Alkylating Agent-Induced ER Stress Overcomes Microenvironmental Resistance to Lymphoma Therapy
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ABSTRACT:
PROVIDER: PRJNA528647 | ENA |
REPOSITORIES: ENA
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Project description:Compartment-specific resistance to cancer therapy remains poorly understood. We utilized orthotopic xenografts of human double-hit lymphoma (DHL) to interrogate responses across involved sites. We identified resistance to multiple chemotherapies and the anti-CD52 antibody Alemtuzumab within the bone marrow (BM) that depended on extensive lymphoma involvement and impaired antibody-dependent cellular phagocytosis. This resistance was overcome by high doses of alkylating agents, including cyclophosphamide (CTX), which exhibited >80-fold in vivo synergy with Alemtuzumab. CTX induced ER stress in BM DHL cells leading to ATF4- mediated paracrine secretion of VEGF-A and massive macrophage infiltration. Macrophages from DHL-engrafted, CTX-treated mice had increased phagocytic capacity for lymphoma cells that was reversed by VEGF-A blockade and required SYK phosphorylation. A subset of these macrophages, defined by surface CD36/FcgRIV and a distinct transcriptional state, were “super- phagocytic.” Together, our findings define a unique mechanism through which high-dose alkylating agents can overcome therapy-resistant niches by ER stress-induced activation of phagocytosis.
2020-03-01 | GSE128728 | GEO
Project description:We are investigating the transcriptional response of mice that are wt or null for Aag in response to alkylating agent MMS; We used microarrays to detail the global programme of gene expression response due to MMS exposure in a DNA repair proficient or deficient mouse Experiment Overall Design: Mice (WT and Aag null) were treated with MMS and livers extracted at 6, 12, 24, and 48 hours
2009-02-21 | E-GEOD-11252 | biostudies-arrayexpress
Project description:Understanding the mechanisms by which cells respond to chemotherapeutics is key to identifying means to improve therapy effiicacy while reducing systemic toxicity of these widely used classes of drugs. While determining the role of NRF2-GSH and ER stress in cells exposed to alkylating compounds such as methyl-methanesulfonate (MMS), we asked if these pathways could also be a general cell damage response relevant to other clinically used chemotherapeutics or if it is an alkylation specific response. With this intent, we performed RNA sequencing of MDA-MB231 breast cancer and U2OS osteosarcoma cells lines treated for 8 hours with a topoisomerase II inhibitor etoposide (20 µM), the antimitotic beta-tubulin-interacting drug paclitaxel (0.2 µM), doxorubicin (1 µM) and compared to MMS (40 µg/mL) treated cells. Doses represent IC50 level after 72 hours exposure. We observed that even though non-alkylating drugs, especially etoposide, caused an increase in the mRNA expression of some NRF2 and ER stress signaling markers, the number and magnitude of upregulation of genes markers in either pathway was more pronounced in alkylation treatments compared to other drugs. This indicates that alterations in NRF2 and ER stress pathways could be more likely associated with differential sensitivity to alkylating chemotherapies.
2017-02-13 | GSE84863 | GEO
Project description:Iodoacetamide is by far the most commonly used agent for alkylation of cysteine during sample preparation for proteomics. An alternative, 2-chloroacetamide, has been recently suggested to reduce the alkylation of residues other than cysteine, such as the N-terminus, Asp, Glu, Lys, Ser, Thr and Tyr. Here, we show that although 2-chloroacetamide reduces the level of off-target alkylation, it exhibits a range of adverse effects. The most significant of these was methionine oxidation, which increases to a maximum of 40% of all Met containing peptides compared to 2-5% with iodoacetamide. Increases were also observed for mono and dioxidised tryptophan. No additional differences between the alkylating reagents were observed for a range of other post-translational modifications and digestion parameters. The adverse impact of 2-chloroacetamide on methionine oxidation suggest it is not the ideal alkylating reagent for proteomics.
2017-08-16 | PXD007071 | Pride
Project description:Pyrrole-imidazole polyamides are versatile DNA minor groove binders and attractive therapeutic options against oncological targets, especially upon functionalization with an alkylating agent such as endo-seco-CBI. These molecules also provide an alternative for oncogenes deemed “undruggable” at the protein level, where the absence of solvent-accessible pockets or structural crevices prevent the formation of protein-inhibitor ligands; nevertheless, the genome-wide effect of pyrrole-imidazole polyamide binding remain largely unclear to-date. Here we propose a next-generation sequencing-based workflow combined with whole genome expression arrays to address such issue using a candidate anti-cancer alkylating agent, KR12, against codon 12 mutant KRAS. Biotinylating KR12 enables the means to identify its genome-wide effects in living cells and possible biological implications via a coupled workflow of enrichment-based sequencing and expression microarrays. The subsequent computational pathway and expression analyses allow the identification of its genomic binding sites, as well as a route to explore a polyamide’s possible genome-wide effects. Among the 3,343 KR12 binding sites identified in the human LS180 colorectal cancer genome, the reduction of KR12-bound gene expressions was also observed. Additionally, the coupled microarray-sequencing analysis also revealed some insights about the effect of local chromatin structure on pyrrole-imidazole polyamide, which had not been fully understood to-date. A comparative analysis with KR12 in a different human colorectal cancer genome SW480 also showed agreeable agreements of KR12 binding affecting gene expressions. Combination of these analyses thus suggested the possibility of applying this approach to other pyrrole-imidazole polyamides to reveal further biological details about the effect of polyamide binding in a genome.
2016-11-01 | GSE86598 | GEO
Project description:Pyrrole-imidazole polyamides are versatile DNA minor groove binders and attractive therapeutic options against oncological targets, especially upon functionalization with an alkylating agent such as endo-seco-CBI. These molecules also provide an alternative for oncogenes deemed “undruggable” at the protein level, where the absence of solvent-accessible pockets or structural crevices prevent the formation of protein-inhibitor ligands; nevertheless, the genome-wide effect of pyrrole-imidazole polyamide binding remain largely unclear to-date. Here we propose a next-generation sequencing-based workflow combined with whole genome expression arrays to address such issue using a candidate anti-cancer alkylating agent, KR12, against codon 12 mutant KRAS. Biotinylating KR12 enables the means to identify its genome-wide effects in living cells and possible biological implications via a coupled workflow of enrichment-based sequencing and expression microarrays. The subsequent computational pathway and expression analyses allow the identification of its genomic binding sites, as well as a route to explore a polyamide’s possible genome-wide effects. Among the 3,343 KR12 binding sites identified in the human LS180 colorectal cancer genome, the reduction of KR12-bound gene expressions was also observed. Additionally, the coupled microarray-sequencing analysis also revealed some insights about the effect of local chromatin structure on pyrrole-imidazole polyamide, which had not been fully understood to-date. A comparative analysis with KR12 in a different human colorectal cancer genome SW480 also showed agreeable agreements of KR12 binding affecting gene expressions. Combination of these analyses thus suggested the possibility of applying this approach to other pyrrole-imidazole polyamides to reveal further biological details about the effect of polyamide binding in a genome.
2016-11-01 | GSE86597 | GEO
Project description:Purpose: T-cell exhaustion limits immunotherapy for the treatment of solid tumors. While immune checkpoint blockade and adoptive T cell therapy (ACT) can mediate tumor regression, their potency is often determined by tumor burden. Here, we identified tumor burden-related pathway changes that are conducive to T-cell exhaustion. We then determined if microenvironmental reprogramming via epigenetic modulation could reverse T-cell exhaustion and improve immunotherapeutic responsiveness. Experimental Design: We developed a murine syngeneic tumor model wherein an increased burden ablated therapeutic responsiveness to ACT, which corresponded with systemic induction of T cell exhaustion. Transcriptome analysis of these large tumors allowed us to characterize changes to immunosuppressive pathway expression during class I histone deacetylase inhibitor (HDACi) MS-275 treatment. We then measured the therapeutic impact of MS-275 during ACT and assessed T-cell exhaustion by transcriptome/phenotypic analysis. Results: ACT durably regressed small tumors but failed to control large tumors, which were associated with systemic T-cell exhaustion and ablation of T-cell responses. Large tumors were defined by an immunosuppressive pathway signature. MS-275 reversed this pathway signature and promoted durable regression of large tumors during ACT. Prototypical exhaustion marker Tim-3 was selectively up-regulated in transferred T cells despite displaying a reduced exhaustion signature. Instead, we observed enhanced activation-dependent signaling correlating with enrichment of the IL2-STAT5 signaling axis. Activated CD8+ T-cell responses were predominantly skewed towards terminal effector cell-like CD44+ Tim-3hi TCF1- CD127- KLRG1+ differentiation. Conclusion: Tumor burden-induced pathway changes can be reversed through epigenetic reprogramming, enabling the conversion from T-cell exhaustion to effector lineage differentiation. We used whole transcriptome arrays (Clariom D) to determine the effect of MS-275 on exhausted T cells in ACT+/-MS-275-treated mice
2023-07-29 | GSE238222 | GEO
Project description:Purpose: T-cell exhaustion limits immunotherapy for the treatment of solid tumors. While immune checkpoint blockade and adoptive T cell therapy (ACT) can mediate tumor regression, their potency is often determined by tumor burden. Here, we identified tumor burden-related pathway changes that are conducive to T-cell exhaustion. We then determined if microenvironmental reprogramming via epigenetic modulation could reverse T-cell exhaustion and improve immunotherapeutic responsiveness. Experimental Design: We developed a murine syngeneic tumor model wherein an increased burden ablated therapeutic responsiveness to ACT, which corresponded with systemic induction of T cell exhaustion. Transcriptome analysis of these large tumors allowed us to characterize changes to immunosuppressive pathway expression during class I histone deacetylase inhibitor (HDACi) MS-275 treatment. We then measured the therapeutic impact of MS-275 during ACT and assessed T-cell exhaustion by transcriptome/phenotypic analysis. Results: ACT durably regressed small tumors but failed to control large tumors, which were associated with systemic T-cell exhaustion and ablation of T-cell responses. Large tumors were defined by an immunosuppressive pathway signature. MS-275 reversed this pathway signature and promoted durable regression of large tumors during ACT. Prototypical exhaustion marker Tim-3 was selectively up-regulated in transferred T cells despite displaying a reduced exhaustion signature. Instead, we observed enhanced activation-dependent signaling correlating with enrichment of the IL2-STAT5 signaling axis. Activated CD8+ T-cell responses were predominantly skewed towards terminal effector cell-like CD44+ Tim-3hi TCF1- CD127- KLRG1+ differentiation. Conclusion: Tumor burden-induced pathway changes can be reversed through epigenetic reprogramming, enabling the conversion from T-cell exhaustion to effector lineage differentiation.
2023-07-29 | GSE238233 | GEO
Project description:Alkylating agents are a frontline therapy for the treatment of several cancers including pediatric glioblastoma, a devastating lethal tumor in children. Unfortunately, many tumors are resistant to this therapy. We sought to identify ways of sensitizing tumor cells to alkylating agents while leaving normal cells unharmed; increasing therapeutic response while minimizing toxicity. Here we report using a siRNA screen targeting over 240 DNA damage response genes identified novel sensitizers to alkylating agents. In particular the base excision repair (BER) pathway, including 3-methylpurine-DNA glycosylase (MPG), as well as ataxia telangiectasia mutated (ATM) were identified in our screen. Interestingly, we identified MPG as a direct novel substrate of ATM. ATM-mediated phosphorylation of MPG was required for enhanced MPG function. Importantly, combined inhibition or loss of MPG and ATM resulted in increased alkylating agent-induced cytotoxicity in vitro and prolonged survival in vivo. The discovery of the ATM-MPG axis will lead to improved treatment of alkylating agent-resistant tumors.
2014-07-24 | GSE59678 | GEO
Project description:Alkylating agents are a frontline therapy for the treatment of several cancers including pediatric glioblastoma, a devastating lethal tumor in children. Unfortunately, many tumors are resistant to this therapy. We sought to identify ways of sensitizing tumor cells to alkylating agents while leaving normal cells unharmed; increasing therapeutic response while minimizing toxicity. Here we report using a siRNA screen targeting over 240 DNA damage response genes identified novel sensitizers to alkylating agents. In particular the base excision repair (BER) pathway, including 3-methylpurine-DNA glycosylase (MPG), as well as ataxia telangiectasia mutated (ATM) were identified in our screen. Interestingly, we identified MPG as a direct novel substrate of ATM. ATM-mediated phosphorylation of MPG was required for enhanced MPG function. Importantly, combined inhibition or loss of MPG and ATM resulted in increased alkylating agent-induced cytotoxicity in vitro and prolonged survival in vivo. The discovery of the ATM-MPG axis will lead to improved treatment of alkylating agent-resistant tumors. Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from snap frozen biopsy and surgical brain tissue from paediatric high grade gliomas Copy number analysis on Affymetrix SNP 250K arrays was performed for 35 paediatric high grade glioma samples and HapMap samples.
2014-07-24 | E-GEOD-59678 | biostudies-arrayexpress