bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3773: Mitochondrial fusion is required to support higher rates of respiration in proliferating cancer cells
Authors: Wang Yahui , Yao Conghui , Stancliffe Ethan , Fowle-Grider Ronald , Wang Rencheng , Wang Cheng , Schwaiger-Haber Michaela , Shriver Leah P. , Weber Jason , Patti Gary J.
Published: June 2022

Proliferating cancer cells exhibit an elevated rate of glucose consumption and lactate excretion, a phenomenon described as the “Warburg effect”. Although increased glycolytic flux in proliferating cells is well established, metabolic changes associated with mitochondria still remain elusive. Here, using various inducible quiescence models and mass spectrometry-based metabolomics, we directly compared mitochondrial respiration and TCA cycle activity from the same cells in the proliferative and quiescent states. We found that mitochondrial respiration and TCA cycle flux is significantly increased during proliferation in both transformed and non-transformed cell lines. This increase was supported by mitochondrial fusion rather than mitochondrial biogenesis. Notably, as cells transitioned from a quiescent state to a proliferative state, the elongation of mitochondria occurred as quickly as 3 hours. Our results reveal that the Warburg effect, which was originally associated with dysfunctional mitochondria, does not necessitate decreased mitochondrial respiration. Rather, in many proliferating cancer cells, mitochondrial respiration is actually elevated, underscoring the importance of mitochondrial respiration during proliferation.

Citation Format: Yahui Wang, Conghui Yao, Ethan Stancliffe, Ronald Fowle-Grider, Rencheng Wang, Cheng Wang, Michaela Schwaiger-Haber, Leah P. Shriver, Jason Weber, Gary J. Patti. Mitochondrial fusion is required to support higher rates of respiration in proliferating cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3773.
Abstract 38: Racial disparities in prostate cancer mortality rates explained by differences in dietary and lifestyle factors
Authors: Rencsok Emily , Sodipo Michelle , Gerke Travis , Stopsack Konrad , Mucci Lorelei
Published: June 2022

Introduction: Black men have 2.1 times higher prostate cancer mortality rates than White men in the United States, yet the reasons for this disparity remain unclear. Several dietary and lifestyle factors may influence the risk of developing lethal prostate cancer. This study evaluated to what extent differences in the prevalence of modifiable risk factors by race could explain racial disparities in prostate cancer mortality.

Methods: Our study utilized data from the Health Professionals Follow-up Study (HPFS, N=51,529 men, 1986-2016) and the National Health and Nutrition Examination Study (NHANES, 2000-2010). First, we used data from NHANES to estimate the prevalence of three factors among Black and White men: smoking, vitamin D, and coffee. Then, using HPFS, we generated relative risk estimates for lethal prostate cancer for each factor. To estimate the relative prostate cancer burden associated with these risk factors, we calculated the population attributable fraction (PAF) for each factor by self-identified race, defined as the reduction in mortality that would be achieved if the population had been entirely unexposed, compared with the current exposure pattern. We then calculated the difference in the PAF between Black and white men to estimate the difference in mortality reduction between the two groups if all men had been entirely unexposed, assuming causality of risk factors and no effect modification by race.

Results: There were notable differences in the prevalence of several risk factors by race. For example, Black men had significantly lower levels of vitamin D[MLA1], were less likely to drink coffee, and had a higher prevalence of smoking compared to white men. We estimated that eliminating these risk factors among Black men could lead to a reduction in prostate cancer mortality that is 4-14% larger compared to eliminating these risk factors among white men.

Conclusions: Modifiable lifestyle and dietary factors are potentially responsible for a proportion of the racial disparity in prostate cancer mortality. Our future work will expand the modifiable factors as well as incorporating more contemporary comparisons of the prevalence of these factors to predict future disparities in prostate cancer.

Citation Format: Emily Rencsok, Michelle Sodipo, Travis Gerke, Konrad Stopsack, Lorelei Mucci. Racial disparities in prostate cancer mortality rates explained by differences in dietary and lifestyle factors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 38.
Abstract 3808: Spatially resolved transcriptomics of cellular architecture in EGFR-mutated lung cancer
Authors: Kerr Daniel L. , Wu Wei , Tamaki Whitney , Urisman Anatoly , Chou Yu-Ting , Gui Philippe , Jablons David M. , Bivona Trever G. , Blakely Collin M.
Published: June 2022

Introduction: Single-cell RNA sequencing of dissociated tumors enables the profiling of cellular states in fine detail, but erases the cellular organization of the analyzed tissue. Spatially resolved transcriptomics (SRT) using 10X Genomics’ Visium platform combines histological staining and RNA sequencing by capturing each transcript across spatially barcoded microarrays. SRT yields gene-expression matrices resolved within 55-micron array spots, and creates opportunities to explore how lung biology is affected by lung adenocarcinoma and how tumor cells and the proximal microenvironment are modulated by targeted therapy.

Methods: SRT reactions (n=8) were performed on surgical specimens from human lung adenocarcinomas driven by kinase domain mutations of EGFR (exon 19 deletion, L858R point mutation, exon 20 insertion). SRT reactions analyzed primary lung cancer (n=5) or paired tumor-adjacent lung tissues (n=3).

Results: 28458 array spots were recorded across all samples, and array spots captured a median of 5800 total transcripts and a median of 2416 unique transcripts. Single marker gene expression and unsupervised clustering across array spots corresponded with histological annotations of lung structures and adenocarcinoma. Integration of single-cell RNA sequencing data from lung adenocarcinoma specimens permitted mapping and quantification of 15 major cell types, including cancer cells, T- and B- lymphocytes, macrophages, dendritic cells, fibroblasts, and endothelial cells. Compared with paired tumor-adjacent lung tissues, adenocarcinoma tissues contained fibroblast-enriched desmoplasia and B-cell enriched tertiary lymphoid structures. In a clinical case with resistance to Osimertinib, the standard tyrosine kinase inhibitor, gene signature analysis of cancer-containing array spots revealed enrichment of gap-junction, fatty acid metabolism, and kynurenine pathway signatures compared to treatment naïve array spots.

Conclusion: This pilot study demonstrates the feasibility of using SRT in lung adenocarcinoma. Paired with a single-cell atlas of lung adenocarcinoma during targeted therapy, this approach enables hypothesis-generation to investigate the alterations of tumor and tumor microenvironmental architecture in relation to targeted therapy.

Citation Format: Daniel L. Kerr, Wei Wu, Whitney Tamaki, Anatoly Urisman, Yu-Ting Chou, Philippe Gui, David M. Jablons, Trever G. Bivona, Collin M. Blakely. Spatially resolved transcriptomics of cellular architecture in EGFR-mutated lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3808.
Abstract 386: Non-intrusive, non-invasive and patient friendly focused ultrasound device as a delivery vehicle for CNS tumors
Authors: Ramamurthy Bhaskar , Keralapura Mallika , Marshall Jack , Need Dan , Dittamore Ryan , Kesari Santosh , Fonkem Ekokobe
Published: June 2022

A major impediment to treatment of brain cancers is the inability to transport drugs across the blood-brain barrier (BBB). The development of an effective, targeted, and non-invasive method to penetrate the BBB to deliver cancer therapeutics is an unmet need in the treatment of brain cancers. Large molecular weight chemo- and immuno-therapies such as doxorubicin and others may be potentially effective against brain cancers if such drugs are sufficiently bioavailable in the brain. Focused ultrasound techniques can safely and transiently open the BBB but current techniques require invasive/intrusive or expensive and high-touch procedures and are not optimal for wide adoption. To address this unmet need, we are developing an innovative technique to non-intrusively, non-invasively and transiently open the BBB in a specified location within the brain using guided ultrasound (US). Our device consists of a proprietary US generator that is controlled by a highly portable system that has a small physical footprint, enabling the US generator and system to be placed in confined spaces such as chemotherapy infusion centers. The US generator is cap-shaped device that is placed on a patient’s head. It includes multiple sets of ultrasound transducers that are distributed within the cap according to the anatomy of the skull. With our proprietary technique, we calculate the position of the cap in relation to the internal anatomy in a real-time manner with a non-intrusive and a non-invasive manner. We have recently concluded an extensive study that resulted in algorithms that can accurately guide the US to various targets within the brain across a spectrum of patients. We also have completed a pilot preclinical study on a large animal demonstrating our ability to open the BBB non-invasively and deposit a drug proxy (gadolinium and Evans Blue).

Citation Format: Bhaskar Ramamurthy, Mallika Keralapura, Jack Marshall, Dan Need, Ryan Dittamore, Santosh Kesari, Ekokobe Fonkem. Non-intrusive, non-invasive and patient friendly focused ultrasound device as a delivery vehicle for CNS tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 386.
Abstract 3864: Cytometric analysis of immune cell populations in clinical tumor biopsy tissue microarrays for immuno-oncology
Authors: Villani Tom , Patel Jigar , Collins Kim , Tunstall Kate Lillard , Krisanits Bradley
Published: June 2022

Over the past decade, a new focus on the interactions between the immune system and the development and progression of cancer has led to a wide array of research into medical treatments which leverage activation or modulation of the immune system to aid in treatment. Research in the field of immuno-oncology and elucidation of the biological mechanisms by which cancer cells suppress immunological function has led to new biological treatments for cancer, such as pembrolizumab, which have shown great clinical success since their advent. Population-level analysis of immune cell populations is critical for understanding the immune-component of disease progression, and flow cytometry is a commonly utilized technique for this purpose. However, flow cytometry requires dissolution of tissue specimens, and is incompatible with formalin-fixed paraffin-embedded (FFPE) samples, the gold standard in tissue archives. In order to allow for cytometric analysis of FFPE tissues, we have developed a digital pathology technique for the quantitative assessment of immune-cell populations within tissue microarrays prepared from archived FFPE clinical tissue specimens from patient biopsies. The technique involves a combination of HALO® image analysis using the Highplex FL, Spatial Analysis, and TMA modules, and custom scripting to develop imaging cytometry scatterplots. Bioinformatics techniques were utilized to assess features correlated with tumor progression from the quantitative data extracted from the tissue microarrays.

Citation Format: Tom Villani, Jigar Patel, Kim Collins, Kate Lillard Tunstall, Bradley Krisanits. Cytometric analysis of immune cell populations in clinical tumor biopsy tissue microarrays for immuno-oncology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3864.
Abstract 3877: Deep ultrahigh-plex spatial phenotyping of human cancer tissues
Authors: Jhaveri Niyati , Nikulina Nadya , Zong Hailing , Ma Ning , Cheikh Bassem Ben , Pratapa Aditya , Kassim Yasmin , Anand Bhaskar , Prater Michael , Basu Subham , Braubach Oliver
Published: June 2022

Spatial phenotyping is poised to revolutionize cancer research and treatment. Technologies like CO-Detection by indEXing (CODEX), which enable the detection of dozens of protein biomarkers in situ, have already transformed immuno-oncology and are expected to play formative roles in future research. To further support the implementation of CODEX in cancer research, we present a comprehensive 100 antibody panel that is aimed at in-depth identification of immune cell lineages, activation states, immune checkpoints, tissue structure, cellular signaling, apoptosis, DNA damage and cellular metabolism. Deployment of this panel is possible at high-throughput on the CODEX platform, which is a fully-integrated spatial biology solution that provides cyclical multiplexed imaging at unprecedented resolution and rapid turn-around time. The key features of this novel spatial biology system include: 1) true single-cell and sub-cellular resolution, 2) unparalleled biomarker imaging depth on whole FFPE samples, 3) wide-ranging compatibility with whole slide FFPE tissues, 4) panel customization and flexibility via dedicated barcodes, 5) uncompromised access to whole tissues, and 6) sophisticated single-cell analytical metrics. In this study, we generate a comprehensive account of the spatial biology in different tumor tissues including lung cancer, colon cancer, and melanoma using the CODEX instrument. Single-cell spatial phenotyping coupled with deep bioinformatic analyses reveals new quantitative phenotypic and spatial information that will be essential to gain further insights into distinct cellular neighborhoods and their role in regulating immune and metabolic functions of certain cancers.

Citation Format: Niyati Jhaveri, Nadya Nikulina, Hailing Zong, Ning Ma, Bassem Ben Cheikh, Aditya Pratapa, Yasmin Kassim, Bhaskar Anand, Michael Prater, Subham Basu, Oliver Braubach. Deep ultrahigh-plex spatial phenotyping of human cancer tissues [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3877.
Abstract 3882: Translating genotype to immunophenotype in HRAS mutated head and neck squamous cell carcinoma (HNSCC) to identify effective Tipifarnib partners for optimal patient outcomes
Authors: Psyrri Amanda , Anastasiou Miriam , Spathis Aris , Doumas Stergios , Economopoulou Panagiota , Kotsantis Ioannis , Pantazopoulos Anastasios , Kyriazoglou Anastasios , Kavourakis George , Rampias Theodoros , Foukas Periklis
Published: June 2022

Background: Tipifarnib is a highly selective farnesyltransferase inhibitor that has shown promising activity in HRAS mutant recurrent/metastatic HNSCC. The incidence of HRAS mutations in HNSCC is 4-10%. Since HRAS mutated HNSCC can be successfully targeted with Tipifarnib, we sought to analyze the immune and genomic landscape of these tumors to guide combinatorial strategies.

Methods: We identified thirteen patients with HRAS mutant tumors. We included in the analysis 50 patients with HRAS wild-type tumors. Genomic DNA from baseline biopsies of tumors was purified and subjected to targeted sequencing (TruSightTM Oncology 500 panel). PD-L1 expression in formalin-fixed tumor samples was assessed using the PD-L1 IHC 22C3 pharmDx assay (Agilent Technologies, Carpinteria, CA, USA) and characterized by the combined positive score (CPS). Assessment of PD-L1+ cells inside and at the periphery of the tumor was performed. HRAS mutational status was correlated with PD-L1 status. The genomic data derived from the targeted gene panel of HRAS mutant tumors were compared to those with wild-type HRAS.

Results: Thirty-seven patients had both genomic and PD-L1 expression data available. The presence of HRAS mutations was associated with higher TPS PD-L1 scores (26% vs 40% vs 100% in TPS <1%, 1-49% and >50% respectively, p=0.043). Similar results were obtained for CPS (28% vs 80% in CPS <10% and =>10%, p=0.047). HRAS mutations were more commonly found with TP53 mutations (16% in WT TP53 and 55.6% in mutated TP53, p=0.073). Higher PD-L1 TPS showed increased survival without reaching significance when measuring either in the margins (64.1 vs 35.8, p=0.066) or the center of the tumor (p=0.097) due to the small sample size.

Conclusions: HRAS mutant HNSCC are associated with high PD-L1 expression and co-occurrence of p53 mutations. Combinations of tipifarnib with an anti-PD1 antibody, an anti-PDL1 antibody, or an anti-CTLA-4 antibody warrant clinical investigation in this rare subset of HNSCC.

Citation Format: Amanda Psyrri, Miriam Anastasiou, Aris Spathis, Stergios Doumas, Panagiota Economopoulou, Ioannis Kotsantis, Anastasios Pantazopoulos, Anastasios Kyriazoglou, George Kavourakis, Theodoros Rampias, Periklis Foukas. Translating genotype to immunophenotype in HRAS mutated head and neck squamous cell carcinoma (HNSCC) to identify effective Tipifarnib partners for optimal patient outcomes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3882.
Abstract 3894: WNT5B in osteosarcoma stem cells
Authors: Perkins Rachel S. , Suthon Sarocha , Miranda-Carboni Gustavo A. , Krum Susan A.
Published: June 2022

Osteosarcoma is a rare but deadly pediatric bone cancer. As it stands, there is no targeted therapy available for osteosarcoma. Therefore, patients are left with chemotherapy and surgical resection as their only treatment options. Further, osteosarcoma frequently metastasizes to the lungs and patients with metastatic disease have a dismal overall survival rate of 20% due to the lack of targeted therapy. Therefore, the goal of this study is to understand the role of the WNT5B signaling pathway in osteosarcoma, as it could be a potential therapeutic target. Using RNA sequencing from publicly available data sets and immunohistochemistry on tumor microarrays, we show that WNT5B is overexpressed in a subset of osteosarcoma patients. In these high expressing patients, WNT5B’s overexpression correlates to metastasis and worse overall survival. Due to the significant increase in WNT5B expression between primary tumors and metastasis, we began looking at the stem cell population of osteosarcoma cells. By selecting for the osteosarcoma stem cells using spheroid assays, we show that both protein and mRNA levels of WNT5B are enhanced in the stem cell population compared to the standard adherent cell population. Then, we looked at stemness markers and we show that WNT5B regulates the stemness gene SOX2. We then performed limiting dilution sphere assays to assess the role of WNT5B in sphere forming. We reveal that WNT5B is directly responsible for the sphere forming efficiency, as evidenced by a 50% reduction in spheres formed between 143B control and 143B WNT5B knockdown cells. Additionally, WNT5B drives proliferation and migration of osteosarcoma cancer stem cells as there is significantly decreased sphere size and migratory distance in WNT5B knockdown cells compared to control cells, and there is significantly increased sphere size and migration with the addition of recombinant WNT5B. Through revealing a novel role for WNT5B in osteosarcoma cancer stem cells, we present the WNT5B pathway as a candidate for therapeutically targeting the clinical implications of cancer stem cells, that is, the development of chemoresistance, metastasis and relapse.

Citation Format: Rachel S. Perkins, Sarocha Suthon, Gustavo A. Miranda-Carboni, Susan A. Krum. WNT5B in osteosarcoma stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3894.
Abstract 3900: Β3 adrenergic receptor blockade impairs compensatory mechanism to overcome nutrient deprivation in malignant rhabdoid tumor of the kidney
Authors: Subbiani Angela , Boaretto Alessia , Buonvicino Daniela , Cecchi Cecilia , Favre Claudio , Calvani Maura
Published: June 2022

In this study, the effects of nutrient deprivation in regulating tumor growth and metabolism in Malignant Rhabdoid Tumor of the Kidney (MRTK) was investigated. MRTK is a rare and highly aggressive tumor occurring in infancy and early childhood. Unfortunately, there are not many studies to date that can help us better understand the involvement of metabolism in MRTK growth and progression. However, it is now well known that nutrient deprivation is an environmental stress factor that can influence the behavior of cancer cells leading to reprogramming in metabolism. Here, we first evaluated the effects on cell survival and metabolic adaptation of MRTK G-401 cells after glutamine or tyrosine/phenylalanine deprivation, or both. Then, the involvement of the β3-adrenergic receptor (β3-AR) in regulating the metabolic changes in G-401 cells exposed to limited conditions was investigated. We demonstrated that deprivation of glutamine alone or in combination with tyrosine/phenylalanine enhanced the expression of the glucose transporter GLUT1, associated with increased glucose uptake, ATP production, and oxidative stress, as a compensatory mechanism following amino acid deprivation partially regulated via β3-AR. Indeed, β3-AR antagonism was able to decrease the expression of GLUT1, glucose uptake and survival in G-401 cells. Furthermore, exposure of G-401 cells to limited conditions induced an improved activity of the TCA cycle enzymes leading to a different and enhanced fumarate production. Overall, our results indicate that tyrosine/phenylalanine and glutamine deprivation in MRTK is able to trigger a compensatory mechanism to sustain tumor growth and proliferation.

Citation Format: Angela Subbiani, Alessia Boaretto, Daniela Buonvicino, Cecilia Cecchi, Claudio Favre, Maura Calvani. Β3 adrenergic receptor blockade impairs compensatory mechanism to overcome nutrient deprivation in malignant rhabdoid tumor of the kidney [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3900.
Abstract 3902: TRIM33 interacts with HSF1 in osteosarcoma development
Authors: Lallier Morgane , Tefaye Robel , Moukengué Brice , Charrier Céline , Thepault Rose-Anne , Royer Benedicte Brounais-Le , Baud'huin Marc , Verrecchia Franck , Ory Benjamin , Lamoureux François
Published: June 2022

Osteosarcoma (OS) is the most common primary malignant bone tumor in oncopediatrics. Despite therapeutic advances, the 5-year survival rate is 30% in bad responders to treatment or when pulmonary metastases are present at diagnosis. It is therefore urgent to better understand the molecular mechanisms involved in the development of OS in order to identify new therapeutic targets. As a major player in the stress response, it has recently been shown that the transcription factor HSF1 coordonates a stress-independent transcriptional program involving it in many pro-tumor biological processes in different cancers. A better understanding of the mechanisms by which HSF1 activity participates in tumor development is therefore essential to consider its therapeutic targeting. The co-partners of HSF1 to DNA, by modulating its transcriptional activity, could explain its pleiotropic effects. Based on this hypothesis, TRIM33 was identified by RIME. TRIM33 is known as a transcriptional cofactor and its involvement in various cancers. In this context, our objective is to study the interaction between TRIM33 and HSF1, their common genomic localization and to investigate the involvement of TRIM33 in the development of OS. Our results show that TRIM33 and HSF1 co-localize in the nucleus in OS TMA and by immuno-fluorescence in tumor cells and preclinical models. Furthermore, we observed by chIP-seq that TRIM33 and HSF1 share common DNA binding sites. Finally, our preliminary results show that TRIM33 invalidation by RNA interference in OS cell lines decreases tumor cell proliferation. Thus, in view of its role as a DNA partner of HSF1, TRIM33 is an interesting therapeutic target in OS.

This work was supported by la Fédération Enfants et Santé, la SFCE et La ligue Contre le Cancer (comités 44, 49)

Citation Format: Morgane Lallier, Robel Tefaye, Brice Moukengué, Céline Charrier, Rose-Anne Thepault, Benedicte Brounais-Le Royer, Marc Baud'huin, Franck Verrecchia, Benjamin Ory, François Lamoureux. TRIM33 interacts with HSF1 in osteosarcoma development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3902.
Abstract 3935: The polymeric fluoropyrimidine CF10 overcomes limitations of 5-FU in pancreatic ductal adenocarcinoma cells through increased replication stress
Authors: DiNiro Roberto , Haber Alex O. , Jeong Kang J. , Park Soon Y. , Mills Gordon B. , Gmeiner William H. , Brody Jonathan R.
Published: June 2022

Pancreatic ductal adenocarcinoma (PDA) is a lethal disease that will soon become the second leading cause of cancer deaths in the US. Beside surgery, current therapies have narrow clinical benefits for this devastating disease, and in the majority of cases, only improve survival by a few months. Moreover, systemic toxicities are a harsh reality of these treatments. FOLFIRINOX is the drug regimen of choice, one component of which is 5-Fluorouracil (5-FU) which causes serious gastrointestinal and hematopoietic toxicities and is vulnerable to resistance due, in part, to thymidylate synthase (TS) overexpression. The Gmeiner lab has pioneered the development of polymeric fluoropyrimidines, named F10 and CF10, which unlike 5-FU, are in principle completely converted to the TS inhibitory metabolite FdUMP, without generating appreciable levels of ribonucleotides that cause systemic toxicities and while displaying much stronger anti-cancer activity. Here, we further confirm the potency of CF10 and investigate enhancement of its efficacy through combination with inhibitors targeting replication stress, a hallmark of PDA cells. We show that, consistent with our previous studies in PDA cells, CF10 is much more potent as a single agent than 5-FU, by an average 308x fold, and was effective in the nM range (GI50 range 3.13 - 336 nM) in 5 PDA cell lines tested. We also found that CF10 induces increased replication stress as assessed by phosphorylation of ATR, which appears as early as 8 hours after treatment and increases in intensity over 48 hours, consistent with the kinetics of FdU being released from CF10 and incorporated into DNA, a process requiring several hours. Importantly, phosphorylation of ATR induced by CF10 was significantly higher than with 5-FU. This is especially striking because compounds for this assay were used at their IC50 concentrations, with for example MiaPaCA2 cells being exposed to 899 time more 5-FU than CF10, yet at this concentration 5-FU induced less ATR phosphorylation. Further, we find that the activity of CF10, but not 5-FU can be enhanced through combination with inhibitors of ATR and Wee1 that regulate the S and G2 damage checkpoints. Our results indicate CF10 has potential to supersede the established benefit of 5-FU in PDA treatment and indicate novel combination approaches that may be beneficial compared to well-established regimens used currently for 5-FU.

Citation Format: Roberto DiNiro, Alex O. Haber, Kang J. Jeong, Soon Y. Park, Gordon B. Mills, William H. Gmeiner, Jonathan R. Brody. The polymeric fluoropyrimidine CF10 overcomes limitations of 5-FU in pancreatic ductal adenocarcinoma cells through increased replication stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3935.
Abstract 3939: Preclinical development of embryonic ectoderm development (EED) inhibitor APG-5918/EEDi-5273 for cancer therapy
Authors: Zhu Saijie , Fang Douglas D. , Li Qiang , Yang Dongmei , Gu Shoulai , Wang Shaomeng , Yang Dajun , Zhai Yifan
Published: June 2022

Three core components constitute the polycomb repressive complex 2 (PRC2), a multiprotein complex that catalyzes methylation of histone H3 at lysine 27 (H3K27): (1) enhancer of zeste homolog 2 (EZH2), (2) EED, and (3) suppressor of zeste 12 protein homolog (SUZ12). Dysregulated function of PRC2 is implicated in the development of various cancer types. With regulatory approval of EZH2 inhibitor tazemetostat, a potentially effective successful cancer therapeutic strategy is now available for patients with epitheloid sarcoma and relapsed or refractory follicular lymphoma. However, EZH2 inhibitor activity might be reduced because of acquired resistance through secondary mutations in EZH2 or its inability to inhibit paralogs of EZH2 (including EZH1). Because binding of EED with trimethylated H3K27 (H3K27me3) is required to activate methyltransferase activity of EZH2, allosterically targeting EED is emerging as a novel approach to inhibit PRC2 (Qi et al, Nat Chem Biol 2017). APG-5918/EEDi-5273 is a well-documented, investigational, novel, bioactive, and potent EED inhibitor (Rej et al, JMC 2021). The aim of this study was to further characterize the anticancer therapeutic activity of APG-5918 in the preclinical setting.

APG-5918/EEDi-5273 showed high-affinity binding to EED protein in a biochemical assay (IC50 = 1.2 nM). It also exerted potent antiproliferative activity in several EZH2 mutant (EZH2mut) diffuse large B-cell lymphoma (DLBCL) cell lines, with IC50 values in the nanomolar range and inhibitory effects that were approximately 5 times lower than those with tazemetostat. In a mouse xenograft model derived from EZH2mut KARPAS-422 DLBCL cells, single-agent APG-5918 conferred potent and dose-dependent antitumor activity, resulting in durable complete tumor regression that correlated with inhibition of H3K27me3, induction of PRC2 target genes, and drug exposure in tumors. In summary, our results provide scientific rationale for the clinical development of APG-5918/EEDi-5273 in EZH2mut lymphomas and, potentially, other hematologic malignancies and solid tumors.

Citation Format: Saijie Zhu, Douglas D. Fang, Qiang Li, Dongmei Yang, Shoulai Gu, Shaomeng Wang, Dajun Yang, Yifan Zhai. Preclinical development of embryonic ectoderm development (EED) inhibitor APG-5918/EEDi-5273 for cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3939.
Abstract 3952: A novel Mcl-1 inhibitor induces cells death in a caspase-dependent manner and increases the efficacies of Venetoclax and ABT-737 in multiple myeloma cells
Authors: Al-Odat Omar S. , Tripathi Rahul S. , Srivastava Sandeep K. , Gowda Krishne , Amin Shantu G. , Budak-Alpdogan Tulin , Jonnalagadda Subash C. , Pandey Manoj K.
Published: June 2022

Multiple myeloma (MM) is a deadly malignancy of the blood, characterized by the uncontrolled proliferation of plasma cells. MM is challenging to diagnose and treat, accounting for approximately 12% of hematologic malignancies. The overexpression of anti-apoptotic proteins, particularly Myeloid cell leukemia 1 (Mcl-1), play a significant role in the pathogenesis of MM. The overexpression of Mcl-1 is associated with acquired resistance and poor prognosis. Thus, inhibition of the Mcl-1 protein is an attractive therapeutic strategy against myeloma cells. We synthesized and developed a novel Mcl-1 inhibitor, KS18. The preclinical testing of this novel agent is investigated in MM cell lines. Through docking studies, we predicted that KS18 interacts with Mcl-1. KS18 down-regulates the expression of Mcl-1 in MM cells in a dose- and time-dependent manner. Suppression of Mcl-1 expression by KS18 correlated with activation of caspases, PARP- cleavage, and apoptosis. However, this molecule had no effect on the expression of Bcl-2 and Bcl-xL proteins. Importantly, KS18 markedly enhances the efficacies of Venetoclax and ABT-737 in MM cells. Our results propose that targeting Mcl-1 by KS18 may represent a new viable strategy for MM treatment.

Citation Format: Omar S. Al-Odat, Rahul S. Tripathi, Sandeep K. Srivastava, Krishne Gowda, Shantu G. Amin, Tulin Budak-Alpdogan, Subash C. Jonnalagadda, Manoj K. Pandey. A novel Mcl-1 inhibitor induces cells death in a caspase-dependent manner and increases the efficacies of Venetoclax and ABT-737 in multiple myeloma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3952.
Abstract 3969: Y842 mutations in the activation loop of FLT3 regulates drug response in FLT3-ITD positive acute myeloid leukemia
Authors: Al Ashiri Lina , Moharram Sausan , Purohit Rituraj , Uddin Julhash Kazi , Rönnstrand Lars
Published: June 2022

Acute myeloid leukemia (AML) is an aggressive form of blood cancer with a poor prognosis. Approximately 30% of AML patients carry mutations in the type III receptor tyrosine kinase FLT3. The most common form of FLT3 mutations includes internal tandem duplication (ITD) mutations, which are also associated with poor clinical outcomes. Several inhibitors of FLT3 have been developed and are under clinical trials. Major problems in targeting FLT3-ITD include acquired resistance to the inhibitors which is mediated by secondary point mutations in FLT3. The activity of many, but not all, tyrosine kinases is dependent on the well-conserved tyrosine residue in the activation loop for receptor activation. For instance, we have recently shown that the KIT receptor is dependent on its activation loop tyrosine for downstream signaling and receptor stability, but not for the activation of the receptor. In many AML patients, FLT3-ITD mutations in combination with mutations in the activation loop tyrosine residue (Y842) result in drug resistance. We therefore stably transduced cells with FLT3-ITD in combination with different Y842 mutants (Y842C and Y842F). Using MM-PBSA, we found that the introduction of the Y842C and Y842F point mutations changed the binding energy of the FLT3 inhibitors quizartinib, sorafenib, and midostaurin in silico. In addition, the mutants displayed a change in drug response and apoptosis levels after treatments with FLT3 inhibitors in vitro. Previous data from our lab also shows that the Y842F mutant resulted in delayed tumor formation in vivo, indicating that the phosphorylation of the activation loop tyrosine is an important event in FLT3-ITD-mediated transformation. All in all, our data suggest that the activation loop tyrosine residue in FLT3-ITD positive cells plays an important role in drug response and tumor formation.

Citation Format: Lina Al Ashiri, Sausan Moharram, Rituraj Purohit, Julhash Kazi Uddin, Lars Rönnstrand. Y842 mutations in the activation loop of FLT3 regulates drug response in FLT3-ITD positive acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3969.
Abstract 3987: Brain penetrant HDAC and PI3K/mTOR inhibitors synergize to induce DIPG cell death
Authors: Barnett Katherine , Kwon Hyuk Jean , Novak Orlandi , Eberhart Charles , Raabe Eric
Published: June 2022

Diffuse intrinsic pontine glioma (DIPG) is a universally fatal pediatric brain tumor. We have previously shown that the brain penetrant class 1 HDAC inhibitor RG2833 (RGFP109) inhibits cell growth (MTS assay for HSJD007 p=0.0004, JHH-DIPG1 p=0.001, SF-7761 p=0.04, SU-DIPG13 p=0.01 by t-test), reduces proliferation (phospho-Rb Western blot, BrdU immunofluorescence HSJD007 p=0.008, JHH-DIPG1 p=0.0002 by t-test), and induces apoptosis (cPARP Western blot, cleaved caspase 3 immunofluorescence HSJD007 p<0.003, JHH-DIPG1 p=0.0026 by t-test) in multiple DIPG cell lines. We have also shown that treatment of a DIPG flank tumor mouse model with RG2833 alone for 1 week suppresses flank tumor growth (p< 0.005 by t-test) and induces apoptosis (Western blot for cPARP). In a DIPG orthotopic mouse model, we confirmed that RG2833 100mg/kg has on target activity in the mouse brain (Western blot for histone 3 acetylation) without causing toxicity. We next sought to identify a rational combination therapy to enhance RG2833 efficacy against DIPG. We found that RG2833 increases expression of negative cell cycle regulators p16, p21, and p27 and decreases expression of anti-apoptotic proteins BCL-2, BCL-XL, and XIAP (assessed by Western blot). Given that PI3K/mTOR pathway is upregulated in DIPG and plays an important role in cell cycle regulation via p21 and the FOXO signaling, we hypothesized that RG2833 would synergize with inhibitors of PI3K/mTOR. The brain penetrant PI3K/mTOR inhibitor Paxalisib is currently being evaluated in a phase 1 clinical trial of children with DIPG. We found that RG2833 has strong synergism with Paxalisib at clinically achievable doses (Loewe synergy scores by SynergyFinder for SF7761 and JHH-DIPG1 >20). Evaluation of combination treatment for apoptotic effects in vitro and in vivo are ongoing. These findings identify RG2833 and Paxalisib as a promising combination therapy for DIPG.

Citation Format: Katherine Barnett, Hyuk Jean Kwon, Orlandi Novak, Charles Eberhart, Eric Raabe. Brain penetrant HDAC and PI3K/mTOR inhibitors synergize to induce DIPG cell death [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3987.
Abstract 4022: ERK1/2 inhibitor ulixertinib demonstrates activity in atypical (non-V600) BRAF mutant models
Authors: Knoerzer Deborah , Reddy Anupama , Derti Adnan , Emery Caroline M.
Published: June 2022

Atypical BRAF (non-V600) alterations comprise approximately half of all BRAF mutations in cancer and can be categorized according to characteristics of molecular signaling (either Class II or III). Atypical BRAF alterations are rare (approximately 3% across all human cancers) and there are currently no approved therapies for this indication. As next-generation sequencing becomes standard clinical practice, oncologists are frequently identifying atypical BRAF alterations in their patients’ tumors. The efficacy of the first-in-class ERK1/2 inhibitor, ulixertinib (BVD-523), was assessed across 10 patient-derived xenograft (PDX) models, which harbored class II or III BRAF alterations (5 models with type II alterations, 4 models with class III, and 1 model with both class II and III). Responses ranging from robust regression to moderate tumor growth delay were observed in 9/10 models. RNA sequencing was performed on tumors from the vehicle-treated and ulixertinib-treated groups with three replicates per PDX model. The samples were collected 2 hours after the last dose of treatment. Expression of the mutant BRAF alleles was readily confirmed from RNA-seq data in all PDX models. In addition, gene expression analysis showed differential expression of MAPK pathway genes in responders compared to the non-responders.

In summary, ulixertinib has exhibited strong pre-clinical activity in a variety of patient-derived xenograft models with atypical BRAF alterations. Ulixertinib has FDA fast-track designation for patients with solid tumors, other than CRC, harboring specific BRAF mutations (G469A, L485W, or L597Q) and is currently under clinical evaluation in patients with tumors harboring any atypical BRAF alteration (NCT04488003).

Citation Format: Deborah Knoerzer, Anupama Reddy, Adnan Derti, Caroline M. Emery. ERK1/2 inhibitor ulixertinib demonstrates activity in atypical (non-V600) BRAF mutant models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4022.
Abstract 4035: Driver-gene dependencies reveal clinically actionable drug repositioning opportunities
Authors: Babak Tomas , Vermeulen Michael , Villa Doris Coto , Craig Andrew
Published: June 2022

Cancer cell lines have numerous characteristics that make them favorable pre-clinical research models, yet they are notoriously poor at predicting drug response in the clinic. Here we sought to investigate the utility of synthetic lethality (SL) interactions discovered from large-scale CRISPR functional screens (i.e. the BROAD and Sanger Cancer Dependency Maps or "DepMap") as predictors of targets that validate in patients. Mutual exclusivity, the phenomenon where two genes are rarely mutated together in the same tumor, is a powerful clinical-stage readout that can be caused by synthetic lethality. We found that SL interactions discovered in DepMap are significantly more likely to be mutually exclusive in TCGA when they include a driver (tumor-suppressor/oncogene). These SL interactions represent high-value targeting opportunities with the advantage of clear patient selection criteria based on their driver mutation status. In an effort to identify drugs that target these proteins as potential repurposing opportunities, we found that pharmacogenomic inhibition rarely invokes the same target dependencies as a genetic deletion of the drug target. Nonetheless, we identified several dozen "clean" drugs with potential for repositioning and validated the top candidates in PDx. Although tumours are more heterogenous than cancer cell lines, we show that cell line viability readouts linked to single-gene/drug perturbations can yield accurate predictions of clinical efficacy when tied to tumor-driver biology.

Citation Format: Tomas Babak, Michael Vermeulen, Doris Coto Villa, Andrew Craig. Driver-gene dependencies reveal clinically actionable drug repositioning opportunities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4035.
Abstract 4038: Study of therapeutic effects of polyisoprenylated cysteinyl amide inhibitors on lung cancer cells of black patients using 3D-Printed alveolar model
Authors: Belony Nadine , Ren Bing , Pham Phuc , Gregory Matthew , Puente Pablo E. , Lamango Nazarius S. , Offringa Ite A. , Huang Yong
Published: June 2022

Lung cancer is one of the top five deadliest cancers, and black Americans have a higher mortality rate when compared to white Americans even though they smoke relatively fewer cigarettes. Lung adenocarcinoma (LUAD) is the most common histological lung cancer subtype in all population groups including blacks. There are no in vitro model systems to study lung cancer in black subjects, and there is only one lung adenocarcinoma cell line from a black patient (NCI-H23). Here we aim to address the lack of in vitro models to study black LUAD. Using three-dimensional (3D) biofabrication technologies, we’ve developed a thick vascularized tissue model to recapitulate physiologically relevant heterogeneous human alveoli. Specifically, the blood-air barrier of the alveolar model is simulated with two perfusable channels embedded in a thick vascularized tissue construct: a helical blood channel and a straight air channel. Spatially, the air channel is in the center and is surrounded by the blood channel. Human endothelial cells are seeded onto the inner surface of the blood channel and NCI-H23 cells (which carry a KRAS mutation) or untransformed alveolar epithelial cells derived from black subjects, are seeded on the inner surface of the air channel. The blood and air channels are perfused using cell medium and oxygenated air, respectively. The model is used for the study of potential LUAD therapeutic agents such as polyisoprenylated cysteinyl amide inhibitors (PCAIs), which target and disrupt the RAS-mediated signaling pathway. KRAS is the most commonly mutated oncogene in LUAD in blacks, and our experiments are aimed at ensuring that PCAIs are effective in black LUAD and may offer better treatment outcomes for ethnically/racially diverse patients in whom the RAS signaling pathway is frequently altered and for whom pharmaceutical therapy is necessitated by the late diagnoses.

Citation Format: Nadine Belony, Bing Ren, Phuc Pham, Matthew Gregory, Pablo E. Puente, Nazarius S. Lamango, Ite A. Offringa, Yong Huang. Study of therapeutic effects of polyisoprenylated cysteinyl amide inhibitors on lung cancer cells of black patients using 3D-Printed alveolar model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4038.
Abstract 4042: Genomic and pharmaco-genomic profiling of pancreatic cancer using patient-derived organoids
Authors: Deschênes Astrid , Belleau Pascal , Plenker Dennis , Habowski Amber , Patel Hardik , Park Youngkyu , Tiriac Hervé , Baker Lindsey A. , Krasnitz Alexander , Tuveson David A.
Published: June 2022

Introduction: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal common malignancies, with little improvement in patient outcomes over the past decades. We have developed a novel methodology to culture organoids from both human healthy pancreatic ductal epithelial tissues and PDAC. A collection of patient-derived organoids (PDO), grown using this protocol, numbers over 100 models to date. The 100% neoplastic purity of the organoid cultures facilitates molecular characterization that has been traditionally challenging in the pauci-cellular state of primary pancreatic tumors. These PDO open new opportunities for deep genomic and transcriptomic studies of the disease, and for individualized drug screens. Here we demonstrate that accurate predictive models of response to pharmacological treatments of PDAC can be developed using data from such screens alongside molecular profiles of the PDO.

Methods: Molecular analysis of the PDO library yielded genomic and transcriptional profiles of the cultures, including those of copy number variation (CNV), mutations in the exome and mRNA expression. From these, we drew features for prediction of drug responses. Using molecular features drawn from these profiles, we developed a panel of predictive models for response to standard-of-care cytotoxic agents and a number of targeted treatments. We employed Random Forest (RF) regression as a machine-learning tool for this purpose.

Results: PDO are faithful models of PDAC, whose molecular features closely resemble those of PDAC tumor specimens. Using a subset of these features, we were able to accurately learn PDO responses to cytotoxic agents: for each of the five agents considered, the predicted drug response correlated strongly (p < 10-7) with the observed value. A similar accuracy of prediction was achieved for a number of targeted agents.

Conclusion: PDOs are a valuable resource for molecular and pharmacological characterization of PDAC, with a potential to guide clinical decisions with regard to treatment.

Citation Format: Astrid Deschênes, Pascal Belleau, Dennis Plenker, Amber Habowski, Hardik Patel, Youngkyu Park, Hervé Tiriac, Lindsey A. Baker, Alexander Krasnitz, David A. Tuveson. Genomic and pharmaco-genomic profiling of pancreatic cancer using patient-derived organoids [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4042.