CN115029325A - 一种重组溶瘤腺病毒及其应用 - Google Patents
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Abstract
本发明涉及肿瘤生物治疗领域,具体涉及一种重组溶瘤腺病毒及其应用,该重组溶瘤腺病毒的基因组E1和E3区域缺失,缺失区域被稳定地插入胆固醇调控元件和免疫调节元件的外源核酸序列;其中,所述的胆固醇调控元件为包含APOA1基因或其简并序列在内的功能元件;所述的免疫调节元件为包含选自GM‑CSF、IFN‑γ、IL2、IL7、IL12、IL15、IL16、IL18、IL21、IL22、IL27、IL28和IL29基因或其简并序列在内的功能元件;所述的胆固醇调节元件和免疫调节元件的基因以非融合方式表达;所述的胆固醇调节元件和免疫调节元件是以可操作地连接方式连接至外源调控序列,包括启动子序列、增强子序列和PA序列。该重组溶瘤病毒携载的功能元件能够协同稳定肿瘤微环境中的T细胞功能。
Description
技术领域
本发明涉及肿瘤生物治疗领域;具体涉及一种能有效抑制肿瘤生长、侵袭及转移的溶瘤性腺病毒载体的制备,该载体可携带并表达胆固醇代谢调节分子和免疫共刺激分子的基因。
背景技术
一些野生减毒或基因改造的病毒株,因其具备在肿瘤中优势复制的能力,并发挥直接溶瘤作用及诱导免疫活化作用,而被广泛地应用到抗肿瘤治疗领域中。然而,溶瘤病毒(OVs)单独应用的治疗效果不佳。为解决这个技术问题,现阶段主要的技术方案是:1)优化OVs的骨架结构(如感染及复制相关基因)或材料包裹等方式以减少中和抗体对其清除,增加其对肿瘤组织感染、复制及传播的病毒学表现;2)以OVs作为基因治疗的载体,表达功能基因,发挥OVs和功能基因的协同作用(如OVs表达GM-CFS、IL-2、IL-12、IL-15等免疫共刺激分子,或/和PD1、PD-1L、CTLA-4、TIGIT等免疫检查点调节剂);3)OVs联合放疗、化疗及其他免疫疗法等第二疗法(Nat Rev Cancer,2018,18(7):419-432;Curr Opin Biotechnol,2020,65:25-36)。
近年来研究发现,肿瘤微环境(TME)中肿瘤细胞对某些代谢底物的优势竞争,以及代谢产物的排出堆积能够抑制免疫系统正常功能的发挥(Cell,2015,162(6):1229-41)。由此,延伸出一系列新的免疫治疗靶点:代谢-免疫调控轴。特别是以OVs所介导的免疫治疗,活病毒感染肿瘤细胞后,子代病毒的复制及功能基因的表达均加速TME中营养素的消耗和代谢副产物的堆积。这些由肿瘤发展及OVs干预所诱导的代谢环境恶化,削弱了OVs及其携载的免疫调节剂所介导的抗肿瘤免疫应答(Mol Ther,2020,28(6)1417-1421;Front CellInfect Microbiol,2019,9:95)。因此,修正失常的TME中代谢稳态成为OVs疗法亟需解决的问题。
目前,靶向代谢重编程以提高OVs疗效的技术方案主要集中在修正糖代谢TCA循环的失常,如乳酸堆积,OVs联合PDK抑制剂DCA,重组溶瘤痘苗病毒(oVV)表达leptin(oVV-leptin)(Cancer Res,2019,79(15):3824-3836;Immunity,2019,51(3):548-560)。2019年,《Cell Metabolism》杂志研究报道,TME微环境中胆固醇堆积能够诱导抗肿瘤细胞毒性T淋巴细胞(CTL)上的免疫检查点上调,从而加速诱导CTL功能耗竭(Cell Metab,2019,30(1):143-156)。申请者在此研究的基础之上,在溶瘤腺病毒Ad5上创造性地整合了胆固醇代谢调控元件APOA1,获得重组APOA1溶瘤腺病毒Ad5(Ad5-APOA1)。与Ad5-con相比,该Ad5-APOA1通过APOA1修正了TME中的富集的胆固醇,而协同抑制乳腺癌等肿瘤生长和转移(PCT/CN2020/078360)。然而,Ad5-APOA1方案对某些肿瘤中应答不够完全。为了解决这个问题,申请人采取进一步优化技术方案,旨在Ad5-APOA1骨架上进一步整合免疫调节因子,即共表达胆固醇调控元件及免疫调节元件。
发明内容
针对现有技术的不足,本发明的目的是提供一种重组溶瘤腺病毒及其应用。本发明的技术方案免疫应答效应温和,有利于溶瘤细胞在肿瘤细胞内早期的复制和功能基因的表达,而且胆固醇调控元件与免疫调控元件能够产生协同促进长效免疫记忆的形成。
为实现上述发明目的,本发明采用了下列发明内容:
一种重组溶瘤腺病毒,其基因组E1和E3区域缺失,缺失区域被稳定地插入胆固醇调控元件和免疫调节元件的外源核酸序列;其中,
所述的胆固醇调控元件为包含APOA1基因或其简并序列在内的功能元件;
所述的免疫调节元件为包含选自GM-CSF、IFN-γ、IL2、IL7、IL12、IL15、IL16、IL18、IL21、IL22、IL27、IL28和IL29基因或其简并序列在内的功能元件;
所述的胆固醇调节元件和免疫调节元件的基因以非融合方式表达;
所述的胆固醇调节元件和免疫调节元件是以可操作地连接方式连接至外源调控序列,包括启动子序列、增强子序列和PA序列。
作为本申请的优选技术方案,所述腺病毒选自C血清型亚群,包括人2型和5型腺病毒。
作为本申请的优选技术方案,所述启动子序列选自组成型、组织特异型或诱导型启动子;优选的,所述组成型启动子选自CMV、SV40或EF1α启动子。
作为本申请的优选技术方案,所述胆固醇调节元件和免疫调节元件为人源化的。
优选的,其腺病毒为人5型腺病毒,所述胆固醇调节元件为APOA1基因,所述免疫调节元件为IL15基因。
作为本申请的优选技术方案,所述胆固醇调节元件APOA1的核酸序列选自(a)或(b):
(a)如SEQ ID NO:1所示;
(b)与SEQ ID NO:1具有60%、65%、70%、75%、80%、85%、90%、95%、98%或99%的同源性的核酸序列。
作为本申请的优选技术方案,所述免疫调节元件IL15的核酸序列选自(c)或(d):
(c)如SEQ ID NO:2所示;
(d)与SEQ ID NO:2具有60%、65%、70%、75%、80%、85%、90%、95%、98%或99%的同源性的核酸序列。
一种重组溶瘤Ad5,其包含修饰的Ad5基因组;该修饰包含(i)缺失野生型Ad5基因组的E1,E3区序列,但包括早期复制所需的E1A序列;(ii)编码胆固醇代谢调节剂和/或免疫调节剂的外源性核酸序列,其中所述的外源性核酸序列被稳定地加入到至少是所述经修饰的Ad5基因组的被缺失区域。
优选的,所述的编码胆固醇代谢调节剂的外源性核酸序列如SEQ ID NO:1所示;所述的编码免疫调节剂的外源性核酸序列如SEQ ID NO:2所示。
申请人在Ad5-APOA1骨架上进一步整合温和型免疫调节因子(如IL-15等),即共表达APOA1及IL-15的新型溶瘤Ad5-APOA1-IL15。第一,该技术方案免疫应答效应温和,有利于OVs在肿瘤细胞内早期的复制和功能基因的表达;第二,APOA1与IL-15能产生协同促进长效免疫记忆的形成;第三,与武装IL-2、IL-6、GM-CFS或免疫检查点抑制剂等短期强效的OVs技术方案相比,本方案长期温和的免疫活化效应能够适用于某些重要功能脏器肿瘤的免疫治疗(如肝脏、胰腺、颅内等)。
本发明还保护前文任一所述的重组溶瘤腺病毒、前文所述的重组溶瘤Ad5在制备治疗或缓解癌症相关疾病的药物中的应用。
作为本申请的优选技术方案,本发明还保护前文任一所述的重组溶瘤腺病毒、前文所述的重组溶瘤Ad5,与第二治疗药物在制备治疗或缓解癌症相关疾病的药物中的应用。
更优选的,所述第二治疗药物为PD1抗体。
作为本申请的优选技术方案,所述癌症选自胆囊癌、基地细胞瘤、肝外胆管癌、结肠癌、子宫内膜癌、子宫肌瘤、食道癌、尤因肉瘤、前列腺癌、胃癌、肝癌、肝细胞癌、霍奇金淋巴瘤、喉癌、肺癌、黑色素瘤、间皮瘤、胰腺癌、直肠癌、肾癌、甲状腺癌、神经胶质瘤、恶性周围神经细胞肿瘤、恶性外周神经鞘瘤、皮肤和丛状神经纤维瘤、平滑肌瘤、平滑肌肉瘤、纤维瘤、乳头状腺瘤、甲状腺未分化癌、甲状腺髓样癌、甲状腺滤泡癌、hurthle细胞癌、甲状腺癌、腹水、恶性腹水、唾液腺肿瘤、唾液腺粘液表皮样癌、唾液腺腺泡细胞癌、胃肠道基质肿瘤、导致身体潜在空间积液的肿瘤、胸腔积液、心包积液、腹膜积液、巨细胞瘤、骨巨细胞瘤、色素沉着绒毛结节性滑膜炎、腱鞘巨细胞瘤和其他肉瘤中的任一种。
优选的,所述癌症选自神经胶质瘤、胰腺癌或肝癌。
本发明还请求保护一种药物组合物,其含有前文任意一项所述的重组溶瘤腺病毒、前文所述的重组溶瘤Ad5,以及药学上可接受的载体或赋形剂。
优选的,该组合物可被用于配制成瘤内注射制剂。
本发明还涉及一种治疗癌症的方法,所述方法包括向适用对象注射有效量的本发明所述的重组溶瘤Ad5或所述药物组合物。
此外,本发明还涉及所述重组溶瘤Ad5在治疗癌症的方法中的应用。
本发明还涉及前文所述的重组溶瘤Ad5或药物组合物在制备治疗抗癌药物中的应用。
有益效果
本发明提供的一种重组溶瘤腺病毒及其应用,与现有技术相比,具有如下有益效果:
(1)本发明的技术方案免疫应答效应温和,有利于溶瘤细胞在肿瘤细胞内早期的复制和功能基因的表达;
(2)本发明胆固醇调控元件与免疫调控元件能够产生协同促进长效免疫记忆的形成;
3)与武装IL-2、IL-6、GM-CFS或免疫检查点抑制剂等短期强效的OVs技术方案相比,本方案长期温和的免疫活化效应能够适用于某些重要功能脏器肿瘤的免疫治疗(如肝脏、胰腺、颅内等)。
附图说明
本发明的各个方面及优点可从下面参考附图的详细描述中获得。
图1.重组溶瘤Ad5病毒的示意图。实施例中以腺病毒E1区域插入目的基因序列,表达人和鼠的APOA1、GM-CSF、IL7、IL15功能元件。
图2.重组溶瘤Ad5病毒APOA1、GM-CSF、IL7和IL15的表达。ELISA检测重组溶瘤Ad5病毒感染293T细胞后上清中目的蛋白的表达情况,****表示P<0.0001。
图3.体外溶瘤试验。实例重组溶瘤Ad-APOA1-IL15感染人和鼠肿瘤细胞系后的溶瘤情况,MOI=0、1、10、100。
图4.Ad5-APOA1处理后肿瘤组织内胆固醇含量检测试验。检测肿瘤组织在经过Ad5-APOA1处理后组织中胆固醇富集情况,*表示P<0.05,**表示P<0.01。
图5.Ad5-GM-CSF、Ad5-IL7和Ad5-IL15介导的免疫活化试验。ELISA法检测处理后上清中IFN-γ和TNF-α的表达情况反映免疫调控元件分子的免疫活化功能,**表示P<0.01,***表示P<0.001。
图6.Ad5-APOA1联合Ad5-GM-CSF、Ad5-IL7或Ad5-IL15介导的免疫活化试验。体外模拟APOA1修正胆固醇富集对CTL的耗竭作用,恢复免疫调节因子活化CTL细胞的效应,*表示P<0.05,**表示P<0.01,NS表示没有统计学意义。
图7.GL261模型体内抗肿瘤活性试验。原位GL261模型测试重组溶瘤病毒Ad5-mAPOA1和Ad5-mGM-CSF或Ad5-mIL15之间的联合作用,CR表示完全治愈,每组5只C57bl/6小鼠,病毒以2×108PFU剂量在肿瘤移植第7天,瘤内注射1次。GL261皮下瘤模型,病毒以2×108PFU剂量在肿瘤体积>50mm3时,瘤内隔天注射,连续3次,验证Ad5-mAPOA1-mIL15的抗肿瘤作用。
图8.H22模型体内抗肿瘤活性试验。H22皮下瘤模型,验证Ad5-mAPOA1-mIL15的抗肿瘤作用,病毒以2×108PFU剂量在肿瘤体积>50mm3时,瘤内隔天注射,连续3次,*表示P<0.05,**表示P<0.01。
图9.Panc02模型体内抗肿瘤活性试验。Panc02皮下瘤模型,验证Ad5-mAPOA1-mIL15的抗肿瘤作用,病毒以2×108PFU剂量在肿瘤体积>50mm3时,瘤内隔天注射,连续3次,*表示P<0.05,NS表示没有统计学意义。
图10.抗肿瘤rechallenge免疫记忆试验。GL261治愈小鼠rechallenge后7d测T细胞免疫记忆强度,CD44+CD62L+表示中央记忆T细胞,*表示P<0.05,**表示P<0.01,***表示P<0.001。
具体实施方式
发明详述
术语“一”或“一个”实体是指该实体中的一个或多个;例如,“重组溶瘤Ad5”可被理解为表示一个或多个重组溶瘤Ad5病毒。因此,术语“一个”、“一个或多个”和“至少一个”可以在本文中互换使用。
“同源性”或“同一性”或“相似性”是指两个肽之间或两个核酸之间的序列相似性。
一个多核苷酸或多核苷酸区域与另一个序列具有特定百分数,例如60%、65%、70%、75%、80%、85%、90%、95%、98%或99%的“序列同一性”是指,当序列对比时,两个序列该百分比的碱基(或氨基酸)是相同的。该对比和百分比同源性或序列同一性可以使用本领域已知的软件来确定。
术语“癌症”或“肿瘤”是指可根据本公开内容治疗并涉及异常细胞生长的一组疾病,其可能侵入或扩散至身体。并非所有的肿瘤都是癌性的;良性肿瘤不会侵袭转移到其他身体部位。本公开内容优选适用于实体瘤。肿瘤或癌症的非限制性实施例包括胆囊癌、基地细胞瘤、肝外胆管癌、结肠癌、子宫内膜癌、子宫肌瘤、食道癌、尤因肉瘤、前列腺癌、胃癌、肝癌、肝细胞癌、霍奇金淋巴瘤、喉癌、肺癌、黑色素瘤、间皮瘤、胰腺癌、直肠癌、肾癌、甲状腺癌、神经胶质瘤、恶性周围神经细胞肿瘤、恶性外周神经鞘瘤(MPNST)、皮肤和丛状神经纤维瘤、平滑肌瘤、平滑肌肉瘤、纤维瘤、乳头状腺瘤、甲状腺未分化癌、甲状腺髓样癌、甲状腺滤泡癌、hurthle细胞癌、甲状腺癌、腹水、恶性腹水、唾液腺肿瘤、唾液腺粘液表皮样癌、唾液腺腺泡细胞癌、胃肠道基质肿瘤(GIST)、导致身体潜在空间积液的肿瘤、胸腔积液、心包积液、腹膜积液、巨细胞瘤(GCT)、骨GCT、色素沉着绒毛结节性滑膜炎(PVNS)、腱鞘巨细胞瘤(TGCT)和其他肉瘤。在优选的实施方式中,本发明用于制备治疗神经胶质瘤、胰腺癌、肝癌的药物中。
术语“治疗”是指治疗性治疗和预防性措施,目的是预防或减缓(减轻)不正常的生理变化或紊乱,例如癌症的进展。有益的或期望的临床结果包括,但不限于,缓解症状、减少疾病程度、稳定(即不恶化)疾病状态、延缓或减缓疾病进展、改善或缓解疾病状态、以及症状消失(部分或全部)、无论是可检测的还是无法检测的。“治疗”也意味着与未接受治疗时所预期的生存期相比延长生存期。需要治疗的病人包括那些已经患有疾病或病症的人、以及那些容易患有疾病或病症的人,或那些预防疾病或病症的人。
术语“对象”、“个体”、“动物”、“患者”或“哺乳动物”是指期望进行诊断、预后或治疗的任何对象,特别是哺乳动物对象。哺乳动物对象包括人、家畜、农场动物、动物园动物、竞技动物或宠物,如狗、猫、豚鼠、兔、大鼠、小鼠、马、牛等。
本领域普通技术人员还应该看、理解,可以修饰如本文所公开的修饰的基因组,使得它们在核苷酸序列上与它们所衍生出的修饰的多核苷酸不同。例如,从指定的DNA序列衍生的多核苷酸或核苷酸序列可以是相似的,例如与起始序列具有一定的百分比同一性,例如它可以与起始序列60%、70%、75%、80%、85%、90%、95%、98%或99%相同。
此外,可以进行核苷酸或氨基酸取代、缺失或插入,以在“非必需”区域进行保守取代或改变。例如,衍生自指定蛋白质的多肽或氨基酸序列,除了一个或多个单独的氨基酸取代、插入或缺失(例如1、2、3、4、5、6、7、8、9、10、15、20或更多个氨基酸取代、插入或缺失)之外,其余部分可以与起始序列相同。在某些实施方案中,衍生自指定蛋白的多肽或氨基酸序列相对于起始序列具有1至5个、1至10个、1至15个、1至20个单独氨基酸取代、插入或缺失。
术语“调节元件”旨在包括启动子、增强子和其他表达调控元件。启动子是指导RNA聚合酶结合DNA并启动RNA合成的DNA序列。一个强大的启动子能够引起mRNA的高频率启动。用于在真核细胞中加工的合适元件是聚腺苷酸化信号。调节元件序列包括在许多细胞中指导核苷酸序列表达的调节元件序列(如,组织特异性调节元件序列)。调节元件也包括绝缘子(insulator),其包括在细胞染色体上发现的一类DNA元件,其保护染色体的一个区域中的基因免受另一个区域的调节影响,如CTCF基序。
本领域普通技术人员可以基于例如期望的组织特异性和表达水平来选择合适的调节元件。例如,细胞类型特异性或肿瘤特异性启动子可用于将基因产物的表达限制于特定的细胞类型。可用于本技术的组织特异性启动子的实例包括前列腺特异性抗原(PSA)启动子,其对前列腺细胞是特异性的;肌间线蛋白启动子,其对肌细胞是特异性的;烯醇化酶启动子,其对神经元是特异性的;β球蛋白启动子,其对红细胞是特异性的;tau-globin启动子,其对红细胞也是特异性的;生长激素启动子,其对垂体细胞是特异性的;胰岛素启动子,其对胰腺β细胞具有特异性;胶质纤维酸性蛋白启动子,其对星形胶质细胞是特异性的;酪氨酸羟化酶启动子,其对儿茶酚胺能神经元是特异性的;淀粉样前体蛋白启动子,其对神经元是特异性的;多巴胺β-羟化酶启动子,其对去甲肾上腺素能和肾上腺素能神经元是特异性的;色氨酸羟化酶启动子,对5-羟色胺/松果体细胞是特异性的;reg(胰结石蛋白)启动子,其对结肠和直肠肿瘤以及胰腺和肾细胞是特异性的;和甲状旁腺激素相关肽(PTHrP)启动子,其对肝和盲肠肿瘤以及神经鞘瘤、肾细胞、胰腺细胞和肾上腺细胞是特异性的。
在肿瘤细胞中特异性起作用的启动子的实施例包括对乳腺癌细胞特异的基质溶素3启动子;对非小细胞肺癌细胞特异的表面活性蛋白A启动子;表达SLPI的癌特异性的分泌性白细胞蛋白酶抑制剂(SLPI)启动子;对黑色素细胞特异的酪氨酸酶启动子;对纤维肉瘤/致瘤细胞特异的应激诱导的grp78/Bip启动子;对脂肪细胞特异的AP2脂肪增强子;对肝细胞特异的α-1抗胰蛋白酶转甲状腺素蛋白启动子;对多形性成胶质细胞瘤特异的白细胞介素-10启动子;对胰腺、乳腺、胃、卵巢和非小细胞肺细胞特异的c-erbB-2/3/4启动子;对脑肿瘤细胞的特异的α-B-晶状体蛋白/热休克蛋白27启动子;对神经胶质瘤和脑膜瘤细胞特异的碱性成纤维细胞生长因子启动子;对鳞状细胞癌、神经胶质瘤和乳腺肿瘤细胞具有特异的表皮生长因子受体启动子;对乳腺癌细胞具有特异的粘蛋白样糖蛋白(DF3、MUC1)启动子;对于转移性肿瘤特异的mts1启动子;对甲状腺癌细胞特异的甲状腺球蛋白启动子;对肝癌细胞特异的甲胎蛋白(AFP)启动子;对胃癌细胞特异的villin启动子;和对肝癌细胞特异的白蛋白启动子。在优选的实施方式中,本发明选择神经胶质瘤、肝癌、胰腺癌组织特异性启动子。
除了使用组织特异性启动子之外,病毒的局部施用可实现局部的表达和效应。可以使用的非组织特异性启动子,如早期巨细胞病毒(CMV)启动子,人延长因子1α(EF1α)启动子。例如,在一些实施例中,外源性的核苷酸序列可操作地连接至启动子,如CMV或EF1α启动子。
术语“APOA1”在NCBI中基因ID:335。该基因编码载脂蛋白A-I,是血浆中高密度脂蛋白(HDL)的主要蛋白组分。编码的前体蛋白经蛋白水解处理,生成成熟蛋白,促进胆固醇从组织外排到肝脏排泄,是卵磷脂胆固醇酰基转移酶(LCAT)的辅助因子,LCAT是一种负责大多数血浆胆固醇酯形成的酶。该基因与11号染色体上的另外两个载脂蛋白基因密切相关。该基因的缺陷与高密度脂蛋白缺乏有关,包括丹吉尔病,以及系统性非神经性淀粉样变性。选择性剪接导致多个转录本变异,其中至少有一个编码一个前蛋白。APOA1基因有多个转录本,分别为NM_000039.2/NP_000030.1(cDNA序列/蛋白序列),NM_001318017.2/NP_001304946.1,NM_001318018.2/NP_001304947.1,NM_001318021.1/NP_001304950.1。
术语“IL15”在NCBI中基因ID:3600。该基因编码白细胞介素15是一种多效性细胞因子,具有激活T细胞、B细胞和NK细胞,并可介导这些细胞的增殖和存活的功能。此外,IL15能激活、维持和扩增CD8+记忆性T细胞,而不激活调节性T淋巴细胞(Tregs,具有免疫抑制功能)。IL15的作用主要是通过与表达在抗原提呈细胞上的高亲和力α受体(IL15Rα)结合,以此将IL15递呈给IL2/15Rβγ二聚体形成三元复合物来激活胞内JAK和STAT型号通路,最终促进靶细胞增殖与活化、IFN-γ、TNF-α分泌水平提升,以及Bcl-2、Bcl-XL(抗凋亡蛋白)的上调、Bim、Puma(促凋亡蛋白)的下调--减弱凋亡信号。
术语“E1A”。病毒的E1区基因可以进一步分为E1A和E1B。E1A主要由两种成分构成,分别为289R(或13S)和243R(或12S)。腺病毒基因组进入细胞核后,细胞转录因子首先与E1A区上游的增强子结合,表达E1A蛋白,该蛋白的作用是调节细胞代谢,使病毒DNA更易于在细胞中复制。E1A蛋白还可以激活其他早期基因(E1B、E2A、E2B、E3和E4)的启动子,其中E2B驱动另外三个与病毒复制有关的早期基因转录单位末端蛋白前体(pTP,precursor terminalprotein)、单链DNA结合蛋白(ssDBP,single-stranded DNA binding proteins)以及DNA聚合酶(DNA pol,DNA polymerase)的表达,这三个基因的表达产物紧密地结合成一个复合物,与至少三种细胞蛋白相互作用,启动病毒基因组的复制。本发明中E1A前包含一个独立的CMV启动子。GFP和E1A中间由2A连接序列连接。
不拘于理论限制,作为连接序列的2A本身只有翻译后蛋白剪切位点,2A肽段将保留在2A前的蛋白后。使用2A连接GFP和E1A,使得早起复制元件和GFP标签蛋白分离。
不拘于理论限制,在腺病毒血清型选择方面已发现100余个血清型,其中人腺病毒有52种,分为A、B、C、D、E和F六个亚群(subgroup),它们对宿主细胞的亲嗜性、致瘤性以及疾病史各不相同。基因治疗常用人2型及5型腺病毒在血清学上均属于C亚群,在DNA序列上有95%的同源性。腺病毒感染细胞的过程是从腺病毒纤毛的头节区粘附到细胞表面的特异性受体开始的。因为人腺病毒主要与柯萨奇B病毒共用一种受体,因此这种受体被称为柯萨奇/腺病毒受体即CAR(coxsackie/adenovirus receptor)。低水平表达CAR的无疑制约腺病毒的转导效率。然而,由于C亚群腺病毒载体已经被成功应用临床,其安全性已在人体中广泛测试,作为载体安全性高。本发明重组溶瘤腺病毒能在肿瘤细胞内复制。
本发明所述的药物,应用包括预防制品,治疗制品,诊断制品三个方面。优选地,应用为治疗制品。治疗为单独治疗,辅助治疗或者联合治疗。
本发明所述的药物的给药途径,包括但不限于口服、直肠、透黏膜、经肠给药,或者局部、经皮、吸入、肠胃外、舌下、阴道内、鼻内、眼内、腹膜内、肌内、皮下、静脉内、肿瘤原位给药。优选的给药途径是静脉注射、肿瘤原位给药。
重组人5型腺病毒
根据本发明任一项所述的复制型溶瘤腺病毒载体,所述的溶瘤腺病毒包括A、B、C、D、E和F亚类,优选C亚类腺病毒;更优选地,人5型腺病毒Ad5、重组溶瘤Ad5病毒。
本发明提供一种能够分泌胆固醇调控元件以及免疫调控元件的重组溶瘤腺病毒。具体地,该复制型溶瘤腺病毒载体的构建方案包括以下步骤:在E1和E3区域缺失的腺病毒骨架载体上插入本发明目的序列。目的序列可选择插入在E1或E3区域其中之一或分配到E1和E3区域;第一组成型启动子CMV控制腺病毒早期复制元件(E1A)和检测标签(EGFP);第二组成启动子EF1α控制的胆固醇调控元件(APOA1)和免疫调控元件(如IL-15)。例如,本发明第一组成启动子和第二组成启动子调控序列可共同分配至E1区或E3区,也可分别分配至E1或E3区,如第一组成启动子分配至E1区,第二组成启动子调控区分配至E3区。E1或E3区内第一组成启动子调控序列和/或第二组成启动子调控序列内部和结构之间组合不分先后,包括2种第一及第二调控序列前后的组合,2种启动子变化组合,2种启动子后功能序列的变化组合。例如,第一及第二调控区同在E1区域的CMV-E1A-EGFP/EF1α-APOA1-IL15,也可替换成CMV-EGFP-E1A/EF1α-IL15-APOA1这类启动子后功能区的变换方式,EF1α-E1A-EGFP/CMV-EGFP-E1A这类启动子替换方式,或EF1α-APOA1-IL15/CMV-E1A-EGFP这类两个功能区的替换方式等。第一组成启动子序列调控区域和第二组成启动子调控区域接受反向表达或背靠表达的基因操纵方式。
第一和第二启动子,除常见组成型启动子(如CMV、EF1α、SV40等)外,可替换成本领域内常见的条件特异性启动子或组织特异性启动子。第一和第二启动子之间的互换不包括相同类型的启动子。
理论上,用于检测病毒滴度的标签序列(如EGFP)和E1A在同一个启动子的控制之下,实验用途的重组溶瘤Ad5包括检测标签序列,药物用途的重组溶瘤Ad5的EGFP标签序列可被替换成其他示踪标签或删除。实验用途的标签序列但不限于本领域内常用的其他化学发光、自发荧光或His等标签序列。
E1A和EGFP或APOA1和IL15中间由PA2或IRES2等类似功能序列连接。
在一个实施例中,重组溶瘤Ad5包含编码胆固醇代谢调控的外源核酸序列,所述的胆固醇代谢调控序列选自人APOA1序列(在后续实施例中,前缀带有h,为人源)。在另一个实施例中,所述的胆固醇代谢调控序列选自鼠APOA1序列(在后续实施例中,前缀带有m,为鼠源)。
在一个实施例中,重组溶瘤Ad5包含编码免疫调控的外源核酸序列,所述的免疫调控序列选自GM-CSF、IFN-γ、IL2、IL7、IL12、IL15、IL16、IL18、IL21、IL22、IL27、IL28和IL29等细胞因子或MIP-1、MCP-1等趋化因子。在优选实施例中,所述的免疫调控序列选自IL7或IL15。在更优选的实施例中,所述的免疫调控序列是人IL15或人源化IL15(在后续实施例中,前缀带有h,为人源)。在优选实施例中,所述的免疫调控序列是鼠IL15(在后续实施例中,前缀带有m,为鼠源)。
在一个实施例中,重组溶瘤Ad5包含编码胆固醇代谢调控序列和免疫调控的外源核酸序列。例如,在一个实施例中胆固醇代谢调控序列为APOA1,免疫调控序列为IL15。所引入的外源序列,优选插入E1区域,并置于E1A或E1A-EGFP序列之后。例如,在一个实施例中,重组溶瘤Ad5为Ad5-△E1(CMV-E1A-EGFP/EF1α-APOA1-IL15);次选插入到E3区域,例如Ad5-△E1(CMV-E1A-EGFP)-△E3(EF1α-APOA1-IL15)。
可以理解的是,将一个或多个外源核酸序列被稳定地整合到修饰的Ad5基因组中并不干扰天然Ad5基因(Ad5-con)的表达,并且外源核酸序列被稳定地整合到修饰的Ad5基因组,使得可以预期外源核酸序列的功能性表达。
编码胆固醇代谢调节元件和/或免疫调节元件的重组基因含有编码蛋白质的核酸以及用于蛋白表达的调节元件。通常,存在于重组基因中调节元件与待表达的核酸序列可操作地连接,并且基于待用于表达的宿主细胞来选择,可包括转录启动子、核糖体结合位点和终止子。在重组表达载体内,“可操作地连接”旨在表示感兴趣的核酸序列以允许核苷酸序列表达(例如在体外转录/翻译系统中表达,或当病毒被引入宿主细胞时在宿主细胞中表达)的方式与调节元件序列连接。
溶瘤病毒疗法最主要是通过病毒诱导的DAMP模式分子和PAMP模式分子来突破瘤内“冷”的TME,增加抗肿瘤效应T细胞的浸润、增殖和分化。肿瘤间质内富集的胆固醇使得CTL细胞内的ER stress上调而诱导PD1等免疫检查点上调,使其表现为耗竭表型。清除瘤内富集的胆固醇有助于延长CTL和肿瘤周围抗原呈递细胞上MHC-I-抗原表位复合物的瞬时接触作用时向,增加IFN-γ、TNF-α和Gzm-B等细胞因子的释放。
组合物
OVs可以在合适的药学上可接受的载体或赋形剂中制备。在通常的储存和使用条件下,这些制剂含有防腐剂以防止微生物的生长。适用于注射使用的药物形式包括无菌水溶液或分散液和用于临时制备无菌注射溶液或分散液的无菌粉末。在所有情况下,制剂必须是无菌的,并且必须是流体以便容易注射。在生产和贮存条件下必须稳定,并且防止细菌和真菌等微生物的污染。
载体可以是包含例如水、乙醇、多元醇(例如甘油、丙二醇和液体聚乙二醇等)、其合适的混合物和/或植物油的溶剂或分散介质。例如通过使用诸如卵磷脂的包衣,通过在分散的情况下维持所需的粒度和通过使用表面活性剂来保持适当的流动性。可以通过各种抗菌剂和抗真菌剂,例如对羟基苯甲酸酯、氯丁醇、苯酚、山梨酸等来预防微生物的作用。在许多情况下,优选包括等渗剂,例如糖或氯化钠。可以通过在组合物中使用延长吸收的试剂(例如单硬脂酸铝或明胶)来延长可注射组合物的吸收。
对于在水溶液中的胃肠道外给药,例如,溶液应适当地缓冲(如果需要的话),并且首先用足够的盐水或葡萄糖使液体稀释剂等渗。这些特定的水溶液特别适用于静脉内、肌内、皮下、肿瘤内和腹膜内给药。就此而言,根据本发明公开的内容,可以使用的无菌水性介质将是本领域技术人员已知的。例如,可以将一个剂量溶解在1mL的等渗NaCl溶液中,并且将其添加到1000mL的皮下灌注液中,或者在所建议的输注位置注射。根据所治疗的受试者的状况剂量必然会发生一些变化。无论如何,负责给药的人员将确定个体受试者的适当剂量。此外,对于人体给药,制剂应答满足FDA生物制品标准所要求的无菌性、无热原性、一般安全性和纯度要求。
通过将活性化合物以需要的量与上面举例的各种其他成分按需要合并在合适的溶剂中,然后过滤灭菌水来制备无菌可注射溶液。通常,通过将各种灭菌的活性成分掺入含有基本分散介质和来自上述举例的所需其他成分的无菌载体中来制备分散液。在用于制备无菌注射溶液的无菌粉末的情况下,优选的制备方法是真空干燥和冷冻干燥技术,其从无菌过滤溶液中生产出包含活性成分及任何另外的所需成分的粉末。
本文所用,“载体”包括任何和所有溶剂、分散介质、载体、包衣、稀释剂、抗菌剂和抗真菌剂、等渗剂和吸收延迟剂、缓冲剂、载体溶液、混悬剂、胶体等。用于药物活性物质的样的介质和药剂在本领域是公知的。除了与活性成分不相容的任何常规的介质或试剂之外,预期其他介质或试剂可以用于所述治疗组合物中。补充的活性成分也可以掺入组合物中。
短语“药学上可接受的”是指当施用于人时不产生过敏或类似的不良反应的分子实体和成分。含有蛋白质作为活性成分的水性组合物的制备在本领域中是充分理解的。典型地,这样的组合物被制备成注射剂,无论作为液体溶液或混悬剂;也可以制备适用于在注射之前溶解或悬浮于液体中的固定形式。
疗法
本发明公开了一种重组溶瘤Ad5病毒。另一方面,也公开提供了用其治疗或缓解癌症的方法,其包括向有需要的受试者施用有效量的重组溶瘤Ad5病毒或包含上述的重组溶瘤Ad5病毒的药物组合物。
一方面公开了重组溶瘤Ad5在制备治疗或缓解癌症相关疾病的药物中的应用。另一方面,公开了一种所述的重组溶瘤Ad5的药物组合物。
在一些实施例中,所述的重组溶瘤Ad5病毒或药物组合物以瘤内方式施用。在一些实施例中,将重组溶瘤Ad5病毒或药物组合物以可注射溶液的形式直接注射到肿瘤内。
在一些实施例中,可将携载胆固醇调控元件和/或免疫调控元件基因的重组溶瘤Ad5病毒与其他癌症治疗剂组合。例如,癌症的治疗可用重组溶瘤Ad5病毒和其他第二疗法来实施,如手术、PD1抗体、CAR-T或放化疗等。该过程可能涉及使细胞与表达构建题和试剂或多种因子同时接触。这可以通过使细胞与包含两种药剂额单一组合物或药理学制剂接触,或通过使细胞与两种不同的组合物或制剂同时接触来实现,其中一种组合物包含表达构建体,而另一种包含第二试剂。
可以在其他药物治疗之前或之后几分钟到几周的时间间隔内进行病毒治疗。在将其他药剂和OVs分别适用于细胞的实施例中,通常将确保在每次递送时间之间不间隔相当长的一段时间,以使得药剂和病毒仍然能够有利地施加对细胞的联合作用。在这样的情况下,预期可以在彼此间隔12-24小时之内使细胞与两种疗法接触。但是,在某些情况下,当施用间隔几天到几周的时间时,可能需要显著延长治疗的时间。
重组溶瘤Ad5病毒的构建
本发明所使用的重组溶瘤Ad5病毒是在ViraPowerTMAdenoviral系统的基础上进行改造。将病毒复制的关键元件E1A基因插入到非复制型腺病毒载体,从而恢复重组溶瘤腺病毒的复制功能。复制型重组溶瘤Ad5病毒的构建分为5个步骤:1)构建连接目的基因的穿梭质粒pShuttle;2)穿梭质粒与病毒载体同源重组;3)溶瘤腺病毒的拯救;4)病毒的扩增与纯化;5)病毒的滴度检测。
全合成CMV-E1A-P2A-EGFP-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-con;
全合成CMV-E1A-P2A-EGFP-EF1α-hAPOA1-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-hAPOA1;
全合成CMV-E1A-P2A-EGFP-EF1α-mAPOA1-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-mAPOA1;
全合成CMV-E1A-P2A-EGFP-EF1α-mGM-CSF-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-mGM-CSF;
全合成CMV-E1A-P2A-EGFP-EF1α-mIL7-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-mIL7;
全合成CMV-E1A-P2A-EGFP-EF1α-hIL15-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-hIL15;
全合成CMV-E1A-P2A-EGFP-EF1α-mIL15-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-mIL15;
全合成CMV-E1A-P2A-EGFP-EF1α-hAPOA1-IRES2-hIL15-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-hAPOA1-hIL15;
全合成CMV-E1A-P2A-EGFP-EF1α-mAPOA1-IRES2-mIL15-BGH polyA基因片段连接到pShuttle(pENTER/D-TOPO)上的attL1和attL2之间构成Ad5-mAPOA1-mIL15。
DH5α感受态细胞转化,挑选单克隆,质粒小提(菌种保存),PCR鉴定质粒DNA(E1A)。
使用LR Clonase II enzyme mix重组试剂盒将pShuttle质粒整合到pAd/PL-DEST腺病毒骨架基因组载体上。DH5α感受态细胞转化,挑选单克隆,质粒小提(菌种保存),PacI酶切PCR鉴定质粒DNA片段长度,测序鉴定序列正确。
正确重组的质粒进行质粒大提,PacI酶切线性化病毒基因组、转染293T细胞完成病毒的包装、扩增和纯化。各重组溶瘤Ad5病毒结构如图1所示,hAPOA1核酸序列如SEQ IDNO:1所示,hIL15核酸序列如SEQ ID NO:2所示。
体外实验
1)体外重组溶瘤Ad5病毒APOA1、GM-CSF、IL7和IL15的表达
本发明所构建的重组溶瘤Ad5病毒以MOI值=10感染2×106个293T细胞,48h后收集细胞培养上清ELISA检测APOA1、GM-CSF、IL7和IL15的表达情况。结果如图2所示,各构建的重组溶瘤Ad5病毒均能表达相应的功能蛋白。
2)体外溶瘤实验
96孔板中培养人胶质瘤细胞系U87、人胶质瘤细胞系U251、人肝癌细胞系HepG2、人胰腺癌细胞系PANC1、鼠胶质瘤细胞系GL261、鼠肝癌细胞系H22、鼠胰腺癌细胞系Panc02、鼠结肠癌细胞系MC38,每孔接种5×103个细胞,隔夜待细胞贴壁后没空接种不同MOI的重组溶瘤Ad5-hAPOA1-hIL15或Ad5-mAPOA1-mIL15病毒,MOI值=0、1、10、100。在感染24h、48h和72h后,CCK8法分别检测细胞活力水平,结果如图3所示。
功能实验
1)APOA1促进肿瘤组织中胆固醇富集减少的功能实验
取皮下接种GL261、H22和Panc02肿瘤细胞株,待肿瘤体积达到100mm3以上,瘤内分别注射PBS、1×108PFU的Ad5-con和Ad5-mAPOA1。2d后取各组肿瘤组织,10mg肿瘤组织进行裂解,并提取总胆固醇,Amplex Red(invitrogen)检测各组胆固醇含量。结果如图4所示,携载APOA1的重组溶瘤Ad5病毒,能够减少肿瘤组织中的胆固醇含量。
2)GM-CSF、IL7和IL15活化免疫细胞的功能实验
GL261细胞以每孔5×104个细胞数量接种于24孔板,待细胞贴壁后Ad5-con病毒和分别携载mGM-CSF、mIL7和mIL15的重组溶瘤Ad5病毒以MOI=10感染细胞24h,取免疫正常小鼠脾脏,研磨,过滤取淋巴细胞与感染肿瘤细胞1:20共培养,48h后取上清ELISA检测IFN-γ、TNF-α的表达,结果如图5所示。
3)胆固醇富集环境中APOA1联合GM-CSF、IL7和IL15活化免疫细胞的功能实验
0.75ug/ml胆固醇培养基条件下,单独携载mGM-CSF、mIL7和mIL15的重组溶瘤Ad5病毒(MOI=10)或1:1联合重组溶瘤Ad5-mAPOA1病毒(MOI=5:MOI=5)感染GL261细胞24h,再取免疫正常小鼠脾脏,研磨,过滤取淋巴细胞与感染肿瘤细胞1:20共培养,48h后取上清ELISA检测IFN-γ、TNF-α的表达,结果如图6所示。
体内实验
1)重组溶瘤Ad5病毒活体抗肿瘤实验
GL261、H22和Panc02细胞皮下和颅内移植瘤用于评估重组溶瘤Ad5病毒的抗肿瘤活性。GL261采用颅内移植瘤方式,评估的重组溶瘤Ad5病毒涉及Ad5-con、Ad5-mAPOA1、Ad5-mIL15、Ad5-mGM-CSF、Ad5-mAPOA1+Ad5-mIL15联合和Ad5-mAPOA1+Ad5-mGM-CSF联合;GL261采用皮下移植瘤方式,评估Ad5-mAPOA1-mIL15的体内抗肿瘤活性,结果如图7所示。H22和Panc02采用皮下移植瘤方式,评估的重组溶瘤Ad5-mAPOA1-mIL15,结果如图8和9所示。相比单独携载mAPOA1或mIL15组的重组溶瘤Ad5病毒,Ad5-mAPOA1-mIL15具有明显的抗肿瘤优势。本发明的实施例得出的技术启示,使得领域内技术人员很容易获得Ad5-hAPOA1-hIL15在人源化动物模型中的抗肿瘤效果。
2)Ad5携载的mAPOA1和mIL15具有促进免疫记忆效应的协同效应
采用治愈的GL261小鼠,进行rechallenge实验,7d后取小鼠脾脏进行流式细胞术检测CD8+T细胞免疫记忆biomarker(CD44和CD62L)指标的变化,结果如图10所示。共表达APOA1和IL15的溶瘤Ad5组,CD62L和CD44具有显著提高,显示出协同促进免疫记忆的效应。
本领域技术人员将容易意识到,本发明非常适合于获得所提及以及本文固有的目的和优点。本文中作为目前代表性的优选实施方式描述的方法、变化形式和组合物仅是示例性的,并非对本发明的范围的限制。对本领域技术人员来说,可发生改变和其他用途,但这也被包括在本发明的由权利要求的范围界定的本发明的实质精神内。
本文示例性地描述的发明可适当地在不存在未在本文具体公开的任何一个或多个元素、一个或多个限制条件的情况下实施。所使用的术语和表达方式被用作描述而非限制,并非有意图在使用这些术语和表达方式时将所显示或描述的特征或其一部分的任何等同物排除在外,而要认识到,各种修改都可能在本发明的范围内。因此,要理解的是,虽然本发明已通过优选实施例和任选的特征被具体公开,但本领域技术人员仍可对本发明公开的概念作出修改和变化,而且这些修改和变化被认为在由所附权利要求限定的本发明的范围内。
序列表
<110> 南京惟亚德生物医药有限公司
<120> 一种重组溶瘤腺病毒及其应用
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 804
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
atgaaagctg cggtgctgac cttggccgtg ctcttcctga cggggagcca ggctcggcat 60
ttctggcagc aagatgaacc cccccagagc ccctgggatc gagtgaagga cctggccact 120
gtgtacgtgg atgtgctcaa agacagcggc agagactatg tgtcccagtt tgaaggctcc 180
gccttgggaa aacagctaaa cctaaagctc cttgacaact gggacagcgt gacctccacc 240
ttcagcaagc tgcgcgaaca gctcggccct gtgacccagg agttctggga taacctggaa 300
aaggagacag agggcctgag gcaggagatg agcaaggatc tggaggaggt gaaggccaag 360
gtgcagccct acctggacga cttccagaag aagtggcagg aggagatgga gctctaccgc 420
cagaaggtgg agccgctgcg cgcagagctc caagagggcg cgcgccagaa gctgcacgag 480
ctgcaagaga agctgagccc actgggcgag gagatgcgcg accgcgcgcg cgcccatgtg 540
gacgcgctgc gcacgcatct ggccccctac agcgacgagc tgcgccagcg cttggccgcg 600
cgccttgagg ctctcaagga gaacggcggc gccagactgg ccgagtacca cgccaaggcc 660
accgagcatc tgagcacgct cagcgagaag gccaagcccg cgctcgagga cctccgccaa 720
ggcctgctgc ccgtgctgga gagcttcaag gtcagcttcc tgagcgctct cgaggagtac 780
actaagaagc tcaacaccca gtga 804
<210> 2
<211> 489
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
atgagaattt cgaaaccaca tttgagaagt atttccatcc agtgctactt gtgtttactt 60
ctaaacagtc attttctaac tgaagctggc attcatgtct tcattttggg ctgtttcagt 120
gcagggcttc ctaaaacaga agccaactgg gtgaatgtaa taagtgattt gaaaaaaatt 180
gaagatctta ttcaatctat gcatattgat gctactttat atacggaaag tgatgttcac 240
cccagttgca aagtaacagc aatgaagtgc tttctcttgg agttacaagt tatttcactt 300
gagtccggag atgcaagtat tcatgataca gtagaaaatc tgatcatcct agcaaacaac 360
agtttgtctt ctaatgggaa tgtaacagaa tctggatgca aagaatgtga ggaactggag 420
gaaaaaaata ttaaagaatt tttgcagagt tttgtacata ttgtccaaat gttcatcaac 480
acttcttga 489
Claims (10)
1.一种重组溶瘤腺病毒,其特征在于:其基因组E1和E3区域缺失,缺失区域被稳定地插入胆固醇调控元件和免疫调节元件的外源核酸序列,其中,
所述的胆固醇调控元件为包含APOA1基因或其简并序列在内的功能元件;
所述的免疫调节元件为包含选自GM-CSF、IFN-γ、IL2、IL7、IL12、IL15、IL16、IL18、IL21、IL22、IL27、IL28和IL29基因或其简并序列在内的功能元件;
所述的胆固醇调节元件和免疫调节元件的基因以非融合方式表达;
所述的胆固醇调节元件和免疫调节元件是以可操作地连接方式连接至外源调控序列,包括启动子序列、增强子序列和PA序列。
2.根据权利要求1所述的重组溶瘤腺病毒,其特征在于:所述腺病毒选自C血清型亚群,包括人2型和5型腺病毒。
3.根据权利要求1所述的重组溶瘤腺病毒,其特征在于:所述启动子序列选自组成型、组织特异型或诱导型启动子;优选的,所述组成型启动子选自CMV、SV40或EF1α启动子。
4.根据权利要求1所述的重组溶瘤腺病毒,其特征在于:所述胆固醇调节元件和免疫调节元件为人源化的。
5.根据权利要求1或4所述的重组溶瘤腺病毒,其特征在于:其腺病毒为人5型腺病毒,所述胆固醇调节元件为APOA1基因,所述免疫调节元件为IL15基因。
6.根据权利要求5所述的重组溶瘤腺病毒,其特征在于:所述胆固醇调节元件APOA1的核酸序列所述胆固醇调节元件APOA1的核酸序列选自(a)或(b):
(a)如SEQ ID NO:1所示;
(b)与SEQ ID NO:1具有60%、65%、70%、75%、80%、85%、90%、95%、98%或99%的同源性的核酸序列;
优选的,所述免疫调节元件IL15的核酸序列选自(c)或(d):
(c)如SEQ ID NO:2所示;
(d)与SEQ ID NO:2具有60%、65%、70%、75%、80%、85%、90%、95%、98%或99%的同源性的核酸序列。
7.一种重组溶瘤Ad5,其包含修饰的Ad5基因组,其特征在于;该修饰包含(i)缺失野生型Ad5基因组的E1,E3区序列,但包括早期复制所需的E1A序列;(ii)编码胆固醇代谢调节剂和/或免疫调节剂的外源性核酸序列,其中所述的外源性核酸序列被稳定地加入到至少是所述经修饰的Ad5基因组的被缺失区域;优选的,所述的编码胆固醇代谢调节剂的外源性核酸序列如SEQ ID NO:1所示;所述的编码免疫调节剂的外源性核酸序列如SEQ ID NO:2所示。
8.权利要求1-6任一所述的重组溶瘤腺病毒、权利要求7所述的重组溶瘤Ad5在制备治疗或缓解癌症相关疾病的药物中的应用;优选的,权利要求1-6任一所述的重组溶瘤腺病毒、权利要求7所述的重组溶瘤Ad5,与第二治疗药物在制备治疗或缓解癌症相关疾病的药物中的应用;更优选的,所述第二治疗药物为PD1抗体。
9.根据权利要求8所述的应用,其特征在于:所述癌症选自胆囊癌、基地细胞瘤、肝外胆管癌、结肠癌、子宫内膜癌、子宫肌瘤、食道癌、尤因肉瘤、前列腺癌、胃癌、肝癌、肝细胞癌、霍奇金淋巴瘤、喉癌、肺癌、黑色素瘤、间皮瘤、胰腺癌、直肠癌、肾癌、甲状腺癌、神经胶质瘤、恶性周围神经细胞肿瘤、恶性外周神经鞘瘤、皮肤和丛状神经纤维瘤、平滑肌瘤、平滑肌肉瘤、纤维瘤、乳头状腺瘤、甲状腺未分化癌、甲状腺髓样癌、甲状腺滤泡癌、hurthle细胞癌、甲状腺癌、腹水、恶性腹水、唾液腺肿瘤、唾液腺粘液表皮样癌、唾液腺腺泡细胞癌、胃肠道基质肿瘤、导致身体潜在空间积液的肿瘤、胸腔积液、心包积液、腹膜积液、巨细胞瘤、骨巨细胞瘤、色素沉着绒毛结节性滑膜炎、腱鞘巨细胞瘤和其他肉瘤中的任一种,优选为神经胶质瘤、胰腺癌或肝癌。
10.一种药物组合物,其含有权利要求1-7中任意一项所述的重组溶瘤腺病毒或权利要求7所述的重组溶瘤Ad5,以及药学上可接受的载体或赋形剂;优选的,所述的组合物被配制用于瘤内施用。
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Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1706955A (zh) * | 2004-06-07 | 2005-12-14 | 成都康弘科技实业(集团)有限公司 | 肿瘤细胞专一表达免疫调节因子gm-csf的溶瘤性腺病毒重组体的构建及其应用 |
US20050282280A1 (en) * | 2003-08-28 | 2005-12-22 | Ennist David L | Oncolytic adenoviral vectors encoding GM-CSF |
CN105177045A (zh) * | 2014-05-21 | 2015-12-23 | 晏阳 | 表达人白介素15的重组溶瘤腺病毒及其构建方法 |
CN112391412A (zh) * | 2019-08-19 | 2021-02-23 | 南京诺惟生物科技有限公司 | 一种调控脂质代谢的复制型溶瘤腺病毒及其应用 |
-
2021
- 2021-03-08 CN CN202110248832.5A patent/CN115029325A/zh active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20050282280A1 (en) * | 2003-08-28 | 2005-12-22 | Ennist David L | Oncolytic adenoviral vectors encoding GM-CSF |
CN1706955A (zh) * | 2004-06-07 | 2005-12-14 | 成都康弘科技实业(集团)有限公司 | 肿瘤细胞专一表达免疫调节因子gm-csf的溶瘤性腺病毒重组体的构建及其应用 |
CN105177045A (zh) * | 2014-05-21 | 2015-12-23 | 晏阳 | 表达人白介素15的重组溶瘤腺病毒及其构建方法 |
CN112391412A (zh) * | 2019-08-19 | 2021-02-23 | 南京诺惟生物科技有限公司 | 一种调控脂质代谢的复制型溶瘤腺病毒及其应用 |
Non-Patent Citations (1)
Title |
---|
XINGZHE MA ET AL.: "Cholesterol induces CD8+ T-cell exhaustion in the tumor microenvironment", 《CELL METAB.》, vol. 30, no. 1, 2 July 2019 (2019-07-02), pages 143 - 156 * |
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