bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 122: The subclonal dynamics of gross ploidy alterations in TNBC cells under chemotherapeutic pressure
Authors: Gardner Andrea L. , Zheng Lan , Parker Patrik , Howland Kennedy , Morgan Daylin , Brock Amy
Published: April 2023

Solid tumors such as triple-negative breast cancer (TNBC) have characteristically high rates of aneuploidy and polyploidy. Chromosome-level aneuploidies are known to arise through errors in cytokinesis, but many of the mechanisms which drive large-scale ploidy alterations such as whole genome doubling (WGD) are still under investigation. WGD and subsequent ploidy reduction has been implicated as a key event in cancer evolution across patient samples. Further, in vitro studies have suggested that cells may undergo polyploidization to survive certain chemotherapies. Here, we apply lineage tracing technologies to test the hypotheses that some cells are primed to undergo WGD through endocycling or cell-cell fusion and that active polyploidization is a mechanism to resist chemotherapy. To deconvolve WGD by endocycling from WGD by cell-cell fusion, we transduced the TNBC cell line HCC1806 to create parallel populations with transcribed DNA barcodes on H2B-GFP or H2B-mCherry. All experiments were performed with homotypic cocultures containing equal proportion of cells from each fluorescent barcode population. With this method, ploidy alterations due to endoreplication were identifiable as monofluorescent single-barcoded cells with increased ploidy, while ploidy changes due to fusion were identifiable as double positive (GFP+mCherry+) double-barcoded cells with increased ploidy. The cell population was treated with an LD75 dose of doxorubicin and parallel plates were monitored by live-cell imaging. At each timepoint (pre-treatment, then 3, 5, and 14 days post-treatment), triplicate plates were stained with Hoechst and sorted into two fractions for lysis and DNA extraction: Near diploid (~2N) and ii) Hyperploid (> 4N). Barcode composition of cells from each fraction and each timepoint was determined through Illumina sequencing of barcode amplicons. Here, we will present preliminary results revealing the subclonal dynamics of gross ploidy alterations through normal culture conditions and chemotherapeutic perturbation. This study will elucidate whether pre-existing or de novo polyploidy promotes cell survival during chemotherapy, and determine if gross ploidy alterations occur randomly or through deterministic selection.

Citation Format: Andrea L. Gardner, Lan Zheng, Patrik Parker, Kennedy Howland, Daylin Morgan, Amy Brock. The subclonal dynamics of gross ploidy alterations in TNBC cells under chemotherapeutic pressure [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 122.
Abstract 1259: Disturbing the Siglec-Sialoglycan axis to target myeloid- derived suppressor cells in the tumor microenvironment
Authors: Wieboldt Ronja , Zingg Andreas , Carlini Emanuele , Börsch Anastasiya , Läubli Heinz , Manutano Natalia Rodrigues
Published: April 2023

Background: Overexpression of sialic acids on glycans, called hypersialylation is a common alteration in cancer that drives immune evasion via interaction with Sialic acid-binding immunoglobulin-like lectin (Siglec) immunoregulatory receptors on tumor-infiltrating immune cells. Myeloid derived suppressor cells (MDSCs) generated by cancer-induced aberrant myelopoiesis are highly immunosuppressive cells inhibiting immune cells via direct interaction or secretion of suppressive cytokines. Targeting the complex suppressive tumor microenvironment (TME) remains a challenge.

Methods: Siglec expression of murine and human MDSCs in healthy conditions and tumor setting was assessed by flow cytometry. Functional analysis of Siglec-E knockout on MDSCs in mice was evaluated using SigE-LysMCre mice and suppressive capacity was tested in vitro in combination with SiglecE blocking and cleavage of Sialoglycans using Sialidase. Results were confirmed in the human setting using an in vitro assay to generate MDSC-like cells including RNA-Sequencing and a MDSC suppression assay with cancer-derived MDSCs.

Results: MDSCs express multiple inhibitory Siglec receptors in humans and mice. Knockout of SiglecE in the myeloid compartment in vivo resulted in prolonged survival and increased T cell infiltration compared to litter mates expressing Siglec-E. Of note, desialylation of MDSCs and Siglec-E blocking in vitro reduced MDSCs suppressive function against T cells. Similarly, reducing Sialoglycans by Sialidase in human MDSC generated in vitro resulted in increased suppressive capacity and downregulation of functional MDSC markers on RNA level. Findings were validated with lung cancer patient derived MDSCs.

Conclusion: Our results provide first insights into the importance of the Siglec-Sialoglycans axis to modulate MDSCs in the TME. Targeting glycosylation of MDSCs with Sialidase reduces their suppressive capacity making it a powerful approach to improve cancer immunotherapy. Further studies are needed to reveal the underlying mechanisms.

Citation Format: Ronja Wieboldt, Andreas Zingg, Emanuele Carlini, Anastasiya Börsch, Heinz Läubli, Natalia Rodrigues Manutano. Disturbing the Siglec-Sialoglycan axis to target myeloid- derived suppressor cells in the tumor microenvironment [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 1259.
Abstract 1348: Neutrophil pro-tumorigenic role in progression and therapy resistance in pancreatic ductal adenocarcinoma
Authors: Sturgeon Reegan , Goel Paran , Molczyk Caitlin , Batra Surindar K. , Singh Rakesh
Published: April 2023

Pancreatic cancer (PC) continues to be a challenge for today’s therapeutics, with a meager 5-year survival rate of 11%. Due to the pancreas’ location, patients often receive diagnosis late, leaving them with limited treatment options. Often, they fail to respond to chemotherapy regimens or develop resistance to those therapies. Tumor-promoting inflammation is a hallmark of cancer that contributes to tumor cells’ survival and proliferation. Infiltrating leukocytes and pro-inflammatory cytokines released into the tumor microenvironment (TME) often cause this inflammation. Previously we have determined that inflammation and CXCR2 ligand expression increases with pancreatic ductal adenocarcinoma (PDAC) progression. In the present study, we examined the role of neutrophil recruitment in PDAC progression using Kras-dependent KC and KPC murine models. Our data suggest a positive association between tumor-associated neutrophils (TANs) and PDAC progression. Furthermore, the frequency of neutrophil extracellular traps (NETs) also increased as the disease progressed. In vitro, we observed PDAC cell condition-media enhanced survival of neutrophils in a concentration-dependent manner. Moreover, increased survival was associated with disease aggressiveness and therapy resistance. Additionally, we observed that PDAC-conditioned media modulated the expression of immunoregulatory cytokines and chemokines in neutrophils. Together, our data suggest that increased recruitment of TANs supports PDAC progression, and PDAC cell-neutrophil interaction modulates neutrophil survival and immunoregulatory phenotype.

Citation Format: Reegan Sturgeon, Paran Goel, Caitlin Molczyk, Surindar K. Batra, Rakesh Singh. Neutrophil pro-tumorigenic role in progression and therapy resistance in pancreatic ductal adenocarcinoma [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 1348.
Abstract 1363: Breast pericytes: A newly identified driver of tumor cell proliferation
Authors: Toro Katelyn Del , Licon-Munoz Yamhilette , Hines William C.
Published: April 2023

A crucial problem limiting our progress to a cure for breast cancer is understanding what drives tumor growth. While we are aware that potent regulators of tumor development include non-malignant cells coexisting in tumors, not all cell types are easily recognized—nor have they been examined rigorously as potential contributors to malignancy. An obstacle in approaching this problem has been an inability to isolate and purify each cell type from the tissue. Recently our laboratory developed models of each cell type in the breast, allowing us the opportunity to investigate the contributions of primary cell types to tumor cell growth through co-culture. We have discovered that pericytes secrete a potent factor that drives tumor cell expansion directly— and dramatically more than controls (isogenic fibroblasts). The effects of this factor are dose-dependent, and we find that it is protease- and heat-inactivated. Furthermore, tissue immunostaining reveals that pericytes expand into tumor stroma, where they may interact directly with tumor cells. Single-cell RNA sequencing uncovers the heterogeneity of pericytes, which exist on a perivascular spectrum.

Citation Format: Katelyn Del Toro, Yamhilette Licon-Munoz, William C. Hines. Breast pericytes: A newly identified driver of tumor cell proliferation [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 1363.
Abstract 1366: HMGB1 localization and its effect on the immune response in the lung tumor microenvironment
Authors: Jafri Kisa , Andre Nicole M. , Simmons Glenn E.
Published: April 2023

Lung adenocarcinoma is the leading cause of cancer mortality. High Mobility Group Box 1 (HMGB1) is a master regulator of innate immunity that is elevated in lung cancer tissues. Furthermore, HMGB1 has been previously demonstrated to suppress functional anti-tumor immune responses in this context. However, beyond demonstrating that HMGB1 is elevated and modulates inflammation, studies to date have not comprehensively investigated the influence of HMGB1 on the immune response to lung cancer. HMGB1 contributes to PD-L1 expression in melanoma and promotes secretion of inhibitory cytokines TGF-α and IL-10, which have roles in the differentiation of regulatory T cells. Therefore, HMGB1 sits atop an immunomodulatory signaling cascade in the tumor microenvironment. While treatment of lung cancers with immune checkpoint inhibitors (ICI) (anti-PD-1/PD-L1) has had success, there are limitations. One major limitation is the failure of patients with advanced or refractory disease to generate a strong anti-tumor immune response. Recently, we have identified HMGB1 as a protein that is functionally dependent on monounsaturated fatty acid (MUFA) availability. Utilizing a combination of molecular assays and 3-dimensional cultures, we now show that HMGB1 is sequestered intracellularly, rather than being secreted extracellularly in a MUFA-dependent fashion. We hypothesized that MUFAs promote an anti-tumor immune response via modulating HMGB1 release from cancer cells. Thus, by restricting extracellular HMGB1 levels, HMGB1-responsive proinflammatory/tumor signaling pathways (e.g., NF-kB) are suppressed. Taken together, these observations suggest a novel approach to decreasing maladaptive immune and inflammatory signaling from innate immune cells. Exploiting the MUFA/HMGB1 signaling axis could be a novel approach to improve the efficacy of therapies in populations that are not currently benefiting from current strategies for eliminating tumors.

Citation Format: Kisa Jafri, Nicole M. Andre, Glenn E. Simmons. HMGB1 localization and its effect on the immune response in the lung tumor microenvironment [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 1366.
Abstract 1416: Genomic features of renal cell carcinoma developed during end-stage renal disease and dialysis
Authors: Maekawa Shigekatsu , Fujita Masahi , Takata Ryo , Ikarashi Daiki , Matsuura Tomohiko , Kato Renpei , Kato Yoichiro , Kanehira Mitsugu , Sugimura Jun , Abe Takaya , Nakagawa Hidewaki , Obara Wataru
Published: April 2023

Introduction: Patients with end-stage renal disease (ESRD) or receiving dialysis have a much higher risk for renal cell carcinoma (RCC), but carcinogenic mechanisms and genomic features remain little explored and undefined. This study’s goal was to identify the genomic features of ESRD RCC and characterize them for associations with tumor histology and dialysis exposure. In this study, we obtained 33 RCCs, with various histological subtypes, that developed in ESRD patients receiving dialysis and performed whole-genome sequencing and transcriptome analyses.

Materials and Methods: Frozen kidney samples were obtained from 48 patients who were undergoing dialysis treatment and developed RCC at Iwate Medical University Hospital. Among them, we selected 38 subjects for study from whom matched tumor and adjacent non-tumor tissue samples could be obtained and from which DNA (n=38) or RNA (n=26) could be isolated. We performed whole genome sequencing of genomic DNA using MGIEasy FS DNA Library Prep Set and short read sequencer DNBSEQ-G400. mRNA Sample were sequenced using HiSeq. Driver events, copy-number alteration (CNA) analysis, and mutational signature profiling was performed using an analysis pipeline that integrated data from germline and somatic SNVs, Indels, and structural variants as well as CNAs, while transcriptome data was analyzed for differentially expressed genes and through gene set enrichment analysis.

Results: ESRD related clear cell RCCs' driver genes and mutations mirrored those in sporadic ccRCCs. Longer dialysis periods significantly correlated with a rare mutational signature SBS23, whose etiology is unknown, and increased mitochondrial copy number. All ACD (acquired cystic disease)-RCCs, which developed specifically in ESRD patients, showed chromosome 16q amplification. Gene expression analysis suggests similarity between certain ACD-RCCs and papillary RCCs and in TCGA papillary RCCs with chromosome 16 gain identified enrichment for genes related to DNA repair, as well as pathways related to reactive oxygen species, oxidative phosphorylation, and targets of Myc.

Conclusion: This analysis suggests that ESRD or dialysis could induce types of cellular stress that impact some specific types of genomic damage leading to oncogenesis.

Citation Format: Shigekatsu Maekawa, Masahi Fujita, Ryo Takata, Daiki Ikarashi, Tomohiko Matsuura, Renpei Kato, Yoichiro Kato, Mitsugu Kanehira, Jun Sugimura, Takaya Abe, Hidewaki Nakagawa, Wataru Obara. Genomic features of renal cell carcinoma developed during end-stage renal disease and dialysis [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 1416.
Abstract 1441: p27Kip1 is upregulated in UMSCV-4 cells corresponding to entering a state of quiescence when treated with clobetasol
Authors: Minnuto Gianna , North Luke , Ogden Mack , Haering Kia , Lewis Jani E.
Published: April 2023

Quiescence, the temporary exit from the cell cycle, presents therapeutic challenges to cancer management since it allows evasion of chemotherapy and radiotherapy treatment. Essential to the study of quiescence in carcinogenesis is an established model system. Our studies have found that clobetasol treatment of the vulvar cancer cell line, UMSCV-4 causes these cells to enter a state of dormancy. Subsequent removal of the cells from clobetasol show a return to normal cell proliferation, even after dormancy for 3 months. p27Kip1 protein has been used as marker for entry into quiescence. Using both antibodies against p27Kip1 and transfection of a modified p27Kip1 containing plasmid into the UMSCV-4 cells we found that p27Kip1 is upregulated by addition of clobetasol, reflecting that quiescence is being activated in the treated UMSCV-4 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 p27Kip1 upon re-exposure to clobetasol. This establishes the UMSCV-4 cells as a good model system for the study of clobetasol induced quiescence in vulvar squamous epithelial cells. The importance of this work is underscored by the observation that 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.

Citation Format: Gianna Minnuto, Luke North, Mack Ogden, Kia Haering, Jani E. Lewis. p27Kip1 is upregulated in UMSCV-4 cells corresponding to entering a state of quiescence when treated with clobetasol [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 1441.
Abstract 149: An organoid model for understanding the pathogenesis of Lynch syndrome
Authors: Guild Caroline , Madden-Hennessey Kirby , Heinen Chris
Published: April 2023

Lynch syndrome (LS) is an inherited predisposition for developing cancer caused by germline variants in mismatch repair (MMR) genes. It is estimated that LS is responsible for 1 out of every 35 colorectal cancers and 1 out of every 56 endometrial cancers. Loss of MMR is also seen in many sporadic cancers. Still, the mechanism of tumorigenesis in these patients is unknown. Loss of MMR increases genomic mutation rate, but it is unclear if increased mutational burden alone leads to tumorigenesis. Recent studies of LS patient colon samples show fields of morphologically normal tissue in which cells have lost expression of an MMR protein, suggesting that loss of MMR in these cells is sufficient to confer an advantage over heterozygous cells still proficient in MMR. However, whether MMR loss provides a direct advantage to cell types associated with cancer in LS has not been directly tested. We hypothesize that loss of MMR is sufficient to confer a selective advantage to colonic stem cells and is the first step towards tumorigenesis in LS patients.

To test this, we use a human colonic organoid model to examine competition between MMR-deficient (dMMR) and MMR-proficient (pMMR) cells over time. The colonic organoids are differentiated from human embryonic stem cells.

To track cell populations, dMMR stem cells are labeled with mCherry and pMMR stem cells are labeled with GFP. We have initially observed that dMMR stem cells are much more efficient at generating colonic organoids. Thus, the cells are mixed so that resulting organoids initially contain a small population of dMMR cells in order to mimic the spontaneous loss of heterozygosity that occurs in LS patients’ colonic crypts. Live fluorescent imaging of individual organoids is used to quantify changes in cell population over time. Preliminary data indicate that dMMR cells can outgrow pMMR cells resulting in a shift in the ratio of green to red cells over a two-week time period. These results suggest that loss of MMR function imparts an immediate selective advantage. Furthermore, this assay could be used to find conditions or compounds that decelerate population shifts with the ultimate goal of finding a way to reduce dMMR fields in LS patients. In these ways, this assay can be leveraged for gaining new insights into MMR function, pathogenesis of LS, and potential therapies for LS.

Citation Format: Caroline Guild, Kirby Madden-Hennessey, Chris Heinen. An organoid model for understanding the pathogenesis of Lynch syndrome [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 149.
Abstract 1494: PBRM1-deficient PBAF complexes target de novo genomic loci to activate NF-κB pathway in clear cell renal cell carcinoma
Authors: Yao Xiaosai , Hong Jing Han , Nargund Amrita , Ng Michelle Shu Wen , Heng Hong Lee , Li Zhimei , Guan Peiyong , Sugiura Masahiro , Chu Pek Lim , Wang Loo Chien , Ye Xiaofen , Yauch Robert , Chang Kenneth Tou-En , Sobota Radoslaw M. , Tan Patrick , Teh Bin Tean
Published: April 2023

PBRM1 is an accessory subunit of the PBAF subclass of the SWI/SNF chromatin remodeler and the inactivation of PBRM1 is the second most frequent mutational event in kidney tumorigenesis. However, the impact of PBRM1 loss on chromatin remodeling, especially pertaining to kidney tumorigenesis, has not been examined previously. Here we show that in VHL-deficient ccRCC tumors, PBRM1 deficiency results in altered PBAF complexes that retain the association between SMARCA4 and ARID2 but disengage BRD7 from SMARCA4. The PBRM1-deficient PBAF complexes redistribute from promoter proximal regions to distal enhancer regions containing motifs of pro-tumorigenic factors including NF-κB. Subsequently, PBRM1-deficient cells display heightened NF-κB activity across multiple cell models. The ATPase function of SMARCA4 maintains chromatin occupancy of both pre-existing and newly acquired RELA specific to PBRM1 loss, and activates downstream target gene expression. Proteasome inhibitor bortezomib reverses NF-κB activation by reducing RELA occupancy and delays growth of PBRM1-deficient tumors. In conclusion, PBRM1 safeguards the chromatin by repressing aberrant liberation of pro-tumorigenic NF-κB target genes by residual PBRM1-deficient PBAF complexes.

Citation Format: Xiaosai Yao, Jing Han Hong, Amrita Nargund, Michelle Shu Wen Ng, Hong Lee Heng, Zhimei Li, Peiyong Guan, Masahiro Sugiura, Pek Lim Chu, Loo Chien Wang, Xiaofen Ye, Robert Yauch, Kenneth Tou-En Chang, Radoslaw M. Sobota, Patrick Tan, Bin Tean Teh. PBRM1-deficient PBAF complexes target de novo genomic loci to activate NF-κB pathway in clear cell renal cell 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 1494.
Abstract 1498: Melanoma antigen gene A6 and androgen receptor interaction in bladder cancer
Authors: Kushwaha Prem Prakash , Verma Shiv , Gupta Sanjay
Published: April 2023

The Melanoma Antigen Gene (MAGE) protein family is a large and highly conserved group of proteins that share a common MAGE homology domain. Interestingly, several MAGE proteins are limited in expression to reproductive tissues but are aberrantly expressed in various types of human malignancies. We have recently demonstrated overexpression of MAGE family member A6 in advance-stage bladder cancer (Cancers 13:23, 5931, 2021). However, the mechanisms involved in MAGE gene overexpression and its coregulators are currently unknown. One hypothesis is that MAGEA6 and androgen receptor (AR) signaling is involved in the etiology of bladder cancer progression. In the present study, we determined the functional relationship between MAGEA6 and AR in bladder cancer. Treatment with DHT, an AR agonist, increased AR and MAGEA6 expression in UMUC3 and J82 bladder cancer cells at the protein and transcript level. On the contrary, exposure of cells to enzalutamide, an antiandrogen, resulted in simultaneous reduction of AR and MAGEA6 more prominently in UMUC3 cells, compared to J82 harboring low MAGEA6 transcript. Dual-sgRNA CRISPR/Cas9 knockout of MAGEA6 in UMUC3 and J82 cells resulted in inhibition of cell proliferation, reduced survival, and migration potential in both cell lines. An induction of S-phase and G0/G1 phase cell cycle arrest was noted in UMUC3 and J82 cells. In silico analysis showed Arg20 and Cys557 amino acid residue of AR interacts with Asp236 and Gln106 amino acid residue of MAGEA6 through hydrogen bonds. Further confirmation of physical interaction between AR and MAGEA6, performed by immunoprecipitation of AR and probed for MAGEA6 and vice versa established interaction of AR-MAGEA6 with each other which was further confirmed by mass spectrometry analysis. In conclusion, these findings confirm the oncogenic role of MAGEA6 and its physical interaction with AR may be involved in bladder cancer progression.

Citation Format: Prem Prakash Kushwaha, Shiv Verma, Sanjay Gupta. Melanoma antigen gene A6 and androgen receptor interaction in bladder 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 1498.
Abstract 151: Optimization of screening conditions using patient-derived organoids for diagnostic purposes
Authors: Verissimo Carla S. , Smabers Lidwien , Wensink Emerens , Koedoot Esmee , Pitsa Katerina-Chara , Doorn Mayke , van Weersch Inez , Warda Hamdy , Barón Celia Higuera , ten Hag Gerben , Vries Robert G. , Roodhart Jeanine , Boj Sylvia F.
Published: April 2023

Patient-derived organoids (PDOs) are increasingly adopted in preclinical research. PDOs, or HUB Organoids®, are directly established from patient tissue, faithfully recapitulate patient disease, and effectively bridge the gap between the lab and the clinic, bringing a “patient in the lab®”. PDOs can also revolutionize clinical research because they can predict patient response to standard-of-care (SoC) compounds. However, there is an urgent need to refine the assay conditions to evaluate the activity of SoC drugs with different mechanisms of action (topoisomerase inhibitors, alkylating agents, targeted agents) and various types of molecules (small molecules, antibodies) and achieve optimal results from the PDO-based assay. Specifically, for metastatic colorectal cancer (mCRC), we have optimized the media composition required for drug screening assays to test response to SoC, such as oxaliplatin and panitumumab. Our results indicate that optimal media conditions are essential to observe an in vitro drug response in PDOs similar to the patient response in the clinic. In addition, an image-based readout was developed that enables precise quantification of organoid numbers ensuring high assay quality. Optimizing screening conditions to evaluate SoC compounds in PDOs is a required step to accelerate the implementation of the HUB Organoid Technology as a diagnostic platform to accurately predict treatment response and improve clinical outcomes for cancer patients.

Citation Format: Carla S. Verissimo, Lidwien Smabers, Emerens Wensink, Esmee Koedoot, Katerina-Chara Pitsa, Mayke Doorn, Inez van Weersch, Hamdy Warda, Celia Higuera Barón, Gerben ten Hag, Robert G. Vries, Jeanine Roodhart, Sylvia F. Boj. Optimization of screening conditions using patient-derived organoids for diagnostic purposes [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 151.
Abstract 1561: Preclinical examination of PLK1 inhibitors for the treatment of diffuse midline gliomas
Authors: Hayden Elisha , Franshaw Laura , Upton Dannielle , Liu Jie , Nguyen Hieu , Joshi Swapna , Tsoli Maria , Dolman Emmy , Rayner Benjamin , Ziegler David
Published: April 2023

Diffuse Midline Gliomas (DMGs) are devastating and incurable pediatric brain tumours. DMGs are characterised by epigenetic aberrations in the form of histone modifications, coupled with mitotic abnormalities, which together drive a stem-like transcriptional cell state that is critically dependent on the cell cycle. Polo-like kinase 1 (PLK1) is a pivotal regulator of mitosis and cell cycle progression. To determine an effective therapeutic strategy against DMG, we characterised the anti-tumour potential of PLK1 inhibition, either alone or in combination with radiotherapy. We show that targeting PLK1 with small molecule inhibitors BI2536 and BI6727 (volasertib) reduced DMG colony formation and cell proliferation, and induced G2/M checkpoint arrest and apoptosis in vitro at physiologically achievable, clinically relevant concentrations. When combined with radiotherapy, PLK1 inhibitors acted as potent radiosensitisers with synergistic suppression of DMG survival. We further show that PLK1 inhibitors exert potent anti-tumour effects in highly aggressive orthotopic DMG models, effectively extending the median survival of tumour harbouring mice both as a single agent and enhanced in combination with radiotherapy. We further demonstrate a potential novel role for PLK1 in defining an oncogenic transcriptional state in DMG by acting as a conduit between cell cycle progression and the DMG epigenome through modulation of polycomb repressive complex 2 (PRC2) activity and its resultant histone methyltransferase function. Together, these data indicate that targeting PLK1 is a highly promising therapeutic strategy to arrest DMG tumorgenicity. This data has formed the basis for a planned international Phase 1/2 trial of the PLK1 inhibitor volasertib in combination with radiotherapy in paediatric patients with DMG.

Citation Format: Elisha Hayden, Laura Franshaw, Dannielle Upton, Jie Liu, Hieu Nguyen, Swapna Joshi, Maria Tsoli, Emmy Dolman, Benjamin Rayner, David Ziegler. Preclinical examination of PLK1 inhibitors for the treatment of diffuse midline gliomas [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 1561.
Abstract 1569: OXC-101 kills cancer by disturbing microtubule polymerization and inhibiting MTH1
Authors: Berglund Ulrika Warpman , Gad Helge , Sanjiv Kumar , Jemth Ann-Sofie , Mortusewics Oliver , Brautigam Lars , Rantala Juha , Helleday Thomas
Published: April 2023

Many cancers suffer from replication stress, oxidative stress, a dysfunctional redox status, and high levels of endogenous oxidative DNA damage. OXC-101 (karonudib, TH1579) targets these vulnerabilities by stopping the cancer cells in mitosis, generating ROS which increases oxidative stress and oxidative DNA damage, thereby killing the cancer cells. The aim of this study was to further investigate the underlying mechanism of OXC-101 and the cancer specific effects in solid cancers. Using e.g. microtubule polymerization assays, co-immunoprecipitation and immunofluorescence microscopy, we show a dual mechanism of action of OXC-101, disturbing microtubule function and inhibiting the nucleotide pool sanitizing enzyme MTH1. Furthermore, we demonstrate the importance of a functional spindle assemble checkpoint (SAC) for the response of OXC-101, since knocking down MAD2,using siRNA, or inhibiting SAC with reversine, results in treatment resistance. We show that the excess of oxidized nucleotides is cytotoxic when inhibiting or knocking out MTH1 in zebrafish embryos, and that pretreatment with ROS scavenger rescues the effect of OXC-101 in cancer cells. These data indicate that the increased incorporation of oxidized nucleotides into DNA is, together with the mitotic arrest, an important part of the mechanism of action and response. OXC-101 has a broad anti-cancer effect. In an ex vivo drug sensitivity study of 157 primary biopsies from solid cancer patients, OXC-101 was efficacious in a large proportion of the patient derived samples, with best responses in endometrial (73%), HNSCC (73%), bladder (70%) and prostate (83%) cancer samples. Biomarker searches for genetic alterations explaining responders/non-responders are presently on-going. In summary, the dual mechanism of action of the mitotic MTH1 inhibitor OXC-101 explains its strong anti-cancer effect.

Citation Format: Ulrika Warpman Berglund, Helge Gad, Kumar Sanjiv, Ann-Sofie Jemth, Oliver Mortusewics, Lars Brautigam, Juha Rantala, Thomas Helleday. OXC-101 kills cancer by disturbing microtubule polymerization and inhibiting MTH1 [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 1569.
Abstract 1595: Thyroid hormone drives the terminal differentiation of tumor cells in medulloblastoma
Authors: Yang Zeng-Jie , Yang Yijun Yang , Heller Allie
Published: April 2023

Hypothyroidism is commonly detected in patients with medulloblastoma (MB), a pediatric brain tumor, which is generally considered as a treatment-related complication. Although reduced levels of thyroid hormone (TH) significantly correlate with poor survival of patients with MB, the possible link between TH signaling and MB pathogenicity is unknown. Here, we found that TH plays a critical role in MB pathogenicity by regulating terminal differentiation of tumor cells. Elimination or reduction in the levels of TH frees the unliganded TH receptor, TRα, to bind to EZH2 and repress expression of NeuroD1, a transcription factor that drives terminal differentiation of neuronal progenitors as well as MB cells. However, increased TH reverses EZH2-mediated repression of NeuroD1 by abrogating the binding of EZH2 and TRα, thereby stimulating terminal differentiation of tumor cells and reducing MB growth. Moreover, TH promotes extensive differentiation and reduced proliferation of tumor cells from multiple molecular subtypes of MB including the hedgehog (HH) group as well as group 3 (G3) MB, indicating that TH-induced differentiation is not restricted by the oncogenic driver mutations in tumor cells. Consequently, TH treatment significantly inhibits the in vivo growth of SHH- and G3-MB by promoting tumor cell differentiation, with no obvious increase in tumor cell death, indicating that TH signaling represents a novel therapeutic entry-point for broad treatment of MB. These findings elucidate the mechanisms underlying terminal differentiation of MB cells, and they establish an unprecedented association between TH signaling and MB progression. Our studies provide compelling evidence for a promising tumor therapeutic modality targeting tumor cell differentiation.

Citation Format: Zeng-Jie Yang, Yijun Yang Yang, Allie Heller. Thyroid hormone drives the terminal differentiation of tumor cells in medulloblastoma [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 1595.
Abstract 1657: Discovery of a first-in-class multifunctional TYMS non-classical antifolate inhibitors with potent in vivo activity that prolongs survival
Authors: Dib Peter Elias , Guijarro Maria , Kellish Patrick , Paciaroni Nick , Nawab Akbar , Andring Jacob , Kulemina Lidia , Leas Derek , Borrero Nick , Modenutti Carlos , Bennett Richard , Licht Jonathan , McKenna Robert , Roitberg Adrian , Kaye Frederic , Zajac-Kaye Maria
Published: April 2023

Thymidylate synthase (TYMS/TS) is overexpressed in many cancer types and is commonly associated with advanced disease. TYMS catalyzes the de novo synthesis reaction of deoxythymidine monophosphate (dTMP), one of the building blocks of DNA. Although TYMS inhibitors have served as components of chemotherapy regimens, the currently available inhibitors induce TYMS overexpression or alter folate transport/metabolism feedback pathways that tumor cells exploit for drug resistance limiting overall benefit. Here we report a small molecule TYMS inhibitor and its two analogues that exhibit i) enhanced anti-tumor activity as compared to current fluoropyrimidines and antifolates without inducing TYMS overexpression, ii) are structurally distinct from classical antifolates, iii) extend survival in both pancreatic xenograft tumor models and hTS/Ink4a/Arf null genetically engineered mouse tumor model, iv) and are well tolerated with equal efficacy using either intraperitoneal or oral administration. Mechanistically, we confirm the compound and its analogues are a multifunctional non-classical antifolate, and we identify structural features allowing direct TYMS inhibition while also maintaining the ability to inhibit dihydrofolate reductase (DHFR). Collectively, this work identifies new non-classical antifolate inhibitors that optimize inhibition of thymidylate biosynthesis with a favorable safety profile highlighting potential for enhanced cancer therapy.

Citation Format: Peter Elias Dib, Maria Guijarro, Patrick Kellish, Nick Paciaroni, Akbar Nawab, Jacob Andring, Lidia Kulemina, Derek Leas, Nick Borrero, Carlos Modenutti, Richard Bennett, Jonathan Licht, Robert McKenna, Adrian Roitberg, Frederic Kaye, Maria Zajac-Kaye. Discovery of a first-in-class multifunctional TYMS non-classical antifolate inhibitors with potent in vivo activity that prolongs survival [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 1657.
Abstract 1681: The alternative mechanism of Src inhibition in colon cancer cells
Authors: Lu Yi-Wen , Hou Xiang-Ling , Lu Yen-Tse , Lin Chia-Wen , Huang Chien-Chi , Chao Wei-Ting
Published: April 2023

In colon cancer, Src showed the high expression in tumor cells and was known as an important migration marker that plays the role in coordinate with many important cellular signal pathways. According to previous studies, cell viability and migration ability can be inhibited by Src inhibitor, Dasatinib. However, the clinical study demonstrated the survival rate has no significant increase in Dasatinib treatment. Therefore, we want to figure out what is the real function of Src in colon cancer cell in this study. HT-29, colon cancer cell with epithelial cell morphology was used in the study. After treated with Dasatinib, cells were followed by reattachment experiment. Then the cell growth, migration ability, signaling pathway transition, molecular localization and expressions were analyzed. The results showed Src inhibition can reduce EGFR activation on cell-cell contacts and increased Akt phosphorylation upon EGF stimulation. We further found Dasatinib has limited effect on cell toxicity, however, can be the selector to separate two cell populations, Dasatinib sensitive (DSTS) and Dasatinib resistance (DSTR) cells, interestingly, the DSTS cells were grown as colony phenotype efficiently and majorly through EGFR/AKT pathway. Meanwhile, DSTR was demonstrated the promotion of collective cell migration ability by EGFR/Ecad pathway. The results of this study suggested Src activity plays novel role in switch EGFR/Akt and EGFR/Ecad pathways in colon cancer cells and harboring different strategies to survive, therefore, the alternative resistance mechanism of Src inhibitor in colon cancer cells may need be concerned in clinical application.

Citation Format: Yi-Wen Lu, Xiang-Ling Hou, Yen-Tse Lu, Chia-Wen Lin, Chien-Chi Huang, Wei-Ting Chao. The alternative mechanism of Src inhibition in colon 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 1681.
Abstract 1683: Ether phospholipids metabolism is a latent vulnerability for pancreatic cancer resistance
Authors: Chen Ziheng , Ho I-Lin , Soeung Melinda , Yen Er-Yen , Rose Johnathon , Srinivasan Sanjana , Deem Angela , Gao Sisi , Ying Haoqiang , Draetta Giulio
Published: April 2023

Mitochondria are hubs where bioenergetics, redox homeostasis, and anabolic metabolism pathways integrate through a tightly coordinated flux of metabolites. The essential contributions of mitochondrial metabolism to fuel tumor growth and resistance to therapy are now evident in multiple contexts. However, targeting of mitochondrial metabolism in the clinical setting has repeatedly failed, owing to a narrow therapeutic window and the remarkable ability of tumor cells to activate compensatory metabolic programs. Thus, it is essential to uncover additional mechanisms that may render cancer cells selectively susceptible to treatment and those that enable them to survive when exposed to mitochondrial metabolism-targeted drugs. Here, using pancreatic ductal adenocarcinoma (PDAC) as a disease model, we report that cellular and, in particular, mitochondrial lipid composition affects the sensitivity of cancer cells to pharmacological inhibition of electron transport chain complex I. Metabolomic, including lipidomic, analyses of patient-derived PDAC models uncovered a critical role for monounsaturated fatty acids (MUFAs) and demonstrated that MUFA-linked ether phospholipids have a critical role to sustain homeostasis of mitochondrial reactive oxygen species (ROS). Blocking de novo ether phospholipid biosynthesis in the peroxisomes, sensitized PDAC cells to mitochondrial complex I inhibition by inducing mitochondrial ROS and lipid peroxidation. Biochemical analysis revealed a role of ether phospholipids in the assembly of mitochondrial supercomplexes and ROS production. Together, our data identify an ether phospholipid-dependent mechanism that regulates mitochondrial redox control and contributes to the sensitivity of PDAC cells to complex I inhibition. These findings might lead to novel approaches to target mitochondrial metabolism in cancer cells.

Citation Format: Ziheng Chen, I-Lin Ho, Melinda Soeung, Er-Yen Yen, Johnathon Rose, Sanjana Srinivasan, Angela Deem, Sisi Gao, Haoqiang Ying, Giulio Draetta. Ether phospholipids metabolism is a latent vulnerability for pancreatic cancer resistance [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 1683.
Abstract 1737: Genome-scale metabolic modeling reveals a metabolic switch that restores sensitivity to anticancer chemotherapy in drug-resistant breast cancer cells
Authors: Lim JinA , Jung Hae Deok , Lee Kyung-min , Jeon Moonhyeon , Ryu Han Suk , Kim Hyun Uk , Kim Yoosik
Published: April 2023

Anticancer chemotherapy is an essential part of cancer treatment, but the emergence of resistance due to continued drug usage remains a major hurdle. Among the multiple mechanisms behind the acquisition of drug resistance, metabolic plasticity is receiving increasing attention. Recent proteomic and transcriptomic studies identified metabolic enzymes as potential targets for overcoming the resistance. Yet, these studies mainly focused on differentially expressed genes and overlooked the complexity of the metabolic network, especially its ability to reorganize the metabolic state in response to gene downregulation. Here, we employ the genome-scale metabolic model (GEM) to simulate the metabolic flow and identify candidates that can act as a metabolic switch for drug resistance. Using adriamycin- and paclitaxel-resistant MCF-7 breast cancer cells as model systems, we perform proteomics analysis followed by GEM construction to analyze the differential metabolic flow in the resistant cells. Moreover, we perform GEM simulations after individually knocking out every metabolic enzyme and identify a metabolic switch whose knockout restores the metabolic state of drug-sensitive MCF-7 cells. Notably, downregulating the candidate metabolic switch predicted by our model results in sensitization to the drug. We further show that the cotreatment of adriamycin or paclitaxel with a small chemical inhibitor of the metabolic switch synergistically induces cell death in drug-resistant cells. Collectively, our study demonstrates the application of GEM simulation and subsequent modulation of metabolic state as a strategy to restore drug sensitivity to overcome drug-resistant cancer.

Citation Format: JinA Lim, Hae Deok Jung, Kyung-min Lee, Moonhyeon Jeon, Han Suk Ryu, Hyun Uk Kim, Yoosik Kim. Genome-scale metabolic modeling reveals a metabolic switch that restores sensitivity to anticancer chemotherapy in drug-resistant breast 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 1737.
Abstract 1748: Mcl-1 inhibitor KS18 overcomes bortezomib, venetoclax, and ABT-737 resistance in multiple myeloma
Authors: Odat Omar S. Al , Gowda Krishne , Amin Shantu G. , Alpdogan Tulin Budak- , Jonnalagadda Subash C. , Pandey Manoj K.
Published: April 2023

Recent advancements in multiple myeloma (MM) therapy have not changed the fact that it is incurable, and all patients experience relapse or become refractory. Thus, more effective treatments are needed to prolong the survival of MM patients. Anti-apoptotic myeloid cell leukemia 1 (Mcl-1) overexpression is correlated with poor prognosis and drug resistance in MM. Nonetheless, there is a lack of effective Mcl-1 inhibitors. To address this unmet clinical need, we developed KS18, a Mcl-1 inhibitor. We generated respective resistant cells to investigate whether KS18 may overcome acquired resistance against various available drugs in the clinic including bortezomib, venetoclax, and ABT-737. KS18 was effective against all MM cells with IC50 values ranging between 1.5 to 2.5µM. KS18 showed significant Mcl-1 inhibition in various MM cells including resistant cells. Importantly, KS18 re-sensitizes resistant cells to bortezomib and other chemotherapeutic drugs. Moreover, KS18 was able to enhance the anti-myeloma activity of venetoclax and ABT-737 treatments toward bortezomib-resistant cells. These results indicate that KS18 possesses the potential to overcome resistance by targeting Mcl-1. Our future studies in patient samples and in vivo model are proposed for further preclinical assessment of this exciting molecule.

Citation Format: Omar S. Al Odat, Krishne Gowda, Shantu G. Amin, Tulin Budak- Alpdogan, Subash C. Jonnalagadda, Manoj K. Pandey. Mcl-1 inhibitor KS18 overcomes bortezomib, venetoclax, and ABT-737 resistance in multiple myeloma [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 1748.