bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2908: PT217, an anti-DLL3/anti-CD47 bispecific antibody, exhibits anti-tumor activity through novel mechanisms of action
Authors: Jia Haiqun(John) , Fang Dorthy , Lobsinger Molly , Long Yan , Chen Kelan , Lam Albert , Guo Yangyang , Selesniemi Kaisa , Wang Ming , Zou Hui
Published: June 2022

Small cell lung cancer (SCLC) is the most lethal subtype of lung cancers with few patients surviving more than 5 years from diagnosis. While SCLC initially responds well to standard chemotherapy treatment, chemotherapy-resistant recurrence is characteristic and available treatment options for recurrent and refractory diseases are limited. Immunotherapy as an alternative or additional treatment has been very successful in treating many cancers, yet it has shown disappointingly limited benefits in SCLC and notable treatment related toxicities have been observed. Delta like 3 (DLL3) is an attractive target for SCLC immunotherapy, as its expression is highly restricted to SCLC and can therefore be used for targeting purposes. On the other hand, CD47 is also strongly expressed on the surface of SCLC cells, where it inhibits macrophage-mediated tumor cell killing. We have developed PT217, a DLL3/CD47 bispecific antibody with effector effect competent Fc to specifically target SCLC cells via two mechanisms. First, PT217 blocks the CD47-SIRPα interaction and stimulates phagocytosis of tumor cells by macrophage. Second, PT217 exhibits robust effector effect (ADCC by NK cells and ADPC by macrophage) mediated by the functional Fc of PT217. Interestingly, it appears that the tumor cells targeted by NK cells are subsequently engulfed by macrophage. Elevated phagocytosis of the tumor cells by macrophages and dendritic cells is expected to increase the presentation of tumor neoantigens that may eventually lead to T cell-mediated killing of tumor cells that do not expression DLL3. In vivo, PT217 demonstrated tumor suppression activity in xenograft mouse models. In addition to the strong tumor killing activity, PT217 also demonstrated good safety profile in NHP studies. Furthermore, PT217 exhibits regular mAb-like manufacturability in process development and CMC. We expect to initiate a phase 1 trial to target SCLC in the first half of 2022.

Citation Format: Haiqun(John) Jia, Dorthy Fang, Molly Lobsinger, Yan Long, Kelan Chen, Albert Lam, Yangyang Guo, Kaisa Selesniemi, Ming Wang, Hui Zou. PT217, an anti-DLL3/anti-CD47 bispecific antibody, exhibits anti-tumor activity through novel mechanisms of action [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 2908.
Abstract 2922: Targeting triple negative breast cancer with a VHL recruiting EZH2 protein degrader
Authors: Dale Brandon , Anderson Chris , Park Kwang-su , Kaniskan H. Ümit , Yu Xufen , Jin Jian
Published: June 2022

EZH2 (enhancer of zeste homolog 2) is the main catalytic subunit of the polycomb repressive complex 2 (PRC2) that catalyzes methylation of histone H3K27. EZH2 is overexpressed in a broad spectrum of cancers including prostate, myeloma and lymphoma, and high expression correlates with poor prognosis. However, EZH2 inhibitors, which do not affect EZH2 protein levels, are ineffective at inhibiting growth of TNBC and other breast cancer cell lines with EZH2 overexpression even though they effectively inhibit the catalytic activity of EZH2/PRC2. It has also been shown that overexpression of EZH2 is a major driver for breast cancer development and progression, and knockdown of EZH2 inhibits proliferation of TNBC and other breast cancer cells. These results together suggest that EZH2 overexpression, but not the methyltransferase activity of EZH2, is critical for driving breast cancer progression. Bivalent inhibitor technologies such as PROTACs (proteolysis targeting chimeras) and hydrophobic tagging have been successfully applied to selective degradation of multiple protein targets. CRBN-recruiting (E7) and VHL-recruiting EZH2 (YM281) EZH2 targeting PROTACs have been reported, but do not show potent efficacy in targeting TNBC. Here, we characterize MS8815, a VHL-recruiting EZH2 degrader, that we created. MS8815 displayed nanomolar efficacy at EZH2 degradation and potent in vitro effects on TNBC cell lines, no off-target interactions and utilized the canonical PROTAC mechanism of action. Based on these promising preliminary results, we report MS8815 as the best-in-class EZH2 PROTAC degrader that is able to efficiently degrade EZH2 and kill TNBC cells.

Citation Format: Brandon Dale, Chris Anderson, Kwang-su Park, H. Ümit Kaniskan, Xufen Yu, Jian Jin. Targeting triple negative breast cancer with a VHL recruiting EZH2 protein degrader [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 2922.
Abstract 2948: A multimodal, label-free tool for gentle and efficient cell separation
Authors: Travers Kevin , Stevens Chris , Jett Susana , Weaver Suzanne , de Groot Teddy , Ha Stefan , Facer Geoff , Lazaruk Kathy
Published: June 2022

LevitasBio has developed the LeviCell™ platform, a powerful new technology for cell separation and characterization that utilizes magnetic fields to levitate cells. Unlike other methods, cell separation via magnetic levitation does not require dyes, antibodies or specific markers, and the cells are not modified or perturbed in any fashion. However, magnetic levitation also does not preclude the use of these tools to further enhance separation of cells. In its simplest form, magnetic levitation requires only the addition of an inert paramagnetic compound to the media in which the cells are suspended. The cells are subsequently introduced into an externally applied magnetic field while contained within a single-use cartridge, which causes the cells to levitate in solution. The cells’ intrinsic properties, including density and magnetic susceptibility, determine the height to which the cells levitate. As different cell types often have different such properties, they may often levitate to different heights, facilitating their separation. Continuous imaging allows for complete sample analysis and characterization with the benefit of real-time control over the cell separation. The simplicity of magnetic levitation enables cells to be treated gently without use of high pressure or other perturbations that commonly lead to increased cellular stress responses, specific cell type activation, or even cell death. The direct and gentle flow path leads to high yields of live cells even with very low input cell numbers.

This novel cell separation technology enables a variety of applications, including: the enrichment of viable cells from dead/dying cells and debris; separation of different cell types possessing distinct densities; and, in cases where the intrinsic density changes, the separation of differentiated from undifferentiated cells. Given that the levitation chamber is flanked by two magnets on the top and bottom, the levitation technology also inherently allows for the use of magnetic particles to deplete undesired cells within a cell population. Here we illustrate the performance of anti-CD45 antibody-conjugated magnetic particles for CD45+ cell depletion with the LeviCell system. CD45+ depletion rates of greater than 99% are readily obtained across a wide range of total CD45+ input cell numbers, from 105 to 107 total cells. Furthermore, CD45+ cell depletion is performed simultaneously with live cell enrichment, allowing for the collection of a high viability CD45- live cell population.

Citation Format: Kevin Travers, Chris Stevens, Susana Jett, Suzanne Weaver, Teddy de Groot, Stefan Ha, Geoff Facer, Kathy Lazaruk. A multimodal, label-free tool for gentle and efficient cell separation [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 2948.
Abstract 2950: Coexpression networks of miRNAs and gene transcripts coinciding with lung adenocarcinoma progression and survival
Authors: Young Corey D. , Dammer Eric B. , Griffen Ti'ara L. , Nunez Sha'kayla K. , Dill Courtney , Carey Kaylin M. , Lillard James W.
Published: June 2022

Lung & bronchus cancer is the second most diagnosed cancer, causing the most cancer-related deaths in the United States. A lung and bronchus subtype, NSCLC (non-small cell lung cancer), accounts for over 75% of new lung cancer diagnoses of which lung adenocarcinoma (LUAD) is the most common. Here, we studied the role of microRNAs and the transcriptome expression profile in LUAD prognosis, progression, and overall patient survival. We analyzed RNA-sequence data from a cohort of LUAD patients from The Cancer Genome Atlas (TCGA). The correlations of microRNA gene expression to transcription networks identified by weighted gene co-expression network analysis (WGCNA) were further explored in an upstream regulator/signaling pathway analysis, gene ontology (GO) process enrichment, and differential expression analysis. Mature microRNA abundance was also integrated with the mRNA network via correlation analysis. Our analysis revealed 18 clusters (modules) of highly correlated gene transcripts. The turquoise and red modules were of particular interest, as they were correlated negatively with overall survival, positively with stage 1 status, and negatively to later pathologic stages (3-4). Additionally, high select miRNA correlation to the red module transcripts (e.g., C16Orf89, ADGRF5, CFAP221, and SELENBP1; all with module eigengene correlation rho ≥ 0.75), and to the turquoise module (e.g., DDX39B and LENG8; each with rho ≥ 0.85) suggests these are miRNA-mRNA coregulatory networks of which both the gene products and potentially upstream miRNAs may serve as promising mechanistic targets and prognostic markers for LUAD, meriting further study.

Citation Format: Corey D. Young, Eric B. Dammer, Ti'ara L. Griffen, Sha'kayla K. Nunez, Courtney Dill, Kaylin M. Carey, James W. Lillard. Coexpression networks of miRNAs and gene transcripts coinciding with lung adenocarcinoma progression and survival [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 2950.
Abstract 2956: Superenhancer-drvien SMC4 promote cell growth and proliferation in multiple myeloma
Authors: Jia Yunlu , Zhou Jianbiao , Tan Tze-King , Chung Tae-Hoon , Chen Yongxia , Zhang Enfan , Cai Zhen , Sanda Takaomi , Chng Wee-Joo
Published: June 2022

Background Multiple myeloma (MM) is an incurable malignancy characterized by complex heterogeneous cytogenetic abnormalities. Defined as large clusters of cis-acting enhancers, super-enhancers (SEs) could promote oncogenic transcription to which cancer cells highly addicted. Structural maintenance of chromosome 4 (SMC4) is a core subunit of condensin complexes, which contributes to chromosome condensation and segregation. Here, we identified SMC4 as a novel SE-associated oncogene of myeloma cells, whose functional roles in MM remain largely elusive.

Methods We performed H3K27ac ChIP-seq and RNA-seq on primary MM patient samples, MM cell lines, normal CD138+ plasma cell and lymphoma cell lines as the controls, then the ROSE analysis was used to annotate SEs and their associated genes. THZ1 as a transcriptional CDK7 inhibitor was used to further filter THZ1-responsive genes. Combined analysis of THZ1-sensitive and SE-associated gene uncovered a number of promising MM oncogenes as SMC4. The public data mining, RNA interference, CRISPR/Cas9 gene editing system, overexpression and a variety of cellular functional assays were performed to determine the oncogenic effects of SMC4 on MM malignant. Transcriptome analysis with SMC4 silencing and overexpression, and the underlying molecular mechanism of SMC4 oncogenic role in MM are ongoing.

Results We have demonstrated SMC4 as one of the SE-associated genes specific to MM patient samples and cell lines. SMC4 was related to myelomagenesis with increased expression from MGUS, SMM to MM cases, and overexpression of SMC4 predicted poor overall survival of MM patients, suggesting its clinical relevance. SMC4 silencing in MM cells suppressed cell proliferation and lead to cell apoptosis. Repression of the SMC4-SE region also led to impairment of cell viability and cell growth, consistent with an oncogenic function.

Conclusion In summary, our integrative approached by combing H3H27ac ChIP-seq, RNA-seq and THZ1-sensitive transcripts identified the landscape of SEs and novel oncogenic transcripts as SMC4 in MM. Mapping these acquired SEs and their associated genes may provide novel insight into understanding MM biology, and SMC4 may serve as novel therapeutic targets in MM.

Citation Format: Yunlu Jia, Jianbiao Zhou, Tze-King Tan, Tae-Hoon Chung, Yongxia Chen, Enfan Zhang, Zhen Cai, Takaomi Sanda, Wee-Joo Chng. Superenhancer-drvien SMC4 promote cell growth and proliferation in multiple myeloma [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 2956.
Abstract 2957: Inhibition of epigenetic regulator menin is a novel therapeutic approach for high-risk neuroblastoma
Authors: Chilamakuri Rameswari , Agarwal Saurabh
Published: June 2022

Controlling epigenetic factors such as histone methyltransferases and demethylases has recently emerged as an attractive therapeutic approach for different cancers. KMT2 family protein KMT2A or MLL1 is a H3K4 methyltransferase, which is known to play an essential role in cancer development. MLL1 forms a COMPASS complex to function as an epigenetic regulator and Menin is one of the specific and required binding partner for the activation and function of this epigenetic enzyme. Menin acts as a tumor suppressor protein, and inhibition of Menin-MLL1 interaction has been shown to destabilize the MLL1 epigenetic complex, inhibits H3K4 methyltransferase activity, and hinder the oncogenic potential of various cancers. High-risk neuroblastoma (NB) is the most common extracranial solid pediatric tumor and accounts for almost 15% of all pediatric cancer-related deaths. Recent reports indicate that aberrant activation of epigenetic regulators may drive NB progression and relapse. Therefore, in the present study, we used a specific small-molecule inhibitor, MI-503 to disrupt Menin-MLL1 interaction, and to further understand the role of Menin in pediatric NB. We used different NB cell lines including MYCN-non-amplified (SH-SY5Y, SK-N-AS, CHLA-255) and MYCN-amplified (NGP, LAN-5, IMR-32), and performed cytotoxicity and Colonogenic assays to determine the effect of Menin inhibition on NB proliferation. Results showed that MI-503 significantly inhibits NB proliferation and colony formation capacity in a dose-dependent manner in contrast to controls. Further, we performed apoptosis and cell cycle assays and observed that MI-503 significantly induces apoptosis as detected by the increase of Annexin-V positive early apoptotic cells, and blocks cell cycle progression at the S phase in all NB cell lines tested. Additionally, Western blot assays demonstrated an overall inhibition of H3K3me3 levels in NB cells in response to MI-503. To further determine the effects of Menin inhibition on NB tumor growth, we developed a NB 3D spheroid tumor model that recapitulate in vivo tumor growth for solid tumors. Treatment with MI-503 significantly inhibits spheroidal tumor growth by inducing tumor cell death in a dose-dependent manner. Overall, these data highlight the role of epigenetic regulation, MLL1 complex, and of Menin in NB growth and tumorigenicity. In our future efforts, we will further elucidate the effects of MI-503 and the role of Menin in NB by using in vivo tumor models. Epigenetic inhibitor such as MI-503 holds significant potential for further clinical development and as a novel therapeutic approach for NB.

Citation Format: Rameswari Chilamakuri, Saurabh Agarwal. Inhibition of epigenetic regulator menin is a novel therapeutic approach for high-risk neuroblastoma [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 2957.
Abstract 2960: KAT6A and ENL form an epigenetic transcriptional control module to drive critical leukemogenic gene expression programs
Authors: Blanco Andres
Published: June 2022

Acute myeloid leukemia (AML) is a poor-prognosis hematological malignancy that is characterized by differentiation blockade and uncontrolled proliferation. Epigenetic programs influence AML cell fate decisions, and their mis-regulation can promote differentiation arrest. Itis therefore of paramount importance to identify the regulators of AML cell differentiation and understand the mechanisms by which they repress myeloid transcriptional programs. Here, we performed a cell-fate selection CRISPR screen for chromatin factors whose inhibition promotes AML cell differentiation. We found that the histone acetyltransferase KAT6A is a critical driver of the differentiation block, and that both genetic and chemical inhibition of KAT6A markedly de-repressed latent myeloid maturation programs, most commonly in MLL-rearranged AML. Inhibition of KAT6A catalytic activity reduced self-renewal, induced cell cycle arrest, and extinguished long-term proliferative capacity of AML cells. Through ChIP-seq, RNA-seq, and extensive analysis of clinical datasets, we found that KAT6A is the likely initiator of a newly-described "writer-reader" module that drives oncogenic gene expression in AML. In this model,KAT6A catalyzes promoter H3K9 acetylation, which is bound by the H3K9ac reader ENL, leading to Super elongation complex (SEC) recruitment and release of paused RNA PolII at oncogenic loci such as MYC. KAT6A is elevated in human AMLs compared to matched normal tissue, and its downregulation correlates with monocytic differentiation transcriptional programs in clinical AML datasets. These findings suggest the potential of targeting both KAT6A and ENL to disrupt this writer-reader module and ablate downstream MYC transcriptional programs inMLL-rearranged AML.

Citation Format: Andres Blanco. KAT6A and ENL form an epigenetic transcriptional control module to drive critical leukemogenic gene expression programs [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 2960.
Abstract 2984: ARHGAP17, a Cdc42-specific GAP, localizes to invadopodia and regulates their turnover as part of an ARHGAP17/Cdc42/CIP4 complex
Authors: Kreider-Letterman Gabriel , Castillo Abel , Rabino Agustin , Mahlandt Eike K. , Goedhart Joachim , Goicoechea Silvia , Garcia-Mata Rafael
Published: June 2022

One of the hallmarks of aggressive cancer is its ability to invade into surrounding tissues and metastasize. Many invasive cancers form invadopodia, which are protrusive, degradative structures that are regulated by the Rho family of GTPases. Rho GTPases commonly have aberrant activity in invasive cancers caused by changes in their upstream regulators: RhoGEFs and RhoGAPs. We have identified the RhoGAP ARHGAP17 (RICH1/NADRIN) as a negative regulator of invadopodia in triple negative breast cancer. Using confocal and STORM super-resolution microscopy techniques, we show that ARHGAP17 localizes to the regulatory ring surrounding invadopodia where it acts to promote disassembly by decreasing invadopodia lifetime through the inactivation of the Rho GTPase Cdc42. We further support these results in live cells by using a new localization-based Cdc42 biosensor, which reveals that ARHGAP17 suppresses Cdc42 activity specifically at the invadopodia ring. We have also identified the Cdc42 effector CIP4 as part of a complex with ARHGAP17 and Cdc42 that we propose is key in the regulation of invadopodia. Finally, using spheroid and inverse invasion assays, we show that the invadopodia phenotype correlates with increased invasive properties. In conclusion, we have identified ARHGAP17 as a key regulator of invadopodia and invasion in triple negative breast cancer by localizing to invadopodia and inactivating Cdc42 as part of a complex with CIP4. In future studies, we will further characterize the molecular mechanisms that regulate the ARHGAP17/Cdc42/CIP4 complex and their function during invadopodia formation.

Citation Format: Gabriel Kreider-Letterman, Abel Castillo, Agustin Rabino, Eike K. Mahlandt, Joachim Goedhart, Silvia Goicoechea, Rafael Garcia-Mata. ARHGAP17, a Cdc42-specific GAP, localizes to invadopodia and regulates their turnover as part of an ARHGAP17/Cdc42/CIP4 complex [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 2984.
Abstract 2996: Pre-activation of autophagy impacts PARP inhibitor sensitivity of prostate cancer cells
Authors: Cahuzac Maxime , Péant Benjamin , Fleury Hubert , Mes-Masson Anne-Marie , Saad Fred
Published: June 2022

Introduction: Advanced prostate cancer (APC) is one of the most common causes of cancer-related mortality in males largely related to the development of resistance to hormonal and chemotherapeutic based regimens. Recently, the PARP inhibitor (PARPi) olaparib was shown to improve progression-free and overall survival in patients with APC although resistance remains a barrier to cure. Therapeutic resistance in cancers can be mediated by autophagy, a cellular recycling process. Here we addressed the role of autophagy in the development of olaparib resistance in APC.

Methods: Autophagy levels and olaparib sensitivity were evaluated in prostate cancer (PC) cell lines (LNCaP, C4-2B and PC-3) by western blots and clonogenic assays, respectively. We introduced a double-tagged (mCherry-GFP) LC3 protein in cells to monitor autophagic flux by confocal microscopy and evaluated the temporal importance of autophagy activation by rapamycin on olaparib sensitivity. We measured cell proliferation (Incucyte), cell cycle (flow cytometry) and DNA repair (microscopy). Finally, we confirmed the role of autophagy in olaparib sensitivity using CRISPR/Cas9 Atg16L1 knockout cell lines.

Results: We found a correlation between the basal level of autophagy and olaparib sensitivity in PC cell lines. PC-3 cells were olaparib resistant and had a higher basal level of autophagy compared to LNCaP and C4-2B cells, which were PARPi sensitive. When autophagy was pre-activated before olaparib treatment, PC cell lines became more resistant by increasing homologous recombination activity. This autophagy-mediated resistance was partially regulated by a balance between SQSTM1 and FLNA protein levels in the nucleus. Inhibition of autophagy or post-activated autophagy prevented development of PARPi resistance.

Conclusion: The autophagic flux is associated with PARPi resistance in PC cell lines by regulating DNA repair. Inhibiting autophagy may lead to new strategies to increase PARPi efficacy. Understanding the role of autophagy in PC progression may lead to strategies that prevent the development of PARPi resistance.

Citation Format: Maxime Cahuzac, Benjamin Péant, Hubert Fleury, Anne-Marie Mes-Masson, Fred Saad. Pre-activation of autophagy impacts PARP inhibitor sensitivity of prostate 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 2996.
Abstract 3022: Tryptophan metabolism is associated with obesity and triple negative breast cancer risk in black and white women
Authors: Smith Alana , Gu Qingqing , Amos-Abanyie Ernestine Kubi , Tolley Elizabeth , Lu Lu , Lyn-Cook Beverly , Starlard-Davenport Athena
Published: June 2022

Non-Hispanic black (NHB) women are more likely to be diagnosed and die from triple negative breast cancer (TNBC) than non-Hispanic white (NHW) women. We previously showed that TNBC tissues display a distinct microbial signature. Obesity which contributes to chronic low-grade inflammation and gut microbial dysbiosis further exacerbates TNBC outcomes for NHB women. However, it is unclear the extent to which microbial differences in breast tumor tissues may be driven by alterations in microbial metabolites as a result of obesity status. Therefore, the aim of this global metabolomic profiling study was to investigate alterations in microbial metabolism pathways in breast tissues of NHB and NHW obese women (avg. BMI = 35.7) relative to non-obese women (avg. BMI = 23.4) with and without TNBC. In this study, we analyzed 984 metabolites derived from a total of 51 NHB and NHW women, including 15 patients with TNBC, 12 patients with less aggressive Luminal A-type breast cancer, and 24 healthy controls for comparison using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) to identify microbial metabolites and metabolomic signatures that differ by breast tumor subtype and obesity status. We detected 501 significantly increased metabolites and 33 significantly decreased metabolites in TNBC patients compared to controls. Principal component analysis and hierarchical clustering of the global metabolite profiling data revealed separation between the metabolic signatures of normal and breast tumor tissue. Random forest analysis revealed a unique biochemical signature associated with elevated L-Tryptophan (Trp) and Kynurenine (Kyn) metabolites and lower levels of microbial-derived metabolites critical for controlling inflammation and immune response in obese individuals and those with TNBC. TCGA analysis further revealed that expression of key Trp enzymes was significantly associated with worse survival outcomes in TNBC patients compared to other breast tumor subtypes. Our findings highlight a potential role for Trp metabolism through the Kyn pathway in obesity and TNBC risk. Further investigations into the Kyn metabolomic pathway may prove to be a novel therapeutic target for TNBC.

Citation Format: Alana Smith, Qingqing Gu, Ernestine Kubi Amos-Abanyie, Elizabeth Tolley, Lu Lu, Beverly Lyn-Cook, Athena Starlard-Davenport. Tryptophan metabolism is associated with obesity and triple negative breast cancer risk in black and white women [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 3022.
Abstract 3027: Elucidating the role of glutamine metabolism in head & neck squamous cell carcinoma
Authors: Allevato Michael , Yokoyama Yumi , Wild Robert , Gutkind J. Silvio
Published: June 2022

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide, resulting in over 14,600 deaths each year in the United States alone. HNSCC is associated with human papillomavirus (HPV) infection, tobacco use, and abusive alcohol intake. Without truly effective targeted therapies, surgery and radiotherapy represent the primary treatment options for most patients. Unfortunately, these treatments are associated with significant morbidity and a reduction in quality of life. Immunotherapies have recently revolutionized HNSCC treatment, but <20% of patients exhibit clinical responses, albeit often not durable (Saddawi-Konefka et al. Frontiers in Oncology 2021). This highlights the unmet need to identify novel therapeutic options and biomarkers predicting a more favorable response to maximize the efficacy of targeted cancer strategies for HNSCC treatment. Glutamine is a conditionally essential amino acid for rapidly proliferating cancer cells making glutamine pathway inhibition an attractive approach for anti-cancer therapy. We found that treatment with the broad glutamine antagonist sirpiglenastat (DRP-104), which irreversibly inhibits all known enzymes involved in glutamine metabolism, results in metabolically halted cell growth in a large panel (n=8) of HPV- and HPV+ HNSCC cell lines (IC50 of 0.2-25uM). Interestingly, HNSCC cells bearing genetic alterations in PIK3CA and PTEN were significantly more sensitive to glutamine antagonism than unaltered HNSCC cells. The dependence of glutamine in HNSCC growth and the increased sensitivity of PIK3CA/PTEN aberrant cells was also observed in orosphere assays and HNSCC tumor xenografts in mice in vivo. We next explored the mechanism of glutamine suppression in HNSCC by integrating the results from genome-wide CRISPR-Cas9 knockout library screens and broad-spectrum metabolomics analysis. Both approaches converged on the identification of a dysregulated metabolic pathway that represents a synthetic lethal vulnerability that can be exploited via targeted therapies. Our data suggest that broad glutamine antagonism using sirpiglenastat (DRP-104) has therapeutic potential in HNSCC by dismantling cancer metabolism and sensitizing cells to additional perturbations leading to specific cell death. A clinical trial of sirpiglenastat (DRP-104) is currently ongoing (NCT04471415).

Citation Format: Michael Allevato, Yumi Yokoyama, Robert Wild, J. Silvio Gutkind. Elucidating the role of glutamine metabolism in head & neck squamous cell carcinoma [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 3027.
Abstract 3033: Glucose deprivation promotes lung adenocarcinoma de-differentiation due to unbalanced EZH2 activity
Authors: Saggese Pasquale , Pandey Aparamita , Fung Eileen , Giurato Giorgio , Weisz Alessandro , Dubinett Steven M. , Scafoglio Claudio
Published: June 2022

Lung cancer is the most frequent cancer-related cause of death, and adenocarcinoma (LUAD) is the most frequent type. Despite the recent success of immunotherapies, survival of lung cancer patients has not significantly improved in the last decades. New therapies are necessary. We have previously identified sodium-glucose transporter 2 (SGLT2) as the major responsible for glucose uptake in LUAD, and we have showed that treatment with SGLT2 inhibitors significantly delays LUAD development and prolongs survival in murine models. However, our data shows that SGLT2 inhibitors also induce de-differentiation of LUAD cells, leading to a more aggressive phenotype and increased resistance to cisplatin. Glucose deprivation causes reduced αKG levels, leading to reduced activity of αKG-dependent histone demethylases and consequent histone hypermethylation. Supplementation of αKG or inhibition of the histone methyltransferase EZH2 reverse this phenotype, suggesting that this de-differentiated phenotype depends on insufficiency of αKG-dependent histone demethylases and unbalanced EZH2 activity. Consistently, double treatment with an SGLT2 inhibitor and an EZH2 inhibitor significantly reduces the tumor burden in a genetically engineered murine model of LUAD. We further characterized the effect of low glucose-induced tumor de-differentiation, identifying stabilization of hypoxia inducible factor 1α (HIF1α) as a major pathway responsible for the acquisition of a more aggressive phenotype following glucose deprivation. Finally, we identified an HIF1α-dependent transcriptional signature with prognostic significance in human LUAD. Our studies further our knowledge of the relationship between glucose metabolism and cell differentiation in cancer, characterizing the epigenetic adaptation of cancer cells to nutrient deprivation and identifying novel targets to prevent the development of resistance to metabolic therapies.

Citation Format: Pasquale Saggese, Aparamita Pandey, Eileen Fung, Giorgio Giurato, Alessandro Weisz, Steven M. Dubinett, Claudio Scafoglio. Glucose deprivation promotes lung adenocarcinoma de-differentiation due to unbalanced EZH2 activity [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 3033.
Abstract 3048: CSI-Microbes: Identifying cell-type specific intracellular microbes from single-cell RNA-seq data
Authors: Robinson Welles , Schischlik Fiorella , Gertz E. Michael , Lee Joo S. , Zhu Kaiyuan , Sahinalp S. Cenk , Patro Rob , Leiserson Mark D. , Schaffer Alejandro A. , Ruppin Eytan
Published: June 2022

Despite recent reports of microbes living within tumor cells, the identification of intracellular microbes remains an open and important challenge. Here we introduce CSI-Microbes (computational identification of Cell type Specific Intracellular Microbes), a computational approach for the discovery of cell-type specific intracellular microbial taxa from single-cell RNA sequencing of host cells. It is first validated on two gold-standard datasets of human immune cells exposed to Salmonella. Next, CSI-Microbes is tested on Merkel cell and colorectal carcinoma where it identifies both reported and previously unknown tumor-specific intracellular microbes. Finally, CSI-Microbes is applied to analyze the intracellular microbiome of thirteen lung tumors where it identifies four tumors with bacterial taxa enriched in tumor cells and two tumors with bacterial taxa enriched in stroma or immune cells. Notably, the infected tumor cells down-regulate pathways associated with anti-microbial response and antigen processing and presentation, testifying to the functional significance of bacterial presence.

Citation Format: Welles Robinson, Fiorella Schischlik, E. Michael Gertz, Joo S. Lee, Kaiyuan Zhu, S. Cenk Sahinalp, Rob Patro, Mark D. Leiserson, Alejandro A. Schaffer, Eytan Ruppin. CSI-Microbes: Identifying cell-type specific intracellular microbes from single-cell RNA-seq data [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 3048.
Abstract 3071: A reconstruction of evolutionary trajectories in primary tissue and PDTOs in a patient with metastatic osteosarcoma
Authors: Davarifar Ardy , Yanagawa Jane , Al Shihabi Ahmad , Nguyen Huyen T. , Liu Lydia Y. , Salcedo Adriana , Gonzalez Alfredo , Yamaguchi Takafumi N. , Bernthal Nicholas , Nelson Scott D. , Federman Noah , Boutros Paul C. , Soragni Alice
Published: June 2022

Metastatic osteosarcoma is a rare and highly lethal cancer that affects children and young adults. The rarity of osteosarcoma and its highly heterogeneous genetic composition have resulted in a lack of curative treatments for recurrent and metastatic disease.

In this study, we present the functional and molecular characterization of a case of a pediatric patient with left distal femur osteosarcoma and pulmonary metastases at presentation. Through longitudinal sampling over the course of the patient’s illness, we have procured tissue from the initial treatment-naïve diagnostic biopsy, primary tumor resection and multiple subsequent lung metastasectomies, and have developed patient-derived tumor organoids (PDTOs) from the viable samples. PDTOs are useful pre-clinical models that are representative of the source tissue and allow further interrogations of the biology of tumors. These models can also help with the identification of effective personalized therapies (Al Shihabi et al, 2021). We applied our mini-ring organoid screening technology (Phan et al, 2019; Nguyen et al, 2020) to perform high-throughput drug screening with different pharmacologic agents. We also performed deep whole-genome sequencing on all collected samples and corresponding organoids to reconstruct their evolutionary history, identify driver mutations and quantify heterogeneity in mutational processes across time and space.

Our preliminary data show that the subclonal content of the tumor and derivative organoids as well as the PDTO drug responses vary with the patient’s treatment history. We identify the genomic makeup of the subclones and posit changes that may be responsible for chemoresistance. Furthermore, our data shows correlations between the subclonal genomic composition of the PDTOs and source tissue, thereby reinforcing that PDTOs are capable of accurately recapitulating the subclonal heterogeneity of the parental tumor.

Citation Format: Ardy Davarifar, Jane Yanagawa, Ahmad Al Shihabi, Huyen T. Nguyen, Lydia Y. Liu, Adriana Salcedo, Alfredo Gonzalez, Takafumi N. Yamaguchi, Nicholas Bernthal, Scott D. Nelson, Noah Federman, Paul C. Boutros, Alice Soragni. A reconstruction of evolutionary trajectories in primary tissue and PDTOs in a patient with metastatic osteosarcoma [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 3071.
Abstract 3077: Leveraging a 3D organoid library to identify novel therapeutic targets during ESCC initiation and progression
Authors: Flashner Samuel , Shimonosono Masataka , Takada Satoshi , Matsuura Norihiro , Tomita Yasuto , Chen Xiao , Taylor Alison , Klein-Szanto Andres , Momen-Heravi Fatameh , Diehl J. Alan , Lu Chao , Nakagawa Hiroshi
Published: June 2022

Esophageal squamous cell carcinoma (ESCC) is a devastating and common disease worldwide. Despite recent advances annotating the genetic and epigenetic changes underlying the initiation and development of ESCC and its precursor lesion, esophageal squamous dysplasia, more work is needed to translate these findings into novel therapeutic strategies. This gap is due in part to the lack of tractable models of ESCC initiation and development that can then be manipulated ex vivo for preclinical testing. Currently, the large-scale studies characterizing the aforementioned molecular changes have used primary tissue for their analyses and have subsequently been unable to test the specific hypotheses that arise from their data using the original tissue that they profiled. To address this gap, we have generated a large (n = 54) three-dimensional organoid library from the esophagi of mice treated with the carcinogen 4-Nitroquinoline N-oxide (4-NQO) representing each stage of esophageal squamous cell initiation and progression from early intraepithelial dysplasia through lung metastasis. We have determined that these organoids faithfully recapitulate the molecular biology, histology, and oncogenic phenotype of their tissue of origin. Through integrative analysis of both RNA sequencing and whole exome sequencing data, we have identified several novel therapeutic targets that contribute to ESCC initiation and development including the DNA damage response and NOTCH1 signaling. Together, our work capitalizes on the intersection of large-scale multiomics and sophisticated 3D organoid models to identify novel therapeutic targets for the treatment of a devastating disease.

Citation Format: Samuel Flashner, Masataka Shimonosono, Satoshi Takada, Norihiro Matsuura, Yasuto Tomita, Xiao Chen, Alison Taylor, Andres Klein-Szanto, Fatameh Momen-Heravi, J. Alan Diehl, Chao Lu, Hiroshi Nakagawa. Leveraging a 3D organoid library to identify novel therapeutic targets during ESCC initiation and progression [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 3077.
Abstract 3081: High-speed live cell interferometry captures heterogeneity in bioprinted tumor organoids
Authors: Tebon Peyton J. , Wang Bowen , Markowitz Alexander L. , Murray Graeme , Davarifar Ardalan , Nguyen Huyen T. , Tavanaie Nasrin , Nguyen Thang L. , Boutros Paul C. , Teitell Michael A. , Soragni Alice
Published: June 2022

As interest grows in using tumor organoids as models for drug discovery and precision medicine, platforms for the reliable functional testing and analysis of these organoids are needed. Existing functional approaches in precision medicine face difficulties in the creation, scaling, and analysis of physiological tumor organoid models. We present a method for the automated bioprinting of tumor organoids coupled with real-time biomass quantification using high-speed live cell interferometry (HSLCI). This workflow enhances consistency, preserves sample viability, and assesses response to treatment with single-organoid resolution. First, we describe the bioprinting protocol used to achieve the deposition of organoids in uniformly thin three-dimensional layers of Matrigel. Then we show that bioprinting preserves the histological and molecular characteristics of manually seeded organoids. In drug screening experiments, we demonstrate the simultaneous monitoring of thousands of individual organoids and show that HSLCI can rapidly identify drug sensitivity and resistance to treatment. We observe significant heterogeneity in the drug responses of single organoids and discuss the potential implications for use of this pipeline in the clinical setting.

Citation Format: Peyton J. Tebon, Bowen Wang, Alexander L. Markowitz, Graeme Murray, Ardalan Davarifar, Huyen T. Nguyen, Nasrin Tavanaie, Thang L. Nguyen, Paul C. Boutros, Michael A. Teitell, Alice Soragni. High-speed live cell interferometry captures heterogeneity in bioprinted tumor organoids [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 3081.
Abstract 3086: Organoids standardized to a clinically validated drug response assay for truly predictive in vitro drug response profiling
Authors: Millard Melissa , Williams Natalie A. , Elrod Ashley K. , DesRochers Teresa M.
Published: June 2022

Unlike cell lines, organoids maintain most of the biological properties of the parental tissue from which the starting cells were isolated including the histology and gene expression. When organoids include clinical annotation, they become a useful, renewable tool for clinical correlation studies, but to be truly predictive the drug profiling assays utilized to screen organoid response must have measurable correlation with patient response. 3D Predict™ is a highly accurate assay that is 89% and 85% predictive of response in first-line ovarian cancer and high-grade gliomas (HGG) respectively. We have developed a panel of organoids that are clinically annotated, include correlative primary tissue 3D Predict™ drug response data, and have been assessed for the recapitulation of primary tissue histology and genomics. Additionally, our organoid models incorporate matched immune cells, a key component of the tumor microenvironment, making them an ideal model for immune-oncology studies. Here we present data on 15 available organoid models across HGG, breast, colorectal and bladder cancer. We have applied these models to drug response studies, including checkpoint inhibitors and shown correlation to primary patient response. The assurance of predictive capacity is unique to KIYATEC’s organoids and is significant because it avoids the pitfalls of comparing drug responses across non-concordant assay platforms while providing assurance that the models are reflective of individual patient response and outcomes.

Citation Format: Melissa Millard, Natalie A. Williams, Ashley K. Elrod, Teresa M. DesRochers. Organoids standardized to a clinically validated drug response assay for truly predictive in vitro drug response profiling [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 3086.
Abstract 3087: The identification and characterization of FOLFOX chemotherapy resistant cell lineages in colorectal cancer organoids
Authors: Morris Julia M. , Thorne Curtis A.
Published: June 2022

Resistance to chemotherapy drugs is a well-documented issue inhibiting the treatment of a wide variety of cancers. Colorectal cancer (CRC) is the third most common neoplasm worldwide and has the third highest mortality rate. However, the longtime gold-standard FOLFOX chemotherapy regimen, consisting of 5-FU and Oxaliplatin, results in only 70% survival rates in Stage 3 CRC patients, worse in Stage 4. More recent targeted therapies have marginally improved on this number, but the fact remains that CRC represents a highly heterogeneous neoplasm and resistance to standard chemotherapeutic regimens poses a significant barrier in treating this devastating disease.

There is an immediate need to better understand how this resistance arises. We hypothesize that a small number of cells with high phenotypic plasticity (HPP) are able to remain dormant during the initial chemotherapy treatment then reenter the cell cycle to cause recurrence and metastasis months or even years after treatment has concluded. Patient-derived organoids (PDOs) are a recently developed ex vivo model that allows for the study and extensive characterization of such HPP cell populations that may be found in patient tumors. We are developing and expanding various PDO populations to examine them genotypically and phenotypically on the single-cell level through RNA sequencing and high throughput imaging, respectively. By identifying these HPP cell populations, we then discover drugs that prevent the development of drug-resistant subpopulations. In the long term, we use our tumor heterogeneity profile to develop models that predict the responsiveness of tumors, including their drug-resistant HPP subpopulations, to well-documented and clinically used oncology drugs, streamlining treatment and improving survival outcomes.

Citation Format: Julia M. Morris, Curtis A. Thorne. The identification and characterization of FOLFOX chemotherapy resistant cell lineages in colorectal cancer organoids [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 3087.