Ferroptosis is an iron-dependent cell death mechanism characterized by the accumulation of toxic lipid peroxides and cell membrane rupture. GPX4 (glutathione peroxidase 4) prevents ferroptosis by reducing these lipid peroxides into lipid alcohols. Ferroptosis induction by GPX4 inhibition has emerged as a vulnerability of cancer cells, highlighting the need to identify ferroptosis regulators that may be exploited therapeutically. Through genome-wide CRISPR activation screens, we identify the SWI/SNF (switch/sucrose non-fermentable) ATPases BRM (SMARCA2) and BRG1 (SMARCA4) as ferroptosis suppressors. Mechanistically, they bind to and increase chromatin accessibility at NRF2 target loci, thus boosting NRF2 transcriptional output to counter lipid peroxidation and confer resistance to GPX4 inhibition. We further demonstrate that the BRM/BRG1 ferroptosis connection can be leveraged to enhance the paralog dependency of BRG1 mutant cancer cells on BRM. Our data reveal ferroptosis induction as a potential avenue for broadening the efficacy of BRM degraders/inhibitors and define a specific genetic context for exploiting GPX4 dependency.
<p>Figure S1. Consequences of KEAP1 mutations on Nrf2 expression and ubiquitination Figure S2-4. Inducible knockdown of Nrf2 in KEAP1 mutant and WT cells and tumors</p>
Abstract In response to diverse stimuli, tumor cells can undergo a process resembling the epithelial-mesenchymal transition (EMT) observed during development. During this transition, epithelial-like tumor cells acquire mesenchymal-like physiologic features, become more motile and invasive, and acquire insensitivity to many therapeutic agents. While the importance of this phenomenon to cancer treatment and patient outcomes is well known, considerable heterogeneity exists across experimental models of EMT, and diverse mechanisms have been proposed to explain its various associated malignant phenotypes. Consequently, the extent to which these processes are shared uniformly across instances of EMT, as well as their relative importance in treatment contexts, remains unclear. We have used RNAseq, ATACseq, and parallelized high-throughput cell viability screening to identify differences in gene expression levels, chromatin accessibility, and drug sensitivity that correlate with EMT in a series of cell line models. By contrasting these changes across different genetic backgrounds and mechanisms of EMT induction, we are able to identify sets of elements in the transcribed and noncoding genome that are characteristic of different modes of EMT induction. Then, by comparing these candidate sets with observed differences in acquired drug sensitivity in the respective models, we were able to define a set of genetic elements whose biologic activity is likely to influence the efficacy of these compounds. Finally, we used a series of targeted CRISPR-based knockout screens to assess the relative importance of these elements to the viability of isogenic epithelial-like and mesenchymal-like cells, alone and in the presence of multiple therapeutic drugs. Using this approach, we were able to validate known mechanisms of acquired drug resistance and identify new candidate effectors. Notably, we identify multiple candidate noncoding elements containing CTCF binding sites that appear to meaningfully influence cell viability in a drug-specific manner, potentially implicating genome conformational changes in acquired therapeutic resistance. Citation Format: Russell O. Bainer, Catherine Wilson, Marinella Callow, Siyu Feng, Michael Costa, Colin Watanabe, Oleg Mayba, Eva Lin, Scott Martin, Bob Yauch, Richard Bourgon, Christiaan Klijn. Functional coding and noncoding drivers of EMT-mediated acquired drug resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 398.
Abstract Xenotransplantation of porcine organs, tissues, and cells inherits a risk for xenozoonotic infections. Viable tissues and cells intended for transplantation have to be considered as potentially contaminated non‐sterile products. The demands on microbial testing, based on the regulatory requirements, are often challenging due to a restricted shelf life or the complexity of the product itself. In E urope, the regulatory framework for xenogeneic cell therapy is based on the advanced therapy medicinal products ( ATMP ) regulation (2007), the EMA CHMP Guideline on xenogeneic cell‐based medicinal products (2009), as well as the WHO and Council of Europe recommendations. In the USA , FDA guidance for industry (2003) regulates the use of xenotransplants. To comply with the regulations, validated test methods need to be established that reveal the microbial status of a transplant within its given shelf life, complemented by strictly defined action alert limits and supported by breeding in specific pathogen‐free ( SPF ) facilities. In this review, we focus on assays for the detection of the porcine endogenous retroviruses PERV ‐A/‐B/‐C, which exhibit highly polymorphic proviral loci in pig genomes. PERV s are transmitted vertically and cannot be completely eliminated by breeding or gene knock out technology. PERV s entail a public health concern that will persist even if no evidence of PERV infection of xenotransplant recipients in vivo has been revealed yet. Nevertheless, infectious risks must be minimized by full assessment of pigs as donors by combining different molecular screening assays for sensitive and specific detection as well as a functional analysis of the infectivity of PERV including an adequate monitoring of recipients.
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