bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1851: Mechanisms underlying focused ultrasound-induced immunogenicity in melanoma
Authors: Pelka Sandra , Bandyopadhyay Sanmay , Pandey Sanjay , Guha Chandan , Macian Fernando
Published: June 2023

Advanced melanoma is an aggressive skin cancer characterized by low survival rates. The generation of a robust anti-tumor response relies on dendritic cells (DCs) to process tumor antigens, migrate to draining lymph nodes, and activate T-cells. Within the tumor microenvironment, immunosuppressive factors impede DC differentiation and activation, and the subsequent immature DC phenotype is a major mechanism underlying the ineffective T-cell response in melanoma patients. We have shown that low-intensity focused ultrasound (LOFU), a novel therapy that delivers mechanical and thermal stresses locally at the tumor site, may enhance DC function in a B16 murine melanoma model. We hypothesize that LOFU enhances the dendritic cell-mediated anti-tumor response by inducing changes in tumor cell and dendritic cell autophagy and increasing pro-immunogenic factors in the TME.

Autophagy is a lysosome-dependent process of cellular content degradation that has previously been implicated in the regulation of multivesicular body formation/degradation, and thus in the secretion of extracellular vesicles. Autophagy may subsequently modify the cargo of tumor-derived extracellular vesicles, such as exosomes, in a pro-immunogenic manner. We have found that LOFU treatment is associated with an increase in autophagic flux in melanoma cells and have identified at least 82 proteins within melanoma-derived extracellular vesicles that are significantly enriched or reduced with LOFU treatment, with enriched proteins partaking in essential processes including antigen processing, intracellular trafficking, and cellular metabolism. DCs pulsed with vesicles from LOFU-treated melanoma cells have shown enhanced activation of both CD4+ and CD8+ T-cell activation, suggesting that the LOFU-induced increase in tumor cell autophagy and modification of vesicular cargo may promote a DC-activated anti-tumor antigen activation response. Characterization of other mechanisms by which LOFU may enhance DC function, such as direct changes in DC autophagy, is critical to elucidate DC-mediated pathways involved in the anti-tumor response. Further understanding of DC involvement in the anti-tumor response will provide novel targets for the development of immunotherapies to improve survival and quality of life in advanced melanoma patients.

Citation Format: Sandra Pelka, Sanmay Bandyopadhyay, Sanjay Pandey, Chandan Guha, Fernando Macian. Mechanisms underlying focused ultrasound-induced immunogenicity in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1851.
Abstract 2013: Oxidized anthracycline payloads induce anti-tumor immunogenic cell-death and show linker-dependent tolerability when delivered as ADCs
Authors: Hamilton Joseph Z. , Klussman Kerry , Mahzereh Rebecca , Simmons Jessica , Ulrich Michelle , Gardai Shyra J. , Senter Peter D. , Burke Patrick J.
Published: June 2023

Novel payloads with orthogonal mechanisms of action (MOAs) to established ADC technologies are important for expanding the utility of antibody-drug conjugates (ADCs) in cancer treatment. Anthracyclines, a class of cytotoxic small molecules that include important clinical chemotherapeutics such as doxorubicin, have long been of interest to the ADC field due to their high potency and unique MOA. Interest in anthracyclines has increased in recent years due to their status as canonical inducers of immunogenic-cell death (ICD), a feature that may lead to more durable tumor regressions and increased synergy with immune checkpoint blockade. In this work, we investigate several drug-linkers based on oxidized derivatives of the highly potent anthracycline analogue PNU-159682. Resultant ADCs are potent and immunologically specific with linker-dependent bystander activity in co-culture cytotoxicity assays displaying heterogeneous antigen (Ag+/Ag-) expression. Rodent in vivo anti-tumor activity and tolerability are shown with significant differences emerging between the toxicity profile of the cleavable and non-cleavable analogues. In vitro, tumor cells treated with anthracycline ADCs showed key hallmarks of ICD including markers of endoplasmic reticulum (ER) stress and the induction of ICD damage-associated molecular patterns (DAMPs) such as HMGB1 and ATP. In vivo mouse xenograft studies show increased tumoral F4/80+ macrophage infiltration by IHC in anthracycline ADC treated animals relative to controls. ADCs delivering monomethyl auristatin E (MMAE) show comparable activity in ICD assays, confirming MMAE as an ICD inducing ADC payload alongside canonical inducers such as anthracyclines. Our results indicate there is a therapeutic window in rodents alongside ICD induction by anthracycline ADCs, positioning anthracyclines as a potentially impactful class of ADC payloads.

Citation Format: Joseph Z. Hamilton, Kerry Klussman, Rebecca Mahzereh, Jessica Simmons, Michelle Ulrich, Shyra J. Gardai, Peter D. Senter, Patrick J. Burke. Oxidized anthracycline payloads induce anti-tumor immunogenic cell-death and show linker-dependent tolerability when delivered as ADCs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2013.
Abstract 2024: High resolution limited proteolysis (HR-LiP), a novel structural proteomics approach for the prediction of small molecule-protein binding events
Authors: Beaton Nigel , Bruderer Roland , Feng Yuehan , Adhikari Jagat , Cornella-Taracido Ivan , Reiter Lukas
Published: June 2023

High resolution profiling of drug-protein interactions and binding mechanisms remains a major hurdle during lead selection and optimization. A key milestone in structure-based drug design is compound binding site identification and characterization. Structure-activity relationship (SAR) studies traditionally utilize time and cost intensive techniques such as nuclear magnetic resonance (NMR), x-ray crystallography (X-ray) and cryo-electron microscopy (cryo-EM). To date, hydrogen-deuterium exchange (HDX) is the only mass spectrometry-based approach that has been extensively utilized. Further, SAR studies are often complicated by protein size (large proteins and/or oligomers) and location (membrane proteins), which can lead to protocol adaptations that can introduce artifacts. High resolution limited proteolysis (HR-LiP) is a high-throughput, high-resolution approach based on LiP-Quant that utilizes peptide-level resolution to characterize drug-protein interactions. Two well-characterized drug target proteins, bromodomain-containing protein 4 (BRD4) and transitional endoplasmic reticulum ATPase (VCP), were selected for analysis as they represent both difficult targets for structural biology (size and oligomerization respectively). BRD4 was targeted with a well characterized inhibitor (JQ1), while VCP was investigated for binding with a known autophagy activator. Using HR-LiP we identify the binding site of the BRD4 inhibitor JQ1 in the full-length protein, which is typically too large to be used directly in with conventional methods. Further, we confirm binding of the autophagy activator to VCP at a site known to increase VCP activity. For both compounds tested, our data are in good accordance with orthogonal data obtained by other structural protein approaches including HDX-MS and “click” chemistry-based chemoproteomics. We demonstrate that HR-LiP can be used to dissect small molecule-protein binding events, including compound binding site prediction for protein targets classically considered to be difficult and time intensive. Further, we show that this data can be used to shed light on the biological mechanisms driving a phenotypic response. Given its biological power, broad applicability and ease of implementation, we envision the use of HR-LiP as a routine approach for target validation and lead optimization in small molecule drug discovery pipelines.

Citation Format: Nigel Beaton, Roland Bruderer, Yuehan Feng, Jagat Adhikari, Ivan Cornella-Taracido, Lukas Reiter. High resolution limited proteolysis (HR-LiP), a novel structural proteomics approach for the prediction of small molecule-protein binding events [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2024.
Abstract 2092: Consensus analysis of associated target-biomarker pairs in cancers
Authors: Jeong Sumin , Park Yuna , Jeong Eunah , Yoon Sukjoon
Published: June 2023

The heterogeneity of cancers limits the reproducibility of target-biomarker relationships in diverse cohorts or sample groups. Here, we carried out consensus analyses of associated data pairs to improve the consistency (reproducibility) of prognostic power of biomarkers and anticancer efficacy of targets. All of RNA expression, patient survival, CRIRSPR/shRNA screening data were integrated from TCGA, GDSC and CCLE datasets. While >90% of prognostic RNA expression found in given sample sets were not reproduced (i.e., not significant) in other test sets, those hits exhibiting consensus prognostic powers in diverse subtypes or lineages, were more significant in test sets than non-consensus hits. In the analysis of the association between CRISPR knockout and RNA expression data, the lineage consensus of the association increased the reproducibility in the validation using sh/siRNA knockdown and qPCR data. Further experimental validations reveal that ITGAV is an effective anticancer target in diverse lineages, significantly associated with prognostic RNA expression of several consensus biomarker genes. In conclusion, the consensus analysis was useful to prioritize reproducible targets and biomarkers from omics data. It has been implemented in Q-omics software (http://qomics.io) for general cancer research.

Citation Format: Sumin Jeong, Yuna Park, Eunah Jeong, Sukjoon Yoon. Consensus analysis of associated target-biomarker pairs in cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2092.
Abstract 2106: Importance of alveolar epithelial cell type 2 populations in non-small cell lung cancer progression from localized to metastatic disease
Authors: Suvilesh Kanve Nagaraj
Published: June 2023

Introduction: Non-small cell lung cancer (NSCLC) is by far the leading cancer killer. Identifying personalized treatment strategies to prevent and treat metastases remains an unmet clinical need. Alveolar epithelial type II (AT2) cells are self-renewing epithelial stem cells that have been identified to initiate a majority NSCLCs. Previously, we have reported the presence of multiple AT2 cell types in micrometastatic circulating tumor cell-derived xenograft (CDX) models and patient metastatic tumors. These findings provide a rationale to further study whether and how AT2 cells contribute to metastatic progression.

Hypothesis: We hypothesize that AT2 cell types have a functional role in metastasis tumorigenicity of NSCLC.

Methods: A library of NSCLC patient-derived circulating tumor cell (CTC) lines, xenograft (PDX) models, and publicly available data sets of patients’ NSCLC tumor tissues were subjected to single cell/bulk RNA sequencing and analysis to determine the frequencies and expression patterns of AT2 subclones. Using magnetic bead sorting AT2 cells were enriched and AT2-associated tumorigenicity, migratory and invasion capacity determined using 3D culture and PDX models. The effects of AT2 depletion on PDX tumor characteristics were evaluated histologically. Frequencies of AT2 phenotypes in primary and metastatic tumors were validated in human tissue microarrays and patient CTCs.

Results: An additional AT2 population was identified in NSCLC brain metastases and primary lung tumors that had metastasized already. However, this additional AT2 cell population was absent in primary lung tumors of patients that had not metastasized and in patients with lymph node, pleural and liver metastases. AT2-depleted NSCLC cell lines had reduced tumorigenicity in PDX models in comparison to tumors derived from whole cell populations. In addition, tumors derived from AT2 depleted cell lines were highly necrotic. AT2-depleted cell lines showed decreased migratory and invasion capacity in 3D models. Further, AT2 phenotypes were identified at high rates in NSCLC patients’ metastases and CTCs.

Conclusion: Findings indicate an important role of AT2 cells in metastatic progression, beyond their known role in NSCLC initiation. The role of AT2 in metastasis migration, invasion and growth offers unique opportunities to identify novel therapeutic targets. Expansion of translational findings on AT2 cells has high potential for clinical impact to prevent or treat metastatic disease in NSCLC patients.

Citation Format: Kanve Nagaraj Suvilesh. Importance of alveolar epithelial cell type 2 populations in non-small cell lung cancer progression from localized to metastatic disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2106.
Abstract 2113: The age-dependent intratissue accumulation of cholesterol, resulting in reduced squalene epoxidase, is associated with a high colorectal cancer risk population
Authors: Jun Soo Young , Yoon Ji-Yong , Lee Jeong-Ju , Kim Nam-Soon
Published: June 2023

Objective: we recently reported that cholesterol accelerates CRC aggravates via the dissociation of GSK3β/p53 by the degradation of squalene epoxidase (SQLE). However, despite its clinical reports, the underlying mechanisms between age, cholesterol, and CRC aggravation have never been evaluated in human cohorts.

Design: tissue lysates and human specimens collected prospectively from CRC patients and controls were grouped according to CRC grade and divided into two groups based on age 50, the average risk for CRC. The intra-tissue content of total cholesterol and cholesteryl ester were measured using CRC lysates. We then used fluorescence-based multiplex immunohistochemistry with digital image analysis and machine learning to measure the quantitative expression of SQLE, GSK3β, p53wt, and p53mt.

Results: through an epidemiological study, we estimated age-dependent increases in CRC incidence and the percentage of a group with high cholesterol around age 50. Considering this age-dependency, we demonstrated age and CRC-grade-dependent manner cholesterol accumulated inside CRC tissues, SQLE, p53wt, and p53mt degraded, and GSK3β inactivated. SQLE and GSK3β were clinically associated with increased risk and showed good diagnostic and prognostic performance in CRC patients older than 50. Furthermore, the combined value for four variables exhibited superior diagnostic performance in discriminating from CRC patients after age 50 to controls.

Conclusion: this study provides new biological perceptions explaining the effects of age in the progression of CRC aggravation and potential biomarkers associated with a high CRC-risk population.

Citation Format: Soo Young Jun, Ji-Yong Yoon, Jeong-Ju Lee, Nam-Soon Kim. The age-dependent intratissue accumulation of cholesterol, resulting in reduced squalene epoxidase, is associated with a high colorectal cancer risk population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2113.
Abstract 2302: Detection of tumor-associated methylation and mutation signatures for early colorectal cancer diagnosis
Authors: Bains Pushpinder , Kulak Mikhail , Cui Zhen , Babu Alisha , Tran Stella , Lee Bongyong , Zhang Aiguo , Sha Michael
Published: June 2023

Introduction: The ability to identify epigenetic signatures, together with genetic alterations, has expanded the translational utility of genomic sequencing in clinical settings. Epigenetic changes may appear at very early stages before diagnosis. Thus, they could be used as epigenetic biomarkers for early detection or risk of cancer development. Circulating cell-free DNA (cfDNA) profiling has emerged as a promising biomarker for cancer diagnosis and prognosis. Herein, we developed targeted next-generation sequencing (NGS) based assays for assessing multiple DNA methylated and mutated regions in both low-quantity (plasma cfDNA) and low-quality formalin-fixed paraffin-embedded (FFPE) tissue for colorectal cancer (CRC) detection.

Methods: For the OptiSeqTM CRC methylation panel, we assessed the methylation status of CpG dinucleotides in 42 genes. With this panel, we profiled 6 CRC samples (matching cfDNA and FFPE), 2 primary colon tumors with adjacent normal, and 6 advanced adenomas (AA) FFPE tissue DNA. For healthy individuals, we profiled 12 normal colon FFPE samples and 31 plasma cfDNA (26 matching buffy coat DNA samples). For the OptiSeqTM CRC mutation panel, the mutation status was assessed in 43 genes. Here, we profiled 6 CRC samples (matching cfDNA and FFPE), 19 CRC plasma cfDNA, 4 AA FFPE and 42 healthy individuals’ plasma cfDNA (26 matching buffy coat DNA samples). The analytical and clinical performance for both panels (separately and combined) were calculated using advanced adenoma (AA), CRC, and cancer-free individuals.

Results: For CRC cfDNA samples, clinical samples testing showed that the methylation panel had a sensitivity of 50% (95% CI: 13.9-86.0) and specificity of 100% (95% CI: 86.2-100) while the mutation panel yielded the sensitivity 88% (95% CI: 67.7-96.8) and specificity 100% (95% CI: 89.5-100). Combining the mutation panel with the methylation panel, the sensitivity increased to 100% (95% CI: 51.6-100) and specificity to 100% (95% CI: 65.1-93.3). For AA FFPE samples, the methylation panel analysis yielded a sensitivity of 100% (95% CI: 39.5-100.0) and a specificity of 100% (95% CI: 67.8-100).

Conclusion: We have developed and validated the targeted NGS-based panels to detect clinically relevant mutation and methylation patterns and demonstrate its potential as a highly sensitive assay for early CRC diagnosis.

Citation Format: Pushpinder Bains, Mikhail Kulak, Zhen Cui, Alisha Babu, Stella Tran, Bongyong Lee, Aiguo Zhang, Michael Sha. Detection of tumor-associated methylation and mutation signatures for early colorectal cancer diagnosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2302.
Abstract 2653: ABN501, a novel anti-claudin-3 antibody, shows potent anti-cancer activity in vitro and in vivo
Authors: Lee Saehyung , Choi Jae , Hong Sunghyun , Sim Euni , Kim Jaemun , Lee Sangin , Hong Sungyoul , Yang Hobin , Jo Jiwon , Kim Na Young , Shin Young Kee , Choi Jun Young
Published: June 2023

Claudin-3, one of the tight junction proteins, could be a potential therapeutic target and biomarker. Since it is not only overexpressed in various types of malignant solid cancers but also exposed externally during tumorigenesis. Previously, we developed a fully human monoclonal antibody (ABN501) with sub-nanomolar affinity against the native claudin-3 protein. In this study, we confirmed that ABN501 is highly specific for the extracellular loop (ECL) 2 domain of claudin-3 using stable cell lines expressing two chimeric claudins fused to claudin-3 and claudin-4. Followed by, we evaluated in vitro ADCC activity of ABN501 using NK cells expressing CD16a (NK-92MI-CD16a), in various cancer cell lines, expressing claudin-3. ABN501 exhibited potent in vitro ADCC activity in diverse cancer cell lines. We also confirmed in vivo anti-tumor activity of ABN501 in xenograft mice bearing claudin-3-expressing tumors. In summary, we suggest that ABN501 can be used as promising tissue-agnostic drug candidates as therapeutics for various types of claudin-3-positive cancers. ABN501 is currently developed with various antibody engineering platforms.

Citation Format: Saehyung Lee, Jae Choi, Sunghyun Hong, Euni Sim, Jaemun Kim, Sangin Lee, Sungyoul Hong, Hobin Yang, Jiwon Jo, Na Young Kim, Young Kee Shin, Jun Young Choi. ABN501, a novel anti-claudin-3 antibody, shows potent anti-cancer activity in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2653.
Abstract 2869: The role of regulatory T cells in NASH-mediated HCC development
Authors: Yang Ming , Kimchi Eric T. , Staveley-O’Carroll Kevin F. , Li Guangfu
Published: June 2023

Objective Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease, ranging from simple steatosis to an advanced form of non-alcoholic steatohepatitis (NASH) without effective treatment. NASH can progress to liver cirrhosis and hepatocellular carcinoma (HCC) in association with increased production of liver-resident regulatory T cells (Treg). This study aims to identify the role of Tregs in NASH-driven HCC development.

Methods Wild-type C57BL/6 mice were fed with a choline-low high-fat high-sugar diet (CL-HFS), which recapitulates the Western-pattern diet, to induce NASH, then received transplantation of oncogenic hepatocytes by intrasplenic injection to induce an orthotopic HCC. Wild-type mice fed with a normal diet were used for control. Hepatic immune cell profiling was characterized by flow cytometry to identify the cellular basis mediating CL-HFS-induced NASH. The metagenomic assay was applied to identify NASH-relevant gut microbiota in the liver tissue.

Results CL-HFS caused successful development of NASH and liver fibrosis in wild-type mice. Fast orthotopic tumor growth was detected in NASH-bearing mice compared to wild-type mice. Strongly positive staining of glypican 3 (GPC3), increased frequency of Tregs, and increased expression of Col1α1, Col4α1, and α-SMA were detected in livers and tumors from NASH-bearing mice compared to wild-type mice by IHC and flow cytometry. Metagenomic assay detected the lipid-producing bacterial genus Bacillus in NASH liver, which was demonstrated to increase the frequency of Tregs in healthy spleen cells.

Conclusion CL-HFS-caused NASH promotes HCC growth likely by increasing liver-resident Tregs and liver-infiltrating Bacillus.

Keywords: Non-alcoholic fatty liver disease (NAFLD), Non-alcoholic steatohepatitis (NASH), choline-low high-fat high-sugar diet (CL-HFS), regulatory T cells (Tregs), Bacillus, hepatocellular carcinoma (HCC).

Citation Format: Ming Yang, Eric T. Kimchi, Kevin F. Staveley-O’Carroll, Guangfu Li. The role of regulatory T cells in NASH-mediated HCC development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2869.
Abstract 2914: iPSC-derived CAR-NK cell therapy: nominating clinical candidate clones through integrated multi-functional analysis
Authors: Hall Matthew S. , Manor Ohad , Campion Liam , Bullaughey Chiamin , Morse Barry A. , Gurung Buddha , Borges Luis
Published: June 2023

CNTY-101 is an induced Pluripotent Stem Cell (iPSC) derived Chimeric Antigen Receptor (CAR) NK cell clinical candidate for the treatment of B-cell malignancies. It incorporates six gene edits. A CAR targeting CD19 mediates tumor cell engagement and killing, the homeostatic cytokine IL-15 improves functional persistence, and EGFR acts as a safety switch, providing an antibody target for elimination of the engineered cells, if ever necessary. Further edits are designed to reduce graft rejection due to alloreactivity, including disruption of beta-2-microglobulin (B2M) to prevent major histocompatibility complex class I (MHC-I) surface expression, disruption of class II major histocompatibility complex transactivator (CIITA), and expression of HLA-E fused with B2M. Our data indicate that iPSC lines show significant functional variability dependent on the original somatic cell and donor used for reprogramming. Additional functional variability is induced when the iPSC are genetically edited and differentiated to NK cells. This complexity necessitates careful methodology for nominating clinical candidate clones with desirable functional attributes and presents an opportunity for future refinement. Here, we evaluate 28 clinical candidate clones for CNTY-101, integrating in-process data from genetic engineering, differentiation, and a battery of functional assays into a unified analysis framework. We find that the donor and the number of integrated transgene alleles (ex. monoallelic vs biallelic) have a large impact on clone function. In silico correction for donor and allele number reveals that transgene surface expression correlates with functions including short-term killing, serial-killing, and persistence. We show that IL-15 transgene expression, serial-killing, and persistence correlate along one functional axis, while CAR expression, short-term killing, and cytokine secretion correlate on a second functional axis. In addition, we identify biomarkers measured during the differentiation process that are correlated with functional performance, such as the level of CD94 surface expression and tumor killing. Finally, we discuss the application of functional multi-omics to candidate clone nomination for Century Therapeutics’ pipeline of future products.

Citation Format: Matthew S. Hall, Ohad Manor, Liam Campion, Chiamin Bullaughey, Barry A. Morse, Buddha Gurung, Luis Borges. iPSC-derived CAR-NK cell therapy: nominating clinical candidate clones through integrated multi-functional analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2914.
Abstract 2915: Intratumoral co-injections of NK cells and neutralizing antibodies against the NKG2A inhibitory pathway achieve efficacious anti-tumor responses in a CD8 T cell and cDC1-dependent manner
Authors: Alvarez Maite , Ochoa Maria C. , Molina Carmen , Teijeira Alvaro , Garasa Saray , Sanchez-Gregorio Sandra , Olivera Irene , Glez-Vaz Javier , Cirella Asunta , Gomis Gabriel , Gonzalez-Gomariz Jose , Berraondo Pedro , Melero Ignacio
Published: June 2023

Local intratumoral NK cell activity is known to foster adaptive T cell-mediated immunity against cancer, that conceivably is inhibited by the interactions of the inhibitory receptor NKG2A with the MHC class I molecule HLA-E in humans or Qa-1b in mice. In this study, we found that intratumoral delivery of NK cells significantly attains therapeutic effects only if co-injected with anti-NKG2A and anti-Qa1b blocking monoclonal antibodies (mAb) in two solid mouse tumors models. Such therapeutic activity was contingent on endogenous CD8 T cells and on type-1 conventional dendritic cells (cDC1), which mediate cross-priming of tumor antigens. Moreover, the anti-tumor effects can be enhanced upon combination with anti-PD-1 mAb. This combination achieves at least partial abscopal efficacy against distant untreated tumors. In xenografted mice with HLA-E-expressing human cancer cells, intratumoral co-injection of allogeneic human NK cells and the clinical-grade anti-NKG2A mAb monalizumab synergistically achieved therapeutic effects and gave rise to stronger antibody-dependent cellular cytotoxicity (ADCC) in conjunction with cetuximab. In conclusion, these studies provide evidences for the therapeutic potential of on-site NK cell-based immunotherapies that may recruit endogenous T-cell responses.

Citation Format: Maite Alvarez, Maria C. Ochoa, Carmen Molina, Alvaro Teijeira, Saray Garasa, Sandra Sanchez-Gregorio, Irene Olivera, Javier Glez-Vaz, Asunta Cirella, Gabriel Gomis, Jose Gonzalez-Gomariz, Pedro Berraondo, Ignacio Melero. Intratumoral co-injections of NK cells and neutralizing antibodies against the NKG2A inhibitory pathway achieve efficacious anti-tumor responses in a CD8 T cell and cDC1-dependent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2915.
Abstract 3085: Role of amino acid sequences within the N terminal domain in regulating AF1-mediated glucocorticoid receptor activity
Authors: Khan Shagufta H. , Kumar Raj
Published: June 2023

The steroid hormone receptors (SHRs) play an important role in many physiological and pathological processes including endocrine cancers. However, the precise mechanisms by which these SHRs pass signals from a hormone to control gene expression remain a central unresolved problem, which has limited SHRs therapeutic utilities in a cell/tissue specific manner. Role of the activation function-1 (AF1) in this process is of immense importance as it provides platforms for interactions of specific co-regulatory proteins, and thereby dictates the final outcome SHR-regulated target gene expression. The AF1 located in the N-terminal domain (NTD) plays a critical role in regulating cell/tissue-specific effects of SHRs. However, the exact mechanism of this action is not known. It has been suggested that cell/tissue-specific activities of SHRs may be due to AF1 conformations and subsequent coactivator interactions. The SHRs’ NTD/AF1 has been reported to exist in an intrinsically disorder conformation and undergoes disorder-order transition through intra- and inter- molecular allosteric regulations. In the study, we tested whether the flanking sequences around AF1 affect glucocorticoid receptor (GR) activity due, in part, to conformational changes in otherwise ID AF1 that can facilitate its interaction with critical coregulatory proteins such as TATA Box-Binding Protein (TBP) and steroid receptor coactivator-1 (SRC-1). Using biophysical techniques, we found that placing AF1 immediately upstream from the DNA binding domain (DBD) results into acquisition of a relatively ordered conformation in GR AF1. Further analyses revealed that removing amino acid sequences (1-76) prior to AF1 or removal of amino acids (263-420) between AF1 and DBD differentially regulates AF1-mediated GR activity. The results from this study may provide a potential mechanism for cell-dependent differing GR activities as well as other SHRs and thereby, our results may provide additional SHR selectivity needed to target cell-tissue specific gene regulations in current endocrine-based therapies.

Citation Format: Shagufta H. Khan, Raj Kumar. Role of amino acid sequences within the N terminal domain in regulating AF1-mediated glucocorticoid receptor activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3085.
Abstract 3328: Using Luminex approach to develop a panel of anti-TAA autoantibodies for the detection of prostate cancer
Authors: Qiu Cuipeng , Wang Xiao , Batson Serina , Francia Giulio , Zhang Jian-Ying
Published: June 2023

Autoantibodies to tumor-associated antigens (TAAs) have been demonstrated as reporters in early carcinogenesis. Luminex technology enables the development of simultaneous detection of multiple autoantibodies. This study aimed to use Luminex technology to evaluate and develop an optimal panel for the detection of prostate cancer (PCa) based on autoantibodies to fourteen TAAs (p53, p16, IMP2, IMP3, SOX2, BIRC5, HIF1-α, HSP60, ENO1, CTAG1B, MUC1, Her2, GAL1, and GAL3) associated with cancers. A total of 163 samples including 91 PCa sera and 72 normal control sera were screened to evaluate the expression of fourteen autoantibodies by multiple autoantibody bead panel. Except for autoantibodies to Her2 and IMP3, twelve autoantibodies showed significantly high frequencies ranging from 19.8% to 51.6% in the PCa group at 95.8% specificity for normal controls. The area under the curve (AUC) of these twelve autoantibodies ranged from 0.609 to 0.868. After logistic regression modeling and leave-one-out cross-validation, an optimal panel with the combination of three autoantibodies to p16, IMP2, and HSP60 was developed, achieving an AUC of 0.909 with 71.4% sensitivity and 95.8% specificity. Autoantibody to HSP60 exhibiting the best performance with an AUC of 0.868 in the autoantibody bead panel was further validated by Enzyme-linked immunosorbent assay (ELISA) with a larger size sample containing 200 PCa sera and 137 normal control sera, it also showed an AUC of 0.848 to identify PCa from normal controls. Therefore, the autoantibody bead panel showed similar results to ELISA. In summary, we developed an optimal autoantibody bead-based panel for the detection of PCa. It could be used as a high throughput tool to facilitate Pca detection.

Citation Format: Cuipeng Qiu, Xiao Wang, Serina Batson, Giulio Francia, Jian-Ying Zhang. Using Luminex approach to develop a panel of anti-TAA autoantibodies for the detection of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3328.
Abstract 3384: Evaluation of a tumor informed MRD assay with contrived breast cancer samples
Authors: Russell Hugh , LaBrousse Paul , Stetson Daniel , Jones Greg , Marsico Giovanni , Forshew Tim , McEwen Robert , De Bruin Elza , Abbosh Christopher , Hodgson Darren , Dougherty Brian , Hadfield James , Barrett Carl
Published: June 2023

The non-invasive detection of circulating tumor DNA (ctDNA) from plasma has been shown to have clinical value for detection of minimal residual disease (MRD), emergence of resistance, and predicting treatment response. The sensitive detection of MRD following curative treatment allows for the identification of patients destined to recur. Higher assay sensitivities enable the longest lead times to clinical recurrence. Evaluations using several contrived pan-cancer samples have shown that the Inivata RaDaR tumor-informed personalized assay has a high sensitivity to detect ctDNA. The tumor-informed analysis utilizes sequencing of the tumor tissue to identify up to 48 clonal somatic variants, which are selected to generate a personalized panel to track the variants in patient plasma. To evaluate this technology in a disease specific manner, we generated a set of contrived plasma samples from commercial breast cancer patients and healthy individuals. The biological materials allowed us to generate a plasma sample with defined range of tumor content between 0.2 and 0.001% variant allele fraction (VAF). The sample dilutions were intended to recapitulate challenging cell-free DNA inputs that can be encountered in a clinical setting (median input: 5.76 ng [range 0.22 to 103.73]. Through our pan-cancer and breast cancer evaluations, we were able generate both sensitivity metrics along with a small limit of detection (LOD) study. The assay successfully detected tumor fractions in samples tested at 1X and 1.5X of the established LOD95% of 0.0011% VAF. RaDaR was able to detect the majority of MRD+ samples at 0.5X LOD. We will present data showing the level of sensitivity based on using both 2000 and 20000 genomic equivalents. The RaDaR assay showed high sensitivity for pan-cancer as well as ER+ breast cancer, which is known to have a low TMB compared to other subtypes. In this albeit small LoD study the Inivata assay was highly sensitive for the detection of ctDNA which is imperative to confidently detect ctDNA in advance of overt clinical recurrence.

Citation Format: Hugh Russell, Paul LaBrousse, Daniel Stetson, Greg Jones, Giovanni Marsico, Tim Forshew, Robert McEwen, Elza De Bruin, Christopher Abbosh, Darren Hodgson, Brian Dougherty, James Hadfield, Carl Barrett. Evaluation of a tumor informed MRD assay with contrived breast cancer samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3384.
Abstract 339: Exploring the roles of GSDMD in mammary tumorigenesis
Authors: Zhang Derek Yisen
Published: June 2023

Breast cancer (BC) is the most frequently diagnosed cancer worldwide, with an ever-increasing incidence rate. While screening and treatment methods have improved markedly over the last three decades, BC remains the leading cause of malignant death in women annually. Therefore, there is an urgent need to identify new biomarkers and therapeutic targets for BC. Recently, dysregulated gasdermin D (GSDMD), a central player of inflammation-induced cell death called pyroptosis, has been shown to be implicated in cancer. However, it is unclear whether GSDMD plays important roles in BC. In addition, the molecular mechanism underlying GSDMD-involved tumorigenesis remains unknown. Here, we discovered that GSDMD is significantly upregulated in BC. We have shown that overexpression or knockout of GSDMD increases or decreases BC cell proliferation, respectively. In addition, we have identified and validated several novel GSDMD-interacting proteins that play important roles in breast cancer. These present findings will not only shed light on the roles of GSDMD in BC tumorigenesis but also provide new preclinical data for future successful treatments of BC patients.

Citation Format: Derek Yisen Zhang. Exploring the roles of GSDMD in mammary tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 339.
Abstract 380: BaF3-BTK engineering cell lines as a useful platform for novel BTK inhibitor discovery
Authors: Wang Guoqian , Chen Hongling , Wang Yu , Xu Jingxiao , Zhou Lin , Tang Yao , Ning Jinying , Hao Feng
Published: June 2023

Introduction: Bruton's tyrosine Kinase (Btk) plays an essential role in BCR signaling and has been a hot new target in molecular targeted therapy in the past few years, led by the clinical success of ibrutinib and acalabrutinib, two first approved BTK inhibitors, that greatly improved the outcome of patients with relapsed chronic lymphocytic leukemia (CLL). Currently, there are 5 approved BTK inhibitors including additionally zanubrutinib, orelabrutinib and velexbru. Up to 24 BTK inhibitors are under active clinical evaluation as antineoplastic drugs against chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), B-cell malignancies and mantle cell lymphoma (MCL) in different countries. However, the acquired resistance due to the C481S mutation in BTK remains an essential challenge to improve the poor prognosis of the patient. Subsequently, several next-generation BTK inhibitors are under clinical evaluation targeting the BTK-C481S mutation, such as LOXO-305, a highly selective and non-covalent BTK inhibitor which potently inhibits the cellular activity of BTK C481S/T/R mutations.

Methods: Our group has generated 9 Ba/F3-BTK engineering cell lines as useful cell line and xenograft models for novel BTK inhibitor discovery and development. They harbor wild type BTK or ibrutinib resistance BTK mutations such as the C481S and C481S-double mutations, and the T474M/I mutations. Moreover, we generated Ba/F3-ETV6-BTK-F517L and Ba/F3-ETV6-BTK-C481S-F517L cell lines using a dose-escalation method after treating cells with a mutagenic agent. The BTK-F517L mutation is highly resistant to both ibrutinib and LOXO-305 treatment. Crystal structure analysis and 3D modeling revealed that the F517L mutation may affect the conformation of the BTK binding pocket to ibrutinib, indicating novel BTK inhibitors may be needed to overcome this resistance mutation.

Conclusion: The Ba/F3-BTK engineering cell lines can be a useful tool to develop and evaluate next-generation BTK inhibitors and to explore new acquired resistance BTK mutations.

Citation Format: Guoqian Wang, Hongling Chen, Yu Wang, Jingxiao Xu, Lin Zhou, Yao Tang, Jinying Ning, Feng Hao. BaF3-BTK engineering cell lines as a useful platform for novel BTK inhibitor discovery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 380.
Abstract 3801: Identification of circular RNAs associated with the ferroptosis of pancreatic cancer cells
Authors: Paladines Jessica Jazmin Pena , Wang Shao Weng , Wong Chi Hin , Chen Yang Chao
Published: June 2023

The five-year survival rate for pancreatic cancer patients is significantly low which is predominately due to the lack of early-detection biomarkers, and poor response to currently available treatments that include drug resistance development. Recent studies suggest cancer cells can be highly sensitive to a newly defined type of cell death, ferroptosis, which is related to the development of resistance to standard chemotherapy drugs for pancreatic cancer such as gemcitabine and 5-Fluorouracil. However, the role of circular RNAs (circRNAs) in ferroptosis remains largely unexplored. Hence, this project aims to identify circRNAs in ferroptosis to uncover further components and mechanisms, and potentially provide specific effective targets for anticancer therapies. To achieve this, different ferroptosis inducers were tested, where the cystine-depletion method demonstrated higher efficiency based on cell viability and ferroptosis markers such as iron and lipid peroxidation levels. This method was used to induce ferroptosis for 24 hours in pancreatic cancer cells for subsequent circRNA sequencing, and to establish a ferroptosis-resistant cell line. From the sequencing data, 31 differentially expressed (DE) circRNAs were obtained, and eight were selected for further validation. Three circRNAs; circPPP2R2B, circNDUFAB1, and circNLN, showed consistent differential expression in ferroptosis non-resistant (downregulated) and resistant (upregulated) cells after cystine-depletion treatment. The knockdown of these circRNAs decreased the viability, invasion, migration, and colony formation ability of PDAC cells. Simultaneously, lipid peroxidation and intracellular iron levels, which are ferroptosis markers, were increased. Additionally, bioinformatic analysis of these circRNAs demonstrated their possible interaction with proteins and miRNAs with reported ferroptosis-related functions. In summary, these findings suggest that circPPP2R2B, circNDUFAB1, and circNLN may be critically involved in PDAC progression and the evasion of ferroptosis which supports the further investigation of their oncogenic and ferroptosis-related action mechanism to propose a potential target for anticancer therapies.

Citation Format: Jessica Jazmin Pena Paladines, Shao Weng Wang, Chi Hin Wong, Yang Chao Chen. Identification of circular RNAs associated with the ferroptosis of pancreatic cancer cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3801.
Abstract 3813: Role of CDKN2B antisense RNA 1 in Epstein-Barr virus associated gastric carcinoma
Authors: Torres Keila , Olivares Wilda , Carrasco Gonzalo , Rodriguez Andres , Corvalan Alejandro H.
Published: June 2023

Background: Epstein-Barr Virus (EBV)-associated gastric carcinoma (GC) (EBVaGC) is a subtype of GC which represents approximately 10% of this tumor. CDKN2B antisense RNA 1 (CDKN2B-AS1) is a long non-coding RNA associated with chronic diseases like cardiovascular disease and cancer. This study aimed to evaluate if the loss of CDKN2B-AS1 shifted the balance of the competing endogenous RNA (ceRNA) networks favoring an apoptosis resistant phenotype through ebv-miR-BART4-5p/BID in EBVaGC.

Material and Methods: Seven gastric cancer cell lines were used to determine the expression of CDKN2B-AS1. Specific cell lines were used for CDKN2B-AS1 inhibition assays, the transfection of ebv-miR-BART4-5p and the binding between CDKN2B-AS1 and ebv-miR-BART4-5p. The detection of BID protein was determined by Western Blot. To validate the in vitro results, we detected CDKN2B-AS1, BID, and ebv-miR-BART4-5p expression levels by RT-qPCR in four EBVaGC positive and five negative cases. Apoptosis was detected by in situ hybridization (ISH).

Results and Discussion: CDKN2B-AS1 is expressed at higher levels in NCI-N87 and SNU-1 cell lines and at lower levels in AGS and KATO-III. A significant decrease in BID protein expression was observed after co-transfection with ebv-miR-BART4-5p and CDKN2B-AS1 inhibitor. However, these results were not significant at mRNA level. The luciferase assay validated the predicted binding between ebv-miR-BART4-5p and CDKN2B-AS1. In EBVaGC samples, a significant decrease in BID expression and cleaved caspase-3 in the presence of miR-BART4-5p was observed. Our results show that decreasing CDKN2B-AS1 expression favors viral miR binding on BID mRNAs through ceRNA networks. Also, these results could explain why positive EBVaGCs have lower apoptosis rate than EBVaGC negative cases.

Grant support: ANID/FONDECYT/POSTDOCTORADO/3201028, CONICYT-FONDAP-15130011, FONDECYT-1191928

Citation Format: Keila Torres, Wilda Olivares, Gonzalo Carrasco, Andres Rodriguez, Alejandro H. Corvalan. Role of CDKN2B antisense RNA 1 in Epstein-Barr virus associated gastric carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3813.