CN113004408B - 抗人程序死亡因子-1单克隆抗体 - Google Patents
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Abstract
本发明提供了人PD‑1抗体、其抗原结合片段及其医药用途。还提供了包含所述抗体CDR区的嵌合抗体、包含人PD‑1抗体及其抗原结合片段的药物组合物,以及所述抗体在制备用于治疗疾病或病症的药物中的用途。
Description
技术领域
本申请涉及免疫学领域。具体涉及鼠抗人程序死亡因子-1(PD1)单克隆抗体及其应用。
背景技术
程序死亡因子-1(PD1)是CD28家族成员,在活化的B细胞,T细胞及髓样细胞上表达。人PD1由基因Pdcd1编码,位于2q37.3,全长9.6kb,由5个外显子和4个内含子组成,其上游包含663bp的启动子。PD1为55KDa的I型跨膜蛋白,分子结构由胞外区,跨膜区及胞内区组成,胞外区含有一个免疫球蛋白可变区IgV结构域,胞内区含有免疫受体酪氨酸抑制基序(ITIM)和免疫受体酪氨酸转换作用模块(ITSM)。PD-1胞外区氨基酸序列和CTLA-4有24%的同源性,与CD28有28%的同源性。在T细胞被激活后,PD-1主要通过ITIM将酪氨酸磷脂酶SHP2集合,导致下游效应分子去磷酸化。
PD-1有两个配体:PD-L1和PD-L2。PD-L1和PD-L2都是B7同源物,PDL基因位于人染色体9P24.2位点,大小为42kb,其分子结构均包含一个免疫球蛋白样可变区结构域,一个恒定区样结构域,一个跨膜区和一个短的胞质尾巴。
PD-1与PD-L1和PD-L2结合后可下调T细胞的活化。PD-L1表达于多种肿瘤细胞表面,这些肿瘤细胞包括:肺癌,胃癌,肝癌,食道癌,肾癌,卵巢癌,宫颈癌,乳腺癌,皮肤癌,结肠癌,膀胱癌,神经胶质癌,头颈癌,口腔鳞状细胞癌。而且这些癌症周边发现大量表达PD-L1的CD8+T细胞,临床结果统计显示,PD-L1在肿瘤细胞上的高表达水平与癌症患者不良预后有关。
发明内容
本发明要解决的技术问题是克服现有抗体结合PD-1的亲和力和特异性不高,不能有效阻断PD-1与PD-L1、PD-L2结合的缺陷和不足,提供了新的能高效结合PD-1并能阻断PD-1与PD-L1和PD-L2结合的抗体,鼠单抗在SEB超抗原刺激PBMC及混合淋巴细胞反应实验(MLR)中,更能促进T细胞分泌细胞因子IL2和IFNγ。
本发明提供了抗PD1的单克隆抗体。具体地,本发明提供了以下几个方面:
1.抗PD1的抗体或其抗原结合片段,包含选自以下各项组成的组的重链CDR1-3和轻链CDR1-3:
(1)HCDR1:由SEQ ID NO:8的序列表示;HCDR2:由SEQ ID NO:9的序列表示;HCDR3:由SEQ ID NO:10的序列表示;
LCDR1:由SEQ ID NO:20的序列表示;LCDR2:由SEQ ID NO:21的序列表示;LCDR3:由SEQ ID NO:22的序列表示;
(2)HCDR1:由SEQ ID NO:12的序列表示;HCDR2:由SEQ ID NO:13的序列表示;HCDR3:由SEQ ID NO:14的序列表示;
LCDR1:由SEQ ID NO:24的序列表示;LCDR2:由SEQ ID NO:25的序列表示;LCDR3:由SEQ ID NO:26的序列表示;
(3)HCDR1:由SEQ ID NO:16的序列表示;HCDR2:由SEQ ID NO:17的序列表示;HCDR3:由SEQ ID NO:18的序列表示;
LCDR1:由SEQ ID NO:28的序列表示;LCDR2:由SEQ ID NO:29的序列表示;LCDR3:由SEQ ID NO:30的序列表示。
2.上述1所述的抗体或其抗原结合片段,其中所述抗体包括:
(1)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:7所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:19所示的氨基酸序列;
(2)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:11所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:23所示的氨基酸序列;
(3)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:15所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:27所示的氨基酸序列。
3.上述1-2任一项所述的抗体或其抗原结合片段,其中所述抗体还包含重链恒定区和轻链恒定区,优选来自人IgG或IgM,更优选IgG4。
4.上述1-2任一项所述的抗体或其抗原结合片段,其中所述抗原结合片段选自Fab,scFv,Fab′,dAb,F(ab′)2,Fv或Fab/c。
5.上述1-2任一项所述的抗体或其抗原结合片段,其中所述抗体是嵌合抗体或多特异性抗体(例如双特异性抗体)。
6.编码上述1-5任一项所述的抗体或其抗原结合片段的多核苷酸。
7.药物组合物,其包含上述1-5任一项所述的抗体或其抗原结合片段;可选地,其还包括药学上可接受的载体和/或赋形剂。
8.如上述1-5任一项所述的PD-1抗体或其抗原结合片段、如上述7所述的药物组合物在制备用于治疗PD-1介导的疾病或病症的药物中的用途。
9.如上述1-5任一项所述的抗体或其抗原结合片段、如上述7所述的药物组合物在预防和/或治疗和/或辅助治疗和/或诊断治疗肿瘤疾病或在制备预防和/或治疗和/或辅助治疗和/或诊断治疗肿瘤疾病的药物中的用途,优选地,所述肿瘤选自肺癌,胃癌,肝癌,食道癌,肾癌,卵巢癌,宫颈癌,乳腺癌,皮肤癌,结肠癌,膀胱癌,神经胶质癌,头颈癌,口腔鳞状细胞癌。
10.上述7所述的药物组合物或上述9所述的用途,所述药物组合物或药物为适于注射的形式,优选是适于通过皮下注射、皮内注射、静脉内注射、肌内注射或病灶内注射施用的形式。
如本发明中所使用的,术语“轻链”包括全长轻链和其具有可变区序列来赋予结合特异性的片段。全长轻链包括可变区结构域VL和恒定区结构域CL。轻链的可变区结构域在多肽的氨基末端。轻链包括κ链和λ链。
如本发明中所使用的,术语“重链”包括全长重链和其具有可变区序列来赋予结合特异性的片段。全长重链包括可变区结构域VH和3个恒定区结构域CH1、CH2和CH3。VH结构域在多肽的氨基末端,并且CH结构域在羧基末端,CH3最靠近多肽的羧基末端。重链可具有任何同种型,包括IgG(包括IgG1、IgG2、IgG3和IgG4亚型)、IgA(包括IgA1和IgA2亚型)、IgM和IgE。
如本发明中所使用的,术语“Fab片段”由一条轻链和CH1以及一条重链的可变区组成。Fab分子的重链不能与另一条重链分子形成二硫键。
如本发明中所使用的,术语“Fc”区含有两个包含抗体的CH1和CH2结构域的重链片段。两个重链片段通过两个或更多个二硫键以及通过CH3结构域的疏水相互作用保持在一起。
如本发明中所使用的,术语“Fab’片段”含有一条轻链和一条重链的部分(其含有VH结构域和CH1结构域以及还有CH1与CH2结构域之间的区域的部分),以便可在两个Fab’片段的两条重链之间形成链间二硫键以形成F(ab’)2分子。
如本发明中所使用的,术语“F(ab’)2片段”含有两条轻链和两条含有CH1与CH2结构域之间的恒定区的部分的重链,以便在两条重链之间形成链间二硫键。F(ab’)2片段从而由通过两条重链之间的二硫键保持在一起的两个Fab’片段组成。
如本发明中所使用的,术语“Fv区”包含来自重链和轻链的可变区,但缺乏恒定区。
如本发明中所使用的,术语“Fd”片段意指由VH和CH1结构域组成的抗体片段(Ward等人,Nature 341:544-546(1989))。
如本发明中所使用的,术语“dAb”片段(Ward等人,Nature 341:544-546(1989))由VH结构域组成。
如本发明中所使用的,术语“Fab/c”片段是免疫球蛋白经胃蛋白酶消化形成的抗体裂解中间产物,其兼有Fab和Fc区的优点,即具有扩散能力强,体内代谢清除慢的特点,并能保持高度亲和力(刘建军,《细胞与分子免疫学杂志》,1989(4):29-29)。
如本发明中所使用的,术语“单链抗体”是其中重链与轻链可变区通过柔性接头连接来形成单条多肽链(其形成抗原结合区)的Fv分子(参见,例如,Bird等人,Science.242:423-426(1988)和Huston等人,Proc.Natl.Acad.Sci.USA.90:5879-5883(1988))。
如本发明中所使用的,术语“结构域抗体”是只含有重链的可变区或轻链的可变区的免疫功能性免疫球蛋白片段,包括多价结构域抗体或双价结构域抗体。在一些情况下,两个或更多个VH区通过肽接头共价地连接,以生成多价结构域抗体(特别是双价结构域抗体)。双价结构域抗体的两个VH区可靶向相同或不同的抗原。
如本发明中所使用的,术语“多特异性抗原结合蛋白”或“多特异性抗体”是靶向不止一种抗原或表位的抗原结合蛋白或抗体。
如本发明中所使用的,术语“双特异性”、“双重特异性”或“双功能性”抗原结合蛋白或抗体是分别具有两个不同的抗原结合位点的杂交抗原结合蛋白或抗体。双特异性抗体是一种多特异性抗原结合蛋白或多特异性抗体,并且可通过多种方法产生,包括,但不限于杂交瘤的融合或Fab’片段的连接。参见,例如,Songsivilai和Lachmann,1990,Clin.Exp.Immunol.79:315-321;Kostelny等人,1992,J.Immunol.148:1547-1553。双特异性抗原结合蛋白或抗体的两个结合位点将结合两个不同的表位,所述表位存在于相同或不同的蛋白质靶标上。
如本发明中所使用的,术语“表位”是指,抗原上被免疫球蛋白或抗体特异性结合的部位。“表位”在本领域内也称为“抗原决定簇”。表位或抗原决定簇通常由分子的化学活性表面基团例如氨基酸或碳水化合物或糖侧链组成并且通常具有特定的三维结构特征以及特定的电荷特征。例如,表位通常以独特的空间构象包括至少3,4,5,6,7,8,9,10,11,12,13,14或15个连续或非连续的氨基酸,其可以是“线性的”或“构象的”。参见,例如,EpitopeMapping Protocols in Methods in Molecular Biology,第66卷,G.E.Morris,Ed.(1996)。在线性表位中,蛋白质与相互作用分子(例如抗体)之间的所有相互作用的点沿着蛋白质的一级氨基酸序列线性存在。在构象表位中,相互作用的点跨越彼此分开的蛋白质氨基酸残基而存在。
如本文中使用的,术语“相似性”或“序列相似性”、“同一性”是指两个或更多个蛋白质或多肽分子的序列之间的关系,如通过比对和比较序列测定的。“百分比同一性”意指被比较的分子中的氨基酸之间的相同残基的百分比,并且可基于待比较的最小的分子的大小来计算。为了进行这些计算,必须通过特定的数学模型或计算机程序(即,“算法”)来解决比对中的缺口(如果有的话)。当用于多肽时,术语″大体上的同一性″,是指两个肽序列,当例如使用程序GAP或BESTFIT,利用程序提供的缺省缺口权重,进行最佳对齐时,共有至少70%、75%或80%的序列同一性,至少90%或95%的序列同一性,和至少97%、98%或99%的序列同一性。在某些情况下,不相同的残基位点相异在于保守氨基酸置换。″保守氨基酸置换″是这样的置换,即其中氨基酸残基被具有拥有相似化学性质(例如,电荷或正在水性)的侧链R基团的另一个氨基酸残基置换。一般地,保守氨基酸置换将基本上不改变蛋白质的功能性质。在其中两个或更多个氨基酸序列彼此相异在于保守置换的情况下,可上调百分比序列同一性以就置换的保守性质进行修正。用于进行该调整的方法对于本领域技术人员来说是熟知的。参见例如Pearson,Methods Mol.Biol.243:307-31(1994)。具有拥有相似化学性质的侧链的氨基酸组的实例包括1)脂肪族羟基侧链:甘氨酸、丙氨酸、缬氨酸、亮氨酸和异亮氨酸:2)脂肪族羟基侧链:丝氨酸和苏氨酸:3)含酰胺侧链:天冬酰胺和谷氨酰胺:4)芳香族侧链:苯丙氨酸、酪氨酸和色氨酸:5)碱性侧链:赖氨酸、精氨酸和组氨酸:6)酸性侧链:天冬氨酸和谷氨酸;和7)含硫侧链:半胱氨酸和甲硫氨酸。例如,保守氨基酸置换组是缬氨酸-亮氨酸-异亮氨酸-甘氨酸-丙氨酸、苯丙氨酸-酪氨酸、苏氨酸-丝氨酸、赖氨酸-精氨酸、谷氨酸-天冬氨酸和天冬酰胺-谷氨酰胺。
可选择地,保守置换是在Gonnet等人,Science 256:1443-45(1992)(通过引用合并入本文)中公开的PAM250对数似然矩阵(PAM250log-likelihood matrix)中具有正值的任何变化。“中度保守的”置换是在PAM250对数似然矩阵中具有非负值的任何变化。
附图说明
图1:不同浓度的PD-1-112-C2对IL-2/IFN-γ分泌的影响。
图2:不同浓度的PD-1-97-C2对IL-2/IFN-γ分泌的影响。
图3:不同浓度的PD-1-76-C2对IL-2/IFN-γ分泌的影响。
图4:不同浓度的PD-1-97-C2对T细胞增殖和T细胞分泌细胞因子IL-2的影响。
图5:不同浓度的PD-1-112-C2对T细胞增殖和T细胞分泌细胞因子IL-2的影响。
图6:不同浓度的PD-1-76-C2对T细胞增殖和T细胞分泌细胞因子IL-2的影响。
图7:不同浓度的PD-1-97-C2对T细胞增殖和T细胞分泌细胞因子IFN-γ的影响。
图8:不同浓度的PD-1-112-C2对T细胞增殖和T细胞分泌细胞因子IFN-γ的影响。
图9:不同浓度的PD-1-76-C2对T细胞增殖和T细胞分泌细胞因子IFN-γ的影响。
具体实施方式
下面将结合实施例对本发明的实施方案进行详细描述,但是本领域技术人员将会理解,下列实施例仅用于说明本发明,而不应视为限定本发明的范围。实施例中未注明具体条件者,按照常规条件或制造商建议的条件进行。所用试剂或仪器未注明生产厂商者,均为可以通过市购获得的常规产品。
实施例1:抗PD-1抗体的制备
1.免疫源
人工合成人PD-1序列(NCBI NP 005009),上游引物CCGCAAGCTTGCCGCCACCATG(SEQID NO:1)和下游引物CCGGAATTCTCATTAATGGTGATGGTGATGATGCTGGAACTGGCCGGCAGGTC(SEQID NO:2),PCR扩增胞外端,经Hind III和EcoRI双酶切后克隆到pCDNA3.4A真核表达系统,以此质粒转染293细胞,收获上清后纯化获得人PD-1重组蛋白(hPD-1)。
2.免疫动物
将125ug浓度为1.23mg/ml的hPD-1重组蛋白作为抗原与等量免疫佐剂弗氏佐剂(Sigma-Aldrich F5881)混合,取5只6周雌性BAL b/C小鼠进行皮下免疫,每只小鼠免疫抗原量为25ug。在初次免疫以后,每周进行一次相同剂量的加强免疫。总共5次免疫后,通过采集小鼠尾血来监测免疫应答。通过FACS筛选(如下所述),使用具有足够抗hPD-1免疫球蛋白滴度的小鼠来进行融合。用抗原进行腹腔加强免疫后3天,处死小鼠并取出脾进行细胞融合。
3.选择产生抗hPD-1抗体的BAL b/C小鼠
为了选择产生抗hPD-1抗体的BAL b/C小鼠,通过FACS对经免疫的小鼠血清进行测试。来自hPD-1重组蛋白免疫小鼠的血清稀释液与转染了hPD1的CHO细胞在4摄氏度孵育30分钟,以PBS洗涤3次后,加入0.4ug/ml的PE山羊抗小鼠IgG(Biolegend 405307)并在4摄氏度育孵30分钟。以PBS洗涤3次后,将样品放入Beckman Coulter公司流式细胞仪(CytoFLEXA00-1-1102)检测以验证其是否可以与转染了hPD1的CHO细胞结合,筛出产生抗hPD-1抗体的BAL b/C小鼠,然后进行细胞融合。
4.生成产生针对hPD-1的鼠单克隆抗体杂交瘤
将经免疫过BAL b/C小鼠的脾细胞与小鼠骨髓瘤细胞融合,然后对得到的杂交瘤进行抗原特异性抗体的筛选。用PEG 1500(Roche 10783641001)将来自经免疫过小鼠的脾细胞的单细胞悬浮液与五分之一数目且不分泌免疫球蛋白的小鼠骨髓瘤细胞(SP2/0,ATCCCRL1581)进行细胞融合。将融合细胞以约1x105个/孔铺在96孔细胞培养板中,放入培养箱(Panasonic MCO-18AIC),培养条件为37℃,5%CO2。随后在HAT选择性培养基中培养大约一周,所述培养基在1640培养基中含1X青链霉素双抗(Gibco 15140122),1X HAT(SigmaCRLP-7185)和有20%胎牛血清(Royacel RY-F11-01)。1周后,再用HT培养基(1640培养基含1X青链霉素双抗(gibco 15140122),1X HT(gibco 11067030)和20%胎牛血清(RoyacelRY-F11-01))替换HAT的培养基进行培养,然后通过FACS检测融合板的细胞培养上清液,筛出分泌可结合hPD-1蛋白抗体的杂交瘤。对分泌可结合hPD-1蛋白抗体的杂交瘤重新铺板,再次进行筛选。对筛出的结合hPD-1蛋白抗体阳性的杂交瘤,通过有限释法亚克隆至少两次。然后体外培养稳定亚克隆并生成小量抗体用于进一步分析。选择杂交瘤克隆PD1-112-C2,PD1-97-C2,PD-76-C2进行下一步的分析。
实施例2:抗PD-1鼠单抗亲和力表征
根据常规方法,利用重组技术制备在细胞表面表达重组人PD-1的CHO(中国仓鼠卵巢细胞)细胞系(CHO-hPD1),表达猴PD1(Uniprot:B0LAJ2)的CHO细胞系(CHO-cynoPD1),表达小鼠PD1(Uniprot:Q02242)的CHO细胞系(CHO-mousePD1),这三种细胞系将用于流式细胞计量术(FCM)测定抗PD-1鼠单抗PD-1-76-C2,PD-1-97-C2,PD-1-112-C2的结合表征。
为了评估抗PD-1鼠单抗跟CHO-hPD1的结合,在96孔板里加入2×105CHO-hPD1细胞以及经浓度梯度稀释(初始浓度为10μg/ml,三倍梯稀释)的抗PD-1鼠单抗,4℃孵育30分钟,缓冲液(PBS含3%BSA)洗涤细胞一次后加入用PE标记的抗鼠IgG(Fc)Ab(Biolegend)荧光二抗,4℃孵育30分钟后缓冲液洗涤细胞一次后PBS重悬,而后细胞悬液经CytoFlex(贝克曼流式细胞仪)进行流式细胞分析,根据染色的平均荧光强度(MFI)来测量结合到细胞上的抗体量;同样的方法用于评估此抗PD-1鼠单抗与CHO-cyno细胞、CHO-mousePD1(在本发明中有时简写为“CHO-mPD1)细胞的结合。结果见表1,数据表明,抗PD-1鼠单抗PD-1-76-C2,PD-1-97-C2,PD-1-112-C2都能较高亲和力结合CHO-hPD1细胞以及CHO-cyno细胞;同时,三株鼠单抗都不结合CHO-mousePD1细胞(数据略)。
实施例3:PD-1抗体与活化的PBMC的结合
新鲜人外周血单个核细胞(PBMC)在PHA(Sigma)的刺激下,淋巴细胞能活化增殖,并在第三天最高丰度表达PD1,可以用来进行PD-1抗体与活化的淋巴天然表达PD1的结合实验。
新鲜人的外周血通过淋巴分离液梯度离心方法得到PBMC后,调整密度为1×106细胞/ml接种到T75中,同时加入终浓度为1μg/ml PHA-L(Sigma)刺激淋巴细胞增殖,37℃,5%CO2培养箱中静置3天后,取出细胞悬液,离心去掉上清,缓冲液(PBS含3%BSA)重悬,按照2*105个/孔加入到96孔U型板中,再加入从30μg/ml起始,3倍梯度稀释,共10个浓度梯度的抗PD1抗体,4℃孵育30分钟后300g离心5分钟,缓冲液洗涤细胞一次,加入PE标记的山羊抗人IgG荧光抗体(Biolegend),4℃孵育30分钟;离心洗涤细胞一次后PBS重悬细胞,而后进行CytoFlex流式细胞仪分析,检测结合到PBMC上的抗体量。结果如表1,抗PD1抗体能高亲和力的与活化的淋巴细胞结合。
实施例4:抗PD-1鼠单抗结合特异性
将抗PD-1鼠单抗与四种不同CD28家族成员蛋白结合来验证抗体结合PD-1的特异性。利用标准ELISA方法,将浓度1μg/ml的PD-1,CD28,CTLA-4,ICOS(ACRO)固定在ELISA板上,加入浓度为10μg/ml的抗人PD-1鼠单抗,将偶联了过氧化物酶(HRP)的抗小鼠IgG作为二抗(Sigma)。TMB显色,终止后,酶标仪读值。结果如表2显示,抗PD-1鼠单抗PD-1-76-C2,PD-1-97-C2,PD-1-112-C2都能特异性结合PD-1,而不结合CD28其他家族成员。
实施例5:生物层干涉(BLI)法测定抗人PD-1鼠单抗的亲和力
ForteBio(Octet Qke)亲和力测定:通过将浓度为5μg/ml的PD-1-his(ACRO)重组蛋白装载在HISIK生物传感器上120秒,然后将装载好的传感器在标准缓冲液(PBST,PBS+0.02%Tuween20)中平衡120秒,之后传感器转移到抗PD-1鼠单抗稀释液中停留180秒测量结合速率,再转移至标准缓冲液中停留20分钟测量解离速率。最后利用动力学模型来进行分析,数据处理结果如表3。
实施例6:抗PD-1鼠单抗阻断配体PD-L1/PD-L2与CHO-hPD1结合
通过流式细胞仪分析抗PD-1鼠单抗对于阻断配体与经转染CHO细胞表面稳定表达PD-1的结合能力。实验中用到的配体蛋白为重组的PD-L1/PD-L2胞外段连接上human IgG1Fc段融合蛋白:PD-L1-hFc(ACRO),PD-L2-hFc(ACRO)。
CHO-PD1细胞用缓冲液(PBS含3%BSA)重悬,调整密度为2×106细胞/ml,100μl/孔细胞悬液加入到96孔U型板中,300g离心5分钟后,去掉上清。
后续过程可分为两种阻断模式进行:模式一,向细胞孔中加入浓度为3μg/ml的PD-L1-hFc/PD-L2-hFc,4℃孵育30分钟后,再加入从30μg/ml起始,3倍梯度稀释,共10个浓度梯度释的抗PD-1鼠单抗,4℃孵育30分钟;模式二,向细胞孔中加入从30μg/ml起始,3倍梯度稀释,共10个浓度梯度的抗PD-1鼠单抗,4℃孵育30分钟后,再加入浓度为3μg/ml的PD-L1-hFc/PD-L2-hFc蛋白,4℃孵育30分钟。
300g离心5分钟,缓冲液洗涤细胞一次,加入PE标记的山羊抗人IgG荧光抗体(Biolegend),4℃孵育30分钟。离心洗涤细胞一次后PBS重悬细胞,而后进行CytoFlex流式细胞仪分析,检测结合到细胞上的配体蛋白量,计算得到PD-1抗体结合阻断的IC50值。结果如表4,三株抗PD-1鼠单抗:PD-1-76-C2,PD-1-97-C2,PD-1-112-C2在两种模式中都能有效的阻断PD-L1/PD-L2与细胞CHO-PD1的结合。
实施例7:抗PD-1抗体对SEB刺激的PBMC细胞的细胞因子释放的影响
此实施例中,过夜培养的外周血单个核细胞(PBMC)通过加入超抗原金黄色葡萄球菌肠毒素B(SEB)刺激时,在抗PD-1抗体存在或缺失情况下,检测细胞因子分泌的影响。
新鲜的外周单个核细胞(PBMC)用含10%FBS的X-VIVO 15培养基(LONZA)重悬后,添加到T25培养瓶,37℃,5%CO2静置过夜培养,次日,取悬浮细胞,离心后,新鲜X-VIVO(含10%FBS)培养基重悬,并添加终浓度为200ng/ml的SEB超抗原(Toxin technology),然后按照每孔1×105细胞添加到96孔平板中,同时添加不同浓度的抗PD-1抗体,另外设置同型对照抗体(mIgG1同型对照抗体(Biolegend);hIgG4同型对照抗体(Biolegend)),无抗体对照孔。3天后,样本孔中取样,使用IL2/IFN-γHuman Uncoated ELISA Kit(eBioscience)试剂盒测量IL-2/IFN-γ水平,结果显示于图1-3,抗PD-1抗体以浓度依赖性方式提高IL-2/IFN-γ的分泌。这些结果表明,在SEB超抗原刺激PBMC中,抗PD-1抗体:PD-1-76-C2,PD-1-97-C2,PD-1-112-C2能进一步促进T细胞分泌细胞因子。
实施例8:抗PD-1抗体在混合淋巴细胞反应中的影响
在混合淋巴细胞反应(MLR)中,抗PD-1抗体的存在与否能证明PD1信号被阻断情形下的T细胞增殖情况和T细胞分泌细胞因子水平高低。
用CD14 MicroBeads,human(Miltenyi)从新鲜PBMC中分离得到CD14+单核细胞(monocyte),在GM-CSF/IL-4存在下,诱导6天后,添加TNF-α,3天后诱导DC的成熟;实验当天,用EasySepTM Human T Cell Enrichment Kit(StemCell)纯化PBMC中的T细胞,1×104的DC细胞与1×105的T细胞混合培养,并且向混合细胞中添加不同浓度梯度的抗PD-1抗体,另外设置同型对照抗体(mIgG1同型对照抗体和hIgG4同型对照抗体(Biolegend)),无抗体对照孔。混合培养3天后,取上清进行IL-2的检测,再培养2天后,取上清进行IFN-γ的检测。结果显示于图4-9,抗PD-1抗体:PD-1-76-C2,PD-1-97-C2,PD-1-112-C2在MLR实验中,能以抗体浓度依赖性方式的阻断PD1与配体的结合,抑制PD1信号通路,从而促进T细胞增殖,促进T细胞分泌IL-2,IFN-γ细胞因子。
实施例9:候选抗体可变区序列
将候选抗体杂交瘤细胞培养到1*107个细胞,300g离心10分钟,使用MagExtractor-RNA-试剂盒(东洋纺/NPK-201F)提取RNA,将此RNA变性后进行逆转录获得cDNA,以cDNA为模板分别对重链和轻链进行PCR。Light Chain基因以Universal PrimerA Mix(UPM)、Nested Universal Primer A(NUP)和mkR引物进行扩增,Heavy Chain基因以UniversalPrimer A Mix(UPM)、Nested Universal Primer A(NUP)和mHR引物进行扩增。其中LightChain的引物对扩增出0.8KB左右的目的条带,Heavy Chain的引物对扩增出1.4KB左右的目的条带。
Universal Primer A Mix(UPM):
5’>CTAATACGACTCACTATAGGGCAAGCAGTGGTATCAACGCAGAGT<3’(SEQ ID NO:3);
Nested Universal Primer A(NUP):
5’>AAGCAGTGGTATCAACGCAGAGT<3’(SEQ ID NO:4)
mkR:5’>TTTTCCTTTTGAATTCCTAACACTCATTCCTGTTGAAGC<3’(SEQ ID NO:5);
mHR:5’>TTTTCCTTTTGAATTCTCATTTACCAGGAGAGTGGGAGA<3’(SEQ ID NO:6)。
反应总体积为50μL,含PCR 2×缓冲液25μL,DNA模板1μL,KOD DNA Polymerase 1μL,dNTP(25mM)10μL,以及特异性上、下引物(终浓度200nmol/L)各1μL,并加消毒双蒸水至总体积50μL。
反应条件为94℃预变性5分钟,然后依次94℃变性30秒,接着56℃退火30秒,72℃延伸40秒分钟,共30个循环。
将扩增产物测序后,得到杂交瘤克隆76,97和112的重链和轻链的可变区为:
PD-1-76-C2
重链可变区:SEQ ID NO:7
HCDR1:SEQ ID NO:8
HCDR2:SEQ ID NO:9
HCDR3:SEQ ID NO:10
PD-1-97-C2
重链可变区:SEQ ID NO:11
HCDR1:SEQ ID NO:12
HCDR2:SEQ ID NO:13
HCDR3:SEQ ID NO:14
PD-1-112-C2
重链可变区:SEQ ID NO:15
HCDR1:SEQ ID NO:16
HCDR2:SEQ ID NO:17
HCDR3:SEQ ID NO:18
PD-1-76-C2
轻链可变区:SEQ ID NO:19
LCDR1:SEQ ID NO:20
LCDR2:SEQ ID NO:21
LCDR3:SEQ ID NO:22
PD-1-97-C2
轻链可变区:SEQ ID NO:23
LCDR1:SEQ ID NO:24
LCDR2:SEQ ID NO:25
LCDR3:SEQ ID NO:26
PD-1-112-C2
轻链可变区:SEQ ID NO:27
LCDR1:SEQ ID NO:28
LCDR2:SEQ ID NO:29
LCDR3:SEQ ID NO:30
表1
表2
表3
待测抗体 | kon(1/Ms) | kdis(1/s) | KD(M) |
Opdivo(ABA0333) | 1.38E+06 | 3.63E-06 | 2.62E-12 |
PD-1-97-C2 | 4.68E+05 | <1.0E-07 | <1.0E-12 |
PD-1-112-C2 | 7.32E+05 | 3.18E-05 | 4.35E-11 |
PD-1-76-C2 | 7.71E+05 | <1.0E-07 | <1.0E-12 |
表4
Claims (13)
1.抗PD1的抗体或其抗原结合片段,其特征在于包含选自以下各项组成的组的重链CDR1-3和轻链CDR1-3:
(1)HCDR1:由SEQ ID NO:8的序列表示;HCDR2:由SEQ ID NO:9的序列表示;HCDR3:由SEQ ID NO:10的序列表示;
LCDR1:由SEQ ID NO:20的序列表示;LCDR2:由SEQ ID NO:21的序列表示;LCDR3:由SEQID NO:22的序列表示;
(2)HCDR1:由SEQ ID NO:12的序列表示;HCDR2:由SEQ ID NO:13的序列表示;HCDR3:由SEQ ID NO:14的序列表示;
LCDR1:由SEQ ID NO:24的序列表示;LCDR2:由SEQ ID NO:25的序列表示;LCDR3:由SEQID NO:26的序列表示;
(3)HCDR1:由SEQ ID NO:16的序列表示;HCDR2:由SEQ ID NO:17的序列表示;HCDR3:由SEQ ID NO:18的序列表示;
LCDR1:由SEQ ID NO:28的序列表示;LCDR2:由SEQ ID NO:29的序列表示;LCDR3:由SEQID NO:30的序列表示。
2.权利要求1所述的抗体或其抗原结合片段,其中所述抗体包括:
(1)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:7所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:19所示的氨基酸序列;
(2)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:11所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:23所示的氨基酸序列;
(3)重链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:15所示的氨基酸序列,和
轻链可变区,其包含下述序列或由下述序列组成:
SEQ ID NO:27所示的氨基酸序列。
3.权利要求1-2任一项所述的抗体或其抗原结合片段,其中所述抗体还包含重链恒定区和轻链恒定区。
4.权利要求3所述的抗体或其抗原结合片段,其中所述重链恒定区来自人IgG或IgM。
5.权利要求3所述的抗体或其抗原结合片段,其中所述重链恒定区来自人IgG4。
6.权利要求1-2任一项所述的抗体或其抗原结合片段,其中所述抗原结合片段选自Fab,scFv,Fab',F(ab')2,Fv或Fab/c。
7.权利要求1-2任一项所述的抗体或其抗原结合片段,其中所述抗体是嵌合抗体或多特异性抗体。
8.编码权利要求1-7任一项所述的抗体或其抗原结合片段的多核苷酸。
9.药物组合物,其包含权利要求1-7任一项所述的抗体或其抗原结合片段。
10.权利要求9所述的药物组合物,其还包括药学上可接受的载体和/或赋形剂。
11.如权利要求1-7任一项所述的抗体或其抗原结合片段、如权利要求9或10所述的药物组合物在制备预防和/或治疗和/或诊断肿瘤疾病的药物中的用途,其中所述肿瘤选自肺癌,胃癌,肝癌,食道癌,肾癌,卵巢癌,宫颈癌,乳腺癌,皮肤癌,结肠癌,膀胱癌,神经胶质癌,头颈癌,口腔鳞状细胞癌。
12.权利要求9或10所述的药物组合物或权利要求11所述的用途,所述药物组合物或药物为适于注射的形式。
13.权利要求9或10所述的药物组合物或权利要求11所述的用途,所述药物组合物或药物是适于通过皮下注射、皮内注射、静脉内注射、肌内注射或病灶内注射施用的形式。
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Publication number | Priority date | Publication date | Assignee | Title |
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CN105026428A (zh) * | 2013-12-12 | 2015-11-04 | 上海恒瑞医药有限公司 | PD-l抗体、其抗原结合片段及其医药用途 |
WO2017019846A1 (en) * | 2015-07-30 | 2017-02-02 | Macrogenics, Inc. | Pd-1-binding molecules and methods use thereof |
CN106632674A (zh) * | 2015-10-30 | 2017-05-10 | 中山康方生物医药有限公司 | 一种抗pd-1单克隆抗体、其药物组合物及其用途 |
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