bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 967: Phosphodiesterase 4D depletion enhances anti-tumor effects of tyrosine kinase inhibitor in renal cell carcinoma involving CRAF-ERK pathway
Authors: Cao Minghua , Ajay Amrandra K. , Gasser Martin , Hsiao Li-LI , Waaga-Gasser Ana Maria
Published: July 2021

Clear cell renal cell carcinoma (ccRCC) is the most lethal form of kidney cancer. Small molecule Vascular Endothelial Growth Factor Receptor (VEGFR) inhibitors are widely used but are not curative and various resistant mechanisms have been described. Previous studies have indicated a crosstalk between the cAMP pathway and the MAPK pathway, and the elevation of cAMP levels by Phosphodiesterase 4 (PDE4) inhibition can result in growth arrest and/or cell death. In this study we focused on the effects of PDE4 in ccRCC. We showed for the first time that PDE4D is the dominant subtype of PDE4 in kidney. PDE4D depletion in ccRCC, Caki-1 cells, using CRISPR Knock Out (KO) enhanced sensitivity of Tyrosine Kinase Inhibitor (TKI), sorafenib and increased cytotoxicity. We also showed synergistic effects of PDE4 inhibitor, roflumilast and sorafenib, in increasing cytotoxicity of ccRCC in the absence of PDE4D. Our results further demonstrated that PDE4D deficiency increased intracellular cAMP levels and resulted in downregulation of MAPK but not PI3K/AKT signaling in a CRAF dependent manners. In conclusion, we provide a preclinical rationale for dual PDE4/VEGFR inhibition in patients with ccRCC. While the MAPK signaling pathway is activated in ccRCC, dual inhibitions may improve the anti-tumor effects in patients with proven CRAF signaling activation.

Citation Format: Minghua Cao, Amrandra K. Ajay, Martin Gasser, Li-LI Hsiao, Ana Maria Waaga-Gasser. Phosphodiesterase 4D depletion enhances anti-tumor effects of tyrosine kinase inhibitor in renal cell carcinoma involving CRAF-ERK pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 967.
Abstract 977: Mcl-1 levels critically determines the sensitivities of human colon cancer cells to APG1244, a novel Bcl-2 and Bcl-XLinhibitor, that induces Bax- and caspase-8- dependent apoptosis
Authors: Yao Weilong , Sun Shi-Yong
Published: July 2021

Colorectal cancer (CRC) is the third leading cause of cancer-related deaths worldwide due to its poor prognosis. Most of the patients receive surgical resection when diagnosed as CRC, whereas the recurrence rate is very high. Therefore, additional treatment options, such as targeted therapy, are still urgently needed. The novel BH3 mimetic, APG1244 (a prodrug for APG1252), is a highly potent Bcl-2 and Bcl-XL dual inhibitor and being tested in clinical trials as a potential cancer therapeutic agent. The current study focuses on demonstrating the therapeutic efficacies of APG1244 against colorectal cancer and understanding the underlying mechanisms. A panel of CRC cell lines with different levels of Bcl-2, Bcl-XL and Mcl-1 possessed different sensitivities to APG1244. APG1244 effectively decreased the survival of CRC cell lines, particularly those expressing relative low levels of Mcl-1, with induction of apoptosis. High levels of Mcl-1 in these cell lines were significantly correlated to decreased sensitivity to APG1244. APG1244, when combined with a Mcl-1 inhibitor, synergistically decreased the survival and induced apoptosis of APG1244-insensitive cell lines with high levels of Mcl-1. This combination could further decreased the survival and enhanced apoptosis even in the cell lines with relatively low levels of Mcl-1, whereas enforced expression of ectopic Mcl-1 in these cells abrogated APG1244's effects on decreasing cell survival and inducing apoptosis, which can be reversed by Mcl-1 inhibition. In our study, it appears that APG1244 quickly activated caspase-8 and caspase-3 and induced PARP cleavage with induction of cytochrome C and Smac release from mitochondria. Deficiency of Bax, Smac, Bim or PUMA, but not FADD, in CRC cells protected (BAX-KO) or partially protected cells from APG1244-induced apoptosis, indicating that APG1244 induces Bax-dependent apoptosis. Furthermore, blockage of caspase-8 with a caspase-8 specific inhibitor abolished cleavage of caspase-3 and PARP including attenuation of cytochrome C and Smac release induced by APG1244, suggesting a caspase-8-dependent mechanism. The current study thus demonstrates the potential of APG1244 as a monotherapy in the treatment of CRC, particularly those with low Mcl-1 expression, or in combination with a Mcl-1 inhibitor for CRC treatment, warranting further evaluation in vivo and in the clinic.

Citation Format: Weilong Yao, Shi-Yong Sun. Mcl-1 levels critically determines the sensitivities of human colon cancer cells to APG1244, a novel Bcl-2 and Bcl-XLinhibitor, that induces Bax- and caspase-8- dependent apoptosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 977.
Abstract 996: Cold atmospheric plasma therapy selectively targets triple negative breast cancer cells
Authors: Wang Peiyu , Zhou Renwu , Zhou Rusen , Gunter Jennifer , Simpson Fiona , Ostrikov Kostya (Ken) , Richard Derek , Dai Xiaofeng , Thompson Erik (Rik) Walter
Published: July 2021

Background: Cancer cells notoriously escape radiation therapy, chemotherapy, hormonal therapy, and other anti-cancer therapies. Among various breast cancer subtypes, triple negative breast cancers (TNBCs) are known for their poor clinical outcome and dearth of effective targeted therapies due to the absence of druggable receptors. Cold atmospheric plasma (CAP) holds promise as a cancer-specific treatment agent that selectively kills basal-like/TN breast cancer cells.

Method: In this study, we used plasma-activated media (PAM) to capture and store the multi-modal chemical species of CAP, which significantly expands the treatment scope and flexibility of plasma medicine as a clinical therapeutic approach. The PAM was diluted into different doses and cells were treated for 12 or 24 hours. The effects of PAM on cell viability and ROS levels in 9 different cell lines; normal cells (MCF-10A and HEK-293T), luminal breast cancer cell lines (MCF-7, T47D, HCC70), and TNBC cell lines (MDA-MB-468, SUM-159PT, SUM-149PT, MDA-MB-231), were assessed.

Results: The results showed that TNBC cells were more highly sensitive, in a dose-dependent manner, to PAM treatment. PAM selectively caused apoptosis of TNBC cell lines compared to luminal lines and had little effect on the normal cell lines. It has been hypothesised that the cancer-selective CAP and PAM cytotoxicity is due in part to reactive oxygen (ROS) and nitrogen species (RNS). Constitutive RONS levels, combined with those present in and/or induced by PAM, can combine to push the cells over a cytotoxic threshold. In support of this, we found a tight correlation (R2=0.7502) between the constitutive ROS level of different cell lines used and their response (cell viability) to 100% PAM treatment, and found that ROS levels selectively increased further in the TNBC>luminal>normal cells in response to PAM. We have also identified a strong synergistic molecular partner that further promotes sensitivity and selectivity of PAM for TNBC cells.

Conclusion: We demonstrated that treatment with PAM promotes apoptosis n cell lines representing the most aggressive TNBC subtype. This discovery opens new TNBC-selective opportunities for simultaneous inhibition of diverse (parallel) cancer development factors through synergistic interactions enabled through plasma activated biochemical media.

Citation Format: Peiyu Wang, Renwu Zhou, Rusen Zhou, Jennifer Gunter, Fiona Simpson, Kostya (Ken) Ostrikov, Derek Richard, Xiaofeng Dai, Erik (Rik) Walter Thompson. Cold atmospheric plasma therapy selectively targets triple negative breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 996.
Abstract CT156: Radiologic scoring method to define the therapeutic response and surgical candidacy
Authors: Rahmanuddin Syed , Von Hoff Daniel , Cridebring Derek , Brase Jordyn , Fong Yuman , Motarjem Pejman , Korn Ron , Choong Kevin , Boswell William , Borazanci Erkut
Published: July 2021

Purpose: The purpose of this study is to define the best CT perfusion analytical method for the surgical candidacy of pancreatic cancer patients in Neo-Adjuvant treated clinical trial population. Experimental Design: Study was designed to assess tumor perfusion status. This prospective study involved 24 adult patients with pancreatic adenocarcinoma (PDAC). Inclusion criteria included, patients with histologically or cytologically confirmed resectable, borderline resectable, or locally advanced (unresectable) PDAC. All patients received neoadjuvant therapy (paricalcitol, paclitaxel protein bound, cisplatin and gemcitabine) for up to 6 months Multi-phase CT was performed for most patients on a GE scanner using a standard pancreatic cancer protocol including a pre-injection phase, arterial phase, and porto-venous phase. The iodinated contrast agent Isovue 370 was dosed at 150 cc + 40 cc saline, and injected at a rate of 5 cc/sec into the antecubital vein. Volumetric analysis of images from 24 patients was conducted using 3D imaging software and perfusion scoring assessment was done on each case using 2D and 3D methodology. Regions of interest were drawn to define the volumetric tumor quantification. This work looked at the difference in density between arterial and venous phases, using the Marley equation.

This data was then used to assess treatment response and ultimately whether or not a patient would be a surgical candidate. This surgical candidacy of these patients was defined by the “R” score. Results: The R score defines the number of vessels with tumor involvement. Each vessel that had any amount of tumor involvement was scored with a 1. Any patient with a score of 6 or higher was deemed not eligible for surgery while a score of 5 or less was eligible for surgery. 3D Tumor volumetric assessments were performed for the assessment of the treatment response that linked directly with the tumor perfusion.Conclusion: CT vessel perfusion using the R score method could be helpful to define the surgical candidacy of pancreatic cancer patients along with 3D tumor volumetric quantification.

Citation Format: Syed Rahmanuddin, Daniel Von Hoff, Derek Cridebring, Jordyn Brase, Yuman Fong, Pejman Motarjem, Ron Korn, Kevin Choong, William Boswell, Erkut Borazanci. Radiologic scoring method to define the therapeutic response and surgical candidacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT156.
Abstract CT212: Expanded phase 1/2a study of PLX8394, a novel next generation BRAF inhibitor in patients with advanced, unresectable solid tumors with alterations in BRAF
Authors: Janku Filip , Sherman Eric , Yaeger Rona , Parikh Aparna , Sullivan Ryan , Feun Lynn , Fuente Macarena De La , Tsai Frank Yung-Chin , Gordon Michael , Allen Carl , Rudoltz Marc S. , Balinski Kathe , Averbuch Steven , Vidne Michael , Tarcic Gabi
Published: July 2021

The authors did not submit an updated abstract. The original abstract should be considered final.

Citation Format: Filip Janku, Eric Sherman, Rona Yaeger, Aparna Parikh, Ryan Sullivan, Lynn Feun, Macarena De La Fuente, Frank Yung-Chin Tsai, Michael Gordon, Carl Allen, Marc S. Rudoltz, Kathe Balinski, Steven Averbuch, Michael Vidne, Gabi Tarcic. Expanded phase 1/2a study of PLX8394, a novel next generation BRAF inhibitor in patients with advanced, unresectable solid tumors with alterations in BRAF [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT212.
Abstract CT227: A phase 1 multiple ascending dose study to investigate the safety, pharmacokinetics, pharmacodynamics and preliminary efficacy of KD033 in subjects with metastatic or locally advanced solid tumors
Authors: Luke Jason J. , Olszanski Anthony J. , Puzanov Igor , Lu Dan , Hackett Adrian , Martomo Stella , Patel Jeegar , Schueller Olivier , Mora Alessandro , Schlitt Miranda L. , Li Linghui
Published: July 2021

Background: IL-2 and IL-15 signal through the shared IL-2/15 βγ receptor, but unlike IL-2, IL-15 does not expand regulatory T cells (Tregs). In addition, unlike IL-2, IL-15 does not mediate activation-induced cell death and is expected to have an improved therapeutic index compared to IL-2. KD033 is a fusion antibody molecule generated by combining a fully human, high affinity anti-human Programmed Death Ligand 1 (PD-L1) IgG1 antibody with the human IL-15 receptor alpha (IL15Rα) sushi domain and human IL-15 (IL-15). KD033 (or its mouse cross reactive surrogate molecule, srKD033) has been extensively characterized in multiple in vitro and in vivo nonclinical studies to understand the pharmacology, pharmacokinetic (PK) and toxicology properties. The fusion of anti-PD-L1 antibody to IL-15 significantly increases the maximal-tolerated dose (MTD) of srKD033 in mice compared to free-IL-15. The increased safety and efficacy of PD-L1-targeted IL-15 observed in pre-clinical studies warranted evaluation of KD033 in humans. Methods: This is a phase 1, open-label, multiple ascending dose, multi-center clinical trial being conducted in patients with metastatic or locally advanced solid tumors (NCT04242147). The primary objective is to determine the safety and tolerability and the MTD of KD033. KD033 is administered as an IV infusion over 30 minutes every 14 days (i.e., a cycle equals 2 weeks). The starting dose of KD033 is 3 µg/kg, with step-up dosing to 12.5 µg/kg in the first cohort of 3 subjects. After the first cohort, the dose escalation will use a standard 3+3 design with planned doses from 25 µg/kg to 600 µg/kg. The dose escalation phase will be followed by one expansion cohort, which will enroll patients who received PD-1/PD-L1 inhibitor but have either progressed or are refractory to therapy. Secondary objectives include characterization of PK and immunogenicity, evaluation of immune correlates that will potentially differentiate the impact of KD033 versus monotherapy as well as investigate IL-15 biology and determine best overall response and duration of response. (NCT04242147)

Citation Format: Jason J. Luke, Anthony J. Olszanski, Igor Puzanov, Dan Lu, Adrian Hackett, Stella Martomo, Jeegar Patel, Olivier Schueller, Alessandro Mora, Miranda L. Schlitt, Linghui Li. A phase 1 multiple ascending dose study to investigate the safety, pharmacokinetics, pharmacodynamics and preliminary efficacy of KD033 in subjects with metastatic or locally advanced solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT227.
Abstract LB091: Long-term statin use and risk of colorectal cancer: A prospective cohort study
Authors: Zhang Yin , Chan Andrew T. , Meyerhardt Jeffrey A. , Giovannucci Edward L.
Published: July 2021

Importance Statin use has been examined as a potential chemopreventive strategy against colorectal cancer (CRC), with conflicting findings reported. Prior studies have not been able to investigate this topic with adequate follow-up time. Objective To examine prospectively the association between long-term statin use and CRC risk. Design setting and participants The Nurses' Health Study (NHS) and Health Professionals Follow-Up Study (HPFS) are large prospective cohorts of US women and men. Participants who were alive and free of cancer at the time when information on their statin use was first assessed were eligible for inclusion (100,300 women and 47,991 men), and were followed for up to 26 years (NHS: 1994-2016; HPFS: 1990-2016). Exposures Overall status and duration of statin use. Main outcome and measures Cox proportional hazards models were used to estimate hazard ratios (HRs) and 95% confidence intervals (95% CIs) for exposures and CRC risk. Results We documented 2,924 incident CRC cases during follow-up. In fully-adjusted analyses, compared to participants who never used statins, longer duration of statin use was associated with higher risk of overall CRC (HR, 95%CI was 1.09, 0.98-1.20 for ever use; 1.06, 0.94-1.19 for 1 to 5 years; 1.10, 0.96-1.27 for 6 to 10 years; 1.10, 0.86-1.41 for 11 to 15 years; 1.56, 1.13-2.16 for >15 years; vs. never users, P=.02 for trend). The risk elevation was driven by proximal colon cancer (1.22, 1.05-1.41 for ever use; 1.17, 0.99-1.39 for 1 to 5 years; 1.21, 0.99-1.48 for 6 to 10 years; 1.27, 0.91-1.78 for 11 to 15 years; 2.22, 1.49-3.30 for >15 years; vs. never users, P<.001 for trend) rather than cancers of distal colon or rectum. The results became more robust in sensitivity analyses among participants with hypercholesterolemia. Conclusion and relevance This study does not support benefit of statin use in CRC chemoprevention. Long-term statin use was associated with increased CRC risk (predominantly driven by proximal colon cancer).

Citation Format: Yin Zhang, Andrew T. Chan, Jeffrey A. Meyerhardt, Edward L. Giovannucci. Long-term statin use and risk of colorectal cancer: A prospective cohort study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB091.
Abstract LB117: Identification of novel SOS1-K-Ras disruptors for NSCLC and colon cancers with K-Ras mutation
Authors: Sasmal Sanjita , Ramanathan Anuradha , Boorgu Venkatesham
Published: July 2021

KRAS along with other RAS genes represents the most prevalent oncogene in human cancers. Atypical Ras signaling has been identified in more than 30% of all human cancers with the most common being lung, colon, and pancreatic cancers. In particular, K-Ras has been identified as the most important Ras protein in cancer research, implicated in over 21% of human cancers. Current strategies drug only specific K-Ras mutants, leaving open an unmet need for new agents that can address a broader patient population. A new opportunity is emerging for the development of a therapeutic Pan Ras inhibitor by targeting the upstream guanine nucleotide exchange factor (GEF) protein SOS (Son of sevenless). Disruption of SOS1- K-Ras mutant interaction is a good strategy to inhibit activation of mutant K-Ras. SOS1 inhibition through the protein-protein interaction (PPI) disruption assists in keeping K-Ras in the GDP inactive, rather than its active GTP-bound state. Inhibition of SOS1 results in blockade of the RAS-MEK-ERK pathway and lowering proliferation, in both WT and all mutant Ras forms. Here we discuss our approach to target SOS1 to control aberrant Ras signaling in NSCLC and colon cancers. We report the identification of novel potent SOS1-K-Ras disruptors using the SOS1-K-Ras PPI TR-FRET assay. Multiple series have been identified, with a good SAR trend. We have identified low nanomolar potent compounds that translate to modulation of p-ERK in cell based assays in K-Ras mutant cell lines. These compounds exhibited good ADME properties like solubility and metabolic stability. Further profiling of these compounds is ongoing.

Citation Format: Sanjita Sasmal, Anuradha Ramanathan, Venkatesham Boorgu. Identification of novel SOS1-K-Ras disruptors for NSCLC and colon cancers with K-Ras mutation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB117.
Abstract LB141: Assessment of natural killer (NK) cell activity for immunotherapy using a novel flow cytometry-based killing assay
Authors: Bonnevier Jody , Goetz Christine , Peng Li , Van Etten Jamie , Faulkner Bora , Hammerbeck Christopher , Miller Ariel , Perell Gabriella , Johnson Christopher , Lomakin Joseph , Hermanson David , Flynn Kevin
Published: July 2021

Natural killer (NK) lymphocytes exhibit potent responses against transformed and pathogen-infected cells. NK cells can act rapidly as they monitor the bloodstream and tissues for tumorigenic and infected cells that lack “self” markers of MHC class I. Upon recognition of a foreign invader or tumor cell, NK cells release cytotoxic proteins such as interferon-gamma and granzyme B. Because of their ability to recognize stressed cells lacking “self” markers, human NK cells have shown promise as therapeutics in clinical trials to reduce disease burden in patients with therapy-resistant or advanced-stage blood cancers. With more than one hundred NK cell- and induced NK cell-based clinical trials ongoing for blood cancers and refractory solid tumors, there is an ever-increasing need for NK cell therapies to treat cancer and immune diseases. We sought to develop novel, clinically relevant, and robust methods for human NK cell expansion. In addition, to simplify the processing of cells for characterization, we developed a flow cytometry based killing assay to monitor NK activity. Therefore, within the same experiment, we can analyze NK cell phenotype profiles with specific antibody panels while also assessing NK cell killing activity. Using optimized serum- and xeno-free conditions with minimal cell manipulations, we have expanded highly purified NK cells up to 500-fold in 14 days from human peripheral blood mononuclear cells (PBMCs) using optimized cytokines and Cloudz CD2/NKp46 microspheres. To characterize the resulting NK cell population, we used a unique antibody panel to phenotype the NK cells by flow cytometry. Within the same experiment, NK killing assays against K562 cells can be performed using a flow cytometry-based killing assay, which has the advantage of using one instrument for all analyses, shortened processing times, and reproducibility. Using this workflow, we can identify different culture conditions that adversely affect NK activity. Together, this data presents a comprehensive and clinically relevant workflow for the expansion of highly purified NK cells and assessing their killing potential. Furthermore, this workflow can be used for screening the activity of chimeric antigen receptor (CAR) expressing immune cells against specific cancer antigens.

Citation Format: Jody Bonnevier, Christine Goetz, Li Peng, Jamie Van Etten, Bora Faulkner, Christopher Hammerbeck, Ariel Miller, Gabriella Perell, Christopher Johnson, Joseph Lomakin, David Hermanson, Kevin Flynn. Assessment of natural killer (NK) cell activity for immunotherapy using a novel flow cytometry-based killing assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB141.
Abstract LB170: HER2-targeted interferon-beta-1a mutein, a potent immunocytokine for the treatment of HER2-positive cancers
Authors: Lee Chan Gyu , Kim Tae Eun , Hong Sungyoul , Chu Jongwan , Kang Ju Eun , Jeong Hae Min , Shin Young Kee
Published: July 2021

Background: Interferon beta (IFN-β), a promising potent cytokine, has been attracting attention for treatment of cancer. The pleiotropic antitumor effects of IFN-β have been studied, with specific reference to their direct role on cancer cells and indirect action through the immune effector cells. However, its systemic toxicities and poor biophysical properties prevent IFN-β from being widely used for cancer therapy. Since the potential of cytokine therapies are impaired due to dose-limiting systemic toxicities, novel treatment methods should be developed to safely deliver effective drug quantities at the tumor sites. Like antibody-drug conjugates (ADCs), immunocytokine can be attempted to induce cytokine's organ-targeting and alleviate its systemic side effects. Here, we designed recombinant IFN-β mutein immunocytokines that comprise a HER2-targeting antibody and IFN-β mutein, and evaluate the antitumor properties against HER2-positive cancers.Method: A panel of human gastric cancer cell lines was treated with trastuzumab-IFN-β mutein to evaluate direct antitumor effect. In addition, to test the immune cell-mediated antitumor effect, cancer cells were co-cultured with effector cells (e.g. PBMC) in the absence or drug. The antitumor efficacy of trastuzumab-IFN-β mutein in vivo was tested in HER2-positive cancer xenograft models using nude mice or humanized mice. Result: Trastuzumab-IFN-β mutein directly inhibited the growth of HER2-positive gastric cancer cell lines and was more effective than trastuzumab or IFN-β mutein alone. Trastuzumab-IFN-β mutein also displayed enhanced immune cell-mediated cytotoxicity. Collectively, trastuzumab-IFN-β mutein may have indirect immune cell-mediated antitumor effects and direct cell growth inhibitory effects. Moreover, trastuzumab-IFN-β mutein significantly suppress tumor growth in HER2-positive cancer xenograft models. Tumor-infiltration of lymphocytes was enhanced by trastuzumab-IFN-β mutein, implying that the tumor-targeting IFN-β may have an antitumor effect through increased immune response. Conclusion: We have characterized and evaluated the antitumor properties of trastuzumab-IFN-β mutein in HER2-expressing cancers. Since IFN-β activates antitumor immune responses, it is expected to be administered in combination with immunotherapeutic drugs. Therefore, the study suggests that trastuzumab-IFN-β mutein is a promising candidate for the treatment of HER2-positive carcinoma.

Citation Format: Chan Gyu Lee, Tae Eun Kim, Sungyoul Hong, Jongwan Chu, Ju Eun Kang, Hae Min Jeong, Young Kee Shin. HER2-targeted interferon-beta-1a mutein, a potent immunocytokine for the treatment of HER2-positive cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB170.
Abstract LB189: Three-dimensional genome organization maps in normal haematopoietic stem cells and acute myeloid leukemia
Authors: Wang Benny , Kong Lingshi , Babu Deepak , Choudhary Ruchi , Fam Winnie , Tng Jia Qi , Goh Yufen , Liu Xin , Song Fang Fang , Chia Priscella , Chan Ming Chun , An Omer , Tham Cheng Yong , Benoukraf Touati , Yang Henry , Wang Wilson , Chng Wee Joo , Tenen Daniel , Fullwood Melissa J.
Published: July 2021

Many oncogenes, such as MEIS1 and HOXA9, are overexpressed in some but not all cancers. We identified two key epigenetic mechanisms underlying this heterogeneity in oncogene expression in Acute Myeloid Leukemia. Acute Myeloid Leukemia (AML) is a highly lethal blood cancer arising due to aberrant differentiation of haematopoietic stem cells. MEIS1 and HOXA9 regulate stemness-related transcriptional programs in normal haematopoietic stem cells and AML. Here we obtained 3D genome organization maps in the CD34+ haematopoietic stem cells from healthy individuals and individuals with AML. The MEIS1 oncogenic transcription factor is regulated by a Frequently Interacting Region (FIRE). This FIRE is present in normal bone marrow samples, and an AML sample with high MEIS1 levels. However, it is absent in two AML samples that show low MEIS1 levels. CRISPR excision of the FIRE led to loss of MEIS1 and reduced cell growth. Moreover, MEIS1 can bind to the promoter of HOXA9. HOXA9 can also auto-regulate by binding to its own promoter as well as an Acute Myeloid Leukemia-specific super-enhancer that interacts with the HOXA9 promoter via chromatin interactions. The mechanisms elucidated here could be potentially exploited to utilize epigenetic inhibitors to specifically target oncogene expression in cancer.

Citation Format: Benny Wang, Lingshi Kong, Deepak Babu, Ruchi Choudhary, Winnie Fam, Jia Qi Tng, Yufen Goh, Xin Liu, Fang Fang Song, Priscella Chia, Ming Chun Chan, Omer An, Cheng Yong Tham, Touati Benoukraf, Henry Yang, Wilson Wang, Wee Joo Chng, Daniel Tenen, Melissa J. Fullwood. Three-dimensional genome organization maps in normal haematopoietic stem cells and acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB189.
Abstract LB191: Downregulation of Stem-loop binding protein by nicotine and its role in nicotine-induced cell transformation in human lung epithelial cells
Authors: Sun Qi , Chen Danqi , Jin Chunyuan
Published: July 2021

The use of electronic-cigarettes has increased substantially in recent years among the younger population in particular. The liquid nicotine is the main component of the e-cigarette. Mice exposed to e-cigarette smoke (ECS) generated by heating the liquid nicotine developed lung adenocarcinoma and bladder hyperplasia. This implicated a potential role for ECS/nicotine in cancer development, while the underlying mechanisms remain unclear. Here we report that nicotine reduces the protein level of Stem-loop binding protein (SLBP), a critical factor for the stability, processing and translation of canonical histone pre-mRNAs, in human lung epithelial BEAS-2B cells. It has been known that canonical histone mRNAs typically do not end in a poly(A) tail and that depletion of SLBP results in polyadenylation of canonical histone mRNAs that appeared to be carcinogenic. Indeed, nicotine exposure induced polyadenylation of mRNAs for histones H2A, H2B, H3.1, H3.2, and H4. Interestingly, the inhibitor for nAChRs (nicotinic acetylcholine receptors) reversed nicotine-induced SLBP depletion. Moreover, the depletion of SLBP was attenuated not only by both the inhibitors and siRNAs specific for CDK1/2 and CK2, but also by inhibition of AKT signal pathway. Importantly, nicotine-induced anchorage-independent cell growth was rescued by overexpression of SLBP. These results suggest that activation of nAChR and a series of signal transduction pathways are involved in nicotine-mediated SLBP reduction and they may play an important role in nicotine-induced cell transformation and carcinogenesis.Keywords: Nicotine, Stem-loop binding protein, Cell transformation, nAChR

Citation Format: Qi Sun, Danqi Chen, Chunyuan Jin. Downregulation of Stem-loop binding protein by nicotine and its role in nicotine-induced cell transformation in human lung epithelial cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB191.
Abstract LB192: Sulforaphane suppression of Prmt5 and Mep50 function suppresses the mesothelioma cancer stem cell phenotype
Authors: Ezeka Geraldine C. , Adhikary Gautam , Kandasamy Sivaveera , Friedberg Joseph , Eckert Richard
Published: July 2021

Mesothelioma is an aggressive and fatal cancer of the mesothelial lining which is caused by asbestos exposure. The current treatment strategy is surgical resection followed by chemotherapy, but this is marginally successful and leads to drug resistant and recurrent disease. Mesothelioma cancer stem cells (MCS cells) comprise an important subpopulation of highly aggressive mesothelioma tumor cells that maintain tumor formation and mediate drug resistance. Our goal is to identify proteins that maintain the MCS cell and non-stem cancer cell survival as therapy targets. Protein arginine methyltransferase 5 (PRMT5) functions with the MEP50 cofactor to catalyze symmetric dimethylation of specific arginine resides in histones 3 and 4 to silence tumor suppressor gene expression and enhance cancer cell survival. Our studies show that loss of PRMT5 or MEP50 reduces H4R3me2s formation in MCS cells and that this is associated with a reduction in MCS cell spheroid formation, invasion and migration. Additionally, treatment with sulforaphane (SFN), a promising diet-derived cancer prevention and therapy agent, reduces PRMT5/MEP50 level and H4R3me2s formation, and suppresses the MCS cell phenotype. Moreover, forced expression of PRMT5/MEP50 antagonizes SFN suppression of the MCS cell phenotype, suggesting that loss of PRMT5/MEP50 is required for SFN action. SFN treatment also markedly reduces tumor formation and this is associated with reduced PRMT5/MEP50 expression and H4R3me2s formation. These findings suggest that SFN suppresses PRMT5/MEP50 function to attenuate the MCS cell phenotype and reduce tumor formation, and that SFN may be a useful mesothelioma prevention and therapy agent.

Citation Format: Geraldine C. Ezeka, Gautam Adhikary, Sivaveera Kandasamy, Joseph Friedberg, Richard Eckert. Sulforaphane suppression of Prmt5 and Mep50 function suppresses the mesothelioma cancer stem cell phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB192.
Abstract LB205: The IRF8-MEF2D transcription factor circuit regulated by a druggable multiple post-translational modification (PTM) reader ZMYND8 in acute myeloid leukemia
Authors: Cao Zhendong , Budinich Krista A. , Huang Hua , Lu Bin , Zhang Zhen , Ren Diqiu , Zhou Yeqiao , Huang Yuhan , Pingul Bianca , Kingsley Molly C. , Lenard Alexandra K. , Qi Jun , Carroll Martin P. , Blobel Gerd A. , Faryabi Robert B. , Bernt Kathrin M. , Berger Shelley L. , Shi Junwei
Published: July 2021

The transformed state in acute leukemia requires gene regulatory programs involving transcription factors (TFs) and chromatin modulators. Here, we uncover an IRF8-MEF2D TF regulatory circuit as an acute myeloid leukemia (AML)-specific dependency. We discover and characterize the mechanism by which the chromatin ‘reader' ZMYND8 directly activates IRF8 in parallel with the MYC proto-oncogene through their AML-specific enhancers. ZMYND8 is essential for AML proliferation in vitro and in vivo and associates with MYC and IRF8 enhancer elements that we define in cell lines and in patient samples. ZMYND8 occupancy at IRF8 and MYC enhancers requires BRD4, a transcriptional coactivator essential for AML proliferation. We show that ZMYND8 binds to the ET domain of BRD4 via its chromatin reader cassette, which in turn is required for proper chromatin occupancy and maintenance of leukemic growth in vivo. Our results rationalize ZMYND8 as a potential selective therapeutic target for modulating essential transcriptional programs in AML.

Citation Format: Zhendong Cao, Krista A. Budinich, Hua Huang, Bin Lu, Zhen Zhang, Diqiu Ren, Yeqiao Zhou, Yuhan Huang, Bianca Pingul, Molly C. Kingsley, Alexandra K. Lenard, Jun Qi, Martin P. Carroll, Gerd A. Blobel, Robert B. Faryabi, Kathrin M. Bernt, Shelley L. Berger, Junwei Shi. The IRF8-MEF2D transcription factor circuit regulated by a druggable multiple post-translational modification (PTM) reader ZMYND8 in acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB205.
Abstract LB212: The VGLL4 tumor suppressor inhibits transglutaminase-2 dependent YAP1-TEAD signaling to attenuate the cancer phenotype
Authors: Mickle McKayla B. , Adhikary Gautam , Xu Wen , Shrestha Suruchi , Eckert Richard L.
Published: July 2021

Epidermal squamous cell carcinoma (SCC) is among the most common and aggressive forms of cancer. We previously showed that the pro-cancer transcription factor, YAP1, drives an aggressive SCC cancer phenotype by binding to nuclear TEAD transcription factors to activate pro-cancer gene expression. Vestigial-like protein 4 (VGLL4) is a tumor suppressor that is downregulated in epithelial cancers. It binds to TEAD factors to displace YAP1 binding to reduce YAP1-dependent transcription and target gene expression. The present study shows that VGLL4 overexpression reduces SCC cell TEAD-dependent transcription to reduce expression of mRNA and protein encoding YAP1-target genes (CTGF, CYR61, CCND1), and pro-cancer collagen genes (COL1A2, COL3A1) leading to reduced cell proliferation, spheroid formation, invasion and migration. Identical results were observed in response to transient and stable VGLL4 overexpression. Similarly, VGLL4 expression reduces YAP1 activity in tumors leading to reduced YAP1 target and pro-cancer collagen gene expression and reduced and EMT marker proteins levels which is associated with reduced tumor formation. Loss of YAP1 responsive gene expression appears to be essential for VGLL4 attenuation of the aggressive phenotype, as forced overexpression of CCND1 or COL1A2 antagonizes VGLL4 action. These findings suggest VGLL4 functions as a tumor suppressor in SCC by repressing YAP1/TEAD signaling and consequently attenuating the SCC phenotype.

Citation Format: McKayla B. Mickle, Gautam Adhikary, Wen Xu, Suruchi Shrestha, Richard L. Eckert. The VGLL4 tumor suppressor inhibits transglutaminase-2 dependent YAP1-TEAD signaling to attenuate the cancer phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB212.
Abstract LB241: Genomic and molecular features of metachronous metastasis in clear cell renal cell carcinoma
Authors: Naeini Marjan Mojtabavi , Rohatgi Neha , Ghoshdastider Umesh , Hakimi Ari A. , Skanderup Anders Jacobsen
Published: July 2021

Molecular features associated with risk of metachronous metastases (MM) in clear cell renal cell carcinoma (ccRCC) are poorly defined. Here, we performed systematic genomic and molecular analysis of 192 ccRCC primary tumors with extended clinical follow-up for development of MM. Whole-exome sequencing analysis revealed no association between somatic mutations and development of MM. However, chromosomal deletions at 1p33 were more prevalent in tumors that later progressed with MM. We inferred immune cell infiltrates and found that depletion of regulatory T cells and M1 macrophages were associated with development of MM. Next, we used bulk tumor transcriptome deconvolution to estimate cancer (malignant) and stromal (non-malignant) cell gene expression differences between MM and indolent primary tumors. Downregulation of the Crumbs complex component PATJ (1p31) in cancer cells of MM tumors suggested loss of apicobasal cell polarity as a risk factor in metastatic progression. We also identified downregulation of fatty acid degradation in cancer cells of MM tumors. Overall, our results nominate novel genomic and molecular features underpinning metachronous metastasis in ccRCC.

Citation Format: Marjan Mojtabavi Naeini, Neha Rohatgi, Umesh Ghoshdastider, Ari A. Hakimi, Anders Jacobsen Skanderup. Genomic and molecular features of metachronous metastasis in clear cell renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB241.
Abstract LB244: The role of stromal Per2 in regulation of tumorigenesis
Authors: Mayer Shimrit , Shaashua Lee , Lior Chen , Lavon Hagar , Novoselsky Alexander , Scherz-Shouval Ruth
Published: July 2021

The tumor microenvironment (TME) comprised of multiple proteins, extracellular matrix(ECM) and various cell types that together promote tumor initiation, progression andhomeostasis in a cell-cell communication manner. In this study, we set to understand howthe stromal circadian clock supports the cancer cells, specifically the contribution of thecore clock gene Per2 to tumor initiation and progression. The circadian clock is anendogenous, evolutionally conserved and ubiquitously expressed pacemaker, consisting ofcell autonomous clocks and a central pacemaker located in the hypothalamussuperchiasmatic nucleus (SCN). Together these clocks synchronize numerous biologicalprocesses between the organism and its environment. Amongst these processes are DNAdamage repair, metabolism, and cell cycle. Several studies showed that disruption ofcircadian rhythms has been associated with various forms of cancer in humans and mice.While Per2 was previously suggested to be a tumor suppressor, in this study we found thatthe core clock gene Per2 in the TME is essential for tumor initiation and metastaticcolonization. Another core gene, Per1, is dispensable. We further showed that lossof Per2 in the TME leads to transcriptional rewiring at early stages of metastasesformation, and suppresses subsequent metastatic tumor progression. Thus, our resultsunravel an unexpected protumorigenic role for the core clock gene Per2 in the TME. Thesefindings may imply a non-circadian role for Per2, differentiating it from Per1 withpotential implications for therapeutic dosing strategies and treatment regimens.

Citation Format: Shimrit Mayer, Lee Shaashua, Chen Lior, Hagar Lavon, Alexander Novoselsky, Ruth Scherz-Shouval. The role of stromal Per2 in regulation of tumorigenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB244.
Abstract LB245: TRIM29 mediates metastasis though hypoxia-regulated proteins in microsatellite-stable and p53-mutated colorectal cancers
Authors: Agarwal Sumit , Behring Michael , Bajpai Prachi , Elkohly Amr , Kim Hyung-Gyoon , Chandrashekar Darshan S. , Gupta Nirzari , Al Diffalha Sameer , Varambally Sooryanarayana , Manne Upender
Published: July 2021

In the USA, colorectal cancer (CRC) is a leading cause of cancer deaths in men and women. Two primary molecular subtypes of CRCs are those with microsatellite instability (MSI) and those that are microsatellite stable (MSS). MSI-CRCs have a dysfunction of the mismatch repair (MMR) genes and are less aggressive; MSS-CRCs have active mismatch repair genes, account for 85% of sporadic CRCs, and are more aggressive. CRCs with MSI have a favorable prognosis and are candidates for immunotherapy; those with MSS have a poor prognosis and are resistant to various therapies. Our previous gene expression profiling studies of CRCs showed an association of high expression of tripartite motif-containing 29 (TRIM29) with MSS and p53 mutations. TRIM29, also known as ataxia telangiectasia group D-complementing (ATDC) protein, binds to p53 and inhibits its function. Since point mutations of p53 occur in 50% of CRCs and are associated with the MSS phenotype and with aggressiveness of CRCs, we assessed TRIM29 expression in CRC cells exhibiting various microsatellite and p53 status. With RNA sequencing and qPCR validation, we found higher expression of TRIM29 in MSS and p53-mutated types of CRC cells (HT29 and SW480) as compared to cells exhibiting MSI and the p53 wild-type (HCT116, LOVO, RKO and LS174T) as well as cells with MSS and the p53 wild-type (T84). TRIM29 knockdown studies with HT29 and SW480 CRC cells showed that TRIM29 contributes to the aggressive progression of cells with MSS and p53 mutations. Preclinical studies with experimental animals demonstrated that MSS and p53-mutated CRC cells (HT29) with high expression of TRIM29 metastasized more extensively to liver and bone relative to cells with stably inhibited TRIM29. Further, gene expression profiling of stably inhibited TRIM29 cells showed downregulation of P4HA1, ALDOC, and hexokinase2, enzymes that are involved in the hypoxia pathway. In summary, the present studies show that overexpression of TRIM29 contributes to CRC proliferation, invasion, migration, tumor growth, and experimental metastasis. Thus, targeting of TRIM29 with small molecule inhibitors could be an effective strategy for treatment of CRCs with MSS and p53 mutations. This work was supported in part by an NIH grant (3U54CA118948) and by a grant from the ELKUS foundation.

Citation Format: Sumit Agarwal, Michael Behring, Prachi Bajpai, Amr Elkohly, Hyung-Gyoon Kim, Darshan S. Chandrashekar, Nirzari Gupta, Sameer Al Diffalha, Sooryanarayana Varambally, Upender Manne. TRIM29 mediates metastasis though hypoxia-regulated proteins in microsatellite-stable and p53-mutated colorectal cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB245.
Abstract LB248: Implantation factors assist in the growth of gynecologic tissues in ectopic locations
Authors: Sunde Jan , O'Neil Derek , Katz Tiffany , Wasickanin Morgan
Published: July 2021

OBJECTIVE: Develop a model of endosalpingiosis and endometriosis to investigate the mechanisms which facilitate implantation of exfoliated cells in the peritoneum.

BACKGROUND: Endosalpingiosis (ES) and endometriosis (EM) are the growth of tubal and endometrial epithelium respectively, outside of their site of origin and have been associated with ovarian cancer. Why this growth happens has not been elucidated since Sampson first proposed the concept of retrograde menstruation nearly 100 years ago. We hypothesized there is retrograde flow of uterine implantation factors which drive ectopic growth. To test this hypothesis, we developed a transgenic fluorescent murine model of ES and EM that successfully demonstrated peritoneal implantation in mice. Using the model, we explored possible influences on implantation.

METHODS: tdTomato (tdT) transgenic fluorescent mice were used as donors to allow for visualization of tissues in recipient mice. A progesterone knockout (PKO) of the tdT mouse that blocks implantation factors was created by giving 0.1 mg/kg progesterone on postnatal days 2-10. Metestrus endometrium was induced hormonally in donor tdT and wild-type (WT) female C57BL/6J mice. The tdT oviductal cells and endometrium were then harvested for intraperitoneal injection into synchronized recipient WT mice, receiving minced endometrium alone, minced oviduct alone, or a combination of minced oviduct and WT endometrial tissue (for implantation factors) from non-treated or PKO tdT donors. Ectopic lesions were identified using fluorescence in-vivo imaging and UV light.

RESULTS: PKO tdT oviduct had significantly reduced implantation (4 lesions/6 mice, p<0.02) compared to tdT oviduct without treatment (28 lesions/8 mice). Additionally, PKO tdT oviduct implantation (10 lesions/3 mice) may be restored when implanted with WT endometrium (p=0.07). Chi square was used to evaluate for statistical significance.

ConclusionImplantation factors required for blastocyst implantation are crucial to initiate ectopic tissue growth. We have developed a novel mouse model of ectopic growth of gynecologic tissues which demonstrates that the removal of implantation factors from endometrium and oviduct reduces ectopic lesions. This model allows us to study the initial step of implantation of these lesions that could give rise to malignancies which will ultimately allow for the development of new prevention and treatment modalities.

Citation Format: Jan Sunde, Derek O'Neil, Tiffany Katz, Morgan Wasickanin. Implantation factors assist in the growth of gynecologic tissues in ectopic locations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB248.