Biology subjects

Demichelis, F.

Publications and source records attributed to Demichelis, F..

14 recordsLinked to original sources

bioRxiv2026

The RNA cargo of plasma-derived extracellular vesicles in mCRPC patients captures cancer cells and tumor microenvironment signals

Circulating analytes in cancer patients capture tumor-related signals. We profiled matched extracellular vesicle (EV) total RNA and cell-free DNA (cfDNA) from the same plasma aliquots of chemo-naive metastatic castration-resistant prostate cancer (mCRPC) patients treated with Enzalutamide (n=54 patients, n=119 longitudinal samples; NCT06981377) at four Italian clinical centers and interrogated data from >10,000 cancer patients' and healthy individuals' samples. Transcript integrity analysis identified coding and non-coding species with fragmentation patterns varying across RNA biotypes. EV-RNA data deconvolution revealed signal from immune populations, including fractions classified as CD4+ T cells, whose abundance increased with disease progression in plasma EVs and mCRPC tissues. By leveraging tissue data-informed mining, we established a novel prostate cancer-related EV-RNA signature that resulted in an independent predictor of poor prognosis and captured tumor microenvironment-derived signals. Integrating EV-RNA and ctDNA information improved patient stratification for progression-free survival. These findings suggest a multifaceted role for plasma EVs as a source of cancer biomarkers.

cancer biology↗

bioRxiv2025

T2T-CHM13 reference genome reduces mapping bias and enhances alignment accuracy at disease-associated variants

1The T2T-CHM13v2.0 reference genome added previously uncharacterized genomic sequences and improved the accuracy of repetitive stretches compared to former human genome assemblies. By comprehensive allelic variation analysis and read mapping statistics from sequencing reads aligned to hg38 and T2T-CHM13 assemblies in samples encompassing different sequencing designs and ethnicity groups, we observed that T2T-CHM13v2.0 assembly significantly reduces the reference mapping bias (RMB) and increases read mapping precision at clinically relevant sites, including BRCA1 pathogenic variants. Further, we report the presence of sequence dissimilarities among reference genomes in the proximity of ClinVar annotated variants, suggesting the need for data re-analysis and potential redesign of probes targeting clinically relevant regions. Overall, these findings support the implementation of T2T-CHM13 reference for the improvement of sequencing data analyses in the clinical genomic setting.

genomics↗

bioRxiv2024

Extracellular Vesicles heterogeneity through the lens of multiomics

Extracellular vesicles (EVs) are heterogenous in size, biogenesis, cargo and function. Beside small EVs, aggressive tumor cells release a population of particularly large EVs, namely large oncosomes (LO). This study provides the first resource of label-free quantitative proteomics of LO and small EVs obtained from distinct cancer cell types (prostate, breast, and glioma). This dataset was integrated with a SWATH Proteomic assay on LO, rigorously isolated from the plasma of patients with metastatic prostate cancer (PC). Proteins enriched in LO, which were identified also at the RNA level, and found in plasma LO significantly correlated with PC progression. Single EV RNA-Seq of the PC cell-derived LO confirmed some of the main findings from the bulk RNA-Seq, providing first evidence that single cell technologies can be successfully applied to EVs. This multiomics resource of cancer EVs can be leveraged for developing a multi-analyte approach for liquid biopsy.

cancer biology↗

bioRxiv2024

Sensitive tumor detection, accurate quantification, and cancer subtype classification using low-pass whole methylome sequencing of plasma DNA

The analysis of circulating tumor DNA (ctDNA) is increasingly used for monitoring disease in patients with metastatic cancer. Here, we introduce a robust and reproducible strategy combining low-pass whole methylome sequencing of plasma DNA with METER, a novel computational tool. Engaging prediction models trained on independent available datasets, METER enables the detection and quantification of tumor content (TC) and performs molecular cancer subtyping. Applied to plasma methylomes from early metastatic breast cancer patients, our method demonstrated reliable quantification, sensitive tumor detection below 3% of TC, and the ability to perform accurate Estrogen Receptor (ER) subtyping. METER provided clinically relevant predictions, underscored by associations with relevant prognostic factors, robust correlation with matched circulating tumor cells, and highly correlated with patients outcomes in challenging scenarios as TC<3%. Our strategy provides comprehensive and sensitive analysis of plasma samples, serving as a valuable yet cost-effective precision oncology tool.

cancer biology↗

bioRxiv2024

ANP32E drives vulnerability to ATR inhibitors by inducing R-loops-dependent Transcription Replication Conflicts in Triple Negative Breast Cancer

Oncogene-induced replicative stress (RS) plays a central role in tumor progression, leading to genomic instability by eliciting transcription replication conflicts (TRCs), which represent the major source of R-loops, that ultimately favors the onset of the DNA damage response (DDR). We investigated the pathogenic contribution of chromatin factors in increasing TRCs and R-loop frequencies in cancer. We found that in breast cancer patients the concomitant upregulation of MYC and the H2A.Z-specific chaperone ANP32E correlated with an increase genome instability. Genome-wide profiling revealed that the ANP32E-dependent increases turnover of H2A.Z altered RNApol II processivity, leading to accumulation of long R-loops at TRCs. We showed that ANP32E upregulation increases TRCs and activates an ATR-dependent DDR, which predispose cancer cells to R-loop-mediated genomic fragility. By exploiting the vulnerability of ANP32E-expressing cancer cells to ATR inhibitors, we found that tumors relied on this DDR pathway, whose inhibition halted their pro-metastatic capacity.

cancer biology↗

bioRxiv2023

Integrating extracellular vesicle and circulating cell-free DNA analysis on a single plasma aliquot from breast cancer patients improves the detection of HER2 positivity

BackgroundMulti-analyte liquid biopsies represents an emerging opportunity for non-invasive cancer assessment. We developed ONCE (ONe Aliquot for Circulating Elements), a novel multi-analytes liquid biopsy approach for the isolation of extracellular vesicles (EVs) and cell-free DNA (cfDNA) from a single aliquot of blood. MethodsWe assessed ONCE performance to classify HER2-positive early-stage breast cancer (BrCa) patients by combining RNA and DNA signals on n=64 healthy donors (HD) and non-metastatic BrCa patients. Specifically, we investigated EVs-derived RNA (EV-RNA) and cfDNA by next-generation sequencing (NGS) and by digital droplet PCR (ddPCR). Additionally, we utilized imaging flow cytometry to evaluate EVs as potential carriers of the HER2 protein. ResultsWestern blot analysis and immunocapture assay revealed that EVs-enriched proteins were detected at similar levels among the HER2+ and HER2- subtypes. Sequencing of cfDNA and EV-RNA from HER2- and HER2+ patients demonstrated concordance with in situ molecular analyses of matched tissues. Combined analysis of the two circulating analytes by ddPCR showed increased sensitivity in ERBB2/HER2 detection compared to single nucleic acid components. Multi-analyte liquid biopsy prediction performance was comparable to tissue-based sequencing results from TCGA. Also, we observed HER2 protein on the surface of EVs isolated from the HER2+ BrCa plasma, thus corroborating the potential relevance of studying EVs as companion analyte to cfDNA. ConclusionsThis data confirms the relevance of combining cfDNA and EV-RNA analytes for cancer assessment and supports the ONCE approach as a valuable tool for multi-analytes liquid biopsies clinical implementation.

cancer biology↗

bioRxiv2022

Bladder cancer organoids as a functional system to model different disease stages and therapy response

Bladder Cancer (BLCa) inter-patient heterogeneity is considered the primary cause of tumor reoccurrence and treatment failure, suggesting that BLCa patients could benefit from a more personalized treatment approach. Patient-derived organoids (PDOs) have been successfully used as a functional model for predicting drug response in different cancer types. In our study, we established BLCa PDO cultures from different BLCa stages. BLCa PDOs preserve the histological and molecular heterogeneity of the parental tumors, including their multiclonal genetic landscapes. BLCa PDOs consistently share key genetic alterations detected in parental tumors, mirroring tumor evolution in longitudinal sampling. Our drug screening pipeline was implemented using BLCa PDOs, testing both standard-of-care and additional FDA-approved compounds for other solid tumors. Integrative analysis of drug response profiles with matched PDO genomic analysis was used to determine enrichment thresholds for candidate markers of therapy resistance and sensitivity. By assessing the clinical history of longitudinally sampled cases, the clonal evolution of the disease could be determined and matched with drug response profiles. In conclusion, we have developed a clinically relevant pipeline for drug response profile assessment and discovery of candidate markers of therapy resistance.

cancer biology↗

bioRxiv2020

Cancer methylomes characterization enabled by Rocker-meth

Differentially DNA methylated regions (DMRs) inform on the role of epigenetic changes in cancer. We present Rocker-meth, a computational method exploiting a heterogeneous hidden Markov model to detect DMRs across multiple experimental platforms. Its application to more than 6,000 methylation profiles across 14 tumor types provides a comprehensive catalog of tumor type-specific and shared DMRs, also amenable to single-cell DNA-methylation data. In depth integrative analysis including orthogonal omics shows the enhanced ability of Rocker-meth in recapitulating known associations, further uncovering the pan-cancer relationship between DNA hypermethylation and transcription factor deregulation depending on the baseline chromatin state.

cancer biology↗

bioRxiv2020

CHD8 Suppression Impacts on Histone H3 Lysine 36 Trimethylation and Alters RNA Alternative Splicing.

Disruptive mutations in the chromodomain helicase DNA binding protein 8 (CHD8) have been recurrently associated with Autism Spectrum Disorders (ASD). In normal cellular physiology, CHD8 co-purifies with MLL1 and MOF transcriptional activation complex, with elongating RNAPII and directly binds to DNA promoters and enhancers regions, thus a regulatory role in transcriptional initiation and elongation could be postulated. Here we investigated how chromatin landscape reacts to CHD8 suppression by analyzing a panel of histone modifications in induced pluripotent stem cell-derived neural progenitors. We interrogated transcriptionally active and repressed regions, as well as active and poised enhancers. CHD8 suppression led to significant reduction (47.82%) in histone H3K36me3 peaks at gene bodies, particularly impacting on transcriptional elongation chromatin states. H3K36me3 reduction specifically affects highly expressed, CHD8-bound genes. Histone H3K36me3 reduction associated to CHD8-suppression does not functionally impact on global transcriptional levels, but correlated with altered alternative splicing patterns of [~] 2000 protein coding genes implicated in "RNA splicing", "mitotic cell cycle phase transition" and "mRNA processing", especially affecting alternative first exon and exon skipping events. In summary, our results point toward broad molecular consequences of CHD8 suppression, implicating altered histone deposition/maintenance and RNA processing regulation as important regulatory processes in ASD.

genetics↗

bioRxiv2020

Role of Specialized mSWI/SNF Complexes in Prostate Cancer Lineage Plasticity

Advanced prostate cancer initially responds to hormonal treatment, but ultimately becomes resistant and requires more potent therapies. One mechanism of resistance observed in [~]10% of these patients is through lineage plasticity, which manifests in a partial or complete small cell or neuroendocrine prostate cancer (NEPC) phenotype. Here, we investigate the role of the mammalian SWI/SNF (mSWI/SNF) chromatin remodeling complex in NEPC. Using large patient datasets, patient-derived organoids and cancer cell lines, we identify mSWI/SNF subunits that are deregulated in NEPC and demonstrate that SMARCA4 (BRG1) overexpression is associated with aggressive disease. We also show that SWI/SNF complexes interact with different lineage-specific factors in NEPC compared to prostate adenocarcinoma. These data suggest a role for mSWI/SNF complexes in therapy-related lineage plasticity, which may be relevant for other solid tumors.

cancer biology↗

bioRxiv2019

Integrative genomic, transcriptomic, and epigenomic analyses of benign prostatic hyperplasia reveal new options for therapy

Benign prostatic hyperplasia (BPH), a nonmalignant enlargement of the prostate, is one of the most common diseases affecting aging men, but the underlying molecular features of BPH remain poorly understood, and therapeutic options are limited. Here we employed a comprehensive molecular investigation of BPH, including genomic, transcriptomic and epigenetic profiling of 18 BPH cases. At the molecular level, we found no evidence of neoplastic features in BPH: no evidence of driver genomic alterations, including low coding mutation rates, mutational signatures consistent with aging tissues, minimal copy number alterations, and no genomic rearrangements. Similarly at the epigenetic level, we found global hypermethylation was the dominant process (unlike most neoplastic processes). By integrating transcriptional and methylation signatures, we identified two BPH subgroups with distinct clinical features and associated signaling pathways, which were validated in two independent cohorts. Finally, our analyses nominated mTOR inhibitors as a potential subtype-specific therapeutic option. Supporting this, a cohort of men exposed to mTOR inhibitors showed a significant decrease in prostate size. Our results demonstrate that BPH consists of distinct molecular subgroups, with potential for subtype-specific precision therapy via mTOR inhibition.

genomics↗

bioRxiv2018

Proteomic and Genomic Signatures of Repeat-instability in Cancer and Adjacent Normal Tissues

Repetitive sequences are hotspots of evolution at multiple levels. However, due to technical difficulties involved in their assembly and analysis, the role of repeats in tumor evolution is poorly understood. We developed a rigorous motif-based methodology to quantify variations in the repeat content of proteomes and genomes, directly from proteomic and genomic raw sequence data, and applied it to analyze a wide range of tumors and normal tissues. We identify high similarity between the repeat-instability in tumors and their patient-matched normal tissues, but also tumor-specific signatures, both in protein expression and in the genome, that strongly correlate with cancer progression and robustly predict the tumorigenic state. In a patient, the hierarchy of genomic repeat instability signatures accurately reconstructs tumor evolution, with primary tumors differentiated from metastases. We find an inverse relationship between repeat-instability and point mutation load, within and across patients, and independently of other somatic aberrations. Thus, repeat-instability is a distinct, transient and compensatory adaptive mechanism in tumor evolution.

cancer biology↗

bioRxiv2017

Discovery and reporting of clinically-relevant germline variants in advanced cancer patients assessed using whole-exome sequencing

PurposeIn precision cancer care, WES-based analysis of tumor-normal samples helps reveal somatic alterations but can also identify cancer-associated germline variants important for disease surveillance, treatment choice and cancer prevention. WES can also identify germline secondary findings impacting risk of cardiac, neurodegenerative or metabolic diseases. In patients with advanced cancer, the frequency of reportable secondary findings encountered with WES is not well defined.\n\nMethodsTo address this question, we analyzed a cohort of 343 patients with advanced, metastatic cancer for whom we have performed tumor and germline WES interrogating more than 21,000 genes using a CLIA/CLEP approved assay.\n\nResults17% of patients in our cohort have one or more reportable germline variants, including patients with pathogenic variants in the BRCA1 and BRCA2 genes. The frequency of non-cancer clinically relevant germline variants (8.8%) was within the range of two control non-cancer cohorts (11.0% and 6.5%). The frequency of variants in cancer-associated genes was significantly higher (p<0.0005) in our advanced cancer cohort (8.2%) compared to control cohorts (2.7% and 3.8%). More than 50% of patients with reportable germline cancer variants had a family history of cancer.\n\nConclusionthese results stress the importance of returning germline results found during somatic genomic tumor testing.

genomics↗

bioRxiv2016

EthSEQ: ethnicity annotation from whole exome sequencing data

Whole exome sequencing (WES) is widely utilized both in translational cancer genomics studies and in the setting of precision medicine. Stratification of individual's ethnicity is fundamental for the correct interpretation of personal genomic variation impact. We implemented EthSEQ to provide reliable and rapid ethnicity annotation from whole exome sequencing individual's data and validated it on 1,000 Genome Project and TCGA data demonstrating high precision (>99%). EthSEQ can be integrated into any WES based processing pipeline and exploits multi-core capabilities. Source code, manual and other data is available at http://demichelislab.unitn.it/EthSEQ.

bioinformatics↗