Chatbots and COVID-19: Taking Stock of the Lessons Learned.

While digital innovation in health was already rapidly evolving, the COVID-19 pandemic has accelerated the generation of digital technology tools, such as chatbots, to help increase access to crucial health information and services to those who were cut off or had limited contact with health services. This theme issue titled "Chatbots and COVID-19" presents articles from researchers and practitioners across the globe, describing the development, implementation, and evaluation of chatbots designed to address a wide range of health concerns and services. In this editorial, we present some of the key challenges and lessons learned arising from the content of this theme issue. Most notably, we note that a stronger evidence base is needed to ensure that chatbots and other digital tools are developed to best serve the needs of population health.

Evaluation of Endobronchial Ultrasound-Guided Transbronchial Needle Aspiration (EBUS-TBNA) Samples from Advanced Non-Small Cell Lung Cancer for Whole Genome, Whole Exome and Comprehensive Panel Sequencing.

Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is often the only source of tumor tissue from patients with advanced, inoperable lung cancer. EBUS-TBNA aspirates are used for the diagnosis, staging, and genomic testing to inform therapy options. Here we extracted DNA and RNA from 220 EBUS-TBNA aspirates to evaluate their suitability for whole genome (WGS), whole exome (WES), and comprehensive panel sequencing. For a subset of 40 cases, the same nucleic acid extraction was sequenced using WGS, WES, and the TruSight Oncology 500 assay. Genomic features were compared between sequencing platforms and compared with those reported by clinical testing. A total of 204 aspirates (92.7%) had sufficient DNA (100 ng) for comprehensive panel sequencing, and 109 aspirates (49.5%) had sufficient material for WGS. Comprehensive sequencing platforms detected all seven clinically reported tier 1 actionable mutations, an additional three (7%) tier 1 mutations, six (15%) tier 2-3 mutations, and biomarkers of potential immunotherapy benefit (tumor mutation burden and microsatellite instability). As expected, WGS was more suited for the detection and discovery of emerging novel biomarkers of treatment response. WGS could be performed in half of all EBUS-TBNA aspirates, which points to the enormous potential of EBUS-TBNA as source material for large, well-curated discovery-based studies for novel and more effective predictors of treatment response. Comprehensive panel sequencing is possible in the vast majority of fresh EBUS-TBNA aspirates and enhances the detection of actionable mutations over current clinical testing.

Immune and genomic biomarkers of immunotherapy response in cancer of unknown primary.

BACKGROUND: Cancer of unknown primary (CUP) is a heterogeneous group of metastatic cancers where a primary tissue of origin (TOO) is uncertain. Most patients with CUP have limited treatment options and poor survival outcomes. Immune checkpoint inhibitors (ICIs) can be efficacious in some patients with CUP, but the optimal predictive biomarkers are unknown. We therefore assessed immune and genomic biomarkers as well as predicted TOO in patients with CUP, including a subset treated with ICIs. METHODS: Patients with CUP were subject to gene-expression profiling (GEP) and DNA panel sequencing. Immune and stromal-related gene expression was explored by NanoString, including genes associated with immunotherapy response (IR) in other solid malignancies. ICI responsive cancer types were assigned based on Food and Drug Administration-approved indications, and either detection of a latent primary tumor or the TOO was suspected based on genomics informed pathology review. Tumor mutation burden (TMB) and gene mutations were also assessed. RESULTS: A total of 219 patients with CUP were included, 215 assessed for TOO in a previous study, with the majority (163) receiving both RNA and DNA tests. Of GEP profiled cases, 33% (59/175) had a high IR gene-expression score. Of the DNA sequenced cases, 16% (32/203) had high TMB (>10 mutations/Mb), including two with mismatch repair deficiency. Low correlation was observed between TMB and an IR score (R=0.26, p<0.001). Among 110 CUPs with a latent primary or suspected TOO, 47% (52/110) belonged to ICI-responsive cancer types. More than half of the CUPs had at least one feature that may predict ICI response (high IR score, high TMB, ICI-responsive cancer type). Among patients with CUP treated with ICIs, 8/28 (29%) responded (2 complete responses and 6 partial responses). Among non-responders, 9 had stable and 11 had progressive disease. All responders had a high IR score (7/8) and/or high TMB (3/8), while most (5/8) belonged to ICI-responsive cancer types. These features were detected at a lower frequency in non-responders and mostly in patients with stable disease. CONCLUSIONS: A significant fraction of CUP tumors had genomic features previously associated with ICI response. High IR score was the most sensitive predictive feature of ICI response, warranting evaluation in a larger patient series.

A comparison of DNA sequencing and gene expression profiling to assist tissue of origin diagnosis in cancer of unknown primary.

Cancer of unknown primary (CUP) is a syndrome defined by clinical absence of a primary cancer after standardised investigations. Gene expression profiling (GEP) and DNA sequencing have been used to predict primary tissue of origin (TOO) in CUP and find molecularly guided treatments; however, a detailed comparison of the diagnostic yield from these two tests has not been described. Here, we compared the diagnostic utility of RNA and DNA tests in 215 CUP patients (82% received both tests) in a prospective Australian study. Based on retrospective assessment of clinicopathological data, 77% (166/215) of CUPs had insufficient evidence to support TOO diagnosis (clinicopathology unresolved). The remainder had either a latent primary diagnosis (10%) or clinicopathological evidence to support a likely TOO diagnosis (13%) (clinicopathology resolved). We applied a microarray (CUPGuide) or custom NanoString 18-class GEP test to 191 CUPs with an accuracy of 91.5% in known metastatic cancers for high-medium confidence predictions. Classification performance was similar in clinicopathology-resolved CUPs - 80% had high-medium predictions and 94% were concordant with pathology. Notably, only 56% of the clinicopathology-unresolved CUPs had high-medium confidence GEP predictions. Diagnostic DNA features were interrogated in 201 CUP tumours guided by the cancer type specificity of mutations observed across 22 cancer types from the AACR Project GENIE database (77,058 tumours) as well as mutational signatures (e.g. smoking). Among the clinicopathology-unresolved CUPs, mutations and mutational signatures provided additional diagnostic evidence in 31% of cases. GEP classification was useful in only 13% of cases and oncoviral detection in 4%. Among CUPs where genomics informed TOO, lung and biliary cancers were the most frequently identified types, while kidney tumours were another identifiable subset. In conclusion, DNA and RNA profiling supported an unconfirmed TOO diagnosis in one-third of CUPs otherwise unresolved by clinicopathology assessment alone. DNA mutation profiling was the more diagnostically informative assay. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Single-nuclei and bulk-tissue gene-expression analysis of pheochromocytoma and paraganglioma links disease subtypes with tumor microenvironment.

Pheochromocytomas (PC) and paragangliomas (PG) are rare neuroendocrine tumors associated with autonomic nerves. Here we use single-nuclei RNA-seq and bulk-tissue gene-expression data to characterize the cellular composition of PCPG and normal adrenal tissues, refine tumor gene-expression subtypes and make clinical and genotypic associations. We confirm seven PCPG gene-expression subtypes with significant genotype and clinical associations. Tumors with mutations in VHL, SDH-encoding genes (SDHx) or MAML3-fusions are characterized by hypoxia-inducible factor signaling and neoangiogenesis. PCPG have few infiltrating lymphocytes but abundant macrophages. While neoplastic cells transcriptionally resemble mature chromaffin cells, early chromaffin and neuroblast markers are also features of some PCPG subtypes. The gene-expression profile of metastatic SDHx-related PCPG indicates these tumors have elevated cellular proliferation and a lower number of non-neoplastic Schwann-cell-like cells, while GPR139 is a potential theranostic target. Our findings therefore clarify the diverse transcriptional programs and cellular composition of PCPG and identify biomarkers of potential clinical significance.

Finding the facts in an infodemic: framing effective COVID-19 messages to connect people to authoritative content.

The public's need for timely and trusted COVID-19 information remains high. Governments and global health agencies such as the WHO have sought to disseminate accurate and timely information to counteract misinformation and disinformation that has arisen as part of an 'infodemic'-the overabundance of information on COVID-19-some accurate and some not. In early 2020, WHO began a collaboration with Google to run online public service announcements on COVID-19, in the form of search ads displayed above results of Google Search queries. Web-based text ads can drive online searchers of COVID-19 information to authoritative COVID-19 content but determining what message is most effective is a challenge. WHO wanted to understand which message framing, that is, the way in which ad information is worded for the public, leads searchers to click through to WHO content. WHO tested 71 text ads in English across four COVID-19 topics using a mix of message frames: descriptive, collective, gain, loss, appeals to values and emphasising reasons. Between 11 September 2020 and 23 November 2020, there were 13 million views of the experimental WHO text ads leading to 1.4 million click-throughs to the WHO website. Within the set of 71 ads, there was a large spread between the most effective and least effective messages; for messages on COVID-19, the best performing framings were more than twice as effective as the worst performing framings (18.7% vs 8.5% engagement rate). Health practitioners can apply the messaging tactics WHO found to be successful to rapidly optimise messages for their own public health campaigns and better reach the public with authoritative information. Similar collaboration between big technology companies and governments and global health agencies has the potential to advance public health.

A randomised study of comfort during bronchoscopy comparing conscious sedation and anaesthetist-controlled general anaesthesia, including the utility of bispectral index monitoring.

BACKGROUND: The difference in patient comfort with conscious sedation versus general anaesthesia for bronchoscopy has not been adequately assessed in a randomised trial. This study aimed to assess if patient comfort during bronchoscopy with conscious sedation is noninferior to general anaesthesia. METHODS: 96 subjects were randomised to receive conscious sedation or general anaesthesia for bronchoscopy. The primary outcome was subject comfort. Secondary outcomes included willingness to undergo a repeat procedure if necessary and level of sedation assessed clinically and by bispectral index (BIS) monitoring. RESULTS: There was no significant difference between subject comfort scores (difference -0.01, 95% CI -0.63-0.61 on a 10-point scale; p=0.97) or willingness to undergo a repeat procedure (97.7% versus 91.8%, 95% CI -4.8-15.5%; p=0.37). Deeper levels of sedation in the general anaesthesia cohort was confirmed with both clinical and BIS monitoring. There was no significant difference in diagnostic accuracy (conscious sedation 93.9%, 95% CI 80.4-98.3% versus general anaesthesia 86.5%, 95% CI 72.0-94.1%; p=0.43). There were more complications (29.6%, 95% CI 18.2-44.2% versus 6.1%, 95% CI 2.1-16.5%; p<0.01) in the general anaesthesia group. There was no relationship between high BIS scores and subject discomfort. BIS levels <40 during a procedure were associated with increased complications. CONCLUSION: Conscious sedation is not inferior to general anaesthesia in providing patient comfort during bronchoscopy, despite lighter sedation, and is associated with fewer complications and comparable diagnostic accuracy. BIS monitoring may have a role in preventing complications associated with deeper sedation.

Annexin A1 Is Required for Efficient Tumor Initiation and Cancer Stem Cell Maintenance in a Model of Human Breast Cancer.

Triple-negative breast cancer (TNBC) has a poor outcome compared to other breast cancer subtypes, and new therapies that target the molecular alterations driving tumor progression are needed. Annexin A1 is an abundant multi-functional Ca2+ binding and membrane-associated protein. Reported roles of Annexin A1 in breast cancer progression and metastasis are contradictory. Here, we sought to clarify the functions of Annexin A1 in the development and progression of TNBC. The association of Annexin A1 expression with patient prognosis in subtypes of TNBC was examined. Annexin A1 was stably knocked down in a panel of human and murine TNBC cell lines with high endogenous Annexin A1 expression that were then evaluated for orthotopic growth and spontaneous metastasis in vivo and for alterations in cell morphology in vitro. The impact of Annexin A1 knockdown on the expression of genes involved in mammary epithelial cell differentia tion and epithelial to mesenchymal transition was also determined. Annexin A1 mRNA levels correlated with poor patient prognosis in basal-like breast tumors and also in the basal-like 2 subset of TNBCs. Unexpectedly, loss of Annexin A1 expression had no effect on either primary tumor growth or spontaneous metastasis of MDA-MB-231_HM xenografts, but abrogated the growth rate of SUM149 orthotopic tumors. In an MMTV-PyMT driven allograft model of breast cancer, Annexin A1 depletion markedly delayed tumor formation in both immuno-competent and immuno-deficient mice and induced epithelial to mesenchymal transition and upregulation of basal markers. Finally, loss of Annexin A1 resulted in the loss of a discrete CD24+/Sca1- population containing putative tumor initiating cells. Collectively, our data demonstrate a novel cell-autonomous role for Annexin A1 in the promotion of tumor-forming capacity in a model of human breast cancer and suggest that some basal-like TNBCs may require high endogenous tumor cell Annexin A1 expression for continued growth.

γδ T Cells in Merkel Cell Carcinomas Have a Proinflammatory Profile Prognostic of Patient Survival.

Merkel cell carcinomas (MCC) are immunogenic skin cancers associated with viral infection or UV mutagenesis. To study T-cell infiltrates in MCC, we analyzed 58 MCC lesions from 39 patients using multiplex-IHC/immunofluorescence (m-IHC/IF). CD4+ or CD8+ T cells comprised the majority of infiltrating T lymphocytes in most tumors. However, almost half of the tumors harbored prominent CD4/CD8 double-negative (DN) T-cell infiltrates (>20% DN T cells), and in 12% of cases, DN T cells represented the majority of T cells. Flow cytometric analysis of single-cell suspensions from fresh tumors identified DN T cells as predominantly Vδ2- γδ T cells. In the context of γδ T-cell inflammation, these cells expressed PD-1 and LAG3, which is consistent with a suppressed or exhausted phenotype, and CD103, which indicates tissue residency. Furthermore, single-cell RNA sequencing (scRNA-seq) identified a transcriptional profile of γδ T cells suggestive of proinflammatory potential. T-cell receptor (TCR) analysis confirmed clonal expansion of Vδ1 and Vδ3 clonotypes, and functional studies using cloned γδ TCRs demonstrated restriction of these for CD1c and MR1 antigen-presenting molecules. On the basis of a 13-gene γδ T-cell signature derived from scRNA-seq analysis, gene-set enrichment on bulk RNA-seq data showed a positive correlation between enrichment scores and DN T-cell infiltrates. An improved disease-specific survival was evident for patients with high enrichment scores, and complete responses to anti-PD-1/PD-L1 treatment were observed in three of four cases with high enrichment scores. Thus, γδ T-cell infiltration may serve as a prognostic biomarker and should be explored for therapeutic interventions.See related Spotlight on p. 600.

Clinical, FDG-PET and molecular markers of immune checkpoint inhibitor response in patients with metastatic Merkel cell carcinoma.

BACKGROUND: Metastatic Merkel cell carcinoma (mMCC) is an aggressive neuroendocrine malignancy of the skin with a poor prognosis. Immune checkpoint inhibitors (ICIs) have shown substantial efficacy and favorable safety in clinical trials. METHODS: Medical records of patients (pts) with mMCC treated with ICIs from August 2015 to December 2018 at Peter MacCallum Cancer Centre in Australia were analyzed. Response was assessed with serial imaging, the majority with FDG-PET/CT scans. RNA sequencing and immunohistochemistry for PD-L1, CD3 and Merkel cell polyomavirus (MCPyV) on tumor samples was performed. RESULTS: 23 pts with mMCC were treated with ICIs. A median of 8 cycles (range 1 to 47) were administered, with treatment ongoing in 6 pts. Objective responses (OR) were observed in 14 pts (61%): 10 (44%) complete responses (CR) and 4 (17%) partial responses (PR). Median time to response was 8 weeks (range 6 to 12) and 12-month progression-free survival rate was 39%. Increased OR were seen in pts aged less than 75 (OR 80% vs 46%), no prior history of chemotherapy (OR 64% vs 50%), patients with an immune-related adverse event (OR 100% vs 43%) and in MCPyV-negative tumors (OR 69% vs 43%). Pts with a CR had lower mean metabolic tumor volume on baseline FDG-PET/CT scan (CR: 35.7 mL, no CR: 187.8 mL, p=0.05). There was no correlation between PD-L1 positivity and MCPyV status (p=0.764) or OR (p=0.245). 10 pts received radiation therapy (RT) during ICI: 4 pts started RT concurrently (OR 75%, CR 50%), 3 pts had isolated ICI-resistant lesions successfully treated with RT and 3 pts with multisite progression continued to progress despite RT. Overall, 6 pts (26%) had grade 1-2 immune-related adverse events. CONCLUSION: ICIs showed efficacy and safety in mMCC consistent with trial data. Clinical and imaging predictors of response were identified.

CUP-AI-Dx: A tool for inferring cancer tissue of origin and molecular subtype using RNA gene-expression data and artificial intelligence.

BACKGROUND: Cancer of unknown primary (CUP), representing approximately 3-5% of all malignancies, is defined as metastatic cancer where a primary site of origin cannot be found despite a standard diagnostic workup. Because knowledge of a patient's primary cancer remains fundamental to their treatment, CUP patients are significantly disadvantaged and most have a poor survival outcome. Developing robust and accessible diagnostic methods for resolving cancer tissue of origin, therefore, has significant value for CUP patients. METHODS: We developed an RNA-based classifier called CUP-AI-Dx that utilizes a 1D Inception convolutional neural network (1D-Inception) model to infer a tumor's primary tissue of origin. CUP-AI-Dx was trained using the transcriptional profiles of 18,217 primary tumours representing 32 cancer types from The Cancer Genome Atlas project (TCGA) and International Cancer Genome Consortium (ICGC). Gene expression data was ordered by gene chromosomal coordinates as input to the 1D-CNN model, and the model utilizes multiple convolutional kernels with different configurations simultaneously to improve generality. The model was optimized through extensive hyperparameter tuning, including different max-pooling layers and dropout settings. For 11 tumour types, we also developed a random forest model that can classify the tumour's molecular subtype according to prior TCGA studies. The optimised CUP-AI-Dx tissue of origin classifier was tested on 394 metastatic samples from 11 tumour types from TCGA and 92 formalin-fixed paraffin-embedded (FFPE) samples representing 18 cancer types from two clinical laboratories. The CUP-AI-Dx molecular subtype was also independently tested on independent ovarian and breast cancer microarray datasets FINDINGS: CUP-AI-Dx identifies the primary site with an overall top-1-accuracy of 98.54% in cross-validation and 96.70% on a test dataset. When applied to two independent clinical-grade RNA-seq datasets generated from two different institutes from the US and Australia, our model predicted the primary site with a top-1-accuracy of 86.96% and 72.46% respectively. INTERPRETATION: The CUP-AI-Dx predicts tumour primary site and molecular subtype with high accuracy and therefore can be used to assist the diagnostic work-up of cancers of unknown primary or uncertain origin using a common and accessible genomics platform. FUNDING: NIH R35 GM133562, NCI P30 CA034196, Victorian Cancer Agency Australia.

The impact of concordance with a lung cancer diagnosis pathway guideline on treatment access in patients with stage IV lung cancer.

BACKGROUND: Timely access to treatment of lung cancer is dependent on efficient and appropriate patient assessment and early referral for diagnostic workup. This study assesses the impact of Cancer Care Ontario (CCO) Lung Cancer Diagnostic Pathway Guideline (LCDPG) concordance on access to treatment of stage IV lung cancer patients referred to the Diagnostic Assessment Program (DAP) at a Canadian tertiary cancer centre. METHODS: This retrospective cohort study includes patients diagnosed with clinical stage IV lung cancer referred to the DAP at a Canadian tertiary cancer centre between November 1, 2015 and May 31, 2017. Referral concordance was determined based on CCO LCDPG. The primary outcome; time to treatment from initial healthcare presentation; was compared between the concordant and discordant referrals. RESULTS: Two hundred patients were referred for clinical stage IV lung cancer during the study period. Of these referrals, 151 (75.5%) were assessed and referred in concordance with LCDPG. Guideline concordant referrals were associated with reduced time to treatment from first healthcare presentation compared with guideline discordant referrals (55.3 vs. 108.8 days, P<0.001). Time to diagnostic procedure (32.2 vs. 86.7 days, P<0.001) and decision to treat (38.5 vs. 93.8 days, P<0.001) were also reduced with guideline concordance. The most common reason for discordant assessment and referral was delayed or inadequate investigation of symptoms in a high risk patient (32.7% of discordant referrals). CONCLUSIONS: Guideline concordant assessment and referral of stage IV lung cancer patients results in reduced time to diagnosis and treatment. Future research and education should focus on improving factors that delay DAP referral.

A xenograft and cell line model of SDH-deficient pheochromocytoma derived from Sdhb+/- rats.

Tumors caused by loss-of-function mutations in genes encoding TCA cycle enzymes have been recently discovered and are now of great interest. Mutations in succinate dehydrogenase (SDH) subunits cause pheochromocytoma/paraganglioma (PCPG) and syndromically associated tumors, which differ phenotypically and clinically from more common SDH-intact tumors of the same types. Consequences of SDH deficiency include rewired metabolism, pseudohypoxic signaling and altered redox balance. PCPG with SDHB mutations are particularly aggressive, and development of treatments has been hampered by lack of valid experimental models. Attempts to develop mouse models have been unsuccessful. Using a new strategy, we developed a xenograft and cell line model of SDH-deficient pheochromocytoma from rats with a heterozygous germline Sdhb mutation. The genome, transcriptome and metabolome of this model, called RS0, closely resemble those of SDHB-mutated human PCPGs, making it the most valid model now available. Strategies employed to develop RS0 may be broadly applicable to other SDH-deficient tumors.

FGF13 promotes metastasis of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) represents 10-20% of all human ductal adenocarcinomas and has a poor prognosis relative to other subtypes, due to the high propensity to develop distant metastases. Hence, new molecular targets for therapeutic intervention are needed for TNBC. We recently conducted a rigorous phenotypic and genomic characterization of four isogenic populations of MDA-MB-231 human triple-negative breast cancer cells that possess a range of intrinsic spontaneous metastatic capacities in vivo, ranging from nonmetastatic (MDA-MB-231_ATCC) to highly metastatic to lung, liver, spleen and spine (MDA-MB-231_HM). Gene expression profiling of primary tumours by RNA-Seq identified the fibroblast growth factor homologous factor, FGF13, as highly upregulated in aggressively metastatic MDA-MB-231_HM tumours. Clinically, higher FGF13 mRNA expression was associated with significantly worse relapse free survival in both luminal A and basal-like human breast cancers but was not associated with other clinical variables and was not upregulated in primary tumours relative to normal mammary gland. Stable FGF13 depletion restricted in vitro colony forming ability in MDA-MB-231_HM TNBC cells but not in oestrogen receptor (ER)-positive MCF-7 or MDA-MB-361 cells. However, despite augmenting MDA-MB-231_HM cell migration and invasion in vitro, FGF13 suppression almost completely blocked the spontaneous metastasis of MDA-MB-231_HM orthotopic xenografts to both lung and liver while having negligible impact on primary tumour growth. Together, these data indicate that FGF13 may represent a therapeutic target for blocking metastatic outgrowth of certain TNBCs. Further evaluation of the roles of individual FGF13 protein isoforms in progression of the different subtypes of breast cancer is warranted.

Topconfects: a package for confident effect sizes in differential expression analysis provides a more biologically useful ranked gene list.

Differential gene expression analysis may discover a set of genes too large to easily investigate, so a means of ranking genes by biological interest level is desired. p values are frequently abused for this purpose. As an alternative, we propose a method of ranking by confidence bounds on the log fold change, based on the previously developed TREAT test. These confidence bounds provide guaranteed false discovery rate and false coverage-statement rate control. When applied to a breast cancer dataset, the top-ranked genes by Topconfects emphasize markedly different biological processes compared to the top-ranked genes by p value.

EBUS-TBNA Cytology Specimens are Predictive of Occupational Dust Exposure in Patients With Bilateral Mediastinal and Hilar Lymphadenopathy.

OBJECTIVES: The aim of this study was to describe the cytological features of patients with significant occupational dust exposure presenting with benign bilateral mediastinal and hilar lymphadenopathy (BHL). METHODS: A retrospective cohort study including patients undergoing EBUS-TBNA for investigation of benign BHL. Patient characteristics, dust exposure history, radiology, and cytology samples from EBUS-TBNA were assessed. RESULTS: EBUS-TBNA cytology in patients with exposure showed a significant increase in the presence of birefringent fibers (60.7% vs 19.2%, P = 0.001) and intracellular carbon pigment (75.0% vs 28.9%, P = 0.001) compared with patients without exposure. The presence of these two features together yielded a sensitivity of 53.6% and a specificity of 88.5%. CONCLUSION: In patients with BHL and a history of occupational dust exposure, the presence of birefringent fibers and intracellular carbon pigment in EBUS-TBNA cytology samples may assist in a diagnosis of lymphadenopathy due to occupational dust exposure.