Differential role of glucocorticoid receptor based on its cell type specific expression on tumor cells and infiltrating lymphocytes.

BACKGROUND: The glucocorticoid receptor (GR) is frequently expressed in breast cancer (BC), and its prognostic implications are contingent on estrogen receptor (ER) status. To address conflicting reports and explore therapeutic potential, a GR signature (GRsig) independent of ER status was developed. We also investigated cell type-specific GR protein expression in BC tumor epithelial cells and infiltrating lymphocytes. METHODS: GRsig was derived from Dexamethasone treated cell lines through a bioinformatic pipeline. Immunohistochemistry assessed GR protein expression. Associations between GRsig and tumor phenotypes (proliferation, cytolytic activity (CYT), immune cell distribution, and epithelial-to-mesenchymal transition (EMT) were explored in public datasets. Single-cell RNA sequencing data evaluated context-dependent GR roles, and a cell type-specific prognostic role was assessed in an independent BC cohort. RESULTS: High GRsig levels were associated with a favorable prognosis across BC subtypes. Tumor-specific high GRsig correlated with lower proliferation, increased CYT, and anti-tumorigenic immune cells. Single-cell data analysis revealed higher GRsig expression in immune cells, negatively correlating with EMT while a positive correlation was observed with EMT primarily in tumor and stromal cells. Univariate and multivariate analyses demonstrated the robust and independent predictive capability of GRsig for favorable prognosis. GR protein expression on immune cells in triple-negative tumors indicated a favorable prognosis. CONCLUSION: This study underscores the cell type-specific role of GR, where its expression on tumor cells is associated with aggressive features like EMT, while in infiltrating lymphocytes, it predicts a better prognosis, particularly within TNBC tumors. The GRsig emerges as a promising independent prognostic indicator across diverse BC subtypes.

Proneural-mesenchymal antagonism dominates the patterns of phenotypic heterogeneity in glioblastoma.

The aggressive nature of glioblastoma (GBM) - one of the deadliest forms of brain tumors - is majorly attributed to underlying phenotypic heterogeneity. Early attempts to classify this heterogeneity at a transcriptomic level in TCGA GBM cohort proposed the existence of four distinct molecular subtypes: Proneural, Neural, Classical, and Mesenchymal. Further, a single-cell RNA sequencing (scRNA-seq) analysis of primary tumors also reported similar four subtypes mimicking neurodevelopmental lineages. However, it remains unclear whether these four subtypes identified via bulk and single-cell transcriptomics are mutually exclusive or not. Here, we perform pairwise correlations among individual genes and gene signatures corresponding to these proposed subtypes and show that the subtypes are not distinctly mutually antagonistic in either TCGA or scRNA-seq data. We observed that the proneural (or neural progenitor-like)-mesenchymal axis is the most prominent antagonistic pair, with the other two subtypes lying on this spectrum. These results are reinforced through a meta-analysis of over 100 single-cell and bulk transcriptomic datasets as well as in terms of functional association with metabolic switching, cell cycle, and immune evasion pathways. Finally, this proneural-mesenchymal antagonistic trend percolates to the association of relevant transcription factors with patient survival. These results suggest rethinking GBM phenotypic characterization for more effective therapeutic targeting efforts.

Perspectives on polarity - exploring biological asymmetry across scales.

In this Perspective, Journal of Cell Science invited researchers working on cell and tissue polarity to share their thoughts on unique, emerging or open questions relating to their field. The goal of this article is to feature 'voices' from scientists around the world and at various career stages, to bring attention to innovative and thought-provoking topics of interest to the cell biology community. These voices discuss intriguing questions that consider polarity across scales, evolution, development and disease. What can yeast and protists tell us about the evolution of cell and tissue polarity in animals? How are cell fate and development influenced by emerging dynamics in cell polarity? What can we learn from atypical and extreme polarity systems? How can we arrive at a more unified biophysical understanding of polarity? Taken together, these pieces demonstrate the broad relevance of the fascinating phenomenon of cell polarization to diverse fundamental biological questions.

An endosomal acid-regulatory feedback system rewires cytosolic cAMP metabolism and drives tumour progression.

Although suppressed cAMP levels have been linked to cancer for nearly five decades, the molecular basis remains uncertain. Here, we identify endosomal pH as a novel regulator of cytosolic cAMP homeostasis and a promoter of transformed phenotypic traits in colorectal cancer (CRC). Combining experiments and computational analysis, we show that the Na+/H+ exchanger NHE9 contributes to proton leak and causes luminal alkalinization, which induces resting [Ca2+], and in consequence, represses cAMP levels, creating a feedback loop that echoes nutrient deprivation or hypoxia. Higher NHE9 expression in cancer epithelia is associated with a hybrid epithelial-mesenchymal (E/M) state, poor prognosis, tumour budding, and invasive growth in vitro and in vivo. These findings point to NHE9-mediated cAMP suppression as a pseudo-starvation-induced invasion state and potential therapeutic vulnerability in CRC. Our observations lay the groundwork for future research into the complexities of endosome-driven metabolic reprogramming and phenotype switching and the biology of cancer progression. Implications: Endosomal pH regulator NHE9 actively controls cytosolic Ca2+ levels to downregulate the adenylate cyclase-cAMP system, enabling colorectal cancer cells to acquire hybrid epithelial-mesenchymal characteristics and promoting metastatic progression.

Loss of p53 epigenetically modulates epithelial to mesenchymal transition in colorectal cancer.

Epithelial to Mesenchymal transition (EMT) drives cancer metastasis and is governed by genetic and epigenetic alterations at multiple levels of regulation. It is well established that loss/mutation of p53 confers oncogenic function to cancer cells and promotes metastasis. Though transcription factors like ZEB1, SLUG, SNAIL and TWIST have been implied in EMT signalling, p53 mediated alterations in the epigenetic machinery accompanying EMT are not clearly understood. This work attempts to explore epigenetic signalling during EMT in colorectal cancer (CRC) cells with varying status of p53. Towards this, we have induced EMT using TGFß on CRC cell lines with wild type, null and mutant p53 and have assayed epigenetic alterations after EMT induction. Transcriptomic profiling of the four CRC cell lines revealed that the loss of p53 confers more mesenchymal phenotype with EMT induction than its mutant counterparts. This was also accompanied by upregulation of epigenetic writer and eraser machinery suggesting an epigenetic signalling cascade triggered by TGFß signalling in CRC. Significant agonist and antagonistic relationships observed between EMT factor SNAI1 and SNAI2 with epigenetic enzymes KDM6A/6B and the chromatin organiser SATB1 in p53 null CRC cells suggest a crosstalk between epigenetic and EMT factors. The observed epigenetic regulation of EMT factor SNAI1 correlates with poor clinical outcomes in 270 colorectal cancer patients taken from TCGA-COAD. This unique p53 dependent interplay between epigenetic enzymes and EMT factors in CRC cells may be exploited for development of synergistic therapies for CRC patients presenting to the clinic with loss of p53.

Protocol for inferring epithelial-to-mesenchymal transition trajectories from single-cell RNA sequencing data using R.

The epithelial-to-mesenchymal transition (EMT) provides crucial insights into the metastatic process and possesses prognostic value within the cancer context. Here, we present COMET, an R package for inferring EMT trajectories and inter-state transition rates from single-cell RNA sequencing data. We describe steps for finding the optimal number of EMT genes for a specific context, estimating EMT-related trajectories, optimal fitting of continuous-time Markov chain to inferred trajectories, and estimating inter-state transition rates.

The exostosin glycosyltransferase 1/STAT3 axis is a driver of breast cancer aggressiveness.

The epithelial-mesenchymal transition (EMT) program is crucial for transforming carcinoma cells into a partially mesenchymal state, enhancing their chemoresistance, migration, and metastasis. This shift in cell state is tightly regulated by cellular mechanisms that are not yet fully characterized. One intriguing EMT aspect is the rewiring of the proteoglycan landscape, particularly the induction of heparan sulfate proteoglycan (HSPG) biosynthesis. This proteoglycan functions as a co-receptor that accelerates cancer-associated signaling pathways through its negatively-charged residues. However, the precise mechanisms through which EMT governs HSPG biosynthesis and its role in cancer cell plasticity remain elusive. Here, we identified exostosin glycosyltransferase 1 (EXT1), a central enzyme in HSPG biosynthesis, to be selectively upregulated in aggressive tumor subtypes and cancer cell lines, and to function as a key player in breast cancer aggressiveness. Notably, ectopic expression of EXT1 in epithelial cells is sufficient to induce HSPG levels and the expression of known mesenchymal markers, subsequently enhancing EMT features, including cell migration, invasion, and tumor formation. Additionally, EXT1 loss in MDA-MB-231 cells inhibits their aggressiveness-associated traits such as migration, chemoresistance, tumor formation, and metastasis. Our findings reveal that EXT1, through its role in HSPG biosynthesis, governs signal transducer and activator of transcription 3 (STAT3) signaling, a known regulator of cancer cell aggressiveness. Collectively, we present the EXT1/HSPG/STAT3 axis as a central regulator of cancer cell plasticity that directly links proteoglycan synthesis to oncogenic signaling pathways.

Mutually exclusive teams-like patterns of gene regulation characterize phenotypic heterogeneity along the noradrenergic-mesenchymal axis in neuroblastoma.

Neuroblastoma is the most frequent extracranial pediatric tumor and leads to 15% of all cancer-related deaths in children. Tumor relapse and therapy resistance in neuroblastoma are driven by phenotypic plasticity and heterogeneity between noradrenergic (NOR) and mesenchymal (MES) cell states. Despite the importance of this phenotypic plasticity, the dynamics and molecular patterns associated with these bidirectional cell-state transitions remain relatively poorly understood. Here, we analyze multiple RNA-seq datasets at both bulk and single-cell resolution, to understand the association between NOR- and MES-specific factors. We observed that NOR-specific and MES-specific expression patterns are largely mutually exclusive, exhibiting a "teams-like" behavior among the genes involved, reminiscent of our earlier observations in lung cancer and melanoma. This antagonism between NOR and MES phenotypes was also associated with metabolic reprogramming and with immunotherapy targets PD-L1 and GD2 as well as with experimental perturbations driving the NOR-MES and/or MES-NOR transition. Further, these "teams-like" patterns were seen only among the NOR- and MES-specific genes, but not in housekeeping genes, possibly highlighting a hallmark of network topology enabling cancer cell plasticity.

Characterizing heterogeneity along EMT and metabolic axes in colorectal cancer reveals underlying consensus molecular subtype-specific trends.

Colorectal cancer (CRC) is highly heterogeneous with variable survival outcomes and therapeutic vulnerabilities. A commonly used classification system in CRC is the Consensus Molecular Subtypes (CMS) based on gene expression patterns. However, how these CMS categories connect to axes of phenotypic plasticity and heterogeneity remains unclear. Here, in our analysis of CMS-specific TCGA data and 101 bulk transcriptomic datasets, we found the epithelial phenotype score to be consistently positively correlated with scores of glycolysis, OXPHOS and FAO pathways, while PD-L1 activity scores positively correlated with mesenchymal phenotype scoring, revealing possible interconnections among plasticity axes. Single-cell RNA-sequencing analysis of patient samples revealed that that CMS2 and CMS3 subtype samples were relatively more epithelial as compared to CMS1 and CMS4. CMS1 revealed two subpopulations: one close to CMS4 (more mesenchymal) and the other closer to CMS2 or CMS3 (more epithelial), indicating a partial EMT-like behavior. Consistent observations were made in single-cell analysis of metabolic axes and PD-L1 activity scores. Together, our results quantify the patterns of two functional interconnected axes of phenotypic heterogeneity - EMT and metabolic reprogramming - in a CMS-specific manner in CRC.

Spatiotemporal modulation of growth factors directs the generation of multilineage mouse embryonic stem cell-derived mammary organoids.

Ectodermal appendages, such as the mammary gland (MG), are thought to have evolved from hair-associated apocrine glands to serve the function of milk secretion. Through the directed differentiation of mouse embryonic stem cells (mESCs), here, we report the generation of multilineage ESC-derived mammary organoids (MEMOs). We adapted the skin organoid model, inducing the dermal mesenchyme to transform into mammary-specific mesenchyme via the sequential activation of Bone Morphogenetic Protein 4 (BMP4) and Parathyroid Hormone-related Protein (PTHrP) and inhibition of hedgehog (HH) signaling. Using single-cell RNA sequencing, we identified gene expression profiles that demonstrate the presence of mammary-specific epithelial cells, fibroblasts, and adipocytes. MEMOs undergo ductal morphogenesis in Matrigel and can reconstitute the MG in vivo. Further, we demonstrate that the loss of function in placode regulators LEF1 and TBX3 in mESCs results in impaired skin and MEMO generation. In summary, our MEMO model is a robust tool for studying the development of ectodermal appendages, and it provides a foundation for regenerative medicine and disease modeling.

Development of adaptive anoikis resistance promotes metastasis that can be overcome by CDK8/19 Mediator kinase inhibition.

Anoikis resistance or evasion of cell death triggered by cell detachment into suspension is a hallmark of cancer that is concurrent with cell survival and metastasis. The effects of frequent matrix detachment encounters on the development of anoikis resistance in cancer remains poorly defined. Here we show using a panel of ovarian cancer models, that repeated exposure to suspension stress in vitro followed by attached recovery growth leads to the development of anoikis resistance paralleling in vivo development of anoikis resistance in ovarian cancer ascites. This resistance is concurrent with enhanced invasion, chemoresistance and the ability of anoikis adapted cells to metastasize to distant sites. Adapted anoikis resistant cells show a heightened dependency on oxidative phosphorylation and can also evade immune surveillance. We find that such acquired anoikis resistance is not genetic, as acquired resistance persists for a finite duration in the absence of suspension stress. Transcriptional reprogramming is however essential to this process, as acquisition of adaptive anoikis resistance in vitro and in vivo is exquisitely sensitive to inhibition of CDK8/19 Mediator kinase, a pleiotropic regulator of transcriptional reprogramming. Our data demonstrate that growth after recovery from repeated exposure to suspension stress is a direct contributor to metastasis and that inhibition of CDK8/19 Mediator kinase during such adaptation provides a therapeutic opportunity to prevent both local and distant metastasis in cancer.

A systems-level analysis of the mutually antagonistic roles of RKIP and BACH1 in dynamics of cancer cell plasticity.

Epithelial-mesenchymal transition (EMT) is an important axis of phenotypic plasticity-a hallmark of cancer metastasis. Raf kinase-B inhibitor protein (RKIP) and BTB and CNC homology 1 (BACH1) are reported to influence EMT. In breast cancer, they act antagonistically, but the exact nature of their roles in mediating EMT and associated other axes of plasticity remains unclear. Here, analysing transcriptomic data, we reveal their antagonistic trends in a pan-cancer manner in terms of association with EMT, metabolic reprogramming and immune evasion via PD-L1. Next, we developed and simulated a mechanism-based gene regulatory network that captures how RKIP and BACH1 engage in feedback loops with drivers of EMT and stemness. We found that RKIP and BACH1 belong to two antagonistic 'teams' of players-while BACH1 belonged to the one driving pro-EMT, stem-like and therapy-resistant cell states, RKIP belonged to the one enabling pro-epithelial, less stem-like and therapy-sensitive phenotypes. Finally, we observed that low RKIP levels and upregulated BACH1 levels associated with worse clinical outcomes in many cancer types. Together, our systems-level analysis indicates that the emergent dynamics of underlying regulatory network enable the antagonistic patterns of RKIP and BACH1 with various axes of cancer cell plasticity, and with patient survival data.

Multi-modal transcriptomic analysis unravels enrichment of hybrid epithelial/mesenchymal state and enhanced phenotypic heterogeneity in basal breast cancer.

Intra-tumoral phenotypic heterogeneity promotes tumor relapse and therapeutic resistance and remains an unsolved clinical challenge. It manifests along multiple phenotypic axes and decoding the interconnections among these different axes is crucial to understand its molecular origins and to develop novel therapeutic strategies to control it. Here, we use multi-modal transcriptomic data analysis - bulk, single-cell and spatial transcriptomics - from breast cancer cell lines and primary tumor samples, to identify associations between epithelial-mesenchymal transition (EMT) and luminal-basal plasticity - two key processes that enable heterogeneity. We show that luminal breast cancer strongly associates with an epithelial cell state, but basal breast cancer is associated with hybrid epithelial/mesenchymal phenotype(s) and higher phenotypic heterogeneity. These patterns were inherent in methylation profiles, suggesting an epigenetic crosstalk between EMT and lineage plasticity in breast cancer. Mathematical modelling of core underlying gene regulatory networks representative of the crosstalk between the luminal-basal and epithelial-mesenchymal axes recapitulate and thus elucidate mechanistic underpinnings of the observed associations from transcriptomic data. Our systems-based approach integrating multi-modal data analysis with mechanism-based modeling offers a predictive framework to characterize intra-tumor heterogeneity and to identify possible interventions to restrict it.

Dynamical hallmarks of cancer: Phenotypic switching in melanoma and epithelial-mesenchymal plasticity.

Phenotypic plasticity was recently incorporated as a hallmark of cancer. This plasticity can manifest along many interconnected axes, such as stemness and differentiation, drug-sensitive and drug-resistant states, and between epithelial and mesenchymal cell-states. Despite growing acceptance for phenotypic plasticity as a hallmark of cancer, the dynamics of this process remains poorly understood. In particular, the knowledge necessary for a predictive understanding of how individual cancer cells and populations of cells dynamically switch their phenotypes in response to the intensity and/or duration of their current and past environmental stimuli remains far from complete. Here, we present recent investigations of phenotypic plasticity from a systems-level perspective using two exemplars: epithelial-mesenchymal plasticity in carcinomas and phenotypic switching in melanoma. We highlight how an integrated computational-experimental approach has helped unravel insights into specific dynamical hallmarks of phenotypic plasticity in different cancers to address the following questions: a) how many distinct cell-states or phenotypes exist?; b) how reversible are transitions among these cell-states, and what factors control the extent of reversibility?; and c) how might cell-cell communication be able to alter rates of cell-state switching and enable diverse patterns of phenotypic heterogeneity? Understanding these dynamic features of phenotypic plasticity may be a key component in shifting the paradigm of cancer treatment from reactionary to a more predictive, proactive approach.

Synthetic Gene Circuits Combining CRISPR Interference and CRISPR Activation in E. coli: Importance of Equal Guide RNA Binding Affinities to Avoid Context-Dependent Effects.

Gene expression control based on clustered regularly interspaced short palindromic repeats (CRISPR) has emerged as a powerful approach for constructing synthetic gene circuits. While the use of CRISPR interference (CRISPRi) is already well-established in prokaryotic circuits, CRISPR activation (CRISPRa) is less mature, and a combination of the two in the same circuits is only just emerging. Here, we report that combining CRISPRi with SoxS-based CRISPRa in Escherichia coli can lead to context-dependent effects due to different affinities in the formation of CRISPRa and CRISPRi complexes, resulting in loss of predictable behavior. We show that this effect can be avoided by using the same scaffold guide RNA structure for both complexes.

Effects of microRNA-mediated negative feedback on gene expression noise.

MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression post-transcriptionally in eukaryotes by binding with target mRNAs and preventing translation. miRNA-mediated feedback motifs are ubiquitous in various genetic networks that control cellular decision making. A key question is how such a feedback mechanism may affect gene expression noise. To answer this, we have developed a mathematical model to study the effects of a miRNA-dependent negative-feedback loop on mean expression and noise in target mRNAs. Combining analytics and simulations, we show the existence of an expression threshold demarcating repressed and expressed regimes in agreement with earlier studies. The steady-state mRNA distributions are bimodal near the threshold, where copy numbers of mRNAs and miRNAs exhibit enhanced anticorrelated fluctuations. Moreover, variation of negative-feedback strength shifts the threshold locations and modulates the noise profiles. Notably, the miRNA-mRNA binding affinity and feedback strength collectively shape the bimodality. We also compare our model with a direct auto-repression motif, where a gene produces its own repressor. Auto-repression fails to produce bimodal mRNA distributions as found in miRNA-based indirect repression, suggesting the crucial role of miRNAs in creating phenotypic diversity. Together, we demonstrate how miRNA-dependent negative feedback modifies the expression threshold and leads to a broader parameter regime of bimodality compared to the no-feedback case.

Phenotypic heterogeneity drives differential disease outcome in a mouse model of triple negative breast cancer.

The triple negative breast cancer (TNBC) subtype is one of the most aggressive forms of breast cancer that has poor clinical outcome and is an unmet clinical challenge. Accumulating evidence suggests that intratumoral heterogeneity or the presence of phenotypically distinct cell populations within a tumor play a crucial role in chemoresistance, tumor progression and metastasis. An increased understanding of the molecular regulators of intratumoral heterogeneity is crucial to the development of effective therapeutic strategies in TNBC. To this end, we used an unbiased approach to identify a molecular mediator of intratumoral heterogeneity in breast cancer by isolating two tumor cell populations (T1 and T2) from the 4T1 TNBC model. Phenotypic characterization revealed that the cells are different in terms of their morphology, proliferation and self-renewal ability in vitro as well as primary tumor formation and metastatic potential in vivo. Bioinformatic analysis followed by Kaplan Meier survival analysis in TNBC patients identified Metastasis associated colon cancer 1 (Macc1) as one of the top candidate genes mediating the aggressive phenotype in the T1 tumor cells. The role of Macc1 in regulating the proliferative phenotype was validated and taken forward in a therapeutic context with Lovastatin, a small molecule transcriptional inhibitor of Macc1 to target the T1 cell population. This study increases our understanding of the molecular underpinnings of intratumoral heterogeneity in breast cancer that is critical to improve the treatment of women currently living with the highly aggressive TNBC subtype.

Dynamical modeling of proliferative-invasive plasticity and IFNγ signaling in melanoma reveals mechanisms of PD-L1 expression heterogeneity.

BACKGROUND: Phenotypic heterogeneity of melanoma cells contributes to drug tolerance, increased metastasis, and immune evasion in patients with progressive disease. Diverse mechanisms have been individually reported to shape extensive intra-tumor and inter-tumor phenotypic heterogeneity, such as IFNγ signaling and proliferative to invasive transition, but how their crosstalk impacts tumor progression remains largely elusive. METHODS: Here, we integrate dynamical systems modeling with transcriptomic data analysis at bulk and single-cell levels to investigate underlying mechanisms behind phenotypic heterogeneity in melanoma and its impact on adaptation to targeted therapy and immune checkpoint inhibitors. We construct a minimal core regulatory network involving transcription factors implicated in this process and identify the multiple 'attractors' in the phenotypic landscape enabled by this network. Our model predictions about synergistic control of PD-L1 by IFNγ signaling and proliferative to invasive transition were validated experimentally in three melanoma cell lines-MALME3, SK-MEL-5 and A375. RESULTS: We demonstrate that the emergent dynamics of our regulatory network comprising MITF, SOX10, SOX9, JUN and ZEB1 can recapitulate experimental observations about the co-existence of diverse phenotypes (proliferative, neural crest-like, invasive) and reversible cell-state transitions among them, including in response to targeted therapy and immune checkpoint inhibitors. These phenotypes have varied levels of PD-L1, driving heterogeneity in immunosuppression. This heterogeneity in PD-L1 can be aggravated by combinatorial dynamics of these regulators with IFNγ signaling. Our model predictions about changes in proliferative to invasive transition and PD-L1 levels as melanoma cells evade targeted therapy and immune checkpoint inhibitors were validated in multiple RNA-seq data sets from in vitro and in vivo experiments. CONCLUSION: Our calibrated dynamical model offers a platform to test combinatorial therapies and provide rational avenues for the treatment of metastatic melanoma. This improved understanding of crosstalk among PD-L1 expression, proliferative to invasive transition and IFNγ signaling can be leveraged to improve the clinical management of therapy-resistant and metastatic melanoma.

p53 amyloid pathology is correlated with higher cancer grade irrespective of the mutant or wild-type form.

p53 (also known as TP53) mutation and amyloid formation are long associated with cancer pathogenesis; however, the direct demonstration of the link between p53 amyloid load and cancer progression is lacking. Using multi-disciplinary techniques and 59 tissues (53 oral and stomach cancer tumor tissue samples from Indian individuals with cancer and six non-cancer oral and stomach tissue samples), we showed that p53 amyloid load and cancer grades are highly correlated. Furthermore, next-generation sequencing (NGS) data suggest that not only mutant p53 (e.g. single-nucleotide variants, deletions, and insertions) but wild-type p53 also formed amyloids either in the nucleus (50%) and/or in the cytoplasm in most cancer tissues. Interestingly, in all these cancer tissues, p53 displays a loss of DNA-binding and transcriptional activities, suggesting that the level of amyloid load correlates with the degree of loss and an increase in cancer grades. The p53 amyloids also sequester higher amounts of the related p63 and p73 (also known as TP63 and TP73, respectively) protein in higher-grade tumor tissues. The data suggest p53 misfolding and/or aggregation, and subsequent amyloid formation, lead to loss of the tumor-suppressive function and the gain of oncogenic function, aggravation of which might determine the cancer grade.

An androgen receptor regulated gene score is associated with epithelial to mesenchymal transition features in triple negative breast cancers.

BACKGROUND: Androgen receptor (AR) is considered a marker of better prognosis in hormone receptor positive breast cancers (BC), however, its role in triple negative breast cancer (TNBC) is controversial. This may be attributed to intrinsic molecular differences or scoring methods for AR positivity. We derived AR regulated gene score and examined its utility in BC subtypes. METHODS: AR regulated genes were derived by applying a bioinformatic pipeline on publicly available microarray data sets of AR+ BC cell lines and gene score was calculated as average expression of six AR regulated genes. Tumors were divided into AR high and low based on gene score and associations with clinical parameters, circulating androgens, survival and epithelial to mesenchymal transition (EMT) markers were examined, further evaluated in invitro models and public datasets. RESULTS: 53% (133/249) tumors were classified as AR gene score high and were associated with significantly better clinical parameters, disease-free survival (86.13 vs 72.69 months, log rank p = 0.032) when compared to AR low tumors. 36% of TNBC (N = 66) were AR gene score high with higher expression of EMT markers (p = 0.024) and had high intratumoral levels of 5α-reductase, enzyme involved in intracrine androgen metabolism. In MDA-MB-453 treated with dihydrotestosterone, SLUG expression increased, E-cadherin decreased with increase in migration and these changes were reversed with bicalutamide. Similar results were obtained in public datasets. CONCLUSION: Deciphering the role of AR in BC is difficult based on AR protein levels alone. Our results support the context dependent function of AR in driving better prognosis in ER positive tumors and EMT features in TNBC tumors.