bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A081: Patient-derived xenograft models from hematological malignancies for preclinical drug development and biomarker research
Authors: Siegert Antje , Brzezicha Bernadette , Fuhrmann Stephan , Janz Martin , Schmitt Clemens , Keller Ulrich , Rupp Christian , Walther Wolfgang , Hoffmann Jens
Published: December 2023

Background: Acute leukemias and lymphomas encompass a diverse group of hematologic malignancies, posing significant challenges in clinical practice. They often exhibit resistance to standard-of-care (SoC) treatments and have a high recurrence rate. Recent advancements in molecular profiling have identified potential drivers for various leukemia and lymphoma subtypes, offering new therapeutic targets. However, the validation of these targets and the development of drugs rely heavily on preclinical models that faithfully represent the different clinical subtypes. These models play a crucial role in refining target selection and facilitating drug development projects.

Methods: AML- and ALL-PDX were derived from bone marrow aspirates or peripheral blood samples, from primary or relapsed acute leukemia patients. Purified cells were transplanted either intravenously (i.v.) and/or subcutaneously (s.c.) into immunodeficient mice. Some mice developed a systemic AML, which was monitored by flow cytometric analysis of blood samples. Non-Hodgkin (NHL) - or Hodgkin lymphoma (HL)-PDX were derived from peripheral blood, lymph node extirpations or core needle biopsies, and were usually transplanted subcutaneously into immunodeficient mice.

Results: More than 45 PDX models from AML, ALL, NHL and HL have been successfully established and characterized. Highly individual response rates to SoC and investigational drugs were observed and correlated with mutation and gene expression data.  An individual Human Leucocyte Antigen (HLA) profile of each PDX was determined by RNA-seq and a comprehensive HLA matching analysis of the models and peripheral blood mononuclear cell (PBMC) donors was performed to enable personalized, preclinical immuno-oncology studies.

Conclusions: Our extensively characterized PDX models derived from hematologic malignancies serve as valuable tools for evaluating novel targeted and immunological therapies. These models provide an exceptional platform for identifying and validating new targets, as well as preclinical screening of compounds and combinations for translational research projects in this field.

Citation Format: Antje Siegert, Bernadette Brzezicha, Stephan Fuhrmann, Martin Janz, Clemens Schmitt, Ulrich Keller, Christian Rupp, Wolfgang Walther, Jens Hoffmann. Patient-derived xenograft models from hematological malignancies for preclinical drug development and biomarker research [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A081.
Abstract A104: ADAR1-associated metabolic vulnerabilities in triple-negative breast cancer
Authors: Kung Che-Pei , Shriver Leah P , Terzich Nick , Ilagen Ma Xenia G , Prinsen Mike , Ryu Sua , Kladney Raleigh D , Patti Gary J , Maggi Leonard B , Weber Jason D
Published: December 2023

Triple-negative breast cancer (TNBC), lacking expression of estrogen, progesterone, and HER2 receptors, is aggressive and lacks targeted treatment options. Therefore, the mission to identify actionable targets remains the top priority. We recently demonstrated that an RNA editing enzyme, adenosine deaminase acting on RNA 1 (ADAR1), plays important roles in TNBC’s ability to initiate and maintain tumorigenesis. We posit that ADAR1 functions as a homeostatic factor protecting TNBC from stresses when otherwise kept in check, contributing to tumorigenesis. By both targeted and unbiased approaches, we identified two metabolic vulnerabilities, 1) iron-dependent cell death pathway, ferroptosis, and 2) serine addiction, associated with ADAR1 in TNBC. By performing single-reaction monitoring-based liquid chromatography coupled to mass spectrometry (LC-MS) to measure intracellular lipid contents, we showed that ADAR1 loss in TNBC cells increased the abundance of polyunsaturated fatty acid phospholipids (PUFA-PL), of which peroxidation is the primary driver of ferroptosis. Transcriptomic analysis and phenotypic drug screen using a ferroptosis-focused library were performed to identify underlying mechanisms and drug candidates synergizing with ADAR1 loss through ferroptosis, respectively. Meanwhile, an unbiased screen using a FDA-approved compound library revealed that ADAR1 loss also sensitizes TNBC cell line MDA-MB231 to inhibition of the serine biosynthesis pathway. We showed that ADAR1 knockdown leads to altered serine transport, and a metabolic flux analysis using labeled (U-13C) glucose will be performed to determine if ADAR1 loss affects de novo serine biosynthesis and one-carbon metabolism. By testing the hypothesis that ADAR1 regulates the metabolic fitness of TNBC by desensitizing ferroptosis and serine addiction, we aim to leverage these metabolic vulnerabilities to inform basic, pre-clinical, and clinical studies to develop novel therapeutic strategies for TNBC.

Citation Format: Che-Pei Kung, Leah P Shriver, Nick Terzich, Ma Xenia G Ilagen, Mike Prinsen, Sua Ryu, Raleigh D Kladney, Gary J Patti, Leonard B Maggi, Jason D Weber. ADAR1-associated metabolic vulnerabilities in triple-negative breast cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A104.
Abstract A109: Targeting pyrimidine biosynthesis as a metabolic vulnerability in brain metastasis
Authors: Mobilio Daniel , Singh Sheila K , Chafe Shawn C
Published: December 2023

Metastasis, the spread of cancer cells from one part of the body to another, is the leading cause of death among cancer patients. Lung-to-Brain metastasis remains a largely understudied disease that possesses a dismal prognosis due to the lack of effective therapies. We previously identified purine metabolism as a metabolic vulnerability of brain metastases.  Here, we sought to identify whether pyrimidine biosynthesis is also a metabolic dependency of metastatic brain tumours.  We identify dihydroorotate dehydrogenase (DHODH) as a cancer-selective targetable vulnerability.  DHODH is an essential enzyme that is located within the inner membrane of the mitochondria which has a primary role in producing pyrimidine metabolites through the de novo uridine biosynthesis pathway and also contributes to the proper functioning of the electron transport chain.  Knock-out of DHODH with CRISPR-Cas9 or pharmacological inhibition with BAY2402234 decreased cell viability, sphere formation, and proliferation of patient-derived lung-to-brain (LBM) metastasis cells in vitro. Furthermore, suppression of DHODH expression or BAY2402234 treatment reduced growth of intracranial LBM tumours and significantly extended survival.  Mechanistically, we identify interfering with DHODH activity abrogates normal mitochondrial function leading to cleavage of gasdermin E and induction of pyroptosis. Taken together, this study highlights the promise of DHODH inhibitors in targeting brain metastasis.

Citation Format: Daniel Mobilio, Sheila K Singh, Shawn C Chafe. Targeting pyrimidine biosynthesis as a metabolic vulnerability in brain metastasis [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A109.
Abstract A118: Somatic mutations in the PIK3CA gene and its prognostic implications among Ethiopian breast cancer patients
Authors: Woldesonbet Zelalem Desalegn , Yohannes Meron , Addissie Adamu , Abebe Tamrat , Kantelhardt Eva Johanna , Vetter Martina
Published: December 2023

Purpose: The phosphatidylinositol-4, 5-bisphosphate 3-kinase (PIK3CA), catalytic subunit alpha (c) is frequently mutated in breast cancer (BC). Although the biological impact of PIK3CA mutations in BC has been studied in various parts of the world, limited data is available from sub-Saharan Africa. In this study, the frequency and the prognostic role of PIK3CA mutations were investigated among BC patients from Ethiopia.

Methods: Formalin-fixed paraffin-embedded blocks and fresh frozen BC tissue samples were collected from patients newly diagnosed with BC. Histopathological evaluation was carried out using immunohistochemistry (IHC). PIK3CA mutation profiling was carried out using TaqMan® Mutation detection assay. The association between PIK3CA mutation status and clinical and histopathological features was evaluated. A survival regression (Cox’s model) was employed to assess the effect of PIK3CA mutation and other variables on survival.

Results: The prevalence of the PIK3CA mutations was 20.4%.  Mutation of the PIK3CA gene was mainly observed among younger (24%), early pT stage (19.4%), and G1/G2 tumors (22.3%). The proportion of mutation was 24.4%, 27.3% and 20% in ER+, PR+ and HER2- tumors, respectively. A marked difference was observed in the PIK3CA mutation frequency across the four major tumor subgroups. However, most of the mutated tumors were found among HR+/HER2− (24.1%) followed by HR+/HER2+ tumors (27.8%). In his study, only TNBC was negatively associated with the proportion of mutation in the PIK3CA gene (p-value= 0.02) in comparison to HR+/HER2- tumors. In multivariate Cox’s regression analysis, the advanced pT stage and HR−/HER2- IHC group independently predicted worse overall survival. However, PIK3CA mutation status was not associated with survival.

Conclusions: The PIK3CA mutation was relatively high in HR+ tumors. Mutation rates of PIK3CA in Ethiopian BC tumors are similar to those reported for other populations.

Citation Format: Zelalem Desalegn Woldesonbet, Meron Yohannes, Adamu Addissie, Tamrat Abebe, Eva Johanna Kantelhardt, Martina Vetter. Somatic mutations in the PIK3CA gene and its prognostic implications among Ethiopian breast cancer patients [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A118.
Abstract A142: Fragment based discovery of inhibitors of the eIF4E:eIF4G interaction
Authors: Richardson Caroline J , Vincovic Mladen , East Charlotte , Wallis Nicola , Ward George , Woodhead Andrew
Published: December 2023

eIF4E is the rate limiting factor for cap-dependent protein translation, forming an essential part of the translation initiation complex, eIF4F and is a key signalling convergence point for the major oncogenic pathways MAPK and PI3K/AKT/MTOR. eIF4E has been shown to be overexpressed in cancer, which has been associated with poor prognosis and resistance to chemotherapeutics and targeted agents such as mTOR and MAPK inhibitors. eIF4E is therefore a promising target for cancer treatment but has so far been difficult to target with small molecules due to a lack of druggable pockets. Fragment based crystallographic screening is a powerful technique for probing the surface of proteins to identify potentially druggable binding sites. Here, we describe our fragment-based approach and hit optimisation campaign to generate potent compounds binding to eIF4E  For the fragment screen, we generated an engineered form of eIF4E, where the N-terminal (residues 1-35) were removed and replaced a short, tethered peptide based on the canonical binding sequence of the eIF4E binding protein 4E-BP1, and screened 1400 fragments using a combination of Xray crystallography and NMR. Several low affinity (mM) fragment hits were identified, binding to both the mRNA cap- binding site and at an additional binding site of unknown functional relevance. Using a combination of biophysical and biochemical assays, subsequent rounds of iterative structure-based drug design of yielded potent lead molecules against both binding sites. For the cap-binding site, lead compounds of low nM potency were generated but due to the polar nature of the binding site, the molecules had very poor permeability properties and did not inhibit cell proliferation. For the non-canonical binding site, lead compounds with potency of 100nM alongside less potent enantiomers were generated which provided useful tools for probing the function of the binding site in vitro. We developed a MSD (Meso-scale discovery) assay to determine that the lead compound and not the inactive enantiomer could disrupt the eIF4E:eIF4G interaction in HeLa cell lysates with an IC50 of 1uM. Using a HeLa extract in vitro translation assay, we could also demonstrate that both the cap site and non-canonical site lead compounds could inhibit cap-dependent but not IRES driven protein translation. This work demonstrates the power of fragment screening to identify novel functional pockets on difficult to drug proteins such as eIF4E.

Citation Format: Caroline J Richardson, Mladen Vincovic, Charlotte East, Nicola Wallis, George Ward, Andrew Woodhead. Fragment based discovery of inhibitors of the eIF4E:eIF4G interaction [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A142.
Abstract A167: Cellular fitness of MYC-driven cancer cells to genetic and pharmacologic perturbations in normoxia, hypoxia and 3D culture
Authors: Yang Jun
Published: December 2023

MYC and hypoxia gene clusters are two most commonly recurring transcriptional programs in a heterogenous tumor. Understanding the cellular fitness of MYC-driven cancer cells in normoxia and hypoxia may lead to identification of cancer cell vulnerabilities. Using genome-wide CRISPR screenings of MYC-driven liver cancer cells cultured in 21% and 1% oxygen in monolayer, and 21% oxygen in 3D spheroid over 4 weeks, we not only identify many targetable essential genes under all three conditions but also context-specific fitness genes and pathways. In parallel, we perform drug screening under three conditions. We find that cells respond to 125 FDA approved oncology drugs in different ways in normoxia, hypoxia and 3D. Our resource study highlights unique epigenetic and metabolic dependency of MYC-driven cancer cells in different tumor environments.

Citation Format: Jun Yang. Cellular fitness of MYC-driven cancer cells to genetic and pharmacologic perturbations in normoxia, hypoxia and 3D culture [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A167.
Abstract A168: T cell receptor-based bispecific molecules targeting KRAS neoantigen cancer driver mutations
Authors: Whale Andrew D , Aleksic Milos , Karuppiah Vijaykumar , Moureau Sylvie , Chillakuri Chandramouli
Published: December 2023

KRAS is the most frequently mutated oncogene. Tumour specific neoantigen peptides derived from KRAS are presented by cell surface human leucocyte antigens (HLA) and form a class of shared, public tumour-specific antigens for T cell receptors (TCR) that are attractive targets for immunotherapy. We have previously reported engineering of a soluble T cell engaging ImmTAC (Immune mobilising monoclonal TCR Against Cancer) molecule, IMC-KRASG12D specific for KRASG12D peptide presented by HLA-A*11:01. IMC-KRASG12D was extensively characterised and demonstrated potent redirection of T cell cytotoxicity towards target KRASG12D presenting cancer cells, while sparing normal cells. Here, we focus on TCRs isolated from healthy donors that specifically recognise KRASG12V peptides presented in the context of HLA-A*11:01 or HLA-A*03:01. The affinity of several TCRs was enhanced by phage display, retaining the remarkable ability to distinguish between KRASG12V and KRASWT peptides. In cell-based assays, ImmTAC molecules containing these TCR mediate T cell activation in response to cells expressing KRASG12V but not KRASWT. Interestingly, although structural, biochemical, and computational approaches can explain the molecular mechanism underlying TCR selectivity for KRASG12D pHLA-A*11complexes, it remains unclear how TCR selectivity to KRASG12V is achieved. These studies demonstrate that affinity enhanced TCR can maintain selectivity for target peptides that differ from self-antigens by only a single amino acid, to enable targeting of public neoantigens.

Citation Format: Andrew D Whale, Milos Aleksic, Vijaykumar Karuppiah, Sylvie Moureau, Chandramouli Chillakuri. T cell receptor-based bispecific molecules targeting KRAS neoantigen cancer driver mutations [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A168.
Abstract B017: Clinical utility of next generation sequencing in advanced colorectal cancer: The earlier the better
Authors: An Ho Jung , Kim Hyunho , Park Hyung Soon , Cho Hyeon-Min , Kye Bong-Hyeon , Yoo Ri Na , Moon Ji-Yeon , Kim Sung Hwan , Lee Jonghoon , Lee Hyo Chun
Published: December 2023

Background. Since next generation sequencing (NGS) was approved in solid tumor in Korea, it has been usually performed when patients were progressed on standard chemotherapy in advanced colorectal cancer (CRC).  Recently, new targeted agents have been developed rapidly, so the NGS is recommended as early as possible. In this study, we analyzed NGS results and investigated clinical benefit in advanced CRC.

Patient and method. Between October 2018 to December 2022, NGS data in 85 advanced CRC patients in St. Vincent’s Hospital, Suwon, Korea were retrospectively analyzed.

Results. Seven patients had metachronous, and others had synchronous metastasis. The median age was 58 (34-84), and 59 (69.4%) were male. Fourteen (16.5%) were right-side, 40 (47.1%) were left-side colon, and 30 (35.3%) were rectum. The median time from diagnosis of advanced CRC to NGS was 13 (0-81) months, and median time to death after NGS was 8 (0.3-43.0) months. NGS platform were ‘Oncomine comprehensive assay Plus’ in 71 (83.5%), and the others were various platforms in K-master project. NGS analyses revealed new therapeutic target in 17 (20%): high tumor mutational burden (n = 2), KRAS G12C (n = 4), KRAS (n = 1) or BRAF (n = 2) mutation not routinely screened at initial, BRAF class II mutation (n = 2), Her2/3 mutation (n = 2), and NTRK fusion (n = 1). Among those, two patients with KRAS G12C mutation enrolled in clinical trial of KRAS G12 inhibitor, and one patient with NTRK fusion received NTRK inhibitor (3.5% of total). Among RAS wild type population (n=30), we identified hyperselected patients (n = 18, [60%], absence of either KRAS, NRAS, PTEN, and extracellular domain EGFR mutations, HER2 and MET amplifications, and ALK, RET or NTRK1 fusions by PARADIGM trial), which showed longer survival in cetuximab-treated population (p<0.001). Specific mutation including KRAS, NRAS, BRAF, TP53, or APC did not impact on prognosis. High TMB (≥10Mut/Mb) were associated with longer survival (p = 0.020). For patients with oligometastasis, hyperselected patients (p = 0.062) showed good prognosis.

Conclusion. The NGS results provided additional predictive or prognostic information in advanced CRC patients. Application of NGS as early as possible might help to make a clinical decision.

Citation Format: Ho Jung An, Hyunho Kim, Hyung Soon Park, Hyeon-Min Cho, Bong-Hyeon Kye, Ri Na Yoo, Ji-Yeon Moon, Sung Hwan Kim, Jonghoon Lee, Hyo Chun Lee. Clinical utility of next generation sequencing in advanced colorectal cancer: The earlier the better [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B017.
Abstract B019: RNA transcriptome profiling in microsamples of blood
Authors: Chenchik Alex , Kobzik Lester , Makhanov Mikhail , Diehl Paul
Published: December 2023

Multi-omic analysis of microsamples of lancet-induced blood drops allows frequent capture and quantitation of numerous metabolites, lipids, cytokines, and proteins. Microsample-based transcriptome profiling would facilitate use of RNA biomarkers in diagnosis and treatment of immunotherapy patients, but a suitable method has not been available. We tested a targeted  sequencing protocol for this purpose in human blood microsamples. We tested normal blood collected either by phlebotomy into standard vacutainer tubes or by absorption of 30 uL of blood onto a Mitra device pre-treated with an RNA-stabilization reagent. We tested detection of gene expression changes by incubating anti-coagulated blood for 18 hours with endotoxin followed by isolation of identical volumes of blood using a standard method (Qiagen kit) or from Mitra devices air dried for 24 h to mimic a home-use scenario. RNA was extracted from the microsamplers. After RNA purification, a panel of 274 immune/inflammatory genes was quantified by NGS after PCR using targeted primers following the Cellecta DriverMap protocol, which avoids counting of abundant rRNA and mitochondrial sequences. The targeted sequencing results were normalized to counts per million and used to compare results in standard vs microsamples.  We found robust detection of gene expression and correlation using the two methods in both unstimulated (r = 0.94) and endotoxin-stimulated blood. The differentially expressed genes (DEGs) identified in both standard and microsample methods showed high overlap with DEGs reported in public datasets in similar experiments. We conclude that RNA transcriptome profiling using targeted sequencing allows sensitive detection of gene expression levels in normal and activated blood samples. Because microsamples can be air-dried and mailed in for analysis, this approach has great promise for simple and repeated monitoring of RNA biomarkers in immunotherapy.

Citation Format: Alex Chenchik, Lester Kobzik, Mikhail Makhanov, Paul Diehl. RNA transcriptome profiling in microsamples of blood [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B019.
Abstract B052: Small molecule RPA inhibitors abrogate the ATR kinase signaling pathway
Authors: Jordan Matthew R , Ainembabazi Diana , VanderVere-Carozza Pamela S , Gavande Navnath S , Pawelczak Katherine S , Turchi John J
Published: December 2023

Replication Protein A (RPA) is the major single-stranded DNA (ssDNA) binding protein in the cell and plays a role in replication, recombination, and the DNA damage response (DDR) and repair. RPA functions to protect ssDNA from degradation and also serves as an interaction hub to recruit replication and DDR machinery. Under normal cellular conditions, RPA protein abundance is in excess to that of the ssDNA generated during S phase replication so as to protect from replication fork collapse and replication catastrophe in the event of elevated levels of replication stress. RPA inhibition is thus a promising cancer therapeutic strategy that targets oncogenic cells with increased replication stress due to rapid cell proliferation, genotoxic stressors, DDR inhibition, etc, and therefore an increased ssDNA burden. We have previously developed the RPA inhibitor (RPAi) NERx329 that disrupts RPA-ssDNA binding and exhibits single agent anti-cancer activity in vitro and in vivo presumably due to chemical RPA exhaustion. When a cell experiences extensive replication stress, elevated levels of RPA-bound ssDNA are recognized by the kinase ataxia telangiectasia and Rad2-related (ATR) through the ATR interacting protein (ATRIP). ATR is then activated through interactions with DNA topoisomerase 2-binding protein 1 (TopBP1) and phosphorylates downstream targets largely to direct the G2/M checkpoint signaling pathway through checkpoint kinase 1 (CHK1). In this work, we biochemically reconstitute this signaling pathway in order to evaluate an RPAi mechanism of action. We utilize a ssDNA substrate that mimics a stalled replication fork and demonstrate the RPA-dependence of ATR activation by monitoring phosphorylation of multiple ATR substrates: RPA, TopBP1, and p53. We then show that RPAi disrupt ATR kinase activity. Collectively, this works describes a novel mechanism of action of RPAi whereby RPA inhibition results in replication catastrophe by blocking the ATR activity and the G2/M checkpoint pathway.

Citation Format: Matthew R Jordan, Diana Ainembabazi, Pamela S VanderVere-Carozza, Navnath S Gavande, Katherine S Pawelczak, John J Turchi. Small molecule RPA inhibitors abrogate the ATR kinase signaling pathway [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B052.
Abstract B060: Dual blockade of BRD4 and the ATR/WEE1 pathway exploits ARID1A loss in clear cell ovarian cancer
Authors: Kinose Yasuto , Xu Haineng , Kumar Sushil , Kim Hyoung , Medvedev Sergey , Wang Xiaolei , George Erin , Shan Xiaoyin , Gitto Sarah , D'Andrea Kurt , Wubbenhorst Bradley , Whicker Margaret , Hallberg Dorothy , Schwartz Lauren , Nathanson Katherine L , Mills Gordon , Velculescu Victor E , Wang Tian-Li , Brown Eric , Drapkin Ronny , Simpkins Fiona
Published: December 2023

Clear Cell Ovarian Cancer (CCOC) is one of the most challenging subtypes of ovarian cancer to treat as advanced stages are often chemo-resistant compared to the more common high-grade serous ovarian cancer (HGSOC). Limited treatment options are available for this devastating disease. Understanding the resistance mechanism and identifying effective therapeutic options for CCOC are clearly needed. Exploiting genetic alterations common in CCOC, such as ARID1A, is a rational strategy for combination therapies. ARID1A, an epigenetic tumor suppressor, is the most common genetic mutation in CCOC, with about 50% of CCOC demonstrating loss of ARID1A. ARID1A regulates the transcription of specific genes (e.g. c-myc) by altering the chromatin structure. We hypothesize that ARID1A loss increases CCOC cell dependency on chromatin remodeling and DNA repair pathways for survival. Through a drug screen, BET inhibitors and DNA damage response inhibitors were the most active monotherapies in ARID1A mutant CCOC. However, monotherapy efficacy needs to be further improved. We demonstrate in our study that the combination of BRD4 inhibitor (BRD4i) with DNA damage response (DDR) inhibitors (ATR or WEE1 inhibitors; e.g. BRD4i-ATRi) were synergistic at low doses leading to decreased survival, and colony formation in an ARID1A dependent manner.  Combination BRD4i-ATRi caused significant tumor regression and increased overall survival in ARID1AMUT not ARID1AWT patient-derived xenograft (PDX) models. Combination BRD4i-ATRi significantly increased gH2AX, and decreased RAD51 foci and BRCA1 expression suggesting loss in the ability to repair DNA double-strand-breaks by homologous recombination (HR), more so than with monotherapy in ARID1AMUT cells. These studies demonstrate BRD4i-ATRi is an effective treatment strategy that capitalizes on synthetic lethality with ARID1A loss in CCOC warranting further study.

Citation Format: Yasuto Kinose, Haineng Xu, Sushil Kumar, Hyoung Kim, Sergey Medvedev, Xiaolei Wang, Erin George, Xiaoyin Shan, Sarah Gitto, Kurt D'Andrea, Bradley Wubbenhorst, Margaret Whicker, Dorothy Hallberg, Lauren Schwartz, Katherine L Nathanson, Gordon Mills, Victor E Velculescu, Tian-Li Wang, Eric Brown, Ronny Drapkin, Fiona Simpkins. Dual blockade of BRD4 and the ATR/WEE1 pathway exploits ARID1A loss in clear cell ovarian cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B060.
Abstract B076: Discovery of BBT-176 as fourth generation EGFR tyrosine kinase inhibitor
Authors: Duggirala Krishna Babu , Lee Kwangho
Published: December 2023

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) improve overall survival rate in patients with EGFR mutated non-small cell lung cancers (NSCLC). However, upon treatment with third-generation EGFR TKIs (Osmertinib) develops C797S resistance[KS1] [u2]  in the 20 – 30% of the patients. To date, there is no approved drug for third-generation resistance NSCLC patients. There is an unmet medical need to develop fourth generation EGFR TKIs[KS3]  targeting C797S mutation. Extensive structure–activity relationship (SAR) studies led to the discovery of BBT-176 as a reversible fourth generation EGFR TKI.  BBT-176 shows promising results with highly potent and anti-cancer efficacy in Osmertinib-resistant in vitro and in vivo cell lines. In mouse models, BBT-176 exhibited potent inhibition of tumor growth, leading to tumor regression in certain cases. Furthermore, promising results were observed in the clinical trial, where two patients with EGFR Del[KS4] 19/T790M/C797S mutations in their blood demonstrated tumor shrinkage and improvements in radiological assessments.   BBT-176, a fourth-generation EGFR inhibitor, demonstrates significant preclinical activity against non-small cell lung cancer (NSCLC) that is resistant to current EGFR tyrosine kinase inhibitors (TKIs). This promising compound has shown early signs of clinical efficacy and safety in initial trials.

Citation Format: Krishna Babu Duggirala, Kwangho Lee. Discovery of BBT-176 as fourth generation EGFR tyrosine kinase inhibitor [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B076.
Abstract B081: Aurora kinase A inhibition overcomes adaptive resistance to KRAS G12C inhibitor by G1-checkpoint induced apoptosis in KRAS non-small cell lung cancer
Authors: Li Chendi , Chang Jeremy , Graser Christopher J , Syed Usman M , Nimbalkar Anahita , Shen Yi , Altschuler Steve , Wu Lani , Michor Franziska , Gong Xueqian , Hata Aaron N
Published: December 2023

Cancer cells gain drug-tolerant states and evade therapy. In response to KRAS G12C inhibitor (G12Ci), KRAS mutant non-small cell lung cancer (NSCLC) cells maintain a drug-tolerant state by aurora kinase A (AURKA). AURKA can phosphorylate c-Raf to maintain new KRAS signaling, however, how AURKA becomes activated to cause KRAS G12Ci resistance is unclear. We show here that KRAS G12C + AURKA inhibition cause synthetic lethality in KRAS G12C NSCLC cells. LY3499446 (KRAS G12Ci) and LSN3321213 (aurora kinase A inhibitor) induced apoptosis that is independent of inhibited MAPK reactivation. Using high-content imaging that tracks single-cell fate during treatment, we observed that single-agent KRAS G12Ci induces G1 arrest in a sub-population of cells. Upon co-treatment with G12C + AURKAi, cells that are halted in G1 phase undergo early apoptosis, while those that initially evade G1 arrest and proceed through G2/M undergo apoptosis in subsequent G1. These data suggest the hypothesis that AURKA inhibition may increase the probability of G1 checkpoint-induced apoptosis by facilitating chromosomal misalignment and genomic instability in G2. In summary, we provide pre-clinical rationale for clinical testing of KRAS G12C + AURKA inhibitors. We also suggest a novel mechanism explaining the dependency of KRAS G12C resistant subpopulations on AURKA, leading to the opportunity to investigate the role of genomic instability in conferring KRAS G12Ci adaptive resistance.

Citation Format: Chendi Li, Jeremy Chang, Christopher J Graser, Usman M Syed, Anahita Nimbalkar, Yi Shen, Steve Altschuler, Lani Wu, Franziska Michor, Xueqian Gong, Aaron N Hata. Aurora kinase A inhibition overcomes adaptive resistance to KRAS G12C inhibitor by G1-checkpoint induced apoptosis in KRAS non-small cell lung cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B081.
Abstract B083: Secondary KRAS mutations lead to acquired resistance to KRASG12D inhibitor in colorectal cancer
Authors: Lieb Simone , Jurado Sabine , Nowoshilow Sergej , Zak Krzysztof M. , Hofmann Marco H.
Published: December 2023

KRAS is the most frequently mutated oncogene, with high prevalence in tumor types with a significant unmet clinical need, such as colorectal cancer (CRC). Activating mutations in the KRAS gene leading to amino acid exchange at position 12 from Glycine to Asparagine, G12D, occur in approximately 12.5% of CRC. The approval of mutant specific KRASG12C inhibitors MRTX849 and AMG510 by the FDA raised the question if KRAS targeted therapy approaches could provide benefit to cancer patients with other mutant alleles.  Several KRASG12D inhibitors have recently been described and MRTX1133, a selective KRASG12D inhibitor from Mirati Therapeutics, has now entered clinical trial, recruiting patients with advanced solid tumors harboring a KRAS G12D mutation. Alongside the great hopes placed in KRAS targeting therapy, resistance is likely to occur, as it was observed in patients relapsing in response to KRASG12C inhibitors.  We aimed to use this pre-clinical study to predict and understand potential resistance mechanisms that may arise during targeted treatment against KRASG12D in the CRC setting. To this aim, the colorectal cancer cell line GP2d was continuously treated with MRTX1133 to generate resistance and individual outgrowing clones were subsequently profiled in a series of assays to identify potential mechanism(s). These assays included analysis of gene expression changes, mutation calling, as well as qPCR and protein-based assays. Interestingly, we identified two individual secondary on-target mutations in the KRAS gene, which could affect binding of MRTX1133 to the pocket in KRAS. These findings were confirmed in an orthogonal assay, by screening for resistance in Ba/F3 cells transduced with the corresponding KRAS double mutant. Combination assays were then performed in parental and resistant cells to study whether additional vertical or alternative pathway inhibition could address the resistance. Collectively our pre-clinical study identified secondary KRAS mutations in the compound binding pocket as a mechanism of resistance to KRAS G12D inhibition by MRTX1133 in colorectal cancer cells.

Citation Format: Simone Lieb, Sabine Jurado, Sergej Nowoshilow, Krzysztof M. Zak, Marco H. Hofmann. Secondary KRAS mutations lead to acquired resistance to KRASG12D inhibitor in colorectal cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B083.
Abstract B088: VT3989, the first-in-class and first-in-human TEAD auto-palmitoylation inhibitor, enhances the efficacy and durability of multiple targeted therapies of the MAPK and P13K/AKT/mTOR pathways
Authors: Tang Tracy T , Post Leonard
Published: December 2023

TEAD transcription factors are the major effectors of the Hippo-YAP/TAZ-TEAD pathway involved in the regulation of cell proliferation, survival, and cell migration. TEAD auto-palmitoylation has been shown to be required for TEAD interaction with coactivator YAP/TAZ and hence activation of transcriptional activity. There are multiple reports that YAP/TEAD activation provides the essential survival signal in drug-tolerant persister/dormant cells in cancer in response to treatment with targeted therapies, leading to drug resistance and cancer relapse in BRAF-mutant, KRAS-mutant, EGFR-mutant, and ALK-rearranged non–small cell lung cancers (NSCLC). The TEAD inhibitor VT3989, currently in clinical testing, was evaluated for effect on emerging drug tolerant persister/dominant cancer cells and drug resistance. VT3989, with similar biological activity in vitro and in vivo as the potent small molecule TEAD auto-palmitoylation inhibitors reported (Tang et al, 2021, Mol Cancer Ther. 20(6):986-998), disrupts YAP/TAZ-TEAD protein interaction, suppresses TEAD transcriptional activity, and selectively blocks proliferation of NF2-deficient mesothelioma in vitro and inhibit NF2 mutant xenograft tumor growth in vivo. In in vitro assays as well as in vivo studies using cell line derived (CDX) and patient derived (PDX) xenograft models, VT3989 enhanced the efficacy and durability of clinically approved EGFR inhibitors (osimertinib, lazertinib), MET inhibitors (savolitinib, capmatinib), EGFR-MET bispecific antibody (amivantinib), KRAS-G12C inhibitors (sotorasib, adagrasib), BRAF inhibitors (encorafenib, dabrafenib), MEK inhibitors (trametinib, cobimetinib), and mTORC inhibitor (everolimus).  With the addition of VT3989, combination anti-tumor effect was observed and/or tumor regrowth was delayed compared to monotherapy alone. Therefore, in addition to Hippo-YAP/TAZ-TEAD pathway driven cancers, such as mesothelioma, where VT3989 can show monotherapy efficacy, future VT3989 clinical studies will include combination therapy with approved targeted therapies to enhance the efficacy and durability of treatment.

Citation Format: Tracy T Tang, Leonard Post. VT3989, the first-in-class and first-in-human TEAD auto-palmitoylation inhibitor, enhances the efficacy and durability of multiple targeted therapies of the MAPK and P13K/AKT/mTOR pathways [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B088.
Abstract B098: Unbiased in vitro and in vivo drug anchor screens identify mechanisms of resistance and sensitization for MTA-cooperative PRMT5 inhibitors in MTAP-deleted cancer models
Authors: Lombardo Steven , Tonini Matthew R , Grove Lauren , Fenoglio Silvia , Tepper James , Shen Binzhang , Stowe Hannah , Liu Shangtao , Meier Samuel R , Choi Ashley H , Khendu Tenzing , Yu Yi , Cottrell Kevin M , Maxwell John P , Andersen Jannik N , Huang Alan , Briggs Kimberly J , Teng Teng
Published: December 2023

Homozygous deletion of the MTAP gene is one of the most common genetic alterations in cancer, affecting 10-15% of all human cancer.  Clinical MTA-cooperative PRMT5 inhibitors, including TNG908 and TNG462, were developed to leverage the well-characterized synthetic lethal interaction between PRMT5 inhibition and MTAP deletion. Indeed, the first disclosed clinical data for an MTA-cooperative PRMT5 inhibitor demonstrated the ability of TNG908 to selectively inhibit PRMT5 in MTAP-deleted tumors while sparing adjacent normal, MTAP-proficient cells in patients. To further explore the mechanism of action of MTA-cooperative PRMT5 inhibitors in MTAP-deleted cells, we employed multiple CRISPR-based editing platforms (CRISPRn, CRISPRi and CRISPRa) to conduct unbiased MTA-cooperative PRMT5 inhibitor anchor screens in vitro and in vivo.  A panel of cancer models representing multiple histologies revealed potential PRMT5 inhibitor sensitization and resistance pathways that also include mechanisms that are specific to MTA-cooperative inhibitors. Specifically, the results of these screens identify 1) the effect of MTA/SAM metabolism on MTA-cooperative PRMT5 inhibitors, and 2) identification of potential sub-stratification strategies for patients with MTAP-deleted cancer, and 3) the activity of PRMT5 in anti-apoptotic pathways and synergy between PRMT5 and BCLxL inhibition.

Citation Format: Steven Lombardo, Matthew R Tonini, Lauren Grove, Silvia Fenoglio, James Tepper, Binzhang Shen, Hannah Stowe, Shangtao Liu, Samuel R Meier, Ashley H Choi, Tenzing Khendu, Yi Yu, Kevin M Cottrell, John P Maxwell, Jannik N Andersen, Alan Huang, Kimberly J Briggs, Teng Teng. Unbiased in vitro and in vivo drug anchor screens identify mechanisms of resistance and sensitization for MTA-cooperative PRMT5 inhibitors in MTAP-deleted cancer models [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B098.
Abstract B102: Predicting small cell lung cancer liver metastasis
Authors: Chrisafis George , Takahashi Nobuyuki , Thomas Anish
Published: December 2023

Small cell lung cancer (SCLC) is a highly aggressive, treatment-resistant form of lung cancer which carries an exceptionally poor prognosis. Patients with SCLC often present at an advanced stage due the propensity of SCLC for early metastasis, with common sites including the brain, liver, adrenal glands, and bone. Liver metastasis has been identified as an independent prognostic factor and is correlated with an unfavorable response to chemotherapy, but specific steps in the liver metastatic cascade have yet to be described. Here, we seek to better characterize SCLC liver metastases by analyzing gene expression data from cell lines and patient biopsy samples. We start by analyzing data published in MetMap (Jin et al., 2020), which calculated organ-specific ‘metastatic potential’ values for cell lines in the Cancer Cell Line Encyclopedia (CCLE). Gene set enrichment analysis of 20 high-grade neuroendocrine differentiated cancer cell lines (8 SCLC, 12 large cell carcinoma) revealed that lines with high liver metastatic potential exhibit significant positive enrichment of genes involved in MYC signaling, oxidative phosphorylation, and xenobiotic metabolism; genes involved in the mitotic spindle and E2F targets are negatively enriched. This hallmark signature appears unique to these cells compared with other non-small cell lung cancer and colorectal cancer. We identify ~45 genes from these hallmarks which are differentially expressed in cell lines with high vs. low liver metastatic potential and create a multiple linear regression model to predict liver metastatic potential based on the expression of these genes. When applied to clinical SCLC samples, the model predicts samples obtained from the liver to have a significantly higher metastatic potential than samples from all other biopsy sites. A higher estimated metastatic potential also correlates with poorer overall survival. Genes and pathways identified by this analysis, following experimental validation, could serve as potential therapeutic targets to disrupt the liver metastatic cascade.

Citation Format: George Chrisafis, Nobuyuki Takahashi, Anish Thomas. Predicting small cell lung cancer liver metastasis [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B102.
Abstract B110: β3-adrenergic receptor as a new molecular target in neuroblastoma treatment
Authors: Putino Rossana , Boaretto Alessia , Bruno Gennaro , Mattei Gianluca , Tondo Annalisa , Favre Claudio , Calvani Maura
Published: December 2023

Purpose of study:  β3-Adrenergic receptor correlation with tumor growth and progression in preclinical and clinical studies in neuroblastoma.

Experimental procedures: Knock-out of β3-Adrenergic Receptor (β3-AR) in Neuroblastoma (NB) cell lines using CRISPR-Cas9 technology. In vivo experiment using β3-AR knock-out cells in order to evaluate the tumor growth in mice. RNA Sequencing of tumor masses derived from mice to investigate β3-AR Dependent tumor signaling pathways.  Flow Cytometry analysis to observe the expression level of β3- Adrenergic Receptor on circulating tumor cells (CTCs) in peripheral blood and on disseminated tumor cells (DTCs) in bone marrow from NB patients with low-risk and high-risk prognosis.

Summary of data:  CRISPR/Cas9 technology was performed to obtain ADRB3-/- clones from human and murine NB cell lines. ADRB3-/- knocked-out cells were inoculated in murine models NU-Foxn1nu and A/J mice, showing an absence of tumor growth compared to wild type NB cells. Subsequently, RNA sequencing on tumor masses was performed to investigate β3-AR-dependent tumor signaling pathways. RNA-sequencing analysis showed many pathways were altered. Notably, downregulation of genes involved in epithelial–mesenchymal transition (EMT) pathway and genes involved in cell adhesion were underlined in Knock-out vs control, correlated with the absence of tumor growth. Cytofluorimetric analysis of β3-AR level expression were conducted on circulating tumor cells (CTCs) in peripheral blood and on disseminated tumor cells (DTCs) in bone marrow from NB patients with low-risk and high-risk prognosis to evaluating the expression levels of β3-AR. As expected, most of the high-risk NB patients showed high percentage of β3-AR+ cells among CTCs and DTCs, while the majority of low-risk patients showed lower percentage.

Statement of the conclusions: Ours data showed that β3-AR could be a potential target therapy for neuroblastoma treatment. In fact, his deletion results in an absence of tumor growth and progression as demonstrate in mice in vivo experiments. In addition, NB patients with lower β3-AR level, showed a better prognosis . Furter studies should be focused on the pathways related to β3-AR. Moreover, new drugs will be tested in order to normalize the level of β3-AR in NB patients. 

Citation Format: Rossana Putino, Alessia Boaretto, Gennaro Bruno, Gianluca Mattei, Annalisa Tondo, Claudio Favre, Maura Calvani. β3-adrenergic receptor as a new molecular target in neuroblastoma treatment [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B110.