bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 139: Exploiting interactions between ploidy and host physiology
Authors: Andor Noemi
Published: April 2023

Whole-genome doubling facilitates rapid tumor evolution and is a hallmark of many cancers, including Glioblastoma (GBM) where it has an incidence of ~14%. GBM remains uniformly lethal, with a median overall survival of 14-16 months, despite aggressive therapy. In people aged 65 and above, prognosis is even more dismal, with median survival of less than 7 months. While comorbidities explain some of the increased age-related risk, age-related changes in the tumor microenvironment likely also contribute. Aging brains have higher incidence of polyploidy (Pearson r=0.80; P=0.031; Nandakumar et al., 2021), hypoxia related gene expression (Pearson r =0.22, P =0.008) and Glucose levels (Ding et al., 2021) as well as decreased stiffness (Hiscox et al., 2018). The average ploidy of 20 different cancer types correlates with the Oxygen levels recorded in their respective tissue of origin (Spearman r = 0.66, P = 0.002). Together these results suggest that availability of metabolic substrates in the GBM environment drives different cell fate decisions in cancer cells of different ploidy and shapes GBM disease initiation.

We address this hypothesis with integrated in-vitro and in-silico experiments and multi-omic and imaging analysis of primary and recurrent GBM. We genetically engineered cell lines to obtain diploid and tetraploid lineages for each and tested how exposure to variable gradients of glucose, hypoxia and phosphate affect their migratory patterns. We performed RNA-seq and WGS of multiple regions obtained from two patient-matched primary and recurrent GBM. Pre- and post-surgery MRI data was available from both surgeries and registered to a T1 map Atlas to quantify stiffness and oxygenation in the brain. This data was used to calibrate a Stochastic State-Space Model (S3MB) we developed to recapitulate re-growth of enhancing primary tumor margins into the respective recurrent tumor. The model domain is initiated from MRI data and consists of 4mm3 voxels. Each voxel state is defined by 8+ variables that are updated over time: Stiffness, Oxygen, Phosphate, Glucose, Vasculature, Dead cells, migrating cells and proliferating cells of various ploidies.

In-vitro results, multi-omics results and similarity between observed and simulated recurrent tumors were used to infer model parameters (resource consumption-, migration-, death and proliferation rates). Running the model backwards in time, we predict the local point of origin (LPO) of each GBM. We found that resource availability in the predicted LPO was correlated to WGD status, suggesting conditions in the local environment in which a GBM originates shape the outcome of an early competition between diploid and tetraploid cells.

Citation Format: Noemi Andor. Exploiting interactions between ploidy and host physiology [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 139.
Abstract 1397: Collagen COL26A1 promotes papillary thyroid cancer invasion and metastasis and correlates with poor survival and clinical outcomes
Authors: Carnazza Michelle , Quaranto Danielle , DeSouza Nicole , Dadafarin Sina , Moscatello Augustine , Geliebter Jan , Tiwari Raj K.
Published: April 2023

Papillary thyroid cancer (PTC) is the most common cancer in women under 25 years of age and is increasing in incidence. While PTC is often slow-growing and surgically resectable, recurrence years to decades after initial surgery occurs in ~20% of patients (now only in their 40s/50s), with mortality of 38-69%. The identification of both prognostic biomarkers and actionable therapeutic targets for high-risk PTC at diagnosis is critically important and an unmet need. RNASeq analysis of 45 PTC and matched normal patient tissues from our biobank identified collagen 26A1 (COL26A1) as significantly upregulated in patients with higher risk extrathyroidal extension, lymph node metastasis, and multifocal tumors, compared to those with localized disease. Kaplan Meier survival curves of The Cancer Genome Atlas (TCGA) data confirmed that increased COL26A1 expression significantly decreases survival probability by 25% (p=0.0086). Further analyses of TCGA data indicated that COL26A1 expression significantly correlated with clinical attributes including MACIS score (q=0.001), differentiation score (q=0.025), and tumor stage (q=0.025). Data from our in vitro culture PTC model system indicated that COL26A1 expression was increased 3X in PTC cell line K1, compared to immortalized “normal” thyroid epithelial cells, NThy-ori-3-1. Two CRISPR guides designed to target COL26A1 introduced into K1 resulted in repression of COL26A1 RNA and protein levels by 50% and 70%, respectively. COL26A1 repression decreased critical tumor phenotypes of aggressiveness and metastasis including proliferation (30%), clonogenicity (33%), anchorage-independent growth (37%), cell motility (43%), invasion (30%) and migration (33%). Cell-to-cell adhesion and cell-matrix adhesion also decreased (36% and 25%, respectively). Thus, COL26A1 may have use as a prognostic marker and actionable target for small molecule inhibitors in PTC.

Citation Format: Michelle Carnazza, Danielle Quaranto, Nicole DeSouza, Sina Dadafarin, Augustine Moscatello, Jan Geliebter, Raj K. Tiwari. Collagen COL26A1 promotes papillary thyroid cancer invasion and metastasis and correlates with poor survival and clinical outcomes [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 1397.
Abstract 1404: A functional genetic polymorphism of mitochondrion-associated DNAJA3 in nonalcoholic steatohepatitis-related hepatocellular carcinoma
Authors: Shiode Yuto , Chang Ching-Wen , Wang Xin Wei
Published: April 2023

Nonalcoholic fatty liver disease (NAFLD) is the most common form of liver disease worldwide and may progress to nonalcoholic steatohepatitis (NASH). Although emerging evidence suggests that NASH would be an important driver of hepatocellular carcinoma (HCC), molecularly what drives the transition of NAFLD to NASH and then HCC has not been clearly defined. Since NASH is a mitochondrial disease, we aimed to determine genetic variants associated with mitochondrial dysfunction linked to NASH and NASH-related HCC. We performed an integrative genomics analysis by analyzing a large-scale genome-wide association study of the body fat distribution-related variants in 344,369 individuals from multiple ancestries. And we linked these results to the top 24 significant variants to NASH and NASH-related HCC in 1,009 individuals from the NCI-UMD cohort. We also performed the expression quantitative trait loci (eQTL) analysis on candidate variants. We analyzed the 3D genome architecture especially chromatin topology based on UCSC Genome Browser that influences the expression of candidate variants. As a result, we identified rs3747579-TT, an eQTL to a mitochondrial chaperone Hsp40, also known as DNAJA3 rather than a coding variant, as a risk allele linked to NASH-related HCC. HCC patients with rs3747579-TT variant had a poor prognosis compared to HCC patients with rs3747579-CT or CC variant. We confirmed that HCC with rs3747579-TT had a reduced expression of DNAJA3, encoding a protein that maintains mitochondrial homeostasis. Consistently, HCC patients with low DNAJA3 expression had a poor prognosis. We found a specific site based on chromatin topology that may influence rs3747579 to regulate the expression of DNAJA3. In summary, we identified rs3747579-TT as a functional polymorphic variant of DNAJA3 that may be responsible for the development of NASH and NASH-related HCC.

Citation Format: Yuto Shiode, Ching-Wen Chang, Xin Wei Wang. A functional genetic polymorphism of mitochondrion-associated DNAJA3 in nonalcoholic steatohepatitis-related hepatocellular 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 1404.
Abstract 1414: The role of chromosome arm 3p and human papillomavirus on proliferation, differentiation, and metastasis in squamous cancers
Authors: Mallick Samyukta , Taylor Alison M.
Published: April 2023

Squamous Cell Carcinoma (SCC) is an aggressive form of cancer that develops in the mucosal epithelium of the digestive and respiratory tracts as well as on the skin. Two major risk factors for development of SCC are carcinogen exposures, such as alcohol and tobacco, and Human Papillomavirus (HPV) infection. Viral- and carcinogen-driven SCCs have differential responses to radiation, chemotherapy, and immunotherapy, with HPV+ SCCs responding better to these treatments than HPV- carcinomas. Analysis of aneuploidy patterns of genomic samples from The Cancer Genome Atlas (TCGA) have shown that squamous cancers cluster together and are characterized by gain of chromosome arm 3q gain and loss of chromosome arm 3p. However, while HPV+ SCCs show similar rates of 3q gain, they have significantly lower rates of 3p loss, suggesting that 3p deletion and HPV infection might have overlapping roles in cancer initiation and/or progression. In addition, the degree to which these two paths of tumor development contribute to differences in disease progression and outcome has been largely unexplored. We have identified a strong correlation between 3p deletion and high epithelial-mesenchymal transition (EMT) gene signature across all cancer types. Interestingly, interferon signaling has been found to be upregulated in non-squamous cancers upon 3p deletion but downregulated in squamous cancers with 3p deletion. SCCs positive for HPV infection show decreased expression of EMT genes and increased expression of genes in interferon signaling and cell cycle checkpoint pathways. Together, the results of these TCGA analyses suggest mechanisms by which chromosome 3p deletion and HPV infection may affect cancer progression and are currently being validated in vitro in a panel of squamous cancer cell lines. Additionally, we will use a targeted CRISPR-Cas9 system developed in our lab to delete chromosome 3p, as well as transfect HPV oncoproteins, in normal keratinocytes to create isogenic models. With these models, we will study the role of individual aneuploidy events in tumorigenesis, tumor development, and metastasis in the context of HPV positive and negative SCC.

Citation Format: Samyukta Mallick, Alison M. Taylor. The role of chromosome arm 3p and human papillomavirus on proliferation, differentiation, and metastasis in squamous cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1414.
Abstract 1415: The search for molecular biomarkers of relapse in early stage lung adenocarcinoma
Authors: Ke Honggang , Jing Zhongying
Published: April 2023

Background: Recurrence cannot be completely prevented by resection or chemotherapy. Molecular biomarkers reported in previous studies have the potential to predict the recurrence risk of lung adenocarcinoma (LUAD), but the results were controversial.

Methods: We retrospectively analyzed the molecular characteristics of the 218 early-stage patients with lung adenocarcinoma based on 599 gene-panel target sequencing. All the patients received no neoadjuvant therapy and no EGFR-TKI treatment. The follow-up information on recurrence was complete. Univariate and multivariate analyses were performed using the Cox proportional hazard models to investigate whether the common alterations in EGFR and KRAS were prognostic factors for relapse-free survival(RFS).

Results: Among all the enrolled patients, the mutation rates of EGFR and KRAS were 47.6% and 9.3%, respectively. EGFR mutation was not proved to be a prognostics signature of relapse for lung adenocarcinoma regardless univariate and multivariate analysis (univariate: HR, 1.11; 95%CI, 0.82-1.50, P = 0.49; multivariate: HR, 1.18; 95%CI, 0.86-1.64, P = 0.31). The univariate and multivariate analysis identified KRAS alteration as an independent prognostic factor for a shorter RFS (univariate: HR, 1.64; 95%CI, 0.75-2.43, P < 0.05; multivariate: HR, 1.66; 95%CI, 1.19-2.32, P <0.05). For patients in pathological stage I, the patients with KRAS alterations showed significant recurrent risk compared with wild-type patients (HR, 2.02; 95%CI, 1.38-2.96; P <0.0.01).

Conclusion: KRAS alterations rather than EGFR alterations were prognostic biomarkers of relapse for early-stage lung adenocarcinoma, especially for patients in pathological stage I.

Citation Format: Honggang Ke, Zhongying Jing. The search for molecular biomarkers of relapse in early stage lung 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 1415.
Abstract 1420: The non-canonical role of Rb in enhancers and insulators
Authors: Lee Hanjun , Gkotinakou Ioanna M. , Sanidas Ioannis , Lawrence Michael S. , Dyson Nicholas J.
Published: April 2023

Despite the great significance of the tumor suppressor Rb in cancer biology, the full spectrum of its binding pattern in human cells was yet to be established. Here, we report that rather than exclusively associating with E2F-bound promoters, chromatin-bound Rb associates with three distinct types of loci—E2F1-bound promoters, AP-1-bound enhancers, and CTCF-bound insulators. The binding pattern of Rb correlated with cohesin binding and chromatin accessibility in all three types of Rb-bound regions. When cells entered the S phase after the G1/S checkpoint, Rb redistributed from promoters to enhancers, leading to distinct transcriptional changes. These Rb-bound enhancers were cell-type-specific, in contrast to Rb-bound promoters that were constitutively present throughout most human cell types. Overall, our study provides a comprehensive view on the role of the tumor suppressor Rb, which has a well-preserved role in the G1 phase of regulating transcription at E2F1-bound promoters, while having a cell-type-specific role in the S phase when cells are determined to replicate.

Citation Format: Hanjun Lee, Ioanna M. Gkotinakou, Ioannis Sanidas, Michael S. Lawrence, Nicholas J. Dyson. The non-canonical role of Rb in enhancers and insulators [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 1420.
Abstract 1423: Regulation of SKP2 expression by methyltransferase-independent function of EZH2 in tumor growth and progression
Authors: Colon Tania Marlyn , Dai Wei , Choi Byeong Hyeok
Published: April 2023

Skp2, an F box protein and a substrate-binding component of the SCFSkp2 ubiquitin E3 ligase complex, promotes ubiquitination and degradation of cyclin-dependent kinase inhibitors (CKIs), including p21, p27, and p57. Aberrant Skp2 expression is associated with many human cancers. EZH2 (Enhancer of Zeste Homologue 2) is a core catalytic subunit of the polycomb repressive complex 2 (PRC2) that can regulate gene expression by mediating the methylation of lysine 27 of histone H3 (H3K27me3). Accumulating evidence shows that EZH2 could bind to transcription factors and activate gene expression independently of its methyltransferase activity, leading to the activation of oncogenic signaling pathways and promoting the proliferation of tumor cells. Although the methyltransferase-independent function of EZH2 is important in cell proliferation, its mechanism in cell cycle regulation is still unclear. In this study, we demonstrate that EZH2 positively regulates Skp2 gene expression. We show that suppression of EZH2 expression resulted in a significant decrease of Skp2 protein levels, while the EZH2 enzymatic inhibitors, EPZ6438, had no impact on protein levels of Skp2, which suggests that EZH2 regulates Skp2 expression independently of its methyltransferase activity. Consistently, EZH2 depletion reduced the mRNA levels of Skp2, while EPZ6438 inhibited EZH2 methyltransferase activity without affecting the mRNA levels of Skp2. Importantly, EZH2 expression is positively correlated with the expression of EMT (Epithelial-mesenchymal transition) regulators (YAP1/TAZ, ZEB1, and Snail) and immune checkpoint proteins (FGL1 and Siglec-15) in cervical and pancreatic cancer cells, but reversely correlated with the expression of CKIs (p21, p27, and p57). Taken together, our findings suggest a novel mechanism by which EZH2 regulates tumorigenesis through its methyltransferase-independent function by regulating Skp2 expression. Targeting the methyltransferase-independent function of EZH2 may have therapeutic efficacy for cancer patients.

Citation Format: Tania Marlyn Colon, Wei Dai, Byeong Hyeok Choi. Regulation of SKP2 expression by methyltransferase-independent function of EZH2 in tumor growth and progression [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 1423.
Abstract 1448: The YAP1/TEAD and NF-Kappa B transcription factors may cooperate to mediate prostate cancer progression
Authors: Cinar Bekir , Al-Mathkour Marwah M. , Dwead Abdulrahman M. , Boston Ava M. , Lewis Michael S. , Moreno Carlos S.
Published: April 2023

Metastatic castration-resistant prostate cancer is morbid and lethal. The transcriptional coregulator YAP1 (Yes-associated protein 1) is a nuclear effector of the Hippo pathway. The Hippo pathway regulates cell growth, motility, and carcinogenesis. Here, we investigated biochemical and functional interactions between YAP1 and the nuclear factor (NF) kappa B/RELA subunit in prostate cancer cell models. We demonstrated that endogenous YAP1 and RELA form protein complexes in the cell, as revealed by co-immunoprecipitation and western blotting. Compared with control, we found that combined treatment of cells with androgen and SDF1a (stromal cell-derived factor-1 alpha) or RANKL (receptor activator of NF-kappa Β ligand) enhanced the protein-protein interaction between YAP1 and RELA, as showed by proximity ligation assay. Our confocal microscopy experiment further showed that combined SDF1a and androgen treatment augmented the YAP1 and RELA colocalization relative to single-agent. Moreover, our promoter-reporter and RNAi experiments showed that genetic silencing YAP1 or TEAD protein, a key mediator of the YAP1 transcription, significantly reduced the NF-Kappa B promoter-reporter gene activity. Also, disruption of YAP1 activity by genetic and small molecule attenuated endogenous protein-protein interaction between TEAD and RELA. Furthermore, controlled expression of MST1/STK4, a potent inhibitor of YAP1, suppressed the NF-Kappa B-promoter reporter activity. Additionally, our unbiased bioinformatics analysis of the exiting ChIP-seq (chromatin immunoprecipitation-sequencing) data identified several genes likely co-regulated by the TEAD1-4 and NF-Kappa B transcription factors. Besides, our computational analysis of the TCGA and Stand Up To Cancer (SU2C) prostate cancer data sets indicated that expression of YAP/TEAD and NF-kappa B/RELA correlates in clinical cases. These findings suggest that cooperative androgen and cytokine signaling regulates Hippo/YAP and NF-Kappa B interaction and their functions. These may have critical roles in advanced prostate cancer downstream of the Hippo pathway.

Citation Format: Bekir Cinar, Marwah M. Al-Mathkour, Abdulrahman M. Dwead, Ava M. Boston, Michael S. Lewis, Carlos S. Moreno. The YAP1/TEAD and NF-Kappa B transcription factors may cooperate to mediate prostate cancer progression [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 1448.
Abstract 1460: Co-expression network analysis reveals male breast cancer specific transcriptional module
Authors: Gou Xuxu , Wang Junkai , Lei Jonathan T. , Jaehnig Eric J. , Anurag Meenakshi
Published: April 2023

Introduction: According to the American Cancer Society, An estimated 2,710 men are diagnosed with breast cancer annually of which about 530 die due to the disease annually in the United States. Most male breast tumors are estrogen receptor positive. Risk factors include age, radiation exposure, BRCA1/2 gene mutations, Klinefelter syndrome, testicular disorders, diabetes, gynecomastia (enlarged breast) etc. Due to low testing and incidence rate and lack of bioinformatics approaches focused on identification of drivers of rare cancer, little is known about this disease in men than in women. Because of this knowledge-gap there is no therapy that is specifically tailored for male breast cancer, leading to nineteen percent higher mortality in male than female with breast cancer.

Method: In the past, gene expression analysis has revealed differential pathways in male than in female breast cancer, but these studies compared male and female tumors without considering innate (sex-specific) differences in the mammary tissues between the two sexes. To overcome these challenges, we have utilized publicly available tumor (E-TABM-810, TCGA-Breast) and normal (GTEX - mammary) datasets to identify pathways and gene co-expression networks enriched in specific transcription factor (TF) targets that are specific to male breast tumors . Findings from this analysis is further validated in male breast tumor patient derived xenograft (PDX) model.

Result: Using differential pathway analysis corrected by sex, Type I diabetes mellitus is the only Hallmark pathway signature that is significantly downregulated in male tumors compared to both female tumors and normal mammary tissue. No pathway is distinctively up-regulated in male tumors. Co-expression network analysis of gene expression datasets identified unique TF-driven networks that are specific to male breast tumors. This analysis identified a network of ELK1 and DMRT1 TF targets that are unregulated and downregulated in male breast tumors respectively. This was further validated in PDX dataset.

Conclusion: Based on our results, we hypothesize that loss of DMRT1 TF activity is potentially associated with occurrence of male breast tumors. The role of DMRT1 in sex differentiation and gonadal differentiation has been explored, but its role in male breast tumor formation is not as well characterized. Thus, our analysis provides novel insight in transcription factors associated with incidence of this rare form of cancer in men.

Citation Format: Xuxu Gou, Junkai Wang, Jonathan T. Lei, Eric J. Jaehnig, Meenakshi Anurag. Co-expression network analysis reveals male breast cancer specific transcriptional module [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 1460.
Abstract 1490: Exon exclusion of NELF-E produces an anti-tumorigenic truncated isoform in hepatocellular carcinoma
Authors: Reynolds Laura , Lamm Ryan , Barry Anna , Grim Meghan , Zhang Kai , Dang Hien
Published: April 2023

Overexpression of negative elongation factor E (NELF-E), a subunit of the negative elongation factor (NELF) complex involved in transcription pausing, has been implicated in poor hepatocellular carcinoma (HCC) outcome. We recently discovered an intronic heterozygous single nucleotide polymorphism (SNP), rs79208225 (G>A), near a spliceosome binding site in less than 1% of the general population but ~4% of our HCC cohort. HCC patients with the SNP had significantly longer times till recurrence (p=0.001) and better overall survival (p=0.02) compared to patients without the SNP. Using the minigene assay, we found the SNP causes exon 10 exclusion, which resulted in lower levels of full length NELF-E and the presence of a truncated NELF-E isoform that was not degraded via RNAi surveillance. We hypothesize NELF-E isoform switching is anti-tumorigenic in HCC. To investigate the effects of exon 10 exclusion, we employed antisense oligonucleotides (ASOs). HCC cells treated with ASOs had decreased cell proliferation and colony formation. Moreover, exon 10 exclusion enriched for the truncated NELF-E isoform and decreased full length mRNA and protein expression. Genetic models (CRISPR/cas9) showed similar phenotypes, suggesting that the truncated NELF-E is anti-tumorigenic. Currently, we are investigating the functional role of the truncated NELF-E protein in transcription regulation and the effects of isoform switching in HCC cells as a potential therapeutic strategy for HCCs with elevated NELF-E levels.

Citation Format: Laura Reynolds, Ryan Lamm, Anna Barry, Meghan Grim, Kai Zhang, Hien Dang. Exon exclusion of NELF-E produces an anti-tumorigenic truncated isoform in hepatocellular 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 1490.
Abstract 1492: Interaction between Argonaute-2 and Negative Elongation Factor-E in hepatocellular carcinoma
Authors: Ruiz Brittany Natalie , Lucci Alvaro , Barry Anna , Zhang Kai , Dang Hien
Published: April 2023

Hepatocellular carcinoma (HCC) continues to be one of the leading cause of cancer-related deaths globally, whose incidence is on the rise in the United States. We and others have shown that high levels of AGO2 is associated with a worse prognosis in HCC patients. Specifically, we showed AGO2 promotes HCC progression by modulating MYC mRNA stability in an RNAi-independent manner. In addition to its canonical role as an RNA binding protein, AGO2 has been demonstrated to regulate gene expression by affecting RNA polymerase II (Pol II) pausing through the Negative Elongation Factor (NELF) complex in Drosophila. It remains elusive how AGO2 modulates the chromatin to affect transcription. Here, we hypothesize AGO2/NELF interactions modulate chromatin accessibility to promote HCC. Immunoprecipitation assays were employed to detect protein-protein interactions between AGO2 and the NELF complex in HCC cells. To determine the effects of AGO2 RNAi-silencing on their interaction, DICER-knockout (via CRISPR) HCC cell lines were also probed for protein-protein interactions. We found AGO2 and NELFE, a member of the NELF complex, interact in both wild-type and DICER-knockout HCC cell lines. These data suggest AGO2 may play an important role in modulating transcription through NELF. Currently, we are investigating the potential interplay between AGO2 and NELFE in modulating chromatin accessibility to promote hepatocarcinogenesis. Understanding the effects of the AGO2/NELF interaction may help elucidate transcriptional regulation in HCC and lead to better therapies for HCC patients.

Citation Format: Brittany Natalie Ruiz, Alvaro Lucci, Anna Barry, Kai Zhang, Hien Dang. Interaction between Argonaute-2 and Negative Elongation Factor-E in hepatocellular 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 1492.
Abstract 1493: ROS inhibits enzymatic activation of PRMT5 and increases the protein stability of tumor suppressors as non-histone substrates in liver cancer
Authors: Lee Ji Min , Kim Hyunkyung , Im Hyuntae , Baek Hee-ji
Published: April 2023

Retinoic acid receptor-related orphan receptor α (RORα) and p53 are transcription factors involved in nuclear gene expression and known tumor suppressors. RORα was the first identified substrate of lysine methylation-dependent degradation. However, the mechanisms of other post-translational modifications (PTMs) that occur in RORα and p53 remain largely unknown, especially in liver cancer. Arginine methylation is a common PTM in arginine residues of nonhistone and histone proteins and affects substrate protein function and fate. We found an analogous amino acid disposition containing R37 at the ROR N-terminus compared to histone H3 residue, which is arginine methylated. Here, we provide evidence that R37 methylation-dependent degradation is carried out by protein arginine methyltransferase 5 (PRMT5). In addition, we discovered that PRMT5 regulated the interaction between the E3 ubiquitin ligase ITCH and RORα through RORα arginine methylation. Arginine methylation-dependent ubiquitination-mediated RORα degradation reduced downstream target gene activation. H2O2-induced reactive oxygen species (ROS) decreased PRMT5 protein levels, consequently increasing RORα protein levels in liver cancer in vitro and in vivo. Furthermore, the enzymatic activities of PRMT5 were directly affected by ROS via the dissociation of PRMT5-Mep50 octamer in the methylosome complex. Taken together, ROS inhibited liver cancer progression by inducing apoptosis via PRMT5-mediated methylation of RORα combined with other histone and non-histone substrates and the ITCH axis. Our results potentiate PRMT5 as an elimination target in cancer therapy, and this additional regulatory level within ROS signaling may help identify new targets for therapeutic intervention in liver cancer.

Citation Format: Ji Min Lee, Hyunkyung Kim, Hyuntae Im, Hee-ji Baek. ROS inhibits enzymatic activation of PRMT5 and increases the protein stability of tumor suppressors as non-histone substrates in liver 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 1493.
Abstract 157: Patient in the lab: Down-scaling patient-derived organoid screening for diagnostic purposes
Authors: Koedoot Esmee , van Weersch Inez , Harada Gakuro , Watanabe Masahiko , Warda Hamdy , Kyan Hideaki , Abouleila Yasmine , Kumagai Takahiko , Hikichi Yuichi , Overmeer René , Roodhart Jeanine , Matsuno Kiyotaka , Verissimo Carla , Boj Sylvia F.
Published: April 2023

Colorectal cancer (CRC) is the third leading cause of cancer and cancer-related death. A large fraction of the CRC patients diagnosed with de novo metastatic disease do not benefit from standard of care and experience substantial side effects. Therefore, there is an urgent need for preclinical models that help predict patient response in the clinic. Patient-derived organoids (PDOs or HUB Organoids®) represent a significant breakthrough as preclinical models as they are directly established from patient tissue and faithfully recapitulate patient disease. HUB Organoid Technology can already be applied to preclinical drug screening, however, to provide direct patient benefit the turnaround time between patient diagnosis and PDO-based results must be shortened. In collaboration with Yamaha Motor, the Yamaha CELL HANDLERTM has been validated to automatically pick-and-place with high accuracy a significantly reduced number of organoids per screening plate, compared to standard procedures. In addition, an image-based readout was developed that enables a precise quantification of organoid number ensuring high assay quality. Organoid responses to chemotherapy and targeted agents were validated using this newly automated system, confirming comparable PDO-patient drug sensitivity profiles, and known drug responses based on genetic dependencies. In summary we describe the development of an automated workflow that combines patient-representative HUB Organoids and state-of-the-art robotics by Yamaha CELL HANDLER. This workflow down-scales the number of organoids needed per screening well and allows to efficiently predict patient response, thereby reducing the diagnostic turnaround time and increasing patient benefits.

Citation Format: Esmee Koedoot, Inez van Weersch, Gakuro Harada, Masahiko Watanabe, Hamdy Warda, Hideaki Kyan, Yasmine Abouleila, Takahiko Kumagai, Yuichi Hikichi, René Overmeer, Jeanine Roodhart, Kiyotaka Matsuno, Carla Verissimo, Sylvia F. Boj. Patient in the lab: Down-scaling patient-derived organoid screening 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 157.
Abstract 1571: The feasibility and pharmacological mechanism of cucurbitacin E for the treatment of EGFR-TKI-primary resistant NSCLC
Authors: Chen Chun-Han
Published: April 2023

Lung cancer ranks the leading cause of cancer death, with an estimated 1.8 million deaths according to the latest global cancer statistics. In the United States, there will be an estimated 236,740 new cases diagnosed, and 130,180 people will die from lung cancer in 2022. In non-small-cell lung cancer (NSCLC) patients, targeted therapies are available for patients with EGFR exon 19 deletion and L858R mutations. However, tumors harboring wild-type EGFR or Kras mutation are not eligible for targeted therapies. Therefore, the development of new drugs to treat primary resistant NSCLC remains urgent needs. Cucurbitacin E (CuE) is a tetracyclic triterpenoid compound extracted from the family of Cucurbitaceae. The data showed that CuE exhibits a significant antiproliferative effect with a GI50 value of 0.03±0.01 μM. Cell cycle analysis revealed that CuE induces G2/M arrest and down-regulates cell cycle regulatory proteins in A549 cells. CuE treatment further increased the expression of cleaved caspase-3/8/9/PARP, and altered the expression of DNA-damaged-related biomarkers. Meanwhile, protein levels of LC3BII and p62, and ROS production were upregulated by CuE. Pharmacological Inhibition of ATM reversed the effects of CuE on autophagy-related proteins. In summary, CuE is a potential drug for the treatment of primary resistant NSCLC.

Citation Format: Chun-Han Chen. The feasibility and pharmacological mechanism of cucurbitacin E for the treatment of EGFR-TKI-primary resistant NSCLC [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 1571.
Abstract 1573: Inhibition of acetyl-CoA carboxylase induced spindle abnormalities
Authors: Fang Chieh-Ting , Kuo Hsiao-Hui , Liu Sih-Long , Yao Jhong-Syuan , Yih Ling-Huei
Published: April 2023

Acetyl-CoA carboxylase (ACC), the rate-limiting enzyme in the de novo fatty acid synthesis pathway, is required for successful cell division in yeast, indicating that fatty acid synthesis may be regulated in order to properly safeguard nuclear and cell division. ACC is also phosphorylated during mitosis and this inhibitory phospho-ACC is accumulated at the spindle pole at metaphases. In this study, we set out to understand the role of ACC in the regulation of mitotic spindle assembly in human cells. We found that inhibition of ACC with TOFA induced spindle defects and cell death. The spindle defects induced by TOFA could be rescued by exogenous palmitic acid (PA), the immediate products of fatty acid synthesis. Inhibiting protein palmitoylating or depalmitoylating enzymes could induced spindle defects. These data indicate that well-regulated cellular PA levels and protein palmitoylation are required to assure accurate spindle assembly. In addition, the spindle microtubules comprised of α-tubulins mutated at the reported palmitoylation site exhibited disrupted dynamic instability. These results reveal that ACC may modulate tubulin palmitoylation during mitosis to delicately control MT dynamic instability and spindle assembly, thereby safeguarding nuclear and cell division.

Citation Format: Chieh-Ting Fang, Hsiao-Hui Kuo, Sih-Long Liu, Jhong-Syuan Yao, Ling-Huei Yih. Inhibition of acetyl-CoA carboxylase induced spindle abnormalities [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 1573.
Abstract 1582: Inhibition of autophagy by CAGE targeting peptide retard tumor growth in non-small cell lung cancer of EGFR TKI resistance
Authors: Shin Jung-Young , Kim Min-Young , Lee Mi-Ran , Lee Hankyu , Jeon Doyong , Baek Seung Ryel , Il Kim Joo , Koh Jaemoon , Jeoung Dooil , Kang Jin Hyoung
Published: April 2023

Epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI) is the standard of care as 1st-line treatment for NSCLC patients with activating EGFR mutations (exon 19 deletion or L858R mutation). However, EGFR-TKI therapy eventually leads to acquired drug resistance approximately 1-2 years from the beginning of treatment. Multiple acquired resistance mechanisms including on-target EGFR mutations, phenotype transformation, and activation of bypass signaling molecules have been identified, but over 35% still remains unknown. CAGE, a cancer/testis antigen, was originally isolated from the sera of patients with gastric cancer and is known to regulate the autophagic flux in acquired resistance to EGFR TKI in NSCLC. Herein, the expression of CAGE was found in more than 50% of NSCLC patients using immunohistochemistry (IHC). Interestingly, the expression level was increased especially for the patients acquiring resistance to EGFR-TKIs. In addition, both PC-9/ER (Erlotinib-resistance cells) and H1975/OR (Osimertinib-resistant cells) cells demonstrated increased CAGE expression and autophagic flux, when compared with their parental cells. Thus, we developed the peptide drugs targeting the DEAD box domain of CAGE. This novel peptide effectively inhibited tumor cell proliferation in EGFR-TKI resistant cell line. Moreover, in Erlotinib-resistant patient-derived xenograft (PDX) mice, the peptide drug retarded tumor growth and enhanced anti-tumor efficacy. Collectively, our data suggest that CAGE plays a crucial role in the tumor progression and acquired resistance of EGFR TKI by the modulation of autophagic flux. Furthermore, we provided evidence that the inhibition of autophagy through CAGE- derived peptide could overcome the acquired resistance to EGFR TKIs in advanced/metastatic NSCLC patients.

Citation Format: Jung-Young Shin, Min-Young Kim, Mi-Ran Lee, Hankyu Lee, Doyong Jeon, Seung Ryel Baek, Kim Joo Il, Jaemoon Koh, Dooil Jeoung, Jin Hyoung Kang. Inhibition of autophagy by CAGE targeting peptide retard tumor growth in non-small cell lung cancer of EGFR TKI 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 1582.
Abstract 1599: Targeting tumor synthetic vulnerabilities and immune evasion mechanisms with CO-1002
Authors: Ciznadija Daniel , Degaga Gemechis , Mondal Abdul , Hillbert Maxwell , Chen Cong , Debnath Bikash , Mehrotra Sonam , Jones Brandon , Rivero Ralph , Komurov Kakajan
Published: April 2023

Cancer immunotherapy with immune checkpoint inhibitors (ICI) has shown durable clinical benefits in patients with immune-hot tumors. However, the majority of cancers are characterized by an immune-cold tumor microenvironment, which are refractory to ICI therapy. Targeting of tumor synthetic vulnerabilities due to genomic alterations is another promising approach that has proven effective in the clinic (e.g. PARP inhibitors in BRCA-mutant tumors). Here, we aimed to identify therapeutic targets that have dual liabilities in tumor cell survival and the evasion of anti-tumor innate immune responses. Applying machine learning approaches to a manually curated compendium of public and proprietary datasets, we identified and validated several genes with a high potential of conferring tumor-intrinsic cell survival and blocking of innate pro-inflammatory responses in immune-cold tumors. The targeting of these genes induced robust cell killing and activation of innate immune response pathways in specific molecularly-defined tumor populations, including immune-cold cells. A series of highly potent small molecule compounds were established against these targets using structure-based design, which shows robust in vitro and in vivo target engagement and favorable cytotoxic and phenotypic profiles across a large panel of cancer cell lines and PDXs. Collectively, this therapeutic program shows significant promise in treating a multitude of solid tumors as a monotherapy and in combination with currently approved immune therapies.

Citation Format: Daniel Ciznadija, Gemechis Degaga, Abdul Mondal, Maxwell Hillbert, Cong Chen, Bikash Debnath, Sonam Mehrotra, Brandon Jones, Ralph Rivero, Kakajan Komurov. Targeting tumor synthetic vulnerabilities and immune evasion mechanisms with CO-1002 [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 1599.
Abstract 1602: Co-targeting mitochondria and nucleotide metabolism in T315I-BCR-ABL myeloid leukemia for therapy
Authors: Huang Chang-Yu , Chung Yin-Hsuan , Wu Sheng-Yang , Wang Hsin-Yang , Lin Chih-Yu , Yang Tsung-Jung , Fang Jim-Ming , Chen Yu-Ju , Hu Chun-Mei , Chang Zee-Fen
Published: April 2023

T315I mutation of Bcr-Abl in chronic myeloid leukemia (CML) leads to therapeutic resistance. It is known that Bcr-Abl transformation causes ROS-induced DNA damage, which can be exploited for anti-nucleotide treatment. We developed a small compound, JMF4073, which inhibits pyrimidylate kinases. Intriguingly, treatment of JMF4073 selectively eliminated WT-, but not T315I-, Bcr-Abl-transformed cells. We found that the higher level of cellular glutathione(GSH) in T315I-Bcr-Abl cells confers replication fork progression and sustains dTTP pool, leading to JMF4073 insusceptibility. Blocking mitochondrial pyruvate carrier (MPC) by UK-5099 reduced GSH and induced DNA replication stress in T315I-Bcr-Abl cells, thus increasing JMF4073 sensitivity in vitro and in vivo. We also observed the increases in glutamine flux to GSH production in T315I-Bcr-Abl cells, and reductive carboxylation of glutamate metabolism, together with mitochondrial dysfunction. The proteomic assay revealed the deficiencies in respiration complex I, II, and IV proteins in T315I-Bcr-Abl cells. Most interestingly, calcineurin inhibition remarkably elevated mitochondrial oxidative stress to cause cytotoxicity specifically in T315I-Bcr-Abl cells. Our results demonstrate the metabolic shift by T315I mutation of Bcr-Abl CML and provide the therapeutic options by co-targeting mitochondria and pyrimidylate kinases.

Citation Format: Chang-Yu Huang, Yin-Hsuan Chung, Sheng-Yang Wu, Hsin-Yang Wang, Chih-Yu Lin, Tsung-Jung Yang, Jim-Ming Fang, Yu-Ju Chen, Chun-Mei Hu, Zee-Fen Chang. Co-targeting mitochondria and nucleotide metabolism in T315I-BCR-ABL myeloid leukemia for therapy [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 1602.
Abstract 1649: Chemically induced chaperone-mediated protein degradation (CHAMP) of KRAS(G12C)
Authors: Foley Kevin P. , Ying Chenghao , Wang Yaya , Dai Yan , Wang Zhiyong , Li Jinhua , Yang Zimo , Sun Yuetong , Zhou Hao Xin , Prince Thomas L. , Wang Guoqiang , Ying Weiwen
Published: April 2023

Covalent KRAS(G12C) inhibitors have recently shown promising efficacy in the clinic. However, the rapid development of drug resistance compromises their long-term benefits, indicating the need for additional approaches to treat KRAS-mutated cancers. Targeted protein degradation represents one such potential approach. We have therefore explored applying Chaperone-mediated Protein Degradation (CHAMP) technology to degrade KRAS(G12C) protein. CHAMPs are heterobifunctional small molecules that mediate protein degradation by inducing proximity between a target protein and the HSP90 chaperone complex, resulting in target ubiquitination and degradation by the proteasome. Due HSP90 being highly activated in cancer cells, CHAMPs also preferentially accumulate in tumors relative to normal tissues, resulting in an improved therapeutic index relative to typical inhibitors. In order to chemically induce KRAS(G12C) degradation, CHAMP compounds were synthesized by covalently coupling KRAS(G12C)- and HSP90-binding moieties through a short linker. CHAMP treatment of KRAS(G12C)-mutated cell lines resulted in formation of a KRAS(G12C)-CHAMP-HSP90 ternary complex and subsequent proteasome-dependent KRAS(G12C) degradation and inhibition of cell proliferation. In contrast, combination treatment with KRAS(G12C) and HSP90 inhibitors did not result in substantial KRAS(G12C) degradation. Importantly, CHAMPs retained potency in cell line models of resistance to covalent KRAS(G12C) inhibitors. Further, in mouse xenograft models, CHAMPs displayed prolonged pharmacokinetics in tumors relative to plasma and normal tissues and strongly inhibited tumor growth at tolerated doses. Based on these findings, KRAS(G12C)-degrading CHAMPs represents a promising approach to the treatment of KRAS(G12C)-mutated cancers.

Citation Format: Kevin P. Foley, Chenghao Ying, Yaya Wang, Yan Dai, Zhiyong Wang, Jinhua Li, Zimo Yang, Yuetong Sun, Hao Xin Zhou, Thomas L. Prince, Guoqiang Wang, Weiwen Ying. Chemically induced chaperone-mediated protein degradation (CHAMP) of KRAS(G12C) [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 1649.