Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A137
Iliana Kyriaki, S. Mangsbo
The mouse bladder 49 (MB49) syngeneic tumors respond to various immunotherapies, both stimulatory immunotherapies and check-point blockers; when tumors are cured it is often an antitumor memory response established (previous publications by our group and others). Herein we present data suggesting that bladders of orthotopic MB49 tumor-bearing mice hold a reservoir of lymphocytes surrounding the tumor, wherein they migrate rapidly upon immune adjuvant stimulation. The tumors appear to form an accessible immune microenvironment rich in vessels that mediate the homing of leukocytes to the tumor site. This is in contrast to MB49 tumors that grow subcutaneously and do not appear to provide the same rapid immune cell infiltration capacity. Herein we further examined the immune cell populations of tumors locally treated with CpG ODN 1668, a type B CpG and a strong immunostimulatory agent or dilution buffer, as a control. Interestingly, we identified markers of high-endothelial venules (HEVs) in the vessels present in tumors from both untreated and treated animals. Thus we conclude that HEVs develop naturally in the orthotopic MB49 model and we wished to explore whether manipulation of the tumor microenvironment (TME) can induce HEVs and how this stimuli affect the infiltration of immune cells. Immunohistochemical profiling revealed that CD11c+ dendritic cells, CCL21+ cells along with B and T lymphocytes are present in various ways in different zones of the tumor with HEVs and CD31+ vessels. A correlation with vessel density and the presence of HEVs was identified in untreated animals. In immune-stimulated tumors, this correlation was lost in the early phase post stimuli. We could also identify that CpG treated bladder displayed increased vascularization around the tumor 24 hours post treatment. Interestingly, it appeared as CCL21 expression intratumorally aligned with increased recruitment of CD11c+ DCs in the TME. Intratumorally it was a clear correlation between CCL21 expression with both CD11c+ and CD8+ cell infiltration. As CD11c+ dendritic cells migrate towards gradients of CCL21 via their CCR7 receptor, enabling antigen presentation, it is plausible that antigen-presentation takes place in the TME in relation to this finding. By these preliminary data we conclude that in the orthotopic MB49 tumor model, CCL21 appears to play a role in the induction of adaptive immune responses through the CCL21/CCR7 axis. The role of HEVs, or alternatively lymphatic vessels, for this recruitment is yet to be determined. Currently we are investigating multiplex analyses on bladder tumors, urine and blood from tumor-bearing animals to identify cytokines in the TME or systemically that are correlated the formation of an adaptive immune response. Our future goal is to examine what is the key to raise an effective immune response to the so called immune "hot" orthotopic MB49 tumor. Citation Format: Iliana Kyriaki, Sara M. Mangsbo. The innate/adaptive immune response tr
小鼠膀胱49 (MB49)同基因肿瘤对刺激免疫疗法和检查点阻断剂等多种免疫疗法均有应答;当肿瘤被治愈时，通常是建立了抗肿瘤记忆反应(我们小组和其他人之前的出版物)。在此，我们提供的数据表明，原位MB49荷瘤小鼠的膀胱在肿瘤周围有一个淋巴细胞储存库，在免疫佐剂刺激下，淋巴细胞迅速迁移。肿瘤似乎形成了一个可接近的免疫微环境，富含介导白细胞归巢到肿瘤部位的血管。这与皮下生长的MB49肿瘤形成对比，MB49肿瘤似乎不提供相同的快速免疫细胞浸润能力。在这里，我们进一步研究了用CpG ODN 1668局部治疗的肿瘤的免疫细胞群，CpG是一种B型CpG和一种强免疫刺激剂或稀释缓冲液作为对照。有趣的是，我们在未治疗和治疗的动物的肿瘤血管中都发现了高内皮小静脉(HEVs)的标记物。因此，我们得出结论，hev在原位MB49模型中自然发展，我们希望探索操纵肿瘤微环境(TME)是否可以诱导hev，以及这种刺激如何影响免疫细胞的浸润。免疫组织化学分析显示，CD11c+树突状细胞、CCL21+细胞以及B淋巴细胞和T淋巴细胞以不同的方式存在于hev和CD31+血管肿瘤的不同区域。在未经治疗的动物中发现了与血管密度和hev存在的相关性。在免疫刺激的肿瘤中，这种相关性在刺激后的早期阶段消失。我们还发现，CpG治疗后24小时膀胱肿瘤周围血管化增加。有趣的是，CCL21在肿瘤内的表达与TME中CD11c+ DCs的增加募集一致。瘤内CCL21的表达与CD11c+和CD8+细胞浸润有明显的相关性。由于CD11c+树突状细胞通过其CCR7受体向CCL21梯度迁移，使抗原呈递成为可能，因此抗原呈递发生在TME中与这一发现有关。通过这些初步数据，我们得出结论，在原位MB49肿瘤模型中，CCL21似乎通过CCL21/CCR7轴在诱导适应性免疫应答中发挥作用。hev或淋巴管在这种招募中的作用尚未确定。目前，我们正在研究对荷瘤动物膀胱肿瘤、尿液和血液的多重分析，以确定TME或系统中与适应性免疫反应形成相关的细胞因子。我们未来的目标是研究什么是提高对所谓的免疫“热”原位MB49肿瘤有效免疫反应的关键。引文格式:Iliana Kyriaki, Sara M. Mangsbo。原位生长的膀胱癌肿瘤局部免疫治疗引发的先天/适应性免疫反应[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;癌症免疫学杂志2019;7(2增刊):摘要nr A137。
{"title":"Abstract A137: The innate/adaptive immune response triggered in response to local immunotherapy of orthotopically growing bladder cancer tumors","authors":"Iliana Kyriaki, S. Mangsbo","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A137","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A137","url":null,"abstract":"The mouse bladder 49 (MB49) syngeneic tumors respond to various immunotherapies, both stimulatory immunotherapies and check-point blockers; when tumors are cured it is often an antitumor memory response established (previous publications by our group and others). Herein we present data suggesting that bladders of orthotopic MB49 tumor-bearing mice hold a reservoir of lymphocytes surrounding the tumor, wherein they migrate rapidly upon immune adjuvant stimulation. The tumors appear to form an accessible immune microenvironment rich in vessels that mediate the homing of leukocytes to the tumor site. This is in contrast to MB49 tumors that grow subcutaneously and do not appear to provide the same rapid immune cell infiltration capacity. Herein we further examined the immune cell populations of tumors locally treated with CpG ODN 1668, a type B CpG and a strong immunostimulatory agent or dilution buffer, as a control. Interestingly, we identified markers of high-endothelial venules (HEVs) in the vessels present in tumors from both untreated and treated animals. Thus we conclude that HEVs develop naturally in the orthotopic MB49 model and we wished to explore whether manipulation of the tumor microenvironment (TME) can induce HEVs and how this stimuli affect the infiltration of immune cells. Immunohistochemical profiling revealed that CD11c+ dendritic cells, CCL21+ cells along with B and T lymphocytes are present in various ways in different zones of the tumor with HEVs and CD31+ vessels. A correlation with vessel density and the presence of HEVs was identified in untreated animals. In immune-stimulated tumors, this correlation was lost in the early phase post stimuli. We could also identify that CpG treated bladder displayed increased vascularization around the tumor 24 hours post treatment. Interestingly, it appeared as CCL21 expression intratumorally aligned with increased recruitment of CD11c+ DCs in the TME. Intratumorally it was a clear correlation between CCL21 expression with both CD11c+ and CD8+ cell infiltration. As CD11c+ dendritic cells migrate towards gradients of CCL21 via their CCR7 receptor, enabling antigen presentation, it is plausible that antigen-presentation takes place in the TME in relation to this finding. By these preliminary data we conclude that in the orthotopic MB49 tumor model, CCL21 appears to play a role in the induction of adaptive immune responses through the CCL21/CCR7 axis. The role of HEVs, or alternatively lymphatic vessels, for this recruitment is yet to be determined. Currently we are investigating multiplex analyses on bladder tumors, urine and blood from tumor-bearing animals to identify cytokines in the TME or systemically that are correlated the formation of an adaptive immune response. Our future goal is to examine what is the key to raise an effective immune response to the so called immune \"hot\" orthotopic MB49 tumor. Citation Format: Iliana Kyriaki, Sara M. Mangsbo. The innate/adaptive immune response tr","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"47 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"85622443","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A136
E. Kandimalla, S. Nallagatla, Bart R. Anderson, R. Kang
Spherical nucleic acids (SNA) are a novel class of therapeutic agents in which oligonucleotides are densely packed and radially arranged on the surface of liposomal nanoparticles providing a 3D-architecture. This architecture provides increased cellular uptake and nuclease stability compared to linear oligonucleotides that are not in SNA format. AST-008 is an SNA configuration of a Toll-like receptor 9 (TLR9) agonist that induces potent Th1-type immune responses in vitro, and in mice and nonhuman primates. Subcutaneous (SC), intratumoral (IT) and intravenous (IV) administration of AST-008 has shown potent antitumor activity as a monotherapy and enhanced checkpoint inhibitor (CPI) activity in a number of mouse tumor models. Our previous studies have shown that AST-008 induces potent innate and adaptive immune responses and increases tumor infiltrating lymphocytes, interferon-inducible gene expression, and activation and expansion of CD8+ T-cells with reduced T-regulatory cells in the tumor microenvironment facilitating increased effectiveness of CPI. In the present studies, we have evaluated i) sequence-specific antitumor effects of AST-008 following SC delivery in an EMT-6 breast tumor model, ii) abscopal antitumor effects of a murine version of the TLR9 agonist SNA following SC delivery in a two-flank EMT-6 tumor model, and iii) abscopal antitumor effects of AST-008 following SC and IT administrations in an MC38 colon cancer model. Treatment of EMT-6 breast tumors in mice with SC administration of AST-008 showed dose-dependent tumor growth inhibition that was statistically significant compared with vehicle-treated tumor-bearing mice. A control oligonucleotide-SNA had no effect on tumor growth compared with the vehicle group. In a two-flank EMT-6 breast tumor model, SC administration of TLR9 agonist SNA showed about 59% and 37% tumor growth inhibition at the injected and distant tumors, respectively, compared to the vehicle-treated mice. The tumor growth inhibition was statistically significant at both the injected and uninjected tumors compared to vehicle-treated tumor-bearing mice. In a two-flank MC38 colon cancer model, either SC or IT administration of AST-008 resulted in 69% and 68% tumor growth inhibition, respectively, at the injected tumor. In this study, distant tumor that was not treated with AST-008 showed 59% and 46% tumor growth inhibition for SC and IT routes of administration, respectively. The tumor growth inhibition at both the treated and untreated tumors was statistically significant compared to the vehicle-treated tumors. These data demonstrate sequence-specific antitumor activity of AST-008 and that SC or IT administration at one tumor lesion leads to systemic antitumor activity at distant uninjected tumors. The current results, together with our previous studies in combination with CPI in several tumor models, support the potential utility of the TLR9-mediated innate immune stimulator AST-008 in combination with CPI in cance
球形核酸(SNA)是一类新型的治疗剂，其中寡核苷酸密集堆积并呈放射状排列在脂质体纳米颗粒表面，提供3d结构。与非SNA格式的线性寡核苷酸相比，这种结构提供了更高的细胞摄取和核酸酶稳定性。AST-008是toll样受体9 (TLR9)激动剂的SNA构型，在体外、小鼠和非人灵长类动物中诱导强效th1型免疫反应。在许多小鼠肿瘤模型中，皮下(SC)、肿瘤内(IT)和静脉(IV)给药AST-008显示出强大的抗肿瘤活性，并增强了检查点抑制剂(CPI)活性。我们之前的研究表明，AST-008诱导了强效的先天和适应性免疫反应，增加了肿瘤浸润淋巴细胞、干扰素诱导的基因表达，以及肿瘤微环境中CD8+ t细胞的激活和扩增，减少了t调节细胞，从而提高了CPI的有效性。在目前的研究中，我们评估了1)在EMT-6乳腺肿瘤模型中，SC给药后AST-008的序列特异性抗肿瘤作用，2)在两侧EMT-6肿瘤模型中，SC给药后TLR9激动剂SNA的小鼠版本的体外抗肿瘤作用，以及3)在MC38结肠癌模型中，SC和IT给药后AST-008的体外抗肿瘤作用。SC给药AST-008治疗小鼠EMT-6乳腺肿瘤显示出剂量依赖性肿瘤生长抑制，与载瘤小鼠相比具有统计学意义。对照寡核苷酸- sna对肿瘤生长无影响。在双侧EMT-6乳腺肿瘤模型中，SC给药TLR9激动剂SNA对注射肿瘤和远处肿瘤的肿瘤生长抑制作用分别为59%和37%。与载体处理的荷瘤小鼠相比，注射和未注射肿瘤的肿瘤生长抑制均有统计学意义。在双侧MC38结肠癌模型中，SC或IT给药AST-008分别在注射肿瘤处产生69%和68%的肿瘤生长抑制。在本研究中，未使用AST-008治疗的远处肿瘤，SC和IT给药途径分别显示59%和46%的肿瘤生长抑制。治疗组和未治疗组的肿瘤生长抑制均比药物治疗组有统计学意义。这些数据证明了AST-008的序列特异性抗肿瘤活性，并且在一个肿瘤病变处给予SC或IT可导致远端未注射肿瘤的全身抗肿瘤活性。目前的结果，以及我们之前在几种肿瘤模型中联合CPI的研究，支持tlr9介导的先天免疫刺激剂AST-008联合CPI在癌症患者中的潜在效用。AST-008目前处于临床开发阶段。引文格式:Ekambar R. Kandimalla, SubbaRao Nallagatla, Bart R. Anderson, Richard Kang。新型TLR9激动剂SNA AST-008在小鼠肿瘤模型中诱导体外抗肿瘤作用[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 A136。
{"title":"Abstract A136: AST-008, a novel TLR9 agonist SNA, induces abscopal antitumor effects in mouse tumor models","authors":"E. Kandimalla, S. Nallagatla, Bart R. Anderson, R. Kang","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A136","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A136","url":null,"abstract":"Spherical nucleic acids (SNA) are a novel class of therapeutic agents in which oligonucleotides are densely packed and radially arranged on the surface of liposomal nanoparticles providing a 3D-architecture. This architecture provides increased cellular uptake and nuclease stability compared to linear oligonucleotides that are not in SNA format. AST-008 is an SNA configuration of a Toll-like receptor 9 (TLR9) agonist that induces potent Th1-type immune responses in vitro, and in mice and nonhuman primates. Subcutaneous (SC), intratumoral (IT) and intravenous (IV) administration of AST-008 has shown potent antitumor activity as a monotherapy and enhanced checkpoint inhibitor (CPI) activity in a number of mouse tumor models. Our previous studies have shown that AST-008 induces potent innate and adaptive immune responses and increases tumor infiltrating lymphocytes, interferon-inducible gene expression, and activation and expansion of CD8+ T-cells with reduced T-regulatory cells in the tumor microenvironment facilitating increased effectiveness of CPI. In the present studies, we have evaluated i) sequence-specific antitumor effects of AST-008 following SC delivery in an EMT-6 breast tumor model, ii) abscopal antitumor effects of a murine version of the TLR9 agonist SNA following SC delivery in a two-flank EMT-6 tumor model, and iii) abscopal antitumor effects of AST-008 following SC and IT administrations in an MC38 colon cancer model. Treatment of EMT-6 breast tumors in mice with SC administration of AST-008 showed dose-dependent tumor growth inhibition that was statistically significant compared with vehicle-treated tumor-bearing mice. A control oligonucleotide-SNA had no effect on tumor growth compared with the vehicle group. In a two-flank EMT-6 breast tumor model, SC administration of TLR9 agonist SNA showed about 59% and 37% tumor growth inhibition at the injected and distant tumors, respectively, compared to the vehicle-treated mice. The tumor growth inhibition was statistically significant at both the injected and uninjected tumors compared to vehicle-treated tumor-bearing mice. In a two-flank MC38 colon cancer model, either SC or IT administration of AST-008 resulted in 69% and 68% tumor growth inhibition, respectively, at the injected tumor. In this study, distant tumor that was not treated with AST-008 showed 59% and 46% tumor growth inhibition for SC and IT routes of administration, respectively. The tumor growth inhibition at both the treated and untreated tumors was statistically significant compared to the vehicle-treated tumors. These data demonstrate sequence-specific antitumor activity of AST-008 and that SC or IT administration at one tumor lesion leads to systemic antitumor activity at distant uninjected tumors. The current results, together with our previous studies in combination with CPI in several tumor models, support the potential utility of the TLR9-mediated innate immune stimulator AST-008 in combination with CPI in cance","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82114993","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A142
Y. Leshem, E. King, Y. Reiter, I. Pastan
Background: SS1P and LMB-100 are anti-mesothelin immunotoxins composed of a targeting antibody fragment genetically fused to a truncated pseudomonas exotoxin A. We have previously repotrted a synergistic antitumor effect when SS1P or LMB-100 were injected locally into tumors of murine breast cancer in combination with antibodies that block the immune check point CTLA-4. In this study our goal is to explore if this treatment can be applied in a murine mesothelioma model. Method: The mouse mesothelioma AE17M cell line, expressing human mesothelin, was treated in culture with anti-mesothelin immunotoxins. We collected data on the cytotoxic activity and ability to induce markers of immunogenic cell death. In vivo effects were performed on AE17M tumors growing in C57/Bl6 mice. Results: AE17M cells are sensitive to both SS1P and LMB-100 in culture, but when injected IV, the immunotoxins did not affect AE17M tumor growth rate. To overcome obstacles in drug penetration we injected immunotoxins directly into AE17M tumors and found that they only increased the survival of mice from 16 days to 20 days. In addition, anti-CTLA-4 monotherapy failed to control AE17M tumor growth rate in most mice. But when locally injected SS1P or LMB-100 was combined with i.p. anti-CTLA-4, tumor regressions occurred and the median survival increased to 80 days or longer. In search for possible mechanisms enabling immunotoxins to contribute to anti-CTLA-4 activity, we examined their ability to induce markers of immunogenic cell death. We found that both LMB-100 and SS1P increased release of ATP from AE17M cells. In addition, LMB-100 induced calreticulin expression on the surface of both AE17M cells and KLM-1 cells. These results point to possible pathways allowing immunotoxins to promote anti-tumor immunity and sensitize tumors for anti-CTLA-4 effect. Conclusion: Our findings in the AE17M model provide additional support for the use of local immunotoxins in combination with anti-CTLA-4. Citation Format: Yasmin Leshem, Emily King, Yoram Reiter, Ira Pastan. Anti-mesothelin immunotoxins induce markers of immunogenic cell death and when injected locally into AE17M mesothelioma tumors enhance the effect of CTLA-4 blockade [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 A142.
{"title":"Abstract A142: Anti-mesothelin immunotoxins induce markers of immunogenic cell death and when injected locally into AE17M mesothelioma tumors enhance the effect of CTLA-4 blockade","authors":"Y. Leshem, E. King, Y. Reiter, I. Pastan","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A142","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A142","url":null,"abstract":"Background: SS1P and LMB-100 are anti-mesothelin immunotoxins composed of a targeting antibody fragment genetically fused to a truncated pseudomonas exotoxin A. We have previously repotrted a synergistic antitumor effect when SS1P or LMB-100 were injected locally into tumors of murine breast cancer in combination with antibodies that block the immune check point CTLA-4. In this study our goal is to explore if this treatment can be applied in a murine mesothelioma model. Method: The mouse mesothelioma AE17M cell line, expressing human mesothelin, was treated in culture with anti-mesothelin immunotoxins. We collected data on the cytotoxic activity and ability to induce markers of immunogenic cell death. In vivo effects were performed on AE17M tumors growing in C57/Bl6 mice. Results: AE17M cells are sensitive to both SS1P and LMB-100 in culture, but when injected IV, the immunotoxins did not affect AE17M tumor growth rate. To overcome obstacles in drug penetration we injected immunotoxins directly into AE17M tumors and found that they only increased the survival of mice from 16 days to 20 days. In addition, anti-CTLA-4 monotherapy failed to control AE17M tumor growth rate in most mice. But when locally injected SS1P or LMB-100 was combined with i.p. anti-CTLA-4, tumor regressions occurred and the median survival increased to 80 days or longer. In search for possible mechanisms enabling immunotoxins to contribute to anti-CTLA-4 activity, we examined their ability to induce markers of immunogenic cell death. We found that both LMB-100 and SS1P increased release of ATP from AE17M cells. In addition, LMB-100 induced calreticulin expression on the surface of both AE17M cells and KLM-1 cells. These results point to possible pathways allowing immunotoxins to promote anti-tumor immunity and sensitize tumors for anti-CTLA-4 effect. Conclusion: Our findings in the AE17M model provide additional support for the use of local immunotoxins in combination with anti-CTLA-4. Citation Format: Yasmin Leshem, Emily King, Yoram Reiter, Ira Pastan. Anti-mesothelin immunotoxins induce markers of immunogenic cell death and when injected locally into AE17M mesothelioma tumors enhance the effect of CTLA-4 blockade [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 A142.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":" 7","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91411202","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A160
F. V. Maldegem, Karishma Valand, V. Tsang, E. Mugarza, D. Caswell, P. Hobson, J. Downward
With the advancement of immunotherapies for lung cancer, it has become apparent how insufficient our knowledge is surrounding the interactions between the tumor and the immune system. In particular in the context of combining immunotherapy with conventional chemotherapy or novel targeted therapies, it is of the utmost importance that we understand the effects that those therapies have on the tumor immune infiltrate and how that would counteract or synergize with immunotherapy. We have used multiplex flow cytometry to characterise the basal tumor immune infiltrate of various spontaneous and orthogonous syngeneic mouse models for lung cancer, with different levels of immunogenicity. With imaging mass cytometry, visualizing and quantifying >30 markers simultaneously in mouse tumor tissue sections, we studied the localization and interactions of the immune cells within and surrounding the tumors. This has highlighted the level to which tumors are regulating their microenvironment, actively excluding all potential effector cells while attracting protumoral myeloid cells, and how this is amplified as the tumors progress. We have observed differences between the various models and seen how the tumors make adaptations when they are intrinsically more immunogenic. Furthermore, we have started to address how these balances are perturbed by various immune and chemotherapies in short- and long-term experiments and preliminary data will be presented. Citation Format: Febe van Maldegem, Karishma Valand, Victoria Tsang, Edurne Mugarza, Deborah Caswell, Philip Hobson, Julian Downward. Deep immunoprofiling of mouse lung cancer models in steady state and upon drug treatment. [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 A160.
随着肺癌免疫疗法的进步，我们对肿瘤和免疫系统之间相互作用的认识明显不足。特别是在将免疫疗法与传统化疗或新型靶向治疗相结合的情况下，我们了解这些疗法对肿瘤免疫浸润的影响以及如何与免疫疗法相互抵消或协同作用是至关重要的。我们使用多重流式细胞术来描述各种自发和正交同源肺癌小鼠模型的基础肿瘤免疫浸润，具有不同水平的免疫原性。在小鼠肿瘤组织切片中，我们采用成像细胞术，同时对30多个标记物进行可视化和定量，研究了肿瘤内和肿瘤周围免疫细胞的定位和相互作用。这突出了肿瘤调节其微环境的水平，积极排除所有潜在的效应细胞，同时吸引原瘤髓样细胞，以及随着肿瘤的进展，这种调节如何被放大。我们观察了不同模型之间的差异，并观察了肿瘤在本质上更具免疫原性时是如何适应的。此外，我们已经开始在短期和长期实验中解决各种免疫和化疗如何扰乱这些平衡，并将提供初步数据。引文格式:Febe van Maldegem, Karishma Valand, Victoria Tsang, Edurne Mugarza, Deborah Caswell, Philip Hobson, Julian Downward。稳态和药物治疗小鼠肺癌模型的深度免疫谱分析。[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;癌症免疫学杂志，2019;7(2增刊):摘要nr A160。
{"title":"Abstract A160: Deep immunoprofiling of mouse lung cancer models in steady state and upon drug treatment.","authors":"F. V. Maldegem, Karishma Valand, V. Tsang, E. Mugarza, D. Caswell, P. Hobson, J. Downward","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A160","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A160","url":null,"abstract":"With the advancement of immunotherapies for lung cancer, it has become apparent how insufficient our knowledge is surrounding the interactions between the tumor and the immune system. In particular in the context of combining immunotherapy with conventional chemotherapy or novel targeted therapies, it is of the utmost importance that we understand the effects that those therapies have on the tumor immune infiltrate and how that would counteract or synergize with immunotherapy. We have used multiplex flow cytometry to characterise the basal tumor immune infiltrate of various spontaneous and orthogonous syngeneic mouse models for lung cancer, with different levels of immunogenicity. With imaging mass cytometry, visualizing and quantifying >30 markers simultaneously in mouse tumor tissue sections, we studied the localization and interactions of the immune cells within and surrounding the tumors. This has highlighted the level to which tumors are regulating their microenvironment, actively excluding all potential effector cells while attracting protumoral myeloid cells, and how this is amplified as the tumors progress. We have observed differences between the various models and seen how the tumors make adaptations when they are intrinsically more immunogenic. Furthermore, we have started to address how these balances are perturbed by various immune and chemotherapies in short- and long-term experiments and preliminary data will be presented. Citation Format: Febe van Maldegem, Karishma Valand, Victoria Tsang, Edurne Mugarza, Deborah Caswell, Philip Hobson, Julian Downward. Deep immunoprofiling of mouse lung cancer models in steady state and upon drug treatment. [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 A160.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"259 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77139508","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A157
J. Tan, Jie Chen, E. Ginn, D. Ashok, A. Anderson, J. Banuelos, Kristen Zhang, Amber T. Pham, Tim Park, Ada Chen, Xiaoning Zhao, K. Lawson, J. Jeffreys, J. Kalisiak, M. Leleti, M. Walters, J. Powers
Introduction: Extracellular adenosine (ADO), present at high concentrations in the tumor microenvironment (TME), suppresses immune function via inhibition of T-cell, natural killer (NK) cell, and dendritic cell (DC) activation. Intratumoral generation of ADO depends on the sequential catabolism of ATP by two ecto-nucleotidases CD39 (ATP→AMP) and CD73 (AMP→ADO). Inhibition of CD73 eliminates a major pathway of ADO production in the TME and can reverse ADO-mediated immune suppression. Here we present the preclinical characterization of AB680, a novel, highly potent, reversible and selective small-molecule inhibitor of CD73, currently in preclinical development as a potential antitumor agent. The link between CD73 levels present in different tissues, efficacy in mouse tumor models, plasma and tumor exposure, and projected human pharmacokinetic (PK) profile can be combined to provide an expected AB680 dosing strategy for the upcoming first-in-human clinical trial. Methods: The potency of AB680 against human CD73 was determined using CHO-CD73 cells, blood CD8+ T-cells, recombinant human CD73, and serum/plasma using either malachite green assay, AMP-Glo assay, or LCMS/MS. The selectivity of AB680 against related ecto-nucleotidases was also assessed using similar methods. Quantitation of soluble CD73 in mouse and human serum, and mouse tumor homogenates, was performed via in-house developed and validated ELISA or Western blot methods. Syngeneic mouse tumor models were established to assess the efficacy of AB680 at multiple doses. AB680 levels in plasma and tumor associated with each dosing regimen were determined via LCMS/MS. The potency of AB680 in an intratumoral setting was determined using various biochemical methods. The effects of AB680 on syngeneic tumor volumes were assessed in prophylactic and therapeutic settings. The PK properties of AB680 were evaluated in multiple preclinical species and a projected human dosing schedule for AB680 was determined via allometric scaling. Results: AB680 is a highly potent, reversible and selective inhibitor of CD73 activity (IC50 Citation Format: Joanne B.L. Tan, Jie Chen, Elaine Ginn, Devika Ashok, Amy E Anderson, Jesus Banuelos, Kristen Zhang, Amber Pham, Timothy Park, Ada Chen, Xiaoning Zhao, Kenneth K.V. Lawson, Jenna Jeffreys, Jarek Kalisiak, Manmohan R. Leleti, Matthew J. Walters, Jay P. Powers. Preclinical pharmacokinetic and pharmacodynamic characterization of AB680, a small-molecule CD73 inhibitor for cancer immunotherapy [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 A157.
细胞外腺苷(ADO)，高浓度存在于肿瘤微环境(TME)中，通过抑制t细胞、自然杀伤细胞(NK)细胞和树突状细胞(DC)的激活来抑制免疫功能。肿瘤内ADO的生成依赖于两种外核苷酸酶CD39 (ATP→AMP)和CD73 (AMP→ADO)对ATP的顺序分解代谢。CD73的抑制消除了TME中ADO产生的主要途径，并可以逆转ADO介导的免疫抑制。AB680是一种新型的、高效的、可逆的、选择性的CD73小分子抑制剂，目前作为一种潜在的抗肿瘤药物处于临床前开发阶段。不同组织中存在的CD73水平、小鼠肿瘤模型中的疗效、血浆和肿瘤暴露以及预计的人体药代动力学(PK)谱之间的联系可以结合起来，为即将到来的首次人体临床试验提供预期的AB680给药策略。方法:利用CHO-CD73细胞、血CD8+ t细胞、重组人CD73和血清/血浆，采用孔雀石绿法、AMP-Glo法或LCMS/MS法测定AB680对人CD73的效价。AB680对相关外核苷酸酶的选择性也用类似的方法进行了评估。通过自行开发和验证的ELISA或Western blot方法，定量小鼠和人血清以及小鼠肿瘤匀浆中的可溶性CD73。建立小鼠同基因肿瘤模型，评估AB680多剂量的疗效。通过LCMS/MS检测与各给药方案相关的血浆和肿瘤中AB680的水平。采用多种生化方法测定AB680在肿瘤内的效价。在预防和治疗环境下评估AB680对同基因肿瘤体积的影响。在多个临床前物种中评估了AB680的PK特性，并通过异速计量确定了AB680的预计人体给药计划。结果:AB680是一种高效、可逆和选择性的CD73活性抑制剂(IC50引用形式:Joanne B.L. Tan, Jie Chen, Elaine Ginn, Devika Ashok, Amy E Anderson, Jesus Banuelos, Kristen Zhang, Amber Pham, Timothy Park, Ada Chen, Xiaoning Zhao, Kenneth K.V. Lawson, Jenna Jeffreys, Jarek Kalisiak, Manmohan R. Leleti, Matthew J. Walters, Jay P. Powers)。肿瘤免疫治疗小分子CD73抑制剂AB680的临床前药代动力学和药效学表征[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 A157。
{"title":"Abstract A157: Preclinical pharmacokinetic and pharmacodynamic characterization of AB680, a small-molecule CD73 inhibitor for cancer immunotherapy","authors":"J. Tan, Jie Chen, E. Ginn, D. Ashok, A. Anderson, J. Banuelos, Kristen Zhang, Amber T. Pham, Tim Park, Ada Chen, Xiaoning Zhao, K. Lawson, J. Jeffreys, J. Kalisiak, M. Leleti, M. Walters, J. Powers","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A157","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A157","url":null,"abstract":"Introduction: Extracellular adenosine (ADO), present at high concentrations in the tumor microenvironment (TME), suppresses immune function via inhibition of T-cell, natural killer (NK) cell, and dendritic cell (DC) activation. Intratumoral generation of ADO depends on the sequential catabolism of ATP by two ecto-nucleotidases CD39 (ATP→AMP) and CD73 (AMP→ADO). Inhibition of CD73 eliminates a major pathway of ADO production in the TME and can reverse ADO-mediated immune suppression. Here we present the preclinical characterization of AB680, a novel, highly potent, reversible and selective small-molecule inhibitor of CD73, currently in preclinical development as a potential antitumor agent. The link between CD73 levels present in different tissues, efficacy in mouse tumor models, plasma and tumor exposure, and projected human pharmacokinetic (PK) profile can be combined to provide an expected AB680 dosing strategy for the upcoming first-in-human clinical trial. Methods: The potency of AB680 against human CD73 was determined using CHO-CD73 cells, blood CD8+ T-cells, recombinant human CD73, and serum/plasma using either malachite green assay, AMP-Glo assay, or LCMS/MS. The selectivity of AB680 against related ecto-nucleotidases was also assessed using similar methods. Quantitation of soluble CD73 in mouse and human serum, and mouse tumor homogenates, was performed via in-house developed and validated ELISA or Western blot methods. Syngeneic mouse tumor models were established to assess the efficacy of AB680 at multiple doses. AB680 levels in plasma and tumor associated with each dosing regimen were determined via LCMS/MS. The potency of AB680 in an intratumoral setting was determined using various biochemical methods. The effects of AB680 on syngeneic tumor volumes were assessed in prophylactic and therapeutic settings. The PK properties of AB680 were evaluated in multiple preclinical species and a projected human dosing schedule for AB680 was determined via allometric scaling. Results: AB680 is a highly potent, reversible and selective inhibitor of CD73 activity (IC50 Citation Format: Joanne B.L. Tan, Jie Chen, Elaine Ginn, Devika Ashok, Amy E Anderson, Jesus Banuelos, Kristen Zhang, Amber Pham, Timothy Park, Ada Chen, Xiaoning Zhao, Kenneth K.V. Lawson, Jenna Jeffreys, Jarek Kalisiak, Manmohan R. Leleti, Matthew J. Walters, Jay P. Powers. Preclinical pharmacokinetic and pharmacodynamic characterization of AB680, a small-molecule CD73 inhibitor for cancer immunotherapy [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 A157.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77953195","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A135
Sarra Idri, G. Pawelec, Y. Barnett, G. Pockley
In addition to being less toxic and having a lower impact on a patient’s quality of life, evidence suggests that low-dose or metronomic chemotherapy modulates adaptive and innate antitumor immune responses. In this study, we examined whether the treatment of breast cancer cells and animals bearing breast cancer cell-derived tumors with low, nontoxic doses of chemotherapies promotes sensitivity to natural killer (NK) cells, based on target cell killing in vitro and the control of tumor growth and metastasis in vivo. We have developed a low-dose doxorubicin treatment protocol that sensitizes triple-negative breast cancer (TNBC) cells (MDA-MB-231, MDA-MB-468) to killing by NK cells. Specifically, low-dose doxorubicin treatment induced a “senescent-like” state in breast cancer cells concomitant with an impaired proliferative capacity, as shown by a decreased expression of Ki-67. Treatment of breast cancer cells also upregulated their expression of ligands for activatory NK cell receptors (e.g., MICA/B, ULBP1, ULBP2, ULBP3) and markedly increased their sensitivity to lysis by donor-derived, primary NK cells and the NK-92 cell line, as assessed using an in vitro flow cytometry-based assay. The cytotoxicity of donor-derived NK cells was markedly increased by prior stimulation with IL-2. The therapeutic potential of low-dose chemotherapy alone and in combination with adoptively transferred resting and activated NK cells was evaluated using immunodeficient NSG mice bearing MDA-MB-231/RFP/LUC-derived human TNBC xenografts. Although low-dose doxorubicin treatment alone reduced the growth of the primary tumors, as assessed using caliper measurements and in vivo imaging, the growth of the primary tumor was more markedly reduced following the adoptive transfer of healthy-donor derived NK cells (mean tumor volume: 70.56mm3 versus 228.8mm3 in control groups at day 49). Importantly, although treatment with low-dose doxorubicin alone also delayed the onset of metastasis by 7 days (day 49 in control animals, day 56 in treated animals), no signs of metastasis were observed in healthy-donor derived NK cell-treated animals at the time of culling on day 50. The expression of MICA/B was 8.5 times greater on tumor cells derived from mice that received the combination treatment (p=0.0001) and these cells were more sensitive to killing by NK-92 cells in vitro (3-fold increase at E:T 1:5). Furthermore, in vitro studies demonstrating that the treatment of MDA-MB-231 cells with low-dose doxorubicin reduces the number of CD44High/CD24−/low/EpCAM+ Cancer Initiating Cells (CICs) were confirmed by there being a significantly lower number of CICs in tumors derived from treated mice. These novel findings indicate that this approach has the capacity of preventing the primary tumor “fueling” the dissemination of aggressive, metastatic disease. Taken together, these studies demonstrate that the treatment of breast cancer with low, nontoxic doses of chemotherapies promotes their sensit
除了毒性更小、对患者生活质量的影响更小之外，有证据表明，低剂量或节奏化疗可以调节适应性和先天抗肿瘤免疫反应。在这项研究中，我们基于体外靶细胞杀伤和体内肿瘤生长和转移的控制，研究了用低剂量、无毒的化疗治疗乳腺癌细胞和携带乳腺癌细胞源性肿瘤的动物是否促进了对自然杀伤(NK)细胞的敏感性。我们已经开发了一种低剂量的阿霉素治疗方案，使三阴性乳腺癌(TNBC)细胞(MDA-MB-231, MDA-MB-468)对NK细胞的杀伤变得敏感。具体来说，低剂量阿霉素治疗在乳腺癌细胞中诱导了“衰老样”状态，并伴有增殖能力受损，这可以通过Ki-67的表达降低来证明。通过体外流式细胞术检测，乳腺癌细胞的治疗也上调了激活NK细胞受体的配体的表达(如MICA/B、ULBP1、ULBP2、ULBP3)，并显著提高了它们对供体来源的原代NK细胞和NK-92细胞系裂解的敏感性。供体NK细胞的细胞毒性在IL-2刺激下明显增加。采用MDA-MB-231/RFP/ luc来源的人类TNBC异种移植物，对免疫缺陷NSG小鼠进行了低剂量化疗单独和联合过继转移静息和活化NK细胞的治疗潜力进行了评估。尽管单独使用低剂量阿霉素治疗可以减少原发肿瘤的生长，但通过卡尺测量和体内成像评估，原发肿瘤的生长在健康供体来源的NK细胞的继代转移后更显着减少(第49天，对照组的平均肿瘤体积为70.56mm3，而对照组为228.8mm3)。重要的是，尽管单独使用低剂量阿霉素治疗也将转移的发生推迟了7天(对照组动物第49天，治疗动物第56天)，但在第50天剔除时，在健康供体NK细胞治疗的动物中未观察到转移的迹象。MICA/B在接受联合治疗的小鼠肿瘤细胞中的表达增加了8.5倍(p=0.0001)，并且这些细胞对NK-92细胞的体外杀伤更敏感(E:T: 1:5时增加了3倍)。此外，体外研究表明，用低剂量阿霉素处理MDA-MB-231细胞可以减少cd44高/CD24−/低/EpCAM+癌症起始细胞(CICs)的数量，并证实了处理小鼠肿瘤中CICs的数量显著减少。这些新发现表明，这种方法有能力防止原发肿瘤“助长”侵袭性转移性疾病的传播。综上所述，这些研究表明，在体外和体内，用低剂量、无毒的化疗治疗乳腺癌，可促进乳腺癌对NK细胞杀伤的敏感性。我们已经证明，将人类NK细胞给予患有人类乳腺肿瘤的小鼠，这些肿瘤已经接受了低剂量阿霉素治疗，可以减少肿瘤的生长和转移，并消除“喂养”这一过程的侵袭性细胞。正在进行的研究正在进一步探讨这种方法对乳腺癌转移的影响。总之，该策略为更有效地治疗侵袭性三阴性乳腺癌患者提供了一个有希望的机会。引文格式:Sarra Idri, Graham Pawelec, Yvonne Barnett, Graham Pockley。低剂量化疗联合NK细胞免疫疗法治疗三阴性乳腺癌[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 A135。
{"title":"Abstract A135: Combining low-dose chemotherapy with an NK cell-based immunotherapy as a treatment for triple-negative breast cancer","authors":"Sarra Idri, G. Pawelec, Y. Barnett, G. Pockley","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A135","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A135","url":null,"abstract":"In addition to being less toxic and having a lower impact on a patient’s quality of life, evidence suggests that low-dose or metronomic chemotherapy modulates adaptive and innate antitumor immune responses. In this study, we examined whether the treatment of breast cancer cells and animals bearing breast cancer cell-derived tumors with low, nontoxic doses of chemotherapies promotes sensitivity to natural killer (NK) cells, based on target cell killing in vitro and the control of tumor growth and metastasis in vivo. We have developed a low-dose doxorubicin treatment protocol that sensitizes triple-negative breast cancer (TNBC) cells (MDA-MB-231, MDA-MB-468) to killing by NK cells. Specifically, low-dose doxorubicin treatment induced a “senescent-like” state in breast cancer cells concomitant with an impaired proliferative capacity, as shown by a decreased expression of Ki-67. Treatment of breast cancer cells also upregulated their expression of ligands for activatory NK cell receptors (e.g., MICA/B, ULBP1, ULBP2, ULBP3) and markedly increased their sensitivity to lysis by donor-derived, primary NK cells and the NK-92 cell line, as assessed using an in vitro flow cytometry-based assay. The cytotoxicity of donor-derived NK cells was markedly increased by prior stimulation with IL-2. The therapeutic potential of low-dose chemotherapy alone and in combination with adoptively transferred resting and activated NK cells was evaluated using immunodeficient NSG mice bearing MDA-MB-231/RFP/LUC-derived human TNBC xenografts. Although low-dose doxorubicin treatment alone reduced the growth of the primary tumors, as assessed using caliper measurements and in vivo imaging, the growth of the primary tumor was more markedly reduced following the adoptive transfer of healthy-donor derived NK cells (mean tumor volume: 70.56mm3 versus 228.8mm3 in control groups at day 49). Importantly, although treatment with low-dose doxorubicin alone also delayed the onset of metastasis by 7 days (day 49 in control animals, day 56 in treated animals), no signs of metastasis were observed in healthy-donor derived NK cell-treated animals at the time of culling on day 50. The expression of MICA/B was 8.5 times greater on tumor cells derived from mice that received the combination treatment (p=0.0001) and these cells were more sensitive to killing by NK-92 cells in vitro (3-fold increase at E:T 1:5). Furthermore, in vitro studies demonstrating that the treatment of MDA-MB-231 cells with low-dose doxorubicin reduces the number of CD44High/CD24−/low/EpCAM+ Cancer Initiating Cells (CICs) were confirmed by there being a significantly lower number of CICs in tumors derived from treated mice. These novel findings indicate that this approach has the capacity of preventing the primary tumor “fueling” the dissemination of aggressive, metastatic disease. Taken together, these studies demonstrate that the treatment of breast cancer with low, nontoxic doses of chemotherapies promotes their sensit","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"166 4 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75519596","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A151
N. Saavedra-Ávila, Shalu Sharma, Christopher T. Johndrow, Tony W. Ng, C. Gravekamp, S. Porcelli
Listeria monocytogenes (Lm) is a gram-positive bacterium associated with gastrointestinal infections. Initially, infects epithelial cells in the small intestine, then migrates to other organs where it is taken up mainly by phagocytic cells. Once phagocytized, Lm may then be processed in the phagosome to generate peptides presented on MHC Class II for activation of Lm-specific CD4+ T-cell responses. Lm can also escape the phagosome, leading to secretion of protein antigens into the cytosol. These are processed and presented by MHC Class I molecules to CD8+ T-cells, generating strong protective immunity that can clear the infection and prevent subsequent reinfection. Lm has been exploited for its use as a vaccine vector, particularly for the delivery of tumor-associated antigens. Our previous work showed that the efficacy of Lm as an anti-umor therapeutic vaccine could be significantly improved by incorporation of synthetic glycolipid activators of CD1d-restricted invariant natural killer T-cells (iNKT-cells) into the bacteria. Using an attenuated Lm strain expressing the tumor-associated antigen Mage-B, this approach gave improvements in prevention of metastasis in the 4T1 breast carcinoma model in BALB/c mice, and also gave modest reductions in the size of primary tumors. Currently, our efforts are directed at further improving this approach to more effectively target primary tumors, and also at validating the effects in a more humanized experimental models. To this end, we are using two models: 1. human CD1d knock-in (hCD1dKI) mouse line in which the mouse CD1d protein has been replaced by human CD1d, and 2. VaKI mouse, which is a new model made by our lab that is a hCD1dKI mouse that has as a transgene the α-chain of the human iNKT TCR (Vα24Jα18) and is deficient of endogenous murine iNKT-cells due to a murine Jα18 KO. These mice models have diminished percentages of iNKT-cells closely resembling what is seen in most humans, and thus mount iNKT-cell responses that are comparable to those seen in humans. We found that incorporation of iNKT-cell activating glycolipids into Lm leads to improved iNKT-cell-dependent immune responses in our models, and induces synergistic antitumor immune mechanisms against implanted primary syngeneic tumors (B16 F10 and MC38). Besides, administration of iNKT-cell activating glycolipids in association with Lm did not induce anergy or exhaustion of the responding cells, thus enabling repeated treatment to boost antitumor immune responses. Citation Format: Noemi Alejandra Saavedra-Avila, Shalu Sharma, Christopher T. Johndrow, Tony Ng, Claudia Gravekamp, Steven A. Porcelli. Immunotherapy against cancer using iNKT ligands in combination with an attenuated Listeria monocytogenes in humanized mouse models [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
单核增生李斯特菌(Lm)是一种与胃肠道感染相关的革兰氏阳性细菌。最初感染小肠上皮细胞，然后转移到其他器官，主要被吞噬细胞吸收。一旦被吞噬，Lm可能会在吞噬体中被加工，产生在MHC II类上呈现的肽，以激活Lm特异性CD4+ t细胞反应。Lm也可以逃离吞噬体，导致蛋白抗原分泌到细胞质中。这些被MHC I类分子处理并提呈给CD8+ t细胞，产生强大的保护性免疫，可以清除感染并防止随后的再感染。Lm已被用作疫苗载体，特别是用于递送肿瘤相关抗原。我们之前的工作表明，通过将cd1限制性不变性自然杀伤t细胞(inkt细胞)的合成糖脂激活剂掺入细菌中，可以显著提高Lm作为抗肿瘤治疗疫苗的功效。使用表达肿瘤相关抗原Mage-B的减毒Lm菌株，该方法在BALB/c小鼠4T1乳腺癌模型中改善了转移的预防，并且还适度减少了原发肿瘤的大小。目前，我们正在努力进一步改进这种方法，以更有效地靶向原发性肿瘤，并在更人性化的实验模型中验证其效果。为此，我们使用了两个模型:1。人类CD1d敲入(hCD1dKI)小鼠系，其中小鼠CD1d蛋白已被人类CD1d取代;VaKI小鼠是我们实验室新建立的hCD1dKI小鼠模型，该模型具有人iNKT TCR (Vα24Jα18) α-链的转基因，由于小鼠Jα18 KO导致内源性小鼠iNKT细胞缺失。这些小鼠模型减少了与大多数人类相似的inkt细胞百分比，因此产生了与人类相似的inkt细胞反应。我们发现，在我们的模型中，将inkt细胞激活糖脂掺入Lm可改善inkt细胞依赖的免疫反应，并诱导针对植入式原发性同基因肿瘤(B16 F10和MC38)的协同抗肿瘤免疫机制。此外，给药inkt细胞激活与Lm相关的糖脂不会诱导应答细胞的能量或衰竭，因此可以重复治疗以增强抗肿瘤免疫反应。引文格式:Noemi Alejandra Saavedra-Avila, Shalu Sharma, Christopher T. Johndrow, Tony Ng, Claudia Gravekamp, Steven A. Porcelli。在人源化小鼠模型中使用iNKT配体联合减毒单核细胞增生李斯特菌进行抗癌免疫治疗[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 A151。
{"title":"Abstract A151: Immunotherapy against cancer using iNKT ligands in combination with an attenuated Listeria monocytogenes in humanized mouse models","authors":"N. Saavedra-Ávila, Shalu Sharma, Christopher T. Johndrow, Tony W. Ng, C. Gravekamp, S. Porcelli","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A151","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A151","url":null,"abstract":"Listeria monocytogenes (Lm) is a gram-positive bacterium associated with gastrointestinal infections. Initially, infects epithelial cells in the small intestine, then migrates to other organs where it is taken up mainly by phagocytic cells. Once phagocytized, Lm may then be processed in the phagosome to generate peptides presented on MHC Class II for activation of Lm-specific CD4+ T-cell responses. Lm can also escape the phagosome, leading to secretion of protein antigens into the cytosol. These are processed and presented by MHC Class I molecules to CD8+ T-cells, generating strong protective immunity that can clear the infection and prevent subsequent reinfection. Lm has been exploited for its use as a vaccine vector, particularly for the delivery of tumor-associated antigens. Our previous work showed that the efficacy of Lm as an anti-umor therapeutic vaccine could be significantly improved by incorporation of synthetic glycolipid activators of CD1d-restricted invariant natural killer T-cells (iNKT-cells) into the bacteria. Using an attenuated Lm strain expressing the tumor-associated antigen Mage-B, this approach gave improvements in prevention of metastasis in the 4T1 breast carcinoma model in BALB/c mice, and also gave modest reductions in the size of primary tumors. Currently, our efforts are directed at further improving this approach to more effectively target primary tumors, and also at validating the effects in a more humanized experimental models. To this end, we are using two models: 1. human CD1d knock-in (hCD1dKI) mouse line in which the mouse CD1d protein has been replaced by human CD1d, and 2. VaKI mouse, which is a new model made by our lab that is a hCD1dKI mouse that has as a transgene the α-chain of the human iNKT TCR (Vα24Jα18) and is deficient of endogenous murine iNKT-cells due to a murine Jα18 KO. These mice models have diminished percentages of iNKT-cells closely resembling what is seen in most humans, and thus mount iNKT-cell responses that are comparable to those seen in humans. We found that incorporation of iNKT-cell activating glycolipids into Lm leads to improved iNKT-cell-dependent immune responses in our models, and induces synergistic antitumor immune mechanisms against implanted primary syngeneic tumors (B16 F10 and MC38). Besides, administration of iNKT-cell activating glycolipids in association with Lm did not induce anergy or exhaustion of the responding cells, thus enabling repeated treatment to boost antitumor immune responses. Citation Format: Noemi Alejandra Saavedra-Avila, Shalu Sharma, Christopher T. Johndrow, Tony Ng, Claudia Gravekamp, Steven A. Porcelli. Immunotherapy against cancer using iNKT ligands in combination with an attenuated Listeria monocytogenes in humanized mouse models [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","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"95 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83695308","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A158
J. Tavernier
An ideal generic cancer immunotherapy should mobilize the immune system to destroy tumor cells without harming healthy cells and remain active in case of recurrence. Furthermore, it should preferably not rely on tumor-specific surface markers, as these are only available in a limited set of malignancies. Despite approval for treatment of various cancers, clinical application of cytokines is still impeded by their multiple toxic side effects. Type I interferon (IFN), for instance, has a long history in the treatment of cancer, but its multifaceted activity pattern and complex side effects prevent its optimal clinical use. Here we develop AcTakines (Activity-on-Target cytokines), optimized (mutated) immunocytokines that are up to 1000-fold more potent on targeT-cells, allowing specific signaling in selected cell types only. As conventional dendritic cells (cDC) are essential for IFN antitumor efficacy, we targeted type I IFN-derived “AcTaferon (AFN)” to Clec9A+ cDC. Clec9A-AFN therapy displayed strong antitumor activity in murine melanoma (B16), breast carcinoma (4T1) and lymphoma models (A20), as well as against human RL lymphoma in immunodeficient NSG mice reconstituted with a human immune system. In sharp contrast to wild-type IFN therapy, the antitumor efficacy of Clec9A-AFN was not accompanied by any detectable toxicity, assessed by body weight and several hematologic parameters. Clec9A-AFN effects were lost in CD8-depleted or Batf3-/- mice, and depended on IFN signaling in cDCs but not in T lymphocytes. Combined with anti-PDL1 immune checkpoint blockade, Treg-depleting anti-CTLA4 + anti-OX40 therapy, immunogenic chemotherapy, or low-dose TNF, complete tumor regressions and long-lasting tumor immunity (memory) were obtained, still without any adverse effects. Our findings thus indicate that DC-targeted AFN provides a highly efficient, off-the-shelf and safe cancer immunotherapy, with possible application in a broad range of malignancies. Citation Format: Jan H. Tavernier. Delivering type I interferon to dendritic cells empowers tumor eradication and immune combination treatments [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 A158.
一种理想的通用癌症免疫疗法应该动员免疫系统在不伤害健康细胞的情况下摧毁肿瘤细胞，并在复发的情况下保持活性。此外，它最好不依赖于肿瘤特异性表面标志物，因为这些仅在有限的恶性肿瘤中可用。尽管已被批准用于治疗多种癌症，但细胞因子的临床应用仍然受到其多重毒副作用的阻碍。例如，I型干扰素(IFN)在治疗癌症方面有着悠久的历史，但其多方面的活性模式和复杂的副作用阻碍了其最佳临床应用。在这里，我们开发了AcTakines(活性靶细胞因子)，优化(突变)免疫细胞因子，其对靶细胞的效力高达1000倍，仅在选定的细胞类型中允许特定的信号传导。由于传统的树突状细胞(cDC)对IFN的抗肿瘤作用至关重要，我们将I型IFN衍生的“AcTaferon (AFN)”靶向Clec9A+ cDC。Clec9A-AFN治疗在小鼠黑色素瘤(B16)、乳腺癌(4T1)和淋巴瘤模型(A20)中显示出较强的抗肿瘤活性，在用人类免疫系统重建的免疫缺陷NSG小鼠中也显示出较强的抗人RL淋巴瘤活性。与野生型IFN治疗形成鲜明对比的是，通过体重和几个血液学参数评估，Clec9A-AFN的抗肿瘤疗效没有任何可检测到的毒性。在cd8缺失或Batf3-/-小鼠中，Clec9A-AFN的作用消失，并且依赖于cDCs中的IFN信号，而不依赖于T淋巴细胞。联合抗pdl1免疫检查点阻断、treg消耗抗ctla4 +抗ox40治疗、免疫原性化疗或低剂量TNF，获得肿瘤完全消退和持久的肿瘤免疫(记忆)，仍无任何不良反应。因此，我们的研究结果表明，针对dc的AFN提供了一种高效、现成和安全的癌症免疫疗法，可能应用于广泛的恶性肿瘤。引用格式:Jan H. Tavernier。将I型干扰素输送到树突状细胞使肿瘤根除和免疫联合治疗成为可能[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;癌症免疫学杂志，2019;7(2增刊):摘要nr A158。
{"title":"Abstract A158: Delivering type I interferon to dendritic cells empowers tumor eradication and immune combination treatments","authors":"J. Tavernier","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A158","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A158","url":null,"abstract":"An ideal generic cancer immunotherapy should mobilize the immune system to destroy tumor cells without harming healthy cells and remain active in case of recurrence. Furthermore, it should preferably not rely on tumor-specific surface markers, as these are only available in a limited set of malignancies. Despite approval for treatment of various cancers, clinical application of cytokines is still impeded by their multiple toxic side effects. Type I interferon (IFN), for instance, has a long history in the treatment of cancer, but its multifaceted activity pattern and complex side effects prevent its optimal clinical use. Here we develop AcTakines (Activity-on-Target cytokines), optimized (mutated) immunocytokines that are up to 1000-fold more potent on targeT-cells, allowing specific signaling in selected cell types only. As conventional dendritic cells (cDC) are essential for IFN antitumor efficacy, we targeted type I IFN-derived “AcTaferon (AFN)” to Clec9A+ cDC. Clec9A-AFN therapy displayed strong antitumor activity in murine melanoma (B16), breast carcinoma (4T1) and lymphoma models (A20), as well as against human RL lymphoma in immunodeficient NSG mice reconstituted with a human immune system. In sharp contrast to wild-type IFN therapy, the antitumor efficacy of Clec9A-AFN was not accompanied by any detectable toxicity, assessed by body weight and several hematologic parameters. Clec9A-AFN effects were lost in CD8-depleted or Batf3-/- mice, and depended on IFN signaling in cDCs but not in T lymphocytes. Combined with anti-PDL1 immune checkpoint blockade, Treg-depleting anti-CTLA4 + anti-OX40 therapy, immunogenic chemotherapy, or low-dose TNF, complete tumor regressions and long-lasting tumor immunity (memory) were obtained, still without any adverse effects. Our findings thus indicate that DC-targeted AFN provides a highly efficient, off-the-shelf and safe cancer immunotherapy, with possible application in a broad range of malignancies. Citation Format: Jan H. Tavernier. Delivering type I interferon to dendritic cells empowers tumor eradication and immune combination treatments [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 A158.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"65 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83447762","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-IA18
A. Bardelli
When metastatic cancers are challenged with targeted agents almost invariably a subset of cells insensitive to the drug emerges. As a result, in most instances, targeted therapies are only transiently effective in patients. Strategies to prevent or overcome resistance are therefore essential to design the next generation of clinical trials. How can we overcome the near-certainty of disease recurrence following treatment with targeted agents? Addressing this question means considering as a target not “only” individual oncogenes but also the evolving nature of human tumors. We used colorectal cancer (CRC) as a model system to test the hypothesis that by understanding tumor’s evolution, the emergence of drug resistance can be controlled. We find that to have long-term efficacy, the use of targeted therapies must take into account the continuous evolution of cancer cells, that is to say, therapies must adapt to tumor evolution. Another approach is to unleash the ability of the immune system to recognize drug resistant cells. We tested this possibility in syngeneic mouse models of CRC sensitive to targeted therapies. Our findings indicate that inactivation of DNA repair mechanisms and manipulation of mutational loads can trigger immune surveillance and prolonged therapeutic responses. We postulate that rationally combined targeted and immuno-therapies can restrain tumor evolution, and can limit the emergence of drug resistance thus leading to long-term responses. Citation Format: Alberto Bardelli. Inactivation of DNA repair to improve immune surveillance [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 IA18.
{"title":"Abstract IA18: Inactivation of DNA repair to improve immune surveillance","authors":"A. Bardelli","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-IA18","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-IA18","url":null,"abstract":"When metastatic cancers are challenged with targeted agents almost invariably a subset of cells insensitive to the drug emerges. As a result, in most instances, targeted therapies are only transiently effective in patients. Strategies to prevent or overcome resistance are therefore essential to design the next generation of clinical trials. How can we overcome the near-certainty of disease recurrence following treatment with targeted agents? Addressing this question means considering as a target not “only” individual oncogenes but also the evolving nature of human tumors. We used colorectal cancer (CRC) as a model system to test the hypothesis that by understanding tumor’s evolution, the emergence of drug resistance can be controlled. We find that to have long-term efficacy, the use of targeted therapies must take into account the continuous evolution of cancer cells, that is to say, therapies must adapt to tumor evolution. Another approach is to unleash the ability of the immune system to recognize drug resistant cells. We tested this possibility in syngeneic mouse models of CRC sensitive to targeted therapies. Our findings indicate that inactivation of DNA repair mechanisms and manipulation of mutational loads can trigger immune surveillance and prolonged therapeutic responses. We postulate that rationally combined targeted and immuno-therapies can restrain tumor evolution, and can limit the emergence of drug resistance thus leading to long-term responses. Citation Format: Alberto Bardelli. Inactivation of DNA repair to improve immune surveillance [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 IA18.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"109 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77636845","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A144
A. Mackiewicz, Partycja Czerwinska, M. Ruciński, K. Gryska, Iga Grządzielewska, A. Jaworska, J. Mackiewicz
Melanoma (MM) cells leave primary tumors early and evolve at different sites in parallel. However, timing in human MM is unknown. Lymphatic dissemination is fast shortly after the dermal invasion, but metastases may occur late. Such delay in metastases formation suggests possible host defense mechanism, or particular nature of non-actively proliferative (dormant) MM stem cell (MSC) population. T lymphocytes may also induce tumor dormancy. The proposed mechanism involves a cytostatic mode of tumor silencing resulting from the suppression of both neoplastic cell proliferation and cell cycle progression. Significant progress in the development of novel advanced MM therapies was made. However, not all patients benefit from the therapy, developing late metastases decades after treatment ends. Moreover, using the immunotherapy-based model of MM dormancy in mouse, vaccination against MM prevented tumor growth but failed to prevent tumor cell dissemination or elimination of all MM cells. We have developed therapeutic gene modified allogenic MM vaccine (AGI-101H) that has been tested since 1997 and resulted in long-term survival of a substantial fraction of patients. Genetic modification of vaccine cells to express designer cytokine cDNA encoding Hyper-IL-6 protein (comprising of IL-6 linked with a soluble IL-6 receptor) has altered AGI-101H cells phenotype towards MSC-like with high activity of aldehyde dehydrogenase isoenzyme (ALDH1A1). We have found significant induction of anti-MSCs response in immunized patients by a generation of functionally active cytotoxic CD8+T-cells and antibodies specific for ALDH1A1 in circulation. However, the AGI-101H mode of action is complex: besides inducing a direct anticancer response to MSC population, long-term vaccination may induce MM dormancy through T-cell mediated immunity. The above hypothesis is based on clinical observations of AHI-101H treated patient who was progression-free for many years, but injury induced metastases formation in the bruise. This patient developed skin MM in 1999. In 2002 in transit metastases appeared (resected). The patient was enrolled into AGI-101H program. In 2003 metastases in liver (resected), 2004 metastasis in ovary (resected). 11 years (2004-2015) no progression. In Nov. 2015 she had bicycle accident, trauma, and MM metastasis presenting with a bruise (resected). The treatment continues with no progression. The aim was to determine the molecular mechanisms of peripheral T-cell-mediated induction of MM dormancy induced by vaccination. Therefore, T-cell mRNA expression profiling of long-term surviving patients was carried out. PBMCs were isolated from treated patients (18), untreated MM patients (13) and healthy controls (8). Untouched T-cells were separated by magnetic beads and total RNA isolated. The transcriptome profiling was performed with Affymetrix HG U219 microarray (19,285 markers). Differential gene expression (DGE) analysis between all groups was conducted and validat
黑色素瘤(MM)细胞早期离开原发肿瘤，并在不同部位平行进化。然而，人类MM的发病时间尚不清楚。淋巴传播在皮肤浸润后很快，但转移可能发生较晚。这种转移形成的延迟提示可能的宿主防御机制，或非活跃增殖(休眠)MM干细胞(MSC)群体的特殊性质。T淋巴细胞也可诱导肿瘤休眠。提出的机制涉及抑制肿瘤细胞增殖和细胞周期进展导致的肿瘤沉默的细胞抑制模式。新型高级MM治疗方法的开发取得了重大进展。然而，并不是所有的患者都能从治疗中获益，在治疗结束后几十年发生晚期转移。此外，在基于免疫治疗的小鼠MM休眠模型中，接种MM疫苗可以阻止肿瘤生长，但不能阻止肿瘤细胞的播散或消除所有MM细胞。我们开发了治疗性基因修饰的同种异体MM疫苗(AGI-101H)，自1997年以来一直进行测试，并导致相当一部分患者长期生存。对疫苗细胞进行基因修饰以表达编码Hyper-IL-6蛋白(包括与可溶性IL-6受体连接的IL-6)的设计细胞因子cDNA，使AGI-101H细胞表型改变为具有高活性的醛脱氢酶同工酶(ALDH1A1)的msc样细胞。我们发现，在免疫患者中，通过循环中一代功能活跃的细胞毒性CD8+ t细胞和特异性ALDH1A1抗体，可以显著诱导抗mscs反应。然而，AGI-101H的作用方式是复杂的:除了诱导对间充质干细胞群体的直接抗癌反应外，长期接种疫苗可能通过t细胞介导的免疫诱导MM休眠。上述假设是基于AHI-101H治疗患者的临床观察，该患者多年无进展，但损伤引起的瘀伤转移形成。该患者于1999年出现皮肤MM。2002年出现转移瘤(切除)。患者被纳入AGI-101H项目。2003年肝脏转移(切除)，2004年卵巢转移(切除)。11年(2004-2015)无进展。2015年11月，她发生自行车事故、外伤和MM转移，并伴有瘀伤(已切除)。治疗继续，无进展。目的是确定外周t细胞介导的疫苗诱导MM休眠的分子机制。因此，对长期存活患者进行t细胞mRNA表达谱分析。从接受治疗的患者(18例)、未接受治疗的MM患者(13例)和健康对照(8例)中分离PBMCs。用磁珠分离未受影响的t细胞并分离总RNA。转录组分析使用Affymetrix HG U219微阵列(19,285个标记)进行。采用RT-qPCR和FACS对各组进行差异基因表达(DGE)分析。利用基因集富集分析(GSEA)工具进一步分析转录组学结果。DGE分析比较了接种AGI-101H疫苗(AV)和未接种AGI-101H疫苗(C)的患者，发现538个差异表达基因(DEGs)，其中373个在AV (adj p1)中下调，165个在AV (adj p1)中上调，目前正在用RT-qPCR和FACS验证。此外，GSEA分析显示，在接种AGI-101H的患者中，“TNFa_signaling_via_NFkB”，“TGFb_signaling”和“G2/M_checkpoint”标志过程(MSigDB标志基因集)显著富集，这表明相当大的抗肿瘤反应激活和预防MM休眠细胞转化为增殖转移。免疫AGI-101H的长期存活MM患者的外周t细胞转录组分析表明，激活了维持MM细胞处于非主动增殖休眠状态的分子机制。引文格式:Andrzej A. Mackiewicz, Partycja Czerwinska, Marcin Rucinski, Katarzyna Gryska, Iga Grzadzielewska, Anna Jaworska, Jacek Mackiewicz。在接受同种异体黑色素瘤疫苗治疗的患者中，维持黑色素瘤细胞休眠状态的外周t细胞的转录组学特征[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 A144。
{"title":"Abstract A144: The transcriptomic profile of peripheral T-cells that maintain dormant state of melanoma cells in patients treated with allogenic melanoma vaccine","authors":"A. Mackiewicz, Partycja Czerwinska, M. Ruciński, K. Gryska, Iga Grządzielewska, A. Jaworska, J. Mackiewicz","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A144","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A144","url":null,"abstract":"Melanoma (MM) cells leave primary tumors early and evolve at different sites in parallel. However, timing in human MM is unknown. Lymphatic dissemination is fast shortly after the dermal invasion, but metastases may occur late. Such delay in metastases formation suggests possible host defense mechanism, or particular nature of non-actively proliferative (dormant) MM stem cell (MSC) population. T lymphocytes may also induce tumor dormancy. The proposed mechanism involves a cytostatic mode of tumor silencing resulting from the suppression of both neoplastic cell proliferation and cell cycle progression. Significant progress in the development of novel advanced MM therapies was made. However, not all patients benefit from the therapy, developing late metastases decades after treatment ends. Moreover, using the immunotherapy-based model of MM dormancy in mouse, vaccination against MM prevented tumor growth but failed to prevent tumor cell dissemination or elimination of all MM cells. We have developed therapeutic gene modified allogenic MM vaccine (AGI-101H) that has been tested since 1997 and resulted in long-term survival of a substantial fraction of patients. Genetic modification of vaccine cells to express designer cytokine cDNA encoding Hyper-IL-6 protein (comprising of IL-6 linked with a soluble IL-6 receptor) has altered AGI-101H cells phenotype towards MSC-like with high activity of aldehyde dehydrogenase isoenzyme (ALDH1A1). We have found significant induction of anti-MSCs response in immunized patients by a generation of functionally active cytotoxic CD8+T-cells and antibodies specific for ALDH1A1 in circulation. However, the AGI-101H mode of action is complex: besides inducing a direct anticancer response to MSC population, long-term vaccination may induce MM dormancy through T-cell mediated immunity. The above hypothesis is based on clinical observations of AHI-101H treated patient who was progression-free for many years, but injury induced metastases formation in the bruise. This patient developed skin MM in 1999. In 2002 in transit metastases appeared (resected). The patient was enrolled into AGI-101H program. In 2003 metastases in liver (resected), 2004 metastasis in ovary (resected). 11 years (2004-2015) no progression. In Nov. 2015 she had bicycle accident, trauma, and MM metastasis presenting with a bruise (resected). The treatment continues with no progression. The aim was to determine the molecular mechanisms of peripheral T-cell-mediated induction of MM dormancy induced by vaccination. Therefore, T-cell mRNA expression profiling of long-term surviving patients was carried out. PBMCs were isolated from treated patients (18), untreated MM patients (13) and healthy controls (8). Untouched T-cells were separated by magnetic beads and total RNA isolated. The transcriptome profiling was performed with Affymetrix HG U219 microarray (19,285 markers). Differential gene expression (DGE) analysis between all groups was conducted and validat","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"18 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"78453789","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
京公网安备 11010802042870号
京ICP备2023020795号-1
扫码关注我们
求助内容：
应助结果提醒方式：