Biology subjects

Pruneri, G.

Publications and source records attributed to Pruneri, G..

12 recordsLinked to original sources

bioRxiv2026

ENPP3 expressed by HER2-positive breast cancer cells is associated with good prognosis by restraining epithelial-to-mesenchymal phenotype

Background Ectonucleotide pyrophosphatase/phosphodiesterase 3 (ENPP3/CD203c) is largely studied as a marker of mast cells and basophils. By depleting extracellular ATP, it prevents excessive activation of mast cells and basophils, hence reducing inflammation and allergic reactions. Recent findings have also shown that Enpp3 can deplete cGAMP, another molecule involved in STING activation and IFN-mediated pro-inflammation. Little is still known regarding the role of Enpp3 in non-immune cells although a few reports have described its expression in healthy tissues and tumors. Methods In silico analysis were performed to investigate the expression levels and the prognostic value of Enpp3 in breast cancer, together with ovarian, prostate and colon carcinoma. ENPP3 expression was evaluated in formalin-fixed, paraffin-embedded tumor samples of breast cancer patients by immunohistochemistry, and in mouse mammary cancer cell lines by western blots. Cells were treated with EGFR ligands to stimulate the EGFR/HER2 axis. A mouse-derived mammary cancer cell line was engineered by CRISPR/Cas9 to introduce a GFP sequence under the control of the Enpp3 promoter. GFP-positive and -negative cells were sorted and analyzed by gene expression profiling to identify genes and pathways associated with Enpp3 expression. Finally, wild type and Enpp3 knockout cells were injected in the fat pad of Wsh mice, which do not have mast cells, to evaluate the growth of the tumors which were further analyzed by immunohistochemistry. Results We provide evidence that HER2-positive cells express higher levels of ENPP3 in samples of breast cancer patients. Moreover, in vitro models confirmed that HER2 expression and EGFR stimulation result in up-regulation of Enpp3. We identified pathways that can concur to Enpp3 expression and showed that in vivo the absence of Enpp3 promotes tumor growth and development of tumors with a marked epithelial-to-mesenchymal phenotype. Finally, in a small cohort of HER2-positive breast cancer patients, we found that ENPP3 expression correlates with increased relapse-free survival. Conclusions Despite its potential immunosuppressive role, our findings support the notion that ENPP3 expression is promoted by HER2 in breast cancer, and that it is endowed with a positive prognostic value.

cancer biology↗

bioRxiv2024

Loss of the tumor suppressor NUMB drives aggressive bladder cancer through hyperactivation of a RhoA/ROCK/YAP signaling circuitry

Bladder cancer (BCa) is one of the most challenging and costly cancers to treat, yet little progress has been made on the development of predictive biomarkers and targeted therapies. Here, we uncover a critical function of Numb as a tumor suppressor in the bladder, identifying loss of Numb expression as a causal alteration in BCa that underlies biological aggressiveness and disease progression. Through retrospective cohort studies, we established that a Numb-deficient tumor status correlates with worse overall survival in post-cystectomy muscle-invasive bladder cancer (MIBC) patients and increased risk of MIBC progression in non-muscle-invasive bladder cancer (NMIBC) patients. The prognostic value of Numb loss can be attributed to its crucial role as a determinant of aggressive bladder tumorigenesis, as demonstrated in mouse and human models. Targeted Numb ablation in the basal layer of the urothelium was alone sufficient to trigger spontaneous bladder tumorigenesis and drive progression from preneoplastic to preinvasive and, ultimately, overtly invasive tumors. Additionally, Numb ablation sensitized the urothelium to other oncogenic insults, accelerating tumor onset and progression. Using 3D-Matrigel organoid cultures to recapitulate bladder tumorigenesis in vitro, we found that Numb loss heightens the proliferative and invasive potential of both mouse and human BCa cells. Integrative transcriptomic and functional analyses revealed that downregulation of the canonical Hippo pathway, resulting in enhanced YAP transcriptional activity, underlies the biological aggressiveness of Numb-deficient BCa. These molecular events are dependent on the activation of RhoA/ROCK signaling subsequent to Numb loss. Thus, a dysfunctional Numb-RhoA/ROCK-Hippo/YAP regulatory network is at play in aggressive Numb-deficient BCa and represents a therapeutic vulnerability. A 27-gene prognostic signature capable of identifying high-risk Numb-deficient patients could provide the basis of a clinical tool to stratify patients for innovative RhoA/ROCK/YAP targeted therapies. One Sentence SummaryNumb loss-directed hyperactivation of RhoA/ROCK/YAP underlies aggressive bladder cancer biology.

cancer biology↗

bioRxiv2024

B cell receptor silencing reveals the origin of high-grade B cell lymphomas with MYC and BCL2 rearrangements

The B cell receptor (BCR) is essential for mature B cell lymphomas, serving as therapeutic target. Here, we show that high-grade B cell lymphomas with MYC and BCL2 rearrangements (HGBCL-DH-BCL2) predominantly exhibit immunoglobulin heavy (IGH) chain silencing, leading to BCR shutdown. HGBCL-DH-BCL2 with undetectable IGH (IGHUND) differ from IGH-expressing counterparts for germinal center-zone gene programs, MYC expression and T cell infiltration. While IGH+ HGBCL-DH-BCL2 prefer IGM/IG-Kappa expression, IGHUND counterparts have completed IGH class-switching, favoring IG-Lambda (IGL) light chains. IGHUND HGBCL-DH-BCL2 preserve IGHV gene integrity, overcoming antigen-driven selection. IGH silencing precedes onset and shapes evolution of HGBCL-DH-BCL2 from Follicular Lymphoma (FL) or FL/HGBCL-DH-BCL2 common precursor. In FL/HGBCL-DH-BCL2 pairs and HGBCL-DH-BCL2 models, BCR silencing promoted RAG1/2-dependent IG light chain editing, causing t(8;22)(q24;q11)/IGL::MYC. IGH silencing protected HGBCL-DH-BCL2 models from killing by CD79B-targeting Polatuzumab-Vedotin. Collectively, HGBCL-DH-BCL2 primarily originate from BCR-silenced isotype-switched t(14;18)/IGH::BCL2-positive (pre)FL cells acquiring IGL::MYC translocations during IG light chain revision, with clinical implications. SignificanceThese findings link BCR silencing in isotype-switched t(14;18)+ Follicular Lymphoma cells (or their precursors) to RAG1/2 re-expression, promoting IGL::MYC translocations responsible for transformation into high-grade B cell lymphomas (HGBCL). Predominant silencing of the BCR complex in HGBCL with MYC and BCL2 rearrangements protects tumor cells from CD79B-directed Polatuzumab-Vedotin killing.

cancer biology↗

bioRxiv2022

Landscape of immune-related signatures induced by targeting of different epigenetic regulators in melanoma: implications for immunotherapy

BackgroundInnovative cancer immunotherapy approaches aim at combining immune checkpoint inhibitors with other immunomodulatory agents. Epigenetic regulators can control immune-related genes, therefore targeting them with specific inhibitors may be a potential way forward. Here we identified immune-related signatures induced by four classes of epigenetic drugs in human melanoma cells to define the most promising agent and to understand its biological activity in-vitro, in-vivo and in clinical samples. MethodsHuman melanoma cell lines were characterized for mutational and differentiation profile and treated with inhibitors of DNA methyltransferases (guadecitabine), histone deacetylases (givinostat), bromodomain and extraterminal domain proteins (JQ1 and OTX-015) and enhancer of zeste homolog 2 (GSK126). Drug-specific gene signatures were identified by Clariom S and Nanostring platforms. Modulation of 14 proteins was determined by quantitative western blot. Ingenuity Pathway Analysis (IPA) identified Upstream Regulator (UR) molecules explaining changes in gene expression and biological activity of drugs. Gene set enrichment and IPA were used to test modulation of guadecitabine-specific gene and UR signatures, respectively, in on-treatment tumor biopsies from melanoma patients enrolled in the Phase Ib NIBIT-M4 Guadecitabine + Ipilimumab Trial. ResultsDrug-specific gene and UR signatures were identified for each of the four inhibitors. Immune-related genes were frequently upregulated by guadecitabine, to a lesser extent by givinostat, but downregulated by JQ1 and OTX-015. GSK126 was the least active drug. Treatment of melanoma cells with combination of two epigenetic drugs revealed a dominant effect of guadecitabine and JQ1 on immune-related gene modulation. Drug-specific modulatory profiles were confirmed at the protein level. The guadecitabine-specific UR signature was characterized by activated molecules of the TLR, NF-kB, and IFN innate immunity pathways and was induced in drug-treated melanoma, mesothelioma, hepatocarcinoma cell lines and human melanoma xenografts. Most of the guadecitabine-specific signature genes (n>160) were upregulated in on-treatment tumor biopsies from NIBIT-M4 trial. Progressive activation of guadecitabine UR signature molecules was observed in on-treatment tumor biopsies from responding compared to non-responding patients. ConclusionsGuadecitabine was the most promising immunomodulatory agent among those investigated. This DNA methyltransferases inhibitor emerged as a strong inducer of innate immunity pathways, supporting the rationale for its use in combinatorial immunotherapy approaches.

immunology↗

bioRxiv2022

Genetic and epigenetic driven variation in regulatory regions activitycontribute to adaptation and evolution under endocrine treatment

Comprehensive profiling of hormone-dependent breast cancer (HDBC) has identified hundreds of protein-coding alterations contributing to cancer initiation1, 2, but only a handful have been linked to endocrine therapy resistance, potentially contributing to 40% of relapses1, 3-9. If other mechanisms underlie the evolution of HDBC under adjuvant therapy is currently unknown. In this work, we employ integrative functional genomics to dissect the contribution of cis-regulatory elements (CREs) to cancer evolution by focusing on 12 megabases of non-coding DNA, including clonal enhancers10, gene promoters, and boundaries of topologically associating domains11. Massive parallel perturbation in vitro reveals context-dependent roles for many of these CREs, with a specific impact on dormancy entrance12, 13 and endocrine therapy resistance9. Profiling of CRE somatic alterations in a unique, longitudinal cohort of patients treated with endocrine therapies identifies non-coding changes involved in therapy resistance. Overall, our data uncover actionable transient transcriptional programs critical for dormant persister cells and unveil new regulatory nodes driving evolutionary trajectories towards disease progression.

cancer biology↗

bioRxiv2021

Decellularized Normal and Tumor Scaffolds for Cancer Organoid Cultures as a Model of Colorectal Peritoneal Metastases

Peritoneal metastases (PM) from colorectal cancer (CRC) are associated with poor survival. The extracellular matrix (ECM) plays a fundamental role in modulating the homing of CRC metastases to the peritoneum. The mechanisms underlying the interactions between metastatic cells and the ECM, however, remain poorly understood and the number of in vitro models available for the study of the peritoneal metastatic process is limited. Here, we show that decellularized ECM of the peritoneal cavity allows the growth of organoids obtained from PM, favoring the development of three-dimensional nodules that maintain the characteristics of in vivo PM. Organoids preferentially grow on scaffolds obtained from neoplastic peritoneum, which are characterized by greater stiffness than normal scaffolds. A gene expression analysis of organoids grown on different substrates reflected faithfully the clinical and biological characteristics of the organoids. An impact of the ECM on the response to standard chemotherapy treatment for PM was also observed. SignificanceEvidence of the value of ex vivo 3D models obtained by combining patient-derived extracellular matrices depleted of cellular components and organoids to mimic the metastatic niche, to be used as a tool to develop new therapeutic strategies in a biologically relevant context, to personalize treatments and increase their efficacy.

cancer biology↗

bioRxiv2020

VGLL1-directed TEAD activation drives endocrine therapy resistance in estrogen receptor positive breast cancer

Resistance to endocrine therapies (ET) is common in estrogen receptor (ER) positive breast cancer and most relapsed patients die with ET-resistant disease. While genetic mutations provide explanations for some relapsed patients1, mechanisms of resistance remain undefined in many cases. Drug-induced epigenetic reprogramming provides possible routes to resistance2. By analysing histone H3 lysine 27 acetylation (H3K27ac) profiles in models of ET resistance, we discovered that selective ER down-regulators (SERDs) such as fulvestrant promote epigenetic activation of VGLL1, a co-activator for TEAD transcription factors. We show that VGLL1, acting via TEADs, promotes expression of genes that drive growth of fulvestrant-resistant breast cancer cells. Pharmacological disruption of VGLL1/TEAD4 interaction inhibits VGLL1/TEAD transcriptional programmes to block growth of the resistant cells and prevents growth. Among the VGLL1/TEAD-regulated genes, we identify EGFR, whereby VGLL1-directed EGFR upregulation sensitises fulvestrant-resistant breast cancer cells to EGFR inhibitors. Taken together, our findings identify VGLL1 as a transcriptional driver in ET resistance and advance new therapeutic possibilities for relapsed ER+ breast cancer patients.

cancer biology↗

bioRxiv2018

Single-cell Transcriptomics reveals multi-step adaptations to endocrine therapy

Resistant tumours are thought to arise from the action of Darwinian selection on genetically heterogenous cancer cell populations. However, simple clonal selection is inadequate to describe the late relapses often characterising luminal breast cancers treated with endocrine therapy (ET), suggesting a more complex interplay between genetic and non-genetic factors. Partially, this is due to our limited understanding on the effect of ET at the single cell level. In the present study, we dissect the contributions of clonal genetic diversity and transcriptional plasticity during the early and late phases of ET at single-cell resolution. Using single-cell RNA-sequencing and imaging we disentangle the transcriptional variability of plastic cells and define a rare sub-population of pre-adapted (PA) cells which undergoes further transcriptomic reprogramming and copy number changes to acquire full resistance. PA cells show reduced oestrogen receptor activity but increased features of quiescence and migration. We find evidence for sub-clonal expression of this PA signature in primary tumours and for dominant expression in clustered circulating tumour cells. We propose a multi-step model for ET resistance development and advocate the use of stage-specific biomarkers.

cancer biology↗

bioRxiv2018

A cell-of-origin epigenetic tracer reveals clinically distinct subtypes of high grade serous ovarian cancer

High grade serous ovarian cancer (HGSOC) is a major unmet need in oncology. The persistent uncertainty on its originating tissue has contributed to hamper the discovery of oncogenic pathways and effective therapies. Here we define the DNA methylation print that distinguishes the human fimbrial (FI) and ovarian surface epithelia (OSE) and develop a robust epigenetic cell-of-origin tracer that stratifies HGSOC in FI-and OSE-originated tumors across all available cohorts. We translate this origin-based stratification into a clinically actionable transcriptomic signature, demonstrating its prognostic impact on patients survival and identifying novel network level dysregulations specific for the two disease subtypes.

cancer biology↗

bioRxiv2018

SREBP1 drives KRT80-dependent cytoskeletal changes and invasive behavior in endocrine resistant ERα breast cancer

Approximately 30% of women diagnosed with ER breast cancer relapse with metastatic disease following adjuvant treatment with endocrine therapies1,2. The connection between acquisition of drug resistance and invasive potential is poorly understood. In this study, we demonstrate that the type II keratin topological associating domain (TAD)3 undergoes epigenetic reprogramming in cells that develop resistance to aromatase inhibitors (AI), leading to keratin 80 (KRT80) upregulation. In agreement, an increased number of KRT80-positive cells are observed at relapse in vivo while KRT80 expression associates with poor outcome using several clinical endpoints. KRT80 expression is driven by de novo enhancer activation by sterol regulatory element-binding protein 14 (SREBP1). KRT80 upregulation directly promotes cytoskeletal rearrangements at the leading edge, increased focal adhesion maturation and cellular stiffening, which collectively promote cancer cell invasion. Shear-wave elasticity imaging of prospective patients shows that KRT80 levels correlate with stiffer tumors in vivo. Collectively, our data uncover an unpredicted and potentially targetable direct link between epigenetic and cytoskeletal reprogramming promoting cell invasion in response to chronic AI treatment.

cancer biology↗

bioRxiv2018

WDR5 regulates epithelial-to-mesenchymal transition in breast cancer cells via TGFB

Even if the mortality rate in breast cancer (BC) has recently decreased, development of metastases and drug resistance are still challenges to successful systemic treatment. The epithelial-to-mesenchymal transition (EMT), as well as epigenetic dynamic modifications, plays a pivotal role in invasion, metastasis, and drug resistance. Here, we report that WDR5, the core subunit of histone H3 K4 methyltransferase complexes, is crucial in coordinating EMT and regulating epigenetic changes that drive metastasis. We show that silencing of WDR5 in BC up-regulates an epithelial signature in triple negative and luminal B like patients by transcriptional repression of mesenchymal genes and reduction of the metastatic properties of these cells. Moreover, we demonstrate that this regulation is mediated by inhibition of the TGF{beta} signaling both at the transcriptional and post-translational level, suggesting an active role of WDR5 in guiding tumor plasticity upon oncogenic insults, regardless of the pathological BC subtypes.\n\nWe therefore suggest that WDR5 inhibition could be a successful pharmacologic approach to inhibit EMT and sensitize breast cancer cells to chemotherapy.

cancer biology↗

bioRxiv2017

Enhancers mapping uncovers phenotypic heterogeneity and evolution in patients with luminal breast cancer.

The degree of intrinsic and interpatient phenotypic heterogeneity and its role in tumour evolution is poorly understood. Phenotypic divergence can be achieved via the inheritance of alternative transcriptional programs1,2. Cell-type specific transcription is maintained through the activation of epigenetically-defined regulatory regions including promoters and enhancers1,3,4. In this work, we annotated the epigenome of 47 primary and metastatic oestrogen-receptor (ER)-positive breast cancer specimens from clinical samples, and developed strategies to deduce phenotypic heterogeneity from the regulatory landscape, identifying key regulatory elements commonly shared across patients. Highly shared regions contain a unique set of regulatory information including the motif for the transcription factor YY1. In vitro work shows that YY1 is essential for ER transcriptional activity and defines the critical subset of functional ER binding sites driving tumor growth in most luminal patients. YY1 also control the expression of genes that mediate resistance to endocrine treatment. Finally, we show that H3K27ac levels at active enhancer elements can be used as a surrogate of intra-tumor phenotypic heterogeneity, and to track expansion and contraction of phenotypic subpopulations throughout breast cancer progression. Tracking YY1 and SLC9A3R1 positive clones in primary and metastatic lesions, we show that endocrine therapies drive the expansion of phenotypic clones originally underrepresented at diagnosis. Collectively, our data show that epigenetic mechanisms significantly contribute to phenotypic heterogeneity and evolution in systemically treated breast cancer patients.

cancer biology↗