ATRX/DAXX in EUS-FNB Specimens of Pan-NETs

NCT ID: NCT06406387

Last Updated: 2026-01-13

Basic Information

• Recruitment Status: ACTIVE_NOT_RECRUITING
• Total Enrollment: 100 participants
• Study Classification: OBSERVATIONAL
• Study Start Date: 2024-06-01
• Study Completion Date: 2029-11-30

Brief Summary

P-NENs are classified as functional (F-) or non-functional (NF-) depending on the presence or absence of a clinical hormonal hypersecretion syndrome. Moreover, the WHO 2017 classification of pNENs distinguishes between well-differentiated pancreatic neuroendocrine tumors (pNETs) and poorly differentiated pancreatic neuroendocrine carcinomas (pNECs). pNETs are then divided according to a grading scheme based on Ki67 index in pNETs-G1 (Ki67 index ≤3%) and pNETs-G2 (Ki67 index between 4% and 20%). pNECs are all G3, with a Ki67 index >20%. Endoscopic ultrasound with fine-needle biopsy (EUS-FNB) demonstrated safe and effective preoperative grading based on the Ki-67 proliferative index. However, downstaging rate is not neglectable, reaching 15% in a recent metanalysis. Moreover, recent whole-exome and whole genome sequencing studies revealed that the mutually exclusive inactivating mutations in death domain-associated protein (DAXX) and/or in α-thalassemia/mental retardation X-linked (ATRX) chromatin remodeling genes are associated with more aggressive disease. In a retrospective study, the investigators recently evaluated the correspondence of DAXX/ATRX expression on 41 EUS-FNB samples with corresponding surgical specimens demonstrating a 95.1% (almost perfect agreement, κ = 0.828; p < 0.001) and 92.7% (substantial agreement, κ = 0.626; p < 0.001) concordance for DAXX and ATRX expression, respectively. This study aims to evaluate the potential clinical/prognostic role of DAXX/ATRX expression as implementation of the currently used Ki67-based grading, evaluated on EUS-FNB samples in a prospective cohort of patients with NF-pNETs

Conditions

Pancreatic Neuroendocrine Tumor

Study Design

• Observational Model Type: COHORT
• Study Time Perspective: PROSPECTIVE

Study Groups

ATRX/DAXX loss

The "positive" group including patients with loss of DAXX or ATRX expression evaluated on immunohistochemistry on endoscopic ultrasound biopsy specimens

ATRX/DAXX immunohistochemistry

Intervention Type: DIAGNOSTIC_TEST

Immunohistochemistry will be performed using an Autostainer Leica (Leica Biosystems) according to the manufacturer's instructions. Four μm formalin-fixed paraffin-embedded sections will be immunostained with antibodies for Cytokeratin AE1/AE3 (AE1-AE3, 1:100 dilution, Novocastra/United Kingdom) Chromogranin A (DAK-A3, 1:2500, Dako/Denmark), and Synaptophysin (27G12, 1:100, Novocastra), Ki67 (MIB1, 1:100, Dako/Denmark), ATRX (1:400, Sigma-Aldrich), DAXX (1:200, Sigma-Aldrich). After antigen retrieval, immunostaining will be performed in an automated Bond instrument (Vision-Biosystem, Leica, Milan, Italy) using a sensitive peroxidase-based 'Bond polymer Refine' detection system.

ATRX/DAXX preserved

The "negative" group including those with preserved DAXX and ATRX expression evaluated on immunohistochemistry on endoscopic ultrasound biopsy specimens

ATRX/DAXX immunohistochemistry

Intervention Type: DIAGNOSTIC_TEST

Immunohistochemistry will be performed using an Autostainer Leica (Leica Biosystems) according to the manufacturer's instructions. Four μm formalin-fixed paraffin-embedded sections will be immunostained with antibodies for Cytokeratin AE1/AE3 (AE1-AE3, 1:100 dilution, Novocastra/United Kingdom) Chromogranin A (DAK-A3, 1:2500, Dako/Denmark), and Synaptophysin (27G12, 1:100, Novocastra), Ki67 (MIB1, 1:100, Dako/Denmark), ATRX (1:400, Sigma-Aldrich), DAXX (1:200, Sigma-Aldrich). After antigen retrieval, immunostaining will be performed in an automated Bond instrument (Vision-Biosystem, Leica, Milan, Italy) using a sensitive peroxidase-based 'Bond polymer Refine' detection system.

Interventions

ATRX/DAXX immunohistochemistry

Immunohistochemistry will be performed using an Autostainer Leica (Leica Biosystems) according to the manufacturer's instructions. Four μm formalin-fixed paraffin-embedded sections will be immunostained with antibodies for Cytokeratin AE1/AE3 (AE1-AE3, 1:100 dilution, Novocastra/United Kingdom) Chromogranin A (DAK-A3, 1:2500, Dako/Denmark), and Synaptophysin (27G12, 1:100, Novocastra), Ki67 (MIB1, 1:100, Dako/Denmark), ATRX (1:400, Sigma-Aldrich), DAXX (1:200, Sigma-Aldrich). After antigen retrieval, immunostaining will be performed in an automated Bond instrument (Vision-Biosystem, Leica, Milan, Italy) using a sensitive peroxidase-based 'Bond polymer Refine' detection system.

Intervention Type: DIAGNOSTIC_TEST

Eligibility Criteria

Inclusion Criteria

• Age ≥18 years
• Cyto/histologic diagnosis of pNETs
• Signed informed consent

Exclusion Criteria

• Functional pNETs
• Multiple pancreatic nodules
• Diagnosis of MEN-1 or Von-Hippel Lindau
• Mixed types (e.g., mixed neuroendocrine-acinar/adenocarcinoma) or neuroendocrine carcinomas
• Predominantly cystic lesions (more than 50% of the volume).
• Metastatic tumors at the time of diagnosis
• Known bleeding disorder that cannot be sufficiently corrected with co-fact or fresh frozen plasma
• Use of anticoagulants that cannot be discontinued
• INR >1.5 or platelet count <50.000
• Pregnancy or breastfeeding
• Failure to sign the patient's or closest relative's informed consent

• Minimum Eligible Age: 18 Years
• Eligible Sex: ALL
• Accepts Healthy Volunteers: No

Sponsors

Azienda Ospedaliera Universitaria Integrata Verona

OTHER

Sponsor Role: lead

Responsible Party

Responsibility Role: SPONSOR

Locations

University Hospital of Verona

Verona, , Italy

Countries

Italy

Other Identifiers

FORESEE

Identifier Type: -

Identifier Source: org_study_id