Usefulness of an RNA extraction-free test for the multiplexed detection of ALK, ROS1, and RET Gene Fusions in Real Life FFPE Specimens of Non-Small Cell Lung Cancers.

BACKGROUND: ALK, ROS1 and RET rearrangements occur, respectively, in 5%, 2%, and 1% non-small cell lung cancers (NSCLC). ALK and ROS1 fusion proteins detection by immunohistochemistry (IHC) has been validated for rapid patient screening, but ROS1 fusions need to be confirmed by another technique and no RET IHC test is available for clinical use. RESEARCH DESIGN AND METHODS: We report herein the usefulness of the HTG EdgeSeq Assay, an RNA extraction-free test combining a quantitative nuclease protection assay with NGS, for the detection of ALK, ROS1 and RET fusions from 'real-life' small NSCLC samples. A total of 203 FFPE samples were collected from 11 centers. They included 143 rearranged NSCLC (87 ALK, 39 ROS1, 17 RET) and 60 ALK-ROS1-RET negative controls. RESULTS: The assay had a specificity of 98% and a sensitivity for ALK, ROS1 and RET fusions of 80%, 94% and 100% respectively. Among the 19 HTG-assay false negative samples, the preanalytical conditions were identified as the major factors impacting the assay efficiency. CONCLUSIONS: Overall, the HTG EdgeSeq assay offers comparable sensitivities and specificity than other RNA sequencing techniques, with the advantage that it can be used on very small and old samples collected multicentrically.

Multiomic analysis of malignant pleural mesothelioma identifies molecular axes and specialized tumor profiles driving intertumor heterogeneity.

Malignant pleural mesothelioma (MPM) is an aggressive cancer with rising incidence and challenging clinical management. Through a large series of whole-genome sequencing data, integrated with transcriptomic and epigenomic data using multiomics factor analysis, we demonstrate that the current World Health Organization classification only accounts for up to 10% of interpatient molecular differences. Instead, the MESOMICS project paves the way for a morphomolecular classification of MPM based on four dimensions: ploidy, tumor cell morphology, adaptive immune response and CpG island methylator profile. We show that these four dimensions are complementary, capture major interpatient molecular differences and are delimited by extreme phenotypes that-in the case of the interdependent tumor cell morphology and adapted immune response-reflect tumor specialization. These findings unearth the interplay between MPM functional biology and its genomic history, and provide insights into the variations observed in the clinical behavior of patients with MPM.

Micronodular thymic carcinoma with lymphoid hyperplasia: relevance of immunohistochemistry with a small panel of antibodies for diagnosis-a RYTHMIC study.

Micronodular thymic carcinoma with lymphoid hyperplasia (MNTCLH) is a rare form of thymic carcinoma. We present the experience of RYTHMIC, the French national network devoted to the treatment of thymic epithelial tumors through multidisciplinary tumor boards with a review of all tumors by pathologists for classification and staging. Six cases of MNTCLH were diagnosed during a review of 1007 thymic epithelial tumors. Histologically, epithelial cells with atypia and mitoses formed micronodules that were surrounded by an abundant lymphoid background with follicles. There was neither obvious fibro-inflammatory stroma nor necrosis. Spindle cells areas were common. Initial diagnosis was micronodular thymoma in two cases, cellular atypia being overlooked, eclipsed by the micronodular pattern. Immunohistochemistry with a panel of five antibodies showed that cytokeratins (AE1-AE3) and p63-positive epithelial cells also expressed CD5 and that there was no TdT-positive cells within the tumors. CD20 highlighted the lymphoid hyperplasia. Additionally epithelial cells also expressed CD117 and diffusely Glut 1. Twenty-seven micronodular thymomas with lymphoid stroma diagnosed during the same period did not show the CD5 and CD117 positivities seen in MNTCLH and contained TdT-positive lymphocytes. Three of the 6 patients with MNTCLH had adjuvant radiotherapy. Three patients with follow-up information were alive without recurrence at 38, 51, and 95 months. Our study shows that immunohistochemistry, such as that used in the RYTHMIC network with a small panel of antibodies, may easily help to confirm the correct diagnosis of MNTCLH, a rare and low-aggressive form of thymic carcinoma, and avoid the misdiagnosis of micronodular thymoma.

Impact of expert pathologic review of thymic epithelial tumours on diagnosis and management in a real-life setting: A RYTHMIC study.

BACKGROUND: Classification of thymic epithelial tumours (TETs) is known to be challenging; however, the level of discordances at a nationwide level between initial and expert diagnosis and their clinical consequences are currently unknown. RYTHMIC is a national network dedicated to the management of TET based on initial histological diagnosis, followed by an additional expert review of all cases. Our aim was to evaluate the discordances between initial and expert diagnoses and whether they would have led to different clinical management. PATIENTS AND METHODS: We conducted a retrospective analysis of the cohort of patients discussed at RYTHMIC tumour board from January 2012 to December 2016. Assessment of disagreement was made for histological typing and for staging. The discordances were classified as major or minor based on whether they would have changed or not the proposed therapeutic strategy, respectively. Follow-up of the patients with major discordances was conducted until December 2018. RESULTS: Four hundred sixty-seven patients were reviewed, and 183 (39%) discordances were identified either related to histological subtype (132) and/or stage (72). Major discordances were identified in 27 patients (6%). They included 16 patients with TET for whom treatment recommendation based on the central review would have been post-operative radiotherapy, whereas it had not been the case. However, follow-up did not show any progression among the 15 patients with high-grade histology and/or stage resected thymomas. On the other hand, among the remaining 11 patients including 7 with a diagnosis other than TET, the overall management or follow-up would have been completely different with the expert diagnosis. CONCLUSION: Our real-life cohort reveals a high level of discordances considering TET diagnosis and supports expert review for optimal clinical management.

[Thymoma and squamous thymic carcinoma diagnosis; experience from the RYTHMIC network]. / Diagnostic des thymomes et carcinomes thymiques malpighiens ; expérience du réseau RYTHMIC.

The RYTHMIC network, supported by the French National Cancer Institute is dedicated to the management of patients with thymic epithelial tumors through regional and national multidisciplinary tumor boards. Tumor board decisions are based on the initial pathology diagnoses. However, following clinical inclusion in the network, a central pathology review is organized, implicating a panel of pathologists, for histotype and stage classification, which is different from a classical second opinion from pathologist to pathologist for a difficult case. Thanks to the participation of all French pathologists, more than 1000 cases have been reviewed by the panel. The aim of this review is to share with the French pathology community, the experience of the group. It underlines the importance of macroscopy and surgeon-pathologist involvement to allow a good central review, the main histopathological and immunophenotypical patterns of the most frequent thymomas and thymic carcinoma types, the differential diagnoses, as well as the difficulties for the panel to reproducibly assess on slides, stage, for some cases.

COVID-19 pneumonia: relationship between inflammation assessed by whole-body FDG PET/CT and short-term clinical outcome.

PURPOSE: [18F]-2-Fluoro-2-deoxy-D-glucose PET/CT (FDG PET/CT) is a sensitive and quantitative technic for detecting inflammatory process. Glucose uptake is correlated with an increased anaerobic glycolysis seen in activated inflammatory cells such as monocytes, lymphocytes, and granulocytes. The aim of the study was to assess the inflammatory status at the presumed peak of the inflammatory phase in non-critically ill patients requiring admission for COVID-19. METHODS: Patients admitted with COVID-19 were prospectively enrolled. FDG PET/CT was performed from day 6 to day 14 of the onset of symptoms. Depending on FDG PET/CT findings, patients' profiles were classified as "inflammatory" or "low inflammatory." FDG PET/CT data were compared with chest CT evolution and short-term clinical outcome. All inflammatory sites were reported to screen potential extra-pulmonary tropism. RESULTS: Thirteen patients were included. Maximum standardized uptake values ranged from 4.7 to 16.3 in lungs. All patients demonstrated increased mediastinal lymph nodes glucose uptake. Three patients (23%) presented mild nasopharyngeal, two patients (15%) bone marrow, and five patients (38%) splenic mild increase in glucose uptake. No patient had significant digestive focal or segmental glucose uptake. There was no significant physiological myocardial glucose uptake in all patients except one. There was no correlation between PET lung inflammatory status and chest CT evolution or short-term clinical outcome. CONCLUSION: Inflammatory process at the presumed peak of the inflammatory phase in COVID-19 patients is obvious in FDG PET/CT scans. Glucose uptake is heterogeneous and typically focused on lungs. TRIAL REGISTRATION: NCT04441489. Registered 22 June 2020 (retrospectively registered).

Inequity in access to personalized medicine in France: Evidences from analysis of geo variations in the access to molecular profiling among advanced non-small-cell lung cancer patients: Results from the IFCT Biomarkers France Study.

In this article, we studied geographic variation in the use of personalized genetic testing for advanced non-small cell lung cancer (NSCLC) and we evaluated the relationship between genetic testing rates and local socioeconomic and ecological variables. We used data on all advanced NSCLC patients who had a genetic test between April 2012 and April 2013 in France in the frame of the IFCT Biomarqueurs-France study (n = 15814). We computed four established measures of geographic variation of the sex-adjusted rates of genetic testing utilization at the "départment" (the French territory is divided into 94 administrative units called 'départements') level. We also performed a spatial regression model to determine the relationship between département-level sex-adjusted rates of genetic testing utilization and economic and ecological variables. Our results are the following: (i) Overall, 46.87% lung cancer admission patients obtained genetic testing for NSCLC; département-level utilization rates varied over 3.2-fold. Measures of geographic variation indicated a relatively high degree of geographic variation. (ii) there was a statistically significant relationship between genetic testing rates and per capita supply of general practitioners, radiotherapists and surgeons (negative correlation for the latter); lower genetic testing rates were also associated with higher local poverty rates. French policymakers should pursue effort toward deprived areas to obtain equal access to personalized medicine for advanced NSCLC patients.

Randomized Phase II Trial Evaluating Treatment with EGFR-TKI Associated with Antiestrogen in Women with Nonsquamous Advanced-Stage NSCLC: IFCT-1003 LADIE Trial.

PURPOSE: The incidence of lung cancer has dramatically increased in women. Preclinical data have suggested that combining EGFR-tyrosine kinase inhibitor (TKI) with an antiestrogen may overcome resistance to EGFR-TKI. PATIENTS AND METHODS: The IFCT-1003 LADIE trial was a 2 × 2 arms parallel open-label randomized phase II trial. EGFR-TKI-naïve postmenopausal women with advanced lung cancer were treated with gefitinib (G) versus gefitinib + fulvestrant (G+F) in the EGFR-mutated group (EGFR+) or with erlotinib (E) versus erlotinib + fulvestrant (E+F) in the EGFR wild-type group (EGFR-WT). The primary objective was progression-free survival (PFS) at 3 and 9 months for EGFR-WT and EGFR+ patients. RESULTS: Overall, 204 patients (gefitinib 104 and G+F 100) and 175 patients (erlotinib 87 and E+F 88) were enrolled in the EGFR+ and EGFR-WT cohorts. In the EGFR+ cohort, the primary endpoint was reached, with 58% of the G+F group patients being nonprogressive at 9 months. Adding fulvestrant to gefitinib was not associated with improved PFS (9.9 vs 9.4 months) or overall survival (OS; 22.1 vs 28.6 months). In the EGFR-WT cohort, the primary endpoint was also achieved (33.7% of the patients were nonprogressive at 3 months). Adding fulvestrant to erlotinib was not associated with improved outcome (PFS 1.8 vs 2.0 and OS 10.3 vs 7.3 months). No PFS difference was observed regarding estrogen receptor alpha expression. The tolerance was as expected with no treatment-related death. CONCLUSIONS: Adding fulvestrant to EGFR-TKI is feasible, but not associated with prolonged PFS regardless of EGFR status. The lack of benefits while combining fulvestrant to EGFR-TKI does not support its future development in an unselected population.

Comprehensive Molecular and Pathologic Evaluation of Transitional Mesothelioma Assisted by Deep Learning Approach: A Multi-Institutional Study of the International Mesothelioma Panel from the MESOPATH Reference Center.

INTRODUCTION: Histologic subtypes of malignant pleural mesothelioma are a major prognostic indicator and decision denominator for all therapeutic strategies. In an ambiguous case, a rare transitional mesothelioma (TM) pattern may be diagnosed by pathologists either as epithelioid mesothelioma (EM), biphasic mesothelioma (BM), or sarcomatoid mesothelioma (SM). This study aimed to better characterize the TM subtype from a histological, immunohistochemical, and molecular standpoint. Deep learning of pathologic slides was applied to this cohort. METHODS: A random selection of 49 representative digitalized sections from surgical biopsies of TM was reviewed by 16 panelists. We evaluated BAP1 expression and CDKN2A (p16) homozygous deletion. We conducted a comprehensive, integrated, transcriptomic analysis. An unsupervised deep learning algorithm was trained to classify tumors. RESULTS: The 16 panelists recorded 784 diagnoses on the 49 cases. Even though a Kappa value of 0.42 is moderate, the presence of a TM component was diagnosed in 51%. In 49% of the histological evaluation, the reviewers classified the lesion as EM in 53%, SM in 33%, or BM in 14%. Median survival was 6.7 months. Loss of BAP1 observed in 44% was less frequent in TM than in EM and BM. p16 homozygous deletion was higher in TM (73%), followed by BM (63%) and SM (46%). RNA sequencing unsupervised clustering analysis revealed that TM grouped together and were closer to SM than to EM. Deep learning analysis achieved 94% accuracy for TM identification. CONCLUSION: These results revealed that the TM pattern should be classified as non-EM or at minimum as a subgroup of the SM type.

Correction to: Use of combined cardiac and lung ultrasound to predict weaning failure in elderly, high-risk cardiac patients: a pilot study.

The original version of this article unfortunately contained a mistake. Two of the authors forgot to mention recent collaborations in their COI. The correct COI would have been: Dr Silvia Mongodi received feed for lectures from General Electrics; and Professor Francesco Mojoli received feed for lectures from General Electrics, Hamilton Medical and SEDA SpA. The Authors apologise for the missing information.

Use of combined cardiac and lung ultrasound to predict weaning failure in elderly, high-risk cardiac patients: a pilot study.

PURPOSE: Weaning failure from mechanical ventilation may be due to lung de-recruitment or weaning-induced pulmonary oedema (WIPO). Both can be diagnosed by lung ultrasound (LUS) and transthoracic echocardiography (TTE), respectively. We conducted a prospective observational study, combining TTE and LUS, to determine if LUS alone may identify elderly patients at high risk of weaning or extubation failure. METHODS: Before and at the end of spontaneous breathing trials (SBT) in 40 elderly patients, we prospectively performed LUS and TTE. Extubation was decided by an independent operator. LUS included global and anterolateral LUS score. TTE included measurement of E/A and E/Ea ratios to determine LV filling pressures. SBT LUS scores for prediction of weaning outcome and for the diagnosis of WIPO were studied. RESULTS: Weaning or extubation failure was observed in 45% (95% CI 28-61) of patients. ROC analysis for ability of global SBT LUS to predict weaning/extubation failure and extubation failure found AUC of 0.80 and 0.81, respectively. AUC for anterolateral SBT LUS to predict weaning/extubation failure and extubation failure was 0.79 and 0.81, respectively. Increased LV filling pressure during SBT was observed without increase of anterolateral LUS score. Inversely, increase of anterolateral LUS was observed without increased filling pressure and was associated with extubation failure. Global and anterolateral SBT LUS were not correlated to E/Ea. CONCLUSION: In elderly patients, global and anterolateral LUS scores were associated with weaning and extubation failures while echocardiographic indices of filling pressures were not. CLINICAL TRIAL NUMBER AND REGISTRY URL: ClinicalTrials.gov No. NCT03261440.

Outcomes of Patients With Advanced NSCLC From the Intergroupe Francophone de Cancérologie Thoracique Biomarkers France Study by KRAS Mutation Subtypes.

INTRODUCTION: KRAS mutations are detected in 20% to 30% of NSCLC. However, KRAS mutation subtypes may differently influence the outcome of patients with advanced NSCLC. METHODS: In the Biomarkers France study, 4894 KRAS mutations (26.2%) were detected in 4634 patients from the 17,664 enrolled patients with NSCLC. Survival and treatment data on noncurative stage III to IV NSCLC were available for 901 patients. First- and second-line treatment effects on progression-free survival and overall survival were analyzed according to the KRAS mutations subtype. RESULTS: Over 95% of patients with KRAS mutation were smokers or former smokers who were white (99.5%), presenting with adenocarcinoma (82.5%). The most common KRAS mutation subtype was G12C (374 patients; 41.5%), followed by G12V (168; 18.6%), G12D (131; 14.5%), G12A (62; 6.9%), G13C (45; 5.0%), G13D (31; 3.4%), and others (10; 1%). Approximately 21% of patients had transition mutation and 68.2% had a transversion mutation. G12D and transition mutations were predominant in never-smokers. The median overall survival for patients with KRAS-mutated NSCLC was 8.1 months (95% confidence interval [CI]: 7.5-9.5), without any differences according to the different KRAS subtypes mutations. The median progression-free survival was 4.6 months (95% CI: 4.2-5.1) for first-line treatment and 4.8 months (95% CI: 4.3-6.8) for second-line treatment, without any differences according to the different KRAS subtypes mutations. CONCLUSIONS: KRAS mutation subtypes influenced neither treatment responses nor outcomes. The KRAS G12C mutation was detected in 41.5% of patients, who are now eligible for potent and specific G12C inhibitors.

Validation of the QR1 Antibody for the Evaluation of PD-L1 Expression in Non-Small Cell Lung Adenocarcinomas.

The evaluation of Programmed cell Death Ligand 1 (PD-L1) expression in the tumor cells with immunohistochemistry is a mandatory diagnostic step in the treatment of lung cancer. It is important to utilize validated antibodies that can reliably detect PD-L1 positive cells. Different antibodies have already been studied. In this present study, we compared a new clone (QR1, Quartett) with reference clones to determine if it can be used in place of previously identified reference clones. We built a tissue micro array (TMA) from 110 lung adenocarcinomas and compared it using immunohistodetection of four different clones: QR1, 22c3, Sp263, and E1L3N. We analyzed the correlation between the sample duplicates for each clone and then a correlation and the concordance between the clones were calculated. A total of 101 patients were exploitable; the duplicates for each clone had a strong correlation. The correlation was the strongest (r=0.82) between QR1 and 22c3 and less strong with the other clones. Totals of 78%, 79%, and 97% of the QR1 cases were concordant with 22c3 for the thresholds of <1%, 1% to 49%, and ≥50%, respectively. The sensitivities and specificities of QR1, compared with 22c3, were >75% and 81%, respectively. PD-L1 expression, analyzed in lung adenocarcinomas with QR1, is highly correlated and concordant with the main reference clone used in most laboratories (22c3). It can be used to replace the latter in clinical routine.

Immune biomarkers in thymic epithelial tumors: expression patterns, prognostic value and comparison of diagnostic tests for PD-L1.

BACKGROUND: Immunotherapy is currently under investigation in B3 Thymoma (TB3) and Thymic Carcinoma (TC). PD-L1 expression has been evaluated on a limited number of patients with selected antibodies. We aimed to analyze cohort of TB3 and TC with a panel of antibodies to assess the prevalence of PD-L1 expression, its prognostic value and to set up a reproducible test. METHODS: We retrospectively studied 103 patients samples of FFPE histologically confirmed TB3 (n = 53) and TC (n = 50) by expert pathologists within the RYTHMIC national network. We compared PD-L1, PD1, CD8 and PD-L2 expression and performed correlation with tumor types and patients outcomes. Four PD-L1 antibodies were tested, three of them validated as companion tests in lung cancer, one tested on two automates on whole section of tumors. We evaluated the percentage and intensity of both epithelial and immune stained cells. RESULTS: TB3 epithelial cells had a higher and more diffuse expression of PD-L1 than TC regardless the antibodies tested (p < 0.0001). Three out of four antibodies targeting PD-L1 tested on the DAKO autostainer gave similar staining. Concordance between antibodies was lower for PD-L1 staining on immune cells with no significant difference between TB3 and TC except on E1L3N antibody. PD-L2 antibody stained no tumor epithelial cells. High PD-L1 expression was correlated with a better overall survival for TB3 and was not correlated with tumor staging. CONCLUSION: Frequent PD-L1 expression, particularly in TB3, paves the way for immunotherapy in TET (Thymic Epithelial Tumor). Otherwise, we have set up three reproducible LDT (laboratory-developed test) for four PD-L1 antibodies.

Redefining malignant pleural mesothelioma types as a continuum uncovers immune-vascular interactions.

BACKGROUND: Malignant Pleural Mesothelioma (MPM) is an aggressive disease related to asbestos exposure, with no effective therapeutic options. METHODS: We undertook unsupervised analyses of RNA-sequencing data of 284 MPMs, with no assumption of discreteness. Using immunohistochemistry, we performed an orthogonal validation on a subset of 103 samples and a biological replication in an independent series of 77 samples. FINDINGS: A continuum of molecular profiles explained the prognosis of the disease better than any discrete model. The immune and vascular pathways were the major sources of molecular variation, with strong differences in the expression of immune checkpoints and pro-angiogenic genes; the extrema of this continuum had specific molecular profiles: a "hot" bad-prognosis profile, with high lymphocyte infiltration and high expression of immune checkpoints and pro-angiogenic genes; a "cold" bad-prognosis profile, with low lymphocyte infiltration and high expression of pro-angiogenic genes; and a "VEGFR2+/VISTA+" better-prognosis profile, with high expression of immune checkpoint VISTA and pro-angiogenic gene VEGFR2. We validated the gene expression levels at the protein level for a subset of five selected genes belonging to the immune and vascular pathways (CD8A, PDL1, VEGFR3, VEGFR2, and VISTA), in the validation series, and replicated the molecular profiles as well as their prognostic value in the replication series. INTERPRETATION: The prognosis of MPM is best explained by a continuous model, which extremes show specific expression patterns of genes involved in angiogenesis and immune response.

[Guidelines for the macroscopic management of surgically resected lung carcinoma]. / Recommandations SFP pour la prise en charge macroscopique des pièces de résections de tumeurs pulmonaires.

Gross examination is an essential step for pathological report of a surgical sample. It includes the description of the surgical specimen and their disease(s), the precise and exhaustive sampling of tumoral and adjacent tumoral tissue areas. This examination requires a good knowledge of the updated pTNM classification. Pathologists from the PATTERN group have collaborated with thoracic surgeons, under the auspices of the Sociéte française de pathologie, to propose guidelines for resected specimen management. This approach fits into the context of the elaboration of structured pathological report proposed by the société française de pathologie, which is necessary for a standardized management of patients.

Targeted sequencing of plasma cell-free DNA to predict response to PD1 inhibitors in advanced non-small cell lung cancer.

OBJECTIVES: Tumor mutational burden is an emerging biomarker of response to immune checkpoint inhibitors (ICI), whose clinical adoption is challenging. We hypothesized that targeting limited but relevant genetic alterations in plasma cell-free DNA along with early monitoring may non-invasively predict response to ICI in advanced non-small cell lung cancer (NSCLC). MATERIAL AND METHODS: Plasma samples from patients with progressive NSCLC collected before ICI initiation and at 1 month were profiled from responders (R: PFS > 6 months) and non-responders (NR: progressive disease at first evaluation) using amplicon sequencing of hotspots and coding regions from 36 genes. The molecular profile of ctDNA, and its early kinetics were analyzed. RESULTS: 97 patients were analyzed, of which 86 (39 R, 47 NR) were evaluable. Alterations in ctDNA were detectable in 67/86 baseline samples (78%). The detection of a targetable oncogenic driver was associated with a 2 months PFS. The presence of a PTEN or STK11 mutation was correlated with early progression (HR 8.9, p = 0.09 for PTEN, HR 4.7, p = 0.003 for STK11), while transversion mutations (Tv) in KRAS and TP53 predicted better outcomes (HR 0.36, p = 0.011 for TP53 Tv; HR 0.46, p = 0.11 for KRAS Tv). Patients with a low "immune score" (driver and/or PTEN or STK11 mutation and/or without KRAS or TP53 Tv) derived poor outcomes (median PFS 2 months), compared with patients with a high immune score (no driver, no PTEN or STK11 and with KRAS or TP53 Tv (median PFS 14 months, p = 0.0001, HR 2.96). Early changes in the ctDNA allele fraction (AF) of 65 specimens were correlated with clinical outcomes (14 months PFS if AF decreases vs. 2 months if AF increases, p < 0.0001). CONCLUSION: Targeted sequencing of plasma ctDNA and monitoring its early variations can predict response to ICI.

Multicenter Evaluation of the Fully Automated PCR-Based Idylla EGFR Mutation Assay on Formalin-Fixed, Paraffin-Embedded Tissue of Human Lung Cancer.

Before initiating treatment of advanced non-small-cell lung cancer with tyrosine kinase inhibitors (eg, erlotinib, gefitinib, osimertinib, and afatinib), which inhibit the catalytic activity of epidermal growth factor receptor (EGFR), clinical guidelines require determining the EGFR mutational status for activating (EGFR exons 18, 19, 20, or 21) and resistance (EGFR exon 20) mutations. The EGFR resistance mutation T790M should be monitored at cancer progression. The Idylla EGFR Mutation Assay, performed on the Idylla molecular diagnostics platform, is a fully automated (<2.5 hours turnaround time) sample-to-result molecular test to qualitatively detect 51 EGFR oncogene point mutations, deletions, or insertions. In a 15-center evaluation, Idylla results on 449 archived formalin-fixed, paraffin-embedded tissue sections, originating from non-small-cell lung cancer biopsies and resection specimens, were compared with data obtained earlier with routine reference methods, including next-generation sequencing, Sanger sequencing, pyrosequencing, mass spectrometry, and PCR-based assays. When results were discordant, a third method of analysis was performed, when possible, to confirm test results. After confirmation testing and excluding invalids/errors and discordant results by design, a concordance of 97.6% was obtained between Idylla and routine test results. Even with <10 mm2 of tissue area, a valid Idylla result was obtained in 98.9% of the cases. The Idylla EGFR Mutation Assay enables sensitive detection of most relevant EGFR mutations in concordance with current guidelines, with minimal molecular expertise or infrastructure.

Shorter Survival in Malignant Pleural Mesothelioma Patients With High PD-L1 Expression Associated With Sarcomatoid or Biphasic Histology Subtype: A Series of 214 Cases From the Bio-MAPS Cohort.

BACKGROUND: Anticancer immune responses are negatively regulated by programmed cell death 1 (PD-1) T-cell membrane protein interaction with its ligand, programmed death ligand 1 (PD-L1), on cancer cells. We sought to assess the prognostic role of PD-L1 expression in tumor samples from patients enrolled onto the IFCT-0701 MAPS randomized phase 3 trial (NCT00651456). PATIENTS AND METHODS: Tumor samples were analyzed by immunohistochemistry for percentages of PD-L1 membrane-stained tumor cells using the E1L3N clone, and data were correlated to survival by multivariate Cox models including stratification variables. RESULTS: PD-L1 staining was assessed in 214 (47.75%) of 448 patients. Epithelioid subtype represented 83.7% (179/214). Absence of PD-L1 staining occurred in 137 (64.1%) of 214 malignant pleural mesothelioma (MPM) samples, while 77 (35.9%) of 214 were PD-L1 positive, with 50 (64.9%) of 77 showing < 50% PD-L1-expressing tumor cells. Sarcomatoid/biphasic subtypes were more commonly PD-L1 positive than epithelioid subtype (P < .001). In patients with 1% or more PD-L1-stained tumor cells, median overall survival (OS) was 12.3 months versus 22.2 months for other patients (hazard ratio [HR] = 1.25; 95% confidence interval [CI], 0.93-1.67; P = .14). OS did not differ according to PD-L1 positivity in multivariate analyses (adjusted HR = 1.10; 95% CI, 0.81-1.49; P = .55). With a 50% cutoff, PD-L1-positive patients displayed a 10.5 months median OS versus 19.3 months for patients with lower PD-L1 expression (HR = 1.93; 95% CI, 1.27-2.93; P = .002). OS did not significantly differ in adjusted Cox models (adjusted HR = 1.20; 95% CI, 0.74-1.94; P = .47). In the 179 epithelioid MPM patients, high PD-L1 staining (≥ 50% of tumor cells) negatively affected OS, although not significantly, showing a 12.3-month median OS (95% CI, 4.3-21.6) versus 23-month (95% CI, 18.5-25.2) for patients with tumor PD-L1 staining in < 50% cells (P = .071). The progression-free survival (PFS) differences were statistically significant, with a longer 9.9-month median PFS in patients with low PD-L1 staining (< 50% cells) compared to 6.7 months of median PFS in patients with high PD-L1 expression (≥ 50% cells) (P = .0047). CONCLUSION: Although high PD-L1 tumor cell expression was associated with poorer OS in MPM patients from the MAPS trial, its prognostic influence was lost in multivariate analyses in the whole cohort, while PD-L1 expression was strongly associated with the sarcomatoid/biphasic subtypes. In the epithelioid MPM subset of patients, high PD-L1 tumor expression (≥ 50%) negatively affected OS and PFS, with this prognostic influence remaining statistically significant for PFS after adjustment in multivariate Cox model.

Ultra-sensitive EGFR T790M Detection as an Independent Prognostic Marker for Lung Cancer Patients Harboring EGFR del19 Mutations and Treated with First-generation TKIs.

PURPOSE: The detection of preexisting EGFR T790M subclones and the assessment of their clinical significance in the pretreatment of patients with EGFR T790M non-small cell lung cancer (NSCLC) remain unclear. EXPERIMENTAL DESIGN: A total of 179 tumor samples from patients treated or not with a first-generation tyrosine kinase inhibitor (TKI) was analyzed. The presence of ultra-low levels of preexisting EGFRT790M mutation was evaluated using ultra-sensitive droplet digital PCR (ddPCR) and the clinical implication of these mutations on first-generation TKI efficiency assessed. RESULTS: With a ddPCR linear performance of 0.999 and an analytical sensitivity of approximately 0.001%, we observed a 66% (99/150) overall incidence of ultra-low EGFR T790M mutation. Among 82 patients harboring EGFR activating mutations, the presence of a preexisting EGFR T790M mutation prior to any treatment was significantly associated with a longer progression-free survival (PFS; P = 0.009; log-rank test). Interestingly, longer PFS was linked to concomitant EGFR del19 and ultra-low EGFR T790M mutations. Moreover, the presence of both EGFR del19 and ultra-low EGFR T790M mutations was identified as the best fit for predicting the clinical outcome of patients treated with TKI compared with an ultra-low EGFR T790M mutation status or an activating mutation alone (P = 0.042 and P = 0.0071, respectively). CONCLUSIONS: We demonstrate that the detection of the ultra-low EGFR T790M mutation in TKI-naïve patients is not a rare event. We suggest that ddPCR should be used in clinical practice to distinguish patients who may respond to first- or third-generation TKIs.