bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract IA14: Circulating tumor DNA in early-stage NSCLC: A lung TRACERx study
Authors: Abbosh Christopher
Published: February 2019

The Cancer Research UK-funded study TRACERx (TRAcking Cancer Evolution through therapy [Rx]) aims to recruit patients undergoing surgery for early-stage non-small cell lung cancer (NSCLC). Patients donate blood pre- and postoperatively alongside multi-region sampled primary tumor tissue. Up to 40% of patients undergoing surgery for NSCLC experience relapse of their disease. At this point patients are approached for relapse tissue sampling. TRACERx therefore provides a strong platform for evaluating circulating tumor DNA (ctDNA) in early-stage NSCLC and a minimal residual disease (MRD) setting.

In this presentation we will describe data arising from a phylogenetic, patient-specific approach to ctDNA evaluation in the first 100 patients analyzed as part of TRACERx. We will discuss clinicopathologic factors associated with ctDNA detection in early-stage NSCLC and suggest the existence of distinct tumor phenotypic characteristics that predict the presence of ctDNA at measurable quantities in plasma. Additionally, we will discuss tumor volume limit of detection analyses and highlight potential implications of our findings within the context of early cancer detection. We will then examine data arising from the ctDNA profiling of postoperative plasma samples taken in the adjuvant setting. These data demonstrate that ctDNA detection is specific for NSCLC relapse and that ctDNA detection precedes clinical diagnosis of NSCLC relapse (median lead-time of 77 days, range 10 to 346 days). We will provide examples of using phylogenetic ctDNA profiling to characterize the subclonal nature of relapsing NSCLC as a potential method to personalize relapse therapeutic intervention. Finally, we will highlight potential advantages of patient-specific ctDNA profiling in an adjuvant MRD setting and suggest ways that this technology can be used to fuel adjuvant drug development in NSCLC by driving novel study designs.

Citation Format: Christopher Abbosh. Circulating tumor DNA in early-stage NSCLC: A lung TRACERx study [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr IA14.
Abstract IA36: Regulatory T cells, polymorphisms, and response to checkpoint blockade: From mechanisms to potential biomarkers
Authors: Quezada Sergio
Published: February 2019

In mice, anti-CTLA-4 monoclonal antibodies (mAbs) require Fc-gamma receptor (FcγR)-mediated depletion of intratumoral regulatory T (Treg) cells to promote tumor rejection. However, the relevance of this mechanism in the human setting remains controversial. Using a mouse model expressing human FcγRs, and analysis of clinical samples from ipilimumab-treated melanoma patients, we investigate the potential role of Treg depletion in the activity of antibodies targeting human CTLA-4.

Citation Format: Sergio Quezada. Regulatory T cells, polymorphisms, and response to checkpoint blockade: From mechanisms to potential biomarkers [abstract]. In: Proceedings of the Fifth AACR-IASLC International Joint Conference: Lung Cancer Translational Science from the Bench to the Clinic; Jan 8-11, 2018; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(17_Suppl):Abstract nr IA36.
Abstract A013: Haploidentical stem cell transplantation and subsequent immunotherapy with antiGD2 antibody for patients with relapsed metastatic neuroblastoma
Authors: Lang Peter , Flaadt Tim , Ebinger Martin , Schlegel Patrick , Lode Holger , Ladenstein Ruth , Lang Anne-Marie , Ambross Peter , Schaefer Juergen , Fuchs Joerg , Loibner Hans , Schwinger Wolfgang , Handgretinger Rupert
Published: February 2019

Background: Pediatric patients with relapsed metastatic neuroblastomas have a poor prognosis and additional strategies are needed. We present results of a phase I/II-trial with subsequent immunotherapy with an anti-GD2mAb (CH14.18/CHO) after HLA-mismatched, haploidentical stem cell transplantation (SCT). Methods: T- and B-cell depleted stem cells from parental donors were used in combination with melphalan 140mg/m², thiotepa 10mg/kg, fludarabin 160mg/m² and ATG-F. CH14.18/CHOmAb was started on day 60-180 post-transplant: 6 cycles with 20mg/m²/day x 5; in cycles 4-6, 1x106 U/m² Interleukin 2 was given additionally. Disease status was evaluated with whole body MRI, MIBG scan and MRD detection in bone marrow aspirates. Primary endpoint was success of treatment, defined as a patient receiving the full protocol treatment, still alive 180 days after end of treatment without progression/unacceptable toxicity or severe GvHD. Results: 56 patients with 1st or ≥2nd metastatic relapse were enrolled. Disease status prior to antibody infusions was: CR (n=19), PR (n=31), mixed (n=6). 41% of patients with measurable tumor burden responded and reached CR after treatment. 90% of patients with initial CR could maintain this status. In total, success of treatment was observed in 60%. 3-year OS and EFS was 58% and 45%, respectively. Causes of death were: progression/relapse (25%) or TRM (7%). Factors of influence on EFS were (1) remission status prior to SCT (with CR, PR or mixed response resulting in 70%, 45% and 11% 3yEFS) and (2) bone marrow involvement prior to CH14.18 treatment (no MRD detectable: 60% 3yEFS vs. any MRD detectable: 20% 3yEFS). Frequent side effects were pain, fever, CRP elevation; rare side effects comprised SIRS/capillary leak syndrome, seizures, accommodation disturbances. Transient acute severe GvHD occurred in 1 patient. Conclusions: CH14.18/CHO infusions after haploidentical SCT are feasible with acceptable toxicity. Our results suggest an antitumor effect of the new, donor-derived immune system in combination with CH14.18 treatment.

Citation Format: Peter Lang, Tim Flaadt, Martin Ebinger, Patrick Schlegel, Holger Lode, Ruth Ladenstein, Anne-Marie Lang, Peter Ambross, Juergen Schaefer, Joerg Fuchs, Hans Loibner, Wolfgang Schwinger, Rupert Handgretinger. Haploidentical stem cell transplantation and subsequent immunotherapy with antiGD2 antibody for patients with relapsed metastatic neuroblastoma [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 A013.
Abstract A029: Unexpected antagonism between oncolytic virus derived type I interferon and EGFRvIII CAR T-cells
Authors: Evgin Laura , Huff Amanda L. , Wongthida Phonphimon , Thompson Jill , Kottke Timothy , Sampson John , Perez Luis Sanchez , Vile Richard
Published: February 2019

Although the chimeric antigen receptor (CAR) T-cell platform has experienced clinical success in patients with hematologic malignancies, CAR T-cells specific to solid tumor targets have met with more limited efficacy. The highly inflammatory nature of oncolytic viruses and their ability to remodel the tumor microenvironment suggested to us that they would provide a complementary mechanism of action to both recruit and potentiate the functionality of CAR T-cells. VSVmIFNβ injection into B16EGFRvIII tumors increased the expression of chemokines such as CXCL10 and CCL5, which we would expect to recruit CXCR3+and CCR5+ EGFRvIII specific murine CAR T-cells. However, we did not observe an increase in overall survival or tumor control using the combination strategy compared to monotherapy with CAR T-cells. We recovered fewer viable CD8+ CAR T-cells from tumors injected with VSVmIFNβ early after adoptive transfer, and observed a similar reduction in the number of CD8+ CAR T-cells which persisted long term in the blood. We have shown that type I interferon increases the expression of the CAR from the retroviral LTR and in turn sensitizes these cells to tonic signaling mediated exhaustion and apoptosis. Correspondingly, CAR T-cells prepared from IFNAR KO T-cells were protected from the deleterious effect of type I IFN in vivo. We are currently investigating strategies to overcome the interference between these two modalities and to uncouple the regulation of the expression of the CAR from type I IFN signaling.

Citation Format: Laura Evgin, Amanda L. Huff, Phonphimon Wongthida, Jill Thompson, Timothy Kottke, John Sampson, Luis Sanchez Perez, Richard Vile. Unexpected antagonism between oncolytic virus derived type I interferon and EGFRvIII CAR T-cells [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 A029.
Abstract A044: Mutated NPM1 as target for immunotherapy of acute myeloid leukemia
Authors: Lee Dyantha I. van der , Reijmers Rogier M. , Honders M. Willy , Hagedoorn Renate S. , Jong Rob. M. de , Kester Michel G.D. , Steen Dirk M. van der , Ru Arnoud H. de , Kweekel Christiaan , Jedema Inge , Veelken Hendrik , Heemskerk Mirjam M. , Veelen Peter A. van , Falkenburg J.H. Frederik , Griffioen Marieke
Published: February 2019

The most frequent subtype of acute myeloid leukemia (AML) is defined by mutations in the nucleophosmin (NPM1) gene. Mutated NPM1 is an attractive target for immunotherapy, since it is an essential driver gene and 4 base pair frameshift insertions in exon 12 occur in 30-35% of AML, resulting in a novel C-terminal alternative reading frame of 11 amino acids. By searching in the HLA class I ligandome of primary AML, we identified multiple peptides derived from mutated NPM1. For one of these peptides, i.e., HLA-A*02:01-presented CLAVEEVSL, we searched for specific T-cells in AML patients and healthy individuals using peptide-MHC tetramers. Tetramer-positive CD8 T-cell clones were isolated and analyzed for reactivity against primary AML with mutated NPM1. From one selected clone with superior antitumor reactivity, we isolated the T-cell receptor (TCR) and demonstrated specific recognition and lysis of HLA-A*02:01-positive AML with mutated NPM1 in vitro after retroviral transfer to CD8 and CD4 T-cells. In vivo antitumor efficacy of TCR-transduced CD8 and CD4 T-cells was confirmed in immunodeficient mice engrafted with a human AML cell line expressing mutated NPM1. These data show that mutated NPM1-derived peptides are presented on AML and that CLAVEEVSL is a neoantigen that can be efficiently targeted on AML with mutated NPM1 by TCR gene transfer in a co-receptor independent fashion. Immunotherapy targeting mutated NPM1 may therefore contribute to treatment of AML.

Citation Format: Dyantha I. van der Lee, Rogier M. Reijmers, M. Willy Honders, Renate S. Hagedoorn, Rob. M. de Jong, Michel G.D. Kester, Dirk M. van der Steen, Arnoud H. de Ru, Christiaan Kweekel, Inge Jedema, Hendrik Veelken, Mirjam M. Heemskerk, Peter A. van Veelen, J.H. Frederik Falkenburg, Marieke Griffioen. Mutated NPM1 as target for immunotherapy of acute myeloid leukemia [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 A044.
Abstract A062: TIM-3 plays distinct roles in different immune cells to regulate antitumor immune responses
Authors: Dai Jie , Pei Jerry , Mohrs Markus , Thurston Gavin , Ioffe Ella
Published: February 2019

T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) was originally identified as an inhibitory receptor that is expressed on Th1 T-cells to dampen T-cell immunity upon interaction with several putative ligands. The inhibitory role of TIM-3 is supported by multiple preclinical disease models, such as viral and bacterial infections, experimental autoimmune encephalitis, and allograft rejection. Recent evidence suggests that induction of TIM-3 expression on T-cells may promote resistance to cancer immunotherapy, including in response to treatment with PD-1/PD-L1 inhibitors. Thus, TIM-3 represents a putative novel immuno-oncology target. Here we report that in naïve mice TIM-3 is absent on T-cells, but is constitutively expressed on myeloid cells, including dendritic cells and macrophages. In tumor-bearing mice, TIM-3 expression is highly enriched on PD-1 positive CD4 and CD8 T-cells in the tumor microenvironment, but not on T-cells in draining lymph nodes or peripheral blood. Prophylactic or therapeutic treatment with anti-PD-1 blocking antibody delays tumor growth in wild-type mice. However, unexpectedly, TIM-3 genetic deficiency reduced overall survival of tumor-bearing mice treated with anti-PD-1 compared to wild-type control mice. Using a series of in vitro functional cell-based assays, we found that blocking TIM-3 function by either genetic knock-out or an inhibitory Ab increased proliferation of, and IFN-γ production by, effector CD8 T-cells following direct antigen stimulation. By contrast, TIM-3 gene knockout in bone marrow-derived macrophages did not impact responses to stimulation with Toll-like receptor ligands, and TIM-3 blockade reduced phagocytosis of apoptotic tumor cells by a macrophage cell line. Taken together, our results suggest TIM-3 may play opposite roles in T-cells and macrophages (inhibitory vs activating, respectively), and highlight the pleiotropic roles of TIM-3 in different immune cells in tumor immunology.

Citation Format: Jie Dai, Jerry Pei, Markus Mohrs, Gavin Thurston, Ella Ioffe. TIM-3 plays distinct roles in different immune cells to regulate antitumor 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 A062.
Abstract A070: Virotherapy eradicates established melanoma by reprogramming the tumor microenvironment and engaging the adaptive immunity
Authors: Flatz Lukas , Ring Sandra , Bomze David , Onder Lucas , Cupovic Jovana , Schmidt Sarah , Orlinger Klaus , Besse Andrej , Besse Lenka , Driessen Christoph , Cheng Hung-Wei , Lercher Alexander , Speiser Daniel , Bald Tobias , Bergthaler Andreas , Ludewig Burkhard
Published: February 2019

Immunotherapy revolutionized the treatment of cancer patients. However, the lack of tumor specific T-cells and the immunosuppressive tumor microenvironment remain the major obstacles in curing treatment-resistant tumors. Here, we show that a novel, propagating noncytopathic virotherapy expressing the tumor-associated antigen TRP2 can eradicate established tumors. Interestingly, this was dependent on the route of treatment. Systemic administration of gene-based virotherapy induced a high number of tumor-infiltrating TRP2 specific CD8+ T-cells but was not able to cure established tumors. Moreover, localized tumor therapy in the periphery cured also distant metastasis in the lung, indicating that the locally induced immune response generates a systemic antitumor effect. Localized virotherapy predominantly infects tumor cells and tumor-associated fibroblasts, resulting in a proinflammatory reprogramming of the tumor microenvironment. Our data reveal that this immune activation is dependent on type I IFN signaling on the host but not on the tumor cell. These results have important clinical implications: i) our data explain why T-cell transfer or T-cell vaccines alone do not cure established tumors; ii) intratumoral gene-based cancer vaccination is superior to systemic treatment; and iii) a successful local antitumor response is associated with an efficient systemic antitumor response. Directly cancer targeting noncytopathic gene-based vaccines may be a promising approach by simultaneously supercharging the suppressive tumor microenvironment and inducing an adaptive immune response against selected tumor antigens.

Citation Format: Lukas Flatz, Sandra Ring, David Bomze, Lucas Onder, Jovana Cupovic, Sarah Schmidt, Klaus Orlinger, Andrej Besse, Lenka Besse, Christoph Driessen, Hung-Wei Cheng, Alexander Lercher, Daniel Speiser, Tobias Bald, Andreas Bergthaler, Burkhard Ludewig. Virotherapy eradicates established melanoma by reprogramming the tumor microenvironment and engaging the adaptive immunity [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 A070.
Abstract A077: Microenvironmental factors shape resistance patterns to immune checkpoint blockade
Authors: Gu Shengquing Stan , Hu Xihao Sherlock , Wang Xiaoqing Shawn , Li Ziyi , Traugh Nicole , Bu Xia , Xing Xiaofang , Freeman Gordon , Brown Myles , Liu Xiaole Shirley
Published: February 2019

Immune checkpoint blockade (ICB) therapy has dramatically improved the prognosis of several types of cancer. However, only a small proportion of patients respond. Although multiple biomarkers/mechanisms of resistance to ICB have been identified, it remains elusive to what extent the cellular/molecular mechanisms contribute to the heterogeneity of ICB response. To this end, we applied clonal barcoding for lineage tracing of cancer cells following control IgG or ICB treatment in a transplantation mouse model. We identified significant clonal heterogeneity within cancer cells in response to ICB, suggestive of a minority of pre-existing ICB-resistant clones prior to treatment. Furthermore, counter-intuitively, ICB-responding tumors harbored a higher proportion of ICB-resistant clones than nonresponding tumors post-treatment, indicating that the tumor microenvironment might dictate the initial response to ICB. Integrated gene expression and immune repertoire analyses of the tumor microenvironment identified more T-cell and B cell infiltration in the ICB responders, and found that BCR class switch is associated with ICB response. Our study established a system to assess the contributions of cancer cell-intrinsic and microenvironmental factors in response to ICB treatment, and identified B cell infiltration and repertoire constitution as a novel biomarker for ICB response.

Citation Format: Shengquing Stan Gu, Xihao Sherlock Hu, Xiaoqing Shawn Wang, Ziyi Li, Nicole Traugh, Xia Bu, Xiaofang Xing, Gordon Freeman, Myles Brown, Xiaole Shirley Liu. Microenvironmental factors shape resistance patterns 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 A077.
Abstract A097: Complement system mutations in cancer: Uncovering new relationships between tumor hypoxia and complement
Authors: Olcina Monica M. , Balanis Nikolas G. , Kim Ryan K. , Thompson Michael J. , Graeber Thomas G. , Giaccia Amato J.
Published: February 2019

The complement system has been proposed to facilitate cancer hallmarks such as increased metastatic potential, proliferation and apoptosis evasion. Despite the association between complement and tumor progression, a detailed characterization of cancer genetic alterations in the complement system has not been performed to date. Here, we report a number of previously unappreciated mutations in complement system genes. Taken together as a pathway, mutations in complement genes occur at a relatively high frequency and across a number of cancer types. Notably, when grouping complement mutations into functionally relevant subgroups according to gene function, mutations and copy number alterations in genes within these subgroups are associated with changes in overall survival outcomes in a range of cancers. We use specific complement component mutations in colorectal cancer to uncover and experimentally validate crosstalk between complement and hypoxia, providing new associations between this innate immunity pathway and a prevalent component of the tumor microenvironment. Our data highlight the complex mechanism employed by cancers to manipulate the innate immune system and point to the potential use of complement system mutations in successful patient stratification into clinically and biologically relevant groups.

Citation Format: Monica M. Olcina, Nikolas G. Balanis, Ryan K. Kim, Michael J. Thompson, Thomas G. Graeber, Amato J. Giaccia. Complement system mutations in cancer: Uncovering new relationships between tumor hypoxia and complement [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 A097.
Abstract A102: Rescue of lost skin dendritic cells in melanoma is key for the resuscitation of antitumor T-cell responses
Authors: Prokopi Anastasia , Tripp Christoph H. , Tummers Bart , Komenda Kerstin , Hutter Katharina , Cappellano Giuseppe , Bellmann Lydia , Efremova Mirjana , Trajanoski Zlatko , Chen Suzie , Clausen Bjorn E. , Green Douglas R. , Stoitzner Patrizia
Published: February 2019

One major characteristic of skin-related cancers is the loss of skin-resident dendritic cell populations. In the tg(Grm1)EPv mouse model, ectopic expression of the metabotropic glutamate receptor-1 in melanocytes leads to a highly proliferative and antiapoptotic phenotype, resulting in melanoma formation within the dermis. The slow progression of these tumors allows for the in depth analysis of the immune infiltrate in growing tumors. We here show that total DCs (CD11c+ cells) are gradually lost as the tumor progresses. Mainly the dermal CD11b+ DCs are affected, whereas epidermal Langerhans cells remain unchanged. We hypothesized that extrinsic cell death of the DCs may be induced within the growing tumor. Indeed, the tumor tissue upregulated the expression of Fas ligand (FasL) that can induce extrinsic apoptosis in immature DCs that express Fas (CD95). Fas-mediated apoptosis depends on the activation of caspase-8 and mice in which the autoproteolytic cleavage site D387 is mutated to alanine (casp8D387A/casp8D387A) show strong resistance to Fas-mediated death. We compared the apoptosis induction in the different DC subsets in casp8WT/casp8WT and in casp8D387A/casp8D387A upon intradermal administration of an agonistic anti-CD95 antibody and we found that, indeed, CD11b+ DCs in WT mice are susceptible to apoptosis via CD95. In order to retrieve the intratumoral DCs, we treated tg(Grm1)EPv mice carrying tumor lesions with Flt3L and the percentages of the CD11b+ subset could be restored back to the levels of tumor-free mice. At the same time, T-cells from both the tumor and the tumor-draining lymph node were able to produce more cytotoxic cytokines. Our data thus indicate that the loss of DCs in melanoma depends on induction of apoptosis within the tumor; rescue of skin DCs may be a promising strategy in order to enhance the efficacy of existing immunotherapeutic strategies against skin-related cancers.

Citation Format: Anastasia Prokopi, Christoph H. Tripp, Bart Tummers, Kerstin Komenda, Katharina Hutter, Giuseppe Cappellano, Lydia Bellmann, Mirjana Efremova, Zlatko Trajanoski, Suzie Chen, Bjorn E. Clausen, Douglas R. Green, Patrizia Stoitzner. Rescue of lost skin dendritic cells in melanoma is key for the resuscitation of antitumor T-cell 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 A102.
Abstract A106: Higher numbers of cancer stem cells in the peripheral blood of children with B-ALL after chemotherapy
Authors: Salem Mohamed Labib , Attia Mohamed , Abdou Said , Zidan Abdel-Aziz A. , Zidan Mona F.
Published: February 2019

Background: Acute lymphocytic leukemia (ALL) is biologically and clinically considered as a heterogeneous neoplasm of lymphoid progenitor cells in the bone marrow (BM). 15- 20 % of children with ALL who achieve an initial remission, will show relapse. One potential mechanism behind this relapse could be the emergence of cancer stem cells (CSCs), which are considered the driving force of tumorigenesis due to their ability of self-renewal as well as the emergence of immune regulatory cells including myeloid-derived suppressor cells (MDSCs) and regulatory T-cells (Treg). Aim: The main aim of this study was to analyze the numbers of CSCs and correlate these numbers with the numbers of blasT-cells, MDSCs and Treg cells in children with B-ALL before and after induction of chemotherapy. Materials and Methods: CSCs were defined as CD45dimCD19+CD10+CD34+CD38-, MDSCs were defined as Lin-HLA-DR-CD33+CD11b+ and Treg cells were defined as CD4+CD25+CD127-. The frequencies of these cells were analyzed in the peripheral blood of B-ALL patients before (n= 10) and after (n= 10) induction of chemotherapy using flow cytometry. Results: Significant increases in the numbers of CSCs were shown in B-ALL patients after induction of chemotherapy as compared to newly diagnosed patients (7.6± 8.3 vs. 2.7± 2.4, P<0.05). The numbers of CSCs in ALL patents before and after induction of chemotherapy inversely correlated with the numbers of the blasT-cells. Additionally, the numbers of MDSCs and Treg cells were higher and lower, respectively, in patients after induction of chemotherapy as compared to before chemotherapy. Conclusion: Our results indicate that chemotherapy of B-ALL patients results in emergence of high numbers of CSCs and MDSCs, which might be contributing, respectively, to tumor relapse and creation of systemic immune suppression. This pilot study opens a new avenue to investigate the mechanism mediating the emergence of these cells on larger population of B-ALL patients at different treatment stages.

Citation Format: Mohamed Labib Salem, Mohamed Attia, Said Abdou, Abdel-Aziz A. Zidan, Mona F. Zidan. Higher numbers of cancer stem cells in the peripheral blood of children with B-ALL after chemotherapy [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 A106.
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: Leshem Yasmin , King Emily , Reiter Yoram , Pastan Ira
Published: February 2019

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.
Abstract A146: Systematic discovery of immune regulatory mechanisms in tumor cells
Authors: Pan Deng , Kobayashi Aya , Jiang Peng , Yuan Guo-Cheng , Liu X. Shirley , Doench John , Qiu Xintao , Rao Prakash , Long Henry , Brown Myles A. , Wucherpfennig Kai W. , Andrade Lucas Ferrari de , Tay Rong En , Luoma Adrienne M. , Tsoucas Daphne , Lim Klothilda
Published: February 2019

Many human cancers are resistant to immunotherapy for reasons that are poorly understood. We used a genome-scale CRISPR/Cas9 screen to identify mechanisms of tumor cell resistance to killing by cytotoxic T-cells, the central effectors of antitumor immunity. Inactivation of >100 genes sensitized mouse B16F10 melanoma cells to killing by T-cells, including Pbrm1, Arid2 and Brd7, which encode components of the PBAF form of the SWI/SNF chromatin remodeling complex. Loss of PBAF function increased tumor cell sensitivity to interferon-γ, resulting in enhanced secretion of chemokines that recruit effector T-cells. Treatment-resistant tumors became responsive to immunotherapy when Pbrm1 was inactivated. In many human cancers, expression of PBRM1 and ARID2 inversely correlated with expression of T-cell cytotoxicity genes, and Pbrm1-deficient murine melanomas were more strongly infiltrated by cytotoxic T-cells.

Citation Format: Deng Pan, Aya Kobayashi, Peng Jiang, Guo-Cheng Yuan, X. Shirley Liu, John Doench, Xintao Qiu, Prakash Rao, Henry Long, Myles A. Brown, Kai W. Wucherpfennig, Lucas Ferrari de Andrade, Rong En Tay, Adrienne M. Luoma, Daphne Tsoucas, Klothilda Lim. Systematic discovery of immune regulatory mechanisms in tumor cells [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 A146.
Abstract A167: Prevalence of TREG cells and effects of their inhibition on growth of oral cancer cells
Authors: Aggarwal Sadhna , Sharma Suresh C. , Das Satya N.
Published: February 2019

Oral squamous cell carcinoma (OSCC) is one of the major cancers affecting Asian countries. The main causative factor has been tobacco habit. It has been reported that immune dysfunction in these patients is one of the major factors for tumor growth and dissemination that affects disease-free survival of the patients. We assessed the phenotypic and functional characteristics of regulatory T (Treg) CD4+CD25+FoxP3+subsets in patients with OSCC by multicoloured flow cytometry. Subsequently we investigated the effects their inhibition via TDG on growth of OSCC cell lines in vitro. An increased (p<0.05) prevalence of Treg phenotypes (CD4+CD25+, CD4+FoxP3+, CD8+FoxP3+, CD4+CD25+FoxP3+) was observed in the peripheral circulation of OSCC patients that positively correlated with clinicopathologic features. The increased frequency of CD4+CD8+CD25+FoxP3+, a unique T-cell subset, CTLA4+, GITR+, NrP1+ and granzyme B+ (GzmB) Tregs also showed a significantly higher prevalence in OSCC patients. Functionally, CD4+FoxP3+ Tregs showed skewed expression of IL2, IL10 and IL35 in patients as compared with the normal controls. Higher expression of TGFβ in tumor tissues suggests their dominant role in the up regulation of differentiation of Tregs from naive T-cells in the tumor bearing host. Further, enhanced expression of CCR5 and CCR7 on Tregs with up regulation of their ligands (CCL5, CCL19 and CCL21) in tumor cells indicates efficient recruitment and trafficking of Tregs to the tumor site. Treatment with βGBP showed growth-promoting effects on Tregs and oral cancer cells. However, the treatment with its inhibitor TDG resulted in inhibition of Treg subsets and also decreased the frequency of IL10+ and IL35+ Tregs, indicating its immunomodulatory effects. Hence, it seems reasonable to assume that modulation of functional dynamics of selective Treg subsets may be useful in enhancing antitumor immunity and developing immunotherapeutic strategies for patients with oral squamous cell carcinoma.

Citation Format: Sadhna Aggarwal, Suresh C. Sharma, Satya N. Das. Prevalence of TREG cells and effects of their inhibition on growth of oral cancer cells [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 A167.
Abstract A176: Overcoming CD8 T-cell suppression in the tumor microenvironment
Authors: Cartwright Adam N.R , Jiang Peng , Saadatpour Assieh , Yuan Guo-Cheng , Liu Shirley X. , Wucherpfennig Kai W.
Published: February 2019

CD8 T-cell-mediated antitumor immunity is required for the control and elimination of tumors. However, tumors are able to overcome immune response resulting in immunosuppression, tumor growth, and metastatic spread. CD8 T-cells are controlled through a homeostatic network of positive and negative feedback loops. These negative signals are exploited by the tumor and associated suppressive cell populations in the tumor microenvironment. Immunotherapies targeted to receptors that control these negative signals, termed immune checkpoint inhibitors, have provided robust and durable responses to a number of cancers. Unfortunately, only a subset of patients will respond to current immunotherapies. This is due to the accruement of suppressive and inhibitory mechanisms employed by the tumor and its microenvironment. These include expression of inhibitory ligands, release of immune-suppressive factors, and the recruitment of suppressive cell populations. Furthermore, the administration of immune checkpoint inhibitors can enhance tumor acquisition of a more suppressive phenotype, diminishing immune responses through additional inhibitory pathways. We therefore propose that further CD8 T-cell-intrinsic negative regulators most likely exist, which can be targeted to improve antitumor responses. Here, we have used functional genomic approaches to identify pathways that can be targeted to advance anti-tumor responses, either in combination with anti-PD-1 or to overcome suppressive mechanisms employed by the tumor microenvironment. Using both in vivo and in vitro assays, we have identified both a novel target that provides an enhancement to CD8 T-cell effector function when in combination with PD-1 checkpoint blockade, and CD8 T-cell-intrinsic pathways that are modulated by the presence of suppressive cells associated with the tumor microenvironment.

Citation Format: Adam N.R Cartwright, Peng Jiang, Assieh Saadatpour, Guo-Cheng Yuan, Shirley X. Liu, Kai W. Wucherpfennig. Overcoming CD8 T-cell suppression in the tumor microenvironment [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 A176.
Abstract A177: Impact of regulatory T-cells on carcinogenesis of oral squamous cell carcinoma
Authors: Chao Jaime L. , Savage Peter A.
Published: February 2019

Although some head-and-neck squamous cell carcinomas (HNSCCs) are associated with human papillomavirus (HPV) infection, a large proportion of HNSCCs are HPV-negative and are characterized by mutational signatures associated with smoking, suggesting that HPV-negative HNSCCs are induced by carcinogen exposure. In many human cancers, including HNSCC, regulatory T (Treg) cells are found at elevated densities and are thought to be a major barrier to the generation of robust anti-tumor T-cell responses. However, recent studies indicate that Treg cells can serve diverse functions in non-lymphoid sites. Therefore, it is critical to elucidate the function of Treg cells infiltrating HNSCC lesions. To study the functional impact of Treg cells on the development and progression of HPV-negative HNSCC, we utilized an autochthonous mouse model of carcinogen-induced oral squamous cell carcinoma (SCC), in which mice are exposed long-term to the carcinogen 4-nitroquinoline-1-oxide (4-NQO) in the drinking water. Immunohistochemistry and flow cytometric analysis revealed elevated densities of CD3+ T-cells and Foxp3+ Treg cells in premalignant dysplastic lesions and SCC lesions within the tongue epithelium, suggesting that many lesions exhibit a T-cell-inflamed phenotype. Single-cell TCR sequencing demonstrated the presence of recurrent Treg cell clones within carcinogen-induced lesions, suggesting clonal expansion of Treg cells in the tumor microenvironment. Surprisingly, the systemic depletion of Treg cells at the later stages of carcinogen exposure did not induce tumor regression, but instead induced the opposite effect, driving increased incidence and burden of SCC. These findings suggest that Treg cells may play an unexpected role in restricting the progression of carcinogen-induced SCC in the oral cavity, prompting a re-examination of the common paradigm that Treg cells promote tumorigenesis by suppressing antitumor immunity. Clinically, our findings suggest that novel therapies could be developed to combat HNSCC by triggering the recruitment or activation of tumor-associated Treg cells, and suggest that checkpoint blockade therapies that induce intratumoral Treg cell depletion may have unintended adverse consequences.

Citation Format: Jaime L. Chao, Peter A. Savage. Impact of regulatory T-cells on carcinogenesis of oral squamous cell carcinoma [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 A177.
Abstract A189: Targeting T-cell epigenetic programs to enhance the efficacy of immune checkpoint blockade
Authors: Ghoneim Hazem E. , Fan Yiping , Moustaki Ardiana , Abdelsamed Hossam , Dash Pradyot , Dogra Pranay , Carter Robert , Awad Walid , Neale Geoff , Thomas Paul , Youngblood Ben
Published: February 2019

Immune checkpoint blockade (ICB)-mediated rejuvenation of exhausted T-cells has emerged as a promising approach for treating various cancers and chronic infections. However, T-cells that become fully exhausted during prolonged antigen exposure remain refractory to ICB-mediated rejuvenation. Given that many of the impaired effector properties of terminally exhausted CD8 T-cells appear to be heritably maintained even in the absence of antigen, we investigated the role of de novo DNA methylation programming as a T-cell-intrinsic mechanism for establishing the ICB-nonresponsive state of T-cell exhaustion. Whole-genome bisulfite sequencing of antigen-specific murine CD8 T-cells at the effector and exhaustion stages of an immune response identified progressively acquired heritable de novo DNA methylation programs. Blocking de novo DNA methylation in activated CD8 T-cells allowed them to retain their effector functions despite persistent stimulation during chronic viral infection. We found that these de novo epigenetic programs are not only critical for establishing T-cell exhaustion, but also restrict T-cell expansion and clonal diversity during PD-L1 blockade treatment. Moreover, these exhaustion-associated DNA methylation programs were acquired in tumor-infiltrating PD-1hi CD8 T-cell. Therapeutic approaches to reverse these programs enhanced antitumor CD8 T-cell responses and tumor control during PD-L1 blockade therapy. These data establish de novo DNA methylation programming as a major T-cell-intrinsic barrier of ICB therapy. Targeting T-cell epigenetic programs provides great potential for fostering more powerful T-cell immunotherapies. Reference: Ghoneim, et al. Cell 2017.

Citation Format: Hazem E. Ghoneim, Yiping Fan, Ardiana Moustaki, Hossam Abdelsamed, Pradyot Dash, Pranay Dogra, Robert Carter, Walid Awad, Geoff Neale, Paul Thomas, Ben Youngblood. Targeting T-cell epigenetic programs to enhance the efficacy of 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 A189.
Abstract A215: Thymocyte selection-associated HMG box protein TOX is a master regulator of tumor-specific T-cell dysfunction
Authors: Scott Andrew C. , Camara Steven , Lauer Peter , Synder Alexandra , Zamarin Dmitriy , Walther Tyler , Levy Olivier , Glickman Michael , Kaye Jonathan , Philip Mary , Schietinger Andrea
Published: February 2019

Tumor-specific CD8 T-cells in cancers enter a state of dysfunction characterized by the expression of inhibitory receptors and failure to produce effector cytokines and cytotoxic molecules. Here we identify the nuclear factor, Thymocyte selection-associated HMG box protein, TOX, as a master regulator of tumor-specific T-cell dysfunction. TOX is uniquely expressed in dysfunctional CD8 T-cells from mouse and human tumors but absent in functional T-cells. TOX expression is driven by continuous TCR stimulation and NFAT activity. Forced expression of TOX in functional effector T-cells was sufficient to induce a transcriptional program of dysfunction through the concerted expression of genes encoding numerous inhibitory receptors and dysfunction-associated transcription factors. Notably, TOX-deficient tumor-infiltrating T-cells did not upregulate inhibitory receptors such as PD1, LAG3, CD38, or CD39 and maintained high TCF1 expression. Surprisingly, despite their normal, “non-exhausted” phenotype, TOX-deficient T-cells failed to make effector cytokines, suggesting that loss of effector function in tumor-specific T-cells is uncoupled from inhibitory receptor expression. Furthermore, TOX-deficient T-cells failed to persist in tumors, ultimately undergoing activation-induced cell death. We propose that the TOX-induced transcriptional program of hyporesponsiveness is a physiologic negative feedback mechanism that prevents overstimulation; thus TOX is absolutely required for T-cell survival in the setting of chronic antigen stimulation as in cancers.

Citation Format: Andrew C. Scott, Steven Camara, Peter Lauer, Alexandra Synder, Dmitriy Zamarin, Tyler Walther, Olivier Levy, Michael Glickman, Jonathan Kaye, Mary Philip, Andrea Schietinger. Thymocyte selection-associated HMG box protein TOX is a master regulator of tumor-specific T-cell dysfunction [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 A215.
Abstract A216: Functionally specialized subsets of exhausted CD8+ T-cells mediate tumor control and response to checkpoint blockade
Authors: Sen Debattama R. , Miller Brian C. , Abosy Rose Al , Bi Kevin , LaFleur Martin W. , Yates Kathleen B. , Lako Ana , Felt Kristen D. , Naik Girish S. , Manos Michael , Gjini Evisa , Virkud Yamini V. , Hodi Stephen , Rodig Scott J. , Sharpe Arlene H. , Haining W. Nicholas
Published: February 2019

T-cell dysfunction in the tumor microenvironment (TME) is a hallmark of many cancers. Reinvigoration of T-cell function by PD-1 checkpoint blockade can result in striking clinical responses, but is effective only in a minority of patients. The basis for T-cell dysfunction in the TME, as well as the mechanisms by which anti-PD-1 therapy acts on dysfunctional T-cells are not fully understood. Here we show that anti-PD-1 therapy acts on a specific subpopulation of CD8+ tumor-infiltrating lymphocytes (TILs) in melanoma mouse models as well as patients with melanoma. We find that dysfunctional CD8+ TILs possess canonical epigenetic and transcriptional features of T-cell exhaustion, mirroring those seen in chronic viral infection. Similar to chronic viral infection, exhausted CD8+ TILs contain a subpopulation of “stem-like exhausted” T-cells that have a distinct regulatory state. Stem-like exhausted TILs also have critical functional attributes that are not shared by the majority “terminally exhausted” TILs: they retain more polyfunctionality, persist following transfer into tumor-bearing mice, and differentiate to repopulate terminally exhausted TILs in the TME. As a result, stem-like exhausted CD8+ TILs are better able to control tumor growth than terminally exhausted cells. Stem-like exhausted, but not terminally exhausted, CD8+ TILs can respond to anti-PD-1 therapy without reversion of their exhausted epigenetic state. CD8+ T-cells with a stem-like exhausted phenotype can be found in human melanoma samples and patients with a higher fraction of this subpopulation in their tumors have a significantly longer duration of response to combination checkpoint blockade therapy. Responsiveness to checkpoint blockade is therefore restricted to a subpopulation of exhausted TILs that retain specific functional properties which enable them to control tumors. Approaches to expand stem-like exhausted CD8+ T-cells in the tumor microenvironment may be an important component of improving checkpoint blockade response.

Citation Format: Debattama R. Sen, Brian C. Miller, Rose Al Abosy, Kevin Bi, Martin W. LaFleur, Kathleen B. Yates, Ana Lako, Kristen D. Felt, Girish S. Naik, Michael Manos, Evisa Gjini, Yamini V. Virkud, Stephen Hodi, Scott J. Rodig, Arlene H. Sharpe, W. Nicholas Haining. Functionally specialized subsets of exhausted CD8+ T-cells mediate tumor control and response to 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 A216.