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Invasive breast carcinoma is a combination of heterogeneous diseases with distinct molecular and clinical features. Some subsets of breast cancer present major clinical challenges, including triple-negative, metastatic/recurrent disease and rare breast histologies. Previously, we developed a unique resource of 37 hard-to-treat breast cancer patient-derived xenografts (PDX). This set included mainly triple negative breast cancer (TNBC) patients that presented poor response to neoadjuvant chemotherapies (Savage et al. 2020 - PMID: 32546838). PDXs accurately reproduce the molecular heterogeneity of the primary tumors and show that multi-drug chemoresistance was retained upon xenotransplantation. Here, we present the characterization of PDX 3-dimensional cultures organoids (8) and PDX derived epithelial cell lines (11). Using single-cell RNAseq we showed that an organoid cultured for several passages (P8) maintained the heterogeneity of the matched PDX. Although in different proportions, all cancer cell populations found in the PDX were retained in matched organoids supporting that organoids are suitable models that recapitulates the tumor heterogeneity and are therefore a suitable model for drug screening. Among our new PDXs models (30), we have developed four PDXs from rare metaplastic breast cancers (MpBC), an aggressive subtype of breast cancer that present the poorest response to standard of care chemotherapy. We also developed one male breast cancer PDX with matched organoid. Omic analysis on our pre-clinical models and patient primary tumor and metastasis will inform development of therapeutic opportunities. This molecular information will guide selection of compounds that will be validated using our high throughput organoid drug screening pipeline. This will allow rapid screens of thousands of approved drugs, enhancing drug repurposing with potential for rapid clinical translation. The combined use of 3D tumor organoids and PDXs, is an important opportunity poised to transform identification of new therapeutic options for hard-to-treat lethal breast cancers. Citation Format: Hellen Kuasne, Anne-Marie fortier, Sandrine Busque, Simon Mathien, Paul Savage, Constanza Martinez Ramirez, Anie Monast, Margarita Soleinova, Atilla Omeroglu, Jamil Asselah, Nathaniel Bouganim, Sarkis Meterissian, Mark Basik, Morag Park. Development and molecular characterization of hard-to-treat breast cancer pre-clinical models to enhance precision medicine [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-08-09.

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Tumor progression upon treatment arises from preexisting resistant cancer cells and/or adaptation of persister cancer cells committing to an expansion phase. Here, we show that evasion from viral mimicry response allows the growth of taxane-resistant triple-negative breast cancer (TNBC). This is enabled by an epigenetic state adapted to taxane-induced metabolic stress, where DNA hypomethylation over loci enriched in transposable elements (TE) is compensated by large chromatin domains of H3K27me3 to warrant TE repression. This epigenetic state creates a vulnerability to epigenetic therapy against EZH2, the H3K27me3 methyltransferase, which alleviates TE repression in taxane-resistant TNBC, leading to double-stranded RNA production and growth inhibition through viral mimicry response. Collectively, our results illustrate how epigenetic states over TEs promote cancer progression under treatment and can inform about vulnerabilities to epigenetic therapy.Significance:Drug-resistant cancer cells represent a major barrier to remission for patients with cancer. Here we show that drug-induced metabolic perturbation and epigenetic states enable evasion from the viral mimicry response induced by chemotherapy in TNBC. These epigenetic states define a vulnerability to epigenetic therapy using EZH2 inhibitors in taxane-resistant TNBC.See related commentary by Janin and Esteller, p. 1258.This article is highlighted in the In This Issue feature, p. 1241

Many control methods target agricultural pests’ reproductive traits and so, knowledge of these traits is crucial. The seedcorn maggot, Delia platura (Diptera: Anthomyiidae), is reported as a cosmopolitan polyphagous pest species which can be found in high numbers in numerous crops. Two morphologically identical genetic lines of D. platura (H- and N- lines) with distinct distribution ranges were recently discovered. While many biological traits have been described for D. platura, no study to date has been conducted on the life history strategies and reproductive behaviors of its two lines. Using laboratory-reared colonies originating from the Montérégie region in Québec, this project investigates the effect of group composition (sex-ratio and density) on the mating success and pre-oviposition period of the two D. platura lines. We found a substantial increase in mating success with increasing proportion of males within mating groups for both lines while group density had negligeable effects. These results corroborate reports of D. platura high-density swarms in which sex ratios are usually male-biased. The pre-oviposition period decreased as the ratio of males to female increased at low density only for the N-line while the opposite trend was observed at high density for both lines. These results suggest differences between the mating systems of these two lines, with the H-line females being choosier towards potential mates than those of the N-line. We also describe reproductive traits of both lines along with their implications for integrated pest management strategies such as the sterile insect technique or the release of individuals carrying pathogens, both of which must take into account the high degree of sexual selection present in such group contexts. Keywords: reproductive biology, mating dynamics, seedcorn maggot, swarms

Large chromosomal alterations are common in cancer and often show preferential gain or loss across many cancer types indicating their selective advantage. Triple negative breast cancer (TNBC) exhibits complex mutational spectrum without common oncogenic drivers yet displays consistent loss of large chromosomal regions. Here, we characterize selection pressures that maintain a recurrently deleted region of chromosome 4p in TNBC. We used bulk WGS phylogenetic analysis of TNCB PT/PDX panel to show that chr4p deletion is an early event in tumor evolution. We used scRNAseq gene expression and inferred copy number analysis to show that chr4p loss is associated with a proliferative state. This finding was confirmed by a combination of RNA in situ hybridization and immunofluorescence. We then tested the dosage sensitivity of genes residing within this region by individual and dual overexpression in TNBC PDX-derived cell lines and control normal cell line by assessing their effect on cell proliferation. The overexpression of genes within chr4p elicited a strong cell proliferation defect in cancer but not normal cell line models. We also characterized an unknown gene within chr4p region as a novel member of the STRIPAK complex. Genome-wide pooled ORFeome library screens identified a global pattern of background-specific dosage sensitive regions. Our study shows that large chromosomal deletions are maintained due to evolutionary early genetic network rewiring rendering multiple genes within such regions to be dosage sensitive. Ultimately, this work enhances our understanding of genetic events that modulate TNBC. Citation Format: Elena Kuzmin, Jean Monlong, Mathieu Bourgey, Tom Lesluyes, Toby Barker, Genevieve Morin, Dongmei Zou, Michael Schwartz, Yang Yang, Alain Pacis, Constanza Martinez, Hellen Kuasne, Anne-Marie Fortier, Rui Li, Claudia Kleinman, Sidong Huang, Peter van Loo, Jiannis Ragoussis, Guillaume Bourque, Morag Park. Evolution of large copy number variants in breast cancer through genetic network rewiring [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1512.

Tumor progression upon treatment arises from preexisting resistant cancer cells and/or adaptation of persister cancer cells committing to an expansion phase. Here, we show that evasion from viral mimicry response allows the growth of taxane-resistant triple-negative breast cancer (TNBC). This is enabled by an epigenetic state adapted to taxane-induced metabolic stress, where DNA hypomethylation over loci enriched in transposable elements (TE) is compensated by large chromatin domains of H3K27me3 to warrant TE repression. This epigenetic state creates a vulnerability to epigenetic therapy against EZH2, the H3K27me3 methyltransferase, which alleviates TE repression in taxane-resistant TNBC, leading to double-stranded RNA production and growth inhibition through viral mimicry response. Collectively, our results illustrate how epigenetic states over TEs promote cancer progression under treatment and can inform about vulnerabilities to epigenetic therapy. Significance: Drug-resistant cancer cells represent a major barrier to remission for patients with cancer. Here we show that drug-induced metabolic perturbation and epigenetic states enable evasion from the viral mimicry response induced by chemotherapy in TNBC. These epigenetic states define a vulnerability to epigenetic therapy using EZH2 inhibitors in taxane-resistant TNBC. See related commentary by Janin and Esteller, p. 1258. This article is highlighted in the In This Issue feature, p. 1241

Although cancer-associated fibroblasts (CAFs) have gained increased attention for supporting cancer progression, current CAF-targeted therapeutic options are limited and failing in clinical trials. As the largest component of the tumor microenvironment (TME), CAFs alter the biochemical and physical structure of the TME, modulating cancer progression. Here, we review the role of CAFs in altering drug response, modifying the TME mechanics and the current models for studying CAFs. To provide new perspectives, we highlight key considerations of CAF activity and discuss emerging technologies that can better address CAFs; and therefore, increase the likelihood of therapeutic efficacy. We argue that CAFs are crucial components of the cancer drug discovery pipeline and incorporating these cells will improve drug discovery success rates.Recent advances in cancer research have improved our understanding of disease progression; however, the number of drugs failing in clinical trials remains high and therefore, present a critical challenge for cancer drug discovery. Although the interactions of the tissue surrounding the tumor, the tumor microenvironment, are now considered key targets for new interventions in cancer, the role of microenvironment is largely absent in drug discovery pipelines. Here we explore the role of the most prominent cell type in the tumor microenvironment, cancer-associated fibroblasts (CAFs), in altering cancer therapy response and ultimately patient outcome. To provide new perspectives for future studies, we draw attention to key complications of CAF biology and highlight emerging technologies that could be used to address this. We believe including CAFs in drug discovery, whether for targeting cancer cells or the microenvironment, will allow for a better understanding of therapeutic efficacy and ultimately improve clinical outcome.

Transmembrane glycoprotein NMB (GPNMB) is a prognostic marker of poor outcome in patients with triple-negative breast cancer (TNBC). Glembatumumab Vedotin, an antibody drug conjugate targeting GPNMB, exhibits variable efficacy against GPNMB-positive metastatic TNBC as a single agent. We show that GPNMB levels increase in response to standard-of-care and experimental therapies for multiple breast cancer subtypes. While these therapeutic stressors induce GPNMB expression through differential engagement of the MiTF family of transcription factors, not all are capable of increasing GPNMB cell-surface localization required for Glembatumumab Vedotin inhibition. Using a FACS-based genetic screen, we discovered that suppression of heat shock protein 90 (HSP90) concomitantly increases GPNMB expression and cell-surface localization. Mechanistically, HSP90 inhibition resulted in lysosomal dispersion towards the cell periphery and fusion with the plasma membrane, which delivers GPNMB to the cell surface. Finally, treatment with HSP90 inhibitors sensitizes breast cancers to Glembatumumab Vedotin in vivo, suggesting that combination of HSP90 inhibitors and Glembatumumab Vedotin may be a viable treatment strategy for patients with metastatic TNBC.

Despite major advances in the treatment of breast cancer (BC), it remains the most diagnosed and second most deadly cancer among American women. BC is a heterogeneous disease consisting of distinct subtypes, including hormone receptor-positive, HER2 receptor-positive and cancers that lack these receptors categorized as triple-negative breast cancers (TNBC). The TNBC subtype presents the worst outcome and the highest rates of recurrence and metastasis. The absence of targets prevents the use of established precision therapies in TNBC, and the standard of care remains neoadjuvant chemotherapy. While this is effective in some patients, about 50% develop resistance, leading to the development of metastasis. It is known that selective pressures exerted by chemotherapy treatment can promote the outgrowth of resistant tumor subclones. However, the diverse intra-tumoral population and the mechanisms that lead to chemotherapy resistance in TNBC are still poorly understood. We hypothesized that therapeutic regimens influence tumor plasticity by exerting selective pressures leading to the outgrowth of resistant subpopulations with the greatest survival advantage. To evaluate this hypothesis, we aimed to generate in vivo models of chemotherapy resistance and to investigate the plasticity of tumor cell subpopulations challenged with standard-of-care chemotherapies. To this end, we selected a multi-drug resistant (Doxorubicin, Cyclophosphamide, Cisplatin, and Paclitaxel) BC patient and developed a patient-derived xenograft (PDX) from the primary tumor and the lung metastasis. The metastasis PDX model was initially responsive to Gemcitabine (as observed in the BC patient) but eventually developed resistance. We challenged this metastasis PDX with several cycles of Gemcitabine and obtained residual, rebound, and resistant tumor samples. We performed single-cell RNA sequencing (scRNAseq) of these models using a droplet-based technology from 10X Genomics. This scRNAseq data was used to compare the changes in the proportions of cellular subpopulations in each model. Interestingly, our data shows that the rebound model presents greater similarity to the untreated metastasis, while the resistant model has significant differences in cell population expression profiles. We identified a hypoxic population in the primary tumor and its matched metastasis. This population was validated in these models by Nanostring GeoMx Digital Spatial Profiler. Our recent analyses have identified that this hypoxic population persists in the residual, rebound and resistant models. In addition, we have identified other populations that vary in these models. We are currently investigating their cellular mechanisms and gene expression patterns. Using scRNA-sequencing to understand the clonal expansion of resistant subpopulations following chemotherapy reveals distinctive resistant cell features enabling the identification of the vulnerabilities of these tumors. Citation Format: Sandrine Busque, Constanza Martinez Ramirez, Hellen Kuasne, Paul Savage, Anne-Marie Fortier, Anie Monast, Atilla Omeroglu, Jamil Asselah, Nathaniel Bouganim, Sarkis Meterissian, Claudia Kleinman, Mark Basik, Morag Park. Identification of gemcitabine-resistant populations using scRNA-sequencing in triple negative breast cancer patient-derived xenograft [abstract]. In: Proceedings of the AACR Special Conference: Cancer Metastasis; 2022 Nov 14-17; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_2):Abstract nr A033.

Simple Summary There is increasing evidence that obesity and high circulating cholesterol levels are associated with an increased risk of recurrence and a higher mortality rate in breast cancer patients via altering the metabolic programming in breast cancer cells. However, the underlying molecular mechanism by which high cholesterol levels reprogram the metabolic pathways in breast cancer cells is not well-understood. We have previously demonstrated that cholesterol acts as an endogenous agonist of estrogen-related receptor α (ERRα), a strong regulator of cellular metabolism. The aim of the current study is to demonstrate whether cholesterol/obesity mediates its pathogenic effect in breast cancer cells via altering metabolic pathways in an ERRα-dependent manner. The findings of this study provide mechanistic insights into the link between cholesterol/obesity and metabolic reprogramming in breast cancer patients and reveal the metabolic vulnerabilities in such breast cancer patients that could be therapeutically targeted. Abstract The molecular mechanism underlying the metabolic reprogramming associated with obesity and high blood cholesterol levels is poorly understood. We previously reported that cholesterol is an endogenous ligand of the estrogen-related receptor alpha (ERRα). Using functional assays, metabolomics, and genomics, here we show that exogenous cholesterol alters the metabolic pathways in estrogen receptor-positive (ER+) and triple-negative breast cancer (TNBC) cells, and that this involves increased oxidative phosphorylation (OXPHOS) and TCA cycle intermediate levels. In addition, cholesterol augments aerobic glycolysis in TNBC cells although it remains unaltered in ER+ cells. Interestingly, cholesterol does not alter the metabolite levels of glutaminolysis, one-carbon metabolism, or the pentose phosphate pathway, but increases the NADPH levels and cellular proliferation, in both cell types. Importantly, we show that the above cholesterol-induced modulations of the metabolic pathways in breast cancer cells are mediated via ERRα. Furthermore, analysis of the ERRα metabolic gene signature of basal-like breast tumours of overweight/obese versus lean patients, using the GEO database, shows that obesity may modulate ERRα gene signature in a manner consistent with our in vitro findings with exogenous cholesterol. Given the close link between high cholesterol levels and obesity, our findings provide a mechanistic explanation for the association between cholesterol/obesity and metabolic reprogramming in breast cancer patients.

This afterword addresses four broad questions raised by this special issue: uncertainty as a mode of governance, the ontological politics of naturalisation, the citizen-noncitizen distinction, and performative (anxious) states. First, taking uncertainty as a mode of neoliberal governance as the starting point of analysis, this afterword invites the scrutiny of the ways in which the artifice and uncertainty of citizenship are concealed or rendered irrelevant in naturalisation processes. Second, the contributions to this special issue consider naturalisation as a social and political process, rather than solely as a legal status. Pushing this conception further, this afterword considers naturalisation as transactional in two ways: on the one hand, migrants navigate a number of formal and informal requirements and ‘tests’, where some transactions are needed along the way, be they financial, practical, or symbolic. On the other hand, transactions will also occur in the translation of political ideology into policy. Third, naturalisation regimes both blur and reify the citizen-noncitizen and the citizen-migrant distinctions. Distinctions which this afterword unpacks by unravelling the assumed separation between citizenship and migration. How are citizens and migrants migratised? How are migrants and citizens citizenised? Fourth, a further element of the analysis concerns how state-citizen relations are enacted and by extension, how the state itself is ‘made up’ and ‘anxious’. The affective politics of ‘anxious states’ are telling of the frames of desire of naturalisation, which are founded on a threefold principle: the desirability of citizenship, the desire for desirable citizens, and the desirability of the state itself.

Large chromosomal alterations are common in cancer and often show preferential gain or loss across many cancer types indicating their selective advantage. Triple negative breast cancer (TNBC) exhibits complex mutational spectrum without common oncogenic drivers yet displays consistent loss of large chromosomal regions. Here, we characterize selection pressures that maintain a recurrently deleted region of chromosome 4p in TNBC. We used single cell and bulk WGS phylogenetic analysis of TNCB PT/PDX panel to show that chr4p deletion is an early event in tumor evolution. We used scRNAseq gene expression and inferred copy number analysis to show that chr4p loss is associated with a proliferative state. This finding was confirmed by a combination of RNA in situ hybridization and immunofluorescence. We then tested the dosage sensitivity of genes residing within this region by individual and dual overexpression in TNBC PDX-derived cell lines and control normal cell line by assessing their effect on cell proliferation. The overexpression of genes within chr4p elicited a strong cell proliferation defect in cancer but not normal cell line models. We also characterized an unknown gene within chr4p region as a novel member of the STRIPAK complex. Genome-wide pooled ORFeome library screens identified a global pattern of background-specific dosage sensitive regions. Our study shows that large chromosomal deletions are maintained due to evolutionary early genetic network rewiring rendering multiple genes within such regions to be dosage sensitive. Ultimately, this work enhances our understanding of genetic events that modulate TNBC. Citation Format: Elena Kuzmin, Jean Monlong, Mathieu Bourgey, Jarry Barber, Tom Lesluyes, Toby Baker, Genevieve Morin, Dongmei Zou, Michael Schwartz, Yang Yang, Alain Pacis, Constanza Martinez, Hellen Kuasne, Anne-Marie Fortier, Rui Li, Claudia Kleinman, Sidong Huang, Peter van Loo, Quaid Morris, Jiannis Ragoussis, Guillaume Bourque, Morag Park. Evolution of large copy number variants in breast cancer through genetic network rewiring [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 44.

Triple Negative Breast Cancer (TNBC) is characterized by the absence of common oncogenic drivers, limiting its treatment options; however, it exhibits recurrent large chromosomal deletions. We previously showed that chromosome 4p (chr4p) loss is a frequently observed large copy number variant in TNBC and is associated with poor prognosis. We also showed that chr4p deletion is an early event in tumor evolution and confers on cells a proliferative advantage. Here, we set out to uncover the genetic vulnerabilities associated with chr4p deletion in TNBC to identify novel therapeutic avenues for TNBC and enhance our understanding of the genetic mechanisms that maintain chr4p deletion in the genome. Whole genome sequence analysis of our TNBC Primary Tumor(PT)/Patient-Derived Xenograft (PDX) panel identified samples with copy neutral and deletion status of chr4p. These deletion regions span a large fraction of the chr4p arm. RNAseq analysis revealed that chr4p deletion is functionally significant since gene expression of ~80% of genes was reduced upon chr4p deletion. Chr4p deletion was associated with global transcriptomic changes and differentially expressed genes were enriched for proliferation, DNA replication, cell migration, activation of the innate immune response and protein translation. PDX-derived cell models from these samples showed lentiviral infectivity based on a control lentivirus expressing GFP. Additionally, these PDX cell models were shown to be editable using CRISPR-Cas9 through the targeting of core essential genes. We will use a pooled CRISPR-Cas9 approach to systematically screen for genetic vulnerabilities in TNBC PDX-derived cell models harbouring chr4p copy neutral or deletion state. To further investigate the genetic mechanisms buffering chr4p loss, we have leveraged publicly-available CRISPR-Cas9 genome-wide genetic screen data from the Cancer DepMap to identify putative synthetic lethal (pSL) partners with chr4p deletion in TNBC. This was accomplished by developing regression models and integrating them with results from established methods such as drugZ and MAGeCK. We identified pSL partners for chr4p at gene, segmental and arm levels. pSLs were enriched for mitochondrial, protein translation and proliferation pathways. We will validate the top candidates from these analyses in our cohort of chr4p deletion and chr4p copy neutral TNBC PDX-derived cell models and integrate them with the pooled CRISPR screens. Together, this work aims to reveal potential TNBC-specific therapeutic avenues for precision oncology. Citation Format: Michael Schwartz, Rohan Dandage, Lynn Karam, Alain Pacis, Hellen Kuasne, Anne-Marie Fortier, Sidong Huang, Guillaume Bourque, Traver Hart, Elena Kuzmin, Morag Park. Identifying genetic vulnerabilities of chromosome 4p large copy number variants in triple negative breast cancer [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 47.

Triple negative breast cancer (TNBC) is a deadly form of breast cancer due to the development of resistance to chemotherapy affecting over 30% of patients. New therapeutics and companion biomarkers are urgently needed. Recognizing the elevated expression of glucose transporter 1 (GLUT1, encoded by SLC2A1) and associated metabolic dependencies in TNBC, we investigated the vulnerability of TNBC cell lines and patient-derived samples to GLUT1 inhibition. We report that genetic or pharmacological inhibition of GLUT1 with BAY-876 impairs the growth of a subset of TNBC cells displaying high glycolytic and lower oxidative phosphorylation (OXPHOS) rates. Pathway enrichment analysis of gene expression data suggests that the functionality of the E2F pathway may reflect to some extent OXPHOS activity. Furthermore, the protein levels of retinoblastoma tumor suppressor (RB1) strongly correlate with the degree of sensitivity to GLUT1 inhibition in TNBC, where RB1-negative cells are insensitive to GLUT1 inhibition. Collectively, our results highlight a strong and targetable RB1-GLUT1 metabolic axis in TNBC and warrant clinical evaluation of GLUT1 inhibition in TNBC patients stratified according to RB1 protein expression levels., Triple negative breast cancer is a deadly form of breast cancer with limited therapeutic options. Here the authors show the efficacy of GLUT1 pharmacological inhibition against a subset of tumors expressing RB1, thereby identifying RB1 protein level as a biomarker of sensitivity to anti-GLUT1 therapy.

The seedcorn maggot Delia platura (Meigen), and the bean seed maggot Delia florilega (Zetterstedt) can cause considerable feeding damage to a wide range of cultivated crops. The recent discovery of two distinct genetic lines of D. platura, each with a unique distribution pattern overlapping only in eastern Canada, suggests the presence of a new cryptic species for the group. The reliable identification of the three species/lines in the seedcorn maggot complex is crucial to our understanding of their distribution, phenology, and respective contribution to crop damage as well as to the development of specific integrated pest management approaches. As these taxa are morphologically indistinguishable in the immature stages, we developed a high-resolution melting PCR (HRM) assay using primers amplifying a variable 96-bp PCR product in the CO1 mitochondrial gene for rapid and economical identification of specimens. The three species/lines exhibited distinguishable melting profiles based on their different Tm values (between 0.4 and 0.9°C) and identification results based on HRM and DNA sequencing were congruent for all specimens in the validation data set (n = 100). We then used the new, highly sensitive HRM assay to identify survey specimens from the seedcorn maggot complex collected in Quebec, Canada, between 2017 and 2019. Progress curves developed to document the temporal occurrence patterns of each species/lines indicate differences between taxa, with the N-line (BOLD:AAA3453) of D. platura appearing approximately 17 d before D. florilega (BOLD:ACR4394) and the H-line (BOLD:AAG2511) of D. platura.

Triple negative breast cancer (TNBC) is a deadly form of breast cancer due to the development of resistance to chemotherapy affecting over 30% of patients. New therapeutics and companion biomarkers are urgently needed. Recognizing the elevated expression of glucose transporter 1 (GLUT1, encoded by SLC2A1) and associated metabolic dependencies in TNBC, we investigated the vulnerability of TNBC cell lines and patient-derived samples to GLUT1 inhibition. We report that genetic or pharmacological inhibition of GLUT1 with BAY-876 impairs the growth of a subset of TNBC cells displaying high glycolytic and lower oxidative phosphorylation (OXPHOS) rates. Pathway enrichment analysis of gene expression data implicates E2F Targets pathway activity as a surrogate of OXPHOS activity. Furthermore, the protein levels of retinoblastoma tumor suppressor (RB1) are strongly correlated with the degree of sensitivity to GLUT1 inhibition in TNBC, where RB1-negative cells are insensitive to GLUT1 inhibition. Collectively, our results highlight a strong and targetable RB1-GLUT1 metabolic axis in TNBC and warrant clinical evaluation of GLUT1 inhibition in TNBC patients stratified according to RB1 protein expression levels.

Tumor growth is regulated by complex interactions of tumor cells with the tumor microenvironment (TME) which consists of cancer associated fibroblasts (CAFs), various immune cells including tumor infiltrating lymphocytes (TILs) and extracellular matrix components (1). Accurately modeling the TME in vitro is challenging as there is significant genetic (2, 3) and spatial (4) heterogeneity within TME components. Triple-negative breast cancer (TNBC) patients that have tumors with TILs restricted to the margins are called immune “cold” and have poor prognosis (5, 6). Interfering with the mechanisms that exclude TILs from these tumors may improve immunotherapy response and TNBC patient outcome. A major barrier to anti-cancer drug development is the availability of in vitro models that accurately reflect the in vivo TME. Our goal is to establish and validate a 3D bioprinted tumor model that reproduces the spatial relationship between tumor, CAFs and TILs. We have generated a biobank of TNBC patient-derived organoids and tissue-matched TME components. The tumor immune microenvironment molecular subtypes were assigned to each tumor sample using transcriptomic data and immunohistochemistry analysis. We have used microfluidic 3D bioprinting (RX1™ bioprinter, Aspect Biosystems) to print a core-shell fibre design that recapitulates the immune “cold” TME and permits monitoring of immune cell infiltration into the cancer cell core of the fibre. Clinically proven checkpoint inhibitor therapy will be applied to our model to validate its biological function. This innovative bioprinted tumor model will enable phenotypic screening to efficiently identify new targets that transform hard-to-treat immune “cold” tumors into a state where immune cells can infiltrate the tumor. Using patient-derived tissue-matched TME components increases the clinical relevance of the bioprinted tumor model and enables personalized anti-cancer drug screening in vitro to accurately predict patient response to therapy. 1. Balkwill FR, Capasso M, Hagemann T. The tumor microenvironment at a glance. J Cell Sci. 2012; 125: 5591-5596. 2. Finak G, Bertos N, Pepin F, et al. Stromal gene expression predicts clinical outcome in breast cancer. Nat Med. 2008; 14: 518-527. 3. Lehmann BD, Bauer JA, Chen X, et al. Identification of human triple-negative breast cancer subtypes and preclinical models for selection of targeted therapies. J Clin Invest. 2011; 121: 2750-2767. 4. Gruosso T, Gigoux M, Manem VSK, et al. Spatially distinct tumor immune microenvironments stratify triple-negative breast cancers. J Clin Invest. 2019;129(4):1785-1800. 5. Ali HR, Provenzano E, Dawson SJ, et al. Association between CD8+ T-cell infiltration and breast cancer survival in 12,439 patients. Ann Oncol. 2014; 25: 1536-1543. 6. Salgado R, Denkert C, Demaria S, et al. The evaluation of tumor-infiltrating lymphocytes (TILs) in breast cancer: recommendations by an International TILs Working Group 2014. Ann Oncol. 2015; 26: 259-271. Citation Format: Anne-Marie Fortier, Erin Bedford, Salvador Flores Torres, Spiro Getsios, J. Matt Kinsella, Sam Wadsworth, Morag Park. Heating up the fight against cancer using 3D bioprinting [abstract]. In: Proceedings of the AACR Virtual Special Conference on the Evolving Tumor Microenvironment in Cancer Progression: Mechanisms and Emerging Therapeutic Opportunities; in association with the Tumor Microenvironment (TME) Working Group; 2021 Jan 11-12. Philadelphia (PA): AACR; Cancer Res 2021;81(5 Suppl):Abstract nr PO006.

It is increasingly clear that triple negative breast cancer (TNBC) is a heterogeneous disease with variable clinical presentation, histological features and response to therapy. While these differences have been partially explained by inter- and intra-tumoral heterogeneity, spatial heterogeneity represented by the tumor architecture plays a critical role in clonal evolution and displays a landscape for different compartment-specific processes and cell-to-cell specific interactions. To better understand the impact of spatial heterogeneity on gene-expression-defined cell populations, we have coupled single-cell RNA sequencing with multiplex immunofluorescence (IF). Multiplex immunofluorescence using specific markers for each cell cluster, integrated with computational image analyses and neighborhood maps, has revealed spatial zonation of single cell subpopulations. Zonation of single cell populations was coupled to hypoxia and overlayed distinct metabolic tumor zones and defined areas of differential stress and cell plasticity that give rise to tumor cells with enhanced fitness and aggressivity. Citation Format: M Park, C Martínez Ramirez, Y Yang, A Blanchet-Cohen, H Kuasne, A Fortier, J Ragoussis, P Savage, A Omeroglou, S Meterissian, S Costantino, C Kleinman. Spatial variance signatures/Intra-tumor zonation in TNBC [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr SP112.

This article empirically grounds the ‘psychic life of power’ (Butler, 1997) by demonstrating the psychic form that power takes as immigrants or agents of the state make their way through the British ‘citizenisation’ policy – i.e. the ‘integration’ policy that requires noncitizens to acquire ‘citizen-like’ skills and values in view of seeking citizenship or other statuses (e.g. settlement). The framing argument is that an ambivalent relationship between desire and anxiety mediates the state-citizen relationship (following Honig, 2001). Taking this argument further, the article offers an in-depth analysis of how citizenisation policy’s frames of desire (the assumed desirability of citizenship and the desire for desirable citizens) also take the form of anxieties. Drawing on a multi-sited study of citizenisation in Britain, the article explores some of the different forms anxiety takes: fetishisation, enervation, and uncertainty. The analysis reveals how the uneven distribution of anxiety between agents of the state and immigrants not only mediates the state–citizen relationship but also variously enacts the state itself. Attending to the psychosocial dynamics of citizenisation reveals how hierarchies are (re)produced not only discursively and materially, but also through different ‘anxious states’.

This article examines the colonial legacies shaping current language requirements for immigrants applying for settlement or citizenship in Britain. The article argues that common sense understandings of ‘national language’ and monolingualism/multilingualism were developed in the context of imperial expansion, the legacies of which resonate today in a disdain for multilingualism and other Englishes conceived as hampering cohesion. Put simply, other languages and other English are spoken here because English was there. Drawing on interviews with applicants and English teaching professionals, the article discusses how participants variously experience English language requirements. The analysis shows how the colonial legacies supporting the rise of English as a ‘world language’ cast it as the locus of a regime of audibility that establishes a hierarchy between ‘the English’ and the ‘anglicised’. In today’s Britain, the multilingualism of the other is not external and prior to Britain, but rather speaks volumes to and about contemporary Britain.

Latente au début, puis manifeste au fil des ans, la figure d'Arthur Rimbaud a accompagné René Char tout au long de son œuvre, de 1927 à 1988. Elle a aussi joué un rôle central dans le débat sur l'esthétique de la poésie moderne auquel Char a participé. À partir de ses textes et en tenant compte des autres interprétations de la geste rimbaldienne qui ont ponctué le XXe siècle, cet essai retrace l'évolution du rapport de Char à Rimbaud. Il dégage, bien plus qu'une influence, le nouveau texte rimbaldien que Char a élaboré pour revendiquer l'héritage du poète aux semelles de vent. Exister pour la postérité. Mais on l'assimile toujours à ses contemporains. Giuseppe Pontiggia, Le jardin des Hespérides

This article takes the naturalisation process as a vantage point to consider how citizenship constitutes a site of emotional investment not only on the part of applicants and ‘new’ citizens but also on the part of the state. The premise of this article is that naturalisation is more than solely the admission of foreigners to the position and rights of citizenship, and it approaches naturalisation as a state practice that needs to be understood within a politics of desire. The article asks three questions: what makes naturalisation a thinkable and desirable means of acquiring citizenship? Second, what do practices of naturalisation tell us about ‘the state's attachment to particular embodiments of desirable citizens’ (S. Somerville, 2005, Notes toward a queer history of naturalization. American quarterly, 7 (3), 661)? Third, ‘who may desire the state's desire’ (J. Butler, 2002. Is kinship always already heterosexual? Differences, 13 (1), 22)? Using policy documents and auto-ethnographic material, the artic...

Keratins 8 and 18 (K8/18) are simple epithelial cell-specific intermediate filament proteins. Keratins are essential for tissue integrity and are involved in intracellular signaling pathways that regulate cell response to injuries, cell growth, and death. K8/18 expression is maintained during tumorigenesis; hence, they are used as a diagnostic marker in tumor pathology. In recent years, studies have provided evidence that keratins should be considered not only as markers but also as regulators of cancer cell signaling. The loss of K8/18 expression during epithelial-mesenchymal transition (EMT) is associated with metastasis and chemoresistance. In the present study, we investigated whether K8/18 expression plays an active role in EMT. We show that K8/18 stable knockdown using shRNA increased collective migration and invasiveness of epithelial cancer cells without modulating EMT markers. K8/18-depleted cells showed PI3K/Akt/NF-κB hyperactivation and increased MMP2 and MMP9 expression. K8/18 deletion also increased cisplatin-induced apoptosis. Increased Fas receptor membrane targeting suggests that apoptosis is enhanced via the extrinsic pathway. Interestingly, we identified the tight junction protein claudin1 as a regulator of these processes. This is the first indication that modulation of K8/18 expression can influence the phenotype of epithelial cancer cells at a transcriptional level and supports the hypothesis that keratins play an active role in cancer progression.

After briefly discussing the ‘affective turn’ in citizenship studies, this afterword discusses the political mobilisation of affect to consider when, where, and how affect may be connected to citizenship. It asks: What does it mean and do to speak of affective acts as acts of affective citizenship? I argue that the phrase ‘affective citizenship’ attaches affect to a very specific object: citizenship. Studying affective citizenship requires attention to how some feelings attach themselves to citizenship and to how citizenship itself can evoke certain feelings. But affective citizenship does not occur ‘naturally’: it arises from, requires and/or produces knowledge, labour, and (new) ‘feeling rules’ (Hochschild in de Wilde and Duyvendak). I conclude with a call for more research into the dynamics of affective citizenship that go beyond a simple opposition between those simply conceived of as agents of disciplinary power and those seen as (resisting) subjects of disciplinary power.

In 2007, Channel 4 screened Face of Britain, a documentary about the genetic mapping of Britain. Face of Britain promised to reveal ‘who we really are’ by tracing genetic links back to ancient Celts, Anglo-Saxons, Vikings and Normans. This article situates Face of Britain within the wider racial and national politics that it is invariably caught up in, and examines how ‘race’ and racial thinking are reconfigured in ways that are both ‘old’ and ‘new’. Face of Britain constitutes an interesting case study to examine how ideas of indigeneity are produced and naturalised in scientific discourses and practices. Here, indigeneity is mobilised through three sub-narratives: the ‘vanishing indigene’; the promise of facial recognition; and DNA and national relatedness. The analysis reveals how some people of the British Isles are naturalised as indigenous by virtue of their ancestral presence in this land through a combination of genetic and photographic technologies. In short, blood and soil are intertwined, with ...

Traditionally, oxytocin (OT) is well known to play a crucial role in the regulation of cyclic changes in the uterus, implantation of the embryo, and parturition. Recently, an additional role for OT has been identified in several types of cancer cells in which OT acts as a growth regulator. In endometrial cancer cells, OT is known to efficiently inhibit cellular proliferation. In the present study, we show that OT increases invasiveness of human endometrial carcinoma (HEC) cells, which are otherwise resistant to the growth-inhibiting effects of OT. Using pharmacological inhibitors, invasion assay, RNA interference, and immunofluorescence, we found that OT enhances the invasive properties of HEC cells through up-regulation of X-linked inhibitor of apoptosis protein (XIAP), matrix-metalloproteinase 2 (MMP2), and matrix-metalloproteinase 14 (MMP14). In addition, we show that OT-mediated invasion is both cyclooxygenase 1 (PTGS1) and cyclooxygenase-2 (PTGS2) dependent via the phosphatidylinositol 3-kinase/AKT (PIK3/AKT) pathway. PTGS2 knockdown by shRNA resulted in XIAP down-regulation. We also show that OT receptor is overexpressed in grade I to III endometrial cancer. Taken together, our results describe for the first time a novel role for OT in endometrial cancer cell invasion.

Akt/PKB kinases are central mediators of cell homeostasis. There are three highly homologous Akt isoforms, Akt1/PKBα, Akt2/PKBβ and Akt3/PKBγ. Hyperactivation of Akt signaling is a key node in the progression of a variety of human cancer, by modulating tumor growth, chemoresistance and cancer cell migration, invasion and metastasis. It is now clear that, to understand the mechanisms on how Akt affects specific cancer cells, it is necessary to consider the relative importance of each of the three Akt isoforms in the altered cells. Akt1 is involved in tumor growth, cancer cell invasion and chemoresistance and is the predominant altered isoform found in various carcinomas. Akt2 is related to cancer cell invasion, metastasis and survival more than tumor induction. Most of the Akt2 alterations are observed in breast, ovarian, pancreatic and colorectal carcinomas. As Akt3 expression is limited to some tissues, its implication in tumor growth and resistance to drugs mostly occurs in melanomas, gliomas and some breast carcinomas. To explain how Akt isoforms can play different or even opposed roles, three mechanisms have been proposed: tissue-specificity expression/activation of Akt isoforms, distinct effect on same substrate as well as specific localization through the cyto-skeleton network. It is becoming clear that to develop an effective anticancer Akt inhibitor drug, it is necessary to target the specific Akt isoform which promotes the progression of the specific tumor.

‘Community cohesion’ is the preferred framework for managing ‘race relations’ and ‘conflict’ in contemporary Britain. Initially adopted in government policy following civil disturbances in the summer of 2001, ‘community cohesion’ combined visions of shared belonging with strategies of managing diversity. More recent versions still place a strong emphasis on ideas of shared belonging but these are combined with strategies of managing migration and identity which are deployed in view of securing local communities against threats posed by extremism, deprivation, diversity and feelings of ‘white unease’. This article examines how the community cohesion agenda relies on strategies of governance that seek to design particular kinds of human behaviours such as ‘mixing’ and ‘meaningful interaction’, in view of ‘delivering’ cohesion in the community. I analyse the cohesion agenda as a form of governance that operates through mechanisms of subjectivation. However, instead of privileging the responsible, discerning,...

In this paper I examine how new forms of multicultural intimacy are imagined in contemporary Britain, and how they are invested with particular ideals of mixing, loving thy neighbour, and feelings for the nation. I trace these discursive themes in a myriad of social locations and forms—a television documentary titled The Last White Kids, reviews of the documentary in the press, and government policies on community cohesion. A key point of this paper is that racial, ethnic, and cultural relations are not only negotiated and ‘managed’ in literal spatial locations, but also imagined through specific emotional and ethical injunctions, such as ‘embracing the other’ and loving they neighbour. Moreover, these injunctions are imagined in the ambivalent spatial terms of obligations to, and dangers of, proximity—an ambivalence that is inflected with articulations of ‘race’, class, and gender. The analysis thus explores how the imperative of neighbourly love refers to both the desires for and anxieties about what the intimacy stems from and fosters. Who gets close to whom, and under what circumstances, is not left to chance.