Introduction

Pancreatic ductal adenocarcinoma (PDAC) is notorious for its aggressive nature and grim prognosis, underscoring the urgent need for personalized pre-clinical models to develop targeted therapies. Patient-derived organoids (PDOs) are three-dimensional models of human tumours that have emerged as promising research tools.

Aims & Methods

Our study aimed to assess the feasibility of endoscopic ultrasound-guided fine needle biopsy (EUS-FNB) for the generation of PDAC-PDOs. A single-centre prospective database was established to collect data from suspected PDAC patients undergoing EUS-FNB for histological confirmation between May and October 2023. All biopsies were performed using a 22G-needle. Two to three biopsies were taken for histological examination before performing a single-pass FNB to establish PDO cultures. The primary outcome was the successful propagation of PDAC-PDOs for at least three passages (P3). Secondary outcomes included sample adequacy, diagnostic rate, and safety.

Results

A total of 22 patients (M:F 13:9, median age 66.5 years, range 55-86) were enrolled. One case was subsequently diagnosed as pancreatic lymphoma and excluded. PDAC-PDOs were successfully established in 9 out of 21 PDAC cases, resulting in a success rate of 43% (please see Table 1 for further details on singular included patients). Sample adequacy and diagnostic rates were both achieved in 100% of subjects. The presence of a biliary stent during EUS-FNB did not impact the success rate of PDAC-PDO generation (n=1 in the generated PDAC-PDOs group vs n=3 in the non-generated PDAC-PDOs group; p>0.05). No adverse events related to the procedure occurred.

Patient	Gender	Age	Lesion Size (cm)	Pancreatic Location	Sample Adequacy	Adverse Events
1	Male	62	3.5	Head	Yes	No
2	Female	85	3.0	Head	Yes	No
3	Male	61	1.2	Head	Yes	No
4	Male	68	3.5	Tail	Yes	No
5	Female	59	4.0	Head	Yes	No
6	Male	56	4.0	Body	Yes	No
7	Female	64	4.5	Tail	Yes	No
8	Female	60	5.0	Head	Yes	No
9	Female	58	4.0	Neck	Yes	No

Conclusion

This study demonstrates that PDAC-PDOs can be successfully generated through EUS-FNB sampling with a single-pass fine needle biopsy, without compromising the safety profile of the procedure.

References

Tiriac H, Bucobo JC, Tzimas D, Grewel S, Lacomb JF, Rowehl LM, Nagula S, Wu M, Kim J, Sasson A, Vignesh S, Martello L, Munoz-Sagastibelza M, Somma J, Tuveson DA, Li E, Buscaglia JM. Successful creation of pancreatic cancer organoids by means of EUS-guided fine-needle biopsy sampling for personalized cancer treatment. Gastrointest Endosc. 2018 Jun;87(6):1474-1480. doi: 10.1016/j.gie.2017.12.032.

Introduction

Tumor biology studies of pancreatic ductal adenocarcinoma (PDAC) used surgical specimens, even though most PDACs are unresectable.

Aims & Methods

The aims of the study were to 1) establish PDAC organoids platform from endoscopic ultrasonography (EUS)-guided fine needle biopsy (FNB), 2) investigate clinicopathologic and genomic characterization, and 3) find out the clinical applicability of the patient-derived PDAC organoids model from PDAC diagnosis to treatment timeline The patients with suspected PDACs were enrolled prospectively, and PDAC minimal specimens were acquired from EUS-guided FNB at Samsung Medical Center.

Results

Ninety-four PDAC organoids were successfully established (94/113, 83.2%) via EUS-guided FNB within 8.2±2.6 days. Clinical factors were also analyzed associated with the success rate in establishing PDAC organoids; specimens with high cellularity (>10 clusters) were an independent factor in multivariate analysis. Whole exome sequencing showed the high concordance between primary PDAC and its organoid and more enriched genetic alterations in organoids. In addition, the high-throughput screening (HTS) drug test revealed that the clinical correlation with real-world data was 12 out of 15 between PDAC organoids and the chemotherapeutic response of the study patients. Whole-transcriptome sequencing identified genes and pathways associated with the drug responses.

Conclusion

We have achieved the PDAC organoid platform in a high success rate via EUS-guided FNB, and the concordance between primary PDAC and its organoid seems very high. More importantly, the HTS drug test showed that PDAC organoids may well reflect clinical chemotherapeutic response and may also be used as the “Patient Avatar Model” in clinical practice on time.

Clinical Case Summary

A 51-year-old male presented to the emergency department with acute back pain, suspicious for renal colic. An abdominal CT scan incidentally revealed a 30 mm hypodense lesion in the pancreatic uncinate process with central calcifications (see Fig. 1).
An endoscopic ultrasound (EUS) demonstrated a hypoechoic, heterogeneous mass in the uncinate process, with irregular margins and a huge posterior acoustic shadowing. No signs of vascular or ductal involvement were noted. Therefore, EUS-guided fine needle biopsy (FNB) with a 22-G Franseen-tip needle was performed via transduodenal and transgastric approaches, obtaining an adequate histological sample after macroscopic on-site examination (MOSE).
Histological analysis revealed spindle cells positive for CD34, BCL2, STAT6, and CD99, and negative for CKIT, DOG1, desmin, actin, S100, and cytokeratins, with Ki-67 index > 5%, suggesting for solitary fibrous tumor (SFT) of the pancreas (see Fig.2). Immunohistochemical confirmation was obtained at a tertiary referral pathology laboratory. A multidisciplinary tumor board advised conservative follow-up based on the benign histology and absence of local invasion.
Conclusion: Pancreatic SFTs are extremely rare, with only 34 cases reported to date. They are more frequently located in the pancreatic head and are slightly more common in females. Most are discovered incidentally. This case highlights the critical diagnostic value of EUS-FNB, which enabled accurate tissue acquisition for advanced immunohistochemical analysis, informing a non-surgical, surveillance-based approach. Because of the non-specific clinical presentation and radiological features of pancreatic SFTs, diagnosis is particularly challenging using preoperative imaging and laboratory tests alone. Therefore, histopathological and immunohistochemical evaluation is essential for definitive diagnosis.

References

References and Illustrative Material:

Figure 1: Abdominal CT scan showing a pancreatic mass in the uncinate process with central calcification.

Figure 2: On the left, tissue sample stained with hematoxylin and eosin. Spindle cell proliferation with disorganized architecture. The lesion is hypercellular, composed of elongated mesenchymal cells with scant cytoplasm and oval-to-tapered nuclei, suggesting a spindle cell neoplasm, such as SFT. On the right, the same tissue sample was analyzed using immunohistochemistry: strong and diffuse nuclear positivity for STAT6 was found in tumor cells.

Selected References:

1. Ronchi A et al. Solitary fibrous tumors: an update on diagnosis and management. Pathol Oncol Res.

2020;26(2):597-605.

2. Wang WW, Zhou SP, Wu X, Wang LL, Ruan Y, Lu J, Li HL, Ni XL, Qiu LL, Zhou XH. Imaging, pathology, and diagnosis of solitary fibrous tumor of the pancreas: A case report and review of literature. World Journal of Clinical Cases. 2024;12(5):995–1003.

Introduction

Insulinomas are rare, functioning pancreatic neuroendocrine neoplasms (pNEN), whose gold standard therapy is surgical resection. Endoscopic ultrasound-guided radiofrequency ablation (EUS-RFA) has emerged as a minimally invasive therapeutic option for pancreatic lesions. This study aims at describing a series of patients with pancreatic insulinoma treated with EUS-RFA.

Aims & Methods

This single-center, retrospective study includes all consecutive patients with pancreatic insulinoma undergoing EUS-RFA because unfit for surgery or refusing surgery. EUS-RFA was performed by using a 19G RFA needle, and a dedicated current generator system with a RFA energy of 20 to 50W. Technical success (i.e., the achievement of complete ablation), adverse events rate, and radiologic response within 3 months after EUS-RFA (i.e., complete tumor necrosis) were evaluated. Follow-up was assessed by clinical monitoring and blood test evaluation at least every 6 months for the first year after the procedure. A descriptive statistical analysis was performed for all the variables.

Results

From March 2017 to September 2022, 10 patients (mean age: 67.1 ± 10.1 years; F:M 7:3) were included. The mean size of insulinomas was 11.9 ± 3.3mm. The median number of passes was 6 (1-14) with a median ablation duration of 15 (6-20) seconds per pass. Technical success, immediate post-procedural euglycemia, and complete radiological response by abdominal CT scan were achieved in all cases (100%). Mild procedure-related early adverse are shown in Table 1. Persistent euglycemia was assessed at 6 and 12 months for each treated patient.

Patients, n	10
Technical success, n (%)	10 (100)
Early adverse events, n (%) Mild abdominal pain Bleeding	2 (20) 1 (10)
Late adverse events, n (%)	0 (0)
Euglycaemia within 24 hours	10 (100)
Radiologic complete response within 90 days, n (%)	10 (100)
Clinical remission at 6 months, n (%)	10 (100)
Clinical remission at 12 months, n (%)	9/9 (100)

Conclusion

Data from this case series suggest that EUS-RFA is a feasible and safe therapeutic approach for pancreatic insulinomas with mid-term efficacy.

References

Waung, J.A.; Todd, J.F.; Keane, M.G.; Pereira, S.P. Successful management of a sporadic pancreatic insulinoma by endoscopic ultrasound-guided radiofrequency ablation. Endoscopy 2016, 48 (Suppl. S1), E144–E145
Goldberg SN, Mallery S, Gazelle GS, Brugge WR. EUS-guided radiofrequency ablation in the pancreas: results in a porcine model. Gastrointest Endosc. 1999 Sep;50(3):392-401. doi: 10.1053/ge.1999.v50.98847.
Tsoli M, Chatzellis E, Koumarianou A, Kolomodi D, Kaltsas G. Current best practice in the management of neuroendocrine tumors. Ther Adv Endocrinol Metab. 2018 Oct 31;10:2042018818804698. doi: 10.1177/2042018818804698.

Introduction

Insulinomas are rare, functioning neuroendocrine neoplasms that cause non-ketotic hypoglycemia due to hyperinsulinism. Surgery is considered the gold-standard treatment, although endoscopic ultrasound-guided radiofrequency ablation (EUS-RFA) has been recently described as a less invasive, efficient therapeutic option for these tumors. In this study, we aim to compare post-procedural outcomes of surgical resection versus EUS-RFA for pancreatic insulinomas.

Aims & Methods

All patients who underwent surgical resection or EUS-RFA for pancreatic insulinoma in our Institution were included in the study, and data from all procedures were retrospectively recorded and statistically analyzed. EUS-RFA was indicated for all subjects either unfit for surgery or unwilling to undergo surgical treatment. Patients’ baseline characteristics, location, size of insulinomas, procedural time, and intra- and postoperative outcomes were compared.

Results

From April 2012 to July 2022, 11 and 10 patients with pancreatic insulinoma underwent surgical resection and EUS-RFA, respectively. The median age was 65.5 (51-84) and 37 years (28-72, p<0.001) for the EUS-RFA and surgery cohorts, respectively. As a whole, 15 (71.4%) out of 21 were female. The median tumor size was 15mm (8-30) for the surgical group and 11mm (8-19) for the EUS-RFA group (p=0.17). The majority of the insulinomas were located in the pancreatic body (28.6%) and tail (28.6%), with no statistical difference between the two groups (p=0.16). Operative time was significantly longer in the surgical population (280min, 120-372) as compared to the endoscopic one (28min,18-40, p<0.001). Clinical success, defined as the normalization of glucose blood levels, was reached in all cases. The complication rate was significantly higher in the surgical group compared to the endoscopic group (7-63.6% vs 1-10%, p=0.01). Specifically, 7 patients in the surgical cohort developed a pancreatic fistula while one patient in the EUS-RFA population had self-limiting bleeding after the procedure. One patient of the surgical cohort underwent reoperation for an infected abdominal fluid collection.

Table 1. Characteristics of patients classified by treatments.

Variable	EUS-RFA N=10	Surgical resection N=11	p value
Median age-adjusted CCI, points	4.5 (2-6)	2.0 (2-5)	0.002
MEN-1, n (%)	2 (20.0)	0 (0.0)	0.119
Hypoglycemic symptoms, n (%)	10 (100)	11 (100)	/
Positive fasting test, n (%)	9 (90.0)	10 (90.9)	0.943
Median operative time, min (range)	28 (18-40)	280 (120-372)	<0.001
Procedure-related AE, n (%)	1 (10.0)	7 (63.3)	<0.001
Postoperative euglycemia, n (%)	10 (100)	11 (100)	/
Median follow-up time, months (range)	15 (6-62)	51 (6-126)	0.670

Legend: AE: adverse events; EUS-RFA: endoscopic ultrasound-guided radiofrequency ablation; CCI: Charlson Comorbidity Index; MEN-1: multiple endocrine neoplasia type 1.

Conclusion

As compared to surgical resection, EUS-RFA for pancreatic insulinomas may be considered a safe and efficient treatment option with a shorter operative time and a lower rate of related complications. Prospective comparative studies are needed to ascertain the role of EUS-RFA as a treatment of choice for selected patients with pancreatic insulinoma.

References

Fahmawi Y, Mehta A, Abdalhadi H et al. Efficacy and safety of endoscopic ultrasound-guided radiofrequency ablation for management of pancreatic lesions: a systematic review and meta-analysis. Transl Gastroenterol Hepatol. 2022 Jul 25;7:30.
Zhao YP, Zhan HX, Zhang TP et al. Surgical management of patients with insulinomas: Result of 292 cases in a single institution. J Surg Oncol. 2011 Feb;103(2):169-74.

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is the seventh leading cause of cancer related death worldwide, with a progressive increasing incidence¹ and 5 years overall survival (OS) of 12.5% irrespective of the disease stage². Given the tumor biological aggressiveness and the limited response to chemotherapy, it arises the need to develop preclinical study models able to understand the underlying mechanism of tumor progression and chemoresistance³. These models need to be easily generable, reproducible, representative of the entire population affected by PDAC and to have the ability of mimicking the cancer behavior. Patients derived organoids (PDOs) are models of epithelial cells with self-regeneration and self-organization abilities able to behave and mimic the tumors of origin⁴. Liquid biopsy has also gained importance in tailoring the treatment for oncological patients, although the role of bile as biological fluid in patients affected by biliopancreatic malignancies is still uncertain.

Aims & Methods

We performed a prospective observational study aiming to evaluate the feasibility to create tridimensional PDOs from fine needle biopsy (FNB) samples of patients affected by PDAC and to correlate PDOs with native cancer histology. The secondary endpoint of our study was to assess the feasibility of using bile as biological fluid for cell free DNA (cfDNA) and to investigate the landscape of cfDNA in bile and plasma of patients affected by PDAC.
Patients with strong suspicion or recent diagnosis of PDAC and concomitant jaundice were included. Each patient underwent EUS guided FNB (EUS-FNB) for histological diagnosis and organoid collection was performed. By the time of biliary drainage, bile and plasma were collected for DNA extraction and analysis.

Results

From June 2023 to September 2023, 36 patients with neoplastic jaundice were evaluated. Eighteen patients with final diagnosis of PDAC have been included in the study. Eighteen underwent biliary drainage with collection of bile and plasma. Sixteen underwent EUS-FNB for organoid collection.
Mean tumor size was 33.7± 9.6 mm. Tumor was located in the head of pancreas in 94.4% of cases. Nine patients (50%) were metastatic by the time of diagnosis, eight patients (44%) had locally advanced disease and only one patient was considered resectable. Tissue acquisition was performed using FNB needles. Organoids establishment rate was successful in 100% of cases. No EUS-FNB related adverse events were observed.
At a quality concordance pathological analysis between PDAC and PDOs, morphological similarities have been observed in all cases (100%). Cellular morphology and nuclei atypia were the most consistent similarities. Organoids showed a glandular over expression compared to native PDAC. Extracted cfDNA was significantly higher in bile supernatant than in plasma (2.9 ng/ml vs 19.8 ng/ml p=0.0001). No significant differences were found for plasma and bile cfDNA among disease stages. Higher level of bile cfDNA in locally advanced disease and a trend of higher level of plasma cfDNA in metastatic stage. Bile cfDNA showed longer DNA fragment compared to plasma.

Conclusion

Organoids establishment is feasible and safe with a single passage of EUS-FNB. PDOs maintain the morphological architecture of native cancer, although the absence of dense stromal reaction leads to a remarkable organoids ability to create glandular structures. Qualitative and quantitative composition of ctDNA is superior in bile rather than in plasma, making this fluid an important tool for NGS analysis and a better comprehension of tumor mutations.

References

1. Sung H, Ferlay J, Siegel RL, et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J Clin. 2021;71(3):209-249. doi:10.3322/caac.21660
2. Arnold M, Rutherford MJ, Bardot A, et al. Progress in cancer survival, mortality, and incidence in seven high-income countries 1995-2014 (ICBP SURVMARK-2): a population-based study. Lancet Oncol. 2019;20(11):1493-1505. doi:10.1016/S1470-2045(19)30456-5
3. Baker LA, Tiriac H, Clevers H, Tuveson DA. Modeling pancreatic cancer with organoids. Trends Cancer. 2016;2(4):176-190. doi:10.1016/j.trecan.2016.03.004
4. Moreira L, Bakir B, Chatterji P, Dantes Z, Reichert M, Rustgi AK. Pancreas 3D Organoids: Current and Future Aspects as a Research Platform for Personalized Medicine in Pancreatic Cancer. Cell Mol Gastroenterol Hepatol. 2017;5(3):289-298. Published 2017 Dec 16. doi:10.1016/j.jcmgh.2017.12.004

Introduction

Pancreatic ductal adenocarcinoma (PDAC) is characterized by intense fibrosis which enhances the tumor growth and drives an epithelial-mesenchymal transition. The consequence is the uncontrolled proliferation of cancer cells, including pancreatic cancer stem cells (PCSC), which may enhance the metastatic potential.

Aims & Methods

The aim of this study is to assess the prognosis of CD44, CD24 and EpCAM expression, on pancreatic endoscopic ultrasound-guided fine-needle biopsy (EUS-FNB) specimens.
A total of 35 patients diagnosed with PDAC after EUS-FNB within the Research Center of Gastroenterology and Hepatology of Craiova, were included in the study, and the samples were sent for immunohistochemical assessment. A total of three passes were performed on every PDAC tumor, to harvest enough tissue for analysis. Anti-Human CD 24 (policlonal, Abcam, ab 199140) and EpCAm (Epr 20532-225, Rb monoclonal, Abcam, ab 223582) and Human CD 44 (clone DF 1485, Agilent Tehnologies Denmark, cod M7082) were used. The slides were incubated with primary antibodies for 30 minutes at room temperature. Immunolabeling in the study group was considered positive if more than 10% of tumor cells showed a present signal and negative if less than 10% of tumor cells were immunolabeled. The intensity of immunolabeling in positive cases was assessed as weak (1), moderate (2), and intensely positive (3).

Results

We assessed a total of 300 fields and 54 (18%) were assessed as triple positive, however when all three were positive no correlations were found. The expression of CD44+ ⁄ CD24+ expression was significantly correlated with poorer outcomes (P = 0.05), whereas CD44+ ⁄EpCAM+, CD24+ ⁄EpCAM+, CD44+, CD24+, or Ep-CAM+ expression did not correlate with patient’s evolution. Among the patient characteristics, T stage and N stage were significantly correlated with poorer prognosis. CD44 and CD24 were significantly correlated with poor differentiation and high proliferation which might suggest their association with cancer aggressiveness, whereas EpCAM was correlated with well differentiation and low proliferation. When comparing patients with metastasis at the time of diagnosis, CD44+ ⁄ CD24+ expression was more intense when M1 was present (p<0.005), suggesting their prognostic role. Survival analysis showed there was no correlation with the PCSC staining.

Conclusion

EUS-FNB PSCS with CD44 + /CD24+ testing might be considered for PDAC assessment as their positivity might be correlated with a poorer outcome. They might be considered for prognosis purposes at the time of diagnosis as EUS-FNB might provide significant tissue for analysis, thus suggesting their possible therapeutic value in a clinical setting.

Disclosure

No conflicts of interest