bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3224: TTFields-prolonged the PFS of epithelioid glioblastoma patient: a case report
Authors: Ding Yuxuan , Wang Qiang , Wang Feijiang , Wu Nan , Li Jianrui , He Xia , Pan Hao , Wang Lijun
Published: April 2023

Epithelioid glioblastoma (EGBM, classified as glioblastoma, wild type, grade IV according to 2021 WHO CNS5 classification of neurological malignancies) is a highly aggressive malignancy, the median progression-free survival (mPFS) of which is about 4.0 months. Although, the introduction of tumor treating fields (TTFields, possessing anti-cancer capability via anti-mitotic effects) in the maintenance of TMZ chemotherapy was demonstrated beneficial for prolonging the mPFS of GBM patients to 6.9 months in the EF-14 clinical trial, studies centering on the significance of TTFields in EGBM treating are very limited due to the rarity of EGBM. Therefore, a 28-year-old patient (male, recurrent left-sided limb twitching for 1 month and dizziness for 1 week) was chosen as a clinical trial participant. A right frontal lobe occupancy was detected based on magnetic resonance imaging (MRI), and total tumor resection was performed on the patient. Meanwhile, postoperative histopathology test, including immunohistochemical and molecular characterization, was conducted and the results revealed a BRAF V600E mutation, no codeletion of 1p and 19q, and negative MGMT promoter methylation, which confirmed EGBM. In this context, chemoradiotherapy was conducted on the patient at first, and then concurrent TTFields and TMZ was performed. Notably, a long-term PFS of 33 months and KPS score of 90 were achieved over the patient upon TTFields combined TMZ, whose average daily usage of TTFields was higher than 90%. This clinical case corroborated the importance of TTFields in EGBM treating, and the higher compliance of TTFields is more beneficial for EGBM patient without additional side effect. YD, QW contributed equally to this work.

Citation Format: Yuxuan Ding, Qiang Wang, Feijiang Wang, Nan Wu, Jianrui Li, Xia He, Hao Pan, Lijun Wang. TTFields-prolonged the PFS of epithelioid glioblastoma patient: a case report [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 3224.
Abstract 324: Nickel exposure alters extracellular vesicles biogenesis to carry NUPR1 and mediate cancer progression
Authors: Liu Shan , Ortiz Angelica , Iovanna Juan , Costa Max
Published: April 2023

Nickel (Ni), a group I human carcinogen by the International Agency for Research on Cancer (IARC) alters expression of various oncogenic molecules in cells resulting in adoption of cancer cell phenotypes, such as anchorage-independent growth, migration, and invasion. Cancer cell phenotypes are altered by internal genetic or epigenetic changes or in response to external signals mediated by soluble proteins or extracellular vesicles. However, few studies have linked extracellular vesicles (EVs) in response to environmental pollutants, such as Ni. Nuclear Protein 1 (NUPR1) is a stress response protein that is upregulated in cancer cells and it is also upregulated in cells following chronic Ni exposure. However, the role of NUPR1 in EV biogenesis or EV-mediated cancer progression is unknown. To explore the potential role of EVs in Ni-induced cancer progression, this study examined the effect of Ni-exposure on EV signature. Sizes and concentration of EVs from BEAS-2B cells, 6 weeks-Ni-treated BEAS-2B cells (Ni-6W) and Ni-transformed BEAS-2B cells (Ni-3) were identified by nanoparticle-tracking analysis (NTA) and transmission electron microscopy (TEM). In the results, showed that Ni-6W and Ni-3 released more EVs compared to the control. The transcription expression levels of EVs biogenesis markers, CD63, CD82 and Flotillin-1 (FLOT) were coincidentally increased in Ni-6W and Ni-3 compared to the BEAS-2B. We further identified that inhibition of NUPR1 by using the shRNA or NUPR1 inhibitor ZZW-130 can rescue the increased releasing of EVs induced by Ni exposure. Here we demonstrated that Ni exposure elevated EV release and that these Ni-altered EVs contributed to a pro-inflammatory environment associated with tumorigenesis and cancer progression. Moreover, we provided a new method of regulating EVs biogenesis by targeting NUPR1, this would be crucial for EV-based cancer therapy.

Citation Format: Shan Liu, Angelica Ortiz, Juan Iovanna, Max Costa. Nickel exposure alters extracellular vesicles biogenesis to carry NUPR1 and mediate 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 324.
Abstract 3251: The immunostimulatory effect of 9-ING-41, a small molecule GSK-3 inhibitor, in sarcomas
Authors: MacDonald William J. , Lew Samuel , Huntington Kelsey E. , Lulla Rishi R. , DeNardo Bradley D. , El-Deiry Wafik S.
Published: April 2023

Due to the rarity and genomic disparity of soft tissue and bone sarcomas, actionable therapeutic targets have been elusive. However, inhibition of GSK-3β has emerged as a potentially promising therapy that could be of great benefit to pediatric and adult sarcoma patients. Saos-2 osteosarcoma and 93T449 liposarcoma cell lines were pretreated at IC5072 with small molecule GSK-3β inhibitor 9-ING-41 (elraglusib). The cells were harvested for western blot analysis after 48 hours of treatment. Additionally, both cancer cell lines as well as TALL-104 T-cells and NK-92 NK cells were treated with 0.5 μM 9-ING-41 for 24 hours and harvested for Luminex cytokine profiling. The western blots demonstrated an increase in cPARP, an apoptotic marker, in both cancer cell lines after 9-ING-41 treatment. Additionally, an increase in PD-L1 expression was observed. The cytokine analysis revealed stimulation of immune cell activity in response to 9-ING-41. Treated T-cells had an increase in CXCL11, which is associated with T-cell recruitment, as well as an increased level of IL-18, which is shown to induce increased IFN-γ in Th1 cells. Additionally, NK-92 cells demonstrated an increase in IL-8 chemokine and an increase in soluble TRAIL (TRAIL/TNFSF10). The cancer cell lines showed a homogenous increase in growth factor TGF-ɑ, however, only the Saos-2 osteosarcoma cell line demonstrated an increase of IL-6. We are further validating the data with follow-up cytokine profiling. The increase in immunostimulatory cytokines as well as the increased expression of PD-L1 suggest a rationale for combining 9-ING-41 with immune checkpoint blockade therapy. The potential synergistic effect of these two therapies is currently under investigation with co-culture experiments of sarcoma and immune cells. The treatment cohorts for these experiments include 9-ING-41 combined with either anti-PD-L1, anti-PD-1, or anti-CTLA-4 immune checkpoint inhibitors. Our results suggest a promising combination therapeutic strategy for patients with soft tissue and bone sarcomas and future work will strive to better elucidate the mechanisms of efficacy.

Citation Format: William J. MacDonald, Samuel Lew, Kelsey E. Huntington, Rishi R. Lulla, Bradley D. DeNardo, Wafik S. El-Deiry. The immunostimulatory effect of 9-ING-41, a small molecule GSK-3 inhibitor, in sarcomas [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 3251.
Abstract 3309: Genomic features, evolutionary patterns, and minimal residual disease at surgical margins as novel prognostic/predictive biomarkers in locally advanced rectal cancer
Authors: Li Li , He Kewen , Li Aijie , Xu Yang , Pang Jiaohui , Mu Dianbin , Ma Jie , Ge Hong , Maleki Aurian , Qin Xueting , Zhang Xian , Ou Qiuxiang , Tang HaiMeng , Shao Yang , Yu Jinming , Yuan Shuanghu
Published: April 2023

Background: Locally advanced rectal cancers (LARC) are treated with neoadjuvant chemoradiotherapy (nCRT), total mesorectal excision (TME), and adjuvant chemotherapy, and identifying reliable prognostic/predictive biomarkers is currently warranted for LARC.

Methods: Comprehensive genomic profiling was performed in 76 LARC patients who received nCRT plus TME. All 76 patients had baseline tissue biopsies, 72 had paired baseline tissue biopsies and surgical tumor samples, and 55 had paired surgical tumor and margin samples. Post-nCRT tumor regression grade (TRG) and post-surgical disease-free survival (DFS) were used to assess treatment outcomes.

Results: Baseline KRAS mutation (P=0.022) and MYC amplification (P=0.047) appeared to be independent prognostic factors for nCRT, with KRAS/MYC-mutated patients having higher tumor regression grade (TRG) and lower immune infiltration. Intriguingly, high radiation doses during nCRT were correlated with improved nCRT response in KRAS/MYC-positive patients. Additionally, aberrations in multiple signaling pathways, especially JAK-STAT (P=0.022), were enriched in TRG3 patients after nCRT, implying that they might be resistance mechanisms that impair nCRT efficacy. The post-surgical FBXW7 mutation was significantly associated with shorter disease-free survival (P=0.01), while treatment-induced branched tumor evolutionary pattern and the molecular level of residual tumor at the surgical margin were significantly correlated with both poorer nCRT response and higher post-surgical recurrence risk.

Conclusions: Comprehensive genomic profiling helps identify multiple molecular prognostic and/or predictive biomarkers that could predict patients’ clinical outcomes to nCRT and TME and direct nCRT treatment regimen, thus facilitating more accurate prognostic estimation, a better balance between treatment efficacy and quality of life, and timely adjusted treatment decisions.

Citation Format: Li Li, Kewen He, Aijie Li, Yang Xu, Jiaohui Pang, Dianbin Mu, Jie Ma, Hong Ge, Aurian Maleki, Xueting Qin, Xian Zhang, Qiuxiang Ou, HaiMeng Tang, Yang Shao, Jinming Yu, Shuanghu Yuan. Genomic features, evolutionary patterns, and minimal residual disease at surgical margins as novel prognostic/predictive biomarkers in locally advanced rectal 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 3309.
Abstract 3315: Evaluating biomarker potential of germline genomic factors for predicting clinical outcomes in prostate cancer
Authors: Zeltser Nicole , Houlahan Kathleen E. , Al-Hiyari Sarah M. , Eng Stefan E. , Patel Yash , Yamaguchi Takafumi N. , Tao Shu , Huang Rong Rong , Reiter Robert E. , Ye Huihui , Kinnaird Adam S. , Boutros Paul C.
Published: April 2023

Prostate cancer is the second-most diagnosed cancer and the second leading cause of cancer death in American men. Early detection is common, but is followed by the more challenging task of prognosing a highly variable clinical course. Current clinical risk-assessment strategies such as serum abundance of prostate specific antigen (PSA), tumor size & extent, and tumor grade based on biopsy are highly imprecise: over a third of patients are over-treated. An improved method of risk stratification may lie in hereditary factors. Prostate cancer is one of the most strongly inherited (h2 = 57%), with accumulating evidence associating rare variants, common variants, and genetic ancestry to clinical outcomes. We have performed germline sequencing on blood from thousands of patients diagnosed with localized prostate cancer and with extensive follow-up data. We quantify the interactions of rare and common variants, and demonstrate that germline features provide insights into patient outcomes and optimal management strategies.

Citation Format: Nicole Zeltser, Kathleen E. Houlahan, Sarah M. Al-Hiyari, Stefan E. Eng, Yash Patel, Takafumi N. Yamaguchi, Shu Tao, Rong Rong Huang, Robert E. Reiter, Huihui Ye, Adam S. Kinnaird, Paul C. Boutros. Evaluating biomarker potential of germline genomic factors for predicting clinical outcomes in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3315.
Abstract 337: Antitumor effect of oncomagnetic therapy in glioblastoma
Authors: Pandey Arvind , Hambarde Shashank , Baskin David S. , Helekar Santosh A.
Published: April 2023

Background: Glioblastoma (GBM) is the most lethal primary malignant brain tumor with a median survival of 15-20 months. Concurrent temozolomide (TMZ) chemotherapy and radiation (XRT) remain the current standard of care (SOC) treatment for newly diagnosed GBM. The addition of tumor treating fields (TTFs) improves median survival from 16 to 20.9 months. This modest improvement underscores the urgent need to identify a new treatment modality as standalone therapy. We have reported recently that brief 2 - 6 h of daily brain stimulation with a new noninvasive, non-contact, magnetic stimulation device causes >30% reduction of contrast-enhanced tumor (CET) volume in an end-stage recurrent GBM patient after a ~30-day treatment.

Purpose: The noninvasive Oncomagnetic device developed in our laboratory selectively kills glioblastoma (GBM) and other cancer cells while sparing normal developing neurons and astrocytes in vitro. The spinning oscillating magnetic field (sOMF) generated by this device disrupts electron transport in the mitochondrial respiratory chain leading to a marked increase in reactive oxygen species (ROS) and consequent cancer cell death. However, the downstream molecular mechanism of sOMF action is under investigation. We studied the cellular and molecular effects of sOMF treatment on GBM cell lines and investigated the response to Oncomagnetic monotherapy (OMT) in a syngeneic mouse model. We treated LN229 and U87 GBM cells with sOMF with and without a sub-therapeutic dose of TMZ in vitro. To study the anticancer effect of sOMF in vivo, we developed syngeneic GBM tumors in BALB/c mice by stereotactic intracranial injection of GL261WT cells. Tumor growth was monitored periodically by MRI scans.

Results: Our results showed that sOMF significantly reduce cell proliferation and cell survival in vitro. We observed that sOMF induces DNA damage and arrests cells in G1 phase of cell cycle. OMT of tumor-bearing mice using a whole-body sOMF stimulation method caused a substantial retardation of tumor growth and reduction of the contrast-enhanced tumor volume in 9.4 T MRI scans. OMT also showed significant increase in overall survival in these mice.

Conclusion: These experiments contribute to unraveling the mechanism of action underlying the response to OMT in GBM and provide a strong rationale for a standalone Oncomagnetic therapy for GBM.

Citation Format: Arvind Pandey, Shashank Hambarde, David S. Baskin, Santosh A. Helekar. Antitumor effect of oncomagnetic therapy in glioblastoma [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 337.
Abstract 3413: RTK/SHP2 dependency and feedback adaptation to MEK inhibition defines sensitivity of RAS-mutant proteins to SHP2 inhibitor-based therapies
Authors: Baars Beau , Adamopoulos Christos , Orive-Ramos Ana , Aaronson Stuart A. , Poulikakos Poulikos I.
Published: April 2023

Activating RAS mutations constitute the majority of recurrent mutations found in MAPK-driven human tumors. However, RAS mutants have long been considered undruggable, and therefore efforts have been predominantly focused on targeting downstream MAPK signaling. We and others have shown that, MAPK inhibition using MEK inhibitors (MEKi) have limited effectiveness in such tumors due to adaptive resistance. A major mechanism of adaptive resistance involves relief of negative feedback that promotes reactivation of RTK signaling, which in turn activates RAS and results in MAPK rebound. The recent development of direct (i.e. RAS(G12C)) and indirect (i.e. SHP2i, SOSi) inhibitors of RAS have uncovered the dependence of activity of RAS(G12C) and of other RAS mutants on upstream signaling.

We and others have previously characterized sensitivity of various RAS mutants in cancer to either SHP2 inhibition (SHP2i), or to combination of SHP2i with MEKi, but a systematic characterization of most RAS mutants commonly expressed in cancer has been lacking. In this study, by combining biochemical and chemical genetic approaches we identified two classes of RAS-mutants, based on their degree of dependency on RTK/SHP2 signaling and adaptation to MEK inhibition. One class of RAS mutants, including most RASQ61X, show complete independence from RTK/SHP2 signaling and are fully resistant to either SHP2i alone or in combination with MEKi. The other class includes the majority of RAS mutants and is characterized by relative resistance to SHP2i, or the combination of SHP2i plus MEKi, compared to cells expressing wild-type RAS, including normal cells. Early clinical data using single agent SHP2i or combinations of SHP2i with MEKi for the treatment of patients with RAS-mutant tumors suggested modest activity as well as dose-limiting on-target (i.e. MAPK pathway related) toxicities. Our findings predict that achieving an acceptable therapeutic index by either approach for treatment of RAS-mutant tumors will be challenging, and they support instead SHP2i-based therapeutic efforts in patients with MAPK-driven tumors expressing wild-type RAS, or in combination with RAS-mutant selective inhibitors.

Collectively this work provides a framework for the design of SHP2i-based treatments for patients with MAPK-driven tumors, the majority of which currently have no effective targeted therapeutic options.

Citation Format: Beau Baars, Christos Adamopoulos, Ana Orive-Ramos, Stuart A. Aaronson, Poulikos I. Poulikakos. RTK/SHP2 dependency and feedback adaptation to MEK inhibition defines sensitivity of RAS-mutant proteins to SHP2 inhibitor-based therapies [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 3413.
Abstract 3415: Preclinical characterization of a brain penetrant RAF inhibitor, BDTX-4933, targeting oncogenic BRAF Class I/II/III and RAS mutations
Authors: Han Yoon-Chi , Ng Pui-Yee , Ogawa Luisa Shin , Yang Shao Ning , Chen Miao , Ishiyama Noboru , Lin Tai-An , Buck Elizabeth
Published: April 2023

Mutations in BRAF and RAS are often oncogenic and lead to a constitutively active MAPK pathway that promotes aberrant cell proliferation and tumor growth. Currently approved BRAF inhibitors are selective against monomeric BRAF V600 mutants. These drugs are largely inactive against non-V600 dimeric BRAF mutants and have poor brain penetration. Although there is an FDA-approved KRAS G12C mutant-selective inhibitor, there are no approved inhibitors for cancer patients who harbor other (non-G12C) KRAS and NRAS mutations which promote tumor growth likely through constitutively active RAF dimers. There remains a high unmet clinical need for a highly CNS penetrant oral RAF inhibitor that targets a broad spectrum of BRAF mutations and constitutively active RAF dimers without paradoxical activation of the MAPK signaling pathway. BDTX-4933 is a potent, reversible, CNS penetrant RAF MasterKey inhibitor designed to target a large family of oncogenic BRAF mutations including BRAF monomers and RAF dimers. The compound inhibits not only all classes (I, II, and III) of BRAF mutations but also targets constitutively active RAF dimers promoted by upstream oncogenic MAPK pathway alterations, such as RAS mutations. In a panel of cancer cell lines that endogenously express BRAF or RAS mutations, BDTX-4933 demonstrates inhibition of the MAPK pathway signaling without paradoxical activation, resulting in potent inhibition of cellular proliferation. BDTX-4933 shows target engagement, inhibiting ERK phosphorylation, in tumor models in vivo, achieving strong anti-tumor efficacy and tumor regression across tumor models driven by either BRAF or RAS mutations. Furthermore, BDTX-4933 exhibits high CNS exposure leading to dose-dependent tumor growth inhibition, and survival benefit in mice implanted intracranially with xenograft BRAF mutant tumors. BDTX-4933 has a best-in-class profile to treat cancer patients harboring BRAF mutations or RAF dimer-promoting upstream genetic alterations. IND-enabling studies for BDTX-4933 are on-going.

Citation Format: Yoon-Chi Han, Pui-Yee Ng, Luisa Shin Ogawa, Shao Ning Yang, Miao Chen, Noboru Ishiyama, Tai-An Lin, Elizabeth Buck. Preclinical characterization of a brain penetrant RAF inhibitor, BDTX-4933, targeting oncogenic BRAF Class I/II/III and RAS mutations [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 3415.
Abstract 3522: DICER1 Platform domain missense variants led to tumor susceptibility via failure of precursor miRNA binding
Authors: Pelletier Dylan , Chong Anne-Laure , Albert Sophie , Fabian Marc , Foulkes William
Published: April 2023

DICER1 is a key protein in the micro(mi)RNA biogenesis pathway responsible for cleaving precursor (pre)-miRNA stemloops with its RNase IIIa and IIIb domains into mature single stranded miRNAs. DICER1 syndrome is a pleiotropic tumor predisposition syndrome in which clinical manifestations are normally first seen during childhood. This genetic disorder is defined by the presence of inherited or de novo germline pathogenic variants (GPVs) in the DICER1 gene. While many germline loss of function variants fail to generate DICER1 protein, a subset of patients has germline missense variants that are predicted to produce full-length DICER1 protein that cluster in the DICER1 Platform domain, a region that plays a role in binding to the 5’ phosphate of pre-miRNA stemloops. Due to the uncertainty surrounding the pathogenicity of missense variants in DICER1 syndrome, clinical actionability can be limited. To determine how Platform mutations predispose to DICER1 syndrome we characterized a number of these variants using in vitro pre-miRNA binding and cleavage assays. Interestingly, four of these variants displayed impaired pre-miRNA binding and processing, with three of them localizing to a hydrophobic pocket in the Platform domain. In keeping with in vitro results, DICER1 mutants displayed impaired miRNA processing when introduced into mesenchymal stem cells. Furthermore, we identified a second somatic hit in the RNase IIIb domain of DICER1 in thyroid nodular hyperplasia DNA from a patient affected by one of these variants. Taken together, these results identify a key region in the DICER1 protein that is essential for pre-miRNA processing and help the classification of these germline variants as pathogenic to ensure proper clinical management of affected patients. Additionally, these results provide insight on the molecular basis involved in DICER1 syndrome predisposition.

Citation Format: Dylan Pelletier, Anne-Laure Chong, Sophie Albert, Marc Fabian, William Foulkes. DICER1 Platform domain missense variants led to tumor susceptibility via failure of precursor miRNA binding. [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 3522.
Abstract 3600: MET-mediated STING activation regulates EMT and NF-ΚB pathway in non-small cell lung cancer
Authors: Mun Seungchan , Han Bogyeong , Ahn HyunKyung , Koh Jaemoon , Kim Sehui , Chung DooHyun , Jeon YoonKyung
Published: April 2023

The cyclic GAMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway is known to have anti-tumor activity by activating the innate immune response against cytosolic double-stranded DNA (dsDNA). The regulating mechanism of STING remains unclear, and a pro-tumorigenic effect of STING pathway was recently reported. In this study, we investigated the expression of STING by MET and a novel role of the STING pathway in non-small cell lung cancer (NSCLC). The Cancer Genome Atlas (TCGA) and Cancer Cell Line Encyclopedia (CCLE) analysis showed that cGAS/STING pathway is enriched and STING expression is higher in MET-overexpressing tumors. Western blot for NSCLC cells and immunohistochemistry for NSCLC tumor tissues showed positive correlation between MET and STING expression in NSCLC cells. MET-overexpression and a MET agonist (HGF) treatment increased STING expression and CXCL10 and IFNβ expression and secretion, whereas MET-knockdown and a MET inhibitor (capmatinib) treatment decreased STING expression in NSCLC cells. In MET-positive and STING-positive cells, a STING agonist (poly-dAdT and diABZI) treatment led to increased activation of STING signaling pathway. In EBC-1 cells, MET knockdown decreased STING expression and STING-agonist-induced STING-pathway activation and cytokine production. To further investigate the role STING in NSCLC, EBC-1 cells were transfected with STING siRNA and submitted to RNS-seq. Gene set enrichment analysis revealed that several pathways were affected by STING expression. Epithelial-mesenchymal transition (EMT) and TNFα-signaling via NF-κB signaling were significantly downregulated in STING-knockdowned cells. In vitro experiments using EBC-1 cells, STING overexpression increased SLUG and TWIST1 expression and migration of cells, whereas STING knockdown decreased SLUG and TWIST1 expression and migration of cells. In summary, this study demonstrated that MET and STING expression were positively correlated with each other in NSCLC and MET overexpression increased STING expression and subsequent activation upon STING agonist stimulation. In addition, STING contributed to EMT in NSCLC.

Citation Format: Seungchan Mun, Bogyeong Han, HyunKyung Ahn, Jaemoon Koh, Sehui Kim, DooHyun Chung, YoonKyung Jeon. MET-mediated STING activation regulates EMT and NF-ΚB pathway in non-small cell lung 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 3600.
Abstract 3629: Calcium signalling and the breast cancer microenvironment
Authors: Bong Alice H. , Gieniec Krystyna A. , Sadras Francisco , Robitaille Mélanie , Roberts-Thomson Sarah J. , Davis Felicity M. , Monteith Gregory R.
Published: April 2023

Calcium is a key regulator of a variety of pathways important in breast cancer progression, including those involved in cell proliferation and invasiveness. Calcium signaling is implicated in processes relevant to the breast cancer tumor microenvironment and altered calcium influx is a feature of cancer associated fibroblasts. In these studies, simultaneous use of genetically encoded calcium indicators with distinct fluorescence spectral properties (GCaMP6m or JRCaMP1b), were used to explore the cross-talk between breast cancer cells and fibroblasts via calcium signaling and changes in different cellular compartments. HMF3S human fibroblast cells co-cultured with breast cancer cells expressing GCaMP6m or JRCaMP1b were assessed in 2D and 3D co-culture using automated epi-fluorescence or confocal microscopy. Mitochondrial calcium levels were assessed using targeted GCaMP6m (2mtGCaMP6m) in HMF3S cells expressing JRCaMP1b. Results showed distinct spatial and temporal changes in free calcium levels between both fibroblasts and breast cancer cells and the mitochondria and cytosol during activation. These methodological advances provide an opportunity to better understand calcium signaling in cancer associated fibroblasts. These techniques will also help identify the calcium channels and pumps that could be targeted to manipulate the breast cancer tumor microenvironment to inhibit pathways important in breast cancer progression.

Citation Format: Alice H. Bong, Krystyna A. Gieniec, Francisco Sadras, Mélanie Robitaille, Sarah J. Roberts-Thomson, Felicity M. Davis, Gregory R. Monteith. Calcium signalling and the breast cancer microenvironment. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3629.
Abstract 3672: RNA epigenetic regulation of cholesterol metabolism in neuroblastoma
Authors: Bansal Mohit , Gupta Anamika , Ding Han-Fei
Published: April 2023

Pseudouridylation (conversion of uridine to pseudouridine) is a common epigenetic modification of RNA which increases its structural stability. RNA pseudouridylation is catalyzed by the family of pseudouridine synthases (PUS). The functional significance and biological relevance of mRNA pseudouridylation are poorly understood. Here we report that the oncogene MYCN, which drives the development of high-risk neuroblastoma, transcriptionally upregulates the PUS1 gene. Further, examination of gene expression profiling data from neuroblastoma patients (the SEQC dataset) reveals that higher PUS1 expression is associated with poor prognosis. The transcriptional upregulation of PUS1 expression in MYCN amplified cell lines is of functional significance, as our studies reveals that knockdown of PUS1 expression in neuroblastoma cell lines suppresses cell survival and proliferation. Our RNA-seq analysis reveals that a key downstream target of PUS1 is cholesterol metabolism. PUS1 knockdown reduces the expression of key enzymes in the mevalonate pathway for cholesterol synthesis. Mechanistically, PUS1 may targets SREBP2, a master regulator of cholesterol metabolism responsible for transcriptional upregulation of enzymes for cholesterol biosynthesis. We showed that PUS1 expression is essential for maintaining SREBP2 expression in MYCN-amplified neuroblastoma cells and for statin-induced feedback activation of the mevalonate pathway, a commonly used system for assessing the transcriptional activity of SREBP2. These findings suggest that a function of PUS1 in sustaining the proliferation of neuroblastoma cells is to promote cholesterol metabolism, which could be exploited as a therapeutic strategy against high-risk neuroblastoma.

Citation Format: Mohit Bansal, Anamika Gupta, Han-Fei Ding. RNA epigenetic regulation of cholesterol metabolism in neuroblastoma. [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 3672.
Abstract 3695: Mitochondrial uncoupling reverses reductive carboxylation in cancer
Authors: Jiang Haowen , He Clifford Jiajun , Li Albert M. , He Bo , Li Yang , Ye Jiangbin
Published: April 2023

When electron transport chain (ETC) activity is inhibited upon hypoxia or when cultured in suspension, reductive carboxylation plays an essential role in supporting tumor growth through converting α-ketoglutarate (α-KG) to citrate, which is converted to acetyl-coenzyme A (acetyl-CoA) for lipogenesis. However, there is no effective way to target reductive carboxylation for tumor therapy. In this study, we use the mitochondrial uncoupler niclosamide ethanolamine (NEN) and BAM15 to reverse reductive carboxylation in cancer cells. Uncoupler treatment activates ETC and increases the cellular NAD+/NADH ratio. Using U-13C-glutamine and 1-13C-glutamine tracers, we discovered that uncoupler treatment accelerates the oxidative TCA cycle and blocks reductive carboxylation, particularly under hypoxia and spheroid culture condition. Uncoupler treatment significantly reduces tumor cell proliferation, survival and sphere formation. In addition, the intracellular acetyl-CoA levels are significantly decreased by NEN treatment. Furthermore, lipidomics analysis showed that NEN treatment reduced fatty acid abundance and altered the lipid composition under both normoxia and hypoxia. Together, these data demonstrate mitochondrial uncoupling redirects the α-KG flux from reductive carboxylation back to the oxidative TCA cycle, highlighting that the ETC activity and NAD+/NADH ratio is the key switch that determines the metabolic fate of α-KG. Reversing reductive carboxylation could be one key mechanism that mitochondrial uncoupler inhibits tumor growth.

Citation Format: Haowen Jiang, Clifford Jiajun He, Albert M. Li, Bo He, Yang Li, Jiangbin Ye. Mitochondrial uncoupling reverses reductive carboxylation in 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 3695.
Abstract 3697: Modulation of Nicotinamide N-Methyl Transferase (NNMT) expression in TNBC is associated with altered cell membrane cholesterol levels
Authors: Chee Woei-Yaw , Dolcen Deniz Nesli , Bennett Lynda , Conzen Suzanne D.
Published: April 2023

Introduction: Cancer cells often develop a program of metabolic adaptability and epigenetic remodelling to promote gene expression changes causing cancerous cell proliferation and invasion. The metabolic-epigenetic axis as a an underlying mechanism of carcinogenesis could lead to new approaches to cancer treatment. Nicotinamide N-Methyl Transferase (NNMT) is a metabolic enzyme that catalyzes the transfer of a methyl group from S-Adenosyl methionine (SAM) to nicotinamide, thereby decreasing the overall methyl pools for RNA, histone and DNA methylation. High NNMT expression has been previously associated with a poor outcome in TNBC.

Hypothesis: We hypothesized that high versus low NNMT activity and resulting oncogenic gene expression in TNBC may alter histone methylation is association with altered metabolic pathway activation.

Methods: We used two TNBC cell lines models, MDA-MB 231 and SUM159, to study the association between high versus low NNMT activity and metabolic pathway activation. Surprisingly, we identified gene expression pathways suggestive of increased free cholesterol biosynthesis gene expression with low NNMT, and then used biochemical assays and filipin to measure free cholesterol in both total high and low NNMT TNBC cell lysates and relative cholesterol expression in the High vs Low NNMT cell membranes.

Results: In vitro, NNMT knockdown resulted in increased cholesterol synthetic pathway enzyme expression in both TNBC cell lines. In vivo studies also showed that protein expression of cholesterol synthesis enzymes was also increased in NNMT-depleted MDA-MB 231 xenografted tumors. Interestingly, the increased cholesterol level upon NNMT depletion was mainly localized at the plasma membrane via Filipin III staining, suggesting a possible association with less cellular membrane fluidity and a decreased oncogenic phenotype. In ongoing experiments, we are evaluating whether changes in histone methylation in high vs low NNMT cells are associated with differential expression of cholesterol biosynthetic genes, and free cholesterol production. We hypothesize that NNMT might epigenetically decrease the methylation of histones regulating cholesterol production in TNBC.

Citation Format: Woei-Yaw Chee, Deniz Nesli Dolcen, Lynda Bennett, Suzanne D. Conzen. Modulation of Nicotinamide N-Methyl Transferase (NNMT) expression in TNBC is associated with altered cell membrane cholesterol levels. [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 3697.
Abstract 3702: Serine promotes aggressive phenotype in renal cancer
Authors: Karki Suman , Kundu Anirban , Brinkley Garret , Nam Hyeyoung , Goliwas Kayla , Deshane Jessy , Sudarshan Sunil
Published: April 2023

Tumor cells are known to have increased demand for nutrients to support proliferation. Among the most highly utilized nutrients by tumor cells are nonessential amino acids (NEAAs). While the role of NEAAs in supporting tumor growth is well-established, their role in supporting aggressive phenotypes such as migration and invasion is poorly understood. Yet, the major cause of morbidity from malignancies such as renal cell carcinoma (RCC) is metastasis. Through an unbiased approach, we assayed the role of NEAAs in promoting the migration of RCC cells. We demonstrate that exogenous serine and glutamine are essential and sufficient to support aggressive phenotypes in RCC. Moreover, the availability of both NEAAs is critical for the expression of the proinvasive transcription factor SNAIL. Our data converge on the role of these two NEAAs in supporting the translation of this factor. Perturbations that limit the availability and/or synthesis of either result in translational reprogramming through the integrated stress response (ISR) that leads to loss of SNAIL1 expression and attenuates aggressive phenotypes. Our findings associate nutrient status to the ISR including ER stress. Collectively, our study highlights new insights for NEAA metabolism in supporting aggressive tumor phenotypes and could reveal novel approaches to mitigate metastasis by targeting NEAA availability and/or utilization.

Citation Format: Suman Karki, Anirban Kundu, Garret Brinkley, Hyeyoung Nam, Kayla Goliwas, Jessy Deshane, Sunil Sudarshan. Serine promotes aggressive phenotype in renal 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 3702.
Abstract 371: Integrin-mediated lipolysis by pancreatic cancer-derived extracellular vesicles in cancer-associated cachexia
Authors: Shibata Chikako , Otsuka Motoyuki , Fujishiro Mitsuhiro
Published: April 2023

Introduction: Cancer-associated cachexia (CAC) is a life-threating condition frequently observed in cancer patients, especially those with pancreatic cancer. CAC is a multifactorial disease, with the notable characteristic of systemic lipolysis. Although humoral factors have been implicated in its pathogenesis, critical factors remain to be determined. Since even localized pancreatic cancer frequently induce lipolysis systemically, we examined the molecular mechanisms in vitro and in vivo based on the hypothesis that extracellular vesicles (EVs) systemically released from pancreatic cancer cells induce lipolysis.

Procedures: EVs derived from the supernatants of cultured pancreatic cancer cell lines and from the sera of pancreatic cancer patients were characterized. The effects of EVs on adipocytes were determined using mature human adipocytes induced from human adipose-derived mesenchymal stem cells. To confirm the in vitro findings, lipolysis induction was examined in mice by intravenous injection of EVs. EV characteristics inducing lipolysis were determined. Those characteristics were confirmed by knockout studies and examining specifically isolated pancreatic cancer-derived EVs from heterogenous EV population in patients’ sera.

Results: Lipolysis was triggered by specific EVs from pancreatic cancer cell line via the cAMP inside and uptake of more EVs into adipocytes was crucial for higher levels of lipolysis. In vivo studies, database search, and knockout studies revealed that integrin β1 and integrin α6 on EVs were the determinants for the attachment and uptake of the EVs to adipocytes, resulting in inducing more lipolysis. There was no difference in the percentage of these integrins positive EVs between heterogenous EV population in cancer patients’ sera and EVs in cancer-free controls’ sera. However, pancreatic cancer cell-derived EVs specifically isolated by targeting CA19-9, an aberrant glycosylation, from heterogenous EV population in patients’ sera, confirmed that pancreatic cancer-derived EVs have more such integrins.

Conclusions: Cancer cell-derived EVs in the sera of pancreatic cancer patients have more ITGB1 and ITGA6 levels, which may explain the systemic induction of lipolysis in cancer cachexia from the early stages and could be novel therapeutic targets against CAC. In addition, specific isolation of cancer-derived EVs in sera from heterogenous EV population is crucial for elucidating cancer-related pathogenesis.

Citation Format: Chikako Shibata, Motoyuki Otsuka, Mitsuhiro Fujishiro. Integrin-mediated lipolysis by pancreatic cancer-derived extracellular vesicles in cancer-associated cachexia [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 371.
Abstract 3726: Histone acetylation axis in the control of RNA Pol2 clusters
Authors: Gryder Berkley E. , Chin Diana , Kim Hyunmin , Osman Issra
Published: April 2023

Core regulatory transcription factors (CR TFs) orchestrate the placement of super enhancers (SEs) to activate transcription of cell-identity specifying gene networks and are critical in promoting cancer. We defined the core regulatory circuitry of fusion positive rhabdomyosarcoma (FP-RMS, a cancer of childhood) in primary tumors and cell lines, which includes PAX3-FOXO1 (P3F), MYOD1, SOX8, MYCN and others. To find chemical probes able to selectively inhibit CR TF transcription, we screened the Structural Genomics Consortium epigenetic probe set by RNA-seq. We found that chemical probes along the acetylation-axis, and not the methylation-axis, are able to cause selective disruption of CR TF transcription. Inhibitors of HDACs (acetylation erasers), BRD4 (acetylation readers) and CBP/p300 (acetylation writers) were all able to selectively halt CR TF transcription. For HDACs, this raised a conundrum: why would too much histone acetylation, an active chromatin mark, stop transcription at CR TFs? ChIP-seq showed that CR TFs build SEs that have the largest quantities of histone acetylation and the enzymes that write acetylation (i.e., p300), yet paradoxically also harbor the highest amounts of the opposing histone deacetylases (HDACs). To investigate the architectural effects of disabling HDACs and causing hyper acetylation, we developed Absolute Quantification of Architecture (AQuA) HiChIP, revealing erosion of native SE contacts at CR TFs, and extensive aberrant contacts. This did not cause an elongation defect, but rather removed RNA Pol2 from core regulatory genetic elements and eliminated RNA-Pol2 phase condensates in 20 minutes. We further dissected the contribution of HDAC isoforms using a set of HDAC selective inhibitors, finding HDAC1/2/3 are co-essential to CR transcription. Using HAT inhibitors/degraders, we discovered a profound dependence on CBP/p300 for clustering of Pol2 loops that connect P3F to its target genes. In the absence of CBP/p300, Pol2 long range enhancer loops collapse, Pol2 accumulates in CpG islands and fails to exit the gene body. These results reveal a potential novel axis for therapeutic interference with P3F in FP-RMS and clarify the molecular relationship of P3F and CBP/p300 in sustaining active Pol2 clusters essential for oncogenic transcription. Overall, our data reveals a SE-specific need for balancing histone acetylation states to maintain SE architecture, Pol2 clustering in 3D, and CR TF transcription.

Citation Format: Berkley E. Gryder, Diana Chin, Hyunmin Kim, Issra Osman. Histone acetylation axis in the control of RNA Pol2 clusters. [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 3726.
Abstract 3777: MicroRNA-inducible CRISPR/Cas9 for cell type-specific genome regulation in cancer
Authors: Oh Seung Ja , Shin Cheol-Hee , Lee Ji Min , Lee Juyong , Park Su Chan
Published: April 2023

MicroRNA-dependent mRNA decay plays an important role in gene silencing by facilitating posttranscriptional and translational repression. Inspired by this intrinsic nature of microRNA-mediated mRNA cleavage, here, we describe a microRNA-targeting mRNA as a switch platform called mRNA bridge mimetics to regulate the translocation of proteins. We applied the mRNA bridge mimetics platform to Cas9 protein to confer it the ability to translocate into the nucleus via cleavage of the nuclear export signal called CRISPR Self Check-In. This system performed programmed gene editing in vitro and in vivo. Combinatorial treatment with cisplatin and miR-21-EZH2 axis-targeting CRISPR Self Check-In improved sensitivity to chemotherapeutic drugs in vivo. Using the endogenous microRNA mediated mRNA decay mechanism, our platform is able to remodel a cell’s natural biology to allow the entry of precise drugs into the nucleus, devoid of non-specific translocation. The mRNA bridge mimetics strategy is promising for applications in which the reaction must be controlled via intracellular stimuli and modulates Cas9 proteins to ensure safe genome modification in diseased conditions.

Citation Format: Seung Ja Oh, Cheol-Hee Shin, Ji Min Lee, Juyong Lee, Su Chan Park. MicroRNA-inducible CRISPR/Cas9 for cell type-specific genome regulation in 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 3777.
Abstract 382: Synergistic anti-tumor effect of ABN401 in combination with KRAS inhibitor in a c-MET-altered KRASG12C non-small cell lung cancer model
Authors: Lee Saehyung , Cho Sunghwan , Kim Na Young , Nirmal Rajasekaran , Park Kyung Eui , Choi Jun Young
Published: April 2023

ABN401, a highly selective c-MET inhibitor, is being evaluated for the treatment of non-small cell lung cancer (NSCLC) in clinical trials. NSCLC is a heterogeneous in nature and leading cause in cancer related mortality. Numerous studies are still ongoing to identify the key oncogenic mutations that promote tumor progression. Various actionable genetic mutations such as EGFR, ALK, BRAF, MET, RET, and NTRK/ROS1 have been identified, and the targeted therapies are being developed. It is known that targeted therapies can control disease progression in NSCLC patients and drug resistance due to their heterogeneity remains a clinical hurdle. Among the targets, KRAS gene mutations, an unactionable and is now considered a promising therapeutic approach for NSCLC. Currently, co-occurrence of KRASG12C mutation has been reported in NSCLC patients with MET alterations. Moreover, oncogenic KRAS protein can induce c-MET expression levels by signaling pathways. In this study, we aim to provide an alternative therapeutic strategy for NSCLC patients with KRASG12C mutation harboring MET alterations. We have performed several in vitro and in vivo efficacy studies for the combination of ABN401, c-MET inhibitor, and sotorasib, KRASG12C variant inhibitor, including a PDX model of c-MET-altered KRASG12C NSCLC. Our results suggest that the combination of ABN401 and KRAS inhibitor could be an alternative treatment option for c-MET-altered KRASG12C NSCLC patients.

Citation Format: Saehyung Lee, Sunghwan Cho, Na Young Kim, Rajasekaran Nirmal, Kyung Eui Park, Jun Young Choi. Synergistic anti-tumor effect of ABN401 in combination with KRAS inhibitor in a c-MET-altered KRASG12C non-small cell lung cancer model [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 382.