In recent years, there has been tremendous therapeutic progress for advanced lung cancer, leading to the identification of a multitude of therapeutic targets and significantly expanding the list of potential target genes to be tested. However, precision oncology requires knowledge of the exact biology of the tumor through sequencing of the cancer genome, which is hampered by limited tissue availability in thoracic malignancies. Liquid biopsy, namely plasma cell-free DNA (cfDNA) analysis, has expanded these opportunities and is now firmly established in the diagnostic algorithm of patients with lung cancer with metastatic disease. However, the full potential of this powerful tool has been largely underexplored. In this issue of Cancer Research, Nair and colleagues provide evidence of the clinical utility of bronchoalveolar lavage (BAL) cfDNA profiling through an ultra-deep next-generation sequencing approach. The study findings support further development of BAL cfDNA analysis for tumor genotyping, besides the current gold standard sources (tissue and plasma), as a complementary tool in patients undergoing diagnostic bronchoscopy for tumor diagnosis and/or rebiopsy, increasing the success rates of genomic analyses. In addition, BAL cfDNA profiling might represent an important diagnostic tool in early-stage lung cancer, outperforming plasma cfDNA in stage I–II and detecting field cancerization signs, potentially identifying tumors before their clinical appearance. Further studies should confirm the full potential of BAL cfDNA profiling in lung cancer and its place in the large family of liquid biopsies.

See related article by Nair et al., p. 2838

Precision oncology tailors the peculiar biological features of the tumor, selectively targeting the genetic vulnerabilities of cancer cells. Therefore, it is of paramount importance to obtain knowledge of the exact biology of the tumor through sequencing the cancer genome. Tissue availability for tumor genotyping is a clinically relevant issue in lung cancer and liquid biopsy represents an important resource in patients with metastatic disease. Accumulating evidence suggest that plasma cell-free DNA (cfDNA) analyses can be not only an alternative source when tissue for tumor genotyping is scant or inadequate, but also can be used upfront given the lower turnaround time, potentially increasing the detection rates of clinically relevant oncogenic drivers (1, 2). For these reasons, this “plasma-first” approach has been included in international guidelines for the optimal use of liquid biopsy in lung cancer, allowing the use of cfDNA analysis either sequentially or concurrently with tissue genotyping (3). Robust evidence supports a good concordance rate for plasma and tissue genotyping using commercially available and FDA-approved cfDNA NGS platforms (1, 4), but unfortunately in some cases, such as in metastatic disease limited to the thorax and isolated central nervous disease progression, tumor DNA shedding in the circulation can be low (5), limiting the clinical utility of these methodologies. In addition, the use of plasma circulating tumor DNA (ctDNA) analyses in very early-stage lung cancer (T1a-b) for early cancer detection is challenged by the low abundance of ctDNA in the plasma of these patients, which in most cases exhibit a mutant allele frequency lower than the limits of detection of currently available technologies (6).

Lung cancer diagnosis often requires a bronchoscopy for tumor confirmation and rebiopsies are increasingly used for tracking clonal evolution of the tumors, identifying targetable mechanisms of acquired resistance to targeted therapies. Bronchoalveolar lavage (BAL) performed during bronchoscopic procedures might represent a precious source for cfDNA analyses, in addition to the applications it is currently used for like cytology and diagnosis of infections and interstitial lung diseases. DNA from cell-free lavage supernatants from patients with lung cancer can be isolated and used for genomic alterations detection (7), but the performance of BAL cfDNA tumor profiling as compared with plasma cfDNA analysis is largely unknown. In this issue of Cancer Research, Nair and colleagues (8) explored the potential of BAL cfDNA analyses for genotyping and detection of lung cancer, using an ultra-deep next-generation sequencing (NGS) approach. The study enrolled two distinct cohorts (Fig. 1), including 38 patients with lung cancer and 21 high-risk controls without cancer undergoing lung cancer screening. Using a tumor-informed approach, the authors showed that detection of tumor-derived mutations by Cancer Personalized Profiling by Deep Sequencing (CAPP-Seq) of BAL cfDNA is more sensitive than plasma profiling, with tumor-derived variants identified in 81% and 47% of the samples, respectively (8). These results suggest that BAL fluid might represent an additional source for tumor genotyping in addition to tissue testing and plasma cfDNA analysis in lung cancer patients both at diagnosis and at the time of rebiopsy. A complementary tissue and BAL approach might increase the adequacy of samples obtained from bronchoscopy, especially when plasma cfDNA analysis is expected to fail due to low ctDNA levels. Instances where cfDNA analysis may be unsuitable include when disease is limited to sanctuary sites or during oligo-progression. This is critical issue in clinical practice, as transbronchial biopsy and endobronchial ultrasound-guided transbronchial needle aspiration might be associated with a non-negligible rate of genomic analysis failure, requiring a repeated bronchoscopy (9). Concurrent use of BAL cfDNA and tumor tissue profiling could increase the diagnostic yield, minimizing the number of patients requiring repetitive invasive procedures and increasing savings of costs and time.

Figure 1.

Schematic representation of the study by Nair and colleagues (8) and potential clinical applications of BAL cfDNA analyses. CAPP-Seq, Cancer Personalized Profiling by Deep Sequencing; LDCT, low-dose CT; PDWB, plasma-depleted whole blood; STAMP, Stanford Actionable Mutation Panel.

Figure 1.

Schematic representation of the study by Nair and colleagues (8) and potential clinical applications of BAL cfDNA analyses. CAPP-Seq, Cancer Personalized Profiling by Deep Sequencing; LDCT, low-dose CT; PDWB, plasma-depleted whole blood; STAMP, Stanford Actionable Mutation Panel.

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In addition to tumor genotyping of advanced/metastatic lung cancer, BAL cfDNA analyses can potentially impact the diagnostic path of subjects with suspicious lung nodules undergoing bronchoscopy. In the accompanying study, Nair and colleagues (8) showed that BAL cfDNA tumor profiling can better identify lung cancer than plasma cfDNA with a significantly higher detection of tumor variants (P < 0.001 in all stages), including stage I–II tumors that are usually detected only in a minor fraction of the cases when using ultrasensitive plasma cfDNA profiling methods (6). In addition, the authors also performed a tumor uniformed analysis (i.e., without prior knowledge of the tumor mutation profile) demonstrating a better performance in BAL cfDNA (12/18, 67%) than plasma cfDNA in stage I–II disease (2/18, 11%; ref. 8). With the growing diffusion of low-dose computed tomography screening, the number of diagnostic bronchoscopies for suspicious early-stage lung cancers is destined to increase and BAL cfDNA profiling could increase the accuracy of currently used diagnostic procedures. Further studies confirming these findings and testing integrated approaches combining both BAL cfDNA and plasma cfDNA analyses are warranted. Finally, the evidence of mutations in cancer driver genes in BAL cfDNA without confirmation in the matching tumor specimens and the detection of these mutations also in high-risk controls without cancer further support the concept of field cancerization in lung cancer and might be exploited for intercepting cancer cases before their clinical evidence, a concept known as cancer interception (10).

Albeit promising, the results of this pivotal study should be confirmed in further larger studies, which will demonstrate the full potential of BAL cfDNA profiling in lung cancer and its place in the large family of liquid biopsies.

C. Rolfo reports grants from Lung Cancer Research Foundation-Pfizer Grant 2019, NCI SeroNet 2020, NIH SBIR, American Cancer Society; personal fees from MSD, GuardantHealth, AstraZeneca, Roche, Inivata, ArcherDx, EMD Serono, Novartis, BMS, Boston Pharmaceuticals, Eisai, BluePrint, CORE2, Pfizer; and other support from GuardantHealth outside the submitted work. U. Malapelle reports personal fees from Boehringer Ingelheim, Roche, MSD, Amgen, Thermo Fisher Scientific, Eli Lilly, Diaceutics, GSK, Merck and AstraZeneca, Janssen, Diatech, Novartis, Hedera outside the submitted work. A. Russo reports personal fees from MSD, Novartis, Pfizer, and personal fees from AstraZeneca outside the submitted work.

Figure 1 was created with BioRender.com.

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