CN115466328A - 抗pd-1人源化抗体或其抗原结合片段及其应用 - Google Patents
抗pd-1人源化抗体或其抗原结合片段及其应用 Download PDFInfo
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- CN115466328A CN115466328A CN202210643978.4A CN202210643978A CN115466328A CN 115466328 A CN115466328 A CN 115466328A CN 202210643978 A CN202210643978 A CN 202210643978A CN 115466328 A CN115466328 A CN 115466328A
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Abstract
本发明涉及生物医药技术领域,具体而言,涉及抗PD‑1人源化抗体或其抗原结合片段及其应用。该抗体能高效、特异性结合PD‑1,并能有效阻断PD‑1与PD‑L1、PD‑L2结合;因此,该抗体或其抗原结合片段、及其相关的核酸、载体、细胞或药物组合物能够制备用于治疗PD‑1介导的疾病或病症的药物。
Description
技术领域
本发明属于生物医药技术领域。更具体地,涉及抗PD-1人源化抗体或其抗原结合片段及其应用。
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的缺陷和不足,提供抗PD-1人源化抗体或其抗原结合片段及其应用。
本发明的目的是提供抗PD-1人源化抗体或其抗原结合片段,所述抗体包含轻链CDR区和重链CDR区,重链CDR区由HCDR1、HCDR2、HCDR3组成,轻链CDR区由LCDR1、LCDR2、LCDR3组成,HCDR1、HCDR2、HCDR3的氨基酸序列依次如SEQ ID NO:8~10所示,LCDR1、LCDR2、LCDR3的氨基酸序列依次如SEQ ID NO:11~13所示;所述抗体的重链可变区的氨基酸序列如SEQ ID NO:3~5任一项所示。
所述抗体的轻链可变区的氨基酸序列如SEQ ID NO:6~7任一项所示。
本发明还涉及所述抗体或其抗原结合片段相关的核酸、载体、细胞或药物组合物。
本发明还涉及所述抗体或其抗原结合片段、及其相关的核酸、载体、细胞或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。
附图说明
图1是不同浓度的PD-1-112-C2对IL-2/IFN-γ分泌的影响结果图。
图2是不同浓度的PD-1-112-C2对T细胞增殖和T细胞分泌细胞因子IL-2的影响结果图。
图3是不同浓度的PD-1-112-C2对T细胞增殖和T细胞分泌细胞因子IFN-γ的影响结果图。
图4是抗PD-1人源化抗体c53对肿瘤体积的影响结果图。
图5是抗PD-1人源化抗体c53对小鼠生存期的影响结果图。
具体实施方式
以下结合具体实施例来进一步说明本发明,但实施例并不对本发明做任何形式的限定。除非特别说明,本发明采用的试剂、方法和设备为本技术领域常规试剂、方法和设备。
除非特别说明,以下实施例所用试剂和材料均为市购。
本发明涉及抗PD-1人源化抗体或其抗原结合片段,所述抗体包含轻链CDR区和重链CDR区,重链CDR区由HCDR1、HCDR2、HCDR3组成,轻链CDR区由LCDR1、LCDR2、LCDR3组成,HCDR1、HCDR2、HCDR3的氨基酸序列依次如SEQ ID NO:8~10所示,LCDR1、LCDR2、LCDR3的氨基酸序列依次如SEQ ID NO:11~13所示,所述抗体的重链可变区的氨基酸序列如SEQ IDNO:3~5任一项所示;所述抗体的轻链可变区的氨基酸序列如SEQ ID NO:6~7任一项所示。
本发明采用Kabat编号系统标示CDR区,但其他方法标示的CDR区也属于本发明的保护范围。
在一些实施方式中,所述抗体的重链可变区的氨基酸序列如SEQ ID NO:3、轻链可变区的氨基酸序列如SEQ ID NO:6所示,或所述抗体的重链可变区的氨基酸序列如SEQ IDNO:4、轻链可变区的氨基酸序列如SEQ ID NO:7所示,或所述抗体的重链可变区的氨基酸序列如SEQ ID NO:5、轻链可变区的氨基酸序列如SEQ ID NO:7所示。
在一些实施方式中,所述抗体含有重链恒定区和轻链恒定区,所述重链恒定区为IgG1、IgG2、IgG3、IgG4、IgA、IgD、IgE或IgM中的任一种或几种;所述轻链恒定区为κ链或λ链。
在一些实施方式中,所述重链恒定区和轻链恒定区的种属来源选自人、鼠或猴。
在一些实施方式中,所述抗体为嵌合抗体或多特异性抗体(例如双特异性抗体)。
在本发明中,术语“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。
在本发明中,术语“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。
在本发明中,术语“特异性结合”或其类似表述是指抗体或其抗原结合片段对预先确定的抗原上的表位的结合。通常,抗体或其抗原结合片段以大约小于10-6M,例如大约小于10-7M、10-8M、10-9M或10-10M或更小的亲和力(KD)结合。KD是指解离速率与结合速率的比率(koff/kon),该量可通过本领域技术人员熟悉的方法测量。
在一些实施方式中,所述抗原结合片段为F(ab’)2、Fab、scFv、Fv及单域抗体中的任一种或几种。
在本发明中,术语“F(ab’)2”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。
在本发明中,术语“Fab”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。
在本发明中,术语“scFv”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))。
在本发明中,术语“Fv”包含来自重链和轻链的可变区,但缺乏恒定区。
在本发明中,术语“单域抗体”只包含一个重链可变区(VHH)和两个常规的CH2与CH3区,但却不像人工改造的单链抗体(scFv)那样容易相互沾粘,甚至聚集成块。更重要的是单独克隆并表达出来的VHH结构具有与原重链抗体相当的结构稳定性以及与抗原的结合活性,是已知的可结合目标抗原的最小单位。
本发明还涉及一种核酸,编码所述抗PD-1人源化抗体或其抗原结合片段。
在优选实施方式中,所述核酸包括:编码所述抗体或其抗原结合片段的重链可变区的第一核酸,和/或,编码所述抗体或其抗原结合片段的轻链可变区的第二核酸。
在本发明中,核酸通常是RNA或DNA,核酸分子可以是单链或双链的,但优选是双链DNA。当将核酸与另一个核酸序列置于功能关系中时,核酸是“有效连接的”。例如,如果启动子或增强子影响编码序列的转录,那么启动子或增强子有效地连接至所述编码序列。当其连入载体时优选采用DNA。此外,鉴于抗体为膜蛋白,所以核酸通常带有信号肽序列。
本发明还涉及一种载体,所述载体携带所述核酸。
在本发明中,术语“载体(vector)”是可将多聚核苷酸插入其中的一种核酸运载工具。当载体能使插入的多核苷酸编码的蛋白获得表达时,载体称为表达载体。载体可以通过转化,转导或者转染导入宿主细胞,使其携带的遗传物质元件在宿主细胞中获得表达。载体是本领域技术人员公知的,包括但不限于:质粒;噬菌粒;柯斯质粒;人工染色体,例如酵母人工染色体(YAC)、细菌人工染色体(BAC)或P1来源的人工染色体(PAC);噬菌体如λ噬菌体或M13噬菌体及动物病毒等。可用作载体的动物病毒包括但不限于,逆转录酶病毒(包括慢病毒)、腺病毒、腺相关病毒、疱疹病毒(如单纯疱疹病毒)、痘病毒、杆状病毒、乳头瘤病毒、乳头多瘤空泡病毒(如SV40)。
本发明还涉及一种细胞,所述细胞携带所述核酸、含有所述载体或能够表达所述抗体或其抗原结合片段。
本发明还涉及一种药物组合物,所述组合物含有所述抗体或其抗原结合片段、所述核酸、所述载体或所述细胞。
在本发明中,术语“药物组合物”是以允许活性成分的生物学活性有效的形式存在,并且不包含对将施用所述组合物的对象具有不可接受的毒性的另外的成分。
在一些实施方式中,所述药物组合物还包括药学上可接受的载体和/或赋形剂。
在本发明中,术语“药学可接受的载体”可以包括生理学上相容的任何和所有溶剂、分散介质、包衣、抗细菌剂和抗真菌剂、等渗剂和延迟吸收剂等,用来延长抗体的保存限期或效力。
另外,所述抗体或其抗原结合片段、所述核酸、所述载体、所述细胞或所述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,也应在本发明的保护范围之内。
在一些实施方式中,所述药物组合物或药物为适于注射的形式。
在优选实施方式中,所述药物组合物或药物为适于通过皮下注射、皮内注射、静脉内注射、肌内注射或病灶内注射施用的形式。
本发明具有以下有益效果:
本发明提供的抗PD-1人源化抗体或其抗原结合片段能高亲和力结合CHO-hPD1细胞、CHO-cyno细胞和活化的PBMC,其亲和力相比阳性对照显著提高,能高效、特异性结合PD-1,有效阻断配体PD-L1/PD-L2与CHO-hPD1结合,在MLR中,能够阻断PD-1与配体的结合,抑制PD-1信号通路,从而促进T细胞增殖和分泌IL-2、IFN-γ细胞因子;因此,该抗体或其抗原结合片段、及其相关的核酸、载体、细胞或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应用前景广泛。
实施例1抗PD-1抗体的制备
1.免疫原
人工合成人PD-1序列(NCBI NP 005009),上游引物:5’-CCGCAAGCTTGCCGCCACCATG-3’(SEQ ID NO:1)、下游引物:5’-CCGGAATTCTCATTAATGGTGATGGTGATGATGCTGGAACTGGCCGGCAGGTC-3’(SEQ ID 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1×105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),1×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),1×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),细胞系将用于流式细胞计量术(FCM)测定抗PD-1鼠单抗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。
结果如表1所示,数据表明,抗PD-1鼠单抗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,鼠单抗都不结合CHO-mousePD1细胞。
表1
实施例3抗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所示,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OD450值。
结果如表2所示,抗PD-1鼠单抗PD-1-112-C2都能特异性结合PD-1,而不结合CD28其他家族成员。
表2
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所示。
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所示,抗PD-1鼠单抗: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。
表4
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。
不同浓度的PD-1-112-C2对IL-2/IFN-γ分泌的影响结果如图1所示,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-1-112-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。
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的影响结果如图2所示,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的影响结果如图3所示,图2和图3的结果表明抗PD-1抗体: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抗PD-1鼠单抗人源化
对以上获得的抗PD-1鼠单抗PD-1-112-C2(其HCDR1、HCDR2、HCDR3的氨基酸序列依次如SEQ ID NO:8~10所示,LCDR1、LCDR2、LCDR3的氨基酸序列依次如SEQ ID NO:11~13所示,重链可变区序列如SEQ ID NO:14所示,轻链可变区序列如SEQ ID NO:15所示)进行人源化,具体方法为:
人工合成人PD-1序列(NCBI NP 005009),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。将获得的人PD-1重组蛋白进行皮下免疫雌性BAL b/C小鼠,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。筛出的结合hPD-1蛋白抗体阳性的杂交瘤,通过有限稀释法亚克隆至少两次后,体外培养稳定亚克隆并生成小量抗体在进一步筛选后得到PD-1-112-C2克隆。
SEQ ID NO:8:TYYMY
SEQ ID NO:9:GINPSNGGTNFNEKFKS
SEQ ID NO:10:RDSNYDGGFDY
SEQ ID NO:11:RASKSVSTSGYSYMH
SEQ ID NO:12:LAYHLES
SEQ ID NO:13:QHSWELPIT
SEQ ID NO:14:
QVQLQQPGAELVKPGASVKLSCKASGYTFTTYYMYWVKQRPGQGLEWIGGINPSNGGTNFNEKFKSKATLTVDKSSSTAYMQLSSLTSEDSAVYYCTRRDSNYDGGFDYWGQGTTLTVSS
SEQ ID NO:15:
DIVLTQSPASLAVSLGQRATISCRASKSVSTSGYSYMHWYQQKPGQPPKLLIYLAYHLESGVPARFSGSGSGTDFTLNIHPVEEEDAATYYCQHSWELPITFGSGTKLEIKR
参照SEQ ID NO:14和SEQ ID NO:15,在Germline数据库中选取与其非CDR区匹配最好的人源化模板。其中抗体重链的模板为IGHV1,抗体的轻链模板为IGKV1,在以不影响抗体结构稳定性,不影响抗体与抗原结合,不引入糖基化,磷酸化等蛋白修饰位点,不引入已被氧化氨基化等位点,增强结构稳定性的原则下,设计重链人源化序列为VH1-5,设计轻链人源化序列为VL1-3,同时轻链以同源性最高的IGKV7-3*01为另一组人源化设计模板,设计人源化序列VL-4。设计轻重链配对形式为IGHV1/IGKV2为常见配对形式,得到人源化抗体。
根据各人源化抗体轻链和重链氨基酸序列合成基因,用Hind III(NEB)和EcoRI(NEB)双酶切后,将基因片段采用T4 DNA连接酶(TAKARA 2011A)通过Hind III(NEB)/EcoRI(NEB)酶切位点插入到PCDNA3.4A表达载体(Invitrogen)上。将表达载体和转染试剂PEI(Poly science,Inc.Cat.NO.23966)以1:2的比例转染HEK293细胞(Life TechnologicsCat.NO.11625019),并置于CO2培养箱培养5-7天。表达的抗体通过离心回收后按照常规方法进行抗体纯化,得到本发明的抗PD-1人源化抗体(c11、c21、c31、c41、c51、c12、c22、c32、c42、c52、c43、c53、c44、c54);其中,3株抗PD-1人源化抗体(c22、c43、c53)的氨基酸序列如表5所示。
表5 3株抗PD-1人源化抗体(c22、c43、c53)的氨基酸序列
SEQ ID NO:3:
QVQLVQSGAEVKKPGASVKVSCKASGYTFTTYYMYWVRQAPGQGLEWMGGINPSNGGTNFNEKFKSRVTMTVDKSTSTAYMELSSLRSEDTAVYYCTRRDSNYDGGFDYWGQGTTVTVSS
SEQ ID NO:4:
QVQLVQSGAEVKKPGASVKVSCKASGYTFTTYYMYWVRQAPGQGLEWIGGINPSNGGTNYAEKFKGRVTLTVDTSTSTAYMELSSLRSEDTAVYYCTRRDSNYDGGFDYWGQGTTVTVSS
SEQ ID NO:5:
QVQLVQSGAEVKKPGASVKVSCKASGYTFTTYYMYWVRQAPGQGLEWMGGINPSNGGTNYAQKFQGRATMTVDTSTSTAYMELSSLRSEDTAVYYCTRRDSNYDGGFDYWGQGTTVTVSS
SEQ ID NO:6:
DIQLTQSPSSLSASVGDRATITCRASKSVSTSGYSYMHWYQQKPGKAPKLLIYLAYHLESGVPSRFSGSGSGTDFTLTISSVQPEDFATYYCQHSWELPITFGQGTKLEIKR
SEQ ID NO:7:
DIQMTQSPSSLSASVGDRVTITCRASKSVSTSGYSYMHWYQQKPGKAPKLLIYLAYHLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQHSWELPITFGQGTKLEIKR
实施例10抗PD-1人源化抗体亲和力表征
1、实验方法
利用重组技术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的CHO细胞系(CHO-cynoPD1),这两种细胞系将用于流式细胞计量术(FCM)测定抗PD-1人源化候选单抗(c22、c43、c53)的结合表征。具体方法为:
为了评估抗人源化抗体跟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30μg/ml,三倍梯稀释)的人源化抗体,4℃孵育30分钟,缓冲液(PBS含3%BSA)洗涤细胞一次后加入用PE标记的抗人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人源化抗体与CHO-cyno细胞。
2、实验结果
抗PD-1人源化抗体亲和力表征如表6所示,结果显示本发明抗PD-1人源化抗体都能较高亲和力结合CHO-hPD1细胞以及CHO-cynoPD1细胞。
表6抗PD-1人源化抗体亲和力表征
实施例11抗PD-1人源化抗体与活化的PBMC的结合
1、实验方法
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,可以用来进行抗PD-1人源化抗体(c22、c43、c53)与活化的淋巴天然表达PD1的结合实验。具体方法为:
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1×106cells/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2E5/孔加入到96孔U型板中,再加入不同浓度梯度的人源化抗体,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。
2、实验结果
抗PD-1人源化抗体与活化的PBMC的结合能力测定结果如表6所示,结果显示本发明抗PD-1人源化抗体能高亲和力的与活化的淋巴细胞结合。
与实施例2和3中的抗PD-1鼠单抗PD-1-112-C2(表1)相比,本发明抗PD-1人源化抗体(c22、c43、c53)与CHO-hPD1细胞、CHO-cyno细胞和活化的PBMC的结合能力与其相当。
实施例12抗PD-1人源化抗体结合特异性
1、实验方法
将本发明抗PD-1人源化抗体(c22、c43、c53)与四种不同CD28家族成员蛋白结合来验证抗PD-1人源化抗体结合PD-1的特异性。利用标准ELISA方法,将浓度1μg/ml的PD-1(Acro),CD28(Acro),CTLA-4(Acro),ICOS固定在ELISA板上,加入浓度为10μg/ml的人源化抗体,将偶联了过氧化物酶(HRP)的抗人IgG(Fab)作为二抗。TMB显色,终止后,酶标仪读OD450值。
2、实验结果
抗PD-1人源化抗体结合特异性结果如表7所示,结果显示本发明抗PD-1人源化抗体都能特异性结合PD-1,而不结合CD28其他家族成员。
表7抗PD-1人源化抗体结合特异性结果
实施例13抗PD-1人源化抗体的亲和力测定
1、实验方法
ForteBio(Octet Qke)亲和力测定:通过将浓度为5μg/ml的PD-1-his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人源化抗体(c22、c43、c53)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。以Opdivo(ABA0333)为阳性对照。
2、实验结果
抗PD-1人源化抗体的亲和力测定结果如表8所示,结果显示本发明的抗PD-1人源化抗体均能高亲和力结合PD-1。
与实施例5中的抗PD-1鼠单抗PD-1-112-C2(表3)相比,本发明抗PD-1人源化抗体的亲和力与其相当;和阳性对照Opdivo相比(表3),本发明抗PD-1人源化抗体的亲和力显著提高。
表8抗PD-1人源化抗体的亲和力测定结果
实施例14抗PD-1人源化抗体阻断配体PD-L1/PD-L2与CHO-hPD1结合
1、实验方法
通过流式细胞仪分析抗PD-1人源化抗体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mouseIgG1 Fc段融合蛋白:PD-L1-mFc,PD-L2-mFc。
CHO-PD1细胞用缓冲液(PBS含3%BSA)重悬,调整密度为2×106cells/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。向细胞孔中加入浓度为0.2μg/ml的PD-L1-mFc/PD-L2-mFc,4℃孵育30分钟后,再加入浓度梯度稀释的抗PD-1人源化抗体(c22、c43、c53),4℃孵育30分钟。
300g离心5分钟,缓冲液洗涤细胞一次,加入PE标记的山羊抗鼠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。
2、实验结果
抗PD-1人源化抗体的亲和力测定结果如表9所示,结果显示本发明抗PD-1人源化抗体都能有效的阻断PD-L1/PD-L2与细胞CHO-PD1的结合。
表9抗PD-1人源化抗体的亲和力测定结果
实施例15抗PD-1人源化抗体在混合淋巴细胞反应中的影响
1、实验方法
在混合淋巴细胞反应(MLR)中,抗PD-1人源化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。具体方法为:
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,1×104cells/孔的DC细胞与1×105cells/孔的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人源化抗体(c22、c43、c53),另外设置同型对照抗体,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。
3、实验结果
抗PD-1人源化抗体在混合淋巴细胞反应中的影响结果如表10所示,结果显示本发明抗PD-1人源化抗体在MLR实验中,能阻断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。
表10抗PD-1人源化抗体在混合淋巴细胞反应中的影响结果
实施例16抗PD-1人源化抗体对小鼠结肠癌细胞的体内抗肿瘤药效评价
1、实验方法
实验目的:测定抗PD-1人源化抗体(c53)对小鼠结肠癌细胞(MC38细胞)的体内抗肿瘤活性,同时设置同型对照组(Isotype)、阳性对照组(Sintilimab)。
实验材料:hPD1 Knock in小鼠,雌性,6-8周(C57BL/6背景,来源:北京维通达生物技术有限公司);MC38细胞(国家实验细胞共享资源平台);FBS(Gibco,10091-148),0.25%胰蛋白酶-EDTA(Gibco,25200056),DMSO(Sigma,D2650),DPBS(Hyclone,SH30028.02),青霉素-链霉素(Gibco,15140122),DMEM高糖培养基(Gibco,11965084)。胎牛血清(Gibco),谷氨酰胺(Gibco),
仪器设备:电子天平(上海舜宇恒平科学仪器有限公司,JA12002),游标卡尺(上海美耐特实业有限公司,MNT-150T),显微镜(重庆奥特光学仪器有限公司,BDS200),医用离心机(湖南湘仪实验室开发有限公司,L530R),数显恒温水浴锅(普瑞斯机械有限公司,HH-S),二氧化碳培养箱(日本松下健康医疗器械株式会社,MCO-18AC),双人垂直型超净台(无锡易净化设备有限公司,SW-CJ-VS2)。
实验步骤:
细胞培养:将MC38细胞培养在含有10%胎牛血清、1%谷氨酰胺与1%青霉素-链霉素(1:1)的DMEM高糖培养基中。
接种:收集对数生长期的MC38细胞,调节细胞浓度为3×106/mL。取雌性hPD1小鼠40只,皮下接种MC38细胞,接种体积为0.1mL/小鼠,即3×105/小鼠。
给药:接种当天记为第0天(D0),第7天,将小鼠按瘤体积随机分为3组,每组8只,开始给药(MC38肿瘤模型给药剂量、方式以及频率如表11所示)。
记录:D7开始测量肿瘤体积并记录,之后每周2次用游标卡尺测量肿瘤长径和短径。以公式:(1/2)×长径×(短径)2计算肿瘤体积。每只小鼠达到实验终点时(肿瘤体积超过2000mm3达到仁慈终点),颈椎脱臼法处死小鼠,记录生存曲线。
2、实验结果
抗PD-1人源化抗体对肿瘤体积的影响结果如表12和图4所示,可以看出,与Isotype组相比,抗PD-1人源化抗体(c53)对MC38肿瘤模型的肿瘤生长有显著的抑瘤作用(TGI=104.44%,肿瘤全消小鼠7只),与Sintilimab组抗肿瘤作用相当(TGI=105.81%,肿瘤全消小鼠7只)。
抗PD-1人源化抗体对小鼠生存期的影响结果如图5所示,可以看出,与Isotype组相比,抗PD-1人源化抗体(c53)能够明显延长小鼠生存期。
表12抗PD-1人源化抗体对肿瘤体积的影响结果(mm3)
以上结果表明:本发明提供的抗PD-1人源化抗体(c53)能够显著抑制MC38细胞的生长,有效延长小鼠生存期,对治疗小鼠结肠癌具有显著疗效。
上述实施例为本发明较佳的实施方式,但本发明的实施方式并不受上述实施例的限制,其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化,均应为等效的置换方式,都包含在本发明的保护范围之内。
SEQUENCE LISTING
<110> 广东菲鹏制药股份有限公司
<120> 抗PD-1人源化抗体或其抗原结合片段及其应用
<130> 2021
<160> 15
<170> PatentIn version 3.5
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Gly Gly Ile Asn Pro Ser Asn Gly Gly Thr Asn Tyr Ala Gln Lys Phe
50 55 60
Gln Gly Arg Ala Thr Met Thr Val 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
Thr Arg Arg Asp Ser Asn Tyr Asp Gly Gly Phe Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Thr Val Thr Val Ser Ser
115 120
<210> 6
<211> 112
<212> PRT
<213> 人工序列
<400> 6
Asp Ile Gln Leu Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly
1 5 10 15
Asp Arg Ala Thr Ile Thr Cys Arg Ala Ser Lys Ser Val Ser Thr Ser
20 25 30
Gly Tyr Ser Tyr Met His Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro
35 40 45
Lys Leu Leu Ile Tyr Leu Ala Tyr His Leu Glu Ser Gly Val Pro Ser
50 55 60
Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser
65 70 75 80
Ser Val Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln His Ser Trp
85 90 95
Glu Leu Pro Ile Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg
100 105 110
<210> 7
<211> 112
<212> PRT
<213> 人工序列
<400> 7
Asp Ile Gln Met Thr Gln Ser Pro Ser Ser Leu Ser Ala Ser Val Gly
1 5 10 15
Asp Arg Val Thr Ile Thr Cys Arg Ala Ser Lys Ser Val Ser Thr Ser
20 25 30
Gly Tyr Ser Tyr Met His Trp Tyr Gln Gln Lys Pro Gly Lys Ala Pro
35 40 45
Lys Leu Leu Ile Tyr Leu Ala Tyr His Leu Glu Ser Gly Val Pro Ser
50 55 60
Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Thr Ile Ser
65 70 75 80
Ser Leu Gln Pro Glu Asp Phe Ala Thr Tyr Tyr Cys Gln His Ser Trp
85 90 95
Glu Leu Pro Ile Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg
100 105 110
<210> 8
<211> 5
<212> PRT
<213> 人工序列
<400> 8
Thr Tyr Tyr Met Tyr
1 5
<210> 9
<211> 17
<212> PRT
<213> 人工序列
<400> 9
Gly Ile Asn Pro Ser Asn Gly Gly Thr Asn Phe Asn Glu Lys Phe Lys
1 5 10 15
Ser
<210> 10
<211> 11
<212> PRT
<213> 人工序列
<400> 10
Arg Asp Ser Asn Tyr Asp Gly Gly Phe Asp Tyr
1 5 10
<210> 11
<211> 15
<212> PRT
<213> 人工序列
<400> 11
Arg Ala Ser Lys Ser Val Ser Thr Ser Gly Tyr Ser Tyr Met His
1 5 10 15
<210> 12
<211> 7
<212> PRT
<213> 人工序列
<400> 12
Leu Ala Tyr His Leu Glu Ser
1 5
<210> 13
<211> 9
<212> PRT
<213> 人工序列
<400> 13
Gln His Ser Trp Glu Leu Pro Ile Thr
1 5
<210> 14
<211> 120
<212> PRT
<213> 人工序列
<400> 14
Gln Val Gln Leu Gln Gln Pro Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Thr Tyr
20 25 30
Tyr Met Tyr Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Gly Ile Asn Pro Ser Asn Gly Gly Thr Asn Phe Asn Glu Lys Phe
50 55 60
Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Thr Arg Arg Asp Ser Asn Tyr Asp Gly Gly Phe Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Thr Leu Thr Val Ser Ser
115 120
<210> 15
<211> 112
<212> PRT
<213> 人工序列
<400> 15
Asp Ile Val Leu Thr Gln Ser Pro Ala Ser Leu Ala Val Ser Leu Gly
1 5 10 15
Gln Arg Ala Thr Ile Ser Cys Arg Ala Ser Lys Ser Val Ser Thr Ser
20 25 30
Gly Tyr Ser Tyr Met His Trp Tyr Gln Gln Lys Pro Gly Gln Pro Pro
35 40 45
Lys Leu Leu Ile Tyr Leu Ala Tyr His Leu Glu Ser Gly Val Pro Ala
50 55 60
Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Asn Ile His
65 70 75 80
Pro Val Glu Glu Glu Asp Ala Ala Thr Tyr Tyr Cys Gln His Ser Trp
85 90 95
Glu Leu Pro Ile Thr Phe Gly Ser Gly Thr Lys Leu Glu Ile Lys Arg
100 105 110
Claims (10)
1.抗PD-1人源化抗体或其抗原结合片段,所述抗体包含轻链CDR区和重链CDR区,重链CDR区由HCDR1、HCDR2、HCDR3组成,轻链CDR区由LCDR1、LCDR2、LCDR3组成,HCDR1、HCDR2、HCDR3的氨基酸序列依次如SEQ ID NO:8~10所示,LCDR1、LCDR2、LCDR3的氨基酸序列依次如SEQ ID NO:11~13所示,其特征在于,所述抗体的重链可变区的氨基酸序列如SEQ IDNO:3~5任一项所示。
2.根据权利要求1所述抗体或其抗原结合片段,其特征在于,所述抗体的轻链可变区的氨基酸序列如SEQ ID NO:6~7任一项所示。
3.根据权利要求2所述抗体或其抗原结合片段,其特征在于,所述抗体的重链可变区的氨基酸序列如SEQ ID NO:3、轻链可变区的氨基酸序列如SEQ ID NO:6所示,或所述抗体的重链可变区的氨基酸序列如SEQ ID NO:4、轻链可变区的氨基酸序列如SEQ ID NO:7所示,或所述抗体的重链可变区的氨基酸序列如SEQ ID NO:5、轻链可变区的氨基酸序列如SEQID NO:7所示。
4.根据权利要求1~3任一所述抗体或其抗原结合片段,其特征在于,所述抗体含有重链恒定区和轻链恒定区,所述重链恒定区为IgG1、IgG2、IgG3、IgG4、IgA、IgD、IgE或IgM中的任一种或几种;所述轻链恒定区为κ链或λ链。
5.根据权利要求1~4任一所述抗体或其抗原结合片段,其特征在于,所述抗体为嵌合抗体或多特异性抗体;所述抗原结合片段为F(ab’)2、Fab、scFv、Fv及单域抗体中的任一种或几种。
6.一种核酸,其特征在于,编码所述抗PD-1人源化抗体或其抗原结合片段;
优选地,所述核酸包括:编码所述抗体或其抗原结合片段的重链可变区的第一核酸,和/或,编码所述抗体或其抗原结合片段的轻链可变区的第二核酸。
7.一种载体,其特征在于,所述载体携带权利要求6所述核酸。
8.一种细胞,其特征在于,所述细胞携带权利要求6所述核酸、含有权利要求7所述载体或能够表达权利要求1~5任一所述抗体或其抗原结合片段。
9.一种药物组合物,其特征在于,所述组合物含有权利要求1~5任一所述抗体或其抗原结合片段、权利要求6所述核酸、权利要求7所述载体或权利要求8所述细胞。
10.权利要求1~5任一所述抗体或其抗原结合片段、权利要求6所述核酸、权利要求7所述载体、权利要求8所述细胞或权利要求9所述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。
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Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN110637031A (zh) * | 2017-03-04 | 2019-12-31 | 湘潭腾华生物科技有限公司 | 程序性死亡蛋白1(pd-1)的重组抗体及其用途 |
CN111349162A (zh) * | 2018-12-21 | 2020-06-30 | 神州细胞工程有限公司 | 人源化抗pd-1抗体及其用途 |
CN111670197A (zh) * | 2017-12-05 | 2020-09-15 | 普莱戈斯瑞恩癌症有限责任公司 | 治疗癌症的抗前胃泌激素抗体与免疫疗法的组合疗法 |
Family Cites Families (13)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2010029434A1 (en) * | 2008-09-12 | 2010-03-18 | Isis Innovation Limited | Pd-1 specific antibodies and uses thereof |
JOP20200094A1 (ar) * | 2014-01-24 | 2017-06-16 | Dana Farber Cancer Inst Inc | جزيئات جسم مضاد لـ pd-1 واستخداماتها |
US10513558B2 (en) * | 2015-07-13 | 2019-12-24 | Cytomx Therapeutics, Inc. | Anti-PD1 antibodies, activatable anti-PD1 antibodies, and methods of use thereof |
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US20190225689A1 (en) * | 2018-01-22 | 2019-07-25 | Janssen Biotech, Inc. | Methods of treating cancers with antagonistic anti-pd-1 antibodies |
WO2019148410A1 (en) * | 2018-02-01 | 2019-08-08 | Merck Sharp & Dohme Corp. | Anti-pd-1 antibodies |
WO2019148412A1 (en) * | 2018-02-01 | 2019-08-08 | Merck Sharp & Dohme Corp. | Anti-pd-1/lag3 bispecific antibodies |
GB201803745D0 (en) * | 2018-03-08 | 2018-04-25 | Ultrahuman Eight Ltd | PD1 binding agents |
WO2020127366A1 (en) * | 2018-12-21 | 2020-06-25 | Ose Immunotherapeutics | Humanized anti-human-pd-1 antibody |
AR118619A1 (es) * | 2019-04-10 | 2021-10-20 | Bio Thera Solutions Ltd | Anticuerpos de unión a pd-1 |
TW202112825A (zh) * | 2019-09-06 | 2021-04-01 | 大陸商北京拓界生物醫藥科技有限公司 | 抗pd-1單域抗體、其衍生蛋白及其醫藥用途 |
CN113004414B (zh) * | 2019-12-20 | 2022-09-06 | 广东菲鹏制药股份有限公司 | 抗PD1和TGFβ的双功能抗体及其制备方法,以及含有其的药物组合物 |
CN113004408B (zh) * | 2019-12-20 | 2022-07-01 | 广东菲鹏制药股份有限公司 | 抗人程序死亡因子-1单克隆抗体 |
-
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Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN110637031A (zh) * | 2017-03-04 | 2019-12-31 | 湘潭腾华生物科技有限公司 | 程序性死亡蛋白1(pd-1)的重组抗体及其用途 |
CN111670197A (zh) * | 2017-12-05 | 2020-09-15 | 普莱戈斯瑞恩癌症有限责任公司 | 治疗癌症的抗前胃泌激素抗体与免疫疗法的组合疗法 |
CN111349162A (zh) * | 2018-12-21 | 2020-06-30 | 神州细胞工程有限公司 | 人源化抗pd-1抗体及其用途 |
Non-Patent Citations (1)
Title |
---|
李斌;刘朝奇;王见之;史继静;覃晓琳;吕佰瑞;王梦瑶;韩琴;: "人PD-L1胞外区基因的克隆、原核表达及抗体制备", 解放军医学杂志, no. 08, 1 August 2010 (2010-08-01) * |
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