bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 228: Recurrent mitochondrial mutations in thyroid cancer
Authors: Hopkins Julia F. , Fleischmann Zoe , Jin Dexter X. , Killian Jonathan Keith , Montesion Meagan
Published: April 2023

Despite the importance of mitochondrial function on the bioenergetics and metabolism of the cell, understanding the functional impact of somatic mitochondrial mutations has been difficult. Additionally, small cohorts, as used in previous studies, lack the power to estimate the prevalence of specific recurrent mutations.

We evaluated 177540 advanced cancer patients, across 84 different tumor types, who were sequenced as part of routine clinical care on the FoundationOne®CDx platform. Off-target sequencing reads were mapped to the mitochondrial genome (rCRS) and variant calling was performed. We had a median read depth of 18 and filtering by allele frequency (<= 95% and >= 5%) led to the identification almost 400,000 predicted somatic mitochondrial variants. Over 80% of patients had at least one mitochondrial variant detected, with an average of 2.2 mitochondrial variants detected per patient.

We next assessed each recurrent mitochondrial short variant to look for associations with specific cancers. The alteration with the strongest enrichment was G3224A, a mutation in the mitochondrial leucine tRNA gene (MT-TL1), which had a prevalence of 3.6% (51/1426) in thyroid cancer samples, as compared to 0.05% (85/176114) in all other disease types (OR=76.8, P<2.2e-16). The G3244A mutation has been previously identified as occurring in thyroid cancer, specifically Hurthle cell carcinomas, as well as in the mitochondrial disease MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes). Thyroid cancer samples in our dataset with G3244A were found to be associated with thyroid follicular carcinomas (7/124, 5.6%) and Hurthle cell carcinomas (7/57, 12.2%). Additional recurrent MT-TL1 mutations were identified in the thyroid cancer cohort at positions 3242 (9/1426) and 3248 (5/1426).

We also identified a mutation in the NADH dehydrogenase subunit MT-ND1. G3380A has been previously shown to occur in thyroid and colorectal cancer and results in a R25Q mutation that is thought to affect ND1 function. G3380A was identified in 26/1426 (1.8%) of thyroid cancer patients. The majority of G3380A variants occurred in thyroid anaplastic carcinomas 23/26 (88.5%).

Our dataset has allowed us to identify and determine the prevalence of rare mitochondrial short variant alterations in cancer. Further research is needed to investigate associations of these alterations with other clinical features.

Citation Format: Julia F. Hopkins, Zoe Fleischmann, Dexter X. Jin, Jonathan Keith Killian, Meagan Montesion. Recurrent mitochondrial mutations in thyroid 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 228.
Abstract 2280: BG34-200 immunotherapy of advanced melanoma
Authors: Roche Veronique P. , Sandoval Victor , Senders Zachary , Lyons Joshua , Wolford Claire , Zhang Mei
Published: April 2023

Background: High levels of myeloid derived cells are characteristic of tumor microenvironment (TME) of advanced melanoma. These cells work with the tumor cells to suppress host development of anti-tumor immune responses, regulate tumor metastasis, and drive cancer drug resistance to virtually all types of therapy. Therefore, methods to disrupt tumor-associated myeloid cell function are actively being sought to find a cure.

Goals: Our team has recently developed a plant-derived carbohydrate molecule, BG34-200, that modulates tumor-associated myeloid cells by targeting the cell surface receptor CD11b. In this study, we investigated the immune activation induced by BG34-200-CD11b engagement in the context of advanced melanoma and use the knowledge to develop BG34-200-based combination immunotherapy.

Methods: Wild type B57Bl6/J mice were injected with 1.5million B16F10 cells and treated intravenously with BG34-200. Tumor volume and animals survival were monitored, and whole blood was drawn for FACS analysis.

Results: We found that BG34-200 IV administration could significantly inhibit tumor growth and improve survival in B16F10 mice with advanced melanoma. Our data supported a model that the entry of BG34-200 into circulating melanoma tumor-associated inflammatory monocytes (TAIMs) could trigger a sequential immune activation: the BG34-200+ TAIM subsets migrated to tumor and differentiated into monocyte-derived dendritic cells (mo-DCs); then, the BG34-200+ mo-DCs migrated to tumor draining lymph nodes where they triggered the generation of tumor antigen specific T cells. Based upon these results, we combined BG34-200 treatment with adoptive transfer of TdLN-derived T-cell to treat advanced melanoma, which significantly improved animal survival and helped tumor-free survivors be resistant to a second tumor cell challenge.

Conclusions: The scientific findings from this study will allow us to develop new technology to apply BG34-200-based immunotherapy to patients with advanced melanoma who have not responded to current standard of care therapies with and without immunotherapy.

Citation Format: Veronique P. Roche, Victor Sandoval, Zachary Senders, Joshua Lyons, Claire Wolford, Mei Zhang. BG34-200 immunotherapy of advanced melanoma [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 2280.
Abstract 2339: Exploration of multiple immunotherapy modalities in ostesarcoma
Authors: Ismail Aliaa A. , Patton Daniel J. , Higgins Theresa A. , Sandey Maninder , Smith Bruce F. , Agarwal Payal
Published: April 2023

Osteosarcoma (OS) is the most common primary bone malignancy affecting long bones in children and the third most frequent in adults in the United States. It is highly invasive and metastatic. The survival rates over the last 20 years are unchanged. Therefore, it is essential to develop novel, targeted therapeutic approaches to treat OS. In pursuit of that goal, novel models are needed. Due to the high similarity in cancer genetics, the spontaneous occurrence of tumors, and the presence of an intact immune system, dogs are deemed an excellent cancer immunotherapy model. OS is also the most common malignant bone tumor (80%) in dogs. Monotherapies like oncolytic viruses and immune checkpoint inhibitors have been shown to be insufficient for effective cancer treatment. Therefore, we propose a combination of immune checkpoint inhibition and oncolytic virotherapy to target osteosarcoma tumor cells. In this combined therapy, the oncolytic virus will serve to both stimulate an inflammatory response and transmit the checkpoint inhibitor, which in turn will block PD-1-directed down-regulation of T cells, allowing the adaptive immune system to respond better to tumors. In order to evaluate this approach in the canine model, we have developed an oncolytic virus (CAV2-AU-M2) based on canine adenovirus type 2 and armed with an immune checkpoint inhibitor, an anti-canine PD1 single domain camelid antibody (sdAb). Four canine osteosarcoma cell lines, D17, D22, MCKOS, and CF11, were cultured in two-dimensional (2D) and three-dimensional (3D) cultures and were infected with CAV2-AU-M2 to analyze the infective and lytic effects of the virus. CAV2-AU-M2 infected and lysed the cells in 2D. The virus also produced anti-PD1 sdAb upon infecting the cells. Similarly, results were evident in 3D cultures. The virus infected the spheroids in the outer layers on Day 4 and slowly diffused towards the core in 7 days. The spheroids infected started showing some cell death on day 4, which progressed significantly by Day 7. This approach to oncolytic virotherapy makes the tumor more visible to the immune system and abrogates tumor-induced immune suppression.

Citation Format: Aliaa A. Ismail, Daniel J. Patton, Theresa A. Higgins, Maninder Sandey, Bruce F. Smith, Payal Agarwal. Exploration of multiple immunotherapy modalities in ostesarcoma [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 2339.
Abstract 2390: Claudin-4 probe identifies triple negative breast cancer using multispectral optoacoustic tomography
Authors: Parsons Laycee , MacCuaig William M. , McNally Lacey R.
Published: April 2023

Background: Triple negative breast cancer (TNBC) is the most aggressive subtype of breast cancer. The cancer is known for the lack of estrogen and her2 receptors. TNBC has a 5-year survival rate of 77% as for the other breast cancers averaging out at 93%. The CL-750 probes are made to identify the claudin-4 that is expressed in TNBC, making it easy to identify the cancer quickly so we can treat quickly.

Methods: TNBC, aggressive pancreatic and kidney cancer cell lines were used to assess the expression of claudin-4 via western blot. Based upon the C-terminal fragment of Clostridium perfingens, we synthesized a 30 aa peptide. CL peptide was conjugated to amine reactive HiLite 750 nm fluorescent dye. The CL-750 probes were purified using dialysis to remove unattached dye. MDA-MB-468 (TNBC), MDA-MB-231 (TNBC), S2VP10 (pancreatic), and 293 (kidney) cells were incubated with CL-750 probes for 30 minutes and evaluated with near infrared-fluorescence using Odyssey imaging software. Using the treated cells were then scraped into agar tissue mimicking phantoms, the probes were assessed for optoacoustic activity using the MSOT (Multispectral optoacoustic tomography).

Results: Using western blot to show expression of claudin-4 within cell lines the S2VP10 pancreatic (positive control) showed the most expression of claudin-4, while the 293 kidney cells showed a lesser amount. Both MDA-MB-468 and MDA-MB-231 TNBC cells showed the lowest amount of claudin-4 nonetheless both showed enough to continue with experiments. Cell line S2VP10 showed an extreme amount of binding to probe, while as the probe also bound to both smaller amount of claudin positive breast cancer cell line (MDA-MB-231) and larger amount of claudin breast cancer cell line (MDA-MB-468). Both CL-Short-750 and CL-Long-750 showed signal using MSOT. The CL-Long-750 showed 2x as much signal as CL-Short-750 making the Cl-long-750 probe more efficient in identifying claudin positive cell TNBC (p<0.05).

Conclusion: Both CL-750-probes have the capability of identifying claudin-4 positive cell lines, but CL-Long-750 was more effective and had greater binding.

Citation Format: Laycee Parsons, William M. MacCuaig, Lacey R. McNally. Claudin-4 probe identifies triple negative breast cancer using multispectral optoacoustic tomography [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 2390.
Abstract 2397: An advanced approach technique for tracking point between immune check point inhibitor and cancer cell mechanism in immune response motility by infiltration a breast cancer cell
Authors: Kim Hyunseok , Ahn Soyeon , Kwon Hyungjin , Chen Hsuan-Ju , Park Inwon , Kim Pilhan , Kubra Akyildiz
Published: April 2023

In Breast Cancer, Tumor-infiltrating lymphocyte (TIL) motility has been reported that it is reflecting tumor microenvironment response by adoptive cell therapy (ACT). ACT is well-known to be best applied for suppressing cancer cell growth with immune response and it also was well-known the immunotherapy. However, ACT response tracking by general analysis method included immunohistochemistry and flow cytometry was provided limited information for cancer therapy in recent years. Therefore, by assessing relationship immunotherapy marker, PD-L1, known as immune check point inhibitor and TIL using intravital microscopy in breast cancer model, we also imaged whether TIL response was related to immunotherapy following dependent on time point in a various breast cancer cell implanted mouse model such as 4T1 (mouse breast cancer cell), MDA-MB-231 (human triple negative breast cancer cell) and MCF7 (human breast cancer cell). These images result in PD-L1 expression was decreased in cancer cell implanted until 10days, whereas TIL expression increased in a various breast cancer cell when it is measuring displacement length, track mean velocity and confinement ratio called tumor microenvironment. These results indicated that we can imaged immune response related to TIL response in breast cancer cell line by intravital microscopy instead of general microscopy. It means that intravital microscope was an advanced approach technique to understand cancer mechanism accompanied by the immune response and established the method of high-resolution intravital cellular visualization compared to a general histopathological method for evaluating immune response in cancer tissue

Citation Format: Hyunseok Kim, Soyeon Ahn, Hyungjin Kwon, Hsuan-Ju Chen, Inwon Park, Pilhan Kim, Akyildiz Kubra. An advanced approach technique for tracking point between immune check point inhibitor and cancer cell mechanism in immune response motility by infiltration a breast cancer cell [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 2397.
Abstract 240: Comprehensive analytical solution to measure HRD genomic scars from SNP arrays, pan-cancer NGS panels, and optical genome mapping (OGM)
Authors: Saul Daniel , Roytman Megan , Wasnikar Viren , Burnside Rachel , Hastie Alex , Chaubey Alka
Published: April 2023

Homologous recombination deficiency (HRD) status has been shown to be an effective pharmacogenetic biomarker for determining the efficacy of PARP inhibitor therapy across various tumor types. A functional assessment of the genomic instability associated with HRD is critical as more than half of HRD positive tumors do not have a known causative mutation. Genomic instability is commonly achieved through assessment of three characteristic genomic scars: Loss of heterozygosity (LOH), Telomeric Allelic Imbalance (TAI), & Large-Scale State Transitions (LST) but is challenging due to the need for robust detection of copy number and allelic events genome-wide. As the technology commonly used to analyze tumors varies, we sought to compare the concordance of commonly used platforms to measure genomic instability.

Bionano VIA™ software provides a standardized analytical measurement of the genomic scars across technologies, including SNP arrays, pan-cancer gene panels, and optical genome mapping (OGM). The software is uniquely capable to profile solid tumors from pan-cancer sequencing panels, such as the Illumina TruSight Oncology 500 (TSO500) panel, by employing a dynamic binning approach of sequencing data to detect copy number variant (CNV) and LOH events. OGM can detect additional classes of structural rearrangements missed by other technologies to provide new insights into genomic instability. In this study, tumor samples were run on multiple technologies and analyzed in VIA to provide a measurement of the genomic scars. A comparative analysis of each technology’s performance to detect genomic instability through a standardized analytical approach will be described to provide insight into the most effective technologies to assess HRD.

Citation Format: Daniel Saul, Megan Roytman, Viren Wasnikar, Rachel Burnside, Alex Hastie, Alka Chaubey. Comprehensive analytical solution to measure HRD genomic scars from SNP arrays, pan-cancer NGS panels, and optical genome mapping (OGM) [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 240.
Abstract 2495: Investigating pancreatic cancer-derived extracellular vesicle signaling in the tumor microenvironment
Authors: Finan Jennifer M. , Saugstad Julie , Brody Jonathan
Published: April 2023

In many solid tumors, including pancreatic ductal adenocarcinoma (PDAC), the secretion of extracellular vesicles (EVs) has been shown to support tumor progression, chemotherapeutic resistance, and metastasis. Importantly, PDAC tumors are composed of up to 80% non-tumor cells and an extracellular matrix, yet no studies have been performed to determine whether PDAC EVs are imported preferentially by specific non-tumor cell types. To improve our understanding of this signaling, we aim to define the recipient cells of PDAC EVs and evaluate the functional importance of this internalization. Utilizing an in vitro live imaging system, we found that cancer-associated fibroblasts (CAFs) import more PDAC EVs than normal pancreas epithelial, endothelial, or PDAC cells, suggesting PDAC EV trafficking may be preferential to CAFs. We have generated human PDAC and mouse KrasG12D Trp53R172H driven (KPC) PDAC cell lines to express an extracellular particle reporter labeling the inner leaflet of cell membranes with GFP-nLuc. We are currently using this model to characterize recipient cells of the GFP-nLuc-positive EVs in an immunocompetent orthotopic PDAC mouse model. We developed flow cytometry and immunofluorescence panels to assess PDAC EV import by CAFs, endothelial cells, macrophages, B cells, and T cells in developed tumors. Utilizing immunofluorescence staining for a panel of phenotypic markers, we will assess the functional importance of EV internalization. Future studies will begin to determine how EV mRNA cargoes are important for recipient cell phenotypic and functional changes. This study is the first to ascertain the fate of PDAC EVs within the tumor microenvironment in a physiologically relevant model.

Citation Format: Jennifer M. Finan, Julie Saugstad, Jonathan Brody. Investigating pancreatic cancer-derived extracellular vesicle signaling in the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2495.
Abstract 2518: Heterogeneity of cytokine secretion in single senescent fibroblasts is impacted by ONC201 and ABT-263
Authors: Zhou Lanlan , Zhang Shengliang , Huntington Kelsey E. , Carneiro Benedito A. , El-Deiry Wafik S.
Published: April 2023

Cellular senescence and the associated secretory phenotype (SASP) promote cancer. During the process of aging, cellular and molecular changes that occur in non-cancerous cells can alter responses to cancer therapy and immune response in the tumor microenvironment (TME). Targeting senescent cancer-associated fibroblasts (CAFs) by means of removal, modulation of the SASP, or through cellular reprogramming represent therapeutic avenues for treating cancer. Heterogeneity of CAFs is emerging as an important area of investigation in cancer therapy. To better elucidate the interplay between cancer cells, immune cells, and senescent fibroblasts from different tissues and determine their impact on tumor growth and response to cancer therapy, we investigated the heterogeneity of cytokine secretion in senescent fibroblasts by a single cell cytokine assay. Control and treated proliferating and senescent fibroblasts were loaded onto IsoPlexis IsoCode chips coated with multiplexed antibodies which allows detection of 32 cytokines at a single-cell level for frequency and intensity. The Polyfunctional Strength Index (PSI) showed that stimulatory and chemo-attractant cytokines were dominant in the senescent fibroblasts. The Polyfunctional heat map showed that there are subpopulations of cells which have polyfunctional cytokine secretion (>2 different cytokines) in live senescent IMR90 cells. Treatment with ONC201 reduced cytokine polyfunctionality in live senescent IMR90 cells. Although ABT-263 (small molecule inhibitor of Bcl-2, Bcl-XL) did not show a significant reduction of polyfunctionality in live senescent IMR90, the frequency showed that ABT-263 reduced the secretion frequency of live senescent IMR90 cells except for GM-CSF. Combining ONC201 and ABT-263 diminished polyfunctionality and secretion frequency of senescent IMR90 cells except for MIF which was detected in 1.5% of the cells. Single cell cytokine profiling of live cells in the complex tumor microenvironment is providing insights into tumor progression, therapy resistance, and the effects of senolytic drugs and drug combinations.

Citation Format: Lanlan Zhou, Shengliang Zhang, Kelsey E. Huntington, Benedito A. Carneiro, Wafik S. El-Deiry. Heterogeneity of cytokine secretion in single senescent fibroblasts is impacted by ONC201 and ABT-263 [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 2518.
Abstract 2584: Overcoming cetuximab resistance in colorectal cancer through inhibition of HGF processing
Authors: Jones Vivian Truong , Graves-Deal Ramona , Bogatcheva Galina , Cao Zheng , Higginbotham James , Harmych Sarah , Ayers Gregory , Ramirez Marisol , Liu Qi , Janetka James , Singh Bhuminder
Published: April 2023

Cetuximab is a monoclonal antibody against epidermal growth factor receptor (EGFR) and can prolong survival in metastatic colorectal cancer (CRC) patients. However, therapeutic resistance can limit its efficiency and use. Our lab has previously identified that the activation of receptor tyrosine kinases (RTKs) MET and RON contributed to cetuximab resistance. In the absence of activating mutations of MET and RON, the activity of these RTKs depends on the availability of their ligands, hepatocyte growth factor (HGF) and hepatocyte growth factor-like (HGFL), respectively. To become biologically active, the inactive precursors of HGF and HGFL must be processed by serine proteases (HGFA, matriptase, and hepsin). A survey of TCGA datasets indicated that MET and HGF were overexpressed in several CRC CMS subtypes. However, we did not observe any significant overexpression of the serine proteases. Hepatocyte growth factor activator inhibitor type 1 and 2 (HAI-1/2) are two Kunitz-type serine protease inhibitors that displayed reduced expression in TCGA datasets. The addition of recombinant human HGF conferred cetuximab resistance to cetuximab-sensitive cells and cetuximab-sensitive cells engineered to overexpress HGF also exhibited cetuximab-resistant phenotype. Cetuximab resistance induced by HGF addition or overexpression could be overcome by downstream inhibition with crizotinib, a multi-targeted RTK inhibitor. ZFH7116 is a small molecule inhibitor of the serine proteases that mimics HAI-1 activity. ZFH7116 suppressed HGF cleavage and maturation in our cetuximab-resistant SC cells. We observed cooperative effect of ZFH7116 with cetuximab to reduce SC cell growth in 3D collagen cultures. We are currently testing if upstream inhibition with recombinant human HAI-1 addition can suppress HGF cleavage and overcome cetuximab resistance induced by HGF overexpression. Collectively, our findings suggest that inhibiting HGF cleavage and processing may be a unique strategy for overcoming EGFR-targeted therapy resistance in colorectal cancer.

Citation Format: Vivian Truong Jones, Ramona Graves-Deal, Galina Bogatcheva, Zheng Cao, James Higginbotham, Sarah Harmych, Gregory Ayers, Marisol Ramirez, Qi Liu, James Janetka, Bhuminder Singh. Overcoming cetuximab resistance in colorectal cancer through inhibition of HGF processing [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 2584.
Abstract 2585: Mutant p53 gains oncogenic functions through a cytosolic DNA response
Authors: Zhao Mei , Wang Tianxiao , Gleber-Netto Frederico O. , Chen Zhen , McGrail Daniel J. , Gomez Javier A. , Ju Wutong , Gadhikar Mayur A. , Ma Wencai , Shen Li , Tang Ximing , Pathak Sen , Raso Maria G. , Burks Jared , Lin Shiaw-Yih , Wang Jing , Multani Asha S. , Pickering Curtis R. , Chen Junjie , Myers Jeffrey N. , Zhou Ge
Published: April 2023

TP53 mutations are the most common cancer driver mutations among all cancers. Although some TP53 mutations lead to a loss of function of wild-type p53, many other TP53 mutations confer gain-of-function (GOF) activities, which promote cancer cell metastasis and pro-tumorigenic inflammation. Despite that many functional models of GOF mutant p53 (mutp53) have been proposed previously, the mechanisms involved in mutp53 GOF still remain largely elusive. Here we show that by directly targeting minichromosome maintenance complex component 5 (MCM5), a component of the hexametric DNA helicase MCM2-7 complex, GOF mutp53 predisposes cancer cells to replication stress and chromosomal instability, which leads to a tumor cell-autonomous and stimulator of interferon genes (STING)-dependent cytosolic DNA response that activates downstream non-canonical nuclear factor kappa light chain enhancer of activated B cell (NC-NF-κB) signaling. Furthermore, our results demonstrate that GOF mutp53-activated tumor cell-intrinsic STING-NC-NF-κB signaling not only stimulates tumor cell metastasis, but also promotes tumor immune resistance through fostering an immunosuppressive tumor microenvironment. Therefore, our findings that mutp53 exerts its GOF role through pro-tumorigenic MCM5-CIN-STING-NC-NF-κB signaling highlight the importance of TP53 and its inactivation in cancer genome evolution of genomic instability that drives tumor development and progression.

Citation Format: Mei Zhao, Tianxiao Wang, Frederico O. Gleber-Netto, Zhen Chen, Daniel J. McGrail, Javier A. Gomez, Wutong Ju, Mayur A. Gadhikar, Wencai Ma, Li Shen, Ximing Tang, Sen Pathak, Maria G. Raso, Jared Burks, Shiaw-Yih Lin, Jing Wang, Asha S. Multani, Curtis R. Pickering, Junjie Chen, Jeffrey N. Myers, Ge Zhou. Mutant p53 gains oncogenic functions through a cytosolic DNA response [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 2585.
Abstract 2593: A MYCN-independent mechanism mediating secretome reprogramming and metastasis in MYCN-amplified neuroblastoma
Authors: Zhang Hai-Feng , Delaidelli Alberto , Javed Sumreen , Turgu Busra , Hughes Christopher S. , Yang Xiaqiu , Lizardo Michael , Martinez Daniel , Maris John M. , Bosse Kristopher R. , Williams Karla C. , Sorensen Poul H.
Published: April 2023

MYCN amplification (MNA) is a defining feature of high-risk neuroblastoma (NB) that predicts poor prognosis. However, whether genes within or in close proximity to the MYCN amplicon also contribute to aggressiveness in MNA+ NB remains poorly understood. Here we identify that GREB1, a transcription factor encoding gene neighboring the MYCN locus, is frequently co-expressed with MYCN, and promotes gene expression independently of MYCN in MNA+ NB. We found that GREB1, as a crucial estrogen receptor (ER) regulatory factor, is highly expressed in MNA+ NB comparable to ER+ breast cancer. Among GREB1-regulated genes altered in MNA+ NB, we uncover that Myosin 1B (MYO1B) promotes aggressive features in MNA+ NB, including invasive capacity in vitro, as well as extravasation and distant metastasis in vivo. Global secretome and proteome profiling uncover MYO1B as a major regulator of secretome reprogramming in MNA+ NB cells. Moreover, we identify the cytokine MIF as an important pro-invasive and pro-metastatic mediator of MYO1B activity. Together, we have identified a putative GREB1-MYO1B-MIF axis as an unconventional mechanism that promotes the aggressiveness of MNA+ NB, and independently of MYCN. Furthermore, we found that MYO1B is also upregulated in association with other oncoproteins during cellular transformation, and is dramatically increased in multiple human cancer types, suggesting a crucial role of MYO1B in cancers in addition to MNA+ NB.

Citation Format: Hai-Feng Zhang, Alberto Delaidelli, Sumreen Javed, Busra Turgu, Christopher S. Hughes, Xiaqiu Yang, Michael Lizardo, Daniel Martinez, John M. Maris, Kristopher R. Bosse, Karla C. Williams, Poul H. Sorensen. A MYCN-independent mechanism mediating secretome reprogramming and metastasis in MYCN-amplified 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 2593.
Abstract 2594: Alcohol drinking inhibits NOTCH-PAX9 signaling in esophageal squamous epithelial cells
Authors: Xiong Zhaohui , Shi Menghan , Ren Shuang , Chen Hao , Li Jing , Huang Caizhi , Li Yahui , Han Yuning , Li Yong , Sun Zheng , Chen Xiaoxin
Published: April 2023

Alcohol drinking has been established as a major risk factor for esophageal diseases, such as gastroesophageal reflux disease (GERD) and esophageal squamous cell carcinoma (ESCC). Certain mechanisms of alcohol-induced esophageal injury are supported by experimental data, whereas most are speculative or extrapolated from studies on cancers of other organ sites. Our previous study showed that ethanol exposure inhibited PAX9 expression in human esophageal squamous epithelial cells in vitro and in vivo, and promoted ESCC. In this study, we aimed to investigate the molecular pathways through which alcohol drinking suppresses PAX9 in esophageal squamous epithelial cells. Gene microarray, qRT-PCR, ChIP-PCR, Western blotting, and immunostaining were used to identify and validate inhibition of NOTCH-PAX9 signaling in human esophageal squamous epithelial cells and mouse esophagus after ethanol exposure. Human tissue samples were analyzed to further validate NOTCH inhibition by alcohol drinking. We first demonstrated the inhibition of NOTCH by ethanol exposure in vitro. NOTCH regulated PAX9 expression in esophageal squamous epithelial cells in vitro and in vivo. RBPJ and NOTCH intracellular domain (NICD1) ChIP-PCR confirmed Pax9 as a direct downstream target of NOTCH signaling in mouse esophagus. NOTCH inhibition by alcohol drinking was further validated in mouse esophagus and human tissue samples. In conclusion, ethanol exposure inhibited NOTCH signaling and thus suppressed PAX9 expression in esophageal squamous epithelial cells in vitro and in vivo. Our data

support a novel mechanism of alcohol-induced esophageal injury through the inhibition of NOTCH-PAX9 signaling. As a result, chemical NOTCH activators may be used to prevent or treat alcohol-associated esophageal diseases such as GERD and ESCC.

Citation Format: Zhaohui Xiong, Menghan Shi, Shuang Ren, Hao Chen, Jing Li, Caizhi Huang, Yahui Li, Yuning Han, Yong Li, Zheng Sun, Xiaoxin Chen. Alcohol drinking inhibits NOTCH-PAX9 signaling in esophageal squamous epithelial 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 2594.
Abstract 2629: Single and combined inhibition of PAK and Hippo pathway in NF2-deficient schwannoma
Authors: Benton Dorothy , Chernoff Jonathan
Published: April 2023

Neurofibromatosis 2 (NF2) Syndrome is a rare, but devastating genetic condition that leads to non-cancerous tumors of the nervous system. NF2 patients are likely to develop Schwannomas on the eighth cranial nerve that lead to progressive deafness and spinal cord compression as well as meningiomas and peripheral nerve tumors. Effective treatment for these patients is lacking, with risky surgery being the most reliable approach and pharmacotherapies having inadequate results. The hallmark mutation of this condition is inactivation of NF2, which codes for the protein Merlin. Merlin plays several roles in the cell, two of the most consequential being inhibition of PAK1 and activation of the Hippo tumor suppressor pathway. However, drugs targeting these pathways have not yet been investigated in NF2-deficient Schwannoma. In this study, we explored the use of YAP-TEAD inhibitors as well as the combination of Hippo family and Group I PAK inhibition in NF2-deficient Schwannoma cells. We used several Schwannoma cells lines of human, mouse, and rat origin as well as a human Schwann cell line with NF2 knocked out via CRISPR compared to its wild-type control line. We investigated the role of YAP, the main effector of the Hippo pathway, and PAK1/2 in NF2 Schwannoma cells via genetic and pharmacologic inhibition. We also combined YAP-TEAD inhibitors with several PAK family inhibitors to induce a synergistic reduction in cell viability and proliferation as well as an increase in apoptosis. We found that genetic inhibition of both YAP and Group I PAK proteins led to partial reduction in Schwannoma growth. Using several YAP-TEAD binding inhibitors (TED-347, NSC682769, IK-930), Group I PAK inhibitors (FRAX-1036, G555) and PAK1-specific inhibitor (NVS-1-PAK1) we found that YAP-TEAD binding inhibition combined with Group I PAK inhibition has the greatest reduction on Schwannoma cell viability, proliferation, and increase in cell death. Taken together, our findings indicate that both the Hippo pathway and PAK proteins may be viable targets for Schwannoma therapies. Furthermore, combining the two may lead to a synergistic effect and further prevent growth of NF2-deficient Schwannomas.

Citation Format: Dorothy Benton, Jonathan Chernoff. Single and combined inhibition of PAK and Hippo pathway in NF2-deficient schwannoma [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 2629.
Abstract 2643: BL-M07D1, a novel HER2-targeting ADC, demonstrates potent anti-tumor efficacy in preclinical pharmacodynamic models
Authors: Wan Weili , Zhao Shuwen , Zhuo Shi , Zhang Yong , Chen Lan , Li Gangrui , Khalili Jahan Salar , Xiao Sa , Yan Yongqi , Shen Xuejiao , Zhu Yi
Published: April 2023

HER2 is a member of the epidermal growth factor receptor family. Activation of the HER2 signaling pathway has been shown to strongly promote carcinogenesis. It is therapeutically targeted in cancer owing to its overexpression and pathway dependence in a variety of human carcinomas, especially human breast cancers. We created a promising therapeutic anti-tumor agent, BL-M07D1, an anti-HER2-Ed-04 ADC. It is comprised of a humanized anti-HER2 antibody Trastuzumab, a cathepsin B cleavable linker, and a novel topoisomerase I inhibitor agent (Ed-04), which is a derivative of the alkaloid camptothecin, driving cell cycle arrest at the S phase and subsequent apoptosis. The BL-M07D1 drug-to-antibody-ratio is 8:1 (DAR=8), similar to Trastuzumab Deruxotecan (DS-8201), while possessing a more stable linker.

To evaluate the pharmacological potential of BL-M07D1, xenograft tumor inhibition assays were used to compare BL-M07D1 with the commercialized HER2-targeting ADCs, T-DM1 and DS-8201, which have been approved worldwide for patients with HER2-expressing tumors. Results from the in vivo murine studies show that BL-M07D1 has strong tumor inhibition effects in multiple cell line-derived xenograft (CDX) tumor models. 1) BL-M07D1 exhibited better anti-tumor efficacy than DS-8201 in CDX with low HER2 expression, human epidermal cancer A431 and human non-small cell lung cancer NCI-H1975. Both models are considered be T-DM1-insensitive. 2) BL-M07D1 exhibited better anti-tumor efficacy in comparison to either T-DM1 or DS-8201 in a CDX with JIMT-1, a HER2-positive human breast cancer cell line. 3) BL-M07D1 exhibits potent bystander effects in a heterogeneous xenograft model of HER2-positive and HER2-negative tumor cells composed of NCI-N87 and MDA-MB-468 cells. In this model, BL-M07D1 exhibited stronger tumor inhibition than T-DM1, consistent with bystander effects that are also exhibited by DS-8201.

In conclusion, in vivo studies suggest that BL-M07D1, a novel HER2-targeting ADC, is potentially more efficacious in a broader patient population than T-DM1, and mediate superior anti-tumor efficacy than DS-8201. The clinical phase I is under way and the available data exhibit excellent efficacy in breast cancer therapy with acceptable tolerability.

Citation Format: Weili Wan, Shuwen Zhao, Shi Zhuo, Yong Zhang, Lan Chen, Gangrui Li, Jahan Salar Khalili, Sa Xiao, Yongqi Yan, Xuejiao Shen, Yi Zhu. BL-M07D1, a novel HER2-targeting ADC, demonstrates potent anti-tumor efficacy in preclinical pharmacodynamic models [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 2643.
Abstract 2648: In vivo efficacy and preclinical safety profile of NN3201, an anti-cKIT antibody-drug conjugate, for treating SCLC and AML
Authors: Ko Han-Jik , Kim Sung Tae , Cho Jin Gu , Kim Jin-Ock , Lee Jung Tae , Park Sang Gyu
Published: April 2023

Overexpression and activating mutations of cKIT are associated with tumor aggressiveness and poor prognosis in various cancer patients such as small cell lung cancer (SCLC), gastrointestinal stromal tumor (GIST), acute myeloid leukemia (AML), and mastocytoma. Since small molecule inhibitors including imatinib frequently have a limited therapeutic application due to eventual drug resistance and severe adverse events, we developed the cKIT-directed antibody-drug conjugate (ADC), NN3201, for the treatment of wild-type and mutant cKIT-positive cancers. NN3201 is an ADC conjugate of the antimitotic payload MMAE to the fully human 2G4 antibody, directed against cKIT, via the cysteine re-bridging technology ThioBridge® at a drug-to-antibody ratio (DAR) of 4. NN3201 displayed preferable biophysical properties and broadly potent in vitro antitumor activities across various cKIT-positive tumor cell lines. NN3201 exhibited potent in vivo antitumor activities in cKIT-positive tumor cell line xenograft and patient-derived tumor xenograft models of SCLC and AML, representing robust and durable antitumor responses. Furthermore, in a single-dose toxicity study with cynomolgus monkey, no significant or severe clinical toxicity signs were observed in animals dosed at 1, 2, and 3 mg/kg. Even though a decrease in reticulocyte, red cell mass (hemoglobin and red blood cell), neutrophil, and monocyte counts, along with bone marrow recovery and reticulocyte compensatory mechanism, red blood cell and white blood cell recovery were observed from day 14 in the low and medium dose groups and day 21 in the high dose group, dose-dependently. These preclinical results suggest that NN3201 is a potential therapeutic agent for the treatment of cKIT-positive cancers and support the further clinical evaluation in SCLC and AML.

Citation Format: Han-Jik Ko, Sung Tae Kim, Jin Gu Cho, Jin-Ock Kim, Jung Tae Lee, Sang Gyu Park. In vivo efficacy and preclinical safety profile of NN3201, an anti-cKIT antibody-drug conjugate, for treating SCLC and AML [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 2648.
Abstract 2675: Vactosertib combined treatment with T1-44, a PRMT5 activity inhibitor, improves the survival and inhibits tumor invasion of murine pancreatic cancer model
Authors: Hong Eunji , Park Sujin , Park Seok Hee , La Thangue Nick B. , Kim Seong-Jin
Published: April 2023

Pancreatic cancer is the most lethal type of cancer and the third leading cause of cancer death with the lowest 5-year survival rate. Heterogeneity, difficulty in diagnosis, and rapid metastatic progression are the causes of high mortality in pancreatic cancer. Recent studies have shown that Protein arginine methyltransferase 5 (PRMT5) is overexpressed in pancreatic cancers, and these patients have a worse prognosis. Therefore, PRMT5 as an anti-cancer target has gained considerable interest. In this study, we investigated whether inhibition of PRMT5 activity was synergistic with blockade of TGF-β1 signaling, which plays an important role in the construction of the desmoplastic matrix and invasiveness in pancreatic cancer and induces therapeutic vulnerability. Compared with T1-44, a selective inhibitor of PRMT5 activity, the combination of T1-44 with the TGF-β1 signaling inhibitor vactosertib significantly reduced tumor size and surrounding tissue invasion and significantly improved long-term survival. RNA sequencing analysis of mouse tumors revealed that the combination of T1-44 and vactosertib significantly altered the expression of genes involved in cancer progression, such as cell migration, extracellular matrix, and apoptotic processes. These data demonstrate that the combination therapy of T1-44 with vactosertib is synergistic for pancreatic cancer, suggesting that this novel combination therapy has value in the treatment strategy of patients with pancreatic cancer.

Citation Format: Eunji Hong, Sujin Park, Seok Hee Park, Nick B. La Thangue, Seong-Jin Kim. Vactosertib combined treatment with T1-44, a PRMT5 activity inhibitor, improves the survival and inhibits tumor invasion of murine pancreatic 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 2675.
Abstract 2682: Targeting phosphoglycerate dehydrogenase (PHGDH) in combination with tamoxifen reduces hormone receptor positive breast cancer growth in vivo
Authors: Clem Brian F. , Imbert-Fernandez Yoannis , Petri Belinda , Klinge Carolyn
Published: April 2023

Breast cancer afflicts thousands of individuals every year and is clinically diagnosed in various subtypes, including hormone receptor positive (HR+ BC). Significant strides have been made in treating HR+ BC through the development of endocrine therapies, such as tamoxifen, fulvestrant, and aromatase inhibitors. While these have improved the survival outcomes of these patients, many eventually relapse and, ultimately succumb to metastatic endocrine resistant disease. Subsequent work has identified various cellular pathways contributing to endocrine resistance, including alterations in cellular metabolism. We previously demonstrated that de novo serine synthesis (SSP) is increased in endocrine resistant cell types and genetic disruption of SSP enzymes impacts tamoxifen sensitivity. Further, elevated transcript expression is inversely correlated with survival outcomes in HR+ BC patients treated with tamoxifen. From this, we aimed to assess whether pharmacological targeting of SSP activity in combination with tamoxifen affects the growth of HR+ BC in vivo. We confirmed the presence of ER-alpha in three distinct patient derived xenograft (PDX) models and found varying levels of both phosphoglycerate dehydrogenase (PHGDH) and phosphoserine aminotransferase (PSAT1), which are the first and second enzymatic steps in the SSP pathway, respectively. In our efficacy studies, we initially found differential sensitivity to tamoxifen administration across the PDX models. While treatment with a PHGDH inhibitor (PHGDHi) had no anti-tumor activity against an endocrine-sensitive PDX, it moderately reduced the growth of endocrine resistant PDX tumors. Notably, combinatorial treatment resulted in a substantial decrease in tumor growth in all three models compared to single agents alone. Together, these data suggest that targeting of SSP activity in combination with endocrine therapies may be a promising therapeutic strategy in HR+ BC.

Citation Format: Brian F. Clem, Yoannis Imbert-Fernandez, Belinda Petri, Carolyn Klinge. Targeting phosphoglycerate dehydrogenase (PHGDH) in combination with tamoxifen reduces hormone receptor positive breast cancer growth in vivo [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 2682.
Abstract 2692: The combination of ulixertinib (ERK1/2 Inhibitor) and KRASG12C inhibition demonstrates significant efficacy in preclinical models
Authors: Knoerzer Deborah , Reddy Anupama , Box Jessica A. , Groover Anna , Kreider Brent , Teresk Martin , Emery Caroline M.
Published: April 2023

Ulixertinib (BVD-523) is a first-in-class and best-in-class small molecule inhibitor of ERK1/2 currently being investigated in several oncology clinical trials, both as a single agent and in combination with other therapeutics. Ulixertinib has demonstrated efficacy in patients with tumors harboring alterations within the RAS-MAPK pathway. KRAS is the most frequently mutated oncogene in cancer and is a key mediator of the RAS-MAPK signaling pathway resulting in proliferation and cellular growth. The specific KRASG12C mutation occurs in approximately 10% of non-small cell lung cancer (NSCLC), 3% of colorectal cancer, and 1-2% across all other tumor types. KRASG12C mutant-inhibitors, including AMG-510 (sotorasib), MRTX849 (adagrasib), and JDQ443 have demonstrated efficacy in KRASG12C-mutant cancers, including NSCLC. Clinically described mechanisms of acquired resistance to KRASG12C inhibitors converge on reactivation of the RAS-MAPK pathway. We hypothesized combining ulixertinib with a KRASG12C inhibitor would circumvent resistance to single agent KRASG12C inhibition, generating increased magnitude and duration of response compared to either single agent alone.

The efficacy of ERK1/2 inhibitor, ulixertinib, in combination with KRASG12C inhibitor, adagrasib, was assessed in cell line derived xenograft models harboring KRASG12C mutations. Models were selected based on response to single agent adagrasib, ranging from sensitive to partially responsive. Combination treatment resulted in superior tumor growth inhibition compared to dosing of either single agent. RNA sequencing was performed on tumor samples that were collected 2 hours after the last dose of treatment. Expression of the mutant KRAS alleles were readily confirmed from RNA sequencing data in all models. Gene expression analysis showed differential expression of MAPK pathway genes in monotherapy versus combination therapy treated groups.

In summary, ulixertinib combined with adagrasib exhibited robust pre-clinical activity in a variety of xenograft models with KRASG12C and should be further evaluated.

Citation Format: Deborah Knoerzer, Anupama Reddy, Jessica A. Box, Anna Groover, Brent Kreider, Martin Teresk, Caroline M. Emery. The combination of ulixertinib (ERK1/2 Inhibitor) and KRASG12C inhibition demonstrates significant efficacy in preclinical models [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 2692.