Friday, September 22, 2023

Close-to-lesion transbronchial biopsy: a novel technique to improve suitability of specimens for genetic testing in patients with peripheral pulmonary lesions – Scientific Reports


This study enrolled 155 patients with peripheral pulmonary lesions that underwent bronchoscopy using a thin bronchoscope with RP-EBUS and guide sheath or an ultrathin bronchoscope with RP-EBUS from September 2021 through November 2022 (Fig. 1). The Ethics Committee for Clinical Investigation of Matsusaka Municipal Hospital approved the study protocol (Approval No.: j-241-230519-1-1), and the study was performed following the Principles of the Helsinki Declaration. Written informed consent was obtained from all study participants.

Figure 1

Study selection criteria. The clinical records of 352 patients that underwent bronchoscopy for parenchymal lesions were evaluated. Of these, 155 were eligible for the study after excluding patients with central or intermediate lung segmental lesions, ground glass nodules, cavitary lesions, patients undergoing bronchoscopy using thick bronchoscopes, and those lost to follow-up.

Study design

The current work is a retrospective and a single-institution study. The following variables were evaluated: patients’ gender and age, computed tomography (CT) bronchus sign, overall and histopathological diagnoses, bronchoscope tip close-to-lesion biopsy, and lesion size, RP-EBUS image, chest X-ray finding, fluoroscopy visualization, and lung distribution (2.5 cm from the hilum/intermediate/2.5 cm from the pleura)9.


All patients underwent CT (TSX-101A/QA, Canon Medical Corporation, Tochigi, Otawara, Japan) prior to surgery, and VBN images were generated using CT-Digital Imaging and Communications in Medicine (CT-DICOM) data (DirectPath; Cybernet Systems, Tokyo, Japan). Bronchoscopy was performed using the thin flexible bronchoscope type P290 (outer diameter of 4.2 mm; working channel diameter of 2.0 mm; Olympus Medical Systems, Tokyo, Japan) or the ultrathin bronchoscope type MP290F (outer diameter of 3.0 mm; working channel diameter of 1.7 mm; Olympus Medical Systems, Tokyo, Japan). Topical lidocaine was used for pharyngeal anesthesia, while intravenous midazolam and fentanyl were administered for patient sedation. The airways were nebulized with lidocaine using a 1.9-mm spray catheter (Spray Catheter; Olympus, Tokyo, Japan), and additional intravenous fentanyl was administered as needed. Bronchoscopy with a thin or ultrathin bronchoscope was conducted with VBN, fluoroscopy, and 1.4-mm RP-EBUS (UM-S2-17S, Olympus, Tokyo, Japan) with or without a 1.95-mm guide sheath (SG-200C, Olympus, Tokyo, Japan). Biopsy was performed using a 1.5-mm (FB-233D; Olympus, Tokyo, Japan) or a 1.9-mm forceps (FB-231D, Olympus, Tokyo, Japan). Bronchial brushing and washing with 20 mL of physiological saline were performed after a close-to-lesion transbronchial biopsy (CL-TBB). As previously described, RP-EBUS images were categorized as within or adjacent to the lesion or invisible3.

Transbronchial biopsy and NGS analysis

A tissue sampling was defined as a CL-TBB when the bronchoscope tip was within or adjacent to the lesion by RP-EBUS and ≤ 10 mm from the lesion under fluoroscopy (Fig. 2). We assessed the NGS suitability rate, the NGS analysis success rate, and the overall diagnostic yield of the technique. The NGS suitability rate was the proportion of samples that met the criteria for genetic analysis, and the NGS analysis success rate was the proportion of samples that allowed both DNA and RNA analysis. We fixed the biopsy specimens immediately with 10% neutral-buffered formalin at room temperature for 12–24 h and then embedded them in paraffin. An experienced pathologist evaluated the number of tumor cells in specimens stained with hematoxylin and eosin. The sample was suitable for genetic analysis when the number of tumor cells in the specimen was ≥ 200 or the proportion of tumor cells in the specimen was ≥ 30%. The attending physician judged the suitability of the sample for NGS analysis based on the clinical findings when the number of tumor cells was between 100 and 200. We sent the samples to LSI Medience Laboratories (Tokyo, Japan) to perform NGS analysis using Oncomine Dx Target Test (ODxTT; Ion Torrent PGM Dx Sequencer; Thermo Fisher Scientific)10.

Figure 2
figure 2

Criteria for CL-TBB. The distance from the RP-EBUS probe tip to the ultrasound transducer is 5 mm. “CL-TBB” was considered when the distance from the bronchoscope tip to the lesion was ≤ 10 mm under fluoroscopy and RP-EBUS images.


The final diagnosis in all cases was made based on histopathological, cytological, microbiological, culture, and clinical findings. Samples collected by bronchoscopy (lung tissue, bronchial washing, or brushing), CT-guided lung biopsy, or surgery were used for pathological diagnosis. Lung cancer was diagnosed when any of these samples confirmed malignancy. Organizing pneumonia was diagnosed when the bronchoscopy showed no significant findings, CT-guided lung biopsy or surgery was not performed, or a subsequent CT study confirmed the disappearance or shrinkage of the lesion.

CL-TBB using a thin bronchoscope

The thin bronchoscope was advanced through the bronchus segment and positioned as close to the lesion as possible under the airway lumen and VBN image guidance. Then, the RP-EBUS with the guide sheath was inserted through the 2-mm working channel of the thin bronchoscope until the RP-EBUS-guide sheath tip reached the lesion, and this can be visualized and confirmed by RP-EBUS. When the RP-EBUS image showed within or adjacent to the lesion, the bronchoscope tip was further advanced as close to the lesion as possible using the RP-EBUS-guide sheath as a rail under RP-EBUS and fluoroscopy guidance (Fig. 3, Supplementary Video). During this close approach to the lesion, an assistant holds the RP-EBUS-guide sheath while the bronchoscopist further advances the bronchoscope tip toward the lesion of interest using the RP-EBUS-guide sheath as a rail. After confirming that the bronchoscope tip was close to the lesion and within or adjacent to the lesion by RP-EBUS image, the RP-EBUS was removed, leaving the guide sheath in position. A 1.5-mm forceps was then inserted through the guide sheath, following the same route toward the lesion to collect five pieces of the tissue sample. After collecting the samples and confirming that the lesion was visualized by fluoroscopy and the RP-EBUS image was within or adjacent to the lesion, the guide sheath was removed to introduce a 1.9-mm forceps and collect five additional biopsy samples. Therefore, a total of 10 visible tissue samples were collected.

Figure 3
figure 3

CL-TBB using a thin bronchoscope. (a) A patient with a peripheral pulmonary lesion. (b) The bronchoscope is inserted, and the RP-EBUS image shows that the probe is adjacent to the lesion. (c) The bronchoscope tip is moved closer to the lesion under RP-EBUS and fluoroscopy guidance. (d) The bronchoscope tip is pushed further using the RP-EBUS-guide sheath as a rail while checking the RP-EBUS image for the lesion position. The bronchoscope is moved in different directions to get the RP-EBUS image within the lesion. (e) The RP-EBUS is removed, and a 1.5-forceps is inserted through the guide sheath to collect 5 tissue samples. The guide sheath is removed, and a standard 1.9-mm forceps is inserted to collect 5 more tissue samples.

CL-TBB using an ultrathin bronchoscope

The thin bronchoscope was substituted by the ultrathin bronchoscope when the close-to-lesion approach was impossible because the lesion was invisible by RP-EBUS or because the area of contact with the lesion was less than 180° when the RP-EBUS image was adjacent to the lesion. Neither the guide sheath (1.95-mm) nor the standard (1.9-mm) forceps can be used with the ultrathin flexible bronchoscope because its working channel is only 1.7-mm in diameter. Therefore, the biopsy was performed after bronchoscope substitution using the 1.5-mm forceps under RP-EBUS (1.4-mm) and fluoroscopy guidance. The technique for approaching close to the lesion was similar to that used with the thin bronchoscope. The ultrathin bronchoscope was advanced through the bronchial segments toward the lesion under RP-EBUS guidance until the bronchoscope tip was as close to the lesion as possible. The RP-EBUS was then inserted through the bronchoscope working channel until the RP-EBUS tip reached the lesion area, and the lesion could be visualized and confirmed by fluoroscopy and RP-EBUS. After confirming the RP-EBUS image was within or adjacent to the lesion, the RP-EBUS was withdrawn to introduce a 1.5-mm forceps and collect ten tissue samples.

Evaluation and prevention of complications

Medical records were examined for procedure-related complications within one month after the bronchoscopy. After the transbronchial biopsy, the forceps was withdrawn, but the bronchoscope tip or guide sheath was kept securely in the bronchial branch for 3 min for compressive hemostasis to prevent bleeding.

Statistical analysis

SPSS Statistics version 23 software (SPSS Inc., Chicago, IL, USA) was used for statistical analysis. The chi-square test was used to compare proportions between groups, and the Mann–Whitney U test was used to analyze continuous variables. Logistic regression analysis was used to identify factors that can predict the suitability of collected specimens for NGS analysis. A two-sided P < 0.05 was considered significant.

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