Sampling Instruments

Whatever guidance system is used, several sampling instruments can be employed for the transbronchial approach to PPLs, alone or in association, to obtain diagnostic cells or tissue (Table 20.6). It must be emphasized that there are no standardized guidelines on the type of sampling tool or tools to be used. The use of one or more sampling instruments is linked to the operator habits or to the availability in each institution. This is one of the reasons that may explain the heterogeneity of results between studies reported in the literature with most of the guidance systems.

Furthermore, in the era of tailored therapy of lung cancer, the evaluation of diagnostic yield, as reported in many studies, should be associated with the assessment of tool-speci c adequacy for histotype de nition and for complete molecular testing for tumor genotyping. It would be desirable that all future studies on the value of diagnostic techniques in the eld of PPLs take this aspect into consideration when analyzing the results.

Bronchoalveolar lavage (BAL) or bronchial washing is a diffuse technique in the transbronchial approach to PPLs, especially in centers that are not provided by a guidance system. It is performed by introducing an amount of fuid (150 mL in case of BAL) through the segmental bronchus leading to the lesion and using the recovered liquid for a cytological evaluation.

In a study on 55 patients with peripheral lung cancer, BAL was diagnostic in only 28% of

Table 20.6  Sampling instrument utilized in the transbronchial approach to PPLs

Bronchoalveolar lavage (BAL)

Curette

Brushing

Catheter

Forceps biopsy

Flexible transbronchial needles (TBNA)

Triple needle cytology brush

GenCut core biopsy system

Criobiopsy

patients with nodular lesions. Diagnostic yield was higher (40%) in cancer with an in ltrative pattern, like bronchoalveolar carcinoma or carcinomatosis lymphangitis. On the contrary, BAL provided a high diagnostic yield in diseases other than lung cancer (e.g., tuberculosis of other infectious diseases) [52]. The low diagnostic accuracy of BAL and/or washing in case of localized peripheral lesions does not support their routine use as the only means of sampling, except cases where there is a suspicion of infectious disease.

There are few randomized studies comparing the diagnostic yield of different sampling instruments.

In a prospective randomized trial on 218 patients affected by PPL, Trisolini et al. [53] compared the diagnostic value of forceps biopsy (FB), transbronchial needle and bronchial washing. TBNA was more sensitive (65%) than either FB (45%) and bronchial washing (22%). Furthermore, TBNA was the only diagnostic procedure in 21% of patients with malignant lesions, and 27% of PPLs without bronchus sign.

In a systematic review and meta-analysis evaluating the accuracy of TBNA under fuoroscopic guidance in the diagnosis of PPL and comparing its diagnostic yield with FB, Mondoni et al. [54] included 18 studies. The overall diagnostic yield of TBNA was statistically higher when compared to FB (60% vs. 45%). The subgroup analyses documented a higher TBNA yield when the bronchus sign was present (70% vs. 51%), in the case of malignant lesions (55% vs. 17%) and for lesions >3 cm (81% vs. 55%).

The better results obtained by TBNA are due to the ability of the needle to penetrate the lesion even if it does not involve the mucosal surface or it is located in the peri-bronchial area (Fig. 20.6).

Despite this evidence, the employment of TBNA in the transbronchial approach to PPLs is still underutilized.

From the practical point of view, there are some tricks that must be considered to optimize the results of TBNA in this clinical setting. First of all, if the needle is used in association with

Fig. 20.6  Possible relationship between the nodule and the airways in which the needle may be the only sampling instrument able to penetrate the lesion. In (a) the lesion

compresses but not involves the mucosal surface. In (b) the lesion is located adjacent to a bronchial spur

forceps or brushing, it should be employed rst, since the previously performed samplings could induce perilesional bleeding that may allow to aspirate an increased amount of blood, thus reducing the diagnostic value of TBNA. The second trick is to choose a kind of needle with a great fexibility, allowing the insertion into the most angulated bronchi, like the peripheral airways of the upper lobes. In our experience, the most appropriate needles for peripheral lesions are those with a metallic sheath, that are very fexible and that remain straight after insertion, allowing to maintain their direction during progression in the periphery [20]. However, new very fexible plastic sheath needles that can be used also with ultrathin bronchoscopes, have been recently introduced into the market [55].

Another important point that must be emphasized is the way to manage the TBNA samples. The material obtained by TBNA is expelled using a syringe lled by air onto one or more slides (Fig. 20.7a). Sometimes it is not possible to expel the sample by pushing with the syringe, because the material coagulates inside the needle. In such cases, the needle stylet can be used to push out the content of the needle (Fig. 20.7b). While specimens retrieved with forceps biopsy are managed in a standard way (formalin xing, paraf n embedding), samples obtained with needle aspiration procedures provide material which can be processed in different ways. In our routine practice, the material obtained with the rst needle pass is expelled on a slide and smeared. If there is an abundant sample, it is possible to obtain two

or more slides with the same material, just imprinting one slide on another (Fig. 20.8a). The technique for smearing slides is shown in Fig. 20.8b, c. One slide is immediately stained for rapid on-site evaluation. If some tissue cores or clots are present on the slide, these are retrieved with a syringe needle and put in formalin.

Advantages to having smear cytology is that it is suitable for ROSE and it can also provide good material for immunocytochemistry (histological de nition) and also for next-generation ­sequencing (genotyping of the tumor). If ROSE is positive showing that the target has been centered, other TBNA passes are performed and the material is directly fushed into a formalin vial for cell block or tissue core evaluation (Fig. 20.9). This material is useful for some tumor markers,

like PDL1, and it could be treated as a histological sample, facilitating pathologists that are more con dent with tissue evaluation than with smeared cytology.

In the majority of studies on the transbronchial approach to PPLs, whatever guidance method was used, a higher diagnostic yield was obtained with the association of more than one sampling instrument.

In post-hoc analysis of patients included in the NAVIGATE trial [48] (1215 enrolled subjects that underwent EMN bronchoscopy for diagnosis of PPL), Gildea et al. compared the results obtained using a restricted number of tools (only biopsy forceps, standard cytology brush, and/or BAL) with an extensive multimodal strategy (biopsy forceps, cytology brush, TBNA, triple

Fig. 20.8  Methods to prepare cytological slides. (a) The material can be passed on other slides, just imprinting one slide on another (A2), in this way obtaining two slides with similar material (A3). (b) The material is smeared using a second slide placed over and gently swiped. Note

Fig. 20.9  Material obtained by TBNA, fushed directly into a formalin vial for cell block evaluation. A tissue core for histological evaluation is obtained by a 21 G needle

that the second slide is held slightly inclined and parallel to the other. (c) In this “fat” technique, the second slide is placed on top of the other and the two slides are gently pulled in opposite direction

needle cytology, core biopsy system, BAL). 86.8% of true positive diagnoses were obtained using multimodal strategies and, among the different instruments used, positive rates were highest for biopsy forceps and TBNA [56].

Since it is not feasible to use all the sampling instruments, based on the above-mentioned considerations, the most appropriate association is the employment of TBNA with forceps biopsy. TBNA provides a better sensitivity for malignant lesions, while biopsy is reported to provide a better yield in cases of benign lesions [28].

Among the sampling instruments that can be used for the transbronchial diagnosis of PPLs, it must be mentioned also cryobiopsy, that in the last decade has gained large diffusion as a commonly used tool in the diagnostic pathway of in ltrative lung diseases. Recently, several studies have been performed, using cryobiopsy for the diagnosis of PPLs. The aim of these studies was to verify if cryoprobes can provide a better sensitivity and a greater amount of histological material in comparison to conventional forceps biopsy.

Even if cryoprobe can be used with different guidance systems, the majority of the studies employed rEBUS to localize the lesion. A meta-­ analysis recently published by Sryma et al. [57] identi ed nine studies performed on 300 patients using cryobiopsy and rEBUS. Most of the studies used a 1.9 mm cryoprobe and a freezing time between 3 and 5 s. Even if some studies included in this meta-analysis showed a better diagnostic yield with cryobiopsy in eccentrically and adjacently orientated lesions, the overall pooled sensitivities of cryobiopsy and of forced biopsy were similar (77% vs. 72%) and not statistically different. One severe bleeding and three pneumothorax requiring tube placement were reported with cryobiopsy (major complication rate = 1.8%).

Further multicenter randomized studies are needed to establish the utility of cryobiopsy for PPLs in a real-life setting, and if such tool could provide advantages in terms of sample adequacy for tumor genotyping.

Rapid On-Site Cytological

Evaluation (ROSE)

Rapid on-site evaluation of the samples (ROSE) is a cytological diagnostic procedure that allows assessment of the adequacy of the material obtained during bronchoscopy. ROSE can be performed during transbronchial approach to PPLs, whatever guidance system is used.

Usually, ROSE is done when cytological sampling instruments like TBNA are employed, but it is possible to perform ROSE even with material obtained by forceps biopsy, using the so-called “squashing” (the biopsy fragment is squeezed onto a glass slide) or “rolling” (the biopsy fragment is rolled repeatedly over the slide, on which cellular material will be deposited) techniques.

Slides are immediately xed (95% alcohol or air dried, according to the type of stain used) and one is stained for ROSE. A variety of quick stain systems can be utilized, depending on the pathologist preference (Diff-Quick, Haemacolor Merk, rapid Papanicolaou, hematoxylin and eosin).

While there are several prospective randomized studies that evaluate the role of ROSE during transbronchial needle aspiration for mediastinal lesions, both with conventional TBNA or EBUSguided TBNA, there are few reports to assess the value of ROSE in the transbronchial approach to PPLs.

In 1995 our group published a study on 1027 patients affected by PPL that were approachedrst by fuoroscopic-guided transbronchial needle aspiration and biopsy. ROSE was performed in all cases and, if negative, immediately after patients underwent a percutaneous needle aspiration [28]. In this study, which showed the possibility to integrate transbronchial and percutaneous approaches during the same session, ROSE played a crucial role to indicate if the bronchoscopic samplings were diagnostic, avoiding in this way the unnecessary transthoracic puncture.

More recently, a randomized trial was performed to evaluate the role of ROSE during rEBUS [58]. 84 patients received rEBUS-guided biopsy with ROSE and 74 without