bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3228: An online tool to balance baseline covariates for mouse clinical trials
Authors: Mao Binchen , Ouyang Davy , Li Henry , Guo Sheng
Published: September 2020

INTRODUCTION: Randomization is a cornerstone of controlled studies such as mouse clinical trials (MCT). However, simple randomization schemes often lead to unbalanced group allocations, therefore reducing the statistical power of experimental studies. This issue becomes more relevant and critical in MCTs where it is usually necessary to allocate a large number of mice into multiple groups/arms, so that all groups have close tumor volumes (measured by average and variance) as well as other covariates such as body weight, while at the same time also using as few mice as possible. Possible allocations grow exponentially by mouse and group number, and it is computationally intractable to identify optimal allocation schemes; instead, heuristic methods are used to search for near-optimal solutions. Studylog® systems can balance tumor volume with at most one baseline covariate. Randmice, a newly developed tool by Stimunity, shows good performance in balancing bilateral tumors, but it cannot balance more than two baseline covariates. We have developed a fast, in-house, online tool to balance any number of baseline covariates and, in addition, users can assign different weights to each baseline covariate based on levels of importance.

METHODS: We used a two-step method to balance designated baseline covariates for MCTs. First, we used a matched algorithm to find sub-matches from a population of mice, by grouping together mice with similar values of baseline covariates. Based on these sub-matches, we then used an optimization method to minimize the imbalance of baseline covariates guided by a weighted score function. Once the optimization was complete, a summary statistic table and a diagnostic plot for the top 10 group allocation solutions were displayed for picking the most suitable solution.

RESULTS: We used our baseline covariate balancing tool for a benchmark set of mouse clinical trials with more than one baseline covariate to be balanced out, with the difference of key baseline covariates among groups well under control with close means. The tool completed the allocations in a few minutes by running on a single-CPU computer.

CONCLUSIONS: We have developed a fast and powerful online tool to help preclinical researchers balance key baseline covariates in their studies, particularly in mouse clinical trials.

Citation Format: Binchen Mao, Davy Ouyang, Henry Li, Sheng Guo. An online tool to balance baseline covariates for mouse clinical trials [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3228.
Abstract 3232: In vitro cell based cytotoxicity and T cell activation assays to assess safety and efficacy of engineered T cell therapies
Authors: Holt Sanne , Vermond Sophie , Hazenoot Monique Hazenoot , Overman Jeroen , Aarbiou Jamil Aarbiou , DeGroot Jeroen DeGroot
Published: September 2020

Chimeric antigen receptor (CAR) and T cell receptor (TCR) engineered T cells are part of a big wave of immunotherapies showing great promise in cancer clinical trials. With the first T cell based therapies targeting hematological malignances now approved, their next challenge is solid tumors. Solid tumors create an additional challenge due to the lack of tumor specific target antigens, posing significant safety risks, i.e. on-target on-tumor, on-target off-tumor and off-target toxicities. Toxic effects previously reported vary from mild-severe cytokine release syndrome (CRS) to neurotoxicity and death.

We have developed in vitro assays utilizing primary human cells from healthy tissue and/or differentiated iPCS-derived cells to assess on-target off-tumor and/or off-target cytotoxicity for engineered T cell therapies. The presence of CAR-T cell mediated cytotoxicity was measured through co-culture with healthy human primary cells to assess unwanted CAR-T reactivity as well as a high target antigen expressing control cancer cell line to confirm CAR-T functionality. The human primary cell type was selected based on its potential safety risk by establishing low level protein expression of the target antigen. Readouts included IFNγ production determined by MesoScale Discovery platform as a measure of T cell activation and Hoechst/PI staining of target cells by flow cytometry.

Our study generated high quality data of the CAR-T cell, confirming functionality by showing consistent T cell activation and killing against a positive control cell line. Moreover, we were able establish a clear absence of activity against the primary human cells thus providing insight into the safety of the CAR-T therapy.

The developed safety assays provide a robust and rapid platform to assess on-target off-tumor and off-target effects within immuno-oncology therapies, either TCR or CAR-T cells, in both early stage development or late stage testing of the therapeutic product. Through inclusion of a wide range of human primary cells, both high risk tissues and major organs at risk of off-target toxicity, a clear safety profile can be generate in vitro for these novel T cell therapies. Safety risks associated with cell based IO-therapies is the biggest challenge for the success of these therapies, performing a thorough safety screen on healthy primary human tissues is therefore crucial.

Citation Format: Sanne Holt, Sophie Vermond, Monique Hazenoot Hazenoot, Jeroen Overman, Jamil Aarbiou Aarbiou, Jeroen DeGroot DeGroot. In vitro cell based cytotoxicity and T cell activation assays to assess safety and efficacy of engineered T cell therapies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3232.
Abstract 3247: Development of CAR-T cell therapy for B-ALL using a point-of-care approach
Authors: Abdo Luiza , Barros Luciana Rodrigues , Viegas Mariana Saldanha , Marques Luisa Vieira , de Sousa Ferreira Priscila , Chicaybam Leonardo , Bonamino Martín Hernán
Published: September 2020

Introduction: CD19-targeted CAR-T cell immunotherapy is now an approved treatment for B cell leukemias and lymphomas. Current protocols for producing CAR-T cells use viral vectors for genetic modification and take 12-15 days to expand sufficient numbers of T cells, constituting an important bottleneck for the widespread use of this therapy. The development of non-viral, plasmid-based systems like Sleeping Beauty (SB) transposons, coupled with the emergence of large scale, closed electroporation devices, present the opportunity of generating high numbers of CAR-T cells in a short period of time and transforming this in a point-of-care (POC) therapy.

Methods and Results: In this work, mononuclear cells were isolated using Ficoll and electroporated using Nucleofector IIb electroporator combined with plasmids encoding 19BBz CAR (in the pT3 SB transposon backbone) and SB100x transposase. Cells were rested for 24h and used for in vitro and in vivo experiments. The phenotype was assessed by flow cytometry. The in vitro cytotoxicity assay was performed using Calcein-AM dye on target cells incubated with different ratios of effector cells. 8-12-week-old-female-NSG were injected iv. 5 × 106 RS4;11 GFP or 105 Nalm-6 GFP and after 3 days were treated with different doses of recently electroporated CAR-T cells. CAR expression on day 1 following electroporation ranged between 5-15% and in vitro cytotoxic activity against RS4;11 or Nalm-6 CD19+ B cell precursor leukemia lines was low or absent. NSG mice engrafted with RS4;11 and treated with 1 × 105 19BBz+ cells showed improved survival when compared to mice treated with mock electroporated cells. POC approach was also efficient against Nalm-6, where 19BBz+CAR-T cells (7 × 105 per mice) improved the survival of mice. In both models, decreased tumor burden in blood and spleen was observed in mice treated with 19BBz. Head to head comparison of 19BBz cells used in POC approach or expanded for 8-12 days in vitro showed similar antitumor activity in vivo against RS4;11 cells, leading to equivalent improvements in mice survival.

Conclusion: In summary, we show that the POC strategy is a viable approach for the use of CAR-T cells, obviating the need for T cell expansion while having the potential to greatly decrease the total cost and time of CAR-T cell therapy manufacturing.

Citation Format: Luiza Abdo, Luciana Rodrigues Barros, Mariana Saldanha Viegas, Luisa Vieira Marques, Priscila de Sousa Ferreira, Leonardo Chicaybam, Martín Hernán Bonamino. Development of CAR-T cell therapy for B-ALL using a point-of-care approach [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3247.
Abstract 326: Differences of drug sensitivities between 2-dimensional and 3-dimensional culture systems in triple negative breast cancer cell lines
Authors: Muguruma Masako , Teraoka Saeko , Miyahara Kana , Ueda Ai , Kawate Takahiko , Ishikawa Takashi
Published: September 2020

[BACKGROUND] In vitro chemosensitivity tests are not always consistent to clinical results (Hutchinston L et al., Nat Rev Clin Oncol. 2011), partly because of lacking extracellular matrix-cell interactions in 2-dimensional (2D) cell culture system (Antoni D et al., Int J Mol Sci. 2015). Three-dimensional cell culture (3D) system maintains cells with a higher degree of structural complexity and homeostasis, analogous to tissues and organs compared with 2D cell culture system. Thus, 3D cell culture system will be expected for alteration of the animal experiment in chemosensitivity test (Bokhari M et al., J Anat. 2007). Cytotoxic agents are crucial for treating triple negative breast cancer (TNBC).[AIM] To clarify differences of drug sensitivities between 2D and 3D culture in TNBC cell lines, we compared IC50s for Cisplatin (CDDP), Docetaxel (DTX), Epirubicin (EPI) and Bromodomain inhibitor (JQ1).

[METHODS] Thirteen TNBC cell lines were examined; HCC1599, HCC1937, HCC38, MDA-MB-468, HCC1806, HCC70, MDA-MB-436, MDA-MB-231, Hs578T, BT549, MDA-MB-453, BT20, and HCC1395. For 2D culture, cells were plated on general 96 well cell culture plate for 24 hours. For 3D culture, cells were plated on the PrimeSurface® 96 well plate (SUMITOMO BAKELITE) for 72 hours. Then, those were incubated in media including CDDP, DTX, EPI or JQ1 for 72 hours. The cell viability was quantified using CellTiter-Glo 2.0 and CellTiter-Glo 3D cell viability Assay (Promega), respectively.

[RESULTS] For CDDP, IC50s were highly correlated between 2D and 3D culture (r=0.928) . The mean of IC50 value showed higher in 3D culture (27.67±8.40μM) than 2D culture (12.86 ± 17.23μM). For EPI, the correlation was r=0.593. The mean value showed higher in 3D culture (1.41±1.30μM) than 2D culture (0.62±0.72μM). For DTX and JQ1, the mean values were 1.78 ± 2.05nM and 4.31 ±4.72μM in 2D culture, respectively. However, IC50 failed to be obtained in several TNBC cell lines (HCC1937, HCC38, HCC1806, Hs578T and HCC1395 for JQ1 and MDA-MB-436, MDA-MB-453 and BT20 for DTX) in 3D culture.

[CONCLUSION] IC50 value was higher in 3D culture than in 2D culture for CDDP and EPI. The correlation of IC50s between 2D and 3D depended on drugs with the highest in CDDP. We are under investigation on reasons why sensitivity tests for DTX and JQ1 failed in 3D culture system.

Citation Format: Masako Muguruma, Saeko Teraoka, Kana Miyahara, Ai Ueda, Takahiko Kawate, Takashi Ishikawa. Differences of drug sensitivities between 2-dimensional and 3-dimensional culture systems in triple negative breast cancer cell lines [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 326.
Abstract 3265: Anti-GITR agonist antibody for cancer immunotherapy
Authors: Yang Teddy , Dai Ken , Wang Dongxu , Qi Qianqian , Liu Hu
Published: September 2020

Glucocorticoid-induced TNFR-related (GITR) is a gene coding for a member of the TNF receptor superfamily. GITR is expressed at low levels on resting responder T lymphocytes and is up-regulated in T regulatory cells (Treg cells) and in activated T cells. GITR agonist which upregulated co-stimulatory signal to T cells or inhibition of Treg induced suppression which reverts Treg proliferation, is likely to enhance immunity in response to infection and tumor immunity. Therefore, GITR is currently of interest to immunologists as a co-stimulatory immune checkpoint molecule. In this study, various types of GITR immunogens, including gene, protein and cell line, are used for H2L2 mice immunization. Then we generate and characterize panels of anti-GITR monoclonal antibodies which not only could specifically bind to human GITR, but also show cross-reactivity to cyno GITR. Such anti-GITR antibody can increase the expression of NF-κB gene by cross-linking by anti-Fab, and induce the release of INF-γ from T cell. In harsh pH or temperature condition, the anti-GITR antibody can keep the stability. And at high concentration, the viscosity of the anti-GITR antibody is good. Moreover, in vivo study showed that the anti-GITR antibody can effectively inhibit tumor growth. Above results demonstrated that the anti-GITR antibody is expected to develop further for clinical study single or combine with other immune checkpoint antibody.

Citation Format: Teddy Yang, Ken Dai, Dongxu Wang, Qianqian Qi, Hu Liu. Anti-GITR agonist antibody for cancer immunotherapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3265.
Abstract 3298: Eosinophilia and improved survival in non-hematologic malignancies treated with immune checkpoint inhibitors
Authors: Mitchell Jasmine L. , Moyers Justin , Cao Huynh
Published: September 2020

Background

Immune checkpoint inhibitors (ICI) have become a mainstay treatment of a variety of advanced malignancies, prolonging survival. There are no serologic markers to predict response to ICI, but some small series have suggested eosinophilia predicts improved survival.

Methods

This is an IRB approved single-center retrospective observational study including all patients (n= 80) from 2014-2019 who received ICI for non-hematologic malignancies at Loma Linda University Medical Center. Data were collected via chart review. Clinically significant increase of absolute eosinophil count was defined as a count > 0.5 bil/L on at least two occasions during treatment. Kaplan-Meier survival estimates were used to estimate overall survival.

Results

In non-small cell lung cancer (n= 35), patients with eosinophilia (n=5) had no death events compared to the median survival of 17.6 months in those without eosinophilia (p = 0.02). Similarly, in patients with renal cell carcinoma (n=11), those with eosinophilia (n=2) had no death events while the median survival for those without was 16.9 months (p= 0.24). In the metastatic melanoma cohort (n=9), those with eosinophilia (n= 5) had a median overall survival of 45.4 vs 9.9 months in those without (p= 0.02).

Conclusions

In both NSCLC and metastatic melanoma, eosinophilia during ICI therapy was associated with significantly increased overall survival, while in RCC there was a trend towards improved survival. Interestingly, there were no death events in those with eosinophilia in either NSCLC or RCC cohorts.

Citation Format: Jasmine L. Mitchell, Justin Moyers, Huynh Cao. Eosinophilia and improved survival in non-hematologic malignancies treated with immune checkpoint inhibitors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3298.
Abstract 33: Epidemiology and clinicopathology of T-cell intravascular lymphoma (TIVL): Making the case for extra-nodal NK-T cell lymphoma-angiotropic type as a provisional entity
Authors: Haddad Philip A. , Gallagher Kevin , Hammoud Dalia
Published: September 2020

TIVL is a rare and often aggressive NK/T-cell lymphoma characterized by the presence of neoplastic lymphocytes that are restricted to the lumen of small vessels. It comprises 8-10% of all IVL. By the time a diagnosis is made, TIVL tends to be widely disseminated across multiple extra-nodal sites with frequent catastrophic consequences. In order to study the demographic characteristics, presentation and progression patterns, survival, and prognostic factors, we constructed a database that pooled cases between 1986 and 2019. Kaplan-Meier survival curves were constructed. Cox proportional-hazards model and Log-rank tests were used to assess the influence of demographic and clinical factors on survival. A total of 81 patients with confirmed TIVL were identified. The median age was 53 (17-87) years with a peak incidence between ages 50 and 60. There was a slight male predominance with a male to female ratio of 1.5. Comprising 43% of reported cases, Asians were the most afflicted ethnic group. Skin (71%) was the most common site involved with TIVL followed by CNS (23%), and pulmonary and bone marrow involvement (8% each). The median overall survival of the whole group was 7 months. Sex and age did not seem to affect survival rates. When compared to supportive care, chemotherapy significantly improved survival (HR:0.15, p<0.0001). Skin only involvement had the highest survival rates whereas CNS and pulmonary involvement had the worst. When skin only involvement was associated with constitutional symptoms survival rates significantly declined. In a multivariate analysis, skin only involvement (HR:0.43, p=0.009) and not receiving treatment (HR:3.45, p=0.001) continued to have a significant association with survival. CD56+, CD30-, and TCR clonality each seems to adversely impact survival. However, they were of borderline non-statistical significance. Only the detection of EBER in the sample was found to significantly impact survival in a negative manner (p=0.016). Our results indicate that TIVL is an NK/T cell entity with distinct angiotropic pathology and aggressive clinical behavior and often poor outcomes. TIVL's B-cell counterpart is already recognized by the WHO lymphoma classification as a separate B-cell entity. It is our hope that these results serve as the basis for establishing Extra-Nodal NK/T Cell Lymphoma-Angiotropic Type as a provisional entity in order to further study and characterize this rare disease.

Citation Format: Philip A. Haddad, Kevin Gallagher, Dalia Hammoud. Epidemiology and clinicopathology of T-cell intravascular lymphoma (TIVL): Making the case for extra-nodal NK-T cell lymphoma-angiotropic type as a provisional entity [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 33.
Abstract 3320: Humanized NSG-Tg(Hu-IL15) mice support preclinical immune-oncology efficacy for testing of NK cells based immunology
Authors: Yao Li-Chin , Cheng Mingshan , Cai Danying , Shultz Leonard D. , Keck James G.
Published: September 2020

The JAX® Onco-Hu® platform utilizes humanized mice engrafted with tumors to enable in vivo investigation of the interactions between the human immune system and human cancer. A major avenue of our investigation is to generate humanized mouse models containing a more complete human hematopoietic system and robust innate immune cell population. Nature killer (NK) cells are important players of innate defense against cancerous cells by a number of mechanisms, including antibody-dependent cell mediated cytotoxicity (ADCC). To overcome limited NK cell development in humanized NSG™ mice, we have developed NSG-Tg(Hu-IL15) mice constitutively expressing human IL15 in a physiological level. Following CD34+ HSC-engraftment of NSG-Tg(Hu-IL15) mice, significantly higher levels of human NK cells are detectable in peripheral blood as compared to NSG mice and the engraftment is stable for over six months. Levels of circulating human CD45+ cells, CD3+ T cells, and CD33+ myeloid cells are similar between the HSC-engrafted NSG-Tg(Hu-IL15) and NSG mice. Minimal baseline activation of NK cells is detected in the spleen and peripheral blood. Previously we have described that the human NK cells developing in HSC-engrafted NSG-Tg(Hu-IL15) mice are functional and can lyse the NK-sensitive target cells K562 by in vitro cytotoxicity assay. We also showed that NK cells directly limited growth of a PDX melanoma xenograft and NK cell depletion by antibody blockage largely abrogated the delayed PDX growth. Here we extended these observations by evaluating tumor growth of multiple PDX and CDX models in humanized NSG-Tg(Hu-IL15) mice. We found that not all tumors were able to grow and for those that grew some spontaneous regression might still occur. In ex vivo experiments NK cells in the PBMCs isolated from humanized NSG-Tg(Hu-IL15) mice could be activated by Her2+ human cancer cells in the presence of anti-human H2 antibody (Trastuzumab). Furthermore we showed in vivo that humanized NSG-Tg(Hu-IL15) mice mediated efficient ADCC against Daudi B lymphoma cells using anti-human CD20 antibody (Rituximab). Administration of rituximab resulted in significant tumor growth inhibition as measured by both tumor volume and tumor weight. Together these data demonstrate that HSC-engrafted NSG-Tg(Hu-IL15) mice support enhanced development of functional human NK cells and that this mouse model enables preclinical evaluation of NK cell-targeted cancer immunotherapy in vivo.

Citation Format: Li-Chin Yao, Mingshan Cheng, Danying Cai, Leonard D. Shultz, James G. Keck. Humanized NSG-Tg(Hu-IL15) mice support preclinical immune-oncology efficacy for testing of NK cells based immunology [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3320.
Abstract 3348: Exogenous DNA upregulates DUOX2 expression in pancreatic cancer via activation of the cGAS-STING signaling pathway
Authors: Wang Stephen L. , Wu Yongzhong , Antony Smitha , Juhasz Agnes , Meitzler Jennifer , Jiang Goujian , Dahan Iris , Lu Jiamo , Roy Krishnendu , Doroshow James
Published: September 2020

Dual oxidase 2 (DUOX2) is a transmembrane protein that produces hydrogen peroxide in the extracellular environment, mediates innate immunity at mucosal surfaces, contributes to chronic inflammation-related tissue injury and angiogenesis, and can promote the initiation of epithelial-derived cancers. In pancreatic adenocarcinomas (PDAC), the composition of the tumor microbiome significantly affects patient outcomes. However, it remains unclear how microbes in the tumor microbiome affect PDAC on a cellular level. In recent years, the cGAS-STING signaling pathway has been identified as a crucial mediator of anti-viral defense by recognizing the presence of DNA in the cytosol and subsequently producing Type I interferons (IFNs) as well as a host of other anti-viral genes. Using Western blot and qPCR, we show that the introduction of exogenous DNA into the cytosol significantly upregulates DUOX2 expression in human PDAC cells after activating cGAS-STING signaling. Expression of DUOX2 occurs most strongly 2 days post-transfection of DNA, while cGAS-STING signaling is activated within hours. Treatment of BxPC-3 cells with exogenous cyclic GMP-AMP (cGAMP) for 24 hrs increases DUOX2 expression in a concentration-dependent manner, suggesting the necessity of cGAS activity for DUOX2 induction. Furthermore, Western analysis of phospho-NF-κB p65 in BxPC-3 cells demonstrates activation of NF-κB that mirrors the time-dependent induction of DUOX2. In CFPAC-1 cells, another PDAC line, Jak-Stat signaling is activated along with the components of cGAS-STING. These data suggest that DNA-induced DUOX2 upregulation in PDAC cells may be mediated in a cell context-specific fashion, beyond the canonical components of the cGAS-STING pathway. Upregulation of DUOX2 by exogenous DNA is not observed in colon cancer cell lines. In summary, these data suggest that extracellular DNA could contribute to a DUOX2-induced, H2O2-mediated pro-inflammatory milieu that specifically contributes to the pathogenesis of PDAC, in part through activation of the cGAS-STING pathway.

Citation Format: Stephen L. Wang, Yongzhong Wu, Smitha Antony, Agnes Juhasz, Jennifer Meitzler, Goujian Jiang, Iris Dahan, Jiamo Lu, Krishnendu Roy, James Doroshow. Exogenous DNA upregulates DUOX2 expression in pancreatic cancer via activation of the cGAS-STING signaling pathway [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3348.
Abstract 3379: Cancer dependency signatures for line of sight genomic alterations in squamous cell lung cancer yield rational targets
Authors: Ahdesmäki Miika , Armenia Joshua , Bulusu Krishna , Sidders Ben , Dry Jonathan , McDermott Ultan
Published: September 2020

The Cancer Dependency Map Portal enables combined analyses of genomic and phenomic data such as genomic alterations and CRISPR gene dependencies but focuses on single genes or cell lines. This is well suited for verifying previously generated hypotheses such as the dependency of cells on PRMT5 in the setting of loss of function mutations in MTAP. To identify additional actionable de-novo vulnerabilities in-silico, we refined the genomic status of 1000 cell lines by 1. focusing on variants (driver mutations and copy number alterations) that have higher prevalence in a given cancer type in TCGA and 2. reclassifying variants as actionable only if they cause loss of function in known tumour suppressor genes or gain of function in oncogenes. This gives a clear line of sight, a more manageable set of variants and is analogous to Mendelian randomization. This approach was applied to lung squamous cell carcinoma to form a binary event matrix for the shortlisted genes and copy number alterations across a panel of lung squamous cell lines. For each of the variants, we used machine learning approaches to define a dependency signature from CRISPR dependencies for the matching cell lines that distinguish altered vs non-altered cell lines. The top ranking dependencies were further examined for biological relevance and potential toxicity, yielding novel targets to follow up on. The approach generalises well and can be applied to any other cancers with unmet needs.

All data in this study was derived from public databases.

By combining TCGA cancer specific prevalent genomic alterations in tumor suppressor and oncogenes we drew clear line of sight and associated these alterations to CRISPR based vulnerabilities. This yielded a manageable set of alterations, and for each of these we identified a set of essential genes/pathways to use in machine learning. These signatures were further refined by biological enrichment, targetability and toxicity in order to build a process for rational novel target ID generation.

Citation Format: Miika Ahdesmäki, Joshua Armenia, Krishna Bulusu, Ben Sidders, Jonathan Dry, Ultan McDermott. Cancer dependency signatures for line of sight genomic alterations in squamous cell lung cancer yield rational targets [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3379.
Abstract 3380: Predicting the effects of small molecules on transcriptome of cancer cell lines using deep learning
Authors: Jeon Junhyeok , Lee Sang Mi , Lee GaRyoung , Kim Hyun Uk
Published: September 2020

Developing a computational model that can predict the effects of small molecules on biological status of a cell can be very useful for various applications, including candidate drug compound screening and toxicity testing in silico. In this study, we developed a deep learning model that predicts the effects of small molecules on gene expression levels of a cell. This deep learning model takes two inputs: the structure of a chemical, presented in SMILES, and a protein sequence encoded by a gene of interest. The model processes these two input data using a fingerprinting method for the input chemical structure and a convolutional neural network for the input protein sequence. As a result, the model predicts whether a given molecule upregulates or downregulates a gene of interest where the molecule and gene are both provided as inputs. The deep learning model was developed by using so called L1000 profiles covering gene expression levels of various cancer cell lines under a large number of perturbation conditions [1]. L1000 is a cost-effective transcriptome technology that can accurately infer the expression levels of 9,196 genes on the basis of expression levels of 978 ‘landmark' genes that are directly measured. In this study, the deep learning model was developed for three different cancer cell lines, HA1E (kidney), HCC515 (lung), and HEPG2 (liver), by using L1000 profiles covering expression levels of the landmark genes individually perturbed with 5,213 molecules for the HA1E cell line, 5,171 molecules for the HCC515, and 3,529 for HEPG2. The deep learning models developed in this study will be useful for a wide range of studies that examine the effects of small molecules, for example in drug development and toxicity testing. [1] Subramanian et al. A next generation connectivity map: L1000 platform and the first 1,000,000 profiles. Cell 171, 1437-1452 (2017)

Citation Format: Junhyeok Jeon, Sang Mi Lee, GaRyoung Lee, Hyun Uk Kim. Predicting the effects of small molecules on transcriptome of cancer cell lines using deep learning [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3380.
Abstract 3394: Brisk walking and hepatocellular carcinoma risk in US men and women
Authors: Luo Xiao , Yang Wanshui , Ma Yanan , Simon Tracey , Meyerhardt Jeffrey , Meyerhardt Jeffrey , Meyerhardt Jeffrey , Chan Andrew , Giovannucci Edward , Zhang Xuehong
Published: September 2020

Emerging evidence indicates a potential beneficial effect of vigorous physical activity on hepatocellular carcinoma (HCC) by improving energy balance, immune response, and insulin sensitivity. However, the association between moderate physical activity, such as brisk walking, and the risk of HCC remains largely unknown. Two prospective cohorts of 77,535 women from the Nurses' Health Study and 44,779 men from the Health Professionals Follow-up Study were included. Weekly time spent on brisk walking and other physical activity were updated biennially. The Cox proportional hazard regression model was used to calculate multivariable hazard ratios (HRs) and 95% confidence intervals (95%CIs). After an average 23.1-year follow-up, a total of 138 incident HCC cases were identified. In the multivariable-adjusted model, engaging in brisk walking was significantly associated with a lower risk of HCC (over 1 hour/week vs. non-brisk walking; HR: 0.44, 95%CI: 0.27-0.70; Ptrend<0.001). Further adjustment for body mass index (BMI) and type 2 diabetes (T2D) yielded similar results (HR: 0.51; 95%CI: 0.31-0.83; Ptrend=0.006). The association between brisk walking and HCC risk was generally consistent across all subgroups, including age, BMI, T2D, smoking status, aspirin use, and alcohol consumption (all P>0.05 for interaction). In conclusion, brisk walking was associated with a significantly lower risk of developing HCC among US men and women. If confirmed, brisk walking might serve as a feasible way for some individuals for HCC primary prevention.

Citation Format: Xiao Luo, Wanshui Yang, Yanan Ma, Tracey Simon, Jeffrey Meyerhardt, Jeffrey Meyerhardt, Jeffrey Meyerhardt, Andrew Chan, Edward Giovannucci, Xuehong Zhang. Brisk walking and hepatocellular carcinoma risk in US men and women [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3394.
Abstract 3417: A phase II basket trial of dual anti-CTLA-4 and anti-PD-1 blockade in rare tumors (DART) SWOG S1609: The small bowel tumor cohort
Authors: Chae Young Kwang , Othus Megan , Patel Sandip Pravin , Zalupski Mark , Kasi Anup , Khalil Maged , Kalyan Aparna , Polite Blase , Fenton Sarah , Gurung Sewan , McLeod Christine M. , Giles Francis , Chen Helen X. , Sharon Elad , Mayerson Edward , Plets Melissa , Ryan Christopher W. , Blanke Charles D. , Kurzrock Razelle
Published: September 2020

Background: The potential therapeutic benefits of checkpoint inhibitors and their application to many different tumor types has been a key factor in the advancement of medical oncology since their first approval for metastatic melanoma in 2011. However, their efficacy in rare tumor types remains to be seen. This paper presents the results of combination therapy with both anti-CTLA and anti-PD-1 in the small bowel cohort of SWOG S1609 Dual Anti-CTLA-4 & Anti-PD-1 blockade in Rare Tumors (DART).

Methods: This study is designed as a prospective, open-label, multicenter phase 2 clinical trial of ipilimumab (1mg/kg intravenously every 6 weeks) plus nivolumab (240mg intravenously every 2 weeks) in rare tumors. Here we report the outcomes from patients diagnosed with small bowel cancer (SBC). The primary endpoints included overall response rate (ORR) (RECIST v1.1) (complete response (CR) and partial responses (PR)); progression-free survival (PFS), overall survival (OS), stable disease >6 months, and toxicity were the secondary endpoints.

Results: Twenty five patients were registered to the cohort and twenty three received therapy. The duodenum was the primary site of origin in 52% (N=11), 14% (N=3) arose in the ileum and 14% (N=3) arose in the jejunum. The primary site of origin was unknown in 19% (N=4). The overall response rate was 8% (CR, 4%, N= 1; PR, 4%, N= 1). The median PFS was 2 months; 6-month OS was 48% and median OS 6 months. The most common toxicities were diarrhea and fatigue (both 17%) followed by dyspnea (13%) with diarrhea, increased bilirubin, colitis and elevated lipase (all 4.3%) as the most common grade 3-5 immune-related adverse events.

Conclusions: Combination therapy with ipilimumab plus nivolumab in small bowel tumors resulted in an overall response rate of 8% with one partial and one complete response in twenty three treated patients.

Citation Format: Young Kwang Chae, Megan Othus, Sandip Pravin Patel, Mark Zalupski, Anup Kasi, Maged Khalil, Aparna Kalyan, Blase Polite, Sarah Fenton, Sewan Gurung, Christine M. McLeod, Francis Giles, Helen X. Chen, Elad Sharon, Edward Mayerson, Melissa Plets, Christopher W. Ryan, Charles D. Blanke, Razelle Kurzrock. A phase II basket trial of dual anti-CTLA-4 and anti-PD-1 blockade in rare tumors (DART) SWOG S1609: The small bowel tumor cohort [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3417.
Abstract 3429: Targeting gain-of-function of mutant p53 C-terminal and oligomerization domains to disrupt activated DNA replication in breast cancer
Authors: Lundine Devon , Annor George , Ellison Viola , Xiao Gu , Prives Carol , Bargonetti Jill
Published: September 2020

We recently documented that gain-of-function (GOF) mutant p53 (mtp53) R273H in triple negative breast cancer (TNBC) cells interacts with replicating DNA and PARP1 (Xiao et. al., Cancer Research 2019). The missense R273H GOF mtp53, without a functional central DNA binding domain, has the capacity to interact tightly with chromatin but it is unclear what regions of mtp53 are responsible for the chromatin-based DNA replication activities. GOF mtp53 proteins possess an intact C-terminal domain (CTD) and oligomerization domain (OD). In wild-type p53 the CTD and OD domain regulate non-specific DNA binding, post-translational modification, and protein tetramerization. We are targeting the CTD and OD regions of mtp53 to test if their disruption limits oncogenic GOF activity. We generated CRISPR/Cas9 edited R273H CTD and OD in the TNBC cell line MDA-MB-468, and also exogenously expressed such dual mutant proteins in p53 null cancers, to evaluate the roles played by oligomerization and non-specific DNA binding in mtp53 interactions with replicating DNA and PARP1. We determined the oligomerization state of R273H mtp53 using the glutaraldehyde crosslinking assay and observed that GOF mtp53 R273H mtp53 in breast cancer cells exists as a tetramer. Experiments are in progress to evaluate cell cycle progression, replication initiation, and mtp53 chromatin interactions in cells expressing missense mtp53 proteins with co-associated mutated CTDs or ODs. Our preliminary data suggests that disrupting the mtp53 CTD inhibits its interactions with chromatin and progression through the S-phase. Unpublished data on the roles played by the mtp53 CTD and OD in DNA replication will be presented at the 2020 AACR meeting. Acknowledgements: This work was funded by grants from the Breast Cancer Research Foundation BCRF-18-011 and BCRF-19-011 to Jill Bargonetti. Devon Lundine and Gu Xiao also received project support from TUFCCC/HC Regional Comprehensive Cancer Health Disparity Partnership, Award Number U54 CA221704(5) from the National Cancer Institute. Special thanks to Ella Freulich for help with CRISPR/Cas9 work and James J. Manfredi for mutant p53 plasmids to exogenously express protein.

Citation Format: Devon Lundine, George Annor, Viola Ellison, Gu Xiao, Carol Prives, Jill Bargonetti. Targeting gain-of-function of mutant p53 C-terminal and oligomerization domains to disrupt activated DNA replication in breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3429.
Abstract 3444: Intratumor regulatory T cells prevent IFN-gamma-dependent tumor vessel regression and can be selectively targeted by anti-CD25 near-infrared photoimmunotherapy
Authors: Kurebayashi Yutaka , Choyke Peter L. , Kobayashi Hisataka , Sato Noriko
Published: September 2020

Regulatory T cells (Tregs) play pivotal roles in inhibiting anti-tumor immunity at different phases and locations (priming phase in lymph node and effector phase in tumor tissue) through their ability to suppress the function of T, NK, and antigen-presenting cells. However, our knowledge about the roles of Tregs in tumor immunity has largely derived from analyses after systemic depletion of Tregs or systemic manipulation of Treg functions in murine tumor models. The specific roles that intra- and extra-tumor Tregs play remain unclear due to technical difficulties to deplete or manipulate Tregs in a spatially specific manner. In order to achieve spatially specific depletion of Tregs in a clinically applicable manner, we have previously developed an anti-CD25 F(ab')2 near-infrared photoimmunotherapy (NIR-PIT), in which an intravenous injection of IR700-conjugated anti-CD25 F(ab')2 followed by irradiation of near-infrared light on tumors achieves selective depletion of intratumor Tregs and suppresses tumor growth. The suppression of tumor growth by the anti-CD25 F(ab')2 NIR-PIT depends on CD8 T and NK cells and their production of IFN-γ; however, the exact target of IFN-γ and the mechanisms of tumor regression remain to be elucidated. In this study, we aimed to dissect the mechanisms of the anti-tumor effect of intratumor Treg depletion by anti-CD25 F(ab')2 NIR-PIT using a series of Ifngr1 knockout mice models including bone-marrow chimera and conditional knockout models. Unexpectedly, we found that the target of IFN-γ in anti-CD25 F(ab')2 NIR-PIT was endothelial cells of tumor vessels. In both MC38 and EO771 tumor models (murine colon and breast cancer, respectively), CD8 T cells, NK cells, and Tregs were mainly located along the tumor vessels, and depletion of intratumor Tregs by anti-CD25 F(ab')2 NIR-PIT induced IFN-γ expression from these perivascular CD8 T and NK cells for up to 6 hours. This transient but synchronized induction of IFN-γ/STAT1 signaling along tumor vessels caused rapid vessel regression, intratumor ischemia, and apoptotic death of tumor cells. Combination with IL-15 treatment further eradicated residual tumor cells to achieve complete remission of MC38 and EO771 tumors.

Citation Format: Yutaka Kurebayashi, Peter L. Choyke, Hisataka Kobayashi, Noriko Sato. Intratumor regulatory T cells prevent IFN-gamma-dependent tumor vessel regression and can be selectively targeted by anti-CD25 near-infrared photoimmunotherapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3444.
Abstract 3516: Commercial gene expression tests for prostate cancer outcome risk in men of West African ancestry
Authors: Gaston Sandra M. , Kittles Rick A. , Stoyanova Radka , Giret Teresa M. , Ansari Saba A. , Punnen Sanoj , Pollack Alan
Published: September 2020

Both incidence and mortality data show that the burden of prostate cancer (PrCa) is greater for African Americans (AA) than for European Americans (EAs), with AAs about 1.7 times more likely to be diagnosed with PrCa than EAs and about 2.4 times more likely to die of this disease. Molecular testing of prostate cancer tissue is increasingly being incorporated into patient management. Current National Comprehensive Cancer Network (NCCN) guidelines recommend three commercially available gene expression based tests for PrCa prognosis: Decipher (GenomeDX), Oncotype GPS (Genomic Health) and Prolaris (Myriad). All three of these tests were developed in predominantly EA patient populations and subsequent studies in AA patients have not included ancestry genotyping to assess genetic admixture. In a pilot study we used the Genomics Resource Information Database (GRID) to compare the research versions of these three commercial gene expression tests for PrCa outcome risk in 157 cancer positive biopsy cores from 55 patients. We also performed ancestry genotyping to estimate each patients European, West African and Native American Ancestry. The overall distribution of scores predicting average, higher than average and lower than average outcome risk differed for the three tests, with higher risk scores in 17%, 7% and 27% and lower risk scores in 69%, 73% and 43% of the cores for the Decipher, GPS and Prolaris tests, respectively. Interestingly, 25%, 37%, and 16% of 56 Cores with high risk pathology (grade group 3-5) were classified as low risk by the Decipher, GPS and Prolaris tests, respectively. In the 21 cores from men with > 70% WA ancestry, 100%, 80% and 30% of cores with intermediate and high risk pathology (grade group 2-5) were paradoxically classified as low risk by the research Decipher, GPS and Prolaris tests, respectively. These preliminary results are consistent with the evidence recently published by Creed et al (2019) suggesting that the currently available commercial genomic tests may underestimate prostate cancer outcome risk in AAs. Ancestry genotyping may be useful in rescaling genomic prostate cancer outcome risk tests that were developed in predominantly EA cohorts for use in AA patients. Reference: Creed JH et al. Cancer Epidemiol Biomarkers Prev, published online 2019.

Citation Format: Sandra M. Gaston, Rick A. Kittles, Radka Stoyanova, Teresa M. Giret, Saba A. Ansari, Sanoj Punnen, Alan Pollack. Commercial gene expression tests for prostate cancer outcome risk in men of West African ancestry [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3516.
Abstract 3529: Detection of genomic structural variations associated with drug sensitivities and resistance in AML using novel whole genome imaging
Authors: Finlay Darren , Murad Rabi , Hong Karl , Lee Joyce , Pang Andy , Lai Chi-Yu , Burian Carol , Mason James , Hastie Alex , Yin Jun , Vuori Kristiina
Published: September 2020

Acute myeloid leukemia (AML) is a particularly devastating collection of hematological cancers and whilst somewhat rare, the patient survival rate is abysmal without bone marrow transplantation. Traditional chemotherapies administered to AML patients cause significant and devastating side effects. Additionally, more than 30% of patients fail to respond to these initial treatments and most patients that do respond will eventually relapse within 5 years. As such, understanding the evolution of AML to identify novel targets, and therefore drug treatment regimens, is a significant medical need.

Genomic rearrangements and other Structural Variations (SVs) have long been known to be causative and pathogenic in multiple cancers, including leukemias. Indeed the discovery of the “Philadelphia chromosome” (eventually identified as a BCR-ABL translocation) as causative in Chronic Myeloid Leukemia has prompted much research into SVs in cancers, including the development of targeted therapeutics against oncogenic proteins resulting from genomic rearrangements. These SVs may be involved in cancer initiation, progression, clonal evolution, and drug resistance, and a better understanding of SVs from individual AML patients may help guide therapeutic options.

Here we show utilization of an innovative whole genome imaging technology to detect known, and novel, SVs in AML patients' samples. Importantly, this new technology provides an unprecedented level of granularity and quantitation unavailable to other current techniques and it allows an unbiased detection of novel SVs, which may be relevant for disease pathogenesis and/ or drug resistance. Coupled with standard gene expression analyses we have also assessed the chemosensitivities of these samples to 120 FDA approved oncology drugs and 335 epigenetic modulating agents. Here we show how integrative analysis of these diverse datasets is used to associate the detected genomic rearrangements with drug sensitivity profiles, potentially identifying novel therapeutic targets for individual AML patients.

Citation Format: Darren Finlay, Rabi Murad, Karl Hong, Joyce Lee, Andy Pang, Chi-Yu Lai, Carol Burian, James Mason, Alex Hastie, Jun Yin, Kristiina Vuori. Detection of genomic structural variations associated with drug sensitivities and resistance in AML using novel whole genome imaging [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3529.