Pub Date : 2026-01-03DOI: 10.1016/j.jlb.2025.100454
Annarita Nappi , Felice Crocetto , Paolo Conforti , Serena Sagliocchi , Annunziata Gaetana Cicatiello , Federica Restolfer , Lucia Acampora , Silvia Del Mastro , Rosa Sirica , Lorenzo Spirito , Francesco Del Giudice , Roberto La Rocca , Daniela Terracciano , Monica Dentice , Caterina Miro
Background
D2 overexpression has emerged as a recurrent molecular feature across multiple cutaneous malignancies, where it contributes to aberrant Thyroid Hormone (TH) activation and tumor-associated metabolic reprogramming. Liquid biopsy approaches based on circulating cell-free RNA (cfRNA) is emerging as non-invasive strategy to profile gene expression alterations and support dynamic monitoring of transcriptional changes during disease progression.
Methods
We analyzed 54 plasma samples from patients with BLadder CAncer (BLCA) alongside an equivalent cohort of healthy control individuals. Circulating D2 transcripts were quantified after RNA isolation using a modified phenol-chloroform extraction protocol adapted for low-input plasma samples to maximize retrieval of circulating RNA.
Results
D2 transcripts were readily detectable in plasma and showed significantly higher levels in BLCA patients compared with healthy controls. Circulating expression of classical urothelial markers GATA3 and UPK3A, as well as Epithelial-to-Mesenchymal Transition (EMT)-related genes (E-Cadherin, N-Cadherin, Vimentin), was likewise increased in BLCA plasma. However, correlation analyses revealed that D2 expression varied independently from GATA3 and UPK3A across both tumor and non-tumor groups.
Conclusions
These findings demonstrate that D2 is detectably elevated in the circulation of BLCA patients and captures tumor-associated transcriptional alterations that are independent of established urothelial markers. The distinct, non-redundant behavior of circulating D2 supports its potential value as a complementary biomarker for minimally invasive molecular profiling of BLCA. Further studies are required to define its diagnostic performance and clinical applicability.
{"title":"Cell-free RNA profiling uncovers non-canonical circulating D2 transcript elevation in Bladder Cancer plasma","authors":"Annarita Nappi , Felice Crocetto , Paolo Conforti , Serena Sagliocchi , Annunziata Gaetana Cicatiello , Federica Restolfer , Lucia Acampora , Silvia Del Mastro , Rosa Sirica , Lorenzo Spirito , Francesco Del Giudice , Roberto La Rocca , Daniela Terracciano , Monica Dentice , Caterina Miro","doi":"10.1016/j.jlb.2025.100454","DOIUrl":"10.1016/j.jlb.2025.100454","url":null,"abstract":"<div><h3>Background</h3><div>D2 overexpression has emerged as a recurrent molecular feature across multiple cutaneous malignancies, where it contributes to aberrant Thyroid Hormone (TH) activation and tumor-associated metabolic reprogramming. Liquid biopsy approaches based on circulating cell-free RNA (cfRNA) is emerging as non-invasive strategy to profile gene expression alterations and support dynamic monitoring of transcriptional changes during disease progression.</div></div><div><h3>Methods</h3><div>We analyzed 54 plasma samples from patients with BLadder CAncer (BLCA) alongside an equivalent cohort of healthy control individuals. Circulating D2 transcripts were quantified after RNA isolation using a modified phenol-chloroform extraction protocol adapted for low-input plasma samples to maximize retrieval of circulating RNA.</div></div><div><h3>Results</h3><div>D2 transcripts were readily detectable in plasma and showed significantly higher levels in BLCA patients compared with healthy controls. Circulating expression of classical urothelial markers GATA3 and UPK3A, as well as Epithelial-to-Mesenchymal Transition (EMT)-related genes (E-Cadherin, N-Cadherin, Vimentin), was likewise increased in BLCA plasma. However, correlation analyses revealed that D2 expression varied independently from GATA3 and UPK3A across both tumor and non-tumor groups.</div></div><div><h3>Conclusions</h3><div>These findings demonstrate that D2 is detectably elevated in the circulation of BLCA patients and captures tumor-associated transcriptional alterations that are independent of established urothelial markers. The distinct, non-redundant behavior of circulating D2 supports its potential value as a complementary biomarker for minimally invasive molecular profiling of BLCA. Further studies are required to define its diagnostic performance and clinical applicability.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"11 ","pages":"Article 100454"},"PeriodicalIF":0.0,"publicationDate":"2026-01-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145979155","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2026-01-03DOI: 10.1016/j.jlb.2026.100455
Tobin E. Groth, Andrew A. Mishin, Varsha Rao, Ruby Tibet, Christopher J. Troll
Cell-free DNA methylation sequencing provides insight into tissue of origin and chromatin structure. In some workflows, generating libraries includes end-repair. Using matched single-stranded and double-stranded libraries prepared from the same cfDNA extracts, we show that end-repair in double-stranded DNA libraries reduces globally inferred CpG methylation leading to decreased tissue of origin accuracy. Trimming read termini partially mitigates this bias but decreases coverage and removes fragmentomic information compared to single-stranded DNA libraries, which forego end-repair.
{"title":"End-repair causes methylation underestimation in cell-free DNA sequencing libraries","authors":"Tobin E. Groth, Andrew A. Mishin, Varsha Rao, Ruby Tibet, Christopher J. Troll","doi":"10.1016/j.jlb.2026.100455","DOIUrl":"10.1016/j.jlb.2026.100455","url":null,"abstract":"<div><div>Cell-free DNA methylation sequencing provides insight into tissue of origin and chromatin structure. In some workflows, generating libraries includes end-repair. Using matched single-stranded and double-stranded libraries prepared from the same cfDNA extracts, we show that end-repair in double-stranded DNA libraries reduces globally inferred CpG methylation leading to decreased tissue of origin accuracy. Trimming read termini partially mitigates this bias but decreases coverage and removes fragmentomic information compared to single-stranded DNA libraries, which forego end-repair.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"11 ","pages":"Article 100455"},"PeriodicalIF":0.0,"publicationDate":"2026-01-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145928179","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-24DOI: 10.1016/j.jlb.2025.100453
Valerio Gristina , Umberto Malapelle , Gennaro Daniele , Giovanni Maria Iannantuono , Tancredi Didier Bazan Russo , Rossana Berardi , Giordano Domenico Beretta , Ettore Domenico Capoluongo , Marcello Ciaccio , Romano Danesi , Marzia Del Re , Matteo Fassan , Giuseppe Giuffrè , Stefania Gori , Lorena Incorvaia , Antonio Marchetti , Nicola Normanno , Carmine Pinto , Daniele Santini , Andrea Sartore Bianchi , Antonio Russo
Liquid biopsy (LB) offers a minimally invasive alternative to tissue biopsy by detecting tumor-derived analytes in biological fluids. Nonetheless, its adoption is limited by variability in methodologies. Therefore, this study used a modified RAND/UCLA approach involving 23 experts of the field, providing two questionnaires that assessed agreement on 22 items. Consensus was reached for all the pre- and post-analytical phase items, agreeing on the pivotal role of plasma cfDNA. Conversely, opinions varied regarding other biomarkers and biological samples. Furthermore, turnaround time and disease setting resulted as two of the most important analytical parameters for choosing testing methodology. Particularly, a complementary tissue-liquid approach with sampling interval ≤2 weeks was preferred. To conclude, a strong consensus on sample handling, biomarker prioritization, and clinical applications is achieved. However, significant heterogeneity remains regarding novel biomarkers, sampling strategies, and costs. Standardization and validation are needed to enhance the clinical adoption of LB.
{"title":"Liquid biopsy in Oncology: Results of a Delphi consensus study endorsed by the AIOM-SIAPEC/IAP-SIBioC-SIF Italian scientific societies","authors":"Valerio Gristina , Umberto Malapelle , Gennaro Daniele , Giovanni Maria Iannantuono , Tancredi Didier Bazan Russo , Rossana Berardi , Giordano Domenico Beretta , Ettore Domenico Capoluongo , Marcello Ciaccio , Romano Danesi , Marzia Del Re , Matteo Fassan , Giuseppe Giuffrè , Stefania Gori , Lorena Incorvaia , Antonio Marchetti , Nicola Normanno , Carmine Pinto , Daniele Santini , Andrea Sartore Bianchi , Antonio Russo","doi":"10.1016/j.jlb.2025.100453","DOIUrl":"10.1016/j.jlb.2025.100453","url":null,"abstract":"<div><div>Liquid biopsy (LB) offers a minimally invasive alternative to tissue biopsy by detecting tumor-derived analytes in biological fluids. Nonetheless, its adoption is limited by variability in methodologies. Therefore, this study used a modified RAND/UCLA approach involving 23 experts of the field, providing two questionnaires that assessed agreement on 22 items. Consensus was reached for all the pre- and post-analytical phase items, agreeing on the pivotal role of plasma cfDNA. Conversely, opinions varied regarding other biomarkers and biological samples. Furthermore, turnaround time and disease setting resulted as two of the most important analytical parameters for choosing testing methodology. Particularly, a complementary tissue-liquid approach with sampling interval ≤2 weeks was preferred. To conclude, a strong consensus on sample handling, biomarker prioritization, and clinical applications is achieved. However, significant heterogeneity remains regarding novel biomarkers, sampling strategies, and costs. Standardization and validation are needed to enhance the clinical adoption of LB.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"11 ","pages":"Article 100453"},"PeriodicalIF":0.0,"publicationDate":"2025-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145852439","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01DOI: 10.1016/j.jlb.2025.100448
Christian D. Rolfo , Moh'd M. Khushman , Alessandro Russo , Roberto Borea , Bernard Herrman , Jing Wang , Jiemin Liao , Carin R. Espenschied , Sean Gordon , Katie Quinn , Kimberly C. Banks , Arielle J. Medford
{"title":"Corrigendum to “Chemotherapy response monitoring with DNA methylation-based ctDNA tumor fraction: Evidence from a real-world cohort of patients with advanced common solid malignancies” [J Liq Biopsy, 10 (2025), 100442]","authors":"Christian D. Rolfo , Moh'd M. Khushman , Alessandro Russo , Roberto Borea , Bernard Herrman , Jing Wang , Jiemin Liao , Carin R. Espenschied , Sean Gordon , Katie Quinn , Kimberly C. Banks , Arielle J. Medford","doi":"10.1016/j.jlb.2025.100448","DOIUrl":"10.1016/j.jlb.2025.100448","url":null,"abstract":"","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"10 ","pages":"Article 100448"},"PeriodicalIF":0.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145799753","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-12-01DOI: 10.1016/j.jlb.2025.100449
Nadia Ghazali , Ana Ortega-Franco , Diego de Miguel-Perez , Erick F. Saldanha , Konstantinos Venetis , Pasquale Pisapia , Surbhi Singhal , Benjamin A. Bleiberg , Carolina Reduzzi , Roberto Borea
The role of liquid biopsy in oncological care continues to expand, with multiple studies presented at the International Association for the Study of Lung Cancer (IASLC) 2025 World Conference on Lung Cancer (WCLC 2025). This review summarizes recent advances in liquid biopsy for thoracic oncology, encompassing both non–small cell lung cancer (NSCLC) and small cell lung cancer (SCLC).
In early detection and screening, proteomic profiling has identified potential biomarkers predictive of future lung cancer risk. The integration of proteomics with clinical and imaging data can improve pulmonary nodule malignancy prediction. In resectable NSCLC, tumour-informed whole-genome sequencing (WGS) assay demonstrated high sensitivity for minimal residual disease (MRD) detection, with MRD clearance following neoadjuvant osimertinib or chemo-immunotherapy associated with favorable outcomes.
In advanced NSCLC, longitudinal liquid biopsy analyses reveal dynamic subclonal evolution driving early treatment resistance. Circulating tumor DNA (ctDNA) clearance following targeted therapy in MET exon 14 skipping and BRAF-mutated tumors was associated with improved clinical outcomes. Emerging biomarkers such as ctDNA tumour fraction and circulating microRNA signatures are promising for radiotherapy stratification and prediction of immunotherapy-related toxicities.
In SCLC, MRD monitoring enables earlier detection of disease progression and supports ctDNA-guided selection of patients for consolidation immunotherapy following chemotherapy. Overall, these advances demonstrate the expanding role of liquid biopsy in improving early detection, guiding treatment, and improving disease monitoring in lung cancer.
{"title":"Cancer in a drop: Liquid biopsy highlights from the World Conference on Lung Cancer (WCLC) 2025","authors":"Nadia Ghazali , Ana Ortega-Franco , Diego de Miguel-Perez , Erick F. Saldanha , Konstantinos Venetis , Pasquale Pisapia , Surbhi Singhal , Benjamin A. Bleiberg , Carolina Reduzzi , Roberto Borea","doi":"10.1016/j.jlb.2025.100449","DOIUrl":"10.1016/j.jlb.2025.100449","url":null,"abstract":"<div><div>The role of liquid biopsy in oncological care continues to expand, with multiple studies presented at the International Association for the Study of Lung Cancer (IASLC) 2025 World Conference on Lung Cancer (WCLC 2025). This review summarizes recent advances in liquid biopsy for thoracic oncology, encompassing both non–small cell lung cancer (NSCLC) and small cell lung cancer (SCLC).</div><div>In early detection and screening, proteomic profiling has identified potential biomarkers predictive of future lung cancer risk. The integration of proteomics with clinical and imaging data can improve pulmonary nodule malignancy prediction. In resectable NSCLC, tumour-informed whole-genome sequencing (WGS) assay demonstrated high sensitivity for minimal residual disease (MRD) detection, with MRD clearance following neoadjuvant osimertinib or chemo-immunotherapy associated with favorable outcomes.</div><div>In advanced NSCLC, longitudinal liquid biopsy analyses reveal dynamic subclonal evolution driving early treatment resistance. Circulating tumor DNA (ctDNA) clearance following targeted therapy in MET exon 14 skipping and BRAF-mutated tumors was associated with improved clinical outcomes. Emerging biomarkers such as ctDNA tumour fraction and circulating microRNA signatures are promising for radiotherapy stratification and prediction of immunotherapy-related toxicities.</div><div>In SCLC, MRD monitoring enables earlier detection of disease progression and supports ctDNA-guided selection of patients for consolidation immunotherapy following chemotherapy. Overall, these advances demonstrate the expanding role of liquid biopsy in improving early detection, guiding treatment, and improving disease monitoring in lung cancer.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"10 ","pages":"Article 100449"},"PeriodicalIF":0.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145693792","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Liquid biopsy can expose spatially segregated resistance biology that is invisible to single-site tissue testing, particularly across the blood–brain barrier. We report how cerebrospinal fluid (CSF) circulating tumor DNA (ctDNA) clarified therapeutic direction in EGFR-mutated non-small-cell lung cancer (NSCLC) with leptomeningeal involvement. A 67-year-old woman with EGFR exon 19–deleted adenocarcinoma received afatinib followed by long-term osimertinib. After three years, progression of the primary lesion prompted rebiopsy, which revealed a CCDC6-RET fusion with strong RET immunoreactivity. Selpercatinib monotherapy yielded minor thoracic shrinkage at 1 month but was followed by dizziness and MRI evidence of leptomeningeal enhancement at 3 months. CSF analysis showed pleocytosis without malignant cells. Critically, CSF ctDNA demonstrated the EGFR E746_A750 deletion at 29.2 % variant allele frequency by amplicon sequencing, whereas the CCDC6–RET fusion was undetectable by targeted sequencing and highly sensitive single-plex qPCR using junction-specific primers. Re-challenging with osimertinib rapidly improved symptoms and led to resolution of leptomeningeal enhancement. These data indicate clonal divergence at acquired resistance: a RET-fusion clone dominated the thoracic compartment while EGFR-addicted clones predominated in the leptomeninges. The leptomeningeal flare after discontinuing EGFR inhibition highlights the risk of switching to RET inhibitor monotherapy when CNS disease is driven by the original EGFR mutant clone. CSF liquid biopsy provided actionable, compartment-specific genotyping that outperformed cytology and guided effective retreatment. Incorporating CSF ctDNA into routine evaluation may improve therapeutic alignment across sanctuary sites; when feasible, maintaining EGFR blockade should be considered when CNS involvement is suspected in EGFR-mutated NSCLC in routine practice.
{"title":"Cerebrospinal fluid ctDNA clarifies clonal divergence: leptomeningeal flare of EGFR-mutant disease after switch to selpercatinib for acquired RET fusion","authors":"Kei Kunimasa , Motohiro Tamiya , Takako Inoue , Nobuaki Mamesaya , Tsunehiro Tanaka , Kiyohide Komuta , Shun Futamura , Keiichiro Honma , Kazumi Nishino","doi":"10.1016/j.jlb.2025.100450","DOIUrl":"10.1016/j.jlb.2025.100450","url":null,"abstract":"<div><div>Liquid biopsy can expose spatially segregated resistance biology that is invisible to single-site tissue testing, particularly across the blood–brain barrier. We report how cerebrospinal fluid (CSF) circulating tumor DNA (ctDNA) clarified therapeutic direction in <em>EGFR</em>-mutated non-small-cell lung cancer (NSCLC) with leptomeningeal involvement. A 67-year-old woman with <em>EGFR</em> exon 19–deleted adenocarcinoma received afatinib followed by long-term osimertinib. After three years, progression of the primary lesion prompted rebiopsy, which revealed a <em>CCDC6-RET</em> fusion with strong RET immunoreactivity. Selpercatinib monotherapy yielded minor thoracic shrinkage at 1 month but was followed by dizziness and MRI evidence of leptomeningeal enhancement at 3 months. CSF analysis showed pleocytosis without malignant cells. Critically, CSF ctDNA demonstrated the <em>EGFR</em> E746_A750 deletion at 29.2 % variant allele frequency by amplicon sequencing, whereas the <em>CCDC6–RET</em> fusion was undetectable by targeted sequencing and highly sensitive single-plex qPCR using junction-specific primers. Re-challenging with osimertinib rapidly improved symptoms and led to resolution of leptomeningeal enhancement. These data indicate clonal divergence at acquired resistance: a <em>RET</em>-fusion clone dominated the thoracic compartment while <em>EGFR</em>-addicted clones predominated in the leptomeninges. The leptomeningeal flare after discontinuing EGFR inhibition highlights the risk of switching to RET inhibitor monotherapy when CNS disease is driven by the original <em>EGFR</em> mutant clone. CSF liquid biopsy provided actionable, compartment-specific genotyping that outperformed cytology and guided effective retreatment. Incorporating CSF ctDNA into routine evaluation may improve therapeutic alignment across sanctuary sites; when feasible, maintaining EGFR blockade should be considered when CNS involvement is suspected in <em>EGFR</em>-mutated NSCLC in routine practice.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"10 ","pages":"Article 100450"},"PeriodicalIF":0.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145750061","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-29DOI: 10.1016/j.jlb.2025.100451
A. Manzi , C. De Luca , F. Conticelli , B. Zizolfi , M. Guida , G. Troncone , A. Di Spiezio Sardo , M.C. De Angelis
Introduction
Endometrial cancer (EC) is the sixth most frequent female malignancy worldwide. Atypical endometrial hyperplasia (AEH), also termed endometrial intraepithelial neoplasia (EIN), represents the recognized precursor lesion. While hysteroscopy with biopsy remains the diagnostic gold standard, the lack of non-invasive biomarkers for early detection and follow-up is a major clinical limitation. MicroRNAs (miRNAs), small non-coding RNAs regulating gene expression, have emerged as potential diagnostic and prognostic tools in gynecological oncology.
Objective
We aimed to determine the accuracy of the predictive value of a selected panel of miRNAs (miR-504-5p and miR-429) obtained on endometrial samples, in detecting EC and EIN, and to explore their role along the neoplastic continuum.
Methods
A prospective observational study was conducted at the University of Naples “Federico II.” Thirty-seven patients were enrolled: EC (n = 15), EIN (n = 15), and controls with normal endometrium (n = 7). All underwent hysteroscopy with grasp biopsy or “Visual D&C.” Formalin-fixed paraffin-embedded samples were processed for RNA extraction, and miRNA expression was analyzed by RT-PCR (Taqman Advanced miRNA Assays).
Results
Both miRNAs were successfully amplified in most cases. In EC, miR-504-5p was detected in 93.3 % and miR-429 in 100 % of samples, with mean Ct values of 32.2 and 29.7, respectively. AEH showed intermediate expression (93.3 % and 86.7 % detection rates; mean Ct 28.3 and 29.8). Normal endometrium displayed the highest expression (100 % and 85.7 %; mean Ct 25.5 and 26.8). A progressive downregulation of both miRNAs from normal tissue to AEH and EC was observed.
Conclusion
Our preliminary findings suggest that reduced expression of miR-504-5p and miR-429 characterizes the transition from EIN to EC, supporting their potential role as tumor suppressors in this setting. This two-miRNA panel could complement histopathology in distinguishing precursor lesions from carcinoma, addressing a key diagnostic challenge. Larger studies, including minimally invasive liquid biopsy approaches, are warranted to validate their clinical utility.
{"title":"Tissue expression of miR-504-5p and miR-429 as diagnostic biomarkers for endometrial cancer and endometrial intraepithelial neoplasia: a pilot study","authors":"A. Manzi , C. De Luca , F. Conticelli , B. Zizolfi , M. Guida , G. Troncone , A. Di Spiezio Sardo , M.C. De Angelis","doi":"10.1016/j.jlb.2025.100451","DOIUrl":"10.1016/j.jlb.2025.100451","url":null,"abstract":"<div><h3>Introduction</h3><div>Endometrial cancer (EC) is the sixth most frequent female malignancy worldwide. Atypical endometrial hyperplasia (AEH), also termed endometrial intraepithelial neoplasia (EIN), represents the recognized precursor lesion. While hysteroscopy with biopsy remains the diagnostic gold standard, the lack of non-invasive biomarkers for early detection and follow-up is a major clinical limitation. MicroRNAs (miRNAs), small non-coding RNAs regulating gene expression, have emerged as potential diagnostic and prognostic tools in gynecological oncology.</div></div><div><h3>Objective</h3><div>We aimed to determine the accuracy of the predictive value of a selected panel of miRNAs (miR-504-5p and miR-429) obtained on endometrial samples, in detecting EC and EIN, and to explore their role along the neoplastic continuum.</div></div><div><h3>Methods</h3><div>A prospective observational study was conducted at the University of Naples “Federico II.” Thirty-seven patients were enrolled: EC (n = 15), EIN (n = 15), and controls with normal endometrium (n = 7). All underwent hysteroscopy with grasp biopsy or “Visual D&C.” Formalin-fixed paraffin-embedded samples were processed for RNA extraction, and miRNA expression was analyzed by RT-PCR (Taqman Advanced miRNA Assays).</div></div><div><h3>Results</h3><div>Both miRNAs were successfully amplified in most cases. In EC, miR-504-5p was detected in 93.3 % and miR-429 in 100 % of samples, with mean Ct values of 32.2 and 29.7, respectively. AEH showed intermediate expression (93.3 % and 86.7 % detection rates; mean Ct 28.3 and 29.8). Normal endometrium displayed the highest expression (100 % and 85.7 %; mean Ct 25.5 and 26.8). A progressive downregulation of both miRNAs from normal tissue to AEH and EC was observed.</div></div><div><h3>Conclusion</h3><div>Our preliminary findings suggest that reduced expression of miR-504-5p and miR-429 characterizes the transition from EIN to EC, supporting their potential role as tumor suppressors in this setting. This two-miRNA panel could complement histopathology in distinguishing precursor lesions from carcinoma, addressing a key diagnostic challenge. Larger studies, including minimally invasive liquid biopsy approaches, are warranted to validate their clinical utility.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"11 ","pages":"Article 100451"},"PeriodicalIF":0.0,"publicationDate":"2025-11-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145979156","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Adrenal cortical carcinoma (ACC) is a rare and aggressive endocrine tumor that originates from the adrenal cortex. Radical tumor resection remains the most effective therapy since survival dramatically drops when metastases are present at diagnosis. Hence, there is an urgent need for reliable biomarkers to enable early diagnosis, monitor minimal residual disease (MRD), and assess chemotherapy response. Circulating tumor cells (CTCs) detected via blood liquid biopsy may represent a valuable oncological marker in ACC patients. However, CTC isolation methods so far applied to ACC patients lack specificity, reproducibility and standardization, thus preventing the potential application of CTCs in the management of these patients.
In this study, we present a novel method for the specific and reproducible detection and analysis of single CTCs in ACC patients. This approach combines size- and mechanical property-based enrichment via the Parsortix® system with immunofluorescent detection and isolation of single CTCs using DEPArray® technology, targeting the steroidogenic factor-1 (SF1) nuclear adrenal cortex marker. Isolated CTCs undergo low-pass copy number alteration (CNA) analysis. This is the first report of a nuclear antigen-based method for isolating single CTCs in suspension, overcoming the limitations of membrane and cytokeratin markers commonly used in other solid tumors. By exploiting the large nuclear size of CTCs, this strategy provides an alternative and more standardized approach for single-cell isolation in tumors lacking specific surface markers.
{"title":"An innovative nuclear antigen-based approach for single-cell isolation of circulating tumor cells in adrenal cortical carcinoma","authors":"Giulia Cantini , Francesca Salvianti , Roberta Armignacco , Arianna Pia Propato , Letizia Canu , Tonino Ercolino , Anna Aurora Dedonno , Chiara Lepri , Massimo Mannelli , Gabriella Nesi , Serena Pillozzi , Giuseppe Defazio , Pamela Pinzani , Michaela Luconi","doi":"10.1016/j.jlb.2025.100447","DOIUrl":"10.1016/j.jlb.2025.100447","url":null,"abstract":"<div><div>Adrenal cortical carcinoma (ACC) is a rare and aggressive endocrine tumor that originates from the adrenal cortex. Radical tumor resection remains the most effective therapy since survival dramatically drops when metastases are present at diagnosis. Hence, there is an urgent need for reliable biomarkers to enable early diagnosis, monitor minimal residual disease (MRD), and assess chemotherapy response. Circulating tumor cells (CTCs) detected via blood liquid biopsy may represent a valuable oncological marker in ACC patients. However, CTC isolation methods so far applied to ACC patients lack specificity, reproducibility and standardization, thus preventing the potential application of CTCs in the management of these patients.</div><div>In this study, we present a novel method for the specific and reproducible detection and analysis of single CTCs in ACC patients. This approach combines size- and mechanical property-based enrichment via the Parsortix® system with immunofluorescent detection and isolation of single CTCs using DEPArray® technology, targeting the steroidogenic factor-1 (SF1) nuclear adrenal cortex marker. Isolated CTCs undergo low-pass copy number alteration (CNA) analysis. This is the first report of a nuclear antigen-based method for isolating single CTCs in suspension, overcoming the limitations of membrane and cytokeratin markers commonly used in other solid tumors. By exploiting the large nuclear size of CTCs, this strategy provides an alternative and more standardized approach for single-cell isolation in tumors lacking specific surface markers.</div></div>","PeriodicalId":101235,"journal":{"name":"The Journal of Liquid Biopsy","volume":"10 ","pages":"Article 100447"},"PeriodicalIF":0.0,"publicationDate":"2025-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145579312","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-05DOI: 10.1016/j.jlb.2025.100443
Emma Holjak , Tia Brasoveanu , Saurav Verma , Saqib Khan , Morgan Black , Inderdeep Dhaliwal , Michael Mitchell , Rahul Nayak , Mehdi Qiabi , Richard Inculet , Dalilah Fortin , Richard Malthaner , Matthew Cecchini , Anna Lapuk , Daniel Breadner
Liquid biopsy (LB) is a useful tool in patients with advanced non-small cell lung cancer (aNSCLC) to detect actionable molecular alterations and thereby allow genotype-matched therapies. Currently, LB is recommended for individuals diagnosed with aNSCLC who have an insufficient tissue sample or difficult-to-reach tumour tissue. Despite the potential advantages of LB, its incorporation into the standard diagnostic work-up for all newly diagnosed patients with aNSCLC is lacking. Our study aimed to evaluate whether addition of plasma circulating tumor DNA (ctDNA) next generation sequencing (NGS) testing early in the diagnostic work-up for patients with aNSCLC can improve the time to molecular results and treatment initiation. This was a single-centre quality improvement initiative with two cohorts of patients. Patients in the ‘ctDNA cohort’ had plasma ctDNA testing in addition to the standard diagnostic work-up. The ‘reference cohort' was a parallel group of patients who had the standard work-up (no LB). Tissue biopsy and reflex tissue NGS testing were done in both cohorts. ctDNA testing shortened the time to molecular results in the ctDNA cohort compared to the reference cohort (median, 14 vs 35 days; p = 0.01), the time from first respirology/thoracic surgery consult to molecular results (median, 22 vs 48 days respectively; p = 0.01), and the time from medical oncology consultation to initiation of first-line treatment (median, 12 vs 22 days; p = 0.01). In conclusion, in a publicly funded and single-payer healthcare system, ctDNA testing as part of the standard work-up for patients with aNSCLC provides molecular results significantly faster than tissue-based testing and shortens the time to treatment initiation.
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Pub Date : 2025-11-01DOI: 10.1016/j.jlb.2025.100337
Dr. Lara Paracchini , Dr. Lorena Ancona , Dr. Luca Beltrame , Dr. Sonia Ismari , Dr. Micaela Piemontese , Dr. Laura Mannarino , Dr. Francesca Chemi , Paolo Zucali (Prof.) , Paolo Bossi (Prof.) , Domenica Lorusso (Prof.) , Dr. Alberto Puccini , Maurizio D'Incalci (Prof.) , Dr. Sergio Marchini
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