Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A141
H. Lee, D. Oh
Tumor immunotherapy aims to overcome the immunosuppressive microenvironment within tumors, and various approaches have been developed. Tumor-associated T regulatory cells (Tregs) suppress the activation and expansion of tumor antigen-specific effector T-cells, thus providing a permissive environment for tumor growth. Therefore, optimal strategies need to be established to deplete tumor-infiltrated Tregs because systemic depletion of Tregs can result in reduced anti-tumor effector cells and autoimmunity. Here, to deplete Tregs selectively in tumors, we intratumorally injected anti-CD25 antibodies conjugated to Chlorin e6 (Ce6), a photosensitizer that absorbs light to generate reactive oxygen species. Local depletion of tumor-associated Tregs by photodynamic therapy (PDT) inhibited tumor growth, which was likely due to the altered tumor immune microenvironment that was characterized by increased infiltration of CD8+ effector T-cells and the expression of IFN-γ and CD107a, which is cytolytic granule exocytosis marker, in tumor tissues. Furthermore, PDT-induced intratumoral Treg depletion did not influence adaptive immune responses in a murine influenza infection model. Thus, our results show that intratumoral Treg-targeted PDT could specifically modulate tumor microenvironments by depleting Tregs and could be used as a novel cancer immunotherapy technique. Citation Format: Heung Kyu Lee, Dong Sun Oh. Intratumoral depletion of regulatory T-cells using CD25-targeted photodynamic therapy induces antitumoral immune responses [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 A141.
肿瘤免疫治疗的目的是克服肿瘤内的免疫抑制微环境，各种方法已经发展。肿瘤相关T调节细胞(Tregs)抑制肿瘤抗原特异性效应T细胞的激活和扩增，从而为肿瘤生长提供了一个宽松的环境。因此，需要建立最佳策略来消耗肿瘤浸润的Tregs，因为Tregs的全身消耗会导致抗肿瘤效应细胞和自身免疫的减少。在这里，为了选择性地消耗肿瘤中的Tregs，我们在瘤内注射了与氯e6 (Ce6)结合的抗cd25抗体，氯e6是一种吸收光产生活性氧的光敏剂。通过光动力疗法(PDT)局部消耗肿瘤相关Tregs抑制肿瘤生长，这可能是由于肿瘤免疫微环境的改变，其特征是CD8+效应t细胞的浸润增加以及肿瘤组织中IFN-γ和CD107a(细胞溶解颗粒胞外分泌标志物)的表达增加。此外，在小鼠流感感染模型中，pdt诱导的肿瘤内Treg耗竭并不影响适应性免疫反应。因此，我们的研究结果表明，肿瘤内靶向treg的PDT可以通过消耗treg特异性调节肿瘤微环境，并可作为一种新的癌症免疫治疗技术。引文格式:Heung Kyu Lee, Dong Sun Oh。肿瘤内使用cd25靶向光动力疗法耗竭调节性t细胞可诱导抗肿瘤免疫反应[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，2019;7(2增刊):摘要nr A141。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-IA20
S. Loi, P. Savas
Immunotherapy has revolutionized cancer treatment, but the benefits are not ubiquitous or absolute. In breast cancer, higher levels of lymphocytic infiltrate confer significant prognostic advantage, predict response to chemotherapy in early stage disease and are associated with benefit from PD-1 inhibition. To enhance the modest benefits of immunotherapy seen in breast cancer thus far, it is expected that a greater understanding of the immune subpopulations in the tumor microenvironment will be necessary, emphasizing that the quality and quantity of tumor infiltrating lymphocytes are important. We have profiled the T cell infiltrate in primary and metastatic breast cancers, using flow cytometry, single cell mRNASeq, multiplex immunohistochemistry and bulk RNASeq on flow sorted CD8 populations. In this work, we found that CD8 T cells display tissue resident specialization. In particular, we confirmed the presence of a CD8 T cell population with tissue resident memory features. This population was notable for high expression of T cell checkpoint receptors and cytotoxic markers, active proliferation, and a distinct T cell receptor repertoire in contrast to CD8 effector memory T cells. A gene signature derived from this population was found to be prognostic in early stage breast cancer and associated with benefit from PD-1 checkpoint inhibition in melanoma. We find that single cell profiling in human samples is a powerful technique which unveils tissue resident aspects of the anti-cancer immune response. This approach may lead to better immunotherapy biomarkers and optimization of immunotherapeutic strategies. Citation Format: Sherene Loi, Peter Savas. Targeting the immune microenvironment in breast cancer [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 IA20.
免疫疗法已经彻底改变了癌症治疗，但它的益处并不是普遍存在的，也不是绝对的。在乳腺癌中，较高水平的淋巴细胞浸润具有显著的预后优势，预测早期疾病对化疗的反应，并与PD-1抑制的益处相关。为了增强迄今为止在乳腺癌中看到的免疫治疗的适度益处，预计有必要对肿瘤微环境中的免疫亚群有更深入的了解，强调肿瘤浸润淋巴细胞的质量和数量是重要的。我们利用流式细胞术、单细胞mRNASeq、多重免疫组织化学和大量RNASeq分析了原发性和转移性乳腺癌中的T细胞浸润。在这项工作中，我们发现CD8 T细胞表现出组织驻留特化。特别是，我们证实了具有组织驻留记忆特征的CD8 T细胞群的存在。与CD8效应记忆T细胞相比，该群体以T细胞检查点受体和细胞毒性标记物的高表达、活跃的增殖和独特的T细胞受体库而闻名。来自该人群的一个基因标记被发现在早期乳腺癌中具有预后作用，并且与黑色素瘤中PD-1检查点抑制的益处相关。我们发现人体样本中的单细胞分析是一种强大的技术，它揭示了抗癌免疫反应的组织驻留方面。这种方法可能导致更好的免疫治疗生物标志物和免疫治疗策略的优化。引用格式:Sherene Loi, Peter Savas。靶向免疫微环境治疗乳腺癌[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;癌症免疫学杂志，2019;7(2增刊):摘要1 - 20。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A123
Cécile Alanio, B. Bengsch, J. R. Gies, S. Henrickson, N. Weng, Janáe A. Ritz-Romeo, M. O'Hara, J. Melenhorst, S. Lacey, R. Young, C. June, E. Wherry
To understand potential immune system alterations in newly diagnosed, untreated, pancreatic cancer patients and provide a foundation for immunotherapy, we profiled PBMC from pancreatic ductal adenocarcinoma (PDA) patients and age-matched healthy controls using high-dimensional CyTOF analysis. We developed two immune profiling panels: a broad profiling panel that includes 45 phenotypic markers that together permit the identification and enumeration of the main innate and adaptive immune cell subsets in humans, and a deep profiling panel that includes 45 features focusing on T-cell phenotype and biology. We report a 2-fold increase in monocytes, more regulatory T-cells, and more plasmocytes in circulation in pancreatic cancer patients as compared to age-matched controls, as well as a bias towards cytokine-producing NK cells. Using high-dimensional approaches, we observed skewed T-cell differentiation in pancreatic cancer patients, with peripheral blood CD8 T-cells being biased towards more CD45RA-positive CD27-positive CCR7-positive CD95-positive CD49d-positive stem cell memory cells (P=7x10-5), more CD45RA-negative CD27-positive CCR7-negative effector memory cells (P=0.002) and less CD45RA-positive CD27-negative CCR7-negative late effector memory cells (P=0.003) as compared to age-matched controls. Strikingly, when examinating T-cell differentiation in human spleens, we found increased proportions of late effector memory T-cells in the splenic CD8 T-cell compartment of pancreatic cancer patients as compared to age-matched controls. These results suggest an impaired T-cell trafficking in pancreatic cancer patients, with late memory T-cells being retained in the spleen. We are now investigating the mechanisms underlying these observations, as well as their impact on T-cell immunity of the cancer patients. Our goal is to understand the nature of the skewing and how any changes in baseline immune health of the T-cell compartment relate to disease progression and/or response to therapy. These studies should provide a foundation for improving therapy in pancreatic cancer patients. Citation Format: Cecile Alanio, Bertram Bengsch, Josephine R. Gies, Sarah Henrickson, Nan Ping Weng, Janae A Ritz-Romeo, Mark O9Hara, Joseph J Melenhorst, Simon Lacey, Regina M Young, Carl H June, E. John Wherry. Skewed CD4 and CD8 T-cell differentiation in pancreatic cancer patients [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 A123.
为了了解新诊断、未经治疗的胰腺癌患者潜在的免疫系统改变，并为免疫治疗提供基础，我们使用高维细胞tof分析了胰腺导管腺癌(PDA)患者和年龄匹配的健康对照者的PBMC。我们开发了两个免疫分析面板:一个广泛的分析面板，包括45个表型标记，这些标记一起允许鉴定和枚举人类主要的先天和适应性免疫细胞亚群，以及一个深度分析面板，包括45个特征，重点是t细胞表型和生物学。我们报道，与年龄匹配的对照组相比，胰腺癌患者循环中的单核细胞、调节性t细胞和浆细胞增加了2倍，并且倾向于产生细胞因子的NK细胞。使用高维方法，我们观察到胰腺癌患者的t细胞分化偏斜，与年龄匹配的对照组相比，外周血CD8 t细胞偏向于更多的cd45ra阳性cd27阳性ccr7阳性cd95阳性cd49d阳性干细胞记忆细胞(P=7x10-5)，更多的cd45ra阴性cd27阳性ccr7阴性效应记忆细胞(P=0.002)和更少的cd45ra阳性cd27阴性ccr7阴性晚期效应记忆细胞(P=0.003)。引人注目的是，当检查人类脾脏中的t细胞分化时，我们发现胰腺癌患者脾CD8 t细胞区迟效应记忆t细胞的比例比年龄匹配的对照组增加。这些结果表明胰腺癌患者的t细胞运输受损，晚期记忆t细胞保留在脾脏中。我们现在正在研究这些观察结果背后的机制，以及它们对癌症患者t细胞免疫的影响。我们的目标是了解这种倾斜的本质，以及t细胞区室基线免疫健康的任何变化与疾病进展和/或治疗反应的关系。这些研究为改善胰腺癌患者的治疗提供了基础。引文格式:Cecile Alanio, Bertram Bengsch, Josephine R. Gies, Sarah Henrickson, Nan Ping Weng, Janae A Ritz-Romeo, Mark O9Hara, Joseph J Melenhorst, Simon Lacey, Regina M Young, Carl H June, E. John Wherry。胰腺癌患者CD4和CD8 t细胞分化偏斜[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 A123。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A159
L. Hooren, A. Vaccaro, Maria Georganaki, Mohanraj Ramachandran, Hua Huang, J. Lau, A. Smits, M. Essand, A. Dimberg
Cancer immunotherapy of glioma, the most common primary brain tumor in adults, is constrained by an immunosuppressive microenvironment. Tertiary lymphoid structures (TLSs) are ectopic lymphoid structures that mediate immune responses and provide a local site for tumor antigen presentation. Here, we show for the first time that TLSs are present in human low-grade glioma and that they can be induced in murine glioma models. Agonistic CD40 antibodies induced lymphotoxin α expression in B cells and enhanced the formation of TLSs in mice bearing orthotopic GL261 or CT-2A gliomas. TLSs consisting of B cells, T-cells and antigen presenting cells formed around CD31+ vessels proximal to the tumor tissue and adjacent to the meninges. Agonistic CD40 antibody treatment improved the CD8+ T-cell / Treg ratio in tumor-bearing mice, but there was no survival benefit. A decreased CD69 activation status on CD8+ T-cells, an increase of B cells with a CD1d+CD5+ regulatory phenotype and a high infiltration of Tregs in TLSs indicate that agonistic CD40 antibodies induce an immunosuppressive microenvironment in glioma. Consistent with this, the efficacy of both PD-1 and CTLA-4 checkpoint blockade was significantly decreased by co-administration of agonistic CD40 antibodies. These results demonstrate that TLSs are present and can be induced in glioma, but that they at least under certain conditions are associated with immunosuppression, shedding light on new mechanisms of immune regulation in the brain microenvironment. Citation Format: Luuk van Hooren, Alessandra Vaccaro, Maria Georganaki, Mohanraj Ramachandran, Hua Huang, Joey Lau, Anja Smits, Magnus Essand, Anna Dimberg. Agonistic CD40 antibody therapy induces formation of tertiary lymphoid structures in glioma and inhibits the response to immune-checkpoint 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 A159.
神经胶质瘤是成人最常见的原发性脑肿瘤，其免疫治疗受到免疫抑制微环境的限制。三级淋巴样结构(TLSs)是异位淋巴样结构，介导免疫反应并提供肿瘤抗原呈递的局部部位。在这里，我们首次表明TLSs存在于人类低级别胶质瘤中，并且它们可以在小鼠胶质瘤模型中诱导。在原位GL261或CT-2A胶质瘤小鼠中，激动性CD40抗体诱导淋巴素α在B细胞中的表达，并增强TLSs的形成。由B细胞、t细胞和抗原提呈细胞组成的TLSs在肿瘤组织近端和脑膜附近的CD31+血管周围形成。激动性CD40抗体治疗提高了荷瘤小鼠的CD8+ t细胞/ Treg比率，但没有生存益处。CD8+ t细胞上CD69激活状态的降低，CD1d+CD5+调节性表型的B细胞的增加以及tls中Tregs的高浸润表明，激动性CD40抗体在胶质瘤中诱导了免疫抑制微环境。与此一致的是，PD-1和CTLA-4检查点阻断的有效性在联合使用激动性CD40抗体后显著降低。这些结果表明，TLSs在胶质瘤中存在并可被诱导，但至少在某些条件下它们与免疫抑制有关，从而揭示了脑微环境中免疫调节的新机制。引文格式:Luuk van Hooren, Alessandra Vaccaro, Maria Georganaki, Mohanraj Ramachandran, Hua Huang, Joey Lau, Anja Smits, Magnus Essand, Anna Dimberg。激动性CD40抗体治疗诱导胶质瘤中三级淋巴结构的形成并抑制对免疫检查点封锁的反应[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，2019;7(2增刊):摘要nr A159。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-PR07
A. Young, V. Nguyen, J. Kang, Sadaf Mehdizadeh, Amy Mei, K. Sheehan, D. Serreze, Yi-Guang Chen, R. Schreiber, J. Bluestone
With increasing use of cancer immunotherapies, it is evident that some patients develop immunotoxicity, termed immune-related adverse events (irAEs). Many of these irAEs are a consequence of a peripheral breakdown in self-tolerance but the basis, be it genetic, epitope spreading or environment, remains unclear. This highlights an urgent need to develop preclinical models to profile autoimmunity together with antitumor immunity to accurately assess the etiology by which autoimmune pathologies are induced in immunotherapy-treated cancer patients. We generated transplantable syngeneic tumor cell lines in the autoimmune-prone NOD background from spontaneous tumors and methylcholanthrene (MCA)-induced fibrosarcomas. We investigated whether the presence of tumor and level of tumor immunogenicity altered the severity and frequency of autoimmunity in response to cancer immunotherapies. Notably, individual and combination immunotherapies displayed distinct autoimmune pathologies, with induction of certain treatment-associated immunotoxicities, such as type 1 diabetes (T1D), associated with improved tumor control. Following, we investigated genetic and therapeutic strategies to abrogate immunotherapy-induced autoimmunity without impeding antitumor immunity. Multiple loci that control genetic susceptibility to autoimmunity in NOD mice have been identified including a major histocompatibility complex, as well as insulin-dependent diabetes (Idd) susceptibility loci localized to relevant immune checkpoints. Using NOD mice, congenic for individual IDD loci, we determined that NOR- or C57BL/6-derived gene loci provided significant protection from anti-PD-1/PD-L1-induced type 1 diabetes. In some cases, immune tolerance towards autoimmunity limited cancer immune surveillance; however, NOD mice congenic for NOR-Idd9 displayed both superior tumor control and significantly reduced incidence of T1D in comparison to NOD mice. This result suggests that in fact by abrogating the pathogenic autoimmune response, it is possible for improved tumor control to be afforded, representing an exciting area for mechanistic exploration. Together, these preclinical models provide a platform to assess safety profiles for cancer immunotherapies, identifying cellular mechanisms and therapeutic interventions that inhibit the development of autoimmunity while preserving antitumor immunity. Citation Format: Arabella Young, Vinh Nguyen, Jee Hye Kang, Sadaf Mehdizadeh, Amy Mei, Kathleen C. F. Sheehan, David V. Serreze, Yi-Guang Chen, Robert D. Schreiber, Jeffrey A. Bluestone. Developing syngeneic NOD tumor models to profile immunotoxicity and antitumor immunity in response to cancer immunotherapies in autoimmune-prone mice [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 PR07.
随着癌症免疫疗法使用的增加，很明显，一些患者出现免疫毒性，称为免疫相关不良事件(irAEs)。这些irae中有许多是自我耐受性外周破坏的结果，但其基础，无论是遗传、表位扩散还是环境，仍不清楚。这突出了迫切需要开发临床前模型来分析自身免疫和抗肿瘤免疫，以准确评估免疫疗法治疗的癌症患者诱导自身免疫病理的病因。我们从自发性肿瘤和甲基胆蒽(MCA)诱导的纤维肉瘤中获得了可移植的同基因肿瘤细胞系。我们研究了肿瘤的存在和肿瘤免疫原性水平是否改变了癌症免疫治疗反应中自身免疫的严重程度和频率。值得注意的是，单独和联合免疫疗法显示出不同的自身免疫病理，诱导某些与治疗相关的免疫毒性，如与肿瘤控制改善相关的1型糖尿病(T1D)。接下来，我们研究了在不妨碍抗肿瘤免疫的情况下消除免疫治疗诱导的自身免疫的遗传和治疗策略。在NOD小鼠中已经发现了多个控制自身免疫遗传易感性的基因座，包括一个主要的组织相容性复合体，以及位于相关免疫检查点的胰岛素依赖型糖尿病(Idd)易感性基因座。在NOD小鼠中，我们确定了单个IDD基因座的同源性，NOR-或C57BL/6衍生的基因座对抗pd -1/ pd - l1诱导的1型糖尿病具有显著的保护作用。在某些情况下，对自身免疫的免疫耐受限制了癌症免疫监测;然而，与NOD小鼠相比，携带NOR-Idd9基因的NOD小鼠表现出更好的肿瘤控制能力，并显著降低了T1D的发病率。这一结果表明，事实上，通过废除致病性自身免疫反应，有可能改善肿瘤控制，这代表了一个令人兴奋的机制探索领域。总之，这些临床前模型提供了一个平台来评估癌症免疫疗法的安全性，确定细胞机制和治疗干预，抑制自身免疫的发展，同时保持抗肿瘤免疫。引文格式:Arabella Young, Vinh Nguyen, Jee Hye Kang, Sadaf Mehdizadeh, Amy Mei, Kathleen C. F. Sheehan, David V. Serreze, Yi-Guang Chen, Robert D. Schreiber, Jeffrey A. Bluestone。在自身免疫易感小鼠中建立同基因NOD肿瘤模型，分析肿瘤免疫治疗对免疫毒性和抗肿瘤免疫的反应[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 PR07。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A124
A. Anderson, A. Becker, Fangfang Yin, Hema Singh, Xiaoning Zhao, L. Seitz, Rick Stanton, N. Walker, J. Tan
TIGIT (T-cell immunoreceptor with Ig and ITIM domains) is an inhibitory receptor that is expressed on natural killer (NK) cells, CD8+ T-cells, and immunosuppressive regulatory T-cells (Treg). DNAM-1 (DNAX Accessory Molecule-1; CD226) is an activating receptor found on NK cells, monocytes and a subset of T-cells. TIGIT and DNAM-1 are paired receptors that compete for shared ligands CD155 (PVR) and CD112 (Nectin-2) expressed by tumor and antigen-presenting cells. TIGIT binding to CD155 or CD112 results in immune suppression, whereas binding of DNAM-1 to the same ligands mediates immune activation. As malignancies progress, high TIGIT expression often occurs alongside the upregulation of other immune checkpoint proteins and markers of T-cell exhaustion such as PD-1 (Programmed Death-1). We have developed AB154 to inhibit TIGIT and shift the balance in the tumor microenvironment towards a more productive anticancer response. Blockade of multiple immune checkpoint proteins can confer effective and durable responses in the treatment of cancer. Data assembled from TCGA (The Cancer Genome Atlas) identified numerous tumor types in which TIGIT is co-expressed with PD-1. In these tumors, TIGIT and PD-1 were significantly upregulated compared to normal adjacent tissue. Immunophenotyping performed on human tumor infiltrating lymphocytes demonstrated a strong correlation between TIGIT and PD-1 co-expression on specific immune cells including CD8+ T-cells and Treg cells. AB154 is a fully humanized antibody that blocks human TIGIT with sub-nanomolar affinity, as determined using a CHO.hTIGIT over-expressing cell line and primary human T-cells. Functional consequences of blocking TIGIT/CD155 interactions in combination with anti-PD-1 or anti-PD-L1 were evaluated using mixed lymphocyte reactions (MLR). Briefly, we show here that co-cultures of GM-CSF/IL-4-differentiated CD155+ PD-L1+ monocytes and TIGIT+ CD4+ T-cells, in the presence of AB154, significantly increased IFN-gamma secretion when combined with anti-PD-1 or anti-PD-L1 blocking antibodies relative to each monotherapy. Understanding pharmacokinetic (PK) and pharmacodynamic (PD) relationships enables the choice of a dosing regimen that provides adequate target coverage. To evaluate the PD effects of AB154 in clinical samples, we developed a multicolor flow cytometry-based assay that utilizes an anti-TIGIT antibody that is competitive with AB154 to determine receptor occupancy. In human whole blood, ex vivo addition of AB154 achieved complete inhibition of TIGIT. Analysis of blood mononuclear cells, including CD8+ and CD4+ T-cells, Treg and NK cells, demonstrated target engagement by AB154 suitable for clinical development. In addition, we examined TIGIT receptor occupancy of AB154 (added to whole blood ex vivo) in a small cohort of non-small cell lung cancer (NSCLC) patients treated with anti-PD-1 antibody pembrolizumab. In these samples, TIGIT receptor occupancy by AB154 was comparable to that obtained wi
{"title":"Abstract A124: Preclinical characterization of AB154, a fully humanized anti-TIGIT antibody, for use in combination therapies","authors":"A. Anderson, A. Becker, Fangfang Yin, Hema Singh, Xiaoning Zhao, L. Seitz, Rick Stanton, N. Walker, J. Tan","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A124","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A124","url":null,"abstract":"TIGIT (T-cell immunoreceptor with Ig and ITIM domains) is an inhibitory receptor that is expressed on natural killer (NK) cells, CD8+ T-cells, and immunosuppressive regulatory T-cells (Treg). DNAM-1 (DNAX Accessory Molecule-1; CD226) is an activating receptor found on NK cells, monocytes and a subset of T-cells. TIGIT and DNAM-1 are paired receptors that compete for shared ligands CD155 (PVR) and CD112 (Nectin-2) expressed by tumor and antigen-presenting cells. TIGIT binding to CD155 or CD112 results in immune suppression, whereas binding of DNAM-1 to the same ligands mediates immune activation. As malignancies progress, high TIGIT expression often occurs alongside the upregulation of other immune checkpoint proteins and markers of T-cell exhaustion such as PD-1 (Programmed Death-1). We have developed AB154 to inhibit TIGIT and shift the balance in the tumor microenvironment towards a more productive anticancer response. Blockade of multiple immune checkpoint proteins can confer effective and durable responses in the treatment of cancer. Data assembled from TCGA (The Cancer Genome Atlas) identified numerous tumor types in which TIGIT is co-expressed with PD-1. In these tumors, TIGIT and PD-1 were significantly upregulated compared to normal adjacent tissue. Immunophenotyping performed on human tumor infiltrating lymphocytes demonstrated a strong correlation between TIGIT and PD-1 co-expression on specific immune cells including CD8+ T-cells and Treg cells. AB154 is a fully humanized antibody that blocks human TIGIT with sub-nanomolar affinity, as determined using a CHO.hTIGIT over-expressing cell line and primary human T-cells. Functional consequences of blocking TIGIT/CD155 interactions in combination with anti-PD-1 or anti-PD-L1 were evaluated using mixed lymphocyte reactions (MLR). Briefly, we show here that co-cultures of GM-CSF/IL-4-differentiated CD155+ PD-L1+ monocytes and TIGIT+ CD4+ T-cells, in the presence of AB154, significantly increased IFN-gamma secretion when combined with anti-PD-1 or anti-PD-L1 blocking antibodies relative to each monotherapy. Understanding pharmacokinetic (PK) and pharmacodynamic (PD) relationships enables the choice of a dosing regimen that provides adequate target coverage. To evaluate the PD effects of AB154 in clinical samples, we developed a multicolor flow cytometry-based assay that utilizes an anti-TIGIT antibody that is competitive with AB154 to determine receptor occupancy. In human whole blood, ex vivo addition of AB154 achieved complete inhibition of TIGIT. Analysis of blood mononuclear cells, including CD8+ and CD4+ T-cells, Treg and NK cells, demonstrated target engagement by AB154 suitable for clinical development. In addition, we examined TIGIT receptor occupancy of AB154 (added to whole blood ex vivo) in a small cohort of non-small cell lung cancer (NSCLC) patients treated with anti-PD-1 antibody pembrolizumab. In these samples, TIGIT receptor occupancy by AB154 was comparable to that obtained wi","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"30 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84831158","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-A156
K. Sullivan, Y. D. Seo, Xiu-yun Jiang, Teresa Kim, R. Yeung, V. Pillarisetty
Introduction: Pancreatic ductal adenocarcinoma (PDA) is the fourth most common cause of cancer death in the United States. Despite the fact that the human PDA microenvironment contains a mixture of infiltrating immune cells, including activated effector CD8+ T-cells, attempts to apply immune checkpoint inhibitor therapy have not yet been successful in achieving a clinical response (1). Combining blockade of the immune checkpoint receptor PD-1 with blockade of the chemokine receptor CXCR4 in a mouse model of PDA produced T-cell-mediated tumor cell killing (2). We therefore hypothesized that, in a human PDA slice culture model system that maintains the intact tumor microenvironment (3), combination of CXCR4 blockade with PD-1 blockade would lead to anti-tumor T-cell activation. Methods: Cores (6 mm) were taken from freshly resected, sterile human PDA specimens and cut into 250 µm thick slices using a vibratome. The slices were treated with either IgG antibody isotype control, AMD3100 (a CXCR4 blocking small molecule drug) plus antibody isotype control, anti-PD-1 antibody, or a combination of anti-PD-1 antibody and AMD3100 for two days. The slices were then stained with fluorescently labelled antibodies for CD8 and EpCAM, as well as with SR-FLICA, a reagent that produces a fluorescent signal when bound to activated Caspase 3 and 7 enzymes. Live slices were maintained in a humidified, temperature-controlled CO2 chamber for time-lapse confocal imaging for 1 hour. Similarly, untreated slices were imaged for 90 minutes prior to and following combined anti-PD-1 and AMD3100 drug treatment. Each slice was imaged at 3 different positions to observe CD8+ T-cells, EpCAM+ tumor cells, and SR-FLICA+ apoptotic cells throughout the slice. Each cell type was counted at each position over the entire time course imaged. Results: Compared to antibody isotype control or monotherapy, slices treated with combined PD-1 and CXCR4 blockade contained a greater fraction of EpCAM+ tumor cells that were undergoing apoptosis, indicated by SR-FLICA positivity. This treatment also increased the proportion of EpCAM+ tumor cells with a CD8+ T-cell within 20 µm, and EpCAM+ cells had increased evidence of apoptosis when a CD8+ cell was nearby. When individual slices were imaged before and after combined PD-1 and CXCR4 blockade, a greater proportion of EpCAM+ cells were found to have a CD8+ cell nearby after treatment compared to before treatment, and the tumor cells with a CD8+ cell nearby were more like to be undergoing apoptosis compared to before treatment. Conclusion: Direct visualization of live pancreatic cancer slice cultures demonstrates that combined PD-1 and CXCR4 blockade enhances both the migration of CD8+ T-cells towards epithelial tumor cells and their anti-tumor cytotoxic activity. References: 1. Shibuya KC, Goel VK, Xiong W, Sham JG, Pollack SM, Leahy AM, et al. Pancreatic ductal adenocarcinoma contains an effector and regulatory immune cell infiltrate that is altered
简介:胰腺导管腺癌(PDA)是美国第四大最常见的癌症死亡原因。尽管人类PDA微环境包含浸润性免疫细胞的混合物，包括活化的效应CD8+ t细胞，但应用免疫检查点抑制剂治疗的尝试尚未成功实现临床应答(1)。在PDA小鼠模型中，联合阻断免疫检查点受体PD-1和趋化因子受体CXCR4可产生t细胞介导的肿瘤细胞杀伤(2)。因此，我们假设，在维持完整肿瘤微环境的人PDA切片培养模型系统中(3)，CXCR4阻断与PD-1阻断联合可导致抗肿瘤t细胞活化。方法:从新鲜切除的无菌人PDA标本中取芯(6mm)，用振动刀切成250µm厚的切片。切片用IgG抗体同型对照、AMD3100(一种阻断CXCR4的小分子药物)加抗体同型对照、抗pd -1抗体或抗pd -1抗体与AMD3100联合处理2天。然后用CD8和EpCAM的荧光标记抗体以及SR-FLICA对切片进行染色，SR-FLICA是一种与活化的Caspase 3和7酶结合时产生荧光信号的试剂。活切片在加湿、温控的CO2室中保存1小时，进行延时共聚焦成像。同样，在联合抗pd -1和AMD3100药物治疗前后90分钟，对未处理的切片进行成像。每张切片在3个不同位置成像，观察整个切片中CD8+ t细胞、EpCAM+肿瘤细胞和SR-FLICA+凋亡细胞。在整个成像过程中，在每个位置对每种细胞类型进行计数。结果:与抗体同型对照或单药治疗相比，PD-1和CXCR4联合阻断治疗的切片中EpCAM+肿瘤细胞的凋亡比例更高，SR-FLICA阳性。该处理还增加了20µm内含有CD8+ t细胞的EpCAM+肿瘤细胞的比例，并且当CD8+细胞附近时，EpCAM+细胞凋亡的证据增加。在PD-1和CXCR4联合阻断前后对单个切片进行成像时，发现治疗后EpCAM+细胞附近有CD8+细胞的比例比治疗前更高，并且附近有CD8+细胞的肿瘤细胞比治疗前更容易发生凋亡。结论:胰腺癌切片直接可视化显示，PD-1和CXCR4联合阻断可增强CD8+ t细胞向上皮肿瘤细胞的迁移和抗肿瘤细胞毒活性。引用:1。Shibuya KC, Goel VK，熊伟，Sham JG, Pollack SM, Leahy AM，等。胰腺导管腺癌包含效应性和调节性免疫细胞浸润，可通过多模式新辅助治疗改变。科学通报，2014;9(5):96565。2. 张建军，张建军，张建军，等。从表达fap的癌相关成纤维细胞靶向CXCL12与抗pd - l1免疫治疗在胰腺癌中的协同作用中国科学:自然科学版，2013;11(5):20212-7。3.姜欣，徐彦东，常建辉，盖弗勒A，尼杰·恩，潘生，等。长期胰腺导管腺癌切片培养使免疫微环境的精确研究成为可能。Oncoimmunology 2017; 6(7)。引文格式:Kevin M. Sullivan, Yongwoo David Seo, Xiuyun Jiang, Teresa David Kim, Raymond S.W. Yeung, Venu G. Pillarisetty。联合抗pd -1阻断和CXCR4阻断后的PDA肿瘤细胞死亡是CD8+ t细胞的直接作用[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 A156。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A133
Amanda L. Huff, Phonphimon Wongthida, T. Kottke, Jill Thompson, C. Driscoll, M. Schuelke, Kevin G. Shim, R. Harris, Amy M. Molan, J. Pulido, P. Selby, K. Harrington, A. Melcher, Laura Evgin, R. Vile
Tumor cells are able to evade cytotoxic therapies through the accumulation of genomic mutations and rapid evolution. In the case of oncolytic virotherapy, understanding the mechanisms by which cancer cells develop resistance to infection and lysis is critical to the development of more effective viral-based platforms. Here, we identify APOBEC3 as an important factor involved in the restriction of vesicular stomatitis virus (VSV). We show that VSV infection of B16 murine melanoma cells upregulated APOBEC3 in an IFNβ-dependent manner, which was responsible for the evolution of virus-resistanT-cell populations and suggested that APOBEC3 expression promoted the acquisition of a virus-resistant phenotype. ShRNA knockdown of APOBEC3 in B16 cells diminished their capacity to develop resistance to VSV infection in vitro, and enhanced the therapeutic effect of VSV in vivo. Similarly, overexpression of human APOBEC3B promoted the acquisition of resistance to oncolytic VSV in murine melanoma and glioblastoma lines, as well as in a human melanoma cell line in vitro and in vivo. Finally, we demonstrate that progeny virus obtained after passage through APOBEC3B overexpressing cells contained more defective interfering particles, thereby indicating that APOBEC3B directly affected the fitness of VSV, an RNA virus that has not previously been identified to be restricted by APOBEC3B. This research identifies APOBEC3 enzymes as key players to target in order to improve the efficacy of oncolytic viruses as well as broader nucleic acid-based therapeutic platforms. Citation Format: Amanda L. Huff, Phonphimon Wongthida, Timothy Kottke, Jill Thompson, Christopher B. Driscoll, Matthew Schuelke, Kevin G. Shim, Reuben S. Harris, Amy Molan, Jose S. Pulido, Peter J. Selby, Kevin J. Harrington, Alan Melcher, Laura Evgin, Richard Vile. APOBEC3 confers resistance to oncolytic VSV therapy [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 A133.
肿瘤细胞能够通过基因组突变的积累和快速进化来逃避细胞毒性治疗。就溶瘤病毒治疗而言，了解癌细胞对感染和裂解产生抗性的机制对于开发更有效的基于病毒的平台至关重要。在这里，我们确定APOBEC3是参与限制水疱性口炎病毒(VSV)的一个重要因素。我们发现VSV感染B16小鼠黑色素瘤细胞以ifn β依赖的方式上调APOBEC3，这与病毒抗性细胞群体的进化有关，并表明APOBEC3的表达促进了病毒抗性表型的获得。ShRNA敲低B16细胞的APOBEC3，在体外降低了B16细胞对VSV感染的抗性，在体内增强了VSV的治疗效果。同样，人APOBEC3B的过表达促进了小鼠黑色素瘤和胶质母细胞瘤细胞系以及人黑色素瘤细胞系在体外和体内获得对溶瘤性VSV的抗性。最后，我们证明通过APOBEC3B过表达细胞获得的子代病毒含有更多有缺陷的干扰颗粒，从而表明APOBEC3B直接影响VSV的适应度，而VSV是一种以前未被鉴定为受APOBEC3B限制的RNA病毒。本研究确定了APOBEC3酶作为关键靶点，以提高溶瘤病毒的疗效以及更广泛的基于核酸的治疗平台。引文格式:Amanda L. Huff, Phonphimon Wongthida, Timothy Kottke, Jill Thompson, Christopher B. Driscoll, Matthew Schuelke, Kevin G. Shim, Reuben S. Harris, Amy Molan, Jose S. Pulido, Peter J. Selby, Kevin J. Harrington, Alan Melcher, Laura Evgin, Richard Vile。APOBEC3赋予对溶瘤性VSV治疗的抗性[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 A133。
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Pub Date : 2019-02-01DOI: 10.1158/2326-6074.CRICIMTEATIAACR18-A127
Z. Deng, F. Geissmann
Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal disease and frequently metastasize to the liver. The presence of macrophages (F4/80+ cells or CD68+ cells) is reported to correlate with tumor progression and metastasis in pancreatic cancer. Emerging evidence has revised the traditional concept of macrophage ontogeny and made it increasingly apparent that there exists functional diversity between resident macrophages originating from yolk sac erythro-myeloid progenitors (EMPs) and hematopoietic stem cells (HSCs)-derived macrophages. Sophisticated therapeutic intervention requires in-depth understanding of these macrophage subsets in the metastasis microenvironment. However, the role and heterogeneity of macrophages in tumor development, in particular in PDAC liver metastasis, remains poorly understood. Here, we utilized murine PDAC model to inidcate that Kupffer cells and tumor associated macrophages (TAMs) are two developmentally and functionally distinct subsets of macrophages. Removal of Kupffer cells by depletion of Id3 in macrophage progenitors or CSF1R blockade increases tumor burden. These findings reveal that Kupffer cells play a key role in restraining PDAC liver metastasis and show the necessity to understand macrophage origin when developing therapeutics for cancer treatment. Citation Format: Zihou Deng, Frederic Geissmann. Role of macrophage subsets in liver metastasis [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 A127.
{"title":"Abstract A127: Role of macrophage subsets in liver metastasis","authors":"Z. Deng, F. Geissmann","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A127","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A127","url":null,"abstract":"Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal disease and frequently metastasize to the liver. The presence of macrophages (F4/80+ cells or CD68+ cells) is reported to correlate with tumor progression and metastasis in pancreatic cancer. Emerging evidence has revised the traditional concept of macrophage ontogeny and made it increasingly apparent that there exists functional diversity between resident macrophages originating from yolk sac erythro-myeloid progenitors (EMPs) and hematopoietic stem cells (HSCs)-derived macrophages. Sophisticated therapeutic intervention requires in-depth understanding of these macrophage subsets in the metastasis microenvironment. However, the role and heterogeneity of macrophages in tumor development, in particular in PDAC liver metastasis, remains poorly understood. Here, we utilized murine PDAC model to inidcate that Kupffer cells and tumor associated macrophages (TAMs) are two developmentally and functionally distinct subsets of macrophages. Removal of Kupffer cells by depletion of Id3 in macrophage progenitors or CSF1R blockade increases tumor burden. These findings reveal that Kupffer cells play a key role in restraining PDAC liver metastasis and show the necessity to understand macrophage origin when developing therapeutics for cancer treatment. Citation Format: Zihou Deng, Frederic Geissmann. Role of macrophage subsets in liver metastasis [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 A127.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"61 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":"90122318","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-A152
Darin Salloum, B. Santomasso, R. Brentjens, H. Wendel
Chimeric antigen receptor T (CAR-T) cell therapies are rapidly becoming first in line therapies for treatment of hematologic malignancies. This is due to the efficacy of the treatment and long-lasting tumor responses. However, CAR-T therapies are associated with unique set of acute toxicities— cytokine release syndrome (CRS) and neurologic toxicities—the cause of which is not clear. As other immune therapies are becoming more widely used, these toxicities are observed in patients undergoing bispecific T-cell engaging antibodies (BiTEs) and checkpoint blockade therapies. Here, we sought to define metabolic changes in patients undergoing chimeric antigen receptor (CAR) T-cell therapy for B-acute lymphoblastic leukemia. We performed metabolic analysis of amino acids in serum samples, and identified significant up-regulation of tryptophan-kynurenine metabolism in patients with high-grade neurotoxicity. In-depth look at tryptophan-kynurenine pathway in cerebrospinal fluid patient samples determined quinolinic acid up-regulation to correlate with low- and high-grade neurotoxicity, indicating local production of this metabolite. We show that quinolinic acid is produced by activated monocytes and microglia in response to Il1beta and TNF. Importantly, tryptophan metabolite—kynurenine— stimulates Il1beta production and further activation of monocytes, possibly exacerbating CSR and neurotoxicity. This study suggests that inhibition of tryptophan-kynurenine pathway may offer some benefit to manage neurotoxicity and CRS associated with CAR-T-cell therapy treatment. Citation Format: Darin Salloum, Bianca Santomasso, Renier J. Brentjens, Hans-Guido Wendel. Investigating metabolic basis of neurotoxicity during CAR-T therapies [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 A152.
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