Abstract A119: Combating primary and secondary checkpoint blockade resistance using immunostimulatory CD40L/4-1BBL-encoding oncolytic virotherapy for melanoma

J. Wenthe, Mantas Šilanskas, E. Eriksson, A. Loskog
{"title":"Abstract A119: Combating primary and secondary checkpoint blockade resistance using immunostimulatory CD40L/4-1BBL-encoding oncolytic virotherapy for melanoma","authors":"J. Wenthe, Mantas Šilanskas, E. Eriksson, A. Loskog","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A119","DOIUrl":null,"url":null,"abstract":"In recent years, cancer immunotherapies significantly advanced the clinical management of metastatic melanoma. In particular, treatment with immune checkpoint inhibitors increased survival rates compared with standard therapy. However, not all patients benefit from checkpoint blockade. Primary treatment resistance is connected to poor T-cell infiltration in the tumors. This can be due to limited activation of antigen-presentation cells such as dendritic cells (DCs) and the high threshold of activating low-affinity T-cells that may respond to the tumor antigens. CD27 signaling in T-cells during activation lowers the T-cell receptor signaling threshold, which may be important to activate tumor-targeting T-cells. Secondary resistance to checkpoint blockade therapy includes loss of MHC-I on the tumor cells. Hence, inducing natural killer (NK) cell activation in parallel to antitumor T-cells may be crucial. LOAd703 was designed to optimize antitumor immune activation. LOAd703 is an oncolytic adenovirus (serotype Ad5/35) carrying two immunostimulatory transgenes; a trimerized membrane-bound CD40 ligand (TMZ-CD40L) and the full-length 4-1BB ligand (4-1BBL) (patent filed: PCT/EP2015/057489). The viral replication is restricted to tumor cells with a hyperphosphorylated retinoblastoma pathway due to a deletion in E1A, but transgenes are expressed under the control of a cytomegalovirus (CMV) promoter, which enables transgene expression even in the surrounding tumor microenvironment. Herein, we investigated LOAd703 in a melanoma model and its immunostimulatory effect on DC maturation to induce antigen-specific T-cell responses. LOAd703 infected the human melanoma cell line 526-mel and efficiently induced tumor cell death as evaluated by MTS viability assay. The viability of infected cells was reduced to 15% at 72 hours post infection compared to uninfected cells. Transgene expression of both TMZ-CD40L and 4-1BBL was confirmed by flow cytometry post infection. To evaluate the immunostimulatory capacity of LOAd703, immature DCs were differentiated from CD14+ monocytes using granulocyte-macrophage colony-stimulating factor and interleukin-4 and infected with virus. LOAd703-infected DCs upregulated the expression of maturation markers, such as CD83, CD86 and MHC molecules as analyzed by flow cytometry. Interestingly, CD70 that is required for CD27 stimuli of T-cells was highly upregulated on the DCs using LOAd703. Furthermore, the chemokine receptor CCR7 and the adhesion molecule ICAM-1 were increased upon LOAd703 infection, which are crucial for lymph node homing and the initiation of a systemic response. Next, the functional capacity of LOAd703-matured DCs to induce antigen-specific T-cell responses was assessed in a CMV model, in which CMV-peptide pulsed DCs are utilized to induce expansion of CMV-specific T-cells. LOAd703-matured DCs from CMV+ donors were pulsed with CMV-peptide and co-cultured with autologous peripheral blood mononuclear cells for 11 days. LOAd703-matured DCs were able to expand CMV-specific T-cells, but to a lower degree than the positive control Poly I:C matured DCs. However, the CMV-specific T-cells expanded from the positive control had higher expression of PD-1 compared to the LOAd703 group, indicating that LOAd703 leads to the expansion of less exhausted and potentially more functional T-cells. Moreover, LOAd703 also induced a massive expansion of NK cells, which is probably driven by 4-1BBL. In conclusion, LOAd703 killed human melanoma cells by oncolysis and induced the expression of TMZ-CD40L and 4-1BBL in infected cells. Furthermore, LOAd703 infection activated DCs to express costimulatory molecules including high levels of CD70 and CCR7, which in turn could promote the expansion of antigen-specific T-cells with low PD-1 expression, as well as the potent expansion of NK cells. Citation Format: Jessica Wenthe, Mantas Silanskas, Emma Eriksson, Angelica Loskog. Combating primary and secondary checkpoint blockade resistance using immunostimulatory CD40L/4-1BBL-encoding oncolytic virotherapy for melanoma [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A119.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Tackling the Tumor Microenvironment: Beyond T-cells","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A119","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

Abstract

In recent years, cancer immunotherapies significantly advanced the clinical management of metastatic melanoma. In particular, treatment with immune checkpoint inhibitors increased survival rates compared with standard therapy. However, not all patients benefit from checkpoint blockade. Primary treatment resistance is connected to poor T-cell infiltration in the tumors. This can be due to limited activation of antigen-presentation cells such as dendritic cells (DCs) and the high threshold of activating low-affinity T-cells that may respond to the tumor antigens. CD27 signaling in T-cells during activation lowers the T-cell receptor signaling threshold, which may be important to activate tumor-targeting T-cells. Secondary resistance to checkpoint blockade therapy includes loss of MHC-I on the tumor cells. Hence, inducing natural killer (NK) cell activation in parallel to antitumor T-cells may be crucial. LOAd703 was designed to optimize antitumor immune activation. LOAd703 is an oncolytic adenovirus (serotype Ad5/35) carrying two immunostimulatory transgenes; a trimerized membrane-bound CD40 ligand (TMZ-CD40L) and the full-length 4-1BB ligand (4-1BBL) (patent filed: PCT/EP2015/057489). The viral replication is restricted to tumor cells with a hyperphosphorylated retinoblastoma pathway due to a deletion in E1A, but transgenes are expressed under the control of a cytomegalovirus (CMV) promoter, which enables transgene expression even in the surrounding tumor microenvironment. Herein, we investigated LOAd703 in a melanoma model and its immunostimulatory effect on DC maturation to induce antigen-specific T-cell responses. LOAd703 infected the human melanoma cell line 526-mel and efficiently induced tumor cell death as evaluated by MTS viability assay. The viability of infected cells was reduced to 15% at 72 hours post infection compared to uninfected cells. Transgene expression of both TMZ-CD40L and 4-1BBL was confirmed by flow cytometry post infection. To evaluate the immunostimulatory capacity of LOAd703, immature DCs were differentiated from CD14+ monocytes using granulocyte-macrophage colony-stimulating factor and interleukin-4 and infected with virus. LOAd703-infected DCs upregulated the expression of maturation markers, such as CD83, CD86 and MHC molecules as analyzed by flow cytometry. Interestingly, CD70 that is required for CD27 stimuli of T-cells was highly upregulated on the DCs using LOAd703. Furthermore, the chemokine receptor CCR7 and the adhesion molecule ICAM-1 were increased upon LOAd703 infection, which are crucial for lymph node homing and the initiation of a systemic response. Next, the functional capacity of LOAd703-matured DCs to induce antigen-specific T-cell responses was assessed in a CMV model, in which CMV-peptide pulsed DCs are utilized to induce expansion of CMV-specific T-cells. LOAd703-matured DCs from CMV+ donors were pulsed with CMV-peptide and co-cultured with autologous peripheral blood mononuclear cells for 11 days. LOAd703-matured DCs were able to expand CMV-specific T-cells, but to a lower degree than the positive control Poly I:C matured DCs. However, the CMV-specific T-cells expanded from the positive control had higher expression of PD-1 compared to the LOAd703 group, indicating that LOAd703 leads to the expansion of less exhausted and potentially more functional T-cells. Moreover, LOAd703 also induced a massive expansion of NK cells, which is probably driven by 4-1BBL. In conclusion, LOAd703 killed human melanoma cells by oncolysis and induced the expression of TMZ-CD40L and 4-1BBL in infected cells. Furthermore, LOAd703 infection activated DCs to express costimulatory molecules including high levels of CD70 and CCR7, which in turn could promote the expansion of antigen-specific T-cells with low PD-1 expression, as well as the potent expansion of NK cells. Citation Format: Jessica Wenthe, Mantas Silanskas, Emma Eriksson, Angelica Loskog. Combating primary and secondary checkpoint blockade resistance using immunostimulatory CD40L/4-1BBL-encoding oncolytic virotherapy for melanoma [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A119.
A119:利用免疫刺激CD40L/4- 1bbl编码的溶瘤病毒疗法对抗黑色素瘤的原发性和继发性检查点阻断耐药性
近年来,癌症免疫疗法显著推进了转移性黑色素瘤的临床治疗。特别是,与标准治疗相比,免疫检查点抑制剂治疗增加了生存率。然而,并非所有患者都能从检查点阻断中获益。原发性治疗耐药与肿瘤中t细胞浸润不良有关。这可能是由于抗原呈递细胞(如树突状细胞)的激活有限,以及激活可能对肿瘤抗原有反应的低亲和力t细胞的高阈值。t细胞活化过程中的CD27信号传导降低了t细胞受体信号传导阈值,这可能对激活肿瘤靶向t细胞很重要。对检查点阻断治疗的继发性耐药包括肿瘤细胞上mhc - 1的丢失。因此,诱导自然杀伤(NK)细胞激活与抗肿瘤t细胞激活可能至关重要。LOAd703被设计用于优化抗肿瘤免疫激活。LOAd703是一种溶瘤腺病毒(血清型Ad5/35),携带两种免疫刺激转基因;三聚膜结合CD40配体(TMZ-CD40L)和全长4-1BB配体(4-1BBL)(专利申请:PCT/EP2015/057489)。由于E1A缺失,病毒复制仅限于具有高磷酸化视网膜母细胞瘤通路的肿瘤细胞,但转基因在巨细胞病毒(CMV)启动子的控制下表达,这使得转基因甚至可以在周围的肿瘤微环境中表达。在此,我们在黑色素瘤模型中研究了LOAd703及其对DC成熟的免疫刺激作用,以诱导抗原特异性t细胞反应。LOAd703感染人黑色素瘤细胞系526-mel后,MTS活性试验证实其能有效诱导肿瘤细胞死亡。感染后72小时,与未感染的细胞相比,感染细胞的存活率降低至15%。感染后用流式细胞术证实TMZ-CD40L和4-1BBL的转基因表达。为了评估LOAd703的免疫刺激能力,使用粒细胞-巨噬细胞集落刺激因子和白细胞介素-4将未成熟的dc从CD14+单核细胞分化并感染病毒。流式细胞术分析发现,load703感染的dc上调了成熟标志物CD83、CD86和MHC分子的表达。有趣的是,使用LOAd703后,CD27刺激t细胞所需的CD70在dc上被高度上调。此外,趋化因子受体CCR7和粘附分子ICAM-1在LOAd703感染后增加,这对于淋巴结归巢和系统反应的启动至关重要。接下来,在CMV模型中评估load703成熟dc诱导抗原特异性t细胞反应的功能能力,在CMV模型中,利用CMV肽脉冲dc诱导CMV特异性t细胞扩增。来自CMV+供者的load703成熟dc用CMV肽脉冲,与自体外周血单个核细胞共培养11天。load703成熟dc能够扩增cmv特异性t细胞,但扩增程度低于阳性对照Poly I:C成熟dc。然而,与LOAd703组相比,从阳性对照扩增的cmv特异性t细胞具有更高的PD-1表达,这表明LOAd703导致较少耗尽且可能更有功能的t细胞扩增。此外,LOAd703还诱导NK细胞大量扩增,这可能是由4-1BBL驱动的。综上所述,LOAd703通过溶瘤杀伤人黑色素瘤细胞,并诱导感染细胞中TMZ-CD40L和4-1BBL的表达。此外,LOAd703感染激活dc表达共刺激分子,包括高水平的CD70和CCR7,这反过来可以促进低PD-1表达的抗原特异性t细胞的扩增,以及NK细胞的有效扩增。引文格式:Jessica Wenthe, Mantas Silanskas, Emma Eriksson, Angelica Loskog。利用免疫刺激CD40L/4- 1bbl编码的溶瘤病毒疗法对抗黑色素瘤的原发性和继发性检查点阻断耐药性[摘要]。第四届CRI-CIMT-EATI-AACR国际癌症免疫治疗会议:将科学转化为生存;2018年9月30日至10月3日;纽约,纽约。费城(PA): AACR;癌症免疫,2019;7(2增刊):摘要nr A119。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信