bluuebunny/crossref_metadata_2025 huggingface dataset:
Investigation of Conditions for Preparation of Ordered Nanohole Arrays by Anodization of Iron Substrates with Depression Patterns
Authors: Osada Yuga , Yanagishita Takashi
Published: June 2023

Ordered iron oxide nanohole arrays were fabricated by the anodization of iron substrates with depression patterns formed by Ar ion milling with alumina masks in an ethylene glycol electrolyte containing NH4F and H2O. It was found that the optimization of anodization voltage, NH4F and H2O concentrations in the electrolyte, and electrolyte temperature is necessary to achieve straight pore growth induced from the depression patterns in the depth direction. The optimization of the anodization conditions enabled the formation of ordered iron oxide nanohole arrays with aspect ratios exceeding 10. The resulting ordered iron oxide nanohole arrays with high aspect ratios are expected to be applied to various functional devices such as photocatalysts and solar cells.
Ni/NiO Reference Electrode Potential Measurements in Molten CaCl2–CaO
Authors: Dale Olivia R. , Felling Forest , Gonzalez Mario , Zhang Chao , Simpson Michael F.
Published: June 2023

A direct oxide reduction (DOR) process is used to reduce PuO2 to Pu metal via Ca metal. Molten CaCl2 is commonly used as the electrolyte for this reaction. As the DOR reaction proceeds, CaO accumulates in the salt and must be measured in real-time to know when the reaction has gone to completion. Open circuit potential (OCP) was measured in this study between a reference electrode consisting of Ni/NiO contained in a MgO tube and a tungsten working electrode. Two methods of CaO additions were performed over several experiments, and both methods correlated to a 0.11 to 0.23 V decrease in OCP up to 14.4 wt% CaO. CaO concentrations were separately measured via sampling and acid-base titration. A plot of OCP versus concentration of CaO fits the Nernst equation with the CaO powder additions up to 5.2 wt% and also at the higher end of the CaO pellet concentration range. While the MgO tubes were observed to be structurally stable in the molten salt, scanning electron microscopy and energy dispersive X-ray spectroscopy showed that Mg in the tubes had been partially replaced with Ca.
Validation of the Use of Otoliths to Estimate Age and Growth of Larval Lake Whitefish, Coregonus clupeaformis
Authors: Dunlop Erin S. , Hébert Issac , Taylor Courtney
Published: June 2023

Understanding drivers of recruitment variation in fish populations requires research conducted on early life stages. Examination of fish otoliths provides useful information for estimating hatching dates, growth, and survival rates of larvae and for investigating the relationship between early life stage phenology and variation in environmental factors such as climate and food availability. In the Laurentian (i.e., North American) Great Lakes, significant reductions in the number of young (ages 1–4 years) lake whitefish (Coregonus clupeaformis) recruiting into the population and commercial fishery have raised questions about factors affecting growth and survival of the larval life stage. Here, we investigate the utility of using otoliths to estimate the age and growth of larval lake whitefish. We raised offspring of wild-caught parents from Lake Simcoe (Ontario, Canada) in a hatchery environment and analyzed otoliths of these known age fish for 75 days posthatch. We further examined otoliths of wild-sampled larvae and age 0 lake whitefish from Lake Huron. We found a strong linear relationship between known age and number of postcheck increments on the otolith and between growth of the otolith and fish length. Increments formed at nearly 1 (0.9) per day beginning at day 20 after hatch. Check and subsequent increment formation was associated with disappearance of the yolk sac. Wild fish had more prominent checkmarks and grew slower than hatchery fish. Thus, otolith analysis represents a promising tool to examine dynamics of early life stages of lake whitefish, although further research is required on the effects of environmental conditions on otolith microstructure.
Figure S2 from An Oncolytic Poxvirus Encoding hNIS, Shows Antitumor Efficacy and Allows Tumor Imaging in a Liver Cancer Model
Authors: Chaurasiya Shyambabu , Valencia Hannah , Zhang Zhifang , Kim Sang-In , Yang Annie , Lu Jianming , Woo Yanghee , Warner Susanne G. , Ede Nicholas J. , Fong Yuman
Published: June 2023

HMGB1 staining on tumor sections
Data from An oncolytic poxvirus encoding hNIS, shows anti-tumor efficacy and allows tumor imaging in a liver cancer model
Authors: Chaurasiya Shyambabu , Valencia Hannah , Zhang Zhifang , Kim Sang-In , Yang Annie , Lu Jianming , Woo Yanghee , Warner Susanne G. , Ede Nicholas J. , Fong Yuman
Published: June 2023

Abstract
Oncolytic viruses (OVs) are live viruses that can selectively replicate in cancer cells. We have engineered an OV (CF33) to make it cancer-selective through the deletion of its J2R (thymidine kinase) gene. Additionally, this virus has been armed with a reporter gene, human sodium iodide symporter (hNIS), to facilitate non-invasive imaging of tumors using positron emission tomography (PET). In this study we evaluated the oncolytic properties of the virus (CF33-hNIS) in liver cancer model, and its usefulness in tumor imaging. The virus was found to efficiently kill liver cancer cells and the virus-mediated cell death exhibited characteristics of immunogenic death based on the analysis of 3 damage associate molecular patterns (DAMPs): calreticulin, ATP and HMGB1. Furthermore, local or systemic administration of a single dose of the virus showed anti-tumor efficacy against a liver cancer xenograft model in mice and significantly increased survival of treated mice. Lastly, PET scanning was performed following injection of the radioisotope I-124, for imaging of tumors, and a single dose of virus as low as 1E03 pfu, administered intratumorally (I.T.) or intravenously (I.V.), allowed for PET imaging of tumors. In conclusion, CF33-hNIS is safe and effective in controlling human tumor xenografts in nude mice, and it also facilitates non-invasive imaging of tumors.
Data from An Oncolytic Poxvirus Encoding hNIS, Shows Antitumor Efficacy and Allows Tumor Imaging in a Liver Cancer Model
Authors: Chaurasiya Shyambabu , Valencia Hannah , Zhang Zhifang , Kim Sang-In , Yang Annie , Lu Jianming , Woo Yanghee , Warner Susanne G. , Ede Nicholas J. , Fong Yuman
Published: June 2023

Abstract
Oncolytic viruses (OV) are live viruses that can selectively replicate in cancer cells. We have engineered an OV (CF33) to make it cancer-selective through the deletion of its J2R (thymidine kinase) gene. In addition, this virus has been armed with a reporter gene, human sodium iodide symporter (hNIS), to facilitate noninvasive imaging of tumors using PET. In this study, we evaluated the oncolytic properties of the virus (CF33-hNIS) in liver cancer model, and its usefulness in tumor imaging. The virus was found to efficiently kill liver cancer cells and the virus-mediated cell death exhibited characteristics of immunogenic death based on the analysis of 3 damage-associated molecular patterns: calreticulin, ATP, and high mobility group box-1. Furthermore, local or systemic administration of a single dose of the virus showed antitumor efficacy against a liver cancer xenograft model in mice and significantly increased survival of treated mice. Finally, PET scanning was performed following injection of the radioisotope I-124, for imaging of tumors, and a single dose of virus as low as 1E03 pfu, administered intra-tumorally or intravenously, allowed for PET imaging of tumors. In conclusion, CF33-hNIS is safe and effective in controlling human tumor xenografts in nude mice, and it also facilitates noninvasive imaging of tumors.
Abstract 1082: Effect of doublet treatment versus single-agent treatment in gastric cancers with/without MET amplification or a MET exon 14 skipping mutation
Authors: Zang Dae Young , sohn sung-hwa , Sul Hee Jung , Kim Bum Jun
Published: June 2023

Although conventional combination chemotherapy for advanced gastric cancer (GC) is associated with increased survival, it has major adverse effects, which necessitate the development of more effective, less toxic treatments. Combinations of multiple anti-cancer drugs, such as paclitaxel plus ramucirumab, have recently been used for the second-line treatment of advanced GC. This study evaluated the MET copy number variation (CNV), mutations, and gene and protein expression in human GC cells and the differential susceptibility of GC cells to single (tepotinib, ramucirumab, or paclitaxel) or doublet (tepotinib plus paclitaxel or ramucirumab plus paclitaxel) regimens. Compared to ramucirumab plus paclitaxel, tepotinib plus paclitaxel inhibited the growth of GC cells with a MET exon 14 skipping mutation and those without MET amplification but with phosphorylated MET in a dose-dependent manner with a concomitant induction of cell death. Tepotinib plus paclitaxel and ramucirumab plus paclitaxel similarly inhibited the growth of GC cells without a MET amplification or MET phosphorylation in a dose-dependent manner without inducing cell death. However, tepotinib alone or tepotinib plus ramucirumab was more effective in c-MET-positive GC cells (CNV > 30) than ramucirumab alone, paclitaxel alone, or ramucirumab plus paclitaxel. Our in vitro findings suggest that, compared to ramucirumab plus paclitaxel, tepotinib plus paclitaxel inhibits the growth of c-MET-positive GC cells, GC cells without a MET amplification but with phosphorylated MET, and those with MET mutations. Tepotinib alone was as effective as tepotinib plus paclitaxel in c-MET-positive GC cells. Clinical studies are required to confirm the therapeutic effects of these regimens.

Citation Format: Dae Young Zang, sung-hwa sohn, Hee Jung Sul, Bum Jun Kim. Effect of doublet treatment versus single-agent treatment in gastric cancers with/without MET amplification or a MET exon 14 skipping mutation [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 1082.
Abstract 1083: MEKi and SOS1i as a combination treatment option for KRAS mutant endometrial cancer
Authors: Kostyrko Kaja , Hinkel Melanie , Hofmann Marco H. , Trapani Francesca
Published: June 2023

Endometrial cancer (EC) is the most common cancer of the female genital tract, resulting in over 75,000 deaths per year worldwide. Despite recent advances in medical oncology research, chemo- and radiotherapy are still the standard of care treatments for recurrent or metastatic EC and prognosis for these patients remains poor, with median survival rates of less than one year. Approximately 15-30% of the more common type I EC harbors oncogenic KRAS mutations, which have been linked to disease progression. Preclinical studies have shown that MEK inhibitors exhibit strong anti-tumor activity in KRAS mutant EC models. In the clinical setting however, MEKi monotherapy has proven unsuccessful in KRAS mutant EC patients, suggesting that combinations with other targeted agents may be needed to achieve increased signaling modulation and enhanced therapeutic activity. As suggested by our previous work in KRAS mutant lung, colorectal and pancreatic cancer, the combined inhibition of MEK and SOS1 may overcome adaptive resistance to MEKi by blocking SOS1-mediated reactivation of MAPK signaling. We evaluated whether this hypothesis can also be applied in the context of EC. Here we treated preclinical models of EC with a combination of clinically relevant doses of a MEKi (trametinib) and BI-3406, a potent and selective orally bioavailable SOS1 inhibitor. Treatment of KRAS mutant EC cells resulted in decreased activity of the KRAS-MAPK pathway, as measured by ERK phosphorylation. This effect was further enhanced by co-treatment with BI-3406. In vivo, the combination of MEKi with SOS1i synergistically decreased tumor growth in two KRAS G12D mutant orthotopic patient-derived xenograft models of EC. The anti-tumor effect of the combination was comparable to the effect of paclitaxel, the standard of care treatment in EC. This suggests that the combination of MEKi with SOS1i may be a potential therapeutic approach for the treatment of EC patients with tumors harboring KRAS mutations.

Citation Format: Kaja Kostyrko, Melanie Hinkel, Marco H. Hofmann, Francesca Trapani. MEKi and SOS1i as a combination treatment option for KRAS mutant endometrial 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 1083.
Abstract 1088: Acquired radiation resistance induces thiol-dependent cisplatin cross-resistance
Authors: Jenkins Samir V. , Shah Shruti , Jamshidi-Parsian Azemat , Mortazavi Amir , Boysen Gunnar , Vang Kieng B. , Griffin Robert J. , Rajaram Narasimhan , Dings Ruud P.
Published: June 2023

Purpose. Resistance to radiation, both intrinsic and acquired, remains a significant clinical challenge. It is therefore import to identify the underlying molecular and cellular features involved in this resistance.

Materials and Methods. We generated genetically-matched counterparts of various degrees of radiation sensitivity in non-small cell lung carcinoma (NSCLC) tumor models to define characteristics of acquired resistance through repeated exposures to conventional X-ray radiation.

Results. As assessed by cell viability and clonogenic assays murine Lewis lung carcinoma (LLC) and human A549 cell lines acquired an approximate 1.5 - 2-fold increase in radiation resistance (RR) as compared to its parental counterpart. Morphologically, we found that A549-RR exhibited a greater nucleus-to-cytosol (N/C) ratio as compared to its respective counterpart. Since the N/C ratio has been linked to the differentiation state, we next investigated the epithelial-to-mesenchymal transition (EMT) phenotype and cellular plasticity. We found that A549 have a greater radiation-induced plasticity, as measured by E-cadherin, vimentin and double positive (DP) modulation, as compared to LLC. Additionally, migration was suppressed in A549-RR, as compared to A549. Subsequently, we confirmed that the LLC-RR and A549-RR are also more resistant to radiotherapy than their isogenic-matched counterpart in vivo. Remarkably, we found that the acquired radiation resistance also induced resistance to first-line chemotherapeutic cisplatin, but not carboplatin or oxaliplatin. This cross-resistance was attributed to induced elevated thiol levels. Decreasing the amount of thiols with buthionine sulfoximine (BSO), a gamma-glutamylcysteine synthetase inhibitor, sensitized the resistant cells to cisplatin, to levels similar as parental cells.

Conclusions. By generating genetically-matched radiation resistant NSCLC tumor models we were able to identify and overcome cisplatin cross-resistance. This is an important finding arguing for multi-modality treatment regimens with the potential of improving clinical outcomes in the future.

Citation Format: Samir V. Jenkins, Shruti Shah, Azemat Jamshidi-Parsian, Amir Mortazavi, Gunnar Boysen, Kieng B. Vang, Robert J. Griffin, Narasimhan Rajaram, Ruud P. Dings. Acquired radiation resistance induces thiol-dependent cisplatin cross-resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1088.
Abstract 1097: Surufatinib improved the radiation sensitivity of biliary cancer growth by reprograming of TAM and anti-angiogenesis
Authors: Ma Hong , Huang Ai , Xiao Yong , Yao Jing , Wang Ningyu , Mei Xiangping , Wen Shaoshi , Lv Xing
Published: June 2023

Background: Patients with Biliary tract cancer (BTC) have poor clinical outcomes. Radiation therapy is an adjunct strategy throughout biliary tract cancer treatment. The aim of the study was to explore the efficacy and mechanism of the radiosensitization of surufatinib(SF), an oral tyrosine kinase inhibitor with anti-angiogenic and immunomodulatory activities.

Methods: The NOZ and TFK1 human BTC cell lines were treated with SF and/or radiation. Effects on cell viability and cell cycle progression were measured by CCK8, a clonogenic survival assay, and flow cytometry. Finally, the in vivo effect on the growth of NOZ xenografts was assessed in nude mice. F4/80, CD206, Ki67 and CD31 expression were employed in tumor samples.

Results: SF inhibited the proliferation and enhanced the radiosensitivity in NOZ and TFK1 cells lines. SF combined with radiation increased cell cycle arrest in the G2/M phase compared to the other treatments in the NOZ cell line (P<0.05). Similarly, cell cycle arrest occurred in the TFK1 cell line, (P<0.05). In vivo, SF plus radiation significantly inhibited the tumor growth compared to either agent independently.

Conclusions: SF significantly increased the radiosensitivity of NOZ and TFK1 cells in vitro and in vivo. Radiation combined with SF inhibited tumor proliferation, increased apoptosis, prolonged G2/M arrest. SF decreased the number of blood vessels and reprogrammed the macrophages from M2 to M1. These data support the clinical trials of biologically targeted and conventional therapies in the treatment of cancer.

Citation Format: Hong Ma, Ai Huang, Yong Xiao, Jing Yao, Ningyu Wang, Xiangping Mei, Shaoshi Wen, Xing Lv. Surufatinib improved the radiation sensitivity of biliary cancer growth by reprograming of TAM and anti-angiogenesis [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 1097.
Abstract 1124: PTEN loss confers resistance to anti-PD-1 therapy in NSCLC by increasing tumor infiltration of T regulatory cells
Authors: Exposito Francisco , Redrado Miriam , Houry Maeva , Hastings Katherine , Molero-Abraham Magdalena , Lozano Teresa , Solorzano Jose L. , Ortega Julian Sanz , Andradas Vera , Amat Ramon , Redin Esther , Leon Sergio , Legarra Naroa , García Javier , Serrano Diego , Valencia Karmele , Robles-Oteiza Camila , Foggetti Giorgia , Felip Enriqueta , Lasarte Juan J. , Paz-Ares Luis , Zugazagoitia Jon , Politi Katerina , Montuenga Luis M , Calvo Alfonso
Published: June 2023

Immune checkpoint inhibitors (ICIs) targeting PD-L1/PD-1 have dramatically improved the management of NSCLC patients. Nevertheless, more than 50% patients will present primary resistance or will develop secondary resistance within 12-25 months after treatment initiation. The mechanisms of resistance to ICIs may depend on tumor-intrinsic molecular/genetic alterations that generate an immunosuppressive tumor microenvironment (isTME). Using In silico analyses (TCGA and AACR-GENIE cohorts), and protein expression analysis in our institutional CIMA-CUN cohort, we have shown that >50% LUSC and ~28% LUAD tumors exhibited reduced PTEN mRNA/protein levels. Patients with PTEN-low tumors had higher levels of CXCL10, PD-L1 and PD-L2. In a cohort of stage IV NSCLC patients treated with ICIs, we found that patients with low tumor PTEN levels were significantly associated with worse OS (p=0.021). We have developed and characterized a Pten-null LUSC model using UN680 cells, which was also refractory to anti-PD-1 when implanted in syngeneic mice. Pten loss led to a deregulation of multiple pathways related to immunosuppression, with a positive enrichment in “IL-6/JAK/STAT3” and “TNF-α via NFkB” hallmarks, identified by GSEA. Tumors from the Pten-null LUSC model were highly metastatic and fibrotic, and secreted TGF-β/CXCL10, thus favoring conversion of CD4+ lymphocytes into Tregs. Human/mouse PTEN-low tumors were enriched in Tregs and immunosuppressive genes. Importantly, treatment of mice bearing Pten-null tumors with TLR agonists+anti-TGF-β, aimed to alter this isTME, led to tumor rejection and immunological memory in 100% of mice. Our results demonstrate that lack of PTEN causes immunotherapy resistance in LUSC by establishing an isTME that can be reversed therapeutically.

Citation Format: Francisco Exposito, Miriam Redrado, Maeva Houry, Katherine Hastings, Magdalena Molero-Abraham, Teresa Lozano, Jose L. Solorzano, Julian Sanz Ortega, Vera Andradas, Ramon Amat, Esther Redin, Sergio Leon, Naroa Legarra, Javier García, Diego Serrano, Karmele Valencia, Camila Robles-Oteiza, Giorgia Foggetti, Enriqueta Felip, Juan J. Lasarte, Luis Paz-Ares, Jon Zugazagoitia, Katerina Politi, Luis M Montuenga, Alfonso Calvo. PTEN loss confers resistance to anti-PD-1 therapy in NSCLC by increasing tumor infiltration of T regulatory cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1124.
Abstract 1329: Aging increases malignant peritoneal mesothelioma metastasis
Authors: Miller Mykayla
Published: June 2023

Mesothelioma is a rare cancer with approximately 3,000 new cases each year. There are three types of mesotheliomas including pleural (75%), pericardial (>1%) and peritoneal (20%). Mesothelioma occurs in the mesothelial cells, which form a protective barrier to multiple organs. Malignant peritoneal mesothelioma (MPM) is initiated in the peritoneum, a serous membrane that lines the inner walls of the abdominal cavity and the outside of the visceral organs and is a very receptive environment for malignancy and metastasis. MPM metastasizes to the abdominal cavity, affecting the omentum, liver, stomach, intestines, and kidneys. MPM is extremely difficult to diagnose due to its vague symptoms, such as abdominal distension, abdominal pain, weight loss, and nausea. Due to difficulty in diagnosis, most patients are diagnosed at an advanced disease (stage III or IV) with a very poor prognosis with a 9% 5-year survival rate. Asbestos exposure is a major risk factor for MPM with symptoms taking 20-50 years post- exposure to appear, making MPM prominent in aged adults. Mesothelin (MSLN), a 40 kDa protein, is normally expressed on the surface of mesothelial cells and manifests at abnormally high levels in several carcinomas including malignant peritoneal mesothelioma. The exact role of mesothelin is not yet fully understood as mesothelin knockout mice do not display an abnormal phenotype. However, there is some evidence to suggest that mesothelin is involved in tumor cell adhesion by binding to the tumor antigen MUC16. Mesothelin wild-type (WT) and knockout (KO) mice were used to investigate the role of host mesothelin expression on MPM metastasis. A MPM cell line was tagged with nuclear red fluorescence protein (RFP) and injected into the abdomen of female C57Bl/6, young (3-6 months) and aged (18-23 months), MSLN WT or KO mice. Disease progression was evaluated post injection of tumor cells by fluorescent in vivo imaging prior to end point dissection. The abdominal cavity was imaged in situ and tumor burden was evaluated using Image J. Overall, aged mice had increased abdominal tumor burden regardless of host MSLN expression. Aged MSLN KO mice displayed an increase in abdominal tumor burden compared to aged MSLN WT mice, however this difference was not significant. Interestingly, the immune profile of the abdominal cavity of tumor naive mice displays a decrease in natural killer (NK) cells and peritoneal macrophages in aged mice suggesting a role in regulating metastatic peritoneal mesothelioma and could possibly be targets for new immunotherapies. This study demonstrates host age increases tumor growth and metastatic success in peritoneal mesothelioma.

Citation Format: Mykayla Miller. Aging increases malignant peritoneal mesothelioma metastasis [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 1329.
Abstract 1332: Extracted mRNA from frozen buffy coat samples shows promise for use in downstream sequencing analyses
Authors: Ylitalo Erik Bovinder , Vidman Linda , Harlid Sophia , Van Guelpen Bethany
Published: June 2023

Frozen blood samples are routinely collected and stored in medical biobanks across the world, often for periods of several decades, and later used for medical research such as biomarker studies. Some common biomarker targets have included DNA alterations, protein or antibody levels or metabolomics. However, despite the potential importance of gene-expression alterations in circulating immune cells, transcriptional studies on blood samples from biobanks have been limited due to preanalytical technical difficulties of extracting high-quality RNA. In the absence of an RNA preservative, RNA degrades rapidly during the thawing process. Our aim was, therefore, to investigate the possibility of using biobanked blood samples for transcriptomics analyses. First, we used a CryoXtract CXT 350 to establish a protocol capable of isolating high-quality RNA from frozen core aliquots of buffy coat without thawing the entire sample tube. Then, we performed RNA sequencing on RNA extracted from 20 pre-diagnostic buffy coat samples from colorectal cancer patients and matched controls included in the population-based, prospective Northern Sweden Health and Disease Study (NSHDS). For all samples, two different library preparation methods were used (Illumina TruSeq Stranded mRNA poly-A selection and Illumina Stranded Total RNA with Ribo-Zero Globin). We obtained RNA yields of more than 1 µg from a majority of the NSHDS samples (mean=2.57 µg), and more than 92% of samples had RIN values of 6 or higher, indicating suitability for downstream analyses. Preliminary random forest analyses showed expected discriminatory ability between men and women based on data generated from each of the two library preparation methods. In conclusion, our newly established RNA extraction method enables extraction of RNA from frozen, biobanked buffy coat samples, collected in standard tubes, that is of sufficient quantity and quality for use in downstream whole-genome transcriptomic analyses, such as RNA sequencing.

Citation Format: Erik Bovinder Ylitalo, Linda Vidman, Sophia Harlid, Bethany Van Guelpen. Extracted mRNA from frozen buffy coat samples shows promise for use in downstream sequencing analyses [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 1332.
Abstract 1407: Refining endometrial carcinoma of no-specific molecular subtype by sources of estrogen signaling
Authors: Lam Clarissa , Aghajanian Carol , Abu-Rustum Nadeem , Weigelt Britta , Ellenson Lora , Boroujeni Amir Momeni
Published: June 2023

Introduction: Endometrial carcinomas (ECs) of no-specific molecular profile (NSMP-EC) are a large and heterogenous group of tumors. These cancers, in particular endometrioid carcinomas, are thought to be dependent on or benefit from estrogen receptor (ER) signaling. In this study, we sought to determine the sources of ER signaling in patients with NSMP-EC.

Methods: We compiled a retrospective database of EC patients who received their primary treatment at our institution from 2014 to 2022, and who had ECs of NSMP based on a surrogate of The Cancer Genome Atlas (TCGA) prognostic EC molecular subtype. We used ovarian volume (binarized using a cutoff of 40 cm3), menopausal status, body mass index (BMI>25), and ESR1 hotspot mutations as surrogate markers for ER signaling.

Results: A total of 171 patients with NSMP-EC were included in this study. As expected, menopausal status was correlated with ovarian volume (right ovarian volume median 12.3 cm3 vs 32.8 cm3, p=0.001; left ovarian volume median 10.6 cm3 vs 37.7 cm3, p<0.001). Of these 171 patients, 135 (79%) had an identifiable source of ER signaling. Tumors with ER signaling sources tended to be endometrioid carcinomas (125/135, 93%) whereas a lower proportion of ECs without ER signaling were of endometrioid histology (24/36, 67%; p=0.001). ESR1 hotspot mutations almost exclusively occurred in post-menopausal (11/12 mutated cases) and older patients (median age 69 vs 61 years, p=0.012). PTEN mutations tended to be more frequent in ECs with ER signaling source (70% vs. 50%, P=0.036); in contrast, KRAS hotspot mutations were more common in cases without an identifiable ER source (42% vs. 24%, p=0.067). Additionally, the presence of PTEN mutations was associated with a higher BMI (median 31.1 vs 28.3, p=0.012), whereas FBXW7 mutations were associated with a lower BMI (median 24 vs 30.6, p=0.047). Log-rank survival analysis showed that patients whose ECs lacked an identifiable ER signaling source had a markedly worse survival outcome (p=0.0024).

Discussion: We demonstrate that the majority of NSMP-ECs have an identifiable ER signaling source. The minority of NSMP-ECs lacking an ER signaling source may not only represent a hormonally independent tumorigenesis but are also markedly distinct at the molecular, pathologic, and clinical levels. ER protein immunohistochemistry may help in further defining these groups.

Citation Format: Clarissa Lam, Carol Aghajanian, Nadeem Abu-Rustum, Britta Weigelt, Lora Ellenson, Amir Momeni Boroujeni. Refining endometrial carcinoma of no-specific molecular subtype by sources of estrogen signaling [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 1407.
Abstract 1535: A novel ELISA Kit:highly accuracy and sensitivity for GDF-15 detection
Authors: Liang Meng , He Feng , Zhang Meng , Ye Yinsong , Chen Shukun , Ning Jinying , Hao Feng
Published: June 2023

Growth/differentiation factor 15 (GDF-15) is a promising drug target and biomarker for cancer early screening or immunotherapy. Under normal conditions, GDF-15 is expressed in low concentrations in most organs. Increased protein levels are associated with disease states such as tissue hypoxia, inflammation, acute injury and oxidative stress. The development of GDF-15 detection kit can help pharmaceutical companies in the early stage of drug development. It can also facilitate the early screening of related diseases. Hence, we developed a novel GDF-15 detected ELISA kit based on double-antibody sandwich ELISA. Our kit has ultra-high sensitivity and specificity and the concentration of GDF-15 can be detected accurately in complex biological samples such as serum, plasma or urine. First of all, mice were immunized by GDF-15. After detecting the antibody titers in sera of the immunized mice, hybridoma technologies were used to produce GDF-15 monoclonal antibodies. 57 antibodies were analyzed by epitope competition, ELISA binding and KD binding detection. The specificity of pairwise candidates was screened to obtain optimal coating and detecting antibody pair. The detecting antibody was labeled with biotin which can be conjugated with streptavidin labeled HRP. Biotin-streptavidin labeling detection method can achieve signal amplification which offers an improved detection sensitivity. Our GDF-15 detection kit has better property in sensitivity and specificity. The detection range is from 10 pg/mL to 10000 pg/mL. LOD is 5 pg/mL and LOQ is 10 pg/mL. These parameters are better than many commercial test kits. Moreover, the concentration of GDF-15 can be detected accurately in complex clinical samples. We anticipate that this GDF-15 detection kit can be applied not only for companion diagnosis, but also for related drug development.

Citation Format: Meng Liang, Feng He, Meng Zhang, Yinsong Ye, Shukun Chen, Jinying Ning, Feng Hao. A novel ELISA Kit:highly accuracy and sensitivity for GDF-15 detection [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 1535.
Abstract 1536: Machine learning guided engineering of nivolumab for superior physicochemical properties for subcutaneous injection
Authors: Hamamatsu Norio , Kuwabara Naoyuki , Suzuki Emi , Yamazaki Ryo , Oikawa Misaki , Tawara Tomonori , Ito Tomoyuki , Nakazawa Hikaru , Kataoka Shiro , Umetsu Mitsuo
Published: June 2023

Nivolumab is a programmed death receptor-1 (PD-1) blocking antibody that is used as a type of cancer immunotherapeutic agent, called a checkpoint inhibitor. Nivolumab is administered by intravenous infusion. Given the recent studies and potential demand for maintenance immunotherapy, the next generation of Nivolumab for subcutaneous injection would be beneficial for patients’ quality of life. We studied whether we could create a Nivolumab variant that has superior physicochemical properties required for a subcutaneous formulation. A machine-learning guided protein engineering technique was used to improve structural stability at little cost of affinity. As a result, a variant showed an increased melting temperature by 10 ºC with a comparable affinity to Nivolumab. Consistent with the result, the variant is resistant to aggregation under heat stress. Nivolumab is seriously aggregated after treatment at 60 ºC, while the variant does not show any aggregation at all. We also present a result of risk assessment of aggregation at high concentrations.

Citation Format: Norio Hamamatsu, Naoyuki Kuwabara, Emi Suzuki, Ryo Yamazaki, Misaki Oikawa, Tomonori Tawara, Tomoyuki Ito, Hikaru Nakazawa, Shiro Kataoka, Mitsuo Umetsu. Machine learning guided engineering of nivolumab for superior physicochemical properties for subcutaneous injection [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 1536.
Abstract 1668: Targeting drug-induced tolerant & resistant cell populations by novel ferroptosis inducer
Authors: Dubash Taronish , Munteanu Maria Cristina , Farah Shrouq , Nunez Cristian , Jadin Laurence , Radetich Branko , Schmidt Darby , Gurvich Nadia
Published: June 2023

A subset of tumor cells escapes apoptotic and necroptotic death when subjected to chemo- or targeted therapy thereby entering a transient reversible less-proliferative “tolerant state.” Upon continuous drug treatment, these tumor cells become drug resistant, leading to metastases and cancer relapse. A long-standing goal in oncology has been to develop therapeutics to effectively target both cell states. Growing published evidence suggests that accumulation of reactive oxygen species and lipid remodeling make tolerant and resistant tumor cells more vulnerable to ferroptotic mode of cell death. To this end, we here characterize the activity of a proprietary inhibitor of GPX4, an enzyme that safeguards against ferroptosis, across a panel of tolerant and resistant cell line models. Drug tolerant and resistant cancer cell lines spanning three indications were generated by applying continuous treatments with relevant standard-of-cares (SOC) in vitro for short or long durations, respectively. Age-matched control cell lines were derived in parallel by treatment of parental cells with DMSO. Cells were subjected to high dose of SOC for nine days to generate tolerant lines, or to increased therapeutic doses of SOC over three months to generate resistant lines, after which resistance to SOCs was confirmed. In accordance with our hypothesis, we found that drug tolerant lines demonstrated enhanced sensitivity to GPX4 inhibition compared to age-matched control lines. Furthermore, we noted that resistant tumor cell lines retained exquisite sensitivity to GPX4 inhibition across all indications while demonstrating up to 1000-fold resistance to SOCs. In summary, we found that cell line models which evade cell death upon treatment with chemo- or targeted therapies continued to present vulnerability to ferroptosis induction across both cell states. Given the heterogeneity in responses to drug-induced cell-states, understanding the mechanisms of differential drug sensitivity to GPX4 inhibition may help in formulating successful strategies for targeting this relevant cell population in the clinic, thereby limiting cancer progression and/or improving curative potential.

Citation Format: Taronish Dubash, Maria Cristina Munteanu, Shrouq Farah, Cristian Nunez, Laurence Jadin, Branko Radetich, Darby Schmidt, Nadia Gurvich. Targeting drug-induced tolerant & resistant cell populations by novel ferroptosis inducer [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 1668.
Abstract 1692: mTOR inhibition attenuates chemosensitivity through induction of a persister state
Authors: Liu Yuanhui , Azizian Nancy , Sullivan Delaney , Li Yulin
Published: June 2023

Effective cancer therapeutics often kill a large fraction of tumor cells, while sparing a small surviving population, which can resurface as future tumor recurrence. Mechanisms of resistance in these residual tumors may involve rare preexisting clones with mutations, as well as de novo development of resistance through various genetic and/or non-genetic processes. In the context of acute stress resulting from cancer therapeutics, the initial adaptation is essential for tumor cell survival and allows for subsequent development of resistance through additional genetic and non-genetic mechanisms. As an integrator of environmental signals and cellular response, the mTOR pathway is a critical driver of cellular adaptation to environmental changes. In particular, mTOR regulates embryonic diapause, a reversible dormancy state in mammalian development in response to unfavorable environments. Analogous to its role in regulating embryonic diapause, mTOR inhibition may induce a diapause-like state in cancer cells, leading to drug tolerance and persistence of residual tumors following therapy. In line with this notion, recent studies have reported markedly reduced mTOR activity in residual tumors. However, the causal relationship between mTOR inhibition and chemosensitivity has not been investigated. Through a genome-wide CRISPR knockout library screen in pancreatic cancer cells treated with chemotherapeutic agents, we have identified the mTOR pathway as a prominent determinant of chemosensitivity. Pharmacological suppression of mTOR activity in cancer cells from diverse tissue origins leads to the persistence of a reversibly resistant population, which is otherwise eliminated by chemotherapeutic agents. Conversely, activation of the mTOR pathway increases chemosensitivity and predicts better survival among various human cancers. Survival of the persisters is dependent on the regulation of autophagy, G2/M checkpoint, and cell death pathways, as revealed by a small-molecule chemical library screen. Thus, mTOR plays a causal yet paradoxical role in regulating chemotherapeutic response; inhibition of the mTOR pathway, while suppressing tumor expansion, facilitates the development of a reversible drug-tolerant persister state.

Citation Format: Yuanhui Liu, Nancy Azizian, Delaney Sullivan, Yulin Li. mTOR inhibition attenuates chemosensitivity through induction of a persister state [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 1692.
Abstract 1695: Guanylate binding protein 1 modulate the proteasomal machinery and causes the cytochrome C degradation
Authors: Tailor Dhanir , Garcia-Marques Fernando Jose , Bermudez Abel , Pitteri Sharon J. , Malhotra Sanjay V.
Published: June 2023

Guanylate Binding Protein 1 (GBP1) is known as an interferon γ induced GTPase. It plays a crucial role in innate immunity against pathogenic infections but its role in cancer and therapeutic response is not well understood. Therefore, to study the role of GBP1, we investigated genetically manipulated ovarian cancer cells. We have employed in vitro and in vivo studies along with a proteomics approach to uncover the protein-protein interactions and their crosstalk with GBP1. The data suggest that GBP1 knockdown (KD) inhibits the clonogenic potential of cancer cells. In vivo studies including subcutaneous xenograft and intraperitoneal models revealed that GBP1 overexpressing (OV) tumor cells growing faster and reduce the median survival of animals compared to vehicle control (VC). Whereas GBP1 KD tumors were progressing slowly with longer median survival compared to scrambled control (SC). To study the molecular interactions of GBP1, we performed a proteomics-based Cellular thermal shift assay (CETSA) on GBP1 OV and KD ovarian cancer cells. We found that GBP1 interacts with members of proteasome including PSMB1, PSMB6, PSMA3, and Cytochrome C (CYCS). Protein analysis showed that GBP1 over-expressing cells had less ubiquitinylated proteins compared to VC, whereas GBP1 KD cells accumulate the ubiquitinylated proteins compared to SC. We also found that the GBP1 KD cells are sensitive to paclitaxel treatment and accumulated ubiquitinylated proteins compared to SC. Moreover, GBP1 KD inhibits the overall proteasomal activity. Paclitaxel-treated GBP1 OV cells accumulated the ubiquitination of CYCS, followed by clearance of cytosolic CYCS. Furthermore, global proteome profiling also suggests that GBP1 KD induces apoptotic pathways whereas GBP1 OV induces pathways associated with the stress response. Overall, the results from our studies elucidate the role of GBP1 in chemotherapeutic stress response via the activation of proteasomal machinery.

Citation Format: Dhanir Tailor, Fernando Jose Garcia-Marques, Abel Bermudez, Sharon J. Pitteri, Sanjay V. Malhotra. Guanylate binding protein 1 modulate the proteasomal machinery and causes the cytochrome C degradation [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 1695.
Abstract 1708: Sex-biased function of an oncogene, YAP1, regulates medulloblastoma stem cells and immune evasion
Authors: Abdelfattah Nourhan , Natarajan Sivaraman , Maldonado Jose , Tran Han Nhat , Borland Hannah , McMinimy Rachael , Chen Shu-Hsia , Camargo Fernando , Olson James , George Joshy , Yun Kyuson
Published: June 2023

Sex disparity in both cancer incidence and response to therapeutics, especially immunotherapy, has become an important area of research in recent years. Brain tumors show obvious sex biases in incidence and therapy response. Medulloblastoma (MB) is the most common malignant pediatric brain tumor, and similar to other brain tumors, males have consistently higher incidence across all subgroups and at all ages. Moreover, for patients >3 years of age, females have significantly better outcomes than males. However, molecular drivers of such sex disparity in incidence and prognosis remain unclear.Here, we report a male-biased function of an oncogene, YAP1, in MB formation and immune suppression. Integrated multi-platform genomic, transcriptomic, proteomic, and in vivo transcriptional target analyses show that YAP1 promotes cancer stem cell maintenance through concurrent transcriptional activation of stemness genes, such as Sox2, and repression of differentiation regulators such as NeuroD1 and Zic1/2. YAP1 expression in MB cells also inhibits anti-tumor immune cell infiltration by directly regulating Csf1, Igf1, and Igfbp3 transcription and modulating IL6-JAK-STAT3, TNFR1, TGFb, and CCL5 Pathways. Notably, we found that YAP1 genetic deletion results in disproportionately higher survival of males than females in a spontaneous mouse model of infantile SHH MB, despite equivocal expression levels in both sexes. Consistent with our findings in mouse models, YAP1 downstream genes stratify MB patient survival in males but not females. Finally, we show that a brain-penetrant YAP1 inhibitor, verteporfin, extends survival and enhances immune infiltrates in vivo, mimicking the effects of Yap1 genetic deletion. In summary, this study demonstrates that an oncogene function may be sex-biased and underscores the need to incorporate sex as a variable in cancer mechanisms, tumor immunology, and treatment response studies.

Citation Format: Nourhan Abdelfattah, Sivaraman Natarajan, Jose Maldonado, Han Nhat Tran, Hannah Borland, Rachael McMinimy, Shu-Hsia Chen, Fernando Camargo, James Olson, Joshy George, Kyuson Yun. Sex-biased function of an oncogene, YAP1, regulates medulloblastoma stem cells and immune evasion [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 1708.
Abstract 1709: MAPK pathway alterations are a targetable vulnerability in bladder cancer
Authors: Bekele Raie , Samant Amruta , Hanlon Timothy , Mouw Kent
Published: June 2023

Bladder cancer (BC) is the fourth most common cancer in males and accounts for about 80,000 new cases in the US. Approximately 25% patients with BC will present with muscle-invasive bladder cancer (MIBC) and for the past several decades the standard-of-care for MIBC and metastatic bladder cancer has been cisplatin-based chemotherapy. More recently, multiple immunotherapeutic agents have been approved but only a subset of patients respond to these treatments and therefore novel therapeutic strategies are needed to expand treatment options and improve clinical outcomes for BC patients. Large-scale genomic studies of MIBC have identified numerous recurrent genomic alterations including mutations, copy number alterations, and chromosomal translocations. Using publicly available patient datasets, we identified ~20% of MIBC patients harboring alterations in genes involved in the mitogen-activated protein kinase (MAPK) pathway and their upstream activators. RAF1 copy number amplification was the most predominant alteration followed by activating HRAS mutations. Leveraging our institutional cohorts of bladder cancer specimens, we defined the frequency of these alterations and validated RAF1 amplification in a subset of cases using fluorescence in situ hybridization. To functionally define the role of MAPK pathway alterations in BC, we tested activity of RAF and MEK inhibitors in BC cell line models with diverse MAPK pathway alterations including RAF1 amplification and HRAS/NRAS mutations. We found that these MAPK pathway altered models displayed increased sensitivity to RAF/MEK inhibition compared to MAPK-unaltered models. We further demonstrated the feasibility of preclinically targeting these alterations in mouse models of BC using cell line and patient-derived-xenografts. Additionally, we tested the impact of MAPK pathway modulation on tumor extrinsic properties including changes in the tumor microenvironment that could potentially affect responses to immunotherapeutic agents. Taken together, we have identified MAPK pathway alterations as a novel dependency in BC that can be targeted with emerging RAF inhibitors.

Citation Format: Raie Bekele, Amruta Samant, Timothy Hanlon, Kent Mouw. MAPK pathway alterations are a targetable vulnerability in bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1709.