Ethical approval and patient consent

This study was approved by the Wakayama Medical University Hospital Institutional Review Board (number: W-59). The trial is registered with the Japan Registry of Clinical Trials (JRCT; trial registration: jRCTs052230144).

Study aim and design

This multicenter randomized comparative open-label superiority study will involve 10 tertiary centers across Japan. The main aim is to verify the superiority of a 22G Fork-tip needle over a 22G needle Franseen needle for sample acquisition from patients with SELs measuring ≤ 2 cm.

Patients

The investigators will obtain informed consent from the candidates at each institution. Data are input into an electronic data capture system to confirm whether the candidates meet the eligibility criteria, and to register the candidates with the registration secretariat (Fig. 2). Patient enrollment began on December 13, 2023. First participant was enrolled on January 4, 2024.

Inclusion criteria

Individuals meeting all of the following criteria will be considered for the study:

1.

Patients with suspected solid SELs measuring ≤ 2 cm on prior EUS after SELs is first confirmed by endoscopy or CT

2.

Patients scheduled to undergo EUS-FNB for SELs in all organs

3.

Patients who are at least 18 years old at the time of consent

4.

Patients with Eastern Cooperative Oncology Group (ECOG) performance status 2 or lower

5.

Patients who are able and willing to comply with study procedures and provide their own written consent to participate in the research.

Exclusion criteria

Individuals falling under any of the following criteria will be excluded from the study:

1.

Patients with a tendency to bleed (prothrombin time-international normalized ratio > 1.5 or platelet count < 50,000 μL)

2.

Patients with cystic SELs; patients who are or may be pregnant

3.

Patients deemed by the principal investigator (or sub-investigator) to be unsuitable to participate in the research for any other reason.

Data collection

Data will be collected prospectively from all patients, and will include medical history, physical examination, laboratory data, pathological examinations, clinical information, information about prior procedures, and adverse events. The study allocation, intervention, and assessment procedures are adapted from standard protocol items.

Data monitoring and audit

The following aspects will be monitored: data accumulation, patient eligibility, severe adverse events, protocol deviations, reasons for cessation or expiration of the protocol, background factors of the patients, and other problems concerning study progress and safety. Principal investigator will meet with each municipality to monitor participant safety and data assessment procedures. The coordinating center will organize monthly meetings to review the trial conduct. Project management group checks progress once a month and reports back to the investigator. Independent data monitoring meet to review trial conduct and violence of compliance once every three months and reports back to the investigator. We added these points in method section.

Protocol amendments

When the protocol is changed, notifying sponsor and funder first then the principal investigator will notify the centers and that a copy of the revised protocol will be sent to the principal investigator to add to the 1nvestigator site file.

EUS-FNB needles (Fig. 1)Fig. 1

Photographs of (a) a 22G Franseen needle and (b) a 22G Fork-tip needle. a Franseen needles are made of cobalt–chromium and have a crown-shaped tip with three symmetric prongs. b Fork-tip needles are made of stainless steel and contain a nitinol stylet. The device has a multifaceted opposite bevel tip incorporating two sharp prongs of different lengths

Franseen needles

Franseen needles are made of cobalt–chromium, and have a crown-shaped tip with three symmetric prongs.

Fork-tip needles

Fork-tip needles are made of stainless steel and contain a nitinol stylet. The device has a multifaceted opposite bevel tip incorporating two sharp prongs of different lengths.

Study procedureFlow of study procedure (Fig. 2)

After obtaining written consent in person from patients with suspected SELs measuring ≤ 2 cm and who are scheduled to undergo EUS-FNB, their eligibility criteria will be confirmed through appropriate testing and data collection. Eligible patients will be enrolled in accordance with the procedures for patient enrollment. Enrolled patients will be assigned randomly (at a ratio of 1:1) to the 22G Franseen needle or the 22G Fork-tip needle groups (n = 55 per group). On the day of the study procedure, EUS-FNB and the subsequent study procedures will be performed by a qualified endoscopist.

Fig. 2

Protocol workflows for endoscopic ultrasound-guided fine-needle biopsy of subepithelial lesions measuring ≤2 cm

Investigator and facility criteria

The procedure will be conducted by an endoscopist with experience in performing at least 100 EUS-FNBs. The procedure will be performed at a facility accredited by the Japan Gastroenterological Endoscopy Society.

Study procedure (Table 1)

When the study criteria for are met, the four-step study procedure will be conducted as follows:

1.

Puncture: A puncture needle is inserted into the tumor.

2.

Stroke: The puncture needle is moved back and forth within the lesion at least 10 times while applying negative pressure with a 10 mL syringe; ultrasound is used to observe the puncture needle in real time.

3.

Specimen collection: The puncture needle is removed from the endoscope and a stylet is placed inside the extracted puncture needle to push the collected tissue into a petri dish. Then, the needle is flushed with 2.5 mL of physiological saline solution or preservation solution.

4.

Macroscopic on-site quality evaluation (MOSE): When the endoscopist determines that sufficient white specimens have been collected, the study procedure is terminated. If the endoscopist determines that insufficient white specimen has been collected, the steps 1 through 4 are repeated until sufficient white specimen is collected; however, no more than five punctures shall be performed. The study procedure shall be terminated if no white specimen is collected after the fifth puncture.

Final diagnosis

The final diagnosis will be based on pathological findings from tissue obtained by EUS-FNB and/or surgery. When the pathological diagnosis is “not malignant”, or inconclusive due to insufficient material, follow-up EUS is to be conducted after 12 months from the date of the study procedure. The presence or absence of change is verified, and those with no change are considered benign, while that with change is subjected to comprehensive diagnosis through additional EUS-FNB and imaging modalities.

DefinitionsDefinition of adequate specimen obtained from the first puncture

An adequate specimen is defined as a specimen that contains a sufficient volume of tissue to allow for pathological immunostaining, and is judged by the pathologist to corresponds to category D (see below):

Specimen adequacy evaluation criteria A.

Insufficient for diagnosis (including insufficient specimen; only necrotic tissue collected).

B.

Diagnosis only at the cytology level (tissue architecture cannot be evaluated).

C.

Histological evaluation is possible, but additional tests such as immunostaining cannot be performed

D.

Histological evaluation is possible and additional tests such as immunostaining can be performed.

Study designPrimary endpoint 1.

The superiority of a 22G Fork-tip needle over a 22G Franseen needle for collection of an adequate tissue specimen at the first puncture.

Secondary endpoints 1.

Successful puncture rate

2.

Procedure completion rate

3.

Number of adverse events; diagnostic suitability (sensitivity, specificity, and positive diagnosis rate) of the first puncture specimen for GIST

4.

The number of punctures required until white specimen collection by MOSE.

Concomitantly prohibited drugs, concomitantly prohibited therapies

The use of antithrombotic medications when performing study procedures is in accordance with the Guidelines for Gastrointestinal Endoscopy Practice for Patients on Antithrombotic Medications [7].

Criteria for discontinuation of the entire research

1. When it is deemed difficult to complete the study due to delays in case enrollment, frequent deviations from the research protocol, or other reasons.

2. When the risk-to-benefit ratio is judged to be unacceptable (e.g., safety findings of the study, results of interim analyses, if any).

3. When the safety of the study is judged to be problematic or the continuation of the study is judged to be no longer meaningful as a result of evaluation of relevant information obtained from sources other than the study, such as articles and conference presentations.

Application for clinical research

The principal investigator shall obtain the approval of each institutional Review Board and the permission of the administrator of the implementing medical institution to conduct the research using this research protocol. The principal investigator also submitted the implementation plan to the Minister of Health and publish the research information in jRCT.

Availability of data and materials

Only the principal investigator, local co-investigator, and research staff have access to the final dataset solely for research purposes. The datasets generated and analyzed during the current study will not be made publicly available to protect participant privacy but are available from the corresponding author upon reasonable request.

Statistical analysisSample size calculation

A prior retrospective observational study that compared puncture a Fork-tip needle with a Franseen needle for EUS-FNB for solid SELs measuring ≤ 2 cm, reported an adequate specimen collection rate of 96% for the Fork-tip needle and 74% for the Franseen needle [6]. Similar results are expected from the present study. In this case, when Fisher's exact test is conducted at the significance level α = 0.05 (two-sided alternative hypothesis), the minimum number of cases required to obtain a power of 1-β of 80% or more is 98. The target number of enrolled cases is set at 110 (55 per group), which accounts for a 10% dropout rate and cases that are ineligible.

Randomization process

Randomization was performed using a modified version of Pocock and Simon's minimization method [8]. Additionally, the random number generation was carried out using a linear combination pseudorandom number generator [9].

Evaluation of endpointsPrimary endpoint

When a Full Analysis Set (FAS) is the target, data will be compared using Fisher's exact test. In this case, the significance level α is set at two-sided α = 0.05. If the p-value is below the significance level α, and the point estimation of the adequate specimen collection rate is higher for the Fork-tip 22G needle group than for the Franseen 22G needle group, the conclusion will be that the Fork-tip 22G needle is the more promising device. In addition to the above, the point estimation of the adequate specimen collection rate, and an exact 95% confidence interval using the Clopper-Pearson method, will be calculated for each group. The odds ratio and 95% confidence interval will also be calculated.

Secondary endpoints

When FAS is the target, an exact 95% confidence interval for each group will be calculated using the Clopper-Pearson. The odds ratio and 95% confidence interval will also be calculated. As a reference, the p-value will be calculated using Fisher's exact test.

Interim analyses

No interim analysis is planned.