bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1110: Estrogen receptor β signaling induced radio-resistance in bladder cancer
Authors: Ide Hiroki , Murakami Tetsushi , Kashiwagi Eiji , Kitaoka Sotaro , Komatsuda Akari , Nishimura Chugo , Funakoshi Shinsuke , Oya Mototsugu , Miyamoto Hiroshi
Published: March 2024

Introduction and Objective: Although radiotherapy (RT) combined with chemotherapy shows comparable outcomes to surgery, women tend to have higher mortality from bladder cancer (BC) compared to men. Emerging evidence indicated the involvement of estrogen receptorβ (ERβ) signaling in BC progression and the association of ER signaling with DNA repair pathways which are involved in radio-resistance. Therefore, we evaluated the association between ERβ signaling and the sensitivity to BC cells.

Methods: We compared the inhibitory effect of RT on BC cell viability between ERβ positive [5637 and T24 expressing control short-heparin RNA (shRNA)] and ERβ negative (5637 and T24 expressing ERβ-shRNA) cells. Moreover, we compared DNA double-strand breaks (DSB) via γ-H2AX foci formation, as well as the expression levels of DNA repair genes, in these cells after RT. Finally, in a mouse xenograft model, we evaluated it on the growth of ERβ positive tumor treated with radiation combined with mock or tamoxifen treatment.

Results: In vitro experiment showed that ionizing radiation reduced the numbers of viable cells in ERβ negative cells more significantly than those in ERβ positive cells. Moreover, tamoxifen treatment reduced the numbers of viable cells more than mock treatment in ERβ positive cells, while tamoxifen didn’t show significant effects in ERβ negative cells. The down-regulation of ERβ resulted in a delay in DSB repair 4-24 hours after RT. We then established radio-resistant cells and confirmed considerable elevation of the expression of ERβ as well as DNA repair genes including RAD51 and RAD54. Estradiol treatment induced the expression of these genes and tamoxifen abolished these effects. Finally, in vivo experiment showed that tamoxifen enhanced the cytotoxic effects of RT in xenograft bearing mice.

Conclusions: These results indicated that ERβ signaling inversely correlated with sensitivity to RT in BC. Thus, ERβ inhibition have a potential of enhancing the cytotoxic effects of radiation, especially in ERβ positive BC cells.

Citation Format: Hiroki Ide, Tetsushi Murakami, Eiji Kashiwagi, Sotaro Kitaoka, Akari Komatsuda, Chugo Nishimura, Shinsuke Funakoshi, Mototsugu Oya, Hiroshi Miyamoto. Estrogen receptor β signaling induced radio-resistance in bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1110.
Abstract 1125: TNIK induced chemoradiation resistance in small cell lung cancer
Authors: Chowdhury Dipanwita Dutta , Imada Eddie , Connis Nick , Nguyen Triet , Chang Jinhee , Council Danielle , Lafargue Audrey , Rezaee Mohammad , Tran Phuoc T. , Marchionni Luigi , Hann Christine L.
Published: March 2024

Chemoradiation (CRT) is the standard first-line therapy for limited stage small cell lung cancer (LS-SCLC), which is characterized by early metastasis, intrinsic-acquired CRT resistance and tumor recurrence. CRT confers a 20-25% 5-year overall survival in patients with LS-SCLC. In this study, we utilized patient-derived xenograft (PDX) models to demonstrate CRT resistance of SCLC and identified CRT resistance candidate genes from various SCLC PDX that represent the majority of molecular subtypes (n=4 that include SCLC-A and SCLC-N subtypes). RNA-seq data identified TNIK (Traf2- and Nck-interacting kinase) as one such gene that is consistently upregulated in SCLC PDXs exposed to CRT compared to single modality treatments. Copy number gains of TNIK were also present in human SCLC samples. Overcoming this CRT resistance is crucial in improving treatment outcomes and patient survival. Genetic depletion or pharmacological inhibition of TNIK reduced the in vitro clonogenic survival of TNIKhigh SCLC cells, NCI-H446 and makes them increasingly sensitive to CRT. In vivo, pharmacological inhibition of TNIK with the inhibitor NCB-0846, enhances the CRT sensitivity of NCI-H446 cell line-derived xenografts (CDX) in NOD SCID immunodeficient mice. Furthermore, pharmacological inhibition of TNIK in vivo demonstrated sensitivity to CRT in LX33 PDX. In conclusion, our results indicate that TNIK plays a role in conferring resistance to CRT in vitro in SCLC cell lines and in vivo in SCLC CDX and PDX models and therefore, can be a potential therapeutic target in limited stage SCLC.

Citation Format: Dipanwita Dutta Chowdhury, Eddie Imada, Nick Connis, Triet Nguyen, Jinhee Chang, Danielle Council, Audrey Lafargue, Mohammad Rezaee, Phuoc T. Tran, Luigi Marchionni, Christine L. Hann. TNIK induced chemoradiation resistance in small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1125.
Abstract 1179: A single-cell atlas of human gastric malignancy reveals CD8+ T cell rejuvenation as a mediator of immunotherapeutic response
Authors: Liang Han , Chen Jian , Luo Yikai , Kang Muxing , Liu Wei , Wang Lie
Published: March 2024

Immune checkpoint inhibition (ICI) has emerged as a mainstream treatment for patients with advanced stage gastric cancer (GC). Cellular heterogeneity in the tumor microenvironment (TME) has a profound impact on the therapeutic responses to ICI. Here we characterize the kinetics of tumor-infiltrating immune populations in GC patients treated with anti-PD-1 using single-cell RNA and TCR sequencing (scRNA/TCR-seq), including primary tumors and metastases. Integrated with public GC scRNA-seq datasets, we construct the largest multi-omic atlas of GC cell states to date that captures the full trajectory of GC malignancy and comprises >320,000 cells derived from 73 samples. We demonstrate the rejuvenation of exhausted CD8+ T cells into a proliferative state marked by the loss of LAYN expression as a major mediator of ICI response in the TME. In parallel, we observe enhanced immunogenicity of cancer cells in post-ICI tumors, thus highlighting the coordination between tumor-intrinsic and microenvironmental factors elicited by ICI. Our study provides a valuable resource of investigating the tumor response to anti-PD-1 treatment and suggests novel strategies for combination therapy and patient stratification.

Citation Format: Han Liang, Jian Chen, Yikai Luo, Muxing Kang, Wei Liu, Lie Wang. A single-cell atlas of human gastric malignancy reveals CD8+ T cell rejuvenation as a mediator of immunotherapeutic response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1179.
Abstract 1180: Asparagine deprivations enhance CD8+ T cell antitumor activity and ICB immunotherapy via ROS-mediated NFAT nuclear localization
Authors: Chang Hsuan Chia , Lin Kwang-Huei , Yang Huang Yu
Published: March 2024

Metabolic reprogramming plays a crucial role in the differentiation and activation of T cells. Both oxidative phosphorylation (OXPHOS) and glycolysis are upregulated to meet the high energy demands. However, the impact of amino acid metabolism on T cell activation remains unclear. Our study demonstrates that asparagine deprivation enhances the antitumor activity of CD8+ cytotoxic T lymphocytes (CTLs) and increases the flux of glycolysis and the tricarboxylic acid (TCA) cycle. Mechanistically, asparagine deprivation induces the production of reactive oxygen species (ROS) through mitochondrial complex I deficiency and leads to nuclear localization of NFAT, subsequently affecting cytokine production and T cell activation. In the animal model, we found that the clinical drug leunase exhibits antitumor activity that relies on the immune response of T cells. Leunase treatment promotes the activation of tumor-infiltrating lymphocytes (TILs) in vivo. Furthermore, the combination therapy of leunase and anti-PD1 demonstrates a synergistic effect. In patients with nasopharyngeal carcinoma (NPC), leunase therapy enhances the cytokines production and activation of CD8+ T cell. In summary, our findings offer valuable insights into how T cells adapt to changes in their nutritional environment.

Citation Format: Hsuan Chia Chang, Kwang-Huei Lin, Huang Yu Yang. Asparagine deprivations enhance CD8+ T cell antitumor activity and ICB immunotherapy via ROS-mediated NFAT nuclear localization [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1180.
Abstract 1222: Mechanisms of succinate-mediated drug resistance in ER+ breast cancer
Authors: Hogstrom Jenny , Petrova Boryana , Perea Camila , Ward Madelyn , Wulf Gerburg , Collins Laura , Kanarek Naama , Muranen Taru
Published: March 2024

Breast cancer is the most prevalent cancer among women worldwide with estrogen receptor-positive (ER+) cancer constituting 70% of all breast cancers. Although ER+ breast cancers initially respond well to ER-targeted therapies and CDK4/6 inhibitors, approximately 30% patients acquire resistance to targeted therapies. In addition to intrinsic resistance that tumors develop, cancer-associated fibroblasts (CAFs) residing in the tumor microenvironment, also contribute to treatment-resistance and poor survival. The role of secreted metabolites by CAFs has been poorly studied in the context of targeted drug resistance. Therefore, to investigate the mechanism of stroma-mediated resistance to CDK4/6-inhibitors, we propagated patient-derived organoids (PDOs) and CAFs from treatment-naïve and treatment-resistant ER+ breast tumors. Our data show that the CAF conditioned media containing the polar metabolite fraction stimulates resistance to CDK4/6 inhibitors. Therefore, to identify metabolites that drive resistance, we performed an untargeted metabolomics on CAF conditioned media. We observed that the CAF conditioned media was enriched for TCA cycle intermediates, including succinate. Succinate is an oncometabolite that has an essential function in energy metabolism. Aside from causing metabolic reprogramming, succinate can be transported out of the mitochondria to perform other biological processes, such as epigenetic modifications and pseudohypoxia. Furthermore, succinate in the extracellular space binds to the succinate receptor SUCNR1 and induces G-protein coupled receptor-mediated signaling in cells expressing SUCNR1. To assess mechanism of resistance that these metabolites might induce, we treated breast cancer cells with the most enriched metabolites we had identified in our metabolomics assay. These experiments identified succinate as driver of CDK4/6 inhibitor resistance in ER+ breast cancer cells. Furthermore, treating tumor cells with SUCNR1 antagonist increased their sensitivity to CDK4/6-inhibitors, showing that succinate mediated resistance is conveyed via SUCNR1. This suggests that CAF secreted succinate plays a role in supporting a pro-tumor environment by promoting drug resistance towards CDK4/6 inhibitors. Defining this mechanism of resistance will elucidate potential therapeutic targets to augment patient response to therapy.

Citation Format: Jenny Hogstrom, Boryana Petrova, Camila Perea, Madelyn Ward, Gerburg Wulf, Laura Collins, Naama Kanarek, Taru Muranen. Mechanisms of succinate-mediated drug resistance in ER+ breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1222.
Abstract 123: Establishment of humanized CD3EDG/4-1BB mice for testing novel T cell engagers
Authors: Shang Chengzhang , Li Chong , Zhao Lei , Xu Qingqing , Jin James , Ren Menglian
Published: March 2024

Agonism of the 4-1BB receptor is an attractive therapeutic strategy for tumor cell therapy, due to the ability of this receptor to stimulate activity of T and NK cells for tumor cell killing. Many classes of biologics targeting 4-1BB have been generated to date, including bispecific or multispecific engagers designed to stimulate 4-1BB and CD3 activity on the T cell surface. To provide a preclinical platform to test such therapies, Biocytogen generated a humanized B-hCD3EDG/h4-1BB mouse model. This was achieved by crossing humanized mice expressing chimeric CD3E, CD3D and CD3G with mice expressing the human extracellular region of 4-1BB. Previously, we showed that these humanized CD3EDG mice and humanized 4-1BB mice can be used for validating efficacy of anti-human CD3 and 4-1BB antibody-based therapies, respectively. Here, we confirmed expression of human CD3 and 4-1BB on splenocytes in homozygous humanized B-hCD3EDG/h4-1BB mice by flow cytometry, which also demonstrated lack of murine CD3 and 4-1BB expression. Assessment of leukocyte subpopulations in the spleen and peripheral blood confirmed that humanization of the mice did not significantly alter the development or frequency of blood cell populations. Further evaluation of lymphocyte frequency was also performed, which demonstrates no significant alterations in the CD4/CD8 T cell ratio in spleen, lymph node, or peripheral blood of the animals. Presence of Treg and NK cells was also confirmed. Taken together, these data indicate that the humanized B-hCD3EDG/h4-1BB model is a novel, robust tool for testing novel T cell engagers.

Citation Format: Chengzhang Shang, Chong Li, Lei Zhao, Qingqing Xu, James Jin, Menglian Ren. Establishment of humanized CD3EDG/4-1BB mice for testing novel T cell engagers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 123.
Abstract 1291: ASPIRE: Advocates and Scientists Partner in Research Education
Authors: Betts Laurie , Potter Jennifer , Carlisle Veronica , Spears Patricia A.
Published: March 2024

UNC Lineberger Comprehensive Cancer Center (LCCC) has a robust and growing cancer patient advocacy community. UNC Lineberger is the flagship publicly funded comprehensive cancer center in North Carolina. We are constantly looking for ways to educate patient and community advocates on recent advances in cancer biology research and clinical trials. Our advocates are also valuable partners for our basic and clinical researchers that may be early in their career. We developed and implemented an inaugural research education program for new advocates. We model this program partly on the highly successful and popular AACR Scientist Survivor Program that recently celebrated it’s 25th anniversary. Some of the UNC LCCC advocates participate in the SSP and were motivated to develop our own local program.

We will showcase our pilot 2024 ASPIRE education program that brings together cancer patient and community advocates and cancer researchers to learn from each other. The basic itinerary of the ASPIRE program is:

1.Three separate one-hour online presentations by UNC Lineberger researchers with a patient advocate moderator. The topics covered are:

Cancer 101 : cancer biology, diagnosis, staging, treatment.

Clinical Trials 101: clinical trials, phases, patient protection, progress with for example accrual of representative populations.

Research 101: bench research, bioinformatic, AI, how research works and how it may lead to new treatments. To showcase a tissue donation program being developed by UNC Patient Advocates that will benefit researchers.

2.The lecture series is followed by a breakfast roundtable discussion for all the participants to discuss their experiences. There will be short presentations and discussions in small groups of researchers and cancer patient advocates.

We will summarize our ASPIRE pilot experience, how our participation in the SSP program motivated this local program, and how we plan to incorporate feedback from participants to inform future programs. We hope this will lead to more informed patient advocates that can then be even more powerful in research advocacy.

Citation Format: Laurie Betts, Jennifer Potter, Veronica Carlisle, Patricia A. Spears. ASPIRE: Advocates and Scientists Partner in Research Education [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1291.
Abstract 1330: Universal and potent chimeric antigen receptor-natural killer (upCAR-NK) cells for B-cell malignancy treatment
Authors: Kwon Daekee , Moon Bo Kyung , Han Mijung , Lee Tae-Wook , Lee Ji-Han , Kang Kyung-Sun
Published: March 2024

Patient-specific CAR-T cells treatment is an innovative new drug that has shown excellent therapeutic effects against B cell blood cancer. However, side effects such as cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome and ultra-high treatment costs must be overcome, and off-the-shelf CAR-NK cells can be a good alternative. The purpose of this study was to combine cellular reprogramming, gene editing, and differentiation technologies to produce full-off-the-shelf NK cells derived from iPSCs and verify their efficacy and safety. To this end, somatic cell-derived iPSCs were produced, seven genes were edited using CRISPR/Cas9 technology (5 kinds of knockout and 2 kinds of overexpression), and differentiated into NK cells. upCAR (universal and potent chimeric antigen receptor)-iPSCs with seven genes edited showed bialleic INDEL in the coding sequence of five genes and no off-target effect. In addition, normal transcriptome and genetic stability were maintained. High-efficiency differentiated upCAR-NK cells showed a very high differentiation yield, in vitro proliferation, and freezing/thawing were possible. In addition, upCAR-NK cells secrete interferon-γ when they meet cancer cells, showing cytotoxic effects in vitro and in vivo. upCAR-NK cells themselves showed no obvious toxicity in vivo. In conclusion, this study developed upCAR-iPSCs and upCAR-NK cells platform technologies that are less likely to have side effects and can be economically developed for B cell blood cancer than CAR-T. In the future, this technology can be useful in developing full-off-the-shelf CAR-NK cells anti-cancer immune cell therapy with low side effects, high efficacy, and low price.

Citation Format: Daekee Kwon, Bo Kyung Moon, Mijung Han, Tae-Wook Lee, Ji-Han Lee, Kyung-Sun Kang. Universal and potent chimeric antigen receptor-natural killer (upCAR-NK) cells for B-cell malignancy treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1330.
Abstract 1388: Spatial transcriptome analysis of EMT trait and corresponding immune composition study in HNSCC
Authors: Chung Chih-Hung , Yang Muh-Hwa
Published: March 2024

Head and neck squamous cell carcinoma (HNSCC) is a prevalent malignant cancer worldwide. Conventional treatments for this type of cancer involve surgical resection, radiation therapy, and chemotherapy. However, in cases where the tumor recurs or metastasizes distantly, rendering surgical resection or radiation therapy impractical, the only viable option is the utilization of targeted drugs in conjunction with chemotherapy. Despite such interventions, the prognosis remains grim. Hence, the identification of genetic characteristics or biomarkers associated with easy metastasis or recurrence in the early stages of the tumor has become imperative for effective head and neck cancer treatment. The objective of this study is to analyze tumor specimens from the primary site and lymph node metastasis post-surgical resection in HNSCC patients. We employed the GeoMx Digital Spatial Profiler (DSP) for imaging-based digitized gene expression analysis. The DSP provides gene expression information at specific locations we selected, such as the primary tumor center core, primary tumor invasion borders, and lymph node metastasis specimens. The GeoMx DSP spatial analysis tool illustrates the cancer transcriptome atlas (CTA), encompassing the expression of 1825 genes related to cancer and immunity from the primary site to the metastatic site. We identified tissue-specific EMT gene expression patterns in paired tumor regions of interest (ROIs). ITGB1 emerged as the most significant co-expression marker in the primary outer tumor and metastatic tumor. Distinct EMT signature genes expressed in the HNSCC primary site and lymph node metastatic site were also discovered. Additionally, we conducted CTA profiling in correlated stroma ROIs. Through myeloid activity analysis in HNSCC paired samples, we identified distinct myeloid activation signatures in primary and metastatic site stroma cells. These results will provide crucial insights into head and neck cancer treatment strategies based on different tumor site characteristics.

Citation Format: Chih-Hung Chung, Muh-Hwa Yang. Spatial transcriptome analysis of EMT trait and corresponding immune composition study in HNSCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1388.
Abstract 1431: Generation of transparent transgenic zebrafish strain to study cardiovascular functionin translational cardio-oncology research
Authors: Rajpurohit Surendra K. , Gossman Taitum , Pasi Rohan , Nasanally Sai , Bhatt Keshu , Van Derhei Aiden , Hotwani Simaran , Manhiani Devan-Anmol , Kimmerling Kirk , Arora Vishal , Lokeshwar Balakrishna L.
Published: March 2024

Transgenic zebrafish lines have been extensive use in biomedical research and their primary applications. The zebrafish has proven to be an ideal tool for direct In-vivo observations of developmental processes. In vertebrates, including zebrafish and humans, the ability to obtain high-resolution, in-vivo imaging is limited due to the natural opacification of skin and subdermal structures. To conduct in-vivo whole organism imaging studies, skin transparency is a primary requirement, necessitating blocking of pigmentation. Zebrafish casper mutant maintains skin transparency throughout its life, offering the ideal combination of sensitivity and resolution for in-vivo analyses and imaging. Our Laboratory is developing a novel transparent transgenic zebrafish model to address basic and translational cardio-oncology and experimental therapeutics. The fli1 promoter in zebrafish having an enhanced expression reporter gene of green fluorescent protein, GFP. This model allows for the in-vivo examination of the vascular system by using fluorescent and confocal microscopy. In this study, we developed transparent transgenic strain of Caspe/fli1:EGFP and integrating with recently developed casper/myl7/annexin-5:YFP and casper/fli1:EGFP strain through crossbreeding of casper and transgenic zebrafish. We have generated F1 generation of newly developed strains of Caspe/fli1:EGFP and casper/myl7/annexin-5:YFP and casper/fli1:EGFP. The presence of fluorescence expressions RFP, YFP and GFP, in crossbred progeny identified through the screening of larvae at 72 HPF stage. Expression of these fluorescent proteins determines the presence of the reporter genes. Positive transgenic larvae validated by confocal microscopy. The progenies of the F1 generation are heterozygous in nature and allow them to grow into adults to breed transparent transgenic strain to get F2 generation. This process will be continued to F2, F3, F4 and F5 generation until obtain homozygous progeny of newly developed strain. The development of casper/myl7/annexin-5:YFP x casper/fli1:EGFP will aid new dimensions in Cardio-oncology research for cardiac injury and Vasotoxicity induced by anticancer drug treatment in cancer patients.

Citation Format: Surendra K. Rajpurohit, Taitum Gossman, Rohan Pasi, Sai Nasanally, Keshu Bhatt, Aiden Van Derhei, Simaran Hotwani, Devan-Anmol Manhiani, Kirk Kimmerling, Vishal Arora, Balakrishna L. Lokeshwar. Generation of transparent transgenic zebrafish strain to study cardiovascular functionin translational cardio-oncology research [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1431.
Abstract 1493: Secretory function of neuroendocrine-like prostate cancer cell enhances therapy resistance and metastatic potential of adjacent adenocarcinoma
Authors: Mishra Juhi , Bodas Sanika , Bhattacharya Sreyashi , Das Dipanwita , UnNazir Sheeraz , Dutta Samikshan , Datta Kaustubh
Published: March 2024

Background: Castration-resistant prostate cancer (CRPC) is advanced non curable disease associated with poor prognosis and survival n patients. Androgen receptor pathway inhibitors (ARSIs) are used to treat advanced prostate cancer, but in majority of patients it progresses by developing resistance to AR-directed therapies and is referred to as CRPC. Adenocarcinoma cells in CRPC tumor often undergoes lineage switch and become neuroendocrine-like cancer cells (NEPC) with small-cell histology, characterized by the loss of Tp53 and Rb1 genes. NEPC cells are frequently present as scattered foci along with prostatic adenocarcinomas and are secretory in nature.

Objective: The primary objective is to determine how these NEPC cells influence surrounding adenocarcinoma and aids in disease progression.

Methods: We performed Mass spec and cytokine array experiments from NEPC conditioned media to determine its secretory phenotype. We orthotopically implanted NEPC and adeno cells in the nude mice and performed colony formation assay on flow sorted adeno cells from the tumor. We also checked the effect of NEPC on the adeno cells and TME components.

Results: Our analysis of the RNA-seq data from SU2C-PCF cohort and NE-like cells revealed that functions related to secretion were significantly enriched in NE-like PCa. NE-like cells gain the ability to secrete neuropeptides, growth factors, and cytokines to establish cell-cell communication. This secretory function is tightly regulated by the Neuropilin2 (NRP2) axis. We studied for the effect of NE-like cells on the adenocarcinoma cells in the mice tumor model and found that the presence of NE-like cells facilitated the adenocarcinoma cells to acquire aggressive characteristics. This could be due to both direct and/or through the modulation of the tumor microenvironment (TME). The direct effect could be by enhancing the therapy resistant ability of the surrounding adenocarcinoma, and the indirect effect is through the tumor microenvironment especially through neurons and macrophages that promote metastatic potential.

Conclusions: Overall, our results prove that NE-like cancer cells influence the adjacent adenocarcinoma cells directly by making them resistant to treatment and indirectly make them more metastatic by utilizing macrophage-neuron axis.

Citation Format: Juhi Mishra, Sanika Bodas, Sreyashi Bhattacharya, Dipanwita Das, Sheeraz UnNazir, Samikshan Dutta, Kaustubh Datta. Secretory function of neuroendocrine-like prostate cancer cell enhances therapy resistance and metastatic potential of adjacent adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1493.
Abstract 1534: The tumor immune microenvironment at different stages of non-small cell lung cancer
Authors: Hansen Mackenzie , Zhou Qin , DeRose Yoko S. , Kovacsovics Magdalena , Brintz Benjamin , Witt Benjamin L. , O'Neill Kim L. , Hu-Lieskovan Siwen
Published: March 2024

Introduction: To better understand the immune microenvironment in non-small cell lung cancer (NSCLC), we evaluated immune cell populations within the tumor parenchyma and their interaction at the periphery of the tumors at different stages of disease.

Methods: 27 whole tumor slides were collected from NSCLC patients of all stages after surgical resection. Tissue slides were stained with multiplex Opal immunofluorescence for CD8, CD68, CK, PD-1, PD-L1, FoxP3, and DAPI using a Leica Bond automated stainer. Slides were scanned at 20x magnification by Akoya Polaris whole slide scanner and spectrally unmixed. Spatial analyses were performed on the HALO software.

Results: Density of CD8 T cells, T regulatory (Treg) cells, and macrophages were significantly higher (p=0.001) in the interface region compared to the tumor and stroma. There is abundant CD8 T cell infiltration in the tumor at early stage I and this decreased significantly (p=0.05) at higher stages. PD-1− CD8 T cells significantly dropped with higher stage, while PD-1+ CD8 T cells did not. On the other hand, macrophage infiltration increased as stage increased, with most of the increase driven by PD-L1− macrophages. No statistically significant trends were observed in Treg cells across stages, but they appeared to be higher in stage II. No differences in immune infiltration were observed between the NSCLC subtypes adenocarcinoma and squamous cell carcinoma. No correlation was found between proximity of immune cell subsets to tumor cells and stage.

Conclusion: The tumor microenvironment in NSCLC is rich in CD8+ cells at early stage and their density decreases as the stage of the disease increases, while macrophage infiltration increases as the disease stage progresses.

Citation Format: Mackenzie Hansen, Qin Zhou, Yoko S. DeRose, Magdalena Kovacsovics, Benjamin Brintz, Benjamin L. Witt, Kim L. O'Neill, Siwen Hu-Lieskovan. The tumor immune microenvironment at different stages of non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1534.
Abstract 1547: Melanotransferrin (MELTF, CD228) as a determinant of melanomagenesis: MELTF expression attenuates melanoma progression in the A375-luc2 murine metastasis model and human patients
Authors: Jandova Jana , Wondrak Georg T.
Published: March 2024

The cell-surface glycoprotein melanotransferrin (MELTF, CD228) is a member of the iron-binding transferrin superfamily expressed in human melanocytes and other cell types. Dysregulated MELTF expression in malignant melanoma has been reported before but its specific role in tumorigenesis has remained elusive. Following our interest in the role of the transferrin receptor (TFRC) in melanomagenesis, our initial TCGA-analysis suggested that high MELTF expression is associated with increased melanoma patient survival. In order to explore the mechanistic role of MELTF in melanoma we performed CRISPR/Cas9-based MELTF deletion in A375-luc2 human malignant melanoma cells. ICP/MS-analysis indicated no difference in iron-content between MELTF_WT and MELTF_KO cells. NanoString nCounterTM ('PanCancer-Progression-Panel'; 770 genes) comparative transcriptomic profiling (MELTF_KO versus MELTF_WT) demonstrated an upregulation of EMT-related expression networks in MELTF_KO cells, an observation consistent with phenotypic detection of enhanced matrigel invasiveness; likewise, inclusion of recombinant glycosylated MELTF (Gly20-Gly711 His-tag; 92 kDa; 10-100 nM) attenuated invasiveness of MELTF_KO cells. In SCID mice, subcutaneous growth of MELTF_KO exceeded that of MELTF_WT xenografts, and intracardial injection followed by non-invasive bioluminescent monitoring revealed an increase in lung metastatic burden associated with decreased survival of the MELTF_KO group. These data support a heretofore unrecognized tumor-suppressive function of MELTF expression in melanoma progression, a mechanistic role consistent with a potential survival benefit associated with high MELTF expression observed in human melanoma patients.

Citation Format: Jana Jandova, Georg T. Wondrak. Melanotransferrin (MELTF, CD228) as a determinant of melanomagenesis: MELTF expression attenuates melanoma progression in the A375-luc2 murine metastasis model and human patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1547.
Abstract 1590: The role of mucosal associated invariant T (MAIT) cells in pancreatic cancer liver metastasis
Authors: Toor Jugmohit , Dimitrion Peter , Krevh Rachel , Subedi Kalpana , Zuniga Bobby , Wang Jie , Zhou Li , Mi Qing-Sheng
Published: March 2024

Pancreatic cancer liver metastasis (PCLM) is present in 50% of patients diagnosed with pancreatic ductal adenocarcinoma (PDAC) and leads to a median survival of <6 months. Here, we sought to identify urgently needed therapeutic options for treating PCLM by targeting mucosal associated invariant T (MAIT) cells, an invariant T cell restricted to MHCI related protein (MR1). MAIT cells are a unique subset of T cells that are classified as “protumor” MAIT17 and “antitumor” MAIT1 cells. We hypothesize that the PCLM tumor microenvironment (TME) induces direct MAIT cell activation via MR1 which primes MAIT cells towards a “protumor” phenotype. Using publicly available TCGA data, we found that PDAC tumors were enriched for MR1 gene expression compared to healthy tissue and patients with lower tumor MR1 expression had better overall survival. Next, immunostaining of MR1 on tumor samples from primary PDAC and PCLM revealed that that MR1 protein expression is highly enriched in the TME and localized to the surface of tumor cells. This suggests that MAIT cells in the PCLM TME are being activated directly via an MR1 dependent pathway. Next, we performed hemi-spleen injection of mouse pancreatic cancer cells into WT and MR1 KO (lacks MAIT cells) mice to model PCLM. Interestingly, MAIT cells in the WT PCLM mice were shifted towards the “pro-tumor” MAIT17 phenotype. Furthermore, the lack of MAIT cells in MR1 KO mice significantly reduced tumor burden compared to WT mice. Lastly, we found that MR1 KO mice had a reduction in exhausted NK and NKT cells which likely promoted anti-tumor immunity. Overall, we find that the PCLM TME promotes “pro-tumor” MAIT17 cells via MR1 mediated activation and these MAIT17 cells inhibit anti-tumor cells such as NK and NKT cells. This work identifies targeting MAIT cells as a novel immunotherapy approach for PCLM.

Citation Format: Jugmohit Toor, Peter Dimitrion, Rachel Krevh, Kalpana Subedi, Bobby Zuniga, Jie Wang, Li Zhou, Qing-Sheng Mi. The role of mucosal associated invariant T (MAIT) cells in pancreatic cancer liver metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1590.
Abstract 1639: Telomere dynamics in aging and cancer by nanopore long-read sequencing
Authors: Schmidt Tobias T. , Tyer Carly , Rughani Preeyesh , Haggblom Candy , Jones Jeff , Dai Xiaoguang , Frazer Kelly A. , Gage Fred H. , Juul Sissel , Hickey Scott , Karlseder Jan
Published: March 2024

Telomeres are the protective, nucleoprotein structure at the ends of linear eukaryotic chromosomes. The accurate measurement of both telomeric length and composition of individual telomeres in mammalian cells has been challenged by the length and repetitive nature of telomeres. With the advent of third generation sequencing technologies, it is now technically possible to sequence entire telomeres and map them to individual chromosome arms. Here, we report a reliable method to enrich, sequence and analyze human telomeres using Oxford Nanopore Technologies long-read sequencing. To enrich for telomeric sequences we combine the ligation of adapters complementary to the telomeric G-overhang with restriction enzyme digest to sequence the telomeric C-strand and part of the adjacent subtelomere. The subtelomeric information is harvested to map individual telomeric reads to specific chromosome arms and even alleles. We have measured bulk, chromosome-arm-specific telomere length dynamics during cellular aging of cultured primary cells and in a patient-derived aging cohort. To address the impact of the telomere maintenance mechanism on telomere length and composition, we have sequenced matched pairs of fibroblasts and induced pluripotent stem cells, as well as five well-established telomerase- and ALT-positive cancer cell lines. Our results suggest that based on nanopore telomere long-read sequencing ALT-positive cells can be easily discriminated from normal and telomerase-positive cancer cells. Further, telomere sequencing allows to evaluate the methylation status of the subtelomeric CpG islands adjacent to telomeres.

In summary, nanopore telomere long-read sequencing allows to measure the length and composition of individual telomeres and their mapping to specific chromosome arms. Telomere long read sequencing methods will be valuable tools to study telomere biology during aging and cancer.

Citation Format: Tobias T. Schmidt, Carly Tyer, Preeyesh Rughani, Candy Haggblom, Jeff Jones, Xiaoguang Dai, Kelly A. Frazer, Fred H. Gage, Sissel Juul, Scott Hickey, Jan Karlseder. Telomere dynamics in aging and cancer by nanopore long-read sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1639.
Abstract 1641: Clobetasol differentially effects the vulvar cancer cell line, UMSCV-4, causing both increases in apoptosis while maintaining a subpopulation in quiescence
Authors: Lewis Jani E. , Ogden Mack , Minnuto Gianna , Haering Kia , North Luke
Published: March 2024

Vulvar cancer is a rare but aggressive form of cancer that remains understudied. Our results show that treatment of the vulvar cancer cell line, UMSCV-4, with the glucocorticoid, clobetasol, increases apoptosis. Clobetasol is often used to treat a common inflammatory disease of the vulva known as vulvar lichen sclerosus (VLS) and up to 65% of vulvar carcinomas arise in the background of VLS. This would indicate clobetasol may also decrease the progression of vulvar cancer, if our observations reflect what is happening in vivo. However, our studies also show that apoptosis is not universal for the clobetasol treated UMSCV-4 cells. A subpopulation appears to enter a state of quiescence as evidenced by the return of some of the cells to normal cell proliferation upon removal of clobetasol. We previously showed that p27Kip1 is upregulated in the clobetasol treated cells. Cells that were incubated in clobetasol for three months and then removed from clobetasol (UMSCV-4 LT), allowing them to reenter the cell cycle, no longer expressed high levels of p27Kip1 upon re-exposure to clobetasol. We are now examining the differential phosphorylation states of p27Kip1 in the untreated and treated cells as well as identifying other genes important to clobetasol induced quiescence using RNAseq.

Citation Format: Jani E. Lewis, Mack Ogden, Gianna Minnuto, Kia Haering, Luke North. Clobetasol differentially effects the vulvar cancer cell line, UMSCV-4, causing both increases in apoptosis while maintaining a subpopulation in quiescence [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1641.
Abstract 1690: LSD1 and CoREST2 demethylate STAT3 to promote enteroendocrine cell differentiation in mucinous colorectal cancer
Authors: Ladaika Christopher A. , Ghobashi Ahmed H. , Boulton William C. , O'Hagan Heather M.
Published: March 2024

Across different cancer types, tumor heterogeneity has been shown to drive tumor progression, metastasis, and therapeutic resistance. Adenocarcinoma to neuroendocrine lineage transition is an emergent mechanism of targeted therapy resistance in several cancer types, including lung and prostate cancer. Therefore, understanding the dynamics and mechanisms driving neuroendocrine cell fates in cancer is critical. Mucinous colorectal cancer accounts for upwards of 20% of colorectal cancer cases and is characterized by tumors with mucous accounting for at least 50% of the tumor volume. The normal colon epithelium consists of several specialized cell types, including hormone secreting enteroendocrine cells (EECs), which are the neuroendocrine cell of the intestine. We have previously shown that EEC progenitors are enriched in mucinous colorectal cancer and promote cancer cell survival via secreted factors. Additionally, we have shown that lysine specific demethylase 1 (LSD1) promotes EEC differentiation in these tumors; however, the mechanism by which LSD1 promotes EEC differentiation has remained unknown. Typically to carry out its enzymatic function, LSD1 must be a part of a transcriptional regulatory complex. One such complex is the CoREST complex, which contains LSD1, HDAC1, and one of three CoREST protein family members. Here we report that LSD1 and CoREST2 promote EEC differentiation by demethylating the transcription factor signal transducer and activator of transcription 3 (STAT3) to promote STAT3 chromatin binding. Additionally, we demonstrate that knocking down CoREST2 decreases tumor growth and lung metastases of mucinous colorectal cancer cells injected orthotopically into the colons of immunocompromised mice. Furthermore, we utilized single cell multi-omics that combines single cell RNA sequencing with single cell ATAC sequencing to show that during EEC differentiation there is an increase in chromatin accessibility at regulatory regions of known EEC-promoting transcription factors. Finally, through our single cell multi-omics analysis we identify a new rare cell type with neuron-like but not secretory characteristics. Mucinous colorectal cancer is an aggressive and chemotherapeutically intractable form of CRC that is enriched for EEC progenitors. Our data demonstrates that LSD1 and STAT3 promote EEC differentiation, suggesting that inhibiting LSD1 and or STAT3 may have benefits for patients with mucinous colorectal cancer.

Citation Format: Christopher A. Ladaika, Ahmed H. Ghobashi, William C. Boulton, Heather M. O'Hagan. LSD1 and CoREST2 demethylate STAT3 to promote enteroendocrine cell differentiation in mucinous colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1690.
Abstract 1719: Targeting H3.3 serine 28 phosphorylation as a novel therapeutic strategy for diffuse midline glioma
Authors: Yan Yuqian
Published: March 2024

Background: Diffuse midline glioma (DMG) is a highly aggressive pediatric brain cancer with a poor prognosis [1]. Mechanistically, DMG harbors a lysine-to-methionine substitution at position 27 of histone H3.3 (H3.3K27M), which leads to global hypomethylation and a dramatically altered epigenetic landscape [2, 3].

Aim: This study aims to explore the H3.3K27M mutation from a novel perspective by investigating its effects on H3.3 ubiquitination, with the goal of identifying effective, novel therapeutic targets.

Methods: Mass spectrometry was used to identify potential E3 ligases for H3.3, while immunoprecipitation to verify this interaction. Cell viability and survival were determined by MTS and clonogenic survival assays, respectively. Nucleosome fractionation was performed to detect the distribution of wild-type (WT) or K27M mutant H3.3 histones.

Results: Mass spectrometry identified UBR7, an E3 ligase, as exhibiting a stronger binding affinity with WT H3.3 than its K27M mutant counterpart. Further nucleosome fractionation assay revealed that K27M histones are more likely deposited in nucleosomes than WT H3.3. Further analysis demonstrated that a non-phosphorylation mimic mutation (serine to alanine) at amino acid 28 (S28A) restored the binding affinity of K27M mutant H3.3 with UBR7. Importantly, this mutation disrupted H3.3 phosphorylation and significantly diminished cell survival, whereas the constitutively activated phosphorylation mutation (S to glutamine (E)) on S28 (S28E) mimics K27M mutation in promoting cell proliferation and survival.

Conclusion: These findings suggest that S28ph impairs UBR7-mediated K27M ubiquitination and promotes DMG cell growth, suggesting that targeting S28ph is an effective therapeutic strategy. However, the underlying mechanisms by which K27M interplays with S28ph in promoting tumorigenesis require further investigation.

Citation Format: Yuqian Yan. Targeting H3.3 serine 28 phosphorylation as a novel therapeutic strategy for diffuse midline glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1719.
Abstract 1773: Deciphering the roles of germline predisposition variants and somatic mutations on breast cancer cells and the tumor microenvironment
Authors: Rodrigues Fernanda Martins , Gallant Kelsey , Jayasinghe Reyka , Iglesia Michael , Houston Andrew , Chen Siqi , Lal Preet , Fields Ryan , Gillanders William , Chen Feng , Ding Li
Published: March 2024

While great progress has been made in the understanding and treatment of cancer, tumor heterogeneity remains a diagnostic and prognostic challenge. Single-cell sequencing techniques and high-resolution cellular imaging now afford researchers the ability to investigate the effects of germline variants (especially when compared to somatic mutations) on molecular characteristics of cancer cells and non-cancer cells in the tumor microenvironment (TME). Here, we investigated a breast cancer (BRCA) cohort of over 100 individuals, for which we have generated sc/sn RNA-seq, snATAC-seq, and CODEX imaging data as well as bulk whole exome sequencing and RNA-seq data. We identified 21 BRCA cases carrying pathogenic and likely pathogenic rare germline variants in known cancer predisposition genes, such as BRCA2, BRIP1, ATM, and TP53. These predisposition genes showed diverse expression patterns across stromal and immune cell types, indicating roles in cell types beyond breast epithelia. For cases carrying pathogenic germline variants in such genes, we use snRNA-seq/snATAC-seq to assess gene expression changes and pathway alterations (e.g.,HRD and apoptosis) across all cell types and investigate phenotypic changes associated with pathogenic germline variants in cancer cells compared to stroma and immune cells in the TME. By evaluating chromatin accessibility changes and allele-specific expression, we reveal the connection between epigenetic regulators and differential roles of germline variants across cell types. Finally, the integrated analysis of germline variants and somatic mutations from predisposition genes at single cell resolution broadened our understanding of the role of germline variants in TME during tumorigenesis.

Citation Format: Fernanda Martins Rodrigues, Kelsey Gallant, Reyka Jayasinghe, Michael Iglesia, Andrew Houston, Siqi Chen, Preet Lal, Ryan Fields, William Gillanders, Feng Chen, Li Ding. Deciphering the roles of germline predisposition variants and somatic mutations on breast cancer cells and the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1773.