CN114805607B - EGFRvIII-PDL1-GMCSF肿瘤疫苗及其制备方法和应用 - Google Patents
EGFRvIII-PDL1-GMCSF肿瘤疫苗及其制备方法和应用 Download PDFInfo
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Abstract
本发明公开了一种EGFRvIII‑PDL1‑GMCSF肿瘤疫苗及其制备方法和应用,其中,EGFRvIII‑PDL1‑GMCSF肿瘤疫苗中的融合蛋白的氨基酸序列如SEQ ID NO.1所示。本发明的融合蛋白可与DC负载作为肿瘤疫苗可以提高肿瘤特异性抗原的免疫原性,抗肿瘤免疫耐受性,抗肿瘤免疫逃逸,为高度表达PD‑L1的肿瘤免疫治疗提供了一种新的有效策略,具有良好的安全性和可观的前景,同时也促进以EGFRvIII为靶点的药物开发,为癌症的治疗带来了新的希望。
Description
技术领域
本发明属于医药技术领域,更具体地,本发明涉及一种EGFRvIII-PDL1-GMCSF融合蛋白、由所述EGFRvIII-PDL1-GMCSF融合蛋白制备而得的肿瘤疫苗及其制备方法和应用。
背景技术
肿瘤免疫治疗是指通过激活人体免疫***,依靠自身的免疫机能,对肿瘤细胞和组织进行杀灭的疗法,具有疗效好,无耐药性,有效清除残余肿瘤细胞,巩固疗效,长期作用,预防肿瘤复发等特点。与传统的治疗方法不同,肿瘤免疫治疗的直接作用靶标不是肿瘤组织,而是人体自身的免疫***。近年来,包括免疫检查点抑制剂、过继性细胞免疫疗法、肿瘤疫苗、溶瘤病毒等肿瘤免疫疗法在癌症治疗中取得重大突破,并彻底改变了肿瘤学领域。
基于免疫检查点抑制剂和疫苗介导的免疫治疗被报道逆转免疫抑制的肿瘤环境,刺激抗原提呈,并诱导抗肿瘤T细胞反应的免疫治疗策略。树突状细胞(Dendritic cell,DC)是已知体内功能最强的专职抗原呈递细胞,是引发肿瘤抗原强免疫应答的关键,但肿瘤宿主体内肿瘤DC浸润较少且功能受损,通过将患者体内DC细胞的前体细胞分离出来,在体外培养,并使之负载肿瘤抗原肽段,然后回输到患者体内,继而通过DC细胞激发特异性抗肿瘤T细胞反应,制成DC肿瘤疫苗,是获得肿瘤宿主强免疫应答的有效策略。
迄今为止,肿瘤疫苗在癌症中的应用已完成200多项临床试验,这些研究主要在黑色素瘤、***癌、胶质母细胞瘤或肾细胞癌患者中进行,并且证明了肿瘤疫苗诱导抗癌NK细胞,CD8+和CD4+T细胞反应的临床安全性和有效性。
DC细胞在抗原呈递过程中,主要以MHC-Ⅱ抗原肽复合体的形式将抗原呈递给T淋巴细胞。而粒细胞-巨噬细胞集落刺激因子(Granulocyte-macrophage ColonyStimulating Factor,GM-CSF)是影响DC增殖、成熟和迁移的主要细胞因子,并能增加DC表面的MHC-Ⅱ分子的表达,从而增加抗原被辅助性T淋巴细胞的识别机会,激活B淋巴细胞或细胞毒性T淋巴细胞等效应细胞,增强免疫应答,因此被认为是理想的佐剂。相较于传统的铝盐佐剂和油佐剂等无法诱导有效的细胞毒性T细胞及Th1型反应等缺陷问题,研究表明,分泌GM-CSF的肿瘤疫苗会在接种部位诱导DC的大量积累,进而激活肿瘤特异的T细胞,诱导抗肿瘤反应。
在抗肿瘤免疫反应过程中,肿瘤细胞会通过多种调节机制抑制T细胞活化,减弱细胞毒性T细胞的攻击,导致免疫逃逸或免疫耐受。在这些调节机制中,程序性死亡受体1(PD-1)和程序性死亡配体1(PD-L1)分别是在T细胞和肿瘤细胞上的重要免疫抑制检查点,PD-1与PD-L1的结合导致T细胞免疫反应的抑制。
近年来,作用于PD1/PD-L1通路的免疫疗法以其显著的临床疗效、持久的反应性和低毒疗效等优点受到广泛关注。PD-1/PD-L1单克隆抗体已被开发出作为免疫检查点抑制剂用于癌症治疗,如黑色素瘤和非小细胞肺癌等,以消除免疫***上的“刹车”并恢复T细胞攻击肿瘤细胞的能力。
PD-L1主要表达于胰腺癌等多种实体恶性肿瘤的肿瘤细胞和抗原提呈细胞表面,是肿瘤免疫治疗和疫苗研制的理想肿瘤特异性靶点。临床试验表明,抗PD-1或抗PD-L1抗体的重复给药在一部分癌症患者中产生持久反应,但是很大一部分癌症患者对这些免疫检查点抑制剂没有反应,其原因可能是由于这些癌症患者缺乏足够的抗肿瘤T细胞。有研究表明,负载PDL1-Vax的DC(见PMID:31805690)在免疫小鼠中诱导抗PD-L1抗体和T细胞反应,并且PD-L1特异性CTL对PD-L1+的肿瘤细胞具有溶细胞活性。
胰腺癌是免疫抵抗力最强的肿瘤类型之一,在临床治疗中仍然是一个巨大的挑战。随着发病率的持续增加和死亡率的极小变化,到2030年,胰腺癌预计将成为癌症相关死亡的第二大原因。由于传统放疗、化疗等治疗方法对胰腺癌患者疗效有限,使得特异性分子抑制剂的使用成为胰腺癌治疗中一个极具潜力的研发方向。表皮生长因子受体(epidermalgrowth factor receptor,EGFR)是一种受体酪氨酸激酶,在非小细胞肺癌、转移性结直肠癌、胶质母细胞瘤、头颈癌、胰腺癌和乳腺癌等癌症中普遍上调,是癌症治疗中一个重要的分子靶标。表皮生长因子受体III型突变体(epidermal growth factor receptor variantIII,EGFRvIII)是EGFR最常见的突变型,已有研究表明:EGFRvIII可通过调控Ras/Raf/MEK/细胞外调节蛋白激酶(extracellular regulated protein kinases,ERK)等信号通路,影响肿瘤的发生和发展,尤其是在肿瘤的放疗和化疗过程中,发挥类似免疫逃逸的功能。EGFRvIII在正常组织中不表达,在肿瘤细胞表面高表达,使其成为抗肿瘤免疫治疗的理想靶点。
目前肿瘤疫苗面临的挑战是如何提高肿瘤抗原的免疫原性,从而可以降低肿瘤免疫耐受性,避免肿瘤免疫逃逸。
发明内容
基于此,本发明的目的之一在于提供一种EGFRvIII-PDL1-GMCSF融合蛋白,所述融合蛋白可以做为肿瘤疫苗的一部分,提高抗原的免疫原性,抗肿瘤免疫逃逸。
实现上述发明目的的具体技术方案包括如下:
一种EGFRvIII-PDL1-GMCSF融合蛋白,所述融合蛋白的氨基酸序列如SEQ ID NO.1所示。
一种EGFRvIII-PDL1-GMCSF融合蛋白,所述融合蛋白是由如SEQ ID NO.2所示的核苷酸序列所编码。
本发明还提供了上述EGFRvIII-PDL1-GMCSF融合蛋白的制备方法,所述方法包括以下步骤:
(1)、合成包含了人EGFRvIII、Th表位、PDL1和GMCSF的基因序列的融合基因片段;所述融合基因片段的核苷酸序列如SEQ ID NO.2所示;
(2)、对步骤(1)的融合基因片段和pET-21a质粒载体进行NdeI和XhoI双酶切,切胶纯化试剂盒回收,连接,得到表达质粒pET-21a/EGFRvIII-PDL1-GMCSF;
(3)、将表达质粒pET-21a/EGFRvIII-PDL1-GMCSF转入BL21(DE3)表达菌株中,经IPTG诱导,获得目的蛋白,经纯化、透析,即得EGFRvIII-PDL1-GMCSF融合蛋白。
本发明还提供了上述EGFRvIII-PDL1-GMCSF融合蛋白在制备肿瘤疫苗中的应用。
一种肿瘤疫苗,所述肿瘤疫苗的活性成分为EGFRvIII-PDL1-GMCSF融合蛋白。
在其中一些实施例中,所述肿瘤疫苗的融合蛋白负载量为80μg/ml~120μg/ml。
在其中一些实施例中,所述肿瘤疫苗的融合蛋白负载量为95μg/ml~105μg/ml。
本发明还提供了上述肿瘤疫苗的制备方法。
一种肿瘤疫苗的制备方法,所述方法包括以下步骤:
(1)、在培养有树突状细胞的培养基中,加入80μg/ml~120μg/ml的EGFRvIII-PDL1-GMCSF融合蛋白培养过夜;
(2)、再用45μg/ml~55μg/ml EGFRvIII-PDL1-GMCSF融合蛋白对树突状细胞进行脉冲处理1h~3h,即得。
在其中一些实施例中,所述步骤(1)中的培养基为含有20ng/ml GM-CSF和20ng/ml重组小鼠IL-4的RPMI-1640培养基。
本发明还提供了上述肿瘤疫苗在制备治疗实体瘤的药物中的应用。
在其中一些实施例中,所述实体瘤为胰腺癌、结肠癌、黑色素瘤、白血病、乳腺癌、肺癌、或***癌。
在其中一些实施例中,所述实体瘤为胰腺癌。
与现有技术相比,本发明具有以下有益效果:
1、在本发明中,设计并成功表达了EGFRvIII-PDL1-GMCSF融合蛋白,该融合蛋白可与DC负载作为肿瘤疫苗(以EGFRvIII为靶点,融合PD-L1片段,以GM-CSF作为佐剂),可以提高肿瘤特异性抗原的免疫原性,利用GM-CSF募集肿瘤内DC来激活免疫***杀伤肿瘤,从而抗肿瘤免疫耐受性,抗肿瘤免疫逃逸。
2、以本发明的EGFRvIII-PDL1-GMCSF融合蛋白作为活性成分的肿瘤疫苗,在荷瘤小鼠体内验证了其可以诱导有效的肿瘤特异性细胞毒性T淋巴细胞(CTLs)反应,明显抑制胰腺癌细胞的生长,延长小鼠的生存期,为其他高度表达PD-L1的肿瘤(结肠癌、黑色素瘤、白血病、乳腺癌、肺癌、***癌)等实体瘤免疫治疗提供了一种新的有效策略,具有良好的安全性和可观的前景,同时也促进以EGFRvIII为靶点的药物开发,这些无疑为癌症的治疗带来了新的希望。
附图说明
图1为本发明实施例1构建的pET-21a/EGFRvIII-PDL1-GMCSF表达质粒的结构示意图。
图2为本发明实施例2中EGFRvIII-PDL1-GMCSF融合蛋白的SDS-PAGE图谱。
图3为本发明实施例2中对EGFRvIII-PDL1-GMCSF融合蛋白进行免疫印迹分析(WB)的图谱。
图4为本发明实施例3中EGFRvIII-PDL1-GMCSF融合蛋白纯化过程中,裂解液、穿流液和洗脱部分的SDS-PAGE图谱。
图5为本发明实施例3中,用His标签抗体对透析处理后的EGFRvIII-PDL1-GMCSF融合蛋白进行免疫印迹分析(WB)的图谱。
图6为本发明实施例4中EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗免疫小鼠的流程图。
图7为本发明实施例4中通过细胞表面染色和流式细胞仪检测小鼠CD4+T细胞和CD8+T细胞的增殖情况。
图8为本发明实施例4中通过细胞内染色和流式细胞仪检测CD4+T细胞中产生IL-2的频率。
图9为本发明实施例4中通过细胞内染色和流式细胞仪检测CD4+T细胞中产生IFN-γ的频率。
图10为本发明实施例4中通过细胞内染色和流式细胞仪检测CD8+T细胞中产生IL-2的频率。
图11为本发明实施例4中通过细胞内染色和流式细胞仪检测CD8+T细胞中产生IFN-γ的频率。
图12为本发明实施例4中小鼠接种Panc02的肿瘤生长曲线图(n=4),****P<0.0001。
图13为本发明实施例4中肿瘤疫苗接种后荷瘤小鼠的存活曲线(n=4),**P<0.01。
图14为本发明实施例4中免疫小鼠肝、肾切片的H&E染色分析。
具体实施方式
为了便于理解本发明,下面将对本发明进行更全面的描述。本发明可以以许多不同的形式来实现,并不限于本文所描述的实施例。相反地,提供这些实施例的目的是使对本发明公开内容的理解更加透彻全面。
除非另有定义,本发明所使用的所有的技术和科学术语与属于本发明的技术领域的技术人员通常理解的含义相同。本发明的说明书中所使用的术语只是为了描述具体的实施例的目的,不用于限制本发明。本发明所使用的术语“和/或”包括一个或多个相关的所列项目的任意的和所有的组合。
除非另外指明,本发明的实施例中所使用的实验方法,均为常规实验方法,实施例中所用到的各种试剂耗材,均为市售产品。
本发明的第一方面,提供了一种EGFRvIII-PDL1-GMCSF融合蛋白,所述融合蛋白的融合片段包括人表皮生长因子受体III型突变体(EGFRvIII)、程序性死亡配体1(PD-L1)和粒细胞-巨噬细胞集落刺激因子(GM-CSF),所述融合蛋白的氨基酸序列如SEQ ID NO.1所示,其由如SEQ ID NO.2所示的核苷酸序列所编码。
氨基酸序列(SEQ ID NO.1,453 residues,Expected MW with His-tag:50.73kD)
MLEEKKGNYVVTDHAKFVAAWTLKAAALEEKKGNYVVTDHAKFVAAWTLKAAALEEKKGNYVVTDHGSCVRACGADSYEMEEDGVRKCKKCEGPCRKVCNGIGIGEFKDSLSINATNIKHFKNCTSISGDLHILPVAFRGDSFTHTPPLDPQEFTVTVPKDLYVVEYGSNMTIECKFPVEKQLDLAALIVYWEMEDKNIIQFVHGEEDLKVQHSSYRQRARLLKDQLSLGNAALQITDVKLQDAGVYRCMISYGGADYKRITVKVNAPYNKINQRILVVDPVTSEHELTCQAEGPNGSGSGMWLQSLLLLGTVACSISAPARSPSPSTQPWEHVNAIQEARRLLNLSRDTAAEMNETVEVISEMFDLQEPTCLQTRLELYKQGLRGSLTKLKGPLTMMASHYKQHCPPTPETSCATQIITFESFKENLKDFLLVIPFDCWEPVQELEHHHHHH
核苷酸序列(SEQ ID NO.2)
CATATGCTAGAGGAGAAGAAGGGCAACTACGTTGTGACCGATCACGCCAAGTTCGTGGCCGCCTGGACCCTGAAGGCCGCCGCCTTAGAAGAAAAAAAAGGGAACTATGTCGTGACGGACCATGCGAAATTTGTCGCGGCGTGGACATTGAAAGCGGCGGCGCTGGAGGAAAAGAAAGGTAATTATGTGGTGACAGATCACGGCTCGTGCGTCCGAGCCTGTGGGGCCGACAGCTATGAGATGGAGGAAGACGGCGTCCGCAAGTGTAAGAAGTGCGAAGGGCCTTGCCGCAAAGTGTGTAACGGAATAGGTATTGGTGAATTTAAAGACTCACTCTCCATAAATGCTACGAATATTAAACACTTCAAAAACTGCACCTCCATCAGTGGCGATCTCCACATCCTGCCGGTGGCATTTAGGGGTGACTCCTTCACACATACTCCTCCTCTGGATCCACAGGAATTTACTGTCACGGTTCCCAAGGACCTATATGTGGTAGAGTATGGTAGCAATATGACAATTGAATGCAAATTCCCAGTAGAAAAACAATTAGACCTGGCTGCACTAATTGTCTATTGGGAAATGGAGGATAAGAACATTATTCAATTTGTGCATGGAGAGGAAGACCTGAAGGTTCAGCATAGTAGCTACAGACAGAGGGCCCGGCTGTTGAAGGACCAGCTCTCCCTGGGAAATGCTGCACTTCAGATCACAGATGTGAAATTGCAGGATGCAGGGGTGTACCGCTGCATGATCAGCTATGGTGGTGCCGACTACAAGCGAATTACTGTGAAAGTCAATGCCCCATACAACAAAATCAACCAAAGAATTTTGGTTGTGGATCCAGTCACCTCTGAACATGAACTGACATGTCAGGCTGAGGGTCCGAACGGCAGCGGCAGCGGCATGTGGCTGCAGAGCCTGCTGCTCTTGGGCACTGTGGCCTGCAGCATCTCTGCACCCGCCCGCTCGCCCAGCCCCAGCACACAGCCCTGGGAGCATGTGAATGCCATCCAGGAGGCCCGGCGTCTCCTGAACCTGAGTAGAGACACTGCTGCTGAGATGAATGAAACAGTAGAAGTCATCTCAGAAATGTTTGACCTCCAGGAGCCGACCTGCCTACAGACCCGCCTGGAGCTGTACAAGCAGGGCCTGCGGGGCAGCCTCACCAAGCTCAAGGGCCCCTTGACCATGATGGCCAGCCACTACAAACAGCACTGCCCTCCAACCCCGGAAACTTCCTGTGCAACCCAGATTATCACCTTTGAAAGTTTCAAAGAGAACCTGAAGGACTTTCTGCTTGTCATCCCCTTTGACTGCTGGGAGCCAGTCCAGGAGCTCGAGCACCACCACCACCACCAC
在本发明的第二个方面,提供了EGFRvIII-PDL1-GMCSF融合蛋白的制备方法,包括融合蛋白的表达、纯化、透析步骤。
在本发明的第三个方面,提供了利用EGFRvIII-PDL1-GMCSF融合蛋白在制备肿瘤疫苗中的应用。将EGFRvIII-PDL1-GMCSF融合蛋白与树突状细胞DC脉冲并免疫小鼠,发挥抗肿瘤免疫应答的作用。
在本发明的第四方面,提供了一种肿瘤疫苗,所述肿瘤疫苗的活性成分为EGFRvIII-PDL1-GMCSF融合蛋白。
在本发明的第五方面,提供了上述肿瘤疫苗在制备治疗实体瘤的药物中的应用。所述实体瘤为胰腺癌、结肠癌、黑色素瘤、白血病、乳腺癌、肺癌、或***癌。更优选为胰腺癌。
以下结合附图和具体实施例来详细说明本发明。
实施例1 pET-21a/EGFRvIII-PDL1-GMCSF表达质粒的构建与鉴定
本实施例构建了pET-21a/EGFRvIII-PDL1-GMCSF表达质粒,并进行了鉴定。具体包括以下步骤:
(1)设计一段由人EGFRvIII序列、Th表位序列、PD-L1序列和GM-CSF融合而成的基因序列,交由金斯瑞生物科技股份有限公司合成该基因片段;其核苷酸序列如SEQ ID NO.2所示;
(2)、利用pET-21a质粒载体启动子下游的NdeI和XhoI限制位点,在pET-21a质粒载体中***上述(1)合成的融合基因片段。
(3)、将***了融合基因片段的pET-21a质粒载体的连接液转化DH5a大肠杆菌,挑取10个克隆进行PCR鉴定和测序鉴定。
构建得到的表达质粒pET-21a/EGFRvIII-PDL1-GMCSF的结构如图1所示。
实施例2 pET-21a/EGFRvIII-PDL1-GMCSF表达质粒中融合蛋白的诱导表达
抽提测序正确的表达质粒pET-21a/EGFRvIII-PDL1-GMCSF,溶于适量的TE溶液中备用。将质粒转入BL21(DE3)表达菌株中。具体方法:
(1)、取出在-80℃冻存的E.coli BL21(DE3)感受态细胞(上海唯地),立即放置于冰上,待解冻后转化;
(2)、取1ul质粒加到100uL感受态细胞中,轻轻混匀,在冰上孵育30min,42℃热激90S,立即置冰上3min;
(3)、加入在37℃预热的500ul LB液体无抗培养基,37℃,200rpm摇50min;取100-200μl菌液涂布在含有Amp(50μg/ml)的LB平板上,放入37℃培养箱培养过夜。
(4)、取单个菌落接种于2ml LB液体培养基(含50μg/ml Amp)中,37℃200r/min震摇培养3h至对数生长期,取200ul培养菌液作对照(未诱导),剩余的培养基中加入IPTG至终浓度为1mM后,37℃震摇培养3h,收集培养菌液于离心管中。
(5)、各取200μl菌液于离心管中,离心去上清,分别取40μL H2O重悬(诱后全菌),诱导前全菌样同样处理,加入10μL 5×loading Buffer(还原),混匀,煮沸5分钟,12000g离心5min,取10μL上清进行SDS-PAGE电泳和免疫印迹分析(WB,一抗为Pdcd-1L1 Antibody(H-130)rabbitpolyclonalAntibody 1:1000(OmnimAbs);二抗为Anti-rabbit(800)1:10000(goat anti-rabbit IgG-HRP:sc-2004)(Santa Cruz)。
结果分别如图2和图3所示。结果表明,pET-21a/EGFRvIII-PDL1-GMCSF表达质粒经IPTG诱导或无IPTG的条件下,目的蛋白(即融合蛋白EGFRvIII-PDL1-GMCSF)以包涵体形式表达,其大小约为50.73kDa,与预期分子量一致。
实施例3 EGFRvIII-PDL1-GMCSF融合蛋白的纯化
一、将实施例2步骤(4)中的培养菌液以5000g在4℃离心收集,然后重悬于超声缓冲液中(PBS,1%TritonX-100,1mMEDTA,pH7.4),裂解物在冰上以400W的功率超声200次(每超声3秒,间隔5秒),然后15000g离心30分钟后去上清,保留沉淀,即包涵体蛋白。
二、在变性条件下,根据试剂盒的说明(Qiagen),用Ni-NTA柱纯化EGFRvIII-PDL1-GMCSF融合蛋白(带有His标签),通过SDS-PAGE进一步分析裂解液、穿流液和洗脱液部分,如图4结果所示,经过了Ni柱的纯化,与裂解液和穿流液相比,洗脱液中的杂蛋白含量明显减少,达到了纯化的目的。
三、将纯化后的蛋白取一部分样品做Western blot分析,具体步骤如下:
(1)、以每个泳道50μg的量进行蛋白质电泳(12%SDS-PAGE),电泳后的蛋白用金斯瑞eBlotL1快速湿转仪转膜转移到硝酸纤维膜上(Pall Corporation)。
(2)、膜用5%的脱脂奶粉室温封闭1小时然后用PBST洗3次。
(3)、用抗His标签抗体(Santa Cruz)4℃振荡杂交过夜,次日用PBST洗膜5次后用异硫氰酸荧光素(FITC)标记山羊抗鼠二抗(碧云天A0216)室温杂交2小时再用PBST洗膜5次。
(4)、用Amersham Imager 680曝光分析。
四、为了保持生理一致性,对步骤二纯化后的洗脱部分用透析袋进行进一步透析处理,将溶解蛋白的8M尿素溶液置换成PBS溶液。透析后的蛋白溶液用超滤管超滤浓缩。浓缩后的蛋白溶液用BCA试剂盒(Thermo;IH117217)进行定量分析,检测蛋白浓度。透析后的蛋白进行SDS-Page后考马斯亮蓝分析,如图5所示,透析处理后的EGFRvIII-PDL1-GMCSF融合蛋白的SDS-PAGE结果有一条分子量相当的条带,蛋白纯化成功。
采用本实施例的纯化方法,从1L细菌培养液中获得约10mg高纯度EGFRvIII-PDL1-GMCSF融合蛋白,用于进一步的功能鉴定。
实施例4 EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗对胰腺癌的治疗作用
本实施例首先通过EGFRvIII-PDL1-GMCSF融合蛋白脉冲DC,获得了EGFRvIII-PDL1-GMCSF融合蛋白负载的DC(树突状细胞)疫苗和PBS负载的树突状细胞(作为对照),再用于免疫小鼠,检测小鼠脾脏细胞IL-2和IFN-γ分泌,测量肿瘤的生长情况,并检测分析了抗肿瘤过程中肝肾损伤情况。
一、肿瘤疫苗的制备
方法如下:
1、小鼠骨髓从四肢冲出来,穿过尼龙网,用氯化铵裂解红细胞。用RPMI-1640大面积洗涤后,在添加10%FBS、mGM-CSF(20ng/ml)和重组小鼠IL-4(20ng/ml;peprotech)的RPMI-1640中培养细胞。培养48小时后去除非粘附性粒细胞。每隔一天,用含有20ng/mlrmGM-CSF和20ng/ml重组小鼠IL-4的新鲜RPMI-1640培养基替换上清液。所有培养物在37℃的5%湿化二氧化碳中培养。在最后16小时内以100ng/ml添加细菌脂多糖(LPS;Sigma)进行DC成熟。培养7天后,流式细胞仪(FACS)测定80%以上的细胞表达特征性的DC特异性标记物(CD40、CD80和CD86)。
2、为了制备抗原脉冲DC,用100μg/ml EGFRvIII-PDL1-GMCSF融合蛋白或PBS在补充上述细胞因子的培养基(含有20ng/ml rmGM-CSF和20ng/ml重组小鼠IL-4的RPMI-1640培养基)中培养过夜。次日,再用50μg/ml EGFRvIII-PDL1-GMCSF融合蛋白或PBS再次对树突状细胞进行脉冲处理2h。用PBS洗涤3次后,即得EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗和PBS负载的肿瘤疫苗,用于接下来的实验。
二、肿瘤疫苗对胰腺癌的治疗作用
请参考图6,为使用本实施例中的制备的肿瘤疫苗免疫小鼠的流程图。
将C57BL/6小鼠随机分为2组(每组4只),分别为:
1)、PBS-DCs对照组(即负载PBS的树突状细胞)
2)、EGFRvIII-PDL1-GMCSF-DCs组(即负载EGFRvIII-PDL1-GMCSF融合蛋白的树突状细胞)
第0天和第7天通过足垫分别注射PBS-DCs(100μg/ml,1×106DC细胞/只)和EGFRvIII-PDL1-GMCSF-DCs(100μg/ml,1×106DC细胞/只)。
第二次接种3天后,检测小鼠的机体细胞免疫反应,第二次接种7天后,给小鼠皮下接种小鼠胰腺癌细胞PANC02(2.5×105细胞/只),再观察肿瘤曲线和生存期。
1、EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗诱导小鼠脾脏细胞IL-2和IFN-
γ分泌
为了研究EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗是否能诱导有效的免疫反应,进行了如下实验:
第二次接种3天后,分离免疫小鼠的脾脏,消化成单细胞悬浮液,然后在U形底部96孔板中再次用5μg/ml融合蛋白脉冲树突状细胞6h,细胞表面染色和流式细胞术检测,分析小鼠T细胞增殖情况。所用抗体来自BD Biosciences,包括PE-CY5抗鼠CD4、FITC抗鼠CD8a、PE抗鼠IL-2、APC抗鼠IFN-γ。固定活性染料是从东博生物科学公司购买的。所有数据收集在BD FACSVerse上,并使用Flowjo软件进行分析。
正如预期的那样,与对照相比,注射EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗的小鼠脾细胞中CD4+和CD8+T细胞显著增加(如图7所示)。
用细胞内染色和流式细胞术检测,分析产生IL-2和IFN-γ的CD4+T细胞。如图8和图9所示,T细胞产生IL-2的频率在总CD4+细胞上增加了17倍,产生IFN-γ的频率在总CD4+细胞上增加了15倍。
在CD8+T细胞中,也可以观察到类似的结果,IL-2上调了约27倍,IFN-γ上调约6.8倍(图10和图11)。
这些结果清楚地表明,EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗可促进IFN-γ和IL-2的产生,并可能产生良好的CTL诱导能力。
2、EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗可显著抑制肿瘤生长
为了验证EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗的效力,进行了如下实验:
第二次接种1周后,将稳定表达PD-L1的指数生长的panc02胰腺癌肿瘤细胞系(ATCC)皮下注射到小鼠体内(2.5×105cell/只)。肿瘤细胞接种建立后,每隔5天用卡尺测量一次肿瘤大小,监测肿瘤生长,并计算生长曲线。
肿瘤体积计算如下:(最长直径)×(最短直径)2×0.5
肿瘤生长曲线图如图12所示,从图12可以看出:与PBS-DCs对照组相比,EGFRvIII-PDL1-GMCSF-DCs组能更有效地延缓肿瘤生长。
荷瘤小鼠的存活曲线如图13所示,从图13可以看出:与PBS-DCs对照组相比,EGFRvIII-PDL1-GMCSF-DCs组显著提高了荷瘤小鼠的存活率,50%的治疗小鼠至少存活了80天。
因此,这些数据表明,与传统的非PD-L1 DC靶向蛋白疫苗相比,本发明的EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗可能是一种针对胰腺癌更有效的肿瘤疫苗。
3、EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗抗肿瘤过程中肝肾损伤检测分
析
为了进一步验证EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗在抗肿瘤作用中是否损伤小鼠的肝、肾组织细胞,从免疫组中分离出小鼠肝脏和肾脏,放入异戊烷中,用液氮快速冷冻。组织切片按照标准方案进行。
简单地说,冷冻组织在-20℃下被切割(5μm厚),然后立即转移到保存在干冰上并在-80℃下储存的微型滑盒中。这些滑片被风干,用***固定,然后用石蜡包埋。H&E染色在广州医科大学的病理中心完成。
结果显示,切片分析的H&E染色未在肝脏和肾脏的细胞质中检测到阳性标记(如图14所示),说明本发明的EGFRvIII-PDL1-GMCSF融合蛋白负载的肿瘤疫苗在抗肿瘤过程中未对肝肾造成损伤。
以上所述实施例的各技术特征可以进行任意的组合,为使描述简洁,未对上述实施例中的各个技术特征所有可能的组合都进行描述,然而,只要这些技术特征的组合不存在矛盾,都应当认为是本说明书记载的范围。
以上所述实施例仅表达了本发明的几种实施方式,其描述较为具体和详细,但并不能因此而理解为对发明专利范围的限制。应当指出的是,对于本领域的普通技术人员来说,在不脱离本发明构思的前提下,还可以做出若干变形和改进,这些都属于本发明的保护范围。因此,本发明专利的保护范围应以所附权利要求为准。
序列表
<110> 广州医科大学
<120> EGFRvIII-PDL1-GMCSF肿瘤疫苗及其制备方法和应用
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Ile Asn Ala Thr Asn Ile Lys His Phe Lys Asn Cys Thr Ser Ile Ser
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Claims (9)
1.一种EGFRvIII-PDL1-GMCSF融合蛋白,其特征在于,所述融合蛋白的氨基酸序列如SEQ ID NO.1所示。
2.根据权利要求1所述的EGFRvIII-PDL1-GMCSF融合蛋白,其特征在于,所述融合蛋白是由如SEQ ID NO.2所示的核苷酸序列所编码。
3.权利要求1或2所述的EGFRvIII-PDL1-GMCSF融合蛋白的制备方法,其特征在于,所述方法包括以下步骤:
(1)、合成包含了人EGFRvIII、Th表位、PDL1和GMCSF的基因序列的融合基因片段;所述融合基因片段的核苷酸序列如SEQ ID NO.2所示;
(2)、对步骤(1)的融合基因片段和pET-21a质粒载体进行NdeI和XhoI双酶切,切胶纯化试剂盒回收,连接,得到表达质粒pET-21a/EGFRvIII-PDL1-GMCSF;
(3)、将表达质粒pET-21a/EGFRvIII-PDL1-GMCSF转入BL21(DE3)表达菌株中,经IPTG诱导,获得目的蛋白,经纯化、透析,即得EGFRvIII-PDL1-GMCSF融合蛋白。
4.权利要求1或2所述的EGFRvIII-PDL1-GMCSF融合蛋白在制备肿瘤疫苗中的应用。
5.一种肿瘤疫苗,其特征在于,所述肿瘤疫苗的活性成分为EGFRvIII-PDL1-GMCSF融合蛋白。
6.根据权利要求5所述的肿瘤疫苗,其特征在于,所述肿瘤疫苗的融合蛋白负载量为80μg/ml~120μg/ml。
7.根据权利要求6所述的肿瘤疫苗,其特征在于,所述肿瘤疫苗的融合蛋白负载量为95μg/ml~105μg/ml。
8.权利要求5~7任一项所述的肿瘤疫苗的制备方法,其特征在于,所述方法包括以下步骤:
(1)、在培养有树突状细胞的培养基中,加入80μg/ml~120μg/ml的EGFRvIII-PDL1-GMCSF融合蛋白培养过夜;
(2)、再用45μg/ml~55μg/ml的EGFRvIII-PDL1-GMCSF融合蛋白对树突状细胞进行脉冲处理1h~3h,即得。
9.权利要求5~7任一项所述的肿瘤疫苗在制备治疗胰腺癌的药物中的应用。
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Non-Patent Citations (2)
Title |
---|
A phase II, multicenter trial of rindopepimut (CDX-110) in newly diagnosed glioblastoma: the ACT III study;James Schuster et al.;《Neuro-Oncology》;第854-861页 * |
Discovery of a self-assembly and self-adjuvant lipopeptide as a saccharide-free peptide vaccine targeting EGFRvIII positive cutaneous melanoma;Chen et al.;《Biomaterials Science》;第1-9页 * |
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