐标为针对BCMA的OD450/空白对照的OD450，比值大于2.5定义为阳性；
图5和6为VHH-hfc1的真核表达上清与细胞表面表达的膜蛋白BCMA结合的流式结果图。横坐标为anti BCMA VHH-hfc1结合于膜蛋白BCMA的荧光信号值，纵坐标为商品BCMA抗体结合于膜蛋白BCMA的荧光信号值。
具体实施方式
1.免疫原的制备
我们依据NCBI网站上人BCMA蛋白序列和基因序列信息，分析并设计了可有效诱导羊驼产生针对BCMA蛋白的特异性抗体的多肽sBCMA，在C端连接His-tag(sBCMA-his)或兔Fc(sBCMA-rFc)用于后续纯化及检测。
2.羊驼免疫与抗血清的获得
用250μg sBCMA-rFc蛋白与250μl弗氏完全佐剂的乳化混合物对羊驼进行初免，在第14天、28天、42天用sBCMA-rFc蛋白与250μl弗氏不完全佐剂加强免疫3次，第3次和第4次免疫1周后，采血检测抗血清滴度；第4次免疫1周后，采血100ml用于噬菌体抗体库的构建。
抗血清效价通过ELISA检测，用浓度为0.5μg/ml的BCMA蛋白包被检测板，每孔加入梯度稀释的抗血清或者纯化的抗体100μl(对照为免疫前羊驼血清)，37℃孵育1.5h，洗涤2次，每孔加入1：10000稀释的辣根过氧化物酶标记的Goat anti-Llamma IgG(H+L)二抗，37℃孵育1h，洗涤4-6次后，加100μl TMB底物，37℃孵育10min，50μl 0.2M的H2SO4中止反应，测定OD 450nm。ELISA检测血清效价规定为在OD450是空白对照的2.1倍以上并且大于0.2的最高稀释倍数。
结果如图1所示，3免和4免的抗血清效价分别为1.09×106和3.28×106。由此可见，该抗原可诱导羊驼产生特异性针对BCMA蛋白的高滴度抗血清。
3.VHH噬菌体库构建及淘选
收集100ml免疫后羊驼的外周血，利用淋巴细胞分离液(GE Ficoll-Paque Plus)分离获得羊驼的PBMC，根据TRIzol操作手册，提取RNA，并利用oligo(dT)反转为cDNA,通过引物扩增，以及分子克隆等技术，将羊驼的VHH基因克隆至phagemid质粒，转化TG1细菌，得到VHH噬菌体库。为了进一步鉴定sBCMA-VHH噬菌体库是否构建成功，通过PCR扩增免疫sBCMA羊驼的VHH目的基因，可以看出目的条带为500bp，大小符合预期(图2)，说明该sBCMA-VHH噬菌体抗体文库里含有VHH基因。挑选48个克隆进行测序，测序结果显示，所测序列没有完全一致的重复序列；比对结果显示，差异序列大多在CDR结合区。经检测，该构建了一个sBCMA-VHH噬菌体抗体文库的库容为1.6×109，阳性率为100％，序列多样性(Diversity)为100％，有效插入率(In frame rate)为85.4％。
在M13KO7辅助噬菌体的帮助下，用VHH-phagemid转化的细菌，进行噬菌体抗体库的复苏，并用PEG/NaCl进行沉淀。将包被有50μg/ml的sBCMA-His蛋白进行三次富集噬菌体抗体库。将富集的噬菌体，洗脱、转化、涂板、挑取单克隆进行噬菌体与sBCMA蛋白ELISA的结合鉴定，将结合读值>1.0的克隆进行测序，并克隆至表达载体pVAX1，转染293F细胞表达生产纳米单抗。
淘选后的文库与BCMA蛋白进行结合检测。噬菌体ELISA结果显示，没有富集前的sBCMA-VHH噬菌体文库与sBCMA蛋白的结合读值为0.50，经过一轮、二轮、三轮富集后的噬菌体文库读值分别为1.17、2.78、3.33(图3A)。为了进一步验证富集后的文库中结合sBCMA-VHH蛋白的阳性噬菌体率，从第2轮、第3轮和第4轮富集后的文库里分别挑选48，46，47个克隆进行单个噬菌体ELISA检测。结果显示，第2轮文库里，6.25％的单个噬菌体克隆为阳性，第3轮文库里13.04％的噬菌体克隆为阳性，第4轮文库里34.04％的噬菌体克隆为阳性，而且结合的平均读值在3.0左右(图3B)，通过sBCMA蛋白淘选成功地富集了高结合力的sBCMA-VHH噬菌体文库。
4.VHH原核表达文库的构建及VHH表达
对上述四轮淘选富集后的4th-sBCMA-VHH噬菌体抗体文库进行PCR扩增；获取并纯化抗体库中所有VHH的基因片段，将VHH的基因片段克隆至原核表达载体，转化SS320菌株，构建VHH的原核表达抗体库；将原核表达抗体库涂布平板，过夜培养，次日随机挑选单克隆菌落2016个，使用IPTG诱导表达抗体上清，对抗体上清与sBCMA蛋白进行ELISA结合检测。
结果显示，有细菌上清与sBCMA蛋白结合，同时不与空白对照结合，sBCMA结合的读值/空白对照的读值大于2.5(图4和表1)。将这些序列进行测序比对，剔除重复序列，最终获得114个的VHH抗体序列。进一步的实验证实，这114个VHH抗体以及从VHH抗体得到的CDR均可特异性地结合BCMA蛋白。
表1 114个VHH抗体与sBCMA蛋白的结合值及其CDR序列
Figure BDA0002603752200000111
Figure BDA0002603752200000121
Figure BDA0002603752200000131
5.流式细胞法检测VHH抗体与肿瘤细胞的结合
将上述抗体序列构建为人源化VHH-hfc1抗体表达载体，所得载体转染293TT后，收取抗体上清进行如下流式实验。将293TT细胞转染表达膜蛋白BCMA，将上述BANB抗体与表达BCMA的293TT细胞293TT-BCMA混合孵育，100μl/样品，4℃1h；以0.5％PBSF洗涤两遍后，加入荧光二抗：AlexaFluor 488anti human FC,Alexa Fluor 647anti human BCMA，4℃30min；以0.5％PBSF洗涤两遍后，上机检测。同时做阴性细胞对照组，即未转染的293TT细胞组。流式结果如图5和6所示，有14个VHH抗体均可与293TT-BCMA细胞结合而不与293TT细胞结合，包括，BANB0017、BANB0047、BANB1081、BANB0108、BANB0117、BANB0162、BANB1912、BANB1981、BANB0199、BANB0205、BANB0312、BANB0309、BANB0333、BANB0385。以上抗体在与表达BCMA膜蛋白的RPMI-8226细胞的流式检测实验，同样可见阳性结合。由此可见，上述VHH抗体具有靶向结合膜蛋白BCMA的能力，同时有可能通过靶向肿瘤细胞表面的BCMA分子，促进巨噬细胞的吞噬作用，从而达到治疗或者抑制肿瘤生长的效果，因此这14个VHH抗体均有潜力成为治疗肿瘤的新型抗体药物。
因为上述14个VHH能够识别细胞表面的BCMA分子，因此这些VHH抗体序列也可以应用于CAR(Chimeric Antigen Receptor,嵌合抗原x受体,由VHH序列融合第三代或者第四代CD28-4-1BB-CD3zeta分子序列构成)细胞治疗肿瘤的治疗。另外因为上述VHH能识别肿瘤细胞表面的BCMA分子，因此VHH可以通过偶联药物用于ADC(Antibody-drug conjugate,抗体偶联药物)治疗或者偶联同位素用于依赖抗体的分子影像诊断等，为肿瘤的临床治疗提供潜在的纳米新药。
6.使用AAV病毒载体装载的人源化VHH进行体内实验
腺相关病毒载体(AAV)源于非致病的野生型腺相关病毒，由于其安全性好、宿主细胞范围广(分裂和非分裂细胞)、免疫源性低，在体内表达外源基因时间长等特点，被视为最有前途的基因转移载体之一，在世界范围内的基因治疗和疫苗研究中得到广泛应用。
AAV Helper-Free病毒包装系统购于Cell Biolabs,San Diego USA。将上述VHH的DNA编码序列通过分子克隆技术插入到pAAV-MCS质粒；通过测序证明构建成功后，将构建好的质粒pAAV-Ab与pHelper和pAAV-DJ质粒按照质量比1:1:1的方式使用PEI转染试剂共转染AAV-293T细胞。转染后分别于48、72、96和120小时收集上清，并用5xPEG8000(sigma)进行浓缩，最后用1.37g/ml氯化铯进行纯化。纯化的AAV溶解于PBS里，进行鉴定和分装后保存于-80℃。
ImmunodeficientNOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ(NCG)小鼠购自南京大学模式动物所，与NSG小鼠类似，该小鼠在SCID小鼠基础上缺失了IL2受体基因，导致体内没有小鼠T细胞，B细胞以及极少量的NK细胞。1.0-15x 107PBMC腹腔注射进4-6周的NCG小鼠体内；三周后，采血流式检测人T细胞，通过染色人的CD45+、CD3+、CD4+和CD8+。人CD45阳性细胞的比例到达5％以上，判定为小鼠人源化成功。将上述小鼠腹腔注射RPMI-8226细胞3*106/只，一周后，使小鼠接受AAV-BANB(1x1011gc/100μl)肌肉注射，以AAV-GFP为对照组。结果显示，AAV-BANB对多发性骨髓瘤有治疗作用。
由以上实验结果可知，上述VHH以及人源化后得到的VHH-hfc1抗体均可特异性地识别、结合sBCMA，并且在流式细胞层面也可有效结合BCMA膜蛋白，也可对B细胞淋巴瘤在小鼠体内发展产生抑制作用。说明这些抗体可识别天然构象的细胞表面膜蛋白BCMA，可利用其靶向作用，偶联治疗性药物，用以制备抗体偶联药物ADC(Antibody-drug conjugate)或偶联同位素用于依赖抗体的分子影像诊断等。也可应用于CAR疗法，用于治疗多发性骨髓瘤等B细胞恶性肿瘤，比如CAR-T或CAR-NK疗法，将T细胞或自然杀伤细胞(NK细胞)经修饰后表达抗BCMA CAR，从而达到治疗目的。更进一步的，可将编码上述抗体VHH片段的核酸搭载在AAV系统中，用以制备基因治疗药物，达到治疗目的。
以上所述仅为本发明的较佳实施例，并不用以限制本发明，凡在本发明的精神和原则之内，所作的任何修改、等同替换、改进等，均应包含在本发明的保护范围之内。
序列表
<110> 南京安锐生物科技有限公司
<120> 可结合BCMA的纳米抗体及其应用
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<400> 15
Gly Phe Pro Phe Gly Asp Tyr Ala
1 5
<210> 16
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 16
Gly Phe Arg Leu Gly Asp His Ala
1 5
<210> 17
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 17
Gly Phe Ser Ile Arg Asp Tyr Gly
1 5
<210> 18
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 18
Gly Phe Thr Phe Ala Asp Ser Ala
1 5
<210> 19
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 19
Gly Phe Thr Phe Asp Asp Tyr Ala
1 5
<210> 20
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 20
Gly Phe Thr Phe Asp Gly His Ala
1 5
<210> 21
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 21
Gly Phe Thr Phe Ser Thr Tyr Trp
1 5
<210> 22
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 22
Gly Phe Thr Ser Asp Ala Tyr Gly
1 5
<210> 23
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 23
Gly Phe Tyr Phe Ser Leu Tyr Arg
1 5
<210> 24
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 24
Gly Gly Ala Trp Arg Phe Tyr Asp
1 5
<210> 25
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 25
Gly Gly Ser Ile Ser Thr Ser Tyr Phe His
1 5 10
<210> 26
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 26
Gly Gly Ser Ile Thr Thr Asn Trp Asn Tyr
1 5 10
<210> 27
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 27
Gly Gly Ser Ile Thr Thr Asn Tyr Phe Tyr
1 5 10
<210> 28
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 28
Gly Gly Thr Phe Arg Ala Tyr Val
1 5
<210> 29
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 29
Gly Gly Thr Phe Ser Arg Ser Thr
1 5
<210> 30
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 30
Gly Gly Thr Leu Ser Thr Gly Ala
1 5
<210> 31
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 31
Gly Gly Thr Ser Arg Ala Tyr Val
1 5
<210> 32
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 32
Gly Ile Ile Asp Ser Ile Tyr Thr
1 5
<210> 33
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 33
Gly Ile Ile Gly Ser Ile Lys Ser
1 5
<210> 34
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 34
Gly Ile Ile Arg Arg Met Asn Thr
1 5
<210> 35
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 35
Gly Ile Ser Val Asp Ser Ser Ala
1 5
<210> 36
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 36
Gly Lys Ile Phe Ser Leu Ser Asp
1 5
<210> 37
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 37
Gly Lys Ile Phe Ser Asn Met Ile
1 5
<210> 38
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 38
Gly Leu Ser Phe Ser Thr Tyr Thr
1 5
<210> 39
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 39
Gly Leu Thr Ser Asp Asp Tyr Gly
1 5
<210> 40
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 40
Gly Asn Phe Tyr Asn Ile Asn Ser
1 5
<210> 41
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 41
Gly Asn Ile Phe Asn Ile Asn Asp
1 5
<210> 42
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 42
Gly Asn Ile Phe Gln Val Lys Asp
1 5
<210> 43
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 43
Gly Asn Ile Phe Gln Val Tyr Asp
1 5
<210> 44
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 44
Gly Asn Ser Arg Pro Ile Asn Ile
1 5
<210> 45
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 45
Gly Pro Thr Ser Ser Tyr Arg Ala
1 5
<210> 46
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 46
Gly Gln Ile Ser Glu Thr Ser Thr
1 5
<210> 47
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 47
Gly Arg Ala Phe Ser Asn Tyr Ala
1 5
<210> 48
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 48
Gly Arg Ala Ser Ser Ile Tyr Arg
1 5
<210> 49
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 49
Gly Arg Gly Phe Asp Thr Tyr Thr
1 5
<210> 50
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 50
Gly Arg Ile Glu Asn Thr Tyr Val
1 5
<210> 51
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 51
Gly Arg Ile Glu Ser Thr Tyr Val
1 5
<210> 52
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 52
Gly Arg Ile Phe Asp Thr Tyr Thr
1 5
<210> 53
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 53
Gly Arg Ile Ile Arg Val Tyr Asp
1 5
<210> 54
<211> 6
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 54
Gly Arg Ile Leu Ala Thr
1 5
<210> 55
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 55
Gly Arg Ile Val Ser Leu Asp Asp
1 5
<210> 56
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 56
Gly Arg Ser Phe Ser Asn Tyr Lys
1 5
<210> 57
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 57
Gly Arg Thr Asp Ser Ile Ile Thr
1 5
<210> 58
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 58
Gly Arg Thr Phe Ala Ala Tyr Thr
1 5
<210> 59
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 59
Gly Arg Thr Phe Gly Asn Tyr Lys
1 5
<210> 60
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 60
Gly Arg Thr Phe Ile Thr Gly Ala
1 5
<210> 61
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 61
Gly Arg Thr Phe Ser Asp Tyr Ala
1 5
<210> 62
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 62
Gly Arg Thr Phe Ser Asn Tyr Ala
1 5
<210> 63
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 63
Gly Arg Thr Phe Ser Ser Tyr Arg
1 5
<210> 64
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 64
Gly Arg Thr Phe Ser Ser Tyr Ser
1 5
<210> 65
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 65
Gly Arg Thr Phe Ser Ser Tyr Thr
1 5
<210> 66
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 66
Gly Arg Thr Phe Ser Ser Tyr Val
1 5
<210> 67
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 67
Gly Arg Thr Phe Thr Cys Ser Ala Cys Val
1 5 10
<210> 68
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 68
Gly Arg Thr Asn Tyr Thr Tyr Thr
1 5
<210> 69
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 69
Gly Arg Thr Ser Ala Ile Tyr Thr
1 5
<210> 70
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 70
Gly Ser Phe Phe Ser Ile Asp Ser
1 5
<210> 71
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 71
Gly Ser Phe Val Ser Ile Asp Ala
1 5
<210> 72
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 72
Gly Ser Phe Val Ser Ile Asn Thr
1 5
<210> 73
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 73
Gly Ser Gly Phe Leu Ile Asn Val
1 5
<210> 74
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 74
Gly Ser Gly Phe Ser Ile Tyr Gly
1 5
<210> 75
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 75
Gly Ser Ile Phe Asn Ile Asp Thr
1 5
<210> 76
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 76
Gly Ser Ile Phe Arg Thr Asn Asp
1 5
<210> 77
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 77
Gly Ser Ile Phe Arg Val His Asp
1 5
<210> 78
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 78
Gly Ser Ile Phe Ser Ala Tyr Asp
1 5
<210> 79
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 79
Gly Ser Ile Phe Ser Ile Asn Ala
1 5
<210> 80
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 80
Gly Ser Ile Phe Ser Thr Tyr Trp
1 5
<210> 81
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 81
Gly Ser Ile Phe Ser Tyr Asn Pro
1 5
<210> 82
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 82
Gly Ser Ile Phe Tyr Ser Asp Val
1 5
<210> 83
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 83
Gly Ser Ile Leu Arg Val Asp Asp
1 5
<210> 84
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 84
Gly Ser Ile Arg Asp Ile Asn Val
1 5
<210> 85
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 85
Gly Ser Ile Arg Ser Ile Asp Thr
1 5
<210> 86
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 86
Gly Ser Ile Ser Gly Phe Asn Val
1 5
<210> 87
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 87
Gly Ser Ile Ser Ser Ile Asn Ala
1 5
<210> 88
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 88
Gly Ser Ile Val Ser Phe Asn Val
1 5
<210> 89
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 89
Gly Ser Ile Val Ser Ile Asn Ser
1 5
<210> 90
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 90
Gly Ser Ile Tyr Gly Ile Asn Thr
1 5
<210> 91
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 91
Gly Ser Thr Leu Arg Arg Thr Val
1 5
<210> 92
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 92
Gly Ser Val Ser Ile Ser Pro Arg Ile Asn
1 5 10
<210> 93
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 93
Gly Thr Ile Phe Met Ile Asp Thr
1 5
<210> 94
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 94
Gly Thr Ile Phe Ser Ile Asn Gly
1 5
<210> 95
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 95
Gly Thr Ile Phe Ser Arg Phe Pro
1 5
<210> 96
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 96
Gly Thr Asn Val Ser Leu Asn Ile
1 5
<210> 97
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 97
Gly Trp Tyr Val Ser Ser Asn Ala
1 5
<210> 98
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 98
Lys Val Pro Tyr Ser Phe Asp Arg
1 5
<210> 99
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 99
Arg Ser Ile Ser Glu Ile Asn Thr
1 5
<210> 100
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 100
Arg Val Ala Tyr Ser Phe Asp Arg
1 5
<210> 101
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 101
Ser Ser Ile Phe Met Leu Ser Thr
1 5
<210> 102
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 102
Ser Thr Asp Ile Met Gly Asp
1 5
<210> 103
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 103
Thr Gly Ser Phe Ser Ser Tyr Ala
1 5
<210> 104
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 104
Thr Thr Phe Phe Ser Ile Arg Thr
1 5
<210> 105
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 105
Ile Ser Thr Leu Asp Asp Ser Ser
1 5
<210> 106
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 106
Ile Leu Ser Asn Tyr Glu Ala
1 5
<210> 107
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 107
Ile Asn Lys Leu Pro Gly Ile Ile
1 5
<210> 108
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 108
Ile Ser Trp Asn Gly Gly Gly Thr
1 5
<210> 109
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 109
Leu Ser Trp Lys Gly Pro Thr Pro
1 5
<210> 110
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 110
Ile Thr Gly Thr Gly Thr Ala
1 5
<210> 111
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 111
Ile Thr Gly Ser Gly Arg Thr
1 5
<210> 112
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 112
Ile Thr Ser Ala Gly Ser Thr
1 5
<210> 113
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 113
Gln Tyr Ser Val Gly Gly Thr
1 5
<210> 114
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 114
Val Thr Tyr Ser Asp Ser Thr
1 5
<210> 115
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 115
Ile Ser Trp Ser Gly Gly Ser Thr
1 5
<210> 116
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 116
Ile Thr Asn Asn Asp Gly Asp Thr
1 5
<210> 117
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 117
Met Thr Arg Asn Gly Asp Gly Thr
1 5
<210> 118
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 118
Ile Ser Val Thr Asp Gly Ser Asp Gly Ser Thr
1 5 10
<210> 119
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 119
Ile Ser Gln Arg Ser Thr Thr Thr
1 5
<210> 120
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 120
Ile Ser Val His Glu Arg Ser Thr
1 5
<210> 121
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 121
Val Thr Leu Arg Gly Glu Thr
1 5
<210> 122
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 122
Ile Asn Trp Asn Gly Gly Ser Thr
1 5
<210> 123
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 123
Ile Ser Ser Ser Gly Ser Asp Thr Ser Thr
1 5 10
<210> 124
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 124
Ile Met Trp Asn Gly Ala Ser Ala
1 5
<210> 125
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 125
Ile Asn Ser Gly Gly Ser Ser Ile
1 5
<210> 126
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 126
Ile Ser Asp Val Ala Gly Asn Ser
1 5
<210> 127
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 127
Ile Arg Arg Asp Gly Gly Ala Thr
1 5
<210> 128
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 128
Ile Thr Val Asp Gly Gln Thr
1 5
<210> 129
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 129
Ile Gly Tyr Ala Gly Ser Thr
1 5
<210> 130
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 130
Ile Ala Tyr Ser Gly Ser Thr
1 5
<210> 131
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 131
Ile Thr Tyr Ser Gly Asn Thr
1 5
<210> 132
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 132
Ile Gln Trp Ser Asp Gly Thr Thr
1 5
<210> 133
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 133
Glu Ser Trp Ser Gly Arg Thr
1 5
<210> 134
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 134
Ile Ser Ala Ser Gly Gly Asn Thr
1 5
<210> 135
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 135
Val Thr Ser Ser Ser Asp Lys
1 5
<210> 136
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 136
Ile Ser Asp Gly Asp Ile Thr
1 5
<210> 137
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 137
Ile Thr Ser Gly Gly Val Thr
1 5
<210> 138
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 138
Ile Ser Gly Ser Gly Gly Thr Thr
1 5
<210> 139
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 139
Ile Thr Ser Asp Gly Val Thr
1 5
<210> 140
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 140
Ile Thr Ser Asp Thr Val Thr
1 5
<210> 141
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 141
Ile Leu Trp Ala Gly Gly Ser Thr
1 5
<210> 142
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 142
Phe Ser Thr Gly Asp Ala Ser Thr
1 5
<210> 143
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 143
Ile Thr Arg Gly Gly Glu Thr
1 5
<210> 144
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 144
Ile Thr Arg Gly Gly Arg Thr
1 5
<210> 145
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 145
Ile Gly Phe Arg Gly Glu Pro
1 5
<210> 146
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 146
Ile Gly Phe Arg Gly Asp Thr
1 5
<210> 147
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 147
Val Ser Lys Asn Gly Glu Thr
1 5
<210> 148
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 148
Val Ser Trp Asn Gly Glu Gly Thr
1 5
<210> 149
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 149
Ile Ala Asn Gly Gly Ser Thr
1 5
<210> 150
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 150
Ile Asp Trp Thr Ser Gly Val Ser
1 5
<210> 151
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 151
Ile Gln Tyr Arg Gly Phe Thr Thr
1 5
<210> 152
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 152
Ile Leu Trp Ser Gly Gly Ser Thr
1 5
<210> 153
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 153
Ile Met Trp Ser Asp Ser Ile Thr
1 5
<210> 154
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 154
Leu Ser Ile Ser Gly Arg Ser
1 5
<210> 155
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 155
Ile Ser Trp Asp Val Ser Asp Thr
1 5
<210> 156
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 156
Ile Met Trp Ser Asp Ser Ile Ala
1 5
<210> 157
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 157
Ile Ser Arg His Asn Asp Thr
1 5
<210> 158
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 158
Ile Ser Trp Ser Gly Gly Asn Thr
1 5
<210> 159
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 159
Ile Arg Ser Gly Gly Leu Val
1 5
<210> 160
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 160
Ile Ser Arg Ser Gly Asp Arg Thr
1 5
<210> 161
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 161
Ile Ala Trp Ser Thr Asn Lys Thr
1 5
<210> 162
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 162
Ile Val Trp Ser Thr Asn Lys Thr
1 5
<210> 163
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 163
Ile Leu Trp Thr Arg Ser Thr
1 5
<210> 164
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 164
Ile Arg Trp Ser Gly Gly Gly Arg
1 5
<210> 165
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 165
Ile Asn Trp Asn Gly Gly Ser Asn
1 5
<210> 166
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 166
Ile Thr Trp Asn Gly Gly Thr Thr
1 5
<210> 167
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 167
Ile Ile Trp Asn Gly Gly Asn Thr
1 5
<210> 168
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 168
Ile Thr Trp Ser Ala Gly Asn Thr
1 5
<210> 169
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 169
Ile Asn Trp Ser Gly Gly Ser Thr
1 5
<210> 170
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 170
Ile Ser Arg Phe Gly Gly Thr Thr
1 5
<210> 171
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 171
Ile Ile Trp Ser Asp Gly Ser Thr
1 5
<210> 172
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 172
Leu Lys Trp Asp Asn Ser Pro
1 5
<210> 173
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 173
Leu Val Trp Pro Gly Thr Ser Thr
1 5
<210> 174
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 174
Ile Thr Ser Asp Gly Thr Thr
1 5
<210> 175
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 175
Ile Arg Thr Gly Gly Ser Pro
1 5
<210> 176
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 176
Ile Asn Tyr Ser Gly Arg Thr
1 5
<210> 177
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 177
Ile Thr Arg Asp Gly Ile Thr
1 5
<210> 178
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 178
Ile Ser Ser Gly Gly Ser Thr
1 5
<210> 179
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 179
Ile Gly Lys Gly Gly Thr Thr
1 5
<210> 180
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 180
Ile Arg Arg Asn Gly Glu Thr
1 5
<210> 181
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 181
Ile Asn Asn Ser Asp Gly Gly Thr
1 5
<210> 182
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 182
Ile Thr Thr Ala Gly Gly Thr
1 5
<210> 183
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 183
Ile Ser Thr Gly Asp Leu Thr Thr
1 5
<210> 184
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 184
Ile Ser Asn Ser Gly Met Thr
1 5
<210> 185
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 185
Ile Gly Arg Asp Gly Arg Thr
1 5
<210> 186
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 186
Ile Thr Ser Gly Gly Ser Thr
1 5
<210> 187
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 187
Ile Thr Glu Gly Gly Ser Thr
1 5
<210> 188
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 188
Ile Ser Arg Gly Gly Ser Thr
1 5
<210> 189
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 189
Phe Thr Ser Ala Gly Arg Thr
1 5
<210> 190
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 190
Ile Thr Ser Gly Gly Thr Thr
1 5
<210> 191
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 191
Ile Thr Ser Thr Gly Ser Thr
1 5
<210> 192
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 192
Val Thr Ser Gly Gly Ser Thr
1 5
<210> 193
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 193
Ile Thr Arg Arg Ser Ser Thr
1 5
<210> 194
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 194
Ile Ser Arg Gly Gly Gly Tyr Thr
1 5
<210> 195
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 195
Ile Asp Gly Gly Gly Thr Pro
1 5
<210> 196
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 196
Ile Thr Arg Ser His Lys Ala
1 5
<210> 197
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 197
Ile Thr Ser Asp Gly Ser Thr
1 5
<210> 198
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 198
Val Ser Arg His Gly Asp Thr
1 5
<210> 199
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 199
Ile Ser Ser Tyr Gly Val Arg
1 5
<210> 200
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 200
Ile Ala Lys Ser Gly Gly Ser Thr
1 5
<210> 201
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 201
Ile Ile Asn Gly Gly Ser Ile
1 5
<210> 202
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 202
Leu Thr Ser Asn Asp Asp Ala
1 5
<210> 203
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 203
Ile Ser Asn Gly Gly Thr Ile
1 5
<210> 204
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 204
Ile Ser Asn Gly Gly Thr Val
1 5
<210> 205
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 205
Ile Ser Arg Gly Arg Ser Ala
1 5
<210> 206
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 206
Ile Ser Trp Asp Ala Lys Asn Lys
1 5
<210> 207
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 207
Ile Thr Ser Gly Gly Asp Thr
1 5
<210> 208
<211> 21
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 208
Ala Ile Gly Thr Arg Tyr Thr Gly Ser Tyr Tyr Val Gly Asp Cys Pro
1 5 10 15
Asn Glu Phe Asp Tyr
20
<210> 209
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 209
Asn Leu Glu Trp Ser Val Asn Pro Gly Val Gly Val Leu Arg Asp Phe
1 5 10 15
<210> 210
<211> 22
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 210
Ala Val Arg Ser Phe Tyr Ser Gly Ala Asn Phe Pro Glu Trp Asn Ser
1 5 10 15
Asp Tyr Glu Tyr Asp Tyr
20
<210> 211
<211> 25
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 211
Ala Ile Asp Met Leu Pro Arg Arg Ile Met Ala Thr Pro Glu Leu Pro
1 5 10 15
Pro Asn Ser Ser Gly Ala Phe Glu Tyr
20 25
<210> 212
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 212
Ala Gly Gly Pro Tyr Tyr Thr Glu Ile Glu Arg Asn Tyr Ile His
1 5 10 15
<210> 213
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 213
Thr Ala Asp Leu Val Tyr Met Asn Asp Phe Gly Asp Arg Tyr Arg Pro
1 5 10 15
Leu Asp Leu
<210> 214
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 214
Asn Ile Arg Leu Asp Gly Ala Phe Trp Asn Thr Asn Asn Tyr
1 5 10
<210> 215
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 215
Asn Leu Glu Thr His Asp Leu Leu Ala Gly Ser Leu Asp Arg Tyr Arg
1 5 10 15
Val Val
<210> 216
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 216
Thr Arg Gly Asp Trp Leu Ile Asn Thr Ile Gly Arg Leu Asp Asp Phe
1 5 10 15
Asp Tyr
<210> 217
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 217
Thr Arg Gly Asp Trp Leu Ile Asn Lys Val Asn Arg Leu Asp Asp Phe
1 5 10 15
Asp Ser
<210> 218
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 218
Ala Ser Thr Phe Ser Val Ser Trp Arg Ser Gly Val Arg Tyr Asp Ser
1 5 10 15
<210> 219
<211> 25
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 219
Ala Ala Ala Pro Leu Thr Asp Trp Gly Leu Ser Cys Ser Asp His Thr
1 5 10 15
Leu His Phe Asp Tyr Glu Tyr Leu Tyr
20 25
<210> 220
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 220
Ala Ala Arg Arg Ala Phe Lys Ile Thr Ser Val Arg Ser Thr Asp Tyr
1 5 10 15
Asp Tyr
<210> 221
<211> 22
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 221
Ala Thr Asp Ser Gly Thr Pro Pro Tyr Asp Ser Ser Trp Cys Ser Arg
1 5 10 15
Asn Tyr Gly Met Asp Tyr
20
<210> 222
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 222
Ala Ala Asp Gln Tyr Phe Gly Cys Ser Asp Tyr Gly Phe Gly Ser Tyr
1 5 10 15
Asp Tyr
<210> 223
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 223
Ala Met Glu Arg Tyr Phe Tyr Cys Ser Lys Trp Asp Gly Pro Arg Asp
1 5 10 15
Gly
<210> 224
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 224
Asn Arg Asn Pro Ser Ser Phe Asn Tyr
1 5
<210> 225
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 225
Ala Lys Asp Arg Ser Arg Leu Tyr Ser Gly Ser Tyr Tyr His Tyr Glu
1 5 10 15
Ala Asp Tyr
<210> 226
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 226
Ala Ala Glu Leu Leu Asp Tyr Cys Ile Asn Tyr Glu Asp His Asp Tyr
1 5 10 15
Met Asp Tyr
<210> 227
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 227
Ala Lys Ala Gly Pro Tyr Arg Asp Tyr Thr Glu Glu Tyr Asp Gly
1 5 10 15
<210> 228
<211> 21
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 228
Ala Arg Asp Ile Val Ala Pro Pro Gly Arg Leu Val Leu Ile Thr Thr
1 5 10 15
Gln Leu Arg Gln Arg
20
<210> 229
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 229
Gly Ala Arg Ser Leu Tyr Tyr Cys Ser Asp Trp Leu His Gln Asn Gln
1 5 10 15
Met His His
<210> 230
<211> 20
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 230
Ala Lys Pro Lys Met Asp Thr Ala Tyr Pro Asp Ala Phe Ser Ser Asp
1 5 10 15
Asp Tyr Asp Ser
20
<210> 231
<211> 5
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 231
Gly Ala Arg Gly Tyr
1 5
<210> 232
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 232
Ala Arg Gly Leu Thr Val Gln Gly Gly Gly Pro Ala His Asp Tyr
1 5 10 15
<210> 233
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 233
Ala Gln Asn Gly Ala Met Pro Tyr Asp Phe Gly Ser
1 5 10
<210> 234
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 234
Gly Ala Ala Arg Ser Arg Phe Ser Glu Tyr Glu Tyr
1 5 10
<210> 235
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 235
Val Val Arg Pro Val Val Gly Phe Arg Ser Met Asn Ile
1 5 10
<210> 236
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 236
Val Ala Asp Arg Glu Ser Leu Arg Val Ala Pro Arg Pro Ala His Asn
1 5 10 15
Tyr Glu Phe
<210> 237
<211> 20
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 237
Ala Ala Ala Lys Trp Gly Gly Thr Arg Trp Ser Ile Arg Asp Ser Ser
1 5 10 15
Arg Val Asp Tyr
20
<210> 238
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 238
Asn Val Arg Gly Ser Phe Gly Gln Tyr
1 5
<210> 239
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 239
Arg Val Glu Arg Gly Ile Arg Pro Arg Val Ile Ser
1 5 10
<210> 240
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 240
Thr Tyr Ala Pro Leu Tyr Gln Pro Ser Ser Ala Ser
1 5 10
<210> 241
<211> 22
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 241
Ala Val Gly Phe Phe Ser Ala Arg Ala Gly Cys Gly Leu Ile Asn Arg
1 5 10 15
Pro Asp Leu Phe Arg Ser
20
<210> 242
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 242
Asn Val Gln Gly Arg Asn Trp Trp Arg Ala Ala Phe Asp Ser
1 5 10
<210> 243
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 243
Asn Ala Arg Tyr Thr Phe Tyr Asn Phe
1 5
<210> 244
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 244
Ala Ala Asn Pro Tyr Gly Gly Val Thr Glu Lys Glu Asp Gly Tyr Glu
1 5 10 15
Tyr
<210> 245
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 245
Ala Leu Arg Gly Phe Tyr His Cys Ser Ile Tyr Gly Pro Gly Asp Met
1 5 10 15
Asp Tyr
<210> 246
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 246
Ala Leu Arg His Tyr Tyr His Cys Ser Ile Trp Gly Pro Gly Asp Met
1 5 10 15
Asp Tyr
<210> 247
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 247
His Ala Ile Trp Val Asp Pro Lys Gly Tyr Asn Ile Asp Tyr
1 5 10
<210> 248
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 248
Lys Met Gln Ile Val Arg Ile Gln Arg Asp Ser Phe Phe Ser Thr Asp
1 5 10 15
Tyr
<210> 249
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 249
Tyr Ala Asp Val Gln Thr Phe Arg Ser Asp Ile Leu Asp Glu Tyr Thr
1 5 10 15
Tyr
<210> 250
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 250
Ser Ala Asp Val His Thr Phe Arg Ala Tyr Ile Leu Asp Glu Tyr Thr
1 5 10 15
Tyr
<210> 251
<211> 7
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 251
Asn Val Lys Pro Glu Asn Tyr
1 5
<210> 252
<211> 20
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 252
Ala Ala Gly Ala Leu Tyr Ala Arg Ala Glu Asn Leu Arg Gln Lys Ala
1 5 10 15
Asp Tyr Asn Asn
20
<210> 253
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 253
Tyr Gly Asn Phe Arg Arg Leu Gly Trp Thr Lys Phe Leu Asp Tyr
1 5 10 15
<210> 254
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 254
Ala Ala Arg Asp Gly Tyr Arg Pro Lys Asn Glu Tyr Asp Tyr
1 5 10
<210> 255
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 255
Ala Ala Ala Arg Ser Arg Tyr Ser Glu Tyr Val Tyr
1 5 10
<210> 256
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 256
Ala Ala Asp Gln Trp Gly Ser Val Val Ala Thr Ala Pro Glu Tyr Asp
1 5 10 15
Tyr
<210> 257
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 257
Ala Ala Ala Arg Ile Arg Tyr Ser Glu Tyr Gly Tyr
1 5 10
<210> 258
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 258
Tyr Ala Arg Phe Leu Thr Asp Glu Phe
1 5
<210> 259
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 259
Asn Leu His Ser Thr Trp Pro Ser Tyr Asn Leu
1 5 10
<210> 260
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 260
Ala Ala Ala Arg Ser Arg Tyr Phe Glu Tyr Val Tyr
1 5 10
<210> 261
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 261
Thr Ala Glu Ala Gln Thr Pro Gly Arg Phe Ala Leu Val Asn Ala
1 5 10 15
<210> 262
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 262
Ala Ala Asp Glu Ser Ala Tyr Gly Gly Gly Arg Pro Asn Glu Tyr Asp
1 5 10 15
Tyr
<210> 263
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 263
Asn Ala His Asn Ala Val Asp Phe Arg Asp Tyr
1 5 10
<210> 264
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 264
Asn Ala Val Ala Lys Arg Thr Ile Val Thr Thr Pro Glu Pro Tyr
1 5 10 15
<210> 265
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 265
Ala Ser Ala Ala Phe Asp Ala Gly Ser Trp Pro Gly Gly Tyr Asn Tyr
1 5 10 15
<210> 266
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 266
Asn Ala Asn Thr Ser Pro Tyr Gly Arg Ala Asn Tyr
1 5 10
<210> 267
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 267
Ala Val Asp Val Asp Ser Leu Gly Phe Pro Thr Thr Ala Gln Tyr Tyr
1 5 10 15
Glu Tyr
<210> 268
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 268
Ala Ala Asp Arg Ser Arg Leu Leu Ser Ile Arg Arg Gly Ile Ser Asp
1 5 10 15
Tyr Asp Tyr
<210> 269
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 269
Ala Gln Ser Lys Gly Gly Trp Tyr Thr Val Ser Thr Ser Thr Tyr Thr
1 5 10 15
Phe
<210> 270
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 270
Ala Ala Lys Lys Phe Val Leu Thr Ala Thr Pro His Gln Tyr Asp Tyr
1 5 10 15
<210> 271
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 271
Ala Ala Ala Leu Lys Ile Pro Phe Thr Asn Leu Phe Arg Ser Tyr Asp
1 5 10 15
Tyr
<210> 272
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 272
Ala Thr Asp Pro Leu Asp Gln Leu Arg Val Gly Leu Asp Phe Arg Ser
1 5 10 15
<210> 273
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 273
Ala Ala Asp His Ser Ser Asp Tyr Thr Arg Gly Val Tyr Tyr Gly Met
1 5 10 15
Asp Tyr
<210> 274
<211> 21
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 274
Ala Ala Ser Thr Glu Val Thr Glu Val Phe Ser Gly Leu Leu His Pro
1 5 10 15
Glu Ser Tyr Lys Tyr
20
<210> 275
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 275
Ala Tyr Ala Pro Ser Arg Tyr Glu Glu Tyr His Tyr
1 5 10
<210> 276
<211> 21
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 276
Ala Gly Glu Thr Gln Tyr Arg Lys Thr Trp Tyr Ala Tyr Thr Ile Pro
1 5 10 15
Glu Lys Tyr Asp Arg
20
<210> 277
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 277
Asn Val Glu Gly Arg Arg Phe Pro Trp Glu Ser Arg Arg Ala Tyr
1 5 10 15
<210> 278
<211> 18
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 278
Asn Ala Lys Trp Arg Asp Gly Ile Thr Thr Ile Glu Gly Tyr Glu Tyr
1 5 10 15
Asp Tyr
<210> 279
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 279
Asn Gln Arg Val Thr Arg Trp Asn Asn Val Phe Glu Tyr
1 5 10
<210> 280
<211> 6
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 280
His Tyr Gly Thr Ser Thr
1 5
<210> 281
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 281
Thr Ala Lys Gly Tyr Gly Ser Ile Leu Asp Ala Leu Asn Lys Val Leu
1 5 10 15
Gln Thr Tyr
<210> 282
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 282
Asn Val Ile Phe Ala Thr Tyr Leu Pro
1 5
<210> 283
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 283
Ser Ala Arg Ile Ile Arg Met Gly Ser Ser Gln Glu Tyr Thr Trp
1 5 10 15
<210> 284
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 284
Asn Ala Lys Asn Arg Ile Phe Arg Asn Gln Gly Thr Gly Tyr
1 5 10
<210> 285
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 285
Asn Ala Glu Leu Glu Gly Trp Gly Ser Pro Val Ser
1 5 10
<210> 286
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 286
Ala Arg Ala Arg Gly Ala Pro Ser Phe Thr Gly Arg Arg Phe Asp Tyr
1 5 10 15
Asp Tyr Arg
<210> 287
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 287
Asn Thr Lys Ser Leu Ser Arg Asp Tyr
1 5
<210> 288
<211> 9
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 288
Asn Leu Glu Arg Gly Trp Thr Asp Tyr
1 5
<210> 289
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 289
Asn Ala Lys Ile Val Pro Arg Ala Ile Met Trp Arg Leu Thr Asp Tyr
1 5 10 15
<210> 290
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 290
Asn Ala Tyr Arg Thr Ser Trp Ser Gly Arg His Glu Tyr
1 5 10
<210> 291
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 291
Asn Ala Asp Leu Tyr Gly Trp Gly Gln Leu Thr Leu Gln Asn Tyr
1 5 10 15
<210> 292
<211> 12
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 292
Lys Ile Thr Val Phe Phe Pro Ser Glu Thr Thr Tyr
1 5 10
<210> 293
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 293
Asn Leu Arg Arg Ala Arg Phe Leu Val Tyr
1 5 10
<210> 294
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 294
Asn Ile Ala Leu His Asn Pro Trp Gly Thr Phe Asp Thr Tyr
1 5 10
<210> 295
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 295
Asn Ala Lys Gly Arg Asn Pro Pro Val Trp Gly Leu Asp Tyr Asn Ser
1 5 10 15
<210> 296
<211> 14
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 296
Asn Ala Asp Ile Ala Ala Met Pro Asp Trp Tyr Ser Ala Tyr
1 5 10
<210> 297
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 297
Ala Ala Asp Ser Gly Ser Glu Pro Ser Pro Leu Tyr Tyr Ser Tyr Glu
1 5 10 15
Tyr Asp Tyr
<210> 298
<211> 5
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 298
Thr Ala Leu Asn Tyr
1 5
<210> 299
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 299
Lys Ala Glu Ala Tyr Arg Ser Phe Ser Tyr Tyr Val Ser Arg Arg Leu
1 5 10 15
Asp Asp Tyr
<210> 300
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 300
Gln Met Thr Ile Arg Glu Arg Phe Ile Pro Phe Arg Thr Ser Asp Tyr
1 5 10 15
<210> 301
<211> 19
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 301
Asn Ala Gly Arg Thr Ser Thr Leu His Phe Gly Asp Ile Trp Pro Gly
1 5 10 15
Ile Asp Tyr
<210> 302
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 302
His Gly Arg Asn Ser Glu Ile Phe Pro Pro Arg Glu Tyr
1 5 10
<210> 303
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 303
Ala Ala Ser Glu Tyr Pro Asn Thr Trp Arg Asp Pro Gly Arg Tyr Ala
1 5 10 15
His
<210> 304
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 304
His Leu Tyr Gly Ala Gly Gly Ile Leu Asp Ala
1 5 10
<210> 305
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 305
Asn Leu Val Trp Ser Tyr His Thr Pro Gly His Asn Tyr
1 5 10
<210> 306
<211> 13
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 306
Asn Leu Val Trp Ser Tyr His Thr Pro Gly Gln Asn Tyr
1 5 10
<210> 307
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 307
His Leu Tyr Gly Ala Gly Gly Arg Leu Asp Ala
1 5 10
<210> 308
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 308
Arg Ala Asp Arg Ile Val Ser Gly Ala Tyr
1 5 10
<210> 309
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 309
Phe Ala Gln Ala Phe Asp Gln Gln His Met Tyr
1 5 10
<210> 310
<211> 15
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 310
Thr Ala Gly Phe Asp Pro Asp Arg Asp Trp Ser Arg Tyr Asp Leu
1 5 10 15
<210> 311
<211> 8
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 311
Lys Leu Gly Val Thr Arg Ser Tyr
1 5
<210> 312
<211> 25
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 312
Gln Leu Gln Leu Val Glu Ser Gly Gly Gly Leu Val Gln Ala Gly Asp
1 5 10 15
Ser Leu Arg Leu Ser Cys Ala Ala Ser
20 25
<210> 313
<211> 17
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 313
Met Gly Trp Phe Arg Gln Ala Pro Gly Lys Glu Arg Glu Phe Val Ala
1 5 10 15
Gly
<210> 314
<211> 38
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 314
Ser Tyr Thr Asp Ser Ala Lys Asp Arg Phe Thr Ile Ser Lys Asp Asn
1 5 10 15
Thr Lys Asn Thr Val Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu Asp
20 25 30
Thr Ala Val Tyr Leu Cys
35
<210> 315
<211> 11
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 315
Trp Gly Gln Gly Thr Gln Val Thr Val Ser Ser
1 5 10
<210> 316
<211> 25
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 316
Gln Val Arg Leu Val Glu Ser Gly Gly Gly Ser Val Gln Ala Gly Glu
1 5 10 15
Thr Leu Arg Leu Ser Cys Thr Ala Ser
20 25
<210> 317
<211> 16
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 317
Met Gly Trp Tyr Arg Gln Gly Pro Gly Asn Glu Cys Glu Met Val Ala
1 5 10 15
<210> 318
<211> 36
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 318
Ala Asp Ser Thr Lys Gly Arg Phe Thr Ile Ser Gln Asp Asn Ala Lys
1 5 10 15
His Thr Leu Tyr Leu Gln Met Asn Ser Leu Lys Pro Glu Asp Thr Gly
20 25 30
Val Tyr Tyr Cys
35
<210> 319
<211> 10
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 319
Gly Gln Gly Thr Arg Val Thr Val Ser Ser
1 5 10

Claims (10)

1.一种可结合BCMA的多肽，其特征在于，包括3个互补决定区CDR1-3，CDR1序列为或包括SEQ ID NO:1-104所示序列之一，CDR2序列为或包括SEQ ID NO:105-207所示序列之一，CDR3序列为或包括SEQ ID NO:208-311所示序列之一。
2.根据权利要求1所述的多肽，其特征在于，所述多肽还包括4个框架区FR1-4，所述FR1-4与所述CDR1-3按顺序交错排列。
3.根据权利要求2所述的多肽，其特征在于，所述多肽为单克隆抗体。
4.权利要求1-3中任一项所述的多肽在制备BCMA检测剂或B细胞淋巴瘤治疗药物中的应用。
5.一种核酸，其特征在于，编码权利要求1-3中任一项所述的多肽。
6.权利要求5所述的核酸在制备B细胞淋巴瘤治疗药物中的应用。
7.权利要求5所述的核酸在制备针对BCMA靶点的细胞治疗药物中的应用。
8.权利要求5所述的核酸在制备针对BCMA靶点的基因治疗药物中的应用。
9.一种BCMA检测剂，其特征在于，所述检测抗体和所述包被抗体分别为权利要求1-3中任一项所述的多肽中的一种。
10.根据权利要求9所述的BCMA检测剂，其特征在于，所述BCMA检测剂为流式检测试剂。
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CN112592405A (zh) * 2020-12-08 2021-04-02 博奥信生物技术（南京）有限公司 抗人bcma纳米抗体及其制备方法和应用
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CN112538115A (zh) * 2020-12-08 2021-03-23 博奥信生物技术（南京）有限公司 一种抗人bcma纳米抗体及其制备方法和应用
CN113265001A (zh) * 2021-02-08 2021-08-17 华道(上海)生物医药有限公司 一种抗b细胞成熟抗原的纳米抗体及其应用
CN112812186A (zh) * 2021-02-08 2021-05-18 华道(上海)生物医药有限公司 一种抗b细胞成熟抗原的纳米抗体及其应用
CN112812186B (zh) * 2021-02-08 2022-07-29 华道(上海)生物医药有限公司 一种抗b细胞成熟抗原的纳米抗体及其应用
CN113265001B (zh) * 2021-02-08 2022-07-29 华道(上海)生物医药有限公司 一种抗b细胞成熟抗原的纳米抗体及其应用
CN115109156A (zh) * 2021-03-22 2022-09-27 浙江纳米抗体技术中心有限公司 一种靶向bcma的纳米抗体及其应用
CN115109156B (zh) * 2021-03-22 2024-03-08 浙江纳米抗体技术中心有限公司 一种靶向bcma的纳米抗体及其应用
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CN117430709A (zh) * 2023-12-21 2024-01-23 康维众和(中山)生物药业有限公司 一种纳米抗体及其应用
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