Feasibility of shape-sensing robotic-assisted bronchoscopy for biomarker identification in patients with thoracic malignancies.

OBJECTIVE: Molecular diagnostic assays require samples with high nucleic acid content to generate reliable data. Similarly, programmed death-ligand 1 (PD-L1) immunohistochemistry (IHC) requires samples with adequate tumor content. We investigated whether shape-sensing robotic-assisted bronchoscopy (ssRAB) provides adequate samples for molecular and predictive testing. METHODS: We retrospectively identified diagnostic samples from a prospectively collected database. Pathologic reports were reviewed to assess adequacy of samples for molecular testing and feasibility of PD-L1 IHC. Tumor cellularity was quantified by an independent pathologist using paraffin-embedded sections. Univariable and multivariable linear regression models were constructed to assess associations between lesion- and procedure-related variables and tumor cellularity. RESULTS: In total, 128 samples were analyzed: 104 primary lung cancers and 24 metastatic lesions. On initial pathologic assessment, ssRAB samples were deemed to be adequate for molecular testing in 84% of cases; on independent review of cellular blocks, median tumor cellularity was 60% (interquartile range, 25%-80%). Hybrid capture-based next-generation sequencing was successful for 25 of 26 samples (96%), polymerase chain reaction-based molecular testing (Idylla; Biocartis) was successful for 49 of 52 samples (94%), and PD-L1 IHC was successful for 61 of 67 samples (91%). Carcinoid and small cell carcinoma histologic subtype and adequacy on rapid on-site evaluation were associated with higher tumor cellularity. CONCLUSIONS: The ssRAB platform provided adequate tissue for next-generation sequencing, polymerase chain reaction-based molecular testing, and PD-L1 IHC in >80% of cases. Tumor histology and adequacy on intraoperative cytologic assessment might be associated with sample quality and suitability for downstream assays.

Phase I trial of the TNF-α inhibitor certolizumab plus chemotherapy in stage IV lung adenocarcinomas.

We previously identified a chemotherapy-induced paracrine inflammatory loop that paradoxically mitigates the anti-tumor effect of chemotherapy and triggers metastatic propagation in breast and lung cancer models. Therefore, we sought to further validate and translate these findings into patient care by coupling the anti-TNF-α drug certolizumab pegol with standard cisplatin doublet chemotherapy. Here we first validate the anti-metastatic effect of certolizumab in a liver-metastatic Lewis Lung Carcinoma model. We then evaluate the safety, efficacy, and pharmacodynamic effects of certolizumab with cisplatin and pemetrexed in an open label Phase 1 clinical trial (NCT02120807) of eighteen adult patients with stage IV lung adenocarcinomas. The primary outcome is maximum tolerated dose. Secondary outcomes are response rate and progression-free survival (PFS); pharmacodynamic changes in blood and tumor are evaluated as a correlative outcome. There were nine partial responses among 16 patients evaluable (56%, 95% CI 30 to 80%). The median duration of response was 9.0 months (range 5.9 to 42.6 months) and median PFS was 7.1 months (95% CI 6.3 to NR). The standard 400 mg dose of certolizumab, added to cisplatin and pemetrexed, is well-tolerated and, as a correlative endpoint, demonstrates potent pharmacodynamic inhibition of peripheral cytokines associated with the paracrine inflammatory loop.

Shape-Sensing Robotic-Assisted Bronchoscopy in the Diagnosis of Pulmonary Parenchymal Lesions.

BACKGROUND: The landscape of guided bronchoscopy for the sampling of pulmonary parenchymal lesions is evolving rapidly. Shape-sensing robotic-assisted bronchoscopy (ssRAB) recently was introduced as means to allow successful sampling of traditionally challenging lesions. RESEARCH QUESTION: What are the feasibility, diagnostic yield, determinants of diagnostic sampling, and safety of ssRAB in patients with pulmonary lesions? STUDY DESIGN AND METHODS: Data from 131 consecutive ssRAB procedures performed at a US-based cancer center between October 2019 and July 2020 were captured prospectively and analyzed retrospectively. Definitions of diagnostic procedures were based on prior standards. Associations of procedure- and lesion-related factors with diagnostic yield were examined by univariate and multivariate generalized linear mixed models. RESULTS: A total of 159 pulmonary lesions were targeted during 131 ssRAB procedures. The median lesion size was 1.8 cm, 59.1% of lesions were in the upper lobe, and 66.7% of lesions were beyond a sixth-generation airway. The navigational success rate was 98.7%. The overall diagnostic yield was 81.7%. Lesion size of ≥ 1.8 cm and central location were associated significantly with a diagnostic procedure in the univariate analysis. In the multivariate model, lesions of ≥ 1.8 cm were more likely to be diagnostic compared with lesions < 1.8 cm, after adjusting for lung centrality (OR, 12.22; 95% CI, 1.66-90.10). The sensitivity and negative predictive value of ssRAB for primary thoracic malignancies were 79.8% and 72.4%, respectively. The overall complication rate was 3.0%, and the pneumothorax rate was 1.5%. INTERPRETATION: This study was the first to provide comprehensive evidence regarding the usefulness and diagnostic yield of ssRAB in the sampling of pulmonary parenchymal lesions. ssRAB may represent a significant advancement in the ability to access and sample successfully traditionally challenging pulmonary lesions via the bronchoscopic approach, while maintaining a superb safety profile. Lesion size seems to remain the major predictor of a diagnostic procedure.

Putting morphology to the test: An established classification scheme reliably stratifies salivary gland cytology by risk of malignancy with substantial interobserver agreement.

BACKGROUND: The Milan System for Reporting Salivary Gland Cytopathology describes several salivary gland fine-needle aspiration cytology (SGFNAC) morphologies developed by Griffith et al. Basaloid neoplasms are pleomorphic (PB) or monomorphic with fibrillary (MBFib), hyaline (MBHy), or other (MBOther) matrix. Oncocytoid neoplasms can be pleomorphic (PO), demonstrate granular and/or vacuolated cytoplasm (OGV), or be monomorphic with mucinous (MOMuc), cystic (MOCyst), or other (MOOther) background. In the current study, the authors explore interobserver agreement (IOA) and risk of malignancy (ROM) for these subcategories. METHODS: The study included 169 SGFNAC cases with surgical follow-up. Four reviewers categorized these cases using the criteria of Griffith et al. with consensus determined by majority. For all morphologic categories, IOA (using the Fleiss kappa) and ROM were calculated. RESULTS: ROMs for basaloid categories were: PB: 100% (1 of 1 case); MBHy: 71.4% (5 of 7 cases); MBFib: 50.0% (3 of 6 cases); and MBOther: 47.4% (9 of 19 cases). ROMs for oncocytoid neoplasms were: OGV: 100% (10 of 10 cases); MOMuc: 92.3% (12 of 13 cases); PO: 88.9% (8 of 9 cases); MOOther: 33.3% (5 of 15 cases); and MOCyst: 0 (0 of 1 case). The system demonstrated substantial agreement overall (κ = 0.69). For basaloid neoplasms, the IOA results were: MBHy: κ = 0.59; MBFib: κ = 0.41; MBOther: κ = 0.41; and PB: κ = 0.11. For oncocytoid neoplasms, the IOA results were: MOMuc: κ = 0.88; OGV: κ = 0.67; PO: κ = 0.63; MOOther: κ = 0.57; and MOCyst: κ = 0.18. CONCLUSIONS: The SGFNAC scheme proposed by Griffith et al. and incorporated into the Milan System for Reporting Salivary Gland Cytopathology demonstrated substantial agreement overall, with particularly high agreement for the MOMuc, OGV, PO, and MBHy categories. The PB and MOCyst categories demonstrated slight agreement and may be improved by revised criteria. The PB, PO, MOMuc, and OGV categories demonstrated high ROM, and the latter 2 categories might best be classified as suspicious for malignancy.

Concurrent Mutations in STK11 and KEAP1 Promote Ferroptosis Protection and SCD1 Dependence in Lung Cancer.

Concurrent loss-of-function mutations in STK11 and KEAP1 in lung adenocarcinoma (LUAD) are associated with aggressive tumor growth, resistance to available therapies, and early death. We investigated the effects of coordinate STK11 and KEAP1 loss by comparing co-mutant with single mutant and wild-type isogenic counterparts in multiple LUAD models. STK11/KEAP1 co-mutation results in significantly elevated expression of ferroptosis-protective genes, including SCD and AKR1C1/2/3, and resistance to pharmacologically induced ferroptosis. CRISPR screening further nominates SCD (SCD1) as selectively essential in STK11/KEAP1 co-mutant LUAD. Genetic and pharmacological inhibition of SCD1 confirms the essentiality of this gene and augments the effects of ferroptosis induction by erastin and RSL3. Together these data identify SCD1 as a selective vulnerability and a promising candidate for targeted drug development in STK11/KEAP1 co-mutant LUAD.

SCLC Subtypes Defined by ASCL1, NEUROD1, POU2F3, and YAP1: A Comprehensive Immunohistochemical and Histopathologic Characterization.

INTRODUCTION: Recent studies have identified subtypes of small cell lung carcinoma (SCLC) defined by the RNA expression of ASCL1, NEUROD1, POU2F3, and YAP1 transcriptional regulators. There are only limited data on the distribution of these markers at the protein level and associated pathologic characteristics in clinical SCLC samples. METHODS: The expression of ASCL1, NEUROD1, POU2F3, and YAP1 was analyzed by immunohistochemistry in 174 patient samples with SCLC. Subtypes defined by these markers were correlated with histologic characteristics, expression of classic neuroendocrine markers (synaptophysin, chromogranin A, CD56, INSM1), and other SCLC markers, including the neuroendocrine phenotype-associated markers TTF-1 and DLL3. RESULTS: ASCL1 and NEUROD1 expression had the following distribution: (1) 41% ASCL1+/NEUROD1-; (2) 37% ASCL1+/NEUROD1+; (3) 8% ASCL1-/NEUROD1+; and (4) 14% ASCL1-/NEUROD1-. On the basis of their relative expression, 69% of cases were ASCL1-dominant and 17% were NEUROD1-dominant. POU2F3 was expressed in 7% of SCLC and was mutually exclusive of ASCL1 and NEUROD1. YAP1 was expressed at low levels, primarily in combined SCLC, and was not exclusive of other subtypes. Both ASCL1-dominant and NEUROD1-dominant subtypes were associated with neuroendocrine markerhigh/TTF-1high/DLL3high profile, whereas POU2F3 and other ASCL1/NEUROD1 double-negative tumors were neuroendocrine markerlow/TTF-1low/DLL3low. CONCLUSIONS: This is the first comprehensive immunohistochemical and histopathologic analysis of novel SCLC subtypes in patient samples. We confirm that ASCL1/NEUROD1 double-negative tumors represent a distinct neuroendocrine-low subtype of SCLC, which is either uniquely associated with POU2F3 or lacks a known dominant regulator. The expression profiles of these markers appear more heterogeneous in native samples than in experimental models, particularly with regard to the high prevalence of ASCL1/NEUROD1 coexpression. These findings may have prognostic and therapeutic implications and warrant further clinical investigation.

Insights into pathogenesis of fatal COVID-19 pneumonia from histopathology with immunohistochemical and viral RNA studies.

INTRODUCTION: We describe post-mortem pulmonary histopathologic findings of COVID-19 pneumonia in patients with a spectrum of disease course, from rapid demise to prolonged hospitalisation. METHODS AND RESULTS: Histopathologic findings in post-mortem lung tissue from eight patients who died from COVID-19 pneumonia were reviewed. Immunohistochemistry (IHC) and next-generation sequencing (NGS) were performed to detect virus. Diffuse alveolar damage (DAD) was seen in all cases with a spectrum of acute phase and/or organising phase. IHC with monoclonal antibodies against SARS-CoV-2 viral nucleoprotein and spike protein detected virus in areas of acute but not organising DAD, with intracellular viral antigen and RNA expression seen predominantly in patients with duration of illness less than 10 days. Major vascular findings included thrombi in medium- and large-calibre vessels, platelet microthrombi detected by CD61 IHC and fibrin microthrombi. CONCLUSIONS: Presence of SARS-CoV-2 viral RNA by NGS early in the disease course and expression of viral antigen by IHC exclusively in the acute, but not in the organising phase of DAD, suggests that the virus may play a major role in initiating the acute lung injury of DAD, but when DAD progresses to the organising phase the virus may have been cleared from the lung by the patient's immune response. These findings suggest the possibility of a major change during the disease course of COVID-19 pneumonia that may have therapeutic implications. Frequent thrombi and microthrombi may also present potential targets for therapeutic intervention.

HER2-Mediated Internalization of Cytotoxic Agents in ERBB2 Amplified or Mutant Lung Cancers.

Amplification of and oncogenic mutations in ERBB2, the gene encoding the HER2 receptor tyrosine kinase, promote receptor hyperactivation and tumor growth. Here we demonstrate that HER2 ubiquitination and internalization, rather than its overexpression, are key mechanisms underlying endocytosis and consequent efficacy of the anti-HER2 antibody-drug conjugates (ADC) ado-trastuzumab emtansine (T-DM1) and trastuzumab deruxtecan (T-DXd) in lung cancer cell lines and patient-derived xenograft models. These data translated into a 51% response rate in a clinical trial of T-DM1 in 49 patients with ERBB2-amplified or -mutant lung cancers. We show that cotreatment with irreversible pan-HER inhibitors enhances receptor ubiquitination and consequent ADC internalization and efficacy. We also demonstrate that ADC switching to T-DXd, which harbors a different cytotoxic payload, achieves durable responses in a patient with lung cancer and corresponding xenograft model developing resistance to T-DM1. Our findings may help guide future clinical trials and expand the field of ADC as cancer therapy. SIGNIFICANCE: T-DM1 is clinically effective in lung cancers with amplification of or mutations in ERBB2. This activity is enhanced by cotreatment with irreversible pan-HER inhibitors, or ADC switching to T-DXd. These results may help address unmet needs of patients with HER2-activated tumors and no approved targeted therapy.See related commentary by Rolfo and Russo, p. 643.This article is highlighted in the In This Issue feature, p. 627.

Ultrarapid EGFR Mutation Screening Followed by Comprehensive Next-Generation Sequencing: A Feasible, Informative Approach for Lung Carcinoma Cytology Specimens With a High Success Rate.

INTRODUCTION: For patients with advanced NSCLC, cytologic samples may be the only diagnostic specimen available for molecular profiling. Although both rapid and comprehensive assessment are essential in this setting, an integrated multitest approach remains an important strategy in many laboratories, despite the risks and challenges when working with scant samples. In this study, we describe our experience and high success rate in using a multitest approach, focusing on the clinical validation and incorporation of ultrarapid EGFR testing using the Idylla system followed by comprehensive next-generation sequencing (NGS). METHODS: Cytology samples received for routine molecular testing were included in this study. The performance characteristics of the EGFR Idylla assay were assessed; tissue suitability parameters and interpretation criteria to supplement automated mutation calling were established. The assay performance was monitored for 1 year, comparing the results with those of concurrent NGS testing by MSK-IMPACT (primarily) or MSK-AmpliSeq and MSK-Fusion solid panel in a subset of cases. RESULTS: Overall, 301 samples were studied; 83 samples were included in validation (60.2% [50 of 83] were positive for EGFR mutations). Concordance with the reference method was 96.4% (80 of 83) of the samples with excellent reproducibility. The limit of detection was variable depending on the total tissue input and the specific mutation tested. Unextracted tissue inputs that maintained total EGFR cycle of quantification at less than 23 allowed all mutations to be detected if present at greater than 5% variant allele frequency. Mutations could be detected at 1% variant allele frequency with total EGFR cycle of quantification of 18. During the clinical implementation phase, 218 NSCLC samples were tested by Idylla (24.3% [53 of 218] were EGFR mutation positive). Concurrent NGS testing was requested on 165 samples and successfully performed on 96.4% (159 of 165) of the samples. The Idylla automated results were concordant with those obtained by NGS in 96.2% (153 of 159) of cases and improved to 98.7% (157 of 159) after incorporation of manual review criteria to supplement automated calling, resulting in a diagnostic sensitivity of 95.6% (95% confidence interval: 84.9%-99.5%). In general, 9% (14 of 159) of the cases tested by NGS had EGFR mutations not covered by the Idylla assay, primarily insertions in exon 19 and 20 and minor mutations cooccurring with canonical sensitizing mutations. CONCLUSIONS: Comprehensive molecular testing is feasible and has a high success rate in NSCLC cytology samples when using a multitest approach. Testing with the Idylla system enables rapid and accurate determination of the EGFR status without compromising subsequent NGS testing.

The Milan System at Memorial Sloan Kettering: Utility of the categorization system for in-house salivary gland fine-needle aspiration cytology at a comprehensive cancer center.

BACKGROUND: The Milan System for Reporting Salivary Gland Cytopathology (MSRSGC) provides a standardized reporting system for salivary gland fine-needle aspiration (SGFNA). We review the clinical utility of the MSRSGC at a tertiary care cancer center by assessing the rates of malignancy (ROM) among different categories. METHODS: A retrospective search was performed to retrieve all SGFNA cases performed at our institution between 1/1/07 and 12/31/18. The initial primary diagnoses were recorded and cases were then assigned to appropriate MSRSGC categories. ROM was then calculated for all categories. RESULTS: A total of 976 cases were identified, and 373 with follow-up. The ROM was 19.7% (192/976) for all-comers and 51.3% (192/374) among cases with follow-up. Using MSRSGC, SGFNA showed a sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of 65.6%, 87.4%, 100%, and 72.6%, respectively. ROM for MSRSGC categories I, II, III, IVa, IVb, V, and VI were 20.7%, 30.0%, 45.8%, 3.3%, 50.7%, 100%, and 100%, respectively. Utilizing MSRSGC resulted in a nondiagnostic rate of 14.4%. The nondiagnostic rate was lower when the procedure was performed by pathologists vs nonpathologists (12.9% vs 15.8%) but was comparable when rapid on site evaluation (ROSE) was performed (12.9% vs 11.6%). CONCLUSION: In our patient population, MSRSGC resulted in a perfect PPV and moderate NPV. Utilizing MSRSGC results in a higher nondiagnostic rate due to the inclusion of cases with benign elements or cyst contents only in this category. Performing ROSE is more important in attaining an adequate sample than the specialty of the person performing SGFNA.

Cytologic evaluation of p16 staining in head and neck squamous cell carcinoma in CytoLyt versus formalin-fixed material.

BACKGROUND: The management of high-risk human papillomavirus (HR-HPV)-related oropharyngeal head and neck squamous cell carcinomas (HNSCCs) are distinct from HNSCC linked to smoking and alcohol use. HR-HPV-positive HNSCC frequently presents as a cervical lymph node metastasis. Because fine-needle aspiration (FNA) is often the initial diagnostic procedure, evaluating HR-HPV status in cytology specimens is important. The overexpression of p16 is a surrogate for HR-HPV; however, the evaluation of p16 in FNAs remains controversial. METHODS: From September 2015 to December 2016, cytopathologists performed 25 FNAs of neck lymph nodes that were suspicious for HR-HPV-positive HNSCC. Initial passes produced smears for on-site evaluation and CytoLyt material. Additional passes were formalin-fixed. A CytoLyt cell block (CCB) and a formalin-fixed cell block (FFCB) were prepared, and p16 immunocytochemistry was performed. RESULTS: In 24 of 25 cases, the FFCB had diffuse (≥70% of cells), strong nuclear/cytoplasmic p16 staining. In all 24 of these cases, HR-HPV was detected by in situ hybridization. The corresponding CCB had weak-to-moderate p16 staining in <70% of cells (range, 5%-60% of cells) in 17 cases, 4 had weak-to-moderate diffuse staining, and 4 were acellular. The percentage of p16-positive cells was significantly higher with FFCB than with CCB (formalin: 94% ± 2%; CytoLyt, 38% ± 7%; 2-tailed, paired Student t test; P < .001; Fisher exact test, P < .001). CONCLUSIONS: The fixative used had a drastic impact on p16 staining, which explained the staining variability reported in the literature. FFCBs show a diffuse staining pattern, which correlates with HR-HPV status, whereas CCBs show a weaker and inconsistent staining pattern, which is more difficult to interpret.

Comprehensive Next-Generation Sequencing Unambiguously Distinguishes Separate Primary Lung Carcinomas From Intrapulmonary Metastases: Comparison with Standard Histopathologic Approach.

PURPOSE: In patients with >1 non-small cell lung carcinoma (NSCLC), the distinction between separate primary lung carcinomas (SPLCs) and intrapulmonary metastases (IPMs) is a common diagnostic dilemma with critical staging implications. Here, we compared the performance of comprehensive next-generation sequencing (NGS) with standard histopathologic approaches for distinguishing NSCLC clonal relationships in clinical practice. EXPERIMENTAL DESIGN: We queried 4,119 NSCLCs analyzed by 341-468 gene MSK-IMPACT NGS assay for patients with >1 surgically resected tumor profiled by NGS. Tumor relatedness predicted by prospective histopathologic assessment was contrasted with comparative genomic profiling by subsequent NGS. RESULTS: Sixty patients with NGS performed on >1 NSCLCs were identified, yielding 76 tumor pairs. NGS classified tumor pairs into 51 definite SPLCs (median, 14; up to 72 unique somatic mutations per pair), and 25 IPMs (24 definite, one high probability; median, 5; up to 16 shared somatic mutations per pair). Prospective histologic prediction was discordant with NGS in 17 cases (22%), particularly in the prediction of IPMs (44% discordant). Retrospective review highlighted several histologic challenges, including morphologic progression in some IPMs. We subsampled MSK-IMPACT data to model the performance of less comprehensive assays, and identified several clinicopathologic differences between NGS-defined tumor pairs, including increased risk of subsequent recurrence for IPMs. CONCLUSIONS: Comprehensive NGS allows unambiguous delineation of clonal relationship among NSCLCs. In comparison, standard histopathologic approach is adequate in most cases, but has notable limitations in the recognition of IPMs. Our results support the adoption of broad panel NGS to supplement histology for robust discrimination of NSCLC clonal relationships in clinical practice.

CytoLyt fixation significantly inhibits MIB1 immunoreactivity whereas alternative Ki-67 clone 30-9 is not susceptible to the inhibition: Critical diagnostic implications.

BACKGROUND: The Ki-67 proliferation marker has multiple diagnostic and prognostic applications. Although several clones to the Ki-67 antigen are commercially available, the MIB1 clone is widely recommended in the surgical pathology literature for neuroendocrine tumors. In our cytopathology practice, we have encountered unexpectedly low MIB1 immunoreactivity in CytoLyt-fixed cell blocks (CBs). The current study evaluated the impact of fixatives, CB processing, and immunocytochemical (ICC) procedures on Ki-67 immunoreactivity. METHODS: Test CBs were prepared from freshly resected tumors, and multiple variables in the MIB1 ICC procedure were tested, including CytoLyt versus formalin collection media, MIB1 versus other Ki-67 clones including 30-9, and other variables. MIB1 versus Ki-67 30-9 clones were tested in parallel on CytoLyt-fixed CBs from clinical samples of small cell lung carcinoma (SCLC). RESULTS: In the test CBs (n = 10), the mean MIB1 labeling index was 10% in CytoLyt versus 47% in formalin (P = .0116), with a mean loss of reactivity in matched CBs of 37% (up to 70%). None of the procedure modifications tested in 223 individual ICC reactions recovered MIB1 reactivity in CytoLyt except for switching to the Ki-67 30-9 antibody. In CytoLyt-fixed SCLC samples (n = 14), the Ki-67 30-9 antibody demonstrated expected ranges of reactivity (mean, 83%; range, 60%-100%), whereas MIB1 demonstrated markedly inhibited labeling (mean, 60%; range, 10%-95%) (P = .0058). CONCLUSIONS: CytoLyt fixation substantially inhibits MIB1 immunoreactivity, whereas the Ki-67 30-9 clone is not susceptible to inhibition. Markedly discrepant MIB1 reactivity may present a pitfall in the diagnosis of SCLC and may lead to the incorrect prognostic stratification of other tumor types. For laboratories using CytoLyt, we recommend using the Ki-67 30-9 antibody rather than the MIB1 antibody.

Utility of Core Biopsy Specimen to Identify Histologic Subtype and Predict Outcome for Lung Adenocarcinoma.

BACKGROUND: Lung adenocarcinoma histologic subtype is an important indicator of patient outcomes, so preoperative knowledge of subtype may be helpful to guide surgical planning. We evaluated the sensitivity and prognostic efficacy of specimens from computed tomography-guided core needle biopsies to predict histologic subtype and patient outcome after surgery. METHODS: We retrospectively identified 221 patients with lung adenocarcinoma who underwent computed tomography-guided lung biopsy and subsequent surgical resection. Concordance, accuracy, specificity, and sensitivity of histologic subtypes from core biopsy specimens were compared with surgically resected specimens. Tumor characteristics and biopsy procedural factors were analyzed to determine impact on diagnostic sensitivity. Histologic subtype based on biopsy specimen, clinical, tumor, and treatment variables were also examined in relation to time to progression. RESULTS: Overall concordance of biopsy samples with the predominant subtype from surgical specimens was 77%. Specificity (sensitivity) of detecting a nonaggressive and aggressive subtype were 86% (93%) and 95% (48%), respectively. Length of core specimen and percentage subtype composition in the surgically resected specimen were correlated with improved sensitivity but to a lesser extent with aggressive subtypes. Presence of an aggressive subtype in biopsy specimens was an independent predictor of progression after surgery (subdistribution hazard ratio, 2.51; 95% confidence interval, 1.28-4.94; p = 0.0075). CONCLUSIONS: Specimens from computed tomography-guided core biopsies can predict lung adenocarcinoma progression after surgical resection. Future prospective studies should address the role of core biopsy in preoperative planning.

Outcomes of Stage III NSCLC with occult primary vs. known primary lesions.

OBJECTIVES: Occult primary non-small cell lung cancer (OP-NSCLC) involving mediastinal lymph nodes without an identifiable primary tumor is a rare presentation, with little known about how outcomes compare to typical Stage III NSCLC. We reviewed our experience treating OP-NSCLC with definitive radiotherapy and compared outcomes to a contemporary cohort of stage III NSCLC patients. MATERIALS AND METHODS: We reviewed 605 patients with stage III NSCLC staged with PET-CT and treated with definitive radiotherapy between 1998 and 2013. Overall survival, intrathoracic control, and freedom from distant metastasis were computed using Kaplan-Meier method and logrank comparison. Cox hazard ratios were used to perform univariate and multivariate analyses. RESULTS: Twenty-one patients were identified with OP-NSCLC (3.5%). Patients with OP-NSCLC, as compared to known primary NSCLC, had significantly better 5-year rates of intrathoracic control (83.5% vs. 24.2%, P < 0.001), freedom from distant metastasis (59.0% vs. 26.3%, P = 0.003), and overall survival (61.6% vs. 15.2%, P < 0.001). Multivariate analyses confirmed occult primary as an independent prognostic factor associated with a 70% reduction in risk of intrathoracic failure, a 55% reduction in risk of distant metastasis, and a 70% reduction in risk of death. CONCLUSION: To our knowledge, this is the largest reported series of OP-NSCLC and the first to compare it to a contemporary cohort of Stage III NSCLC with known primary lesion. Definitive radiation therapy was associated with favorable locoregional control and survival, particularly compared with typical stage III NSCLC. This difference suggests that occult primary NSCLC may be a distinct entity with different biology than typical NSCLC.

Ado-Trastuzumab Emtansine for Patients With HER2-Mutant Lung Cancers: Results From a Phase II Basket Trial.

Purpose Human epidermal growth factor receptor 2 ( HER2, ERBB2)-activating mutations occur in 2% of lung cancers. We assessed the activity of ado-trastuzumab emtansine, a HER2-targeted antibody-drug conjugate, in a cohort of patients with HER2-mutant lung cancers as part of a phase II basket trial. Patients and Methods Patients received ado-trastuzumab emtansine at 3.6 mg/kg intravenously every 3 weeks until progression. The primary end point was overall response rate using Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. A Simon two-stage optimal design was used. Other end points included progression-free survival and toxicity. HER2 testing was performed on tumor tissue by next generation sequencing, fluorescence in situ hybridization, immunohistochemistry, and protein mass spectrometry. Results We treated 18 patients with advanced HER2-mutant lung adenocarcinomas. The median number of prior systemic therapies was two (range, zero to four prior therapies). The partial response rate was 44% (95% CI, 22% to 69%), meeting the primary end point. Responses were seen in patients with HER2 exon 20 insertions and point mutations in the kinase, transmembrane, and extracellular domains. Concurrent HER2 amplification was observed in two patients. HER2 immunohistochemistry ranged from 0 to 2+ and did not predict response, and responders had low HER2 protein expression measured by mass spectrometry. The median progression-free survival was 5 months (95% CI, 3 to 9 months). Toxicities included grade 1 or 2 infusion reactions, thrombocytopenia, and elevated hepatic transaminases. No patient stopped therapy as a result of toxicity or died on study. Conclusion Ado-trastuzumab emtansine is an active agent in patients with HER2-mutant lung cancers. This is the first positive trial in this molecular subset of lung cancers. Further use and study of this agent are warranted.

Author Correction: Structure and Distribution of an Unrecognized Interstitium in Human Tissues.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Feasibility of endobronchial ultrasound transbronchial needle aspiration for massively parallel next-generation sequencing in thoracic cancer patients.

INTRODUCTION: Next-generation sequencing (NGS) allows for the identification of a growing number of therapeutic and prognostic molecular targets. However, NGS typically requires greater quantities of DNA than traditional molecular testing does. Endobronchial ultrasound transbronchial needle aspiration (EBUS-TBNA) is a minimally invasive procedure used to sample central thoracic lesions, but it is not well established whether this technique provides sufficient material for NGS. METHODS: We performed a retrospective review of EBUS-TBNA at our institution (3/1/14-9/28/16). NGS was performed using a comprehensive hybrid-capture based assay (MSK-IMPACT) that detects >340 gene mutations. Samples found to be diagnostic for malignancy and for which MSK-IMPACT had been attempted were identified. Pathologic and clinical data were obtained from the medical record, and the results of MSK-IMPACT were examined. RESULTS: In total, 784 EBUS-TBNA procedures were performed during the study period. MSK-IMPACT was requested for 115 malignant samples and was successful for 99 (86.1%), identifying an average of 12.7 mutations at a mean coverage depth of 806X. NGS was performed on paraffin-embedded cell blocks in 93 cases (93.9%) and on cell-free DNA in needle rinse fluid in 6 cases. The success rate of the assay improved significantly from the first third of cases (76.3%), to 92.3% for the final one-third of cases (p < 0.05). CONCLUSIONS: EBUS-TBNA reliably provided adequate tissue for hybrid capture NGS, and is a suitable option for comprehensive NGS testing in patients with thoracic malignancies.

Structure and Distribution of an Unrecognized Interstitium in Human Tissues.

Confocal laser endomicroscopy (pCLE) provides real-time histologic imaging of human tissues at a depth of 60-70 µm during endoscopy. pCLE of the extrahepatic bile duct after fluorescein injection demonstrated a reticular pattern within fluorescein-filled sinuses that had no known anatomical correlate. Freezing biopsy tissue before fixation preserved the anatomy of this structure, demonstrating that it is part of the submucosa and a previously unappreciated fluid-filled interstitial space, draining to lymph nodes and supported by a complex network of thick collagen bundles. These bundles are intermittently lined on one side by fibroblast-like cells that stain with endothelial markers and vimentin, although there is a highly unusual and extensive unlined interface between the matrix proteins of the bundles and the surrounding fluid. We observed similar structures in numerous tissues that are subject to intermittent or rhythmic compression, including the submucosae of the entire gastrointestinal tract and urinary bladder, the dermis, the peri-bronchial and peri-arterial soft tissues, and fascia. These anatomic structures may be important in cancer metastasis, edema, fibrosis, and mechanical functioning of many or all tissues and organs. In sum, we describe the anatomy and histology of a previously unrecognized, though widespread, macroscopic, fluid-filled space within and between tissues, a novel expansion and specification of the concept of the human interstitium.