158P - Single-cell RNA sequencing via Endoscopic Ultrasoundguided Fine-Needle Biopsy (EUS-FNB) Pancreatic Biopsies uncovered an aggressive subclone with a poor prognosis

Background

Single-cell RNA sequencing (scRNA-seq) has the ability to unveil uncommon cell populations. However, due to the high demand for tissue quality and cell viability, currently most scRNA-seq for pancreatic cancer was performed by surgical specimen or biopsy from metastatic sites. This study aims to establish a practical experience to help scientists perform primary pancreatic tumor scRNA-seq using EUS-FNB samples in real-world practice.

Methods

Two punctures from the same lesion using the same needle, without applying suction (Non-suction group) and with a negative pressure of 5 ml (Suction group) were evaluated. Single cell RNA sequencing libraries were prepared with Chromium Single cell 5’ Reagent Kits v2 (10X Genomics, USA) following the manufacturer’s protocol.

Results

A total of 20 patients were enrolled. The median age was 65.1 years old (range 46.6-83.2). Suction group achieved a cell preparation success rate of 80% (16/20) which was significantly higher than the 10% (2/20) success rate in the non-suction group (p<0.001). After the establishment of cell preparation protocol, we generate single-cell RNA transcriptomes for four patients, including two early stage (9,632 cells) and two late stage (4,592 cells). After quality control, 11,950 single cells were feasible for downstream analysis. Overall, 66% of cells (7,842) belonged to early stage and 34% (4,108) belonged to late stage. 12 major cell subtypes were identified across early and late stage. The proportion of cancer cells cluster-4 was significantly higher in late stage. Differentially expressed genes analysis showed UBE2C is the most highly expressed gene in cancer cells cluster-4. As external validation, in TCGA PAAD dataset, we found UBE2C high expression pancreatic cancer had significantly poor survival.

Conclusions

EUS-FNB with a negative pressure of 5 ml is feasible for single-cell sequencing in daily practice. A UBE2C high-expression subclone exists in early-stage pancreatic cancer and correlates with poor prognosis, potentially becoming a new therapeutic target in future studies.

Clinical trial identification

Protocol number: NCT05767697 Release date: 02 March, 2023.

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

National Health Research Institutes.

Disclosure

All authors have declared no conflicts of interest.