Since the outbreak of the COVID-19 pandemic, much has been learned regarding its clinical course, prognostic inflammatory markers, disease complications, and mechanical ventilation strategy. ...Clinically, three stages have been identified based on viral infection, pulmonary involvement with inflammation, and fibrosis. Moreover, low and high elastance phenotypes can be distinguished in mechanically ventilated patients, based on lung mechanics, ventilation-to-perfusion ratio, and CT scans; these two phenotypes have presumed differences in their underlying pathophysiology. Although essential for therapeutic guidance, the pathophysiology of COVID-19 is poorly understood. Here, we systematically reviewed published case reports and case series in order to increase our understanding of COVID-19 pathophysiology by constructing a timeline and correlating histopathological findings with clinical stages of COVID-19. Using PRISMA-IPD guidelines, 42 articles reporting 198 individual cases were included in our analysis. In lung samples (n = 131 cases), we identified three main histological patterns: epithelial (n = 110, 85%), with reactive epithelial changes and DAD; vascular (n = 76, 59%) with microvascular damage, (micro)thrombi, and acute fibrinous and organizing pneumonia; and fibrotic (n = 28, 22%) with interstitial fibrosis. The epithelial and vascular patterns can present in all stages of symptomatic COVID-19, whereas the fibrotic pattern presents starting at ~3 weeks. Moreover, patients can present with more than one pattern, either simultaneously or consecutively. These findings are consistent with knowledge regarding clinical patterns of viral infection, development of hyperinflammation and hypercoagulability, and fibrosis. Close collaboration among medical staff is necessary in order to translate this knowledge and classification of pathophysiological mechanisms into clinical stages of disease in individual patients. Moreover, further research, including histopathological studies, is warranted in order to develop reliable, clinically relevant biomarkers by correlating these pathological findings with laboratory results and radiological findings, thus, increasing our understanding of COVID-19 and facilitating the move to precision medicine for treating patients.
This meeting report from the XV Banff conference describes the creation of a multiorgan transplant gene panel by the Banff Molecular Diagnostics Working Group (MDWG). This Banff Human Organ ...Transplant (B‐HOT) panel is the culmination of previous work by the MDWG to identify a broadly useful gene panel based on whole transcriptome technology. A data‐driven process distilled a gene list from peer‐reviewed comprehensive microarray studies that discovered and validated their use in kidney, liver, heart, and lung transplant biopsies. These were supplemented by genes that define relevant cellular pathways and cell types plus 12 reference genes used for normalization. The 770 gene B‐HOT panel includes the most pertinent genes related to rejection, tolerance, viral infections, and innate and adaptive immune responses. This commercially available panel uses the NanoString platform, which can quantitate transcripts from formalin‐fixed paraffin‐embedded samples. The B‐HOT panel will facilitate multicenter collaborative clinical research using archival samples and permit the development of an open source large database of standardized analyses, thereby expediting clinical validation studies. The MDWG believes that a pathogenesis and pathway based molecular approach will be valuable for investigators and promote therapeutic decision‐making and clinical trials.
This Banff meeting report summarizes the progress of the Banff Molecular Diagnostics Working Group, which generated consensus on a transplant‐specific discovery gene panel and a potential roadmap for its validation for diagnostic application.
Aims
Immunohistochemical programmed death‐ligand 1 (PD‐L1) staining to predict responsiveness to immunotherapy in patients with advanced non‐small cell lung cancer (NSCLC) has several drawbacks: a ...robust gold standard is lacking, and there is substantial interobserver and intraobserver variance, with up to 20% discordance around cutoff points. The aim of this study was to develop a new deep learning‐based PD‐L1 tumour proportion score (TPS) algorithm, trained and validated on a routine diagnostic dataset of digitised PD‐L1 (22C3, laboratory‐developed test)‐stained samples.
Methods and results
We designed a fully supervised deep learning algorithm for whole‐slide PD‐L1 assessment, consisting of four sequential convolutional neural networks (CNNs), using aiforia create software. We included 199 whole slide images (WSIs) of ‘routine diagnostic’ histology samples from stage IV NSCLC patients, and trained the algorithm by using a training set of 60 representative cases. We validated the algorithm by comparing the algorithm TPS with the reference score in a held‐out validation set. The algorithm had similar concordance with the reference score (79%) as the pathologists had with one another (75%). The intraclass coefficient was 0.96 and Cohen’s κ coefficient was 0.69 for the algorithm. Around the 1% and 50% cutoff points, concordance was also similar between pathologists and the algorithm.
Conclusions
We designed a new, deep learning‐based PD‐L1 TPS algorithm that is similarly able to assess PD‐L1 expression in daily routine diagnostic cases as pathologists. Successful validation on routine diagnostic WSIs and detailed visual feedback show that this algorithm meets the requirements for functioning as a ‘scoring assistant’.
Lymph node micrometastases could be one of the reasons for the high recurrence rate after complete surgical resection in stage I–IIIA non‐small cell lung cancer (NSCLC). The standard evaluation of a ...single haematoxylin and eosin (H&E) slide of a paraffin‐embedded section of a lymph node is insufficient for the detection of micrometastases, and there is a need for additional histopathological evaluation. The association of lymph node micrometastases with survival remains as yet unresolved. The aim of this systematic review and meta‐analysis is to investigate if lymph node micrometastases and isolated tumour cells in patients with stage I–IIIA NSCLC, detected with multiple sectioning and/or immunohistochemistry (IHC) and/or reverse transcriptase polymerase chain reaction (RT‐PCR), are associated with overall survival (OS) and disease‐free survival (DFS) after surgical resection. We performed a meta‐analysis of time‐to‐event outcomes based on 15 articles using ancillary techniques to detect micrometastases. We extracted the OS and DFS every 3–6 months after surgery, for patients with and without occult lymph node micrometastasis, from the survival curves published in each article. These data were used to reconstruct OS and DFS for ‘micrometastasis’ and ‘no micrometastasis’ groups. Based on all included studies that used IHC, serial sectioning, or RT‐PCR, we found a 5‐year OS of 55% (micrometastasis) vs. 75% (no micrometastasis), and a 5‐year DFS of 53% (micrometastasis) vs. 75% (no micrometastasis). Patients with stage I–IIIA NSCLC with lymph node micrometastases detected by ancillary histopathological and molecular techniques have a significantly poorer OS and DFS compared to patients without lymph node micrometastases.
Objectives
A minority of NSCLC patients benefit from anti-PD1 immune checkpoint inhibitors. A rational combination of biomarkers is needed. The objective was to determine the predictive value of ...tumor mutational load (TML), CD8
+
T cell infiltration, HLA class-I and PD-L1 expression in the tumor.
Materials and methods
Metastatic NSCLC patients were prospectively included in an immune-monitoring trial (NTR7015) between April 2016-August 2017, retrospectively analyzed in FFPE tissue for TML (NGS: 409 cancer-related-genes) and by IHC staining to score PD-L1, CD8
+
T cell infiltration, HLA class-I. PFS (RECISTv1.1) and OS were analyzed by Kaplan–Meier methodology.
Results
30 patients with adenocarcinoma (67%) or squamous cell carcinoma (33%) were included. High TML was associated with better PFS (
p
= 0.004) and OS (
p
= 0.025). Interaction analyses revealed that patients with both high TML and high total CD8
+
T cell infiltrate (
p
= 0.023) or no loss of HLA class-I (
p
= 0.026), patients with high total CD8
+
T cell infiltrate and no loss of HLA class-I (
p
= 0.041) or patients with both high PD-L1 and high TML (
p
= 0.003) or no loss of HLA class-I (
p
= 0.032) were significantly associated with better PFS. Unsupervised cluster analysis based on these markers revealed three sub-clusters, of which cluster-1A was overrepresented by patients with progressive disease (15 out of 16), with significant effect on PFS (
p
= 0.007).
Conclusion
This proof-of-concept study suggests that a combination of PD-L1 expression, TML, CD8
+
T cell infiltration and HLA class-I functions as a better predictive biomarker for response to anti-PD-1 immunotherapy. Consequently, refinement of this set of biomarkers and validation in a larger set of patients is warranted.
Aims
Mucinous adenocarcinoma arising in congenital pulmonary airway malformation (CPAM) is a rare complication, with little being known about its natural course. The aims of this article are to ...describe a series of mucinous adenocarcinomas arising from CPAMs, and present their clinicopathological features, genetics, and clinical outcome.
Methods and results
Thirty‐seven cases were collected within a 34‐year period, and the subtype of adenocarcinoma and CPAM, tumour location, stage, growth patterns, molecular data and follow‐up were recorded. The cohort comprised CPAM type 1 (n = 33) and CPAM type 2 (n = 4). Morphologically, 34 cases were mucinous adenocarcinomas (21 in situ; 13 invasive), and three were mixed mucinous and non‐mucinous adenocarcinoma. Seventeen cases showed purely extracystic (intra‐alveolar) adenocarcinoma, 15 were mixed intracystic and extracystic, and five showed purely intracystic proliferation. Genetically, nine of 10 cases tested positive for KRAS mutations, four with exon 2 G12V mutation and five with exon 2 G12D mutation. Residual disease on completion lobectomy was observed in two cases, and three cases recurred 7, 15 and 32 years after the original diagnosis. Two patients died of metastatic invasive mucinous adenocarcinoma.
Conclusions
Most adenocarcinoma that arise in type 1 CPAMs, are purely mucinous, and are early‐stage disease. Intracystic proliferation is associated with lepidic growth, an absence of invasion, and indolent behaviour, whereas extracystic proliferation may be associated with more aggressive behaviour and advanced stage. Most cases are cured by lobectomy, and recurrence/residual disease seems to be associated with limited surgery. Long‐term follow‐up is needed, as recurrence can occur decades later.
Aims
COVID‐19 pneumonia is characterized by an increased rate of deep venous thrombosis and pulmonary embolism. To better understand the pathophysiology behind thrombosis in COVID‐19, we performed ...proteomics analysis on SARS‐CoV‐2 infected lung tissue.
Methods
Liquid chromatography mass spectrometry was performed on SARS‐CoV‐2 infected postmortem lung tissue samples. Five protein profiling analyses were performed: whole slide lung parenchyma analysis, followed by analysis of isolated thrombi and endothelium, both stratified by disease (COVID‐19 versus influenza) and thrombus morphology (embolism versus in situ). Influenza autopsy cases with pulmonary thrombi were used as controls.
Results
Compared to influenza controls, both analyses of COVID‐19 whole‐tissue and isolated endothelium showed upregulation of proteins and pathways related to liver metabolism including urea cycle activation, with arginase being among the top upregulated proteins in COVID‐19 lung tissue. Analysis of isolated COVID‐19 thrombi showed significant downregulation of pathways related to platelet activation compared to influenza thrombi. Analysis of isolated thrombi based on histomorphology shows that in situ thrombi have significant upregulation of coronavirus pathogenesis proteins.
Conclusions
The decrease in platelet activation pathways in severe COVID‐19 thrombi suggests a relative increase in venous thromboembolism, as thrombi from venous origin tend to contain fewer platelets than arterial thrombi. Based on histomorphology, in situ thrombi show upregulation of various proteins related to SARS‐CoV‐2 pathogenesis compared to thromboemboli, which may indicate increased in situ pulmonary thrombosis in COVID‐19. Therefore, this study supports the increase of venous thromboembolism without undercutting the involvement of in situ thrombosis in severe COVID‐19.
A decrease in platelet activation pathways in COVID‐19 thrombi suggests a relative increase in venous thromboembolism, as thrombi from venous origin tend to contain fewer platelets than arterial thrombi. In situ thrombi show upregulation of SARS‐CoV‐2 pathogenesis proteins compared to thromboemboli, underscoring the significance of in situ pulmonary thrombosis in COVID‐19.
Malignant mesothelioma (MM) is an aggressive cancer that primarily arises from the pleura (MPM) or peritoneum (MPeM), mostly due to asbestos exposure. This study reviewed the Dutch population-based ...incidence, treatment and survival since the national ban on asbestos in 1993.
Patients with MPM or MPeM diagnosed from 1993 to 2018 were selected from the Dutch cancer registry. Annual percentage change (APC) was calculated for (age-specific and sex-specific) revised European standardised incidence rates (RESR). Treatment pattern and Kaplan-Meier overall survival analyses were performed.
In total, 12 168 patients were included in the study. For male patients younger than 80 years, the MM incidence significantly decreased in the last decade (APC ranging between -9.4% and -1.8%, p<0.01). Among both male and female patients aged over 80 years, the incidence significantly increased during the entire study period (APC 3.3% and 4.6%, respectively, p<0.01). From 2003 onwards, the use of systemic chemotherapy increased especially for MPM (from 9.3% to 39.4%). Overall, 62.2% of patients received no antitumour treatment. The most common reasons for not undergoing antitumour treatment were patient preference (42%) and performance status (25.6%). The median overall survival improved from 7.3 (1993-2003) to 8.9 (2004-2011) and 9.3 months from 2012 to 2018 (p<0.001).
The peak of MM incidence was reached around 2010 in the Netherlands, and currently the incidence is declining in most age groups. The use of systemic chemotherapy increased from 2003, which likely resulted in improved overall survival over time. The majority of patients do not receive treatment though and prognosis is still poor.
Background
Immunohistochemical staining of programmed death‐ligand 1 (PD‐L1) is used to determine which patients with non–small cell lung cancer (NSCLC) may benefit most from immunotherapy. ...Therapeutic management of many patients with NSCLC is based on cytology instead of histology. In this study, concordance of PD‐L1 immunostaining between cytology cell blocks and their histologic counterparts was analyzed. Furthermore, the effect of various fixatives and fixation times on PD‐L1 immunoreactivity was studied.
Methods
Paired histologic and cytologic samples from 67 patients with NSCLC were collected by performing fine‐needle aspiration on pneumonectomy/lobectomy specimens. Formalin‐fixed, agar‐based or CytoLyt/PreservCyt‐fixed Cellient cell blocks were prepared. Sections from cell blocks and tissue blocks were stained with SP263 (standardized assay) and 22C3 (laboratory‐developed test) antibodies. PD‐L1 scores were compared between histology and cytology. In addition, immunostaining was compared between PD‐L1–expressing human cell lines fixed in various fixatives at increasing increments in fixation duration.
Results
Agar cell blocks and tissue blocks showed substantial agreement (κ = 0.70 and κ = 0.67, respectively), whereas fair‐to‐moderate agreement was found between Cellient cell blocks and histology (κ = 0.28 and κ = 0.49, respectively). Cell lines fixed in various alcohol‐based fixatives showed less PD‐L1 immunoreactivity compared with those fixed in formalin. In contrast to SP263, additional formalin fixation after alcohol fixation resulted in preserved staining intensity using the 22C3 laboratory‐developed test and the 22C3 pharmDx assay.
Conclusions
Performing PD‐L1 staining on cytologic specimens fixed in alcohol‐based fixatives could result in false‐negative immunostaining results, whereas fixation in formalin leads to higher and more histology‐concordant PD‐L1 immunostaining. The deleterious effect of alcohol fixation could be reversed to some degree by postfixation in formalin.
Fixation of cytologic specimens in alcohol‐based fixatives results in negative effects on programmed death‐ligand 1 (PD‐L1) immunostaining when using PD‐L1 immunohistochemistry protocols validated on formalin‐fixed, paraffin‐embedded material. The use of formalin results in more histology‐concordant PD‐L1 immunostaining, and the deleterious effect of alcohol fixation potentially may be reversed to some degree by postfixation in formalin.
Chronic lung allograft dysfunction continues to be the main contributor to poor long-term allograft survival after lung transplantation. The restrictive phenotype of chronic lung allograft ...dysfunction carries a particularly poor prognosis. Little is known about the pathogenetic mechanisms involved in restrictive chronic lung allograft dysfunction. In this study, we performed histomorphological and immunohistochemical analysis of restrictive chronic lung allograft dysfunction lungs. Explant lung tissue from 21 restrictive chronic lung allograft dysfunction patients was collected and histopathologic patterns of rejection, fibrosis and vascular changes were scored after routine histochemical stains and additional immunohistochemistry for endothelial markers and C4d. In all, 75% of cases showed evidence of acute cellular rejection; lymphocytic bronchiolitis was absent in most lungs, whereas in 55% there was obliterative bronchiolitis. Almost half of the cases showed a pattern consistent with pleuroparenchymal fibro-elastosis (n=10), and a subset showed nonspecific interstitial pneumonia (n=5) or irregular emphysema (n=5). Fibrinous alveolar exudates were frequently seen in association with fibrosis (n=6), but no diffuse alveolar damage was found. Evidence of microvascular damage was present in most cases. An emphysematous pattern of fibrosis was associated with a better survival (P=0.0030), whereas fibrinous exudates were associated with a worse survival (P=0.0007). In addition to the previously described nonspecific interstitial pneumonia and pleuroparenchymal fibro-elastosis patterns in restrictive chronic lung allograft dysfunction, we are the first to describe a pattern of fibrosis-induced subpleural/paraseptal emphysema. This pattern confers a better survival, whereas fibrinous exudates are associated with a worse survival. We believe that our findings offer a pathogenetic theory for pleuroparenchymal fibro-elastosis in restrictive chronic lung allograft dysfunction, and show that restrictive chronic lung allograft dysfunction is an increasingly heterogeneous disease with presumably different mechanisms of subpattern formation.
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