bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 108: Insulin-like growth factor binding protein 4 exerts differential effects on breast cancer cell proliferation based on the expression status of pregnancy-associated plasma protein A
Authors: Flynn Keenan L. , Houston Kevin D.
Published: June 2022

Insulin-Like Growth Factor Binding Protein 4 (IGFBP-4) is one of six binding proteins involved in sequestering insulin-like growth factors 1 and 2 (IGFs). IGFBP-4 expression is induced by estrogen receptor α (ERα), and its classical function is to prevent activation of IGF-1 receptor (IGF-1R) by sequestration of IGF-1. However, the aggressive triple-negative breast cancer cell line MDA-MB-231 shows high expression of IGFBP-4 along with its protease, pregnancy-associated plasma protein A (PAPP-A). Further, inhibition of IGFBP-4 in MDA-MB-231 cells is sufficient to reduce migration. To better understand the function of IGFBP-4 as a pro-cancer or anti-cancer protein, its anti-proliferative effects in ERα positive cells must be reconciled with its pro-migratory effects in triple negative cells. The role of IGFBP-4 in proliferation across four breast cancer cell lines representing ERα positive and triple negative subtypes was investigated. Stable transfection with IGFBP-4 expression construct in the MCF-7 and T-47D cell lines, which have silenced the PAPP-A gene, shows a reduction in proliferation. Further, MCF-7 parental cells cultured in conditioned media containing IGFBP-4 also show reduced proliferation, suggesting that the anti-proliferative effect of IGFBP-4 is associated with the absence of PAPP-A. Stable IGFBP-4 transfection of triple-negative MDA-MB-468 cell line, which has low extracellular IGFBP-4 when compared to MDA-MB-231 but does express PAPP-A, was sufficient to increase proliferation. Taken together, these results support the conclusion that proliferative effects of IGFBP-4 are dependent upon the expression status of PAPP-A.

Citation Format: Keenan L. Flynn, Kevin D. Houston. Insulin-like growth factor binding protein 4 exerts differential effects on breast cancer cell proliferation based on the expression status of pregnancy-associated plasma protein A [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 108.
Abstract 1084: FlexBodies: Targeting intracellular tumor antigens with T cell redirecting bispecific antibodies
Authors: Haber Lauric , McKay ryan , Finney jennifer , castaneda stephen , spitler kristen , Rizvi sahar , Hwang aidan , Bray kevin , Ramos willy , Krueger pamela , delfino frank , Meagher Craig , huang tammy , shen yang , olson william , Lin john , smith eric
Published: June 2022

Bispecific antibodies engineered to recruit T cells as a means to kill tumor cells are a promising new class of therapeutics in the field of oncology. A challenge for this class of agents is the requirement for cell surface expressed tumor specific targets, as a large number of tumor-specific antigens are only expressed intracellularly. One strategy to access these targets is to take advantage of the natural processing and presentation of intracellular antigens in the context of the major histocompatibility complex (MHC). However, this requires the development of therapeutics that can be active against targets with a very low surface density. Here, we describe the generation and characterization of Flexbodies, a novel format bispecific antibody engineered to target MHC-peptide tumor neoantigens. We demonstrate that the orientation of the targeting arms and the valency of these reagents allow specific T-cell activation and potent cytotoxic activity against cell lines expressing endogenous level of peptide antigens. This bispecific format has the potential to become an important new tool to expand the range of tumors able to be specifically targeted with t-cell redirecting biologics.

Citation Format: Lauric Haber, ryan McKay, jennifer Finney, stephen castaneda, kristen spitler, sahar Rizvi, aidan Hwang, kevin Bray, willy Ramos, pamela Krueger, frank delfino, Craig Meagher, tammy huang, yang shen, william olson, john Lin, eric smith. FlexBodies: Targeting intracellular tumor antigens with T cell redirecting bispecific antibodies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1084.
Abstract 1086: PEGylated talazoparib enhances therapeutic window of its combination with temozolomide in Ewing sarcoma
Authors: Del Pozo Vanessa , Robles Andrew J. , Fontaine Shaun , Liu Qianqian , Michalek Joel E. , Houghton Peter J. , Kurmasheva Raushan
Published: June 2022

Introduction: The Ewing family of sarcomas is the fourth most common highly malignant childhood cancer. Current standard therapy is relatively ineffective for relapsed and metastatic Ewing sarcoma predominantly due to development of resistance to chemotherapeutics. Nanoparticle-formulated drugs are reported to increase uptake into tumor tissue, while reducing drug access to normal tissues, because of reduced permeability of normal vasculature. This potentially leads to lower dose exposure in normal tissues and reduced toxicity. The purpose of this study was to evaluate the antitumor activity of the PEGylated poly(ADP) ribose polymerase 1/2 (PARP1/2) inhibitor talazoparib combined with the DNA alkylating agent temozolomide (TMZ) in Ewing sarcoma xenograft models and other pediatric cancers.

Method:. We evaluated the preclinical efficacy of PEGylated talazoparib (PEG~TLZ), alone or in combination with TMZ, in Ewing sarcoma and glioblastoma patient derived xenograft (PDX) models. Additional solid tumor models, such as rhabdomyosarcoma, Wilms tumor, malignant rhabdoid tumor, osteosarcoma, and synovial sarcoma were evaluated using a novel single mouse testing approach. PEG~TLZ was administered as a single administration intraperitoneally to mice bearing subcutaneous tumors at a single dose of 5 - 20 µmol/kg alone, or at 10 µmol/kg on day 1 combined with TMZ administered orally at 40 mg/kg starting on day 3 or 4 for 5 consecutive days.

Results: Mice bearing Ewing sarcoma and MGMT-deficient glioblastoma xenografts tolerated PEG~TLZ+TMZ with minimal toxicity and achieved maintained complete response (MCR). The time-separated drug administration schedule of the single dose of PEG~TLZ followed by the 5-day TMZ treatment demonstrated objective responses in Ewing sarcoma, rhabdoid tumor, rhabdomyosarcoma, and glioblastoma PDX models.

Conclusion: We evaluated the range of PEG~TLZ+TMZ activity among pediatric solid tumor panels using conventional and single-mouse testing approaches, and demonstrated that PEG~TLZ combined with delayed TMZ administration enhances the therapeutic window of the treatment compared to free TLZ. From the clinical standpoint, the single intravenous administration of PEG~TLZ could be advantageous for treating infants and young children.

Citation Format: Vanessa Del Pozo, Andrew J. Robles, Shaun Fontaine, Qianqian Liu, Joel E. Michalek, Peter J. Houghton, Raushan Kurmasheva. PEGylated talazoparib enhances therapeutic window of its combination with temozolomide in Ewing sarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1086.
Abstract 111: Head and neck cancer cells can differentiate and resemble their tissue of origin
Authors: Oppel Felix , Shao Senyao , Gendreizig Sarah , Kühnel Philipp , Przybycin Vivien , Hain Carsten , Schmidt Pascal , Schürmann Matthias , Goon Peter , Niehaus Karsten , Kalinowski Jörn , Sudhoff Holger
Published: June 2022

Head and neck squamous cell carcinoma (HNSCC) is a highly malignant disease with a mortality rate of about 50%. It represents the sixth most common type of malignant tumor in the world. Risk factors include tobacco and alcohol usage and infection with the human papilloma virus (HPV). HPV-negative HNSCCs frequently display areas of cornification evident by keratin pearls in the tumor tissue. Cornification represents a natural differentiation path of keratinocytes in the normal epidermis and oral mucosa. To investigate the mechanisms of HNSCC cell differentiation we have established spheroid cell cultures from patient-derived HNSCC and parotid gland adenoid cystic carcinoma (ACC) tissue that grow in suspension under serum-free conditions. The use of a specific differentiation medium induced striking adhesion, loss of proliferation, and differentiation in tumor cells. Spheroid cells grew as single cell clones under serum-free conditions with a cloning efficiency of 40-60%, which was fully diminished under differentiating conditions. HNSCC cells cornified as indicated by the formation of lamellar bodies in the cytoplasm of adherent cells and an upregulation of cornification markers SPRR3 and involucrin. ACC cells upregulated parotid gland differentiation markers including α-amylase. RNA-seq analysis in HNSCC cells confirmed an upregulation of signaling pathways associated with cornification and epithelial cell differentiation. Conversely, pathways regulating the three-dimensional organization of the genome were downregulated upon differentiation. This was accompanied by the formation of ATRX-positive heterochromatin foci in the nucleus of differentiated ACC and HNSCC cells resembling those previously described to arise during therapy-induced senescence. Moreover, gas chromatography mass spectrometry analysis revealed a lack of essential amino acids including leucine to be implicated in the differentiation process. Altogether, our spheroid model of HNSCC and ACC cells is suitable to analyze the mechanisms underlying tumor cell differentiation and might lead to new therapeutic approaches that can drive long-term repopulating HNSCC and ACC cells into differentiation.

Citation Format: Felix Oppel, Senyao Shao, Sarah Gendreizig, Philipp Kühnel, Vivien Przybycin, Carsten Hain, Pascal Schmidt, Matthias Schürmann, Peter Goon, Karsten Niehaus, Jörn Kalinowski, Holger Sudhoff. Head and neck cancer cells can differentiate and resemble their tissue of origin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 111.
Abstract 1112: Detection ofHMGB1 and extracellular ATP for the assessment of immunogenic cell death
Authors: Somberg Richard L. , Kupcho Kevin , Niles Andrew , Cali James
Published: June 2022

Immunogenic cell death (ICD) is a form of apoptosis that kills susceptible populations of cancer cells while teaching the immune system to attack the remaining resistant cells. Chemotherapeutics which induce ICD elicit their immune response by inducing tumor cells to display or release damage-associated molecular patterns (DAMPs). There are two key biomarkers for ICD:  1) during the apoptotic process, tumor cells secrete ATP (extracellular ATP or eATP) and 2) during secondary necrosis, tumor cells release HMGB1 (high mobility group box 1). Because the dying tumor cells display these DAMP molecules, they stimulate the recruitment of dendritic cells (DCs) into the tumor bed and ultimately “teach” cytotoxic T-lymphocytes (CTLs) to respond to these tumor specific antigens. These primed CTLs will then kill additional tumor cells through a direct cytotoxic response. Therefore, therapeutics which provoke an ICD response offer a therapeutically desirable outcome for cancer therapy. Current assay methods such as ELISA assays and flow cytometry for measuring these ICD biomarkers are laborious and involve multiple transfer and wash steps. We have developed homogeneous (no wash), single addition assays for measuring the two principle ICD biomarkers. The eATP assay uses optimized luciferase detection chemistry that is applied directly to live cells to measure ATP release over 24 hours. The HMGB1 assay uses a complementary luciferase fragment-labeled antibody approach to measure protein concentration. We demonstrated the use of these two assays with model systems for inducing Immunogenic Cell Death with human and murine cell lines including U2OS, EL4, and U937 cells. The potency and response magnitude of ICD-inducing compounds including doxorubicin, idarubicin and mitoxantrone differs depending on the model cell line used. Further, the timing of extracellular ATP release and HMGB1 release differs between cell lines and inducers. Improved assays to measure two markers of Immunogenic Cell Death will expedite the discovery and development of inducers of Immunogenic Cell Death, including small molecule therapeutics, oncolytic viruses, radiation therapy and other novel therapeutics, by streamlining the workflow, increasing sample throughput and providing real time live cell data.

Citation Format: Richard L. Somberg, Kevin Kupcho, Andrew Niles, James Cali. Detection ofHMGB1 and extracellular ATP for the assessment of immunogenic cell death [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1112.
Abstract 114: Progesterone via integrin αvβ3 induces cell proliferation in PR-negative breast cancer cells
Authors: Lin Hung-Yun , Huang Tung-Yung , Chang Tung-Cheng , Chen Han-Yu , Huang Haw-Ming , Lee Sheng-Yang , Li Zi-Lin , Chu Hung-Ru , Whang-Peng Jacqueline , Wang Kuan
Published: June 2022

Background: Progesterone (P4) stimulates reproductive functions such as ovulation and the initiation and maintenance of pregnancy. P4 regulates biological activities via its nuclear receptor. However, it is not fully understood mechanisms involved in P4 functions in PR-negative cells. Anchor protein integrin αvβ3 plays a vital role in cancer growth and metastasis.

Methods: We investigated the effect of P4 in PR-positive breast cancer MCF-7 cells and PR-negative breast cancer MDA-MB-231 cells. The protein and RNA expression changes induced by P4 were both analyzed in two cell lines. At the same time, the anti-integrin αvβ3 antibody and RGD peptide were employed for investigating the role of integrin αvβ3 in P4 regulation.

Results: P4 inhibited the proliferation in MCF-7 cells but stimulated cell growth in MDA-MB-231 cells. In addition to ERK1/2 activation, P4 stimulated the phosphorylation at Y397 and Y925 in focal adhesion kinase (FAK), a vital kinase in integrin ανβ3 signal pathway that modulated by co-treatment with anti-integrin αvβ3 antibody or RGD peptide. P4 activated the gene expression of integrin ανβ3, CCND1, PCNA, and MMP-9 in MDA-MB-231 cells. P4 also induced PD-L1 and CCND1 accumulation.

Conclusions: Progesterone via integrin αvβ3-mediated signal transduction regulates gene expression, cell proliferation, and migration in TNBC cells. It is the first time to demonstrate that P4 binds with the cell surface anchor protein, integrin αvβ3, to stimulate cancer cell proliferation in PR-negative breast cancer cells.

Keywords: Progesterone, Progesterone receptor, Integrin αvβ3, Breast cancer, Cell proliferation

Citation Format: Hung-Yun Lin, Tung-Yung Huang, Tung-Cheng Chang, Han-Yu Chen, Haw-Ming Huang, Sheng-Yang Lee, Zi-Lin Li, Hung-Ru Chu, Jacqueline Whang-Peng, Kuan Wang. Progesterone via integrin αvβ3 induces cell proliferation in PR-negative breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 114.
Abstract 1145: Therapeutic drug monitoring of Sunitinib in real-world patients
Authors: Ferrer Laurent , Chauvin Jonathan , Deville Dr Jean-Laurent , Ciccolini Joseph
Published: June 2022

Therapeutic Drug Monitoring (TDM) with model-based adaptive dosing is expected to quickly ensure that exposure levels of anticancer agents are in the adequate range of plasma concentrations. Here, we have compared such TDM + in silico approach with empirical changes in dosing performed upon clinical signs in patients with metastatic renal cell carcinoma treated with Sunitinib. A total of 31 patients were monitored in our institute. All patients were treated with a standard starting dose of 50 mg QD following the 4/2 schedule. Sunitinib and n-desethyl sunitinib trough levels were assayed at steady state by a fully validated LC-MS/MS analysis. A PK-pop model with Bayesian estimate implemented on Monolix helped to identify individual PK parameters from a single time-point. Two different target exposures were considered (i.e., trough levels comprised between 50 and 100 ng/ml or 0-24h AUC comprised between 1250 and 2150 ng/ml.h). The PK model enabled to simulate both trough levels and 0-24h AUC from a single sample, regardless of the sampling time. Overall, dosing was modified empirically in about 54% of the patients after treatment started, mostly (46%) for reducing the dosing after that toxicities showed. Clinical benefit (i.e., stable disease + partial response + complete reponse) was achieved in 54% of patients but treatment discontinuation was observed in 58% of the patients eventually, mostly because of severe side-effects. A relationship between baseline exposure levels and early onset treatment-related toxicities was found. Because of the various changes in dosing, no such relationship was found between baseline exposure levels and efficacy evaluated at 3 months. TDM showed that only 45% of the patients were in the right window for trough levels (i.e., 50-100 ng/ml) and only 26% when considering AUC as the target exposure (i.e., 1200-2150 ng/ml.h). Consequently, the PK/PD model would have suggested to rapidly customize dosing in a much larger part of the patients (i.e., 84% with respect to the target AUCs). Importantly, Sunitinib dosing was empirically reduced only in 41% patients who displayed early-onset severe toxicities, whereas modelling would have immediately proposed to cut the dosing in more than 80% of those patients, thus suggesting that the safety of Sunitinib could have been improved when using our model. Although performed on a limited number of patients, this real-world study supports the hypothesis that TDM associated with PK modelling could help the prescriber to identify the best Sunitinib dosing in a quicker way than empirical change in dosing.

Citation Format: Laurent Ferrer, Jonathan Chauvin, Dr Jean-Laurent Deville, Joseph Ciccolini. Therapeutic drug monitoring of Sunitinib in real-world patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1145.
Abstract 1150: Pharmacokinetics and pharmacogenetics of liposomal cytarabine in AML patients treated with CPX-351
Authors: Donnette Melanie , Hamimed Mourad , Ciccolini Joseph , Costello Regis , Delassus Laure , Venton Geoffroy , Fanciullino Raphaelle
Published: June 2022

CPX-351 is a liposomal form encapsulating cytarabine and daunorubicin for treating Acute Myeloid Leukemia (AML) patients. Cytidine Deaminase (CDA) catabolizes free cytarabine in the liver, but to what extent it could affect as well the pharmacokinetics of liposomal cytarabine is yet to be investigated. Here we have studied the pharmacokinetics (PK) of released, liposomal and total cytarabine using a population-modeling approach in adult AML patients treated with CPX-351. Impact of CDA status (i.e., Poor Metabolizer (PM) vs. Extensive Metabolizer (EM)) on cytarabine exposure levels (AUC, trough levels, Cmax) and PK parameters were analyzed. All patients showed febrile neutropenia and one toxic-death was observed. Overall response rate was 75%. The ratio free:total cytarabine monitored in our patients was higher than expected (i.e., 47%). Inter-individual variability on pharmacokinetics parameters and subsequent exposure levels was >60%. A trend towards severe toxicities was observed in patients with higher exposure of cytarabine. Results showed that liposomal CPX-351 led to sustained exposure with reduced clearance (Cl = 0.16 L/h) and prolonged half-life (T1/2 = 28 h). Of note, liposomes were observed transiently in bone marrow on D15, with cytarabine levels 2.3-time higher than in plasma. Sequencing CDA gene was not contributive and CDA status was primarily evaluated upon phenotyping patients. PM status was found in 77% of the patients with a marked impact on cytarabine PK parameters, i.e., PM patients had higher exposure than EM patients (AUC: 5536 vs. 1784 ng/mL.h), prolonged half-life (T1/2: 33.9 vs. 13.7 h), and reduced clearance (Cl: 0.12 vs. 0.29 L/h). This pilot study suggests that despite being encapsulated in a liposomal vehicle as CPX-351, cytarabine fate in the body is highly dependent upon CDA activity, suggesting that liver metabolism is only partly skipped by the nanoparticle. Here, CDA status proved to have a major impact on cytarabine PK and possibly safety in AML patients treated with CPX-351. Indeed PM patients displayed higher exposure levels with higher risk for severe non-hematological toxicities. This study suggests that CDA status could be used as a covariate to customize CPX-351 dosing in AML patients.

Citation Format: Melanie Donnette, Mourad Hamimed, Joseph Ciccolini, Regis Costello, Laure Delassus, Geoffroy Venton, Raphaelle Fanciullino. Pharmacokinetics and pharmacogenetics of liposomal cytarabine in AML patients treated with CPX-351 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1150.
Abstract 1159: Predicting clinical drug responses using a few-shot learning-based interpretable AI
Authors: Park Sungjoon , Singhal Akshat , Silva Erica , Kreisberg Jason F. , Ideker Trey
Published: June 2022

High-throughput screens have generated large amounts of data characterizing how thousands of cell lines respond to hundreds of anti-cancer therapies. However, predictive drug response models trained using data from cell lines often fail to translate to clinical applications. Here, we focus on two key issues to improve clinical performance: 1) Transferability: the ability of predictive models to quickly adapt to clinical contexts even with a limited number of samples from patients, and 2) Interpretability: the ability to explain how drug-response predictions are being made given an individual patient’s genotype. Notably, an interpretable AI model can also help to identify biomarkers of treatment response in individual patients. By leveraging new developments in meta-learning and interpretable AI, we have developed an interpretable drug response prediction model that is trained on large amounts of data from experiments using cell lines and then transferred to clinical applications. We assessed our model’s clinical utility using AACR Project GENIE data, which contains mutational profiles from tumors and the patient’s therapeutic responses. We have demonstrated the feasibility of applying our AI-driven predictive model to clinical settings and shown how this model can support clinical decision making for tumor boards.

Citation Format: Sungjoon Park, Akshat Singhal, Erica Silva, Jason F. Kreisberg, Trey Ideker. Predicting clinical drug responses using a few-shot learning-based interpretable AI [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1159.
Abstract 1171: Genotoxic preventive potential of ethanol leaves extract of Annona muricata on N-Nitroso-N-ethylurea pro-leukaemia carcinogen in mice by bone marrow micronucleus assay
Authors: Akanni Olufemi Emmanuel , Bamisaye Eunice Oluwaseyi , Fashina Sunday Anthony , Abdulraheem Fatai Adesina , Fadiora Solomon Olufemi
Published: June 2022

Background: Genotoxicity which is an important indicator of DNA damage, repair and cell rejuvenation has been a predisposing factor to cancer development evidenced by bone marrow suppression in leukemia and studies have proven the effect of several agents inclusive of natural products to induce, prevent and treat genotoxicity through experimental models and clinical trials.

Aim:The genotoxic preventive potential of Annona muricata ethanol leaves extract on N-nitroso-N-ethylurea (NEU) pro-leukemia carcinogen in mice model using micronuclei formation in bone marrow was assessed.

Materials and Methods: 48 mice weighing 18-24g were randomly divided into six groups of eight mice each comprising of the; mice intravenously administered 20mg/kg of NEU 48 hourly 3 times (A); 80mg/kg of NEU 48 hourly 3 times (B); negative control fed with feed and water only (C); 0.2ml (0.1g/ml) ethanolic leaves extract of Annona muricata for 3 weeks prior to administration of low dose of NEU (D); 0.2ml (0.1g/ml) ethanolic extract of Annona muricata for 3 weeks prior to administration of high dose of NEU (E); and Annona muricata (ethanolic leaves extract) and commercial diet for adverse/toxicity group (F). After six weeks of last exposure, animals were ethically sacrificed, bone marrow harvested, smeared, stained using MayGrumwald and number of micronucleus polychromatic erythrocyte (MNPCEs) were determined microscopically.Result: Groups exposed to various dosage of the NEU yielded significantly increased MNPCEs with group B producing higher MNPCEs than A. The groups treated with extract displayed a significant reduction in the MNPCEs despite the prior exposure concentrations of NEU. The adverse group displayed no difference in MNPCEs when compared with the negative control (Table 1).

Conclusion: The extract displayed profound capacity to prevent genotoxicity thereby alleviating leukemia condition and it is well tolerated.

Week Number Groups (Mean±Standard deviation of MNPCE) A B C D E F 2 14.13±0.56 50.45±2.31 4.52±0.34 6.15±0.49 17.11±1.11 4.53±0.45 3 15.01±0.18 53.77±1.95 4.61±0.67 7.85±0.98 20.34±2.34 4.62±0.23 4 16.11±0.22 56.95±2.01 4.91±0.88 8.01±1.01 25.11±1.67 5.63±0.45 5 19.64±1.68 68.23±1.56 4.82±0.75 10.07±0.67 30.54±2.34 5.92±0.67

Citation Format: Olufemi Emmanuel Akanni, Eunice Oluwaseyi Bamisaye, Sunday Anthony Fashina, Fatai Adesina Abdulraheem, Solomon Olufemi Fadiora. Genotoxic preventive potential of ethanol leaves extract of Annona muricata on N-Nitroso-N-ethylurea pro-leukaemia carcinogen in mice by bone marrow micronucleus assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1171.
Abstract 118: BACH1 is an oxygen-sensitive mediator of the hypoxia response in triple-negative breast cancer
Authors: Dann Christopher M. , Nguyen Long C. , Yang Dongbo , Wynne Joseph P. , Valdespino Andrea , Stock Letícia , Rosner Marsha R.
Published: June 2022

Triple-negative breast cancer (TNBC) is an aggressive solid tumor characterized by a hypoxic phenotype that promotes metastatic progression and resistance to therapy. The canonical cellular response to hypoxia is largely driven by hypoxia inducible factors (HIFs) that stimulate transcription of genes related to angiogenesis, cancer stem cells, cell survival, and glucose and iron metabolism. There is growing evidence that several factors, in addition to HIFs, contribute to the tumor hypoxia response. Thus, identifying novel mechanisms that regulate this cellular response in addition to the HIFs is key to our understanding of hypoxia and could have direct clinical impact. High expression of BTB and CNC homology 1 (BACH1), a heme-regulated bZIP transcription factor, drives metastatic progression in various cancers including TNBC. While BACH1 has been shown to be induced under hypoxia in some cell lines, its regulation and role in hypoxia signaling, specifically with regards to cancer cells, is poorly understood. Here we report a novel regulatory pathway of the tumor hypoxia response in TNBC. We show that BACH1 is post-translationally prolyl-hydroxylated by the HIF prolyl-hydroxylases (PHDs) in an oxygen-dependent and NRF2/HMOX1-independent manner. Hypoxic exposure thus stabilizes BACH1 protein and enhances its DNA binding activity at loci of both canonical and hypoxia-specific downstream targets. We further show that BACH1 regulates the hypoxia response by promoting expression of both HIF-dependent and HIF-independent hypoxia-induced genes in TNBC cells. Our findings describe BACH1 as a novel oxygen-sensitive effector of the hypoxia response in TNBC that represents an alternative target for attenuating hypoxia-induced, pro-metastatic signaling and therapeutic resistance.

Citation Format: Christopher M. Dann, Long C. Nguyen, Dongbo Yang, Joseph P. Wynne, Andrea Valdespino, Letícia Stock, Marsha R. Rosner. BACH1 is an oxygen-sensitive mediator of the hypoxia response in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 118.
Abstract 1198: Discovering novel bispecific antibody targets through the mining of large-scale bulk and single cell RNA-seq databases
Authors: Chekalin Eugene , Paithankar Shreya , Chen Bin
Published: June 2022

Bispecific antibodies (bsAbs) that bind two distinct surface proteins in cancer cells are emerging as an appealing therapeutic strategy in cancer immunotherapy. Among thousands of surface proteins, experimentally identifying the best target pairs that are expressed only in cancer cells, but not in normal cells, is costly and time-consuming. The open bulk RNASeq and single-cell (sc)RNASeq offers a great resource to identify novel bispecific targets. Bulk RNASeq has been widely explored to identify therapeutic targets and biomarkers, resulting in voluminous data for various cancers and normal tissues, but the mixture of cell types in bulk RNASeq could not characterize the precise expression of targets in cancer cells, resulting in considerable false positives. scRNASeq provides a high resolution of target expression in individual cells, however, the challenges in solving the dropout issue and cell type classification hinder the direct use of scRNASeq in bsAbs target identification. Utilizing the OCTAD database consisting of over 20,000 bulk RNASeq samples, we proposed an approach that identifies target pairs that separate tumors from healthy the most, taking into account cluster heterogeneity, the distance between tumors and healthy as well as the angle between potential markers. Among the top pairs in Hepatocellular Carcinoma (HCC), CD33~PLVAP, for example, was a false positive because CD33 is myeloid and lymphoid cell lineage-specific. We thus assembled a scRNAseq database of healthy vital organs containing 39361 cells to aid selection. By comparing their expression with the expression of 18000 malignant cells predicted from 72000 cells of eight HCC samples, we identified target pairs that mostly express on the surface of the malignant HCC cells and had low or zero expression in vital organs. The most promising marker pair was GPC3~MUC13, presenting on the surface of over 30% of malignant HCC cells, with very low expression in vital organs. We further developed an R package to navigate the bsAbs target selection from open bulk RNASeq and scRNASeq.

Citation Format: Eugene Chekalin, Shreya Paithankar, Bin Chen. Discovering novel bispecific antibody targets through the mining of large-scale bulk and single cell RNA-seq databases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1198.
Abstract 1201: GenomeCruzer: A tool for interactive 3D visualization and analysis of multidimensional cancer omics
Authors: Isella Claudio , Corsi Riccardo , Elhasnaoui Jamal , Bertotti Andrea , Vezzadini Luca , Medico Enzo
Published: June 2022

In cancer genomics, integrative analysis of data obtained from biological samples of patient cohorts requires handling large groups of patients, each with molecular data of different type for thousands of genes, such as expression, mutation, copy number, and others.

Currently available tools allow visualization and analysis of such complex data, however forcing the user to work with partial views due to the amount of information. Also, these systems usually involve somewhat laborious procedures, so that for a complete analysis of the information it is often necessary to use many tools in sequence.

Here we present GenomeCruzer, a software for 3D interactive visualization and analysis of multidimensional omic data sets (e.g. gene expression, methylation, copy number alteration). GenomeCruzer is particularly useful to quickly achieve clinical and biological insight, providing easy integration and interactive visualization of genomic data of any kind. GenomeCruzer has the dual objectives of representing large amounts of genomic data in an easily readable way, allowing interactive analysis and providing visual results in real-time. Powerful graphics also allow simultaneous display of different types of values for the same objects, through multiple dimensions and customizable visual metaphors. The user can navigate this 3D environment and interact with the different elements, which constitute a representation of the starting datasets, building analysis paths and using statistical tools e.g. for the comparison of groups to highlight correlations.

In conclusion, GenomeCruzer enables users without bioinformatics background to browse, analyse and interpret complex omics datasets for multiple groups of patients and genes simultaneously, extracting clinical correlates and biological knowledge.

Citation Format: Claudio Isella, Riccardo Corsi, Jamal Elhasnaoui, Andrea Bertotti, Luca Vezzadini, Enzo Medico. GenomeCruzer: A tool for interactive 3D visualization and analysis of multidimensional cancer omics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1201.
Abstract 1219: Unraveling alternative polyadenylation in prostate cancer with CORE-PAD
Authors: Imada Eddie L. , Wilks Christopher , Langmead Ben , Marchionni Luigi
Published: June 2022

Mechanisms that control gene expression at the RNA level are often referred to as post-transcriptional regulation (PTR) mechanisms. Splicing and polyadenylation (PA) are well-known examples of PTR that can regulate not only gene expression but also their function. Alternative Polyadenylation (APA) has already been shown to be essential to many biological processes (e.g., proliferation, cell differentiation, etc) and has also been implicated in the development and progression of multiple diseases (e.g., cancer, hematological and immune disorders, etc.). Although several sequencing methods have been developed to sequence only the transcript termination site (TTS), the number of publicly available data derived from these methods is extremely limited in comparison to traditional RNA-Seq data. To overcome this limitation, we created a new framework - Compositional Regression of Polyadenylation Differences (CORE-PAD) - for the study of differential APA events using traditional bulk RNA-seq data. Through simulated data, we showed that CORE-PAD has higher accuracy than other methods (accuracy = 0.98) in detecting APA events. We applied CORE-PAD across prostate cancer (PCa) samples with impaired CDK12 and “wild” samples. We found multiple genes presented differential PA site usage, including DNA repair genes. Most notably, we notice that many genes that exhibit differential APA were not differentially expressed at gene level, meaning they are potentially “silent drivers” that cannot be capture through standard differential gene expression analysis. These findings highlight the importance of studying APA, which can help shed light into another layer of regulation occurring between transcription and translation. This is especially important since these events can be source of neoantigens or targeted mRNA degradation which could be explore for new treatments. Finally, the CORE-PAD framework was designed to take advantage of our recently published recount3 resource making over 750,000 RNA-Seq samples of human and mouse origin readily available for analysis, enabling studies of APA across thousands of phenotypes in an accurate and accessible way.

Citation Format: Eddie L. Imada, Christopher Wilks, Ben Langmead, Luigi Marchionni. Unraveling alternative polyadenylation in prostate cancer with CORE-PAD [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1219.
Abstract 1263: Comprehensive genomic profiling to identify biomarkers predictive of response to immunotherapy
Authors: Noel Pawan , Hall David W. , Udhane Sameer S. , Wang Min , Alyaqoub Fadel S. , Szelinger Szabolcs , Ozols Audrey A. , LoBello Janine R. , Baehner Frederick L. , Basu Gargi D.
Published: June 2022

Background: Immune-oncology (I-O) drugs show promise across multiple tumor types, and several biomarkers (BM) predict responsiveness to I-O therapy. In this descriptive study of clinical lab experience (2018-2021), we investigated the prevalence of genomic BMs associated with sensitivity and resistance to I-O drugs across solid tumors.

Methods: The GEM ExTra® assay was used to exome-sequence 2,359 paired samples at 180X and 400X coverage for germline and tumor DNA respectively. Reportable alterations were single base substitutions, indels, copy number alterations, fusions, alternate transcripts, as well as tumor mutational burden (TMB) and microsatellite instability (MSI) status. For I-O sensitive BMs we recorded MSI-High status, TMB-High (> 10 mut/Mb) status, and mutations in POLE/POLD1, MLH1, MSH2/3/6, PMS1/2, PD-L1/-L2, CTLA4, CD28, ARID2, and PBRM1. For I-O resistance BMs, IDO1 amplification and mutations in B2M, JAK1/2, IDO1, KRAS and STK11 were tallied. Results: I-O sensitive BMs varied from 0% to 58% (results in Table 1 for selected tumor types). Overall, TMB-High was in 6% of samples and MSI-High in 3%; all MSI-High were also TMB-High. In the 46% of renal cell carcinoma samples with I-O-sensitive BMs, PBRM1 mutation was common (94%). In 3 TMB-High endometrial samples, POLE/POLD1 was mutated, indicative of an ultra-mutated phenotype. Of note, 35% of colorectal cancer and 8% of melanoma samples with I-O sensitive mutations also had loss-of-function mutations in JAK1/2 and B2M, suggestive of I-O resistance. In non-small cell lung cancer, co-mutation of STK11 and KRAS, suggesting inferior response to I-O therapy, was present in 4% of samples with I-O BMs. IDO1 amplification was present in less than 1% of entire cohort.

Conclusions: Our study found I-O sensitive BMs in common and in rare aggressive cancers. The GEM ExTra assay can stratify cancer patients for likely responsiveness to immunotherapy.

Table 1. Immune-oncology sensitive biomarkers observed across select tumors Solid tumor type % of samples with signature All (n = 2,360) 13% Melanoma (n = 50) 54% Other skin cancers (n = 13) 62% Renal cell carcinoma (n = 200) 46% Urothelial (n = 86) 22% Endometrial (n = 88) 20% Gastro-intestinal (n = 360) 12% Sarcoma (n = 90) 11% Prostate (n = 151) 6% Lung (n = 100) 4% Breast (n = 297) 2% Central nervous system (n = 60) 0%

Citation Format: Pawan Noel, David W. Hall, Sameer S. Udhane, Min Wang, Fadel S. Alyaqoub, Szabolcs Szelinger, Audrey A. Ozols, Janine R. LoBello, Frederick L. Baehner, Gargi D. Basu. Comprehensive genomic profiling to identify biomarkers predictive of response to immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1263.
Abstract 1271: Detection of ALK, ROS1 and RET fusion transcripts in FFPE samples of non-small cell lung cancer patients using a novel RT-PCR based assay
Authors: Denis Marc G. , Vallée Audrey , Sagan Christine , Herbreteau Guillaume , Charpentier Sandrine , Rajamani Jaya , Lee Michael , Ordinario Ellen
Published: June 2022

Background and objective: Detection of genomic rearrangements like ALK fusions are mandatory in non-small cell lung cancer (NSCLC) as those alterations can be targeted by an increasing number of drugs. Fluorescence in-situ hybridization (FISH) is the gold standard but multiplexed technologies allow the analysis of several targets simultaneously. We have evaluated a novel RT-PCR based assay for the detection of ALK, ROS1 and RET rearrangement in NSCLC patients in this research study.

Materials and Methods: FFPE tissue sections from 309 patients with late stage NSCLC were screened for ALK and ROS1 status using immunohistochemistry (IHC) and confirmed with FISH. Total nucleic acids were extracted from tissue sections using the Maxwell RSC RNA FFPE kit (Promega). Fusions were detected by RT-PCR using a prototype ALK/RET/ROS1 Fusion Panel assay (Roche). A subset of discordant cases were further analyzed by RNA sequencing using the QIAseq Targeted RNAscan Lung Cancer Panel (Qiagen).

Results: All 309 samples were tested by RT-PCR with a 0% invalid rate. ALK fusions were detected in 39 samples: 29 were ALK FISH+, 5 were ALK FISH not contributive, 3 were ALK FISH- (less than 15% tumor cells presenting a gene fusion), and 2 were ALK IHC- and not tested by FISH. ROS1 fusions were detected in 12 samples: 9 were ROS1 FISH+, 1 was ROS1 FISH-, and 2 were ROS1 IHC- and not tested by FISH. RET fusions were detected in 3 samples. The remaining 255 samples were RT-PCR-: 5 were ALK FISH+ containing fusions not covered by the RT-PCR assay, 1 was ALK FISH+ containing ALK polysomy, 1 was ROS1 FISH+ containing a fusion not covered by the RT-PCR assay, 1 was ROS1 FISH+ containing ROS1 amplification, 1 was ROS1 FISH+ but ROS1 RNAseq-, and 246 samples were IHC- or IHC+/FISH-. The overall concordance rate between RT-PCR and FISH for ALK and ROS1 gene rearrangements was 91.9%. Although the RT-PCR assay did not detect 5 ALK and 1 ROS1 fusions due to design limitations, it did identify the presence of fusions in 9 tumors that were undetected by FISH (8 ALK and 1 ROS1).

Conclusion: Using limited amount of biological material, RT-PCR was able to detect a substantial majority of ALK and ROS1 fusions identified by FISH, as well as fusions that were missed by the initial IHC/FISH screening. This study demonstrates RT-PCR as a feasible approach to detecting fusions in NSCLC.

Citation Format: Marc G. Denis, Audrey Vallée, Christine Sagan, Guillaume Herbreteau, Sandrine Charpentier, Jaya Rajamani, Michael Lee, Ellen Ordinario. Detection of ALK, ROS1 and RET fusion transcripts in FFPE samples of non-small cell lung cancer patients using a novel RT-PCR based assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1271.
Abstract 1276: Nucleolin has prognostic value in neuroblastoma patients
Authors: Cangelosi Davide , Brignole Chiara , Bensa Veronica , Tamma Roberto , Malaguti Fabiana , Carlini Barbara , Calarco Enzo , Perri Patrizia , Ribatti Domenico , Fonseca Nuno A. , Moreira Joao N. , Eva Alessandra , Amoroso Loredana , Conte Massimo , Garaventa Alberto , Sementa Angela R. , Corrias Maria V. , Ponzoni Mirco , Pastorino Fabio
Published: June 2022

Background: Neuroblastoma (NB) represents the most frequent form of extracranial solid tumor of infants, responsible for 15% of childhood cancer deaths. Nucleolin (NCL) prognostic value in NB was here investigated.

Methods: NCL protein expression was evaluated in NB patients at diagnosis and after induction chemotherapy. NCL mRNA prognostic value was first assessed in a cohort of 20 stage M NB patients and confirmed in the MultiPlatform786 including 786 NB patients of all stages. Overall and event-free survival curves were plotted by Kaplan-Meier method and compared by log-rank test.

Findings: NCL protein resulted significantly overexpressed in NB tumors compared to the non tumoral counterpart, and higher in stage M compared to stage L patients. In the stage M cohort and in MultiPlatform786 dataset, patients with high NCL mRNA expression revealed a significant lower survival probability than those with low NCL expression. In MultiPlatform786 dataset, NCL mRNA expression was significantly higher in patients with age >18 months, in stage M and in MYCN amplified tumors than in patients with age ˂18 months and in stage L or MS and with MYCN non amplified tumors, respectively. Multivariate analysis suggested NCL has a significant prognostic value even in the model adjusted for established prognostic markers. NCL significantly stratified patients with age <18 and age >18, stage M, >18 months and stage M tumor, stages L or MS, and with MYCN not amplified. A significant correlation between NCL and MYCN, MYC, and TERT was found in two independent datasets. Gene set enrichment analysis revealed a significant positive enrichment of MYC target genes and genes involved in telomerase maintenance. NCL protein resulted down-modulated after chemotherapy, in association with morphological features of neuroblastic differentiation.

Interpretation: NCL is a novel and independent prognostic marker for NB.

Funding: IMH-EuroNanoMed II-2015 (ER-2015-2360441-Eranet) and AIRC IG n. 24397 to PF.

Citation Format: Davide Cangelosi, Chiara Brignole, Veronica Bensa, Roberto Tamma, Fabiana Malaguti, Barbara Carlini, Enzo Calarco, Patrizia Perri, Domenico Ribatti, Nuno A. Fonseca, Joao N. Moreira, Alessandra Eva, Loredana Amoroso, Massimo Conte, Alberto Garaventa, Angela R. Sementa, Maria V. Corrias, Mirco Ponzoni, Fabio Pastorino. Nucleolin has prognostic value in neuroblastoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1276.