bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 2007: Investigating how RIPK1-mediated mitophagy induces cell death in ECM-detached cancer cells
Authors: Cockfield Jordan Alexandria , Antony Ishan , Ekanayake Samantha , Schafer Zachary T. 
Published: July 2021

Abstract
Lead Contact Metastasis is responsible for 90% of deaths caused by cancer. For cancer cells to begin successful metastasis, they must detach from their extracellular matrix (ECM) and evade intrinsic cell death mechanisms. Detachment from the ECM is known to induce a caspase-dependent cell death program, termed anoikis, that cancer cells have been shown to circumvent. Nonetheless, overcoming caspase-dependent cell death alone is not sufficient to promote survival during ECM-detachment, but cancer cells must also address caspase-independent cell death programs. In our recent studies, we have identified RIPK1 as a critical mediator of ECM-detachment induced caspase-independent cell death via its capacity to promote mitophagy and subsequent accumulation of mitochondrial ROS. More recently, we have begun to examine how ROS levels are elevated as a consequence of mitophagy and how these ROS ultimately promote cell death. During mitophagy, autophagosomes containing mitochondria fuse with lysosomes containing a plethora of hydrolases that could facilitate cell death. As such, we sought to examine the relationship between RIPK1-mediated mitophagy and lysosomal membrane permeabilization (LMP) Using immunoblotting, we found that not only does ECM-detachment promote the expression of the lysosomal protease cathepsin B, but also that this induction is dependent upon RIPK1 activity. Given that mitochondrial ROS (mitoROS) was shown to be a consequence of RIPK1-mediated mitophagy during ECM-detachment, we investigated if mitoROS were required for the observed changes in cathepsin B expression. Indeed, when treated with a mitochondrial ROS (mitoROS) scavenger, cathepsin B protein levels noticeably reduce. Taken together, this data suggests that RIPK1-driven mitoROS production is playing a role in regulating cathepsin B expression during ECM-detachment. Future studies will be aimed at determining whether these RIPK1-mediated alterations in cathepsin B are linked to the induction of caspase-independent cell death.
Citation Format: Jordan Alexandria Cockfield, Ishan Antony, Samantha Ekanayake, Zachary T. Schafer. Investigating how RIPK1-mediated mitophagy induces cell death in ECM-detached cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2007.
Abstract 2010: Pancreatic cancer stem cell exosomes drive a hierarchical intratumor communication network that fuels disease progression through hippo pathway inhibition
Authors: Ruivo Carolina F. , Bastos Nuno , Batista Ines , Adem Barbara , Duraes Cecilia , Melo Carlos A. , Campos-Laborie Francisco J. , Moutinho-Ribeiro Pedro , Mourão Barbara , Cravo Marília , Macedo Guilherme , Carneiro Fátima , Kouzarides Tony , Kalluri Raghu , Machado Jose C. , Melo Sonia 
Published: July 2021

Abstract
Intra-tumor heterogeneity represents a major challenge for cancer treatment. Resistance to therapy and tumorigenic ability varies greatly between distinct subpopulations of cancer cells. Intercellular communication is critical to establish a cooperative environment that maintains tumor heterogeneity and potentiates disease progression. However, how subpopulations of cancer cells interact to support tumor progression is not fully understood. Exosomes have the potential to re-educate recipient cells through the delivery of their cargo, emerging as the main mediators of intercellular communication. Here we demonstrate that subpopulations of pancreatic cancer cells use exosomes to establish a hierarchical and dynamic communication network, the ExoNet. Exosomes from cancer stem cells (CSC) are preferentially taken up by non-CSC (NCSC). Targeted inhibition of CSC exosomes in PDAC orthotopic, genetically engineered and patient-derived xenograft mouse models is sufficient to impair tumor growth and metastasis establishment. A comprehensive comparative proteomic analysis of exosomes derived from distinct subpopulations of pancreatic ductal adenocarcinoma (PDAC) human cell lines identified a significant enrichment of Agrin in CSC exosomes. Blocking exosomes secretion and downregulating Agrin in CSCs significantly diminishes YAP nuclear localization in receiving cells, indicating that CSC exosomes contribute for Hippo pathway inactivation. Agrin+ circulating exosomes correlate with tumor burden in PDAC patients, and most importantly correlate with CD133+ circulating exosomes, indicating that circulating Agrin+ exosomes are from CSC origin. Longitudinal analysis of circulating Agrin+ exosomes in PDAC patients predicts disease progression and response to therapy. Our study describes a novel role of cancer exosomes in the interplay between subpopulations of cancer cells promoting tumor progression. We identified Agrin as a potential non-invasive biomarker to monitor disease progression and predict treatment response.
Citation Format: Carolina F. Ruivo, Nuno Bastos, Ines Batista, Barbara Adem, Cecilia Duraes, Carlos A. Melo, Francisco J. Campos-Laborie, Pedro Moutinho-Ribeiro, Barbara Mourão, Marília Cravo, Guilherme Macedo, Fátima Carneiro, Tony Kouzarides, Raghu Kalluri, Jose C. Machado, Sonia Melo. Pancreatic cancer stem cell exosomes drive a hierarchical intratumor communication network that fuels disease progression through hippo pathway inhibition [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 2010.
Abstract 2027: SETD2 loss in renal carcinoma cells induces the unfolded protein response
Authors: Metz Alexander , Uzzo Robert , Abbosh Philip 
Published: July 2021

Abstract
Introduction: SETD2 encodes a histone H3-K36 methyltransferase which is frequently inactivated in clear cell renal carcinomas (ccRCCs) and papillary RCCs via 3p deletion/LOH and deleterious mutations. Histone H3-K36 trimethylation is facilitated by SETD2, which is necessary for proper pre-mRNA intron splicing. Improperly spliced mature-mRNA may lead to aberrant translation of retained introns (ATaRI), which represents potential therapeutic vulnerabilities. We explored this hypothesis using real world data and RCC models.
Methods and Results: Gene set enrichment analysis comparing SETD2-mutant to WT tumors using samples from the TCGA KIRC data set revealed that the untranslated protein response (UPR) was strongly enriched, as well as several immunotherapy-relevant pathways. This suggested that peptides arising from ATaRI may be present, since they would not be expected to fold properly and thus need to be addressed by the UPR pathway to maintain homeostasis. To investigate this further, we generated Setd2-isogenic RENCA cells using CRISPR. Knockout was confirmed by sequencing and immunoblot. H3K36 trimethylation was decreased or eliminated in monoclonal knockout cell lines, confirming a functional effect. Markers of UPR activation, including cleaved Atf6 and Atf4, were found to be upregulated in Setd2-mutant RENCA cells compared to controls as measured by immunoblot. Cleaved ATF6 is known to translocate to the nucleus to take part in a UPR transcriptional program. Consistent with this, ATF6 was found to localize to the nucleus in Setd2-knockout cells using immunofluorescence. Interestingly, CHOP, another downstream effector of the UPR pathway which predominantly regulates cell death, did not become upregulated in Setd2-knockout cells, suggesting activation of a compensatory cell survival pathway.
Conclusions: We identify activation of the UPR upon Setd2 loss and suggest that activation of part of the UPR pathway may represent a new therapeutic vulnerability for exploitation as a rationale for personalized medicine. We continue to evaluate the generation of peptides arising from ATaRI in Setd2-mutant contexts.
Citation Format: Alexander Metz, Robert Uzzo, Philip Abbosh. SETD2 loss in renal carcinoma cells induces the unfolded protein response [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 2027.
Abstract 2039: The role of HORMAD1 in DNA damage repair in squamous cell carcinomas
Authors: Gantchev Jennifer , Villarreal Amelia Martinez , Ramchatesingh Brandon , Litvinov Ivan V. 
Published: July 2021

Abstract
Genomic instability is a prominent hallmark of cancer, however the dynamic mechanisms that drive and sustain genomic instability remain elusive. Research demonstrates that numerous cancers with increased levels of genomic instability express meiosis genes and undergo a process called meiomitosis. Cancer meiomitosis is the orchestrated activation of both mitotic and meiotic machineries in neoplastic cells that confer a selective advantage. Specifically, meiCT (meiosis-specific cancer/testis) genes that specialize in the reductional division of germ cells in meiosis I are ectopically expressed in several cancers, particularly in squamous cell carcinomas. The meiCT gene, HORMAD1 is a meiosis specific protein that functions to ensure that a sufficient number of DSBs are formed for appropriate meiotic progression and to maintain genome integrity throughout homologous recombination. Studies show that HORMAD1 is significantly upregulated in several cancers and is regarded as a potentially important oncogene that plays a role in sustaining increased genomic instability. With the use of shRNA mediated knockdown of HORMAD1, we evaluated the effects ofHORMAD1 on genomic instability and survival in squamous cell carcinomas (SCCs). Our results demonstrate that HORMAD1 knockdown in SCC cancer cell lines leads to an increase in genomic instability denoted by an increase in double strand breaks, micronuclei formation and ploidy. Furthermore, shHORMAD1 results in increased sensitivity to etoposide treatment leading to increased centrosome formation and reduced cell survival. We also demonstrate thatHORMAD1 modulates a functional and balanced level of genomic instability by eliciting an DNA damage response through homologous recombination mechanisms that allows SCCs to thrive following DNA damage.
Citation Format: Jennifer Gantchev, Amelia Martinez Villarreal, Brandon Ramchatesingh, Ivan V. Litvinov. The role of HORMAD1 in DNA damage repair in squamous cell carcinomas [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 2039.
Abstract 204: Identification of gene expression signatures as potential novel biomarkers in malignant melanoma
Authors: Figueroa Stephanie , Tiwari Raj , Geliebter Jan 
Published: July 2021

Abstract
Melanoma accounts for the vast majority of skin cancer-related deaths due to its ability to metastasize and therapeutic resistance. We aimed to identify expression-based signatures associated with melanoma patient survival by analyzing The Cancer Genome Atlas (TCGA) database to identify potential, prognostic biomarkers of cutaneous melanoma. Genes overexpressed in melanoma, compared with normal skin, as well as genes associated with the survival of melanoma patients were found using the online bioinformatics platform GEPIA2 (Gene Expression Profiling Interactive Analysis). The Metascape bioinformatics tool was then used for clustering of the genes based on processes and Protein-Protein Interaction (PPI) Enrichment Analysis. The genes from the 6 most significantly enriched pathways were selected to create gene signatures, which were then subjected to Kaplan-Meier survival analysis. Melanoma patients from the TCGA database (458) were placed into two groups (229 patients each): a high and a low expression group. There were 139 statistically significantly upregulated genes in melanoma (p&lt;0.01), while 500 genes had expression levels that were significantly associated with the survival of melanoma patients. Overexpressed genes were found to belong to pigment-related pathways (pigmentation, cellular pigmentation, and pigment cell differentiation), while survival-associated genes were found to function in immune system-related pathways (adaptive immune response, lymphocyte activation, and immune response-regulating signaling pathways). Kaplan-Meier survival analysis showed that the most overexpressed genes in melanoma correlated with poorer survival, while higher expression of the most-significantly survival associated gene signatures correlated with better survival. Together, these gene signatures may serve as predictors of survival in melanoma patients.
Citation Format: Stephanie Figueroa, Raj Tiwari, Jan Geliebter. Identification of gene expression signatures as potential novel biomarkers in malignant melanoma [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 204.
Abstract 2049: Cell-cycle dependent regulation of Sp1 at DSBs promotes repair via NHEJ and sensitizes cells to PARP inhibition
Authors: Swift Michelle , Azizkhan-Clifford Jane 
Published: July 2021

Abstract
DNA double-strand breaks (DSBs) are life-threatening lesions that must be repaired to preserve chromosomal integrity. These lesions are repaired by two primary pathways: homologous recombination (HR) and non-homologous end-joining (NHEJ). The factors that mediate pathway choice are critical to maintain genomic integrity. While many of these factors, epigenetic changes and cell cycle stages are known, with HR predominating in S and G2, the underlying mechanisms that determine pathway choice are not well understood. Previously, we found that the transcription factor Sp1 is rapidly recruited to DSBs and is necessary for repair. Here, we demonstrate that Sp1 integrates repair pathway choice with cell cycle by promoting NHEJ in G1. Sp1 localizes to DSBs in G1 to recruit the NHEJ repair factors 53BP1 and Ku70. Sp1 is phosphorylated by Cyclin A/cdk2 upon entry into S phase, evicting both Sp1 and 53BP1 from the break site, thereby permitting BRCA1 binding and HR. Mutation of this phosphorylation site (Sp1-S59A) results in persistence of Sp1 and 53BP1 at DSBs in S phase cells, precluding BRCA1 binding and HR. We also demonstrate that Sp1 is sumoylated in S phase in response to damage. This sumoylation in recognized by the E3-ubiquitin ligase, RNF4, and therefore polyubiquitylates Sp1 and targets it for proteasomal degradation. Expression of a sumoylation deficient Sp1 (Sp1-K16R) results in Sp1 and 53BP1 occupancy at DSBs in S phase, inhibiting BRCA1 binding to DSBs and HR. HR proteins are commonly mutated in cancers, resulting in cellular “addiction” to alternative repair pathways. Exploiting this addiction is the basis of synthetic lethality and is applicable to the treatment of many types of malignancies. Similar to BRCA1 deficiency, expression of Sp1-S59A or Sp1-K16R also increases cell sensitivity to PARP inhibition in BRCA1+/+ cells because of the inability to recruit BRCA1. Alternatively, synthetic lethality in BRCA1-/- cells requires 53BP1, whereas loss of 53BP1 confers PARPi resistance in a BRCA1 deficient cell. Consistent with this, depletion of Sp1 also results in PARPi resistance in BRCA1-/- cells. Interestingly, Sp1 is overexpressed in many tumors and is associated with poor prognosis. These data corroborate our finding that Sp1 is required for NHEJ and provide preliminary evidence that Sp1 may be a biomarker for PARPi sensitivity.
Citation Format: Michelle Swift, Jane Azizkhan-Clifford. Cell-cycle dependent regulation of Sp1 at DSBs promotes repair via NHEJ and sensitizes cells to PARP inhibition [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 2049.
Abstract 2052: TRIP13 amplification confers PARP inhibitor resistance and polymerase theta inhibitor sensitivity
Authors: Patterson-Fortin Jeffrey , Zhou Jia , D'Andrea Alan 
Published: July 2021

Abstract
Genome integrity is critical for cellular survival and is maintained by DNA repair pathways collectively known as the DNA damage response (DDR). Double-stranded breaks (DSBs) are the most deleterious form of DNA damage and can be repaired through homologous recombination (HR), non-homologous end-joining (NHEJ), and microhomology-mediated end-joining (MMEJ). The first step in both HR and MMEJ is resection of the DNA DSB termini to create single-stranded DNA 3' overhangs. In contrast, the 53BP1-Shieldin complex limits DNA end-resection and promotes NHEJ. Defects in HR repair can be present in cancer cells secondary to germline or somatic mutations in BRCA1, BRCA2, or other genes in the HR pathway, and such defects confer sensitivity to inhibitors of poly (ADP-ribose) polymerase (PARP). Resistance to PARP inhibitors can occur if there is loss of the 53BP1-Shieldin complex or amplification of TRIP13, a negative regulator of the 53BP1-Shieldin complex (Clairmont, et al. NCB 22:87, 2020). Here, we show that cells with TRIP13 amplification have increased DNA end-resection and increased expression of polymerase theta (POLQ), the critical enzyme for MMEJ. The resulting increase in POLQ activity limits DNA end-resection and increases MMEJ-mediated DSB repair. In these cells, inhibition of POLQ by a POLQ inhibitor, novobiocin (Zhou, et al. bioRxiv, 2020), resulted in excessive DNA end-resection, replication stress, activation of the ataxia-telangiectasia and Rad3 related (ATR) DNA damage response, and ultimately cell death. Conversely, knockdown of TRIP13 promotes PARP inhibitor sensitivity and POLQ inhibitor resistance. Thus, TRIP13 amplification may function as a biomarker for resistance to PARP inhibitors and sensitivity to a POLQ inhibitor. In summary, these results reveal a mechanism of PARP inhibitor resistance and POLQ inhibitor sensitivity in HR-deficient cells, provide a rationale for using TRIP13 as a biomarker for PARP inhibitor or POLQ inhibitor sensitivity, and may offer novel therapeutic strategies for PARP inhibitor resistant cancers.
Citation Format: Jeffrey Patterson-Fortin, Jia Zhou, Alan D'Andrea. TRIP13 amplification confers PARP inhibitor resistance and polymerase theta inhibitor sensitivity [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 2052.
Abstract 2069: Targeting the DNA damage response in combination with high Z metal nanoparticle radiosensitizers in glioblastoma
Authors: Kerr Brianna 
Published: July 2021

Abstract
Introduction: Glioblastoma (GBM) is a devastating disease with an extremely poor prognosis of approximately 11-15 months following standard-of-care treatments. There is a critical need for the development of clinical unmet need and a need for novel therapies in GBM. Combining novel drugs with radiotherapy has clear potential to considerably improve patient outcomes. Metal nanoparticles (NPs) and inhibitors of the DNA damage response (DDRIs) have been successfully translated to the clinical setting to benefit patients. In this study, we aimed to evaluate a novel multi-modal treatment in GBM by combining radiotherapy with metal NPs and DDRIs.
Materials and methods: GBM cell models (U87 & U251) were screened with different NP formulations and sizes, including gadolinium-, iron- and gold-based NPs, to determine the most effective NP. Cells were transiently exposed to NPs for 1 or 24 hr prior to irradiation by x-rays and cell survival determined. IC50 values were determined for various DDRIs which specifically target PARP, ATR, ATM and DNA-PKcs. Clonogenic cell survival assays were conducted and cells treated with NP and DDRI combinations either 1 or 24 hr prior to irradiation to define the optimum schedule for drugging.
Results and conclusions: We have demonstrated the radiosensitizing potential of Activation and Guidance of Irradiation by X-ray (AGuIX), a gadolinium-based NP, at low-moderate concentrations with the highest concentration tested (2mM) showing the greatest efficacy. A promising lack of cytotoxicity on both GBM cell models was observed. In contrast to AGuIX, U87 and U251 cell survival responses indicated a lack of dose dependency when exposed to varying concentrations of Superparamagnetic Iron Oxide Nanoparticles (SPIONs). Both cell models exhibited differential radiotherapy responses when exposed to ID10 and ID11, gold-based NPs with varying PEG chain lengths. Combination treatment with AGuIX and Olaparib resulted in differential efficacy between cell lines. A non-significant difference in cell survival was observed between differing treatment scheduling. Future work will explore several variations of NP and DDRI treatments in combination with irradiation by x-rays on GBM cell models. The future implications of this research are GBM patients would receive combination therapies tailored to the specific DNA mutations in their tumor.
Citation Format: Brianna Kerr. Targeting the DNA damage response in combination with high Z metal nanoparticle radiosensitizers in glioblastoma [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 2069.