bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1129: Combination of ZEN-3694 with talazoparib is a novel therapeutic approach in ER positive breast cancer resistant to CDK4/6 inhibitors, independent of BRCA status
Authors: Kharenko Olesya A. , Patel Reena , Calosing Cyrus
Published: July 2021

Estrogen receptor positive breast cancer (ER+) remains a very prevalent disease with a high mortality rate despite recent successes with new therapies such as CDK4/6, PARP, and PI3K inhibitors. These therapies have shown to significantly prolong progression free survival of metastatic patients, however resistance develops over time in all of the patients and the need to develop new therapeutic strategies is pressing. The bromodomain and extra-terminal domain (BET) proteins play an important role in the regulation of transcription through interaction with acetylated lysine (AcLys) residues of histones or transcription factors. Moreover, BET proteins directly regulate transcription of the estrogen receptor (ER) as well as cell cycle pathway genes. ZEN-3694 is an orally bioavailable BET inhibitor currently being in triple negative breast cancer patients with WT BRCA phenotype in combination with PARP inhibitor talazoparib (NCT03901469) and the data to date has shown that the combination is active.Here, we report a novel combination strategy in ER+ wildtype BRCA1/2 breast cancer resistant cell lines to either palbociclib or abemaciclib combining ZEN-3694 with talazoparib. We demonstrate that this combination has a potent and synergistic effect on inhibition of proliferation of palbociclib and abemaciclib-resistant ER+ cell lines with a WT BRCA phenotype. We also report that the ZEN-3694 + talazoparib induced DNA damage response reflected in the synergistic increase of γH2AX levels, resulting in the increase of apoptosis of the CDK4/6i sensitive and resistant models. Moreover, this combination led to the impairment of the homologous recombination pathway through downregulation of CtIP and WEE1 expression.Furthermore, our data demonstrate that ZEN-3694 targets several mechanisms of endocrine and CDK4/6i resistance including inhibition of estrogen receptor expression and ER signaling, a significant downregulation of several drivers of CDK4/6i resistance such as CDK6, CDK4 and CCND1, as well as inhibition of alternative pathways such as STAT3 and Erbb2. Our RNAseq analysis revealed that ZEN-3694 leads to a strong downregulation of key genes involved in the BRCA1-mediated DNA damage response pathway, as well as double-strand break repair by non-homologous end joining.We conclude that ZEN-3694 in combination with PARP inhibitors is a novel therapeutic strategy in ER+ breast cancer patients progressing on CDK4/6 inhibitors and a promising approach in HR proficient breast cancer.

Citation Format: Olesya A. Kharenko, Reena Patel, Cyrus Calosing. Combination of ZEN-3694 with talazoparib is a novel therapeutic approach in ER positive breast cancer resistant to CDK4/6 inhibitors, independent of BRCA status [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1129.
Abstract 1149: Nuclear receptor 4A1 (NR4A1) antagonists target PD-L1 in colon cancer
Authors: Mohankumar Kumaravel , Karki Keshav , Safe Stephen , Abdelrahim Maen
Published: July 2021

The nuclear orphan receptor 4A1 (NR4A1, Nur77, TR3) is overexpressed in multiple solid tumors including colorectal tumors and is a negative prognostic factor for patient survival. NR4A1 is expressed in colon cancer cells and exhibit pro-oncogenic activity and results of examination of several colon cancer cell lines show that PD-L1 expression is limited and NR4A1 and PD-L1 are co-expressed in SW480 and RKO colon cancer cell lines. Previous studies in breast cancer cells showed that PD-L1 was regulated by NR4A1 which activates transcription factor Sp1 bound to the PD-L1 gene promoter. Knockdown of NR4A1 or Sp1 by RNA interference or treatment with mithramycin an inhibitor of Sp-mediated transcription decreased expression of PD-L1 in RKO and SW480 colon cancer cell lines. These data coupled with ongoing gene expression and PD-L1 promoter studies demonstrate that PD-L1 expression is regulated by NR4A1/Sp1 in colon cancer cells. Bis-indole derived NR4A1 ligand that act as receptor antagonists have been developed in this laboratory and these compounds block pro-oncogenic NR4A1-regulated genes/pathways. Treatment of RKO and SW480 colon cancer cell lines with a series of potent 1,1-bis(3′-indolyl)-1-(3,5-disubstitutedphenyl) analogs decreased expression of PD-L1. These results show that bis-indole derived NR4A1 antagonists act as small molecule mimics of immunotherapeutics that target PD-L1. In vivo applications of NR4A1 ligands that target PD-L1 and their effects on tumor growth and immune surveillance are currently being investigated.

Citation Format: Kumaravel Mohankumar, Keshav Karki, Stephen Safe, Maen Abdelrahim. Nuclear receptor 4A1 (NR4A1) antagonists target PD-L1 in colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1149.
Abstract 1154: Site-specific gene editing with ABBIE for T-cell therapy
Authors: Park Jihyun , Wang Xiaohong , Mirandola Leonardo , Chiriva-Internati Maurizio
Published: July 2021

Therapeutic T-cell engineering using viral vectors has been extensively explored as the most promising therapeutic purpose. However, virus-mediated gene delivery carries many challenges and limitations, such as random insertion of transgene into chromosome, posing risk of oncogenic transformation of the host cells, and complexity in the manufacturing process.We are developing a gene-editing tool, ABBIE (A Binding-Based Integrase Enzyme) which combines the specificity of Cas9 with the non-destructive DNA integration conferred by HIV integrase. Unlike endonucleases, HIV integrase can efficiently insert LTR-flanked DNA without causing dsDNA breaks. In our system, by use of fusing protein of enzymatically dead Cas9 (Cas9DN) to integrase of HIV-1 or its mutants, we were able to perform the gene knock-out (KO) and knock-in (KI) in one step at the target genome loci. For proof of concept, we generated the ABBIE proteins and donor DNA containing fluorescent gene with LTR sequence and tested the ABBIE mediated knock-in with conversion of fluorescent proteins. The integration of fluorescent donor DNA into genome was observed in the 293 T-cells transfected with both WT and Mut ABBIE proteins. Higher integration rate was observed in the cells expressing Mut ABBIE protein. Further, we applied this strategy to deliver the donor fluorescent DNA or chimeric antigen receptors (CARs) via electroporation into the HPRT1 locus for specific insertion using primary human T cells. We observed an average of ~30% and ~16% knock-in efficiency as measured by flow cytometry of GFP+ and CD34+ expression, respectively. The site-specific transgene integration was tested by treating the cells with 6- thioguanine (6-TG). We will further optimize the electroporation condition to maximize editing efficiency. This novel therapeutic toolkit will provide a modular platform for inserting therapeutic genes safely and efficiently into the target genome, further accelerate the offshore immunotherapy development.

Citation Format: Jihyun Park, Xiaohong Wang, Leonardo Mirandola, Maurizio Chiriva-Internati. Site-specific gene editing with ABBIE for T-cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1154.
Abstract 1167: MCM2-7 complex is a novel druggable target for neuroendocrine prostate cancer
Authors: Stoyanova Tanya Ivanova
Published: July 2021

Background: Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer that rarely develops de novo in primary tumors and is commonly acquired during the development of treatment resistance. NEPC is characterized by gain of neuroendocrine markers and androgen receptor (AR) loss, making it resistant to currently used therapeutic strategies targeting the AR signaling axis.

Methods: We utilized whole proteomic analysis of a previously characterized NEPC model and identified MCM2, MCM3, MCM4, and MCM6 proteins as novel targets to inhibit NEPC growth. High levels of MCM2/3/4/6 were significantly enriched in human NEPC when compared to castration-resistant prostate adenocarcinoma in multiple datasets. Colony formation, 3D Matrigel drop and subcutaneous xenograft models of NEPC were used to test the effect of MCM2-7 inhibition using ciprofloxacin in NEPC.

Results: Here, we report that MCM2, MCM3, MCM4, and MCM6 (MCM2/3/4/6) are elevated in human NEPC and high levels of MCM2/3/4/6 are associated with liver metastasis and poor survival in prostate cancer patients. MCM2/3/4/6 are four out of six proteins that form a core DNA helicase (MCM2-7) responsible for unwinding DNA forks during DNA replication. Inhibition of MCM2-7 helicase activity by treatment with ciprofloxacin inhibits NEPC cell proliferation and migration in vitro, significantly delays NEPC tumor xenograft growth, and partially reverses the neuroendocrine phenotype in vivo.

Conclusions: Our findings uncover that MCM2/3/4/6 are markedly elevated in NEPC and represent new druggable targets for therapeutic intervention. The study further reveals that inhibition of MCM2/3/4/6 complex using ciprofloxacin or other targeted approaches may represent a new effective therapy for NEPC. Our studies warrant further exploration of design and screening for MCM2-7 complex-specific inhibitors.

Citation Format: Tanya Ivanova Stoyanova. MCM2-7 complex is a novel druggable target for neuroendocrine prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1167.
Abstract 1182: The NEDD8 inhibitor pevonedistat blocks the repair of topoisomerase I (TOP1)-induced replication damage and synergizes with TOP1 inhibitors
Authors: Sun Yilun , Baechler Andrea , Factor Valentina , Okamoto Kanako , Pommier Yves
Published: July 2021

Genomic DNA is constantly challenged by various genotoxic substances, resulting in an array of DNA lesions including DNA-protein crosslinks (DPCs), one of the most detrimental forms of DNA damage. Topoisomerase I (TOP1) is the molecular target of anti-cancer agents camptothecin derivatives and indenoisoquinolines, which act by trapping TOP1 on genomic DNA. The resulting TOP1-DPCs interfere with DNA replication and transcription and trigger cell death if left unrepaired. In this work, we found that the ubiquitin-like protein NEDD8 plays a pivotal role in the repair of TOP1-DPCs by activating the cullin 4 (CUL4)-RING ubiquitin ligases (CRL4) for K48-linked polyubiquitylation of TOP1-DPCs and their subsequent proteasomal degradation. We identified DCAF13, an understudied family member of DDB1- and CUL4-asscoiated factors, as the receptor of TOP1-DPCs for CRL4. DCAF13 colocalizes with TOP1-DPCs at nascent replication forks and binds the TOP1 core domain with its WD40 domains, linking TOP1 to DDB1-CRL4 for ubiquitylation upon TOP1-induced replication damage. Prompted by these findings, we hypothesized that targeting this NEDD8-activated repair pathway can be developed as a strategy to overcome resistance to TOP1 inhibitors. To test this hypothesis, we performed pharmacological analyses using CellMiner (http://discover.nci.nih.gov/cellminercdb) and found that the activities of TOP1 inhibitors and pevonedistat, a first-in-class NEDD8-activating enzyme inhibitor in clinical trial, are highly correlated across cancer cell lines and with the expression of SLFN11. We also assessed the sensitivity of cultured cancer cells to combination of clinical TOP1 inhibitors and pevonedistat. The combination exhibited synergy in multiple cell lines particularly in human colon cancer cells HCT116 independently of SLFN11. We next investigated the efficacy of combination with TOP1 inhibitor topotecan and pevonedistat in HCT116 xenografts and observed supra-additive effect on tumor growth inhibition. Taken together, our work not only identifies a novel pathway for the repair of TOP1-DPCs but also provides translational evidence to support pevonedistat in combination with TOP1 inhibitors as a rationale therapeutic combination for cancer treatments.

Citation Format: Yilun Sun, Andrea Baechler, Valentina Factor, Kanako Okamoto, Yves Pommier. The NEDD8 inhibitor pevonedistat blocks the repair of topoisomerase I (TOP1)-induced replication damage and synergizes with TOP1 inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1182.
Abstract 121: A genetic signature of cancer dormancy: implications for human disease derived from a novel murine model
Authors: Casabianca Anthony S. , Dudgeon Crissy , Harris Chris , De Subhajyoto , Shah Mihir , Roberts Arthur , Collisson Eric , Balachandran Vinod , Carpizo Darren
Published: July 2021

Pancreatic cancer (PC) is a highly lethal malignancy characterized by local tumor aggressiveness and early metastatic dissemination. While surgical resection is used to treat localized cancers, the majority of patients do recur suggesting the presence of disseminated tumor cells (DTC) at the time of surgery. These DTC's represent minimal residual disease. While most patients recur early, a subset recur late due to the reactivation of dormant tumor cells. Currently, the mechanisms of cancer dormancy are poorly understood in part due to a lack of animal models that reflect human disease. Here we describe a murine model of PC dormancy that mimics outcomes in resected human patients. Using single-cell transcriptomics and an assay for chromatin accessibility, we found dormancy is a distinct cellular state from pre or post-dormant cancer cells. Mechanisms of dormancy include increased expression of transcriptional repressor Dec2, which functionally drives quiescence, and monoallelic suppression of mutant KRAS by DNA methylation. Pathway analysis of dormant tumor cell transcriptomics indicated an up-regulation of enzymes involved in linoleic acid metabolism among others. Linoleic acid treatment of tumor cells resulted in inhibited cell proliferation, decreased tumor growth, and an increase in Dec2 expression. We have identified a dormancy gene signature that is distinct from non-dormant PC cells and found that this correlates with human resected patients with long term survivals, and inversely in patients with short term survivals. We demonstrate that dormancy is characterized by large scale transcriptomic changes and global chromatin remodeling. We developed methods for isolating DTCs from the livers of early stage PC patients undergoing resection. Using these methods we have found that the murine dormancy signature correlated with the gene expression of the DTCs and not primary tumor cells indicating this dormancy signature has human relevance with potential novel therapeutic avenues.

Citation Format: Anthony S. Casabianca, Crissy Dudgeon, Chris Harris, Subhajyoto De, Mihir Shah, Arthur Roberts, Eric Collisson, Vinod Balachandran, Darren Carpizo. A genetic signature of cancer dormancy: implications for human disease derived from a novel murine model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 121.
Abstract 1231: Identification of selective Brd4 degrader for cancer treatment
Authors: Hu Biao , Hu Jiantao , Xu fuming , Huang Liyue , Wang Mi , Wang Shaomeng
Published: July 2021

Human BET family proteins consist of four members, termed Brd2, Brd3, Brd4 and testis-specific BrdT. They all contain two highly conserved tandem N-terminal bromodomains (BD1 and BD2) and a C-terminal extra-terminal (ET) domain. One of the major functions of the BET proteins is to recognize the acetylated lysine residues on histone proteins and transcriptional factors, thereby regulating the gene expression. Recent studies have implicated the BET proteins as a valid pharmaceutical target for treating cancers. Disrupting the interactions by pan-BET inhibitors have displayed beneficial prognosis in both the preclinical animal models and the clinic trials. Nevertheless, dose limiting adverse effects including mild to moderate gastrointestinal toxicity and fatigue, as well as reversible grade 3 thrombocytopenia, have been observed for some pan-BET inhibitors clinically. While the three BET proteins BRD2, BRD3 and BRD4 exhibit redundant and dynamic patterns in regulating gene transcription, the individual protein may have distinct functions. Thus developing of isoform-selective regulatory reagents is of interest to elucidate the biological function of the BET proteins and their applications in pharmaceutical development. Here, we describe our efforts for the development of selective Brd4 degraders with proteolysis-targeting chimera approaches. Our medicinal chemistry campaign led to the discovery of a series of highly potent Brd4 selective degraders. Treat the tumor cells with these selective Brd4 degraders resulted in rapid, sustained and selective degradation of Brd4. RNA-seq and real time RT-PCR analysis indicates the selective Brd4 degrader exhibits distinct regulation of gene expression in tumor cells comparing to pan-BET degraders and inhibitors. Further tests with flow cytometry analysis and WST-8 assays indicates that selective Brd4 degraders strongly suppress the tumor cells growth and induces the cell death through induction of apoptosis. Strong tumor suppression was also observed with treatment of selective Brd4 degrader in mouse xenograft models. These results suggest that the selective Brd4 degraders are strong candidates for potential tumor treatment.

Citation Format: Biao Hu, Jiantao Hu, fuming Xu, Liyue Huang, Mi Wang, Shaomeng Wang. Identification of selective Brd4 degrader for cancer treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1231.
Abstract 1232: Using mesoporous silica nanoparticles (MSNs) for delivering the mesoionic compound MIH 2.4Bl in treating breast cancer
Authors: Debnath Dipti , Nazzal Sami , Acharya Suchismita , Souza Helivaldo Diógenes da Silva , Filho Petrônio Filgueiras de Athayde , Fudala Rafal , Mathis J Michael
Published: July 2021

Breast cancer is the most frequently diagnosed cancer and the leading cause of cancer death in women worldwide, accounting for approximately 24% of all new cancer cases and is the leading cause of cancer death in over 100 countries. Thus, breast cancer is one of the most critical public health problems in the world facing women. There is a growing interest in studying the biological activity of mesoionic compounds, which possesses a 5-membered heterocyclic aromatic ring associated with a sextet of electrons. Mesoionic compounds have shown promising potential as anti-cancer agents due to their unique structure and reaction properties. We reported the synthesis of a new 1,3-thiazolium-5-thiolate derivative of a mesoionic compound (MIH 2.4Bl) and the characterization of its selective cytotoxicity on a panel of breast cancer cells lines. The results of our studies suggest a possible induction of apoptotic death through mitochondrial dysfunction by the treatment with MIH 2.4Bl. Based on our previous findings, MIH 2.4Bl is a promising candidate for treating breast cancer. However, a major challenge facing cancer therapy is the selective destruction of malignant cells while sparing normal cells to preserve tissue integrity. The development and use of drug delivery systems is a recognized approach to improve the efficacy of chemotherapy agents. Nonetheless, drug delivery systems have been unexplored in the context of mesoionic compounds. Drug delivery based on nanotechnology has flourished as a platform applicable to cancer therapy. Among various types of nanoparticles, mesoporous silica nanoparticles (MSNs) are more attractive nanomaterials for a broad range of functionality including gene/drug delivery, cancer therapy and bioimaging due to their broad surface area, less toxicity, low density, excellent biocompatibility, higher drug loading capacity, and impressive mechanical and thermal stability. In this preliminary work, we present an improved delivery strategy of a newly developed formulation of MIH 2.4BI compound with MSNs as the delivery agent. Also, physico-chemical characterization of the nanoparticles and cytotoxicity analyses using a panel of breast cancer cell lines were performed. These studies support the potential therapeutic use of MIH 2.4Bl in treating breast cancer.

Citation Format: Dipti Debnath, Sami Nazzal, Suchismita Acharya, Helivaldo Diógenes da Silva Souza, Petrônio Filgueiras de Athayde Filho, Rafal Fudala, J Michael Mathis. Using mesoporous silica nanoparticles (MSNs) for delivering the mesoionic compound MIH 2.4Bl in treating breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1232.
Abstract 1238: Treating estrogen receptor (ER)-negative and triple-negative breast cancer by targeting STAT3 signaling with putative STAT3 inhibitors
Authors: Kim Hyejin , Fricke Doerte R. , Xu Jimin , Chen Haiying , Zhou Jia , Shen Qiang
Published: July 2021

Approximately 20% to a quarter of breast cancer cases are estrogen receptor (ER)-negative breast cancer (ENBC), including the most aggressive subtype, triple-negative breast cancer (TNBC), which spreads early and shows poor prognosis. ENBCs do not respond to selective ER modulators such as tamoxifen. Therefore, there is an urgent need for non-ER-based therapies for ENBC. Recently, STAT3 was found to play an important role in maintaining cancer stem cells in vitro and in mouse models of human cancers. In addition, during mammary carcinogenesis, STAT3 is highly activated in mammary epithelial cells involving in mammary epithelial cell transformation, tumor initiation, progression, and metastasis. STAT3 has been traditionally considered as non-targetable or undruggable, and there is currently a lack of FDA-approved STAT3 inhibitors for clinical use. We recently developed a putative STAT3 inhibitor, HJC0152, and a series of HJC0152-based analogues as orally bioavailable STAT3 inhibitors. These small molecules have been evaluated and a number of them showed low micro-molar potency in inhibiting the proliferation of a panel of breast cancer cell lines. Compound JMX0804 showed improved anti-proliferative effects against the highly aggressive TNBC cell line MDA-MB-231 and other ER+ and ER- breast cancer cell lines in comparison to HJC0152. JMX0804 inhibited STAT3 phosphorylation at the Tyr705 residue and induced apoptosis in MDA-MB-231 cells. In vivo, mice treated with JMX0804 at a 10 mg/kg dose via i.p. showed reduced growth of xenograft tumors arisen from MDA-MB-231 cells, with minimal impact on general health and body weight. Our current research supports the notion that JMX0804 suppresses breast cancer growth by inhibiting breast cancer cell proliferation and inducing apoptosis via mediating STAT3 signaling. JMX0804 is currently undergoing further characterization for breast cancer metastasis and mechanisms of action as well as optimization to improve drug properties including potency and safety profile. This work is supported by NIH/NCI grant R01 CA231150.

Citation Format: Hyejin Kim, Doerte R. Fricke, Jimin Xu, Haiying Chen, Jia Zhou, Qiang Shen. Treating estrogen receptor (ER)-negative and triple-negative breast cancer by targeting STAT3 signaling with putative STAT3 inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1238.
Abstract 1304: AbYlinkTM: A site-selective labeling method for preclinical imaging of therapeutic antibodies
Authors: Postupalenko Viktoriia , Marx Léo , Viertl David , Gasilova Natalia , Plantin Mathilde , Gsponer Nadège , Johanssen Alexandre , Denoel Thibaut , Hagens Gerrit , Segura Jean-Manuel , Levy Frédéric , Garrouste Patrick , Prior John , Schottelius Margret , Schaefer Niklaus , Nyanguile Origene
Published: July 2021

Clinical development of therapeutic antibodies relies on robust and thorough preclinical evaluation of multiple pharmacodynamic and pharmacokinetic parameters of antibody candidates. While current methods are available for in vitro characterization of antibodies, in vivo determination of tissue distribution, tumor accumulation and retention and potential off-target effects cannot be performed non-invasively in vivo over a prolonged period of time.

Here, we used AbYlinkTM, a novel site-selective labeling method of native antibodies, enabling the single-step covalent conjugation of a radionuclide chelator to the Fc domain of any IgG antibody isotype. Given the selectivity of this reaction for the Fc region of the immunoglobulin, labeling of the antibody at this location should not compromise binding to the antigen. Chelators that are covalently attached to the antibody allows for the labeling with radioisotopes such as 68Ga, 177Lu, 111In and 89Zr, and for the sensitive and quantitative molecular in vivo measurement by Positron Emission Tomography (PET) or Single Photon Emission Computed Tomography (SPECT). AbYlinkTM also permits the conjugation of fluorescent dyes to antibodies with the aim to measure antibody pharmacodynamic parameters by intravital microscopy. In this work, we have validated the technology by labeling several antibodies and ADCs with different payloads. Importantly, antibodies conjugated with AbYlinkTM retained the same affinity to the antigen as the original, unconjugated antibodies, contrary to the randomly labelled antibodies. We will also present data on the specific radiolabeling of commercially available trastuzumab with 111Indium and the SPECT images obtained after injection of mice bearing HER2-positive tumors with 111In-trastuzumab.

The present data suggest that AbYlinkTM can be used to support preclinical drug discovery tools to assist in the selection of therapeutic antibody candidates and/or for pharmacodynamic assessment of commercial antibodies. Furthermore, the specificity of the chemical conjugation suggests that AbYlinkTM can help to control the chelator distribution on the antibody drug conjugates.

Citation Format: Viktoriia Postupalenko, Léo Marx, David Viertl, Natalia Gasilova, Mathilde Plantin, Nadège Gsponer, Alexandre Johanssen, Thibaut Denoel, Gerrit Hagens, Jean-Manuel Segura, Frédéric Levy, Patrick Garrouste, John Prior, Margret Schottelius, Niklaus Schaefer, Origene Nyanguile. AbYlinkTM: A site-selective labeling method for preclinical imaging of therapeutic antibodies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1304.
Abstract 1306: Primary NK cells and luciferase expressing reporter cell lines for use in developing ADCC assays for immuno-oncology drug screening
Authors: Liu Haiyun , Fera Brian Della , Foulke John , Chen Luping , Tian Fang
Published: July 2021

Immuno-oncology is one of the fastest growing fields in oncology, and cancer immunotherapy in recent years represents a milestone in the treatment of cancer. Natural killer (NK) cells are potent innate effectors capable of targeting and killing virally infected and malignant cells. The use of monoclonal antibodies (mAbs) targeting tumor antigens triggers antibody-dependent cellular cytotoxicity (ADCC) by NK cells, which has shown prominent clinical efficacy in clearing tumors in cancer patients. Therefore, the development of tumor cell targeting mAbs and biologics dictate a clear need for human cell-based models to evaluate antibody efficacy. In this study, human PBMCs and purified NK cells from healthy donors are used to develop an ADCC assay. The NK cells purified from PBMCs are >90% CD45+ CD56+, express high levels of CD16, and NK activation markers such NKG2A, KIR2DL1, NKp44 upon activation. The activated NK cells exhibit both strong direct and antibody-dependent cytotoxicity. Genotyping for FcγRIIIa receptor F and V variant is also performed for cells isolated from different donors, which is the genetic polymorphism that affects ADCC response. In addition, we employ a portfolio of luciferase reporter tumor cell lines to evaluate NK cell direct killing and ADCC targeted killing. These include K562, Wil2S, BT474, A431, A375 parental, and CRISPR edited A375 KRAS mutant cell lines engineered to express luc2 reporter. The commonly targeted cell surface markers, such as CD20, Her2, and EGFR, are highly expressed within this panel of target tumor cell lines. We validate the ADCC assay system through testing the efficacy for some of the well-known mAb biologics such as anti-CD20, anti-HER2, and anti-EGFR. After co-culture of activated primary NK cells with luciferase reporter target cell lines and the addition of dilutions of ADCC and isotype control antibodies, we are able to show target cell specific and dose-dependent killing effects. The luciferase reporter provides great convenience in measuring target cell killing without having to use radioactive 51Cr release assay or pre-label the cells with calcein. This data demonstrates the utility of primary NK cells along with a portfolio of well characterized luciferase reporter tumor cell line to develop “custom” designed ADCC assays for studying NK cell immune response and screening for biological drugs for cancer immunotherapy.

Citation Format: Haiyun Liu, Brian Della Fera, John Foulke, Luping Chen, Fang Tian. Primary NK cells and luciferase expressing reporter cell lines for use in developing ADCC assays for immuno-oncology drug screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1306.
Abstract 1320: NanoGhost: Harnessing the power of stem cell in an off-the-shelf novel targeted nano-delivery platform
Authors: Machluf Marcelle , Machluf Marcelle , Hamias Shani , Levii Lior , Kashtan Osnat Bohana
Published: July 2021

Mesenchymal stem cells (MSC) have been extensively investigated as cell carriers or cell therapies for treating a wide range of malignant and inflammatory diseases. In addition, recent studies show that MSCs' tropism to malignant and inflamed tissues as well as their immunomodulatory capacity, both are largely governed by direct cell-cell interactions facilitated by membrane-bound proteins. Never the less the translation of MSCs into broadly-applicable clinical applications may face obstacles due to MSCs' susceptibility to host-induced changes, limiting their ability to deliver a long-lasting effect. Based on this knowledge we developed a scalable technology for the production of nano-vesicles from the cell membrane of MSCs. These nano-vesicles, termed Nano-Ghosts (NGs), and our data demonstrate that these NGs can be used as a cancer targeted delivery platform, effectively loaded and used to selectively deliver diverse therapeutics including biological drugs, small molecules, and nucleic acids. In several preclinical cancer models (pancreatic, NSCLAC) we have shown selective NG targeting resulting in reduction in tumor growth and prolonged survival. In this study, we extend previous data and demonstrate the ability of NGs to penetrate the BBB in a two mice models for GBM demonstrating high and specific accumulation of NGs in the tumor while no off-target inn normal brain tissue nor filtrating organs. The lethal effect of drug carrying NGs was expanded by using cytotoxic ligand derived from CTLs (TRAIL) genetically engineered on the surface of the MSC-NGs, creating a new class of NGs termed immunotherapeutic NGs (iNGs). The membrane bound TRAIL enable the iNGs to inhibit significantly mice bearing melanoma tumors, without the need of a loaded drug. Our results, clearly demonstrate the translational potential of NGs, both as targeted carriers and as novel immunomodulatory biologics, for oncological and immunological applications.

Citation Format: Marcelle Machluf, Marcelle Machluf, Shani Hamias, Lior Levii, Osnat Bohana Kashtan. NanoGhost: Harnessing the power of stem cell in an off-the-shelf novel targeted nano-delivery platform [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1320.
Abstract 1335: Dual MCT1/4 inhibition promotes anti-tumor immunity in triple-negative breast cancer
Authors: Goreczny Gregory James , Escobedo Jaime , Sandanayaka Vincent
Published: July 2021

Despite the recent FDA approval of Abraxane (nab-paclitaxel) and Atezolizumab (anti-PD-L1) for a subset of Triple-Negative Breast Cancer (TNBC) patients, there remains an unmet need to develop novel therapeutic strategies to effectively combat this disease. Nirogy Therapeutics has developed a first-in-class, orally bioavailable small molecule inhibitor targeting MonoCarboxylate Transporters (MCTs), specifically MCT1 and MCT4, which transport lactate into and out of the cell. MCT1/4 expression are upregulated in TNBC patients, and high MCT4 expression correlates with worse outcome. Although a MCT1-specific inhibitor is currently under investigation in clinical trials for some cancers that express only MCT1, primary resistance to these inhibitors occur via the upregulation of MCT4. Most cancers overexpress both MCT1 and MCT4. Thus, the need for dual MCT1/4 inhibitors (dMCTi) that block both MCT1 and MCT4 is crucial to effectively treat these patients. We have shown that our dMCTi compound, NGY-B, exhibits direct cytotoxicity against a panel of TNBC cell lines with various MCT1/4 expression patterns, and it inhibits lactate import through MCT1 and export through MCT4. Furthermore, NGY-B exhibited a significant tumor growth reduction in an MDA-MB-231 CDX mouse model of TNBC compared to the current standard-of-care chemotherapy. Using the syngeneic 4T1 mouse model of TNBC, we have shown that NGY-B also exhibited a significant reduction of tumor growth, indicating potential impact on anti-tumor immunity as well. Further analysis of the 4T1 model showed that NGY-B significantly enriched the CD8+ tumor infiltrating lymphocyte (TIL) population while exhibiting a concurrent reduction in the macrophage M2/M1 ratio, indicating preferred polarization to M1-phenotype. Also, we observed a significant reduction in granulocytic suppressor cells, gMDSCs in the tumor microenvironment (TME). Further work has demonstrated that NGY-B synergizes with immune checkpoint blockade to promote an anti-tumor immune response. By blocking lactate accumulation in the TME, our compound activates local immunity by overcoming lactate mediated immunosuppression in cytotoxic T-cells while suppressing inhibitory immune populations. Therefore, such a broader effect on multiple immune cells by NGY-B leads to overall immune activation in the TME. Our pre-clinical work shows that NGY-B intervenes two key hallmarks of caner; metabolism and immunity, providing us a novel therapeutic modality to combat TNBC.

Citation Format: Gregory James Goreczny, Jaime Escobedo, Vincent Sandanayaka. Dual MCT1/4 inhibition promotes anti-tumor immunity in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1335.
Abstract 1339: Investigations into rational combination approaches with ivosidenib in a mutant IDH1 acute myeloid leukemia patient-derived xenograft model
Authors: Hudson Christine , Narayanaswamy Rohini , Ronseaux Sebastien , Bowden Chris , Nicolay Brandon
Published: July 2021

Background: Somatic mutations in isocitrate dehydrogenase 1 (IDH1) arise in roughly 6-10% of patients with acute myeloid leukemia (AML). These mutations lead to the production of the oncometabolite 2-hydroxyglutarate at concentrations shown to interfere with epigenetic control of gene expression, which prevents hematopoietic cell maturation. Ivosidenib (IVO; AG-120) is an oral, potent, targeted oral inhibitor of mutant IDH1 (mIDH1) and is FDA-approved for the treatment of mIDH1 relapsed/refractory AML patients and mIDH1 newly diagnosed (ND) AML in adults ≥ 75 years of age or with comorbidities precluding intensive induction chemotherapy.

Methods: Currently, IVO is being investigated in combination with azacitidine (AZA) in the actively enrolling phase 3 AGILE study (NCT03173248). To support this ongoing effort, studies were undertaken to explore the IVO + AZA combination in a mIDH1 AML patient-derived xenograft (PDX) model in vivo. These studies examined the timing of IVO + AZA treatment with respect to disease burden control and durability in mice inoculated with a disseminated mIDH1 AML PDX model, both as a frontline treatment as well as in a maintenance therapy setting following either venetoclax (VEN) + AZA, or VEN + AZA + IVO. In addition, the studies examined the utility of combining IVO + VEN as an alternative frontline treatment approach in the same preclinical mIDH1 AML PDX disseminated model.

Results: The IVO + AZA combination led to a > 99% reduction in hCD45+ cells within the bone marrow and undetectable disease in the peripheral blood of 7/10 mice by Week 10. Similar results were observed with IVO + VEN treated animal bone marrow (99% reduction in hCD45+ cells) and undetectable disease was observed in 8/10 mice by week 10.

Conclusions: Each combination was well tolerated by the mice. The observed disease control with either combination in the animals was greater than any of the three single agents alone. Ongoing follow-up xenograft studies investigating the utility of the combinations with IVO as a maintenance therapy are still ongoing and outcomes will be presented.

Citation Format: Christine Hudson, Rohini Narayanaswamy, Sebastien Ronseaux, Chris Bowden, Brandon Nicolay. Investigations into rational combination approaches with ivosidenib in a mutant IDH1 acute myeloid leukemia patient-derived xenograft model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1339.
Abstract 1400: SWI/SNF chromatin remodeling complex regulation of YAP-dependent enhancers drives therapeutic resistance in triple-negative breast cancer
Authors: Li Yihao , Qiu Xintao , Liu Hui , Wang Xiaoqing , Geck Renee C. , Tewari Alok , Chow Kin-Hoe , Cejas Paloma , Nguyen Quang-Dé , Long Henry , Liu Shirley X. , Toker Alex , Brown Myles
Published: July 2021

Outcomes for patients with triple negative breast cancer (TNBC) remain poor despite significant advances in the treatment of other breast cancer subtypes. We report an epigenetic mechanism leading to the adaptive resistance of TNBC to fibroblast growth factor receptor (FGFR) inhibitors. Prolonged FGFR inhibition suppresses the function of BRG1-dependent chromatin remodeling leading to an epigenetic state that derepresses YAP-associated enhancers. These chromatin changes induce the expression of several amino acid transporters resulting in increased intracellular levels of specific amino acids that reactivate mTORC1. Consistent with this mechanism, addition of mTORC1 or YAP inhibitors to FGFR blockade synergistically attenuated the growth of TNBC patient-derived xenografts (PDX) models. Treatment-induced YAP/TEAD accessible chromatin was observed in a subpopulation of PDX cells by single cell analysis of accessible chromatin. Thus, combinatorial therapies based on the YAP and mTORC1 dependent feedback loop have the potential to improve the efficacy of FGFR inhibitors in TNBC.

Citation Format: Yihao Li, Xintao Qiu, Hui Liu, Xiaoqing Wang, Renee C. Geck, Alok Tewari, Kin-Hoe Chow, Paloma Cejas, Quang-Dé Nguyen, Henry Long, Shirley X. Liu, Alex Toker, Myles Brown. SWI/SNF chromatin remodeling complex regulation of YAP-dependent enhancers drives therapeutic resistance in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1400.
Abstract 1434: Uncovering the mechanism of adaptive resistance to RET inhibitors in RET rearranged thyroid cancer
Authors: Raman Renuka , Villefranc Jacques , Ullmann Timothy , Thiesmeyer Jessica , Anelli Viviana , Pauli Chantal , Bareja Rohan , Eng Kenneth Wha , Dorsaint Princesca , Wilkes David , Beg Shaham , Shaw Reid , Churchill Michael , Gumpeni Naveen , Scognamiglio Theresa , Rubin Mark , Grandori Carla , Mosquera Juan , Mosquera Juan , Mosquera Juan , Dephoure Noah , Sboner Andrea , Elemento Olivier , Houvras Yariv
Published: July 2021

Kinase inhibitors are a critical tool for cancer treatment, but their efficacy is limited by resistance mechanisms. In thyroid and lung cancer RET gene rearrangements result in constitutive MAPK pathway activation and drive malignancy. While treatment with new selective RET inhibitors has been associated with significant clinical responses, a majority of patients experience a partial response or disease stabilization as their best clinical outcome. Resistance to RET inhibitors has emerged as a clinical problem requiring new treatment strategies. Using a combination of drug screening, proteomic, and biochemical profiling we identified a strategy to overcome adaptive resistance to RET inhibitors in human thyroid cancer cell lines and vertebrate animal models. The identification of alternative signaling pathways that reactivates ERK signaling as a mechanism of resistance to RET inhibitors provides an opportunity to anticipate resistance to selective RET inhibitors and use combination therapy that leads to more significant and durable anti-tumor responses in patients with RET rearranged cancers.

Citation Format: Renuka Raman, Jacques Villefranc, Timothy Ullmann, Jessica Thiesmeyer, Viviana Anelli, Chantal Pauli, Rohan Bareja, Kenneth Wha Eng, Princesca Dorsaint, David Wilkes, Shaham Beg, Reid Shaw, Michael Churchill, Naveen Gumpeni, Theresa Scognamiglio, Mark Rubin, Carla Grandori, Juan Mosquera, Juan Mosquera, Juan Mosquera, Noah Dephoure, Andrea Sboner, Olivier Elemento, Yariv Houvras. Uncovering the mechanism of adaptive resistance to RET inhibitors in RET rearranged thyroid cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1434.
Abstract 1448: Combination therapy of anti-sense oligonucleotide targeting TGF-beta2 (TASO) and IL-2 (Proleukin) has anti-cancer effect in solid cancer
Authors: Park Jun-Eui , Koo Jihye , Lee Hong-Kyu , Kim Tae Hun
Published: July 2021

Background: TGF-Β (transforming growth factor-beta) is an essential cytokine for tumor proliferation and metastasis. The expression of TGF-Β correlates with malignancy of various cancers and involves immunosuppression and angiogenesis of a tumor. IL-2 is a major cytokine to proliferate T cells and NK cells which are major players of cancer immunity. However, the toxicity of high dose IL-2 limits its use in cancer therapy. Combination treatment of TGF-Β inhibitor and IL-2 would have an anti-tumor effect by immune cells through diminishing immunosuppression by TGF-Β and enforcement of immune cells by IL-2. Trabedersen is an anti-sense oligonucleotide targeting human TGF-Β mRNA. It is shown that Trabedersen is well tolerable in cancer patients and effective reagent to treat pancreatic cancer, melanoma, and glioblastoma. Proleukin is the only approved IL-2 reagent to treat Renal cell carcinoma and Melanoma.

Methods: Trabedersen and Proleukin activated human PBMC (Peripheral Blood Mononuclear Cell) are treated to several solid cancer cell lines, such as Breast cancer, Pancreatic cancer, Melanoma, Lung cancer, and Colon cancer to see the cytotoxicity effect of combination therapy of Trabedersen and Proleukin. NSG mouse (NOD Scid Gamma mouse, NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ) which are humanized by human PBMC engraftment and the tumor growth of Melanoma and TNBC (Triple Negative Breast Cancer cell) cell line are monitored.

Results: The combination treatment of Trabedersen and low dose Proleukin decreased cancer cell viability in vitro experiment in solid cancer cell lines. Melanoma and TNBC tumor growth was delayed in humanized NSG mouse model by Trabedersen and low dose IL-2 combination therapy and tumor growth delay was statistically significant to Trabedersen alone or IL-2 alone group. Tumor infiltrating lymphocyte population was increased in Trabedersen treated group and FoxP3+ regulatory T cell population in blood and tumor microenvironment was decreased by treatment of Trabedersen.

Conclusion: TGF-Β inhibitor (Trabedersen) and low dose IL-2 (Proleukin) combination treatment is expected to be an effective regimen in solid cancer treatment than individual treatment by alteration of tumor environment. Modulation of the dose of Proleukin expects to help reduce the toxicity of IL-2 and increase the anti-cancer effect by combination with Trabedersen.

Citation Format: Jun-Eui Park, Jihye Koo, Hong-Kyu Lee, Tae Hun Kim. Combination therapy of anti-sense oligonucleotide targeting TGF-beta2 (TASO) and IL-2 (Proleukin) has anti-cancer effect in solid cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1448.